Reduced Krüppel-like factor 2 expression may aggravate the endothelial injury of diabetic nephropathy

Abstract

Kruppel-like Factor 2 (KLF2), a shear-stress inducible transcription factor, has endoprotective effects. In streptozotocin-induced diabetic rats, we found that glomerular Klf2 expression was reduced in comparison to non-diabetic rats. However, normalization of hyperglycemia by insulin treatment increased Klf2 expression to a level higher than that of non-diabetic rats. Consistent with this, we found that Klf2 expression was suppressed by high glucose but increased by insulin in cultured endothelial cells. To determine the role of KLF2 in streptozotocin-induced diabetic nephropathy, we used endothelial cell-specific Klf2 heterozygous knockout mice and found that diabetic knockout mice developed more kidney/glomerular hypertrophy and proteinuria than diabetic wide type mice. Glomerular expression of Vegfa, Flk1, and angiopoietin 2 increased but expression of Flt1, Tie2, and angiopoietin 1 decreased in diabetic knockout compared to diabetic wide type mice. Glomerular expression of ZO-1, glycocalyx, and eNOS was also decreased in diabetic knockout compared to diabetic wide type mice. These data suggest knockdown of Klf2 expression in the endothelial cells induced more endothelial cell injury. Interestingly, podocyte injury was also more prominent in diabetic knockout compared to diabetic wide type mice, indicating a crosstalk between these two cell types. Thus, KLF2 may play a role in glomerular endothelial cell injury in early diabetic nephropathy.

Reduced Krüppel-Like Factor 2 Aggravates Glomerular Endothelial Cell Injury and Kidney Disease in Mice with Unilateral Nephrectomy

“…30 We also recently showed that eNOS expression was reduced in the diabetic Klf2 heterozygous KO mice as compared to diabetic WT mice. 18 Similarly, we observed herein that mRNA levels of eNOS were significantly decreased in the glomeruli of KO-UNX as compared to all other groups ( Figure 4A). Reduction of eNOS specifically in the KO-UNX glomeruli was further confirmed at the protein level by immunostaining (Figure 4, B and C) and by Western blot analyses (Figure 4, DeF).…”

“…Our previous study showed a 50% reduction of Klf2 mRNA expression in isolated glomerular endothelial cells from KO kidneys as compared to those from WT kidneys. 18 In addition, at baseline, KO mice did not exhibit albuminuria, glomerular injury, or tubulointerstitial disease when examined at 6 and 12 months of age (data not shown).…”

“…26 Recently, we reported that reduced KLF2 expression in endothelial cells leads to more severe glomerular endothelial cell injury and kidney damage in early diabetic nephropathy, which is characterized by glomerular hyperfiltration. 18 However, we were unable to assess the relationship between hyperfiltration and KLF2 expression in diabetic mice, because KLF2 expression is also affected by high glucose and insulin. Therefore, we used the UNX model, which allows us to address the role of KLF2 in mediating glomerular hyperfiltrationeinduced kidney injury.…”

“…Specimens were initially baked for 20 minutes in 55 C to 60 C oven and then processed as described previously. 18 Briefly formalin-fixed and paraffin-embedded sections were deparaffinized, and endogenous peroxidase was inactivated with hydrogen peroxide. Sections were then blocked in 2% goat serum in phosphatebuffered saline for 1 hour at room temperature and then incubated with a rabbit anti-KLF2 antibody (1:1000, GenScript, Piscataway, NJ) at 4 C overnight.…”

“…17 Our recent study suggests that KLF2 is up-regulated by both high glucose and insulin and it protects against endothelial cell injury in early diabetic nephropathy. 18 In addition to high glucose and insulin, we hypothesized that the increased shear stress in glomerular endothelial cells during the early stages of diabetic nephropathy would contribute to the up-regulation of KLF2 expression. We were previously unable to address this hypothesis in the diabetic kidney because of the effects of high glucose and insulin on KLF2 expression.…”