The Effects of Lycopene and Insulin on Histological Changes and the Expression Level of Bcl-2 Family Genes in the Hippocampus of Streptozotocin-Induced Diabetic Rats

Abstract

The aim of this study was to evaluate the effects of antioxidants lycopene and insulin on histological changes and expression of Bcl-2 family genes in the hippocampus of streptozotocin-induced type 1 diabetic rats. Forty-eight Wistar rats were divided into six groups of control (C), control treated with lycopene (CL), diabetic (D), diabetic treated with insulin (DI), diabetic treated with lycopene (DL), and diabetic treated with insulin and lycopene (DIL). Diabetes was induced by an injection of streptozotocin (60 mg/kg, IP), lycopene (4 mg/kg/day) was given to the lycopene treated groups as gavages, and insulin (Sc, 1-2 U/kg/day) was injected to the groups treated with insulin. The number of hippocampus neurons undergoing cell death in group D had significant differences with groups C and DIL (p < 0.001). Furthermore, insulin and lycopene alone or together reduced the expression of Bax, but increased Bcl-2 and Bcl-x_L levels in DI, DL, and DIL rats, especially when compared to group D (p < 0.001). The ratios of Bax/Bcl-2 and Bax/Bcl-xL in DI, DL, and DIL rats were also reduced (p < 0.001). Our results indicate that treatment with insulin and/or lycopene contribute to the prevention of cell death by reducing the expression of proapoptotic genes and increasing the expression of antiapoptotic genes in the hippocampus.

Figure 1

The effects of insulin and lycopene on neural cell death of hippocampus. (a) Neuronal cell death was evaluated via hematoxylin-eosin staining at 40x magnification. C: the control group; CL: the control group treated with lycopene; D: untreated diabetic group; DI: diabetic group treated with insulin; DL: diabetic group treated with honey; DIL: diabetic group treated with insulin and honey simultaneously. (b) ^∗∗∗p < 0.001, comparing C, DI, and DIL with D. ^∗p < 0.05, comparing D with DL. (c) ^∗∗∗p < 0.001, comparing C, DI, DL, and DIL with D. (d) ^∗∗∗p < 0.001, comparing C, DI, and DIL with D. ^∗p < 0.01, comparing D with DL. (e) ^∗∗∗p < 0.001, comparing C, DI, and DIL with D. ^∗p < 0.05, comparing D with DL. Bar graphs indicate the mean ± SEM (N = 8).

Figure 2

Semiquantitative RT-PCR analysis of expressions of (a) Bcl-2 gene in the hippocampus of the studied rats. Amplification of the Bcl-2 gene (228 bp) compared with that of the GAPDH gene (461 bp). (b) The effects of insulin and lycopene on expression level of Bcl-2 gene at mRNA level in the hippocampus of the studied rats. †††p < 0.001, comparing (expression level of Bcl-2 gene) the untreated diabetic group (D) with the control group (C). ^∗p < 0.05, comparing (expression level of Bcl-2 gene) the untreated diabetic group (D) with the diabetic groups treated with lycopene (DL). ‡‡p < 0.01, comparing (expression level of Bcl-2 gene) the untreated diabetic group (D) with the diabetic group treated insulin (DI) and insulin and lycopene simultaneously (DIL). (c) Bcl-X_L gene in the hippocampus of the studied rats. Amplification of the Bcl-X_L gene (357 bp) compared with that of the GAPDH gene (461 bp). (d) The effects of insulin and lycopene on expression level of Bcl-X_L gene at mRNA level in the hippocampus of the studied rats. †††p < 0.001, comparing (expression level of Bcl-X_L gene) D with C, DI, and DIL. ^∗p < 0.05, comparing (expression level of Bcl-2 gene) D with DL. (e) Bax gene in the hippocampus of the studied rats. Amplification of the Bax gene (200 bp) compared with that of the GAPDH gene (461 bp). (f) The effects of insulin and lycopene on expression level of Bax gene at mRNA level in the hippocampus of the studied rats. †††p < 0.001, comparing (expression level of Bax gene) D with C, DI, and DIL. ^∗p < 0.05, comparing (expression level of Bax gene) D with DL. Bar graphs indicate the mean ± SEM (N = 8).

Figure 3

The effects of insulin or honey or both on (a) Bax/Bcl-2 ratio at mRNA level in the hippocampus of the studied rats. †††p < 0.001, comparing (expression level of Bax gene) the untreated diabetic group (D) with the control group (C) and the diabetic group treated with insulin (DI), lycopene (DL), and both together (DIL). (b) Bax/Bcl-X_L ratio at mRNA level in the hippocampus of the studied rats. ^∗∗∗p < 0.001, comparing (expression level of Bax gene) the untreated diabetic group (D) with the control group (C) and the diabetic group treated insulin (DI), lycopene (DL), and both together (DIL). Bar graphs indicate the mean ± SEM (N = 8).