Stress granules (SGs) are membraneless ribonucleoprotein (RNP)-based cellular foci formed in response to stress, facilitating cell survival by protecting against damage. Mammalian spermatogenesis should be maintained below body temperature for proper development, indicating its vulnerability to heat stress (HS). In this study, biotin tracer permeability assays showed that the inhibition of heat-induced SG assembly in the testis by 4–8 mg/kg cycloheximide significantly increased the percentage of seminiferous tubules with a damaged blood–testis barrier (BTB). Western blot results additionally revealed that the suppression of heat-induced SG assembly in Sertoli cell line, TM4 cells, by RNA inference of G3bp1/2 aggravated the decline in the BTB-related proteins ZO-1, β-Catenin and Claudin-11, indicating that SGs could protect the BTB against damage caused by HS. The protein components that associate with SGs in Sertoli cells were isolated by sequential centrifugation and immunoprecipitation, and were identified by liquid chromatography with tandem mass spectrometry. Gene Ontology and KEGG pathway enrichment analysis revealed that their corresponding genes were mainly involved in pathways related to proteasomes, nucleotide excision repair, mismatch repair, and DNA replication. Furthermore, a new SG component, the ubiquitin associated protein 2 (UBAP2), was found to translocate to SGs upon HS in TM4 cells by immunofluorescence. Moreover, SG assembly was significantly diminished after UBAP2 knockdown by RNA inference during HS, suggesting the important role of UBAP2 in SG assembly. In addition, UBAP2 knockdown reduced the expression of ZO-1, β-Catenin and Claudin-11, which implied its potential role in the function of the BTB. Overall, our study demonstrated the role of SGs in maintaining BTB functions during HS and identified a new component implicated in SG formation in Sertoli cells. These findings not only offer novel insights into the biological functions of SGs and the molecular mechanism of low fertility in males in summer, but also potentially provide an experimental basis for male fertility therapies.

S100A4 has been suggested to be a critical regulator of tumor metastasis and is implicated in the progression of inflammation. The aim of this study is to investigate the expression and possible role of S100A4 in epididymitis. Using a mouse model of epididymitis induced by the injection of lipopolysaccharide (LPS) in the deferent duct, we found that LPS administration induced an upregulation of S100a4 transcription (P < 0.05) and a recruitment of S100A4 positive cells in the epididymal interstitium of wild type (WT) mice. Co-immunofluorescence showed that S100A4 was mainly expressed by granulocytes, CD4 lymphocytes and macrophages. Deficiency of S100A4 reduced epididymal pathological reaction and the mRNA levels of the pro-inflammatory cytokines IL-1β and TNF-α (P < 0.01), suggesting that S100A4 promotes the progression of epididymitis. Furthermore, S100A4 deficiency alleviated the decline of sperm motility and rectified the abnormal expression of sperm membrane protein AMAD3, which suggested that in the progression of epididymitis, S100A4 aggravates the damage to sperm vitality. In addition, both Ki-67 marked cell proliferation and TUNEL detected cell apoptosis were reduced in S100a4^−/− mice compared with WT mice after LPS treatment, indicating that S100A4 promotes both cell proliferation and cell apoptosis in epididymitis. Overall, these results demonstrate that S100A4 promotes the progression of LPS-induced epididymitis and facilitates a decline in sperm vitality, and its function may be related to the process of cell proliferation and apoptosis during inflammation.

Heat stress impairs spermatogenesis primarily through increased testicular oxidative stress. Astaxanthin, a potent antioxidant, has shown beneficial effects in sperm cryopreservation. However, its potential to mitigate testicular damage induced by elevated temperatures remains uninvestigated. In this study, male mice were administered astaxanthin (10 mg/kg/d, 50 mg/kg/d, and 100 mg/kg/d) via gavage for 21 days, with heat exposure occurring during the final 14 days. Samples were collected after the last treatment or following a recovery period. Spermatozoa in the cauda epididymis were assessed using computer-aided sperm analysis (CASA) or Diff-Quik staining, and serum testosterone levels and oxidative stress markers in both serum and testis were quantified via enzyme-linked immunosorbent assay (ELISA). Heat treatment resulted in significant reductions in sperm motility, viability, and morphological integrity. However, daily supplementation of astaxanthin at 50 mg/kg and 100 mg/kg effectively alleviated these heat-induced impairments. Furthermore, astaxanthin at 50 mg/kg/d notably improved testis weight, testis index, and serum testosterone levels under heat stress. Its antioxidant capacity was validated by significant restoration of total antioxidant capacity (T-AOC), superoxide dismutase (SOD), and reduction in malondialdehyde (MDA) levels in both testis and serum. In conclusion, this study highlights the protective effects of astaxanthin against heat-induced testicular and sperm damage by reducing oxidative stress, supporting its potential use as a nutritional or nutraceutical supplement to promote male reproductive health, particularly in the context of summer subfertility in farm animals.

In order to establish a lighting regime suitable for rabbit farms in East China, the effects of lighting schedule, intensity and colour on the reproductive performance of rabbit does were evaluated by three experiments, respectively. In experiment 1, does were exposed to different lighting schedules: 16L:8D-continuous, 16L:8D-18d (6 d before artificial insemination (AI) to 12 d post-AI), 16L:8D-6d (6 d before AI to the day of AI) and 12L:12D-continuous. In experiment 2, does were exposed to different light intensities: 40 lx, 60 lx, 80 lx and 120 lx. In experiment 3, does were exposed to different light colours: white, yellow, blue and red. For all experiments, conception rate, kindling rate and pre-weaning mortality were calculated; litter size at birth, litter weight at birth, litter size at weaning, litter weight at weaning and individual kit weight at weaning were recorded. Results showed that none of the reproductive parameters of does were affected by the application of 16L:8D-18d lighting schedule compared with the continuous 16L:8D group(P>0.05). Moreover, rabbits does exposed to 80 lx light performed as well as those under 120 lx light in conception rate, kindling rate, litter size (total and alive) at birth and litter weight at birth (P>0.05). Furthermore, the exposures of 60 lx and 80 lx light were beneficial for litter weight at weaning. In addition, red light had a positive effect, as it led to a larger litter size and litter weight at weaning and lower pre-weaning mortality than white light (P<0.05). In summary, a 16L:8D photoperiod with 80 lx red light from 6 d before AI to 12 d post-AI is recommended for use in breeding of rabbit does according to our results.