Korean J Urol.  2007 Feb;48(2):212-218. 10.4111/kju.2007.48.2.212.

Effects of Alpha-lipoic Acid on Nitric Oxide Synthase Expression and Ultrastructural Changes in the Bladder of Rats with Streptozotocin-induced Diabetes

• Kang DI
• Kim SH
• Lee SD
• Kwak HS
• Choi SH
• Kim DR
• Min KS

• KMID: 2139763
• DOI: http://doi.org/10.4111/kju.2007.48.2.212

Abstract

PURPOSE: To evaluate whether alpha-lipoic acid (ALA) is effective at restoring the levels of nitric oxide synthase (NOS) expression and preventing ultrastructural changes in the bladder of rats with streptozotocin- induced diabetes.
MATERIALS AND METHODS
Nine-week-old male Sprague-Dawley rats were used. The experimental groups included a control group (n=6), a diabetes group (n=6), and two groups of diabetic rats treated with intraperitoneal injections of ALA (n=12) at either 50 (ALA50) or 100mg/kg/day (ALA100) for 8 weeks after the induction of diabetes. Diabetic oxidative stress was determined based on evaluation of immunohistochemical staining for 8-hydroxy-2-deoxyguanosine (8-OHdG). The measurements of the levels of eNOS and nNOS expressions, as well as an assessment of the ultrastructural changes in detrusor smooth muscle cells were performed.
RESULTS
The highest expression of 8-OHdG was observed in the diabetes group; whereas, the 8-OHdG expression in the ALA-treated groups was similar to that in the control group. Both eNOS and nNOS were constitutively expressed in the control group. The expression levels of both eNOS and nNOS proteins were higher in the diabetes group, which had experienced increased oxidative stress, than in the ALA50 and ALA100 groups. Compared with the control group, the diabetes group exhibited severe degeneration of the detrusor muscle cells. In the rats treated with ALA, the detrusor muscle cells showed mild to moderate degeneration. The mean numbers of mitochondria per smooth muscle cell in the control, diabetes, ALA50 and ALA100 groups were 12.6+/-1.5, 5.1+/-0.7, 18.3+/-0.7 and 19.3+/-1.3, respectively (p<0.01). CONCLISIONS: Our data suggest that diabetes enhanced the levels of eNOS and nNOS expressions in the bladder, and ALA inhibited the expressions of eNOS and nNOS. ALA had a protective effect against the degeneration of intracellular micro-organelles produced by diabetic oxidative damage in detrusor muscle cells. This study suggests that early treatment with ALA can reduce the damage caused by diabetic oxidative stress.

Keyword

Antioxidants; Bladder; Diabetes mellitus; Oxidative stress

MeSH Terms

Animals
Antioxidants
Diabetes Mellitus
Humans
Injections, Intraperitoneal
Male
Mitochondria
Muscle Cells
Myocytes, Smooth Muscle
Nitric Oxide Synthase*
Nitric Oxide*
Oxidative Stress
Rats*
Rats, Sprague-Dawley
Thioctic Acid*
Urinary Bladder*
Antioxidants
Nitric Oxide
Nitric Oxide Synthase
Thioctic Acid

Figure

• 

  Fig. 1 The expressions of 8-hydroxy-2'-deoxyguanosine in the urothelium and muscle cells of the bladder. Data are given as the means and SEM for the percentage of cells showing 8-hydroxy 2'-deoxyguanosine immunoreactivity. *p<0.01, †p=0.078. ALA50: diabetic rats treated with alpha-lipoic acid at 50mg/kg/day, ALA100: diabetic rats treated with alpha-lipoic acid at 100mg/kg/day. ALA: alpha-lipoic acid.

• 

  Fig. 2 Immunostaining of 8-hydroxy-2'-deoxyguanosine (8-OHdG) in the bladder. The black arrows indicate the nuclei of the urothelium and smooth muscle cells expressing immunoreactive 8-OHdG. The arrowheads indicate negative findings for 8-OHdG. A: control, B: diabetes, C: ALA50, D: ALA100 groups (×200). ALA: alpha-lipoic acid.

• 

  Fig. 3 The expressions of eNOS and nNOS proteins. A: Western blot analyses of eNOS and nNOS protein expression levels, B, C: quantification of data, corrected for beta-actin, in each sample. *: p<0.05. eNOS: endothelial nitric oxide synthase, nNOS: neuronal nitric oxide synthase.

• 

  Fig. 4 Cylindrical profiles of longitudinally sectioned detrusor smooth muscle cells. The black arrows indicate dense bodies, and open arrows electron-dense bands. The asterisk indicates vacuolation. Many mitochondria are seen in the smooth muscle cells of the control (A), whereas few are seen in the diabetes group (B), ALA treatment restored the number of mitochondria in the smooth muscle cells (C, D). A: control, B: diabetes, C: ALA50, D: ALA100 groups (final magnification, ×4,000). ALA: alpha-lipoic acid.

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