Assessment of the iodine deficiency severity in the population by serum thyrotropin levels in newborns

Results. During the analyzed period, there was a positive trend: the median ioduria in pregnant women increased from 89.8 μg/l in 2015 to 140.1 μg/l in 2020, the level of neonatal hyperthyrotropinemia above 5 μIU/l decreased to 5 % in 2020 (P < 0.001), there was an upward trend in the median ioduria among prepubertal children (60.4 ± 9.3 μg/l). The frequency of goiter was different and accounted for 25.3 % among children living in the mountainous area and it was much lower among children living in the plain areas and in Chernivtsi (15.6 % and 13.1 %, respectively, P < 0.05). Thyromegaly among children of the reference group occurred in 15.1 %. 12.3 % of children were characterized by TSH in the range of age standards, which were shifted towards its increase (4.23–5.00 μIU/l).

The problem of iodine deficiency (ID) and its adverse effects on the Ukrainian population, especially on children, adolescents, pregnant and lactating women, is still relevant [1]. Iodine is an important micronutrient necessary for the normal functioning of the thyroid gland, growth and development. Sub-optimal iodine intake causes inadequate secretion of thyroid hormones, which leads to a spectrum of adverse outcomes, collectively referred to as iodine deficiency disorders (IDD) [2]. The most detrimental effects of IDD are observed in the antenatal period and during the first two years of life [3]. Severe iodine deficiency during this period increases the risk of stillbirth, birth defects, perinatal and infant mortality and impairs physical and cognitive development. Brain damage to the fetus and young children is often irreversible, causing mental retardation and reduced school performance [4]. The term IDD was introduced by the World Health Organization (WHO) in 1983. It has become clear that iodine deficiency causes not only thyroid disease, but also many other disorders caused by lack of thyroid regulation [5]. Iodine deficiency in the environment has been reported in large areas of the earth [6,7]. The iodine deficient territories in the world are in all continents [8][9][10][11]. In 1985, the International Council for the Control of Iodine Deficiency Diseases (ICCIDD) was established, which works closely with the WHO and the UNICEF. In 1990, IDD was recognized as an important medical and social problem in 118 countries (WHO/UNICEF/ICCIDD), and a historic resolution called for the elimination of iodine deficiency as a public health problem by 2000 [8]. To eliminate the iodine deficiency in Ukraine, the orders of the Ministry of Health of Ukraine No. 58 dated May 24, 2001 "On introduction of priority measures to overcome iodine deficiency in the population of Ukraine" and No. 67 dated October 25 of the same year "On additional measures to overcome iodine deficiency in the population of Ukraine", and the resolution of the Cabinet of Ministers of Ukraine dated September 26, 2002 "About the statement of the State Program for prevention of iodine insufficiency in the population during 2002-2005" were issued [9]. An important area of this program was the need to monitor iodine supply in all regions of Ukraine. A study on urinary iodine excretion showed generally low degree of iodine deficiency (median ioduria -90.1 μg/l) in Ukraine between 2002-2003 [10]. However, significant differences were found for various clusters (settlements). In particular, the region of Northern Bukovina, which is geographically divided into plain and mountainous zones, belongs to the areas with mild (plain zone) and medium (mountainous zone) degree of iodine deficiency. A largescale national study of iodine deficiency in 2001-2003, which covered all regions of Ukraine, found that there have been significant positive changes in improving the iodine supply among the population. However, the problem of the prophylactic measures for IDD remains unresolved in Ukraine generally. To date, Ukraine lacks a legal framework for the organization of mass iodine prophylaxis by general iodization of table salt, which does not allow achieving a significant reduction in iodine deficiency diseases in children and adults. In recent years, high-risk groups of IDD have been identified (pregnant women, breast-feeding mothers, children of the first two years of life), for whom insufficient iodine intake is especially dangerous due to the possibility of irreversible brain disorders development in a child. For these categories of the population, additional iodine medication intake is obligatory [11]. Since the child's brain and cognitive functions develop most intensively in the first years after birth, pediatricians should competently plan the tactics of postnatal iodine prophylaxis for all young children without exception. In order to assess the degree of iodine deficiency in new socio-economic conditions in Ukraine, in particular in Northern Bukovina, the problem of iodine supply must be addressed according to the WHO criteria, especially the level of neonatal hyperthyrotropinemia (above 5 μIU/l) and goiter in children groups.

Aim
To assess the state of iodine status of the Northern Bukovina population by the level of thyrotropin (TSH) in newborns during the years 2015-2020.

Materials and methods
An analysis of selective screening to determine the blood level of TSH in newborns during 2015-2020 (a total of 47888 results) was performed. To determine the correlations, the results of medical and biological monitoring during this period were used (frequency of iodine-deficient goitre and median ioduria in prepubertal children). For this purpose, a survey, ultrasonographic and hormonal study among 199 prepubertal children living in Northern Bukovina (100 children from the zone of mild iodine deficiency and 99 children from the zone of moderate iodine deficiency) were conducted. Inclusion criteria to enroll prepubertal children in the study: residence in Northern Bukovina from birth, prepubertal age, no diseases of the endocrine system, no use of iodine and thyroid hormones. Exclusion criteria from the study: moving to another place of residence, other thyroid diseases.
Determination of TSH levels in newborns was performed on the 4 th day using a test kit for quantitative determination of TSH in samples of dry blood spots on filter paper (Neonatal hTSH EIA, 960 wells, Neonatal Card (903 Body Fluid Collection Paper) manufactured by Ani Labsystems Ltd. Oy (Vantaa, Finland). Assessment of the morphofunctional state of the thyroid gland in children before puberty was performed according to the indicators of visual-manual examination and ultrasound examination (USE) on the Scanner -100 using a linear sensor with a frequency of 7.5 MHz. According to the parameters obtained via USE, the volume of each thyroid lobe was calculated by the formula: maximum thickness /cm/ × width /cm/ × length /cm/ × 0.478 and compared with the recommended standards of thyroid volumes obtained during the examination of children in iodine-supplied regions of Europe, taking into account age, sex and body area.
The functional state of the pituitary-thyroid system was assessed by clinical symptoms using screening integrated tables, as well as standard kits for RIA Т 3 -"rio-T 3 -PG, Т 4 -"rio-T 4 -PG, (Belarus); the concentration of TSH was determined by standard test kits (Mallinckrodt, Germany). Blood sampling for hormonal examination was implemented using venipuncture of the ulnar vein. Sample counting and result processing were performed using an automatic counter with a microprocessor and computer data processing.
Taking into account that the concentration of iodine in a single portion of urine is well correlated with the level of iodine in daily urine, we took children urine into test tubes of 5 ml of urine at a time. The iodine concentration was determined by cerium-arsenite method with wet ashing of the samples. Based on the data obtained, the median ioduria was calculated, which was expressed in μg of iodine per 1 liter of urine.
The results obtained were processed by the method of statistical variation and correlation analysis. Statistical processing included the calculation of the arithmetic mean for each indicator (M), the standard deviation (σ). The statistical significance of the results was evaluated via determining the mean error of the arithmetic mean (m), the significance of differences in the mean values -by t-Studen's test. A computer program "Biostat" was used for statistical processing.

Results
The survey-based results showed that the proportion of families using iodized salt in food was only about 20 % in Ukraine, the prevalence of endemic goiter among schoolchildren remained high and averaged 41 %, reaching 70 % in some regions. In the structure of thyroid pathology in the population of our country, IDD occupied a leading position -95 % among children. As the main goal of iodine prophylaxis in childhood, the majority (69.4 %) of children chose the prevention of thyroid disease. Only 25 (12.5 %) children correctly indicated the priority task of adequate iodine consumption -prevention of mental deficiency. The study found that the median ioduria in pregnant women did not reach 150 μg/l recommended by the WHO for this population, but during the analyzed period there was a po sitive trend, as evidenced by the improvement in the median ioduria, which increased from 89.8 μg/l in 2015 to 140.1 μg/l in 2020 (Fig. 1). Determination of TSH levels on the 4th day after birth was performed in 47888 cases ( Table 1).
The level of neonatal hyperthyrotropinemia above 5 μIU/l averaged 18.3 %. In the process of screening, there was a statistically significant decrease in the frequency of neonatal TSH above 5 μIU/l to 5 % in 2020 (P < 0.001) characterizing the whole region of Northern Bukovina as the area with mild ID. However, when distributed relative to living in the mountainous or plain area, neonatal TSH levels above 5 μIU/l were distributed as follows (Fig. 2): in the mountainous area, the number of these cases decreased from 38.8 % in 2015 to 19.6 % in 2020, while in the plains -from 32.6 % in 2015 to 17.2 %. Therefore, according to the WHO criteria, these areas belong to the zone with mild iodine deficiency. Since the criteria for the severity of iodine deficiency, defined by the WHO, also include the incidence of goiter and thyromegaly in prepubertal children, we also analyzed these indicators. Determination of the urinary iodine concentration in prepubertal children from different areas of residence showed that in most children, the excretion of iodine in the urine was reduced, but over the past 6 years, there was a tendency to its increase (Fig. 3).
The median ioduria in the examined population as a whole was 60.4 ± 9.3 μg/l, which according to the WHO criteria indicated the presence of mild iodine deficiency. A more detailed analysis showed that among children living in the plains, 14.2 % had ioduria greater than 100 μg/l, i.e. normal iodine supply, while 4.1 % of children had ioduria less than 20 μg/l corresponding to severe iodine insufficiency. The children living in the plains and the children of Chernivtsi demonstrated a mild iodine deficiency (median ioduria was 68.4 ± 3.1 μg/l and 69.1 ± 1.9 μg/l, respectively), and the children of mountainous areasiodine deficiency of moderate severity (median ioduria was 28.9 ± 1.9 μg/l). Ioduria greater than 100 μg/l was found in  The survey findings showed a significant incidence of goiter among children of Bukovіna -17.6 %. In different climatic and geographical zones, the frequency of goiter varied in children living in the mountainous zone (25.3 %) being much lower among children living in the plains and in Chernivtsi (15.6 % and 13.1 %, respectively, P < 0.05). Thyromegaly among children of the reference group occurred in 15.1 %.
As the main characteristic of the thyroid status in the children of prepubertal age, the level of TSH was cho-sen as the most sensitive indicator of the thyroid hormone deficiency presence ( Table 2).
According to this indicator, the examined group of children had no abnormalities in the functional state of the thyroid gland. However, 12.3 % of the children were characterized by TSH in the range of age standards, which was shifted towards its increase (4.23-5.00 μIU/l). When using the indicator TSH/T 4 , we found signs of subclinical hypothyroidism in 2.1 % of children (this indicator was 0.115-0.143). The evaluation of all studied criteria is given in Table 3.

Original research
Correlation analysis between the neonatal TSH above 5 μMo/l frequency with the frequency of iodine deficiency goiter in prepupertal children showed a statistically significant high positive relationship (r = 0.89, P < 0.01). It is known that the main criterion for the state of iodine saturation in the region is the median ioduria.
Thus, analyzing the nature of iodine deficiency in the surveyed areas based on the WHO criteria, we can conclude a slight degree of iodine deficiency in Northern Bukovіna according to the median ioduria in pregnant women and prepupertal children due to the level of neonatal hyperthyrotropinemia above 5 μIU/l/. At the same time, the determination of the goiter frequency by palpation and thyromegaly ultrasonographically indicated the presence of moderate iodine deficiency. Therefore, the incidence of neonatal hyperthyrotropinemia above 5 μIU/l more reflects the presence of iodine deficiency in the cohort of pregnant women than in the general population, which dictates the need to reassess the importance of this indicator as the criterion for iodine deficiency in the general population. However, despite all preventive measures carried out in the region, indicators of neonatal hyperthyrotropinemia above 5 μIU/l occurred with a frequency above the norm of 3 %, recommended by the WHO for iodine-saturated areas. This characterizes the lack of effectiveness of the prevention programs that depends largely on women's awareness.

Discussion
The high frequency of thyroid diseases depends on many reasons, among which the most important are the cessation of centralized iodine prophylaxis, man-made pollution, negative changes in the nutritional structure among the vast majority of the population in the new socio-economic conditions [12]. Prevention of IDD, such as goiter, decreased thyroid function, mental dysfunction in children and adults, congenital anomalies, cretinism, delayed physical and sexual development, miscarriage, infertility, remains an acute problem [13]. According to the WHO, pathological conditions caused by iodine deficiency rank third in the list of the most common non-communicable diseases. In various countries, programs of prevention and elimination of IDD are being implemented at the state level [14,15]. The main coordinators of these programs are the respective national committees, coordinating councils, WHO, UNICEF, ICCIDD which defined criteria of iodine deficiency: goiter frequency according to palpation and thyromegaly according to ultrasound, median urinary iodine excretion, TSH level, serum thyroglobulin level. A separate risk group is pregnant women. The number of pregnant women with this pathology is growing every year, which can be attributed to the lack of timely prevention of existing iodine deficiency in our country.
Although some authors give conflicting values regarding the level of neonatal TSH, this indicator is a common criterion for iodine deficiency. Contradictory data from various authors can be associated with many technical issues that remain unresolved regarding the use of TSH screening in newborns to monitor iodine status, making it questionable as a sensitive and reliable quantitative tool. Additional research is needed to address these issues [16]. The aim of the E. Cortés-Castell et al. study [17] was to analyze possible risk factors for elevated TSH level that may distort its validity as the marker of iodine status. The clinical significance of this issue is that the associated with iodine deficiency factors are known, iodine supplements can be introduced into risk groups both during pregnancy and for newborns [18].
It is probably important to take into account the birth date and birth weight, as well as the nutritional status of a pregnant woman [19]. We support the authors' view that the urinary iodine concentration directly reflects the iodine intake with food and remains the most common indicator used worldwide to assess iodine levels in the population, that are confirmed by studies. Clinical and laboratory studies of thyroid function (including serum concentrations of pituitary hormone thyrotropin) are problematic. Even in regions with severe iodine deficiency, there are a large number of individual changes in the ability of the thyroid gland to adapt. In most institutions and population subgroups, except for newborns, thyroid function tests are not considered as sensitive indicators of the population iodine status.
Obviously, in order to assess the degree of iodine deficiency burden, in addition to the median ioduria, it is necessary to analyze the frequency distribution of urinary iodine concentration in a specific group of subjects. We did not find a significant difference in the urinary iodine concentration in the children depending on age and sex. The inconsistency of certain indicators that serve as criteria for iodine deficiency indicated that the endemic goiter burden in the territory of Northern Bukovіna cannot be explained only by iodine deficiency. It is possible that other strumogenic factors are involved in the goiter occurrence. These can be xenobiotics, imbalance of other micro-and macroelements, etc. Northern Bukovіna is represented by different climatic and geographical zones (mountainous, foothills, plains), being the largest endemic region of Ukraine in terms of goiter prevalence in children, that can be explained by the weakening of control over iodine prophylaxis among children in the region [20].
Our study had some limitations. We had no data on TSH levels and thyroid volume in pregnant women. Not all factors were taken into account (birth weight, whether thyroid hormones were used during pregnancy, mode of delivery), as well as other possible factors that may affect TSH levels in newborns.