Hydroxysteroid Dehydrogenase Research Articles

The Black Box Orchestra of Gut Bacteria and Bile Acids: Who Is the Conductor?

Over the past decades the potential role of the gut microbiome and bile acids in type 2 diabetes mellitus (T2DM) has been revealed, with a special reference to low bacterial alpha diversity. Certain bile acid effects on gut bacteria concern cytotoxicity, or in the case of the microbiome, bacteriotoxicity. Reciprocally, the gut microbiome plays a key role in regulating the bile acid pool by influencing the conversion and (de)conjugation of primary bile acids into secondary bile acids. Three main groups of bacterial enzymes responsible for the conversion of bile acids are bile salt hydrolases (BSHs), hydroxysteroid dehydrogenases (HSDHs) and enzymes encoded in the bile acid inducible (Bai) operon genes. Interventions such as probiotics, antibiotics and fecal microbiome transplantation can impact bile acids levels. Further evidence of the reciprocal interaction between gut microbiota and bile acids comes from a multitude of nutritional interventions including macronutrients, fibers, prebiotics, specific individual products or diets. Finally, anatomical changes after bariatric surgery are important because of their metabolic effects. The heterogeneity of studies, diseases, bacterial species and (epi)genetic influences such as nutrition may challenge establishing specific and detailed interventions that aim to tackle the gut microbiome and bile acids.

Idiopathic hirsutism: Is it really idiopathic or is it misnomer?

Hirsutism, which is characterized by excessive growth of terminal hair in a male pattern, may result from various causes including polycystic ovary syndrome (PCOS), non-classic congenital adrenal hyperplasia, adrenal or ovarian tumors or it may be idiopathic. Idiopathic hirsutism is currently defined as hirsutism associated with normal ovulatory function, normal serum androgen levels and normal ovarian morphology, however, the pathogenesis of idiopathic hirsutism is not clear. The androgens are the main hormones to stimulate growth of body hair, therefore, there should be any form of increased androgen effect irrespective of normal serum androgen levels in any patient with hirsutism. In accordance to this scientific truth, we have previously shown that, although within normal limits, patients with idiopathic hirsutism have relatively higher serum androgen levels (relative hyperandrogenemia) in comparison to healthy subjects which let as to think that is idiopathic hirsutism really idiopathic? In addition to relative hyperandrogenemia, we have previously shown that, in comparison to healthy subjects, women with idiopathic hirsutism demonstrated higher expression of steroid sulphatase and 17-beta hydroxysteroid dehydrogenase mRNA both in the subumbilical region and arm skin, which contributes to local androgen metabolism. Those results support the idea that, in some patients, although the adrenals or ovaries do not secrete increased amount of androgens leading to hyperandrogenemia, pilocebaceous unit locally produce increased amount of androgens leading to hirsutism without ovulatory dysfunction. Upon the demonstration of relative hyperandrogenemia and possible increase in local androgen synthesis in patients with idiopathic hirsutism, we think that idiopathic hirsutism is not idiopathic and it may be named as "normoandrogenic hirsutism". Furthermore, it may not be a different entity but may be an early stage of hyperandrogenic disorders such as PCOS. Clinically, this can be find out by following-up patients with idiopathic hirsutism prospectively.

Investigating the biosynthesis and metabolism of 11β-hydroxyandrostenedione.

The "rediscovery" 11β-hydroxyandrostenedione (11OHA4) placed the spotlight on this unique adrenal-derived hormone with researchers and clinicians once again focusing on the steroid's presence in endocrine pathology. Little was known about the steroid other than its chemical characterisation and that a mitochondrial cytochrome P450 enzyme catalysed the 11β-hydroxylation of 11OHA4. The fact that neither the biosynthesis nor metabolism of 11OHA4 had been fully characterised presented an ideal opportunity to investigate the metabolic pathways. In addition, methodologies and analytical tools have improved vastly since 11OHA4 was first identified in the 1950s. Cell models, recombinant DNA technology and steroid quantification using liquid chromatography mass spectrometry have greatly facilitated investigations in the field of steroidogenesis. Evident from the structure is that 11OHA4 can be metabolised by hydroxysteroid dehydrogenases and reductases acting on the C4/C5 double bond and on functional moieties at specific carbons on the cyclopentane-perhydro-phenanthrene backbone of the steroid. In this chapter, the biosynthesis and metabolism of 11OHA4 is followed using two strategies that complement each another; (i) human cell models either transiently transfected with recombinant DNA or expressing endogenous steroidogenic enzymes and (ii) steroid identification and quantification using high resolution mass spectrometry. These methodologies have proven invaluable in the determination of 11OHA4's metabolic route. Both strategies are presented with the focus on the accurate identification and quantification of steroids using UHPLC-MS/MS and UPC2-MS/MS. The protocols described in this chapter lay a sound foundation which can aid the researcher and be adapted and implement in future studies.

In vivo Drug Screening to Identify Anti-metastatic Drugs in Twist1a-ERT2 Transgenic Zebrafish.

Here, we present an in vivo drug screening protocol using a zebrafish model of metastasis for the identification of anti-metastatic drugs. A tamoxifen-controllable Twist1a-ERT2 transgenic zebrafish line was established to serve as a platform for the identification. By crossing Twist1a-ERT2 with xmrk (a homolog of hyperactive form of the epidermal growth factor receptor) transgenic zebrafish, which develop hepatocellular carcinoma, approximately 80% of the double transgenic zebrafish show spontaneous cell dissemination of mCherry-labeled hepatocytes from the liver to the entire abdomen and tail regions in five days, through induction of epithelial to mesenchymal transition (EMT). This rapid and high-frequency induction of cell dissemination makes it possible to perform an in vivo drug screen for the identification of anti-metastatic drugs targeting metastatic dissemination of cancer cells. The protocol evaluates the suppressor effect of a test drug on metastasis in five days, by comparing the frequencies of the fish showing abdominal and distant dissemination patterns in the test drug-treated group with those in the vehicle-treated group. Our study previously identified that adrenosterone, an inhibitor for hydroxysteroid (11-beta) dehydrogenase 1 (HSD11β1), has a suppressor effect on cell dissemination in the model. Furthermore, we validated that a pharmacologic and genetic inhibition of HSD11β1 suppressed metastatic dissemination of highly metastatic human cell lines in a zebrafish xenotransplantation model. Taken together, this protocol opens new routes for the identification of anti-metastatic drugs. Graphical overview Timing Day 0: Zebrafish spawning Day 8: Primary tumor induction Day 11: Chemical treatment Day 11.5: Metastatic dissemination induction in the presence of a test chemical Day 16: Data analysis.

Isolation of Environmental Bacteria Able to Degrade Sterols and/or Bile Acids: Determination of Cholesterol Oxidase and Several Hydroxysteroid Dehydrogenase Activities in Rhodococcus, Gordonia, and Pseudomonas putida.

Interest about the isolation and characterization of steroid-catabolizing bacteria has increased over time due to the massive release of these recalcitrant compounds and their deleterious effects or their biotransformation derivatives as endocrine disruptors for wildlife, as well as their potential use in biotechnological approaches for the synthesis of pharmacological compounds. Thus, in this chapter, an isolation protocol to select environmental bacteria able to degrade sterols, bile acids, and androgens is shown. Moreover, procedures for the determination of cholesterol oxidase or different hydroxysteroid dehydrogenase activities in Pseudomonas putida DOC21, Rhodococcus sp. HE24.12, Gordonia sp. HE24.4J and Gordonia sp. HE24.3 are also detailed.

JS-ISH-ESH-1: DRUGS AND SUBSTANCES THAT INCREASE BLOOD PRESSURE

The use of herbal medications and substances that have been touted to raise BP is increasing globally with estimates for example 6 to 48 percent of persons use herbal therapies in Europe. Other medications such as NSAIDs are among the most widely used treatments globally. Several of such medications and substances may raise blood pressure or antagonize the BP lowering effects of antihypertensive therapy in individuals. The individual effect of these substances on blood pressure can be highly variable with greater increases noted in the elderly, those with higher baseline blood pressure, use of antihypertensive therapy, and co-existing renal disease. In this discussion, we will review the evidence for the most commonly implicated substances that have been considered to raise blood pressure including NSAIDs, oral contraceptives, hormone replacement therapy, and antidepressants. We will also review the evidence for herbal therapies that may raise BP. Coxibs are associated with a 3.9 mm Hg mean increase in SBP and nonselective NSAIDs 2.8 mm Hg compared with placebo. In meta-analyses, aspirin was not associated with a significant blood pressure increase. High regular acetaminophen dosing is associated with an increased risk of new onset hypertension. Oral contraceptives were associated with new onset hypertension in approximately 5 percent of users of high dose compounds that contain at least 50 mg estrogen but these high estrogen doses are not commonly used in current formulations. SNRI antidepressants were found to have a small increase in systolic blood pressure relative to SSRI antidepressants 1.5 mmHg. Patients prescribed SSRIs showed no significant differences in blood pressure changes compared with placebo. Despite their widespread use, there are few studies on the effects of herbal substances on blood pressure apart from licorice and ephedra or ma huang. Licorice contains glycyrrhizic acid that acts as an 11 beta hydroxysteroid dehydrogenase type 2 inhibitor causing pseudohyperaldosteronism. Given the widespread use of these medications and substances, physicians will have to screen for these medications and substances when assessing patients at risk for and with hypertension.

Wistar 흰쥐에서 안드로겐 수용체 증가를 통한 비수리 추출물의 남성 갱년기 개선 효과

Male menopause, also referred to as late-onset hypogonadism or andropause, is a clinical syndrome characterized by sexual dysfunction, depressive disorder, and insomnia secondary to a testosterone deficiency in older men. Lespedeza cuneata, which belongs to the Fabaceae family, possesses several biological properties, including antiinflammatory, anti-oxidant, and antidiabetic effects. We investigated the inhibitory effect of standardized L. cuneata extract (LCE) on andropause symptoms in vivo. LCE administration significantly increased serum levels of total testosterone without any effects on serum estrogen levels. Additionally, LCE significantly increased serum 17-beta hydroxysteroid dehydrogenase 13 protein levels and reduced levels of sex hormone-binding globulin (SHBG), a protein that blocks free testosterone movement into the cytosol. LCE significantly upregulated the expression of cAMP response element binding protein and androgen receptor (AR), which functions as a transcription factor to stimulate spermatogenesis-associated genes and results in increased numbers of AR-positive cells, such as Leydig and Sertoli cells in the testis. To summarize, LCE administration increases serum testosterone, reduces SHBG levels and upregulates AR expression. Therefore, LCE may be useful as alternative therapy for andropause in older men.

A comparative assessment of artificial and natural energy drinks in the epididymal and testicular milieu

"Artificial and natural energy drinks are both taken for increased energy, physical stamina, and alertness, although they differ in composition. This study investigated the effects of artificial and natural energy drinks on the testicular milieu in male pubertal rats. Eighteen Wistar rats were randomly divided into 3 groups of 6 rats each and all animals had access to food ad libitum. Group 1: (control) received water only; Group 2: (artificial energy drink- AED) received AED; Group 3: (natural energy drink- NED) received NED. A dose of 1.41ml/day/150g animal was administered and this lasted for 28 days. Sperm and testicular variables, biochemical parameters, and hormonal assays were carried out. There were significant decreases in the levels of testosterone, Lactate dehydrogenase, glucose, 3β-Hydroxysteroid dehydrogenase, and 17β- Hydroxysteroid dehydrogenase activities in AED and NED groups when compared to the control group. There was a marked increment in sperm abnormalities in the NED group when compared to AED and control groups. Also, the intake of AED led to an elevated level of Glucose-6-phosphate dehydrogenase compared to the control while a significant reduction was observed in the NED group when compared to the AED group. Artificial and natural energy drinks although consumed for strength and vigor distorted epididymis and testicular integrity via alteration of the testicular metabolism, lowering sperm quality and androgenic hormones in pubertal male Wistar rats. Keywords: energy drink, jaggery, sperm quality, steroidogenic enzymes, lactate dehydrogenase."

Expression Profile of Hydroxysteroid Dehydrogenase-like 2 in Polychaete Perinereis aibuhitensis in Response to BPA.

Hydroxysteroid dehydrogenases (HSDs) play an important role in the metabolism of steroids and xenobiotics. However, the function of HSDs in invertebrates is unclear. In this study, we cloned the hydroxysteroid dehydrogenase-like 2 (HSDL2) gene in Perinereis aibuhitensis, which is 1652 bp in length, encoding 400 amino acids. This sequence contains conserved short-chain dehydrogenase and sterol carrier protein-2 domain, and the alignment analysis showed its close relationship with other invertebrate HSDL2. Further, the tissue distribution analysis of the HSDL2 gene showed it is expressed strongly in the intestine. The expression level of HSDL2 after inducement with bisphenol A (BPA) was also detected both at transcriptional and translational levels. The results inferred that BPA exposure can induce HSDL2 expression, and the inductive effect was obvious in the high-concentration BPA group (100 μg/L). In summary, our results showed the detoxification function of HSDL2 in polychaetes.

Type-2 Diabetes Mellitus: A Review of Current Trends

Diabetes mellitus type 2 is a chronic metabolic condition whose incidence has been continuously growing around the world. As a result of this tendency, it is quickly becoming an epidemic in some parts of the world, with the number of individuals affected anticipated to double in the next decade as the world's population ages, adding to the already existing load on healthcare providers, particularly in poor developing countries. World Health Organization (WHO) guidelines are still used for screening and diagnosis, which include both clinical and laboratory characteristics. There is currently no cure for the disease; however, therapy options include lifestyle changes, obesity management, oral hypoglycemic medications, and insulin sensitizers such as metformin, a biguanide that reduces insulin resistance which is used as first line medication especially for obese patients. The traditional herbal medicine like garlic, neem, bitter gourd, aloe vera, onion, methi, amla, jamun, baheda etc. are also used for the treatment of the diabetes. Other effective medication includes non-sulfonylurea secretagogues, thiazolidinediones, alpha glucoside inhibitors, and insulin. Recent research into the biology of type 2 diabetes has led to the development of new medications such as glucagon-like peptide 1 analogues: dipeptidyl peptidase inhibitors, sodium glucose cotransporter 2 inhibitors, and 11-hydroxysteroid dehydrogenase 1 inhibitors, insulin releasing glucokinase activators. The novel drug delivery improves the treatment of diabetes by the inhaled insulin. Keywords: Type 2 diabetes mellitus, Diabetes complications, Diabetes diagnosis, Diabetes management, Antidiabetic newer drugs, Antidiabetic herbal drugs

Hypothalamic-Pituitary-Adrenal Hormones Impair Pig Fertilization and Preimplantation Embryo Development via Inducing Oviductal Epithelial Apoptosis: An In Vitro Study.

Previous studies show that stressful events after ovulation in sows significantly impaired the embryo cleavage with a significant elevation of blood cortisol. However, the effects of corticotropin-releasing hormone (CRH), adrenocorticotropic hormone (ACTH) and cortisol on fertilization and embryo development remain to be specified, and whether they damage pig embryos directly or indirectly is unclear. This study demonstrated that embryo development was unaffected when pig parthenotes were cultured with different concentrations of CRH/ACTH/cortisol. However, embryo development was significantly impaired when the embryos were cocultured with pig oviductal epithelial cells (OECs) in the presence of CRH/cortisol or cultured in medium that was conditioned with CRH/cortisol-pretreated OECs (CRH/cortisol-CM). Fertilization in CRH/cortisol-CM significantly increased the rates of polyspermy. CRH and cortisol induced apoptosis of OECs through FAS and TNFα signaling. The apoptotic OECs produced less growth factors but more FASL and TNFα, which induced apoptosis in embryos. Pig embryos were not sensitive to CRH because they expressed no CRH receptor but the CRH-binding protein, and they were tolerant to cortisol because they expressed more 11-beta hydroxysteroid dehydrogenase 2 (HSD11B2) than HSD11B1. When used at a stress-induced physiological concentration, while culture with either CRH or cortisol alone showed no effect, culture with both significantly increased apoptosis in OECs. In conclusion, CRH and cortisol impair pig fertilization and preimplantation embryo development indirectly by inducing OEC apoptosis via the activation of the FAS and TNFα systems. ACTH did not show any detrimental effect on pig embryos, nor OECs.

Review article: the role of HSD17B13 on global epidemiology, natural history, pathogenesis and treatment of NAFLD.

Non-alcoholic fatty liver disease (NAFLD) occurs in around a quarter of the global population and is one of the leading causes of chronic liver disease. The phenotypic manifestation and the severity of NAFLD are influenced by an interplay of environmental and genetic factors. Recently, several inactivating variants in the novel 17-Beta hydroxysteroid dehydrogenase 13 (HSD17B13) gene have been found to be associated with a reduced risk of chronic liver diseases, including NAFLD. To review the existing literature on the epidemiology of HSD17B13 and discuss its role in the natural history, disease pathogenesis and treatment of NAFLD. We extensively searched relevant literature in PubMed, Google Scholar, clinicaltrials.gov and the reference list of articles included in the review. HSD17B13 is a liver-specific, lipid droplet (LD)-associated protein that has enzymatic pathways involving steroids, pro-inflammatory lipid mediators and retinol. The estimated prevalence of the best characterised HSD17B13 variant (rs72613567) ranges from 5% in Africa to 34% in East Asia. Loss-of-function variants in HSD17B13 are protective against the progression of NAFLD from simple steatosis to non-alcoholic steatohepatitis (NASH), liver fibrosis, cirrhosis and hepatocellular carcinoma. Emerging data from mechanistic and preclinical studies with RNA interference (RNAi) and small molecule agents indicate that inhibiting HSD17B13 activity may prevent NAFLD progression. The loss-of-function polymorphisms of the newly identified HSD17B13 gene mitigate the progression of NAFLD. It is important to understand the exact mechanism by which these variants exert a protective effect and implement the gathered knowledge in the treatment of NAFLD.

Testosterone promotion effect of Eucommia ulmoides staminate flower via the steroidogenic pathway and potential hormonal mechanism

Eucommia ulmoides staminate flowers (EUF), a newly approved functional food in China, have great potential for hormonal regulation. Herein, we aim to demonstrate the chemical composition and pharmacological activity of EUF in testosterone production and hormonal regulation. EUF extract and its components, kaempferol and geniposidic acid, exhibited a strong stimulating effect by increasing testosterone secretion, reducing ROS production, or promoting viability in Leydig cells. Meanwhile, the increased testosterone production was related to the upregulation of mRNA and protein expression of the steroidogenic pathway, such as steroidogenic acute-regulatory protein (StAR), 3β -hydroxysteroid dehydrogenase type 1 (HSD3B1), 17α-hydroxylase/17,20-lyase (CYP17A1), and nuclear receptor subfamily 5 group A member 1 (NR5A1). However, PKA inhibitor H89 or adenylyl cyclase inhibitor SQ22536 could block their effect. The results of transgenic yeast models showed the androgenic agonistic effects of kaempferol and naringenin and the estrogenic agonistic effects of rutin. These results indicated that the testosterone promotional effect of EUF was related to the activation of the steroidogenic pathway and potential hormonal regulation. Kaempferol and geniposidic acid might be the key active ingredients.

ODP443 Targeting NAD+ availability modulates 11β- Hydroxysteroid dehydrogenase 1 activity in mouse and human cell lines and primary cells

Abstract Four nicotinamide adenine dinucleotide coenzymes – NAD+, NADH, NADP+, and NADPH – are the central catalysts of metabolism. The enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) is NADPH-dependent reductase that catalyses the reduction of the inactive glucocorticoid cortisone) to active cortisol (11-dehydrocorticosterone to corticosterone in rodents). 11β-HSD1 is in the endoplasmic reticulum (ER) lumen, and the level of NADPH in the ER regulates enzyme activity. Little is known about the biochemical mechanisms controlling cytosolic/ER NAD(P)(H) crosstalk. Here we examined how perturbed cellular NAD+ availability modulates 11β-HSD1 enzyme activity in the ER across a range of mouse and human cell types. 11β-HSD1 activity was measured in a series of transformed and primary mouse and human liver (HepG2), muscle, and Human Dermal Fibroblast cells (HDFn) depleted for NAD+ using FK866 to inhibit nicotinamide phosphoribosyl transferase (NAMPT), the rate-limiting enzyme in NAD+ biosynthesis. To replete NAD+ we supplemented cells with 0.5mM of NAD+ precursors nicotinamide riboside (NR) and nicotinamide mononucleotide (NMN). FK866 (100nM, 48h) was effective at depleting NAD+ by approximately 90% in C2C12 myotubes and 50% in HepG2 liver cells. In each cell line, 11β-HSD1 reductase activity was decreased by FK866 compared to untreated controls (80% in C2C12 P<0. 0001, 55% in HepG2, P=0. 03 and 65% in HDFn P=0. 002). Restoring NAD+ level with NR (0.5mM, 6h) in C2C12 and (0.5mM, 24h) in HepG2 and HDFn rescued 11β-HSD1 enzyme activity compared with no precursor treatment (60% in C2C12 P=0. 0007, 52% in HepG2 P=0. 03 and 68% in HDFn P=0. 0006). Human dermal fibroblasts treated with TNFa (10nM, 24h) boosted 11β-HSD1 expression 4-fold, however this was not able to overcome the FK866 induced drop-in reductase activity. These data show that depletion of NAD+ and impaired 11β-HSD1 reductase activity in the ER is a generalised phenomenon across cell types. This suggests a crosstalk between cytosolic and ER NAD(P)(H) pools that can respond rapidly to changes in NAD+ precursor availability. This could provide new insight into the regulation of cellular glucocorticoid metabolism and ER NAD(P)(H) homeostasis. Presentation: No date and time listed

PMON49 Checkpoint Inhibitor Induced Hypophysitis with Growth Hormone Deficiency and Implications for the Treatment of ACTH Deficiency

Abstract Background/Purpose Hypophysitis is a known complication of immune checkpoint inhibitor (ICI) therapies used in the treatment of malignant cancers. In general, central adrenal insufficiency (AI) and hypothyroidism are the most recognized deficiencies following a diagnosis of ICI induced hypophysitis. As a result, other hypothalamic-pituitary axes (HPAs) are either incompletely evaluated or are disregarded from a therapeutic standpoint. Of particular importance is the growth hormone axis which is well established to affect tissue level metabolism of cortisol and cortisone through its effect on 11 beta hydroxysteroid dehydrogenase in target tissues. Our hypothesis is that growth hormone deficiency is also present in ICI induced hypophysitis. Methods We queried current and legacy electronic medical records of patients who received ICI for clinical notes mentioning adrenal insufficiency and concern for hypophysitis using the EMERSE tool. Patients with ICI exposure were included if they had 2 of the following 3 criteria; new onset central AI as defined by a low ACTH and cortisol, pituitary imaging suggestive of hypophysitis, or clinical presentation concerning hypophysitis. Patients were excludedfrom the analysis if there was a history of brain irradiation or in whom the diagnosis was not clear due to high dose steroid exposure prior to biochemical evaluation. Results Twenty-nine patients met inclusion criteria. Out of 29 patients, 16 underwent testing of the growth hormone axis with IGF-1 (11) and/or random growth hormone level (5). Seventeen patients (59%) had biochemical evidence of deficiency of 3 or more HPAs. Twenty-six patients (90%) had biochemical evidence of 2 or more PHAs. Two patients only underwent evaluation of 2 pituitary axes so it is unknown if they indeed suffered from panhypopituitarism. Most patients (28/29) were prescribed hydrocortisone for AI with an average dose of 20 mg per day. Conclusions Our findings are suggestive that growth hormone deficiency may be present in ICI induced hypophysitis. This finding holds implications for cortisol replacement therapy for coexisting AI. When one considers the average adrenal cortisol production is 5.7–7.4 mg/m2/day or 9.5–9.9 mg/day, 20 mg of daily cortisol replacementmay be too high in the face of concomitant growth hormone deficiency. Practice changes may include lower doses ofhydrocortisone replacement or adjustment to cortisone acetate which has been shown to have more favorable effectson tissue level metabolic effects. Presentation: Monday, June 13, 2022 12:30 p.m. - 2:30 p.m.

Depleting NAD+ pools specifically in the endoplasmic reticulum lumen impairs 11[beta]-hydroxysteroid dehydrogenase activity

Searchable abstracts of presentations at key conferences in endocrinology ISSN 1470-3947 (print) | ISSN 1479-6848 (online)

RNA-seq and LC-MS/MS analysis of antiviral effects mediated by cold stress and stress hormone corticosterone in chicken DF-1 cells

Cold stress in poultry is a global problem that causes huge economic losses and threatens the health and welfare of poultry. However, knowledge of chicken responses to virus infection under cold stress is limited. The purpose of this research was to investigate the effects of cold stress on gene expression and viral replication in chicken DF-1 cells in hypothermia. In addition, the characterization of circulating steroid hormone profiles in the plasma of chickens under cold stress was analyzed by liquid chromatography-tandem mass spectrometry. Herein, we performed RNA sequencing to obtain DF-1 cell transcriptional profiles under cold stress. A total of 9499 differentially expressed genes (DEGs) were identified in DF-1 cells. Overexpressed DEGs were related to the proteasome, cell cycle, spliceosome, ribosome biogenesis, and mammalian target of rapamycin (mTOR). Down-regulated DEGs were related to ribosomes, oxidative phosphorylation, apoptosis, and the p53 signaling pathway. Gene set enrichment analysis showed that the DEGs mainly affect host ribosome translation and mitochondrial respiratory electron transport. The principal steroid hormone alterations in chickens subjected to cold stress included dihydrotestosterone, testosterone, β-sitosterol, androstenedione, 7a,27-dihydroxycholesterol,7-ketocholesterol, and desmosterol, which are associated with endocrine resistance, ovarian steroidogenesis, and steroid hormone biosynthesis. In addition, Infectious bronchitis virus (IBV), Newcastle disease virus (NDV), and Influenza A (H9N2) Virus replication in DF-1 cells is significantly inhibited by cold stress. Moreover, the plasma concentrations of corticosterone, an important stress hormone in poultry, were significantly elevated in chickens subjected to cold stress, and we found that IBV and vesicular stomatitis virus (VSV) replication were strongly inhibited in DF-1 cells pretreated with CORT, but NDV and H9N2 replication were unaffected. In conclusion, in response to cold stress, the translation efficiency and mitochondrial respiratory chain are temporarily weakened in DF-1 cells, which affects virus replication. Chickens may regulate aromatase deficiency, androstenedione metabolism, androgen and estrogen metabolism, and 17-beta hydroxysteroid dehydrogenase III deficiency through steroid hormones in response to cold stress. This study provides valuable insights into the molecular regulatory mechanisms of poultry under cold stress and may support further research on the intrinsic link between steroid hormones and virus replication under stress.

11β Hydroxysteroid dehydrogenase type 1 Gene Expression in Obesity and its Complications

11β Hydroxysteroid dehydrogenase type 1 Gene Expression in Obesity and its Complications

Functional Characterization and Synthetic Application of Is2-SDR, a Novel Thermostable and Promiscuous Ketoreductase from a Hot Spring Metagenome.

In a metagenome mining-based search of novel thermostable hydroxysteroid dehydrogenases (HSDHs), enzymes that are able to selectively oxidize/reduce steroidal compounds, a novel short-chain dehydrogenase/reductase (SDR), named Is2-SDR, was recently discovered. This enzyme, found in an Icelandic hot spring metagenome, shared a high sequence similarity with HSDHs, but, unexpectedly, showed no activity in the oxidation of the tested steroid substrates, e.g., cholic acid. Despite that, Is2-SDR proved to be a very active and versatile ketoreductase, being able to regio- and stereoselectively reduce a diversified panel of carbonylic substrates, including bulky ketones, α- and β-ketoesters, and α-diketones of pharmaceutical relevance. Further investigations showed that Is2-SDR was indeed active in the regio- and stereoselective reduction of oxidized steroid derivatives, and this outcome was rationalized by docking analysis in the active site model. Moreover, Is2-SDR showed remarkable thermostability, with an apparent melting temperature (TM) around 75 °C, as determined by circular dichroism analysis, and no significant decrease in catalytic activity, even after 5 h at 80 °C. A broad tolerance to both water-miscible and water-immiscible organic solvents was demonstrated as well, thus, confirming the potential of this new biocatalyst for its synthetic application.

Fructose promotes more than glucose the adipocytic differentiation of pig mesenchymal stem cells.

The goal of this study was to evaluate how glucose and fructose affected the adipose differentiation of pig newborn mesenchymal stem cells (MSCs). Cells were grown with or without inosine in 7.5 mM glucose (substituted with 1.5 or 6 mM fructose). MSCs displayed adipose morphology after 70 days of differentiation. Fructose stimulated the highest levels of PPARγ and C/EBPβ. Fructose at 6 mM, but not glucose at 7.5 mM or fructose at 1.5 mM, promotes differentiation of MSCs into adipocytes and increases 11-hydroxysteroid dehydrogenase (11β-HSD1) and NADPH oxidase 4 (NOX4) mRNA in the absence of hepatic effects (as simulated by the inosine). Fructose and glucose increased xanthine oxide-reductase (XOR) catalytic activity almost 10-fold and elevated their products: intracellular reactive oxygen species (ROS) pool, extracellular H2 O2 pool by 4 orders of magnitude, and uric acid by a factor of 10. Therefore, in our experimental model, differentiation of MSCs into adipocytes occurs exclusively at the blood concentration of fructose detected after ingestion by people on a high fructose diet. PRACTICAL APPLICATIONS: The results of this study provide new evidence for fructose's adipogenic potential in mesenchymal stem cells, a model in which its effects on XOR activity had not been studied. The increased expression of genes such as C/EBPβ, PPARγ, and NOX4, as well as the increased XOR activity and high production of ROS during the differentiation process in the presence of fructose, coincides in pointing to this hexose as an important factor in the development of adipogenesis in young animals, which could have a great impact on the development of future obesity.