The epididymis-specific antimicrobial peptide β-defensin 15 is required for sperm motility and male fertility in the rat (Rattus norvegicus)

Abstract

Spermatozoa acquire forward motility and fertilizing capacity during their transit through the epididymis. The maturation process involves modifications of the sperm surface by different proteins that are secreted by a series of specialized regions in the epididymal epithelium. Here we show that the rat epididymis-specific β-defensin 15 (Defb15) exhibits an androgen-dependent expression pattern, and it can bind to the acrosomal region of caput sperm. Coculture of caput spermatozoa with Defb15 antibody in vitro resulted in a significant decline in sperm motility. Moreover, the total and progressive motility of the spermatozoa dramatically decreased in rats when Defb15 was downregulated by lentivirus-mediated RNAi in vivo. Remarkably, knock down of Defb15 led to a reduction in fertility and embryonic development failure. In addition, the recombinant Defb15 showed antimicrobial activity in a dose-dependent fashion. These results suggest that Defb15 plays a dual role in both sperm maturation and pathogen defense in rat epididymis.

The Novel Human β-Defensin 114 Regulates Lipopolysaccharide (LPS)-mediated Inflammation and Protects Sperm from Motility Loss

“…Although it is still unclear why such a wide range of ␤-defensins are present in the male reproductive tract, their diverse functions can be briefly classified into three types: their common role in innate immunity by exhibiting antimicrobial activity in the reproductive system (28, 39) and non-reproductive systems (19); sperm maturation-related functions (28,40,41); and immunoregulating activity, which has been shown in nonreproductive systems (19). In addition to their expression in reproductive tissues, the majority of human ␤-defensins are also expressed in non-reproductive tissues (19).…”

“…1d). The generation of the antibody followed the protocols described previously (28). The antibody could recognize Ͻ5 ng of antigen protein in Western blot assays.…”

Spink13, an Epididymis-specific Gene of the Kazal-type Serine Protease Inhibitor (SPINK) Family, Is Essential for the Acrosomal Integrity and Male Fertility

“…Identification and characterization of differentially expressed genes in epididymis have provided us additional insight into the mechanisms of sperm maturation. Our previous study demonstrated that several epididymal proteins, such as Bin1b (5), HongrES1 (46), and Defb15 (44), are essential in sperm motility, capacitation, and male fertility, respectively. In the present study, we identified an epididymis-specific gene, Spink13, and demonstrated its physiological function in regulation of acrosome reaction.…”

“…Protocols used for examination of the knockdown efficiency and production of lentiviral particles were as previously described (44). Briefly, the psiRNA constructs and a pcDNA 3.0 vector expressing Spink13 were co-transfected into the mouse epididymal epithelial cells (PC1) using Lipofectamine 2000 (Invitrogen).…”

“…(c) Detection of tyrosine phosphorylation: sperm collected at different capacitation times were treated as previously reported. Western blot analysis was performed by using anti-phosphotyrosine monoclonal antibody as previously described (44). (d) Evaluation of sperm capacitation and acrosome reaction: Chlortetracycline fluorescence staining method, as used in previous studies (46), was conducted to morphologically assess sperm capacitation.…”