Human peritubular cells as a model for the study of pathological processes in the human testis: TNFα induced synthesis of nerve cell growth factor (NGF) and inflammatory mediators

Objectives: In testes of infertile men spermatogenic defects are often associated with tissue remodelling and fibrosis, as well as increased innervation density and the presence of inflammatory cells, e.g. mast cells and macrophages, which are producers of TNFα. To investigate the involvement of TNFα in pathomechanisms associated with male infertility, we examined whether human testicular peritubular cells (HTPCs) are targets for TNFα.

Methods: Isolated HTPCs were chosen as model, because they are supposed to be responsible for peritubular fibrosis, a hallmark of male infertility. Gene arrays, RT-PCR, Western blotting (WB), immunocytochemistry (ICC), as well as zymography and ELISA techniques were employed.

Results: Using ICC, TNF receptors 1 and 2 were localized on HTPCs. Employing WB, we detected phosphorylation of ERK1/2 in HTPCs after stimulation with recombinant human TNFα (5–100ng/ml) for 20 minutes, indicative of MAPK pathway activation. Long-term exposure (7 days) with TNFα resulted in a flattened and elongated cell morphology. However, TNFα did not alter the production of extracellular matrix (ECM) components (e.g. collagen-1, fibronectin) or ECM remodelling enzymes (e.g. MMP-2, TIMP-1), as shown by gene arrays, RT-PCR and zymography. RT-PCR studies indicated induction of inflammatory markers (MCP-1, IL-6), and WB showed a 2-fold increase in cyclooxygenase-2 (COX2) protein after TNFα stimulation. Unexpectedly, HTPCs express the NGF gene and secrete NGF protein under basal conditions. Stimulation with TNFαincreased mRNA levels of NGF (3-fold after 3 hours) and release nearly 5-fold after 24 hours.

Conclusion: TNFα affects human peritubular cells and the subsequent production of COX2 and release of NGF, as well as other inflammatory factors, may in part explain the increase in innervation and inflammatory cells commonly observed in testes of infertile men.

(DFG MA 1080/16–1; First and second author contributed equally).