Down-regulation of the Expression of the Obese Gene by an Antidiabetic Thiazolidinedione in Zucker Diabetic Fatty Rats and db/db Mice

Bei Zhang,Michael P Graziano,Thomas W Doebber,Mark D Leibowitz,Sylvia White-Carrington,Deborah M Szalkowski,Patricia J Hey,Margaret Wu,Catherine A Cullinan,Philip Bailey,Bettina Lollmann,Robert Frederich,Jeffrey S Flier,Catherine D Strader,Roy G Smith

doi:10.1074/jbc.271.16.9455

Abstract

Obese (ob) is a recently identified gene involved in the regulation of energy balance in the mouse. We report here that AD-5075, a potent thiazolidinedione which lowered plasma glucose and triglyceride in Zucker diabetic fatty (ZDF) rats and db/db mice, decreased the expression of the ob gene in these animal models of obesity and non-insulin-dependent diabetes mellitus. The level of adipose ob mRNA in ZDF rats was 3-fold greater than that detected in the Zucker lean littermates. Chronic treatment with AD-5075 elicited a 67 and 70% reduction of ob mRNA in ZDF and control lean rats, respectively. Furthermore, the amount of adipose ob mRNA in db/db mice was 7 times higher than that detected in lean littermates. Treatment of db/db mice with AD-5075 resulted in a 78% reduction of the level of ob mRNA with parallel changes in circulating level of the ob gene product, leptin. The reduction of the ob mRNA in the Zucker lean rats was accompanied by significantly greater food intake and weight gain. However, in ZDF rats and db/db mice, there was profound increase in body weight without hyperphagia. The results demonstrate that the expression of the ob gene is up-regulated in these two rodent models of diabetes compared to their lean counterparts and that such overexpression is attenuated by treatment with an agent that improves insulin sensitivity and glucose homeostasis in vivo.

Highlights

Obesity is a predisposing factor for insulin resistance and non-insulin-dependent diabetes mellitus (NIDDM)1 in both humans and animals [1]
Administration of recombinant mouse leptin resulted in a significant reduction of food intake and body weight as well as a normalization of metabolic status in ob/ob mice (4 – 6), consistent with the hypothesis that leptin is a key hormone in the control of energy intake and expenditure
When Zucker lean littermates were treated with AD-5075, a profound reduction of ob mRNA levels (3-fold) was observed (Fig. 1). These results demonstrate that expression of the ob gene was subject to regulation by the antidiabetic agent in a rodent model of NIDDM, and in the lean controls

Summary

EXPERIMENTAL PROCEDURES

Animals—Male C57BL/KsJ-db/db mice and lean littermates (ϩ/?) were obtained from Jackson Laboratories. A cDNA probe for ob was obtained by cloning the entire coding region of ob using polymerase chain reaction based on the published sequence [3]. Total white adipose RNA was isolated from Swiss-Webster mice and first strand cDNA synthesized. Using polymerase chain reaction the coding region of the ob cDNA was isolated as 2 overlapping fragments using the following primer sets (5Ј-CAGTGAGCCCCAAGAAGAGG-3Ј, 5Ј-TCCAGGTCATTGGCTATCTG-3Ј, and 5Ј-ATTCCTGGGCTTCAGGGGATTCTGAGTTTC-3Ј, 5Ј-GCGTGTACCCACGGAGGAAC-3Ј). The cDNA probe for mouse adipocyte fatty acid-binding protein (FABP) (aP2) was obtained from Dr David Bernlohr (University of Minnesota). The antibody was affinity purified using SulfolinkR antibody purification kit (Pierce) following the manufacturers instruction Western blots using this antibody detect a 16-kDa protein in the media of HEK-293 cells transfected with pCMV vector containing the coding region of the ob cDNA [8].

RESULTS

72 Ϯ 4 55 Ϯ 3*

DISCUSSION