Leydig cells, located in the loose interstitial tissue of seminiferous tubules, are the major site for androgen synthesis and secretion, and play an important role in the reproductively and fertility of males. The dysfunction of Leydig cells may lead to various male diseases, such as primary hypogonadism, cryptorchidism, and hypospadias. This review outlines the recent findings concerning the generation, development and regulation of Leydig cells.