Aim : The standard management of varicocele repair is the subject of ongoing controversy. We retrospectively evaluated three surgical methods of varicocele treatment to determine the minimally invasive and most effective procedure. Methods : We performed 144 varicocelectomies on infertile patients with left clinical varicocele. Of the patients, 50 were treated with retroperitoneal high ligation under lumbar anesthesia, 33 with laparoscopic ligation under general anesthesia, and 61 with subinguinal microscopic ligation under local anesthesia. Operative time, hospital days, and clinical outcomes were compared between these techniques. Results : The operating time and hospitalization period required for subinguinal microscopic ligation was significantly shorter compared to those for the other procedures. All patients treated with subinguinal microscopic ligation could achieve normal activity as soon as they returned to their rooms. Postoperative complications were observed in five (10.0%) cases treated with high ligation and three (9.1%) laparoscopic cases, but were not observed after the subinguinal procedure. There were six cases (12.0%) of recurrence in the high ligation group and six (6.1%) in the laparoscopic group, but none in the subinguinal group. Sperm density was significantly improved in all procedures postoperatively, but sperm motility was not improved. The two-year pregnancy rate calculated by the Kaplan-Meier method was 35.8% for high ligation, 40.4% for laparoscopic ligation and 50.9% for subinguinal microscopic ligation, although there were no statistical differences between the three groups. Conclusion : We concluded that subinguinal microscopic varicocelectomy could be a minimally invasive procedure compared to the other two techniques and a worthy method for treating male infertility due to clinical varicocele.

ABSTRACT. Tubulobulbar complexes (TBCs) are composed of several tubular invaginations formed at the plasma membrane of testicular Sertoli cells. TBCs are transiently formed at the contact region with spermatids at spermatogenic stage VII in rat and mouse, and such TBC formation is prerequisite for spermatid release. Since the characteristic structure of TBCs suggests that the molecules implicated in endocytosis could be involved in TBC formation, we here investigated the localization and physiological roles of endocytic proteins, amphiphysin 1 and dynamin 2, at TBCs. We demonstrated by immunofluorescence that the endocytic proteins were concentrated at TBCs, where they colocalized with cytoskeletal proteins, such as actin and vinculin. Immunoelectron microscopy disclosed that both amphiphysin 1 and dynamin 2 were localized on TBC membrane. Next, we histologically examined the testis from amphiphysin 1 deficient {Amph -/-} mice. Morphometric analysis revealed that the number of TBCs was significantly reduced in Amph -/-. The ratio of stage VIII seminiferous tubules was increased, and the ratio of stage IX was conversely decreased in Amph -/-. Moreover, unreleased spermatids in stage VIII seminiferous tubules were increased in Amph -/-, indicating that spermatid release and the following transition from stage VIII to IX was prolonged in Amph -/-mice. These results suggest that amphiphysin 1 and dynamin 2 are involved in TBC formation and spermatid release at Sertoli cells.

Maspin is a member of the serine protease inhibitors and the maspin gene, a tumor suppressor gene, is down-regulated in a large fraction of prostate cancers. We evaluated the use of adeno-associated virus (AAV, serotype 2) vector encoding maspin as a means for in vivo gene therapy for human prostate cancer. TUNEL assay of subcutaneously formed LNCaP or DU145 tumors in nude mice showed that intratumoral AAV-mediated maspin expression significantly upregulated the number of apoptotic cells compared with AAV-LacZ treatment. Immunofluorescence double staining for maspin protein and apoptosis in LNCaP tumors showed that the percentage of apoptotic cells in AAV-maspin-mediated maspin-expressing cells was significantly high compared with that in AAV-GFP-mediated GFP-expressing cells. Moreover, significantly fewer CD31-positive microvessels were observed in AAV-maspin-treated tumors compared with the control tumors. These therapeutic responses were highly correlated to persistent maspin expression in tumors, confirmed by Western blot analysis until at least day 56 after treatment. Finally, intratumoral delivery of AAV-maspin significantly suppressed growth of LNCaP and DU145 tumors and improved survival of mice. We conclude that AAV-mediated prolonged maspin expression efficiently suppresses human prostate tumor growth in vivo by apoptosis induction and inhibition of angiogenesis.

Testicular germ cell-induced autoimmune orchitis is characterized by inflammatory cell infiltration followed by disturbance of spermatogenesis. Experimental autoimmune orchitis (EAO) is an animal model for human immunological male infertility; delayed-type hypersensitivity (DTH) response plays a key role in its induction. Interleukin-10 (IL-10) is a regulatory cytokine that is critical in preventing organ-specific autoimmune inflammation. To determine the effects on EAO of human IL-10 (hIL-10) gene transfer, C3H/He mice immunized by unilateral testicular injury were administered intramuscular (i.m.) injections of adeno-associated viral (AAV) vector-encoding hIL-10 on the day of immunization. Serum hIL-10 was detected beginning at 1 week postinjection, and peaked at 3 weeks. Histological examinations showed a significantly low incidence of orchitis and disturbance of spermatogenesis in AAV hIL-10-treated mice, and the DTH response to autologous testicular cells was significantly suppressed. Immunohistochemical analysis of IFN-g and IL-2, T-cell-associated cytokines, in the spleen and testes revealed significantly fewer cytokine-expressing cells after treatment. We conclude that a single i.m. administration of AAV hIL-10 significantly suppresses EAO and hypospermatogenesis by regulating cell-mediated immunity in the testes.

Amphiphysin 1 is involved in clathrin-mediated endocytosis. In this study, we demonstrate that amphiphysin 1 is essential for cellular phagocytosis and that it is critical for actin polymerization. Phagocytosis in Sertoli cells was induced by stimulating phosphatidylserine receptors. This stimulation led to the formation of actin-rich structures, including ruffles, phagocytic cups, and phagosomes, all of which showed an accumulation of amphiphysin 1. Knocking out amphiphysin 1 by RNA interference in the cells resulted in the reduction of ruffle formation, actin polymerization, and phagocytosis. Phagocytosis was also drastically decreased in amph 1 (؊/؊) Sertoli cells. In addition, phosphatidylinositol-4,5-bisphosphate-induced actin polymerization was decreased in the knockout testis cytosol. The addition of recombinant amphiphysin 1 to the cytosol restored the polymerization process. Ruffle formation in small interfering RNA-treated cells was recovered by the expression of constitutively active Rac1, suggesting that amphiphysin 1 functions upstream of the protein. These findings support that amphiphysin 1 is important in the regulation of actin dynamics and that it is required for phagocytosis. INTRODUCTIONPhagocytosis is a form of endocytosis in which large molecules are engulfed by the plasma membrane and internalized for digestion (Greenberg and Grinstein, 2002;Niedergang and Chavrier, 2004). In vertebrates, macrophages and testicular Sertoli cells are the predominant phagocytes. Phagocytosis contributes to removal of foreign bodies, bacteria, and microbe-infected and apoptotic cells. Binding of these objects to the receptors, such as Fc␥ receptors or phosphatidylserine receptors, on the surface of the macrophage initiates membrane remodeling and reorganization of the actin cytoskeleton. Cytoskeletal changes cause the cells to extend pseudopods that then engulf the objects (Anderem and Underhill, 1999).Sertoli cells participate in the maturation of germ cells and the release of sperm (Russell et al., 1990). These cells are highly phagocytic and are responsible for eliminating the residual cytoplasm of spermatids (Chemes, 1986;Clermont et al., 1987;Morales and Clermont, 1991). The phagocytosis is also used to remove apoptotic germ cells (Shiratsuchi et al., 1997(Shiratsuchi et al., , 1999Kawasaki et al., 2002). Phagocytosis in Sertoli cells is initiated by the recognition of phosphatidylserine (PS) found on the germ cell surface by PS receptors (Shiratsuchi et al., 1997(Shiratsuchi et al., , 1999Blanco-Rodriguez and Martinez-Garcia, 1999;Kawasaki et al., 2002). The class B scavenger family receptors SR-BI and CD36 are found on the surface of Sertoli cells where they function in PS recognition (Shiratsuchi et al., 1997(Shiratsuchi et al., , 1999Kawasaki et al., 2002;Gillot et al., 2005). In the testis, Sertoli cells express high levels of amphiphysin 1 (Watanabe et al., 2001;Kamitani et al., 2002). The expression of amphiphysin, along with that of dynamin 2, increases with sexual development (Watanabe et al., 2...