SP1 activates AKT3 to facilitate the development of diabetic nephropathy.

BACKGROUND

Diabetic nephropathy (DN) is a severe complication of diabetes mellitus and has the complex pathogenesis. The previous study reported that protein kinase Bγ (AKT3) was involved in DN progression. Our aim was to explore the detailed mechanisms of AKT3 in DN development.

METHODS

RT-qPCR was performed to measure the levels of specificity protein 1 (SP1) and AKT3. Mesangial cells were treated with high glucose (30 mM) to form DN cell model in vitro. Western blot was conducted to detect the protein expression of AKT3, SP1, fibrosis-related proteins, and AKT/mTOR pathway-related proteins. Cell proliferation and inflammation were evaluated via MTT, EdU staining, and ELISA assays, respectively. Oxidative stress was determined via measuring ROS and MDA levels. ChIP and dual-luciferase reporter assays were carried out to verify the relationship between SP1 and AKT3. C57BL/6 mice-treated with streptozotocin for 5 days were used to establish DN mouse model in vivo, and HE and Masson staining were conducted to evaluate pathological changes of mouse kidney tissues.

RESULTS

AKT3 and SP1 were highly expressed in DN kidney tissues and HG-induced mesangial cells. AKT3 depletion could relieve HG treatment-caused cell damage of mesangial cells through repressing cell proliferation, fibrosis, inflammation and oxidative stress. SP1 can bind to the promoter of AKT3 and serve as a translation regulation factor of AKT3. SP1 overexpression worsened HG treatment-caused cell damage of mesangial cells. Moreover, AKT3 upregulation could block the suppressive effects of SP1 depletion on cell proliferation, fibrosis, inflammation and oxidative stress in HG-induced mesangial cells. SP1 depletion reduced AKT3 expression to inactivate the AKT/mTOR pathway in HG-induced mesangial cells. Besides, AKT3 knockdown inhibited the activation of the AKT/mTOR pathway to hamper the development of DN in mice through alleviating fibrosis and inflammation in vivo.

CONCLUSION

Our results indicated that SP1 activated AKT3 and AKT/mTOR pathway to promote mesangial cell proliferation, fibrosis, inflammation and oxidative stress, thereby facilitating DN development.