DOI: 10.5061/dryad.gqnk98skp
title: Unfavorable triglyceride-rich particle profile in subclinical thyroid disease: A cross-sectional analysis of ELSA-Brasil

description: Introduction: Subclinical thyroid disorders have been associated with
 atherosclerosis and increased cardiovascular risk. As triglyceride-rich
 lipoprotein particles (TRLP) have recently emerged as a casual factor for
 atherogenesis, the aim of this study was to evaluate the relationship
 between subclinical hypo and hyperthyroidism and TRLP subfractions.
 Methods: We selected 5,066 participants from the ELSA-Brasil cohort with
 available data of thyroid function and lipid profile measured by NMR.
 Individuals were divided into three groups by baseline thyroid function
 (subclinical hypothyroidism, euthyroidism, and subclinical
 hyperthyroidism). TRLP subfractions were analyzed through Nuclear Magnetic
 Resonance (NMR) spectroscopy. To examine the association between TRLP
 subfractions and thyroid function, we conducted univariate and
 multivariate linear regression models adjusted for demographic
 characteristics, BMI, diabetes, smoking status and alcohol use. Results:
 Of 3,304 individuals, 54% were women, with a mean age of 50.6±8,7 years,
 51% white and 53% with at least college education. Of these individuals,
 92% were euthyroid, whereas 6.8% had subclinical hypothyroidism and 1.2%
 had subclinical hyperthyroidism. The univariate linear regression showed
 that Very Small TRLP (p=0.026) and Very Large TRLP (p=0.008) were
 statistically increased in subclinical hypothyroidism when compared to
 euthyroidism. In subclinical hyperthyroidism, there was a reduction in
 total TRLP (p=0.003), seemingly driven by reduced Very Small-TRLP
 (p=0.067). The findings were confirmed when adjusted for demographic
 characteristics, as well as comorbidities. Conclusions: This study
 suggests that subclinical hypothyroidism is associated to very small and
 very large TRLP, which are related to an unfavorable atherogenic profile.
 Subclinical hyperthyroidism is associated to lower very small TRLP.
description: ELSA-Brasil
 Study The Brazilian
 Longitudinal Study of Adult Health (ELSA-Brasil) is a prospective cohort
 study following 15,105 civil servants from six cities in Brazil (Salvador,
 Belo Horizonte, Vitória, Rio de Janeiro, São Paulo and Porto Alegre) and
 has been previously described(26-28). The main objective of the study is
 to investigate the incidence of cardiovascular diseases and diabetes, as
 well as their associated risk factors(27,29). All participants in this
 study provided written informed consent and each enrolling site obtained
 approval from the Institutional Review Boards (CAAE 0016.1.198.000-06)
 according to the Declaration of Helsinki. We selected 5,066 participants
 from the Investigation Center of São Paulo with available data of thyroid
 function and lipid profile measured by NMR. Participants with overt
 thyroid disease, using statins or medications that could interfere with
 thyroid function and those with previous severe cardiovascular event were
 excluded from the analysis as described in figure 1.
 Thyroid Function
 Definition Thyroid-stimulating
 hormone (TSH) and Free Thyroxine (FT4) were measured using a
 third‐generation immunoenzymatic assay (Siemens, Deerfield, IL, USA) in
 serum obtained from centrifuged venous blood samples after overnight
 fasting.<b> </b>FT4 levels were measured in participants
 exhibiting altered TSH levels. In this study, reference range levels were
 0.55 - 4.78 mcUI/mL for TSH and 0.89 - 1.76 ng/dL for FT4, similar to
 other ELSA‐Brasil studies(30-32). The intra-assay and inter-assay
 coefficients of variation were 3.32% and 9.65% for TSH and 1.40% and 6.40%
 for FT4, respectively. Biochemical data about the performed tests can be
 seen in table 1 of the supplemental material*.
 ELSA‐Brasil
 participants were classified into five categories of thyroid function,
 according to TSH and FT4 levels and information related to the use of
 medication to treat thyroid disorders: overt hypothyroidism (high TSH and
 low FT4 levels, or use of levothyroxine to treat hypothyroidism),
 subclinical hypothyroidism (high TSH levels, normal FT4 levels and no use
 of drugs to treat thyroid diseases), euthyroidism (normal TSH and no use
 of thyroid drugs), subclinical hyperthyroidism (low serum TSH, normal FT4
 levels and no use of drugs to treat thyroid diseases) and overt
 hyperthyroidism (low serum TSH and high FT4 levels or use of medication to
 treat hyperthyroidism). Accordingly, the definition of subclinical thyroid
 disease included only participants who did not use any drugs to treat
 thyroid disorders. Participants with overt thyroid disorders were excluded
 from the analyses. Lipoprotein
 Measurements Blood was collected
 from participants after 8-12 hours nocturnal fasting. The samples were
 centrifuged at the sites and stored in tubes at - 80°C. Conventional lipid
 concentrations (total cholesterol, HDL-cholesterol, and triglycerides)
 were determined by a nonprecipitated colorimetric method using ADVIA 1200
 Siemens equipment. LDL cholesterol was calculated using the Friedewald
 equation(33), except for cases with elevated triglyceride levels (more
 than 400mg/dL), when an enzymatic colorimetric assay was used (ADVIA 1200,
 Siemens). Detailed data about the performed tests can be found at table S1
 of the supplemental material(26,34)*. Triglyceride-rich
 lipoproteins (TRLP) subfractions were measured by new nuclear magnetic
 resonance (NMR) spectroscopy (NMR LipoProfile® 4 test spectra, LabCorp,
 Raleigh, NC). This technique quantifies the size as well as the
 concentration (“number”) of lipoprotein particles expressed as an average
 particle size (in nanometers) or as lipoprotein particle concentration (in
 particle nmol/L). NMR can quantify the concentration of lipoprotein
 subclass particles because of two phenomena: lipoprotein subclasses of
 different size in plasma simultaneously emit distinctive NMR signals whose
 individual amplitudes can be accurately and reproducibly measured; and
 also the measured subclass signal amplitudes are directly proportional to
 the numbers of subclass particles emitting the signal, irrespective of
 variation in particle lipid composition(35). Thus, this method relies on
 the spectroscopic distinctness of particles’ size, and not protein
 composition or metabolic origin(36). Total TRLP comprises particles
 measuring 24 to 240nm in diameter. The mean particle size for TRLP (TRLZ)
 rests between 30 and 100nm. IDL-cholesterol is now named very small TRLP
 (VS-TRLP), comprising particles with estimate diameter from 24 to 29nm.
 Similarly, other NMR fractions are defined as small TRLP (S-TRLP) – 30 to
 36nm, medium TRLP (M-TRLP) – 37 to 49nm, large TRLP (L-TRLP) – 50 to 89nm
 and very large TRLP (VL-TRLP) – 90 to 240nm within the TRLP particles(37).
 Other
 variables Age is presented as a
 continuous variable. Sex is described as percentage female. Race was
 self-reported and categorized as white, brown, black and other (including
 Asian and indigenous). Educational level was stratified as less than high
 school, high school, and college or higher. Anthropometric measures were
 obtained with standard techniques with the participant wearing light
 clothes. Body mass index (BMI) was calculated as weight divided by height
 squared (kg/m<sup>2</sup>). Hypertension was defined as the
 use of antihypertensive drug, systolic blood pressure ≥ 140 mm Hg or
 diastolic blood pressure ≥ 90 mm Hg. The diagnosis of diabetes was based
 on a previous medical diagnosis of diabetes self-report by the participant
 or use of oral antidiabetic agents or insulin therapy, fasting plasma
 glucose ≥ 126 mg/dL, 2-hour post-prandial 75 g glucose test ≥ 200 mg/dL,
 or glycosylated hemoglobin ≥ 6.5% (exams which were evaluated in all
 participants) (38). Smoking status was defined as never, former (withdraw
 for more than 2 years) or current. Participants are considered smokers if
 they smoked more than 100 cigarettes (or five packs) in their life before
 enter the study or if they answered yes to the question “Do you currently
 smoke cigarettes?” independently of the number of cigarettes smoked.
 Alcohol use was defined as never, former or current. Former alcohol users
 were those who stopped using alcohol for more than 2 years. We classified
 participants in the excessive drinking category if they intake more than
 210 g per week if men and more than 140 g per week if women according to
 WHO criteria(39). Statistical
 Analysis Continuous variables
 were presented as descriptive statistics with mean and standard deviation
 for normal distribution and, for non-normal distributions, with median
 (quartiles). The data distribution’s normality was verified through
 Kolmorogov-Smirnov test. We showed categorical variables in absolute and
 relative frequency. TSH, triglycerides and TRLP subfractions were
 log-transformed for use in regression models. Comparison of quantitative
 variables across groups was performed using analysis of variance for
 variables with normal distribution and the Kruskal-Wallis test and
 Spearman correlation for those with non-normal distributions. Categorical
 variables were analyzed by chi-square test and Fisher’s exact test. To
 examine the association between TRLP subfractions and thyroid function, we
 conducted univariate and multivariate linear regression models. For these
 analyses, we standardized total TRLP, VS-TRLP, S-TRLP, M-TRLP, L-TRLP,
 VL-TRLP and mean size of TRLP (TRLPZ). Three multiple linear regression
 models were adjusted as follow: Crude model: univariate; Model 1: race,
 gender and educational level; Model 2: model 2 + BMI, diabetes,
 hypertension, smoking status and alcohol use. Statistical significance was
 defined as p&lt;0.05. Analyses were performed with Stata version 13.1
 (StataCorp, USA).
description: The information in this dataset refers to the Supplemental
 Material of the original article.