Study design and participants
This study is a prospective cohort study, using a multicenter convenience sampling method to select mid pregnancy (24-28W) women recruited from Nantong University Affiliated Hospital, Nantong Maternal and Child Health Hospital, and Nantong Third People's Hospital prenatal clinics from July 2021 to January 2023 according to inclusion and exclusion criteria. Among them, 32W pregnant women with abnormal glucose tolerance were measured for fasting serum insulin before delivery, and all offspring were followed up from birth to 12 months. Investigate the effects of maternal intrauterine blood glucose exposure or insulin resistance on the eruption of deciduous teeth in their offspring. The project has been reviewed by the Ethics Committee of Nantong University Affiliated Hospital (2021-K004-01) and registered with the Chinese Clinical Trial Registration Center (ChiCTR210042814).Obtaining written informed consent from research participants.
Inclusion and Exclusion criteria
We enrolled women that fulfill the following inclusion criteria: 1) Age ≥ 18 years old and ≤40 years old; 2)No serious complications during pregnancy; 3)Singleton pregnancy; 4)Has communication and understanding skills; 5)Informed consent and voluntary participation in this study.
We excluded women if she has any of the following situation: 1)High risk of chromosomal abnormalities in prenatal screening of fetuses (suspicious or confirmed); 2)Women with severe cognitive impairment and mental illness; 3)Women who have used drugs such as tetracyclines that affect the development of fetal teeth and bones during pregnancy.We excluded women if her child have any of the following situation:1)In perinatal period, there were complicated asphyxia, apnea and hypoxic ischemic encephalopathy; 2)Infants with congenital or serious physical illnesses.
Data Collection and Variable Definition
Maternal Characteristics
The investigator obtained sociodemographic data such as the names, ages, places of residence, and educational levels of the mothers included in the study through a data survey form when their mothers were 24-28W pregnant. Among them, GDM pregnant women undergo fasting serum insulin concentration measurement from 32W pregnancy to delivery. With the consent and support of the hospital nursing department director and ward nurse, the investigator informs GDM pregnant women of the purpose of blood extraction. After explaining the research purpose, their consent is obtained, and an informed consent form is filled out. On the basis of the pregnant woman's consent, blood samples are taken to measure fasting serum insulin concentration. Data such as parity, gestational age, prepregnancy BMI, gestational weight gain, delivery method, baby gender and birth weight, were obtained from the maternal hospitalization medical records.
Serological indicator detection methods
- Specimen collection and testing: GDM pregnant women who meet the requirements should draw peripheral venous blood on an empty stomach for at least 8 hours in the late stage of pregnancy (32 weeks to delivery), inject it into EDTA anticoagulant tubes, and send it to the laboratory. Centrifuge the blood sample at 3000rpm for 10 minutes, take 1ml of the upper plasma and store it in a -80 ℃ freezer. Thaw it to 4℃ for insulin measurement and analyze the fasting plasma insulin concentration within 48 hours. The reagent kit was purchased from Roche Elecsys Diagnostic Products Co., Ltd. in Shanghai, China, and insulin detection was performed using electrochemiluminescence.
- Calculation formula: Insulin Resistance Index (IRI)=Fasting Blood Glucose (FPG) mmol/L × Fasting insulin (FINS) μ U/mL/22.5
- GDM and IR diagnostic criteria: Diagnosis is made through the Oral Glucose Tolerance Test (OGTT) at 24-28 weeks of pregnancy. Its diagnostic criteria [22]: When any of the following values are met or exceeded at 24-28 weeks of pregnancy: fasting PG (0h) ≥ 5.10 mmol/L, 1h PG ≥ 10.00 mmol/L, 2h PG ≥ 8.50 mmol/L, any blood glucose value that meets or exceeds the above criteria can be diagnosed as GDM. Pregnant women are divided into GDM group and non GDM group. Insulin resistance rapidly increases from 24 to 28 weeks of pregnancy and reaches its peak after 32 weeks. According to HOMA-IR, the critical value is set to 2.0. If HOMA-IR ≥ 2, it can be diagnosed as IR[23]. The research subjects were ultimately divided into three groups:the group unexposed to diabetes,the group exposed to GDM without IR and the group exposed to GDM with IR.
Offspring Characteristics
The status of infant vitamin D supplementation, daily outdoor activity time, time for adding complementary foods, postpartum breastfeeding, pacifier use, time of first deciduous tooth eruption, and total number of deciduous teeth at 12 months were obtained through telephone or WeChat follow-up. Forming a WeChat group for all pregnant women who meet the inclusion criteria and have informed consent is beneficial for further tracking the development of deciduous teeth in later generations.
Inform parents to take their baby to the children's health clinic for routine physical examination. During the initial physical examination, the nurse from the hospital's children's health department will train parents on the method of examining the eruption of deciduous teeth. The eruption of teeth is judged by any part of the crown appearing on the gums and being visible and palpable in the oral cavity. Confirm mastery through a model test.
Follow up by phone at 4, 6, 9, and 12 months to record the time of the first deciduous tooth eruption. The age identification is accurate to the day (for example, 5 months and 15 days are recorded as 5.5 months).At the age of 12 months, the offspring will undergo a physical examination at the Children's Health Stomatology Department to evaluate their teeth and oral development status, record the number of teeth, and calculate the teeth growth rate (number/month)=12 months of total teeth/(12 months of age - the age of the first deciduous tooth eruption).
Statistical Analysis
The data was double entered using EpiData software and subjected to consistency testing. Statistical analysis was conducted using SPSS 25.0 software. The chi square test and analysis of variance were used to explore the differences in maternal and infant characteristics between three group for categorical and continuous variables, respectively. Continuous variables were expressed as mean±standard deviation(X̅±S), while categorical variables were expressed as frequency (percentage). The correlation between different degrees of blood glucose exposure during pregnancy and the eruption time of the first deciduous teeth in offspring, the total number of teeth at 12 months, and the average monthly teething speed was analyzed using Spearman correlation analysis. Use multiple linear regression to analyze the influencing factors of offspring deciduous tooth eruption. P<0.05 shows statistically significant differences.