Glycogen Synthase Kinase 3β Regulates Podocyte Cytoskeleton: Implications Of The Pathogenesis And Therapy Of Podocytopathy

Glomerular disease is the most common original disease of end-stage-renal-disesase (ESRD) in China. Podocyte, forms the out layer of the glomerular filtration barrier (GFB) and plays a pivotal role in maintaining the normal glomerular filtration. And the podocyte dysfunction is considered to be one of the culprits of proteinuric disease. Thus to explore the mechanism of podocyte injury and the potential therapeutic target is important.Podocyte injury mainly includes foot process effacement and podocyte loss. Podocyte foot process effacement usually related to the actin cytoskeleton which distributed in the foot process and the microtubules which distributed in podocyte cell body and major process mianly control the major process elongation. Our research explored the effect of microfilaments and microtubules in podocyte injury and compensatory repair upon Adriamycin injury and demonstrated the Glycogen synthase kinase 3β (GSK3β) on the one hand mediates podocyte injury by controlling paxillin activity and focal adhesion turnover and thus induces microfilaments disassembling and foot process effacement; and on the other hand mediates microtubule polymerization by controlling microtubule associate proteins and regulates podocyte major process elongation and the compensatory adaptation of podocyte.Aberrant focal adhesion turnover is centrally involved in podocyte actin cytoskeleton disorganization and foot process effacement. The structural and dynamic integrity of focal adhesions is orchestrated by multiple cell signaling molecules, including GSK3β, a multitasking kinase lately coined as a mediator of kidney injury. In adriamycin injured podocytes, lithium, a GSK3β inhibitor and neuroprotective mood stabilizer, obliterated the accelerated focal adhesion turnover, rectified podocyte hypermotility, and restored actin cytoskeleton integrity. Mechanistically, lithium counteracted the adriamycin elicited GSK3β overactivity and hyperphosphorylation and overactivation of paxillin, a focal adhesion associated adaptor protein. Moreover, forced expression of a dominant negative kinase dead mutant of GSK30 highly mimicked, whereas, ectopic expression of a constitutively active GSK3β mutant abolished, the effect of lithium in adriamycin injured podocytes, suggesting that the effect of lithium is mediated, at least in part, through inhibition of GSK3β. Furthermore, paxillin interacted with GSK3β and served as its substrate. In mice with adriamycin nephropathy, a single low dose of lithium ameliorated proteinuria and glomerulosclerosis. Consistently, lithium therapy abrogated GSK3β overactivity, blunted paxillin hyperphosphorylation and reinstated actin cytoskeleton integrity in glomeruli, associated with an early attenuation of podocyte foot process effacement. Thus, the GSK3β modulated focal adhesion dynamics might serve as a novel therapeutic target for podocytopathy.Upon podocyte loss, remnant intact podocytes exhibit a considerable adaptive capacity to extend their cellular processes and cover the denuded glomerular basement membrane, reminiscent of neural repair. Microtubules, one of the principal cytoskeletal components of podocyte major processes, play a crucial role in podocyte morphogenesis and podocyte process outgrowth, branching and elongation. Here, we demonstrated that microtubule associated proteins tau and collapsin response mediator protein (CRMP)2, key regulators of microtubule dynamics, were abundantly expressed by glomerular podocytes in vivo and in vitro, interacted with GSK3β and served as its putative substrates. GSK3β overactivity induced by adriamycin injury or a constitutively active mutant of GSK3β caused hyperphosphorylation of tau and CRMP2, concomitant with microtubule depolymerization, cell body shrinkage and shortening of podocyte processes. Conversely, inhibition of GSK3β by a dominant negative mutant or lithium diminished tau and CRMP2 phosphorylation, resulting in microtubule polymerization, podocyte expansion and lengthening of podocyte processes. In the mouse model of adriamycin induced podocyte depletion and nephropathy, delayed administration of a single low dose of lithium attenuated proteinuria and ameliorated progressive glomerulosclerosis despite no correction of podocytopenia. Mechanistically, lithium therapy obliterated GSK3β overactivity, counteracted hyperphosphorylation of tau and CRMP2 and enhanced microtubule polymerization and stabilization in glomeruli in adriamycin injured kidneys, associated with elongation of podocyte major processes. Collectively, our findings suggest that therapeutic targeting of GSK3P restores podocyte microtubule integrity and reinforces the compensatory adaptation of glomeruli following podocyte depletion.In conclusion, our research demonstrated the mechanism of GSK3β regulated cytokelston change in podocyte injury and repair, suggesting a potential new direction of the treatment of podocytopathy and proteinuric disease.