A cross-sectional survey was undertaken for this study. Areas without high water iodine levels in Tianjin were selected as the survey sites, including the urban areas of Hedong District and the suburban areas of Hangu District. The subjects of this study were individuals with insufficient iodine, adequate iodine, and above requirements iodine levels. The classification of iodine nutritional status is shown in supplemental material. The study population was selected by random cluster sampling. Participants included in the study had lived l ocally for at least five years. Pregnant women, breastfeeding women, and participants who took drugs containing iodine in the last three months were excluded.
PASS (version 15) was used for sample-size calculations. As this study was a cross-over research design, we used the confidence interval (CI) for one proportion for sample size calculation. According to Flores-Rebollar’s study, the prevalence of clinical hypothyroidism, subclinical hypothyroidism and subclinical hyperthyroidism in healthy adults was 1.8%, 5%, 2.8% and the 1.8% was selected for calculation. the allowable error was set at 1% and the α error was set at 0.05. The calculation revealed a minimum sample size of 789.
Baseline demographic information (gender, age, height, weight) was obtained through questionnaires. Body mass index (BMI) was calculated according to the following formula: BMI = weight (kg)/height 2(m2) and body surface area (BSA) was calculated according to the following formula: BSA=weight (kg)0.425 × height (cm)0.725 × 0.007184. Trained professionals collect blood and urine samples on-site. Serum and urine samples were sealed and stored at −80℃ until analysis.
sUIC and UCr
Urinary iodine concentration (sUIC) was detected and analyzed by inductively coupled plasma mass spectrometry ICP-MS (iCAP Q, Thermo Fisher Scientific, Germany) using Te for mass bias correction. A calibration curve was obtained using nine solutions with iodine concentrations within the range of 0–1200 µg/L. The R2 of the standard curve was not less than 0.999. The quality of each batch of urine samples was compared with standard human urine material (Ref. 1403081, Seronorm, Norway). The average concentration of iodine in the standard urine was 292.5±10.3μg/L, which was consistent with the certified value (297 μg/L).
Urinary creatinine (UCr) was measured by a national standard spectrophotometric method. Using this method, the CV for UCr concentration was 0.2 - 3.2% in the laboratory. Two concentrations of urinary creatinine standard were used to correct the urinary creatinine concentration: 0.649g/l (95% CI: 0.489-0.808g/L) and 1.469g/l (95% CI: 1.050-1.887g/L). The relative standard deviation was 3.8% ~ 4.2%.
Thyroid function tests
Serum free triiodothyronine (FT3), free thyroxine (FT4), and thyroid stimulating hormone (TSH) were measured by chemiluminescence immunoassays (Bayer Healthcare, Siemens, Germany). The detection range of FT3, FT4 and TSH were 0.8-30pg/ mL, 4.5-60pg/ mL and 0.2-50mIU/L, respectively. The intra assay coefficients of variation were TSH: 2.1-4.9%, FT4: 1.7-4.2%, FT3: 2.4-3.1%. The coefficients of variation between batches were FT3: 2.8-4.1%, FT4: 1.4-3.5%, TSH: 1.5-4.4%. Thyroid disorders include clinical hyperthyroidism, subclinical hyperthyroidism, clinical hypothyroidism, and subclinical hypothyroidism. Detailed diagnostic criteria are provided in Table 1, which includes the suggested reference ranges as suggested by the Laboratory Department of the General Hospital of Tianjin Medical University.
Serum iodine concentration
Serum iodine concentration was analyzed by inductively coupled plasma mass spectrometry ICP‐MS (iCAP Q, Thermo Fisher Scientific, Germany) using tellurium for mass bias correction. Serum samples were diluted with 1.5% isopropyl alcohol and 7 mmol aqueous ammonium before ICP-MS analysis to determine serum iodine (SI). Protein was precipitated with acetonitrile (acetonitrile: serum = 50μL: 25μL). The quality of each batch of serum samples was compared with standard human serum material (Ref. 201405, Seronorm, Norway). The average concentration of iodine in the standard serum was 71.7±2.2μg/L, which was consistent with the certified value (71.8μg/L). The detection range of serum iodine was 0-1000μg/L.
Microsoft Excel (Win10, 2019) and SPSS (version 22, NCSS Statistical Software) were used for data processing and statistical analyses. The statistical power (1-β error probability) was set at 0.8, the significance level α was set at P < 0.05, The normal distribution variables, including age, height, weight, BMI, BSA, were expressed as mean ± standard deviation. sUIC, UCr, sUIC/UCr, SI and TSH were non-normal distribution variables and were therefore expressed using the median (25th percentile, 75th percentile). Categorical variables, such as the prevalence of thyroid disorders, are expressed as percentages. Comparative analysis of the different groups was performed using a one-way ANOVA or Kruskal-Wallis tests. The trend test was performed to assess the relationship between trends in iodine nutrition indicators and the prevalence of thyroid dysfunctions by SI quartile groups. The relationship between SI and LnTSH was analyzed by local polynomial regression, with fitting on a scatter diagram. Binary and multivariate logistic regression was used to assess the relationship between SI and iodine deficiency, iodine excess, and thyroid dysfunctions.