Metabolite Retinoic Acid Research Articles

Retinoic acid receptor alpha expression in endometrioid heterotopias and the endometrium of patients with endometriosis

BACKGROUND: Vitamin A is a fat-soluble compound, which is a biologically inactive food form and undergoes transformation by cytosolic alcohol dehydrogenases and microsomal retinol dehydrogenases into retinaldehyde, followed by oxidation by retinaldehyde dehydrogenases 1, 2 and 3 into retinoic acid. All-trans-retinoic acid has been identified as an active metabolite of retinoic acid, indirectly participating in various processes: differentiation, proliferation and apoptosis. Based on a number of studies, the role of retinoic acid and its receptors in various diseases has been proven. For example, the destruction of the RAR-β gene by the integration of the hepatitis B virus into its loci can be a factor in the development of human hepatocellular carcinoma. Subsequently, it was described that the characteristic point of interruption of translocation t(15:17), observed in acute promyelocytic leukemia, was located at the locus encoding retinoic acid receptor-α. As a result, in patients with acute promyelocytic leukemia, complete remission of the disease could often be achieved by treatment with high doses of all-trans-retinoic acid. Existing data on the ability of retinol to influence the processes of differentiation, proliferation, and apoptosis indicate that retinol may be involved in the pathogenesis of endometriosis, but the mechanisms of its effect on the disease are subject to further study. AIM: The aim of this study was to evaluate the retinoic acid receptor alpha expression in endometrioid heterotopias and the endometrium of patients with different endometriosis stages according to the Revised American Society for Reproductive Medicine classification. MATERIALS AND METHODS: This was a prospective study to determine the retinoic acid receptor alpha expression in the endometrium and endometrioid heterotopias of patients with endometriosis (n = 28) and of the control group (n = 10). RESULTS: The retinoic acid receptor alpha expression in endometrioid heterotopias was lower compared to the eutopic endometrium of patients with endometriosis and the endometrium of patients in the control group by 3.4 and two times, respectively (p 0.001; the Kruskal – Wallis method). CONCLUSIONS: The data obtained indicate the involvement of retinol in the pathogenesis of endometriosis. Despite the relative increase in retinol level in the peripheral blood and peritoneal fluid in patients with endometriosis compared to the control group, shown in our previous studies, the reduced retinoic acid receptor alpha expression in endometrioid heterotopias does not allow for full binding of vitamin A to the receptor and is accompanied by its improper metabolism.

Retinoic acid and TGF-β orchestrate organ-specific programs of tissue residency.

Tissue-resident memory T (TRM) cells are integral to tissue immunity, persisting in diverse anatomical sites where they adhere to a common transcriptional framework. How these cells integrate distinct local cues to adopt the common TRM cell fate remains poorly understood. Here, we show that whereas skin TRM cells strictly require transforming growth factor β (TGF-β) for tissue residency, those in other locations utilize the metabolite retinoic acid (RA) to drive an alternative differentiation pathway, directing a TGF-β-independent tissue residency program in the liver and synergizing with TGF-β to drive TRM cells in the small intestine. We found that RA was required for the long-term maintenance of intestinal TRM populations, in part by impeding their retrograde migration. Moreover, enhanced RA signaling modulated TRM cell phenotype and function, a phenomenon mirrored in mice with increased microbial diversity. Together, our findings reveal RA as a fundamental component of the host-environment interaction that directs immunosurveillance in tissues.

The pathogenic role of retinoid nuclear receptor signaling in cancer and metabolic syndromes.

The retinoid nuclear receptor pathway, activated by the vitamin A metabolite retinoic acid, has been extensively investigated for over a century. This study has resulted in conflicting hypotheses about how the pathway regulates health and how it should be pharmaceutically manipulated. These disagreements arise from a fundamental contradiction: retinoid agonists offer clear benefits to select patients with rare bone growth disorders, acute promyelocytic leukemia, and some dermatologic diseases, yet therapeutic retinoid pathway activation frequently causes more harm than good, both through acute metabolic dysregulation and a delayed cancer-promoting effect. In this review, we discuss controlled clinical, mechanistic, and genetic data to suggest several disease settings where inhibition of the retinoid pathway may be a compelling therapeutic strategy, such as solid cancers or metabolic syndromes, and also caution against continued testing of retinoid agonists in cancer patients. Considerable evidence suggests a central role for retinoid regulation of immunity and metabolism, with therapeutic opportunities to antagonize retinoid signaling proposed in cancer, diabetes, and obesity.

Boosting of retinol activity using novel lecithin: Retinol acyltransferase inhibitors.

Lecithin:retinol acyltransferase (LRAT) is the main enzyme catalysing the esterification of retinol to retinyl esters and, hence, is of central importance for retinol homeostasis. As retinol, by its metabolite retinoic acid, stimulates fibroblasts to synthesize collagen fibres and inhibits collagen-degrading enzymes, the inhibition of LRAT presents an intriguing strategy for anti-ageing ingredients by increasing the available retinol in the skin. Here, we synthesized several derivatives mimicking natural lecithin substrates as potential LRAT inhibitors. By exploring various chemical modifications of the core scaffold consisting of a central amino acid and an N-terminal acylsulfone, we explored 10 different compounds in a biochemical assay, resulting in two compounds with IC50 values of 21.1 and 32.7 μM (compounds 1 and 2), along with a simpler arginine derivative with comparative inhibitory potency. Supported by computational methods, we investigated their structure-activity relationship, resulting in the identification of several structural features associated with high inhibition of LRAT. Ultimately, we conducted an exvivo study with human skin, demonstrating an increase of collagen III associated with a reduction of the skin ageing process. In conclusion, the reported compounds offer a promising approach to boost retinol abundance in human skin and might present a new generation of anti-ageing ingredients for cosmetic application.

Mould allergen Alt a 1 spiked with the micronutrient retinoic acid reduces Th2 response and ameliorates Alternaria allergy in BALB/c mice.

We investigated the biological function of the mould allergen Alt a 1 as a carrier of micronutrients, such as the vitamin A metabolite retinoic acid (RA) and the influence of RA binding on its allergenicity invitro and invivo. Alt a 1-RA complex formation was analyzed in silico and invitro. PBMCs from Alternaria-allergic donors were stimulated with Alt a 1 complexed with RA (holo-Alt a 1) or empty apo-Alt a 1 and analyzed for cytokine production and CD marker expression. Serum IgE-binding and crosslinking assays to apo- and holo-protein were correlated to B-cell epitope analysis. Female BALB/c mice already sensitized to Alt a 1 were intranasally treated with apo-Alt a 1, holo-Alt a 1 or RA alone before measuring anaphylactic response, serum antibody levels, splenic cytokines and CD marker expression. In silico docking calculations and invitro assays showed that the extent of RA binding depended on the higher quaternary state of Alt a 1. Holo-Alt a 1 loaded with RA reduced IL-13 released from PBMCs and CD3+CD4+CRTh2 cells. Complexing Alt a 1 to RA masked its IgE B-cell epitopes and reduced its IgE-binding capacity. In a therapeutic mouse model of Alternaria allergy nasal application of holo-Alt a 1, but not of apo-Alt a 1, significantly impeded the anaphylactic response, impaired splenic antigen-presenting cells and induced IL-10 production. Holo-Alt a 1 binding to RA was able to alleviate Th2 immunity invitro, modulate an ongoing Th2 response and prevent anaphylactic symptoms invivo, presenting a novel option for improving allergen-specific immunotherapy in Alternaria allergy.

Retinoic acid enhances ovarian steroidogenesis by regulating granulosa cell proliferation and MESP2/STAR/CYP11A1 pathway

IntroductionOvarian steroidogenesis not only affects the embryonic development and pregnancy outcome, but also associates with many diseases in mammals and women. Exploring the nutrients and mechanisms influencing ovarian steroidogenesis is critical to maintaining the optimal reproductive performance, as well as guaranteeing body health. ObjectivesThis research aimed to explore the effect of retinol metabolism on ovarian steroidogenesis and the underlying mechanisms. MethodsComparative transcriptomic analysis of ovaries from normal and low reproductive performance sows were performed to identify the main causes leading to low fertility. The metabolites regulating steroid hormones synthesis were investigated in ovarian granulosa cells. Gene interference, overexpression, dual-luciferase reporter assays, chromatin immunoprecipitation and transcriptome analysis were further conducted to explore the underlying mechanisms of Aldh1a1 mediating ovarian steroidogenesis. ResultsTranscriptome analysis of ovaries from normal and low reproductive performance sows showed the significant differences in both retinol metabolism and steroid hormones synthesis, indicating retinol metabolism probably influenced steroid hormones synthesis. The related metabolite retinoic acid was furtherly proven a highly active and potent substance strengthening estrogen and progesterone synthesis in ovarian granulosa cells. For the first time, we revealed that retinoic acid synthesis in porcine and human ovarian granulosa cells was dominated by Aldh1a1, and required the assistance of Aldh1a2. Importantly, we demonstrated that Aldh1a1 enhanced the proliferation of ovarian granulosa cells by activating PI3K-Akt-hedgehog signaling pathways. In addition, Aldh1a1 regulated the expression of transcription factor MESP2, which targeted the transcription of Star and Cyp11a1 through binding to corresponding promoter regions. ConclusionOur data identified Aldh1a1 modulates ovarian steroidogenesis through enhancing granulosa cell proliferation and MESP2/STAR/CYP11A1 pathway. These findings provide valuable clues for improving ovarian health in mammals.

Subacute effects of the chlorinated flame retardant dechlorane 602 on intestinal microenvironment in mice.

Chlorinated flame retardant Dechlorane 602 (Dec 602) has been detected in daily food, indicating that it may pose a risk to intestinal health. The intestinal microenvironment plays an important role in intestinal health. Intestinal microbiota and metabolites are two important factors for maintaining the microenvironment. However, little is known about the effects of Dec 602 on intestinal microbiota and metabolites. We aimed to probe the effects of Dec 602 on the intestine by revealing the changes that Dec 602 caused to the intestinal microbiota and metabolites. Adult female C57BL/6 mice were exposed to Dec 602 (low/high doses: 1.0/10.0μg/kg body weight per day) orally for 7 consecutive days, and sacrificed after 7days of recovery. The composition of colonic microbiota was measured by 16S rRNA gene sequencing, and the colonic metabolites were determined by LC-ESI-MS/MS. Finally, the effects of Dec 602 on the colon were validated by histopathological analysis. The intestinal microbiota composition was altered toward a pro-inflammatory status after exposure to Dec 602. Dec 602 exposure also up-regulated oxidative metabolites (glutathione disulfide, taurine and retinoic acid) and pro-inflammatory metabolites (prostaglandin E2). On the other hand, antioxidative metabolites (s-adenosylmethionine and 11-cis-retinol) and anti-inflammatory metabolites (alpha-linolenic acid, eicosapentaenoic acid and docosahexaenoic acid) were down-regulated after exposure to Dec 602. Infiltration of lymphocytes in the colonic lamina propria was observed in the mice treated with Dec 602 for 7days, and it was not recovered after another 7days without further treatment. Dec 602 interfered with the colonic microbiota and metabolome, and exhibited inflammatory features. Histopathological studies confirmed that Dec 602 exposure did induce colonic inflammation.

Retinoic acid promotes fibrinolysis and may regulate polyp formation

Patients with aspirin-exacerbated respiratory disease (AERD) regularly exhibit severe nasal polyposis. Studies suggest that chronic rhinosinusitis with nasal polyps (CRSwNP) is characterized by excessive fibrin deposition associated with a profound decrease in epithelial tissue plasminogen activator (tPA). Retinoids, including vitamin Aand its active metabolite retinoic acid (RA), are necessary for maintaining epithelial function and well-known inducers of tPA in endothelial cells. This study sought to determine whether endogenous retinoids are involved in NP pathophysiology and disease severity in patients with CRSwNP and AERD. NP tissue was collected from patients with AERD or CRSwNP, and concentrations of retinoids and fibrinolysis markers were measured using ELISA. Normal human bronchial epithelial cells were stimulated alone or in combination with RA and IL-13 for 24 hours. This study observed lower retinoid levels in nasal polyps of patients with AERD than those with CRSwNP or healthy controls (P< .01). Levels of the fibrin-breakdown product d-dimer were the lowest in AERD polyps (P< .01), which is consistent with lower tPA expression (P< .01). Invitro, all-trans RA upregulated tPA levels in normal human bronchial epithelial cells by 15-fold and reversed the IL-13-induced attenuation of tPA expression in cultured cells (P< .01). RA, a potent inducer of epithelial tPA invitro, is reduced in tissue from patients with AERD, a finding that may potentially contribute to decreased levels of tPA and fibrinolysis in AERD. RA can induce tPA in epithelial cells and can reverse IL-13-induced tPA suppression invitro, suggesting the potential utility of RA in treating patients with CRSwNP and/or AERD.

Regenerative Drug Discovery Using Ear Pinna Punch Wound Model in Mice.

The ear pinna is a complex tissue consisting of the dermis, cartilage, muscles, vessels, and nerves. Ear pinna healing is a model of regeneration in mammals. In some mammals, including rabbits, punch wounds in the ear pinna close spontaneously; in common-use laboratory mice, they remain for life. Agents inducing ear pinna healing are potential regenerative drugs. We tested the effects of selected bioactive agents on 2 mm ear pinna wound closure in BALB/c mice. Our previous research demonstrated that a DNA methyltransferase inhibitor, zebularine, remarkably induced ear pinna regeneration. Although experiments with two other demethylating agents, RG108 and hydralazine, were unsuccessful, a histone deacetylase inhibitor, valproic acid, was another epigenetic agent found to increase ear hole closure. In addition, we identified a pro-regenerative activity of 4-ketoretinoic acid, a retinoic acid metabolite. Attempts to counteract the regenerative effects of the demethylating agent zebularine, with folates as methyl donors, failed. Surprisingly, a high dose of methionine, another methyl donor, promoted ear hole closure. Moreover, we showed that the regenerated areas of ear pinna were supplied with nerve fibre networks and blood vessels. The ear punch model proved helpful in testing the pro-regenerative activities of small-molecule compounds and observations of peripheral nerve regeneration.

Vitamin A Status Is More Commonly Associated With Symptoms and Neurodevelopment in Boys With Autism Spectrum Disorders-A Multicenter Study in China.

BackgroundAutism spectrum disorder (ASD) is a neurodevelopmental disorder, and show a striking male bias in prevalence. Vitamin A (VA) is essential for brain development, and abnormalities in its metabolite retinoic acid are associated with the pathophysiology of ASD. This national multicenter study was conducted to investigate the relationship between serum VA level and core symptoms in ASD children and whether there are still sex differences.MethodA total of 1,300 children with ASD and 1,252 typically-developing (TD) controls aged 2–7 years old from 13 cities in China were enrolled in this study. The symptoms of children with ASD were evaluated by the Autism Behavior Checklist (ABC), Social Responsiveness Scale (SRS), and Childhood autism rating scale (CARS). The neurodevelopmental level of the children was evaluated with the revised Children Neuropsychological and Behavior Scale (CNBS-R2016). The serum level of VA was measured by high-performance liquid chromatography (HPLC).ResultsThe serum VA level in children with ASD was significantly lower than that in TD children, especially in boys with ASD. Furthermore, VA levels in male children with ASD were lower than those in female children with ASD. In addition, we found that serum VA level was negatively correlated the SRS, CARS and communication warming behavior of CBNS-R2016 scores in boys with ASD. In terms of developmental quotients, serum VA level was positively associated with the general quotient, language quotient, gross motor quotient and personal-social quotient of boys with ASD, but no difference was found in girls with ASD.ConclusionsASD children, especially boys, have lower serum VA levels than TD children. Moreover, serum VA status is more commonly associated with clinical symptoms and neurodevelopment in boys with ASD.

Role of Retinoid X Receptors (RXRs) and dietary vitamin A in Alzheimer's disease: Evidence from clinicopathological and preclinical studies

BackgroundVitamin A (VitA), via its active metabolite retinoic acid (RA), is critical for the maintenance of memory function with advancing age. Although its role in Alzheimer's disease (AD) is not well understood, data suggest that impaired brain VitA signaling is associated with the accumulation of β-amyloid peptides (Aβ), and could thus contribute to the onset of AD. MethodsWe evaluated the protective action of a six-month-long dietary VitA-supplementation (20 IU/g), starting at 8 months of age, on the memory and the neuropathology of the 3xTg-AD mouse model of AD (n = 11-14/group; including 4–6 females and 7–8 males). We also measured protein levels of Retinoic Acid Receptor β (RARβ) and Retinoid X Receptor γ (RXRγ) in homogenates from the inferior parietal cortex of 60 participants of the Religious Orders study (ROS) divided in three groups: no cognitive impairment (NCI) (n = 20), mild cognitive impairment (MCI) (n = 20) and AD (n = 20). ResultsThe VitA-enriched diet preserved spatial memory of 3xTg-AD mice in the Y maze. VitA-supplementation affected hippocampal RXR expression in an opposite way according to sex by tending to increase in males and decrease in females their mRNA expression. VitA-enriched diet also reduced the amount of hippocampal Aβ40 and Aβ42, as well as the phosphorylation of tau protein at sites Ser396/Ser404 (PHF-1) in males. VitA-supplementation had no effect on tau phosphorylation in females but worsened their hippocampal Aβ load. However, the expression of Rxr-β in the hippocampus was negatively correlated with the amount of both soluble and insoluble Aβ in both males and females.Western immunoblotting in the human cortical samples of the ROS study did not reveal differences in RARβ levels. However, it evidenced a switch from a 60-kDa-RXRγ to a 55-kDa-RXRγ in AD, correlating with ante mortem cognitive decline and the accumulation of neuritic plaques in the brain cortex. ConclusionOur data suggest that (i) an altered expression of RXRs receptors is a contributor to β-amyloid pathology in both humans and 3xTg-AD mice, (ii) a chronic exposure of 3xTg-AD mice to a VitA-enriched diet may be protective in males, but not in females.

Vitamin A and retinoic acid accelerate the attenuation of intestinal adaptability upon feeding induced by high-fat diet in mice

With its unique cellular plasticity, the small intestinal mucosa exhibits efficient adaptability upon feeding. However, little is known about the effect of high-fat diet (HFD) feeding on this adaption and its underlying mechanism. Herein, we demonstrated that the cell proliferation ability, mitochondrial morphology, and global transcriptomic profile of the small intestine exhibited a prominent discrepancy between the fasted and refed state in mice, which were markedly attenuated by long-term HFD feeding. The retinol (Vitamin A, VA) metabolism pathway was dramatically affected by HFD feeding in the small intestine. Both VA and its active metabolite retinoic acid (RA), with the administration of lipid micelles, promoted the expression of genes involved in lipid absorption and suppressed the expression of genes involved in the cell proliferation of intestinal organoids. Via chip-qPCR and RT-qPCR, genes involved in lipid metabolism and cell proliferation were target genes of RARα/RXRα in small intestinal organoids treated with RA and lipid micelles. The role of VA in the in vivo attenuation of intestinal adaptability, in response to HFD, was evaluated. Mice were fed a normal chow diet, HFD, or HFD diet supplemented with additional 1.5-fold VA for 12 weeks. VA supplementation in HFD accelerated the attenuation of intestinal adaptability upon feeding induced by HFD, promoted lipid absorption gene expression, and increased body weight and serum cholesterol levels. In conclusion, the discrepancy of the small intestine between the fasted and refed state was dramatically attenuated by HFD feeding, in which VA and RA might play important roles.

Normalization of Enzyme Expression and Activity Regulating Vitamin A Metabolism Increases RAR-Beta Expression and Reduces Cellular Migration and Proliferation in Diseases Caused by Tuberous Sclerosis Gene Mutations.

BackgroundMutation in a tuberous sclerosis gene (TSC1 or 2) leads to continuous activation of the mammalian target of rapamycin (mTOR). mTOR activation alters cellular including vitamin A metabolism and retinoic acid receptor beta (RARβ) expression. The goal of the present study was to investigate the molecular connection between vitamin A metabolism and TSC mutation. We also aimed to investigate the effect of the FDA approved drug rapamycin and the vitamin A metabolite retinoic acid (RA) in cell lines with TSC mutation.MethodsExpression and activity of vitamin A associated metabolic enzymes and RARβ were assessed in human kidney angiomyolipoma derived cell lines, primary lymphangioleiomyomatosis (LAM) tissue derived LAM cell lines. RARβ protein levels were also tested in primary LAM lung tissue sections. TaqMan arrays, enzyme activities, qRT-PCRs, immunohistochemistry, immunofluorescent staining, and western blotting were performed and analysed. The functional effects of retinoic acid (RA) and rapamycin were tested in a scratch and a BrDU assay to assess cell migration and proliferation.ResultsMetabolic enzyme arrays revealed a general deregulation of many enzymes involved in vitamin A metabolism including aldehyde dehydrogenases (ALDHs), alcohol dehydrogenases (ADHs) and Cytochrome P450 2E1 (CYP2E1). Furthermore, RARβ downregulation was a characteristic feature of all TSC-deficient cell lines and primary tissues. Combination of the two FDA approved drugs -RA for acute myeloid leukaemia and rapamycin for TSC mutation- normalised ALDH and ADH expression and activity, restored RARβ expression and reduced cellular proliferation and migration.ConclusionDeregulation of vitamin A metabolizing enzymes is a feature of TSC mutation. RA can normalize RARβ levels and limit cell migration but does not have a significant effect on proliferation. Based on our data, translational studies could confirm whether combination of RA with reduced dosage of rapamycin would have more beneficial effects to higher dosage of rapamycin monotherapy meanwhile reducing adverse effects of rapamycin for patients with TSC mutation.

Asthma-Associated Long TSLP Inhibits the Production of IgA.

Thymic stromal lymphopoietin (TSLP) contributes to asthmatic disease. The concentrations of protective IgA may be reduced in the respiratory tract of asthma patients. We investigated how homeostatic short TSLP (shTSLP) and asthma-associated long TSLP (loTSLP) regulate IgA production. B cells from healthy donors were stimulated in the presence or absence of shTSLP or loTSLP; the concentrations of IgA, IgM, IgE, and IgG antibodies were determined in cell culture supernatants; and B cells were analyzed by flow cytometry. LoTSLP, but not shTSLP, suppressed the secretion of IgA but not of IgE. The type 2 cytokine IL-4, which in addition to loTSLP contributes to asthmatic disease, did not affect the production of IgA or the frequency of IgA+ B cells. Instead, IL-4 increased IgG production, especially of the subclasses IgG2 and IgG4. LoTSLP inhibited IgA secretion by sorted memory B cells but not by naïve B cells. Although loTSLP inhibited IgA production, the vitamin A metabolite retinoic acid promoted the secretion of IgA, also in the presence of loTSLP, suggesting that vitamin A may promote IgA production in asthma. Our data demonstrate that asthma-associated loTSLP negatively regulates the secretion of IgA, which may negatively impact the surveillance of mucosal surfaces in asthma.

Multi-species transcriptome meta-analysis of the response to retinoic acid in vertebrates and comparative analysis of the effects of retinol and retinoic acid on gene expression in LMH cells

BackgroundRetinol (RO) and its active metabolite retinoic acid (RA) are major regulators of gene expression in vertebrates and influence various processes like organ development, cell differentiation, and immune response. To characterize a general transcriptomic response to RA-exposure in vertebrates, independent of species- and tissue-specific effects, four publicly available RNA-Seq datasets from Homo sapiens, Mus musculus, and Xenopus laevis were analyzed. To increase species and cell-type diversity we generated RNA-seq data with chicken hepatocellular carcinoma (LMH) cells. Additionally, we compared the response of LMH cells to RA and RO at different time points.ResultsBy conducting a transcriptome meta-analysis, we identified three retinoic acid response core clusters (RARCCs) consisting of 27 interacting proteins, seven of which have not been associated with retinoids yet. Comparison of the transcriptional response of LMH cells to RO and RA exposure at different time points led to the identification of non-coding RNAs (ncRNAs) that are only differentially expressed (DE) during the early response.ConclusionsWe propose that these RARCCs stand on top of a common regulatory RA hierarchy among vertebrates. Based on the protein sets included in these clusters we were able to identify an RA-response cluster, a control center type cluster, and a cluster that directs cell proliferation. Concerning the comparison of the cellular response to RA and RO we conclude that ncRNAs play an underestimated role in retinoid-mediated gene regulation.

Retinoic acid–responsive CD8 effector T cells are selectively increased in IL-23–rich tissue in gastrointestinal GVHD

AbstractGastrointestinal (GI) graft-versus-host disease (GVHD) is a major barrier in allogeneic hematopoietic stem cell transplantation (allo-HSCT). The metabolite retinoic acid (RA) potentiates GI-GVHD in mice via alloreactive T cells expressing the RA receptor-α (RARα), but the role of RA-responsive cells in human GI-GVHD remains undefined. Therefore, we used conventional and novel sequential immunostaining and flow cytometry to scrutinize RA-responsive T cells in tissues and blood of patients who had received allo-HSCT and to characterize the impact of RA on human T-cell alloresponses. Expression of RARα by human mononuclear cells was increased after exposure to RA. RARαhi mononuclear cells were increased in GI-GVHD tissue, contained more cellular RA-binding proteins, localized with tissue damage, and correlated with GVHD severity and mortality. By using a targeted candidate protein approach, we predicted the phenotype of RA-responsive T cells in the context of increased microenvironmental interleukin-23 (IL-23). Sequential immunostaining confirmed the presence of a population of RARαhi CD8 T cells with the predicted phenotype that coexpressed the effector T-cell transcription factor T-bet and the IL-23–specific receptor (IL-23R). These cells were increased in GI- but not skin-GVHD tissues and were also selectively expanded in the blood of patients with GI-GVHD. Finally, functional approaches demonstrated that RA predominantly increased alloreactive GI-tropic RARαhi CD8 effector T cells, including cells with the phenotype identified in vivo. IL-23–rich conditions potentiated this effect by selectively increasing β7 integrin expression on CD8 effector T cells and reducing CD4 T cells with a regulatory cell phenotype. In summary, we have identified a population of RA-responsive effector T cells with a distinctive phenotype that is selectively expanded in human GI-GVHD and that represents a potential new therapeutic target.

Quantification of All-Trans Retinoic Acid by Liquid Chromatography-Tandem Mass Spectrometry and Association with Lipid Profile in Patients with Type 2 Diabetes.

Retinoic acids are vitamin A metabolites that have numerous essential functions in humans, and are also used as drugs to treat acne and acute promyelocytic leukemia. All-trans retinoic acid (atRA) is the major occurring metabolite of retinoic acid in humans. This study provides a sensitive and specific liquid chromatography–tandem mass spectrometry approach in order to quantify atRA in human plasma samples. The isolation of atRA by hyperacidified liquid–liquid extraction using hexane and ethyl acetate resulted in a recovery of 89.7 ± 9.2%. The lower limit of detection was 20 pg·mL−1, and 7 point calibration displayed good linearity (R2 = 0.994) in the range of 50–3200 pg mL−1. Selectivity was guaranteed by the use of two individual mass transitions (qualifier and quantifier), and precision and accuracy were determined intraday and interday with a coefficient variation of 9.3% (intraday) and 14.0% (interday). Moreover, the method could be used to isolate atRA from hyperlipidemic samples. Applying this method to plasma samples from patients with poorly controlled Type 2 diabetes significantly decreased atRA plasma levels as compared to those of the healthy controls. In addition, atRA concentrations were highly associated with increased low-density lipoprotein (LDL) and decreased high-density lipoprotein (HDL) cholesterol levels.

LRAT coordinates the negative-feedback regulation of intestinal retinoid biosynthesis from β-carotene

AbstractThere is increasing recognition that dietary lipids can affect the expression of genes encoding their metabolizing enzymes, transporters, and binding proteins. This mechanism plays a pivotal role in controlling tissue homeostasis of these compounds and avoiding diseases. The regulation of retinoid biosynthesis from β-carotene (BC) is a classic example for such an interaction. The intestine-specific homeodomain transcription factor (ISX) controls the activity of the vitamin A-forming enzyme β-carotene oxygenase-1 in intestinal enterocytes in response to increasing concentration of the vitamin A metabolite retinoic acid. However, it is unclear how cells control the concentration of the signaling molecule in this negative-feedback loop. We demonstrate in mice that the sequestration of retinyl esters by the enzyme lecithin:retinol acyltransferase (LRAT) is central for this process. Using genetic and pharmacological approaches in mice, we observed that in LRAT deficiency, the transcription factor ISX became hypersensitive to dietary vitamin A and suppressed retinoid biosynthesis. The dysregulation of the pathway resulted in BC accumulation and vitamin A deficiency of extrahepatic tissues. Pharmacological inhibition of retinoid signaling and genetic depletion of the Isx gene restored retinoid biosynthesis in enterocytes. We provide evidence that the catalytic activity of LRAT coordinates the negative-feedback regulation of intestinal retinoid biosynthesis and maintains optimal retinoid levels in the body.

Multiple Vaccinations and the Enigma of Vaccine Injury.

A growing number of vaccines are administered at the same time or in close succession, increasing the complexity of assessing vaccine safety. Individual vaccines are assumed to have no other effect than protection against the targeted pathogen, but vaccines also have nonspecific and interactive effects, the outcomes of which can be beneficial or harmful. To date, no controlled trials and very few observational studies have determined the impact of vaccination schedules on overall health. The balance of the risks and benefits from mass vaccination therefore remains uncertain. Recent studies worryingly suggest links between multiple vaccinations and increased risks of diverse multisystem health problems, including allergies, infections, and neuropsychiatric or neurodevelopmental disorders. Here, we propose that, in susceptible persons, multiple vaccinations activate the retinoid cascade and trigger apoptotic hepatitis, leading to cholestatic liver dysfunction, in which stored vitamin A compounds (retinyl esters and retinoic acid) enter the circulation in toxic concentrations; this induces endogenous forms of hypervitaminosis A, with the severity of adverse outcomes being directly proportional to the concentration of circulating retinoids. In very low concentrations, vitamin A and its major metabolite retinoic acid contribute to immune function and to the process of immunization, whereas excess vitamin A increases the risk of adverse events, including common “side-effects” as well as chronic adverse outcomes. The increasing rates of allergy, ear infections, and neurodevelopmental disorders (NDDs) in countries with high rates of vaccination could be related to mass vaccination and to its impact on liver function and vitamin A metabolism, collectively representing endogenous manifestations of hypervitaminosis A. Further studies of health outcomes in vaccinated and unvaccinated groups are urgently needed, to increase understanding of the pathophysiology and treatment of vaccine injury, to identify the risk factors and screen for vaccine injury, to inform public health policy on potential hazards related to vaccination schedules, and to optimize the safety and benefits of vaccines.

Dietary Glucose Consumption Promotes RALDH Activity in Small Intestinal CD103+CD11b+ Dendritic Cells

Retinal dehydrogenase (RALDH) enzymatic activities catalyze the conversion of vitamin A to its metabolite Retinoic acid (RA) in intestinal dendritic cells (DCs) and promote immunological tolerance. However, precise understanding of the exogenous factors that act as initial trigger of RALDH activity in these cells is still evolving. By using germ-free (GF) mice raised on an antigen free (AF) elemental diet, we find that certain components in diet are critically required to establish optimal RALDH expression and activity, most prominently in small intestinal CD103+CD11b+ DCs (siLP-DCs) right from the beginning of their lives. Surprisingly, systematic screens using modified diets devoid of individual dietary components indicate that proteins, starch and minerals are dispensable for this activity. On the other hand, in depth comparison between subtle differences in dietary composition among different dietary regimes reveal that adequate glucose concentration in diet is a critical determinant for establishing RALDH activity specifically in siLP-DCs. Consequently, pre-treatment of siLP-DCs, and not mesenteric lymph node derived MLNDCs with glucose, results in significant enhancement in the in vitro generation of induced Regulatory T (iTreg) cells. Our findings reveal previously underappreciated role of dietary glucose concentration in establishing regulatory properties in intestinal DCs, thereby extending a potential therapeutic module against intestinal inflammation.