Endocrine Research 97
Antiapoptotic and Antioxidant Effects of Carvedilol and Vitamin E Protect Against Diabetic Nephropathy and Cardiomyopathy in Diabetic Wistar Albino Rats
Authors
M. H. Abdel-Raheem1, S. U. Salim1, E. Mosad2, A. Al-Rifaay3, H. S. Salama1, H. Hasan-Ali4
Affiliations
1
Key words
Abstract
▼
Carvedilol is a novel β-adrenoreceptor blocker, with antioxidant properties inhibiting lipid peroxidation and preventing the depletion of endogenous antioxidants. Moreover, carvedilol was reported to enhance the expression of Bcl-2 gene, which has antioxidant and antiapoptotic effects. There are few researches testing the protective effect of carvedilol on the development of diabetic cardiomyopathy and nephropathy. In this study, we induced diabetes mellitus in male Wistar albino rats. We investigated carvedilol, as well as vitamin E, administrated in healthy and diabetic rats for 6 weeks to compare their effects on biochemical parameters and the expression of
Introduction received 12.03.2014 accepted 07.07.2014 Bibliography DOI http://dx.doi.org/ 10.1055/s-0034-1385855 Published online: August 8, 2014 Horm Metab Res 2015; 47: 97–106 © Georg Thieme Verlag KG Stuttgart · New York ISSN 0018-5043 Correspondence H. Hasan-Ali Cardiovascular Medicine Department Assiut University Hospitals 3-Nazih Khalifa St. 71111 Assiut Egypt Tel.: + 20/88/2356 753 Fax: + 20/88/2343 700 [email protected]
▼
Diabetes mellitus represents a serious risk factor for the development of cardiomyopathy and nephropathy [1]. Oxidative stress resulting from hyperglycemia has been implicated as a possible mechanism for such complications [2], where the circulating oxygen free radicals possibly mediate the process of apoptosis [3]. Carvedilol has been widely used in the treatment of heart failure, hypertension with or without diabetes [4]. It was found to inhibit the formation of reactive oxygen radicals and lipid peroxidation, and prevent the depletion of endogenous antioxidants [5, 6]. It has been suggested that carvedilol may provide greater benefit than traditional β-blockers in chronic heart failure because of its antioxidant actions that synergize with its non-specific β-and α-blocking effects [4, 6]. Whether carvedilol can prevent cardiomyopathy or nephropathy in diabetic animal models via antioxidant pathway is not well studied yet. Therefore, in the present study we investigated the protective effect of carvedilol on renal function, myocardial
Bcl-2 protein in both myocardial and renal tissues by immunohistochemistry. The study showed that the diabetic rats not only had renal dysfunction and more myocardial damage, but also showed lower expression of Bcl-2 protein. Carvedilol and vitamin E treatments were associated with better renal function and less myocardial damage, lower blood glucose, and lipid peroxidation, higher antioxidant capacity, better serum lipids, and higher expression of Bcl-2 protein in diabetic rats. These results indicate that carvedilol and vitamin E treatments partly protect against myocardial and renal damage probably via their antioxidant and antiapoptotic properties in diabetic rats.
damage, total antioxidant capacity, lipid profile, lipid peroxidation, histopathologic changes in myocardial and renal tissues, and Bcl-2 protein expression in the diabetic rats. These effects of carvedilol were compared with those of the traditional antioxidant vitamin E.
Materials and Methods
▼
This study was a case control study conducted on adult male Wistar albino rats weighting 180– 220 g aged 8 weeks obtained from the animal house in our institution. Rats were housed 4 per cage and maintained in a 12-h light/dark cycle in a temperature- and humidity-controlled facility. The room temperature was 19–24 °C and relative humidity was 45–60 %. Standard rat chow and acidified tap water (pH 2.5) were provided ad libitum. The experimental protocol flow chart is illustrated in ● ▶ Fig. 1. The rats were divided into the following groups each consisting of 10 rats. Group I: Control non-diabetic rats (saline treated rats); Group II: carvedilol-treated non-diabetic
Abdel-Raheem MH et al. Carvedilol and Vitamin E as Antiapoptotics … Horm Metab Res 2015; 47: 97–106
This document was downloaded for personal use only. Unauthorized distribution is strictly prohibited.
▶ thiobarbaturic acid ● ▶ Bcl-2 ● ▶ total antioxidant status ●
Department of Pharmacology, Faculty of Medicine, Assiut University, Assiut, Egypt Clinical Pathology Department (Immunohistochemistry Lab), South Egypt Cancer Institute, Assiut University, Assiut, Egypt 3 Pathology Department, Faculty of Medicine, Assiut University, Assiut, Egypt 4 Cardiovascular Medicine Department, Faculty of Medicine, Assiut University, Assiut, Egypt 2
Fig. 1 The experimental protocol flow chart. CPK: Creatinine phosphokinase; I.P.: Intraperitoneal; PEG: Polyethylene glycol; STZ: Streptozotocin; TAOS: Total antioxidant status; TBARS: Thiobarbituric acid reactive substances; TUNEL: Terminal deoxynucleotidyl transferase (TdT) dUTP Nick-End Labeling. (Color figure available online only).
rats [10 mg/kg intraperitoneal (I.P.) for 6 weeks]; Group III: vitamin E-treated non-diabetic rats (100 mg/kg orally for 6 weeks); Group IV: control diabetic rats [streptozotocin (STZ)-treated rats]; Group V: polyethylene glycol (PEG)-treated diabetic rats (a vehicle of carvedilol given orally for 6 weeks); Group VI: oiltreated diabetic rats (a vehicle of vitamin E given orally for 6 weeks); Group VII: diabetic rats treated with carvedilol [10 mg/ kg intraperitoneal (I.P.) for 6 weeks]; Group VIII: diabetic rats treated with Vitamin E (100 mg/kg orally for 6 weeks); Group IX: diabetic rats treated with carvedilol (10 mg/kg I.P.) and vitamin E (100 mg/kg orally) for 6 weeks. Diabetes was induced in adult male Wistar albino rats by single intraperitoneal (I.P.) injection of STZ at a dose of 65 mg/kg body weight. The control non-diabetic rats were injected with an equal volume of sterile 0.9 % saline solution. Blood glucose concentration was determined 72 h after STZ injection. The diabetic rats included in the study had blood glucose concentrations > 250 mg/dl [7]. Control diabetic rats were left untreated for 6 weeks for the development of diabetes complications. This 6 weeks period was reported by Cameron and associates to be an appropriate duration for the development of complications of diabetes [7]. The tested drugs were given to diabetic rats within 72 h after STZ injection for 6 weeks to evaluate their protective effects. These
drugs were given simultaneously to non-diabetic treated rats. At the end of the 6 weeks duration, the rats were killed. At the time of euthanasia, rats were anesthetized with sodium pentobarbitone (60 mg/kg body weight, I.P.), and then the cutting down of jugular vein was performed for blood sampling immediately before rats being killed by decapitation. Blood samples collected from neck veins were centrifuged and the serum from each sample was kept in Eppendorf tubes in the deep freezer at − 20 °C for measurement of blood glucose level [estimated by the enzymatic colorimetric method using commercially available standard glucose kits (human)], serum lipogram [estimated by automated chemistry analyzer (integra, Roche)], thiobarbaturic acid [8] as an index of endogenous lipid peroxidation, total antioxidant status (TAOS) [9], creatinine phosphokinase (CPK) as a marker of myocardial damage, serum urea, and serum creatinine were measured by spectrophotometer according to the manufacturer instructions (human kit). Heart and kidney tissues were isolated, formalin fixed, and Hematoxylin and Eosin (H & E) stained for histopathological examination. Bcl-2 protein expression by immunohistochemical method was carried out. Our institutional ethical committee has approved the study and animal care was in accordance with the National Institutes of Health “Principles of Laboratory Animal Care”.
Abdel-Raheem MH et al. Carvedilol and Vitamin E as Antiapoptotics … Horm Metab Res 2015; 47: 97–106
This document was downloaded for personal use only. Unauthorized distribution is strictly prohibited.
98 Endocrine Research
Endocrine Research 99
Immunohistochemical evaluation of Bcl-2 protein expression in renal and heart tissues
Briefly, sections mounted on glass slides were deparaffinized and rehydrated through graded alcohols with water. Endogenous peroxidase activity was blocked with 0.6 % H2O2. Sections were then immersed in the retrieval solution and subjected to heat-induced antigen retrieval. The slides, in plastic Coplin jars containing retrieval solution, were microwaved. Non-specific protein binding was blocked. Sections were then incubated with mouse monoclonal antibodies with dilution (1:50) for 30 min at room temperature (clone 124, IgG1, kappa for Bcl-2; Dako Corporation, CA, USA). A catalyzed signal amplification system (K1500; Dako) was used according to the manufacturer’s instructions. Positive controls: Specimens consisted of lymph nodes (reactive lymphoid hyperplasia). Negative controls: Additional sections, running in parallel but with omission of the primary antibody. Evaluation of Bcl-2 Protein: Expression of Bcl-2 protein was identified as a diffuse brown cytoplasmic staining (Bcl-2). In kidney tissue, cells were counted in serial sections;
the population of positive cells was expressed as the number of reactive cells per glomerular cross-section or myocytes.
Statistical analysis
Analyses of variance were used to evaluate the differences of the means. Post-hoc analysis was done using Dunnet’s t-test. (Statistical Program for Social Sciences, version 14 for windows, 2006, SPSS Inc., Chicago, Illinois, USA). The minimum level of significance was set at a p-value of 0.05.
Results
▼
Compared to non-diabetics, diabetic rats had a worse lipid profile (higher TC, TG, LDL-cholesterol, and a lower HDL-cholesterol), higher thiobarbaturic acid level, CPK level, serum urea, serum creatinine, and lower TAOS. Treatment with carvedilol or vitamin E but not the vehicles (PEG or oil) prevented these harmful changes. Treatment with both carvedilol and vitamin E was associated with even a more significant improvement than ▶ Table 1– 3), ( ▶ Fig. 2c–f, 3, 4a, b). with either alone ( ● ● Treatment of diabetic rats with vitamin E, singly or concurrently with carvedilol, resulted in a significant decrease in the blood glucose level as compared to diabetic rats treated with the vehicles (PEG or oil). However, blood glucose level remained higher ▶ Fig. 2a). In addition, treatment of diabetic than non-diabetics ( ● rats with either carvedilol or vitamin E alone or in combination showed a significant increase in body weight in diabetic treated rats. A greater effect on body weight was noted when using both treatments together than either alone. Again, treatment with ▶ Table 3, ● ▶ Fig. 2b). the vehicles (PEG or oil) had no effect ( ● ▶ Fig. 4c, 5, it was found that apopAs shown in ● ▶ Table 4 and ● totic cells were detected more significantly in diabetic rats than the control. Also, treatments with carvedilol or vitamin E singly or in combination, but not with the vehicles (PEG or oil), produced a significant decrease in the detection of apoptotic cells as well as increase in the expression of Bcl-2 protein in both kidney ▶ Fig. 4c, d, 6). and cardiac tissues ( ●
Table 1 Effect of different treatments on serum lipid profile in rats. Group
Treatment
Dose and Duration
TC (mg/dl)
TG (mg/dl)
HDL (mg/dl)
LDL (mg/dl)
Equal vol. single I.P. 10 mg/kg/day I.P. for 6 weeks 100 mg/kg/day orally for weeks 65 mg/kg single I.P. 65 mg/kg single i.p. Equal vol. I.P. for 6 weeks 65 mg/kg single I.P. Equal vol. orally for 6 weeks 65 mg/kg single i.p. 10 mg/ kg/day I.P. for 6 weeks 65 mg/kg single I.P. 100 mg/ kg/day for 6 weeks orally 65 mg/kg single i.p. 10 mg/ kg/day I.P. for 6 weeks 100 mg/kg orally for 6 weeks
122.5 ± 9.2 119.3 ± 8.1 121.4 ± 9.0 260.5 ± 11.2 * 259.8 ± 10.3 *
37.4 ± 2.1 36.2 ± 1.9 34.3 ± 1.2 26.2 ± 2.1 * 23.1 ± 1.1 *
33.9 ± 2.2 30.5 ± 2.4 36.5 ± 3.0 165.3 ± 6.3 * 167.3 ± 5.1 *
94.5 ± 8.8 95.4 ± 9.1 90.2 ± 9.2 245.0 ± 13.9 * 240.0 ± 10.2 *
265.3 ± 14.5 *
21.0 ± 1.1 *
169.6 ± 6.3 *
246.0 ± 14.2 *
161.1 ± 11.6 †
30.4 ± 1.3 †
71.2 ± 3.1 †
154.3 ± 10.2 †
167.5 ± 12.6 †
31.2 ± 1.5 †
72.7 ± 2.6 †
160.3 ± 11.3 †
151.3 ± 10.7 †‡
34.2 ± 1.0 †‡
64.4 ± 1.9 †‡
112.9 ± 9.0 †‡
I II III IV V
Non-diabetic (control) Non-diabetic (treated) Non-diabetic (treated) Diabetic (control) Diabetic (treated)
Saline Carvedilol Vitamin E STZ STZ PEG
VI
Diabetic (treated)
STZ Oil
VII
Diabetic (treated)
STZ Carvedilol
VIII
Diabetic (treated)
STZ Vitamin E
IX
Diabetic (treated)
STZ Carvedilol Vitamin E
Results represent the mean of 10 rats ± S.E. *p
Antiapoptotic and Antioxidant Effects of Carvedilol and Vitamin E Protect Against Diabetic Nephropathy and Cardiomyopathy in Diabetic Wistar Albino Rats
Authors
M. H. Abdel-Raheem1, S. U. Salim1, E. Mosad2, A. Al-Rifaay3, H. S. Salama1, H. Hasan-Ali4
Affiliations
1
Key words
Abstract
▼
Carvedilol is a novel β-adrenoreceptor blocker, with antioxidant properties inhibiting lipid peroxidation and preventing the depletion of endogenous antioxidants. Moreover, carvedilol was reported to enhance the expression of Bcl-2 gene, which has antioxidant and antiapoptotic effects. There are few researches testing the protective effect of carvedilol on the development of diabetic cardiomyopathy and nephropathy. In this study, we induced diabetes mellitus in male Wistar albino rats. We investigated carvedilol, as well as vitamin E, administrated in healthy and diabetic rats for 6 weeks to compare their effects on biochemical parameters and the expression of
Introduction received 12.03.2014 accepted 07.07.2014 Bibliography DOI http://dx.doi.org/ 10.1055/s-0034-1385855 Published online: August 8, 2014 Horm Metab Res 2015; 47: 97–106 © Georg Thieme Verlag KG Stuttgart · New York ISSN 0018-5043 Correspondence H. Hasan-Ali Cardiovascular Medicine Department Assiut University Hospitals 3-Nazih Khalifa St. 71111 Assiut Egypt Tel.: + 20/88/2356 753 Fax: + 20/88/2343 700 [email protected]
▼
Diabetes mellitus represents a serious risk factor for the development of cardiomyopathy and nephropathy [1]. Oxidative stress resulting from hyperglycemia has been implicated as a possible mechanism for such complications [2], where the circulating oxygen free radicals possibly mediate the process of apoptosis [3]. Carvedilol has been widely used in the treatment of heart failure, hypertension with or without diabetes [4]. It was found to inhibit the formation of reactive oxygen radicals and lipid peroxidation, and prevent the depletion of endogenous antioxidants [5, 6]. It has been suggested that carvedilol may provide greater benefit than traditional β-blockers in chronic heart failure because of its antioxidant actions that synergize with its non-specific β-and α-blocking effects [4, 6]. Whether carvedilol can prevent cardiomyopathy or nephropathy in diabetic animal models via antioxidant pathway is not well studied yet. Therefore, in the present study we investigated the protective effect of carvedilol on renal function, myocardial
Bcl-2 protein in both myocardial and renal tissues by immunohistochemistry. The study showed that the diabetic rats not only had renal dysfunction and more myocardial damage, but also showed lower expression of Bcl-2 protein. Carvedilol and vitamin E treatments were associated with better renal function and less myocardial damage, lower blood glucose, and lipid peroxidation, higher antioxidant capacity, better serum lipids, and higher expression of Bcl-2 protein in diabetic rats. These results indicate that carvedilol and vitamin E treatments partly protect against myocardial and renal damage probably via their antioxidant and antiapoptotic properties in diabetic rats.
damage, total antioxidant capacity, lipid profile, lipid peroxidation, histopathologic changes in myocardial and renal tissues, and Bcl-2 protein expression in the diabetic rats. These effects of carvedilol were compared with those of the traditional antioxidant vitamin E.
Materials and Methods
▼
This study was a case control study conducted on adult male Wistar albino rats weighting 180– 220 g aged 8 weeks obtained from the animal house in our institution. Rats were housed 4 per cage and maintained in a 12-h light/dark cycle in a temperature- and humidity-controlled facility. The room temperature was 19–24 °C and relative humidity was 45–60 %. Standard rat chow and acidified tap water (pH 2.5) were provided ad libitum. The experimental protocol flow chart is illustrated in ● ▶ Fig. 1. The rats were divided into the following groups each consisting of 10 rats. Group I: Control non-diabetic rats (saline treated rats); Group II: carvedilol-treated non-diabetic
Abdel-Raheem MH et al. Carvedilol and Vitamin E as Antiapoptotics … Horm Metab Res 2015; 47: 97–106
This document was downloaded for personal use only. Unauthorized distribution is strictly prohibited.
▶ thiobarbaturic acid ● ▶ Bcl-2 ● ▶ total antioxidant status ●
Department of Pharmacology, Faculty of Medicine, Assiut University, Assiut, Egypt Clinical Pathology Department (Immunohistochemistry Lab), South Egypt Cancer Institute, Assiut University, Assiut, Egypt 3 Pathology Department, Faculty of Medicine, Assiut University, Assiut, Egypt 4 Cardiovascular Medicine Department, Faculty of Medicine, Assiut University, Assiut, Egypt 2
Fig. 1 The experimental protocol flow chart. CPK: Creatinine phosphokinase; I.P.: Intraperitoneal; PEG: Polyethylene glycol; STZ: Streptozotocin; TAOS: Total antioxidant status; TBARS: Thiobarbituric acid reactive substances; TUNEL: Terminal deoxynucleotidyl transferase (TdT) dUTP Nick-End Labeling. (Color figure available online only).
rats [10 mg/kg intraperitoneal (I.P.) for 6 weeks]; Group III: vitamin E-treated non-diabetic rats (100 mg/kg orally for 6 weeks); Group IV: control diabetic rats [streptozotocin (STZ)-treated rats]; Group V: polyethylene glycol (PEG)-treated diabetic rats (a vehicle of carvedilol given orally for 6 weeks); Group VI: oiltreated diabetic rats (a vehicle of vitamin E given orally for 6 weeks); Group VII: diabetic rats treated with carvedilol [10 mg/ kg intraperitoneal (I.P.) for 6 weeks]; Group VIII: diabetic rats treated with Vitamin E (100 mg/kg orally for 6 weeks); Group IX: diabetic rats treated with carvedilol (10 mg/kg I.P.) and vitamin E (100 mg/kg orally) for 6 weeks. Diabetes was induced in adult male Wistar albino rats by single intraperitoneal (I.P.) injection of STZ at a dose of 65 mg/kg body weight. The control non-diabetic rats were injected with an equal volume of sterile 0.9 % saline solution. Blood glucose concentration was determined 72 h after STZ injection. The diabetic rats included in the study had blood glucose concentrations > 250 mg/dl [7]. Control diabetic rats were left untreated for 6 weeks for the development of diabetes complications. This 6 weeks period was reported by Cameron and associates to be an appropriate duration for the development of complications of diabetes [7]. The tested drugs were given to diabetic rats within 72 h after STZ injection for 6 weeks to evaluate their protective effects. These
drugs were given simultaneously to non-diabetic treated rats. At the end of the 6 weeks duration, the rats were killed. At the time of euthanasia, rats were anesthetized with sodium pentobarbitone (60 mg/kg body weight, I.P.), and then the cutting down of jugular vein was performed for blood sampling immediately before rats being killed by decapitation. Blood samples collected from neck veins were centrifuged and the serum from each sample was kept in Eppendorf tubes in the deep freezer at − 20 °C for measurement of blood glucose level [estimated by the enzymatic colorimetric method using commercially available standard glucose kits (human)], serum lipogram [estimated by automated chemistry analyzer (integra, Roche)], thiobarbaturic acid [8] as an index of endogenous lipid peroxidation, total antioxidant status (TAOS) [9], creatinine phosphokinase (CPK) as a marker of myocardial damage, serum urea, and serum creatinine were measured by spectrophotometer according to the manufacturer instructions (human kit). Heart and kidney tissues were isolated, formalin fixed, and Hematoxylin and Eosin (H & E) stained for histopathological examination. Bcl-2 protein expression by immunohistochemical method was carried out. Our institutional ethical committee has approved the study and animal care was in accordance with the National Institutes of Health “Principles of Laboratory Animal Care”.
Abdel-Raheem MH et al. Carvedilol and Vitamin E as Antiapoptotics … Horm Metab Res 2015; 47: 97–106
This document was downloaded for personal use only. Unauthorized distribution is strictly prohibited.
98 Endocrine Research
Endocrine Research 99
Immunohistochemical evaluation of Bcl-2 protein expression in renal and heart tissues
Briefly, sections mounted on glass slides were deparaffinized and rehydrated through graded alcohols with water. Endogenous peroxidase activity was blocked with 0.6 % H2O2. Sections were then immersed in the retrieval solution and subjected to heat-induced antigen retrieval. The slides, in plastic Coplin jars containing retrieval solution, were microwaved. Non-specific protein binding was blocked. Sections were then incubated with mouse monoclonal antibodies with dilution (1:50) for 30 min at room temperature (clone 124, IgG1, kappa for Bcl-2; Dako Corporation, CA, USA). A catalyzed signal amplification system (K1500; Dako) was used according to the manufacturer’s instructions. Positive controls: Specimens consisted of lymph nodes (reactive lymphoid hyperplasia). Negative controls: Additional sections, running in parallel but with omission of the primary antibody. Evaluation of Bcl-2 Protein: Expression of Bcl-2 protein was identified as a diffuse brown cytoplasmic staining (Bcl-2). In kidney tissue, cells were counted in serial sections;
the population of positive cells was expressed as the number of reactive cells per glomerular cross-section or myocytes.
Statistical analysis
Analyses of variance were used to evaluate the differences of the means. Post-hoc analysis was done using Dunnet’s t-test. (Statistical Program for Social Sciences, version 14 for windows, 2006, SPSS Inc., Chicago, Illinois, USA). The minimum level of significance was set at a p-value of 0.05.
Results
▼
Compared to non-diabetics, diabetic rats had a worse lipid profile (higher TC, TG, LDL-cholesterol, and a lower HDL-cholesterol), higher thiobarbaturic acid level, CPK level, serum urea, serum creatinine, and lower TAOS. Treatment with carvedilol or vitamin E but not the vehicles (PEG or oil) prevented these harmful changes. Treatment with both carvedilol and vitamin E was associated with even a more significant improvement than ▶ Table 1– 3), ( ▶ Fig. 2c–f, 3, 4a, b). with either alone ( ● ● Treatment of diabetic rats with vitamin E, singly or concurrently with carvedilol, resulted in a significant decrease in the blood glucose level as compared to diabetic rats treated with the vehicles (PEG or oil). However, blood glucose level remained higher ▶ Fig. 2a). In addition, treatment of diabetic than non-diabetics ( ● rats with either carvedilol or vitamin E alone or in combination showed a significant increase in body weight in diabetic treated rats. A greater effect on body weight was noted when using both treatments together than either alone. Again, treatment with ▶ Table 3, ● ▶ Fig. 2b). the vehicles (PEG or oil) had no effect ( ● ▶ Fig. 4c, 5, it was found that apopAs shown in ● ▶ Table 4 and ● totic cells were detected more significantly in diabetic rats than the control. Also, treatments with carvedilol or vitamin E singly or in combination, but not with the vehicles (PEG or oil), produced a significant decrease in the detection of apoptotic cells as well as increase in the expression of Bcl-2 protein in both kidney ▶ Fig. 4c, d, 6). and cardiac tissues ( ●
Table 1 Effect of different treatments on serum lipid profile in rats. Group
Treatment
Dose and Duration
TC (mg/dl)
TG (mg/dl)
HDL (mg/dl)
LDL (mg/dl)
Equal vol. single I.P. 10 mg/kg/day I.P. for 6 weeks 100 mg/kg/day orally for weeks 65 mg/kg single I.P. 65 mg/kg single i.p. Equal vol. I.P. for 6 weeks 65 mg/kg single I.P. Equal vol. orally for 6 weeks 65 mg/kg single i.p. 10 mg/ kg/day I.P. for 6 weeks 65 mg/kg single I.P. 100 mg/ kg/day for 6 weeks orally 65 mg/kg single i.p. 10 mg/ kg/day I.P. for 6 weeks 100 mg/kg orally for 6 weeks
122.5 ± 9.2 119.3 ± 8.1 121.4 ± 9.0 260.5 ± 11.2 * 259.8 ± 10.3 *
37.4 ± 2.1 36.2 ± 1.9 34.3 ± 1.2 26.2 ± 2.1 * 23.1 ± 1.1 *
33.9 ± 2.2 30.5 ± 2.4 36.5 ± 3.0 165.3 ± 6.3 * 167.3 ± 5.1 *
94.5 ± 8.8 95.4 ± 9.1 90.2 ± 9.2 245.0 ± 13.9 * 240.0 ± 10.2 *
265.3 ± 14.5 *
21.0 ± 1.1 *
169.6 ± 6.3 *
246.0 ± 14.2 *
161.1 ± 11.6 †
30.4 ± 1.3 †
71.2 ± 3.1 †
154.3 ± 10.2 †
167.5 ± 12.6 †
31.2 ± 1.5 †
72.7 ± 2.6 †
160.3 ± 11.3 †
151.3 ± 10.7 †‡
34.2 ± 1.0 †‡
64.4 ± 1.9 †‡
112.9 ± 9.0 †‡
I II III IV V
Non-diabetic (control) Non-diabetic (treated) Non-diabetic (treated) Diabetic (control) Diabetic (treated)
Saline Carvedilol Vitamin E STZ STZ PEG
VI
Diabetic (treated)
STZ Oil
VII
Diabetic (treated)
STZ Carvedilol
VIII
Diabetic (treated)
STZ Vitamin E
IX
Diabetic (treated)
STZ Carvedilol Vitamin E
Results represent the mean of 10 rats ± S.E. *p
