Grapefruit juice ameliorates nephropathy in streptozotocin induced diabetes.

Abstract

Background Diabetic Nephropathy (DN) is the leading cause of end stage renal disease and mortality in diabetic patients. Over the years, medicinal plants have been used to manage diabetes and its complications. Metformin, the synthetic analogue of galegine is used as first line therapy for Diabetes Mellitus (DM) but its use is contraindicated in patients with kidney dysfunction. The management of DN currently is limited to the use of anti-hypertensive agents; ACE inhibitors and angiotensin receptor blockers. Grapefruit juice (GFJ) has shown potential as an anti-diabetogenic agent because of its ability to ameliorate hyperglycaemia and dyslipidemia but its effect on fluid and electrolyte disturbance is not known. This study was designed to investigate the effect of GFJ on renal dysfunction in streptozotocin (STZ) induced male wistar rats. Materials and Methods Male wistar rats weighing between 250-300g were divided into 7 groups (n=7) and kept in cages for the treatment period of 8 weeks. Laboratory conditions of 12 hour light/dark cycle, temperature 25±20C and humidity 50-55 % were maintained throughout the study period. Non-diabetic animals group 1 (Control) were treated orally with 1.0 ml /Kg BW of distilled water, while group 2 (ND-GFJ) were treated orally with 3.0 ml /kg BW of GFJ. The diabetic groups 3, 4, 5, 6 and 7 were starved overnight in preparation for the STZ injection. Fasting blood glucose concentration was obtained via tail prick before 45 mg or 60 mg of STZ was administered via a single injection in the peritoneal cavity. STZ was prepared by dissolving it in 0.2 ml of 0.1 M Citrate buffer at pH 4.5. Groups 3, 4 and 7 received 60.0 mg/ Kg BW of STZ while group 5 and 6 received 45 mg/kg BW of STZ. Three days following STZ induction, the diabetic state was confirmed by measuring fasting blood glucose and animals with glucose concentration greater than 6 mmol/L were included in the study. Group 4 (INS- D60) and group 5 (INS- D45) were additionally treated with 4.0 U/kg BW of insulin via subcutaneous injection (S.C) twice a day while Group 6 (GFJ-D45) and group 7 (GFJ-D60) were treated orally with 3.0ml/Kg BW of GFJ. Group 3 (D-60) were similarly treated with 1.0 ml /Kg BW of distilled water. Fasting blood glucose (FBG) and glucose tolerance tests (GTT) were done on days 1 and 58 respectively in all the treatment groups. Urine was collected for a period of 24 hours on day 59 and on the last day animals were sacrificed by halothane overdose. Blood samples were obtained via cardiac puncture; kidney tissues were removed and preserved in formalin while the liver was snap frozen with liquid nitrogen and stored in a freezer (-800C) before analysis. Results Reduced plasma insulin was accompanied by decrease in body weight and an increase in FBG accompanied by polyuria, polydipsia and glucose intolerance in the non-treated diabetic animals compared to the control. Fasting blood glucose was significantly (p<0.0001) increased in the diabetic groups and treatment with GFJ or insulin lowered FBG in groups (GFJ-D45, GFJ-D60 & INS-D60) compared to the diabetic control (D60). GFJ significantly (p=0.0034) improved glucose intolerance in diabetic animals (GFJ-D60 & GFJ-D45) when compared to diabetic control groups D-60 & D-45 respectively. Hepatic glycogen content was reduced in diabetic animals (P=0.024) and treatment with GFJ significantly (P=0.00016) increased the glycogen concentration. In the non-diabetic group (GFJ-ND) treatment with GFJ significantly (P=0.0013) increased the glycogen concentration when compared to the control group. In the diabetic animals, decreased GFR was accompanied by Na+ retention accompanied by low urinary K+ and Cl- concentration. Treatment with GFJ significantly (p<0.05) increased urinary Na+ and K+ and Cl- in the diabetic group (GFJ-D60) but did not increase urinary Cl- in non-diabetic group and consequently improved GFR in the diabetic group. Renal pathology showed structural changes in the glomerulus and treatment with GFJ had some reno-protective effect. Conclusion GFJ lowered the fasting blood glucose and improved glucose tolerance in the STZ-induced diabetic rats in a comparable manner to insulin treated diabetic rats. GFJ decreased Na+ retention and increased GFR in the diabetic animals. This study suggests that GFJ could ameliorate nephropathy associated with diabetes mellitus. These results are the first to show that GFJ has renoprotective effects in STZ-Induced diabetic rats.