Study design: This single-blind clinical trial was performed on mothers and neonates with a gestational age of 32-37 weeks who were referred with labor pains from February 20th 2021 to June 29th 2021 in the obstetrics and gynecology department and intensive treatment unit of Ghaem Hospital, Mashhad University of Medical Sciences, Iran. Once the Ethics Committee of Mashhad University of Medical Sciences (IR.MUMS.MEDICAL.REC.1399.613 and number 990699) approved the study, the researcher received informed consent from the participating parents. The study was registered in the Iranian Clinical Trial Registration Center (IRCT20110807007244N7) before starting the project.
Inclusion and exclusion criteria: Inclusion criteria included pregnant women with a gestational age of 32-37 weeks, having a risk of preterm delivery, normal fetal prenatal ultrasound, lack of significant medical and obstetric problems for the mother (such as cardiovascular problems, diabetes mellitus, epilepsy, and asthma).
Exclusion criteria, on the other hand, included meconium aspiration syndrome, congenital anomaly, congenital infection, RDS, TTN, intubation in delivery room, death in the delivery room, severe asphyxia, addicted mothers, pneumonia, and persistent respiratory distress after the first 12 h of life.(Figure 1)
Randomization: In this study, blocking is used as a reliable randomization method as it leads to a balanced allocation of individuals in treatment groups at the end of each block. For this purpose, a web-based online method is used to determine the volume and number of random blocks. This online software can create random blocks with desired variable sizes. Twenty blocks with variable sizes of 6, 8, and 10 were introduced for a total of 160 samples using this website. The blocks were marked with numbers 1 to 20. Each block includes an equal number of intervention and control groups according to the size of that block. Following the reference of the first mother, one of the blocks was randomly selected. Then, it was assigned according to the order of the contents of the block in one of the groups. By referring to the next mother, the next item of the same block was selected. The remaining blocks were processed, and these steps were repeated until sampling was completed. This process continued until the completion of the first block. The next block was selected randomly from the remaining blocks, and the above process was repeated until sampling was completed. The neonates were followed by the principal researcher, who did not know the relevant, control, or intervention group regarding the occurrence or non-occurrence of respiratory distress and the time of recovery and diagnosis of respiratory distress. Data were classified as A and B in the SPSS software, and the statistical analyst interpreted the data without knowing the type of grouping.
Intervention: Within 72 h of preterm delivery, a single dose of 50,000 units of intramuscular 25-hydroxy vitamin D was injected into pregnant women in the intervention group who had not taken a 25-hydroxy vitamin D in the last three months.
Laboratory evaluation: A sample containing 1.5 ml of whole blood was taken from the umbilical cord of the infant and mother to assess the level of vitamin D. The prepared samples were centrifuged, and its serum was kept at -20℃ and sent to the central laboratory of Ghaem Hospital for evaluation. The level of vitamin D was measured in ng/mL by ELISA Reader device model RT2100c (Germany) and ELISA Washing device using the ELISA method. The 25-hydroxy vitamin D lower and higher than 30 ng/mL was considered 25-hydroxy vitamin D deficiency and sufficient values. Deficiency values were divided into three groups of severe deficiency (≤10 ng/mL), moderate deficiency (10.1-20 ng/mL), and mild deficiency (20.1-30 ng/mL).
Radiological evaluation: Infants having more than 2 h of respiratory distress and being admitted to the neonatal ward underwent a chest radiographic examination. Next, the cause of neonatal respiratory distress was determined based on radiographic findings and clinical course.
Outcomes: The primary outcome of the neonates in the study group included the incidence of non-specific respiratory distress. The diagnosis criteria included respiratory distress (tachypnea, intercostal muscle retraction, granting, and nasal flaring) more than 2 h and less than 12 h, normal chest radiograph, and negative blood culture.
Data collection: Complete characteristics of infants (birth weight, age, sex, gestational age, Apgar score, and clinical signs), maternal history (age, pregnancy, and delivery problems, and delivery method), and laboratory results were collected and recorded in a questionnaire. The need for resuscitation, duration of respiratory distress, duration of oxygen therapy, duration of hospitalization in the NICU, and mortality were also recorded in the checklist. The levels of calcium and vitamin D of the infant were also recorded.
Patients and Methods
A single-blinded randomized controlled clinical trial was conducted on pregnant mothers with a gestational age (GA) of 32-37 weeks at risk of preterm labor. The mothers were randomly assigned into two groups, including control group (with no injection) and intervention. A single dose of 50,000 units of intramuscular 25-hydroxy vitamin D was injected into pregnant women in the intervention group within 72 h of preterm delivery. A sample containing 1.5 ml of whole blood was taken from the mothers and infants’ umbilical cord to assess the level of vitamin D. Short-term outcomes, including neonatal respiratory findings, and laboratory results of mothers and infants were recorded in checklist.
This was a clinical trial on 169 infants with gestational age of 32-37 weeks at birth. Neonates with more than 2 h and less than 12 h respiratory distress, normal chest radiograph, and negative blood culture met the criteria for the study. The infants were placed under oxyhood with closed monitoring for worsening respiratory symptoms. Umbilical catheter was inserted for infants and ABG according to the patient status was obtained. All infants were monitored continuously; pulse oximetry was performed by a probe at right upper extremity, and respiratory symptoms were examined repeatedly. Blood gas analysis and oxygen saturation were gold standard to detect infants at risk of respiratory failure.
Data analysis: Statistical analysis was performed by the SPSS software (IBM SPSS Statistics, Version 23).The mean of quantitative variables between groups was compared using an independent t-test or Mann-Whitney U test. Chi-square test and Fisher’s exact test were used to compare qualitative variables between the studied groups. Data were expressed as mean ± standard deviation (SD). P <0.05 was considered as the statistically significant level of the data.