Ralph Rühl

Vitamin E activates gene expression via the pregnane X receptor.

Tocopherols and tocotrienols are metabolized by side chain degradation via initial omega-oxidation and subsequent beta-oxidation. omega-Oxidation is performed by cytochrome P450 (CYP) enzymes which are often regulated by their substrates themselves. Results presented here show that all forms of Vitamin E are able to activate gene expression via the pregnane X receptor (PXR), a nuclear receptor regulating a variety of drug metabolizing enzymes. In HepG2 cells transfected with the human PXR and the chloramphenicol acetyl transferase (CAT) gene linked to two PXR responsive elements, CAT activity was most strongly induced by alpha- and gamma-tocotrienol followed by rifampicin, delta-, alpha- and gamma-tocopherol. The inductive efficacy was concentration-dependent; its specificity was underscored by a lower response when cotransfection with PXR was omitted. Up-regulation of endogenous CYP3A4 and CYP3A5 mRNA was obtained by gamma-tocotrienol, the most potent activator of PXR, with the same efficacy as with rifampicin. This points to a potential interference of individual forms of Vitamin E with the metabolism and efficacy of drugs.

The Human Sebocyte Culture Model Provides New Insights into Development and Management of Seborrhoea and Acne

Seborrhoea and acne are exclusively human diseases and sebaceous gland differentiation is species specific. Therefore, fundamental research on human sebaceous cell function and control requires human in vitro models. The human sebocyte culture model, introduced in 1989, has been used in several studies to elucidate sebaceous gland activity and its regulation at the cellular level. Cultured human sebocytes have been shown to preserve important sebocytic characteristics, although they undergo an incomplete terminal differentiation in vitro. In vitro synthesis of free fatty acids without bacterial involvement and marked interleukin 1α expression at the mRNA and protein levels with no further induction by lipopolysaccharides lead to the assumption that human sebocytes may initiate acne lesions by an intrinsic mechanism. Androgens affected sebocyte activity in vitro in a manner dependent on the localization of the sebaceous glands. In vitro stimulation of sebocyte proliferation by androgens could be completely abolished by spironolactone. Cultured sebocytes strongly expressed type 1 5α-reductase and metabolized testosterone to androstenedione, 5α-androstanedione, 5α-dihydrotestosterone, androsterone and 5α-androstanediol, whereas the levels of 5α-reductase activity were probably not feedback regulated. 4,7β-Dimethyl-4-aza-5αcholestan-3-one, a type 1 5α-reductase inhibitor, induced an early, marked down-regulation of 5α-reductase activity in human sebocytes in vitro, while hydrofinasteride, a type 2 inhibitor, required 103-fold higher concentrations to induce similar effects. Stimulation of sebocyte proliferation by insulin, thyroid-stimulating hormone and hydrocortisone indicates that the hormonal control of the sebaceous gland could be a complex mechanism. Retinoids inhibited sebocyte proliferation in a dose-dependent manner and down-regulated lipid synthesis and sebocyte differentiation in vitro. Isotretinoin was the most potent compound. On the other hand, vitamin A was found essential for sebocyte activity and differentiation in vitro and could be partially substituted by synthetic retinoids. The inhibitory effect of isotretinoin on sebocyte proliferation was barely affected by the presence of vitamin A. The low persistent isotretinoin levels or, more likely, the considerably elevated tretinoin concentrations detected in human sebocytes after treatment with isotretinoin in vitro may be responsible for the inhibitory effect of this compound on sebocyte activity.

Transcriptional Regulation of Human CYP27 Integrates Retinoid, Peroxisome Proliferator-Activated Receptor, and Liver X Receptor Signaling in Macrophages

ABSTRACT Cholesterol uptake and efflux are key metabolic processes associated with macrophage physiology and atherosclerosis. Peroxisome proliferator-activated receptor gamma (PPARγ) and liver X receptor alpha (LXRα) have been linked to the regulation of these processes. It remains to be identified how activation of these receptors is connected and regulated by endogenous lipid molecules. We identified CYP27, a p450 enzyme, as a link between retinoid, PPARγ, and LXR signaling. We show that the human CYP27 gene is under coupled regulation by retinoids and ligands of PPARs via a PPAR-retinoic acid receptor response element in its promoter. Induction of the enzymes expression results in an increased level of 27-hydroxycholesterol and upregulation of LXR-mediated processes. Upregulated CYP27 activity also leads to LXR-independent elimination of CYP27 metabolites as an alternative means of cholesterol efflux. Moreover, human macrophage-rich atherosclerotic lesions have an increased level of retinoid-, PPARγ-, and LXR-regulated gene expression and also enhanced CYP27 levels. Our findings suggest that nuclear receptor-regulated CYP27 expression is likely to be a key integrator of retinoic acid receptor-PPARγ-LXR signaling, relying on natural ligands and contributing to lipid metabolism in macrophages.

Astroglia-derived retinoic acid is a key factor in glia-induced neurogenesis

Astroglial cells are essential components of the neurogenic niches within the central nervous system. Emerging evidence suggests that they are among the key regulators of postnatal neurogenesis. Although astrocytes have been demonstrated to possess the potential to instruct stem cells to adopt a neuronal fate, little is known about the nature of the glia‐derived instructive signals. Here we propose that all‐trans reti‐noic acid, one of the most powerful morphogenic molecules regulating neuronal cell fate commitment, may be one of the glia‐derived factors directing astro‐glia‐induced neurogenesis. According to data obtained from several complementary approaches, we show that cultured astrocytes express the key enzyme mRNAs of retinoic acid biosynthesis and actively produce all‐trans retinoic acid. We show that blockage of retinoic acid signaling by the pan‐RAR antagonist AGN193109 prevents glia‐induced neuron formation by noncommitted stem cells. Therefore, we provide strong in vitro evidence for retinoic acid action in astroglia‐induced neuronal differentiation.—Környei, Z., Gócza, E., Rühl, R., Vörös, E., Orsolits, B., Szabo, B., Vágovits, B., Madarász, E. Astroglia‐derived retinoic acid is a key factor in glia‐induced neurogenesis. FASEB J. 21, 2496–2509 (2007)

Docosahexaenoic acid (DHA) supplementation in atopic eczema: a randomized, double‐blind, controlled trial

Background The increasing prevalence of atopic eczema has been linked to the alteration of the Western diet, namely the reduced consumption of omega‐3 (n‐3) polyunsaturated fatty acids (PUFA) and an increased omega‐6 (n‐6) PUFA intake.

Effects of dietary retinoids and carotenoids on immune development.

Carotenoids and retinoids are groups of nutritionally-relevant compounds present in many foods of plant origin (carotenoids) and animal origin (mainly retinoids). Their levels in human subjects vary depending on the diversity and amount of the individuals nutrient intake. Some carotenoids and retinoids have been investigated for their effects on the immune system both in vitro and in vivo. It has been shown that retinoids have the potential to mediate or induce proliferative and differentiating effects on several immune-competent cells, and various carotenoids are known to be inducers of immune function. The immune-modulating effects of retinoids have been well documented, while the effects of carotenoids on the immune system have not been investigated as extensively, because little is known about their molecular mechanism of action. The present review will mainly focus on the molecular mechanism of action of retinoids and particularly carotenoids, their nutritional origin and intake, their transfer from the maternal diet to the child and their effects or potential effects on the developing immune system.

Research resource: transcriptome profiling of genes regulated by RXR and its permissive and nonpermissive partners in differentiating monocyte-derived dendritic cells.

Retinoid X receptors (RXRs) are heterodimerization partners for many nuclear receptors and also act as homodimers. Heterodimers formed by RXR and a nonpermissive partner, e.g. retinoic acid receptor (RAR) and vitamin D receptor (VDR), can be activated only by the agonist of the partner receptor. In contrast, heterodimers that contain permissive partners, e.g. liver X receptor (LXR) and peroxisome proliferator-activated receptor (PPAR), can be activated by agonists for either the partner receptor or RXR, raising the possibility of pleiotropic RXR signaling. However, it is not known to what extent the receptor’s activation results in triggering mechanisms dependent or independent of permissive heterodimers. In this study, we systematically and quantitatively characterized all probable RXR-signaling pathways in differentiating human monocyte-derived dendritic cells (Mo-DCs). Using pharmacological, microarray and quantitative RT-PCR techniques, we identified and characterized gene sets regulated by RXR agonists (LG100268 and 9-cis retinoic acid) and agonists for LXRs, PPARs, RARα, and VDR. Our results demonstrated that permissiveness was partially impaired in Mo-DCs, because a large number of genes regulated by PPAR or LXR agonists was not affected by RXR-specific agonists or was regulated to a lesser extent. As expected, we found that RXR agonists regulated only small portions of RARα or VDR targets. Importantly, we could identify and characterize PPAR- and LXR-independent pathways in Mo-DCs most likely mediated by RXR homodimers. These data suggested that RXR signaling in Mo-DCs was mediated via multiple permissive heterodimers and also by mechanism(s) independent of permissive heterodimers, and it was controlled in a cell-type and gene-specific manner.

Retinoid‐ and carotenoid‐enriched diets influence the ontogenesis of the immune system in mice

Vitamin A (VA) has been identified as an important factor for the development of the immune system, especially during ontogenesis. It has been shown that antibody secretion and proliferation of lymphocyte populations depend on retinoids. In the present study we investigated the influence of a base VA diet and diets enriched with VA, β‐carotene and lycopene, on the ontogenesis of the immune system in mice. We examined the absolute and relative concentrations of splenic B lymphocytes (CD45R/B220), T lymphocytes (CD3+) and their subpopulations (CD4+ and CD8+), and measured serum immunoglobulin G (IgG) concentrations in the offspring of supplemented dams at different ages (1, 3, 5, 7, 14, 21 and 65 days). The experimental diets resulted in higher numbers of T and B lymphocytes after VA and carotenoid enrichment, when compared, at various time‐points, with the base diet. Higher values of total serum IgG were found in the β‐carotene‐enriched diet group on day 7. On days 7 and 14, the enriched diets induced significant alterations in the percentages and total numbers of splenic lymphocytes in comparison to the base diet. Our results confirm that supplementation with VA and carotenoids affect the immune‐cell function during ontogenesis and suggest a possible role of these nutritional factors on the development of the immune system.

PPARγ-mediated and arachidonic acid-dependent signaling is involved in differentiation and lipid production of human sebocytes.

The transcriptional basis of sebocyte differentiation and lipid production is mostly unclear. Peroxisome proliferator-activated receptor gamma (PPARγ), a lipid-activated transcription factor, has been implicated in differentiation and lipid metabolism of various cell types. Here, we show that PPARγ is differentially expressed in normal and pathological human sebocytes and appears to have roles in their differentiation and lipid production. We used laser-microdissected normal and pathological human sebaceous glands (SGs) and SZ95 cells (immortalized sebocyte cell line) analyzed by real-time quantitative PCR and immunohistochemistry. Lipids were analyzed by quantitative fluorimetry- and mass spectrometry-based approaches. We have observed that PPARγ and its target genes, ADRP (adipose differentiation-related protein) and PGAR (PPARγ angiopoietin-related protein), are expressed in sebocytes and show association with their level of differentiation. Also, PPARγ is present in normal and hyperplastic SG, whereas its expression levels are decreased in SG adenoma and SG carcinoma cells, reflecting a maturation-linked expression pattern. Furthermore, in SZ95 sebocytes, naturally occurring lipids, including arachidonic acid and arachidonic acid keto-metabolites (e.g., 5-KETE (5-oxo-6E,8Z,11Z,14Z-eicosatetraenoic acid), 12-KETE (12-oxo-5Z,8Z,10E,14Z-eicosatetraenoic acid)), appear to regulate PPARγ signaling pathways, which in turn modulate phospholipid biosynthesis and induce neutral lipid synthesis. Collectively, our findings highlight the importance of endogenous ligand-activated PPARγ signaling in human sebocyte biology and suggest that PPARγ might be a promising candidate for the clinical management of SG disorders.

Retinoid receptor-activating ligands are produced within the mouse thymus during postnatal development

Vitamin A deficiency is known to be accompanied with immune deficiency and susceptibility to a wide range of infectious diseases. Experimental evidence suggests that the active metabolites of vitamin A that mediate its effects on the immune system are the retinoic acids (RA), which are ligands for the nuclear RA receptor (RAR) family. RA were previously shown both to promote proliferation and to regulate apoptosis of thymocytes. In this study we detected the age‐dependent mRNA expression of retinaldehyde dehydrogenases (RALDH1 and 2), cellular RA binding protein‐II and CYP26A, proteins responsible for the synthesis, nuclear transport and degradation of RA in the postnatally developing thymus. RALDH1 was located in thymic epithelial cells. However, the amount of all‐trans RA in thymic homogenates was close to the detection limit, suggesting that in this tissue all‐trans RA is not the main RAR‐regulating product of retinol metabolism. At the same time, by measuring the induction of a RAR‐responsive transgene in two independent transgenic mouse strains, we demonstrated the production of an RAR‐activating ligand, which was age and RALDH dependent. Our data provide evidence for the existence of endogenous retinoid synthesis in the thymus and suggest that retinoids similar to glucocorticoids might indeed be involved in the regulation of thymic proliferation and selection processes by being present in the thymus in functionally effective amounts.