HEMATOLOGICAL INDICES RELATED TO VITAMIN D DEFICIENCY IN OBESE CHILDREN

Abstract

Introduction

Vitamin D is involved in differentiation and induction of erythropoiesis in bone marrow cells.

Aim

We compared the serum 25(OH) vitamin D level in obese children versus control and found correlations between vitamin D level and hematological indices in obese children.

Materials and methods

25 overweight and obese patients and 15 normal weight children were enrolled in an observational study .

Results

In obese children, the serum level of 25(OH) vitamin D was significantly (p<0.04) lower (20.60 ng/mL) compared with the value from normal weight ones (25.63 ng/mL) and the body fat percentage BFP was higher. We found a positive correlation (r=0.44, p<0.05) between serum vitamin D and hemoglobin level and a negative one between serum vitamin D and the number of platelets (r= -0.43, p<0.05). Also, the serum iron was at the lower normal limit in the obese children and negatively correlated with the percent of the body fat (r= -0,62, p<0.05).

Conclusion

Obese children have vitamin D deficiency. The hemoglobin level and the number of platelets are correlated with the serum level of 25(OH) vitamin D. Supplements with vitamin D may have pleiotropic effects, including those on bone marrow activity.

INTRODUCTION

Obesity is a chronic pathology spread worldwide, which always raises the need for new strategies for prevention and medical treatment (1). Obesity in the pediatric patients has reached alarming rates worldwide, being a complex nutritional disorder in children (2). In Romania, one in four children were found to be either overweight or obese (3). Factors such as: eating habits, genetics, environment and lifestyle play an important role in the development and maintenance of obesity (4).

In recent years, there is a continuous effort to identify those clinical and paraclinical abnormal parameters in obese children and find ways to improve their health.

Vitamin D is known to be the main component of calcium homeostasis and bone mineralization (5), and appears to be associated with the prevention of chronic disease and modulation of immunity, control of insulin secretion and sensitivity, the regulation of cellular growth, and the differentiation and induction of erythropoiesis in bone marrow cells (6-9).

The inert prohormone of vitamin D is hydroxylated to 25-hydroxyvitamin D in the liver and then converted to its final form, calcitriol (1, 25 (OH)₂D), in the kidney cells (10). Serum 25(OH) vitamin D level is the best indicator to evaluate total vitamin D status in the body, as the metabolite is the main circulating form and it reflects both dietary intake and cutaneous synthesis (11). Calcitriol, the active form of vitamin D3, acts through vitamin D receptors (VDRs) located in a large variety of cell types (12).

Data from meta-analyses consistently support an inverse association of vitamin D levels with body weight. Experimental studies have demonstrated that low vitamin D may be implicated in adipose tissue differentiation and growth leading to obesity either by regulation of gene expression or through modulation of parathyroid hormone, calcium, and leptin (13). Important causes of hypovitaminosis D are a higher body fat mass and obesity (14).

It is well-documented that obese individuals express a chronic inflammatory status (15). Moreover, increased levels of inflammatory cytokines, such as tumor necrosis factor alpha (TNF-α) and interleukin (IL-6), have been found in subjects with low 25(OH) vitamin D levels (16). Interestingly, inflammation enhances oxidative stress, and this alteration contributes to the release of immature platelets from bone marrow to the circulatory system. Platelet number is known to reflect the whole activity of releasing cytokines and promoting thrombosis (17,18).

Epidemiological data suggest that vitamin D deficiency, defined as plasma 25(OH) vitamin D levels below 20 ng/mL (12), may become a global risk factor for several multifactorial diseases (19). A meta-analytical study have clearly shown a bidirectional association between vitamin D deficiency and anemia risk in both children and adults (20). Several reports have also revealed the unique role of the vitamin in erythropoiesis (21). However, it should be noted that the active form of vitamin D can affect erythropoiesis through stimulating the erythroid progenitor cells for proliferation and maturation, therefore calcitriol deficiency may affect the iron status (22).

Therefore, this study addressed the association between some haematological indices and 25(OH) vitamin D serum levels in obese pediatric subjects.

SUBJECTS AND METHODS

A total of 25 overweight and obese patients aged 8 to 17 years (with a mean age of 12.2 ± 3.02 years) and 15 normal weight patients (with a mean age of 11.8 ± 3.72 years) were enrolled in an observational study conducted during August 2017- November 2019. The pediatric patients were from Smart Nutrition Clinic, Bucharest and those with a medical history of taking corticosteroids, renal and/or endocrine disease, acute or chronic inflammation were excluded. All subjects were nonsmokers.

The study protocol was approved by the Ethical Commission of the Clinic and a written informed consent was obtained from each parent.

Anthropometric measurements, weight and height were assessed. The BMI was calculated as the ratio between weight (kg) divided by square height (m²). Overweight is defined as 85-95^th BMI percentile and obesity as ≥ 95^th BMI percentile (23). To measure the total weight, the body fat mass (BFM) and the body fat percentage (BFP), a body analyzer with bioimpedance Jawon IOI 353 was used. The unit of measurement of BFM is kg. Determination of the percentage of the body fat (PBF) is the total mass of fat divided by total body weight, all multiplied by 100.

The body analyzer IOI 353 performs measurements of the whole body but also of segments (lower limbs, upper limbs, abdomen) with three frequency channels (5, 50 and 250 kHz) and eight electrodes. The bioimpedance is based on a low intensity electric current that is sent through the body. Resistance between conductors helps to measure body fat between a pair of electrodes. Electric resistance varies between adipose tissue, muscle and skeleton. The fat-free tissue (muscle) is a good conductor because it contains a large amount of water (about 73%) and electrolytes, while the fat is anhydrous and a weak conductor of the electric current. Factors such as food intake, ingestion of fluids and exercise should be controlled because the level of hydration is a source of error in determining the passage of electricity. To start the determination some data must be specified such as: sex, height, date of birth, date of measurement.

Biochemical measurements

Fasting blood samples were collected and adequately stored until measurement. Serum parameters were analyzed: 25(OH) vitamin D, iron, hemoglobin and number of platelets (24). The method used for vitamin D determination was the immunochemical method with electrochemiluminescence detection. The method’s principle is the generation of highly reactive species resulting from stable precursors on the surface of an electrode. These highly reactive species react with each other producing light. The reference range of the total level of 25(OH) vitamin D is 30-100 ng/mL. Levels of 20-29 ng/mL define vitamin D insuficiency, and levels below 20 ng/mL, the deficiency of vitamin D. Recently the value attributed to vitamin D deficiency has changed to 32 ng/mL.

Statistical calculations were performed by using Microsoft Excel 2010.

RESULTS

In the obese children the important anthropometric parameters: the body weight, the body fat mass (BFM), the percent of body fat (PBF) and the Body mass index (BMI) were significantly increased, while the serum 25(OH) vitamin D was significantly decreased (p<0.04). The average value of the serum 25(OH) vitamin D was 20.6 ng/mL, which means insufficiency. High levels for serum triglycerides (p<0.05) and uric acid (p<0.05) were demonstrated in the obese children vs. control group (Table 1).

Table 1.

The average values, the standard deviation for the studied parameters and the test result for the obese versus normal weight children

Parameters	Control group	Obese	p
Average age (years)	12.2±3.02	11.8±3.72	Ns
Body weight (kg)	45.15±16.7	70.6±20.5	(p<0.001)
BFM (kg)	12.15±6.8	22.7±8.8	(p<0.004)
PBF (%)	21.2±8.20	31.27±4.69	(p<0.001)
BMI (kg/m2)	18.5±2.3	27.6±3.6	p<0.001)
25(OH) vitamin D (ng/mL)	25.63±6.09	20.59±7.14	(p<0.04)
Hb(g/dL)	13.8±5.53	13.3±4.61	Ns
PLT (x103)/mL	161.4±22.5	181.8±46.5	Ns
Serum iron (microg/dL)	84.4±9.5	75.4±8.7	Ns
Cholesterol (mg/dL)	162.46± 17.4	179.2± 34.3	Ns
Triglycerides (mg/dL)	68.8±21.7	85.8±38.7	<0.05
HDL-C (mg/dL)	57.3±14.9	50.4±12.1	Ns
LDL-C (mg/dL)	104.23±30.4	112.23±30.4	Ns
Uric Acid (mg/dL)	4.3 ± 0.72	5.4±1.44	<0.05
Glycemia (mg/dL)	80.2 ± 3.4	82.2±7.5	Ns
ALT(UI/L)	13.07 ± 2	18.8±7.5	<0.05
Creatinine (mg/dL)	0.65±0.15	0.68±0.16	Ns

Legend: BFM: body fat mass. PBF: percent body fat. BMI: body mass index. PLT: platetes number; Ns: nonsignificant statistical difference.

The serum iron was at the lower normal limit in the obese children and negatively correlated with the percent of the body fat (r= -0,62, p<0.05) (Fig. 1). We found a positive correlation (r=0.44, p<0.05) between serum vitamin D and hemoglobin level (Fig. 2) and a negative one between serum vitamin D and the number of platelets (r= - 0.43, p<0.05) (Fig. 3).

Figure 1.

Relation between PBF(percent body fat) and serum iron level in obese children.

Figure 2.

Relation between hemoglobin and 25(OH) vitamin D in obese children.

Figure 3.

Relation between the platelets number and 25(OH) vitamin D in obese children.

DISCUSSION

Vitamin D deficiency in childhood obesity is associated with enhanced systemic inflammation, which contributes to the development of insulin resistance and cardiovascular complications later in life (25-27). It is also known that vitamin D increases the sensitivity of bone marrow precursors to erythropoietin and that the systemic inflammation reduces hemathopoesis, acting contrary (28,29). Platelets are involved in hemostasis, thrombosis, immunity and inflammation and their aggregation contributes to atherothrombosis. Vitamin D, by controlling the platelet calcium homeostasis influences megakaryocytopoiesis and platelet activation (30).

Increasing with the severity of the obesity, a high prevalence of vitamin D deficiency in obese children was mentioned in various studies (14,19,20,25). In our study, the values of the studied parameters are shown in Table 1. By studying the percentage of the body fat (PBF) in the two groups of children, obese versus normal weight ones, it is obvious that the difference was strong, p<0.001. Opposite was observed for the serum vitamin D level (Table 1).

There can be multiple explanations for this. The bioavailability of vitamin D, which is a fat-soluble molecule, is decreased in the adipose tissue. The volumetric dilution of vitamin D in the large stores of adipose tissue has been demonstrated (31). Behavioural factors, such as reduced sun exposure and lower consumption of vitamin D fortified foods in obese children, who are sedentary and with an inadequate quality input of nutritional principles is another explanation (32).

In adipose tissue, vitamin D counteracts adipogenesis and changes the profile of adipokines. Therefore, there are current recommendations for the correction of vitamin D deficiency. Thus, in overweight and obese children, the higher dose of cholecalciferol is needed to achieve serum calcifediol targets (26).

Visceral adipose tissue has been shown to be more inflammatory active than subcutaneous adipose tissue. Strong negative correlation between visceral fat and serum 25(OH) vitamin D has been demonstrated (33). The systemic inflammatory reaction is the main cause of suboptimal serum iron level in obesity and the principal culprit is hepcidin. In chronic inflammation, the increased hepcidin contributes to the sequestration of iron in storages and macrophages. The hepatic protein, hepcidin binds itself to the only known iron export protein, ferroportin, inducing its internalization and degradation and thus limiting the amount of iron released in the plasma (28,34).

In our study, the serum iron value was in the normal range, but at the lower limit (Table 1). In the obese group, the serum iron levels were negatively correlated with the body fat percentage (Fig. 1).

In the last years there have been published studies that have shown that suboptimal vitamin D status was associated with increased risk of iron insufficiency. Thus, in a US cohort, Vitamin D deficiency was associated with anemia among African Americans (35). Pregnant adolescents with serum 25(OH) vitamin D concentrations <20 ng/mL) were 4–8 times more likely to be anemic than those with 25OHVitD ≥20 ng/mL (36). In a huge study conducted by the National Center for Health Statistics, Atlanta, on civilian population (31,509 participants), the suboptimal vitamin D status may contribute to anemia via inadequate erythropoietin production, and/or by promoting a persistent inflammatory state (37).

It is known that the serum iron level and the erythropoietin activity influence the hemoglobin level. In our study, the hemoglobin level was positively correlated with serum 25(OH) vitamin D in obese children (Fig 2). Vitamin D has multiple effects on the hematogenous marrow. Vitamin D increases the proliferation of erythrocyte precursors in the bone marrow by increasing their sensitivity to erythropoietin. On the one hand, severe vitamin D deficiency causes hyperparathyroidism, which leads to an inhibition of endogenous erythropoietin production (38). On the other hand, vitamin D has protective effect against anemia by decreasing the pro-inflammatory cytokines and hepcidin (28).

The platelet indices are affected in vitamin D deficiency. In a retrospective study done on almost 900 participants without chronic diseases, low levels of 25(OH) vitamin D were associated with an increased platelet count (39). A Korean study done on more than 3000 adults had shown that the platelet count and the medium platelet volume (MPV) were inversely associated with vitamin D levels. The increased platelet count, as well as low serum levels of active vitamin D metabolites caused high blood pressure and led to progression of metabolic syndrome (40).

We have observed that in our study, the platelet count was negatively correlated with the serum 25(OH) vitamin D in the obese children (Fig. 3).

The combined assessment of platelet indices with vitamin D could predict activation of inflammation more efficiently, in Sjögren’s Syndrome patients (41).

For the first time, Zupo et al. demonstrated that in obese adult subjects, the vitamin D deficiency is associated with a parallel increase in platelet number, which is a leading mechanism to a greater cardiovascular risk (27). Vitamin D deficiency is also linked to increased production and release of inflammatory cytokines that can have a direct negative effect on the cardiac system (42).

At the same time, there are controversial studies showing that vitamin D deficiency can cause autoimmune disorders, including immune-mediated thrombocytopenia. In that case, the number of platelets could be improved by vitamin D administration (43).

There are some limitations of this study, because the number of children enrolled is low. Future clinical study will demonstrate that vitamin D3 has beneficial effects on bone marrow activity.

In conclusion, this study has demonstrated that in obese children, the serum level of vitamin D is lower compared to healthy, normal weight children. The deficit of the vitamin has hematogenous consequences like anemia and increased platelet count. These modifications act in cluster with dyslipidemia and insulin resistance, increasing the risk for cardiovascular disease, later, in young adulthood. The information may be helpful in implementation of prophylaxis of these disorders through the administration of vitamin D.

Conflict of interest

The authors declare that they have no conflict of interest.