review REVIEW Spermatogenesis 1:3, 195-208; July/August/September 2011; © 2011 Landes Bioscience The emerging role of matrix metalloproteases of the ADAM family in male germ cell apoptosis Ricardo D. Moreno,* Paulina Urriola-Muñoz and Raúl Lagos-Cabré Departamento de Fisiología; Pontificia Universidad Católica de Chile; Santiago, Chile Key words: spermatogenesis, apoptosis, metalloprotease, infertility Constitutive germ cell apoptosis during mammalian sperm- Despite that some data suggest that germ cell apoptosis is autono- atogenesis is a key process for controlling sperm output and to mous (independent of the environment), the role of the Sertoli eliminate damaged or unwanted cells. An increase or decrease cell should not be disregarded, because it may provide important in the apoptosis rate has deleterious consequences and leads survival and/or dead clues to germ cells (see below). Germ cell to low sperm production. Apoptosis in spermatogenesis has been widely studied, but the mechanism by which it is induced apoptosis has been shown to play an important role in control- under physiological or pathological conditions has not been ling sperm output in many species, and in humans it has been 5-10 clarified. We have recently identified the metalloprotease related to infertility. Additionally, it has been proposed that ADAM17 (TACE) as a putative physiological inducer of germ the purported decline in global sperm production is caused by cell apoptosis. The mechanisms involved in regulating the environmental xenoestrogens that induce germ cell apoptosis.11 shedding of the ADAM17 extracellular domain are still far In the same way, a modern lifestyle and the use of underwear has from being understood, although they are important in order been proposed to induce elevated scrotal temperature that may to understand cell-cell communications. Here, we review cause germ cell apoptosis, akin to transient heat stress (43°C for the available data regarding apoptosis during mammalian 15 min) in the testis.12,13 Germ cells undergoing meiosis (sper- spermatogenesis and the localization of ADAM proteins in the matocytes) and diploid spermatogonia are sensitive to heat shock, male reproductive tract. We propose an integrative working ionising radiation, growth factor deprivation and chemotherapeu- model where ADAM17, p38 MAPK, protein kinase C (PKC) 14-18 and the tyrosine kinase c-Abl participate in the physiological tic agents. Many studies have shown the relevance of apopto- signalling cascade inducing apoptosis in germ cells. In our sis in regulating spermatozoa output and in eliminating damaged 5,19-24 model, we also propose a role for the Sertoli cell in regulating germ cells, particularly spermatocytes. Moreover, in seasonal the Fas/FasL system in order to induce the extrinsic pathway of breeding mammals, apoptosis plays an important role in the mas- apoptosis in germ cells. This working model could be applied to sive germ cell depletion observed during the non-breeding sea- further understand constitutive apoptosis in spermatogenesis son.25-27 Therefore, apoptosis is important in physiological and and in pathological conditions (e.g., varicocele) or following pathological conditions, and knowledge of the molecular mecha- environmental toxicants exposure (e.g., genotoxicity or nisms underlying this process may help in developing new phar- xenoestrogens). macological tools in order to modulate fertility (Fig. 1). In this review, we will analyze the available evidence regarding the role of different apoptotic pathways in germ cell apoptosis. In addi- tion, we will review the distribution and localization of matrix Introduction metalloproteases of the ADAM family in the male reproductive tract and discuss the recently discovered role of ADAM17 in the In order to ensure fertility, an important step is the proper differ- initiation of the apoptotic programme in germ cells. entiation and function of spermatozoa. Sperm production relies on physiological and environmental factors, which may attenu- Molecular Mechanisms ate or even totally suppress testicular function (Fig. 1). Massive of Germ Apoptosis Execution germ cell death occurs under physiological conditions (constitu- tive apoptosis) during the first round of spermatogenesis, which From primitive multicellular organisms to higher vertebrates, seems to be vital in order to establish a proper interaction between well-orchestrated cell death events are critical for the removal of germ and Sertoli cells and to eliminate unwanted germ cells.1-3 superfluous cells as a vital part of tissue sculpting during devel- Different experimental approaches have pointed out that germ opment. Physiological cell death enables the elimination of cells undergoing meiosis (spermatocytes) are the main target for unwanted extra cells to maintain cellular homeostasis in devel- apoptosis, in addition to a smaller fraction of spermatogonia.1,4 oped adults, as well as the elimination of harmful cells, or cells with serious cellular or genomic damage.28 Apoptosis is pro- *Correspondence to: Ricardo D. Moreno; Email: [email protected] grammed cell death characterized by several hallmarks such as Submitted: 07/29/11; Revised: 08/27/11; Accepted: 08/29/11 internucleosomal DNA fragmentation, caspase activation and http://dx.doi.org /10.4161/spmg.1.3.17894 externalization of phosphatidyl serine.29,30 In this context, it is www.landesbioscience.com Spermatogenesis 195 elongating spermatids and Leydig cells.37 In the human, the DR4 receptor is faintly detected in some myoid cells and intense immunoexpression has been detected in Sertoli cells, whereas strong expression is detected in spermatocytes, round and elon- gated spermatids. The DR5 receptor is highly expressed by sper- matocytes, round and elongated spermatids, but not detected in Leydig or Sertoli cells. In the mammalian testis, death receptor ligands such as Trail (DR4 and DR5 ligand) have been shown to be expressed by all germ cells, as well as Sertoli and Leydig cells. On the other hand, Fas ligand (FasL) is expressed by Sertoli cells,38 although other reports have detected it in germ cells.39 Thus, ligand-induced germ cell apoptosis could be triggered in a juxta/paracrine or autocrine fashion upon ligand binding to its cognate receptor. Activation (trimerization) of death receptors in response to ligand binding induces the formation of a multimeric complex Figure 1. Factors affecting germ cell apoptosis in spermatogenesis. named the death-induced complex (DISC), which allows for the activation of procaspase-8 in mice along with caspase-10 in humans.35,40,41 Caspase-8 and/or 10 initiate apoptosis by induc- important to differentiate between apoptosis induction and apop- ing proteolytic processing of caspases-3, -6 and -7.42,43 Among tosis execution. Apoptosis initiation or induction can be defined these, caspase-3 is the main executioner caspase involved in frag- as the molecular signaling events which trigger the extrinsic or mentation of the Golgi apparatus, nuclear lamina and DNA (due intrinsic apoptotic pathways. In this way, events such as protein in part to proteolysis of a DNAse inhibitor) and the decrease in kinase activation, calcium signaling or oxidative stress could be mitochondrial membrane potential.41,44,45 regard at the initiation step of apoptosis. In this way, the activa- Evidence supporting the extrinsic pathway in physiological tion of signaling cascades and their cross-talk produce activation germ cell apoptosis during the first round of spermatogenesis of apoptotic and anti-apoptotic genes (as a defence mechanism). comes from studies showing an increase in levels of the Fas recep- However, a cell with competence to undergo apoptosis is able tor and activation of caspases -8, -3, -2 and -6 in apoptotic germ to reverse this process thanks to the induction of anti-apoptotic cells.46,47 In addition, pharmacological inhibition of caspase-8 genes until the cell passes the “point of no return” and the exe- reduces by more than 50% the number of apoptotic germ cells, cution program is activated.31 Apoptosis execution involves the suggesting an important role of the extrinsic pathway in this pro- activation of caspases, a family of serine proteases with around cess. Nevertheless, these data do not rule out the involvement 14 members, which are synthesized as inactive zymogens and of other death receptors which are found in testes.36,37 However, become active upon specific death stimuli.32 The common strat- male mice harbouring a spontaneous loss of function mutation egy adopted to achieve apical caspase activation is the formation in the Fas gene (lpr mice) or FasL (gld mice) are fertile with of complexes containing several caspase zymogens.33,34 In this normal spermatogenesis, suggesting that the Fas/FasL path- way, apical caspase activation can be induced by the intrinsic way may be dispensable in germ cell apoptosis.48 Despite this, (mitochondrial) pathway or by the extrinsic (cell death receptor) FasL mutant gld (generalized lymphoproliferative disease) mice pathway. Several cellular mechanisms can be recognized in order that lack a functional Fas signaling pathway have a small but to prevent and/or regulate caspase activation. Among these, we significant increase in testis weight and numbers of spermatid could mention: (1) cellular caspase inhibitor expression; (2) cell heads per testis, as compared with wild type mice. In addition, death receptor activation; (3) mitochondrial membrane perme- gld mice show a slightly higher spontaneous