In order to examine the male toxicity of nonylphenol after it enters the body, the estrogencity and potential mechanisms of mixture of nonylphenol and 17-Estradiol (E2), a typical natural estrin on rat sertoli cells, was studied. The results indicated that nonylphenol and E2 could promote the growth of sertoli cells, showing certain male toxicity, and their mixture showed significant enhancement effect. Furthermore, E2 did not impact the expression of vimentin protein, a key protein in sertoli cells. Nonylphenol increased its expression, and the mixture significantly increased its expression. Mechanism study showed that, among the three pathways of MAPK channel, ERK protein was the key for E2 enhancing nonylphenol male toxicity. Both nonylphenol and E2 decreased the expression of ERK protein, but the mixture increased its expression. Nonylphenol noticeably promoted the expression of p-JNK protein and p38 protein, which was weakened when mixed with E2. Consequently, as the result of ERK channel activation, the mixture of nonylphenol and E2 significantly increased the expression of transcription factors, c-Myc proteins and cyclinD1 proteins.