Conversion Of Retinol Research Articles

Human intestinal stromal cells promote homeostasis in normal mucosa but inflammation in Crohn’s disease in a retinoic acid–deficient manner

Intestinal stromal cells (SCs), which synthesize the extracellular matrix that gives the mucosa its structure, are newly appreciated to play a role in mucosal inflammation. Here we show that human intestinal vimentin+CD90+SMA- SCs synthesize retinoic acid (RA) at levels equivalent to intestinal epithelial cells, a function in the human intestine previously attributed exclusively to epithelial cells. Crohn’s disease SCs (Crohn’s SCs), however, synthesized markedly less RA than SCs from healthy intestine (Normal SCs). We also show that microbe-stimulated Crohn’s SCs, which are more inflammatory than stimulated Normal SCs, induced less RA-regulated differentiation of mucosal DCS (circulating pre-DCs and monocyte-derived DCs), leading to the generation of more potent inflammatory IFN-γhi/IL-17hi T cells than Normal SCs. Explaining these results, Crohn’s SCs expressed more DHRS3, a retinaldehyde reductase that inhibits retinol conversion to retinal, and thus synthesized less RA than Normal SCs. These findings uncover a microbe–SC–DC crosstalk in which luminal microbes induce Crohn’s disease SCs to initiate and perpetuate inflammation through impaired synthesis of RA.

Retinoic acid mitigates the NSC319726-induced spermatogenesis dysfunction through cuproptosis-independent mechanisms

Copper ionophore NSC319726 has attracted researchers' attention in treating diseases, particularly cancers. However, its potential effects on male reproduction during medication are unclear. This study aimed to determine whether NSC319726 exposure affected the male reproductive system. The reproductive toxicity of NSC319726 was evaluated in male mice following a continuous exposure period of 5 weeks. The result showed that NSC319726 exposure caused testis index reduction, spermatogenesis dysfunction, and architectural damage in the testis and epididymis. The exposure interfered with spermatogonia proliferation, meiosis initiation, sperm count, and sperm morphology. The exposure also disturbed androgen synthesis and blood testis barrier integrity. NSC319726 treatment could elevate the copper ions in the testis to induce cuproptosis in the testis. Copper chelator rescued the elevated copper ions in the testis and partly restored the spermatogenesis dysfunction caused by NSC319726. NSC319726 treatment also decreased the level of retinol dehydrogenase 10 (RDH10), thereby inhibiting the conversion of retinol to retinoic acid, causing the inability to initiate meiosis. Retinoic acid treatment could rescue the meiotic initiation and spermatogenesis while not affecting the intracellular copper ion levels. The study provided an insight into the bio-safety of NSC319726. Retinoic acid could be a potential therapy for spermatogenesis impairment in patients undergoing treatment with NSC319726.

LKB1 in Intestinal Epithelial Cells Regulates Bile Acid Metabolism by Modulating FGF15/19 Production.

Background & AimsLiver kinase B1 (LKB1) is a master upstream protein kinase involved in nutrient sensing and glucose and lipid metabolism in many tissues; however, its metabolic role in intestinal epithelial cells (IEC) remains unclear. In this study, we investigated the regulatory role of LKB1 on bile acid (BA) homeostasis.MethodsWe generated mice with IEC-specific deletion of LKB1 (LKB1ΔIEC) and analyzed the characteristics of IEC development and BA level. In vitro assays with small interfering RNA, liquid chromatography/mass spectrometry, metagenomics, and RNA-sequencing were used to elucidate the regulatory mechanisms underlying perturbed BA homeostasis.ResultsLKB1 deletion resulted in abnormal differentiation of secretory cell lineages. Unexpectedly, BA pool size increased substantially in LKB1ΔIEC mice. A significant reduction of the farnesoid X receptor (FXR) target genes, including fibroblast growth factor 15/19 (FGF15/19), known to inhibit BA synthesis, was found in the small intestine (SI) ileum of LKB1ΔIEC mice. We observed that LKB1 depletion reduced FGF15/19 protein level in human IECs in vitro. Additionally, a lower abundance of bile salt hydrolase-producing bacteria and elevated levels of FXR antagonist (ie, T-βMCA) were observed in the SI of LKB1ΔIEC mice. Moreover, LKB1ΔIEC mice showed impaired conversion of retinol to retinoic acids in the SI ileum. Subsequently, vitamin A treatment failed to induce FGF15 production. Thus, LKB1ΔIEC mice fed with a high-fat diet showed improved glucose tolerance and increased energy expenditure.ConclusionsLKB1 in IECs manages BA homeostasis by controlling FGF15/19 production.

Biallelic hypomorphic variants in ALDH1A2 cause a novel lethal human multiple congenital anomaly syndrome encompassing diaphragmatic, pulmonary, and cardiovascular defects.

This study shows a causal association between ALDH1A2 variants and a novel, severe multiple congenital anomaly syndrome in humans that is neonatally lethal due to associated pulmonary hypoplasia and respiratory failure. In two families, exome sequencing identified compound heterozygous missense variants in ALDH1A2. ALDH1A2 is involved in the conversion of retinol (vitamin A) into retinoic acid (RA), which is an essential regulator of diaphragm and cardiovascular formation during embryogenesis. Reduced RA causes cardiovascular, diaphragmatic, and associated pulmonary defects in several animal models, matching the phenotype observed in our patients. In silico protein modeling showed probable impairment of ALDH1A2 for three of the four substitutions. In vitro studies show a reduction of RA. Few pathogenic variants in genes encoding components of the retinoic signaling pathway have been described to date, likely due to embryonic lethality. Thus, this study contributes significantly to knowledge of the role of this pathway in human diaphragm and cardiovascular development and disease. Some clinical features in our patients are also observed in Fryns syndrome (MIM# 229850), syndromic microphthalmia 9 (MIM# 601186), and DiGeorge syndrome (MIM# 188400). Patients with similar clinical features who are genetically undiagnosed should be tested for recessive ALDH1A2-deficient malformation syndrome.

Probiotic modulation of perfluorobutanesulfonate toxicity in zebrafish: Disturbances in retinoid metabolism and visual physiology

Perfluorobutanesulfonate (PFBS), an aquatic pollutant of emerging concern, is found to disturb gut microbiota, retinoid metabolism and visual signaling in teleosts, while probiotic supplementation can shape gut microbial community to improve retinoid absorption. However, it remains unknown whether probiotic bacteria can modulate the toxicities of PFBS on retinoid metabolism and visual physiology. In the present study, adult zebrafish were exposed for 28 days to 0, 10 and 100 μg/L PFBS, with or without dietary administration of probiotic Lactobacillus rhamnosus. Interaction between PFBS and probiotic was examined regarding retinoid dynamics (intestine, liver and eye) and visual stimuli transmission. PFBS single exposures remarkably inhibited the absorption of retinyl ester in female intestines, which were, however, restored by probiotic to normal status. Although coexposure scenarios markedly increased the hepatic storage of retinyl ester in females, mobilization of retinol was reduced in livers by single or combined exposures regardless of sex. In the eyes, transport and catalytic conversion of retinol to retinal and retinoic acid were interrupted by PFBS alone, which were efficiently antagonized by probiotic presumably through an indirect action. In response to the availability of retinal chromophore, transcriptions of opsins and arrestin genes were altered adaptively to control visual perception and termination. Neurotransmission across retina circuitry was changed accordingly, centering on epinephrine and norepinephrine. In summary, the present study found the efficient modulation of probiotic on retinoid metabolic disorders of PFBS pollution, which subsequently impacted visual signaling. A future work is warranted to provide mechanistic clues in retinoid interaction.

Reduced retinoic acid synthesis accelerates prophase I and follicle activation.

In female mammals, reproductive potential is determined during fetal life by the formation of a non-renewable pool of primordial follicles. Initiation of meiosis is one of the defining features of germ cell differentiation and is well established to commence in response to retinoic acid. WIN 18,446 inhibits the conversion of retinol to retinoic acid, and therefore it was used to explore the impact of reduced retinoic acid synthesis on meiotic progression and thus germ cell development and subsequent primordial follicle formation. e13.5 mouse fetal ovaries were cultured in vitro and treated with WIN 18,446 for the first 3 days of a total of up to 12 days. Doses as low as 0.01 µM reduced transcript levels of the retinoic acid response genes Stra8 and Rarβ without affecting germ cell number. Higher doses resulted in germ cell loss, rescued with the addition of retinoic acid. WIN 18,446 significantly accelerated the progression of prophase I; this was seen as early as 48 h post treatment using meiotic chromosome spreads and was still evident after 12 days of culture using Tra98/Msy2 immunostaining. Furthermore, ovaries treated with WIN 18,446 at e13.5 but not at P0 had a higher proportion of growing follicles compared to vehicle controls, thus showing evidence of increased follicle activation. These data therefore indicate that retinoic acid is not necessary for meiotic progression but may have a role in the regulation of its progression and germ cell survival at that time and provide evidence for a link between meiotic arrest and follicle growth initiation.

Characterization of an Acyl‐CoA:retinol Acyltransferase (ARAT) Activity in the Hepatopancreas of the American Lobster (Homarus americanus)

Vitamin A (retinol) is a necessary nutrient in higher animals. In mammals, the pathways by which retinol is transported and metabolized are well described. One important reaction in these pathways is the conversion of retinol into retinyl esters, which occurs in several tissues, including the small intestine, liver, and eye. Little is known about the conversion of retinol into retinyl esters in invertebrates. We have identified an acyl‐CoA:retinol acyltransferase (ARAT) activity in the hepatopancreas of the American lobster (Homarus americanus). This activity shows a linear dependence on time and microsome concentration. It shows temperature dependence and an optimal pH of 6.5. We have characterized its acyl‐CoA preference and determined Km and Vmax values for both retinol and acyl‐CoA.Support or Funding InformationCentre College

Retinoid metabolism and functions mediated by retinoid binding-proteins.

Cellular retinoid-binding proteins (BP) chaperone retinol through esterification, conversion of retinol into retinal, reduction of retinal, conversion of retinal into all-trans-retinoic acid (ATRA), and ATRA to catabolism. They also deliver ATRA to nuclear receptors and mediate non-genomic ATRA actions. These retinoid-specific binding-proteins include: cellular retinol binding-protein, type 1 (Crbp1), cellular retinol binding-protein type 2 (Crbp2), cellular retinol binding-protein type 3 (Crbp3), cellular retinoic acid binding-protein type 1 (Crabp1); cellular retinoic acid binding-protein type 2 (Crabp2). Retinoid BP bind their ligands specifically and with high-affinity. These BP seemingly evolved to solubilize the lipophilic retinoids in the aqueous cellular medium, and allow retinoid access only to enzymes that recognize both the BP and the retinoid. By chaperoning retinoids through cells, retinoid BP provide specificity to retinoids' metabolism and protect the scarce resource from dispersing into cell membranes and/or undergoing catabolism as xenobiotics. Other functions include non-genomic actions of Crabp1, delivery of ATRA to RAR by holo-Crabp2, and stabilization of HuR by apo-Crabp2. In addition to the retinoid-specific BP, Fabp5 also binds ATRA and delivers it to Pparδ. This article describes these BP and their functions, with a focus on experimental protocols to distinguish protein-protein interactions from diffusion-mediated transfer of ligand from BP to enzymes or receptors, and methods for quantifying retinoids.

Astaxanthin sources: Suitability for human health and nutrition

Background: Astaxanthin (AX) has been consumed as a nutritional supplement for approximately twenty years. The primary source has been a natural plant-based supplement from the single-cell alga Haematococcus pluvialis (NAT-AX). Recently, Astaxanthin from other sources has entered the marketplace. The primary alternative source in the human nutritional supplement market has been a synthetic form of Astaxanthin produced from petrochemicals (SYN-AX). Additionally, a very small amount of Astaxanthin from a genetically-manipulated yeast Xanthophyllomyces dendrorhous (former nomenclature Phaffia rhodozyma, still commonly referred to as “Phaffia”) (PH-AX) is also available in some supplement products. The three forms have substantial chemical differences. In addition to the chemical differences between sources of AX, in-vitro research has demonstrated profound differences in antioxidant strength and animal research has revealed fundamental differences in health benefits. In all cases, NAT-AX has proven more biologically active than the other sources. This review is designed to assist readers in understanding which form(s) of AX are suitable for consumers desiring preventive or therapeutic health benefits. Results: In head-to-head antioxidant experiments, NAT-AX demonstrated 14X to 90X greater antioxidant activity than SYN-AX. In numerous animal trials in diverse species, NAT-AX in esterified form has demonstrated superior efficacy in increasing lifespan; treating skin cancer; preventing the formation of gastric ulcers; improving resistance to stress; decreasing reactive oxygen species (ROS); increasing retinol conversion in the liver; augmenting enzyme levels; increasing growth rates; and improving exercise endurance.From a safety perspective, NAT-AX has been the subject of human clinical trials demonstrating safety and a wide variety of health benefits. In addition, no documented adverse events have surfaced during its twenty years of distribution as a food supplement for humans. SYN-AX and PH-AX have not been proven safe for direct human consumption and have not demonstrated any health benefits in clinical trials. Due to these facts, SYN-AX and PH-AX have not been allowed for human consumption by government regulators in many countries while NAT-AX is widely accepted in most countries around the world. Conclusion: Based on our review of the literature below, we recommend NAT-AX as the sole form of AX for human consumption until SYN-AX and PH-AX have been proven safe and efficacious through human clinical research.

Biochemical and histochemical analyses of lecithin:retinol acyltransferase from polar bear (Ursus maritimus) livers

Vitamin A is a fat-soluble vitamin required for many physiological functions. It is known that polar bears (Ursus maritimus) store a large amount of vitamin A within hepatic stellate cells (HSCs). However, the molecular mechanisms for high-capacity storage of vitamin A by polar bear HSCs are unknown. We biochemically and histochemically analyzed lecithin:retinol acyltransferase (LRAT), an enzyme responsible for conversion of retinol to its storage form, retinylesters, in the liver. Surprisingly, LRAT activities of the liver microsome fractions from polar bears weighing 100–500 kg were considerably lower than those obtained from rats (Rattus norvegicus). Although there is a possibility that the quality of polar bear liver specimens was reduced during transport, intrinsic LRAT activity of polar bears might actually be lower than those of other mammals. Fluorescent immunohistochemistry revealed that LRAT and cellular retinol-binding protein (CRBP) I were well co-localized within the liver lobules of polar bears. Taking into account that retinol bound to CRBP I is prevented from degradation and becomes a good substrate of LRAT, we hypothesized that CRBP I effectively conveys retinol to the LRAT, thereby circumventing the low enzymatic activity of LRAT in polar bear livers. As the top predator in the Arctic, polar bears have access to large amounts of vitamin A via the food web. We suggest that polar bears acquired an efficient mechanism of esterification of vitamin A within the liver during the course of their evolution.

Retinol saturase coordinates liver metabolism by regulating ChREBP activity

The liver integrates multiple metabolic pathways to warrant systemic energy homeostasis. An excessive lipogenic flux due to chronic dietary stimulation contributes to the development of hepatic steatosis, dyslipidemia and hyperglycemia. Here we show that the oxidoreductase retinol saturase (RetSat) is involved in the development of fatty liver. Hepatic RetSat expression correlates with steatosis and serum triglycerides (TGs) in humans. Liver-specific depletion of RetSat in dietary obese mice lowers hepatic and circulating TGs and normalizes hyperglycemia. Mechanistically, RetSat depletion reduces the activity of carbohydrate response element binding protein (ChREBP), a cellular hexose-phosphate sensor and inducer of lipogenesis. Defects upon RetSat depletion are rescued by ectopic expression of ChREBP but not by its putative enzymatic product 13,14-dihydroretinol, suggesting that RetSat affects hepatic glucose sensing independent of retinol conversion. Thus, RetSat is a critical regulator of liver metabolism functioning upstream of ChREBP. Pharmacological inhibition of liver RetSat may represent a therapeutic approach for steatosis.

Abstract 4924: Adh1blow/Cyp26b1high activity of CD90+ (myo)fibroblasts supports tumor-promoting inflammation in colorectal cancer

Abstract Introduction. Retinol (RO) and its active metabolite, all-trans retinoic acid (atRA), have been widely studied as cancer chemotherapeutic agents. atRA is a critical immunoregulatory molecule reported to decrease cancer cell proliferation and inhibit tumor-promoting inflammation. However, recent studies report a loss of efficacy of atRA treatment to counteract colorectal cancer (CRC) progress, and the mechanisms underlying this failure are not fully understood. Alcohol dehydrogenase 1B (ADH1B), a major isoform of alcohol dehydrogenase expressed in normal colon, is responsible for the conversion of RO to atRA. We recently reported that stromal (myo)fibroblasts are major ADH1B expressers in normal colonic mucosa and that this expression is lost during the neoplastic transformation process of CRC. Thus, we hypothesized that aberrant retinoid signaling in cancer-associated fibroblasts (CAFs) is among key processes supporting tumor-promoting inflammation in CRC. Methods. Real-time PCR, confocal microscopy and cytokine luminex arrays were used to evaluate alterations in retinoid pathway gene expression and activity to define its relevance to the tumor-associated inflammation in human normal and CRC colonic tissues, and in normal primary (myo)fibroblasts (N-CMFs) and CAFs. Results. We observed a strong downregulation of ADH1B expression in CAFs in situ and in culture along with an increased expression of the tumor growth promoting inflammatory cytokine IL-6. Addition of the ADH1B substrate, RO, or its metabolite, atRA, to N-CMF cultures decreased the LPS-inducible IL-6 expression by N-CMFs. No downregulation of LPS-inducible IL-6 expression by RO was observed in CAFs, while atRA treatment partially inhibits IL-6 expression. Silencing of adh1b gene in N-CMFs led to the increased production of the LPS-inducible IL-6. This suggests that lack of ADH1B expression in CAFs prevents conversion of RO into atRA, resulting in the disruption of the negative regulation of IL-6. A decrease in downstream enzyme ALDH1A expression, which converts ADH1B’s byproduct retinaldehyde, to atRA, was also observed in CAFs. Finally, only a partial inhibition of IL-6 production by exogenous atRA was observed in CAFs. When compared to N-CMFs, CAFs expressed higher levels of CYP26B1, a cytochrome that is responsible for atRA degradation. Treatment of CAFs with atRA resulted in an eight fold higher upregulation of CYP26B1 expression than in N-CMFs. Conclusions. Taken together, our data suggests that CYP26B1 overexpression in CAFs along with ADH1B downregulation support aberrant retinoic acid metabolism in CRC. Adh1blow/Cyp26b1high activity of CAFs leads to a decrease of atRA availability in the tumor microenvironment, supporting IL-6-driven tumor growth promoting inflammation and resistance to exogenous atRA as a chemotherapeutic agent in CRC. Citation Format: Romain Villeger, Gabriella Uribe, Judy A. Trieu, Paul Johnson, Suimmin Qiu, Don W. Powell, Ellen J. Beswick, Iryna V. Pinchuk. Adh1blow/Cyp26b1high activity of CD90+ (myo)fibroblasts supports tumor-promoting inflammation in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4924. doi:10.1158/1538-7445.AM2017-4924

Quantitation of retinaldehyde in small biological samples using ultrahigh-performance liquid chromatography tandem mass spectrometry

We report an ultrahigh-performance liquid chromatography tandem mass spectrometry (UHPLC–MS/MS) method to quantify all-trans-retinal in biological samples of limited size (15–35mg), which is especially advantageous for use with adipose. To facilitate recovery, retinal and the internal standard 3,4-didehydroretinal were derivatized in situ into their O-ethyloximes. UHPLC resolution combined with high sensitivity and specificity of MS/MS allowed quantification of retinal-O-ethyloximes with a 5-fmol lower limit of detection and a linear range from 5fmol to 1pmol. This assay revealed that extraocular concentrations of retinal range from approximately 2 to 40pmol/g in multiple tissues—the same range as all-trans-retinoic acid. All-trans-retinoic acid has high affinity (kd⩽0.4nM) for its nuclear receptors (RARα, -β, and -γ), whereas retinal has low (if any) affinity for these receptors, making it unlikely that these retinal concentrations would activate RAR. We also show that the copious amount of vitamin A used in chow diets increases retinal in adipose depots 2- to 5-fold relative to levels in adipose of mice fed a vitamin A-sufficient diet, as recommended for laboratory rodents. This assay also is proficient for quantifying conversion of retinol into retinal in vitro and, therefore, provides an efficient method to study metabolism of retinol in vivo and in vitro.

Dendritic cells metabolize and activate retinol into all-trans retinoic acid which stimulates antimicrobial activity in M. tuberculosis-infected cells via paracrine interaction (INM3P.406)

Abstract Understanding the innate immune response to the Mycobacterium tuberculosis, is a key step to development of effective immunotherapies for tuberculosis (TB). Epidemiological evidence correlates low serum vitamin A (retinol) levels to active TB; however, retinol is biologically inactive. Previously, we demonstrated that stimulation of M. b-infected macrophages with the active vitamin A hormone, all-trans retinoic acid (ATRA), triggers a NPC2-dependent antimicrobial activity. Therefore, understanding the mechanism by which the immune system metabolizes and activates retinol at the site of infection will solve the mystery of how retinol, despite being biologically inactive, can influence the immune response to infection. Through gene expression and functional analysis of human macrophage subsets and dendritic cells (DCs) we identified that GM-CSF-derived DCs express enzymes capable of the two step conversion of retinol into ATRA including, ALDH1A2 and DHRS9, which is subsequently released from the cell. Conditioned medium, from DCs cultured with retinol, stimulated expression of NPC2 from monocytes which was inhibited by a retinoic acid receptor inhibitor (RARi). Inclusion of DEAB, an ALDH1A2 inhibitor, during DCs and retinol incubation also abrogated this response. Expression levels of vitamin A metabolic enzymes are lower in TB lung tissues compared with normal lung. These results help us incorporate vitamin A into clinical care and therapeutic modalities for TB.

Butyrate stimulates the epithelial potential to produce retinoic acid demonstrated in primary epithelial enteroid systems

Reduced levels of the short chain fatty acid (SCFA) butyrate are associated with microbial dysbiosis and disease development. Butyrate serves as main energy source in colonocytes and is purported to exert anti-inflammatory effects. Using enteroid cultures for primary intestinal epithelia, we studied effects of butyrate on the production of retinoic acid (RA) as intestinal epithelia dictate homeostasis and immune tolerance by stimulating tolerogenic CD103+DCs through release of RA. Retinal dehydrogenases (aldha1a1 and aldh1a3) are gatekeeping enzymes for retinoic acid production. Mouse CMT93, human CACO2 epithelial cell lines, and mouse intestinal enteroids were incubated with SCFA and analyzed for RA metabolic enzyme expression, stemness, and maturation markers. Functional RA conversion was analyzed by Aldefluor activity assays. Butyrate (0-5mM) dose dependently induced aldh1a1 and -3 expression in CACO2 and CMT93 cells (up to 10-30 fold at 5mM for aldh1a1 and -3 resp.) coinciding with increased functional conversion of retinol to RA. Moreover, in enteroids butyrate (2-5mM) upregulated aldh1a3 expression 50 fold over control. RA receptor alpha was upregulated two-fold, indicating enhanced RA signaling. Butyrate (2-5mM) reduced stemness marker (LGR5, OLMF4) expression but upregulated Wnt target genes (CD44, ASCL2, CDKN1), indicating that enhanced RA conversion did not reflect dedifferentiation. We conclude butyrate can support epithelial conversion of dietary retinoic acid thereby enhancing the capacity to generate tolerogenic DCs and drive immune tolerance in the gut.

The Retinol Dehydrogenase Rdh10 Localizes to Lipid Droplets during Acyl Ester Biosynthesis

Rdh10 catalyzes the first step of all-trans-retinoic acid biogenesis physiologically, conversion of retinol into retinal. We show that Rdh10 associates predominantly with mitochondria/mitochondrial-associated membrane (MAM) in the absence of lipid droplet biosynthesis, but also locates with lipid droplets during acyl ester biosynthesis. Targeting to lipid droplets requires the 32 N-terminal residues, which include a hydrophobic region followed by a net positive charge. Targeting to mitochondria/MAM and/or the stability of Rdh10 require both the N-terminal and the 48 C-terminal hydrophobic residues. Rdh10 behaves similarly to cellular retinol-binding protein, type 1, which also localizes to mitochondria/MAM before lipid droplet synthesis, and associates with lipid droplets during acyl ester synthesis (Jiang, W., and Napoli, J. L. (2012) Biochem. Biophys. Acta 1820, 859-8692). LRAT, an ER protein, also associates with lipid droplets upon acyl ester biosynthesis. Colocalization of Rdh10, Crbp1, and LRAT on lipid droplets suggests a metabolon that mediates retinol homeostasis.

Morphological defects in a novel Rdh10 mutant that has reduced retinoic acid biosynthesis and signaling

Retinoic acid (RA) signaling is necessary for proper patterning and morphogenesis during embryonic development. Tissue-specific RA signaling requires precise spatial and temporal synthesis of RA from retinal by retinaldehyde dehydrogenases (Raldh) and the conversion of retinol to retinal by retinol dehydrogenases (Rdh) of the short-chain dehydrogenase/reducatase gene family (SDR). The SDR, retinol dehydrogenase 10 (RDH10), is a major contributor to retinal biosynthesis during mid-gestation. We have identified a missense mutation in the Rdh10 gene (Rdh10(m366Asp) ) using an N-ethyl-N-nitrosourea-induced forward genetic screen that result in reduced RA levels and signaling during embryonic development. Rdh10(m366Asp) mutant embryos have unique phenotypes, such as edema, a massive midline facial cleft, and neurogenesis defects in the forebrain, that will allow the identification of novel RA functions.

Multiple Retinol and Retinal Dehydrogenases Catalyze All-trans-retinoic Acid Biosynthesis in Astrocytes

All-trans-retinoic acid (atRA) stimulates neurogenesis, dendritic growth of hippocampal neurons, and higher cognitive functions, such as spatial learning and memory formation. Although astrocyte-derived atRA has been considered a key factor in neurogenesis, little direct evidence identifies hippocampus cell types and the enzymes that biosynthesize atRA. Here we show that primary rat astrocytes, but not neurons, biosynthesize atRA using multiple retinol dehydrogenases (Rdh) of the short chain dehydrogenase/reductase gene family and retinaldehyde dehydrogenases (Raldh). Astrocytes secrete atRA into their medium; neurons sequester atRA. The first step, conversion of retinol into retinal, is rate-limiting. Neurons and astrocytes both synthesize retinyl esters and reduce retinal into retinol. siRNA knockdown indicates that Rdh10, Rdh2 (mRdh1), and Raldh1, -2, and -3 contribute to atRA production. Knockdown of the Rdh Dhrs9 increased atRA synthesis ∼40% by increasing Raldh1 expression. Immunocytochemistry revealed cytosolic and nuclear expression of Raldh1 and cytosol and perinuclear expression of Raldh2. atRA autoregulated its concentrations by inducing retinyl ester synthesis via lecithin:retinol acyltransferase and stimulating its catabolism via inducing Cyp26B1. These data show that adult hippocampus astrocytes rely on multiple Rdh and Raldh to provide a paracrine source of atRA to neurons, and atRA regulates its own biosynthesis in astrocytes by directing flux of retinol. Observation of cross-talk between Dhrs9 and Raldh1 provides a novel mechanism of regulating atRA biosynthesis.

Tissue-specific increases in endogenous all-trans retinoic acid: possible contributing factor in ethanol toxicity

All-trans retinoic acid (atRA) is required for neurogenesis and dendritic growth in the hippocampus. The toxic effects of ethanol include developmental defects, cognitive dysfunction, and increased risk of cancer and have some similarities to the detrimental effects of excess atRA, the active form of vitamin A. It is therefore possible that disruption of atRA homeostasis would contribute to the deleterious effects of ethanol. However, previous work, using very high exogenous doses of retinol, found that ethanol toxicity led to decreased formation of atRA, apparently due to competitive inhibition of alcohol dehydrogenase, which is purportedly involved in the conversion of retinol to retinal. A new study, using assays that are highly sensitive for endogenous atRA, has reported that ethanol toxicity in mice actually increased atRA concentration in certain tissues, including brain hippocampus, apparently due to a mobilization of hepatic retinyl esters that led to increased retinol and atRA in specific tissues.

Provitamin A from Golden Rice is as effective as provitamin A in oil to children

Spinach and pure β‐carotene (β‐C) in oil capsule are good sources for vitamin A. A new transgenic product, Golden Rice (GR), is enriched with β‐C genetically. To determine its vitamin A value as compared with spinach or pure 2H8 labeled β‐C (M+8), we conducted a study with healthy Chinese school children (age 7–8y). Intrinsically labeled GR and spinach were obtained by growing plants in a nutrient solution containing ~25% 2H2O. The GR β‐C and spinach β‐C were enriched with deuterium with the highest abundance at M+9. An oil capsule of 13C10 ‐ retinyl acetate (M+10) was taken as a reference dose to determine the β‐C to retinol conversion factors. Serum samples collected from the subjects were analyzed using GC/ECNCI‐MS for the enrichments of labeled retinol by monitoring the isotopes of M+4 (derived from 2H8 β‐C), M+5 (from GR or spinach β‐C) and M+10 (from 13C10 ‐ retinyl‐acetate). By using the response to the dose of 13C10 ‐ retinyl acetate (0.5 mg) as a reference, our preliminary results (using area under the curve of percent enrichment at days 1, 3, 7, 14, and 21 after the labeled doses) showed that the conversions of pure β‐C (0.5 mg), GR β‐C (0.6 mg), and spinach β‐C (1.4 mg) to retinol were 2.0, 2.1, and 7.3 to 1 by weight, respectively. These results indicate that GR can be as effective as β‐C in oil in contributing to the daily vitamin A requirements of children, thus, to improve population‐based vitamin A adequacy.