Expression of Vascular Endothelial Growth Factor Receptors During Male Germ Cell Differentiation in the Mouse1

Abstract

Overexpression of vascular endothelial growth factor (VEGF) in the testis of transgenic mice induces infertility, suggesting a potential role for VEGF in the process of spermatogenesis. Spermatogenesis occurs within the confines of the seminiferous tubules, and the seminiferous epithelium lining these tubules consists of Sertoli cells and germ cells in various stages of maturation. We investigated the source of VEGF and VEGF-target cells within the seminiferous tubules of the normal mouse testis. Sections of testes fixed in Bouin solution and embedded in paraffin were subjected to immunofluorescent staining with specific antibodies against VEGF, and its receptors, VEGFR-1 (Flt-1) and VEGFR-2 (Flk-1). Total RNA was extracted from isolated populations of Sertoli cells, type A spermatogonia, pachytene spermatocytes, and spermatids. Primer pairs specific for VEGF and its receptors were designed and reverse-transcriptase polymerase chain reaction (RT-PCR) was performed. Immunofluorescent studies indicated that VEGF is strongly expressed in the cytoplasm of Sertoli cells. VEGFR-1 and VEGFR-2 were not expressed by the Sertoli cell. In contrast, a differential expression of VEGF receptors was observed in germ cells. Although VEGFR-2 was expressed in the cytoplasm of type A spermatogonia, VEGFR-1 was expressed in the acrosomal region of spermatids and spermatozoa. Pachytene spermatocytes did not exhibit any staining. Further, we examined the transcription of VEGF and its receptors by RT-PCR. VEGF was actively transcribed only in Sertoli cells. The transcription of VEGFR-2 was confined to type A spermatogonia. Interestingly, VEGFR-1 was transcribed both in pachytene spermatocytes and round spermatids. The mRNA expression of VEGFR-1 and VEGFR-2 in germ cells was inversely correlated during postnatal development of the mouse testis. Thus, VEGF may play a potential role in regulating the initial stages of the process of spermatogonial proliferation through VEGFR-2 and spermiogenesis through VEGFR-1.

NAMPT (visfatin) in the chicken testis: influence of sexual maturation on cellular localization, plasma levels and gene and protein expression

“…VEGF increases NAMPT expression in the chicken testis vascular endothelial cell proliferation in the testis (Rudolfsson et al 2004) and can therefore affect testicular vascularization. VEGF expression has been detected in mouse Sertoli cells, whereas its receptors are located in spermatocytes and spermatids (Nalbandian et al 2003). Hypoxia increases testosterone release in the mouse TM3 cell line mediated by increased VEGFA production (Hwang et al 2007).…”

Vascular endothelial growth factor regulates germ cell survival during establishment of spermatogenesis in the bovine testis

“…Based on the data presented here, we hypothesize that proliferating undifferentiated germ cells may produce VEGF in response to local hypoxia and oxidative stress to prevent apoptosis in these cells preceding the onset of puberty. Accordingly, testicular germ cells of several species express KDR and FLT1 (Ergun et al 1997, Nalbandian et al 2003, Rudolfsson et al 2004, and although these receptors are expressed in endothelial cells, no active angiogenesis takes place within the postnatal testis. VEGF synthesis by Leydig cells is stimulated in response to hCG/LH (Rudolfsson et al 2004), and FSH triggers Sertoli cell production of VEGF in the adult testis (Marti & Risau 1998, Liu & Yang 2004, Reisinger et al 2007.…”

“…VEGF synthesis by Leydig cells is stimulated in response to hCG/LH (Rudolfsson et al 2004), and FSH triggers Sertoli cell production of VEGF in the adult testis (Marti & Risau 1998, Liu & Yang 2004, Reisinger et al 2007. Interestingly, FSH has been shown to stimulate proliferation of spermatogonia (De Rooij et al 1989), and type A spermatogonia express KDR in neonatal mice (Nalbandian et al 2003). Thus, autocrine and paracrine regulation of germ cell activity by VEGF in the mammalian testis during early development and adulthood may be important for the initiation and maintenance of sperm production.…”

“…Rodent studies have shown that male fetal germ cells express VEGF protein until shortly after birth (Bott et al 2006), and Sertoli cells continue to produce VEGF throughout adult life (Marti & Risau 1998, Nalbandian et al 2003. Interestingly, type A spermatogonia in close association with Sertoli cells also express KDR but do not produce VEGF in mice aged 5 days post partum and older.…”

“…An increasing number of reports suggest that VEGFA has direct effects on nonvascular cells including neuronal cells (Marti & Risau 1998, Wada et al 2006, Nishijima et al 2007), muscle cells (Germani et al 2003, Bryan et al 2008, and granulosa cells (Greenaway et al 2004). Data also suggest potential functional roles for VEGFA in the postnatal testis, as KDR and FLT1 are expressed in testicular germ cells in humans (Ergun et al 1997), rats (Rudolfsson et al 2004), and mice (Nalbandian et al 2003), while somatic Sertoli and Leydig cells produce VEGFA (Liu & Yang 2004). Two studies have shown that overexpression of VEGFA-120 and VEGFA-165 causes detrimental effects on male fertility in transgenic mice (Korpelainen et al 1998, Huminiecki et al 2001.…”