Heat stress upregulates arachidonic acid to trigger autophagy in sertoli cells via dysfunctional mitochondrial respiratory chain function.

As a key factor in determining testis size and sperm number, sertoli cells (SCs) play a crucial role in male infertility. Heat stress (HS) reduces SCs counts, negatively impacting nutrient transport and supply to germ cells, and leading to spermatogenesis failure in humans and animals. However, how HS affects the number of SCs remains unclear. We hypothesized that changes in SC metabolism contribute to the adverse effects of HS. In this study, we first observed an upregulation of arachidonic acid (AA), an unsaturated fatty acid after HS exposure by LC-MS/MS metabolome detection. By increasing ROS levels, expression of KEAP1 and NRF2 proteins as well as LC3 and LAMP2, 100 µM AA induced autophagy in SCs by activating oxidative stress (OS). We observed adverse effects of AA on mitochondria under HS with a decrease of mitochondrial number and an increase of mitochondrial membrane potential (MMP). We also found that AA alternated the oxygen transport and absorption function of mitochondria by increasing glycolysis flux and decreasing oxygen consumption rate as well as the expression of mitochondrial electron transport chain (ETC) proteins Complex I, II, V. However, pretreatment with 5 mM NAC (ROS inhibitor) and 2 µM Rotenone (mitochondrial ETC inhibitor) reversed the autophagy induced by AA. In summary, AA modulates autophagy in SCs during HS by disrupting mitochondrial ETC function, inferring that the release of AA is a switch-like response, and providing insight into the underlying mechanism of high temperatures causing male infertility.

Glutamate rescues heat stress-induced apoptosis of Sertoli cells by enhancing the activity of antioxidant enzymes and activating the Trx1-Akt pathway in vitro.

Heat stress reduces the number of Sertoli cells, which is closely related to an imbalanced redox status. Glutamate functions to maintain the equilibrium of redox homeostasis. However, the role of glutamate in heat treated Sertoli cells remains unclear. Herein, Sertoli cells from 3-week-old piglets were treated at 44 °C for 30 min (heat stress). Glutamate levels increased significantly following heat stress treatment, followed by a gradual decrease during recovery, while glutathione (GSH) showed a gradual increase. The addition of exogenous glutamate (700 µM) to Sertoli cells before heat stress significantly reduced the heat stress-induced apoptosis rate, mediated by enhanced levels of antioxidant substances (superoxide dismutase (SOD), total antioxidant capacity (TAC), and GSH) and reduced levels of oxidative substances (reactive oxygen species (ROS) and malondialdehyde (MDA)). Glutamate addition to Sertoli cells before heat stress upregulated the levels of glutamate-cysteine ligase, modifier subunit (Gclm), glutathione synthetase (Gss), thioredoxin (Trx1) and B-cell leukemia/lymphoma 2 (Bcl-2), and the ratio of phosphorylated Akt (protein kinase B)/total Akt. However, it decreased the levels of Bcl2-associated X protein (Bax) and cleaved-caspase 3. Addition of the inhibitor of glutaminase (Gls1), Bptes (Bis-2-(5-phenylacetamido-1,3,4-thiadiazol-2-yl)ethyl sulfide, 30 µM)to Sertoli cells before heat stress reversed these effects. These results inferred that glutamate rescued heat stress-induced apoptosis in Sertoli cells by enhancing activity of antioxidant enzymes and activating the Trx1-Akt pathway. Thus, glutamate supplementation might represent a novel strategy to alleviate the negative effect of heat stress.

Melatonin alleviates the heat stress-induced impairment of Sertoli cells by reprogramming glucose metabolism.

Sertoli cells (SCs) provide structural and nutritional support for developing germ cells. Normal glucose metabolism of SCs is necessary for spermatogenesis. Melatonin could alleviate the effects of heat stress on spermatogenesis. However, the influences of heat stress on glucose metabolism in SCs remain unclear, and the potential protective mechanisms of melatonin on SCs need more exploration. In this study, boar SCs were treated at 43°C for 30 min, and different concentrations of melatonin were added to protect SCs from heat stress-induced impairment. These results showed that heat stress-induced oxidative stress caused cell apoptosis, inhibited the pentose phosphate pathway, and decreased the ATP content. Furthermore, heat stress increased the expressions of glucose intake- and glycolytic-related enzymes, which enhanced the glycolysis activity to compensate for the energy deficit. Melatonin relieved heat stress-induced oxidative stress and apoptosis by activating the Kelch-like ECH-associated protein 1 (KEAP1)/NF-E2-related factor 2 signaling pathway to increase the capacity of antioxidants. In addition, melatonin enhanced heat-shock protein 90 (HSP90) expression through melatonin receptor 1B (MTNR1B), thereby stabilizing hypoxia-inducible factor-1α (HIF-1α). Activation of the HIF-1α signaling pathway enhanced glycolysis, promoted the pentose phosphate pathway, and increased cell viability. Our results suggest that melatonin reprograms glucose metabolism in SCs through the MTNR1B-HSP90-HIF-1α axis and provides a theoretical basis for preventing heat stress injury.

A new wortmannine derivative from a Tripterygium wilfordii endophytic fungus Talaromyces wortmannii LGT-4.

A new wortmannine derivative named wortmannine E (1) was isolated from Talaromyces wortmannii LGT-4, an endophytic fungus of Tripterygium wilfordii. Its structure was established by 1D and 2D NMR spectra.

Colletotrilactam A-D, novel lactams from Colletotrichum gloeosporioides GT-7, a fungal endophyte of Uncaria rhynchophylla.

Four novel lactams, colletotrilactam A-D (1-4), along with six known compounds (5-10) were isolated from the culture broth of Colletotrichum gloeosporioides GT-7, a fungal endophyte of Uncaria rhynchophylla. The structures of these compounds were elucidated by comprehensive NMR spectroscopy. Isolates were tested for monoamine oxidase (MAO) inhibitory activity and compound 9 showed potent MAO inhibitory activity with IC50 value of 8.93±0.34µg/mL, when the IC50 value of iproniazid as a standard was 1.80±0.5µg/mL.