17beta-Estradiol supplementation reduces tubulointerstitial fibrosis by increasing MMP activity in the diabetic kidney.

We previously reported that supplementation with 17beta-estradiol (E2) attenuates albuminuria, glomerulosclerosis, and tubulointerstitial fibrosis in diabetic nephropathy. The present study examined the mechanisms by which E2 regulates extracellular matrix (ECM) metabolism, a process that contributes to the development of glomerulosclerosis and tubulointerstitial fibrosis. The study was performed in female nondiabetic (ND), streptozotocin-induced diabetic (D), and diabetic with E2 supplementation (D+E2) Sprague-Dawley rats for 12 wk. Diabetes was associated with an increase in the renal expression of collagen alpha type IV [ND, 0.22 +/- 0.02; D, 0.99 +/- 0.09 relative optical density (ROD); P < 0.05] and fibronectin protein (ND, 0.36 +/- 0.08; D, 1.47 +/- 0.08 ROD; P < 0.05), as measured by Western blot analysis. E2 supplementation partially attenuated this increase in collagen alpha type IV (D+E2, 0.47 +/- 0.10 ROD) and fibronectin (D+E2, 0.71 +/- 0.16 ROD) protein expression associated with D. Diabetes was also associated with a decrease in the expression of matrix metalloproteinase (MMP) isoform MMP-2 (ND, 0.79 +/- 0.01; D, 0.62 +/- 0.06 ROD; P < 0.05) and MMP-9 protein (ND, 0.49 +/- 0.02; D, 0.33 +/- 0.03 ROD; P < 0.05). E2 supplementation restored MMP-2 and MMP-9 protein to levels similar or even greater than in the ND kidneys (MMP-2, 0.75 +/- 0.06; MMP-9, 0.73 +/- 0.01 ROD). The activities of MMP-2 (ND, 7.88 +/- 0.44; D, 5.60 +/- 0.54 ROD; P < 0.05) and MMP-9 (ND, 29.9 +/- 1.8; D, 12.9 +/- 2.3 ROD; P < 0.05), as measured by zymography, were also decreased with D. E2 supplementation restored MMP-2 and MMP-9 activity to levels similar to that in ND kidneys (MMP-2, 7.66 +/- 0.35; MMP-9, 21.4 +/- 2.9 ROD). We conclude that E2 supplementation is renoprotective by attenuating glomerulosclerosis and tubulointerstitial fibrosis by reducing ECM synthesis and increasing ECM degradation.

Mechanisms of antioxidant and pro-oxidant effects of alpha-lipoic acid in the diabetic and nondiabetic kidney.

BACKGROUND: alpha-Lipoic acid is a potent antioxidant that improves renal function in diabetes by lowering glycemia, however, the mechanisms by which alpha-lipoic acid exerts its antioxidant effects are not completely understood. METHODS: Metabolic parameters, renal function, and morphology, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity and subunit expression were analyzed in nondiabetic and streptozotocin-induced diabetic rats fed normal rat chow (control) with or without alpha-lipoic acid (30 mg/kg body weight) for 12 weeks. RESULTS: Blood glucose was increased with diabetes (nondiabetic + control 89 +/- 3 mg/dL and diabetic + control 336 +/- 28 mg/dL) and was similar with alpha-lipoic acid treatment (diabetic +alpha-lipoic acid 351 +/- 14 mg/dL). In contrast, alpha-lipoic acid attenuated albuminuria (nondiabetic + control 8.9 +/- 1.3 mg/day; diabetic + control 28.1 +/- 4.6 mg/day; and diabetic +alpha-lipoic acid 17.8 +/- 1.2 mg/day) associated with diabetes. Similarly, alpha-lipoic acid attenuated glomerulosclerosis (nondiabetic + control 0.22 +/- 0.01; diabetic + control 0.55 +/- 0.04; diabetic +alpha-lipoic acid 0.36 +/- 0.03), tubulointerstitial fibrosis (nondiabetic + control 0.42 +/- 0.18; diabetic + control 1.52 +/- 0.05; diabetic +alpha-lipoic acid 1.10 +/- 0.05), superoxide anion (O(.-) (2)) generation (nondiabetic +control 15.8 +/- 1.7; diabetic +control 87.1 +/- 3.5; diabetic +alpha-lipoic acid 25.5 +/- 3.3 RLU/mg protein), and urine 8-isoprostane (8-iso) excretion (nondiabetic + control 7.4 +/- 1.4; diabetic + control 26.0 +/- 4.5; diabetic +alpha-lipoic acid 19.6 +/- 5.6 ng/day) associated with diabetes. alpha-Lipoic acid also reduced kidney expression of NADPH oxidase subunits p22phox and p47phox. Surprisingly, alpha-lipoic acid appears to cause pro-oxidant effects in nondiabetic animals, resulting in increased albuminuria (nondiabetic +alpha-lipoic acid 14.2 +/- 1.2 mg/day), increase in plasma creatinine levels (nondiabetic + control 59 +/- 6; diabetic + control 68 +/- 6; nondiabetic +alpha-lipoic acid 86 +/- 9; diabetic +alpha-lipoic acid 69 +/- 7 mumol/L), exacerbated glomerulosclerosis and tubulointerstitial fibrosis, increased O(.-) (2) generation, up-regulated p22phox and p47phox expression and increased 8-iso excretion. CONCLUSION: We conclude that alpha-lipoic acid improves albuminuria and pathology in diabetes by reducing oxidative stress, while in healthy animals, alpha-lipoic acid may act as a pro-oxidant, contributing to renal dysfunction.

17beta-Estradiol replacement improves renal function and pathology associated with diabetic nephropathy.

The protective factor of female gender appears to be lost in diabetes; the incidence of diabetes and its complications, including diabetic nephropathy, are equal in women and men. This study examined the effects of estrogen deficiency by ovariectomy (OVX) and replacement with 17beta-estradiol (OVX+E2) on renal function and pathology in the nondiabetic (ND) and streptozotocin (STZ)-induced diabetic (D) rat kidneys for 12 wk. Diabetes was associated with an increase in urine albumin excretion (UAE; ND, 0.39 +/- 0.03; D, 5.9 +/- 0.8 mg/day; P < 0.001), decrease in creatinine clearance (CrCl; ND, 0.69 +/- 0.03; D, 0.43 +/- 0.09 mg x min(-1) x 100 g body wt(-1); P < 0.05), increase in the index of glomerulosclerosis [GSI; ND, 0.01 +/- 0.01; D, 0.15 +/- 0.04 arbitrary units (AU); P < 0.01], tubulointerstitial fibrosis (TIFI; ND, 0.04 +/- 0.04; D, 0.68 +/- 0.2 AU; P < 0.01), and transforming growth factor-beta (TGF-beta) protein expression (ND, 0.61 +/- 0.02; D, 1.25 +/- 0.07 AU; P < 0.01). In the D group, the severity of these changes was augmented with OVX (UAE, 8.1 +/- 0.6 mg/day; CrCl, 0.40 +/- 0.04 mg x min(-1) x 100 g body wt(-1); GSI, 0.29 +/- 0.04 AU; TIFI, 0.90 +/- 0.06 AU; TGF-beta, 1.26 +/- 0.10 AU), whereas E2 replacement attenuated these changes (UAE, 6.3 +/- 0.8 mg/day; CrCl, 0.66 +/- 0.03 mg x min(-1) x 100 g body wt(-1); GSI, 0.06 +/- 0.02 AU; TIFI, 0.36 +/- 0.08 AU; TGF-beta, 0.57 +/- 0.08 AU). We conclude that E2 deficiency increases the severity of renal disease in a diabetic animal model and that E2 replacement is renoprotective by attenuating the decline in renal function and pathology associated with diabetes.

Diabetic nephropathy is associated with decreased circulating estradiol levels and imbalance in the expression of renal estrogen receptors.

BACKGROUND: The incidence of cardiovascular and renal disease is lower in premenopausal women than in aged-matched men. However, in the setting of diabetes mellitus (DM), this "female advantage" no longer exists: the incidence and progression of DM and its associated end-organ complications are equal in men and women, regardless of age. We have recently reported that estrogen supplementation attenuates the progression of diabetic nephropathy, suggesting that lack of estrogen may nullify female sex as a protective factor against DM. OBJECTIVE: This study examined circulating levels of estradiol in DM and expression of estrogen receptor subtypes (ERa and ERP) in the nondiabetic (ND) and diabetic (D) kidney. METHODS: : The study was performed in ND and streptozotocin-induced D Sprague-Dawley rats after 2 weeks (male and female) and 12 weeks (female) of DM. The animals (N = 8/group) were kept either intact, ovariectomized (OVX), or OVX with 17beta-estradiol (E(2)) supplementation (OVX + E(2), 5 mug/kg/d). Plasma estradiol levels were measured by enzyme-linked immunosorbent assay, and expression of renal ERalpha and ERbeta was measured by immunohistochemistry and Western blot analysis. RESULTS: DM was associated with reduced circulating estradiol levels (ND: mean [SEM] 37.1 [7.2]; D: 24.5 [9.3] pg/mL; P < 0.05). The diabetic kidney exhibited increased expression of ERalpha protein (ND: 0.82 [0.06]; D: 1.15 [0.09] arbitrary units; P < 0.05), but no differences in ERP were observed. This resulted in an overall increase in the ratio of ERalpha/ERbeta protein expression in the diabetic kidney. No differences in the expression of ERa were observed in either females or males with similar glycemic levels after 2 weeks of DM. CONCLUSIONS: Reduced circulating levels of estradiol and imbalance in the expression of estrogen receptor subtypes in the diabetic kidney may explain why female sex is no longer a protective factor in the setting of DM. Thus, estradiol supplementation may be an effective regimen in attenuating the onset and progression of diabetic renal complications.