Growth hormone exacerbates diabetic renal damage in male but not female rats

Abstract

Background: Human and animal studies support the idea that there are sex differences in the development of diabetic renal disease. Our lab and others have determined that in addition to Ang II (through the AT1R), growth hormone (GH) contributes to renal damage in models of renal failure; however, the impact of sex and GH on the mechanisms initiating diabetic renal disease is not known. This study examined the effect of sex and GH on parameters of renal damage in early, uncontrolled streptozotocin (STZ)-induced diabetes.

Methods: Adult male and female Sprague-Dawley rats were injected with vehicle (control), STZ, or STZ + GH and euthanized after 8 weeks.

Results: Mild but significant glomerulosclerosis (GS) and tubulointerstitial fibrosis (TIF) was observed in both kidneys from male and female diabetic rats, with GH significantly increasing GS and TIF by 30% and 25% in male rats, but not in female rats. STZ increased TGF-β expression in both kidneys from male and female rats; however, while GH had no further effect on TGF-β protein in diabetic females, GH increased TGF-β protein in the male rat's kidneys by an additional 30%. This sex-specific increase in renal injury following GH treatment was marked by increased MCP-1 and CD-68+ cell density. STZ also reduced renal MMP-2 and MMP-9 protein expression in both kidneys from male and female rats, but additional decreases were only observed in GH-treated diabetic male rats. The sex differences were independent of AT1R activity.

Conclusions: These studies indicate that GH affects renal injury in diabetes in a sex-specific manner and is associated with an increase in pro-inflammatory mediators.

Keywords: Gender; Glomerulosclerosis; Inflammation; Renal Injury; Sex Differences; TGF-β.

Figure 1

Effect of sex on GSI in Control and diabetic rats treated with and without GH. (A) STZ treatment significantly increased the GSI in both sexes, while GH treatment significantly increased GSI beyond STZ alone in males only.^*P < 0.01 vs. control of the respective sex, ^#P < 0.05 vs male STZ. (B) Representative pictures of periodic acid Schiff-stained sections. Diabetic kidneys were characterized by moderate glomerulosclerosis, as evidenced by mesangial expansion (arrowheads), and is seen more abundantly in male STZ + GH than STZ alone. Original magnification is × 400, and bar represents 100 μm.

Figure 2

Effect of sex on TIFI in control and diabetic rats treated with and without GH. (A) STZ treatment significantly increases the TIFI in both sexes, and while GH treatment increased the TIFI in both sexes, the increase is greater in males than in females. ^*P < 0.01 vs. control of the respective sex, ^#P < 0.05 vs STZ of the respective sex, ^$P < 0.05 vs female GH. (B) Representative pictures of Masson’s trichrome-stained sections. Diabetic kidneys were characterized by moderate tubule interstitial fibrosis, evidenced by the presence of extracellular matrix deposits (blue staining) and tubular dilation, and these changes (blue staining and larger tubules) are seen more abundantly in STZ + GH-treated animals, and most abundantly in STZ + GH-treated males. Original magnification is × 400, and bar represents 100 μm.

Figure 3

Effects of STZ and GH treatment on renal MCP immunoreactivity. (A) STZ increases the MCP-1 staining in kidneys from diabetic male and female rats; however, GH treatment increased MCP-1 only in kidneys from diabetic male rats. ^*P < 0.01 vs control and ^#P < 0.05 vs female. (B) Representative images show an increase in brown staining on dilated and injured tubules (solid arrows show dilated tubules, and dashed arrows show brown staining). Original magnification is × 400, and bar represents 100 μm.

Figure 4

Effects of STZ and GH treatment on renal CD68 immunoreactivity. (A) No significant changes were observed in CD68 protein expression in kidneys from diabetic female rats. While CD68 staining was not increased in kidneys from STZ-only-treated rats, GH significantly increased CD68 staining in the glomeruli and tubules in diabetic male, but not female, rats. ^*P < 0.01 compared to all other data (B) Representative images of CD68+ cells in kidney sections (dark brown dots). The intensity of staining is significantly increased in the STZ + GH-treated males. Original magnification is × 400, and bar represents 100 μm.

Figure 5

Effect of sex on renal cortical TGF-β protein abundance in control and diabetic rats treated with and without GH. (A) STZ and STZ + GH treatment increased female cortical TGF-β protein levels from control equally. ^*P < 0.01 vs. female controls. (B) STZ treatment increased male TGF-B levels significantly, and STZ + GH significantly increased those levels. ^*P < 0.01 vs. male controls, ^#P < 0.05 vs. male STZ.

Figure 6

Effect of sex on renal cortical MMP-2 and MMP-9 protein abundance in control and diabetic rats treated with and without GH. (A) STZ and STZ + GH treatments significantly decrease MMP-2 levels in female mice compared to control. ^*P < 0.01 vs. female controls. (B) STZ significantly decreases MMP-2 protein levels in male mice compared to control, and STZ + GH treatment is significantly lower than STZ alone. ^*P < 0.01 vs. male control, ^#P < 0.05 vs. male STZ. (C) STZ and STZ + GH treatments significantly decrease female MMP-9 levels than in controls. ^*P < 0.01 vs. female controls. (D) STZ significantly decreases MMP-9 protein levels in male mice compared to control, and STZ + GH treatment is significantly lower than STZ alone. ^*P < 0.01 vs. male control, ^#P < 0.05 vs. male STZ.