Histological chorioamnionitis is associated with an increased risk of wheezing in preterm children less than 34 gestational weeks

Intrauterine events can have a lifelong impact on chronic diseases. Exposure to adverse stimuli can permanently change the structure and function of tissues and organs. Every year, nearly 14.9 million preterm babies are born, accounting for about 10.5% of total births [1]. Studies have revealed that preterm birth is associated with an increased risk of chronic diseases in adulthood and impairs neurodevelopment such as cognitive and motor impairments and hearing loss [2]. Chorioamnionitis is a common gestational complication which results in 40–70% of preterm deliveries. Diagnosis of histological chorioamnionitis (HCA) is mostly based on the infiltration of neutrophils and lymphocytes into the placenta and associated membranes. Many studies have associated chorioamnionitis with various short- and long-term adverse outcomes, such as intrauterine growth restriction, cerebral palsy, chronic lung disease, wheezing or asthma later in life [3, 4]. However, the outcomes are inconsistent across studies. For example, several studies found that chorioamnionitis increased the risk of long-term cognitive and behavioural impairments in preterm children [5, 6], while others did not [7].

Currently, substantial studies about the association between preterm birth and atopic diseases and long-term physical development have been published [8, 9]. However, studies about the effects of chorioamnionitis on wheezing and atopic diseases among preterm children less than 34 gestational weeks remain limited. There is also a lack of hematological indicators in previous studies. Furthermore, systemic measurements of the physiological developments of preterm children less than 34 weeks who were exposed to chorioamnionitis at 2–3.5 years of age have not been reported.

Based on previous reports, we hypothesized that HCA exposure would be associated with an increased impairment of children’s developments and risk of atopic diseases. Thus, in the present study, we followed up the outcomes related to anthropometric indexes, neurodevelopment, wheezing, atopic diseases (eczema, food allergies and allergic rhinitis) and allergic indicators among preterm children less than 34 weeks who were exposed to HCA. The aim of the study was to investigate the relationship between HCA and physiological development, wheezing, and atopic diseases in preterm children.

In this study, a total of 307 eligible mother-children pairs were screened out, of which 115 were successfully enrolled (Fig. 1). Comparison of the maternal and children’s baseline characteristics revealed that there was nearly no significant difference between the recruited and study populations, except the maternal education level (Table S1). Among the 115 children, 47 (40.87%) were in the HCA exposed group and 68 in the unexposed group (Table 1). The mean gestational age was 31.66 ± 1.70 weeks. There were no significant differences between the two groups in maternal age, children’s birth weight, gender, and first degree relatives’ allergic history [16, 17]. Variables that were significantly associated with HCA included a smaller gestation age, preterm premature rupture of the membranes (PPROM), vaginal delivery and the use of antibiotics in the first year of life (Table 1).

During the follow-up (ages 2–3.5 years), outcomes of physical development, blood pressure, and neurodevelopment were similar among the children exposed to HCA and those who were not (Table 2). However, preterm children exposed to HCA were significantly more prone to wheezing than control children (38.30% vs 16.18%; P < 0.05). After adjusting for maternal age, children’s current age, gender, type of delivery and use of antibiotics in the first year of life, multivariate logistic regression analysis revealed that preterm children who were exposed to HCA had a 2.72-fold increased risk of wheezing (aOR, 2.720; 95% confidence interval, CI, 1.02–7.23). Furthermore, neither univariate nor multivariate analyses revealed an association between HCA and eczema, food allergies, or allergic rhinitis (Table 3).

To further investigate the influence of HCA on preterm children, serum metabolomic profiling was conducted, which is a powerful tool for assessing metabolic alterations under a given set of physiological conditions. In total, 163 differential metabolites were identified, including 105 upregulated ones and 58 downregulated ones in preterm children who were exposed to HCA compared to those who were not (Table S2). PCA analysis revealed a clear separation between the HCA and control groups (Fig. 2a). To minimize the influence of intergroup variability and to further improve group separation, PLS-DA and OPLS-DA analyses were conducted. Similarly, the HCA group was clearly separated from the control group (Fig. 2b). In the OPLS-DA analysis, the R² value indicated how well the data were mathematically reproduced, which ranged between 0 and 1 with 1 indicating the best model. The Q² value indicated the percent variation of the response predicted by the model [18]. The results revealed an R² of 0.906 and a Q² of − 0.497, indicating that the model was stable and perfectly fit. The top differentially identified metabolites included 3-hydroxy-2-methyl-[R-(R,S)]G-butanoic acid, annotemoyin 1,and charine (Table 4). KEGG analysis of the differential metabolites revealed an enrichment of several key metabolic pathways, including the glycerophospholipid, alpha-linolenic acid and choline metabolisms (Fig. 2c). Specially, we found that the pathway of pathogenic Escherichia coli infection was involved in the metabolic changes associated with HCA (Fig. 2c), indicating a continuous influence of the intrauterine exposure since gram-negative bacterial infection is the most common cause of HCA.

In chorioamnionitis, local levels of cytokines such as IL-1, IL-6 and TNF-α are increased. This disorder further induces multiple organ injuries [19]. Moreover, elevated proinflammatory mediators and inflammatory cytokines can induce life-long epigenetic modifications, which alter the development and function of cells. In this study, the follow-up demonstrated that HCA was associated with an increased risk of wheezing in preterm children less than 34 gestational weeks. Additionally, there was a significant alteration in serum metabolites. Our results contributed a further stone for the theory of intrauterine original disease.

Since the first report on the association between chorioamnionitis and early lung inflammation by Watterberg et al [20], the prenatal influence on lung development has been extensively studied. A cohort study of 1096 children found a joint effect of chorioamnionitis and prematurity on early children’s recurrent wheezing [21]. Another study of preterm children less than 32 gestational weeks also reported that fetal inflammatory response syndrome increased the risk of early childhood wheezing [22]. In this study, we found a 2.72-fold increased risk of wheezing in preterm children less than 34 weeks with HCA. However, in full-term children, reports indicated there was no such an association [23]. Studies in asthma showed that chorioamnionitis increased the risk of asthma, but the effect would diminish with increased gestational age [15]. Similar characteristics may also exist for the relationship between chorioamnionitis and early childhood wheezing, though further study is required.

Despite the increased risk of wheezing in preterm children exposed to chorioamnionitis, we did not observe significant differences in serum total or specific IgE in this study. There are at least two possible reasons for this: (1) the large variation in the levels of serum IgE among population; (2) the sampling time, since the children with wheezing in both groups were not all in an ongoing status. Nonetheless, we found children with chorioamnionitis were more prone to have a higher level of plasma eosinophils. Besides, metabolomic profiling revealed the alteration of serum metabolites from hypoxia and immune responses, and the lipid metabolism in HCA exposed children (Fig. 2c), which were fingerprints of asthma [24]. Reports indicated there was a linear correlation between early childhood wheezing and asthma among preterm children exposed to chorioamnionitis [25]. Thus, these results reinforce our finding that an increased risk of wheezing occurs in preterm children exposed to HCA.

Increased risk of IUGR and poor early postnatal growth in children exposed to chorioamnionitis has been widely reported. However, limited literatures have been published about the effect of chorioamnionitis on long-term physical development in premature children, and outcomes remain controversial [26, 27]. In this follow-up, we found no significant difference in height, weight, skinfold thickness or blood pressure between the preterm children exposed to HCA or not at the age of 2–3.5 years. Infancy and early childhood are critical periods for physical development. Our findings indicated that there might be no influence of chorioamnionitis on children’s physical development. However, the long-term physical disorders still require further investigations [28].

Chorioamnionitis may induce brain injury in preterm newborns and be linked to later adverse neurodevelopment. Thus, in the present study, children’s neurodevelopment was also evaluated. However, we found that there was no association between HCA and poor neurodevelopment in early childhood. This finding is consistent with recent reports [7, 29], which revealed an adverse association between chorioamnionitis and brain injury or developmental disability [6]. This discrepancy may be due to several factors including: (1) maternal education, which is associated with cognitive and behaviour developments in children (only 4.32% of mothers in our study did not receive an education at or above the high school level), and (2) antenatal steroid administration (91.26% of mothers in the present study received antenatal steroids), which protects the fetal brain from injury and decreases the risk of adverse neonatal outcomes.

Metabolomic profiling can provide insight into active physiological status, which has been proved to be useful in diagnosis of disease, monitoring prognosis and evaluation of therapeutic strategies. For example, formate, hippurate and methanol were proved to be common biomarkers of asthma [30]. In this study, we observed a clear separation of the serum metabolites between the HCA and control groups. Many altered metabolites or altered metabolic pathways have been reported to play key roles in children’s growth and development. For example, linolenic acid metabolism was reported to be associated with neurodevelopment [31]. Levels of metabolites can change before the emergence of phenotype. The minimal impacts observed during this follow-up may not tell the full story. Noteworthy, metabolomic analysis provides the field with a new perspective on the mechanisms underlying chorioamnionitis impacts on long-term chronic diseases. Thus, prompt clinical intervention in HCA cases might reduce the prevalence of wheezing in offspring.

There were some potential limitations that need to be considered when interpreting our data. First, the sample size of present follow-up was relatively small. For example, while the aOR for wheezing was 2.72, the 95% CI stretched across 1.02 to 7.23. Meanwhile, inconsistent with previous studies [7, 23, 29], the other observed children’s outcomes of HCA exposure was modest in this study. Although there was nearly no difference between the recruited and study populations (Table S1), statistical power calculation revealed that the recruited number was lower than the required for the outcomes of low prevalence, such as DQ < 85 and physical retardation. A further limitation to this study was that postnatal nutrition data were not collected. Numerous studies have demonstrated the direct relationship of children’s physical development and nutrition [32]. The sodium salt intake is also strongly associated with blood pressure. Another limitation was the basis of socioeconomic status. More than 70% mother of the present study received education above college, which was much higher than the average of China. Reports have indicated socioeconomic status was strongly associated with children’s developments [14].

In this study, we found that HCA was associated with an increased risk of wheezing among preterm children less than 34 weeks. Moreover, HCA induced a significant change of serum metabolomic profile in preterm children. However, HCA was not linked to atopic diseases, abnormal blood pressure, or physical or neuro- development.