Lipids in Health and Disease

Background: Coronary heart disease is increasing in urban Indian subjects and lipid abnormalities are important risk factors. To determine secular trends in prevalence of various lipid abnormalities we performed studies in an urban Indian population. Methods: Successive epidemiological Jaipur Heart Watch (JHW) studies were performed in Western India in urban locations. The studies evaluated adults ≥ 20 years for multiple coronary risk factors using standardized methodology (JHW-1, 1993–94, n = 2212; JHW-2, 1999–2001, n = 1123; JHW-3, 2002–03, n = 458, and JHW-4 2004–2005, n = 1127). For the present analyses data of subjects 20–59 years (n = 4136, men 2341, women 1795) have been included. In successive studies, fasting measurements for cholesterol lipoproteins (total cholesterol, LDL cholesterol, HDL cholesterol) and triglycerides were performed in 193, 454, 179 and 252 men (n = 1078) and 83, 472, 195, 248 women (n = 998) respectively (total 2076). Age-group specific levels of various cholesterol lipoproteins, triglycerides and their ratios were determined. Prevalence of various dyslipidemias (total cholesterol ≥ 200 mg/dl, LDL cholesterol ≥ 130 mg/dl, non-HDL cholesterol ≥ 160 mg/dl, triglycerides ≥ 150 mg/dl, low HDL cholesterol <40 mg/dl, high cholesterol remnants ≥ 25 mg/dl, and high total:HDL cholesterol ratio ≥ 5.0, and ≥ 4.0 were also determined. Significance of secular trends in prevalence of dyslipidemias was determined using linear-curve estimation regression. Association of changing trends in prevalence of dyslipidemias with trends in educational status, obesity and truncal obesity (high waist:hip ratio) were determined using two-line regression analysis. Results: Mean levels of various lipoproteins increased sharply from JHW-1 to JHW-2 and then gradually in JHW-3 and JHW-4. Age-adjusted mean values (mg/dl) in JHW-1, JHW-2, JHW-3 and JHW-4 studies respectively showed a significant increase in total cholesterol (174.9 ± 45, 196.0 ± 42, 187.5 ± 38, 193.5 ± 39, 2-stage least-squares regression R = 0.11, p < 0.001), LDL cholesterol (106.2 ± 40, 127.6 ± 39, 122.6 ± 44, 119.2 ± 31, R = 0.11, p < 0.001), non-HDL cholesterol (131.3 ± 43, 156.4 ± 43, 150.1 ± 41, 150.9 ± 32, R = 0.12, p < 0.001), remnant cholesterol (25.1 ± 11, 28.9 ± 14, 26.0 ± 11, 31.7 ± 14, R = 0.06, p = 0.001), total:HDL cholesterol ratio (4.26 ± 1.3, 5.18 ± 1.7, 5.21 ± 1.7, 4.69 ± 1.2, R = 0.10, p < 0.001) and triglycerides (125.6 ± 53, 144.5 ± 71, 130.1 ± 57, Published: 24 October 2008 Lipids in Health and Disease 2008, 7:40 doi:10.1186/1476-511X-7-40 Received: 16 July 2008 Accepted: 24 October 2008 This article is available from: http://www.lipidworld.com/content/7/1/40 © 2008 Gupta et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


Introduction
Cardiovascular diseases, especially coronary heart disease, are important public health problems in India and many developing countries [1,2]. There is evidence that the diseases are increasing in these countries in contrast to developed nations of Europe and North America where the incidence has decreased [3,4]. Societal changes as well as individual lifestyle factors are important in driving this cardiovascular epidemic [5]. These changes influence the proximate determinants of atherosclerosis which include smoking and tobacco use, high total and low density lipoprotein (LDL) cholesterol, low high density lipoprotein (HDL) cholesterol, high blood pressure, diabetes and the metabolic syndrome. Trends of these risk factors have been well studied in developed countries and show significant correlation with rise and fall of the coronary heart disease epidemic [5].
There have been only a few studies that have examined trends in cardiovascular risk factors in middle and low income countries [6]. In Seven Countries Study multiple cross sectional surveys were conducted among men aged 40-59 years in Yugoslavia, Italy, Greece, Holland, Finland, Japan and USA [7]. These studies reported that while major coronary risk factors initially stabilized and later declined in many of these countries, in middle income countries such as Yugoslavia the risk factors increased. The WHO-MONICA study reported that population risk factors increased in the Chinese while they declined in North American and Western European cohorts [8,9]. Increasing trends in coronary risk factors has also been reported from many middle income Latin American countries [6]. In Asia, increasing trends in lipids and in prevalence of dyslipidemias (high LDL cholesterol and low HDL cholesterol) has been reported in urban populations of Beijing [10], rural China [11] and South Korea [12].
To our knowledge no single study that systematically evaluated trends in major cardiovascular risk factors in India exists although reviews have reported increasing prevalence of hypertension [13], diabetes [14], and hypercholesterolemia [2], and declining smoking rates among the educated Indians [15]. All these evaluations suffer from multiple biases inherent in compiling studies from different sources and different methodologies [16]. We performed multiple coronary heart disease risk factor epidemiological studies in urban populations in western Indian state of Rajasthan to determine their lifestyle and other determinants [17][18][19][20]. Here we report trends in levels of various lipoproteins (total, LDL, HDL and non-HDL cholesterol, triglycerides) and total-HDL cholesterol ratio and prevalence of dyslipidemias using current definitions.

Methods
A series of cross sectional epidemiological studies using similar tools in the Indian state of Rajasthan over years 1992-2005 were performed to determine cardiovascular risk factors in urban populations [17][18][19][20]. All the studies were approved by the institutional ethics committee and supported financially by different organizations. The studies have been performed in Jaipur, the capital city of Rajasthan state in western India with population in year 2001 of 2.34 million. The first study -Jaipur Heart Watch (JHW)-1 [17], was conducted in years 1993-1994 and randomly selected 1608 men and 1392 women were targeted using stratified cluster sampling on the Voters' lists in six locations in Jaipur city. 2212 subjects (1415 men 88.0%, 797 women 57.3%) were evaluated for various cardiovascular risk factors and attempt for fasting blood sample for cholesterol lipoproteins and triglycerides was in 15%. In the second urban study (JHW-2) [18] we targeted 960 men and 840 women in the same locations as in JHW-1 and could examine 550 men (57.3%) and 573 women (68.2%). In this study we targeted all the participants for the fasting blood sample. The third (JHW-3) [19] and fourth (JHW-4) [20] urban studies targeted at a smaller sample and were designed to gather information on risk factors in middle-class locations. Response rates are shown in Table 1.

Data collection
Methodological details have been previously reported [17]. Briefly, we collected information regarding demographic data, educational level, history of chronic illnesses such as coronary heart disease, hypertension, diabetes or high cholesterol levels, and smoking or tobacco intake. Brief questions were asked to evaluate physical activity and diet but the results were considered inadequate and not included in the analyses. Physical examination was performed to assess height, weight, waist and hip size and blood pressure. BMI was calculated as weight (kg) divided by squared height (m). Waist-to-hip ratio was calculated. Fasting glucose was determined at a central laboratory using glucose peroxidase method and external quality control. Total cholesterol was measured using cholesterol oxidase-phenol 4-aminophenazone peroxidase method and HDL cholesterol using an enzymatic method after precipitating non-HDL cholesterol with a managaneseheparin substrate. Triglycerides were measured using the glycerol phosphate oxidase-peroxidase enzymatic method. Quality control measures were followed for estimation of total cholesterol, high density lipoprotein (HDL) cholesterol and triglycerides while low density lipoprotein (LDL) cholesterol was estimated using the Friedewald formula [21].

Diagnostic criteria
The diagnostic criteria for coronary risk factors have been advised by the American College of Cardiology clinical data standards [22]. Educational level was used as marker for socioeconomic status as reported in an earlier study [23]. More than 5 years of formal education (primary education or more) was taken as acceptable literacy level for analysis. Obesity or overweight was defined as body mass index of ≥ 25 kg/m 2 and truncal obesity was defined by waist:hip ratio of > 0.95 for men and > 0.85 for women [24]. Dyslipidemia was defined by the presence of high total cholesterol (≥ 200 mg/dl), high LDL cholesterol (≥ 130 mg/dl), low HDL cholesterol (< 40 mg/dl), high non-HDL cholesterol (≥ 160 mg/dl), high cholesterol remnants [very low density lipoprotein cholesterol = total -(HDL+LDL) cholesterol ≥ 25 mg/dl] or high triglycerides (≥ 150 mg/dl) according to National Cholesterol Education Program (NCEP) Adult Treatment Panel-3 (ATP-3) guidelines [25]. High total to HDL cholesterol was defined when ratio was either ≥ 5.0 or ≥ 4.0 as reported in an earlier study from India [26].

Statistical analyses
The continuous variables are reported as mean ± 1 SD and ordinal variables in percent. Prevalence rates are reported in percent. Age-stratified prevalence rates and distribution of various risk factors have been reported for decadal intervals from 20 years 59 years. Age-adjustment of various prevalence rates was performed using direct method using the Jaipur urban population according to 2001 census. Correlation of age with lipid values was performed by simple correlation analysis and significance of ageadjusted trends in mean lipoprotein levels was evaluated by 2-stage least squares regression using SPSS 10.0 statistical package (SPSS Inc, Chicago). Significance of trends in prevalence rates was determined using linear curve-estimation regression analysis using the SPSS package. Regression coefficients are reported as multiple R values after age adjustment. Significance of graphical trends was determined by logarithmic regression analysis using the Microsoft Office Power Point (2002) program. Signifi- cance of two-line trends was determined by least squares regression analyses using GB-Stat for Windows ® software 7.0 (Dynamic Microsystems Inc, Silver Spring, MD USA) and reported as r 2 values. The r 2 values of more than 0.10 and p values less than 0.05 were considered significant.
Mean levels of various lipoproteins at different age-groups are shown in Table 2. There is age-associated escalation in total cholesterol, LDL cholesterol, non-HDL cholesterol, remnant cholesterol, total:HDL cholesterol ratio and triglycerides in men and women in all the cohorts. The levels of HDL cholesterol decline with age, the decline being similar in men and women. Correlation of various lipoproteins with age in combined data from JHW studies is shown in Figure 1. There is a significant increase in total cholesterol (r = 0. 16 Table 4).

Discussion
This study shows that there is a high prevalence of various forms of lipoprotein abnormalities in Indian urban subjects. Secular trends reveal increasing mean levels of total-, LDL-, non HDL-, and remnant cholesterol, total:HDL cholesterol ratio and triglycerides and decline in HDL cholesterol. Prevalence of high non-HDL cholesterol, Correlation of various cholesterol lipoproteins and triglycerides with age in combined data from JHW studies  remnant cholesterol, and total-HDL cholesterol ratio increased. These changes correlate significantly with increasing education (socioeconomic status) and truncal obesity. Most of the lipid abnormalities are markers of dietary excess, low physical activity and increasing obesity [27]. The present study confirms that increasing obesity manifest as truncal obesity, due to population-wide sedentary lifestyle and high calorie intake [28,29], leads to increase in multiple dyslipidemias. We have previously reported increase in prevalence of coronary heart disease [2] in urban Indian populations and the present study suggests that increasing non-HDL cholesterol, cholesterol remnants, and total-HDL cholesterol ratio are important risk factors. The importance of these dyslipidemias has been highlighted in multiple prospective studies from other countries [30][31][32].
Rise and fall of cholesterol and other lipoproteins associated with changing cardiovascular mortality and coronary heart disease incidence has been well documented in Trends in age-adjusted prevalence of various dyslipidemias in various Jaipur Heart Watch (JHW) studies  Remna nt Cholester ol >=25 many developed countries [7,8,25]. There is paucity of similar data from developing countries. Data of the present study has significant healthcare policy and pharmacoeconomic implications because more than 40% of the world's population is in India and China. A econo-mies of these countries boom [33] and individual buying capacity increases the lifestyle changes shall lead to massive increase in lipid levels fuelling cardiovascular epidemic as observed in the present study in an Indian urban population. In China two large scale surveys have been carried out to determine prevalence of lipid abnormalities [34,11]. The first survey in 1992 reported greater lipid values in urban as compared to rural populations [34]. Among 9477 subjects the mean ± SD cholesterol at urban sites in China was 181.6 ± 32 to 184.8 ± 38 mg/dl in men and 187.5 ± 33 to 187.6 ± 42 in women. The values were 15-25 mg lower in rural subjects [34]. Prevalence of hypercholesterolemia ≥ 200 mg/dl was 29.1-31.0% in urban subjects and 7.7-20.0% rural subjects. Second survey in 2004 was a population based epidemiological study among 15540 adults and reported mean ± SEM cholesterol of 193.0 ± 0.7 in urban men and 196.4 ± 0.7 in urban women [11]. These levels were significantly greater than the 1992 study. This study also reported lower urban-rural gap in cholesterol levels (10-11 mg/dl more in the urban). Age-adjusted prevalence of hypercholesterolemia was 39.8% in urban men, 44.1% in urban women, 30.2% in rural men and 31.7% in rural women. Age-adjusted prevalence rates for hypercholesterolemia are lower in our study ( Table 3). Prevalence of low HDL cholesterol < 40 mg/dl in Chinese urban subjects were 29.5% in men and 14.6% in women aged 35-74 years which is lower than reported in our subjects. These studies did not report prevalence of hypertriglyceridemia or high total:HDL cholesterol ratios. Another study from China reported changing trends of cardiovascular risk factors in different socioeconomic groups but did not comment on lipid levels [35].  [7]. The decline in total and LDL cholesterol has been attributed to documented decreases in dietary intake of saturated fats and cholesterol [40]. However, recent evidence suggests that the decline in USA may have been due to increased use of medications rather than positive lifestyle changes  [40]. It is also suggested that the slower decline in recent years is likely due to increase in obesity among adults and the observed increase in triglyceride levels is a marker [39]. In the present study too, it is observed that increasing obesity is important determinant of increases in total, non-HDL, and LDL cholesterol and triglycerides. This augurs more adverse lipid profiles worldwide unless the obesity epidemic is controlled.
This study has multiple limitations as well as strengths. The variable and low response rates in some cohorts make the data tenuous but the age-structure of the studied cohorts was similar to the local populations and therefore the data can be generalized for evaluation of risk factor trends. Small number of subjects in each of the studies and age-specific subgroup could also be concern but the sample sizes have been determined using available recommendations for the prevalence of cardiovascular risk factors in a community [41] and are considered appropriate for inter-group comparisons. We have determined both age-adjusted mean levels of various lipoproteins as these could be earliest population-level change and these show significant trends. Prevalence rates are robust evidence of population level change and the present study using simple meta-regression techniques shows significant trends in important lipid abnormalities. This type of meta-regression is used for combining clinical trials as well as epidemiological studies using pre-defined end points [42]. Generalizability of the study results to the local urban population or to the whole country may not be appropriate at this time as socioeconomic structure of the country is so different from locality to locality and town to town [43]. A major strength of the study is use of similar assessment methodologies that make the observations comparable. Another strength is determination of different types of lipoprotein abnormalities and cholesterol ratios which have emerged as important risk factor. The study definitively shows that biological risk factors (lipids) are causally related to increasing obesity and to increasing socioeconomic status as measured by educational status. It has been previously reported that up to a certain level of socioeconomic status (gross national product) the risk factors tend to increase and once a particular per capita income is achieved the risk factors tend to decline with increasing socioeconomic status [44]. Indeed, a more careful assessment of the trends of dyslipidemias ( Figure  3) reveals that risk factors in educated Indian middle-class subjects may be starting to level-off as observed in JHW-3 and JHW-4 studies. This indicates importance of evolving socioeconomic changes as an important driver as well as controller of cardiovascular diseases [44].
Low HDL cholesterol and high total:HDL cholesterol are important cardiovascular risk factors. Multiple prospective studies have identified the importance of low HDL cholesterol as cardiovascular risk factor [25]. Importance of total:HDL ratio has been highlighted in the Physicians Health Study that reported relative risk (RR) of acute myocardial infarction in the top vs. bottom quintile of total:HDL cholesterol ratio was 3.73 (95% confidence interval 1.95-7.12) and was substantially greater than total cholesterol (RR 1.86; 1.05-3.28), HDL cholesterol (0.38; 0.21-0.69), and apolipoprotein B (2.50; 1.31-4.75) [45]. The INTERHEART Study also reported that ratio of apolipoprotein B/A 1 was the most important risk factor for acute myocardial infarction in South Asians [46]. It has also been reported that in patients receiving statin therapy levels of non-HDL cholesterol, apolipoprotein B, and total:HDL cholesterol ratio of ≥ 4.0 were more important than other lipid parameters [47]. Increasing ratio in this Indian urban population along with increasing non-HDL cholesterol and falling HDL cholesterol levels associated with increasing socioeconomic status and obesity shows the appropriate direction for prevention effort. Increasing socioeconomic status of Indians has to be complemented with intensive public health education and policy changes at the national level [2,48] for cardiovascular disease prevention.
In conclusion, this report is first in a low income country -India-that demonstrates cross-sectional and longitudinal trends in dyslipidemia and its causal relationship with adiposity and socioeconomic status. Data analysis of such a cohort reveals non-similarities (increasing non-HDL cholesterol, triglycerides, and total-HDL ratio) as well as similarities (increasing socioeconomic status and obesity) vis-à-vis other developing and developed regions of the world [49]. The inferences that can be translated for implications to health care policies and practice, medical education and research is beyond the scope of this publication.