Cyclic AMP prevents decrease of phosphorylated ezrin/radixin/moesin and chloride intracellular channel 5 expressions in injured podocytes.
BACKGROUND
Our previous in vitro studies suggested that cyclin AMP (cAMP) signaling protects against podocyte injury. However, the molecular mechanisms remain unknown. The aim of the present study was to explore the role of forskolin, an agonist for adenylate cyclase, on ezrin/radixin/moesin (ERM) phosphorylation and chloride intracellular channel 5 (CLIC5) expressions in injured podocytes.


METHODS
ADR nephrosis model were induced by adriamycin (ADR) injection in BalB/C mice. Parts of ADR nephrosis mice were pretreated with forskolin. Albuminuria was estimated by urine Coomassie blue stain. Nephrin, synaptopodin, CLIC5, phosphorylated ERM and podocalyxin were measured by confocal microscopy. CLIC5 and phosphorylated ERM also were studied using western blotting. RhoA and Rac1 were estimated by G-Lisa kit.


RESULTS
We found that forskolin partially alleviated albuminuria and width of foot processes. Nephrin, synaptopodin, phosphorylated-ERM (p-ERM) and CLIC5 expression were decreased in ADR mice, which were improved by forskolin pretreatment. In vitro studies, pretreatment of podocytes with pCPT-cAMP(PKA-selective cAMP analogue)prevented puromycin aminonucleoside (PAN)-induced CLIC5 downregulation. 8-pCPT-2'-O-Me-cAMP (2Me-cAMP, an Epac-selective cAMP analogue) blocked PAN-induced p-ERM downregulation. PAN inhibited RhoA activation in podocytes, which could be prevented by pCPT-cAMP pretreatment. Y-27632, a Rho inhibitor, decreased CLIC5 expression in podocytes.


CONCLUSIONS
Activation cAMP signaling might attenuate albuminuria in ADR-induced nephrosis mice. Different downstream signaling pathway might mediate cAMP protection on CLIC5 and p-ERM expression, respectively.
Renal Division and Molecular Cell Lab for Kidney Disease, Renji Hospital, School of Medicine, Shanghai Jiaotong University, 160#, Pu Jian Road, Shanghai, 200001, China.
Hua
H
Tao
Renal Division and Molecular Cell Lab for Kidney Disease, Renji Hospital, School of Medicine, Shanghai Jiaotong University, 160#, Pu Jian Road, Shanghai, 200001, China.
Xiaoying
X
Li
Renal Division and Molecular Cell Lab for Kidney Disease, Renji Hospital, School of Medicine, Shanghai Jiaotong University, 160#, Pu Jian Road, Shanghai, 200001, China.
Kai
K
Wei
Renal Division and Molecular Cell Lab for Kidney Disease, Renji Hospital, School of Medicine, Shanghai Jiaotong University, 160#, Pu Jian Road, Shanghai, 200001, China.
Kewei
K
Xie
Renal Division and Molecular Cell Lab for Kidney Disease, Renji Hospital, School of Medicine, Shanghai Jiaotong University, 160#, Pu Jian Road, Shanghai, 200001, China.
Zhaohui
Z
Ni
Renal Division and Molecular Cell Lab for Kidney Disease, Renji Hospital, School of Medicine, Shanghai Jiaotong University, 160#, Pu Jian Road, Shanghai, 200001, China. guleyi@aliyun.com.|Renal Division, Kashgar Prefecture Second People's Hospital of Xinjiang Uygur Autonomous Region, 1#, Jian Kang Road, Kashgar, Xinjiang Uygur Autonomous Region, China. guleyi@aliyun.com.
Leyi
L
Gu
D000818:Animals; D000900:Anti-Bacterial Agents; D018118:Chloride Channels / Q000187:drug effects / Q000378:metabolism*; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000494:pharmacology*; D003598:Cytoskeletal Proteins / Q000187:drug effects / Q000378:metabolism*; D004317:Doxorubicin; D008297:Male; D008565:Membrane Proteins / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008840:Microfilament Proteins / Q000187:drug effects / Q000378:metabolism*; D009401:Nephrosis / Q000139:chemically induced / Q000378:metabolism / Q000473:pathology; D010766:Phosphorylation / Q000187:drug effects*; D050199:Podocytes / Q000187:drug effects / Q000378:metabolism*; D014665:Vasodilator Agents / Q000494:pharmacology; D020741:rho GTP-Binding Proteins / Q000378:metabolism; D020742:rhoA GTP-Binding Protein; D000099149:Ezrin
D000900:Anti-Bacterial Agents; D018118:Chloride Channels; D005576:Colforsin; D000242:Cyclic AMP; D003598:Cytoskeletal Proteins; D004317:Doxorubicin; D008565:Membrane Proteins; D008840:Microfilament Proteins; D014665:Vasodilator Agents; D020741:rho GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein; D000099149:Ezrin; C514190:CLIC5 protein, mouse; C071058:moesin; C071776:radixin; C526380:RhoA protein, mouse
Actin; Albuminurine; Cytoskeleton