Azoospermia, which is the absence of spermatozoa in the ejaculate, is not a rare cause of male infertility. Inducible nitric oxide synthase (iNOS) is a calcium-independent NOS, which is present in the testis and involved in spermatogenesis, and apoptosis of Sertoli and germ cells. Twenty idiopathic infertile men presenting nonobstructive azoospermia were enrolled in this study, and testicular sperm extraction procedures were performed. Tissue extracts were dissected, and the fluid samples were investigated to determine the presence of spermatozoa. Histologic evaluation of the spermatozoa-present samples revealed that seminiferous tubules were normal and were lined by Sertoli cells and spermatogenic cells. However, in the spermatozoa-absent samples, the diameter of the seminiferous tubules was small, and Sertoli-cell-only syndrome was determined in most of the tubules. iNOS expression was very weak in Sertoli cells, germ cells, and in Leydig cells in the spermatozoa-present group. In the spermatozoa-absent group, the immunostaining was very intense in Sertoli and Leydig cells. Electron microscopy findings were supported the histologic results. In conclusion, complete germ cell loss and intense expression of iNOS in the Sertoli and Leydig cells in the spermatozoa-absent groups of azoospermic human testis suggest an essential role of iNOS in spermatogenesis.