摘要
Background Intrarenal activation of the renin angiotensin system (RAS) plays an important role in mediating renal fibrosis. Both angiotensin converting enzyme inhibitors (ACEIs) and angiotensin Ⅱ (AngⅡ) receptor antagonists have been shown to exert a protective role against diabetic and non-diabetic nephropathy. However, the exact mechanism of how blocking local RAS prevents renal fibrosis is unclear. The present study was to investigate the influence of a new AngⅡ receptor antagonist, irbesartan (Irb), on AngⅡ-induced hypertrophy in human proximal tubular cell line (HK-2). Methods The cell line, HK-2, was grown in Dulbeccos’s Modified Eagle’s Medium containing 10% heat-inactivated fetal calf serum. After rested in serum-free medium for 24 hours, the effects of Irb on AngⅡ (10 -7 mol/L)-induced [ 3H]-leucine incorporation, total protein content (measured by the Coomassie brilliant blue G250 method), and change in cell size (determined by scanning electron microscopy) were observed. The influence of Irb on the cell cycle was analyzed by fluorescence activated cell sorter (FACS) flow cytometry. Results AngⅡ induced cell hypertrophy in a time and dose dependent manner. Stimulation of cells with AngⅡ for 48 hours resulted in a increase in [ 3H]-leucine incorporation [0 hour: (5584±1016) cpm/10 5cells vs 48 hours: (10741±802) cpm/10 5cells, P<0.05], which was significantly attenuated by treatment with Irb. AngⅡ significantly increased the total protein content in HK-2 cells [control: (0.169±0.011) mg/10 5cells vs AngⅡ group: (0.202±0.010) mg/10 5 cells, P<0.05], which was also markedly inhibited by cotreatment with Irb (P<0.01). Scanning electron microscopy showed that AngⅡ induced an increase in average physical cell size, which was significantly inhibited by Irb [control: (11.92±1.62) μm; AngⅡ group: (20.63±3.83) μm; AngⅡ+Irb group: (13.59±3.15) μm; P<0.01 vs control, respectively]. Furthermore, flow cytometry revealed that AngⅡ arrested cells in the G 0-G 1 phase, which was significantly reversed by treatment with Irb [G 0-G 1 cells in AngⅡ group: (76.09±1.82)%, in AngⅡ+Irb group: (67.00±2.52)%, P<0.05].Conclusion Irb can inhibit AngⅡ-induced hypertrophy in HK-2 cells.
Background Intrarenal activation of the renin angiotensin system (RAS) plays an important role in mediating renal fibrosis. Both angiotensin converting enzyme inhibitors (ACEIs) and angiotensin Ⅱ (AngⅡ) receptor antagonists have been shown to exert a protective role against diabetic and non-diabetic nephropathy. However, the exact mechanism of how blocking local RAS prevents renal fibrosis is unclear. The present study was to investigate the influence of a new AngⅡ receptor antagonist, irbesartan (Irb), on AngⅡ-induced hypertrophy in human proximal tubular cell line (HK-2). Methods The cell line, HK-2, was grown in Dulbeccos’s Modified Eagle’s Medium containing 10% heat-inactivated fetal calf serum. After rested in serum-free medium for 24 hours, the effects of Irb on AngⅡ (10 -7 mol/L)-induced [ 3H]-leucine incorporation, total protein content (measured by the Coomassie brilliant blue G250 method), and change in cell size (determined by scanning electron microscopy) were observed. The influence of Irb on the cell cycle was analyzed by fluorescence activated cell sorter (FACS) flow cytometry. Results AngⅡ induced cell hypertrophy in a time and dose dependent manner. Stimulation of cells with AngⅡ for 48 hours resulted in a increase in [ 3H]-leucine incorporation [0 hour: (5584±1016) cpm/10 5cells vs 48 hours: (10741±802) cpm/10 5cells, P<0.05], which was significantly attenuated by treatment with Irb. AngⅡ significantly increased the total protein content in HK-2 cells [control: (0.169±0.011) mg/10 5cells vs AngⅡ group: (0.202±0.010) mg/10 5 cells, P<0.05], which was also markedly inhibited by cotreatment with Irb (P<0.01). Scanning electron microscopy showed that AngⅡ induced an increase in average physical cell size, which was significantly inhibited by Irb [control: (11.92±1.62) μm; AngⅡ group: (20.63±3.83) μm; AngⅡ+Irb group: (13.59±3.15) μm; P<0.01 vs control, respectively]. Furthermore, flow cytometry revealed that AngⅡ arrested cells in the G 0-G 1 phase, which was significantly reversed by treatment with Irb [G 0-G 1 cells in AngⅡ group: (76.09±1.82)%, in AngⅡ+Irb group: (67.00±2.52)%, P<0.05].Conclusion Irb can inhibit AngⅡ-induced hypertrophy in HK-2 cells.
基金
ThisprojectwassupportedbytheJiangsuProvinceNaturalScienceFund(No BK2 0 0 2 0 5 2 )andtheKeyMedicalTalentTrainingProgramoftheJiangsuProvinceHealthBureau (No RC2 0 0 2 0 72 )
