The effect of zinc supplementation on anthropometric measurements in healthy children over two years: a systematic review and meta-analysis

Background Zinc deficiency is one of the most important micronutrient deficiencies in children that can affect the children’s growth pattern. In this regard, different studies were conducted to assess the effect of zinc supplementation on growth patterns in healthy children. To the best of our knowledge, no systematic review has summarized the results of these studies. So, in the present study, we systematically reviewed the result of the studies that assessed the effect of zinc supplementation on anthropometric parameters in healthy, over 2-year-old children. Methods A systematic search was carried out in PubMed, Scopus, and Web of Science from inception to November 2021. Data were pooled using the random-effects method and were expressed as weighted mean difference (WMD) and 95% confidence intervals (CI). Results The pooled results of eight studies, including 1586 participants, showed that zinc supplementation significantly increases height [(WMD): 0.9, 95% CI: (0.27, 1.52), p < 0.001], weight [(WMD): 0.51, 95% CI: (0.06, 0.97), p < 0.001], height for age (HAZ) [(WMD): 0.07, 95% CI: (0.03, 0.10), p < 0.001]. Also, meta-regression analysis did not reveal any significant association between dose and duration of intervention and anthropometric parameters. Conclusion The present study demonstrates the beneficial effects of zinc supplementation on weight, height, and HAZ. Supplementary Information The online version contains supplementary material available at 10.1186/s12887-023-04249-x.


Background
Child growth is a vital indicator of health status in the human life cycle.Different factors including childhood infections, child undernutrition, and food insecurity could affect the child's growth [1].Child undernutrition is the underlying cause of three million annual deaths worldwide [2].It includes stunting, wasting, and essential micronutrient deficiencies.Failure to properly treat nutritional deficiencies, or even subclinical malnutrition, can have detrimental effects on an individual's health and the nation's overall economic growth [3].
One of the most important micronutrient deficiencies especially in children is attributed to zinc deficiency [4].Zinc insufficiency is a huge issue in terms of public health that affects over two billion people all over the world [5,6].In Iran, it has been estimated that 10% of people suffer from zinc deficiency [7,8].It has been demonstrated that zinc deficiencies cause children to have slower rates of growth.This issue seems more important in children over two years.Because in children under two years of age, maternal breastfeeding, could provide the children`s need for zinc [9].In addition, the body store of zinc from in-utero development could partly compensate for the body's need for zinc [10].
In this context, the World Health Organization (WHO) and the United Nations Children's Fund are presently advocating for the use of a micronutrient powder formulation that contains 4.1-5 mg of zinc together with 14 other micronutrients [11].In terms of zinc supplementation, some meta-analysis studies were conducted to assess its effect on the growth of children.Brown et al., systematically reviewed the effect of zinc supplementation in prepubertal children [12].Imdad et al. assessed the effect of zinc supplementation on the growth pattern of under five-year-old children in developing countries [13].Liu et al. reviewed the effect of zinc supplementation in children with growth retardation.Different original publications also assessed the effect of zinc supplementation on the growth pattern of healthy children and provided mixed results.To the best of our knowledge, there is no systematic review study that summarizes the result of these studies.So, the current meta-analysis was carried out to compile a summary of the evidence that is currently available on the influence of zinc supplementation on anthropometric parameters in healthy children.

Materials and methods
The Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) procedure served as the basis for the planning, execution, and reporting of this work [14].

Search strategy
Online databases including Scopus, Web of Science, and PubMed were searched to find all the relevant clinical trials up to November 2021.The search strategy for each database is provided in Table S1.The following search terms were used: Population: children; Intervention: Zinc, outcome: Anthropometric Measurements.In addition, we conducted a hand search of the reference lists of the relevant papers and review papers to include any other studies that might be suitable.

Inclusion criteria
Clinical trials were included in the present systematic review and meta-analysis if they fulfilled the following inclusion criteria: (a) were randomized with either parallel or crossover designs in healthy children over 2 years; (b) reported anthropometric indices before and after intervention in each group; (c) compared zinc supplementation with the placebo.Studies were excluded if they: (a) do not use zinc alone; (b) did not provide effects sizes on anthropometric factors before and after the trial; (c): Abstracts, conference proceedings or book chapters, and unpublished studies.
Two investigators (ED and FA) selected eligible articles separately by reading titles, abstracts, and whenever required the full text of the publications.Any disagreements in this regard were resolved through discussion with the third researcher (ZF).

Data extraction
Data extraction was done by two researchers independently (VM and AS).The disagreements were settled through discussion and by agreeing with the assistance of a third reviewer (ZN).The following information was extracted: name of the first author, publication year, individuals' characteristics (mean age), study design, sample size, type of zinc prescribed, the dosage of zinc, unit, and duration of intervention.The corrected mean changes and standard deviations of anthropometric measurements were taken during the study for both the intervention group and the control group, as well as the variables that were considered to be confounding.

Risk of bias assessment
The Cochrane quality assessment tool was used for assessing the risk of bias [15].Two reviewers independently assessed the risk of bias (VM and AS) and the risk of bias was divided into high risk, low risk, and unclear risk.

Certainty assessment
The overall certainty of evidence across the studies was graded according to the Grading of Recommendations, Assessment, Development, and Evaluations (GRADE) guideline, and the evidence was divided into high, moderate, low, and extremely low categories [16].

Statistical analysis
The overall effect sizes were calculated using the mean changes in anthropometric measurements and their respective standard deviations (SD).In studies that reported the SEM, the SD was determined using the following formula: SD = SEM × √n, in which the n is the sample size.The I 2 statistic and Cochrane's Q test were used to determine the presence of heterogeneity.Significant heterogeneity was determined if the I 2 value was > 50% or P < 0.05 [17].To find probable sources of heterogeneity, subgroup analyses were performed according to the predefined variables including gender, country region (Asian or Non-Asian), sample size, and duration of studies (> 24,<24).A sensitivity analysis was performed to determine whether or not the overall effect size was dependent on a specific study.The formal test developed by Begg and Egger looked into the possibility of publication bias.Stata, version 14 was utilized for the meta-analysis, and a P-value < 0.05 was considered a significant level.

Study characteristics
The characteristics of included RCTs are illustrated in Table 1.Two studies were performed on boys and two on girls and four on both genders.The sample size of included RCTs varied from 46 to 804 participants, resulting in a total sample size of 1586.The dosage of zinc Fig. 1 Flowchart of study selection for inclusion trials in the systematic review supplements varied from 5 to 15 mg/day, and the duration of intervention ranged from 6 to 28 weeks.

Risk of bias result
The result of the risk of bias is shown in Table 2.Only two studies [36,37] could be considered high-quality studies, two RCTs [42,43] were of moderate quality and others had low-quality.

Grading of evidence
The GRADE protocol was used to assess the certainty of the evidence (Table 3).Accordingly, studies investigating the effects of zinc supplementation on weight and height were regarded as moderate quality due to the high heterogeneity between studies.Moreover, the effects of zinc supplementation on HAZ, and WAZ had moderate quality evidence.HAZ had low heterogeneity and WAZ has high heterogeneity.

Findings from the meta-analysis
Overall, eight RCTs were included in the meta-analysis.These trials had a total sample size of 1586 individuals aged two years and over.

The effect of zinc supplementation on weight
The effects of zinc supplementation on weight were evaluated in seven research, including 1488 people [35,36,[38][39][40][41][42].Considering the presence of high heterogeneity between studies (I 2 = 95.9,P < 0.001), the random-effect model was used.The result indicated that zinc supplementation had a significant impact on weight [(WMD): 0.51, 95% CI: (0.06, 0.97), p < 0.001] (Fig. 2-B).The heterogeneity might be explained by the location of the study, duration of intervention, gender, and sample size.The subgroup analysis showed that the studies conducted in Asian countries (I 2 = 0.0%, p = 0.545) and gender (boy, I 2 = 40.4p = 0.195) were the potential sources of heterogeneity (Table 4).Each study was assessed for risk of bias using the Cochrane Risk of Bias Assessment tool.Each domain was scored as "high risk" if it contained methodological flaws that may have affected the results, "low risk" if the flaw was deemed inconsequential, and "unclear risk" if information was insufficient to determine.If a study got "low risk" for all domains, it considered as a high-quality study with totally low risk of bias

The effect of zinc supplementation on height
The effects of zinc supplementation on height were evaluated in seven research, including 1488 people [35,36,[38][39][40][41][42].Considering the presence of high heterogeneity (I 2 = 93.9,P < 0.001), the random effect model was used.The result showed that zinc supplementation had a significant effect on height [(WMD): 0.9, 95% CI: (0.27,1.52), p < 0.001] (Fig. 2-A).As shown in Table 4, the heterogeneity might be explained by the location of the study, duration of intervention, gender, and sample size.The subgroup analysis showed that the intervention duration of ≥ 24 weeks (I 2 = 0.0%, p = 0.975) was the potential source of heterogeneity.An intervention duration of greater than 24 weeks (WMD 0.78; 95% CI: 0.57 to 0.99, p < 0.000) significantly increased height compared with a shorter intervention duration (WMD: 0.13; 95% CI: 0.10 to 0.16, p = 0.975).Moreover, we found a significant increase in height with zinc supplementation in RCTs with a sample size of less than 100 (Table 4).

The effect of zinc supplementation on WAZ
The effects of zinc supplementation on WAZ were evaluated in four studies (with five effect sizes), including 581 subjects [35][36][37]40].There was high heterogeneity between studies (I 2 = 72.4,P = 0.006). Te result of the random-effect model showed that zinc supplementation had a significant impact on WAZ [(WMD): 0.06, 95% CI: (-0.03, 0.15), p-value = 0.18] (Fig. 2-C).The heterogeneity might be explained by the duration of the intervention, gender, and sample size.The subgroup analysis showed that all subgroups were the potential sources of heterogeneity and we found a significant increase in WAZ with zinc supplementation with duration 24≥, and among non-Asian people (Table 4).

Sensitivity analysis
The results of the sensitivity analysis revealed that the magnitude of the overall effect regarding the association between zinc supplementation and height, WAZ, and HAZ did not depend on a single study.However, in terms of weight, the total impact size depended on some studies [35][36][37][38][39][40][41][42].As a result, when those studies were disregarded, a substantial positive correlation between zinc supplementation and weight was discovered.

Publication bias
As can be seen on the funnel plot (Figure S1 A-D), there were no signs of publication bias in the studies that evaluated the effect of zinc supplementation on weight, height, WAZ, and HAZ.

Meta-regression analysis
Meta-regression was used to investigate the potential linear association between dose and duration of intervention, with absolute changes in anthropometric indices.Accordingly, meta-regression analysis did not reveal any significant association between the dose of supplementation with weight changes (p linearity = 0.

Discussion
Child malnutrition is one of the main public health challenges worldwide and about 45% of under 5-year-old children's death is attributed to undernutrition [44].In this regard, zinc deficiency is considered one of the most important causes of morbidity [45].So, different studies were conducted to assess the effect of zinc supplementation on child growth in healthy children over two years of age.The meta-analysis of these studies showed that zinc supplementation had a significant effect on healthy children's weight and height.Previous meta-analysis in prepubertal children showed a significant, positive effect of zinc supplementation on height and weight [12].In under-five-year-old children in developing countries, Imdad et al. demonstrated the significant effect of zinc supplementation on linear growth [13].In children with growth retardation, Liu et al. showed a significant effect of zinc supplementation on height, and weight [10].However, Ramakrishnan et al. did not show a positive effect of zinc supplementation on weight and height in under five-year-old children [46].The differences between the results of the various meta-analysis may be partly related to the differences in the age group of children.It has been  shown that zinc supplementation had a more positive effect on over two years of old children compared with infant.Moreover, the health status of children can affect the result.For example, Liu et al. conducted a meta-analysis study on children with growth retardation.However, in the present meta-analysis, we included the paper with healthy subjects.The positive effect of zinc supplementation on weight and height may be related to the effect of zinc on the metabolism of growth hormones (GH).Zinc had a positive effect on the secretion and sensitivity of GH [47][48][49].Moreover, zinc had an important role in the binding of GH to its receptors [50].In addition, zinc regulates the expression of GH receptor and insulin-like growth factor 1 (IGF1) genes in the liver [51].Besides, zinc supplementation increases the insulin-like growth factor-1, and insulin-like growth factor binding protein-3 in healthy children [52].
In the present study, we indicated the significant effect of zinc supplementation on HAZ but not WAZ of healthy children over two-year-old.Conversely, Liu et al. reported the significant effect of zinc supplementation on WAZ in children over two years.In the present systematic review, we included studies with healthy participants, however, Liu et al. conducted a systematic review on studies with children with retarded growth.
In subgroup analyses, we found that intervention duration, study location, gender, and sample size were significant sources of heterogeneity.A previous meta-analysis study indicated that a longer dose of zinc intervention resulted in a more effect on the risk factors [53].In addition, gender differences have been observed in the association of zinc and other diseases [54].Moreover, the higher prevalence of zinc deficiency in developing countries may justify the effect of study location on heterogeneity [55].Besides, a previous study showed that studies with small sample sizes are more heterogeneous than large ones [56].

Limitations
In the current meta-analysis, some potential limitations should be addressed.The publication bias could not be fully excluded.There was considerable heterogeneity between the included studies.Although we performed a subgroup meta-analysis for some factors, not all sources of statistical heterogeneity were recognized.In addition, not all included studies are methodologically sound.The potential methodological problems of studies were as follow selection bias, performance bias, detection bias, and reporting bias.

Conclusion
Altogether, the results of this meta-analysis are in favor of the effect of zinc supplementation on weight, WAZ, and height in healthy under-two-year-old children.Considering the limitations of the included studies, these findings should be confirmed by more high-quality randomized clinical trials that focus on the long-term effects of zinc supplementation on anthropometric indices.

Table 1
Characteristic of included studies in meta-analysis

Table 2
Risk of bias assessment for randomized clinical trials included in the current meta-analysis on the effect of zinc supplementation on Anthropometric factors in Healthy children over 2 years1 L: low risk H: high risk U: unclear 1

Table 3
GRADE profile of the effect of zinc supplementation on Anthropometric factors in Healthy children over 2 years WAZ: weight for age; HAZ: height for age a The test for heterogeneity is significant, and the I 2 is high, 93.9% b The test for heterogeneity is significant, and the I 2 is high, 95.9% c The test for heterogeneity is significant, and the I 2 is moderate, 72.4% d The test for heterogeneity is not significant, and the I 2 is low, 33.1% Fig. 2 Forest plot for; A: weight, B: height, C: weight for age z-score (WAZ), D: height for age z-score (HAZ).WMD: weighted mean difference

Table 4
Subgroup analyses the effect of zinc supplementation on Anthropometric factors in Healthy children over 2 years