Elsevier

Pharmacological Research

Volume 52, Issue 4, October 2005, Pages 313-320
Pharmacological Research

Combination of high-fat diet-fed and low-dose streptozotocin-treated rat: A model for type 2 diabetes and pharmacological screening

https://doi.org/10.1016/j.phrs.2005.05.004Get rights and content

Abstract

The objective of the present study was to develop a rat model that replicates the natural history and metabolic characteristics of human type 2 diabetes and is also suitable for pharmacological screening. Male Sprague–Dawley rats (160–180 g) were divided into two groups and fed with commercially available normal pellet diet (NPD) (12% calories as fat) or in-house prepared high-fat diet (HFD) (58% calories as fat), respectively, for a period of 2 weeks. The HFD-fed rats exhibited significant increase in body weight, basal plasma glucose (PGL), insulin (PI), triglycerides (PTG) and total cholesterol (PTC) levels as compared to NPD-fed control rats. Besides, the HFD rats showed significant reduction in glucose disappearance rate (K-value) on intravenous insulin glucose tolerance test (IVIGTT). Hyperinsulinemia together with reduced glucose disappearance rate (K-value) suggested that the feeding of HFD-induced insulin resistance in rats. After 2 weeks of dietary manipulation, a subset of the rats from both groups was injected intraperitoneally with low dose of streptozotocin (STZ) (35 mg kg−1). Insulin-resistant HFD-fed rats developed frank hyperglycemia upon STZ injection that, however, caused only mild elevation in PGL in NPD-fed rats. Though there was significant reduction in PI level after STZ injection in HFD rats, the reduction observed was only to a level that was comparable with NPD-fed control rats. In addition, the levels of PTG and PTC were further accentuated after STZ treatment in HFD-fed rats. In contrast, STZ (35 mg kg−1, i.p.) failed to significantly alter PI, PTG and PTC levels in NPD-fed rats. Thus, these fat-fed/STZ-treated rats simulate natural disease progression and metabolic characteristics typical of individuals at increased risk of developing type 2 diabetes because of insulin resistance and obesity. Further, the fat-fed/STZ-treated rats were found to be sensitive for glucose lowering effects of insulin sensitizing (pioglitazone) as well as insulinotropic (glipizide) agents. Besides, the effect of pioglitazone and glipizide on the plasma lipid parameters (PTG and PTC) was shown in these diabetic rats. The present study represents that the combination of HFD-fed and low-dose STZ-treated rat serves as an alternative animal model for type 2 diabetes simulating the human syndrome that is also suitable for testing anti-diabetic agents for the treatment of type 2 diabetes.

Introduction

Type 2 diabetes mellitus is a heterogeneous disorder characterized by a progressive decline in insulin action (insulin resistance), followed by the inability of beta cells to compensate for insulin resistance (pancreatic beta cell dysfunction). Insulin resistance is a characteristic metabolic defect that precedes overt beta cell dysfunction and is primarily associated with resistance to insulin-mediated glucose disposal at the periphery and compensatory hyperinsulinemia. The beta cells normally compensate insulin resistance by secreting more amounts of insulin to maintain the glucose homeostasis. In the course of time, however, this beta cell function gets impaired leading to deterioration in glucose homeostasis and subsequent development of impaired glucose tolerance and frank diabetes [1], [2]. There occurs only a relative insulin deficiency as the day-long circulating insulin concentrations in patients with type 2 diabetes are almost comparable or slightly elevated in absolute terms to the values in normal individuals. Despite the role of genetic predisposition, aging, obesity and dietetic/sedentary life style are major risk factors involved in the development of type 2 diabetes. Most of the individuals diagnosed with type 2 diabetes are found to be obese [3], [4].

Although there exists a surplus of animal models (spontaneous as well as induced) available for the study of type 2 diabetes, the pattern of disease initiation and development in most of them do not appear to be closely analogous to the clinical situation in humans. However, there are certain genetic models namely Zucker diabetic fatty (ZDF) rat and db/db mouse which develop diabetes spontaneously resembling human type 2 diabetes, the development of diabetes in them is predominantly genetically determined unlike in humans [5], [6]. Moreover, the observations derived from these highly inbred genetic strains may not always be satisfactorily extended to the human population as a whole because of the large heterogeneity in the latter. In addition, these animals are expensive and are not easily available for the investigative purposes as well as regular screening experiments. Further, in induced diabetic models, most of the animals (adult or neonates) requires relatively high dose of streptozotocin (STZ; >50 mg kg−1) [6]. The development of hyperglycemia in these rats following STZ injection is primarily due to the direct pancreatic beta cell destruction, and resulting insulin deficiency rather than the consequence of insulin resistance [6], [7]. Thus, they depict symptoms and characteristics typical more of human type 1 than type 2 diabetes and further are not very responsive to the effects of drugs like insulinotropic (e.g. glipizide, tolbutamide) and insulin-sensitizing compounds (e.g. pioglitazone, rosiglitazone) [8], [9], [10]. In contrast, the rats fed with high-fat diet (HFD) develop obesity, hyperinsulinemia, and insulin resistance and not frank hyperglycemia or diabetes, thus limiting the screening of agents on controlling the blood glucose level [11], [12]. Hence, there exists a continued quest among the investigators for establishing the ideal animal model for type 2 diabetes either by way of modification of the existing methods or by developing new methodologies or a combination of both [6], [13].

Thus, we initiated this study with the objective of developing a suitable type 2 diabetic rat model that would on the one hand closely mimic the natural history of the disease events (from insulin resistance to beta cell dysfunction) as well as metabolic features of human type 2 diabetes and on the other hand would be cheaper, easily available and useful for the investigation as well as preclinical testing of various compounds viz. insulin sensitizers and insulinotropics for the treatment of type 2 diabetes. The materialization of the disease pattern was achieved by combining the feeding of HFD which produced insulin resistance and low dose of STZ treatment that caused the initial beta cell dysfunction and subsequently the frank hyperglycemia in non-genetic, out-bred Sprague–Dawley rats.

Though attempts have been made previously by the other investigators for developing suitable animal models for type 2 diabetes by injecting STZ into genetically insulin-resistant animals (spontaneous hypertensive rats) or by a combination of HFD and STZ treatment in normal rats and mice [13], [14], [15], [16], the rat model described in this paper is unique in so far as the approach adopted towards the development of the model as well as its suitability for pharmacological screening is concerned.

Section snippets

Materials

STZ was purchased from Calbiochem, Germany. The feed ingredients such as casein and cholesterol (both from Hi-media laboratories, Mumbai, India), dl-methionine (Loba Chemie, Mumbai, India), vitamin and mineral mix (Sarabhai chemicals, Baroda, India) and yeast powder (Pet Care, Bangalore, India) were procured from the commercial sources. Lard, insulin (Eli Lilly, Gurgaon, India), heparin (S.D. Fine-Chem Ltd., Mumbai) and sodium carboxy methyl cellulose (Na-CMC) (Loba Chemie, Mumbai) were also

Features of HFD-fed insulin-resistant rats

Table 2 illustrates that the feeding of HFD for 2 weeks resulted in significant increase (p < 0.05) in body weight as well as non-fasting PGL, PI, PTG and PTC levels in rats as compared to NPD-fed rats. On IVIGTT, HFD rats exhibited significant reduction in glucose disappearance rate (K-value, representing the level of insulin sensitivity) as compared to NPD-fed control rats (Table 2).

Effect of STZ on NPD-fed and HFD-fed insulin-resistant rats

Injection of STZ (35 mg kg−1, i.p.) after 2 weeks of dietary manipulation significantly (p < 0.05) increased PGL in

Discussion

This study was initiated with the objective of developing an ideal model for type 2 diabetes that would closely reflect the natural history and metabolic characteristics of human type 2 diabetes and respond to the pharmacological treatments. Further, it should be less expensive, easily available, taking relatively shorter period for development and adequate enough to allow for invasive procedures to the investigators.

Thus, our initial attempts were directed towards finding the threshold dose of

Conclusions

Our study demonstrates that a combination of HFD and low dose of STZ treatment can be effectively used to generate a rat model that mimic the natural history and metabolic characteristics of the common type 2 diabetes in humans. It is cheap, easy to develop and most suited for studying the pathophysiology of type 2 diabetes and is also useful in evaluating the therapeutic compounds for the treatment of type 2 diabetes.

Acknowledgement

The authors are immensely thankful to Prof. K.P.R. Kartha, Department of Medicinal Chemistry, NIPER, for proof-reading this article.

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