Cell-specific expression of X-linked inhibitor of apoptosis in the anterior pituitary of streptozotocin-induced diabetic rats

Abstract

Cell death is increased in the anterior pituitary of poorly controlled diabetic rats, but anti-apoptotic mechanisms are also activated. We hypothesized that specific cell types are selectively protected against diabetes-induced cell death. To determine when anti-apoptotic mechanisms are activated, streptozotocin-induced diabetic rats were killed after 1, 4, 6 and 8 weeks of evolution. Anterior pituitaries were processed for western blot analysis to determine changes in the intrinsic cell death pathway and upstream kinases involved in cell protection mechanisms. An increase in cell death was detected by ELISA at 4 weeks of diabetes. TUNEL labelling demonstrated that this corresponded to death of primarily lactotrophs, a few somatotrophs, and no thyrotrophs, corticotrophs or gonadotrophs. Levels of phosphorylated (p) Akt were increased at 1 week of diabetes, while pERK1/2 levels increased at 4 weeks and pJNK at 6 weeks. Activation of caspase 3 decreased and anti-apoptotic members of the Bcl-2 protein family increased as early as 1 week after diabetes onset. These changes were coincident with increased IGF-I receptor levels. Levels of X-linked inhibitor of apoptosis protein (XIAP) increased significantly after 6 weeks of diabetes, as did activation of nuclear factor (NF)kB. Double immunohistochemistry indicated that XIAP was expressed in less than 1% of lactotrophs and gonadotrophs, approximately 50% of somatotrophs and more than 90% of corticotrophs and thyrotrophs. These results suggest that some cell survival mechanisms are rapidly activated in the anterior pituitary, even before increased cell death can be detected, while others are more delayed. Furthermore, both pituitary cell death and expression of protective mechanisms such as XIAP are celltype specific.

Ghrelin treatment protects lactotrophs from apoptosis in the pituitary of diabetic rats

“…Immunohistochemistry in combination with TUNEL ( Figure 2) indicated that his cell death corresponded primarily to lactotrophs as previously demonstrated (Arroba et al, 2007). Very few TUNEL labeled cells were found in pituitaries of control rats or diabetic rats treated with ghrelin.…”

“…However, we previously reported that no changes in cell turnover of thyrotrophs, corticotrophs or gonadotrophs occur in poorly controlled diabetic rats (Arroba et al, 2006;Arroba et al, 2007), suggesting that changes in the synthesis and secretion by individual cells could explain the observed changes in circulating hormones. In our study pituitary TSH and ACTH content was not modified by either diabetes or ghrelin treatment to diabetic rats.…”

“…Although XIAP is increased in the anterior pituitary of diabetic rats, this protein is expressed in less than 1% of lactotrophs and gonadotrophs, and in a higher proportion in somatotrophs (50%), corticotrophs (90%) and thyrotrophs (90%) (Arroba et al, 2007). This indicates that XIAP protects pituitary cell populations other than lactotrophs.…”

“…Immunohistochemistry/TUNEL assays were performed on frozen 12 µm cryostat pituitary mounted on positively charged slides as previously A c c e p t e d M a n u s c r i p t described (Arroba et al, 2003(Arroba et al, , 2005(Arroba et al, , 2006(Arroba et al, , 2007. The TUNEL assays were performed following the manufacturer's instructions (Roche).…”

“…We previously demonstrated that in poorly controlled diabetes caspase 8, the prototypical initiator caspase of the extrinsic cell death pathway (Boldin et al, 1996, Wajant, 2002, is activated and colocalizes with prolactin (PRL) producing cells (Arroba et al, 2005), possibly explaining their increased death rate. However, pituitary levels of proteins involved in the intrinsic cell death pathway, including members of the Bcl-2 family, X-linked inhibitor of apoptosis (XIAP), and the effector caspases 3 and 6, are either unchanged or balanced towards cell survival (Arroba et al, 2005), with these changes being cell-type specific (Arroba et al, 2007).…”