Cisplatin is a DNA alkylating agent with well proved efficiency in the treatment of a wide range of solid tumors and common adverse effect is nephrotoxicity, mainly due to the apoptotic or necrotic of renal epithelial tubule cells. Recently a new protein HAX-1 was found related to mitochondria or calcium overloading induced apoptosis. From our previous study shown that cisplatin-induced ROS can inhibit HAX-1 expression and induce kidney epithelium cell apoptosis. However, the molecular mechanism physiology role of HAX-1 in cisplatin-induced nephrotoxicity is still obscured in vivo as a result of death of HAX-1 knock out mice as early as five weeks after breath. Based on this reason, the goal of this application is to understand the regulation of HAX-1 and its downstream calcium-dependent signaling pathways during cisplatin-induced kidney failure. Therefore, in this proposal, we aim to elucidate the following problems: (1) What is the physiology role of HAX-1 following cisplatin-induced kidney injury in kidney specific conditional HAX-1 knock out mice (HAX-1KSP-/-) or in WT mice? (2) What are the signal transduction pathways of HAX-1 in kidney following cisplatin induced injury? To reach our goal, three specific aims will be pursued Aim 1: To investigate how HAX-1 is regulated in cisplatin-induced of kidney failure We will first analyze gene or protein level of HAX-1 whether is regulated by cisplatin induced kidney injury. Aim 2. To determine pathology or physiology role of HAX-1 in the cisplatin induced development of kidney failure. Conditional kidney HAX-1 knock out mice (HAX-1 KSP-/-) mice and restoration of HAX-1 in HAX-1 KSP-/- mice by lentivirus (gain of function) both will be generated and subjected to cisplatin injury in kidney. Aim 3: To characterize the signal pathway of HAX-1 after cisplatin treatment We will overexpress different splicing forms of HAX-1 in vitro and in vivo to characterize the relationship between HAX-1 and apoptosis gene like caspase, AKT, eNOS or calcium concentration regulator like SERCA2, RYR. Furthermore, we will also utilize yeast two hybridize system to screen some uncovered gene which associated with HAX-1 to elucidate possible role of HAX-1. This study, our delineation of molecular mechanisms of HAX-1 in kidney epithelium cells will help to understand the physiological role of HAX-1 and lead to developing novel therapeutic approaches for cisplatin-mediated nephrology. Simultaneously, we also anticipate that the successfully generated transgenic mice containing floxed allele of HAX-1 cross with other tissue-specific Cre mice and generate non-kidney tissue-specific-knock-out offspring, providing an excellent tool for studying signaling and functional effects in certain tissues or organs excluding the kidney.
|Effective start/end date||8/1/12 → 7/31/13|