A chronic high-fat diet causes sperm head alterations in C57BL/6J mice.

A chronic-positive energetic balance has been directly correlated with infertility in men, but the involved mechanisms remain unknown. Herein we investigated weather in a mouse model a chronic feeding with a diet supplemented with chicken fat affects sperm head morphology. To accomplish this, we fed mice for 16 weeks with either control food (low-fat diet, LFD) or control food supplemented with 22% chicken fat (high-fat diet, HFD). At the end of the feeding regimen, we measured: redox and inflammatory changes, cholesterol accumulation in testis and analyzed testicular morphological structure and ultra-structure and liver morphology. We found that the mice fed HFD resembled some features of the human metabolic syndrome, including systemic oxidative stress and inflammation, this group showed an increment in the following parameters; central adiposity (adiposity index: 1.07 ± 0.10 vs 2.26 ± 0.17), dyslipidemia (total cholesterol: 153.3 ± 2.6 vs 175.1 ± 8.08 mg/dL), insulin resistance (indirect Insulin resistance index, TG/HDL-c: 2.94 ± 0.33 vs 3.68 ± 0.15) and fatty liver. Increased cholesterol content measured by filipin was found in the testicles from HFD (fluorescence intensity increase to 50%), as well as an alteration of spermiogenesis. Most remarkably, a disorganized manchette-perinuclear ring complex and an altered morphology of the sperm head were observed in the spermatozoa of HFD-fed mice. These results add new information to our understanding about the mechanisms by which systemic oxidative stress and inflammation may influence sperm-head morphology and indirectly male fertility.

SPINK3 modulates mouse sperm physiology through the reduction of nitric oxide level independently of its trypsin inhibitory activity.

Serine protease inhibitor Kazal-type (SPINK3)/P12/PSTI-II is a small secretory protein from mouse seminal vesicle which contains a KAZAL domain and shows calcium (Ca(2+))-transport inhibitory (caltrin) activity. This molecule was obtained as a recombinant protein and its effect on capacitated sperm cells was examined. SPINK3 inhibited trypsin activity in vitro while the fusion protein GST-SPINK3 had no effect on this enzyme activity. The inactive GST-SPINK3 significantly reduced the percentage of spermatozoa positively stained for nitric oxide (NO) with the specific probe DAF-FM DA and NO concentration measured by Griess method in capacitated mouse sperm; the same effect was observed when sperm were capacitated under low Ca(2+) concentration, using either intracellular (BAPTA-AM) or extracellular Ca(2+) (EDTA) chelators. The percentage of sperm showing spontaneous and progesterone-induced acrosomal reaction was significantly lower in the presence of GST-SPINK3 compared to untreated capacitated spermatozoa. Interestingly, this decrease was overcome by the exogenous addition of the NO donors, sodium nitroprusside (SNP), and S-nitrosoglutathione (GSNO). Phosphorylation of sperm proteins in tyrosine residues was partially affected by GST-SPINK3, however, only GSNO was able to reverse this effect. Sperm progressive motility was not significantly diminished by GST-SPINK3 or BAPTA-AM but enhanced by the addition of SNP. This is the first report that demonstrates that SPINK3 modulates sperm physiology through a downstream reduction of endogenous NO concentration and independently of SPINK3 trypsin inhibitory activity.