Peritubular cells in testes of normal and sub-/infertile men produce GDNF and are likely constitutive contributors of the SSC niche in the human testis.

Our results indicate that the tubular wall of the human testis is a reservoir for cells of the adult Leydig cell lineage and that the steroidogenic potential of these cells can be activated in culture.

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.

Lately, amino-functionalized N ω-carbamoylated arginines were introduced as arginine surrogates enabling peptide labeling. However, this approach is hardly compatible with peptides also containing lysine or cysteine. Here, we present the synthesis of an alkyne-functionalized, N ω-carbamoylated arginine building block (7), which is compatible with Fmoc-strategy solid-phase peptide synthesis. The alkynylated arginine was incorporated into three biologically active linear hexapeptides and into a cyclic pentapeptide. Peptide conjugation to an azido-functionalized fluorescent dye via “click” chemistry was successfully demonstrated. In the case of a peptide also containing lysine besides the alkyne-functionalized arginine, this was feasible in a “bioorthogonal” manner.

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