Gender-Related Difference in Relationship between Insulin Resistance and Serum Leptin Level in Japanese Type 2 Diabetic and Non-Diabetic Subjects

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1 Endocrine Journal 2000, 47 (5), Gender-Related Difference in Relationship between Insulin Resistance and Serum Leptin Level in Japanese Type 2 Diabetic and Non-Diabetic Subjects AYAxo FUSAxo HATTORI, IWATA**, KAZUMAsA UEMURA*, HISAYUKI MIURA, MUNETO USDA, NORIKA TAMAYA, MASAHIRO MURAGUCHI**, YASUIcm OHMOTO** AND AKIHISA IGUCHI Department of Geriatrics, Medicine in Growth and Aging, Program in Health and Community Medicine, Nagoya University Graduate School of Medicine,Nagoya , Japan * Third Department of Internal Medicine, Nagoya University School of Medicine, Nagoya , Japan ** Otsuka Pharmaceutical, Co., Ltd., Tokushima , Japan Abstract. It remains controversial whether or not a correlation exists between serum leptin levels and insulin resistance, and, if such a correlation does exist, whether it is independent of adiposity. To investigate the possible existence of an independent correlation, we have assessed serum leptin levels and insulin resistance in Japanese diabetic and non-diabetic subjects by means of Homeostatic Model Assessment (HOMA-R). Sixty-four Japanese patients with Type 2 diabetes mellitus (DM) (33 men and 31 women) and 53 sex-, age-, and body mass index (BMI)-matched non-diabetic adults (29 men and 24 women) were enrolled. The fasting plasma level of glucose (FPG) and the fasting serum levels of immunoreactive insulin (FIRI) and leptin were determined. Multiple linear regression analysis demonstrated that, in both male diabetic and male non-diabetic subjects, HOMA-R and BMI were independently correlated with serum leptin levels. In females, BMI, but not HOMA-R, was correlated to the serum levels of leptin in both groups. There was no statistically significant difference in the partial regression coefficients between male diabetic and male non-diabetic subjects. These results suggest that the correlation of HOMA-R to the serum levels of leptin in females is dependent on BMI. In males, the relationship between serum leptin levels and the insulin resistance was not affected by the extent of glucose intolerance. Key words: Leptin, Insulin resistance, Type 2 diabetes mellitus, Gender-related difference (Endocrine Journal 47: , 2000) LEPTIN is a hormone secreted by the adipocytes that regulates body adiposity by affecting the central nervous system (CNS) via reduction of food intake and increase of energy expenditure [1, 2]. In humans, the serum level of leptin correlates with body mass index (BMI) or body fat mass [3-6], and hyperleptinemia in obesity suggests leptin resistance in the CNS. Received: April 14, 2000 Accepted: August 7, 2000 Correspondence to: Ayako HATTORI, Department of Geriatrics, Medicine in Growth and Aging, Program in Health and Community Medicine, Nagoya University Graduate School of Medicine, 65 Tsuruma-cho, Showa-ku, Nagoya , Japan Obesity is also well known to be closely associated with insulin resistance or hyperinsulinemia, which may cause Type 2 diabetes mellitus (DM) [7-9]. The coexistence of hyperleptinemia and hyperinsulinemia in obesity suggests a possible relationship between them. In fact, several studies have demonstrated a significant positive correlation between plasma leptin and fasting insulin levels that is independent of body adiposity [10, 11]. Segal et al. reported that insulin resistance was related to an increase in the plasma level of leptin independently of the body fat mass, and showed that the plasma level of leptin in insulin-resistant men was higher than that in BMImatched insulin-sensitive men [12]. However, it remains controversial whether or not a

2 616 HATTORI et al. correlation exists between serum leptin levels and insulin resistance, and, if such a correlation does exist, whether it is independent of adiposity. Nyholm et al. assessed the peripheral insulin sensitivity of firstdegree relatives of Type 2 DM patients or non-diabetic subjects using the hyperinsulinemic euglycemic clamp method and reported that the correlation was independent of body adiposity [13], while Mohamed- Ali et al. quantified the insulin sensitivity of Type 2 DM patients using the same technique and demonstrated that the correlation was not significant after controlling for the BMI and gender of patients [14]. Another report used the minimal model technique to assess the insulin sensitivity in non-diabetic subjects and Type 2 DM patients, and concluded that insulin sensitivity contributed to the association between body adiposity and plasma levels of insulin, but not leptin [15]. The HOMA-R index, which is derived from Homeostatic Model Assessment (HOMA), is an index of whole body insulin sensitivity calculated using blood levels of fasting insulin and glucose, and its results have been shown to be well correlated with those of the glucose clamp technique [16]. The advantages of HOMA-R as an index of insulin sensitivity are: 1) the method is convenient because it requires only a blood sample: 2) the method is less invasive: and 3) the method is much less expensive than the insulin suppression test (1ST) or clamp methods. Therefore, HOMA-R is a suitable tool for assessing the whole body human insulin sensitivity of a large number of subjects. Some studies using HOMA-R have reported an independent correlation between serum leptin levels and peripheral insulin resistance [17, 18]. One of the reasons for the discrepant results above may be the variability of insulin resistance among subjects, since some of these studies assessed insulin resistance in only non-diabetic subjects [13, 19] or diabetic subjects [14]. A racial difference may also be operative, since most Japanese Type 2 DM patients are not obese and deficient in insulin secretion like their Caucasian counterparts. In this study, we assessed the serum leptin levels and HOMA-R values of Japanese diabetic and non-diabetic subjects in order to clarify whether or not there is an independent correlation between the serum leptin levels and insulin resistance. Subjects All subjects provided their informed consent in accordance with the guidelines of the ethical committee of the Nagoya University School of Medicine. Diabetic subjects Sixty-four Japanese patients with Type 2 DM (33 men and 31 women; mean age, 61.3 years; range, years) were enrolled. All patients had been admitted to the hospital of the Nagoya University of Medical School to control their blood glucose or had attended a diabetes clinic at a general hospital between January 1997 and November The diagnosis of diabetes was based on World Health Organization criteria [20]. They had been treated by dietary modification alone or by dietary modification plus oral administration of sulfonylurea. Patients who were treated with insulin, who had renal (serum levels of creatinine >_ 1.2 mg/dl) or hepatic dysfunction (abnormal levels of serum AST or ALT), or who had fasting serum levels of insulin lower than 2 au/ml were excluded. Non-diabetic subjects Fifty-three sex-, age-, and BMI-matched nondiabetic adults (29 men and 24 women, mean age 57.6 years) who underwent a medical check-up in a company-based clinic were recruited as controls. They had normal fasting plasma levels of glucose ( mmol/1) and fructosamine ( mmol/1). Blood sampling Methods Blood samples were taken from diabetic subjects in the hospital before breakfast (0730 h) on the morning after admission, from diabetic subjects who attended the diabetes clinic at h, and from non-diabetic subjects at 0800 h on the morning they visited the clinic. All subjects were instructed to fast overnight before the blood sampling. The fasting plasma level of glucose (FPG) and the fasting serum levels of immunoreactive insulin (FIRI) and leptin were determined. HOMA-R was calculated as [16]:

3 INSULIN RESISTANCE AND LEPTIN IN DIABETICS 617 Assays HOMA-R =FIRI x FPG/22.5. FPG was determined enzymatically using the glucose oxidase method. FIRI was assayed by the enzyme immunoassay method (TOSOH, Yamaguchi, Japan). The inter- and intra-assay coefficients of variation were % and %, respectively. Serum leptin levels were measured by a solid phase sandwich enzyme immunoassay. The minimal detectable leptin concentration was 20 pg/ml. The inter- and intra-assay coefficients of variation were 3.9% and 7.2%, respectively. Statistical analysis Results are expressed as means ± SE. Differences between two groups were tested by the unpaired Student's t-test. Regression analysis was performed using the serum level of leptin as a dependent variable and the HOMA-R, BMI, FIRI, or FPG value as an independent variable. Multiple linear regression analysis was used to assess the independency of the correlation of the fasting serum level of leptin to insulin resistance. Logarithmic transformation of the serum levels of leptin, FPG, FIRI and HOMA-R was performed. Regression slopes of male diabetic subjects were compared to those of male non-diabetic subjects, using the Student's t-test, with the serum level of leptin as a dependent variable and HOMA-R as an independent variable. P-values less than 0.05 were considered to be statistically significant. Statistical analyses were performed with the Statview J-4.5 system (Abacus Concepts, Berkeley, CA). Results In the genders, FPG, HOMA-R and the serum levels of leptin were significantly higher in diabetic subjects than in non-diabetic subjects, while there was no significant difference in FIRI between diabetic and non-diabetic subjects. The serum levels of leptin were significantly higher in females than in males in both the diabetic and non-diabetic groups (Table 1). The serum levels of leptin were positively correlated with BMI, FIRI, or HOMA-R in simple linear regression analyses in all groups (Table 2). FPG was positively correlated with the serum levels of leptin only in non-diabetic females. Multiple linear regression analysis demonstrated that, in both male diabetic and male non-diabetic subjects, HOMA-R and BMI, were independently correlated with serum leptin levels. In females, BMI, but not HOMA-R, was correlated to the serum Table 1. Clinical characteristics

4 618 HATTORI et al. Table 2a. Simple linear regression analyses using log transformed serum leptin as the dependent variable in non-diabetic subjects. Table 2b. Simple linear regression analyses using log transformed serum leptin as the dependent variable in diabetic subjects. Table 3a. Multiple linear regression analyses using subjects. log transformed serum leptin as the dependent variable in non-diabetic Table 3b. Multiple linear regression analyses using log transformed serum leptin as the dependent variable in diabetic subjects.

5 INSULIN RESISTANCE AND LEPTIN IN DIABETICS 619 levels of leptin in both groups (Table 3). There was no statistically significant difference in the partial regression coefficients between male diabetic and male non-diabetic subjects. (f3=0.587 vs , respectively, P>0.10). Discussion There have been conflicting reports regarding the relationship between circulating leptin levels and insulin resistance in human. The disparity of results is thought to be at least partially attributable to gender difference. Although some reports have demonstrated that the correlation between serum leptin levels and insulin resistance is significant in both genders [17], Kennedy et al. [21] and Vettor et al. [22] reported that serum leptin levels were correlated with insulin resistance independently of BMI in males, but not in females. In this study, we demonstrated that serum leptin levels were significantly correlated with insulin resistance independently of BMI in both non-diabetic and diabetic males. But in females, only BMI was significantly correlated with the serum levels of leptin. These results suggest that the correlation of HOMA-R to the serum levels of leptin in females is dependent on BMI, since HOMA-R was significantly correlated with BMI (data not shown). Insulin is one of the important regulators to stimulate leptin release from adipose tissue in rat [23]. The hypothesis is supported in human by demonstrating the hyperleptinemia in patients with insulinoma [24]. Therefore, in the present study the increased level of serum leptin was a result of chronic hyperinsulinemia induced by obesity. Overexpression of leptin was reported to increase glucose metabolism and insulin sensitivity in transgenic skinny mice [25]. Coexistence of hyperleptinemia and insulin resistance assessed by HOMA in the present study suggested that the leptin resistance in the central nervous system that regulates glucose metabolism is responsible for the insulin resistance in peripheral tissues. Although it has been well established that there is a gender difference in circulating leptin levels [3, 4], the mechanism of this difference remains unknown. Kennedy et al. reported that the serum leptin levels rose more rapidly as a function of BMI in female than in male subjects, suggesting a physiological effect of sex hormones to modulates the leptin concentrations at the level of effect of systems activated to control body weight as the causes of the difference by gender [20]. Vettor et al. [22] and Hislop et al. [26] reported that, in male subjects, leptin levels were negatively correlated with testosterone levels. The relatively greater contribution of adiposity in determining the serum leptin levels in females in the present study may be attributable to gender-based difference. The above-described discrepancies in the reports on the relationship between circulating leptin levels and insulin resistance may also related to the method used to estimate insulin resistance. To assess the whole body insulin sensitivity, Kennedy et al. used the hyperinsulinemic euglycemic clamp method [21], Vettor et al. the insulin tolerance test (ITT) [22], Carantoni et al. steady-state plasma glucose (SSPG) by insulin suppression test [19], and Schwartz et al. [15] and Dua et al. [27] Bergman's minimal model. The results of the former two reports supported the hypothesis that serum leptin levels are significantly correlated with insulin sensitivity independently of BMI or percent body fat in humans. However, the latter three reports indicated that the correlation between circulating leptin levels and insulin sensitivity was dependent on body fat mass. In this study, we used HOMA-R as an index of insulin resistance. In principle, HOMA-R is calculated in the absence of exogenous substances that could influence the blood levels of insulin or glucose. Some of our subjects were taking sulfonylurea, which might have reduced the reliability of HOMA-R as an index of insulin resistance. However, Nagasaka et al. recently reported the validity of an insulin sensitivity enhancer, troglitazone, for use in HOMA-R assessment of insulin sensitivity in Japanese patients with Type 2 DM. Some of the subjects of their study were administered sulfonylurea, which suggested that HOMA-R could be a suitable index of insulin resistance even in those taking sulfonylurea [28]. Ono et al. [29] reported that insulin resistance measured by HOMA did not correlate with euglycemic clamping in obese diabetic subjects with high FPG values. However, the BMI of our subjects were 24.1 ± 0.5 in male and 25.3 ± 0.8 in female diabetic subjects. Further, when patients who had fasting plasma glucose levels higher than 11.1 mmol/l were (9 males and 4 females) excluded, no sig-

6 620 HATTORI et al. nificantly different results were obtained (data not shown). In male subjects of the present study, multiple linear regression analysis revealed no statistically significant difference in the slope of the regression of HOMA-R for the serum level of leptin between diabetic and non-diabetic subjects. de Courten et al. [17], Schwartz et al. [15] and Kennedy et al. [20] reported that the relationship between serum levels of leptin and serum levels of insulin or insulin resistance was not affected by the extent of glucose intolerance, and that there was no significant difference in correlation between the Type 2 DM patients and normal control subjects, which is consistent with our results. In conclusion, we evaluated the relationship between serum leptin levels and insulin resistance in Japanese diabetic and non-diabetic subjects using HOMA-R as an index of insulin sensitivity. Our primary results were as follows. 1) In male subjects, both HOMA-R and BMI were independently correlated with serum leptin levels irrespective of glucose tolerance, while in female subjects, only BMI was so correlated. 2) The relationship between the serum leptin levels and the insulin resistance in male subjects was not affected by the degree of glucose intolerance. References Pelleymounter MA, Cullen MJ, Baker MB, Hecht R, Winters D, Boone T, Collins F (1995) Effects of the obese gene product on body weight regulation in ob/ob mice. Science 269: Halaas JL, Gajiwala KS, Maffei M, Cohen SL, Chait BT, Rabinowitz D, Lallone RL, Burley SK, Friedman JM (1995) Weight-reducing effects of the plasma protein encoded by the obese gene. Science 269: Rosenbaum M, Nicolson M, Hirsch J, Heymsfield SB, Gallagher D, Chu F, Leibel R (1996) Effects of gender, body composition, and menopause on plasma concentrations of leptin. J Clin Endocrinol Metab 81: Hickey M, Israel RG, Gardiner SN, Considine RV, McCammon MR, Tyndall GL, Houmard JA, Marks RH, Caro JF (1996) Gender differences in serum leptin levels in humans. Biochem Mol Med 59: 1-6. Ma Z, Gingerich RL, Santiago JV, Klein S, Smith CH, Landt M (1996) Radioimmunoassay of leptin in human plasma. Clin Chem 42: Haffner SM, Stern MP, Miettinen H, Wei M, Gingerich RL (1996) Leptin concentrations in diabetic and nondiabetic Mexican-Americans. Diabetes 45: Lillioja 5, Bogardus C (1988) Obesity and insulin resistance: lessons learned from the Pima Indians. Diabetes Metab Rev 4: Reaven GM (1988) Role of insulin resistance in human disease. Diabetes 37: Lillioja S, Mott DM, Spraul M, Ferraro R, Foley JE, Ravussin E, Knowler WC, Bennett PH, Bogardus C (1993) Insulin resistance and insulin secretory dysfunction as precursors of non-insulin-dependent diabetes mellitus. N Engl J Med 329: Larsson H, Elmstahl 5, Ahren B (1996) Plasma leptin levels correlate to islet function independently of body fat in postmenopausal women. Diabetes 45: Malmstrom R, Taskinen MR, Karonen SL, Yki- Jarvinen H (1996) Insulin increases plasma leptin concentrations in normal subjects and patients with NIDDM. Diabetologia 39: Segal KR, Landt M, Klein S (1996) Relationship between insulin sensitivity and plasma leptin concentration in lean and obese men. Diabetes 45: Nyholm B, Fisker S, Lund S, Moller N, Schmitz 0 (1997) Increased circulating leptin concentrations in insulin-resistant first-degree relatives of patients with non-insulin-dependent diabetes mellitus: relationship to body composition and insulin sensitivity but not to family history of non-insulin-dependent diabetes mellitus. Eur J Endocrinol 136: Mohamed-Ali V, Pinkney JH, Panahloo A, Goodrick 5, Coppack SW, Yudkin JS (1997) Relationships between plasma leptin and insulin concentrations, but not insulin resistance, in non-insulin-dependent (Type 2) diabetes mellitus. Diabet Med 14: Schwartz MW, Moore J, Prigeon RL, Morawiecki A, Kahn S, Boyko EJ, Nicolson M, Porte D (1997) Evidence that plasma leptin and insulin levels are associated with body adiposity via different mechanisms. Diabetes Care 20: Matthews DR, Hosker JP, Rudenski AS, Naylor BA, Treacher DF, Turner RC (1985) Homeostasis model assessment: insulin resistance and -cell function from fasting plasma glucose and insulin concentrations in man. Diabetologia 28: de Courten M, Zimmet P, Hodge A, Collins V, Nicolson M, Staten M, Dowse G, Alberti KGMM (1996) Hyperleptinemia: the missing link in the metabolic syndrome? Diabet Med 14:

7 INSULIN RESISTANCE AND LEPTIN IN DIABETICS Kim-Motoyama H, Yamaguchi T, Katakura T, Miura M, Ohashi Y, Yazaki Y, Kadowaki T (1997) Serum leptin levels are associated with hyperinsulinemia independent of body mass index but not with visceral obesity. Biochem Biophysic Res Commun 239: Carantoni M, Abbasi F, Azhar S, Chen YDI, Klebanov M, Wang PW, Warmerdam F, Reaven GM (1998) Plasma leptin concentrations do not appear to decrease insulin-mediated glucose disposal or glucose-stimulated insulin secretion in women with normal glucose tolerance. Diabetes 47: World Health Organization Study Group on Diabetes mellitus (1985) Diabetes mellitus: Report of a WHO Study Group. Geneva, World Health Org. pp Kennedy A, Gettys TW, Watson P, Wallace P, Ganaway E, Pan Q, Garvey WT (1997) The metabolic significance of leptin in humans: gender-based differences in relationship to adiposity, insulin sensitivity, and energy expenditure. J Clin Endocrinol Metab 82: Vettor R, de Pergola G, Pagano C, Englaro P, Laudadio E, Giorgino F, Blum WF, Giorgino R, Federspil G (1997) Gender differences in serum leptin in obese people: relationships with testosterone, body fat distribution and insulin sensitivity. Eur J Clin Invest 27: Hardie LJ, Rayner DV, Holmes S, Trayhurn P (1996) Circulating leptin levels are modulated by fasting, cold exposure, and insulin administration in lean but not Zucker (fa/fa) rats as measured by ELISA. Biochem Biophys Res Commun 233: D'Adamo M, Buongiorno A, Maroccia E, Leonetti F, Barbetti F, Giaccari A, Zorretta D, Tamburrano G, Sbraccia P (1998) Increased ob gene expression leads to elevated plasma leptin concentrations in patients with chronic primary hyperinsulinemia. Diabetes 47: Ogawa Y, Masuzaki H, Hosoda K, Aizawa-Abe M, Suga J, Suda M, Ebihara K, Iwai H, Matsuoka N, Satoh N, Odaka H, Kasuga H, Fujisawa Y, Inoue G, Nishimura H, Yoshimasa Y, Nakao K (1999) Increased glucose metabolism and insulin sensitivity in transgenic skinny mice overexpressing leptin. Diabetes 48: Hislop MS, Ratanjee BD, Soule SG, Marais AD (1999) Effects of anabolic-androgenic steroid use of gonadal testosterone suppression on serum leptin concentration in men. Eur J Endocrinol 141: Dua A, Hennes MI, Hoffmann RG, Maas DL, Krakower GR, Sonnenberg GE, Kissebah AH (1996) Leptin: a significant indicator of total body fat but not of visceral fat and insulin insensitivity in African-American women. Diabetes 45: Nagasaka S, Iwamoto Y, Ishikawa S, Kuzuya T, Saito T (1997) Efficacy of troglitazone measured by insulin resistance index. Lancet 350: Ono T, Shiga N, Taneda Y, Umemura S (1999) The fasting-plasma glucose range in which insulin resistance measured by homeostasis model assessment correlates with euglycemic clamping. J Japan Diab Soc 42: