Reference intervals of thyroid hormones in a previously iodine-deficient area in Darfur,Sudan

Abstract

Background:

Thyroid-stimulating hormone (TSH), thyroxin (T4) and T3 levels are varied in the different settings with disorders of thyroid homeostasis. It is recommended that every setting has to establish its own reference intervals (RIs) for these hormones.

Methods:

A multi-stage stratified sampling method was used to select a representative sample of a Sudanese adult (>20 years of age) in Nyala in western Sudan in the Darfur region during the period between January and June 2016 to establish RIs of thyroid-related hormones (TSH, T4 and T3).

In this study, 1753 serum samples (male and female) with different age groups were investigated. A radioimmunoassay gamma counter was used to measure the level of these hormones.

Results:

The median (95% intervals) of serum TSH, T4 and T3 levels was 1.2 (0.50–3.0) mIU/l, 111.0 (72.0–161.0) nmol/l and 1.5 (0.8–2.8) nmol/l respectively.

While the level of TSH was significantly higher in the age group between 31 and 40 years, both T4 and T3 levels have shown a progressive increase with age. There was no significant difference in the TSH, T4 and T3 level when the RIs were compared between males and females.

Conclusion:

The RIs for TSH, T4 and T3 in this setting were different from the levels provided by the manufacturers. The RIs were different in the different age groups.

Introduction

Thyroid-related hormones [thyroid-stimulating hormone (TSH), total triiodothyronine (T3), free triiodothyronine (FT3), total thyroxine (T4) and free thyroxine (FT4)], are the most common requested biochemical indices to diagnose and follow up thyroid diseases. ¹

In spite of the extensive use of these hormones (TSH, T3 and T4), there is a debate on the appropriate reference intervals (RIs) for these hormones. Perhaps the distribution of these hormones reference range is not normal, and their level is influenced by various genetic and environmental factors.^2,3 Thus, it is recommended that every population has to establish its own RIs for these hormones. Therefore, the RIs for these hormones were set up in many countries with good resources.^4–7

Various forms of epidemiological, etiological studies were carried out in Sudan to assess the effects of iodine deficiency disorders and the different interventions. ⁸ Yet, there are no published data on the RIs of thyrotropin and the other thyroid hormones in Africa, including Sudan. Research on RIs of thyrotropin and thyroid hormones is of paramount importance for health planners as well as for treating physicians. The current study was carried out to assess the RIs for TSH, T3 and T4 in Nyala, which is an iodine-deficient area. ⁸

Methods

A multi-stage stratified sampling method was used to select a representative sample of a Sudanese adult (>20 years of age) in Nyala in western Sudan in the Darfur region between January and June 2016. Nyala is a town 1700 km from Khartoum with 1 million inhabitants. After signing and informed consent, both males and females were included. Pregnant and lactating women, patients with any illness, thyroid disease, taking any medications with potential influences on the thyroid function, such as estrogen, amiodarone, anti-epileptic drugs, glucocorticoids and excess iodine ingestion were excluded. Patients with an enlarged thyroid, as examined clinically, were excluded. Overall, four medical officers, who were trained by an endocrinologist, conducted the physical examinations. Venous blood samples (1992) were collected and allowed to clot in plain tubes (Ningbo Greetmed Medical Instruments Co., Ltd, Ningbo, China), and the serum stored at (−20°C) until analyzed for the measurement of serum TSH, T3 and T4 using a radioimmunoassay gamma counter (Riostad, Germany) and kits provided by Beijing Isotope Nuclear Electronic Co., Beijing, China.

Statistics

Data were entered into a computer using IBM SPSS version 20 for Windows for data analyses. The outliers were detected (Tukey’s method) and were removed as previously described. ⁹ The Kolmogorov–Smirnov test was used to test for normality where TSH, T4 and T3 levels were not normally distributed, Figure 1. The median and 95.0% confidence interval were used to express the values for TSH, T4 and T3. The International Federation of Clinical Chemistry recommended/approved the nonparametric approach to RIs. ¹⁰ Nonparametric tests were used to compare TSH, T4 and T3 between male and females (Mann–Whitney U test) and between the different age groups (Kruskal–Wallis). A value of p < 0.05 was considered significant.

Figure 1.

Histograms of TSH (a), T4 (b) and T3 (c) samples in the reference population. The curves are the not normally distributed curves.

Ethics

The study received ethical clearance from the Research Board at the Sudan Federal Ministry of Health.

Results

The level of all the three investigated hormones was not normally distributed, Figure 1(a–c). The median (95% confidence intervals) of serum TSH, T4 and T3 levels was 1.2 (0.50–3.0) mIU/l, 111.0 (72.0–161.0) nmol/l and 1.5 (0.8–2.8) nmol/l respectively (Table 1). While the level of TSH was significantly higher in the age group between 31 and 40 years, both T4 and T3 levels showed a progressive increase with age. There was no significant difference in the TSH, T4 and T3 level when the RIs were compared between males and females (Table 1).

Table 1.

Median (95% confidence intervals) of serum TSH, T4 and T3 levels according to age and sex.

Age groups, in years	Number	TSH mIU/l	T4 nmol/l	T3 mIU/l
20–30	227	1.2 (0.5–3.2)	74.0 (53.0–83.0)	1.4 (0.8–2.5)
31–40	720	1.3 (0.5–3.0)	99.0 (85.0–113.0)	1.5 (0.8–2.7)
41–50	473	1.2 (0.5–3.0)	124.0 (114.0–135.0)	1.5 (0.9–2.8)
51–60	267	1.1 (0.5–2.6)	147.0 (137.0–162.0)	1.7 (1.0–3.0)
>60	66	0.8 (0.2–2.5)	179.0 (165.0–190.2)	1.8 (1.0–3.1)
p		<0.001	<0.001	<0.001
Sex
Male	778	1.2 (0.5–3.0)	113.0 (72.0–159.0)	1.5 (0.9–2.8)
Female	975	1.2 (0.5–3.0)	110.0 (69.0–164.0)	1.5 (0.8–2.9)
p		0.444	0.093	0.413
Total	1753	1.2 (0.50–3.0)	111.0 (72.0–161.0)	1.5 (0.8–2.8)

TSH, thyroid-stimulating hormone.

Discussion

The current study showed that the new established reference value of TSH (0.50–3.0), T4 (72.0–161.0) and T3 (0.8–2.8) level was lower than the TSH (0.7–5) and higher than T4 (60–160) and T3 (0.8–3.0) levels provided by the manufacturers. Following initiation of the program of Universal Salt Iodization in 1996, China has become iodine sufficient. ¹¹ Therefore it is valid to establish the reference values for the thyroid hormones in the different settings. We have recently shown that TSH RI was 0.776–4.550 among nonpregnant Sudanese women in Khartoum. ¹² To the best of our knowledge, this is the first published study to establish the RIs of thyroid hormones in Africa which was carried out in a previously iodine-deficient, but presently more than iodine adequate, area of Sudan.

Our upper limit of serum TSH level (3.0 mIU/l) was higher than TSH level that reported in Germany (2.12 mIU/l) ⁷ but lower than the TSH level reported in different settings; for example, the TSH level is 0.43–5.51 mIU/l in China, ⁴ 0.44–4.93 mIU/l in Japan, ⁵ and 0.34–5.1 mIU/l in Thailand. ⁶ It is worth mentioning the results of our study and the later ones should be compared carefully because different methods were used to investigate these hormones. While immunoassay was used in some reports, ⁴ liquid chromatography-tandem mass spectrometry is the recommended method for thyroid hormones determination. ¹³ The difference in TSH in the different settings could be explained by the differences of race and iodine nutritional status.

Our results showed a significant difference among all the age groups in all the investigated thyroid hormones (both T4 and T3 levels have shown a progressive increase with age). This is in line with Langén and colleagues’ findings of the significant differences in TSH levels between subgroups for age. ¹⁴ A previous study has shown a significant difference among Chinese adults in the different age groups in FT3 (which decreased with age), but not in TSH or in FT4. ⁴

Furthermore, another study has shown that the upper limit of the serum TSH values of Japanese patients increased with age. ⁵ On the other hand Völzke and colleagues, reported no significant difference in the levels in the different age groups in a previously iodine-deficient area. ⁷

In the current study there was no difference in the hormones between males and females. It has been shown that males and females had a significantly different TSH level. ¹⁴ Recently, Cai and colleagues reported that males had higher mean FT3, FT4 and T3 levels but a lower mean TSH level than females. ⁴ Recently, Hickman and colleagues reported no age or sex-related differences in TSH and FT4 levels. ¹⁵

Some limitations in the present study should be mentioned. Firstly, the clinical method rather than ultrasound was used to detect the thyroid volume and enlargement. Secondly, urinary iodine was not assessed; hence we failed to investigate the association of urinary iodine with the range intervals of thyroid hormones. Thirdly, thyroid antibodies, anti-thyroid peroxidase (anti-TPO) and anti-thyroglobulin (anti-TG) antibodies were not measured. Fourthly, other factors that could have influenced the thyroid hormones levels such as smoking, obesity were not investigated.^16,17 Unlike our study, the total levels of T3 and T4 rather than the free levels of these hormones are measured and taken as RIs. However, it is still reasonable to use our local RIs and further studies in different part of Sudan are needed. A survey is needed to assess whether most of the laboratories in Sudan are using the total T3 and total T4 rather than the FT3 and FT4.

Conclusion

Thus, the RIs for TSH, T4 and T3 in this setting were different from the levels provided by the manufacturers. The RIs were different in the different age groups.