P16ink4a Expression Is Increased through 12-Lipoxygenase in High Glucose-Stimulated Glomerular Mesangial Cells and Type 2 Diabetic Glomeruli

Abstract

Background/Aims: Arachidonic acid-metabolizing enzyme, 12-lipoxygenase (12-LO), is involved in the glomerular hypertrophy of diabetic nephropathy (DN), in which cyclin-dependent kinase inhibitors (CKIs) play important roles. However, it is unclear whether 12-LO regulates the expression of the CKI p16^ink4a in DN. Methods: Primary glomerular mesangial cells (MCs) and glomeruli isolated from rats were used in this study. The rats were fed a high-fat diet and given low-dose streptozotocin to induce type 2 diabetes. The 12-LO product, 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE), was infused through an osmotic minipump. Enzyme-linked immunosorbent assay, Western blot, and morphometric analyses were performed. Results: High glucose (HG) increased the p16^ink4a protein expression in MCs, but this increase was prevented by the 12-LO inhibitor, cinnamyl-3,​4-dihydroxy-α-cynanocinnamate (CDC). The levels of p-p38MAPK and p16^ink4a in MCs were significantly elevated after the 12(S)-HETE treatment, whereas the p38MAPK inhibitor SB203580 prevented these increases. Compared with levels in control MCs, marked increases in p38MAPK activation and p16^ink4a expression were observed in MCs plated on collagen IV, while the CDC treatment prevented these changes. Subcutaneous injection of CDC did not affect glucose levels, but completely attenuated the diabetes-related increases in the 12(S)-HETE content, p16^ink4a expression, p-p38MAPK levels, glomerular volume, and the kidney/body weight ratio. Compared with levels in controls, p16^ink4a and p-p38MAPK in the glomeruli derived from 12(S)-HETE-treated rats were significantly higher. Conclusions: 12-LO-p38MAPK mediates the upregulation of p16^ink4a in HG-stimulated MCs and type 2-diabetic glomeruli, and new therapies aimed at 12-LO inhibition may prove beneficial in ameliorating diabetes-induced glomerular hypertrophy.