Preterm premature rupture of membranes (PPROM) is defined as a spontaneous rupture of the membranes during pregnancy before 37 weeks’ gestation and at least 1 h before the onset of contractions [1]. The incidence of PPROM ranges from 3.0% to 10.0% of all deliveries. Approximately 40% of PPROM cases result in preterm delivery, contributing significantly to increased neonatal morbidity and mortality [2, 3, 4]. The main causes of PPROM are infection/inflammation, decidual bleeding, uterine overdistention (e.g., polyhydramnios, twins), genetic predispositions, and smoking [5]. The risk factors for PPROM are generally similar to those for preterm birth (PTB), including preterm spontaneous labor with intact membranes [6]. The known risk factors for preterm birth are short cervical length, polyhydramnios, PPROM, assisted reproductive technologies (ART) procedure, prior PTB, pregnancy induced hypertension, placenta previa, placental abruption, fetal growth restriction, urinary tract infections, complex autoimmune diseases with polytherapy, maternal anxiety, obesity, and malnutrition [7, 8, 9, 10]. There are a few reports on maternal vitamin D insufficiency or deficiency associated with PTB [11, 12, 13]. Woo et al. [14] reported a positive association between vitamin D deficiency and PTB. In contrast, a prospective cohort study reported no significant difference in vitamin D levels between the PTB group and full-term delivery group [15], this association remains conflicting. However, there are few reports that have studied the relationship between PPROM and vitamin D deficiency so far. Therefore, we investigated the association between PPROM and vitamin D levels in maternal plasma and umbilical cord blood of newborns with possible circumstances excluding factors affecting PTB.

Our study was conducted prospectively, and all participants with risk factors that could affect PPROM were excluded. All participants were recruited in the first trimester of pregnancy, and blood was collected for vitamin D level analysis at predetermined periods (the first, second, third trimester of pregnancy and just before delivery). Additionally, vitamin D levels were measured from the blood taken from the umbilical cord of the newborn. Our study showed that the occurrence of PPROM in cases with vitamin D deficiency (20 ng/mL) in the umbilical cord blood was remarkably higher than that in cases with a normal level of vitamin D. Vitamin D levels in the maternal plasma and umbilical cord blood at birth in the PPROM group were significantly lower than those in the non-PPROM group. The odds ratio of PPROM was 0.907 with an increase of 1 ng/mL of vitamin D level of cord blood. In addition, since the history of preterm labor or PTB can be a risk factor for PPROM, in the subanalysis, only primigravidas were analyzed for the incidence of PPROM. We found that the incidence of PPROM in primigravidas was higher in the vitamin D deficiency group than the normal group. Even when the known risk factors, the history of preterm labor and preterm delivery are excluded, PPROM is frequent in the vitamin D deficiency group. Therefore, vitamin D deficiency can be considered another risk factor for PPROM.

The rate of PPROM in this study was 3.38% (12/355), with the rate in the vitamin D deficiency group being 6.3% (11/176) and 0.6% (1/179) in the non-deficiency group. Of these, 66.7% (8/12) led to PTB. In the United States, the PTB rate is approximately 10% [21]. PPROM is responsible for 30%–40% of PTBs [22]. Similar to in the United States, PTB rate is approximately 7% in South Korea [23]. Therefore, reducing vitamin D deficiency during pregnancy might lead to a reduction of PTB due to PPROM.

There are several hypotheses for how vitamin D deficiency can affect PPROM. First, vitamin D is involved in several processes leading to an inflammatory response. The fetal membrane not only acts as a barrier to the invasion of microorganisms [24, 25, 26], but also provides immune protection [27, 28, 29]. Impairment of these functions of the fetal membrane can lead to microbial invasion in the genital tract as well as triggering of inflammatory processes, resulting in PPROM through collagenolysis [24, 30, 31, 32]. Some studies have reported that inflammation with bacterial infection is the main cause of PPROM [33], and recent studies have reported that sterile inflammation of the fetal membrane without microbial infection is the predisposing cause of PPROM [34]. Liu et al. [35] reported that vitamin D regulates both acquired and innate immune responses at the fetal-maternal interface. Vitamin D could act as an intracrine regulator of cyclic adenosine monophosphate in trophoblasts and regulate innate immune response in the placenta [36, 37]. In the placenta, 1,25-dihydroxyvitamin D induces the production of cathelicidin, an antimicrobial peptide, and consequently is associated with a reduction of the risk of bacterial vaginosis. Dunlop et al. [38] reported that vitamin D deficiency during pregnancy is more likely to cause bacterial vaginosis. Moreover, sufficient vitamin D levels during pregnancy could reduce the risk of bacterial vaginosis, thereby reducing the incidence of PTB [39, 40]. Therefore, maternal vitamin D could function as an immune regulator in pregnancy and have anti-inflammatory effects which reduce the risk of PPROM by decreasing bacterial colonization of the genital tract.

Second, vitamin D is a potent antioxidant. One of the pathophysiologies of PPROM is oxidative stress (OS) mediated by reactive oxygen species (ROS). Collagen is the primary target of ROS. In healthy pregnancies, the ROS and antioxidants are balanced [41, 42, 43, 44, 45]. However, this balance is disturbed in case of increase in ROS (high O${}_2$ demand, e.g., high altitude pregnancy, twin pregnancy), antioxidant deficiency, or microvascular disease [46]. Insufficient vitamin D levels are reportedly associated with increased OS or reduced antioxidant capacity [47, 48]. Vitamin D promotes the expression of nuclear respiratory factor 2 (Nrf2) which inhibits the action of ROS, and the expression of an antioxidative enzyme that eliminates free radicals at the genetic level [49].

Overall, PPROM is associated with an inflammatory response by infection and directly with OS-inducing chorioamniotic collagenolysis. Vitamin D can regulate the immune response in the placenta and decrease bacterial colonies in the genital tract. Furthermore, vitamin D functions as an antioxidant that upregulates antioxidative enzymes, triggering the expression of Nrf2 that suppresses ROS via genetic mechanism.

We collected pre-pregnancy BMI and measured BMI just before delivery, and Table 3 showed that prepregnant BMI and just before birth BMI were statistically significantly higher in the vitamin D deficiency group than in the normal group. As mentioned in several studies, it was reported that the higher the BMI, the lower the vitamin D blood concentration [50], and there is also a research result that the bioavailability of vitamin D is lowered in the obesity group [51].

Our study has several advantages. This study was prospectively conducted. Therefore, it was possible to exclude factors that could affect PPROM. Additionally, to exclude previous PTB and preterm labor history, a second sub-analysis was performed with only primigravidas. Our sample size of 355 pregnant women was relatively larger than that of previous studies. Moreover, this study investigated the association between PPROM and vitamin D deficiency in the umbilical venous cord blood of newborns. To the best of our knowledge, previous studies on this association are scarce.

However, our study has some limitations. The sample size was too small to allow the generalizability of our research outcomes. Additionally, we could not accurately analyze the correlations between the vitamin D level in the maternal plasma, umbilical cord blood, and vitamin D intake (through vitamin D therapy or diet during pregnancy). In the future, further study on PPROM and vitamin D levels, considering fetal infection and vitamin D intake, is required to study the detailed mechanism of PPROM associated with vitamin D deficiency.

PPROM accounts for a considerable portion of PTB, which increases fetal morbidity and mortality. In this study, we found that vitamin D deficiency had a significant association with PPROM. We expect that the incidence of PPROM could be considerably reduced if vitamin D intake is recommended before and during pregnancy. For this to be achieved, further large-scale research needs to be conducted.

HJL—Designed the research study, acquired patient data, and drafted the manuscript; JYH—Conception, design, statistical analysis, created figures and tables, provided guidance, responsible for the accuracy and integrity of the work presented here; JHH—Statistical analysis and data extraction; HYK—Interpretation of data and statistical analysis; HZC—Statistical analysis and data extraction, created figures and tables. All authors contributed to editorial changes in the manuscript. All authors read and approved the final version of the manuscript.

This prospective study was conducted after obtaining ethics approval from the institutional review board of Seoul Medical Center (2017-03-015). All participants provided written informed consent after receiving a complete explanation of this study.

We would like to express our gratitude to all those who helped us during the writing of this manuscript.

This study was supported by the Seoul Medical Center, Korea (17-C09, 18-C04). The sponsor played no role in the design, collection, analysis and interpretation of data, writing the manuscript, or deciding to submit the manuscript for publication. The article contents are solely the responsibility of the authors and do not necessarily represent the official views of the sponsor.

The authors declare no conflict of interest.