Nitric Oxide/Nitric Oxide Synthase, Spermatogenesis, and Tight Junction Dynamics

BIOLOGY OF REPRODUCTION 70, 267–276 (2004) Published online before print 1 October 2003. DOI 10.1095/biolreprod.103.021329 M i n i r ev i ew Nitric Oxide/Nitric Oxide Synthase, Spermatogenesis, and Tight Junction Dynamics1 Nikki P.Y. Lee and C. Yan Cheng2 ABSTRACT membrane must traverse the BTB to gain entry to the adluminal compartment [1] for further development while differentiating into haploid spermatids. Without this timely movement of developing preleptotene and leptotene spermatocytes across the BTB, spermatogenesis is halted, leading to infertility. However, the mechanism(s) that governs BTB dynamics has remained largely obscure until recently. The BTB creates a unique microenvironment for germ cell development; it immunologically segregates most of the germ cell antigens, except those residing on spermatogonia and preleptotene/leptotene spermatocytes, from the systemic circulation; and maintains cell polarity (for reviews, see [2–4]). The significance of the BTB to spermatogenesis has been known for decades, but how the barrier function is regulated has remained obscure until recently. Studies have shown that BTB dynamics in vitro and/ or in vivo are regulated by cytokines, such as transforming growth factor (TGF) b3 and tumor necrosis factor (TNF) a, via two defined signaling pathways, such as the TGFb3/ MEKKs (MAP [mitogen-activated protein]/ERK kinase kinases)/p38 MAP kinase [5, 6] and the TNFa/integrin-linked kinase (ILK)/glycogen synthase kinase (GSK)/p130cas/cJun N-terminal kinase (JNK) MAP kinase [7] signaling pathways. Because the role of cytokines in BTB dynamics has recently been reviewed [8, 9], this is not discussed herein. However, emerging evidence has clearly illustrated the pivotal role of nitric oxide/nitric oxide synthase (NO/NOS) in epithelial barrier function in many organs, including the testis, and in spermatogenesis. Therefore, it is our goal to summarize recent advances in this field and to discuss the possible relevance of these findings regarding male contraceptive development. During spermatogenesis, preleptotene and leptotene spermatocytes, residing in the basal compartment of the seminiferous epithelium, must traverse the blood-testis barrier (BTB) to gain entry to the adluminal compartment for further development at late stage VIII and early stage IX of the epithelial cycle. As such, the timely opening and closing of the BTB is crucial to spermatogenesis. A compromise in this process can lead to infertility. Moreover, the BTB is unique in its relative localization in the seminiferous epithelium compared to the tight junctions (TJs) found in other epithelia. Sertoli cell TJs are situated near the basal lamina in the testis, closest to the basement membrane (a modified form of extracellular matrix [ECM]), unlike TJs found in other epithelia, which are found nearest the apical portion of an epithelium, farthest away from ECM. Needless to say, BTB function in the testis is maintained by intricate regulatory mechanisms. In addition to hormones and cytokines, nitric oxide (NO) was recently shown to be a putative TJ regulator in the testis. Perhaps equally important, TJ dynamics in the testis were shown to be regulated, at least in part, by occludin, a TJ-integral membrane protein, via the NO/soluble guanylate cyclase/ cGMP/protein kinase G signaling pathway. This minireview summarizes recent advances in the field regarding the role of NO in testicular function, with special emphasis regarding its role in TJ dynamics and the likely implications of these studies for male contraceptive development. cyclic adenosine monophosphate, cyclic guanosine monophosphate, nitric oxide, signal transduction, spermatogenesis INTRODUCTION In the mammalian testis, the blood-testis barrier (BTB) is located near the basal lamina, which effectively divides the epithelium into basal and adluminal compartments (Fig. 1). During late stage VIII through early stage IX of the epithelial cycle, preleptotene and leptotene spermatocytes residing in the basal compartment adjacent to the basement CONSTITUENT PROTEINS OF THE BTB Recent studies have shown that tight junctions (TJs) at the site of the BTB are constituted by three classes of TJintegral membrane proteins, namely occludins, claudins, and junctional adhesion molecules (JAMs) (for reviews, see [2, 10, 11]) (Table 1 and Fig. 1). In turn, these transmembrane proteins structurally associate with different adaptors (Table 1), forming a functional TJ complex at the site of the BTB (for reviews, see [2, 8, 12]) (Fig. 1). For instance, the C-terminus of occludin, claudin, or JAM interacts with zonula occludens (ZO)-1 (for reviews, see [12, 13]), which in turn tethers to the underlying actin filaments either directly or via binding to afadin [14] and cingulin [15]. Equally important, these TJ proteins can structurally interact with other regulatory proteins, such as NOS [16], implicating these TJ constituent proteins in the induction of Supported in part by grants from the CONRAD Program (CICCR CIG96-05B, CIG-01-72 to C.Y.C.), National Institutes of Health (NICHD, UOI HD45908 to C.Y.C.; U54 HD29990, Project 3 to C.Y.C.; U54 HD1354120S to C.Y.C.) and the Noopolis Foundation. N.P.Y.L. was supported in part by a Hong Kong University Research Scholarship Award. 2 Correspondence: C. Yan Cheng, Population Council, Center for Biomedical Research, 1230 York Avenue, New York, NY 10021. FAX: 212 327 8733; e-mail: 1 Received: 24 July 2003. First decision: 11 August 2003. Accepted: 23 September 2003. Q 2004 by the Society for the Study of Reproduction, Inc. ISSN: 0006-3363. http://www.biolreprod.org 267 Population Council, 1230 York Avenue, New York, New York 10021 268 LEE AND CHENG downstream signaling events, which in turn can regulate the opening and closing of TJs (for reviews, see [17, 18]). Several recent reviews have discussed the structural and functional features of these TJ-integral membrane proteins and their peripheral binding partners as well as recent advances in the field that investigate the regulation of TJ dy- namics by cytokines using both in vitro and in vivo models (for reviews, see [2, 8, 9]). As such, details of this subject area are not elaborated herein. In brief, the functionality of these TJ-integral membrane proteins is regulated by and large via protein phosphorylation involving putative protein kinases and phosphatases (for a review, see [2]). For in- FIG. 1. A schematic drawing that illustrates the current molecular architecture of the three TJ-integral membrane proteins and their associated peripheral proteins at the site of the BTB as well as the three possible signaling pathways that regulate the opening and closing of the Sertoli cell tight junction. Two other TJ-integral membrane proteins, namely CAR (coxsackie virus and adenovirus receptor) and CRBI (Crumbs homolog 1), found in other epithelia are not shown here since their presence in the testis remains to be identif (...truncated)