α-Linolenic acid-regulated testosterone biosynthesis via activation of the JNK-SF-1 signaling pathway in primary rooster Leydig cells.

As a functional fatty acid, α-linolenic acid (ALA) is essential in promoting animal testosterone biosynthesis. This study investigated the effects of ALA on testosterone biosynthesis and the possible mechanism underlying the signaling pathway in primary Leydig cells of the rooster. METHODS: Primary rooster Leydig cells were treated with ALA (0, 20, 40, or 80 µmol/L) or pretreated with a p38 inhibitor (50 µmol/L), a c-Jun NH2-terminal kinase (JNK) inhibitor (20 µmol/L), or an extracellular signal-regulated kinase (ERK) inhibitor (20 µmol/L) before ALA treatment. Testosterone content in the conditioned culture medium was detected using an enzyme-linked immunosorbent assay (ELISA). The expression of steroidogenic enzymes and JNK-SF-1 signaling pathway factors was detected using real-time fluorescence quantitative PCR (qRT-PCR). RESULTS: Supplementation with ALA significantly increased testosterone secretion within culture media (P < 0.05), and the optimized dose was 40 µmol/L. Compared with the control group, steroidogenic acute regulatory protein (StAR), cholesterol side-chain cleavage enzyme (P450scc), and 3ß-hydroxysteroid dehydrogenase (3ß-HSD) mRNA expression significantly increased (P < 0.05) in the 40 µmol/L ALA group; 17-hydroxylase/c17-20 lyase (P450c17) and p38 mRNA expressions were not significantly different in the 40 µmol/L ALA group; ERK and JNK mRNA expressions were significantly upregulated (P < 0.05) in 40 µmol/L ALA group. In the inhibitor group, testosterone levels were significantly downregulated (P < 0.05). Compared with the 40 µmol/L ALA group, StAR, P450scc, and P450c17 mRNA expressions were significantly decreased (P < 0.05), and 3ß-HSD mRNA expression in the p38 inhibitor group did not change; StAR, P450scc, and 3ß-HSD mRNA expressions were significantly decreased (P < 0.05), and P450c17 mRNA expression in ERK inhibitor group did not change; StAR, P450scc, 3ß-HSD, and P450c17 mRNA expressions were significantly decreased (P < 0.05) in JNK inhibitor group. Additionally, the increased steroidogenic factor 1 (SF-1) gene expression levels induced by ALA were reversed when the cells were pre-incubated with JNK and ERK inhibitors. The levels in the JNK inhibitor group were significantly lower than those in the control group (P < 0.05). CONCLUSION: ALA may promote testosterone biosynthesis by activating the JNK-SF-1 signaling pathway to upregulate StAR, P450scc, 3ß-HSD, and P450c17 expression in primary rooster Leydig cells.

Inhibition of high-fat diet-induced inflammatory responses in adipose tissue by SF1-expressing neurons of the ventromedial hypothalamus.

Inflammation and thermogenesis in white adipose tissue (WAT) at different sites influence the overall effects of obesity on metabolic health. In mice fed a high-fat diet (HFD), inflammatory responses are less pronounced in inguinal WAT (ingWAT) than in epididymal WAT (epiWAT). Here we show that ablation and activation of steroidogenic factor 1 (SF1)-expressing neurons in the ventromedial hypothalamus (VMH) oppositely affect the expression of inflammation-related genes and the formation of crown-like structures by infiltrating macrophages in ingWAT, but not in epiWAT, of HFD-fed mice, with these effects being mediated by sympathetic nerves innervating ingWAT. In contrast, SF1 neurons of the VMH preferentially regulated the expression of thermogenesis-related genes in interscapular brown adipose tissue (BAT) of HFD-fed mice. These results suggest that SF1 neurons of the VMH differentially regulate inflammatory responses and thermogenesis among various adipose tissue depots and restrain inflammation associated with diet-induced obesity specifically in ingWAT.

Conversion of human fibroblasts into functional Leydig-like cells by small molecules and a single factor.

Reprogramming fibroblasts into Leydig cells (LCs) offers a promising source for cell-based therapy for male hypogonadism. Recently, it has been achieved by forced expression of multiple transcription factors (TFs). However, for ultimate safe and convenient application, small molecules would be a revolutionary and desirable method to reduce or eliminate the genetic manipulations. Here, we report a defined small-molecule cocktail that enables the highly efficient conversion of human fibroblasts into functional LCs with only one transcription factor. These induced cells resembled human LCs with respect to morphology, marker gene expression and secretary function of testosterone. This study lays a foundation for future pharmacological reprogramming and provides a unique venue for investigating mechanisms underlying reprogramming.