Characterization of a non-human primate model for the study of testicular peritubular cells-comparison with human testicular peritubular cells.

STUDY QUESTION: Are monkey testicular peritubular cells (MKTPCs) from the common marmoset monkey (Callithrix jacchus) a suitable translational model for the study of human testicular peritubular cells (HTPCs)? SUMMARY ANSWER: MKTPCs can be isolated and propagated in vitro, retain characteristic markers for testicular peritubular cells and their proteome strongly (correlation coefficient of 0.78) overlaps with the proteome of HTPCs. WHAT IS KNOWN ALREADY: Smooth-muscle-like peritubular cells form the wall of seminiferous tubules, transport sperm, are immunologically active, secrete a plethora of factors and may contribute to the spermatogonial stem cell niche. Mechanistic studies are hampered by heterogeneity of human samples. STUDY DESIGN, SIZE, DURATION: We established a culture method for MKTPCs and characterized these cells from six young adult animals (2-3 years). To examine whether they qualify as a translational model we also examined HTPCs from seven men and compared the proteomes of both groups. PARTICIPANTS/MATERIALS, SETTING, METHODS: We used explant cultures to obtain MKTPCs, which express smooth muscle markers (calponin (CNN1), smooth muscle actin (ACTA2)), lack FSH-receptors (FSHR) and LH-receptors (LHCGR), but possess androgen receptors (AR). MKTPCs can be passaged at least up to eight times, without discernable phenotypic changes. Mass-spectrometry-based analyses of the MKTPC and HTPC proteomes were performed. MAIN RESULTS AND THE ROLE OF CHANCE: We established a method for isolation and cultivation of MKTPCs, and provide a comprehensive analysis of their protein repertoire. The results let us conclude that MKTPCs are suitable as a non-human primate model to study peritubular cell functions. LARGE SCALE DATA: List of identified proteins in MKTPCs by liquid chromatography-tandem mass spectrometry is accessible at the ProteomeXchange (identifier PXD009394). LIMITATIONS, REASON FOR CAUTION: This is an in vitro cellular non-human primate model used to provide a window into the role of these cells in the human testis. WIDER IMPLICATIONS OF THE FINDINGS: Previous studies with HTPCs from patients revealed a degree of heterogeneity, possibly due to age, lifestyle and medical history of the individual human donors. We anticipate that the new translational model, derived from young healthy non-human primates, may allow us to circumvent these issues and may lead to a better understanding of the role of peritubular cells. STUDY FUNDING AND COMPETION OF INTEREST(S): This work was supported by grants from the Deutsche Forschungsgemeinschaft (MA 1080/27-1; AR 362/9-1; BE 2296/8-1). The authors declare no competing financial interests.

Microdissection TESE is superior to conventional TESE in patients with nonobstructive azoospermia caused by Y chromosome microdeletions.

Nonobstructive azoospermia is caused in up to 10% by microdeletions of the Y chromosome in the azoospermia factor (AZF) region, which is divided into three nonoverlapping areas (AZFa, AZFb and AZFc). In 25 male patients with AZF microdeletions, the results of two different techniques for surgical sperm retrieval (SR), conventional multilocular TESE and microdissection TESE, were studied retrospectively over a period of 19 years. Conventional multilocular TESE was carried out in 11 patients and microdissection TESE in 14 patients. Successful SR was possible only in patients with isolated AZFc microdeletions, so only the 20 patients with AZFc microdeletions alone were taken into account for the comparison of the both operative techniques. The sperm detection rate for conventional multilocular TESE was 25%, the sperm detection for microdissection TESE was significantly higher with 67%. In all patients, a histological examination of the testicular tissue was carried out, which showed a mixed picture, but Sertoli-cell-only syndrome in most cases. FSH was no prognostic marker for successful SR. In two of six couples performing an intracytoplasmic sperm injection until now, a pregnancy occurred.

Refertilization surgery. A surgeon's experience of over 27 years with nearly 2000 patients.

AIM: The aim of this paper was to present the experiences of microsurgical refertilization in a single-centre study during a period of 27 years. METHODS: Nearly 2000 patients were operated by a single surgeon (JUS). A total of 1708 patients were evaluated in a data base, 1164 were available for a follow-up. Both vasovasostomy (VV) and epididymovasostomy (EV) were carried out in a three-layer technique. Vasectomy reversal (VR) end-to-end VV was performed only if spermatozoa had been demonstrated at the epididymal stump of the vas. In all other cases of VR, EV was done in a preocclusive region of the epididymal tubule. In the cases of postinfectious obstruction (PIO) of seminal pathways, an EV was always carried out. RESULTS: The outpatient procedure of refertilization was associated with a very low complication rate, which underlines its minimal-invasive character. The follow-up rate was 68%, the overall patency rate was 88% for VR and 67% for PIO and the pregnancy rate was 59% for VR and 38% for PIO. Secondary azoospermia was observed in 1% of the patients. CONCLUSION: In relation to the intervals of obstruction, the patency and pregnancy rates were higher after short-term obstruction than after long-term obstruction. There is a significant discrepancy between patency and pregnancy rates that is likely to be caused by a relevant number of patients with postoperative asthenozoospermia.

The role of shortening procedures for the surgical therapy of Peyronie's disease.

In this article we briefly review the current surgical treatment options for Peyronie's disease (PD) in its stable phase. We emphasize the important role of tunical shortening procedures which account for the major share of operations for PD. Shortening procedures provide excellent curvature correction combined with a very low risk of new erectile dysfunction. Since erectile function is already heavily impaired by the disease and its comorbidities in many patients with PD, tunical shortening procedures often are the treatment of choice for the correction of penile curvature. While there is no hard evidence for the superiority of a specific shortening procedure, several authors prefer the classical Nesbit technique over simple plication techniques. We also present our experiences with the Tunica albuginea underlap technique (TAU-technique), a new modification of the Nesbit procedure, that might add further surgical advantages while preserving the strength of the classical Nesbit technique.

Vasectomy reversal using a microsurgical three-layer technique: one surgeon's experience over 18 years with 1300 patients.

The technique and the results of microsurgical vasectomy reversal in a single-centre study over 18 years are presented. Both vasovasostomy (VV) and epididymovasostomy (EV) were carried out in a three-layer technique. With strict adherence to the strategy, end-to-end VV was only performed if spermatozoa had been demonstrated at the epididymal stump of the vas. In all other cases, EV was carried out in a preocclusive region of the epididymal tubule. The outpatient procedure of refertilization was associated with a very low complication rate, which underlines its minimal-invasive character. The follow-up rate was 71%, the overall patency rate was 89% and the pregnancy rate was 59%. Secondary azoospermia was only observed in 1% of the patients. In relation to the intervals of obstruction, the patency and pregnancy rates were higher after short-term obstruction than after long-term obstruction. Correspondingly, higher success rates were found after VV than after EV. This is understandable because the probability for indication of EV increases with longer periods of obstruction. There is a significant discrepancy between patency and pregnancy rates that is likely to be caused by a relevant number of patients with post-operative asthenozoospermia. The duration of obstruction is an important factor concerning epididymal damage, but it only disproportionately affects the results of refertilization if the technology of EV is implemented consistently in case of an epididymal granuloma. Good clinical results are achieved with this strategy, as evidenced by pregnancy rates and semen analyses.

Evidence for an adaptation in ROS scavenging systems in human testicular peritubular cells from infertility patients.

Fibrosis, increased amounts of immune cells and expression of COX-2 in the testes of infertility patients provide circumstantial evidence for a specific testicular milieu, in which reactive oxygen species (ROS) could be increased. If ROS level increase and/or ROS scavengers decrease, the resulting testicular oxidative stress may contribute to human male infertility. Primary peritubular cells of the human testis, from men with normal spermatogenesis (HTPCs) and infertile patients (HTPC-Fs), previously allowed us to identify an end product of COX-2 action, a prostaglandin derivative (15dPGJ2), which acts via ROS to alter the phenotype of peritubular cells, at least in vitro. Using testicular biopsies we now found 15dPGJ2 in patients and hence we started exploring the ROS scavenger systems of the human testis. This system includes catalase, DJ-1, peroxiredoxin 1, SOD 1 and 2, glutathione-S-transferase and HMOX-1, which were identified by RT-PCR/sequencing in HTPCs and HTPC-Fs and whole testes. Catalase, DJ-1, peroxiredoxin 1 and SOD 2 were also detected by Western blots and in part by immunohistochemistry in testicular samples. Western blots of cultured cells further revealed that catalase levels, but not peroxiredoxin 1, SOD 2 or DJ-1 levels, are significantly higher in HTPC-Fs than in HTPCs. This particular difference is correlated with the improved ability of HTPC-Fs to handle ROS, which became evident when cells were exposed to 100 µm H(2)O(2). H(2)O(2) induced stronger responses in HTPCs than in HTPC-Fs, which correlates with the lower level of the H(2)O(2)-degrading defence enzyme catalase in HTPCs. The results provide evidence for an adaptation to elevated ROS levels, which must have occurred in vivo and which persist in vitro in HTPC-Fs. Thus, in infertile men with impaired spermatogenesis elevated ROS levels likely exist, at least in the tubular wall.

Significance of CFTR gene mutations in patients with congenital aplasia of vas deferens with special regard to renal aplasia.

Between 1994 and 2010, a total of 123 patients with obstructive azoospermia due to aplasia of vas deferens (CAVD) were surgically treated. In 110 patients, the condition was bilateral (CBAVD), 13 men had unilateral aplasia (CUAVD), and 10 patients additionally had aplasia of one kidney. All patients underwent CFTR genetic testing, which detected two mutations (homozygous or compound heterozygous condition) in 38%, one mutation in 34% and no mutation in 28% of the patients with CBAVD. Neither the azoospermic patients with congenital unilateral aplasia of vas deferens nor those with CBAVD and renal aplasia were found to have CFTR mutations. The results militate against the assumption that there is an association between the CFTR gene and unilateral aplasia of vas deferens or bilateral aplasia of vas deferens with renal involvement.

Mast cell tryptase stimulates production of decorin by human testicular peritubular cells: possible role of decorin in male infertility by interfering with growth factor signaling.

BACKGROUND: Myofibroblastic, peritubular cells in the walls of seminiferous tubules produce low levels of the extracellular matrix (ECM) protein decorin (DCN), which has the ability to interfere with growth factor (GF) signaling. In men with impaired spermatogenesis, fibrotic remodeling of these walls and accumulation of tryptase-positive mast cells (MCs) occur. METHODS: Human testicular biopsies with normal and focally impaired spermatogenesis (mixed atrophy) were subjected to immunohistochemistry and laser micro-dissection followed by RT-PCR. Primary human testicular peritubular cells (HTPCs), which originate from normal and fibrotically altered testes (HTPC-Fs), were studied by qRT-PCR, western blotting, enzyme-linked immunosorbent assay measurements and Ca(2+) imaging. Phosphorylation and viability/proliferation assays were performed. RESULTS: Immunohistochemistry revealed DCN deposits in the walls of tubules with impaired spermatogenesis. Mirroring the situation in vivo, HTPC-Fs secreted more DCN than HTPCs (P < 0.05). In contrast to HTPCs, HTPC-Fs also responded to the main MC product, tryptase, and to a tryptase receptor (PAR-2) agonist by further increased production of DCN (P < 0.05). Several GF receptors (GFRs) are expressed by HTPCs and HTPC-Fs. DCN acutely increased intracellular Ca(2+)-levels and phosphorylated epidermal GF (EGFR) within minutes. Platelet-derived GF (PDGF) and EGF induced strong mitogenic responses in HTPC/-Fs, actions that were blocked by DCN, suggesting that DCN in the ECM interferes with GF/GFRs signaling of peritubular cells of the human testis. CONCLUSIONS: The data indicate that the increase in testicular DCN found in male infertility is a consequence of actions of MC-derived tryptase. We propose that the increases in DCN may consequently imbalance the paracrine signaling pathways in human testis.

[Current aspects of surgical restoration of fertility]. / Aktuelle Aspekte der Refertilisierungsoperationen.

Obstruction of the seminal ducts is the cause of infertility in about 5% of patients. It can be congenital or arise as the result of secondary changes. The reconstruction of the duct undertaken depends on the site of the obstruction. The introduction of microsurgical techniques has revolutionized the treatment of male infertility.A tubulovasostomy is carried out if the obstruction lies in the region of the epididymis. Such an anastomosis requires, owing to the minute anatomical relationships involved, a microsurgical procedure. For an obstruction of the vas deferens a vasovasostomy is required. Many investigations have shown that microsurgical techniques are also necessary for this procedure if a satisfactory success rate is to be achieved. The double-layer technique is the standard method for vasovasostomy. Transurethral resection of the ejaculatory ducts (TURED) is required for the very rare obstruction in this region, and men with an obstruction here cannot be regarded as forming a homogeneous group.Before advising an infertile couple it is necessary to investigate the individual conditions and possibilities. Because of the high success rate obtainable today by surgical reconstruction of the seminal ducts, this must constitute the first type of treatment to be considered, before any of the procedures of reproductive medicine are undertaken.

Insights into the role of androgen receptor in human testicular peritubular cells.

Contractile smooth muscle-like peritubular cells build the wall of seminiferous tubules in men. They are crucial for sperm transport and complement the functions of Sertoli cells by secreting factors, including glial cell line-derived neurotrophic factor. Previous studies revealed that they also secrete the chemokine C-X-C motif chemokine ligand 12 (CXCL12), which has known roles in spermatogenesis. Peritubular cells express the androgen receptor (AR), which is retained in isolated human testicular peritubular cells. We aimed to explore AR-regulated functions in human testicular peritubular cells. Bearing in mind that infertile men often have high aromatase activity, which may lower intratesticular androgen concentrations, an animal model for male infertility was studied. These mice display an age-dependent loss in spermatogenesis due to high aromatase activity. Human testicular peritubular cells were exposed to dihydrotestosterone or the antiandrogen flutamide. We studied AR, smooth muscle cell markers, glial cell line-derived neurotrophic factor and 15 secreted factors previously identified, including CXCL12. We used qPCR, Western blotting, ELISA or selected reaction monitoring (SRM). In the animal model for male infertility, we employed qPCR and immunohistochemistry. Dihydrotestosterone increased AR and flutamide prevented these actions. The smooth muscle cell markers calponin and smooth muscle actin were likewise increased, while cell size or cellular proliferation was not changed. Dihydrotestosterone did not increase glial cell line-derived neurotrophic factor or CXCL12 secretion but increased levels of serine proteinase inhibitor (SERPIN) E1. The animal model for male infertility with high aromatase activity showed reduced numbers of AR-immunoreactive testicular peritubular cells, suggesting that altered androgen and/or oestrogen levels could influence AR-mediated responses in peritubular cells. Androgens act on human testicular peritubular cells to enhance AR levels, their contractile phenotype and to modulate the secretion of some secreted factors. This study suggests that some aspects of human peritubular cell functions are regulated by androgens.

Sterile inflammation as a factor in human male infertility: Involvement of Toll like receptor 2, biglycan and peritubular cells.

Changes in the wall of seminiferous tubules in men with impaired spermatogenesis imply sterile inflammation of the testis. We tested the hypothesis that the cells forming the wall of seminiferous tubules, human testicular peritubular cells (HTPCs), orchestrate inflammatory events and that Toll like receptors (TLRs) and danger signals from the extracellular matrix (ECM) of this wall are involved. In cultured HTPCs we detected TLRs, including TLR2. A TLR-2 ligand (PAM) augmented interleukin 6 (IL-6), monocyte chemo-attractant protein-1 (MCP-1) and pentraxin 3 (PTX3) in HTPCs. The ECM-derived proteoglycan biglycan (BGN) is secreted by HTPCs and may be a TLR2-ligand at HTPCs. In support, recombinant human BGN increased PTX3, MCP-1 and IL-6 in HTPCs. Variable endogenous BGN levels in HTPCs derived from different men and differences in BGN levels in the tubular wall in infertile men were observed. In testes of a systemic mouse model for male infertility, testicular sterile inflammation and elevated estradiol (E2) levels, BGN was also elevated. Hence we studied the role of E2 in HTPCs and observed that E2 elevated the levels of BGN. The anti-estrogen ICI 182,780 blocked this action. We conclude that TLR2 and BGN contribute to sterile inflammation and infertility in man.

Human testicular peritubular cells secrete pigment epithelium-derived factor (PEDF), which may be responsible for the avascularity of the seminiferous tubules.

Male fertility depends on spermatogenesis, which takes place in the seminiferous tubules of the testis. This compartment is devoid of blood vessels, which are however found in the wall of the seminiferous tubules. Our proteomic study using cultured human testicular peritubular cells (HTPCs) i.e. the cells, which form this wall, revealed that they constitutively secrete pigment epithelium-derived factor, PEDF, which is known to exert anti-angiogenic actions. Immunohistochemistry supports its presence in vivo, in the human tubular wall. Co-culture studies and analysis of cell migration patterns showed that human endothelial cells (HUVECs) are repulsed by HTPCs. The factor involved is likely PEDF, as a PEDF-antiserum blocked the repulsing action. Thus testicular peritubular cells, via PEDF, may prevent vascularization of human seminiferous tubules. Dihydrotestosterone (DHT) increased PEDF (qPCR) in HTPCs, however PEDF expression in the testis of a non-human primate occurs before puberty. Thus PEDF could be involved in the establishment of the avascular nature of seminiferous tubules and after puberty androgens may further reinforce this feature. Testicular microvessels and blood flow are known to contribute to the spermatogonial stem cell niche. Hence HTPCs via control of testicular microvessels may contribute to the regulation of spermatogonial stem cells, as well.

[A comparison of color duplex sonography with selective penile DSA in assessing erectile dysfunction]. / Vergleich der Farbduplexsonographie mit der selektiven penilen DSA bei der Abklärung der erektilen Dysfunktion.

42 patients with suspected vasculogenic impotence were examined via colour duplex sonography. Penile vascular anatomy was mapped and peak velocity was determined in both cavernous arteries before and after intracorporeal injection of 15 mg papaverine and 0.5 mg regitine. In all patients the results could be compared with selective penile pharmacoangiography as the gold standard. In 34/42 patients colour Duplex sonography and angiography led to an identical evaluation of penile blood supply. 4 patients were classified false-negative and 4 patients false-positive. Sensitivity and specificity were calculated for each side separately. Sensitivity was 82.4% for left side angiogram and 83.3% for right side angiogram. Specificity was 88% for left side angiogram and 87.5% for right side angiogram. Peak flow velocity was significantly diminished in pathological angiograms.

[Purulent cavernitis as a complication of self injection into the cavernous body]. / Eitrige Kavernitis als SKAT-Komplikation.

Intracavernous self-injection of vasoactive drugs is commonly prescribed for the treatment of erectile failure. Cavernitis is a serious complication of this treatment. The case of a 63-years-old patient with cavernitis following intracavernous injection of papaverine and subsequent priapism is reported. He had a phlegmonous infection of both corpora cavernosa without infection of the corpus spongiosum. The treatment consisted in surgical debridement of the corpora cavernosa with intracavernous drains inserted for continued irrigation and suction. Complete remission of the infection was ultimately obtained, but fibrosis of the remnants of both corpora cavernosa remained.

[Microsurgical techniques in urology]. / Mikrochirurgische Techniken in der Urologie.

Microsurgery is an established operative technique for refertilization by vasovasostomy, tubulovasostomy and microsurgical epididymal sperm aspiration. Other andrological indications for the use of a microscope are penile revascularization, resection of varicocele and surgical treatment of Peyronie's disease. Autotransplantation of intra-abdominal testicles and hypospadia correction are indications for microsurgical techniques in pediatric urological surgery. Renal transplant, sex-change surgery and replants of penis and scrotum are further indications for use of microsurgery. Microsurgery will move into new fields of urological surgery, especially neurourology.

[Microsurgical therapy of occlusive azoospermia]. / Mikrochirurgische Therapie der Verschlussazoospermie.

From 5/88 to 8/93 148 patients were operated on for obstructive azoospermia using a microsurgical technique. In 68 patients we performed a vasectomy reversal by vasovasostomy (VT), 42 patients underwent a tubulovasostomy (TT), and 8 patients had a VT with contralateral TT. Two patients had implantation of an alloplastic spermatocele, 9 patients had microsurgical epididymal sperm aspiration (MESA), and 19 patients microsurgical exploration. Patency was achieved in 89% of patients after VT and in 32% after TT. A pregnancy occurred in 34% after VT and in 13% after TT. No pregnancy was achieved after MESA or alloplastic spermatocele. A microsurgical procedure is necessary for the treatment of congenital or acquired obstructive azoospermia.

Expression of the oestrogen receptor GPER by testicular peritubular cells is linked to sexual maturation and male fertility.

Besides the two nuclear oestrogen receptors (ESR1/ESR2), the G protein-coupled oestrogen receptor (GPER) was described in the human testis but little is known about testicular GPER during development or male infertility. We performed an immunohistochemical analysis using human and rhesus monkey testicular samples. The results obtained in adult primate testes showed GPER in interstitial and vascular cells as well as in smooth muscle-like peritubular cells, which build the wall of seminiferous tubules. Expression of GPER was also found in cultured human testicular peritubular cells (HPTCs) by Western blotting and RT-PCR/sequencing. Furthermore, as seen in time-lapse videos of cultured cells, addition of a specific GPER agonist (G1) significantly reduced the numbers of HTPCs within 24 h. A GPER antagonist (G15) prevented this action, implying a role for GPER related to the control of cell proliferation or cell death of peritubular cells. Peritubular cell functions and their phenotype change, for example, during post-natal development and in the cases of male infertility. The study of non-human primate samples revealed that GPER in peritubular cells was detectable only from the time of puberty onwards, while in samples from infantile and prepubertal monkeys only interstitial cells showed immunopositive staining. In testicular biopsies of men with mixed atrophy, a reduction or loss of immunoreactive GPER was found in peritubular cells surrounding those tubules, in which spermatogenesis was impaired. In other cases of impaired spermatogenesis, namely when the tubular wall was fibrotically remodelled, a complete loss of GPER was seen. Thus, the observed inverse relation between the state of fertility and GPER expression by peritubular cells implies that the regulation of primate testicular peritubular cells by oestrogens is mediated by GPER in both, health and disease.

Angiotensin II regulates testicular peritubular cell function via AT1 receptor: a specific situation in male infertility.

We observed that peritubular myoid cells in the human testis are immunoreactive for angiotensin II (AngII) receptors (AT1R) and explored AngII actions in cultured human testicular peritubular cells (HTPCs). In response to AngII they contracted within minutes. The AT1R-blocker losartan blocked contraction, implying involvement of AngII and AT1R in intratesticular sperm transport. AngII also significantly increased IL-6 mRNA levels and IL-6 secretion within hours and losartan again prevented this action. This suggests involvement in inflammatory processes, which may play a role in male infertility. AngII can be generated locally by mast cell (MC)-derived chymase (CHY), which cleaves AngI. In testicular biopsies from infertile men we found abundant MCs, which express CHY, within the wall of seminiferous tubules. In contrast, CHY-positive MCs are hardly found in normal human testis. Testicular inflammatory events may fuel processes resulting in impaired spermatogenesis. Therefore therapeutic interference with MCs, CHY or AT1R might be novel options in male infertility.

Are testicular mast cells involved in the regulation of germ cells in man?

Protease activated receptor-2 (PAR-2) is the receptor for the prototype mast cell product tryptase. PAR-2 expression by cells of the human germinal epithelium was reported, but the exact cellular sites of testicular expression remained unknown. That became of interest, because mast cells, expressing tryptase, were found in the walls of seminiferous tubules of patients suffering from sub- and infertility. This location suggested that mast cells via tryptase might be able to influence PAR-2-expressing cells in the germinal epithelium. To explore these points, we used testicular paraffin-embedded sections for immunohistochemistry. PAR-2-positive cells were mostly basally located cells of the seminiferous epithelium, namely spermatogonia. Some stained for the receptor for GDNF (GFRalpha-1), and possibly represent spermatogonial stem cells (SSCs). As true human SSCs could not be examined, we turned to TCam-2 seminoma cells, expressing PAR-2 and stem cell markers, including GFRalpha-1. TCam-2 cells robustly responded to stimulation with a specific PAR-2 agonist (SLIGKV) by increased intracellular Ca(2+) levels. Recombinant tryptase and trypsin, but not a control peptide (VKGILS) evoked this response, implying functional PAR-2. Video imaging and caspase 3/7 assays showed that SLIGKV and tryptase prevented spontaneous apoptosis and increased proliferation of TCam-2 cells. The expression of the marker of pluripotency OCT3/4 was unchanged upon activation of PAR-2, suggesting that the stem cell-like character is not changed. Furthermore, human germ cell cancers were examined. A subset of seminoma and carcinoma in situ samples expressed PAR-2, indicating that yet unknown subgroups exist. Collectively, the descriptive data obtained in human testicular sections, in germ cell cancers and the functional results in TCam-2 cells imply a trophic role of mast cell-derived tryptase for human germ cells. This may be relevant for subtypes of human germ cell cancers, and possibly SSCs. It also raises the possibility that PAR-2 agonists might be useful for the in vitro propagation of human SSCs.

Partial loss of contractile marker proteins in human testicular peritubular cells in infertility patients.

Fibrotic remodelling of the testicular tubular wall is common in human male infertility caused by impaired spermatogenesis. We hypothesized that this morphological change bears witness of an underlying fundamentally altered state of the cells building this wall, that is, peritubular smooth muscle-like cells. This could include a loss of the contractile abilities of these cells and thus be a factor in male infertility. Immune cells are increased in the tubular wall in these cases, hence local immune cell-related factors, including a prostaglandin (PG) metabolite may be involved. To explore these points in the human, we used testicular biopsies, in which tubules with normal spermatogenesis and impaired spermatogenesis are next to each other [mixed atrophy (MA)], normal biopsies and cultured human testicular peritubular cells. Proteins essential for contraction, myosin heavy chain (MYH11), calponin (Cal) and relaxation, cGMP-dependent protein kinase 1 (cGKI), were readily detected by immunohistochemistry and were equally distributed in all peritubular cells of biopsies with normal spermatogenesis. In all biopsies, vascular smooth muscle cells also stained and served as important intrinsic controls, which showed that in MA samples when spermatogenesis was impaired, staining was restricted to only few peritubular cells or was absent. When spermatogenesis was normal, regular peritubular staining became obvious. This pattern suggests complex regulatory influences, which in face of the identical systemic hormonal situation in MA patients, are likely caused by the local testicular micromilieu. The PG metabolite 15dPGJ2 may represent such a factor and it reduced Cal protein levels in peritubular cells from patients with/without impaired spermatogenesis. The documented phenotypic switch of peritubular, smooth muscle-like cells in MA patients may impair the abilities of the afflicted seminiferous tubules to contract and relax and must now be considered as a part of the complex events in male infertility.