Presentation Authors: Filippo Pederzoli*, Irene Locatelli, Paolo Capogrosso, Eugenio Ventimiglia, Manuela Nebuloni, Massimo Clementi, Nicasio Mancini, Ferrarese Roberto, Francesco Montorsi, Andrea Salonia, Massimo Alfano, Milan, Italy
Introduction: As the local environment demonstrated to support spermatogenesis in animal models, we aimed to study the potential modification of 2 relevant extracellular microenvironments - i.e. the extracellular matrix (ECM) and the bacterial microbiome (BM) - in the testicular pulps of idiopathic non-obstructive azoospermia (iNOA) men.
Methods: Testicular tissues and complete clinical data from a cohort of white-Caucasian iNOA primary infertile men who underwent microTESE were used for these analyses. Specimens were further subdivided according to positive (P) vs. negative (N) sperm retrieval at surgery. Non-neoplastic specimens with normal spermatogenesis were collected from men undergoing orchiectomy for non-metastatic seminoma (most distant areas from tumor). Testicular histology was classified according to the Johnsen score and fibrosis evaluated by the Masson staining. Tissue-associated BM was analyzed up to the level of genus by performing amplifications of the V3-V5 region of 16S rRNA by nested PCR. Taxonomy was assigned using the RDP Classifier.
Results: At histology, P specimens showed germ cell arrest with rare foci of preserved spermatogenesis; conversely, N specimens suggested a Sertoli cells only syndrome. The Masson trichrome staining depicted an increased collagen deposition around the seminiferous tubules in iNOA men, which was steadily associated with a decrease of sperm retrieval; collagen thickness >10 microm was associated with the complete absence of sperm inside the seminiferous tubules. An increased amount of 16S DNA was found within testicular specimens from iNOA men (12.1; 5-15). Tissues with normal germline maturation showed detectable amounts of 16S DNA (median; 3.4; IQR; 1.7-7 copies/ng of loaded DNA). Taxonomic diversity of the BM profiling showed significant differences among samples (Î±-diversity) and BM clusterization within the testicular specimens from normozoospermic and iNOA men (Î²-diversity). Actinobacteria, Bacteroidetes, Firmicutes and Proteobacteria were phyla associated with a normal germline; conversely, specimens from iNOA individuals showed decreased taxa richness. P specimens from iNOA men had more Actinobacteria and Firmicutes, whereas N specimens had mainly Actinobacteria.
Conclusions: Current findings suggested that the human testis is not microbiologically sterile. Different BMs are associated with normal vs. iNOA testicular pulps. Moreover, we depicted the association among seminiferous tubules-associated fibrosis, germ cell aplasia and bacterial dysbiosis.