Objective To investigate the serum 25-hydroxyvitamin D (25OHD3) levels in mothers with gestational diabetes mellitus (GDM) and infants,and their association with insulin (FINS) and the expression of inflammatory factors.Methods Totally 629 pregnant women who delivered in department of obstetrics at the hospital from May 2017 to May 2020 were enrolled in the study. The incidence of GDM was counted. According to presence or absence of GDM,the subjects were divided into GDM group and non-GDM group. General data and serum 25OHD3 levels were compared between the two groups. According to the serum 25OHD3 levels in mothers and infants of GDM group,they were divided into deficiency group,insufficiency group and sufficiency group. The levels of FINS and inflammatory factors (interleukin-6 (IL-6),interleukin-7 (IL-7),interleukin-17 (IL-17),tumor necrosis factor-α (TNF-α) were compared among groups,and correlation analysis was conducted. Results Of the 629 pregnant women,62 had GDM (9.86%). The proportion of pregnant women not under 30 years old was significantly higher in GDM group than in non-GDM group (P＜0.05). Maternal and neonatal serum 25OHD3 levels of the GDM group were significantly lower than those of the non-GDM group (P＜0.05). From the maternal aspect,serum FINS significantly increased in turn,while serum IL-6,IL-7,IL-17,TNF-α,CRP,and fasting blood glucose significantly decreased in turn in the deficiency group,insufficiency group and sufficiency group,with significant differences between any two groups (P＜0.05). From the infant aspect,serum FINS significantly increased,while serum IL-6,IL-7,IL-17,TNF-α,CRP,and fasting blood glucose significantly decreased in turn in deficiency group a,insufficiency group a and sufficiency group a,with significant differences between any two groups (P＜0.05). Serum IL-6,IL-7,IL-17,TNF-α and CRP in each group showed no obvious changes at 6,24,and 48 hours after birth (P＞0.05). Pearson analysis indicated that serum 25OHD3 levels of GDM mothers and infants were significantly associated with the levels of FINS and inflammatory factors (P＜0.05). Conclusion The levels of serum 25OHD3 in GDM mothers and infants show a significant decreasing trend,which are significantly positively correlated with FINS,but negatively correlated with the expression levels of inflammatory factors. The above findings indicate that serum 25OHD3 level is expected to provide guidance for the reasonable prevention of GDM.