Spermatogenesis is disrupted easily by modest increase the temperature or by other environmental insults. Cryptorchid testes in most mammals show the arrested spermatogenesis because of the heat. It has been thought that there are two pathways to determine the cell fate after the heat shock to survive for further differentiation or to do cell death. Induced stress by heat shock in cryptorchid testis usually increase the expression of heat shock factors (HSFs) and eliminate the germ cells by apoptosis through expression of T-cell death associated gene 51 (TDAG51) (Hayashida et al., 2006). On the other hands, increased expression levels of HSFs have another role to protect the germ cells from the heat shock by increase the expression levels of heat shock proteins (HSPs) which assist protein folding and inhibit protein denaturation (Parsell and Lindquist, 1993; Young et al., 2004). Furthermore, it is known that the elevated temperature disrupt testicular tight junctions and caused a defect of which may result in the abnormal spermatogenesis in cryptorchidism (Chen et al., 2012). Because the elephant testis shows the normal spermatogenesis in the cryptorchid testis, we postulate that the responses of the elephant testicular cells to the heat shock by molecular level change related to the heat shock proteins, may differ from those of other mammalian species. Thus, the change of the balanced expression level of these molecules related to the heat stress, may induce the arrest of spermatogenesis in the testis. In other words, the response to the heat shock by appropriate molecular level change on these molecules may rescue the spermatogenic cells after heat shock. To know the effect of heat shock in the elephant testis, we investigated the expression levels of heat shock related molecules after application of the heat shock using elephant testicular tissues and fibroblast cells.

Quantitative image analysis of the immuno-staining show the induction of HSF1 by heat shock on both elephant fibroblast cells and elephant spermatogenic cells of testicular tissues under culture condition after heat shock 0-2 hr, indicating that the heat shock response by HSF1 to both somatic cells and germ cells is conserved in the elephant as same as in the other mammals. Furthermore, the expression levels of T-cell death associated gene 51 (TDAG51) increased on both elephant fibroblast cells and elephant spermatogenic cells in the testicular tissues under culture condition after heat shock 1-2 hr, suggesting that the heat shock response to eliminate the defected cells through TDAG 51 expression is conserved in the elephant as same as in the other mammals. On the other hands, HSP70i was induced after increase of HSF1 in elephant fibroblast cells but not in the elephant spermatogenic cells of the testicular tissue, suggesting that there is a different system to protect cells from heat shock between somatic cells and spermatogenic cells in elephant. B-actin expression levels of both elephant fibroblast cells and elephant spermatogenic cells of testicular tissues under culture condition did not show any changes by heat shock.

 Finally, the expression levels of heat shock related molecules among the mouse cryptorchid testis, mouse adult testis and elephant testis were quantitatively analyzed using b-actin expression for normalization between the samples. HSF1 and connexin 43 expression levels in the elephant testis was preserved as similar level as those in the normal mouse adult testis and higher than those in the cryptorchid mouse testis. These results suggest that in the elephant testis, the induction of apoptosis through HSF1/TDAG51 pathway was inhibited and tight junction complex was preserved as same levels as in the mouse adult testis showing normal spermatogenesis, may result in normal spermatogenesis under cryptorchidism in elephant.