Choline is a water-soluble nutrient essential for human life. Gut microbial metabolism of choline results in the production of trimethylamine (TMA), which, upon absorption by the host is converted into trimethylamine-N-oxide (TMAO) in the liver. A high accumulation of both components is related to cardiovascular disease, inflammatory bowel disease, non-alcoholic fatty liver disease, and chronic kidney disease. However, the relationship between the microbiota production of these components and its impact on these diseases still remains unknown. In this review, we will address which microbes contribute to TMA production in the human gut, the extent to which host factors  and diet affect TMA production, and the colonization of these microbes and the reversal of dysbiosis as a therapy for these diseases. Choline is an essential nutrient for humans throughout their life. Although humans can produce choline in small quantities through the hepatic phosphatidylethanolamine N-methyltransferase pathway, most individuals need to increase choline ingestion through their diet, in order to prevent deficiency ,2,3,4. TThe majority of people do not meet the dietary requirements for choline, particularly those on vegetarian or vegan diets and pregnant or lactating mothers . LactatiCholine metabolism can be divided into four main pathways, which are involved in the synthesis of acetylcholine, betaine, phospholipids, and trimethylamine  by host hepatic monooxygenases ,32 such Therefore, choline is important for the function and structure of membranes including their signaling, transport, and repair ,37,38. MIn this review, we will revisit the relationship between gut microbiota populations and circulating TMA and TMAO levels, highlighting not only the microbiota genetics behind these changes but also how host genetics can influence the gut microbiome. We will also discuss the relationship between choline-microbiota changes and their impact on different diseases as well as explore microbial modulation as a potential therapeutic treatment.Firmicutes and Proteobacteria, which are known producers of this metabolite . Sp. Sp51]. rettgeri ,52.Proteobacteria may also be able to metabolize the TMAO ingested via the diet into TMA via TMAO reductase using metabolic retroconversion [Methanomassiliicoccales are able to reduce TMAO to methane [Once TMA is formed, it will be subsequently oxidized into TMAO in the liver through the enzyme FMO3 . It is wnversion . Finally methane . TherefoBacteroides enterotype, whereas a carbohydrate-based diet mainly consumed by agrarian societies, was linked to a Prevotella enterotype. Western diets are characterized by animal products such as liver, pork meat, and eggs, which contain large amounts of choline [Neisseriaceae or Desulfovibrio and TMAO levels, which were also elevated in symptomatic coronary vascular disease (CVD) patients and those consuming an animal-based diet. Another study from Cho et al. [Firmicutes; meanwhile, individuals with lower levels of TMAO exhibited a higher abundance of Bacteroidetes. It has been demonstrated that the composition and diversity of gut microbiota can be influenced by the diet, since it is, in turn , an impo choline  and are  choline  but even choline  observedo et al.  reportedSimilar differences in microbiota and TMAO levels have been reported between vegetarian and omnivorous diets, which were also accompanied by a lesser ability to produce TMA in vegetarians . These oGammaproteobacteria, and Erysipelotrichi members were associated with changes in liver fat during choline depletion [Nevertheless, due to the importance of choline as an essential nutrient, choline-deficient diets may also induce gut microbiota alterations and health problems ,63,64,65epletion . These repletion ,66. In aepletion ,67. The The concentration of TMA/TMAO in plasma has been linked to the composition of intestinal microbiota and differences among microbiota enterotypes  have beeChristensenellaceae (later associated with a low body mass index), the methanogenic Archaea Methanobrevibacter smithii, and the genus Blautia [LCT and the abundance of Bifidobacterium [Over a period of decades, different studies have tried to understand how a host’s genetic background may influence the overall microbiome. The proportion of phenotypic variation in a trait that is attributable to genetic variation among individuals is known as heritability . Among t Blautia ,72. Moreacterium ,74 or thacterium .Turicibacter, which was overlapped with the HCS1 QTL for susceptibility to murine hepatocellular carcinomas [Focusing on how host genetics can influence the gut microbiome in a disease context, several associations between specific single nucleotide polymorphisms or genetic variants of host genes and intestinal bacteria have been identified . These srcinomas . This opMoreover, genotype-associated differences in TMAO and TMA levels were also suggested in a murine model. Romano and colleagues  observedAs previously mentioned, TMAO is correlated with disease and all-cause mortality ,91. Low Average plasma TMAO in humans is around 3.3 µM; however, this can vary greatly after the intake of certain foods that are rich in choline and returns to the baseline within a few hours . CKD sufThe inability to regulate the vascular tone is an early marker of CVD and may lead to atheroma-development. Mice fed a choline-supplemented diet displayed significantly less reverse cholesterol transport with reductions in mRNA of Cyp7a1 and Cyp27a1. This attenuated the conversion of cholesterol into bile acids, resulting in cholesterol build-up . FurtherRises in TMAO related with CKD and CVD have been associated with an increased abundance of TMA-producing bacteria . Romano Choline-deficient diets have also been associated with NASH and may result in obesity and hyperglycemia ,66,108. Choline-deficient diets interfere with and disrupt the intestinal barrier ,111,112.Lipid accumulation may be a result of reductions in fatty acid esterification genes, which prolong the half-lives of injury-inducing lipids ,116. MorDysregulation in the microbiome is correlated with the activation of the Nod-like receptor protein 3 (NLRP3) inflammasome which is necessary for fibrosis in developing NASH pathology . MoreoveAbnormal choline-intake is involved in the pathogenesis of major debilitating diseases such as CKD, CVD, and NASH . DespiteData from associative and mechanistic studies draw a strong link between dysbiosis and pathobiotic bacteria in the gut, the microbiota-dependent production of TMAO, and an increased risk of cardiometabolic and gastrointestinal disorders ,152,153.As we have previously described, diet is an important lifestyle determinant of gut health. This makes dietary modification a potentially easy and relatively risk-free strategy for reducing TMAO-related health risks. Studies investigating the effect of diet on intestinal microbiota and TMAO levels have largely focused on the Mediterranean diet and veganism, both of which are rich in plant-derived polyphenols, high in fiber, and associated with multifaceted health benefits ,155. EpiSimilarly, long-term adherence to a vegan diet is linked with a diminished capacity to biosynthesize TMAO from the dietary TMAO precursor L-carnitine , but stuPrevotella normally associated with non-Western diets low in processed foods [Prevotella abundance was also associated with an increase in short chain fatty acids and a rise in plasma phosphatidylcholine, indicative of a reduced availability of choline for TMA biosynthesis and a potential protective role in promoting metabolic health [‘Functional foods’ can exert beneficial effects on human health beyond their basic nutritional value. One example is given by prebiotics, defined as non-digestible food components selectively degraded by bacteria in the gut  that acted foods . Notablyc health . Likewisc health .Lactobacillus, reduced the levels of TMAO and attenuated the atherosclerosis phenotype of ApoE-/- mice fed a high-choline diet [Lactobacillus-boosting effects, accompanied by a reduction in the carnitine-induced increase in TMAO plasma levels in mice [Polyphenols are phytochemicals produced as secondary metabolites in plants and constitute another class of chemical compounds of dietary origin with extra health benefits, known for their potent gut microbiota modulating properties. Notably, dietary supplementation with resveratrol (a stilbenoid polyphenol) increased the abundance of ine diet . Flavono in mice . A recen in mice . HoweverLactobacillus paracasei F19 protected rats from oxidative stress-induced liver damage by restoring the intestinal barrier and microbiota diversity [M. smithii bacterial species was associated with a reduction in the TMAO concentration and attenuation of atherosclerosis in ApoE-/- mice [Enterobacter aerogenes is another bacterial strain that has been shown to effectively reduce plasma TMAO and cecal TMAO levels by shifting the ratio of commensals and pathobionts in mice on a high-choline diet [Another strategy for targeting TMAO is the use of probiotics  , with stiversity , while g-/- mice . Enterobine diet . While tine diet  and inteClostridioides difficile infection being the only human condition for which FMT is currently approved [C. difficile infection. In this regard, a small FMT clinical study has shown to reverse gut dysbiosis but failed to induce remission in patients with chronic active ulcerative colitis [A more recent approach to reversing dysbiosis is fecal microbiota transplantation (FMT), which is a type of fecal bacteriotherapy that involves complete replacement of the recipient’s indigenous microbiome with healthy microbiota from a carefully selected donor. Despite positive results in pre-clinical studies, the method has yielded limited clinical success with antibiotic-resistant approved . However colitis . Similar colitis . HoweverPharmacological agents designed to disrupt microbial metabolite biosynthesis cascades include a structural analogue of choline 3,3-Dimethyl-1-Butanol (DMB) and the anti-ischemic drug meldonium. DMB works to reduce TMAO levels and attenuate choline diet-induced atherosclerosis by inducing the non-lethal inhibition of TMA lyase in the gut, which is hypothesized to be associated with a reduced risk of resistance compared to antibiotic treatment, which causes considerably more selective pressure . DespiteTogether, dietary interventions, the use of prebiotics/ probiotics, FMT, and pharmacotherapy represent an exciting potential avenue for the treatment of human diseases associated with dysbiosis and elevated TMAO levels. However, the success of microbiome-targeting therapies is contingent on developing an improved mechanistic understanding of interactions between the gut microbiota and disease pathogenesis. There is clearly a need for better designed randomized double-blind human trials with longer follow-up periods and bigger sample sizes. On the other hand, promoting lifestyle modifications that improve gut health is of significance as evidence suggests that long-term dietary habits and the state of the microbiome pre-treatment are important determinants of clinical success.TMAO is a metabolite of dietary choline and is therefore dependent upon the consumption of precursors, host genetics, and gut microbial enzymatic activity. In this review, we described specific microbial enzymes involved in TMA production pathways as well as the microbes carrying the genes for these enzymes . It is wHowever, important questions remain. Archaea has not received the same level of attention as bacteria, despite potentially being important in nutrient metabolism including TMA production, as supported by Fu et al. . RegardiAnother emerging field is the role of choline as an epigenetic modifier of the genome by changing the availability of methyl-donors such as S-adenosylmethionine (SAM). SAM synthesis depends on dietary choline ingestion and modulates neuronal gene expression and brain function. Therefore, the identification and implementation of effective nutritional strategies early in life could improve cognition and mental health .Therefore, the study of the gut microbiome may significantly enhance our understanding of nutrient metabolism and specific pathways by which diet can influence health and cognition. This may facilitate the adoption of a individualized nutrition-based approach to target gut microbial structure and function as well as the potential to alter different metabolites such as TMAO production.


The penultimate effectors of the Hippo signaling pathways YAP and TAZ, are transcriptional co-activator proteins that play key roles in many diverse biological processes, ranging from cell proliferation, tumorigenesis, mechanosensing and cell lineage fate determination, to wound healing and regeneration. In this review, we discuss the regulatory mechanisms by which YAP/TAZ control stem/progenitor cell differentiation into the various major lineages that are of interest to tissue engineering and regenerative medicine applications. Of particular interest is the key role of YAP/TAZ in maintaining the delicate balance between quiescence, self-renewal, proliferation and differentiation of endogenous adult stem cells within various tissues/organs during early development, normal homeostasis and regeneration/healing. Finally, we will consider how increasing knowledge of YAP/TAZ signaling might influence the trajectory of future progress in regenerative medicine. YAP  and TAZ  are two homologous transcriptional co-activator proteins  Figure , which sIt is important to note that under physiological conditions, the distribution of YAP/TAZ is usually partially cytoplasmic and partially nuclear, being dependent on the relative activities of the various components of the core Hippo signaling cascade  and otheAs mentioned earlier, YAP/TAZ controls the expression of target genes primarily by acting as a co-activator of TEAD transcription factors, of which there are four isoforms in mammals (TEAD1-4) . The varUpstream signaling pathways regulating YAP/TAZ activity have been shown to mediate cellular interactions with a broad range of microenvironmental factors including (i) soluble bioactive ligands , (ii) biHence the focus of this review would be on the regulatory mechanisms by which YAP/TAZ regulate stem/progenitor cell differentiation into the major lineages that are of interest for therapeutic applications . The keyStem cells are defined by two key characteristics: (i) potency, which is the ability to give rise to multiple differentiated lineages, and (ii) capacity for self-renewal. Currently, four broad categories of stem cells are recognized: (i) adult stem cells, (ii) fetal stem cells, (iii) embryonic stem cells (ESCs), and (iv) induced pluripotent stem cells (iPSCs) derived by reprogramming somatic cells. Of these, ESCs and iPSCs are considered to be pluripotent and are collectively referred to as pluripotent stem cells. Adult and fetal stem cells, on the other hand, are considered to be only multipotent. It must be noted that adult stem cells are closely associated with cancer, and there is a widely accepted theory that cancer cells are in fact aberrant adult stem cells that have lost their capacity for proper regulation of apoptosis and the cell cycle . Indeed,in vitro and maintains the stem cell phenotype even under differentiation conditions. Subsequently, in vitro culture depend on the maintenance of YAP/TAZ activity through AKAP-Lbc/Rho GTPase signaling, which modulates actin microfilament organization. It must be noted that the self-renewal and maintenance of pluripotency in human ESCs involve a different mechanism based on bFGF (basic fibroblast factor) signaling, as opposed to LIF signaling in mouse ESCs . Hence, in vitro.Perhaps the most compelling evidence of the role of YAP/TAZ in the self-renewal and pluripotency of human pluripotent stem cells comes from studies on reprogramming adult somatic cells into iPSCs. Besides pluripotent stem cells, YAP/TAZ also play key roles in the expansion, self-renewal and maintenance of “stemness” of tissue-specific adult stem cells. Upon disease or injury, the normally quiescent adult stem cells resident within specific tissues are mobilized into a “transit-amplifying” stage, in which they undergo rapid and extensive proliferation as an undifferentiated intermediate, prior to terminal differentiation into functional lineages. Utilizing the continuously growing mouse incisor model, There is abundant evidence for key roles of YAP/TAZ in the osteogenic differentiation pathway of primary osteoblasts and various types of adult stem cells, especially bone marrow-derived MSCs. While TAZ activation is consistently associated with osteogenesis , the rolThe role of TAZ in the osteogenesis of MSCs was first reported by The role of YAP in osteogenesis is ambiguous. While some studies indicated that YAP activation enhances osteogenesis, other studies reported contrary results. In two separate studies by our research group, we found that YAP activation in rat bone marrow-derived MSCs is associated with osteogenesis . Pan et The findings of YAP/TAZ regulation of osteogenic differentiation is stimulated by various physical and biochemical stimuli. The most widely studied of these are substrate stiffness and surface topography. Generally, stiff and rigid substrates that are conducive to cell adhesion and spreading lead to increased formation of FAs and cytoskeletal stress fibers, which in turn facilitate nuclear translocation of YAP/TAZ through both Hippo-dependent and Hippo-independent mechanisms. This in turn promotes osteogenic differentiation whilst inhibiting adipogenesis, through simultaneous activation and inhibition of RUNX2 and PPARγ (peroxisome proliferator-activated receptor gamma) transcription, respectively . On the There is also evidence that a multitude of other signaling pathways are interwoven in substrate-stiffness-induced osteogenesis. per se (not adhesion area) did not have any effect on YAP/TAZ activity or cell differentiation.With regard to substrate surface topography, several studies have demonstrated conclusively that rough, patterned and fibrous substrates are more conducive to YAP/TAZ activation, which in turn enhances osteogenic differentiation. Besides substrate stiffness and surface topography, various biophysical stimuli also modulate osteogenesis via YAP/TAZ. These include mechanical stimuli, such as cyclic stretching , shear sThe regulation of osteogenesis by YAP/TAZ involves various intracellular signaling proteins that interact directly with YAP/TAZ. For example, Smad4  and polyin vitro revealed that YAP overexpression induces a negative feedback mechanism on the Hippo signaling pathway, which leads to suppression of TAZ activity. This in turn enhances PPARγ activation and increases adipogenesis  supplemented in the culture milieu upregulates transcription factor SLUG expression, which in turn activates YAP, thereby maintaining SIX2, PAX2, and SALL1 expression by nephron progenitors of the metanephric mesenchyme. In accordance with the putative key role of YAP in the proliferation and phenotype maintenance of nephron progenitors, Nephrons, the functional filtration units within the kidney, arise from mesenchymal progenitors, and YAP/TAZ and the Hippo signaling pathway control the delicate balance between self-renewal and differentiation of mesenchymal progenitors that give rise to functional nephrons. Nevertheless, the subsequent deactivation of YAP/TAZ through phosphorylation is required for differentiation into mature functional nephrons. Hence, it can be concluded that YAP/TAZ activation is required for proliferative expansion and phenotype maintenance of the nephron progenitor pool during both embryonic development and kidney regeneration, and that YAP/TAZ deactivation through phosphorylation is required for subsequent maturation of these nephron progenitors into functional nephrons.The formation of new blood vessels during the process of angiogenesis or vascularization is an extremely complex multi-step process that involves the coordinated migration and proliferation of endothelial cells (ECs) and smooth muscle cells (SMCs), followed by complex interactions among these cells and junction formation. Loss-of-function studies have confirmed that YAP/TAZ nuclear localization and activation are essential for initiating angiogenesis or vascularization. in vitro and in vivo. Besides promoting the differentiation and angiogenic sprouting of ECs, YAP/TAZ modulate vascular smooth muscle cell (VMSC) proliferation and differentiation . Wen et YAP/TAZ interact with various canonical signaling pathways during angiogenesis and vascularization. The most prominent of these is the VEGF signaling pathway, which has a critical role in angiogenesis/vascularization. Several studies have reported the convergence of YAP/TAZ and VEGF signaling via the actin cytoskeleton . For exain vitro and vascular morphogenesis within fibrin gels subcutaneously implanted into mice, whereas overexpression of PGC1α had the reverse effect. Hence, YAP-TEAD1 signaling induces mitochondrial biogenesis in ECs and stimulates angiogenesis through PGC1α. YAP also regulates angiogenesis via the STAT3 signaling pathway. Besides the VEGF signaling axis, YAP regulates angiogenesis via the peroxisome proliferator-activated receptor gamma co-activator 1-alpha (PGC1α) signaling pathway that controls glucose metabolism within the mitochondria of ECs . MammotoThe functions of the mammalian liver include detoxification and production of bile acids that facilitate the digestive process. The major cell lineages of the liver are hepatocytes that secrete bile acids, and biliary epithelial cells (cholangiocytes) that line the biliary ducts transporting bile acids into the gall bladder, and are responsible for modifying the bile secretions of hepatocytes. There is also a resident pool of adult mesenchymal stem cells within the post-natal liver, referred to as hepatic stellae cells, which contribute to liver regeneration upon disease or injury.In vitro studies have confirmed the in vivo findings on the role of YAP/TAZ in hepatic proliferation, differentiation and maturation. in vitro hepatocyte differentiation assay to show that YAP activity decreases, whereas Hippo pathway kinase activities (LATS1/2) increase, upon hepatic differentiation. Studies on the spatiotemporal pattern of YAP expression during embryonic liver development  and earlYAP/TAZ activation is required for the mobilization and proliferation of hepatic stellae cells, the endogenous resident adult stem cell pool within the liver that contributes to regeneration following disease or injury. Although hepatic stellae cell activation is essential for liver regeneration, it should be noted that excessive YAP activation in stellae cells, particularly during chronic liver damage, results in fibrosis and ultimately cirrhosis of the liver . Indeed,Postnatal skeletal muscle tissue contains a resident pool of adult stem cells known as satellite cells, which are normally quiescent. However, upon muscle injury, muscle satellite cells are activated to proliferate and generate myoblasts, which in turn give rise to terminally differentiated myotubes of muscle fibers. There is evidence that YAP activation in satellite cells and myoblasts involve mechanotransduction. Besides its role in satellite cell activation and myoblast proliferation, YAP activity also plays a role during the early phase of myoblast differentiation, by regulating mitochondrial structural remodeling. Nevertheless, further differentiation and maturation into myotubes require subsequent deactivation of YAP through phosphorylation. Interestingly, although TAZ, like YAP, plays a role in the proliferation of myoblasts, the two proteins have divergent functions in myoblast differentiation. Hence, in conclusion, YAP/TAZ activity is required for satellite cell activation and proliferation, as well as for the early phase of myoblast differentiation. In contrast, the roles of YAP and TAZ diverge during the later phase of myoblast differentiation and maturation into myotubes, which require TAZ activation and YAP deactivation.The role of YAP/TAZ in cardiomyogenesis was first revealed by studies on fetal heart development. Fetal cardiomyocytes undergo extensive proliferation that ends abruptly after birth, and these changes are strongly correlated with YAP activation and deactivation, respectively . PostnatSeveral studies have reported increased YAP/TAZ activation in diseased hearts, suggesting a role in heart repair and regeneration. In vitro stimuli that trigger cardiomyocyte proliferation through YAP activation include substrate rigidity and nanostructure  of the gastrointestinal tract consists of a single-cell layer of simple columnar epithelial cells, which is known as the intestinal epithelium. This serves two major functions: (i) absorption of nutrients and other useful substances into the body, and (ii) preventing the entry of toxic and harmful substances. Endogenous adult stem cells residing within the intestinal epithelium are called intestinal stem cells, and play a crucial role in the homeostasis and regeneration of the intestinal epithelium, which is a rapidly cycling tissue that renews every 4 to 5 days under normal conditions .As in most other tissues, YAP/TAZ are involved in the proliferation and differentiation of intestinal stem cells . Imajo eAs in the cases of other cell lineages, mechanosensing by YAP/TAZ also serves as a switch between quiescence, proliferation and differentiation in intestinal stem cells. Various signaling pathways regulating cell proliferation are known to be downstream targets of YAP/TAZ activation within intestinal stem cells, including the epiregulin/epidermal growth factor receptor (EGFr), β-catenin and Wnt signaling pathways . While tde novo expression of fetal markers, as well as suppression of markers for adult stem cells and differentiated cells. This can be recapitulated in vitro with a collagen 3D matrix supplemented with Wnt ligands, which serve to sustain endogenous YAP/TAZ activation and induce reprogramming of cell fate (Interestingly, ell fate .In recent years, there has been rapid increase in our knowledge of the signaling mechanisms by which YAP/TAZ regulate the development, homeostasis and regeneration of various tissue/organ lineages within the human body. Despite the great interest in the roles of YAP/TAZ in cancer and tumorigenesis, as much attention, if not more, has been focused on how YAP/TAZ maintains the delicate balance between quiescence, self-renewal, proliferation and differentiation of endogenous adult stem cells in various different tissue types during the processes of tissue regeneration and healing. In fact, we cannot really separate these two interrelated fields, given the widely accepted idea that cancers and tumors originate from aberrant adult stem cells .New insights into YAP/TAZ can potentially impact the stem cell and regenerative medicine field in many ways. Firstly, we can exploit our knowledge of the key roles of YAP/TAZ in maintaining stem cell proliferation and self-renewal during the regeneration process, to scale-up cell culture in order to obtain sufficient numbers of cells for therapeutic applications. Assaying YAP/TAZ activity by means of high-throughput screening technology can identify potential drug leads with the ability to enhance the proliferative and self-renewal capacity of various kinds of adult stem cell lineages.in vitro culture systems and protocols for differentiating adult, embryonic and induced pluripotent stem cells into specific lineages. More precise control of cell lineage determination would be advantageous for therapeutic applications in tissue engineering and regenerative medicine, as well as for non-therapeutic applications in pharmacology and toxicology screening assays. For example, based on existing knowledge that the level of YAP/TAZ activity in MSCs is crucial for specification into osteogenic, chondrogenic or adipogenic lineages, it may be possible to design improved in vitro culture systems for enhancing differentiation into a pre-selected lineage through more precise control of YAP/TAZ activity in the cells. This might be achieved by modifying the biomechanical and topographical properties of the substrata, or supplementation of small molecules or growth factors that modulate YAP/TAZ activity.Secondly, a better understanding of the role of YAP/TAZ in cell lineage fate determination would lead to improved in vitro, we may be able to fine-tune various biomechanical properties such as stiffness (Thirdly, we may exploit our new knowledge of YAP/TAZ signaling mechanisms to optimize the therapeutic properties of newly developed biomaterials. By performing YAP/TAZ assays on relevant cell lineages cultured on newly developed biomaterials tiffness  and topotiffness , enablinLastly, a newly emerging research frontier on how YAP/TAZ modulate stem cell function, are their novel roles in effecting epigenetic modifications to chromatin structure  and postBH and XZ contributed equally to writing most of the manuscript sections. DA prepared the figures and figure legends. YB, XL, and YW wrote some of the manuscript sections. MF and XD provided supervision and funding. 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.


The aging process is sexually dimorphic, with males having higher occurrence rates of cancer and facing a greater risk of mortality. Sexual dimorphism in the response to cellular damage may account for distinct phenotypic changes with age as they relate to the accumulation of cellular damage leading to cancer. Cellular senescence triggers permanent cell cycle arrest in order to protect against malignant growth. However, organismal senescence increases with age and is associated with the release of pro-inflammatory signals  known as the ‘senescence-associated-secretory-phenotype’ (SASP) that, if unchecked, accelerates tissue damage and creates a microenvironment ripe for cancer development. In this study, we hypothesized that sexual disparities in mortality and cancer prevalence stems from differences in the rate of accumulation of senescent cells in mice. Male and female C57BL/6J mice were fed ad libitum or subjected to 30% calorie restriction, a nutritional intervention known to delay the onset of various cancers and prevent senescent cell accumulation. Primary skin fibroblasts were collected longitudinally to allow measurement of cell proliferation, wound healing and the release of SASP factors. The results indicate that when compared to males, fibroblasts of CR-fed females showed significant improvements in cell growth rate, wound healing and SASP markers vs. AL controls. Work is underway to determine how sex influences cellular protective pathways. Thus, like other cell processes, cellular senescence is unequal between males and females and CR delays the emergence of the senescence phenotype.


The human brain organoids derived from pluripotent cells are a new class of three-dimensional tissue systems that recapitulates several neural epithelial aspects. Brain organoids have already helped efficient modeling of crucial elements of brain development and disorders. Brain organoids’ suitability in modeling glioma has started to emerge, offering another usefulness of brain organoids in disease modeling. Although the current state-of-the organoids mostly reflect the immature state of the brain, with their vast cell diversity, human brain-like cytoarchitecture, feasibility in culturing, handling, imaging, and tractability can offer enormous potential in reflecting the glioma invasion, integration, and interaction with different neuronal cell types. Here, we summarize the current trend of employing brain organoids in glioma modeling and discuss the immediate challenges. Solving them might lay a foundation for using brain organoids as a pre-clinical 3D substrate to dissect the glioma invasion mechanisms in detail. Glioma is an aggressive form of brain cancer characterized by poor prognosis , 2. DespGSCs growing in adherent 2D cultures allow studying specific aspects of tumor biology. Furthermore, these 2D GSCs cultures offer possibilities to conduct chemical and genetic screens, transcriptomics, proteomics, clonal or single-cell analyses without the influence of extrinsic signals. Interestingly, early-passaged GSCs retain the genetic and transcriptional signature of the parental tumor, and thus low-passaged GSCs are widely used as an accepted model of glioma . HoweverLast decade researches have seen impressive progress in understanding the fundamental aspects of glioma biology in terms of genetic factors that can elevate intra-tumor heterogeneity, differential behaviors of GSCs, and changes in their growth and invasion behaviors –18. NeveBrain organoids are an innovative experimental model of modern reductionists’ approach. It is noteworthy that the early developmental neurobiologists in the last century have already adapted similar deconstructive strategies to deconstruct the complexities of human brain development , 25. In In this modern era of brain organoids, two laboratories stand out to receive the full credit for their pioneering effort in generating the first kind of 3D neural tissues in vitro. Attempts by the Sasai and Vaccarino laboratories have generated self-organized structures from pluripotent cells that recapitulated several neuroretina and telencephalic development –30. ThesUnlike other cancer types, gliomas metastasize rarely but infiltrates diffusively into the brain , 46. ThuBian and colleagues explored the use of genetically engineered brain organoids and expressed oncogenes ectopically to develop tumors. The observations of ectopic growth of tumor-like mass in organoids did not sufficiently help assessing to what extent those cells’ invasion behaviors mirror the biological relevance . More reTo further optimize the GSCs invasion assay, Goranci-Buzhala et al. tested various parameters such as organoid’s age, GSC numbers, and the order of GSCs addition either single cells or spheres . Their aBased on the observations, it is intuitive that GSCs not only invade into the brain but also interact with the neuronal cells to integrate and to exert its effect to disrupt normal brain functions. To test this aspect, Goranci-Buzhala, and colleagues cultured organoid slices that could generate long-range axons growing as neuronal outgrowths . ImportaIn summary, even though brain organoid-based glioma invasion assays still at the level of development, the findings so far are encouraging that brain organoid tissues can serve as a 3D substrate for clinical GSCs. Nevertheless, the field requires significant refinements, which we outline in the following section. Table As described previously, stem cell-derived brain organoids is a reductionist approach containing mostly immature cell types of developing brain. Thus, these organoids perfectly fit to decode the rules that determine neural stem cell proliferation, differentiation, and primitive cortical plate formation in brain development. Glioma is a late-onset disorder, and thus more meaningful glioma modeling requires brain organoids harboring mature cell types of astrocytes, oligodendrocytes, myelinated neurons, and immune defense cell types of microglia. It is particularly important to employ organoids with a myriad of cell types because GSCs require a microenvironment. GSCs communicate, reprogram the surrounding cells, suppress the immune response, and remodel the extracellular matrix, events that all favor tumor progression. 3D tissues merely containing actively proliferating precursor cells may help obtain initial events but will not provide complex interactions occurring in glioma modeling.To date, several protocols that generate brain organoids allow the differentiation of astrocytes, mature neuronal cell types, and even surprisingly microglia-like cells , 55. MicAnother critical missing factor in the brain organoids is endothelial cells. It is known that GSCs reside preferentially in the vascular niches where they establish a spatial connection with endothelial cells that could affect their sensitivity to chemotherapy , 60. WheAnother critical development in organoid technology is to devise a method that can generate a large number of brain organoids with minimum inter-organoid variations. In our opinion, this effort is particularly critical when organoids are used for drug screening and personalized glioma invasion assays, both of which require statistically significant data. In this direction, using a large scRNA-seq dataset, Velasco et al. demonstrated a reliable reproducibility between individual organoids that they generated . AmendinThe outcome of the glioma invasion assay depends on the starting material. As discussed above, low-passaged GSCs are accepted models of glioma . HoweverBrain organoid models are beneficial for bridging the knowledge gap between in vitro cell cultures and in vivo animal studies. Even though GSCs are accepted models of glioma biology, GSCs may not faithfully reflect the characteristics of parent tumors. There are numerous reasons for it: (i) the culturing conditions can potentially out select a population, (ii) the artifacts of culturing conditions may suppress the heterogeneity of initial tumors, (iii) multiple passages may change the behaviors of GSCs. The glioma tumoroids can significantly address these pitfalls. When brain organoid-based glioma invasion assay is optimized with glioma tumoroids, it will be possible to scale up the reproducible assay with a reasonable turnaround time between four to six weeks after the surgery.Besides, high-resolution imaging and computer algorithm-assisted post-processing should be in place to generate a reliable invasion map of a given clinical sample. Establishing these measures will ensure implementing the assay in a clinical setting. It is noteworthy that brain organoid invasion assays already allow distinguishing the behavior of primary and recurrent GSCs . Thus, aBrain organoid-based invasion assays hold promise for personalized drug discovery programs. Traditional treatment for glioma is the surgical resection of naïve tumors followed by radiation in combination with temozolomide (TMZ). It may be possible that organoid-based invasion assays allow us to predict the drug sensitivity of the naïve GSCs before exposure to radiation and TMZ. It is important note that drugs targeting cell cycle pathways might also affect the proliferative cells of immature organoids. To avoid this situation, one could use mature old organoids or organoid-like structures assembled from post mitotic neurons and astrocytes . In the There is also a room for developing tools to conduct autologous glioma invasion assays. Currently used brain organoid-based invasion assays are performed in a homologous manner, meaning that patient-derived GSCs are grafted in different host brain organoids. Thus, another significant development is establishing autologous innovation assays by grafting patient derived GSCs into brain organoids generated from the same patient. Comparative invasion assays between homologous and autologous assays may reveal the role of diverse human brain microenvironments on tumor spread.The recently emerged 3D human brain organoids have unexpectedly offered us studying glioma biology in a new dimension, offering an excellent opportunity to visualize GSCs invasion in human brain-like tissues. With its neural stem cells, early and mature neurons, astrocytes, and microglial cells, brain organoids provide a suitable brain-like microenvironment for GSCs growth. Human brain organoids did not exist in the last decade, and organoid-based GSCs invasion assays were not there just three years ago. Hence, it is an exciting era for glioma researchers as one can map GSCs invasion in ex-vivo. However, as we discussed in the previous chapter, there are significant bottlenecks, and addressing them will allow us to model the vicious cycle of GSCs, such as the aggressive behavior of GSCs along white matter paths of the human brain (like corona radiates), peri-vascular invasion/niching and escape from immunological surveillance. From a clinical standpoint, optimizing the assays is crucial to identify responses on the sensitivity of a given tumor sample to irradiation or drug treatments within a few weeks of surgery. Such early responses will help predict patient survival after standard treatment and, hopefully, tailor personalized therapies. These innovations await future experiments.


Plant grafting is an ancient agricultural practice widely employed in crops such as woody fruit trees, grapes, and vegetables, in order to improve plant performance. Successful grafting requires the interaction of compatible scion and rootstock genotypes. This involves an intricate network of molecular mechanisms operating at the graft junction and associated with the development and the physiology of the scion, ultimately leading to improved agricultural characteristics such as fruit quality and increased tolerance/resistance to abiotic and biotic factors. Bidirectional transfer of molecular signals such as hormones, nutrients, proteins, and nucleic acids from the rootstock to the scion and vice versa have been well documented. In recent years, studies on rootstock-scion interactions have proposed the existence of an epigenetic component in grafting reactions. Epigenetic changes such as DNA methylation, histone modification, and the action of small RNA molecules are known to modulate chromatin architecture, leading to gene expression changes and impacting cellular function. Mobile small RNAs (siRNAs) migrating across the graft union from the rootstock to the scion and vice versa mediate modifications in the DNA methylation pattern of the recipient partner, leading to altered chromatin structure and transcriptional reprogramming. Moreover, graft-induced DNA methylation changes and gene expression shifts in the scion have been associated with variations in graft performance. If these changes are heritable they can lead to stably altered phenotypes and affect important agricultural traits, making grafting an alternative to breeding for the production of superior plants with improved traits. However, most reviews on the molecular mechanisms underlying this process comprise studies related to vegetable grafting. In this review we will provide a comprehensive presentation of the current knowledge on the epigenetic changes and transcriptional reprogramming associated with the rootstock–scion interaction focusing on woody plant species, including the recent findings arising from the employment of advanced—omics technologies as well as transgrafting methodologies and their potential exploitation for generating superior quality grafts in woody species. Furthermore, will discuss graft—induced heritable epigenetic changes leading to novel plant phenotypes and their implication to woody crop improvement for yield, quality, and stress resilience, within the context of climate change. Olea europaea .Grafting is an ancient agricultural propagation technique widely used to improve plant performance, in terms of yield, quality and resilience to abiotic and biotic stresses. It involves the merging of two genetically different plant parts, the rootstock, and the scion, in such a manner that the two parts join and grow as a single plant. Grafting is essentially dependent on the fundamental ability of wound healing in plants . Graftineuropaea , (b) to enotypes , (c) to enotypes . (d) To enotypes , (e) to in vivo techniques) will have a significant impact in the future. Indeed, various studies nowadays focus on adaptation through grafting approaches for example cucumber salinity tolerance of cucumber  or long non-coding RNAs (long ncRNAs). Epigenetic regulation plays a major role in all aspects of plant development such as proper vegetative growth, successful reproduction, fruit development, yield, fruit quality, and tolerance to environmental stresses . Moreovecucumber , while acucumber . CreatioMost of our knowledge on the molecular mechanisms of grafting has arisen from studies with model and vegetable species but in recent years studies on woody plants have been reported as well. Successful grafting requires a compatible and highly intricate interaction between the two parts of the graft, the scion, and the rootstock, which involves the concerted action of nutrient, hormonal, metabolic, transcriptional, and epigenetic pathways , 2. EstaNicotiana tabacum, N. benthamiana, and N. glauca (tree tobacco) and most notably, the transferred plastid DNA was stably inherited in regenerated plants pointing to evolutionary and agronomical implications  transcript -pumpkin (rootstock) graft, indicating graft–induced transfer from the pumpkin rootstock via the phloem to the cucumber scion , from scion to roots enhanced tuber production in potato grafts (BEL11 and BEL29 transcripts suppressed tuber growth in potato roots (AUX/IAA transcripts (encoding regulators of auxin transport) to the root tip, modified tobacco root architecture in Arabidopsis/tobacco hetero-grafts  across graft segments. Small RNAs comprise a category of 21–24 nucleotides (nt) RNAs of different origins that are directed either to chromatin DNA loci in the nucleus, or target homologous mRNA molecules in the cytoplasm. Extensive studies have demonstrated the essential and instrumental role of small RNAs in all aspects of plant growth and development and in both abiotic and biotic stress responses  heterografts were significantly altered depending on rootstock and rootstock-scion genotype combinations  analysis in a d plants . Similarg system . Importag system . Furtherprofiles . Recentlinations  and DNA inations . All theProgress has been achieved in recent years with respect to the effects of grafting on the improvement of woody plants and the mechanisms underlying this process at the physiological, genetic, and epigenetic level. Employment of advanced—omics technologies has contributed greatly on the genome-wide aspects of rootstock-scion interactions, genotype compatibility, and graft-induced transcriptional and epigenetic effects. Some examples describing these investigations in woody species of high economic value are presented below and are summarized in http://www.oiv.int).Viticulture is a vital sector of agriculture in many countries around the world. Global production of grape and wine amounts to 73.3 million tons of grape berry and 279 million hl of wine, annually , a soil-borne aphid that destroyed around four million of own-rooted vineyard hectares. The common commercial rootstocks currently used around the world were developed from native American Vitis species that have co-evolved with phylloxera and, as a result, they display a higher level of resistance  or Paulsen 1,103 rootstocks . InteresPlasmopara viticola as compared to the susceptible combination Gaglioppo/1103P. Interestingly, the 41B rootstock induced biotic response-related genes such as NBS and NBS-LRR type transcription factors and triggered a remarkable increase of stilbene synthase transcripts associated with ROS scavenging activity and oxidative stress prevention  is associated with differential protein accumulation involving photosynthesis, chlorophyll biosynthesis, amino acid metabolism, and precursor metabolite biosynthesis, as compared to the own-rooted plants .Finally, molecular factors determining the success of graft healing and union formation in relation to graft compatibility/incompatibility have been investigated in grapevine. During the process of union formation in Cabernet-Sauvignon heterografts, CS/RG, and CS/1103 Paulsen, transcriptional changes were detected at the graft interphases of both combinations and these were associated to cell wall modification, wound responses, hormonal signaling, and stress responses, all playing a role in the complex mechanism of graft healing and development . A very Rosaceae family members such as apple, pear, peach, and cherry which greatly contribute to a balanced and healthy diet  to the scion of apple cv. Fuji by in situ hybridization in the phloem. The same year the transport of gibberellic acid insensitive mRNA transcripts (GAI) was detected in both directions (upward and downward) from the graft union (Gibberellic acid insensitive transcripts was demonstrated across the graft union from a wild Pyrus betulaefolia cv. Bunge (rootstock) to a traditional local Chinese pear cultivar, Pyrus bretschneideri cv. Yali (scion) in a distance up to 50 cm (2 years old grafting tree) (GAI transcript was successfully tested by grafting a 35S:pear -GAI transgenic tobacco (Nicotiana tabacum L. cv. Samson.) to wild-type tobacco and detecting transmission of GAI mRNA through the graft union occurred soon as 15 days after grafting. Findings in arabidopsis demonstrate that trafficking of GAI RNA transcripts is mediated by specific RNA motifs (The underlying molecular mechanisms by which the rootstock controls scion growth and properties in ft union . Some prft union . In grafft union . These aft union . In a sing tree) . MoreoveA motifs ; it remaFew works have pointed out the involvement of long-distance signal molecules to biotic stress tolerance. Investigation of resistance mechanisms of three apple rootstocks to wooly apple aphid, showed that the resistance factors exist in the phloem tissue, implying the modulation of resistance via a long-distance signal . Kaja etPbWoxT1 from pear (Pyrus betulaefolia) interacts with a polypyrimidine tract binding protein PbPTB3 assisting in long distance transport in the phloem and possibly controlling flower development and growth (HMs) genes in flower induction in two apple varieties with contrasting flowering characteristics. Their expression patterns indicated that their up- or down-regulation contributed to different aspects of flowering. Although the two varieties were both grafted to the same rootstock, the relationship of grafting to the differential expression of the HMs was not investigated. Recent experiments have aimed to unravel the role of transmissible signals and epigenetic mechanisms on flower induction. In a recent study, it was demonstrated that a d growth . Fan et M. robusta rootstock grafted with scions of wild-type (WT) apple (M. spectabilis) and a more-branching (MB) mutant at the branching stage revealed that sugar metabolism and auxin and cytokinin signaling exert major effects on root growth and development of grafted apple  scion variety grafted on two rootstocks with different soil water extraction capacities, Rangpur Lime (RL) and Sunki Maravilha (SM), displayed differential graft-induced DNA methylation which were associated with enhanced acclimatization responses upon recurrent drought stress for the VO/SM combination . SimilarSQUAMOSA promoter binding protein-like (SPL) gene using different combinations . Results demonstrated that scion age significantly influenced grafted tree maturity through the control of miR156-SPL4-miR172 regulatory network. They also showed that the existence of leaves on cutting rootstocks provided grafting success and influence the abundance of miR156 and miR172 in the scion (he scion .Lycium chinense Mill.) to reveal their implication to diverse biological pathways within a distant-grafting system by using Illumina sequencing technology. Significant differences in miRNA expression of shoots and fruits were revealed comparing non-grafted to grafted tomato plants. The setting of fruits was found to be the stage with the highest abundance of miRNA transcripts (123 were up-regulated). Potential targets of differentially expressed miRNAs identified by in silico analysis were found to be mainly transcription factors involved in diverse metabolic and regulatory pathways. Global transcriptome changes have been detected also between compatible and incompatible graft combinations in Litchi chinensis (Carya illinoinensis) (An interesting study conducted by hinensis  and duriinensis) .Grafting has been used empirically also in clonal forestry section as a tool which primary targets transfer of juvenility properties to mature materials by establishing them onto young rootstocks in nurseries . PhenotyCedrela odorata L.) authors showed the significant effect of elite genotype grafting in juvenility induction in mature material . Transgenic cassava rootstocks expressing sRNAs in order to trigger RNA silencing in non-transgenic scions have successfully prevented the spreading of pomovirus Cassava brown streak disease (CBSD) (Prunus necrotic ringspot virus (PNRSV)  . Moreove (PNRSV) . Later o (PNRSV) .However not all of RNAi-based rootstocks can efficiently transfer the silencing molecules to non-transformed scions and confer resistance. In another study, hairpin siRNA derived from transgenic rootstocks of lime plants did not confer resistance to tristeza tree virus (CTV) in the non-transgenic scions, mainly due to the fact that the biggest fraction of siRNA was very quickly degraded .Terminal Flower1 (MdTFL1) gene, in order to induce early transition from juvenile to flowering stage in apple trees. However, when dsRNAiMdTFL1 transgenic rootstocks were grafted onto non-transgenic apple genotype “PinS,” the flower-inducing signal obtained after silencing of MdTFL1 gene was not transmittable  gene in a transgenic apple rootstock, responsible for enhanced freezing tolerance and growth inhibition, provoked reduced scion growth and delayed flowering with no evidence of phloem-transmissible PpCBF1 mRNA  induced early flowering in non-transgenic scions and the mobile signal responsible for this was a transmissible phytohormonal signal. Finally, in some woody tree species such as nut crops transgrafting has been applied very recently but with encouraging results, confirming its emerging role in tree breeding  between adult trees and seedlings from selfing as well as between mature donor trees and newly grafted trees  are more likely to be stably inherited across generations in woody perennials than vegetables due to their way of clonal propagation  which favors the stable transmission of new characters. It is anticipated that in coming years research will expand greatly in uncovering mechanisms associated with woody plant grafting and transgenerational heritable changes. This knowledge would be of utmost importance for generating and selecting superior grafts for woody crop improvement and for transmitting graft-induced beneficial traits to subsequent generations, in order to meet sustainability demands in a changing environment.AK coordinated author contributions, contributed with specific parts, and edited the manuscript. ET, EVA, EMA, MG, SM, and GD contributed with specific parts each. AD had the idea and edited the manuscript. 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.


Large-scale genomics research is costly, requiring significant resources for community engagement, participant recruitment, experimentation, and genomics data generation. Though the costs of genotyping and sequencing are decreasing, the large sample sizes required for genome wide association studies restrict such studies to researchers with significant funding and adequate resources. Until recently, these studies have been performed predominantly in European and other first world country populations, creating a bias in representation of global populations in public databases. Through a change in funding priorities for some major biomedical research funders, and a recognition of the need for diversity in genetic data, the balance has begun to shift, and under-represented populations are increasingly being included in genomics studies. Data from these under-represented populations have the potential to significantly alter our understanding of the genetic basis for human diseases in all populations, as they enable us to complete a picture which previously had major gaps. For example, inclusion of African populations, our oldest and most diverse populations, is providing important insights into human evolution and the origin of disease-related mutations.For this Research Topic, we sought high quality research papers describing novel insights into genetic and environmental factors that impact disease risk, expression, prognosis, and treatment in understudied populations in human genomics research. Topics could include Population genetics, genome wide association studies, epigenetics, pharmacogenomics, environmental risk factors for diseases or gene-environment interactions in diseases. The final topic issue has 19 published articles covering various diseases studied in African and other previously under-represented populations.Shaffer et al. describe capacity development efforts in Mali to increase the number of trained bioinformaticians and data scientists able to analyse and interpret large-scale genomics data on local populations. Some of the papers describe novel methods or evaluation of existing methods for working on complex populations. For example, Schurz, Müller et al. evaluated the accuracy of three different imputation methods for multi-way admixed populations, using the South African Colored population as an example. Their findings demonstrate the importance of using an appropriate imputation software and reference panel containing populations that accurately represent ancestral populations for admixed individuals.Though not reporting specific studies, Fatumo et al. report the first GWAS in a Ugandan population for multivariate blood cell count phenotypes. The authors used both univariate and multivariate approaches and demonstrated that the multivariate approach has larger power and identifies additional loci. They report that performing a joint analysis of correlated phenotype simultaneously can provide new insights into complex traits that may not be identified in separate univariate analyses. However, there was an observation that highly correlated traits may also inflate p-values. New candidate loci for several blood cell count parameters were found illustrating the need for conducting GWAS in non-European populations to better understand the genetics of blood cell physiology.Vorster et al. describe a multiplex ligation-dependent probe amplification analysis to determine the cause of spinal muscular atrophy (SMA) in Black South African patients. Though no significant pathogenic CNVs were detected, they found discordant copy numbers of exons which suggest complex rearrangements that may affect the SMN1 gene. Their study reiterates the fact that the genetic determinants of SMA in some individuals from this population group differ from those identified previously in other populations. Infectious diseases such as HIV and TB are prevalent in African populations, and mental illness is on the increase. Kalungi et al. sought to identify associations between relative telomere length and internalizing mental disorders, such as depressive disorders, anxiety disorders, and post-traumatic stress disorders, among young HIV-infected Ugandan individuals. A longer relative telomere length was found in individuals with these orders than in age- and sex-matched controls, and they concluded that though the length was not the cause of the disorders, the disorders were causing accelerated cellular aging.There are a number of articles reporting on studies in African populations, most commonly on cohorts in South Africa. Kinnear et al. reported the first TB host susceptibility genome-wide association study (GWAS) with a specific focus on sex-stratified autosomal analysis and the X chromosome. Although the results are only nominally indicative of association, it does highlight the significance of the X chromosome in TB susceptibility, and the importance of considering ascertainment bias in genotyping arrays when selecting appropriate genotyping tools for undertaking studies in understudied populations.In another Africa GWAS study, motivated by the epidemiological evidence that males are more affected by tuberculosis (TB) than females, Schurz, Joubert et al. systematically reviewed the important progress and promising opportunities in environmental health research in Africa. Literature describing harmful health effects of metals, pesticides, and dietary mold represented a context unique to Africa. However, cardiovascular and respiratory health endpoints impacted by air pollution were comparable to observations in other countries. Air pollution exposures unique to Africa were dust and specific occupational exposures. Investigations of environmental exposures with distinct routes of exposure, unique co-exposures and co-morbidities, combined with the extensive genomic diversity in Africa in the context of gene-environment studies may lead to the identification of novel mechanisms underlying complex disease and promising potential for translation to global public health.Looking at environmental impact, Boua et al. examined gene-smoking interactions with carotid intima-media thickness (cIMT) to identify potential drivers for atherosclerosis risk in West-African populations of the AWI-Gen Study. They identified new gene-smoking interaction variants for cIMT within the previously described RCBTB1 region and the novel regulatory region of TBC1D8. In silico functional analysis suggested the involvement of genes implicated in biological processes related to cell or biological adhesion and regulatory processes in gene-smoking interactions with cIMT.In line with this prospect, Nagar et al. surveyed pharmacogenomic variants in two populations in Colombia, Antioquia and Chocó, with differing ancestries. They found that some pharmacogenomic variants have unusually high minor allele frequencies and differentiation according to ancestral contributions. These included variants with toxicity and dosing implications. As a result, the authors developed a cost-effective allele-specific PCR assay to test for relevant variants to inform healthcare decisions. In another paper, O'Connell et al. investigated the potential role of regulatory genes in antipsychotic treatment response in South African schizophrenia patients. Seven candidate genes showed significant expression level changes and four variants within these genes were significantly associated with treatment response. Compared to previously reported studies, two of these variants are identified as lying within eQTLs that impact brain gene expression, providing promising evidence that these may potentially serve as biomarkers of antipsychotic treatment response in the future.Precision medicine and pharmacogenomics was a strong theme in several of the featured publications. In a paper from outside the African continent, All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Neighborhoods play a central role in healthy aging, with changes to neighborhoods having a profound impact on older adults’ ability to age in place. Using gentrification as an indicator of neighborhood change and applying the theoretical framework of the Environmental Press model , this study examined the relationship between changing environments, affordable housing, and environmental attributes that support and hinder the health and well-being of older adults. A qualitative, case-study approach was used to interview low-income, majority Black older adults in a gentrifying area of Washington DC. 32 individuals (16 in non-profit and 16 in for-profit affordable housing) aged 55 and older participated in semi-structured interviews on perceptions of gentrification, neighborhood change, and challenges and supports to aging in place. Transcripts were then analyzed using the framework method of analysis. Although participants generally reported that gentrification improved their neighborhood’s built environment, many attributed it to a decline in social capital. Affordable housing provided an ability to age in place, though participants expressed uncertainty over their long-term ability to age in the context of continuing change. These findings suggest that while the physical changes accompanying gentrification may support older adults’ ability to age in place, its detrimental impact on social capital further increases their risk for social isolation. While affordable housing may enable older adults to age in place, fostering a greater sense of permanence and well-being will require additional policies that both increase accessibility to the physical amenities provided by gentrification and preserve older adults’ social capital.


Regulatory RNAs contribute to gene expression control in bacteria. Antisense RNAs (asRNA) are a class of regulatory RNAs that are transcribed from opposite strands of their target genes. Typically, these untranslated transcripts bind to cognate mRNAs and rapidly regulate gene expression at the post-transcriptional level. In this article, we review asRNAs that modulate bacterial fitness and increase virulence. We chose examples that underscore the variety observed in nature including, plasmid- and chromosome-encoded asRNAs, a riboswitch-regulated asRNA, and asRNAs that require other RNAs or RNA-binding proteins for stability and activity. We explore how asRNAs improve bacterial fitness and virulence by modulating plasmid acquisition and maintenance, regulating transposon mobility, increasing resistance against bacteriophages, controlling flagellar production, and regulating nutrient acquisition. We conclude with a brief discussion on how this knowledge is helping to inform current efforts to develop new therapeutics. In bactA (sRNA) . RegulatA (sRNA) . asRNAs A (sRNA) .Initially, asRNAs were thought to be rare in bacteria, and the pervasive antisense transcription observed in microarray-based studies were assumed to be experimental artifacts . Even wiasRNAs have been shown to modulate bacterial pathogenicity by either regulating the expression of virulence genes  or by covia conjugation, a process that involves asRNAs. An example of this is F-like plasmids, which are part of a large group of conjugative plasmids frequently found in Escherichia coli and throughout Enterobacteriaceae . When the temperature rises to 37°C, the motility gene repressor mogR switches off flagella formation  and protection from bacteriophages in the production of live mucosal vaccines  and through the Rackham Merit Fellowship through the University of Michigan. RR is supported by NIH grants AI133023 and DE028409.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The introduction of clinical criteria for the operationalization of psychosis high risk provided a basis for early detection and treatment of vulnerable individuals. However, about two-thirds of people meeting clinical high-risk (CHR) criteria will never develop a psychotic disorder. In the effort to increase prognostic precision, structural and functional neuroimaging have received growing attention as a potentially useful resource in the prediction of psychotic transition in CHR patients. The present review summarizes current research on neuroimaging biomarkers in the CHR state, with a particular focus on their prognostic utility and limitations. Large, multimodal/multicenter studies are warranted to address issues important for clinical applicability such as generalizability and replicability, standardization of clinical definitions and neuroimaging methods, and consideration of contextual factors . Psychotic disorders pose a major challenge for psychiatric practice and public health, affecting several aspects of functioning and quality of life and being among the leading causes of disease burden worldwide . AntipsyIn the 1990s, a paradigm shift in psychosis conceptualization and research led to an increased focus on the early stages of psychotic disorders, including the period before the onset of overt symptoms . The intCurrently, diagnosis of a clinical high-risk state is usually made based on ultra-high-risk (UHR) criteria, requiring the presence of either (a) positive symptoms that are typical of psychotic disorders but of subthreshold severity  or duration , or (b) genetic high risk accompanied by functional decline (GRD) , 6. A coIndividuals meeting high-risk criteria are at substantially higher risk for developing a psychotic disorder compared with the general population , 8. MostGiven the above limitations of clinical high-risk criteria, there have been intensive efforts to improve specificity and predictive accuracy with respect to transition to psychosis using individual symptom profiles , 17, demGray matter abnormalities are probably the best studied neuroimaging biomarker in high-risk patients so far. Early MRI investigations used manual tracing to investigate regions of interest assumed to be affected in psychotic disorders. In contrast, more recent studies have mainly relied on automated approaches, which have the advantage of allowing group comparisons at the whole-brain level rather than specific regions , 21, thuSeveral volumetric differences in gray matter have been reported in high-risk patients compared with healthy controls, the most consistent being volume reductions in hippocampal/parahippocampal areas, cingulate cortex, as well as the medial and lateral frontal cortex and medial parietal cortex , 31–33. Beyond static comparisons, evidence suggests that the dynamic of changes in brain morphology over time might also be informative for predicting transition in CHR patients. CHR-T have been reported to exhibit greater longitudinal volume reduction and/or cortical thinning in frontal areas, including orbitofrontal, superior frontal, middle frontal, and prefrontal cortices , 52–54; White matter density and regional distribution are age-sensitive, and its development continues well into late adolescence and beyond . TherefoThe most widely used technique to assess white matter integrity is diffusion tensor imaging (DTI), which measures the diffusion of water molecules through tissues using T2-weighted images in diverse directions. Although volume-based measures of white matter integrity similar to those for gray matter are available, track-based statistics are much more popular due to their superior accuracy . The mosFA is generally lowered in multiple brain regions in patients with schizophrenia . CHR havUnfortunately, there are few and inconsistent findings regarding diffusivity indices as a complement to FA , 73, 74.Currently, most neuroimaging studies in psychiatry rely on traditional mass-univariate statistical approaches. While these approaches are useful for detecting features that differentiate between groups, they cannot be used to infer whether an individual patient belongs to a certain group  . TherefoFour studies have used support vector machines, a supervised machine-learning technique, to predict transition to psychosis in CHR patients based on neuroanatomical features –85, in pAll of the above studies yielded high classification accuracy (80–88%), with positive predictive values of 78–100% and negative predictive values of 80–90%, suggesting that machine-learning models might be a promising tool in increasing the reliability of transition prediction. However, certain limitations should be kept in mind when interpreting findings. The most important issue is overfitting, which occurs when a statistical model describes noise  in the data rather than genuine effects of interest ; this ren = 190) to develop age prediction models, and subsequently calculated the gap between model-predicted age and chronological age in CHR patients (n = 380); the Personalized Prognostic Tools for Early Psychosis Management study (PRONIA) implemented a similar approach on neurocognitive performance data of 36 healthy individuals and 48 CHR patients  have alsychosis .Electrophysiological methods such electroencephalography (EEG) and magnetoencephalography (MEG) have been less prominent than MRI-based techniques in early CHR research. However, advances in analysis techniques and the use of dense array configurations that help to enhance spatial resolution have renewed interest in EEG and MEG in recent years. Advantages of these methods are their excellent temporal resolution, which enables the investigation of the fast dynamics of neuronal interactions occurring on a millisecond time-scale , and theSeveral neurophysiological biomarkers for schizophrenia have been identified, including early and late event-related potentials (ERP) and other measures of neural oscillatory activity . ASince the introduction of clinical high-risk criteria in the 1990s, a huge body of structural and functional neuroimaging studies has advanced our understanding of mechanisms associated with risk for, and resilience to, psychotic disorders. Recent studies focus on the utility of structural and functional neuroimaging to improve prediction of outcomes in the individual patient. The findings summarized in the current review testify to the potential of neuroimaging tools as a complement to clinical assessment for quantifying the risk of transition to psychosis in CHR patients. However, several outstanding issues remain, which we address below.Neuroimaging data are collected on special equipment and undergo some extent of data preprocessing. Differences in acquisition parameters or preprocessing protocols may have nontrivial consequences on final results. Common examples include the effect of MRI scanner field strength and manufacturer , preprocComprehensive Assessment of At-Risk Mental States (CAARMS) , they ds (SIPS) . Moreoves (SIPS) ] has nots (SIPS) , 177, ors (SIPS) ] may affs (SIPS) . Thus, ds (SIPS) , 8, studAnother point to consider is that, even using the same diagnostic instrument, the CHR state is not a unitary concept. As detailed in “Introduction”, there are several CHR subgroups, which differ in terms of not only phenomenology but also prognosis . FurtherA major challenge in early psychosis neuroimaging research is that patients are at an age that is characterized, even in healthy individuals, by major neurodevelopmental changes in the brain. The most notable changes are gray matter loss and cortical thinning , 181 andTwo recent studies showcase the complex interactions of age with neuroanatomy and transition prediction. As detailed above, a larger gap between “brain age” and chronological age and was predictive of greater risk of transition to psychosis, was observed in the NAPLS 2 study; however, this effect was found only in patients diagnosed with CHR status at a young age , while it was not present in patients diagnosed at a later age . Anotherd = 0.22 to d = 0.58) . HoweveThe past two decades of CHR research were exceptionally productive, leading to substantial insights into illness mechanisms leading to psychotic disorders. Still, several issues need to be addressed to achieve clinical applicability, including standardization of clinical definitions and neuroimaging methods, increased focus on generalizability and replicability, and increased attention to contextual factors such as age and comorbidity. The “second wave” of CHR research we are currently experiencing is characterized by large, multimodal/multicenter studies that make use of advances in computing techniques to generate findings that will be useful for individualized prediction in the context of precision psychiatry.


The etiology of “dual harm”  is under-researched. Risk factors have mostly been investigated for each behavior separately. We aimed to examine adversities experienced between birth and age 15 years among adolescents and young adults with histories of self-harm and violent criminality, with a specific focus on dual harm. Three nested case-control studies were delineated using national interlinked Danish registers; 58,409 cases in total aged 15–35 were identified: 28,956 with a history of violent criminality (but not self-harm), 25,826 with a history of self-harm , and 3987 with dual-harm history. Each case was matched by date of birth and gender to 20 controls who had not engaged in either behavior. We estimated exposure prevalence for cases versus controls for each of the three behavior groups, and incidence rate ratios (IRRs). Experiencing five or more childhood adversities was more prevalent among individuals with dual-harm history  versus self-harm  and violence  histories. The highest IRRs for dual harm were linked with parental unemployment , parental hospitalization following self-harm  or assault , and parental violent criminality . Growing up in environments that are characterized by poverty, violence, and substance misuse, and experiencing multiple adversities in childhood, appear to be especially strongly linked with elevated dual-harm risk. These novel findings indicate potential etiologic pathways to dual harm.  There is a considerable body of published research concerning the characteristics of young people who have harmed themselves, and the determinants of this behavior are well known ,3,4,5,6.There is increasing understanding of risk factors that are common to people who self-harm and those who engage in interpersonal violence. For example, almost all types of parental mental illness were found to be risk factors for self-harm and violent criminality among cohort members in a Danish register-based study that examined both behavioral outcomes in the same national birth cohort . Low parThere is very little understanding of early life risk factors for the co-occurrence of dual harm. Recognition and treatment of early trauma is necessary to address the causes of externalized violence . UnresolUnderstanding the characteristics of people in the general population who engage in dual harm is vital for addressing the specific needs of this risky and vulnerable group. A systematic review of studies that have examined the relationship between violence and self-harm found that the occurrence of either of these harmful behaviors resulted in increased risk of the other one also occurring . That stFrom a national birth cohort we delineated three nested case-control studies, with cases defined as (i) persons with an episode of hospital-treated self-harm but no record of violent criminality, (ii) persons with a record of committing violent crime but not engaging in self-harm, and (iii) those with a history of both harmful behaviors—dual harm. Self-harm episodes and violent criminal offending were identified if they occurred after cohort members’ 15th birthdays. Each case was matched to twenty controls who had no history of either self-harm or violent criminality, selected randomly from all eligible individuals in each risk set. Cases and controls were matched by gender and by date of birth using incidence density sampling procedures ,25. MatcEach Danish resident is assigned a unique personal identification number, a comprehensive system that enables accurate linkage between multiple administrative registers with complete national population coverage. Each of the three case-control datasets was nested in a cohort of persons born in Denmark to native Danish parents during 1980–2000 and who were alive and residing in the country on their 15th birthdays (N = 1.08 million). Measurement of exposure status began at birth and ended on cohort members’ 15th birthdays. Self-harm episodes and violent crimes were measured from age 15 onwards. Therefore, all cohort members were aged between 15 and 35 years during the study’s observation period. th birthday, on the basis that the minimum age for criminal responsibility in Denmark is 15 years. Therefore, we included self-harm episodes and violent offenses from individuals’ 15th birthday and onwards.We were primarily interested in dual harm, which we defined as having a history of both self-harm and violent criminality after reaching age 15. We also examined self-harm  and violent criminality (without self-harm) as “single harm” comparator outcomes. From 1990, hospital-treated self-harm episodes included those resulting in admissions to general hospitals and psychiatric units, and from 1994 onwards it also included general hospital emergency department presentations and episodes treated in psychiatric unit outpatient clinics. These episodes were identified from the National Patient Register  and fromAdverse experiences occurring before a cohort members’ 15th birthday were examined, including factors pertaining to the individual  and those relating to their parents and family environment. Personal exposures were hospitalization following self-harm, assault, or serious accident. Family environment factors included: parental unemployment, low parental income and low parental educational attainment, younger maternal age, older paternal age, residential transience, sibling death, parental death, parental hospitalization due to self-harm, assault and serious accident, parental mental illness, parental violent criminality, and child–parent separation. The exposures were selected based on data availability within the national registers for factors related to childhood and family experiences.We fitted conditional logistic regression models to estimate relative risks for the three outcome categories of interest: (1) self-harm ; (2) violent criminality (but not self-harm); and 3) dual harm —separately within each respective nested case-control study. Cases in each outcome category were compared to controls with no record of self-harm or violent criminality as regards exposure prevalence for each childhood adversity examined. In the nested case-control study design, the exposure odds ratio estimate is equivalent to the incidence rate ratio, IRR . All analyses were performed using Stata Release 15 . This study did not require approval from the Danish National Committee on Health Research Ethics because it was conducted solely using registry data. Approval to conduct this study was granted by the Danish Data Protection Agency, the Danish Health Data Authority, and Statistics Denmark.The national birth cohort from which the three nested case-control studies were delineated included 1,226,589 Danish people. A total of 58,409 cases were identified, including 28,596 people with a history of violent criminality (but not of self-harm), 25,826 who had previously self-harmed , and 3987 with dual harm; i.e., a record of both harmful behaviors. The prevalence of most childhood risk factors was not significantly higher in the dual-harm group compared to the single harm groups . Two excMost of the family risk factors that we measured were more prevalent in the dual-harm group than in either of the single-harm groups, including parental unemployment, low parental income, and low parental educational attainment, younger maternal age, residential transience, parental death from external causes, hospitalization due to self-harm, assault or serious accident, parental substance misuse disorder and other mental disorder, parental violent criminality and child–parent separation . The preThe incidence rate ratios (IRRs) for self-harm and violent criminality were markedly raised for young people experiencing hospitalization for assault and serious accident . The risIncremental increases in the IRRs for violent criminality, self-harm, and dual harm were observed as the number of adversities that individuals were exposed to during their childhood increased . Whilst In this study, we found that young people with histories of violent criminality, self-harm and dual harm had raised prevalence of the adverse childhood experiences investigated. However, those who had engaged in dual harm had considerably higher prevalence for some of the examined risk factors, and they were also much more likely to have experienced multiple adversities whilst they were growing up compared to one or two such risk factors, versus their peers with single-harm histories. In terms of personal adverse experiences, prior hospitalization for assault predicted a particularly large risk elevation for subsequent dual harm. With respect to family adversity, we observed a substantial excess risk of dual harm among children whose parents were unemployed or had been diagnosed with a substance misuse disorder, who had engaged in violent criminality, or who had been hospitalized following self-harm or assault.We observed the highest risks of dual harm among children who had been hospitalized for assault during childhood or had grown up with parents who had subjected other people to violence, who had been the victim of assault or who had engaged in self-harm requiring hospital treatment. These experiences of violence victimization and exposure to parental violence were stronger predictors than other adverse experiences such as suffering a serious accident or the death of a parent from natural causes. Whilst we cannot infer direct causality between childhood exposure to interpersonal violence and subsequent elevated dual-harm risk, it is a marker that could potentially help explain the pathways to dual harm, which is currently a poorly understood phenomenon. Parental and childhood adversity, and especially the accumulation of multiple adverse experiences, increase self-harm risk in young people ,34. SimiOur findings build upon a number of studies. Investigators in the USA found that experiencing multiple and persistent adversities between ages 4 and 16 was associated with greater use of healthcare services and poorer self-reported health at age 18 . ParentaIn a recently published study examining early life predictors of dual harm in adolescents , those wAttempted suicide in under-24-year-olds was found to be associated with increased risks of violent crime and intimate partner abuse perpetration into mid-life, as well as a range of other damaging social and health outcomes . This hiThis is the first study to examine dual harm in the context of multiple psychosocial adverse events spanning birth to 15 years of age and follow-up into mid-adulthood, highlighting the enduring consequences of adverse childhood experiences. We utilized national interlinked Danish registry data, enabling the examination of multiple adverse childhood experiences with minimal risk of recall and reporting bias. A limitation of our study was that we could only identify persons who presented to hospital after harming themselves, ascertaining a relatively small proportion of all self-harm episodes that occur in the community . IndividOur findings have a number of implications. People engaging in dual harm should be identified as early as possible in educational, healthcare, social services, and criminal justice settings. Our findings highlight and further emphasize the need for thorough assessment and early intervention for people presenting to hospital with self-harm , includiChildren growing up alongside violence, poverty, and substance misuse are considerably more likely to engage in dual harm during late adolescence and early adulthood. Experiencing multiple adversities during childhood was associated with elevated risks of violent criminality and self-harm and an even higher dual-harm risk. Further research should examine the relationship between early life predictors of dual harm as certain combinations of experiences may be particularly harmful. Potentially protective factors should also be examined.