Veillonella is commonly enriched in AIH, PBC, and PSC. Apart from gut microbiome, the oral and bile microbiome seem to be associated with these diseases as well. The functional analysis of metagenomics suggests that metabolic pathways changed in the gut microbiome of the patients. Microbial metabolites, including short-chain fatty acids (SCFAs) and microbial bile acid metabolites, have been shown to modulate innate immunity, adaptive immunity, and inflammation. Taken together, the evidence of host–microbiome interactions and in-depth mechanistic studies needs further accumulation, which will offer more possibilities to clarify the mechanisms of AILDs and provide potential molecular targets for the prevention and treatment in the future.Recent studies have identified the critical role of microbiota in the pathophysiology of autoimmune liver diseases (AILDs), including autoimmune hepatitis (AIH), primary biliary cholangitis (PBC), and primary sclerosing cholangitis (PSC). Metagenomic studies reveal significant decrease of gut bacterial diversity in AILDs. Although profiles of metagenomic vary widely,  Autoimmune liver diseases (AILDs) are chronic inflammatory conditions of the liver, including autoimmune hepatitis (AIH), primary biliary cholangitis (PBC), and primary sclerosing cholangitis (PSC) . Poor unSince the microbiota plays an important role in the development of the innate and adaptive immune system , furtherMetagenomics is a powerful tool that is helpful for the analysis of microbial heterogenicity. It mainly includes two sequencing strategies: amplicon sequencing, most often amplifying portions of the hypervariable regions of 16S rRNA; or shotgun sequencing, which sequences all given genomic DNA from a sample . The shoStudies of the microbiota often focus on the bacterial diversity in the feces of the patients. Collectively, alpha diversity of fecal microbial showed a downward trend in most AILDs patients, which was shown in Veillonella, Klebsiella, Streptococcus, and Lactobacillus in AIH compared to HC. Moreover, they created a model including Veillonella, Lactobacillus, Oscillospira, and Clostridiales to distinguish AIH from controls  . Thus, ipatients . Coprobahout IBD . Howevers of PSC . Until ns of PSC . Howevers of PSC .Enterococcus spp., Prevotella spp., Staphylococcus spp., Lawsonella spp., Veillonella dispar, and Cutibacterium was observed  and vitamins to secondary bile acids and neurotransmitters . Colonicepatitis . FurtherVeillonella and Klebsiella have been proved to be negatively correlated with the level of secondary bile acids in serum  as the susceptible gene of PSC . Specifically, the NOD.c3c4, which could spontaneously develop biliary inflammation, is a model of the human biliary disease primary biliary cirrhosis (It is currently believed that chronic bacterial infection might play a part in the initiation or development of the autoimmune status in patients with AILDs . Many stesponses . Zhang eirrhosis . Biliaryirrhosis . NakamotE. coli is molecularly similar to human PDC-E2, the immunodominant target of AMAs in PBC (Novosphingobium aromaticivorans were more similar with human PDC-E2 than that of E. coli (N. aromaticivorans hasn’t been detected in the liver of patients with PBC (N. aromaticivorans is present in approximately 25% of fecal samples from patients and controls (Lactobacillus delbrueckii and Mycobacterium gordonae might also induce loss of tolerance to human mitochondrial proteins in genetically susceptible individuals due to molecular mimicry and immunological cross-reactivity (Yersinia enterocolitica were present in PBC patients (Molecular mimicry between immunodominant epitopes of the pathogen and self-peptides has been hypothesized to be the key event leading to the disease. It may provide clues to explain the relationship between microbes and disease. Investigations have detected that PDC-E2 of s in PBC . Meanwhi E. coli . Althougwith PBC , a studycontrols . Lactobaactivity . Recent patients . Interespatients . It is wIncreasing evidence has highlighted the crucial role of microbiome in AILDs. Research showed various changes of microbiome in metagenomic and metabolomic analyses. Although the studies so far didn’t clearly demonstrate the causation between microbiome and AILDs, they found the associated microbiome and even possible role in the pathogenesis providing direction for further study in the pathogenesis of AILDs. Moreover, mutual authentication among metagenomic and metabolomic of the microbiome could help us to understand the detailed and correct role of microbiota in AILDs. Nevertheless, the integrated analysis of gut microbiome and metabolite is lacking because of limited data of metagenomics and metabolomics in AILDs. With the application of metagenomics and metabolomics, it is possible to identify new microbial diagnostic markers in the early diagnosis and novel treatments of AILDs. The role of the microbiome, not only bacteria, in the mechanisms of AILDs needs more comprehensive and in-depth research to explore in the future.BW and LZ guided the outline and carried out manuscript editing. YZ and YR collected data and drafted the manuscript. HZ performed manuscript review. Each of the co-authors has approved the final draft submitted. All authors contributed to the article and approved the submitted version.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.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Robotic-assisted bronchoscopy is one of the newest additions to clinicians’ armamentarium for the biopsy of peripheral pulmonary lesions in light of the suboptimal yields and sensitivities of conventional bronchoscopic platforms. In this article, we review the existing literature pertaining to the feasibility as well as sensitivity of available robotic-assisted bronchoscopic platforms. The implementation of routine lung cancer screening guidelines, in addition to improvements in imaging modalities, has led to the increased detection of lung nodules. In recent decades, lung cancer has remained one of the leading causes of cancer-associated mortality , with tiTwo common modalities to sample lung lesions include image-guided trans-thoracic needle biopsy as well as endoscopic sampling. The majority of lung nodules are located in the lung periphery ,3,4 and Conversely, endoscopic approaches have traditionally been limited to large and/or central lung nodules. While endoscopy allows for concurrent airway evaluation and mediastinal lymph node staging, inferior diagnostic yields and limited access to smaller PPLs certainly leave room for improvement.Technologic innovations in recent years have led to improved imaging and navigation that has increased the accuracy, reach, and diagnostic yield of endoscopic sampling. The most recent of advances, robotic bronchoscopy, follows a national and global trend of the proliferation of robotic-assisted surgeries for thoracic ,8 and exThe flexible bronchoscope was developed by Dr. Shigeto Ikeda, a Japanese thoracic surgeon in the 1960s. Further revisions to the initial concept added tip angulation features in 1970 and video capabilities in 1987 . The 199Navigational advances came in the form of electromagnetic navigational bronchoscopy (ENB), designed to guide operators to predetermined locations in the bronchial tree based on multi-planar computed tomography (CT) and real-time bronchoscope localization . AlthougAll said, sensitivities for CT-guided biopsy are as high as 92.1%  in the lNumerous factors worsen the sensitivities of the bronchoscopic platform, including CTBD, misconstruing atelectasis for solid pulmonary nodules on rEBUS, as well as stiff biopsy instruments that impair the maneuverability of bronchoscopes. Due to these limitations, the robotic bronchoscopic platform was developed in an attempt to improve upon existing technology and optimizations.Currently, two robotic platforms have gained approval from the Food and Drug Administration, namely the Monarch platform  and the Ion Endoluminal System . Key findings are discussed below and summarized in The Monarch system  consistsBefore the procedure, a CT is obtained with which virtual 3D reconstruction is generated. Potential pathways are mapped  and uploaded to the robot. Patients lay within an electromagnetic field, allowing for the use of ENB for navigation purposes. The operator can control the bronchoscope with a handheld controller, not unlike that of a gaming controller. Once the target is reached, the location can be verified with fluoroscopy or rEBUS before the entire bronchoscope system is locked to maintain its static position while biopsies are obtained.The Monarch system was first assessed in cadavers in the REACH study of 2018 . MonarchThe ACCESS Study  assessedThe first human feasibility study was performed by Rojas-Solano et al. in 2018 . The stuA retrospective post-market study by Chaddha et al. analyzed 167 lesions in 165 patients . The aveThe BENEFIT study was the first prospective multi-centered study with the Monarch system, consisting of 54 patients across 5 centers . Median The Ion system  has a brSimilar to Monarch, the Ion system uses a pre-procedure CT Chest that is uploaded to their PlanPoint program for 3D reconstruction and target pathway planning. Rather than ENB, Ion uses shape-sensing catheter technology as its modality for localization. The operator is able to navigate the bronchoscope using a trackball controller. At the start of the procedure, the vision probe is fed through the shape-sensing catheter during registration and navigation to target lesions. Once the target is reached, localization can be similarly confirmed with fluoroscopy or rEBUS before the bronchoscope is fixed in place. Notably, the vision probe needs to be retracted from the catheter prior to the passing of biopsy equipment and tissue acquisition under fluoroscopy visualization.The first feasibility study for Ion in human subjects included 29 consecutive cases in a single center [In a cadaveric comparative study of the Ion system vs. ENB vs. rEBUS , implantRecently published was a retrospective observational single-center study at a high-volume academic institution . This stIn addition to the localization of peripheral lung lesions, the robotic systems may offer another modality with which to apply therapeutic interventions for the minimally invasive treatment of lung cancer in the near future. Studies will soon start to examine the use of microwave ablation of early-stage lung cancer in patients who are not surgical candidates as well as local injection therapies. These therapies will also take advantage of cone beam CT technology to ensure appropriate positioning of the catheters. Other therapeutic modalities such as photodynamic therapy, radiofrequency ablation, and YAG laser may also be used in the future as well. Finally, the application of dyes and fiducial markers has also been recently performed for improved lesion localization prior to surgical resection as well as stereotactic radiosurgery. That said, the performance of endoluminal procedures with robotic bronchoscopic systems has yet to be documented in the literature and we await future studies to compare efficacy to existing technologies and procedures.The perceived advantages of the two robotic platforms hinge upon improved tip maneuverability and system stability to enable operators to more easily traverse into peripheral lung fields and obtain biopsy samples of PPLs. Notable drawbacks of robotic bronchoscopy are similar to those noted at the early stages of adoption of other robotic-assisted surgeries, such as increased unit cost as compared to traditional approaches and additional training to OR staff to support robotic platforms. Furthermore, the need to dock/un-dock platforms is thought to lengthen procedure times and response latency to peri-procedural complications. That said, what has been demonstrated in the surgical literature is that these disadvantages can often be ameliorated with operator/staff fluency, seen especially at high-volume centers ,35.The existing literature, by way of single and multi-centered feasibility studies, thus far has demonstrated promising results with a comparable safety profile. Studies currently in the pipeline include the multi-centered, prospective, observational TARGET trial designed to assess the clinical safety and diagnostic accuracy of the Monarch platform (NCT04182815), as well as the prospective, multi-center PRECIsE trial, evaluating the clinical utility and performance of the Ion System (NCT03893539).As the new technology makes its way into hospitals and more single-center experiences are reported ,36, the


Betel quid (BQ) is a package of mixed constituents that is chewed by more than 600 million people worldwide, particularly in Asia. The formulation of BQ depends on a variety of factors but typically includes areca nut, betel leaf, and slaked lime and may or may not contain tobacco. BQ chewing is strongly associated with the development of potentially malignant and malignant diseases of the mouth such as oral submucous fibrosis (OSMF) and oral squamous cell carcinoma (OSCC), respectively. We have shown recently that the constituents of BQ vary geographically and that the capacity to induce disease reflects the distinct chemical composition of the BQ. In this review, we examined the diverse chemical constituents of BQ and their putative role in oral carcinogenesis. Four major areca alkaloids—arecoline, arecaidine, guvacoline and guvacine—together with the polyphenols, were identified as being potentially involved in oral carcinogenesis. Further, we propose that fibroblast senescence, which is induced by certain BQ components, may be a key driver of tumour progression in OSMF and OSCC. Our study emphasizes that the characterization of the detrimental or protective effects of specific BQ ingredients may facilitate the development of targeted BQ formulations to prevent and/or treat potentially malignant oral disorders and oral cancer in BQ users. Betel quid (BQ) chewing is a common habit in many parts of Asia and some Pacific Islands; approximately 600 million people worldwide chew some form of BQ regularly ,2. BQ chDifferent BQ ingredients have distinct effects in cells, tissues and living organisms. The formulation of BQ depends on a variety of factors that may include the cultural background of a country, regional backdrop, personal likings and availability of specific ingredients ,2. BQ isWe believe that it is important to gain more insight into the carcinogenicity of BQ ingredients. Very little is known, for example, about whether there is a correlation between the specific chemical components of BQ and the potential for malignant transformation in the oral cavity. To understand the role of different BQ ingredients in the pathogenesis of fibrosis and cancer, as well as to shed light on the mechanisms by which cellular senescence promotes OSMF and its malignant transformation, a structured search strategy was developed. Eligible articles indexed in MEDLINE/PubMed, Scopus, and Web of Science (WoS) were included. The search strategy used in this narrative review combined the search terms related to three categories, namely: BQ composition (1), malignant transformation of OSMF (2), and cellular senescence in OSMF and OSCC (3). The following search terms were utilized for the search: ‘ or (areca and nut) or betel quid or (slaked and lime) or (betel and leaf) or catechu or (betel and inflorescence) plus (1)  not (lesion or effect or disorder or tea or grapevine or wine or lipase or apple)’ or (2) ‘ or  or  or ’ or (3) ‘(cellular senescence) or (fibroblast senescence) or (Senescence-Associated Secretory Phenotype) and  or ’. The articles screened from this search (in the English language only) were used as the basis for the elaboration of the manuscript. Further grey literature was searched as appropriate.This review aimed to survey the major chemical constituents of BQ and to offer a qualitative assessment of their potential role in oral carcinogenesis. The findings may improve our understanding of the pathophysiology of BQ-associated OSCC and OSMF and may guide future chemopreventive and therapeutic strategies.Areca catechu) tree, typically grown in tropical regions. Alkaloids are one of the many well-known chemical compounds associated with areca nut [Piper betle L.) contains betel oil, which consists of phenolic compounds such as eugenol, chavicol and hydroxychavicol [Different types of BQ contain diverse chemical constituents. Areca nut, also known as betel nut, is the seed of the areca palm , whilst levels of the other alkaloids were unchanged [Guvacine is another prominent constituent of areca nut and BQ. It has been reported that guvacaine is the most abundant of the four main alkaloids (1.39−8.16 mg/g), followed by arecoline (0.64−2.22 mg/g), arecaidine (0.14−1.70 mg/g) and guvacoline (0.17−0.99 mg/g) . In the nchanged . Interesnchanged . These fnchanged  who repoWhilst these studies are important and document the presence of guvacine in BQ, there is no clear data regarding how guvacine relates to oral carcinogenesis.In different areca nut products, Jain et al.  identifiNew alkaloids have been identified in Areca catechu including Acatechu A and B  and Arecm/z arecoline in mass spectra of the saliva of BQ chewers with and without the addition of slaked lime. Tobacco is another popular additive and, in the study by Lord et al. [Additives are popular in many BQ preparations worldwide. Kadi et al.  reportedd et al. , nicotind et al.  who demod et al.  and is nPolyphenols are common constituents of BQ ingredients including areca nut and leaf, husk and stem inflorescence (SI). Phenolic acids have been detected mostly in the husk, leaf and SI, but traces have also been identified in areca nuts from BQ mixtures. In a recent study, flavanols (predominantly catechin and epicatechin) were the most abundant polyphenols detected in Indonesian BQ and concentrations appeared to be higher in nuts than leaf, husk and SI . FlavonoPiper betle leaves [Chavibetol is an allyl benzene found in e leaves . It has Areca catechu L.; the compound structures in this study were determined by comparing their spectroscopic data with that reported in the literature. Other authors [Cao and colleagues  isolated authors  also ideSmoking and alcohol are well-known risk factors of OSCC. However, malignant transformation of OSMF involves additional and potentially different pathways that are specific to OSMF. For example, it is established that the onset of fibrosis is critical in the malignant transformation of OSMF  but the Key triggers of subsequent malignant transformation of OSMF could be the inflammatory milieu and the production of reactive oxygen species (ROS). ROS is produced by both auto-oxidation of areca nut polyphenols in saliva and nitrosation of areca alkaloids. Following an arecoline-dependent increase in IL-6, IL-8 and GRO-α, repeated and continuous exposure to ROS has the potential to cause DNA double strand breaks and other pathogenic mechanisms of DNA damage in oral mucosal cells through oxidative stress . IL-6, IBelow, we summarise the evidence supporting the pro- and anti-carcinogenic effects of individual constituents of BQ mixtures.N-nitrosamine metabolites [N-nitrosoguvacoline, N-nitrosoguvacine, 3-(N-nitrosomethylamino)propionaldehyde and 3-(N-nitrosomethylamino) propionitrile) cause DNA single strand breaks and DNA protein cross-links that, in turn, cause a decrease in the survival of cultured human buccal epithelial cells. In addition, nitrosamines interact with other macromolecules of the cell through oxidative stress, thereby contributing to oral carcinogenesis [N-nitrosomethylamino)propionaldehyde being the most cytotoxic and genotoxic derivative [Arecoline and guvacine are the best-studied compounds detected in areca nut and the relative proportions of arecoline and guvacine vary depending on the part of the areca nut that is sampled, as discussed previously. Early studies concluded that arecoline was the main alkaloid associated with the mutagenic potential of BQ because it induces chromosome breaks and causes other cellular aberrations in vivo . Guvacinabolites . The fouogenesis ,45,46. Hogenesis  originalogenesis . Interesogenesis . Conversrivative ,51,52. Arivative ,54,55.N-nitrosamine metabolites through nitrosation, the induction of ROS and the disruption of collagen breakdown causing increased collagen accumulation, but also are also involved in a variety of other processes such as ulceration from mechanical trauma of the coarse areca nut fibres, stimulation of fibroblast proliferation, activation of the coagulation system and cytotoxicity of oral epithelial cells. All of these factors are likely to contribute to the pathogenesis of OSMF and its malignant transformation [In summary, analysis of the existing literature shows that areca nut constituents not only are associated with the formation of ormation .The association between consumption of slaked lime and OSCC was proposed in seminal epidemiological research from Papua New Guinea . The invSlaked lime also increases the proliferation of fibroblasts and causes phenotypic alterations in these cells , a phenoFive main polyphenols are commonly extracted from betel quid mixtures, namely catechins, flavonoids, flavan-3:4-diols, leucocyanidins and hexadroxyflavans . In the Research from Cirillo’s group has shown that the husk contains the widest range of polyphenols that are associated with high antioxidant capacity , suggestIn summary, the majority of the effects attributed to polyphenols are linked to their antioxidant and anti-inflammatory properties which, in turn, impact molecular events and signalling pathways associated with cell survival, proliferation, differentiation, migration, angiogenesis, detoxification enzymes and immune responses . UnfortuAs such, the anti- or pro-oxidant potential of polyphenols depends greatly on the chemical nature, concentration and the micro-environmental conditions  in which they are consumed . These fCatechins, which are abundant in the areca nut , have beChavibetol in piper betel leaf acts as a bioactive molecule because it has anti-oxidant properties that protect against photosensitization-mediated lipid peroxidation in rat liver mitochondria ,70. ChavFlavonoids include flavonols, flavones, isoflavones and anthocyanidins, amongst other compounds. They are present in foods of plant origin . The potAreca catechu are closely related to cancer development [For decades, it has been thought that tannins in elopment . Later, elopment  that tanelopment . Consistelopment . Furtherelopment  showed telopment . Hence, Various genetic and molecular mechanisms impact the development of OSMF and malignant transformation. Alterations occur in both oral keratinocytes and/or fibroblasts and lead to changes in the cell cycle, DNA and keratin formation, cell proliferation and survival, angiogenesis and fibrosis, epithelial-mesenchymal transition (EMT) and tissue hypoxia (reviewed in ). AlthouEarly OSMF is characterised by epithelial atrophy, juxta-epithelial inflammation and the sub-epithelial connective tissues become avascular and thickened leading to collagen accumulation. In advanced disease, the fibrosis extends into the deeper tissues and is associated with an increased inflammatory cell infiltrate . Many ofSeminal work by Parkinson and colleagues has shown that OSMF is characterised by the presence of senescent fibroblasts in the subepithelial mesenchyme of OSMF patients ,93. SpecOverall, these data suggest that fibroblast senescence is central to the development of OSMF. However, do senescent fibroblasts promote malignancy?We have shown that senescent fibroblasts share common characteristics with activated fibroblasts/myofibroblasts  and can In our model, the main initiating event in the pathogenesis of OSMF is the production of ROS by the BQ constituents. This, in turn, leads to oxidative stress and senescence in keratinocytes, fibroblasts and endothelial cells. Senescence in keratinocytes induces epithelial atrophy, whilst senescence in endothelial cells accounts for a decrease in vascularity and leads to the development of a hypoxic state. In fibroblasts, the mechanism is associated with irreparable DNA damage and development of the SASP. There is then a positive feedback because the SASP reinforces ROS production and the cycle is repeated and amplified . FinallyThis model would be amenable to several preventive and therapeutic strategies for OSMF including the use of senostatics, senolytics and anti-oxidants. Inhibiting SASP production/function (senostatics) has been the goal of past research from an early stage, but recent data have shown that the SASP is cell-type specific making this approach less practical. Some potential drug targets, particularly those associated with inhibition of the inflammasome and plasminogen activator inhibitor-1 , can alsSenolytics are a class of drugs that selectively target senescent cells for destruction. These drugs include BCL-XL inhibitors, finestin, ouabain, quecertin and dasatinib . SenolytDrugs or natural compounds with antioxidant properties such as aloe vera, curcumin and lycopene have been used extensively in the treatment of OSMF, unfortunately with inconsistent results . A recenIn patients with OSMF, progression to cancer is thought to be related to smoking and alcohol consumption  but, alsDespite its known potential to cause disease, betel quid chewing plays an important role in religious practices, cultural rituals and social customs throughout Asia and the Pacific rim. The eradication of this habit, therefore, may not be realistically achievable. Rather than impose a change in culture and way of life, it might be possible to identify mixtures that have a greater or lesser risk of OSMF, and, therefore,, by modulating fibrosis through betel quid regulation, malignant transformation in OSMF may be reduced together with the morbidity of OSMF.Our unpublished data suggest that chewing betel quid mixtures with the addition of betel inflorescence (BI) increases the risk of OSMF. Conversely, the use of betel leaf in the mixture is likely to be protective. For all of these reasons, we believe that a better understanding of the effects of each BQ ingredient would help develop a sustainable chemopreventive strategy for this disease. The present review highlights our lack of knowledge in the field and the urgent need for further research. In the future, therefore, we suggest that further work should involve a broad range of geographic regions where different betel quid constituents are consumed. We believe that the use of consistent and contemporary methodology to measure carcinogenic risk is essential. In this way, we would be able to address the impact of changes in the concentrations of different constituents and the consequences of areca nut maturity and processing.Based on our own findings and those of others, we propose a pathogenic loop involving a cross-talk between keratinocytes and fibroblasts that fuels the development of OSMF and, possibly, its malignant transformation. Specifically, the consequence of cellular senescence is the generation of the SASP, the production of ROS and the induction of DNA double strand breaks in keratinocytes. Only when the keratinocytes escape from senescence does malignant transformation ensue, but the myofibroblasts persist because they evade immune clearance. We, therefore, suggest the use of antioxidants and senolytics as novel mechanism-based treatment modalities for OSMF.


The ripple effects of protracted armed conflicts include: significant gender-specific barriers to accessing essential services such as health, education, water and sanitation and broader macroeconomic challenges such as increased poverty rates, higher debt burdens, and deteriorating employment prospects. These factors influence the wider social and political determinants of health for women and a gendered analysis of the political economy of health in conflict may support strengthening health systems during conflict. This will in turn lead to equality and equity across not only health, but broader sectors and systems, that contribute to sustainable peace building.The methodology employed is a multidisciplinary narrative review of the published and grey literature on women and gender in the political economy of health in conflict.The existing literature that contributes to the emerging area on the political economy of health in conflict has overlooked gender and specifically the role of women as a critical component. Gender analysis is incorporated into existing post-conflict health systems research, but this does not extend to countries actively affected by armed conflict and humanitarian crises. The analysis also tends to ignore the socially constructed patriarchal systems, power relations and gender norms that often lead to vastly different health system needs, experiences and health outcomes.Detailed case studies on the gendered political economy of health in countries impacted by complex protracted conflict will support efforts to improve health equity and understanding of gender relations that support health systems strengthening. Gender is a fundamental yet hugely neglected component in the political economy of health in armed conflict.There is limited evidence on the role of gender in post-conflict health systems but not in active conflict and humanitarian crises.Employing a gendered lens to political economy of health analysis will advance gender equity and equality in conflict settings.Women’s inclusion in the political economy of health in conflict has greater dividends for sustainable peace and more equitable social economic recovery in the post-conflict period.Women’s meaningful participation in peace processes creates more legitimate representation of socio-economic and health concerns.The larger the gender gap, defined as the differences in experiences and opportunities between men and women, the more likely a country is to be involved in violent conflict , 2. FurtThe main aim of this study is to consider and develop an understanding of the role of gender, specifically women, in the political economy of health in armed conflict settings. This article combines existing literature from a range of academic disciplines on three main subject areas: gendered and feminist political economy, gender and peace including peacebuilding, and health systems research in active conflict and post-conflict settings. It aims to enhance understanding of the importance of employing gender analysis within the wider literature on the gendered political economy, and furthermore strengthen the nascent literature of the political economy of health in conflict by ensuring it is not gender blind and envisions gender as a core component in its analysis . In contgendered political economy of health in conflict. We define this concept as the gendered analysis of the causes of disease distribution and relative access to health services requiring attention to the political and economic structures, processes and power relations that produce socially constructed patterns, norms and structures  to explain women’s largely subordinate role within these structures [From a health systems perspective, the political economy of health in conflict examines the historical development and policies of health systems, the role and impact of the multiplicity of actors, including government, non-state actors, the private sector, international donors and humanitarian agencies, and their subsequent effects on socioeconomic health policies and health outcomes . Analysiructures , 16. Thiructures . It shouructures . In acknructures . Interseructures . While tructures , 20. Thehttps://healthsystemsglobal.org/thematic-groups/fragile-and-conflict-affected-settings/); NIHR Research Unit on Health in Situations of Fragility ; the WHO Collaborating Centre for health systems in fragile and conflict affected contexts at the Royal Tropical Institute . Relevant grey material was obtained from appropriate multilateral bodies, intergovernmental and non-governmental organisations. Examples of these sources included: World Health Organisation (WHO), other United Nations (UN) agencies , the World Bank, the International Monetary Fund (IMF), the International Labour Organisation (ILO), the World Economic Forum (WEF), CARE, Oxfam, and the Gender and Development Network (GADN).This study is a narrative review comprising both published and grey literature from a range of academic disciplines such as political science, global public health, international political economy, political philosophy and sociology, gender and feminist political economy, humanitarian, conflict and post-conflict studies. Given the limited literature base supporting this topic, a narrative review was deemed most appropriate as it provides a comprehensive overview of the concepts and wider literature contributing to this specific area, incorporating a diverse range of sources and knowledge-bases, to deepen understanding of this complex topic . EnglishSearch terms included combinations of the following: gender, women, men, feminist, masculinity, gender blind, political economy analysis, political economy of health, political economy of health in conflict, political economy of war, war economy, patriarchy, vulnerability, economic vulnerability, determinants of vulnerability, economic shocks, economic stressors, socio-economic stressors, economic recovery and conflict, humanitarian and political economy of conflict, informal economy, human rights, gender rights, health decision making, health systems strengthening, humanitarian crises, armed conflict, and protracted conflict.https://ucdp.uu.se/year/2019) with additional information used from the Armed Conflict Location & Event Data Project (ACLED): https://acleddata.com/data-export-tool/ We also included low and middle-income regions in conflict: the Sahel, Nagorno Karabak, Palestine. These countries have remained low or middle-income during 2000–2020.The inclusion criteria for our analysis were low and middle-income countries in active armed conflict and affected by humanitarian crisis between 2000 and 2020; this also included low and middle-income countries considered as post-conflict. Definition of low and middle-income were informed by World Bank data as of 2021 . Our lisWe excluded high-income countries including those that host refugees as this requires a separate level of analysis.Interpretation and synthesis of the relevant literature was used to capture the results below. Four broad areas emerged from a mixture of theoretical and empirical research: definitions and content of a gendered political economy and why it is important in health and conflict; the widespread health and non-health effects of armed conflict on women; the importance and lessons from work on gendered health systems in conflict and fragile contexts; and lastly, the relation between gender equality and sustainable peace. These are presented in turn below.power and resources and how they are distributed and contested (either through violent or non-violent means) in different contexts: stable, conflict and post-conflict [Within the broader disciplines of political economy and related feminist theories, there are several theoretical frameworks which are useful for understanding what political economy of health in conflict means, how gender fits into this and why it is important in strengthening public systems in conflict and advancing policy reform that supports peacebuilding initiatives . Politicconflict , 24. A gconflict , 25. Piaconflict . Gendereconflict . Alsaba conflict .Conflict effects men and women differently and exacerbates gender inequalities . Men andFrom a health systems perspective, political economy examines historical development and policies, the role of influential stakeholders such as the government , humanitarian agencies and donors, the private sector and community-level organisations . Incorporating gender into the political economy of health systems illustrates how females, males and peoples of other genders live, work and relate to each other at all levels of the health system. Factors such as vulnerability to illness, health seeking behaviour, access to health services, health expenditure and financing, nature of the health workforce (including capacity strengthening), data collection and management, design and use of medical products and technology, and health policies are all heavily influenced by the gendered political economy of health in conflict , 35. TheA worrying trend from the literature is that the socioeconomic and political status of women directly impacts the likelihood of conflict. As highlighted, the larger the gender gap, the more likely a country is to be involved in inter- and intrastate conflict. Melander’s study suggests that more equal societies, measured as female representation in parliament and the ratio of female-to-male higher education attainment, have lower levels of intrastate  armed conflict . StudiesTwenty years ago, the United Nations Security Council (UNSC) passed Resolution 1325 to support women’s participation in peace negotiations, post-conflict reconstruction, and to protect women and girls from wartime sexual violence . Since tOur main findings suggest that although there is emerging global health interest in the broader fields of political economy, there was very little evidence devoted to gender and specifically women in the political economy of health in active conflict and humanitarian settings. The literature furthermore appears sparse with very little case study or empirical evidence to test more developed theoretical perspectives. Very few case studies examine the broader socioeconomic impacts of health in conflict; how men and women’s livelihoods are affected and how this connects to health systems, access to health services and health seeking behaviour. Nevertheless, there was considerable theoretical discussion on what constituted a gendered political economy of health and why it is important. In its application in conflict affected settings, it was notable that most of the literature concentration was on post-conflict settings rather than those embroiled in active conflict with severe on-going gender-related conflict and humanitarian consequences. Three interrelated reflective themes are analysed below, which may provide researchers and policy-makers a platform to advance this area and specifically how it could be applied in protracted conflict settings to support monitoring and implementation efforts , to endorse UNSC 1325 (and related UNSC resolutions), the World Health Organisation’s Commission on the Social Determinants of Health, the Humanitarian Grand Bargain, the Women Leaders in Health and Conflict Initiative and several related gender equality markers.Firstly, a gendered political economy perspective is vital for understanding how power relations and entrenched patriarchy hinder progress on gender equality and equity in countries and regions affected by war. It is well established that conflict has widespread collateral effects such as destruction of the health system, adverse health outcomes for women, generational effects such as poverty . A gendeDespite global declarations and commitments to gender equality in conflict and humanitarian crises, several barriers remain entrenched that prevent these goals from being effectively implemented. Statistics of peace accord content and implementation rates found that agreements with women signatories demonstrate better accord quality and higher implementation rates - Yakin Ertürk, a former UN Special Rapporteur on Violence against Women, argues that “although new international gender equality instruments continue to be adopted, they remain divorced from the structural inequalities that characterise the global political economy. The comeback of the security state, rise of right-wing populism, general erosion of states’ human rights commitments and the shrinking political space for civic activism, leave these instruments deskbound” . InvolviSecondly, there is significant evidence on the gendered nature and burden of conflict and the subsequent impact on health systems. Women suffer more so than men from the damage to the health infrastructure, wider economic damage and forced displacement. In many societies the division of labour tends to be highly gendered with women taking on responsibility of households and communities, a role that is increasingly divested unto women and girls especially in the times of crisis. Women’s gendered role within the household can limit their participation in formal employment which reinforces gendered norms around women’s role as secondary and dependent, not only within the household, but also within public services and markets . Under-pLastly, the health systems and conflict literature features discussions on gender equity and equality but only in very few post-conflict settings. Limited evidence suggests that public health measures including equitable access to basic health care may contribute to peacebuilding in the aftermath of conflict . Kruk’s This study has several limitations. Firstly, while there is very little literature available on the gendered political economy of health in conflict, there is even less country-specific analysis, which would take into account the various typologies of conflict, for example what resources are available in specific countries during active conflict varies greatly. This makes determining the need for a gendered political economy of health complicated, as context specificity would enhance its applicability. Secondly, defining the gendered political economy of health in conflict is open to considerable interpretation. This is compounded by there being no single conceptual framework for political economy analysis, the lack of a working definition of the political economy of health in conflict, as it is a novel concept, and that the feminist political economy and gendered political economy are frequently used interchangeably. Thirdly, this study does not elaborate on the role of men or individuals who identify with non-binary genders within the gendered political economy of health in conflict. While crucial in understanding the nascent concept of the gendered political economy of health in conflict, it requires further investigation which would complement not only this study, but how the concept is understood across all active conflict settings. Fourthly, this study only used search terms in English and Arabic. Further literature may be available in other languages and this may also source literature beyond the regions and countries we found. Most of the relevant studies we found were from the Eastern Hemisphere with the Middle East and North Africa region being overrepresented in the literature compared with other conflict-affected regions. Subsequently, we are unable to make conclusions on all conflict-affected regions. We therefore recommend future studies address this key limitation.The gendered political economy of health in armed conflict is a convoluted term, influenced by various concepts, theories and ontologies. Yet its importance in deepening understanding of the nexus of health, gender and armed conflict is crucial. Gender-responsive analysis remains thin within the wider field of the political economy of armed conflict . Health


As the leading causes of global death, cardiovascular diseases are generally initiated by artery-related disorders such as atherosclerosis, thrombosis, and aneurysm. Although clinical treatments have been developed to rescue patients suffering from artery-related disorders, the underlying pathologies of these arterial abnormalities are not fully understood. Biofabrication techniques pave the way to constructing diseased artery in vitro models using human vascular cells, biomaterials, and biomolecules, which are capable of recapitulating arterial pathophysiology with superior performance compared with conventional planar cell culture and experimental animal models. This review discusses the critical elements in the arterial microenvironment which are important considerations for recreating biomimetic human arteries with the desired disorders in vitro. Afterward, conventionally biofabricated platforms for the investigation of arterial diseases are summarized, along with their merits and shortcomings, followed by a comprehensive review of advanced biofabrication techniques and the progress of their applications in establishing diseased artery models. The vascular system in the human body plays an important role in circulating nutrients, oxygen, and metabolic wastes. In general, the vascular system can be divided into three subsets of blood vessels: arteries, veins, and capillaries . DiffereAlthough an increasing number of newly developed clinical treatments have been demonstrated to be useful for rescuing patients , plenty of the underlying pathologies of these diseases remain unrevealed, limiting the establishment of effective therapies for disease regression and recovery . In addiA plethora of diseased artery platforms have been previously developed based on experimental animals or planar cell culture . HoweverTo establish a viable diseased artery platform, it is essential to define the core features that should be involved. First, the model should recruit all relevant cells types associated with the target disease. Second, the involved multiple types of cells should be organized in a three-dimensional space following the anatomical architectures shown in natural tissues and disease nidus, facilitating the formation of functional tissues and cell–cell interactions. Third, important biomechanical and biochemical stimuli should be incorporated into the model, exposing cells to disease environments that are key factors in the initiation and deterioration of the relevant disorders. Last, the platform should be easy to reproduce via commonly used tissue culture and engineering approaches, and the outcomes should be easily obtained and analyzed. With the development and progress of science and technology, emerging biomanufacturing technologies may be able to fabricate in vitro models that can achieve these objectives. Biologists and engineers have collaborated to model arterial diseases in vitro, producing diverse innovative tools.This review focuses on the biofabrication of diseased artery in vitro models. We first describe the critical elements in the vascular microenvironment, including vascular cells, cell–cell crosstalk, cell–ECM interaction, and physiological dynamics, which are important considerations for recreating biomimetic human arteries with desired disorders in vitro. Afterward, conventional platforms for the investigation of arterial diseases, such as designed apparatuses, transwell systems, and needle-templating bulk hydrogels are summarized along with their merits and shortcomings. Upon the discussions of advanced techniques , the recent progress of diseased artery models  established using these biofabrication approaches is reviewed.The tissue microenvironment refers to a dynamic population of multiple types of cells and non-cellular components , growth factors, and cytokines) which form a multi-faceted regulatory network that helps to maintain the homeostasis of tissues and organs . BesidesIn general, the wall of a human artery is composed of three distinctive layers, namely tunica intima, tunica media, and tunica adventitia, which are sequentially positioned from the luminal to the abluminal side A. The tuThe ECs residing in healthy arteries play an important role in mitigating blood coagulation and regulating molecular transportation. The typical physiological functions of the endothelium include selective permeability, the resistance of platelet and leukocyte adhesion, and stimuli response B. First,The VSMCs are circumferentially arranged in the tunica media, playing an important role in regulating blood pressure and blood distribution by maintaining an intrinsic level of contractile force, known as vascular tone B 16]. U. U16]. UAlthough the adventitia might not be the frontline during the initiation and progression of most arterial disorders, it acts as immune surveillance and warns of impending vascular diseases. Except for the dominated fibroblasts, the external layer of the artery also includes a mixture of progenitor/stem cells, pericytes, myofibroblasts, macrophages, and dendritic cells which respond to endothelial injury and travel into the other two inner layers for the missions of immune cell trafficking and ECM compositional correction B 19]. I. I19]. IAlthough the human artery is distinctively compartmentalized, the representative cells in each layer continuously communicate not only to maintain the vascular homeostasis but also to mediate diseases. The response of the artery toward pathophysiological stimuli highly depends on the crosstalk between ECs and VSMCs. Such intercellular communications are basically achieved by paracrine association via the diffusion/transportation of soluble factors and extracellular vesicles.Soluble factors, including growth factors, small molecules, and cytokines, are commonly synthesized by cells with specific purposes. Since they are capable of binding to cellular receptors and causing permissive or preventive signaling events , the diffusion of soluble factors is an essential process in EC–VSMC crosstalk. In general, the factors secreted by ECs and VSMCs can regulate and control the morphology, phenotype, and function of each other. For instance, since VSMCs are not directly exposed to the biomechanical and biochemical changes in the blood flow, the signal transduction induced by ECs creates a bridge for VSMCs to react. One representative example is that the endothelial-derived factors can affect the contractile condition of VSMCs, leading to the mediation of vascular tone. While the nitric oxide, hyperpolarizing agents, and prostacyclin released from ECs can cause muscular relaxation in the artery, other factors such as angiotensin II or endothelin promote the contraction of VSMCs . Except Another important mechanism of intercellular dialogue lies in the secretion of extracellular vesicles. According to the dimension, lipid composition, or mechanisms of formation and discharge, the extracellular vesicles can be categorized into three types: exosome 30–100 nm), microvesicle (0.1–1 μm), and apoptotic bodies (over 1 μm) . These e–100 nm, Besides the interactions between parenchymal cells in arterial tissue, the crosstalk between arterial tissue cells and the cells circulating in the bloodstream or exiting in perivascular tissues also significantly determines the pathophysiology of the artery. The communication between ECs and leukocytes is a representative example. In normal cases, the ECs tightly contact each other via several types of intercellular junctions, forming a non-adhesive endothelial surface that limits the attachment and extravasation of patrolling white blood vessels. In response to inflammatory cytokines such as tumor necrosis factor α (TNF-α) and interleukin 6, the ECs are locally activated, expressing intercellular adhesion molecules , vascular cell adhesion molecule 1 (VCAM-1), P-selectin, E-selectin, etc.) . These mTherefore, due to the importance and complexity of intercellular communications, it is critical to involve all participating cells when establishing an in vitro model for investigating the pathological events and signaling pathways of artery-related diseases.Toward the replication of vascular tissue microenvironments, the interaction between cells and ECMs is another pivotal consideration because the compositional and structural properties of ECMs significantly direct cell fates. Relying on transmembrane receptors, such as integrins, α-dystroglycan, and syndecans, such messages can be detected by cells, providing a link between the cytoskeletons and the ECMs to enable cell–ECM contact.All parenchymal cells in the arterial tissue secrete matrix components that contribute to the maintenance of the vascular properties and affect adjacent cell functions. ECs are primarily responsible for the synthesis, deposition, and assembly of the basement membrane, which contains laminin, type IV collagen, perlecan, and nidogens. In return, the EC functions are correlated with the formation and degradation of the basement membrane. While the matured basement membrane is a prerequisite for ECs to generate stable endothelium tissues, the dynamic degradation of the basement membrane as a result of enzyme activity  facilitates the migration of ECs, leading to the sprouting of micro-vessels . VSMCs, Varied compositions of the matrix proteins are particularly associated with cellular receptors. Upon ligand binding, specific integrins determine different signal transduction pathways that mediate cellular fates, such as the modulation of the cell cycle, the organization of the intracellular cytoskeleton, and phenotypic switching. For instance, it is known that the phenotypic change of VSMCs is highly dependent on the type of ECM component. Among the primary ECM components present in tunica media, type I collagen and fibronectin induce a synthetic VSMC phenotype, while type IV collagen and assorted proteoglycans  appear to inhibit the proliferation and migration of VSMCs . InteresAs a part of the circulatory system for the transportation of pumped blood, the arteries are exposed to a variety of physiological dynamic signals including stresses and deformations originating from blood flow and pressure.In general, the dynamic stress imparted on the arterial wall by blood flow is named hemodynamics, entailing the shear stresses from the flow of blood, longitudinal stresses from the surrounding tissue, and circumferential stress from the blood pressure. ECs possess diverse receptors that can sense the blood flow and pass hemodynamic signals through mechanotransductive signaling pathways to recipient molecules, leading to phenotypic and functional changes. In healthy arteries with simple straight geometries, the ECs are exposed to laminar flows and high shear stresses. Such a condition induces pro-survival antioxidant signals and maintains the quiescent phenotype of ECs. However, for bifurcated and curved arteries, the flows are usually disturbed, generating slow and oscillating shear stresses, where the ECs show a pro-inflammatory phenotype associated with low NO production, high permeability, pro-adhesiveness, pro-coagulation, and pro-proliferative properties ,37. For In contrast to the shear stresses, stretch deformation directly influences VSMCs. Such a mechanical force is demonstrated to enhance the expression of ECMs and contractile proteins from VSMCs. Previous efforts have proven that the physiological mechanical strain applied to the VSMCs residing in the human adult aorta can increase the synthesis of collagen and fibronectin, as well as the levels of matrix-degrading enzymes, leading to the accumulation of ECMs. Enlightened by this finding, bioreactors that can provide mechanical stimuli have been integrated into the process of tissue engineering to produce artificial blood vessel grafts . BesidesTaken together, similar to other tissues in the human body, the artery possesses an intricate microenvironment entailing multiple types of cells and ECM components. The close associations between cells and cells or cells and ECMs enable the achievement and modulation of arterial physiological functions. Besides, the physiological dynamics originating from blood flows clearly have a major impact on cellular fates and performance. Therefore, to establish an in vitro platform that is viable for recapitulating the pathophysiology of the human artery, all these critical elements should be collaboratively involved.Upon the maturation of the cell culture technique, it is possible to culture vascular cells in planar devices, such as culture dishes and flasks. However, the mono-type of cells cultured statically in two-dimensional space can hardly reflect the microenvironments in their natural counterparts. To overcome the challenges, referring to the working principle of the rheometer, the first generation of in vitro apparatus was designed to understand the response of vascular cells to tunable flow shear stress.The designed apparatus is made up of cones and plates to simulate the viscoelasticity, the fluid shear stress of ECs in vitro, and other properties of blood . For ins2 shear for 72 h when rotating the flow direction 90° in the cone-plate device. By further experiments, the shape of cells was induced by the shear effect. These results also indicated that cell morphological change and orientation are quite sensitive to the applied shear force. Malek and teammates developed a cone-plate viscometer apparatus which was specially designed for studying the effect of fluid shear stress on large populations of adherent cells in vitro [2 for 24 h and 96 h. The experimental results showed that compared with cells under static conditions, there was no difference between BAE cells exposed to 24 h of shear, as shown by the continual integrity of the BAE monolayer and frank alignment of cells along a streamlined direction when BAE cells were exposed to 96 h of constant fluid shear in the steady laminar regimen. With the assistance of the devised apparatus, the investigations of mechanisms in arterial diseases can be promoted to a higher level. Wayne Orr et al. reported that fluid flow, atherogenic flow, oxidized low-density lipo-protein (LDL), and proatherogenic cytokines all stimulated p21-activated kinase (PAK) phosphorylation and recruitment to cell–cell junctions based on large transwell membranes mounted into a modified cone-plate viscometer, as shown in Such a type of design helps to reveal the relationships between EC activity and hemodynamic forces. Dewey et al. developed a cone-plate Couette flow device that was similar to commercial cone-plate viscometers consisting of the rotating cone and the stationary flat plate that first reported the response of ECs to flow shear stress . They foin vitro . They cuAlthough the method mimics blood dynamics, there are huge limitations when building in vitro models of blood vessels simulated by a cone-plate apparatus. Because monolayer cells undergo the largest shear stress gradient from zero in the center to the periphery, this change in shear stress makes cone-plate and double-disc rheometers unsuitable for most biological studies. Cone-plate devices are usually used for monolayer cell studies, but rarely for the co-culture of cells. Therefore, 3D tissue culture is not achievable with this type of device, which limits the applications of this type of device.To meet the requirement of co-culturing multiple types of cells in a single platform, the transwell system emerges as a useful tool, as shown in Benefiting from these advantages, the transwell system has been widely carried out to build diseased vascular in vitro models including atherosclerosis , the bloThe transwell system has been widely used in academia and the pharmaceutical industry because of its simplicity, cost-effectiveness, high and medium-throughput screening characteristics, and versatility. The transwell model not only has access to apical and basolateral  compartments for drug applications and media sampling but also enables the visualization of cells during experiments. Despite the development of new biofabrication technologies, the transwell system still offers a distinct advantage in that it is relatively easy to set up and control and provides a range of endpoint studies. It provides a reliable, non-invasive quantitative measure of barrier integrity that can be repeated over the required time cycle with minimal damage to cells. However, a considerable drawback of the transwell system is the limited ability to offer flow signals to the residing cells. Therefore, it might be difficult to reveal the effects of hemodynamics on the pathological changes of arterial cells.The concept of creating microchannels to replicate vascular microenvironments truly realizes the in vitro models of arterial tissues and diseases on a three-dimensional level. To achieve this objective, needle-templating is a simple and maneuverable approach, which is also called the subtractive method . This meThis approach has been widely used to study diverse pathophysiology in vitro models. Paek and colleagues focused on the preparation of infusible hydrogel cell culture scaffolds using needle-templating . During However, there are many disadvantages and limitations of needle-templating microchannels. For instance, removing rigid needles and flexible rods may result in micrometer-scale defects at the interface, requiring extra care and precision in the removal process . The insOver the past decade, the improvement of micromanufacturing technology has promoted the development of tissue and organ models in vitro to overcome the shortcomings of traditional methods. The innovative bio-manufacturing technology has further contributed to the understanding of the pathology and clinical drug development of human diseases through fabricating sophisticated in vitro models of diseased arteries. The novel technologies have become a popular tool for revealing the pathologies at cellular and molecular levels, especially in the vascular system, whose characteristic mechanics and structures are difficult to generalize in traditional systems. This section reviews advanced biofabrication approaches including tissue-engineered blood vessels, organ-on-a-chip, and 3D bioprinted tissue analogs, focusing on the work principles of these manufacturing methods and discussing the benefits and limitations of these methods in building vascular pathological models in vitro.Translating the tissue-engineered blood vessels (TEBVs) that are commonly used as bypass grafts to understand the pathology of artery-relevant diseases is another strategy. The most representative approach for the development of TEBVs relies on the rolling of multiple vascular cell sheets to mimic the natural arrangement of cells and the structure and function of a variety of tubular tissues occurring naturally in the body ,69 Figu.Due to mimicking and controlling the different cell arrangements , TEBV teUndoubtedly, TEBVs might be the most eligible candidate for emulating the natural arterial function and physiological environment. However, the low reproducibility and production rate are the critical drawbacks of TEBVs as in vitro research models, especially when the applications require high throughput performances, such as drug screening.An organ-on-a-chip (OoC) is a microfluidic cell culture device containing a continuous perfusion chamber with a multi-cellular layer structure, tissue interface, physicochemical microenvironment, and human vascular circulation. It can also be considered as a cell culture biomimetic system that can simulate and reconstruct the physiological functions of human organs, with the ability to adjust key parameters such as concentration gradients, shear forces, cell patterns, tissue boundaries, and tissue-organ interactions. OoC consists of four key components , includiOoC has exhibited distinguished advantages over the traditional 2D cell culture methods, such as realizing 3D cell co-culture, offering dynamic stimulations, and replicating the tissue interface. The presence and integration of multiple cell types could enable the elaborated chip to reflect the natural physiological interplays of cells  in a 3D environment. Besides, the OoC may also reconstitute the biomechanical characteristics of arterial tissue, such as the muscular contraction and the shear force induced by blood flow. In addition, the microchannels established on OoC serve to refresh and transport cell metabolite products . MoreoveBenefiting from these merits, OoC has shown broad potential for the construction of in vitro arterial models as well as applications in pathological investigation and drug screening . Tsai etBesides the shinning points discussed above, OoC may also pave the way for tackling the challenges of bioengineering techniques, such as guiding cellular compartmentalization and organization, as well as integrating bioelectrical and biochemical signals to provide the cells with a habitat comparable to their natural counterparts. For example, smooth muscle cells’ (SMCs) circumferential arrangement in the artery can be realized by preparing rectangular microchannels to investigate the relationship between cellular orientation and their functional contractility . The narTo gain insight into the type and size of the intracellular forces generated by cell–cell and cell–ECM interactions, OoC can stimulate arterial deformation via electrodes, which has a significant impact on cell function in vitro . The in Despite the great potential and achievements, there are limitations of current OoC systems that confine the extent of pathophysiological remodeling in vascular disease. A large number of developed OoC systems are based on the use of polydimethylsiloxane (PDMS). However, a major drawback of PDMS is that the material absorbs small hydrophobic molecules, making it difficult to evaluate the pharmacokinetics of drugs and toxins ,90. AlthThree-dimensional bioprinting represents the frontier technology in the field of biological manufacturing and is used to fabricate diseased vascular models with complex structures. A notable feature of 3D bioprinting is the ability to manufacture a wide variety of tubular structures and sizes, from arterial to arteriolar sizes, as well as branching structures, and even incorporating vascular preprocessing networks in 3D stents for tissue regeneration . Dellaqu6 cells/mL). The applied bioinks can be chemically gelled or photo-crosslinked. Besides, its preparation time is low and print speed is fast, up to 10,000 droplets per second. Cell viability has been reported in the 80–95% range using this method due to temperature and mechanical stress [Inkjet bioprinting (IBP)—consisting of X, Y, and Z, three moving modules with nozzles on the Z-axis for loading bioinks—uses heat or pressure electricity to deposit the bioink drop by drop, so it comes in two forms, including thermal inkjet bioprinting and piezoelectric inkjet bioprinting ,94. The IBP has been used in a variety of applications, including tissue engineering and regenerative medicine, transplantation and clinical applications, drug testing and high-throughput screening, and cancer research ,96. BecaThe advantages of thermal inkjet printers are fast printing speed, low cost, and wide availability. However, the risk of cells and materials being exposed to thermal and mechanical stresses, uneven droplet sizes, frequent clogging of nozzles, and unreliable cell encapsulation bring considerable disadvantages to the use of these printers in 3D bioprinting. The advantages of piezoelectric inkjet printers include the ability to generate and control uniform droplet sizes and jet directions, as well as the ability to protect cells from exposure to heat and pressure stress sources by creating a sound wave inside the print head that breaks liquid into droplets at regular intervals. In addition, the use of an open pool nozzle-free injection system can avoid the pure stress imposed on the wall cells at the tip of the nozzle. This reduces the potential loss of cell vitality and function and avoids the problem of nozzle clogging. Sonic injectors can be combined into multiple injectors in an adjustable array format, making it easy to print multiple units and material types simultaneously ,100. How8 cells/ml with microscale resolution of a single cell per drop using a laser pulse repetition rate of 5 kHz [Laser-assisted bioprinting (LAB) is an on-demand drop method based on a pulsed laser beam incident on a donor slide in contact with the energy absorption layer. When a bioink is placed next to the energy-absorbing layer, a shock wave forms a jet of bioink that is deposited as water droplets on the collector’s slide ,103. Typof 5 kHz ,106.2 solution was packed into the ribbon. The bioink was then printed onto the fibrinogen, and the laser scanning speed was set to 200 mm/s, based on parallel and vertical lines, and the pattern was printed in seconds. Koch et al. used LAB technology to print NIH 3T3 and HaCaT cells in a collagen matrix to demonstrate tissue formation under natural conditions [Given the characteristics of LAB manufacturing technology, Guillotin et al. used LAB to print stable, soft, free-form structures with a resolution compatible with the size of microvessels . They plnditions ,107. DifKey advantages of LAB are high resolution, a wide range of material densities, and the ability to print batteries on solid or liquid substrates. Other benefits include automation, reproducibility, and high throughput. However, this is a very expensive technique and can cause cell damage. The high resolution of LAB requires fast gel dynamics to achieve high shape fidelity, which results in relatively low overall flow rates .7 mPa/s [Extrusion-based bioprinting (EBB), as the most common 3D bioprinting method, uses layer stacking to produce in vitro models by depositing polymers or bioinks  components, and cell-laden hydrogels) ,110. An 7 mPa/s .EBB has broad utility in various application areas , includiEBB technology has led to the development of complex organotypic in vitro models, driving its use in a variety of biomedical applications, such as tissue modeling ,126 and Photopolymerization-based bioprinting (PBB) is a technical method for fabricating three-dimensional patterned scaffolds with micro/nanostructures. It is a typical light-assisted direct printing method for curing photosensitive biologic inks in the layer-by-layer process. It is a projection printing system that uses a light projector to cross-link photocurable bioinks plane-by-plane ,130. LikIn the wide applications of PBB technology in biological manufacturing, as one of the important bio-materials in biological tissue engineering, hydrogel is often used in photocurable 3D printing to manufacture vascular tissue. Ma et al. developed a new triculture model based on 3D hydrogels by using PBB technology, which combined physiologically relevant cells with liver-specific 3D microstructures . They emDue to the ability to prepare complex structures with a high, precise resolution, its applicability to a variety of bioinks, and its computer-aided design and imaging capabilities to generate patient-specific complex tissue , PBB tecOverall, 3D bioprinting opens up promising avenues in regenerative medicine and disease modeling by promoting the versatility of materials and precise spatial resolution and may provide the necessary technologies for personalized medicine applications when combined with patient-derived cells, custom bio-inks, and microfluidics/sensing technology . As moreDiseased arteries, including atherosclerosis, thrombosis, aneurysm, etc., are caused by changes in the structure and function of blood vessels. Despite clinical advances in the treatment of vascular disease, the mechanisms underlying the progression and regression of multifactorial arterial wall disease are still not fully understood ,141. TheAtherosclerosis begins with endothelial injury induced by inflammatory factors, with large amounts of cholesterol deposited in the major and medium arteries  model with a three-layer structure, ECs, SMCs, and fibroblasts . Early aCompared with the scaffold-based approach, 3D bioprinting can pave the way to easily achieving the goal of engineering arterial analogs in vitro. Notably, since the artery is a multi-layer lumen structure, utilizing coaxial bioprinting could effectively establish the model. Gao et al. reported an advanced bioprinting strategy for a triple-layer artery model with lumen-regulating diameters, geometry-tunable by controlling the nozzle moving speed, flow rate, and pneumatic pressure . The triIn addition, an artery/atherosclerosis model on a chip has been established for studying the development of atherosclerosis . Su et aAll in all, regardless of which biotechnology is used to construct in vitro models of atherosclerosis, there are key points that need to be considered to improve the accuracy of in vitro models and gain insight into this systemic disease. First and foremost, cells carrying atheroprone mutations should be considered in the atherosclerosis of in vitro models to better understand each mutation’s peripheral impact on the vasculature. Second, it is important to fabricate an in vitro model of atherosclerosis tailored to the patient for diagnosis, which is conducive to achieving target-oriented treatment. In particular, atherosclerotic models should predict plaque size, location, and growth rate based on patient data to predict disease prognosis and provide a successful impact on specific treatments .In a healthy body, clotting and anticoagulation systems function normally. When vessel wall injury, platelets, tissue factor, and fibrin accumulate, they form a thrombus . Atheros2 solution, added to induce clotting. Ten minutes later, they observed the clotting formation and H&E staining confirmed thrombus formation. They perfused this model with a tissue plasminogen activator, a thrombolytic agent, and 7 days later achieved a good thrombolytic effect. In addition, they explored the fibroblast existence for the formation of fibrotic clots.Thrombosis-on-a-chip is a useful model for studying developing blood clots due to utilizing human ECs and human blood for studying interactions, which is rare in other technology . Zhang eMicrofluidic devices can mimic physiological fluid shear and achieve changeable geometry, which is important for clinical experiments . Jain etIn conclusion, all in vitro models of diseases are designed to accurately express pathological processes to provide targeted treatment options. Thrombosis, a common and fatal blood vessel disease, should be considered precise and reproducible in 3D in vitro models. Key cell populations, ECM components, morphology, and the intravascular topography involved in coagulation cascade and hydrodynamics should be considered in the preparation of its in vitro model to better express the coagulation pathological process of thrombus.The aneurysm is defined as a local globular dilation of a normal artery that is about 1.5 times larger than the diameter of a normal adjacent segmentary artery ,161. CliThe microfluidic platform can intuitively simulate the pathological hemodynamics of aneurysm in vitro models and mimic the influence of aneurysm morphology on aneurysm formation . Kaneko Compared with other methods of in vitro model manufacturing, 3D bioprinting is a more promising method. Three-dimensional bioprinting technology can build a more accurate 3D aneurysm model to better reproduce the direct effect of blood flow on the aneurysm. In this way, the in vitro model of the aneurysm can be established, which is beneficial to further understanding the pathology of aneurysms and predicting the deformation and strain degree of aneurysm vessels. Importantly, 3D bioprinting technology can realize the research of drug targeted therapy for aneurysms. Epshtein et al. established an in vitro model of aneurysms by using photocurable 3D bioprinting technology . They deSummarily, more accurate aneurysm in vitro models should be developed to better reproduce the direct effect of blood flow on the formation of new ECMs. Combining 3D bioprinting and microfluidic technology, further study on the relationship between fluid dynamics and the stress–strain of aneurysm vessels, as well as the remodeling of cells such as ECs at the location of lesion vessels under the action of blood fluid. Importantly, an appropriate in vitro model of aneurysms should also facilitate discussion of what cellular, extracellular, and mechanical signals promote the synthesis and regulation of proteins involved in ECM remodeling.The development of diseased artery in vitro models from 2D static to 3D dynamic is an important embodiment of the cross-era of biotechnology, which is conducive to a better understanding of pathology and the realization of drug therapy. As the basis for the construction of arterial disease in vitro models, it is important to understand the physiological elements in the target tissue, including cells, ECMs, and dynamic signals, as well as the interactions between them. Although a variety of conventional techniques, such as designed apparatuses, transwell systems, and needle-templating microchannels have been developed to model the arterial pathology in vitro, their limited ability to precisely mimic the natural cellular, biomechanical, and biochemical cues confined their applications. To overcome these challenges, advanced biofabrication technologies, microfluidic platforms, organ-on-a-chip, and 3D bioprinted artery analogs emerged and made a qualitative leap in the in vitro models of artery-relevant diseases, including atherosclerosis, thrombosis, and aneurysm. The accumulated achievements provide a better understanding of the pathology of human arterial disorders and suggest a novel tool for drug discovery.In the future, the technology for realizing the in vitro modeling of a human artery is still the hot research in the field of biofabrication, especially for the in vitro modeling of patient-specific diseased tissue. This may require a combination of medical imaging, organ-on-a-chip, and 3D bioprinting to construct a sophisticated system that can mimic natural artery tissues.


Interactions among the plant microbiome and its host are dynamic, both spatially and temporally, leading to beneficial or pathogenic relationships in the rhizosphere, phyllosphere, and endosphere. These interactions range from cellular to molecular and genomic levels, exemplified by many complementing and coevolutionary relationships. The host plants acquire many metabolic and developmental traits such as alteration in their exudation pattern, acquisition of systemic tolerance, and coordination of signaling metabolites to interact with the microbial partners including bacteria, fungi, archaea, protists, and viruses. The microbiome responds by gaining or losing its traits to various molecular signals from the host plants and the environment. Such adaptive traits in the host and microbial partners make way for their coexistence, living together on, around, or inside the plants. The beneficial plant microbiome interactions have been exploited using traditional culturable approaches by isolating microbes with target functions, clearly contributing toward the host plants’ growth, fitness, and stress resilience. The new knowledge gained on the unculturable members of the plant microbiome using metagenome research has clearly indicated the predominance of particular phyla/genera with presumptive functions. Practically, the culturable approach gives beneficial microbes in hand for direct use, whereas the unculturable approach gives the perfect theoretical information about the taxonomy and metabolic potential of well-colonized major microbial groups associated with the plants. To capitalize on such beneficial, endemic, and functionally diverse microbiome, the strategic approach of concomitant use of culture-dependent and culture-independent techniques would help in designing novel “biologicals” for various crops. The designed biologicals (or bioinoculants) should ensure the community’s persistence due to their genomic and functional abilities. Here, we discuss the current paradigm on plant-microbiome-induced adaptive functions for the host and the strategies for synthesizing novel bioinoculants based on functions or phylum predominance of microbial communities using culturable and unculturable approaches. The effective crop-specific inclusive microbial community bioinoculants may lead to reduction in the cost of cultivation and improvement in soil and plant health for sustainable agriculture. Cultivated soils are one of the most diverse microbial ecosystems, harboring bacteria, fungi, archaea, viruses, protists, and many others and supporting various biogeochemical cycles and plant growth. Soil microbial communities are critical to plant health and adapt rapidly to different abiotic and biotic stresses . The soiMicrobial communities associated with plants, presently referred to as the plant microbiome, extend the host plant genome and their functions . Many stvis-a-vis soil fertility  isolates, by employing both culturable and unculturable approaches. The advent of “omics” technologies understandably provides the tools for a broader understanding of microbial ecosystems and their dynamic interaction with their hosts. These techniques and methods enable the screening of large microbial populations and easily identify the individual or groups of taxa with functional capabilities. Large-scale genomic analyses of plant-associated bacteria have indicated that the bacteria from phyla Actinobacteria, Bacteroidetes, Firmicutes, and Proteobacteria are dominant in different plant niches  can mediate the cycling of several elements from the soil and transform them into the more readily available form of nutrients for plant uptake. Not only do the probiotic action of these formulations increase the growth, yield, and quality of plants, but they are also a tool to produce high-quality functional foods. The use of microbial-based agricultural inputs has a long history, beginning with broad-scale rhizobial inoculation of legumes in the early twentieth century . The “FrThe current knowledge on functions, ecological adaptations, host interactions, and putative beneficial traits of microorganisms associated with the host plants mainly revolves around a handful of cultivable rhizospheric and endophytic bacteria or fungi. Many microbial formulations having individual or mixture of strains are developed and used at present. These biological or bioinoculants are nitrogen fixers, phosphate solubilizers, siderophore producers, photohormone producers, and exopolysaccharide producers. Some of them are involved in lytic enzyme production against pests and pathogens, antibiosis, and induced systemic resistance .Azotobacter, Azospirillum, and Rhizobium; phosphate-, potassium-, and zinc-solubilizing bacteria; vesicular–arbuscular mycorrhiza (VAM), and Acetobacter. Crop-specific biofertilizers like Gluconacetobacter diazotrophicus for sugarcane or generic biofertilizers like Pantoea isolates showing multi-PGP activities in several crops have demonstrated benefits in improving crop yield and productivity  within a microbiome . Phylogein situ. Such systems have been developed for animal and plant models including the well-studied plant Arabidopsis thaliana. With established huge volume of data on the metagenome of different crops, there is a need for its translation to certain tailored microbiome-based solutions for promoting plant growth under a range of environmental conditions and increasing resilience to biotic and abiotic stresses. The genomic data with taxonomic status, habitat compatibility, and functional trait knowledge including metabolic potential of plant microbiome communities can be followed as the approach for designing effective microbial inoculants. Here, based on phylogenetic or functional redundancy, two approaches for synthesizing microbial-communities-based bioinoculants are discussed.Synthetic microbial community analysis in gnotobiotic systems is a valuable approach to create reproducible conditions to experimentally test microbial interactions Microbial colonization in the plant root rhizosphere is the outcome of the interplay between roots exuding chemical compounds that microbes capture as signals and on which their survival and perpetuance depend. The differential abundance of colonizing microbes and the establishment of core-microbiome-based microbial communities forms the basis for plant–microbe interactions. The core members remain present throughout the development of the crop, which may be joined by other taxa during the crop growth. The metagenome data about the relative abundance of colonizing phyla/taxa and core microbiome in the plant rhizosphere and endosphere form the basis for developing Community-Based SB (CSB). Microbial isolates representing the abundant phyla can be sourced either from the crop associated culture bank or with targeted culturomics, for developing the synthetic community. The isolates are expected to be rich in community-forming characteristics like motility, chemotaxis ability, quorum sensing, metabolic diversity, and others. This approach is a direct microbiome manipulation where inoculated CSB may serve to reduce the time required for the rhizosphere microbiome to achieve niche saturation and competitive exclusion of pathogens .Brassica napus, rhizosphere core genera found at each growth stage were generally part of the overall core taxa at the 75% prevalence threshold. Arthrobacter, Bradyrhizobium, and an unclassified Acidobacteria in the class Ellin6075 were present in all growth stages, while other genera joined at the flowering or harvesting stage, as the recruitment of the microbiome is governed majorly by the host plant. Metagenome analysis of more than 600 Arabidopsis thaliana plants from eight diverse, inbred accessions growing at different locations indicated that the core endophytic microbiome is less diverse than their corresponding rhizosphere soil microbiomes. The soil types influenced the microbial communities in the A. thaliana rhizosphere, but the endophytic communities were overlapping and less complex with maximum of actinobacteria and selected proteobacteria. Arabidopsis, but in the endophytic compartment, it remained consistent across different soil types. An extensive bacterial culture collection that captures a large part of the natural microbial diversity of healthy A. thaliana plants was established  and Actinobacteria (Microbacterium and Rhodococcus) strains have the greatest potential to affect community structure as keystone species.ablished . CarlstrSimilar influences of maize inbred lines growing in different soils and agri-management systems suggested the substantial variation in α- or β-bacterial diversity and relative abundances of taxa with a small proportion of heritable variation across fields. Despite significant differences between the microbial community profiles of maize inbreeds, the estimated α- and β-diversity could not define the kinship of the 27 maize inbreeds to supplement the diversification history of maize . EdwardsA. thaliana. Using the plant–bacterium binary-association assays, the effect of bacterial community manipulation was observed on the plant response to phosphate (Pi) starvation. This approach might contribute to microbial communities’ rational design and deployment to improve the host response to biotic and nutritional stresses.The functional diversity within microbial communities enables metabolic cooperation toward accomplishing more complex functions than those possibly exhibited by a single organism. The consortium members or communities can communicate by exchanging metabolites or molecular signals to coordinate their activity through temporal and spatial expression and further execution of required functions. In contrast with monocultures, microbial members at the community level can self-organize to form spatial patterns, as observed in biofilms or soil aggregates. This self-organization enables them to adapt to the gradient changes, improve resource interception, and exchange metabolites more effectively . Hence, In vitro techniques have demonstrated that the host genotypes and abiotic factors influence the composition of plant microbiomes. At the in vivo level, it is a challenge to define the mechanisms controlling the community dynamicity, its assembly, and the beneficial effects on the plant hosts. In an earlier study, the host-mediated natural selection of bacteria by maize roots was employed to select a simplified synthetic bacterial community consisting of seven strains  representing the dominant phyla such as Proteobacteria and Actinobacteria  system, mainly known for P transport, is also a carrier of endophytes in the plant system, can induce systemic resistance to pathogens, and assists in moisture conservation  has been adopted by the 193 member states to develop their vision, strategy, and targets for achieving SDGs by effectively making them part of their policies. In its sustainability framework to realize the goal of ending hunger (SDG2), India has several initiatives that include the management of soil health. Successful organic cultivation and integrated agriculture will be highly dependent on the efficient microbiome-based bioinoculants for plant nutrient management and, more importantly, the recycling of crop residues for soil health . In contrast, many other practices affect the abundance of microbial taxa involved in pest and soil disease suppression and nutrient cycling . The impThe sustainability of the modern agriculture system is critical to feed the continuously growing human and animal populations, wherein the guided use of microbiomes has an inevitable role in promoting plant growth, development, productivity, and nutrient value. The current biofertilizers are based on individual bacterial cultures with specific traits such as N fixation or the solubilization of P or K. But with the detailed diversity and functional analyses of plant-associated microorganisms, a better understanding has emerged that the plant-associated microbiomes have a tremendous and so-far untapped potential to improve the acquisition of nutrients and resilience to abiotic and biotic stresses and, ultimately, the crop yields. The options of generating synthetic communities using taxonomy abundance alone or with functionally annotated predominant taxa are now available for the improved use of microbial resources in crop cultivation. Nevertheless, developing any microbial community requires a collection of promising functionally annotated and compatible isolates in hand, rather than only microbiome data. Hence, it will be appropriate to holistically use the knowledge of unculturable microbiome generated through structural and functional genomics tools and culturable approaches to get the common and rare taxa for synthetic community preparations. The rational workflow for developing community and function-based bioinoculant preparations has been described, which can be used for developing formulations with the targeted functions of nutrient supplementation and stress management in sustainable agriculture.AS conceptualized and wrote the manuscript. VG helped in the finalization of tables and figures. KAn and BR gave intellectual input and edited the manuscript. KAs, JS, PS, and DP contributed in data search for the content and table formulation. All authors contributed to the article and approved the submitted version.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.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.