Investigating the effects of bredemolic acid on selected markers of some prediabetes-associated dysfunctions in diet-induced prediabetic rats.

Abstract

Prediabetes is an abnormal glycaemic state between normoglycaemia and chronic hyperglycaemia which is currently prevalent in developing and developed countries due to increased consumption of high caloric diet coupled with sedentary lifestyle. Prediabetes is associated with abnormal glucose metabolism. Additionally, the risk of developing prediabetes-associated complications such as non-alcoholic fatty liver disease (NAFLD), cardiovascular and renal diseases is not only present in overt diabetes mellitus but also in prediabetes. Management of prediabetes involves the combination of dietary and pharmacological interventions, however there is reported low compliance among patients as they tend to become overly dependent on the pharmacological interventions. Consequently, the pharmacological intervention efficacy is reduced as patients still progress to having overt diabetes. Therefore, managing prediabetes with anti-diabetic agents that will remain effective even in the absence of dietary intervention is considered necessary. Triterpenes have been found to have potential as anti-diabetic agents. Bredemolic acid (BA), a pentacyclic triterpene, has been reported to have increased biological activity relative to some other triterpenes. In this study, we sought to investigate the effects of BA on selected markers of some prediabetes-associated dysfunctions such as abnormal glucose homeostasis, hepatic, cardiovascular and renal dysfunctions in a prediabetic rat model in both the presence and absence of dietary intervention. Materials and Methods Thirty six (36) Sprague Dawley male rats that weighed 150 – 180g were divided into two groups: the non-prediabetic (n=6) and the prediabetic groups (n=30) which were fed a normal diet (ND) and high fat high carbohydrate (HFHC) diet respectively for 20 weeks to induce prediabetes. At 20th week, prediabetes was confirmed by assessment of fasting blood glucose (FBG) and oral glucose tolerance test (OGTT). The prediabetic rats were further sub-divided into five groups (n=6) and treated with either BA (80 mg/kg) or metformin (MET, 500 mg/kg) in the presence and absence of diet intervention for 12 weeks. Every 4 weeks of treatment, all the animals were placed in metabolic cages to determine caloric and fluid intake as well as urine output. Also, the body mass index (BMI), waist circumference (WC), blood pressure and heart rate were measured at every 4 weeks of treatment. After the 12 weeks of treatment, the animals were sacrificed, blood samples were collected into EDTA sample bottles and centrifuged to obtain plasma. Also, the skeletal muscle, liver, heart and kidney were collected, weighed, snapped frozen with liquid nitrogen and stored at -80°C before the biochemical analysis of selected markers of glucose homeostasis, hepatic, cardiovascular and renal functions. Results In the first study, the untreated diet-induced prediabetic rats had a significantly increased body weight, increased caloric intake, elevated glycated haemoglobin, increased ghrelin plasma concentration, decreased muscle glycogen concentration, insulin resistance and hyperinsulinaemia compared to the non-prediabetic rats. However, BA treatment with or without diet intervention ameliorated the body weight, caloric intake, glycated haemoglobin, muscle glycogen, glucose tolerance, plasma insulin and increased the expression of glucose transporter 4 (GLUT 4) in the skeletal muscle by comparison to the untreated prediabetic rats. Prediabetic induction in the second study resulted into elevated plasma concentration of liver enzymes, increased liver glycogen and triglyceride concentrations, increased oxidative stress in the liver and decreased sterol regulatory element binding protein (SREBP1c) by comparison to the non-prediabetic animals. Conversely, administration of BA with or without dietary intervention ameliorated liver functions by decreased oxidative stress, decreased liver enzymes, decreased liver glycogen and triglyceride as well as increased hepatic SREBP1c concentration in comparison to the untreated prediabetic animals. The results in the third study showed that the untreated prediabetic rats had a significantly increased body mass index (BMI), waist circumference (WC), blood pressure, heart rate, lipid profile, oxidative stress and inflammatory markers with significantly decreased endothelial nitric oxide synthase (eNOS) by comparison to the non-prediabetic control rats. On the other hand, the administration of BA with or without diet intervention improved cardiovascular functions by a decrease in BMI, WC, total cholesterol concentration, triglyceride concentration, blood pressure, heart rate, oxidative stress and inflammation with significant increase in eNOS plasma concentration in comparison to the untreated prediabetic rats. In the fourth study, the untreated prediabetic rats had a significantly increased fluid intake, urine output, sodium retention, potassium loss, aldosterone concentration, albuminuria, proteinuria, kidney injury molecule (KIM-1) and urinary podocin mRNA expression in comparison to non-prediabetic control and BA treated rats with or without diet intervention. Also, the untreated prediabetic rats presented increased albumin, total protein, urea, uric acid, creatinine and oxidative stress markers concentrations with a significant decrease in glomerular filtration rate (GFR). However, administration of BA with or without diet intervention attenuated oxidative stress, decreased urinary podocin mRNA expression and the aforementioned renal dysfunctions parameters. Conclusion This study showed that long term consumption of high caloric diet-induced prediabetes and resulted in abnormal glucose homeostasis, hepatic, cardiovascular and renal dysfunctions. Also, the results of this study showed that these dysfunctions are not only present during overt type 2 diabetes mellitus but

already present at the prediabetic stage due to insulin resistance or hyperinsulinaemia that triggered oxidative stress in the physiological systems that we examined in this study. However, due to amelioration of insulin resistance via improved insulin sensitivity and earlier reported antioxidant activities that are common to all pentacyclic triterpenes, administration of BA significantly ameliorated the prediabetes-associated dysfunctions (abnormal glucose homeostasis, hepatic, cardiovascular and renal dysfunctions) with or without diet intervention in the prediabetic stage.