Zinc Levels and Birth Weight in Pregnant Women with Gestational Diabetes Mellitus: A Matched Cohort Study in China

Abstract

Zinc (Zn) is an essential element for several biological functions, including protein synthesis, cellular division, and nucleic acid metabolism (1). Zn deficiency in early life can result in growth retardation and cognitive impairment (2). It has been hypothesized that maternal Zn deficiency during pregnancy may compromise fetal development. Animal studies have confirmed that maternal Zn deficiency leads to adverse birth outcomes such as intrauterine growth retardation and central nervous system malformation (3). However, the association between maternal Zn status and birth outcomes in human populations is not yet clear (4,5). Zn deficiency during pregnancy is a global public health issue and it is estimated that over 80% of pregnant women have inadequate Zn intake around the world (6).

The effects of maternal Zn status on birth outcomes may be complicated by gestational diabetes mellitus (GDM) because of the potential relations between Zn and GDM. It has been established that GDM can alter the placental transport of various chemicals including Zn by changing gene expression (7,8), suggesting that identical maternal Zn levels may impact on fetal growth differently in women with GDM compared with those without GDM. Meanwhile, Zn is a component of insulin crystal (9) and both experimental and epidemiological studies suggest that Zn intake may protect against type 2 diabetes (10). Human studies, including 1 prospective study and 2 cross-sectional studies, supported this finding that higher Zn intake was associated with lower prevalence of type 2 diabetes (11-13). In a randomized clinical trial, Zn supplementation in pregnant women with GDM resulted in better metabolic profiles, such as reduced plasma glucose and serum insulin level, than women not receiving Zn supplementation (14).

GDM also poses a burden on the health of mothers and their offspring. The reported prevalence of GDM varies across different populations, with the highest prevalence of 25% in Singapore (15). GDM is associated with adverse health outcomes in both mothers and offspring (15-18). Women with GDM have a higher risk of type 2 diabetes in later life, prolonged labor, and delivering a baby with fetal macrosomia, that is a large baby (birth weight > 4000 g) (16). Excessive fetal growth increases the risk of both immediate and subsequent adverse outcomes for the child, including premature birth, shoulder dystocia, obesity, type 2 diabetes, and cardiovascular diseases (19-21).

Given the complex relationships among maternal Zn level, GDM, and birth outcomes, we hypothesized that the association between maternal Zn level and birth weight of offspring differs between women with and without GDM. We conducted a matched cohort study of GDM in Taiyuan, China, where pregnant women were generally characterized by inadequate Zn intake (22). We evaluated the associations between Zn levels (mg/L) measured in maternal blood (MB) and cord blood (CB) at delivery and birth weight of offspring born to pregnant women with and without GDM. We used MB and CB Zn levels because they can represent the circulating Zn in the body that links mother and fetus.

Method

Study population

We conducted a matched cohort study of pregnant women with or without GDM enrolled from the first affiliated hospital of Shanxi Medical University of Taiyuan, China, between 2012 and 2016 and evaluated the birth outcomes of the offspring in these women. Pregnant women who came to the hospital for delivery, aged 18 years and older, with a gestational age of 20 weeks or more, and without mental illness were eligible for the study. A total of 12 024 eligible women were contacted for participation. In total, 10 320 agreed and completed the in-person interview. After exclusion of women who had stillbirths or birth defects, who had multiple births, or who did not donate MB and CB samples, 6210 women were included in the study. Demographic information, health behaviors, and medical history were collected through in-person interview. Pregnancy complications and birth outcomes including birth weight of the offspring were abstracted from medical records.

A total of 776 women with GDM were identified within this cohort (prevalence 12.4%). GDM diagnosis followed standard clinical guidelines (23). The diagnosis was based on a 75-g oral glucose tolerance test during gestational weeks 24 and 28. Women who met 1 or more of the following criteria were diagnosed with GDM: (1) fasting blood glucose >5.1 mmol/L, (2) 1-h blood glucose >10.0 mmol/L, or (3) 2-h blood glucose >8.5 mmol/L. Only 5 women with GDM reported medical treatment; the others followed diet and exercise instruction to control GDM. A total of 776 women without GDM were randomly selected within the cohort and were frequency matched to women with GDM by residency, age (±2 years), and month of conception. Among the 1552 participants, 1496 women (752 women with GDM and 744 women without GDM) had information for both MB and CB. The 1496 women were included in the final analysis.

The study was approved by the Human Investigation Committees at the Shanxi Medical University. All participants were informed and provided written consent.

Blood Zn level measurement

MB and CB samples were both collected at the time of delivery and stored immediately. MB and CB Zn levels were measured at the National Institute of Environmental Health, Chinese Center for Disease Control and Prevention using inductively coupled plasma mass spectrometry (ICP-MS). Briefly, 300-μL blood samples were mixed with BV-Ⅲ HNO₃ (200 μL) and 30% H₂O₂ (100 μL) and digested using the Mars-5 microwave accelerated reaction system. Digested samples were diluted with deionized water to a nitric acid concentration of 10%. Metal concentrations in the sample solutions were tested using ICP-MS (PerkinElmerSciex ELAN® DRC-II, USA). Each batch (20 samples) contained 1 positive sample (the SeronormTM trace elements whole blood L-2; LOT 1 003 192) and 1 negative sample (deionized water) for quality control purposes. Each batch included samples of both GDM and non-GDM mothers. Laboratory personnel were blinded to GDM status.

Dietary Zn intake assessment

Dietary intake was retrospectively assessed using a validated 33-item food frequency questionnaire through an in-person interview at the hospital either before or after delivery. Participants were asked to report the frequency and consumption of the standard portion size for each food for the following 4 time periods: the year before conception, the first trimester, the second trimester, and the third trimester during pregnancy. Dietary nutrients were estimated from the frequency of consumption and portion size of food items using the Chinese Standard Tables of Food Consumption (24). According to National Institutes of Health, the recommended dietary Zn intake is 11 mg/day for pregnant women (25). We thus used 11 mg/day as a cutoff to determine adequate and inadequate dietary Zn intake.

Statistical analysis

Distributions of all variables were summarized and compared between GDM and non-GDM pregnant women. The chi-square test was used to examine the difference in categorical variables and Student’s t-test was used to examine the difference in maternal age, birth weight, MB Zn level, and CB Zn level.

In the following analyses, MB Zn level and CB Zn level were log₂-transformed due to skewness. They were analyzed as continuous variables using log₂-transformed values or as categorical variables using their tertiles in the overall population as cutoffs. The tertile cutoffs were 5.81 and 7.23 mg/L for MB Zn level, and 2.27 and 2.82 mg/L for CB Zn level. The use of tertiles allowed us to investigate a potential nonlinear relationship while maintaining adequate statistical power.

In order to compare birth weight across different gestational weeks and fetal sexes, the standardized birth weight (z score) was calculated using the formula actual value – mean/standard deviation. The mean values were based on the gestational week and sex-specific birth weight reference in China published in 2014 (26). Additionally, birth weight was classified as low birth weight (LBW; <2500 g) or high birth weight (HBW; >4000 g).

First, we evaluated the association between maternal Zn levels and GDM in this population using logistic regression. Maternal education, family income, prepregnancy body mass index (BMI), nulliparity, smoking, and gestational hypertension were adjusted in the models. Crude and adjusted odds ratio (OR) and 95% confidence interval (CI) were calculated. This analysis evaluates whether GDM might be a mediator in the causal pathway between maternal Zn levels and birth weight.

Second, we evaluated the associations between Zn levels and birth weight of offspring in mothers with or without GDM using multivariate linear regression and multinomial logistic regression. In the multivariate linear regressions, birth weight was treated as a continuous variable. In the multinomial logistic regression, birth weight was treated as a categorical variable and normal birth weight (2500 g ≤ birth weight ≤ 4000 g) was used as the reference group. Maternal age, education, family income, prepregnancy BMI, nulliparity, smoking, gestational hypertension, and gestational weight gain were included in the multivariate linear regression and multinomial logistic regression models. Additionally, the linear regression was performed again in the population stratified by adequate and inadequate dietary Zn intake.

Generalized additive models were used to test the nonlinearity relationship between continuous exposure and outcome measures. We conducted several sensitivity analyses to examine the robustness of our findings. First, for sensitivity analysis we used quartiles of Zn levels (as opposed to tertiles); second, we used birth weight at the gram scale and adjusted for offspring sex and gestational weeks in the models; third, we adjusted the blood glucose in the models; and, fourth, we reran analyses excluding women with gestational hypertension because the medication used for hypertension control has a diuretic effect that can decrease Zn levels in blood.

All statistical analyses were performed using SAS (version 9.4; SAS Institute Inc., Cary, North Carolina, USA).

Results

Table 1 presents the distribution of selected characteristics between GDM and non-GDM pregnant women. Significant differences were observed for prepregnancy BMI and birth weight z score as expected: women with GDM were more likely to have a higher prepregnancy BMI and their offspring were more likely to have a higher birth weight z score. The gestational weight gain was significantly lower in women with GDM than in women without GDM. No significant difference was observed for MB Zn level (6.56 vs 6.62 mg/L) and CB Zn level (2.68 vs 2.72 mg/L) between women with and without GDM. Approximately 90% (N = 1328) of the entire population did not meet the recommended dietary Zn intake level of 11 mg/day. The correlation between MB and CB Zn levels can be found in Table 2. In general, MB and CB Zn levels were correlated but the correlation was weak, especially in women with GDM.

Table 3 shows the association between GDM and Zn levels. Neither MB nor CB Zn levels were associated with GDM status. Because there was no direct relationship between Zn status and birth weight, this eliminates the possibility that GDM mediates the association between maternal Zn status and birth weight.

Table 4 shows effect estimates (β) of Zn levels for birth weight z scores in multivariate linear models. Overall, associations between birth weight and Zn levels varied by GDM status, and bidirectional associations were observed for MB and CB Zn levels. In women without GDM, no significant association was observed between Zn levels and birth weight. In women with GDM, inverse associations were observed for MB Zn level (per doubling: β = –.17, 95% CI, –0.34 to –0.01) while positive associations were observed for CB Zn level (per doubling: β = .38, 95% CI, 0.06-0.70). Compared with the lowest tertile of MB Zn level, the highest tertile of MB Zn level was associated with 0.23 units decrease in birth weight z score (95% CI, –0.45 to –0.01). Compared with the lowest tertile of CB Zn level, the highest tertile was associated with increased birth weight z scores (β = .24, 95% CI, 0.02-0.47). When the analysis was restricted to women having inadequate dietary Zn intake, the associations became slightly stronger for both MB Zn level (per doubling: β = –.21, 95% CI, –0.42 to –0.01) and CB Zn level (per doubling: β = .41, 95% CI, 0.07-0.76). However, no significant association was observed in women having adequate dietary Zn intake, possibly because of small sample size.

Results from multinomial logistic models (Table 5) were generally consistent with those from multivariate linear models. In women without GDM, the MB Zn level appeared to decrease the risk of LBW (per doubling: OR 0.55; 95% CI, 0.30-1.02), while the CB Zn level appeared to decrease the risk of LBW (per doubling: OR 0.68; 95% CI, 0.19-2.43) but increase the risk of HBW (per doubling: OR 2.59; 95% CI, 0.66-10.17). In women with GDM, the MB Zn level was associated with increased risk of LBW (per doubling: OR 2.01; 95% CI, 0.94-7.39), though the association was only borderline significant. Meanwhile, the CB Zn level was significantly associated with HBW (per doubling: OR 2.37; 95% CI, 1.08-5.21).

A nonlinearity relationship was not detected in generalized additive models (P-value for nonlinearity >.10) between Zn levels and birth weight z score. In the sensitivity analyses, using Zn quartiles in the linear regression model, using continuous birth weight in grams, adjusting for blood glucose, and removing women with gestational hypertension yielded similar results (all supplementary material and figures are located in a digital research materials repository (27)).

Discussion

In this cohort of pregnant women in China matched by GDM and with a high prevalence of inadequate dietary Zn intake, we observed complex relationships between birth weight, Zn levels, and GDM. In women without GDM, we observed no significant association between Zn in MB or CB and birth weight, but high maternal Zn level still appears to decrease the risk of LBW. In women with GDM, we observed intriguing findings that the MB Zn level was inversely associated with birth weight, while the CB Zn level was positively associated with birth weight. These results suggest that in the Chinese population characterized by inadequate Zn intake during pregnancy, GDM may modify the effects of Zn on birth weight, and that these measures of Zn status are capturing different features.

One interesting observation in this study is that the gestational weight gain was lower in women with GDM. The difference can be explained by the higher proportion of women with high prepregnancy BMI in the GDM group, since higher prepregnancy BMI above 18.5 is usually correlated with lower gestational weight gain (28).

In previous population studies, the association between Zn status and birth outcomes is mixed. Goldenberg et al. first reported that Zn supplementation in pregnant women significantly increased the birth weight in 1995 (29), but 2 recent systematic reviews challenged this conclusion (4,5). Both reviews concluded that though Zn supplementation during pregnancy was associated with reduced preterm birth, it had no apparent effect on birth weight. When Zn was measured in MB, 3 studies suggested that MB Zn was positively associated with birth weight (30-32), while 6 studies reported no association (33-38). In 1 study using the CB Zn level, the authors concluded no association between CB Zn level and birth weight (39). In contrast to these previous studies, our study suggested that both MB and CB Zn levels were associated with birth weight but only among women with GDM.

The present study is not the first one to report an inverse association between MB Zn level and birth weight among women with GDM, the most intriguing finding in this study. In 2005, an Italian study reported that among 71 women with GDM, a higher maternal serum Zn level was significantly associated with lower birth weight of offspring (40). The Italian researchers attributed this inverse association to Zn deficiency. When using dietary intake as the measure for Zn deficiency, both our study and the Italian study reported lower dietary Zn intake level than the recommended level of 11 mg/day during pregnancy (41). In the present study, the average dietary Zn intake was 7.4 mg/day and only 11.8% (N = 89) of the women with GDM met the recommended level. In the Italian study, the average dietary Zn intake was 8.5 mg/day in the women with GDM.

It is estimated that women with GDM have a 3-fold risk to deliver macrosomia compared with women without GDM (16). While the underlying mechanisms remain unclear, the Pedersen hypothesis provides a possible explanation for macrosomia in women with GDM: maternal hyperglycemia enhances the placental transmission of glucose while exogenous insulin is unable to cross the placenta; meanwhile, the fetus responds to hyperglycemia by secreting insulin. As a result, more fat and protein are accumulated in fetus (16,42). In women with GDM, the higher MB Zn level could strengthen maternal glycemic control and ameliorate GDM complications during pregnancy (43). Therefore, the controlled maternal glycemia resulting from higher MB Zn level would decrease glucose transmission to the fetus, leading to reduced fetal weight. Moreover, the higher CB Zn level may accelerate the metabolism of glucose in the fetus of women with GDM and result in a positive association between CB Zn level and birth weight.

It is also noteworthy that the correlation between MB and CB Zn levels was weak, especially among women with GDM in this study. Because GDM could alter placenta transport and interfere with maternal Zn homeostasis (44), MB Zn might be prevented from reaching the fetus in women with GDM. Therefore, it could be hypothesized that a higher MB Zn level among women with GDM might indicate a greater deficit of Zn. As a result, a negative association between MB Zn level and birth weight was observed. However, more studies are needed to confirm this hypothesis.

Our study had multiple strengths. One highlight is that the matched-cohort design of maternal GDM allows a large sample size to investigate the potential modifying roles of maternal GDM on the associations between maternal and cord Zn levels and birth weight. Additionally, we evaluated the Zn level in MB and CB and differential results were found for birth weight in women with GDM, suggesting these types of biomarker measures may be an important consideration when assessing concentrations of Zn on adverse birth outcomes.

Some limitations should also be considered when interpreting the findings. First, blood samples were only collected once at the time of delivery, so we were unable to evaluate if the effect of Zn level changes during pregnancy. Second, we used blood Zn levels, which may not represent the overall Zn status. In the human body, the majority of Zn (>80%) is stored in muscles and bones (45). However, the blood Zn level is a reliable biomarker that provides sensitivity and analytic convenience to indicate the Zn status at the population level (45) and represents the circulating Zn in bodies that could cross the placenta and participate in metabolic activities, which may be more important for fetal growth. Third, the individual maternal metabolic profile was not included in this study. The maternal metabolic profile is closely associated with maternal nutrition level and fetal growth and thus may be a confounding factor. However, despite strict metabolic control in women with GDM, fetal growth remains a clinical problem (7). Therefore, the omission of maternal metabolic profile would not substantially bias our results.

Results in this study have several implications. First, the associations between Zn status and neonatal birth weight were only observed in women with GDM, suggesting a previously unidentified path of adverse effects of GDM on birth outcomes. Second, the population in this study was characterized by inadequate dietary Zn intake during pregnancy. Third, there was suggestive evidence in this study that higher maternal Zn status might reduce the risk of LBW in women without GDM. Zn is an essential nutrition and its beneficial effects should be recognized. Zn is recommended in the diet by the National Academy of Sciences (46) and is required in total parenteral nutrition fluids by the Food and Drug Administration (47). As Zn deficiency is a global public health issue, more efforts are needed to eliminate Zn deficiency during pregnancy in order to avoid potential adverse effects and improve birth outcomes.

In conclusion, this study found an inverse association between MB Zn level and birth weight, and a positive association between CB Zn level and neonatal birth weight in offspring born to women with GDM characterized by inadequate dietary Zn intake, but no such associations were observed in offspring of women without GDM. The results suggest that GDM may modify the association between maternal Zn level and birth outcomes in Zn-deficient pregnant women. These findings suggest a previously unidentified path of adverse effects of GDM on birth outcomes. More studies are needed to further evaluate the association and understand the underlying mechanisms.

Abbreviations

Abbreviations
 
• BMI

  body mass index

 
• CB

  cord blood

 
• CI

  confidence interval

 
• GDM

  gestational diabetes mellitus

 
• HBW

  high birth weight

 
• LBW

  low birth weight

 
• MB

  maternal blood

 
• OR

  odds ratio

Acknowledgments

We thank all the study personnel from the First Affiliated Hospital of Shanxi Medical University for their exceptional efforts in study subject recruitment.

Financial Support: this work was supported by National Natural Science Foundation of China (No. 81473061); Natural Science Foundation of Shanxi Province (No. 2013021033-2); 10 Talent Program of Shanxi Medical University; China Scholarship Council (No. 201808140187).

Additional Information

Disclosure Summary: The authors declare no actual or potential competing financial interests.

Data Availability: The datasets generated during and/or analyzed during the current study are not publicly available but are available from the corresponding author on reasonable request.

References

Author notes