The upstream causes of the COVID-19 pandemic have received little attention so far in public health and clinical medicine, as opposed to the downstream effects of mass morbidity and mortality. To resolve this pandemic and to prevent even more severe future pandemics, a focus on upstream causation is essential. Convincing evidence shows that this and every other important viral epidemic emerging in the recent past and predictably into the future comes from the same upstream causes: capitalist agriculture, its destruction of natural habitat, and the industrial production of meat. International and national health organizations have obscured the upstream causes of emerging viral epidemics. These organizations have suffered cutbacks in public funding but have received increased support from international financial institutions and private philanthropies that emphasize the downstream effects rather than upstream causes of infectious diseases. Conflicts of interest also have impacted public health policies. A worldwide shift has begun toward peasant agricultural practices: Research so far has shown that peasant agriculture is safer and more efficient than capitalist industrial agricultural practices. Without such a transformation of agriculture, even more devastating pandemics will result from the same upstream causes. While struggling to lessen the many downstream effects of COVID-19, through dangerous and sometimes heroic efforts, we in medicine and public health also must try to maintain a clear and relentless focus upstream on the root causes of the pandemic. Clarity about the upstream causes rarely emerges in current discussions of the pandemic, even in the scientific and public health communities, and hardly ever in pronouncements of the public health institutions on which many of us rely: the World Health Organization (WHO), U.S. Centers for Disease Control and Prevention (CDC), U.S. National Institute of Allergy and Infectious Diseases, Pan American Health Organization, Gates Foundation, and so forth. Identifying the upstream causes of epidemics has been a central goal of epidemiology since the 1840s, when the pathologist Rudolf Virchow did his path-breaking investigation of the typhus epidemic in Upper Silesia. Due to the current pandemic’s magnitude, one would expect that the upstream causes would be crystal-clear for all to see, so we could address them directly, but amazingly this is not the case.Many people attribute the origins of the pandemic to the strange and retro marketing practices of some individuals and groups in Wuhan, China, who were selling wild animals in the market from which the virus spread, eventually worldwide. But such marketing practices have been going on for a long time, probably hundreds of years or more. Why do we have a pandemic now and not earlier?16 In addition to viral epidemics, these and similar agricultural practices also deepen the parallel crises of multi-drug-resistant bacterial infections , climate change (by destruction of rainforest habitats and long-distance transportation of food that requires burning of petroleum), plastic pollution , and other severe environmental problems.This pandemic and every other important emerging viral epidemic in the recent past and predictably into the future come from the same upstream causes: capitalist industrial agriculture, destruction of natural habitat, and production of meat. In recent decades, the intensity and worldwide scale of these practices have increased rapidly. Pioneering microbiological and epidemiological studies have clarified these upstream causes of emerging epidemics, whose effects we now are confronting every day.7 Similar zoonotic sources of transmission from destroyed habitats have happened in China with the previous coronavirus in severe acute respiratory syndrome (SARS); Ebola in Africa; Zika in Africa, Latin America, and elsewhere; and arguably HIV in Africa.810Natural forest habitat previously provided ecological control for microbes such as SARS-CoV-2 and their hosts, such as bats. Clearing habitat for industrial agriculture emerged as a central characteristic of China’s economy as it “liberalized” after Mao Zedong into a bastion of the capitalist world system.25Another practice stemming from the capitalist model of agriculture involves industrial production of meat. Especially for pigs and chickens but also other species, reproduction of offspring, growth to adulthood, slaughter, and packaging increasingly occur under factory conditions that receive little regulatory oversight and control. Worldwide, a small number of large, oligopolistic, multinational corporations dominate factory farming. As a result, viral contamination and mutations to more virulent organisms in unsanitary factory conditions have led to epidemics of swine flu, avian flu, and a variety of emerging influenza viruses.12Do sources like WHO, CDC, and the Gates Foundation provide a complete picture? Some well-motivated people work for these agencies, and much helpful information is available. But mistakes get made, as have occurred multiple times during the COVID-19 pandemic, and more importantly these sources rarely address the upstream causes of epidemics. Many have commented about the devastating funding cutbacks and de-prioritization that have crippled these organizations’ capacity to protect public health. As just one example, the annual program budget of WHO for the whole world is smaller by about half than the operating budget of a large medical center in the United States .13 This is one reason that the global People’s Health Movement produces “WHO Watch” and “Global Health Watch” to monitor WHO critically and to offer alternatives that WHO and its affiliates do not pursue because of their financial dependency on international financial institutions and philanthrocapitalism.14 Partly due to such financial support, international and national health organizations almost always promote reductionist initiatives that focus on so-called magic bullets such as vaccines and antiviral medications, as well as behavioral change at the level of individuals, rather than upstream causes.Into the financial crisis of international health institutions have stepped the World Bank, International Monetary Fund, Gates Foundation, and other agencies of “philanthrocapitalism,” whose financial priorities and ideologies dominate the policies and practices of WHO and its affiliated organizations worldwide.15 In addition, Gates’ investments emphasize pharmaceutical corporations and other companies that profit from intellectual property, which in the realm of computer software creates most of Gates’ wealth. Similarly, CDC and its employees regularly attract criticism based on revelations about conflicts of interest at both the organizational level  and individual level (employees’ and committee members’ investments in and gifts from industry).16Financial conflicts of interest also can distort the organizations’ policies. For instance, the Gates Foundation has invested in and promoted genetically modified crops through such corporations as Monsanto/Bayer. Farmlands for such crops lead largely to the production of animal feeds, required for increased meat production, which causes further loss of forest habitat.So WHO, CDC, Gates, and their affiliates have obscured the upstream causes of emerging viral epidemics not only in COVID-19 but also in all other recent epidemics. An especially disheartening example (I was involved) was the swine flu epidemic of 2009, which began within 1 mile of Smithfield Foods’ notorious industrial pig farm operation in a rural area of Veracruz state in Mexico. Smithfield Foods had outsourced this operation from the United States partly to avoid occupational and environmental cleanup requirements. Although Mexican public health authorities and investigators reported this epidemiological association between swine flu and capitalist industrial agriculture, CDC, WHO, and all other international health organizations pursued reductionist strategies such as a vaccine, rather than confront radical change in the meat processing industry.17 When COVID-19 struck, Smithfield executives reassured U.S. consumers that, despite ownership in Hong Kong, the corporation did not import pork from China but instead exported U.S. pork to China, where prices were higher.18 A Smithfield pork processing plant in Sioux Falls, South Dakota, became one of the largest COVID-19 hotspots in the United States. Rapid spread of the infection to workers because of similarly unsanitary working conditions threatened to close other meat-producing plants as well.19During the COVID-19 pandemic, Smithfield’s practices became even more startling. Less than a decade after the swine flu epidemic, a Hong Kong-based investment corporation, WH Group Ltd, had acquired Smithfield Foods. In 2018 Smithfield executives based at U.S. headquarters in Smithfield, Virginia, welcomed the ongoing epidemic of African swine fever because a reduced global supply of pork would lead to major increases in prices and profitability for the corporation.20 But the impacts of such corporations on emerging epidemics rarely appear in communications or policies of international health organizations or the Gates Foundation.The role of capitalist industrial agriculture through loss of habitat and meat production occasionally does surface in the mainstream media. Such media attention, while limited, happened recently regarding the sources of COVID-19, although the term “capitalist” did not enter the discussion.21 In this “tabletop exercise,” coordinated by the Johns Hopkins Center for Health Security, Gates Foundation, and World Economic Forum, a novel coronavirus pandemic begins at pig farms in Brazil and spreads rapidly around the world, resulting in 65 million deaths and catastrophic effects on the global economy, political stability, and international security. After the COVID-19 epidemic actually began, the sponsors of Event 201 emphasized that they did not predict the timing of COVID-19 and that the projected death toll did not necessarily apply. But they did not say anything about an initiative to eradicate the practices of capitalist industrial agriculture that led to the hypothetical scenario of Event 201, to the current global COVID-19 pandemic, and to the inevitable future pandemics that will occur on a similar scale or even worse.Leaders of these agencies are fully aware that emerging viral epidemics come from capitalist industrial agriculture. They showed this awareness in Event 201 on October 18, 2019, ironically about 2 months before the COVID-19 epidemic began in Wuhan.22 Millions of people worldwide already are making this transition, often because they/we see no other choice. Especially in the context of economic collapse, capitalist agriculture – with its tendency to overproduce and even destroy surplus food while hunger and food insecurity worsen – is ill suited to feed the world’s peoples.Is there an alternative to capitalist industrial agriculture? Yes. Around the world, often against resistance from corporations and governments, farmers are returning to peasant agricultural practices. A whole body of research has shown that peasant agriculture is not only safer than capitalist agriculture but actually is more efficient and productive as well.Changing the upstream causes of epidemics such as COVID-19 and others yet to come becomes a key scientific and practical priority for medicine and public health, considering the future of humanity and other inhabitants of the planet. If that transformation doesn’t happen, we can expect even more devastating pandemics, stemming from the same upstream causes.


Licensed drugs may cause unexpected adverse reactions in patients, resulting in morbidity, risk of mortality, therapy disruptions, and prolonged hospital stays. Officially approved drug package inserts list the adverse reactions identified from randomized controlled clinical trials with high evidence levels and worldwide postmarketing surveillance. Formal representation of the adverse drug reaction (ADR) enclosed in semistructured package inserts will enable deep recognition of side effects and rational drug use, substantially reduce morbidity, and decrease societal costs.This paper aims to present an ontological organization of traceable ADR information extracted from licensed package inserts. In addition, it will provide machine-understandable knowledge for bioinformatics analysis, semantic retrieval, and intelligent clinical applications.Based on the essential content of package inserts, a generic ADR ontology model is proposed from two dimensions (and nine subdimensions), covering the ADR information and medication instructions. This is followed by a customized natural language processing method programmed with Python to retrieve the relevant information enclosed in package inserts. After the biocuration and identification of retrieved data from the package insert, an ADR ontology is automatically built for further bioinformatic analysis.We collected 165 package inserts of quinolone drugs from the National Medical Products Administration and other drug databases in China, and built a specialized ADR ontology containing 2879 classes and 15,711 semantic relations. For each quinolone drug, the reported ADR information and medication instructions have been logically represented and formally organized in an ADR ontology. To demonstrate its usage, the source data were further bioinformatically analyzed. For example, the number of drug-ADR triples and major ADRs associated with each active ingredient were recorded. The 10 ADRs most frequently observed among quinolones were identified and categorized based on the 18 categories defined in the proposal. The occurrence frequency, severity, and ADR mitigation method explicitly stated in package inserts were also analyzed, as well as the top 5 specific populations with contraindications for quinolone drugs.Ontological representation and organization using officially approved information from drug package inserts enables the identification and bioinformatic analysis of adverse reactions caused by a specific drug with regard to predefined ADR ontology classes and semantic relations. The resulting ontology-based ADR knowledge source classifies drug-specific adverse reactions, and supports a better understanding of ADRs and safer prescription of medications. Chemicals and drugs have made a great contribution to human health care. At the same time, they are rarely free from occasional adverse drug reactions (ADRs) , which aPackage inserts  are the primary official papers which accompany most prescribed drugs and over-the-counter medications. Although different countries have diverse requirements for the obligatory contents, the package inserts serve at least two main purposes. They contain informative details regarding the generic names of drugs, active ingredients, indication for use, instructions for use, special warnings, contraindications, and statistical values from clinical trials, including the percentage of people who had side effects, the types of side effects, and additional precautions. Furthermore, the package insert is an easy reference for physicians when prescribing medications, and can help them avoid prescribing drugs that may be contraindicated. The inserts also serve as an easy reference for patients. However, the informative package inserts are generally semistructured and cannot be understood easily by machines. As the number of newly licensed drugs increases, the demand for automatic technology for semantic integration and linkages, as well as bioinformatic analysis of the information (including ADRs) enclosed in package inserts, has become an urgent issue both in biomedical research and the pharmaceutical industry.In information science, an ontology is the formal, explicit specification of a shared conceptualization of a domain . GeneralIn this paper, we propose an ontological organization of traceable ADR information extracted from licensed package inserts, which aims to provide machine-understandable knowledge for bioinformatics analysis, semantic retrieval, and intelligent clinical applications . This enIn recent decades, many studies have identified ADRs through diverse channels of information, including patient reports -14, elecSeveral studies about building ontologies for ADRs for different applications have already been carried out. For instance, the Adverse Drug Reaction Classification System (ADReCS) was developed as a comprehensive ADR ontology database, which enabled standardization and provided hierarchical classification of ADR terms for a molecular understanding of drug safety in the laboratory, and use in bioinformatics and systems biology for toxicological research . AdditioAdverse drug events (ADEs) refer to the injuries resulting from medical interventions related to drugs , which iWe built the ADR ontology using two dimensions: ADR information and drug-related medication instructions, which were further separated into nine subdimensions. The former covers the following: (1) adverse drug reactions, presenting diverse types of ADRs; (2) the occurrence of ADRs, describing the frequencies of ADRs after the administration of a drug in a population; (3) the severity of ADRs, describing a general measure of the subsequent risks of potential ADRs; (4) populations affected by specific ADRs, noting the individual human patients associated with an adverse reaction after the administration of a drug; and (5) the mitigation methods of ADRs, referring to any measure that shortens the duration of an adverse reaction or reduces its severity. The drug-related medication instructions contain generic information about rational drug use: (6) drug names, describing different pharmaceutical agents; (7) dosage forms of drugs, collecting the complete form of the pharmaceutical preparation used to administer the prescribed dose of medication; (8) administration routes of drugs, consisting of the various ways of administering a drug to a patient to allow the chemical to be absorbed into the blood and delivered to the target tissue; and (9) contraindications of drugs, predefining a condition or factor associated with a recipient that makes the use of a specific drug improper or inadvisable. These nine branches determine the fundamental concepts and classes of our ontology. The biomedical terms denoting nine classes within the ADR ontology are much more comprehensive than shown in In addition, two kinds of data properties will be incorporated into the ADR ontology, including a general description of the drugs ; the key codes and definitions from referenced vocabularies containing ARecNO (the PT code) and the SOC code from WHOART, the MedDRA code, the NCIt code; and definitions from CTCAE. Furthermore, the origin of the package inserts will be recorded in the ontology’s annotation property for ontology data identification and traceability.From the viewpoint of ontology construction, the ADR information and medication instructions extracted from package inserts will become the instances of object properties within the ADR ontology. Due to the large amount of data enclosed in package inserts, manual extraction would be a labor-intensive process. We developed a Python NLP-based algorithm to automatically retrieve ADRs and drug-related information, which consists of two steps: named entity recognition (NER) and relation extraction (RE).Briefly, NER will recognize a string of text as an entity  that is already defined in our ADR ontology. The Jieba word segmentation model implemented in Python is adopted to segment the words enclosed in package inserts, while the names of classes and their synonyms from the proposed ontology will function as a domain vocabulary to improve performance. RE is a process that determines whether two entities have a specific relationship . Since formal package inserts have already been separated into several titled sections, these titles will be used to implement the RE task. Specifically, the titles “ADVERSE REACTIONS” and “WARNINGS AND PRECAUTIONS” are converted into semantic relations about ADR information, while the medication instructions for a particular drug would be extracted from the sections titled “DOSAGE AND ADMINISTRATION,” “CONTRAINDICATIONS,” and “USE IN SPECIFIC POPULATIONS.”The information automatically retrieved from package inserts will be passed to downstream biocuration for data identification. The major criteria of the manual review process emphasize the following two points: (1) whether the medication instructions and information about ADRs caused by a specific drug was accurate and complete, without missing data or mistakes; and (2) the frequency and severity of a drug-ADR triple must be explicit; vague descriptions were not recorded. Eventually, the human biocurated semantic information is used to build the ADR ontology.The Ontology Web Language (OWL) is a widely used programing language for defining and instantiating web-based ontologies. It provides a machine-understandable schema to describe classes and their semantic relations in a specific domain. In this work, the OWL application programming interface, a Java interface and implementation for OWL, is used to build the ADR ontology automatically based on content data primarily obtained from package inserts and well-established vocabularies . Moreover, to ensure high-quality results, the ADR ontology will be evaluated and validated by checking the clarity, coherence, extendibility, minimal encoding bias, and minimal ontological commitment, which are the fundamental principles of building a domain ontology .Quinolone drugs have become commonly used antibacterial agents due to their strong and broad-spectrum antibacterial activity, as well as their rapid and complete absorption in humans. As quinolone usage increases, the risk of ADRs increases proportionally. According to the Annual Report of National ADR Monitoring in China, the number of adverse reactions and events from quinolones has continuously been the second-highest among antibacterial drugs. Therefore, the safe administration of quinolones is a serious matter that requires more attention.After half a century of development, quinolones have evolved from the first generation to the fourth generation. We collected 165 specific drug names of quinolones from the China Pharmaceutical Reference and the National Essential Drugs List in China, and further organized them based on their generation. The electronic package inserts of these drugs were then downloaded from the National Medical Products Administration and Yaozhi drug database in China. Although these package inserts were written in Chinese, the Chinese and English synonyms of the class names defined in the proposed ADR ontology will enable information extraction based on a customized Python program. Finally, an ADR ontology for quinolones was automatically built and evaluated for further analysis.Currently, the specialized ADR ontology covers a total of 2879 classes grouped into nine categories: ADRs and their occurrence, severity, population, and mitigation methods, as well as 165 drugs and their dosages, administration routes, and contraindications A. These We compared our work with four other ontologies (described in the Introduction) in terms of the semantic relations defined in the proposed ADR ontology , since tThe ontology of ADRs caused by quinolones consists of 7725 drug-ADR triples retrieved from 165 package inserts. After duplicate removal, 331 ADRs were identified to be caused by quinolones. The 331 ADRs could be further classified into 18 categories that we defined in the ontology . Most orThe ADR ontology proposed in this study has two major applications. The first is the semantic retrieval system that can use the ADR knowledge to integrate various external sources of information. Since the primary ADR terms and their key codes were reused from WHOART and MedDRA, it should be straightforward to integrate our ontology with these controlled vocabularies, as well as other medical terminologies , for semantic knowledge retrieval by providing formally represented ADR information. Another major application is intelligent clinical decision-making support. The proposed ADR ontology provides machine-understandable knowledge, which could be used by artificial intelligence algorithms in biomedicine. To pursue clinical and therapeutic approaches to optimal disease management and rational drug use, it is useful for a physician treating a specific disorder to know all the identified adverse reactions induced by the drugs prescribed for patients with that condition. Manually reading the package inserts to find the ADR information and medication instructions is laborious and time-consuming, as the number of newly approved drugs and reported ADRs increases every year. Conversely, our ontology will aid in the development of an intelligent clinical decision-making system, which would positively affect the drug prescribing patterns of physicians and potentially have a significant socioeconomic impact.We have shown that the ADR ontology can be used to formally represent and organize the ADR information and medication instructions enclosed in official drug packages. In addition, it can provide machine-understandable knowledge for bioinformatic analysis. By collecting 165 package inserts of quinolone drugs, a specialized ADR ontology was built to classify various identified ADRs caused by quinolones. Systematic analysis of the obtained ontology data improves the deep recognition of drug-specific ADRs, making it possible to intelligently guide safe drug use and benefit human health.The proposed ADR ontology can be generalized to organize the ADR information from other channels, not just package inserts. Electronic health records, in which the ADR data are frequently reported, will be acquired for the enrichment of the ADR ontology in the near future.


Emergency personnel face unpredictable and challenging incidents and their resilience and ability to cope influences their well-being. Personality traits, such as neuroticism, are postulated to be robust predictors of health and health behaviours. Despite evidence in the general population that neuroticism can positively impact health and health behaviours; to date neuroticism in emergency personnel has primarily been associated with adverse health outcomes.To assess whether neuroticism has a negative or positive impact on subjective and objective health and health behaviours in emergency personnel.This study used cross-sectional UK Biobank baseline data of emergency personnel . Logistic regression models examined the strength of the associations of neuroticism tertiles with subjective  and objective health  and self-reported smoking, sleeping, alcohol use and exercise levels.n = 2483). The association between neuroticism and chronic disease/s was significant for police in the second  and third  neuroticism tertiles. Neuroticism in firemen was associated with reduced abdominal obesity  and increased exercise .High neuroticism was positively associated with poorer subjective health outcomes in all emergency personnel (We observed positive and negative associations between neuroticism and health outcomes and behaviours. While differences were observed across the emergency personnel groups, more research is needed to better understand how personality traits may impact health in workers with physically and mentally intense jobs. Emergency personnel are constantly exposed to critical operational and organizational risk factors that can make them susceptible to poor mental health and adverse physical health issues.Personality traits, such as neuroticism, are postulated to be robust predictors of health and health behaviours and to influence workers responses to occupational risk factors.To our knowledge, to date no studies have assessed the differential impact of neuroticism on health and health behaviours across emergency personnel professional groups.This study assesses whether the personality trait of neuroticism has a negative or positive association with subjective and objective health and health behaviours in emergency personnel.Overall, high neuroticism was associated with both negative  and positive health outcomes (waist circumference) in emergency personnel and negative  and positive  health behaviours across the different emergency personnel groups.High neuroticism was associated with increased reports of poor/fair self-rated health in all emergency personnel, with the strongest associations among firemen.While differences were observed across the emergency personnel groups, more research is needed to determine whether these differences are meaningful and what can be done to improve the health, safety and work ability in demanding professions.Occupational risks and hazards are widespread and their impact on mental and physical health, well-being and occupational outcomes may be modified by individual differences in personality and genetic liability ,2. The wIndividual coping strategies and processes are developed over working lives and emergency personnel develop a number of traits that allow them to cope effectively (adaptively) with operational and organizational risk factors and attempt to reduce or manage stress within their role ; or inefStudies investigating associations between neuroticism and mortality, for instance, vary by populations and outcomes ,6,10–13.Research on neuroticism in emergency personnel, however, has mainly focussed on negative health and work outcomes ,6,10–13.It has been proposed that emergency personnel choose their careers because they have certain personality traits that differ from the average person  and thatwww.ukbiobank.ac.uk) is a UK-wide, on-going, prospective cohort study established for identifying determinants of disease in middle- and old-age adults [Baseline UK Biobank data collected from 2006 to 2010 were used for the analysis. The UK Biobank , occupational factors  and social support (ascertained from responses to the question ‘how often are you able to confide in someone close to you?’). All health behaviours  were included in each of the models assessing health outcomes.Missing data: Using data for the entire working population at baseline, two approaches were used to account for missing data. First the variable income had approximately 9% missing data, which was socially patterned. To address concerns of non-response bias, and potential systematic underestimation , we inclSample characteristics were summarized using frequencies and means. Logistic regression models for which odds ratios (ORs) and 95% confidence intervals (CIs) were reported examined the strength of association between neuroticism and (i) self-rated health, (ii) chronic health conditions and (iii) abdominal obesity. We also estimated models assessing the strength of association between neuroticism and the health behaviours: (i) smoking, (ii) engaging in exercise, (iii) sleep deprivation and (iv) alcohol use. All analyses were performed using Stata14 MP Software .Overall, our sample was predominantly male (81%), 43% had a gross household income greater or equal to £52 000, 80% lived with a partner and mean age was 48.5 years (SE = 0.12). Approximately 45% of participants were in neuroticism tertile 1, 30.65% in tertile 2 and 24% were in the highest tertile . There wThe logistic regression results examining the association between neuroticism and health outcomes are presented in Our study, for the first time to our knowledge, assesses whether the personality trait of neuroticism has a negative or positive impact on subjective and objective health and health behaviours in emergency personnel. Overall, high neuroticism was associated with both negative  and positive health outcomes (waist circumference) and negative  and positive behaviours . High neuroticism was associated with increased reports of poor/fair self-rated health in all emergency personnel, with the strongest associations among firemen. Examination of neuroticism’s impact on the objective health measure of abdominal obesity, however, showed a decreasing risk of adverse outcomes with high neuroticism in firemen, and no significant association in other personnel. Previous research has reported similar outcomes and indicated stronger relationships between neuroticism and perceptions of health status, than actual health, and therefore affects subjective health more strongly than objective health ,23. AccoWhile the role of chance in shaping these findings cannot be ruled out, our data suggest that there are differences between the emergency personnel groups. We observed a greater risk of negative health outcomes and behaviours among police and paramedics in higher neuroticism tertiles, and more positive objective health outcomes and health behaviours among firemen. The reasons for this finding are unclear, and more research would be needed to determine if it is meaningful. For instance, these differences could possibly reflect differences in workplace settings and work organization, rather than and/or in addition to personality trait differences.Emergency service personnel do physically and mentally demanding work, characterized by irregular and often dangerous incidents ,24, and Some studies have reported lower levels of stress and burnout in firemen relative to police officers, which may in part be attributed to work arrangements and organizational resources and structure ,26. WhilOur findings should be considered in the light of several limitations. Use of self-reported chronic health conditions has often been criticized as it can be influenced by disease awareness and recall bias among other factors. Previous research however has found self-reported doctor-diagnosed outcomes to be highly correlated with medical records . In addiDespite these limitations, our findings are strengthened using a large, national, cohort with relatively large sample size. Personality traits are postulated to be robust predictors of health and health behaviours and may provide more insight into susceptibility to adverse health, and the barriers or facilitators of health-promoting behaviours in different groups and occupational settings . More reThe results of this study highlight the importance of including personality in occupational and public health research. Previous research suggests a significant direct relationship between personality traits and the type of coping strategies police officers use . NeurotiWe acknowledge financial support from the Medical Research Council and Chief Scientist Office  and an MRC Strategic Award (MC_PC_13027 for E.D.).E.D. has previously received part funding to conduct a different research project on police officer well-being.


Individuals with negative attitudes towards own aging (ATOA) experience worse psychological health in later life. At the intersection of sexism and ageism, women are likely to have greater concerns about growing older and hold more negative views of aging than their men counterparts. However, the impact of gender on the relationship between ATOA and psychological health is unclear. Moving forward, the current study aims to examine 1) gender differences in longitudinal changes in ATOA and 2) whether gender moderates the association of ATOA with cognitive function and depressive symptoms. Using three waves  from the Health and Retirement Study, a total of 6,675 adults aged 50+  were analyzed. A series of multilevel growth curve analyses were performed to investigate the 8-year changes in ATOA and within- and between-person effects of ATOA on cognitive function and depressive symptoms. The models controlled for demographic, socio-economic, and physical health characteristics. Women had more negative ATOA at baseline compared to men, but not in rates of change. When levels of ATOA were more negative, both cognitive performance and depressive symptoms were poorer over time between individuals as well as within-person. We found that the detrimental effects of negative ATOA on depressive symptoms were stronger for women, but there were no significant gender differences in relation to cognitive functioning. Our findings demonstrated that women view aging more unfavorably than men, and the effects of endorsing negative ATOA are more pronounced on women’s mental health.


Opioid use by pregnant women represents a major public health concern given the impact of opioid exposure on maternal and neonatal outcomes, including increased risk of maternal mortality, preterm labor, stillbirth, and neonatal opioid withdrawal syndrome (NOWS) . UnfortuCurrently, the most common first-line medication used to treat NOWS is morphine. A minority of infants are treated with methadone, and a very small percentage receive buprenorphine . DespiteToward the Use of Buprenorphine in Infants: Scientific and Practical Considerations [SM (HEAL) Initiative, a trans-agency effort to speed scientific solutions to stem the national opioid public health crisis [To move the field forward, the National Institutes of Health (NIH) hosted a two-day virtual workshop in August 2020 entitled “erations .” The woh crisis . Table 1The first session focused on the state of the science, informed by the Advancing Clinical Trials in Neonatal Opioid Withdrawal Syndrome (ACT NOW) Current Experience study. This study of 1808 infants with NOWS, conducted at 30 U.S. sites, found considerable variability in what drugs infants were exposed to—illicit and prescription opioids, maternal medication for addiction treatment, and polysubstance exposures —and how mothers and infants are clinically managed . FurtherEunice Kennedy Shriver National Institute of Child Health and Human Development and the NIH Environmental influences in Child Health Outcomes Program, is currently conducting three large studies evaluating: (i) how rapidly it is safe to wean infants with NOWS off of morphine or methadone; (ii) whether the Eat, Sleep, Console approach can shorten hospital stay more than using the Finnegan Neonatal Abstinence Scoring Tool (FNAST or modified FNAST); and (iii) what changes happen in brain development and associated neurodevelopment and behavioral outcomes in infants with NOWS. All three studies will be assessing neurodevelopment at 24 months of age. Moreover, the National Institute on Drug Abuse has embarked on the HEAL-supported HEALthy Brain and Child Development Study (HBCD). Although still in the planning phase, the HBCD Study will establish a large cohort of pregnant women from regions of the country significantly affected by the opioid crisis and follow them and their children for at least 10 years.The second session focused on ongoing and upcoming clinical trials. Specifically, the ACT NOW Program, a HEAL-supported collaboration between the The third session focused on feasibility and operational issues for a potential trial including buprenorphine for the treatment of NOWS. Numerous challenges exist. Identifying the most salient research question is challenging, given the variation in experiences for mothers with OUD and infants with NOWS, including what medication-assisted treatment (MAT) the mother received, when and how long she was treated during pregnancy, whether she received comprehensive treatment versus medication only, and if she used multiple substances during her pregnancy. Testable interventions might compare the effectiveness of the primary medications used among all infants with NOWS or compare the effectiveness of matching the infant’s medication with the mother’s MAT medication. Finding sufficient study sites for future trials may also be difficult, as many potential sites may not have equipoise for all study questions. Furthermore, many sites do not have the capacity to conduct these studies, as they do not have research pharmacy access to controlled-substance study drugs 24 hours a day, 7 days a week.The last session focused on next steps. Because current morphine, methadone, and buprenorphine formulations have much different routes of administration , dose intervals, and weaning protocols, reliably masking study drugs is difficult. Selecting clinically meaningful neurodevelopmental and non-neurodevelopmental outcomes that allow feasible trial length and cost poses a formidable challenge as well. Because this study population may be in unstable living situations and infants are often placed in foster care, collecting long-term outcomes requires research staff to develop relationships with families, guardians, and stakeholders  to maintain contact and establish trust. Finally, several statistical considerations include selecting pragmatic, validated assessment instruments sensitive enough to quantify treatment effects and taking into account the myriad confounding factors .In conclusion, considering the gaps in knowledge in whether and when to use buprenorphine to treat NOWS and the potential for better outcomes, a comparative-effectiveness, randomized controlled trial was recommended by the experts at the workshop to inform clinical practice. Numerous issues were identified during the workshop that need to be considered to design and implement this much needed study successfully.The contents of this article represent the authors’ views and do not constitute an official position of the National Institutes of Health or the U.S. Government.


Urbanization and concomitant challenges pose a great threat to sustainable development. Urban and rural development interacts through the flows of people, materials, energy, goods, capital, and information. Without building sound urban–rural linkages, achieving development in one area could compromise it in another area. Achieving sustainable development needs customized policy prioritization and implementation in both urban and rural areas. Much literature exists in the research field of urban–rural linkages, but little has been done via a comprehensive analysis from an interlinkage perspective in the context of the Sustainable Development Goals (SDGs). Sustainable Development Goal 11 on sustainable cities and several targets embedded under other Goals provides a good framework for analyzing the urban–rural linkages. This paper contributes to this novel research perspective using Ghana as a case. The study applied an integrated approach by combining the results from a solution-scanning exercise with an SDG interlinkage analysis to identify the challenges and priority solutions and assess the synergies and trade-offs of the identified solutions. It extends the conventional solution-scanning approach by further assessing the synergies and trade-offs of the solutions from an SDG interlinkage perspective. It also enables a more practical SDG interlinkage analysis through the contributions from the multi-stakeholder consultations conducted in Ghana. The analyses show that prioritizing gender inclusion  will positively affect many social and well-being outcomes, including poverty elimination , hunger reduction , health improvement  and access to quality education  and basic services, such as water . However, gender inclusion could have potential trade-offs in the agricultural sector  in the case that women who dominate agricultural value chains could move to work in other sectors. Lack of proper infrastructure , such as transport, will hinder wide gender inclusion. An integrated approach that considers both the synergies and trade-offs of relevant solutions is critical for effective policymaking, specifically in developing countries. Urban–rural linkages are defined as the cultural, economic, environmental, social, and political connections between communities on the urban and rural divide Tacoli . Braun  and TacoThe concept of urban–rural linkages is significant to poverty alleviation work and the efforts for achieving wider equality made by many development organizations, such as the Organization for Economic Cooperation and Development Tacoli , the Worhttps://1d1f.gov.gh/) is another major strategy aiming to stimulate economic development and rural development across the country. Also important is the National Urban Policy (NUP) Framework and Action Plan, which seeks to promote effective urban–rural linkages by establishing rural service centers and promoting agriculture activities and the development of agro-based industries , 4 , 5 (gender inclusiveness), 6 (water and sanitation), 7 (energy), 8 (economic growth and decent work), 9 , 10 , 12 (sustainable production and lifestyles), 13 (climate actions), and 15 . Serving as an integrated framework for achieving the 2030 Agenda for sustainable development and covering a broad scope of the three dimensions , SDGs provides an appropriate framework for analyzing the urban–rural linkages.SDGs, through the intrinsic relationships among the goals and between the targets, forms an indivisible system. An SDG interlinkage analysis helps in understanding the relationships among the SDGs and thus informing policymaking on the potential effects of achieving one area of development on other areas. When the identified top solutions are mapped with corresponding SDG targets, an SDG interlinkage analysis can be used to analyze the interlinkages of the top solutions with other relevant SDG targets. The SDG interlinkage analysis can identify the synergies (regarding positive relationships) and trade-offs (regarding negative relationships) among the top solutions and between the top solutions and other SDG targets. This will enable customized solutions by maximizing the synergies and minimizing the trade-offs in policymaking. In this study, we applied the SDG Interlinkages Tool (V3.0)  It is comprehensive in covering all 17 goals and 169 targets at the national level for 27 countries in Asia and Africa, including Ghana. (2) It is transparent with all data available online, including the identification of the interlinkages, indicators and their time-series data, and the dashboards on the synergies and trade-offs. (3) The tool is convenient through the online visualization interface which enables to select a country, goals, and relevant targets. (4) It is flexible in modeling newly defined relationships among the 169 targets and additional elements and associated relationships beyond the 169 targets.Step I: the identification of the interlinkages between relevant SDG targets is based on causation, mainly through a comprehensive literature review together with expert opinions and a certain level of stakeholder consultations regarding specific issues.Step II: indicators with trackable data are selected for corresponding SDG targets based on the Global SDG Indicators UNSD  and otheStep III: time-series data (1990–2019) are collected for the indicators in 27 countries in Asia and Africa (including Ghana) and refined for data analysis.Step IV: the identified SDG interlinkages (Step I) are quantified to build the SDG interlinkage model using statistical methods based on the indicator-level time-series data collected for 27 countries (Step III).The SDG Interlinkages Tool was built on the following methods for the identification and quantification of the interlinkages between relevant targets at the national level  show strong linear relationships, and those with a smaller absolute value  show weak linear relationships. In this study, we used positive linkages to indicate potential synergies and negative linkages to indicate potential trade-offs.Based on the mapping of the top four solutions identified by Somanje et al.  with corFor the SDG Interlinkages Tool, a set of indicators of the SDG targets with trackable data was selected based on the Global SDG Indicators UNSD  and otheUsing the SDG Interlinkages Tool for Ghana, the synergies and trade-offs of the top solutions for building effective urban–rural linkages were analyzed, and the results are presented as follows.The SDG interlinkage analysis reaffirms the findings in Somanje et al.  that genAchieving gender inclusiveness also relies on other influencing factors. Figure Meanwhile, certain measures could negatively affect gender-related targets Fig. . For insSignificant urban–rural disparities are observed in basic services, such as access to food, water, energy, housing, and economic infrastructure. Nine basic-services-related SDG targets were identified for improving urban–rural linkages in Ghana Table . Seven oBasic services also depend on the progression of other SDGs, which is captured in the inward linkages Fig. . For exaHowever, several SDG targets could negatively affect the promotion of basic services and infrastructure through negative inward linkages. For instance, inadequate and inefficient transport systems (11.2), degradation in relevant ecological systems, and unsustainable resource use (12.2) in Ghana are causing a scarcity of resources and fiercer competition among the sectors dependent on these resources. This, in turn, generates a dragging effect on the expansion of basic services and related infrastructure. Addressing these lagged areas could remove the drags and release the potential for making further progress in the provision of basic services and infrastructure in Ghana.Agriculture is a critical export sector in Ghana, with cocoa accounting for 25% of its total exports and approximately 20% of global cocoa exports  to maximize synergies and minimize potential trade-offs were identified for policy formulation.An SDG interlinkage analysis, either qualitatively or quantitatively, could provide a comprehensive overview of the interactions among the key elements of the human–environment system and, therefore, it supports a systems approach to problem-solving. It is a powerful tool to help in addressing development issues, such as sound urban–rural development, which cut across multiple spheres of the social, economic, and environmental dimensions. An SDG interlinkage analysis can identify and analyze the potential synergies and trade-offs that help in informing effective priority setting and resource allocation. For example, in building effective urban–rural linkages in Ghana, enhancing gender inclusiveness and improving basic services and infrastructure could be considered together rather than siloed issues because of the many synergies among the improvement in the two areas. By addressing the two issues together, limited financial and human resources can be used more effectively than addressing the two issues separately.Furthermore, understanding the trade-offs of the proposed solutions is critical to avoid lock-in effects in unsustainable pathways and ensure policy integrity and coherence. For example, promoting agriculture productivity improvement to strengthen the urban–rural linkages in Ghana should consider its trade-offs on environmental pollution and associated damages to public health, in both rural and urban areas, and biodiversity loss. Proper pollution prevention and control measures and sustainable agriculture practices are needed through collaboration between urban and rural areas.For future research in the area of SDG interlinkage analysis, multi-stakeholder consultations and engagement should be strengthened in the processes of problem identification, solution screening, identification of the SDG interlinkages, enhancement of indicators, data availability, and calibration of the results and acting on the results. In this study, multi-stakeholder contributions are limited to the screening and ranking of top solutions and SDG mapping in the context of Ghana.The availability of effective indicators and trackable data presents a major challenge to current and future research of an SDG interlinkage analysis, specifically quantitative analysis. For example, under the existing SDG Interlinkages Tool, which was used for the interlinkage analysis of the urban–rural relationships in this paper, among the 169 targets, only 83 have corresponding indicators with trackable data for Ghana. Showed as dotted lines in the visualization results, many linkages cannot be quantified. The big gaps in effective indicators and data availability have constrained conducting a full-fledged quantitative assessment. Improved and localized indicators and data availability can improve the robustness of the quantitative analysis.This study used an integrated approach by combining the results from a solution-scanning exercise with an SDG interlinkages analysis to assess the synergies and trade-offs of the identified top solutions for achieving effective urban–rural linkages in Ghana. Ghana needs to promote effective urban–rural linkages if the country is to achieve the SDGs. Ignoring such linkages will severely inhibit growth and widen spatial inequalities (World Bank


The Global Fund to Fight AIDS, Tuberculosis and Malaria was founded in 2002 as a public-private partnership between governments, the private sector, civil society, and populations affected by the three diseases. A key principle of the Global Fund is country ownership in accessing funding through “engagement of in-country stakeholders, including key and vulnerable populations, communities, and civil society.” Research documenting whether diverse stakeholders are actually engaged and on how stakeholder engagement affects processes and outcomes of grant applications is limited.To examine representation during the 2017 Global Fund application process in the Democratic Republic of the Congo (DRC) and Uganda and the benefits and drawbacks of partnership to the process.We developed a mixed-methods social network survey to measure network structure and assess perceptions of how working together in partnership with other individuals/organizations affected perceived effectiveness, efficiency, and country ownership of the application process. Surveys were administered from December 2017–May 2018, initially to a set of central actors, followed by any individuals named during the surveys (up to 10) as collaborators. Network analyses were conducted using R.Collaborators spanning many organizations and expertise areas contributed to the 2017 applications . Participation from NGOs and civil society representatives was relatively strong, with most of their ties being to different organization types, Uganda (63%), and DRC (67%), highlighting their collaborative efforts across the network. Overall, the perceived benefits of partnership were high, including very strong ratings for effectiveness in both countries. Perceived drawbacks of partnership were minimal; however, less than half of respondents thought partnership helped reduce transaction costs or financial costs, suggesting an inclusive and participatory process may come with short-term efficiency tradeoffs.Social network analysis can be useful for identifying who is included and excluded from the process, which can support efforts to ensure stronger, more meaningful engagement in future Global Fund application processes. The Global Fund to Fight AIDS, Tuberculosis and Malaria was founded in 2002 as a public-private partnership between governments, the private sector, civil society, and populations affected by the three diseases. As a financing mechanism, the Global Fund raises and disburses funds to principal recipients  and is among the largest global health initiatives, disbursing nearly US$4 billion per year in over 100 countries toward its mission of accelerating the end of the three epidemics [Four principles guide the work of the Global Fund: country ownership, partnership, performance-based financing, and transparency . The Glo3789As described in a recent review, there are various technical support partnership models through which the Global Fund operates ; here weThe process for applying for Global Fund funding has evolved substantially throughout the last decade. In 2013, the New Funding Model introduced flexible application windows, simplification through alignment with national strategic plans, predictability of funding for a three-year cycle, and earlier and enhanced Secretariat engagement and support .meaningfully engaged, and how this adds value to the efficiency and effectiveness of decision making during the funding request and grant making process [Country ownership and partnership are critical for promoting harmonization and impactful design of Global Fund investments, but there has been limited evaluation documenting the complexity of stakeholder inclusion, whether and how diverse stakeholders are  process 15. The oBoth the DRC and Uganda are ranked among the bottom 16% of countries on the Human Development Index . In addiThis study is embedded in the Global Fund Prospective Country Evaluation (PCE). The Global Fund’s Technical Evaluation Reference Group (TERG) commissioned the PCE in 2017 to support the independent assessment of implementation and impact of the Global Fund Strategy 2017–2022 2. The PCEThrough the PCE platform, this comparative, mixed-methods social network analysis draws from data collected on two cases: the Global Fund application process in the DRC and Uganda. In 2017, the Global Fund introduced a new application approach to enable a more streamlined and efficient process, allowing countries to opt for less burdensome grant continuation processes, as well as the existing full application  . The DRCGlobal Fund application processes are important cases for case study research and policy analysis because of the magnitude of resources tied to them and their instrumentality as key decision-making processes in low-income countries. Per previous partnership analyses, a “case” can be defined as a process with a specific resulting outcome, a feature that also enables stronger cross-country comparison 14. We coSocial network surveys were administered between December 2017 and May 2018 in Uganda and the DRC by a mixed-methods evaluation team at the Infectious Diseases Research Collaboration (Uganda) and PATH (DRC), the respective PCE country evaluation partners. We defined network members as any actors involved in the Global Fund grant application process. An initial list  of potential survey respondents was developed through ongoing document review and meeting observation, which served to identify a subset of individuals likely to be central to the application network, for example including stakeholders from the CCM, national health programs and other government agencies, technical partners, bilateral funders, and civil society. These stakeholders were contacted for an interview about the funding request process. Following the interview, a network survey was administered in person via paper or via an online link to an electronic survey programmed in SurveyGizmo’s data collection platform . In UganWe adapted a structured social network survey from a similar survey conducted by Kamya and colleagues, which drew from an existing conceptual framework of how development partnerships add value to decision-making processes through efficiency, effectiveness, and country ownership . The teaDuring the survey, the study team first asked respondents to identify which Global Fund applications they worked on , and specifically which aspects of the application they supported. Respondents were then asked to provide the names and organizations of up to 10 individuals they personally collaborated with on the Global Fund grant application, which funding request(s) they collaborated on, and to rate their level of professional trust (a strong predictor of network performance ) with thWe used existing mathematical algorithms to measure common network metrics, including nodes, ties, density, degree centralization, and betweenness centrality  due to the nature of collaboration within partnerships, which are characterized by mutual decision making and reciprocal accountability, distinct from contractual relationships characterized by hierarchical decision making and unilateral accountability . Within Networks were visualized according to several subgroup attributes available for all nodes, including funding request type, organizational affiliation, gender, and national versus sub-national representation (DRC only). All analyses were conducted using the statnet suite of network analysis packages in the R statistical programming language and the associated statnetWeb R Shiny application 34.Through the overall Prospective Country Evaluation, evidence from document review, meeting observation, and interviews on the funding request process  were routinely entered into an analysis matrix, an Excel-based framework for organizing data by thematic area and stakeholder group . To supp36The DRC protocol was approved by the University of Kinshasa School of Public Health Ethics Committee (#ESP/CE/074/2017), and the Uganda protocol was approved by the Makerere School of Medicine Research Ethics Committee (#REC REF 2017-146) and the Uganda National Council for Science and Technology (#SS 4472). This was determined to be non-human subjects research by the University of Washington’s IRB (#STUDY00003643). The purpose of the survey was explained, including confidentiality measures, and participants were asked for their consent to participate.In both the DRC and Uganda, the 2017 funding requests were submitted during the first application window (March 2017) and approved or validated by the Technical Review Panel to move forward to the grant making stage. There was broad consensus in both countries that the 2017 application process was a success in that it led to grant signature within the planned time frame, allowing countries to avoid a funding gap by remaining on track for initial grant disbursement in early 2018. Stakeholders attributed the faster grant processing to application process changes introduced by the Global Fund, better levels of preparation by country stakeholders, more coordination, and increased support and engagement from the Global Fund Country Teams from the beginning of the application cycle 37.The 2017 Global Fund application required a large network of actors spanning many different types of organizations and expertise. The characteristics of identified actors in the DRC and Uganda application networks are compared in Table The DRC’s overall network structure is characterized by a larger, central core and several peripheral collaboration hubs. The peripheral hubs tend to have more representation from provincial-level stakeholders Figure , while tGlobal Fund application network structures were examined by funding request type, with nodes color coded for TB/HIV (blue), malaria (red), or both funding requests (grey) Figure . In UganWhen the full networks are plotted by gender Figure , the strIn both the DRC and Uganda, roughly 70% of the network was composed of individuals representing civil society, technical partners, or Principal Recipients  Table  and therNGOs and civil society (including civil society PRs and SRs) were well represented in the application networks of both the DRC (47%) and Uganda 22%)  and Ministry of Education and Sport (light yellow), is also notable in Uganda given the increased emphasis on HIV programming for adolescent girls and young women ; however, participation was limited (n = 3), peripheral, and did not include representatives with decision-making authority.Survey respondents were asked about perceived benefits and drawbacks of partnership that occurred due to working together with other individuals and organizations in supporting the 2017 Global Fund application process. The percent agreement with each of the potential benefit and drawback statements, organized according to effectiveness, efficiency, and country ownership domains, is shown in Table Across four items, partnership effectiveness benefits were rated higher by respondents in Uganda (93%) than the DRC (79%) on average. In Uganda, 100% of respondents agreed that partnership had improved the quality and technical soundness of the approved Global Fund grants; whereas, in the DRC most respondents (79%) agreed with the statement. This triangulates with evidence that Uganda’s grant applications received minimal comments from the Technical Review Panel and that grant applications were reviewed and approved on time in both countries. Most respondents in the DRC (83%) and Uganda (85%) were also better able to identify the need for, and to acquire, additional technical support. Nearly one third of respondents in the DRC (31%) and Uganda (30%) reported a drawback of partnership was that it created competition and conflict among member organizations.There was general agreement that partnership supported the efficiency of the grant application process in terms of timely execution of planned activities in Uganda (93%), but this benefit was much less frequently perceived to have occurred in the DRC (58%). Most respondents in Uganda agreed that partnership helped to leverage an organization’s competitive advantages (85%) during the grant application, but fewer than half of the DRC respondents (44%) agreed. However, fewer than half responded that partnership helped to reduce transaction costs  or helped reduce the financial cost of the process . Overall, the perceived drawbacks of the partnership on efficiency were minimal: some respondents in the DRC (19%) and Uganda (29%) thought it resulted in making decisions in an unnatural or atypical way, and few respondents reported strained relations, loss of control/decision-making autonomy, or lack of credit as drawbacks that occurred.While nearly all respondents in Uganda (93%) perceived partnership to have contributed to approved grants that were more responsive to country needs, only 42% of respondents in the DRC responded similarly. Over three quarters of respondents in both countries perceived partnership to have resulted in increased inclusiveness of stakeholders, increased accountability and transparency among partners, and increased fairness and legitimacy of decisions.Social network analysis has increasingly been applied to understanding complex health systems and decision-making structures, such as stakeholder roles and relationship composition within networks, including who is included or excluded, which organizations/actors are most influential, and how relationships form network structures that can influence processes and outcomes 38394041.383940The Global Fund grant application process is a resource-intensive undertaking, requiring about nine months of engagement from a variety of stakeholders. Our analyses indicate well over one hundred stakeholders were involved in the development of the funding requests in the DRC and Uganda, representing a diverse set of organizations and suggesting the process was highly inclusive . That tmeaningful ways. Interview data in Uganda highlighted strong participation of key and vulnerable populations and gender and human rights constituencies in the 2017 application phase compared to the previous funding cycle [Given the Global Fund’s strategic objective to “promote and protect human rights and gender equality ,” it is ng cycle . Similarng cycle . Given twho is involved in the application development, including for priority setting and decision making. Our data indicate only 25% of the application network in the DRC was female; whereas, stronger balance  was evident in Uganda’s network—a finding which may reflect prevailing norms in government leadership, with the share of parliament seats held by women higher in Uganda (34.8%) relative to the DRC (8.2%) [Gendered power relations within health policy making processes can affect participation within governance structures, including who is able to engage, their terms of engagement and how accountability is negotiated between actors . A checkC (8.2%) . ThroughThe Global Fund’s ability to reform is recognized as a unique strength of the organization , and rec36Considering the potential downsides of highly participatory processes is also important—despite being difficult to quantify, these issues are felt in terms of the duration of the process and associated opportunity costs. Our evidence suggests alternative models for “right-sizing” participation and inclusion to ensure meaningful engagement is balanced with efficiency is worth further consideration by the Global Fund. Similarly, recent research on the 2017 funding request process in Malawi, Tanzania, and Zimbabwe questions the relevance and effectiveness of Global Fund’s processes for multisectoral inclusion and deliberative engagement and whether a lengthy country dialogue process is necessary to determine spending priorities. In the context of increasingly commoditized grants where much of the allocation is predetermined , such processes may be functioning “as processes for processes’ sake alone .” More rGlobal Fund’s mandate is to end the three diseases, but increasingly, investments will need to support the Universal Health Coverage  and SustThe study was guided by a theoretical framework on partnership, adapted from an evaluation of a large global health initiative 9. UtilizThe survey was undertaken shortly following the grant signing in each country to limit participant recall bias. Potential differences in respondent bias between countries may help explain some of the differences in responses to questions about perceived effectiveness, efficiency, and country ownership of the process. Based on our observation of the funding request process and review of meeting attendance records, we are fairly confident that the survey captured the majority of network members; however, the low survey response rate is a limitation, which reduces our confidence in the true structure of the network given that a higher response rate would yield a more accurate reflection of the network’s density. We assumed all collaborative ties to be undirected ; however, this assumption limited our ability to examine reciprocity, which is a critical consideration in understanding the meaningful engagement and inclusion of civil society representatives in contributing to the application process. A deeper exploration of the nature of participation of key affected populations and civil society representatives within the Global Fund application network, including an examination of power dynamics, is warranted through future research examining the types of collaborative ties and their directionality.Our results indicate the 2017 Global Fund application processes in the DRC and Uganda were inclusive in terms of representation and participation and that respondents perceived the application process to be country-owned and effective; however, data also indicate perceptions of lower efficiency and higher financial costs. As part of process improvement, the Global Fund should consider examining the tradeoffs of a highly inclusive application development process, including striking the right balance between achieving inclusive and meaningful participation, responsiveness to country needs, and improved efficiency.The additional files for this article can be found as follows:10.5334/aogh.2961.s1Supplemental File 1.Description of Global Fund’s differentiated funding request review types.10.5334/aogh.2961.s2Supplemental File 2.Survey tools: Uganda (English) and DRC (French).10.5334/aogh.2961.s3Supplemental File 3.Additional network figures.


Human pluripotent stem cells (PSCs), including embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), have remarkable developmental plasticity and possess the potential for a wide variety of applications. Three-dimensional organs, termed organoids and produced in vitro by PSCs, contain not only epithelium but also mesenchymal tissue and partially recapitulate intestinal functions. Such intestinal organoids have begun to be applied in disease models and drug development and have contributed to a detailed analysis of molecular interactions and findings in the synergistic development of biomedicine for human digestive organs. In this review, we describe gastrointestinal organoid technology derived from PSCs and consider its potential applications.The gastrointestinal system is one of the most complex organ systems in the human body, and consists of numerous cell types originating from three germ layers. To understand intestinal development and homeostasis and elucidate the pathogenesis of intestinal disorders, including unidentified diseases, several  This paper outlines the characteristics of gastrointestinal tract, pluripotent stem cells (PSCs) and intestinal organoids derived from PSCs and demonstrates how intestinal organoids contribute to understanding of the processes of morphogenesis and diseases.To understand intestinal development, homeostasis and reproduction, and elucidate the pathogenesis of intestinal disorders, including infection, malabsorptive diarrhea, inflammatory bowel disease, digestive system neoplasia, and congenital disease, many studies have been conducted to generate tissues of the gastrointestinal tract, one of the complex organ systems in the human body. Researchers recently have made efforts to manufacture three-dimensional (3D) organs, called (1). The morphogenesis of villi and crypts occurs, and the small intestine of a fetus subsequently becomes absorptive by taking in amniotic fluid by the 13th week of pregnancy (2). The length of the intestine increases along with the growth in body length, and a rapid rate of prenatal small intestinal growth ensures an adequate small intestine to satisfy postnatal nutritional requirements for the mature newborn (3).For humans, the primitive gut tube, which has the capacity to differentiate into the mature gastrointestinal tract, appears as a straight tube during the fourth week of pregnancy and is divided into three segments  along the anterior to posterior axis. Accompanied by the development of the fetus, repeated gut rotation and fixation (dynamic movements) and genetic signaling from endoderm and mesoderm tissues help to establish the esophagus, stomach, the small intestine , the large intestine  and rectum (4). PPs and isolated lymphoid follicles form a specialized immune system of the gut, harboring approximately 70% of peripheral immunocytes and comprising the largest lymphoid tissue in the body (5). In submucosa and muscular layers, two major neural plexuses are present . The enteric nervous system (ENS) is found embedded along the full length of the intestinal tract to form meshes. The ENS orchestrates an independent neural network and regulates many functions, such as peristaltic movement, blood flow, secretion, absorption and more (6).Histologically, the gastrointestinal wall consists of three layers from the lumen outwards: mucosa, muscular layer, and serosa. The mucosa is made up of four layers: an epithelial layer of mucosa, lamina propria, lamina muscularis mucosae and submucosa. The muscular layer underlying the submucosa plays a role in peristaltic movements caused by smooth muscle  and enables foodstuffs to be propelled along the gastrointestinal tract to be assimilated as nutrients. In the lamina propria, lymphoid follicles constitute gut-associated lymphoid tissue (GALT) and aggregate to form Peyer’s patches (PPs) 2 that was thought hitherto to measure 260–300 m2, almost equivalent to the area of a tennis court (7). Thus, the wide absorptive area of the small intestine enables efficient nutrient absorption. The lumen of the duodenum is characterized by long and well-developed villi, which become gradually shorter and flatter in the jejunum and ileum and are not found in colon. The small intestine is responsible for not only nutrient absorption but also metabolism; it also functions as an endocrine gland and in immunoregulation. Moreover, a physical barrier formed by intestinal epithelial cells separates the injurious luminal environment from the underlying intestinal layers (8). The mucosal epithelium of the small intestine is one of the tissues with the highest proliferation and regeneration potential in our body. In the mouse, the small intestinal epithelium entirely turns over every 5 days, maintaining dynamic homeostasis and structural integrity.The epithelial surface of the small intestine forms circular folds, where mucosa protrudes into the lumen with submucosal tissue to increase the absorption efficiency of nutrients. Moreover, circular folds consist of numerous microvilli, hair-like structures that amplify the epithelial surface of the intestinal tract interior. This interior is covered with a single mucosal layer and an epithelial surface area of 30 mIt is necessary to elucidate pathophysiology of diseases relating to intestinal disorders by demonstrating differentiation hierarchy in humans, and it is required to construct a species-specific model that mimics the process of gastrointestinal development and the biological tissue structure in an in vitro culture system. Recent advances in organoid technology are expected to deepen understanding of complex intestinal functions and tissue homeostasis and contribute to research on disease mechanisms.(9). Secretory lineages include goblet cells that produce mucus, endocrine cells that secrete hormones, Paneth cells that produce antimicrobial peptides and configure the stem cell niche, tuft cells that have a defensive effect against parasitic infection, and microfold (M) cells that take up intestinal microbial antigens and hand them over to GALT in the mucosal epithelium. The dynamic and continuous regeneration of mucosal epithelium was noted in the 19th century; moreover, a theory that crypt base columnar (CBC) cells between Paneth cells promoted differentiation of mucosal epithelial cells in crypt base compartments, called the unitarian hypothesis, was proposed (11). By contrast, Potten and colleagues advocated that the stem cells retaining DNA labels for a long period located at position +4 relative to the crypt bottom, directly above the Paneth cells (12). After that, a great deal of effort has been devoted to studying and verifying “true” intestinal stem cells. Detailed studies analyzing molecular mechanisms in colorectal cancer have revealed that Wnt signaling and β-catenin/T-cell factor activity are crucial for regeneration and maintaining homeostasis of the mucosal epithelium. Such studies predicted that the expression of leucine-rich repeat-containing G-protein-coupled receptor 5 (Lgr5) was restricted by the Wnt target gene within crypts and was a cell marker expressed in CBC cells (14). Lgr5-positive CBC cells were demonstrated to be the stem cell of the small intestine and colon and have the ability to self-replicate and the multipotency to generate all lineages (15). This discovery had a great impact on our understanding of the stem cell niche at the molecular level. This niche includes the Wnt agonist R-spondin (16), leucine-rich repeats and immunoglobulin-like domain 1 (Lrig1) as a negative feedback regulator of the epidermal growth factor (EGF) receptor family (17), notch signaling essential to the differentiation of enterocytes (18), and bone morphogenetic protein (BMP) signaling involved in crypt–villus formation (19). At the crypt bottom can be found Lgr5-positive CBC cells and adhering Paneth cells, differentiated by Wnt signaling, that secrete Wnt, EGF and notch essential for the regulation of dedifferentiation and differentiation. These signals are involved in the development of the small intestine’s mucosal epithelium. Furthermore, cell-lineage tracking experiments have revealed a cell differentiation hierarchy and that cellular plasticity exists because of the response of the stem cell niche. A dynamic cell differentiation hierarchy is of extreme importance for an appreciation of the complexity of intestinal disorders in humans. Therefore, it is necessary to construct in vitro models recapitulating the living body and developing processes of the digestive tract.The mucosal epithelium of the small intestine consists of villi and crypts, excluding regions adjacent to PPs. In a unit of a villus and a crypt, cells are brought into existence in the crypt component and become subdivided into various types of cells that subsequently migrate to the top of the villus, as on a belt conveyer. In mucosal epithelium, six major differentiated cell lineages exist that can play a variety of functions. Enterocytes that account for 90% of the epithelium perform functions that allow the absorption of nutrients from digested food and protect the integrity of the epithelium through structural properties and antigen uptake (21). Human ESCs and iPSCs have remarkable developmental plasticity and thus possess a great potential for human organogenesis, disease modeling and drug screening. They also have potential applications in regenerative medicine as a source of cell-based therapy (22). Work on PSCs focused on isolating particular cell types from differentiation at a two-dimensional (2D) level. However, in the human body, organ systems function through the integration of each system. Thus, the recapitulation of organogenesis is important for not only understanding fundamental mechanisms of human development but also translational applications in replacement therapy and in disease modeling. Tissue homeostasis is strictly organized under a tissue stem cell hierarchy. One of the well-characterized tissue stem cell models is intestinal stem cells. Active cycling of intestinal LGR5+ stem cells balances the maintenance of stem cells and the regeneration of mucosal epithelium cells to maintain epithelial homeostasis (23). Notable pioneering works by Sasai’s group have allowed the development of 3D culture methods using PSCs to recapitulate tissue organization in a dish (26). The reconstitution of a 3D tissue structure from stem cells has led to the use of the term “organoid.” This refers to 3D structures grown from stem cells that consist of organ-specific cell types that self-organize through cell sorting and spatially restricted lineage commitment (27).In contrast to tissue stem cells that differentiate into a plurality of kinds of cell groups configuring a particular tissue, PSCs have the ability of unlimited proliferation from all three germ layers. Human PSCs, such as embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), have been demonstrated to proliferate indefinitely and differentiate into cells of any of the three germ layers in vivo(28). The initiation of investigations on intestinal organoids were specifically based on the lineage-specific embryological tracking experiments of mice models in Clevers’ laboratory (29). They succeeded in establishing a 3D structure  in vitro that consisted of multifarious types of small intestinal mucosal epithelium differentiated from self-organizing Lgr5-positive CBC cells to reproduce crypt–villus formation (16). Further studies of mucosal epithelial organoids have progressed and enabled the establishment of organoid culture conditions for epithelial stem cells from different species, including humans and mice, and different gastrointestinal sections, such as the stomach, the small intestine and the colon (32).With regard to gastrointestinal organoids, these have two different cell sources: tissue stem cells (LGR5+ stem cells) and PSCs . Sato’s (33). Intestinal organoids exhibit the following features: 1) They are comprised of a variety of cell types that constitute a tissue structure resembling an intestine in vivo, and 2) they indicate a formative process whereby various cells differentiate from stem cells to accumulate and become shaped into a 3D structure on the basis of the self-organization of dividing cells into appropriate sites containing a niche of stem cells and an extracellular matrix. In generating intestinal organoids derived from ESCs and iPSCs, two major differences exist with the mucosal epithelial organoids mentioned above: (i) Undifferentiated PSCs can recapitulate the human intestinal developmental process, and (ii) the differentiation of not only mucosal epithelial tissue but also submucosal tissue becomes possible in theory. However, it is not easy to recapitulate the gastrointestinal tract, including submucosal tissue, because the developmental process from the archenteron is quite complex. In addition, tridermic cells and tissues are interrelated and provide gastrointestinal functionality; therefore, methods mimicking the gradual differentiation of the early embryo were employed for inducing differentiation (34).Subsequently, another method of directly differentiating human ESCs and iPSCs into human intestinal organoids (HIOs) was invented by Spence et al. nodal signaling pathway that controls the expression of crucial transcription factors (35). Definitive endoderm presents as a 2D sheet of cells and subsequently forms a 3D primitive gut tube functionally subdivided along the anterior–posterior axis into a foregut, midgut, and hindgut from which endodermal organs arise. In these processes, smooth muscles and the vascular system are derived from the mesoderm, and the ENS derived from the neural crest penetrates along the gut tube. CDX2 is an obligate posterior marker that allows differentiation into midgut and hindgut, giving rise to the small and large intestines. Therefore, Spence et al. used activin to direct the differentiation of human PSCs into definitive endoderm to induce posterior endoderm patterning and mid- and hindgut specification highly expressing CDX2 through activation of both Wnt and fibroblast growth factor signaling, allowing cells to take up a caudal/hindgut fate and form small spheroids. In addition to exposure to WNT3A/FGF4 as hindgut-specific growth factors, collecting and embedding hindgut patterning spheroids into Matrigel droplets, and passaging and maintaining them in tissue-specific growth factors  enabled the spheroids to differentiate into HIOs (36).Intestinal development begins with the formation of the embryonic endoderm germ layer that gives rise to the intestinal epithelium during gastrulation. This requires the (33). Since deep crypts and the expression of the intestinal stem cell marker, Lgr5, were not found in HIOs, they are regarded as persisting in a transcriptional and functional state similar to fetal human tissue (38). As a feature of cell culture, HIOs are able to be cut into small pieces and passaged with specific growth factors in vitro for up to a year (38). Moreover, the size of HIOs is only approximately 2 mm, which demands cogitating to use HIOs in various evaluations (39).Generating HIOs from human PSCs requires the modification of an additional factor for each stage and 3D suspension and self-assembled culturing in a Matrigel droplet. HIOs contain a luminal component lined with many columnar epithelial cell types of the intestine, including enterocytes, goblet cells, Paneth cells and enteroendocrine cells, as well as supporting mesenchymal clusters, including fibroblasts, myofibroblasts and smooth muscle cells. The coincident differentiation of epithelial and mesenchymal tissues suggested that interaction between these tissues may be important in the developmental process of PSC-derived intestinal organoids. In the closed luminal structure of HIOs, mucosal epithelium exists that has polarity and exhibits introversion; enterocytes having apical microvilli and peptide absorption ability, that is, HIOs, have an epithelium structure and function that resembles the small intestine, although they are not mature in vitro. First, HIOs do not contain the ENS. In the early developmental process, neural crest cells migrate into the mesenchyme along the digestive tube and form the ENS, which consists of more than 100 million enteric neurons and 400 million glial cells (41). The neurons and glial cells are organized into the inner submucosal plexus and the outer myenteric plexus, regulating intestinal peristalsis movement with intestinal cells of Cajal that operate as pacemaker cells (42). Although HIOs contain mesenchymal cell types, including fibroblasts, smooth muscle and myofibroblasts, they do not have nerve plexuses and the interstitial cells of Cajal. The ENS comprises a functional intestinal neuronal–glial epithelial unit with intestinal epithelial cells, becoming a key regulator of the intestinal epithelial barrier function and of bowel movements (43). For in vitro studies on the role of gut microbiota using intestinal organoids, engineering of additional functional HIOs is required.Several limitations exist regarding the use of HIOs as a human small intestine (43). Although HIOs do not contain immune cells, including M cells, their applications as experimental models of mucosal epithelial barrier and intestinal infection have been reported. Thus far, the disruption of the intestinal mucosal epithelial barrier by Clostridium difficile, Salmonella enterica and Escherichia coli O157 is substantiated using HIOs as a bacterial infection model (46). In terms of viral infection models, although a method is lacking to construct efficient models that allow the infection of cultured cells or animal intestine with clinical strains of noroviruses, enteric adenoviruses and rotavirus, a study whereby clinical retrovirus isolates were replicated in HIOs was conducted by Finkbeiner et al. (47). The use of intestinal organoids as transformative new tools for human virus studies will facilitate an understanding of human viral pathogenesis and advance the development of vaccines, antivirals and therapeutics (48).Another missing structure in HIOs is the immune system, which is one of the major functions of the intestinal tract. The trachea and the gastrointestinal tract are organs that contact the outside and in which mucosal lymphatic tissue develops. In the small intestine, PPs consist of lymphatic follicles, T cells, B cells, macrophages, and dendritic and M cells. M cells are a subset of intestinal epithelial cells, reside in the region covering PPs, and take up luminal antigens and present them to PPs for subsequent intestinal immune responses (49). A failure of this intestinal ecosystem has been suggested in the etiology and morbidity of intestinal diseases such as inflammatory bowel disease and other systemic diseases (50). In these fields, intestinal organoids are expected to be used as a new biomodel of the host–microbe interface (51).The interaction between the intestinal microflora, including commensal and symbiotic bacteria and various cell types, in the intestinal tract constitutes not only regional but general homeostasis crucial for human health in vitro have been attracting increasing attention. HIOs were derived from stem cells with a deleted endogenous NEUROG3 candidate gene, responsible for congenital malabsorptive diarrhea and lost intestinal enteroendocrine cells. These demonstrated that NEUROG3 expression contributed to the differentiation of enteroendocrine cells. A decrease in the number of mature enteroendocrine cells is related to the development of this disease (33).Furthermore, applications of model organisms to observations of congenital gut disorders (39). The in vivo transplantation of HIOs under the kidney capsule in mice enabled the maturation of the epithelium and mesenchyme of such HIOs compared to those generated in vitro(52). In addition, HIOs containing a functional ENS were generated by co-culturing with human PSC-derived neural crest cells, which differentiate into glial cells and neurons, that are used for investigations of the cellular and molecular basis of Hirschsprung's disease (53).Taken together, HIOs are expected to be applied to investigations of human intestinal infection, which were not able to be undertaken using experimental animals, as well as in the development of medicines and in disease research. However, problems exist in generating fully functional intestinal tissue containing a mesenchymal tissue structure and the immune system (54). Mini-Gut differs from conventional intestinal organoids in several ways relating to its production and properties. Mini-Gut is directly differentiated from human PSCs under xenogeneic-free conditions, without animal-derived components such as Matrigel and may be preferred in clinical applications. Furthermore, the production of Mini-Gut mimics the functional interaction and self-assembly of the three primary germ layers from an initial stage of culture on micro-patterned substrates. This differs from HIO culture in which endoderm is first generated using the serial induction of growth factors; the differentiation of specific intestinal cell types and other germ layers is subsequently induced. After day 30, Mini-Gut is observed as self-organized cystic spheroids that begin to spontaneously detach from the plate. It has a cystic tissue structure, with the mucosal surface side facing outward and mesenchymal tissue surrounding the lumen on the inside . For the purpose of application to human disease research, although organoid technologies will become extremely valuable, it is essential to understand the characteristics of both intestinal stem cells and PSCs, including their advantages and disadvantages, and to utilize proper evaluating methods ((59); if highly functional intestinal organoids are used, these will become an appropriate biotool. Model systems including culturing devices and techniques have also been developed to reproduce complex in vivo human intestinal tracts and provide a platform for host–microbe crosstalk research and drug discovery applications (60). For instance, to improve accessibility of microbial adhesion and drug addition to the luminal side and operability of reactivity evaluation on the basement membrane side, methods to reconvert 3D structure prepared in Matrigel into 2D (single-layer) structure using TransWell membrane had been established (61). The air–liquid interface methods have been reported to allow long-term culture of various differentiated cell types derived from collected murine intestinal stem cells (62). To expand the use of organoids that form 3D tissue structures and have tissue-specific functions in research and development, new imaging technologies and evaluation techniques are required. Using hitherto 2D-imaging systems, detailed information obtained from organoids cannot be sufficiently utilized. Because organoids exhibit continuous tissue homeostasis, a 4D rather than a 3D-imaging system will help in monitoring developmental and tissue structure changes, including sequential changes and increase the usefulness of intestinal organoids, making these unprecedented biomedical tools carrying disease-associated genetic information. Integrating visualized and disease models using genome editing technologies, intestinal organoids could become in vitro biomodels, surpassing existing models.Presently, intestinal organoids generated from epithelial stem cells or PSCs are used in basic and disease research and in testines . Recentl methods . Moreovein vitro models for diseases such as enteric infections and gastrointestinal cancers. The organoids mimic the physiological condition between enteric pathogens and the host intestine and allow the analysis of the intestinal barrier function (64). The intestinal organoid system could apply for the intestinal mucosal vaccine development. In gastrointestinal cancer research, the organoid culture technique imitates and reflects an in vivo tumor microenvironment. Particularly, the mucosal epithelial organoids enable single primary patient-derived organoids as preclinical cancer models. These cancer organoids could potentially propose a physiologically relevant reaction, compared with existing simple cancer cell lines, and are expected to be applied in individualized medical care and efficient drug development (66).Intestinal organoids and PSC-derived organoids are also expected to be useful (67). As a method that replaces allotransplantation, introducing mucosal epithelial organoids generated from autologous tissue stem cells or human PSCs derived from epithelial/mesenchymal organoids, including HIOs, to patients is anticipated, and research and development on it is being undertaken (69). The generation of HIOs using the polyethylene glycol (PEG) hydrogel resolved a problem of using Matrigel extracted from mouse sarcoma in regenerative medicine (61). As a result of advances of the gastrointestinal tissue engineering, which promotes the maturation of intestinal organoids (53) and transplantation techniques of heterotopically engrafting small intestinal epithelial organoids into the colon (70), it may be possible to provide medical care that complements the lost intestinal function of the short bowel syndrome caused by extensive resection of the intestine due to necrotizing enterocolitis or inflammatory bowel disease.Many types of diseases originate in the intestine and colon, including congenital, genetic and multifactorial diseases. Many diseases, such as inflammatory bowel disease and Hirschsprung’s disease, lack effective or fundamental medical treatments, resulting in either a significant reduction in the quality of life or threaten the life of patients. The intestine affects all parts of an individual organism, including nutrient digestion and absorption, metabolism, immunity and commensal microbiota. Therefore, investigation of the use of intestinal organoids as cell sources in regenerative therapy for a lack of functional intestine is required. Although the total number of performed intestine transplants has decreased over the past decade because of improvements in medical and surgical treatments, intestine transplants will still play an important role in the treatment of intestinal failure, notwithstanding the difficulty of immune activity regulation of the intestine Research on PSCs, including human ESCs and iPSCs, has benefited developmental biological studies. Recently, the range of applications for PSCs has widened, including in disease modeling, pharmacological testing and regeneration, and cell transplantation therapy. Moreover, a combination of understanding the embryonic development of the digestive tract and the mechanisms that maintain homeostasis of intestinal mucosal epithelium have led to the innovative establishment of human gastrointestinal organoids generated from undifferentiated human PSCs. Investigations utilizing gastrointestinal organoids contribute to a detailed analysis of molecular interactions and result in synergistic development in these fields. Although intestinal organoids containing not only epithelium but also mesenchymal tissue that partially recapitulates the complexity of the intestine, more functional intestinal organoids generated from human PSCs are expected to promote further drug developmental studies and clinical regenerative therapies specifically targeted to patients.NoneThis work was partially supported by a National Center for Child Health and Development Seiiku Medical Study Grant (grant number 30-4) and the Danone Institute of Japan Foundation general researchers grant (grant number DIJF H30-036) to HA.The authors thank their funding sources: the Danone Institute of Japan Foundation and the National Center for Child Health and Development. We also thank Tomoyuki Kawasaki for figure illustration and laboratory colleagues for their helpful discussions.Satoru Tsuruta and Hidenori Akutsu made substantial contributions to the conception and design of the work and drafted and revised it critically for important intellectual content. Hajime Uchida made contributions to the conception of the work.This manuscript is a review article and did not need approval by an IRB.