High glucose level and angiotensin II type 1 receptor stimulation synergistically amplify oxidative stress in renal mesangial cells

Tetsuya Akaishi,Sadayoshi Ito,Michiaki Abe,Takaaki Abe,Kota Ishizawa,Hiroshi Okuda,Tadashi Ishii

doi:10.1038/s41598-019-41536-z

Abstract

Oxidative stress in renal mesangial cell causes diabetic glomerular changes. High glucose levels and angiotensin II (Ang II) are known to stimulate superoxide production in renal mesangial cells. However, it has been unclear whether Ang II stimulation and pre-conditioning with high glucose affects the same pathway of superoxide production in renal mesangial cells or not. In this study, we examined the levels of oxidative stress under Ang II stimulation in renal mesangial cells preincubated for six hours at various glucose levels. Intracellular levels of reactive oxidative species (ROS) were measured using dihydroethidium or 5′,6′-chloromethyl- 2′,7′ dichlorodihydro-fluorescein diacetate, which facilitates the detection of intracellular ROS under real-time fluorescent microscope. Ang II-induced elevated intracellular ROS levels were detected only when the cells were pre-incubated with high levels of glucose (13.5 mM, 27.8 mM), but was not detected under normal glucose condition (5.5 mM). Production of Ang II-induced intracellular ROS was higher under pre-treatment with 27.8 mM glucose compared to pretreatment with 13.5 mM glucose level. This ROS production in mesangial cells was induced within several minutes of the initiation of Ang II stimulation under high glucose levels. The production of intracellular ROS was significantly reduced in the presence of angiotensin II type1-receptor (AT1R) antagonist, whereas it was augmented in the presence of angiotensin II type2-receptor antagonist. In conclusion, Ang II-induced oxidative stress was augmented by high glucose levels and ROS levels were further alleviated in the presence of AT1R antagonists.

Highlights

Renal glomerular changes in diabetic patients are recognized as mesangial expansion and fusion of foot processes on podocytes[1,2]
The production of Eth was significantly increased by pre-conditioning with 13.5 mM glucose www.nature.com/scientificreports along with Ang angiotensin II (II) stimulation, compared to that in control; it was not increased after pre-conditioning with 5.5 mM glucose even with angiotensin II (Ang II) stimulation
There was no increase in intracellular reactive oxidative species (ROS) level after pre-treatment with angiotensin II type1receptor (AT1R) antagonist (RNH6720) and it was comparable with the ROS level in control

Summary

Introduction

Renal glomerular changes in diabetic patients are recognized as mesangial expansion and fusion of foot processes on podocytes[1,2] These glomerular pathological changes are triggered by oxidative stress and one of the generator of such changes is mesangial cell (MC). Since microalbuminuria, which is associated with diabetic microvascular complication, can be observed in the early stage of impaired glucose tolerance, there is a possibility that such ROS production occurs in this condition under mildly elevated glucose levels. Since microalbuminuria, which is associated with diabetic microvascular complication, can be observed in the early stage of impaired glucose tolerance, there is a possibility that such ROS production occurs in this condition under mildly elevated glucose levels10 If such a speculation could be validated, it would strongly suggest that hyper-activated ROS production in renal mesangial cells are associated with the pathogenesis of diabetic glomerular injury from the early stages. To investigate the synergistic negative effect of glucose and Ang II, Ang II was administered at different timings in the presence of glucose

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Conclusion