Diabetes Care In Press, published online May 7, 2007
Relationships Between Estimates of Adiposity, Insulin Resistance and Nonalcoholic Fatty Liver Disease in a Large Group of Nondiabetic Korean Adults Received for publication 14 March 2007 and accepted in revised form 26 April 2007. Ki Chul Sung MD1,2*, Marno C Ryan MD2*, Bum Soo Kim MD,1, Yong Kyun Cho,1, Byung Ik Kim MD,1 Gerald M Reaven MD2 *Equal first author Division of Medicine, Kangbuk Samsung Hospital, Sungkyunkwan University School of Medicine, Seoul, South Korea 2 Division of Medicine, Stanford University Medical Center, 300 Pasteur Drive, Stanford, CA, 94305 USA
1
Running title: Adiposity, Insulin Resistance and NA FLD
Correspondence: Dr Ki Chul Sung MD Kangbuk Samsung Hospital, Sungkyunkwan University, Pyung Dong, Jongro-Ku Seoul, Korea 110-746 Email:
[email protected]
Copyright American Diabetes Association, Inc., 2007
Background: Nonalcoholic fatty liver disease (NAFLD) is emerging as a major health problem in parallel with an increasing prevalence of obesity. Insulin resistance, abdominal and overall adiposity are closely associated with NAFLD; however the interplay between these in the relationship with NAFLD is unclear, especially in nondiabetic individuals. Methods: Measurements were made in 56,249 Korean subjects of abdominal ultrasound, fasting plasma insulin (FPI) and lipid concentrations, hepatitis serology, overall obesity (body mass index, BMI), and abdominal obesity (waist circumference, WC). Results: After rigorous exclusion criteria, 36,654 nondiabetic subjects (54% men) were enrolled. Subjects were divided into controls (no fatty liver on ultrasound, serum ALT 30 U/L for men/> 19 U/L for women were classified as “FL-HALT”. These ALT cut points were determined after a review of previous work both in Koreans (12,13), and non-Koreans (14,15), in an effort to use genderspecific cut points that accurately detected abnormal values. Finally, subjects without FL on ultrasound, but an ALT>30 U/L(men)/>19 U/L (women) (5414 subjects) were not included in this analysis, but the mean FPI and TG/HDL-C concentrations for this group were compared with the other groups and are presented. Since specific measures of insulinmediated glucose uptake had not been performed, the two following surrogate estimates of insulin sensitivity were used to assess the
relationship between insulin resistance/hyperinsulinemia and FL. FPI concentrations are significantly correlated with direct measures of insulin resistance (10,16). The plasma concentration ratio of TG/HDL-C is as closely related to direct measures of insulin resistance as is FPI concentration (10), but has been shown to be superior to either TG or HDL-C alone (17). Statistics Statistical analysis of the data was performed using SPSS version 14.0 (SPSS, Point Richmond, CA, USA), and the continuous variable data is presented as mean ± SD. Student’s unpaired t-test was used to compare the means of Grade 1 FL versus Grades 2 and 3 combined. Analysis of variance (ANOVA) was used to evaluate the overall differences in means. ANCOVA was used to assess the effect of weight (BMI and WC) on the relationship between markers of insulin resistance and the presence of FL. For the comparison of nominal variables, the Chi-square method was used for cross-tabulation analysis. Multiple logistic regression analysis was used to analyze relationships between age, BMI, WC and insulin and the TG/HDL-C ratio and the presence of FL-NALT and FL-HALT. P values less than 0.05 were considered to be statistically significant. Results Overall, the 36,654 subjects were relatively young [mean ± SD age 41±9 years (men 41±8, women 41±9 years)], and nonobese with a BMI of 23.0±3.0 kg/m2 (men 24.3±2.8, women 22.2±2.9 kg/m2) and WC of 78±9 cm (men 83±8, women 72±8
cm). Hepatic ultrasonography indicated that 27,632 subjects were without evidence of FL (75.4%), while the remaining subjects had grade 1 (21.4%, 6149 men, 1683 women), grade 2 (3.2%, 977 men, 181 women), or grade 3 (0.09%, 29 men, 3 women) FL respectively. In Table 1, the subjects are separated by FL grade and gender. Subjects with Grades 2 and 3 FL were combined into one category, due to the very small numbers in Grade 3. For the remainder of the presented analyses, the FL grades are not separated, and the data is presented for two groups: FL-NALT and FL-HALT. 1. The presence of FL and elevation of ALT increased with the degree of both overall and abdominal obesity. The relevant clinical characteristics of the 36,654 subjects, divided into the three experimental groups (control, FL-NALT, and FL-HALT) are shown in Table 1, with the two FL groups also divided into FL grades. In general, the FL groups had a higher overall (BMI) and abdominal (WC) obesity than controls, and the FL-HALT group was significantly more obese by both measures than the FL-NALT group. These data also show that there is a significant trend to have worsening metabolic variables (FPI, TG and HDL-C concentrations, and the TG/HDL-C concentration ratio) across the groups, from the control group to the FL-NALT group, and the FL-HALT group. After division of the two FL groups into Grades of FL, the same can be said of men with Grade 2 FL in comparison with those with Grade 1 FL, within both the FL-NALT and the FL-HALT groups. Thus, in men, the degree of steatosis was positively
associated with worsening metabolic variables. For women, although the division into Grades of FL also demonstrated an association between worsening metabolic variables and a higher grade of FL, female subjects with Grade 2/3 FL in the two FL/ALT groups had equivalent mean values for variables. Table 2 presents the mean values for FPI concentration and the TG/HDL-C concentration of the three experimental groups, this time adjusted for differences in age, BMI, and WC. These results indicate that the estimates of insulin resistance increased in magnitude in parallel with presence of FL and ALT level, with each group being significantly different from the other two groups. 2. Odds ratios for features of NAFLD increased with increasing insulin resistance, independently of obesity. In order to quantify the adverse impact of obesity on the degree of liver disease, odds ratios (ORs) were calculated between age, BMI and WC and the presence of FL-NALT or FLHALT (Table 3). In this relatively young population, being older did not seem to have an adverse effect on the presence of NAFLD in men. However, for women, the two FL groups were significantly older than controls. In terms of adiposity, the larger the BMI or WC, the greater the OR of having either FL-NALT or FL-HALT. The increase in ORs ranged from 10% to 30%, with the values for BMI and WC seeming of comparable magnitude. In Table 4, the impact of insulin resistance was assessed with the
calculation of ORs between FPI concentrations and the TG/HDL-C concentration ratio and the presence of FL-NALT or FL-HALT. For this purpose, the experimental population was divided into quartiles on the basis of their FPI concentration or their TG/HDL-C concentration ratio. Age, BMI and WC were included in the regression model for both the FPI and TG/HDL-C analyses, so that the increasing quartiles of the two markers of insulin resistance are independent of these three variables. In terms of FPI concentration, when comparing each of the three highest insulin quartiles with the lowest, ORs for the presence of FL-HALT increased significantly in both men and women, with the OR increasing five to six-fold in the quartile with the highest FPI concentration. For the TG/HDL-C ratio, similar results are seen. When looking at the ORs for FPI and TG/HDL-C concentrations in predicting the presence of FL-NALT, the results are similar for women, albeit of smaller magnitude. However for men, the OR for FL-NALT decreased with the highest FPI quartile, indicating a decrease in the risk with this degree of hyperinsulinemia. This is difficult to explain, but could indicate that in men an increasing FPI concentration is less associated with steatosis alone, and more associated with steatohepatitis. This is with the caveat that ALT is only a moderately accurate marker of inflammatory activity. Thus apart from this one exception, the higher the FPI concentration and TG/HDL-C ratio, the significantly greater the OR of having either FLNALT or FL-HALT.
3. Secondary analyses Excluded from the above analyses were 5414 subjects (3412 female, 2002 male) with an isolated elevated ALT>30 U/L for men and >19 U/L for women (i.e., no FL on ultrasound), as they could not be included. We compared the mean FPI concentrations and TG/HDL-C ratios for this group with those of our other groups, and found that for both of these measures, the mean values for the isolated ALT group were significantly lower than either the FLNALT or the FL-HALT groups after adjusting for age, BMI and WC (data not shown). Discussion Both obesity and insulin resistance have been identified as factors that are associated with deposition of fat in the liver, although it is unclear whether they are a cause of hepatic fat (18-21) or a result of this hepatic fat (22-25). Since obesity and insulin resistance are themselves significantly correlated (26,27), it is difficult to clarify which of these two variables is most closely related to increased hepatic fat content. The problem is further confounded by questions as to the relative adverse impact of overall obesity as compared to abdominal obesity. This study is, to the best of our knowledge, the largest that has been performed analyzing associations between measures of overall and abdominal obesity, insulin resistance, hepatic transaminases and FL, using a comprehensive evaluation including ultrasound, hepatic enzymes and serum viral markers, in a nondiabetic, apparently healthy population.
Based upon the results in Tables 1 and 2, it appears that both surrogate estimates of insulin resistance used in this study (FPI concentration and the TG/HDL-C concentration ratio) are independently associated with severity of liver disease. Thus, although the results in Table 1 indicate that both estimates of obesity (BMI and WC) and the two surrogate estimates of insulin resistance all increased significantly as a function of the presence of steatosis and a raised ALT, it can be seen from Table 2 that FPI concentration and the TG/HDL-C concentration ratio continued to be statistically associated with these features of liver disease when adjusted for differences in age, BMI, and WC. Furthermore, when the subjects were divided into quartiles on the basis of the magnitude of their surrogate estimates of insulin resistance, the OR of having evidence of FL-HALT increased significantly with each successively higher quartile (Table 4). As with the results in Table 2, these findings were independent of differences in age, BMI, and WC, and with the exception of relatively minor quantitative differences were true of both genders. Thus, in answer to the question we posed, the evidence strongly supports the notion that although both overall and abdominal obesity are associated with FL, the specific relationship between insulin resistance and FL is independent of either overall or abdominal obesity. It is of interest that the relationships between either index of obesity and hepatic fat content were quite comparable, so that neither WC nor BMI was more predictive than the other of hepatic fat. However, the
population under study was a very specific population of East Asian ancestry, with a lower mean BMI and WC than seen in Caucasians, and these findings may not be applicable to other ethnic groups. Further, it should be emphasized that the specific cut points that performed the best in this analysis may only be applicable to individuals of Korean ethnicity. Although the results of this study seem relatively straight forward, certain weaknesses should be acknowledged. Firstly, subjects with sonographic evidence of cirrhosis were excluded, and in doing so some NAFLD chronic liver disease or cirrhosis may have also been excluded, explaining why we found such small numbers with Grade 3 FL. Also, NAFLD cirrhosis, where the hepatic fat tends to be greatly diminished, can be mistaken sonographically for normal non-FL (28). Thirdly, and most importantly, our conclusions are based upon the combined use of hepatic ultrasonography and ALT measurements as markers of NAFLD, without morphological examination of
hepatic tissue. We used “FL-HALT” to attempt to measure prevalence of inflammatory activity in the liver, to distinguish this from benign FL. We acknowledge that serum ALT levels have been shown to be at best a moderate predictor of hepatic necroinflammatory damage or steatohepatitis (29-31), however, we would argue that the large number of apparently healthy individuals studied precludes the possibility of obtaining histology. In summary, we demonstrate that although both adiposity and surrogate estimates of insulin resistance (the FPI concentration and the TG/HDL-C ratio) are associated with the estimates of NAFLD in this healthy, nondiabetic Korean population, these measures of insulin resistance predict NAFLD independently of both overall and abdominal overweight/obesity.
Acknowledgements We would like to acknowledge the efforts of the health screening group at Kangbuk Samsung Hospital, Seoul, South Korea.
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Table 1: Clinical Characteristics of the control group and the two FL groups, divided by Grade of FL and gender (Mean ±SD). Men (n=17616) Control
Women (n=13624)
FL-NALT Grade 1
Grade 2/3
N
10461
2815
128
Age (y)
40 ± 8
42 ± 8
40 ± 9
BMI (kg/m2)
22.9 ±2.4
25.2 ± 2.2
WC (cm)
79 ± 7
FPI (pmol/L)
p*
FL-HALT Grade Grade 1 2/3
P p*
3334
878
0.08
40 ± 7
39 ± 7
