This study aims to estimate the relationship between non-alcoholic fatty liver disease (NAFLD) and measures of atherosclerotic cardiovascular disease (ASCVD), and to determine to what extent such relationships are modified by metabolic risk factors.The study was conducted in the population-based Swedish CArdioPulmonary bioImage Study (SCAPIS) pilot cohort . NAFLD was defined as computed tomography liver attenuation ≤40 Hounsfield Units, excluding other causes of liver fat. Coronary artery calcification score (CACS) was assessed using the Agatston method. Carotid plaques and intima media thickness (IMT) were measured by ultrasound. Metabolic status was based on assessments of glucose homeostasis, serum lipids, blood pressure and inflammation. A propensity score model was used to balance NAFLD and non NAFLD groups with regards to potential confounders and associations between NAFLD status and ASCVD variables in relation to metabolic status were examined by logistic and generalized linear regression models.NAFLD was present in 106 (10.4%) of the subjects and strongly associated with obesity-related traits. NAFLD was significantly associated with CACS after adjustment for confounders and metabolic risk factors , but not with carotid plaques and IMT. The strongest association between NAFLD and CACS was observed in subjects with few metabolic risk factors .NAFLD was independently associated with coronary artery calcification but not with measures of carotid atherosclerosis in this cohort. The association between NAFLD and CACS was most prominent in the metabolically healthy subjects. Given the strong effect of obesity on liver fat as well as on a wide range of metabolic risk factors, results can be difficult to interpret if comparison groups are not balanced with regards to adiposity measures. Also, current MetS definitions do not include metabolic aberrations such as hyperinsulinemia and inflammation which theoretically might modify the relationship between NAFLD and ASCVD.Non-alcoholic fatty liver disease (NAFLD) represents a spectrum of liver conditions ranging from isolated fatty liver (steatosis) to fatty liver along with inflammation . Although NAFLD can progress to liver failure and hepatocellular carcinoma , 2, the In the current study we investigated the relationships between NAFLD, assessed by computed tomography (CT), and measures of atherosclerosis in the coronary and carotid arteries in middle-aged subjects in the population-based Swedish CArdioPulmonary bioImage Study (SCAPIS) pilot cohort. The main aim was to investigate the modifying effects of a wide range of obesity-driven metabolic risk factors, using the odds ratio as the effect parameter for associations between NAFLD and dichotomized ASCVD measures and the ratio of means as the effect parameter for associations between NAFLD and continuous ASCVD measures, and to address this aim we used a propensity score model to balance NAFLD and non-NAFLD groups with regards to obesity measurements and potential confounders.The SCAPIS pilot study is a population-based, cross-sectional study conducted at the Sahlgrenska University Hospital in Göteborg, Sweden . In 2012Among the 1111 participants, 1089 underwent CT imaging to assess fatty liver . A totalImaging for coronary artery calcification score (CACS) and fat depots  was performed using a dedicated dual-source CT scanner equipped with a Stellar Detector  as previously described . Liver aet al [The calcium content in each coronary artery was measured and summed to produce a total coronary artery calcification score (CACS) according to McCollough  and Agatet al . In repeet al  was 0.91Ultrasound examination was performed with an ultrasound scanner (SIEMENS S2000) equipped with a linear transducer 9L4. An ECG signal (lead II) was simultaneously recorded to be able to save a frozen image on top of the R-wave to minimize variability. The right and left carotid arteries were scanned from the distal part of the common carotid artery and approximately 10 mm upstream in the internal and external carotid arteries with the aim to identify and record the occurrence of atherosclerotic plaques. Images for measurement of IMT were recorded from the far wall in the CCA and the carotid bulb. At the position of the thickest part , two separately frozen longitudinal images were saved. One motion image loop was saved to assist in interpreting the frozen images . A plaquInformation on education level, alcohol, smoking and physical activity was collected through self-administrated questionnaires. Weight was measured with participants in light clothing, using calibrated scales. The body mass index (BMI) was calculated by dividing the weight (kg) by the square of the height (m). Blood pressure was calculated as the average of three measurements, after 5-minute supine rest, measured in the right brachial artery. Hypertension was defined as self-reported use of antihypertensive medications, or systolic blood pressure ≥140 or diastolic blood pressure ≥90 mmHg . DiabeteThe statistical analysis aimed at examining associations based on a model framework for underlying relationships between NAFLD, confounders, metabolic risk factors and outcomes  . NAFLD smetabolic risk factors for each outcome, including conceivable intermediates that might have been influenced/modified by the presence of NAFLD: diabetes status , hypertension, high LDL cholesterol, low HDL cholesterol, triglycerides, hsCRP and insulin. The continuous variables included as potential confounders and metabolic risk factors were categorized with cut-offs or corresponding to quartiles.We considered the following covariates as or IFG was counted as 1). Heterogeneity of association estimates across subgroups was examined by including in Model 2 the subgroup variable and an interaction term between this variable and NAFLD; a significant interaction term can be interpreted as significant heterogeneity of association across the subgroups. Statistical analyses were performed using SPSS for Windows, version 23.We employed logistic regression to examine the associations between NAFLD and the two dichotomous outcomes  and a generalized linear model to examine the association between NAFLD and IMT (log-transformed outcome data). Firstly, NAFLD was included as the only covariate (Model 1). Confounding was thereafter controlled for by including the propensity score as a covariate (Model 2). We checked for the model relationship between the propensity score and the outcome . For comparison, we also controlled for confounding by a conventional approach by including the potential confounders as covariates (rather than the propensity score as a covariate). Finally, we added the other metabolic risk factors to the set of covariates (Model 3). Inclusion of intermediates may yield over-adjustment; therefore, it is of interest to report estimates provided by Models 2 and 3, respectively. We repeated the analyses based on Models 1 and 2 for subgroups according to each factor among 7 metabolic risk factors. We also analyzed subgroups according to a summary measure: the number of metabolic risk factors present . Subjects with NAFLD were more obese and had more of obesity-related metabolic aberrations than subjects without NAFLD, including impaired glucose homeostasis, hypertension, high triglyceride levels, low HDL and high CRP . In the NAFLD was significantly associated with CACS after confounder adjustment using propensity score in Model 2  . The assSince there was no association between NAFLD and measurements of carotid atherosclerosis, we did not proceed with subgroup analyses for these outcomes. Subgroup analyses regarding CACS indicated that the association between NAFLD and CACS was modified by the metabolic risk factors . When thThe aim of this study was to estimate the relationship between NAFLD and measures of atherosclerotic cardiovascular disease (ASCVD) and to determine to what extent such relationships are modified by metabolic risk factors. Importantly, we wanted to address these questions without the influence of age, gender, obesity and other potential confounders. By using the propensity score model we balanced the NAFLD and non NAFLD groups with regards to adiposity and potential confounders, and subsequently investigated the association between NAFLD and ASCVD in relation to the overall burden of metabolic risk factors. For classification according to metabolic status we used variables that are known to be associated with obesity and/or ASCVD, including diabetes/IFG, hypertension, hyperinsulinemia, dyslipidemia  and elevated hsCRP. We found that the presence of NAFLD is associated with a two-fold increased risk of coronary artery calcification in the overall study population after confounder adjustment. Furthermore, the association between NAFLD and CACS remained significant after adjustment for all of the metabolic risk factors, and subgroup analyses showed that the effect estimate was primarily driven by the metabolically healthy part of the population. The strong association between NAFLD and CACS was contrasted by a lack of association between NAFLD and measures of carotid atherosclerosis (i.e. carotid plaques and IMT).Our finding that NAFLD is associated with coronary artery disease (CAD) is consistent with several previous reports, including population-based cohorts such as the Jackson Heart study , the FraSeveral previous studies reported NAFLD to be an independent predictor of CACS , 6–16, wThe strongest association between NAFLD and CACS was observed in subjects with few metabolic risk factors according to diabetes status, hypertension, LDL, HDL, triglycerides, hsCRP and insulin. In this subgroup we observed that subjects with NAFLD had as much as a 6-fold increased risk of coronary artery calcification as compared to those not having NAFLD. Previous studies in metabolically healthy subpopulations are scarce and therefore our findings should be confirmed in other cohorts before we can firmly conclude that the relative risk is of this magnitude. The present findings in the SCAPIS pilot cohort may be tested in the larger SCAPIS cohort  within a few years from now .1H-MRS to measure liver fat, both of them conducted in T2D patients, did not confirm a positive correlation: Petit et al reported that NAFLD was not associated with carotid IMT [et al found that liver fat was content was negatively associated with carotid plaque burden [In contrast to the strong association between NAFLD and CACS, we did not observe any convincing association with measures of carotid atherosclerosis in this population. The evidence of a positive correlation between NAFLD and carotid atherosclerosis in the literature mainly comes from hospital-based case-control studies and from Asian community-based studies using ultrasonography to assess liver steatosis , 38. Fewotid IMT  and Loffe burden . Taken te burden , 43. SinSeveral mechanisms through which NAFLD promotes ASCVD have been proposed. There is now convincing evidence that NAFLD is tightly linked to the cardiometabolic risk factors studied here, i.e. glucose dysregulation, dyslipidemia, hypertension and elevated CRP , 44. HowAn important strength of the present study is the well-characterized population-based cohort which made it possible to take into account a wide range of covariates in the analyses. Unlike most previous studies we have characterized the relationship between NAFLD and subclinical atherosclerosis in both coronary and carotid arteries. By studying both vascular regions in the same population using the same statistical approach it becomes apparent that NAFLD is more strongly linked to coronary than to carotid atherosclerosis. Our statistical approach using propensity score was primarily chosen to allow for separate handling of confounders and metabolic risk factors including conceivable intermediates that might have been influenced and/or modified by the presence of NAFLD. The propensity score method provides transparency of data analysis, since researchers can thereby assess whether observed confounding has been adequately balanced between the primary risk factor (in this case NAFLD). By comparison with a conventional approach to controlling for confounding , we demonstrated somewhat more pronounced downward adjustments of the effect estimates by the propensity score method.Some limitations warrant mention. The study is observational and cross-sectional, which limits the ability to infer temporal or causal relationships. This is a population-based cohort and therefore the number of subjects with pronounced ASCVD is relatively low. The data on history of liver disease from other causes were obtained from medical records and not an extensive laboratory screening, and we may therefore have included some subjects with secondary steatosis. The NAFLD diagnosis was defined based only on CT examinations and was not biopsy verified since such invasive procedures would not be feasible in a large population-based cohort. Thus, we were not able to distinguish simple steatosis from NASH. CT is a well-established method to noninvasively quantify liver fat and the method has been used in several prior epidemiologic cohorts to assess NAFLD status , 17, 35.In conclusion, our main findings were that NAFLD was significantly associated with our measure of subclinical CAD (i.e. CACS) and that the risk increase was most prominent in subjects with few metabolic risk factors. The results suggest that screening for subclinical CAD may be relevant for all NAFLD patients regardless of their metabolic status. Key questions for future research are to elucidate the mechanisms that link NAFLD to CAD, and to test if NAFLD-treatments can reduce CAD risk.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.


Xenopus to serve as a high throughput vertebrate organism to model patient-driven genetic diseases. This has facilitated the investigation of effects of patient mutations on specific organs and signaling pathways. This approach promises a rapid investigation into novel mechanisms that disrupt normal organ morphology and function. Considering that many disease states are still interrogated in vitro to determine relevant biological processes for further study, the prospect of interrogating genetic disease in Xenopus in vivo is an attractive alternative. This model may more closely capture important aspects of the pathology under investigation such as cellular micro environments and local forces relevant to a specific organ’s development and homeostasis. This review aims to highlight recent methodological advances that allow investigation of genetic disease in organ-specific contexts in Xenopus as well as provide examples of how these methods have led to the identification of novel mechanisms and pathways important for understanding human disease.Frog model organisms have been appreciated for their utility in exploring physiological phenomena for nearly a century. Now, a vibrant community of biologists that utilize this model organism has poised  Xenopus embryo. Recently, next generation sequencing technologies have allowed researchers to rapidly amass a compendium of candidate gene variants that are putatively disease causing .While results for CRISPR based knock-in technologies look promising in Xenopus , knock-iXenopus . AnotherXenopus is well suited for studying heart development as, unlike mice, Xenopus embryos do not require functional blood circulation for early cardiac development. This permits analysis of mutations that would prove embryonic lethal in mice. Additionally, the optical transparency which persists throughout early organogenesis enables assessment of morphological heart defects via multiple live imaging strategies   ; Table 1TCF-GFP) . In the TCF-GFP) . In part the LRO . Left–ri the LRO . Wnt act the LRO . Once ca the LRO . Shortly the LRO . As the  embryos .Xenopus tadpoles  gene which has been shown to have variants that cause human cardiomyopathy  has recently been employed as a reliable and efficient imaging modality to assess cardiac structural anomalies in tadpoles . OCT usetadpoles . OCT canmyopathy  was emplon times . OverallXenopus consists of a pronephros that is simplistic compared to human metanephroi  is the most prevalent class of birth defects leading to high infant mortality in the United States and yet for the vast majority of cases, the underlying molecular mechanisms remain elusive . HoweverRAPGEF5 which encodes a guanine nucleotide exchange factor for Rap-GTPase was found to have an internal duplication which would likely lead to a null allele in a heterotaxy patient. Depletion of Rapgef5 in Xenopus recapitulates the left-right patterning phenotype found in the patient. Unexpectedly, mechanistic studies established that RAPGEF5 regulates left-right patterning via Wnt signaling by regulating the nuclear localization of β-catenin  include a wide variety of patient presentations ranging from abnormal kidney and urinary tract size and morphology to tumor growth within this organ system  in steroid resistant nephrotic syndrome (SRNS) . SRNS iss status . This sts status . These fThough these studies demonstrate the power that patient driven gene discovery has for assigning disease causing variants in human kidney disease, they also begin to show how this approach aids in discovering novel mechanisms important for kidney morphogenesis and function. Moving forward, the many congenital kidney malformations that affect patients early in life may be an efficient source of genetic information to direct the study of kidney morphogenesis.Xenopus including that of the heart and kidney will allow us to unravel mechanisms of disease pathogenesis. Analysis of candidate genes in Xenopus will not only allow for assessing allele pathogenicity, but also promises to expand our understanding of developmental biology.Sequencing technologies continue to improve in speed, accuracy, and cost. Consequently, rather than phenotype driven therapy, patient genotype can be included in decision-making and treatment options tailored to individuals. However, one major challenge to this approach is determining the biological relevance of each variant as many candidate genes have no known role in disease. Therefore, each variant offers the opportunity for molecular function discovery in animal models. Studying organogenesis in All authors wrote and edited the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


To understand how DPSCs sense and respond to the mechanics of their microenvironments, it is essential to determine how these cells convert mechanical and physical stimuli into function, including lineage specification. This review therefore covers some aspects of DPSC mechanoresponsivity with an emphasis on the factors that influence their behavior. An in-depth understanding of the physical environment that influence DPSC fate is necessary to improve the outcome of their therapeutic application for tissue regeneration.Dental pulp is known to be an accessible and important source of multipotent mesenchymal progenitor cells termed dental pulp stem cells (DPSCs). DPSCs can differentiate into odontoblast-like cells and maintain pulp homeostasis by the formation of new dentin which protects the underlying pulp. DPSCs similar to other mesenchymal stem cells (MSCs) reside in a niche, a complex microenvironment consisting of an extracellular matrix, other local cell types and biochemical stimuli that influence the decision between stem cell (SC) self-renewal and differentiation. In addition to biochemical factors, mechanical factors are increasingly recognized as key regulators in DPSC behavior and function. Thus, microenvironments can significantly influence the role and differentiation of DPSCs through a combination of factors which are biochemical, biomechanical and biophysical in nature. Under  Dental pulp tissue contains an accessible source of multipotent mesenchymal progenitor cells, known as dental pulp stromal/stem cells (DPSCs), which participate in dentin and pulp regeneration . In the A key study by The teeth are essential for mastication and are subjected to various mechanical stresses due to jaw movement and occlusal forces which are transmitted to the dental pulp tissue and can subsequently influence DPSC fate . MoreoveThe signals involved in regulating SC fate are not only ECM proteins, adjacent differentiated cells, secreted and cell surface molecules but also mechanical signals. Notably it has been reported that in response to degradation, disruption and mechanical erosion DPSCs can differentiate into odontoblast-like cells to form dentin. Indeed several studies have now demonstrated that mechanical stresses transmitted to the pulp tissue can affect the behavior of DPSCs . TherefoDental pulp stem cells not only play a crucial role in dentinogenesis but also provide a promising source of cells for use in regenerative medicine . DPSCs cin vivo, many studies have investigated the effects of mechanical forces using in vitro models , promotes their osteoclastogenic differentiation which occurs during physiologic root resorption of deciduous teeth . NotablyThe overall aim of this review therefore is to report the effects of mechanical stimuli on the biological behavior of DPSCs as well as describing the associated intracellular signaling and odonto/osteogenic differentiation in DPSCs. The data discussed in this review indicates that appropriate mechanical stresses are important biological stimuli that can effectively promote proliferation and differentiation of DPSCs. The understanding of how these cells respond to mechanical stimuli (mechanosensitivity) is important for bone and tooth tissue engineering applications using DPSCs alone or in conjunction with biomaterials and other bioactive molecules such as growth factors and cytokines.Dental pulp stem cells are mechanosensitive cells that can recognize mechanical changes and transform this information into cellular responses . For exain vitro , equiaxial static tensile strain and micro- and nano-scale surface topographies have also been shown to induce osteogenic differentiation of DPSCs , prostaglandin E2 (PGE2) and cyclooxygenase-2 (COX-2) (of DPSCs . For exaof DPSCs  are able (COX-2) , 2011.As has been observed for osteocytes, the rapid stimulation of NO production by PFF in DPSCs appears to be related to the activity of the enzyme endothelial nitric oxide synthase (eNOS) but not to the activity of inducible NOS (iNOS) . ImportaAn interesting study by in vivo on DPSCs could be used to promote bone formation and limit bone resorption -6 and -1β, tumour necrosis factor (TNF)-α as well as the expression of the antioxidant genes heme oxygenase-1 (HO-1) and superoxide dismutases (SOD). The same authors also showed that mechanical stimulation using cyclic tensile strain induced expression of the odontoblastic markers DSPP, DMP-1, OPN, and BSP. Moreover, they demonstrated that the odontoblastic differentiation of DPSCs was mediated by the NF-E2-related transcription factor 2 (Nrf2)/HO-1 pathway (The biological response of DPSCs to mechanical stimuli occurs not only during orthodontic tooth movement but also during normal mastication. For example, DPSCs can produce growth factors, angiogenic changes and a mild inflammatory-type reaction in response to orthodontic forces , 2010. F pathway . These d pathway . In supp pathway .Dental pulp tissue is subjected to mechanical stress during normal masticatory process and also during pathological trauma or orthodontic tooth movement .Moreover, it is also well known that occlusive force plays a role in physiological root resorption of deciduous teeth, in which dental pulp cells from these mechanically stressed teeth secrete cytokines that are important for odontoclast activation and subsequent root resorption .Therefore, DPSCs located near the roots of teeth are subjected to higher levels of oral mechanical stress by jaw movement and occlusal forces. Moreover, orthodontic forces transfer horizontal stretch to DPSCs, and also during tooth eruption dental pulp tissue is stretched vertically .Furthermore, during physiological mechanical loading of teeth, the dentin is subjected to fluid flow which can activate the nocireceptors and mechanoreceptors on odontoblast processes present in the dentin tubules that regulate the maintenance of tooth integrity .Importantly, To the best of our knowledge, until now only the study by Regenerative medicine is an important application of DPSCs and this necessitates a thorough knowledge of mechanobiology. However, the studies analyzed in this review demonstrate that the response of DSPCs to mechanical stimuli differs according to the type and source of the mechanical forced applied Table . ImportaThe understanding of the mechanical stimuli that regulate DPSC behavior will not only improve our knowledge relating to mechanisms involved in their differentiation but could also provide valuable insights for optimizing DPSC-based therapies. In fact, the particular mechanoresponsiveness of DPSCs to mechanical stimuli may be of utility for future bone and teeth tissue engineering applications.in vivo environment on DPSCs to promote the regeneration of dental and bone tissues.Although physical and mechanical factors are known to play a key role in regulating DPSC fate, further studies are required to elucidate the detailed molecular mechanisms and signaling pathways involved in the mechanosensitive response of DPSCs to various types of forces.Ultimately it may be possible to use mechanical forces that mimic the dynamics of the FP, MM, BC, RS, BS, PC, and MT conceived and wrote the manuscript. All authors reviewed and approved the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


In an era of precision medicine, it seems regressive that we do not use stratified approaches to direct treatment of oral corticosteroids during an exacerbation of chronic obstructive pulmonary disease (COPD). This is despite evidence suggesting that 40% of COPD patients have eosinophilic inflammation and this is an indicator of corticosteroid response. Treatments with oral corticosteroids are not always effective and not without harm, with significant and increased risk of hyperglycemia, sepsis, and fractures. Eosinophils are innate immune cells with an incompletely understood role in the pathology of airway disease. They are detected at increased levels in some patients and can be measured using non-invasive methods during states of exacerbation and stable periods. Despite the eosinophil having an unknown mechanism in COPD, it has been shown to be a marker of length of stay in severe hospitalized exacerbations, a predictor of risk of future exacerbation and exacerbation type. Although limited, promising data has come from one prospective clinical trial investigating the eosinophil as a biomarker to direct systemic corticosteroid treatment. This identified that there were statistically significant and clinically worsened symptoms in patients with low eosinophil levels who were prescribed prednisolone, demonstrating the potential utility of the eosinophil. In an era of precision medicine our patients’ needs are best served by accurate diagnosis, correct identification of maximal treatment response and the abolition of harm. The peripheral blood eosinophil count could be used towards reaching these aims. Eosinophils are inflammatory leukocytes comprising of bi-lobed nuclei and large acidophilic cytoplasm granules. The cationic protein granules bind to acid stains and in particular eosin which allowed them to be identified by Paul Ehrlich in 1879 .. Expression of C-C chemokine receptor type 3 (CCR3) and IL-5 receptor subunit alpha (IL5Rα) on the cell surface is integral to eosinophil recruitment to tissues. The role of the eosinophil in both innate and adaptive immunity is poorly defined, and their response varies depending on the environment and/or stimulus. Resident eosinophils are predominately found in the gastrointestinal tract, although other resident populations are also found in healthy individuals in physiological conditions in the thymus, spleen, lymph nodes, mammary glands and the uterus indicating other potential roles in homeostasis. Their accumulation at these sites has given rise to the eosinophil role extending to local immunity and/or remodeling and repair in health and disease  [Produced in healthy bone marrow derived from CD34+ myeloid progenitors, the number of eosinophils generated is typically low with circulating eosinophils range between 1–4% of the total white blood cell count. Once mature, eosinophils enter the systemic circulation where they can reside for 8–12 h. Unless stimulated, the cells then migrate to tissues . The mecothesis) .Eosinophils play an important role in immune-regulation by priming B cells and maintaining type-2 immunity . In the In asthma, another common airway disease, the utility of the eosinophil to identify a corticosteroid response has been established , leadingThe involvement of the eosinophil in the pathogenesis of chronic obstructive pulmonary disease (COPD) has not been fully elucidated  and theiSince the emergence of the peripheral blood eosinophil as a useful biomarker in COPD, several post-hoc analysis have explored its utility in directing inhaled corticosteroids to impact on exacerbation burden ,28,29,30Exacerbations of COPD are heterogeneous and attempts to delineate the biological heterogeneity using plasma markers and symptoms have been made . UnsuperAt present, there is no ideal method to direct treatment during an exacerbation of COPD. In the presence of dyspnea, exacerbations are usually always treated with systemic corticosteroids . This is3) being an independent predictor of mortality in severe hospitalized exacerbations of COPD [In a post-hoc analysis from severe hospitalized exacerbations, length of hospital stay is significantly shorter in eosinophilic exacerbations treated with systemic corticosteroids  while re of COPD .Clinical trials using biomarkers such as Procalcitonin  or C-reaIn an era of precision medicine , our pat


With its number of employees ranging from 45,310 to 46,000, the textile and apparel industry is the main private sector employer of labour in Lesotho. It has been reported that a third (an estimated 34%) of these workers are living with HIV. There is perception that textile factory workers living with HIV (TFWLWH) in Lesotho indulge in HIV risk-taking behaviours. However, no study has yet investigated or documented factors that influence risk-taking behaviours amongst these workers. Transmitting the disease to others, treatment complications and death consequent to HIV reinfection are complications associated with HIV risk-taking behaviours by seropositive individuals. Using an in-depth, face-to-face, semi-structured interview, this study obtained the perspectives of ten factory workers from three randomly selected textile factories in Maseru, Lesotho on factors that influence HIV-risk taking behaviour amongst TFWLWH in Lesotho. Analysis of the comments given by workers revealed four core themes, namely, peer pressure, communication, cultural norms and societal norms. Determining the predictors of HIV risk-taking behaviours amongst these workers will inform both present and future interventions aimed at supporting textile factory workers living with HIV in Lesotho. This supports the need for continued research to identify HIV risk-taking behaviours by people living with HIV countrywide, to decrease the incidence of new infections and complications arising from reinfection. Since recording its first case of AIDS in 1986, the number of HIV infections in Lesotho has increased exponentially , LesothoThis research was designed as a qualitative study that made use of an in-depth, face-to-face, semi-structured interview to obtain qualitative data.In-depth, face-to-face, semi-structured interviews: participants were interviewed using a pretested schedule.Study settings: study was conducted at the premises of three randomly selected textile factories (names withheld) located in the Maseru district of Lesotho.Study participants: participants in this study were selected using the following eligibility criteria: textile factory workers belonging to the ALAFA support group; male or female factory workers, 18 years or older; workers infected with HIV and receiving antiretroviral treatment.Unit of analysis: a purposive and voluntary sampling technique was used for this study. Eligible individuals who were willing to participate were invited and briefed during an introductory meeting where confidentiality and the voluntary nature of participation was ensured. Nine volunteers (three in each factory) participated in the study.Exploratory interview: an exploratory interview was conducted to test the suitability of the questions in the interview guide. This interview was conducted with one eligible participant, and no changes to the interview schedule were indicated after administration. Findings from the exploratory interview was included in the dataset.Data collection: interviews were conducted according to a schedule of predetermined appointments in a private, quiet room, in the presence of an independent observer and each session lasted about 45-60 minutes. All interviews were conducted in the local language of the respondents (Sesotho) and were audio recorded. Field notes were taken to enable reflection on details of each interview. Immediately after each interview session, all notes and audio recordings were reviewed. This resulted in a process of concurrent data collection, preliminary analysis, reflection and progressive focusing.Data analysis and interpretation: the recordings were transcribed and translated to English by one of the researchers and quality assured by the independent observer. Thematic analysis formed the cornerstone of the analysis, and was done using NVivo 12 software . Specific attention was given to patterns and emerging themes.Ethical considerations: the Health Sciences Research Ethics Committee of the University of the Free State (ECUFS Nr 158/2014) approved this study.Demographic information of participants: ten interviews were conducted (7 women and 3 men). Participants were between the ages of 18 and 31 years.Factors that influence engaging in HIV risk-taking behaviours by workers: analysis of the comments given by the workers revealed four core themes, namely, peer pressure, communication, cultural norms, and societal norms. Each theme is supported by the participants' responses quoted verbatim.Peer pressure: participants stated that peers play a key role in decision-making, encourage risky behaviours and give misleading information. "Under this working environment peers influence their friends' decision-making, especially young people are encouraged to have premarital sex, and they also attract older men" "Young women encourage each other to have sex outside marriage." "There is a growing number of people living with HIV and AIDS who engaged in unprotected sexual activity with HIV negative people, because they get misleading information from peers that when one have unprotected sex with a person who is not infected HIV can be cured". In addition, participants imply that lacking information, placing too much trust in peers and desiring to "fit in" and impress their friends are probable reasons why workers succumb to peer pressure. "If a person do not have the right information, cannot say no sometimes friends will influence us." "Most young people depend too much on the trust they have in friends (because they grew up together) peer pressure justify their practice of unprotected sex and of not using condom" "Because of peer pressure regardless of the fear of HIV/AIDS, they (referring to some colleagues) are still afraid to buy their own condoms, afraid of what their friend will say".Communication: participants in this study stated that (poor) communication influences HIV risk-taking behaviour by workers. Participants reported that they lack facts about HIV in their communities and this leads to risk-taking behaviour by workers. "The lack of HIV facts information in our community results in people not to disclosed their HIV status and those who are not tested not go for voluntary testing". Furthermore, participants referred to poor parental communication about sex and HIV infection. "Poor communications between young people and their parents on the subject of sex create a lot of problems. They are therefore forced to listen to friends." "Our parents do not talk about sex, HIV, or condoms frequently with their children because they considered these topics disrespectful, on the other side".Cultural norms: workers mentioned community-held, socially shared cultural norms and beliefs on condom use, multiple sexual partners, gender inequality and unequal communication practices as factors that influence HIV risk-taking behaviours. Men's reluctance to use and their dislike for condoms, and the belief that only unfaithful woman, would request male partners to use condoms prevents women from practising safe sex. "Married women who want to protect themselves are suspected to be unfaithful if they ask their husbands to use condoms." "In our community we are facing culture of continued reluctance to use condoms in sexual relationships" Communities often revere men who have more than one partner. "Cultural norms are condoning men to have different sexual partners and the same men are refusing to use condoms therefore contribute significantly to spreading HIV and are especially resilient to change in most cases". Community beliefs and cultural norms may perpetuate gender inequality (women are expected to be subservient to men) and shape patterns of communication within intimate relationships  ["I am also aware that culture has a role to play especially among women to make decisions, women cannot negotiate safe sex, the culture of paying lobola (pride price), proofs women's sexual obligation, gives men the power over women that they own them and therefore women cannot negotiates or decide when and how to have sex" "We don't talk to our husband about sex, it's our culture".out sex) . "I am aSocietal norms: participants in this study cited fear of stigma and discrimination, particularly fear of social rejection by others, losing their jobs and being humiliated, as factors that promote HIV risk-taking behaviour."People still get fired from their jobs and people still get kicked out of their houses. Moreover, the fear is there even now most people are not able to disclose their HIV status to their partners. They do this to avoid rejection, refusals to have sex or of being stigmatized or discriminated against." "Community members may not seek testing because of the fear of discovering that they are HIV positive and fear of the resulting stigma and discrimination. Further, family members may encourage relatives with HIV/AIDS to remain silent about their HIV status to prevent gossip, social rejection and HIV-related stigma". This finding highlights the need for continued work in Lesotho to address HIV discrimination and stigmatisation. Factory workers who participated in this study report that low wages/income promote transactional sex by female factory workers. "Transactional sexual relationships is encouraged somehow in our community because of the state of poverty as a result of insufficient income in the family. This leads to high rates of migration, now women left their homes and villages to care for families. These in most cases lead one to have sex partner relationship  in order to receive additional support for food, transport". Moreover, it has been reported that the majority of sex workers in the districts of Maseru and Leribe were originally textile workers who found sex work more profitable than the low wages offered by the factories [actories .Efforts to curtail the spread of HIV have been directed along two distinct, strategic avenues: (a) preventing new infection and (b) early diagnosis and initiation of treatment in a maximum number of HIV-infected persons . In a reWhilst acknowledging the contribution made by current HIV prevention, treatment and support programmes for textile factory workers in Lesotho, this study confirmed the perception that TFWLWH in Lesotho indulge in HIV risk-taking behaviours. It is recommended that further investigations be carried out to establish if the factors associated with HIV risk-taking behaviours as identified by this study are present in the greater Basotho population. This should be done prior to proposing recommendations for decision-makers in health departments, community-based organisations and other stakeholders involved in the prevention of HIV and AIDS in Lesotho.Lesotho has one of the highest HIV burdens in the world;Textile factory workers in Lesotho have the second highest HIV prevalence (42.7%) amongst worst affected groups.This study confirmed the perception that TFWLWH in Lesotho indulge in HIV risk-taking behaviours;This study identified and contextualised factors associated with risk-taking behaviours amongst TFWLWH in Lesotho.The authors declare no competing interest.


Apteronotus leptorhynchus. We summarize known population coding strategies at the peripheral level, which are largely unaffected by noise correlations. We then move on to the hindbrain, where existing data from the electrosensory lateral line lobe (ELL) shows the presence of noise correlations. We summarize the current knowledge regarding the mechanistic origins of noise correlations and known mechanisms of stimulus dependent correlation shaping in ELL. We finish by considering future directions for understanding population coding in the electrosensory pathways of weakly electric fish, highlighting the benefits of this model system for understanding the origins and impact of noise correlations on population coding.The fact that perception and behavior depend on the simultaneous and coordinated activity of neural populations is well established. Understanding encoding through neuronal population activity is however complicated by the statistical dependencies between the activities of neurons, which can be present in terms of both their mean  and their response variability (noise correlations). Here, we review the state of knowledge regarding population coding and the influence of correlated variability in the electrosensory pathways of the weakly electric fish  Understanding the concerted activity of neural populations remains a central problem in systems neuroscience. While simultaneous recordings of multiple neurons  has become increasingly feasible across animal models and brain areas, interpreting these data are often complicated. This is because neuronal activities are often not independent of one another, but rather show correlations. Such correlations have been found almost ubiquitously across species and brain areas .Here we review the current state of knowledge regarding how electrosensory neural populations encode behaviorally relevant stimuli in wave-type weakly electric fish. This model system is well described in terms of its anatomy and single cell physiology and readily accessible for When evaluating the statistical dependency between the spike trains of two neurons, their correlations (raw-correlation) can be decomposed into two types of correlations . First, The neuron pair shown in Figure In Figure The insets in Figures In the absence of stimulation, raw-correlations between neural activities are often termed “baseline correlations.” These baseline correlations represent the limit that noise correlations will tend toward as stimulus amplitude goes to zero . TherefoOne could argue that the detrimental effects of noise correlations toward stimulus discrimination in the example shown Figure  might seRecent studies have shown that noise correlations are not static but can change in magnitude based on various factors such as the animal’s state , the aniWhile correlations, both signal and noise, are found ubiquitously in the CNS, in many cases understanding their mechanistic origins remains elusive. On the one hand, it is clear that signal and noise correlations in pairs of neurons will highly depend on their input connectivity Figure . Signal There has been progress made toward understanding the mechanistic origins of neural correlations and their effects on coding, and it has become clear that such understanding will require detailed knowledge of the anatomical connections. Nonetheless, the connections of neural networks forming circuits in the brain and their inputs and outputs are often numerous, divers, and highly complicated. Toward this end, the electrosensory pathway benefits from a relatively simple and well-characterized anatomy, which should be advantageous for the investigation of the basic mechanisms underlying correlated variability.A. leptorhynchus emit a quasi-sinusoidal electric signal referred to as the “electric organ discharge” (EOD) thereby building up an oscillatory field surrounding their body  . When twDuring social interaction, fish can emit short-term alterations of their EOD frequency with the purpose of active social communication . Such evSo far, we have only considered stimuli that consist of changes in the mean EOD amplitude. These are sometimes referred to as “first-order” stimuli. However, it is clear that rather than being stationary, fish move extensively during social interactions, thereby causing changes in the amplitude of the beat stimuli . Such “sA. leptorhynchus). However, electrosensory behaviors consist of changes in the animal’s electric field and, as such, can be also be elicited in immobilized animals  are detected by about 16,000 tuberous electroreceptors or electrosensory afferents (EA) that are distributed across the animals’ body and embedded in its skin . At baseEAs project to the hindbrain Figure . All thrThere are two major types of descending pathways to ELL that originate from higher brain areas Figure , orange.As described above, chirp waveforms of a given chirp are very heterogeneous in nature depending on the phase of the AM at which they are emitted (“chirp phase”). Nonetheless, chirps occur with equal probability at any phase of the beat during electrocommunication . The chiInterestingly, these results were similar for both simultaneous and non-simultaneous recordings. As explained above, non-simultaneous recordings cannot be used to infer noise correlations but only signal correlations. This suggests that changes in EA’s correlations are primarily driven by changes in signal correlations and that noise correlations between EAs are either negligible or do not affect such coding. This is supported by other studies showing that correlations between EA baseline activities are negligible except at the EOD frequency and higher harmonics . Based oRecent studies have focused on understanding how EA’s respond to envelopes through changes in firing rate . While eAs found during EA correlation encoding of chirps, results were almost independent of whether simultaneous or non-simultaneous recordings were used for the analysis Figure , inset, ELL pyramidal neurons receive convergent input from EA projections: Anatomical studies have shown that up to about 65% of inputs are shared between pyramidal neurons in neighboring columns . As suchUnder baseline conditions, ELL pyramidal neurons fire action potentials in an irregular pattern switching between bursts and single spikes Figure  Bastian with disThe average RF size and overlap between neighboring neurons was estimated to differ between ELL segments and decrease from the lateral segment (LS) over the centro lateral segment (CLS) to the centro medial segment (CMS) . Based oELL RFs are organized in an antagonistic center-surround organization . This meUsing mathematical models and numerical simulations, it was found that changing the relative strength and size of the RF surround relative to that of the RF center impacts the magnitude of baseline correlations Figure . InteresUnder stimulation, pyramidal neurons will typically encode the stimulus waveform through changes in firing pattern while the overall firing rate changes only minimally on average Figure . This isrnoise: -0.35 to 0.39) as compared to the relation between raw and signal correlations . Furthermore, the spread of the data is less . This could be seen as indication that noise correlations more strongly determine raw correlations between ELL spike trains than signal correlations are.We further compared noise and signal correlations and found that the correlation structure in ELL is mostly parallel  Figure . HoweverBased on the observed correlation structure Figure , one wouSimilar to what is reported in other brain areas, noise correlations in ELL are highly plastic . Indeed, it was shown that their magnitude strongly depend on the stimulus’ spatial extent . SpecifiThis effect is timescale specific: on short timescales (<10 ms) correlations slightly increased under global stimulation while on longer timescales (>10 ms), correlations strongly decreased  Figures, arrows.in vivo Remain Poorly Understood”), noise correlations likely arise due to the shared noise in common inputs. For ELL, these are the afferent inputs from EAs  are encoded within the electrosensory pathway. As mentioned above, most studies have focused on how single EA’s or ELL pyramidal cells encode such stimuli. While extrapolations to the population level were attempted, the potential effects of noise correlations were generally neglected . HoweverWith regard to electrosensory envelopes, previous studies have largely focused on understanding the encoding by single ELL pyramidal cells . FurtherFinally, it should be noted that theoretical studies have suggested that noise correlations themselves could directly encode stimuli, therefore forming an independent channel of information transmission in the brain . The docWhile many of the above discussed assessments of neuronal encoding are based on recordings from immobilized animals, it is important to note that weakly electric fish, on top of being able to display electrical behaviors when immobilized, show elaborate behaviors and astonishing cognitive abilities and are getting more and more attention for the study of various aspects of active sensing behaviors . Recent VH and MC conceived the study, reviewed and edited the text and figures. VH curated and re-analyzed the data, prepared the figures, and wrote the initial draft.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


HeteroGenesis simulates more complex and realistic heterogeneous tumour genomes than existing methods, can model different evolutionary dynamics, and enables the creation of multi-region and longitudinal data.Tumour evolution results in progressive cancer phenotypes such as metastatic spread and treatment resistance. To better treat cancers, we must characterize tumour evolution and the genetic events that confer progressive phenotypes. This is facilitated by high coverage genome or exome sequencing. However, the best approach by which, or indeed whether, these data can be used to accurately model and interpret underlying evolutionary dynamics is yet to be confirmed. Establishing this requires sequencing data from appropriately heterogeneous tumours in which the exact trajectory and combination of events occurring throughout its evolution are known. We therefore developed HeteroGenesis: a tool to generate realistically evolved tumour genomes, which can be sequenced using weighted-Wessim (w-Wessim), an https://github.com/GeorgetteTanner, implemented in Python and supported on linux and MS Windows.HeteroGenesis and w-Wessim are freely available under the GNU General Public Licence from Bioinformatics online. The evolution of a tumour affects its clinical course: malignant phenotypes undergo selection resulting in increased growth, invasion, metastases or therapy resistance. Elucidating the mode and course of tumour evolution therefore impacts upon our understanding of disease progression and how to more effectively treat patients . Numerouin silico whole exome sequencing (WES) tool, Wessim  to be incorporated into the genomes for each clone in a tumour, along with a matched germline. It takes as input: (i) a reference FASTA genome sequence, (ii) an optional file containing known germline SNV and InDel locations and minor allele frequencies formatted from dbSNP and (iii) a JSON file containing a set of parameters. It outputs a JSON file with lists of variants for each clone in the simulated tumour and a matched germline, as well as files containing the order that mutations occurred. heterogenesis_varincorp is run separately for each clone, and the germline sample. It takes the lists of variants generated by heterogenesis_vargen and incorporates them into the reference genome sequence, as well as calculating copy numbers and variant frequencies along the genome. freqcalc is then run to combine variant profiles for individual clones from heterogenesis_varincorp outputs to describe one or more bulk samples.HeteroGenesis consists of three consecutive python programmes  include simulated germline sequencing data to mimic the contamination of normal cells in a tumour sample, (ii) create multiple bulk datasets from the same tumour with different proportions of clones to mimic multi-region sampling, an approach that enables more reliable delineation of subclone architectures  as the probes in the BLAT alignment. By filtering these reads for those with a high alignment mapping score and adjusting the stringency of the BLAT alignment, we are able to generate reads with w-Wessim that match the distribution of reads seen in the real data, with 79.7% of bases aligning on or near target and a mode coverage of 28x . A demonbty1063_Supplementary_DataClick here for additional data file.


With interest, we read the paper published by Klingele et al.  in a recBecause citrate metabolism is oxygen dependent via the citric acid cycle and therefore mainly takes place in organs with high amounts of mitochondria, such as the liver, kidney, or muscle , they hyIn neonates with mitochondrial cytopathy , the citIt is well known that in all cells, citrate can be metabolized within the Cori cycle (tricarboxylic acid cycle) . It descIn the setting of citrate anticoagulation, with disturbed microvascular circulation causing altered hepatic function and citrate accumulation , it is a Honore and colleagues report about two neonates suffering from mitochondrial cytopathy being able to metabolize citrate despite citric acid cycle dysfunction. They conclude from this interesting observation that this infers activation of an inducible pathway, and they refer to our study .We reported about long-term continuous renal replacement therapy (RRT) and anticoagulation with citrate (RCA) in critically ill patients with severe liver dysfunction and unexpected low rates of citrate accumulation. Thus, the citrate metabolism in extrahepatic organs with high amounts of mitochondria such as the muscles was mostly sufficient to deal with the amount of citrate delivered by CRRT with RCA. We therefore concluded that a substantial extrahepatic metabolism of citrate seems to exist. However, in case of disturbed microcirculation, both hepatic and extrahepatic citrate metabolisms were clearly disturbed . In a syOur hypothesis was that the risk of citrate accumulation during regional citrate anticoagulation is more dependent on microcirculation than on liver function. Disturbed microcirculation results in elevated lactate due to impaired organic substrate metabolism. Therefore, lactate is a risk factor for impaired citrate metabolism. This is supported by Khadzhynov and colleagues . HoweverHonore and colleagues agree that disturbed microvascular circulation and lactate are a hint for citrate accumulation and that extrahepatic citrate metabolism explains why CRRT with RCA in case of liver dysfunction not necessarily results in citrate accumulation. Moreover, they suggest an inducible pathway which can be activated despite mitochondrial cytopathy and citric acid cycle dysfunction. This is an interesting and novel approach, being able to explain why hepatic function may be less important for citrate metabolism in CRRT with RCA than previously thought.


Nutrition is known to modulate the immune system and may alter neuroinflammatory processes implicated in the pathogenesis of Alzheimer’s disease (AD) and progression of neurodegeneration. Here, we review the evidence for healthy dietary patterns and age-related cognition and discuss potential neuroinflammatory actions of diet on cognitive function.Anti-inflammatory dietary patterns such as the Mediterranean diet (MD) and dietary approaches to stop hypertension (DASH) may be neuroprotective. Several dietary components consumed in the MD and DASH  can inhibit neuroinflammation associated with AD. Anti-inflammatory diets may also attenuate neuroinflammation via indirect immune pathways from the gut microbiome and systemic circulation.Diet may influence cognitive ageing via several inflammatory pathways. However, data from human studies are lacking and the exact mechanisms linking diet to cognitive function remain elusive. Further dietary intervention studies are required to investigate diet-associated neurological change from the earliest through to latest stages of cognitive decline. Furthermore, incorporation of neuroimaging measures in intervention studies would advance current understanding of the mechanistic effects of dietary modification on neuroinflammation in the ageing brain. Worldwide, 50 million people are living with Alzheimer’s disease (AD) and related dementias, and this figure is projected to triple by 2050 unless preventive measures are developed . AddressThe mechanisms governing dietary influences on cognition are not clear, but inflammatory pathways are likely to be involved , 6. InflDiet is known to modulate the immune system ••, and sMechanistic studies in the brain have primarily focused on single nutrients. However, the synergistic effects of nutrients and foods when consumed together as a usual dietary pattern are likely to exert greater effects on inflammatory processes and neurodegeneration during ageing , 27. TheThis review aims to summarise available evidence to evaluate the role of MD and DASH dietary patterns for cognitive health and AD risk during ageing and discuss underlying inflammatory mechanisms of diet on cognitive function in humans and animal models.The MD describes the traditional dietary pattern consumed in Mediterranean countries and is characterised by high intake of fruits, vegetables, wholegrains, nuts and legumes; moderate intake of fish, poultry and alcohol  and low intake of red and processed meats with olive oil used as the main fat source , 50. AdhObservational studies that have evaluated the MD for cognitive health are summarised in Table It is possible that MD contributes to neuronal integrity across the life-course. Emerging evidence from observational studies report a link between greater MD adherence and more favourable brain structures and functions that protect against neurodegeneration including increased cortical thickness , 48, greThe effect of MD on neurocognition has been evaluated in a limited number of randomised controlled trials (RCTs). The PREDIMED study demonstrated a modest beneficial effect of increased MD adherence over 4–6 years on cognitive function in cognitively healthy adults at high CVD risk , 4, 71 pThe DASH diet is an accepted non-pharmacological treatment for hypertension  and, likCollectively, the available evidence suggests that MD and DASH patterns may provide protection against neurodegeneration during ageing, with more consistent associations in favour of the MD, probably owing to a greater number of studies that have examined MD and cognition, relative to DASH or indeed other dietary patterns. Further prospective studies in diverse populations are recommended to determine relations between dietary patterns and clinically relevant measures of cognitive decline as well as incident AD. While few dietary intervention studies have been conducted, beneficial effects of MD on cognition are shown in trials of longer duration (12 months or more) , 60 and The MD and DASH dietary patterns are typically low in saturated fat, sugar and high in antioxidants, fibres and polyphenols, which have potential independent effects on brain health. It is likely that the myriad of bioactive compounds and nutrients consumed at synergistic levels within these dietary patterns exert the potential to reduce neuroinflammatory processes involved in neurodegeneration .KB and inhibiting pro-inflammatory cell signalling pathways [Nutrients have important physiological roles for normal brain functioning and are transported into the brain via the blood–brain barrier (BBB) or from the choroid plexus transport locus of the blood-cerebrospinal fluid barrier by distinct mechanisms, such as facilitated diffusion and active transport . The effpathways , 79. Lonpathways , 80.While these animal studies provide important insights into mechanistic actions of diet on neuroinflammation, it is not clear how well the findings translate to humans, due to potential differences in nutrient bioavailability and metabolism. Indeed, single nutrient supplementation has shown no cognitive benefit among older adults in the few trials that have been conducted so far •, 82•, aI2 = 67.7%) compared to usual diet which tended to be of greater magnitude in trials with longer duration [Increased peripheral inflammatory markers have been associated with neurodegeneration  and are duration . The MD duration , 29. Adoduration . Furtherduration , 88••. HEmerging observational data lend support to systemic inflammation as a driver between diet and neurocognition Table . High prThe human gut microbiome represents the collective genomes of 10–100 trillion microorganisms harboured in the gastrointestinal tract and is considered important for healthy immune function . The ageAlmost 60% of variation in the gut microbiome is attributable to diet ; therefoGiven the lack of effective treatments and projected increased prevalence of AD, there is considerable interest in understanding the contribution of neuroinflammation to the pathogenesis of AD in order to develop effective preventive strategies for cognitive decline.Compelling evidence shows that nutrients and other bioactive dietary compounds influence neuroinflammatory processes leading to neurodegeneration in animals and that nutrients can act synergistically to exert greater biological effects. It is plausible that diets rich in anti-inflammatory components attenuate neuroinflammation via several immune pathways within the brain and indirectly from the gut microbiome and systemic circulation. However, data from human studies are lacking and the exact inflammatory mechanisms linking diet to cognitive function remain elusive.Growing evidence supports a protective effect of anti-inflammatory dietary patterns, especially the MD, against cognitive decline in older persons but causal associations between diet and AD remain uncertain. The evidence base has been recently strengthened by small intervention studies showing improvements in cognitive function in response to MD and calorie-controlled DASH. However, adequately powered intervention studies with larger sample sizes and longer durations are required to examine the effect of dietary modification on clinically relevant cognitive outcomes. Neuroimaging studies have shown direct beneficial associations of MD on preclinical changes associated with AD; therefore, brain biomarkers should be considered as end-points in future intervention studies to investigate diet-associated neurological change from the earliest through to latest stages of cognitive decline. Furthermore, incorporation of PET (positron emission tomography) could allow measurement of in vivo microglia activation  and advaclinicaltrials.gov Reference NCT02817074, NCT03688126) and will generate valuable data to determine whether improving diet quality is an effective strategy to improve brain health in older adults.While most research attention has been given to MD and DASH, there is still much to learn about the ideal combination of foods and nutrients for optimal cognitive health during ageing. More recently, the MIND (Mediterranean-DASH diet intervention for neurodegeneration delay) dietary pattern that incorporates foods based on evidence in the diet-dementia field has shown to be more predictive of cognitive decline than the MD  and has


In diseases such as multiple sclerosis (MS), inflammation can injure the myelin sheath that surrounds axons, a process known as demyelination. The spontaneous regeneration of myelin, called remyelination, is associated with restoration of function and prevention of axonal degeneration. Boosting remyelination with therapeutic intervention is a promising new approach that is currently being tested in several clinical trials. The endogenous regulation of remyelination is highly dependent on the immune response. In this review article, we highlight the cell biology of remyelination and its regulation by innate immune cells. For the purpose of this review, we discuss the roles of microglia, and also astrocytes and oligodendrocyte progenitor cells (OPCs) as they are being increasingly recognized to have immune cell functions. Demyelination is a common feature of disease occurring after conditions such as spinal cord injury . Schwann cells (SCs) are glial cells generally restricted to the PNS that arise from neural crest-derived SC precursors , secreted by microglia/macrophages as well as astrocytes in mouse cuprizone-induced lesions, promotes OPC expansion and remyelination through TNF receptor 2 (TNFR2) on NG2+ cells . The ECM is composed of several molecules which can influence OPC function  seem to show some benefit in secondary progressive MS . Similarly, the chemokine receptor CX3CR1—enriched in microglia—is critical for myelin debris clearance following cuprizone-induced demyelination (Lampron et al., Why myelin debris clearance declines are likely multifaceted? Recently, a CRISPR-Cas9 knockout screen identified CD22 as a negative regulator of microglia phagocytosis that increases with age (Pluvinage et al., Peripheral changes during aging can also impart effects onto innate immune cells such as microglia. For example, peripheral blood from aged mice is sufficient to lower the levels of type II interferon in the choroid plexus, which is linked to cognitive decline, a common symptom observed in MS patients (Franklin et al., Remyelination involves a complex interplay between the oligodendrocyte lineage cells that produce the myelin, the astrocytes and the other innate immune cells that regulate the microenvironment of remyelination. Microglia and CNS-infiltrating macrophages regulate the lesion environment and make it suitable for remyelination by activities such as the removal of inhibitory myelin debris. Astrocytes secrete growth factors and provide key lipids species, but also produce inhibitory ECM molecules and the Notch ligand Jagged. OPCs are the key source of remyelinating oligodendrocytes but are now recognized to also regulate angiogenesis, respond to injury and secrete cytokines suggesting they may also be a brain resident immune cell. Given that boosting remyelination spares venerable axons (Mei et al., CB, KR, GD and JP drafted and reviewed this manuscript. MH and CB constructed the figures. JP supervised the drafting of this manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


A literature review on ultrathin films of cellulose is presented. The review focuses on different deposition methods of the films—all the way from simple monocomponent films to more elaborate multicomponent structures—and the use of the film structures in the vast realm of materials science. The common approach of utilizing cellulose thin films as experimental models is therefore omitted. The reader will find that modern usage of cellulose thin films constitutes an exciting emerging area within materials science and it goes far beyond the traditional usage of the films as model systems. Ultrathin films (thickness <100 nm) represent an important category of modern materials. Constant advances in deposition techniques as well as in micro and nanoscale patterning have significantly contributed in developing the functionality of thin films. Particularly in materials technology, ultrathin films play a major role in products like solar cells, sensors, and displays. It is obvious that the present research prospects reach far and wide.Cellulose—the polysaccharide that is responsible for the structural scaffold of all plant fibers , consisting exclusively of linearly arranged anhydroglucose units . It is tThe sparse solubility of cellulose  with LiCl . Probably the most popular solvent has been NMMO/water or NMMO/DMSO which was introduced by the Wågberg group already in the early 2000s . Depending on its degree of substitution (DS), TMSC dissolves in many common organic solvents  study  Raman spectroscopy also favored the two phase kinetic model  which is widely used for manufacturing viscose fibers and films and therefore a large scale commodity. Thin films were prepared by spin coating CX on silicon and gold substrates followed by regeneration using HCl vapor  became a ubiquitous commodity in research laboratories worldwide  .Nanosized cellulose—by and large—comes in two main types: cellulose nanocrystals (CNCs) and cellulose nanofibers (CNFs)  deposition is another means of film casting able to utilize charge on the CNC surface. Alternating layers of sulfated CNCs and cationic polyallylamine were introduced by Cranston and Gray . Soon afAn account of all-cellulose films was presented by Niinivaara et al.  who spinBesides spin coating, the accounts on alternative deposition techniques for CNC thin films are scattered. LB or LS deposition is not feasible for colloidal nanocellulose under the general setup of an aqueous subphase because nanocellulose dispersions themselves are virtually always dispersed in water. However, the Rojas group has reported LS films of CNCs by utilizing a cationic surfactant, dioctadecyldimethylammonium bromide (DODA) which easily forms a monolayer on water and complexes with CNCs that have been dispersed in the subphase  or threads (CNFs), its orientation has been a pressing question, especially when used in composites as reinforcing materials. Orientation of CNCs in an ultrathin film would make a good precedent for orienting nanocellulose in larger matrices. In this vein, Hoeger et al.  investigAlthough CNFs as materials were introduced in the early 1980s  concentrations , as demonstrated for highly charged TEMPO-oxidized CNFs by Wilson et al. . The worUnlike the vast influence imparted by nanocellulose on materials science  can be easily accomplished. The low non-specific protein adsorption on cellulosic surfaces originates from the high degree of solvation of the cellulose molecules inside the thin film structures, thereby impeding the deposition of proteins having a large amount of water inside their structure as well. In order to realize a non-specific cellulose-protein interaction, water needs to be removed from the system, which is entropically unfavorable. In order to test surfaces toward non-specific protein adsorption, BSA is often used as a probe. It is well known that on neat cellulose thin films , the amount of deposited BSA is very low under various conditions  with cellulose  esters to form a more stable intermediate with higher efficiencies in binding, particularly with NH2-rich molecules. One of the first attempts to exploit these concepts on regenerated cellulose thin films was carried out by Orelma et al. who chemically immobilized avidin on the surfaces using 1-ethyl-3-[3-dimethylaminopropyl]-carbodiimide (EDC) combined with NHS, i.e., so-called EDC/NHS chemistry   have been created in a TMSC matrix. This was exploited in optoelectronic devices  with good performance and a photocurrent efficiency of ca. 1% . They observed that depending on the activation procedure the rate of crystallization can be controlled  of ZnO (Jin et al., On a different note, the Peinemann group published a series of papers on thin film membranes based on cellulose thin films (Puspasari et al., Overall, utilizing the inherent structure of cellulose within the thin films for materials applications is an approach still in its infancy. There are no commercial applications at present but the current scientific literature gives indications as to where some of the business-related interests may lie in the near future. For example, clear proof-of-concept exists of functioning photoresists from cellulose and its derivatives. Cellulose based photoresist overcome several issues of conventional photoresists. For the preparation of photoresists, environmentally harmful and often toxic solvents such as chlorobenzene are used while the photoresist materials consist of persistent polymers that contribute to plastic pollution. Instead, biodegradable cellulose derivatives proposed so far for phototresists can be processed from ecosolvents such as ethyl acetate. European legislation will further push the replacement of conventional, non-biodegradable, non-renewable synthetic polymers by biobased greener solutions in the next decade and this will certainly affect photoresists as well. Still, further developments are required to commercialize the technology into products, involving adhesion on various substrates, efficiency, durability and resistance in harsh environments for instance.Similar issues abound with membrane technology: organic solvents are generally required for preparation of the present commercial membranes and this could be omitted particularly if water-based nanocellulose dispersions are used for membrane manufacture. Ultimately, however, the performance of cellulose thin films will dictate their pliability in relevant applications. Concern about the dimensional stability of cellulose films under water and humidity, for example, is one of the factors that could limit its usage.However, the specific response of cellulose to water and humidity may be seen as an advantageous issue from materials perspective. Recent work on fundamental understanding of the cellulose-water interactions within ultrathin films underline this relationship (Kontturi et al., Biomedical applications of cellulose based thin films have been shown to have a huge potential as well. Here, several proof of concepts from various groups do exist that propose the detection of biomolecules such as DNA and proteins. Proteins seem to have the largest potential for commercialization in the field of enzyme immobilization and antibody detection. Antibody detection is a crucial point when it comes to blood typing and it was already shown that IgG can be effectively detected on such films. Here the combination of patterned cellulose thin films for in combination with microfluidics could be an opportunity to develop new products.This review has shown how the focus on cellulose films has recently shifted to another direction. Ultrathin films of cellulose were previously perceived mainly as tools for experimental modeling, by applying them as model films to probe and interpret fundamental physico-chemical phenomena occurring with cellulose-based materials. Although the modeling aspect is still important, utilization of these films in materials applications has become an equally important activity. This aspect is still in its infancy, but we see a significant potential in the approach where the functional features of cellulose in its native environment could be utilized for its applications within ultrathin films.EK wrote the Introduction and Conclusions as well as the section on Film Deposition. SS was largely responsive for the section on Materials Applications.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Resident tissue macrophages (RTMs) are cells with a high functional plasticity assuming pleiotropic roles in their tissue of residence, from clearance of dead cells and metabolic sensing in steady state to cytokine production and tissue repair during inflammation. The liver has long been considered as only populated by Kupffer cells (KCs), a macrophage population assumed to be in charge of all of these functions. However, we know now that KCs are not the only macrophage population in the liver, that recently was shown to contain also capsular macrophages, monocyte-derived macrophages as well as recruited peritoneal macrophages inherited from previous inflammatory events. These macrophages exhibit different origins, time of establishing residence and locations in the liver, with both ontogenical and environmental factors shaping their identity and functions. Furthermore, liver macrophages reside in a complex environment with a pronounced metabolic zonation. Here, we briefly discuss how these intrinsic and extrinsic factors influence macrophage biology and liver physiology in general. We notably focus on how the recent advances of single cell transcriptomic approaches are changing our understanding of liver macrophages and diseases. The liver is known to assume a large repertoire of diverse functions such as detoxication of numerous metabolites, synthesis of essential proteins, or recycling of iron-containing red blood cells . Such veMacrophages represent by far the most abundant immune cells in the liver. Hepatic macrophages are still often referred as Kupffer cells (KCs) that indeed represent the major fraction of liver macrophages. KCs were first described more than one century ago by Kupffer who initially described them as endothelial cells, components of liver vascular walls . Then, aBesides of the embryonically-derived KC population that represents the vast majority of liver macrophages, at least at steady-state, another population of macrophages residing in the hepatic capsule has been recently described . These lIn addition to these two main macrophage populations, the liver may contain a variable amount of recruited blood monocyte-derived macrophages. Indeed, in several inflammatory conditions and notably when KC depletion occurs, replacement by monocyte-derived macrophages can be observed , 15. Somhi GATA6+ peritoneal macrophages migrated to the site of injury. This non-vascular recruitment was mediated by ATP released from dead cells acting as a damage-associated molecular pattern and involved also the Hyaluronan-CD44 interaction. Recruited peritoneal macrophages are responsible for disassembling the necrotic nuclei of dead cells and authors have shown that depletion of peritoneal macrophages significantly delayed the wound healing process . These pericytes are localized in the space of Disse between hepatocytes and endothelial cells . They arAltogether, even if at a macroscopic level, liver lobules appear homogeneous with a relatively simple tridimensional architecture, they hide a complexity shaped by many factors that should be taken into account to decipher liver macrophage biology and their potential heterogeneity.So far, most of the studies dealing with liver macrophages have used very few but specific markers such as CD45 (pan-leukocyte marker), CD11b (pan-myeloid marker) and F4/80 (pan-macrophage marker in mice) to study macrophages, most often assimilated to KCs. While it has been very useful to extend our knowledge on KC biology, this conventional approach consisting in defining populations of interest based on the expression of limited markers by flow cytometry appears more and more outdated nowadays. Discoveries of the distinct ontogeny of KCs and monocyte-derived macrophages and the complexity of the liver niche have challenged the view of liver macrophages as a uniform F4/80+ cell population.The very recent burst in single cell transcriptomics is indeed profoundly reshaping our approaches to solve key questions in immunology . This tecdkn1a upregulation. This radio-resistance property is lost when native KCs are replaced by their monocyte-derived counterparts, showing clearly that ontogeny contribute to macrophage functional heterogeneity  . AuthorsHerein, we discussed liver macrophage heterogeneity and how the most recent advances in single cell transcriptomics could be used to decipher liver macrophage biology. Clearly, last years of research have revealed an unexpected heterogeneity of liver macrophages, both at ontogeny and environmental levels. We now need to take into account this diversity in future studies focusing on liver diseases, and the use of the most recent and still evolving technologies such as the single cell transcriptomics will be crucial for this.All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The importance of cardiovascular health in relation to pregnancy outcome is increasingly acknowledged. Women who develop certain pregnancy complications, in particular preeclampsia, are at higher risk for future cardiovascular disease. Independent of its outcome, pregnancy requires a substantial adaptive response of the maternal cardiovascular system. In the Hemodynamic Adaptation to Pregnancy and Placenta-related Outcome (HAPPO) study, we aim to examine longitudinal maternal haemodynamic adaptation to pregnancy from the preconception period onwards. We hypothesise that women who will develop adverse pregnancy outcomes have impaired cardiovascular health before conception, leading to haemodynamic maladaptation to pregnancy and diminished uteroplacental vascular development.In this prospective cohort study embedded in the Rotterdam periconception cohort, 200 women with a history of placenta-related pregnancy complications (high-risk group) and 100 women with an uncomplicated obstetric history (low-risk group) will be included. At five moments , women will undergo an extensive examination of the macrocirculatory and microcirculatory system and uteroplacental vascular development. The main outcome measures are differences in maternal haemodynamic adaptation to pregnancy between women with and without placenta-related pregnancy complications. In a multivariate linear mixed model, the relationship between maternal haemodynamic adaptive parameters, (utero)placental vascularisation indices and clinical outcomes  will be studied. Subgroup analysis will be performed to study baseline and trajectory differences between high-risk and low-risk women, independent of subsequent pregnancy outcome.This study protocol was approved by the Medical Ethics Committee of the Erasmus MC, Rotterdam, the Netherlands (MEC 2018–150). Results will be disseminated to the medical community by publications in peer-reviewed journals and presentations at scientific congresses. Also, patient associations will be informed and the public will be informed by dissemination through  media.NL7394  An integrated approach consisting of parallel non-invasive techniques assessing multiple haemodynamic parameters will compile extensive haemodynamic profiles of participants.Haemodynamic adaptation to pregnancy will be measured prospectively from the preconception period onwards to 3 months postdelivery, thereby covering an important window in the reproductive timespan.Simultaneous assessment of maternal haemodynamic adaptation to pregnancy and uteroplacental vascular development will provide new insights into the interaction between both mechanisms and the pathophysiology of pregnancy complications.Since only multiparous women will be included, conclusions about whether maternal haemodynamic maladaptation is a cause or consequence of previous pregnancy outcome cannot be provided.Placenta-related pregnancy complications (PPC), such as preeclampsia and/or fetal growth restriction (FGR), occur in approximately 10%–15% of all pregnancies.Pregnancy requires a physiological adaptive response of the maternal cardiovascular system with a substantial rise in plasma volume and cardiac output (CO).Moreover, in women with complicated pregnancies, endothelial dysfunction and increased vascular stiffness are also more common.Together, this all supports the hypothesis that in women who develop PPC an increased risk for cardiovascular disease is unmasked by the stressed state of pregnancy.The adequate establishment of the maternal–fetal vascular interface early in gestation is essential for a healthy pregnancy outcome. Among others, this requires adaptation of the uterine spiral arteries into remodelled vessels to create a low-resistance uteroplacental circulation.We hypothesise that impaired maternal cardiovascular health leads to haemodynamic maladaptation to pregnancy, diminished uteroplacental vascular development and subsequent pregnancy complications . The oveTo assess haemodynamic parameters before, during and after pregnancy to gain insight in (patho)physiological maternal haemodynamic adaptation mechanisms.To assess longitudinal utero vasculature development using state-of-the-art three-dimensional ultrasound and Virtual Reality techniques combined with in vivo examination of placental bed biopsies.To construct a multivariate model to study the associations between maternal haemodynamic adaptation to pregnancy, utero vascular development and PPC.To study differences in baseline haemodynamic parameters and haemodynamic adaptation to pregnancy between women with and without a history of PPC.The HAPPO study is a single-centre prospective cohort study, embedded in the ongoing Rotterdam periconception cohort, performed at the department of Obstetrics and Gynaecology of the Erasmus MC, University Medical Centre Rotterdam, the Netherlands.et al.We aim to include 200 women with a history of pregnancy complications . Preeclampsia is defined according to the American College of Obstetricians and Gynecologists (ACOG) criteria and FGR is defined conform the Delphi consensus definition described by Gordijn or the previous pregnancy was uncomplicated. Women who are unable or unwilling to provide informed consent will be excluded from participation. Also, if a woman is currently breastfeeding, she will be excluded from participation since this may influence her vascular condition. The inclusion and exclusion criteria are shown in Also, 100 women after uncomplicated pregnancies and (term) deliveries will be included as a low-risk control group. Women are eligible to participate in this study when they have a minimum age of 18 years, the delivery date of their last pregnancy was more than 1 year ago, they have a current wish to become pregnant and the previous pregnancy was either complicated by PPC An overview of all study procedures is provided in Participants of the study will receive a total of three questionnaires: (1) after the preconceptional study visit, participants will receive an online questionnaire on demographic characteristics, medical conditions and lifestyle; (2) preceding the study visit in the first trimester of pregnancy, participants will receive an online questionnaire to address possible changes in medical conditions and lifestyle; (3) after delivery, participants will receive an online follow-up questionnaire on pregnancy and birth outcomes . During the postdelivery visit, the participants will, among others, be asked about current breastfeeding and smoking habits.At each visit, a woman’s height, weight, hip and waist circumference will be measured using standardised, validated methods to compose the classical cardiovascular risk factors body mass index and waist-to-hip ratio.Two blood samples (8.5 mL serum tube and 10 mL EDTA tube) will be obtained at the preconception visit, the first-trimester and third-trimester visit and the postdelivery visit. All samples will be centrifuged and divided into aliquots of serum, plasma, whole blood and buffy coat and then stored in a −80°C freezer at the trial laboratory at our own facility for future assessment of cardiovascular markers  and markers of placental function .Transthoracic echocardiography has become the preferential method for safe, non-invasive cardiac assessment in pregnant women.One of the first manifestations of an impaired cardiovascular risk profile is endothelial dysfunction of small vessels (microcirculation). Postocclusion reactive hyperaemia will be used in this study to assess the endothelial function of the peripheral microvasculature. We use devices from Moor instruments  to measure microvascular blood flow reactivity following arterial occlusion by using a non-invasive laser Doppler flowmetry (LDF) technique. In resting position, after 3 min of arterial occlusion by a cuff surrounding the upper left arm, the reperfusion of microvascular blood in the skin is measured by the LDF skin probe which is attached to the lower arm. The peak response after the release of the occlusion and the time to recover to baseline reflect endothelial regulated microvascular function.A cuff-based oscillometric device  will be used to assess central and peripheral blood pressure, pulse wave velocity and augmentation index (AIx).To assess the haemodynamic response during physical stress, CO will also be measured using signal morphology impedance cardiography (SM-ICG) using PhysioFlow PF07 Enduro . The exercise during which the SM-ICG will be performed is a cardiopulmonary exercise test on a cycle ergometer with a RAMP protocol to 70% of the estimated maximum heart rate according to the Tanaka formula.Studies using conventional two-dimensional Doppler ultrasound have shown that increased vascular resistance of the uterine arteries is associated with an increased risk for subsequent development of PPC.With respect to the risks of performing ultrasound examinations in the first trimester using 3D power Doppler, recommendations as set by the International Society of Ultrasound in Obstetrics and Gynaecology (ISUOG) are followed for the purpose of this study . Transvaginal scanning time (preconception and first trimester) and transabdominal scanning time (second and third trimester) will be kept as short as possible according to the as low as reasonably achievable (ALARA) principles.At the first trimester visit, additional parameters of embryonic growth will be measured, that is, crown-rump length and embryonic volume. The obtained 3D datasets will be visualised in the Barco I-Space or desktop version and offline volume measurements will be performed.At the second and third trimester visit, additional parameters of fetal growth  and pulsed wave Doppler measurements of the umbilical artery and median cerebral artery will be performed by transabdominal ultrasound.At delivery, histological samples of in vivo placental vasculature will be harvested. The placenta will be collected according to standard procedures and four samples of approximately 3 cm × 3 cm are then fixated in 4% paraformaldehyde for storage. The fixated placenta biopsies will be processed and studied as done in routine placental pathology examination.In a subgroup of women who undergo a caesarean section (estimated 30% of our study population) at the Erasmus MC, four punch biopsies of the placental bed will be collected, which consist of decidual and myometrial tissue and contain spiral arteries.The primary outcome measure of the HAPPO study is the difference in maternal haemodynamic adaptation to pregnancy, expressed as the trajectory of CO assessed by echocardiography before, during and after pregnancy, between women who do and do not develop PPC. CO is one of the most frequently described haemodynamic parameters in pregnancy, alongside with blood pressure. By using transthoracic echocardiography, as a validated and safe method to measure CO, we will obtain consecutive and reliable information on maternal haemodynamic adaptation to pregnancy.The secondary outcome measures are differences in additional indices of maternal haemodynamic adaptation to pregnancy and utero vascular development between women who do and do not develop PPC. This includes differences in weight gain during pregnancy, blood pressure, systemic vascular resistance, left ventricular diastolic function, endothelial regulated microvascular function, pulse wave analysis, haemodynamic response to exercise, placental vascular indices, fetal growth parameters (including Doppler measurements) and placenta/placental bed pathology examinations.Since we aim to investigate multiple haemodynamic parameters and literature on this subject is scarce, sample sizes are difficult to calculate. There are a few studies that describe the longitudinal trajectory of CO from preconception until after delivery in uncomplicated pregnancies and a couple of cross-sectional studies that investigated CO in either the first of third trimester of pregnancy.Based on literature, we expect a 25% recurrence rate of PPC in the high-risk group and a 2% incidence of PPC in the low-risk group.Multiparous women who wish to become pregnant who are actively contemplating pregnancy are assumed to have an approximately 85% chance to become pregnant again within 1 year.Patient characteristics, medical conditions and lifestyle data will be presented as means (SD) or numbers (percentage) and compared using t-tests and χ² tests. Haemodynamic parameters will be presented as means (SD) at each time point and compared between groups using t-tests.Multivariate linear mixed models will be used to analyse the trajectories of maternal haemodynamic parameters over time. We will test whether there is a difference between women who do and do not develop PPC by comparing each trajectory using a likelihood ratio test.Subgroup analyses will be performed to study (baseline and trajectory) differences between high-risk and low-risk women, independent of subsequent pregnancy outcome. Bonferroni corrections will be applied to control for multiplicity of secondary outcome parameters.In case of significant differences in patient characteristics between groups, regression analyses will be adjusted for these characteristics.The study will be performed according to written informed consent procedures which ensures personal data protection and confidentiality. Data will be deidentified (pseudoanonymised) in order to guarantee the privacy of the participants, and the key to the code will be safeguarded by the investigators and will remain on site with the local research team.All research data will be retained and stored in the HAPPO-database at a secured server at the research institution and it will remain there for at least 15 years after completion of the project. All imaging data will be stored and handled in the same way and according to the same rules and regulations as other patient-related imaging data at Erasmus MC.Monitoring of data and safety will be performed once a year, according to local protocol for research with negligible risk.http://www.hellp.nl) and the Preeclampsia Foundation (http://www.preeclampsia.org)) were informed and are involved in promoting the HAPPO study.Patient and public were not involved in the design of the study. However, at the earliest moment of recruitment, two large patient associations for women who experienced PE and/or HELLP syndrome  and in accordance with relevant national guidelines, regulations and Acts ). The study protocol was approved by the Medical Ethics Committee of the Erasmus MC (MEC-2018–150). The study is registered in the Dutch Trial Registry (NL7394).The HAPPO study team invites colleagues who are interested in collaboration or data sharing to contact the principal investigators at HAPPO.studie@erasmusmc.nl.Results of the study will be disseminated to healthcare professionals and to scientific and industrial peers through events, professional organisations and publications in peer-reviewed (preferably open-access) journals as well as presentations at scientific congresses. Also, results will be communicated to women who wish to become pregnant and women who experienced pregnancy complications by informing patient associations and by dissemination via  media.The results of this study will provide more understanding of longitudinal maternal haemodynamic adaptation to pregnancy in relation to utero vascular development and its associations with subsequent pregnancy outcome. If we succeed to establish an association between maternal haemodynamic adaptation to pregnancy and subsequent pregnancy outcome, a larger cohort study in nulliparous women will be needed to investigate causal relationships between haemodynamic adaptation to pregnancy and placenta-related outcomes. Hereby, starting points for future possibilities to optimise cardiovascular and placental health by interventions in clinical validation studies will arise. This will enable the development of more accurate, personalised prevention and treatment strategies for women with high-risk pregnancies, from the preconception period onwards. Thereby, we aim to reduce subsequent maternal and perinatal morbidity and mortality in the future, eventually leading to life-long reduced risks and costs of cardiovascular diseases.