The Correlation Analysis Between the Blood Lipids, Glycosylated Protein Level and Inammatory Factors of Pregnant Women with GDM During Pregnancy and the Weight of Newborn

The study was carried to explore the correlation between blood lipids, blood glucose levels, inammatory factor and weight of newborn, to provide reference for control of blood glucose in pregnant women with gestational diabetes mellitus (GDM) and early screening of macrosomia. Fifty pregnant women (give birth to newborn) with GDM were selected as research group, and 55 normal pregnant women (give birth to newborn) as control group. Blood lipid levels of TC, TG, HDL-C, LDL-C, the fasting blood glucose (FPG), HbAlc, glycosylated albumin (GA), and expression of inammatory factor TLR4 of pregnant women in the two groups were monitored. The levels of TG, FPG, HbA1c and GA in pregnant women, the levels of TLR4 in cord blood of newborn, the relative expression of TLR4 protein and TLR4mRNA in the placenta were higher than in control group, and level of HDL-C was lower than the control group. The levels of TC and LDL-C of pregnant women were higher than in control group. Weight of newborn was positively correlated with these all except HDL-C levels (negatively correlated) and no correlation was found with TC and LDL-C. The weight of newborn and incidence of macrosomia in research group were higher compared to control group, and scores of newborns at 1 min and 5 min were lower compared to control group. The results revealed that strengthening the detection of blood lipid and blood glucose during pregnancy can prevent adverse outcomes such as giant babies and improve the quality of birth.


Introduction
A series of changes in the secretion system and hormone levels of pregnant women during pregnancy will trigger changes in glucose and lipid metabolism [1]. The rst occurrence or discovery of abnormal glucose metabolism during pregnancy is called Gestational Diabetes Mellitus (GDM), and its onset is related to insulin resistance and decreased secretion of islet cells [2][3]. Pregnant women with GDM during pregnancy are prone to adverse pregnancy outcomes, which increase the risk of huge babies, neonatal hypoglycemia, deformity, asphyxia and even death. And pregnant women with GDM during pregnancy have a high probability of developing T2DM [4]. Therefore, GDM is more harmful to mothers and babies.
With the development of the economy, the living standards have improved signi cantly, and the gestational age of women has gradually been pushed back. Many pregnant women have supplied excessive nutrition during pregnancy, lack of necessary exercise and other factors, resulting in an increasing trend in the incidence of GDM year by year [5]. Early diagnosis of GDM patients and early intervention are of great signi cance for reducing complications of GDM pregnant women and newborns, and improving adverse pregnancy outcomes.
Disorders of glucose and lipid metabolism are the key to GDM research. GDM pregnant women have higher levels of VLDL, TC, and TG in blood circulation [6]. When pregnant women are diagnosed, they mainly rely on measuring blood glucose at fasting and 2 hours after meal to guide pregnant women to control blood glucose during pregnancy, but the stability of immediate blood glucose is low and blood glucose control is di cult to standardize [7]. The detection of the levels of glycosylated hemoglobin and glycosylated albumin does not require fasting, and it has a good stability. Therefore, it is more suitable as an indicator for the detection of blood glucose in pregnant women with GDM [8][9]. In recent years, domestic and foreign studies have shown that chronic aseptic in ammatory response is also closely related to GDM. When pancreatic islet cell function is impaired, insulin resistance is aggravated, and insulin resistance is one of the recognized pathogenic factors of GDM [10].
Based on a large number of domestic and foreign literatures, this study collected and analyzed the correlation between the levels of blood lipid, glycosylated protein and in ammatory factors of GMD pregnant women and the weight of newborns, and provided theoretical basis for the control of blood glucose in pregnant women with GMD and early screening of macrosomia. The detailed descriptions are demonstrated as follows: 2. Materials And Methods 2.1. General information 50 GDM pregnant women and their newborns who were hospitalized in the obstetrics department of our hospital from August 2020 to January 2021 were selected as the research group, and 55 normal pregnant women and their newborns who were hospitalized during the same period were selected as the control group.
Considering IADPSG as the diagnostic criteria for pregnant women with GDM, 75g OGTT was performed to screen whether pregnant women are GDM at the rst visit between 24-28 weeks of pregnancy [11].
When the pregnant woman's fasting blood glucose is ≥ 5.1mmol/L or 1-hour blood glucose ≥ 10mmol/L or 2 hours' blood glucose ≥ 8.5mmol/L, GDM can be diagnosed when any one of the above is met.
Inclusion criteria: 20-35 years old; Natural conception; Diagnosed with GDM at 24-28 weeks of gestation; No abnormalities such as Down syndrome, malformation, etc. found on various screenings during pregnancy; Follow-up examinations will be performed every 4 weeks after diagnosis until delivery.
Exclusion criteria: Diabetes complicated with pregnancy; Congenital or genetic diseases; Multiple pregnancy; Pregnant women with other chronic diseases during pregnancy.
According to the situation of GDM, 50 GDM pregnant women and their newborns were taken as the research group, and 55 normal pregnant women who were hospitalized during the same period were taken as the control group. The general clinical data of age, gestational week, gestation times and prepregnancy weight in two groups were not signi cantly different (P > 0.05), and they were comparable.
On the basis of obtaining the consent of the pregnant woman and their family members, the informed consent form is signed, and they voluntarily joined the research project. Then we collect relevant information of pregnant women, and the information comes from various clinical examination index values. This study has been reviewed and approved by the ethics committee of our hospital.

Determination of blood lipid
After 12 hours of fasting, the pregnant women of the two groups pumped 10ml of cubital venous blood in the morning. After the blood was drawn, the upper serum was obtained after anticoagulation and centrifugation. Cobas-8000 automatic biochemical analyzer (Roche, Germany) was used to determine the TC, TG, HDL-C, and LDL-C in serum.

Determination of blood glucose
After 12 hours of fasting, the pregnant women of the two groups pumped 10ml of cubital venous blood in the morning. After the blood was drawn, the upper serum was obtained after anticoagulation and centrifugation. MQ-600 glycosylated hemoglobin analyzer (Nanjing Hanyu Medical Technology Co., Ltd.) was used to determine the levels of glycosylated hemoglobin (HbAlc) and glycosylated albumin (GA).

Determination of in ammatory factors
After the umbilical cord was broken at birth, 5ml of cord venous blood was drawn from the newborn. After centrifugation, the upper serum was placed in a cryotube and stored in a refrigerator at -20°C. Enzymelinked immunoassay (ELISA) was used to detect the expression level of TRL4, operate according to the instructions of the kit (Wuhan Huamei Biological Co., Ltd.), repeat twice and take the average value.
The imprinting method of Western Blot was used to detect the protein level of TLR4 in the placenta.
RT-PCR method was used, RNA puri cation and ampli cation was performed through extraction, separation, precipitation, washing, the mRNA expression level of TLR4 in the placenta was detected.

The level of Blood Lipid
Clinically, TC, TG, HDL-C, LDL-C indicators are used to re ect the blood lipid levels of pregnant women. Diabetes during pregnancy can cause fat metabolism disorders, which in turn triggers changes in the levels of TC, TG, HDL-C, and LDL-C [12].

The Level of Blood Glucose
Clinically, the fasting blood glucose (FPG) index is used to re ect the blood glucose level of pregnant women when measured. FPG is the micro-control index of blood glucose.
HbAlc is a ketamine compound formed by the condensation of hemoglobin and glucose in the blood. The condensation process is slow and irreversible. The level of HbAlc is related to the degree of glycosylation.
The measurement of this indicator can re ect the average blood glucose level of pregnant women 2-3 months before measurement [13].
GA is a kind of ketamine compound formed by the non-enzymatic reaction between the 4 Lysines of glucose and the N-terminus of serum albumin. And the structure of GA is relatively stable. The half-life period of GA is the same as that of albumin, which can re ect the overall level of serum FPG. The measurement of this indicator can re ect the average blood glucose level of pregnant women 2-3 weeks before the measurement [14].
The three indicators re ect at different times and have their own characteristics and clinical signi cance.
Thus, the combined detection of the three indicators was used in this paper during pregnancy to re ect the recent and full blood glucose level of the pregnant woman.
2.2.6. The Level of In ammatory Factor TLR TLR4 is the most important receptor in the TLR family. The enhancement of maternal immune system mediated by TLR4 during pregnancy will aggravate insulin resistance [15][16]].

The Apgar Score of Newborn
The ve physical signs of newborn (heartbeat, breathing, muscle tension, stress response and skin color) is analyzed at 1 and 5 minutes after delivery based on the Apgar score sheet. Each item is scored 0-2 points, and the total score is 10 points. The status of the newborn is determined according to the total score. The higher the score is, the better the state of newborn is [17].

Comparison in the general clinical indicators of pregnant women between the two groups
Through comparison, it is found that the difference in general clinical indicators such as age, gestational week, and gestational times of two groups are not statistically signi cant (P > 0.05), and they are comparable (Table 1).

Comparison in the blood lipid levels of pregnant women between the two groups
The TG level of pregnant women in the research group is 3.30 ± 1.09 mmol/L, which was signi cantly higher than the TG level (2.79 ± 1.07) mmol/L of pregnant women in the control group, and the level of HDL-C (1.72 ± 0.46) mmol/L was signi cantly lower than that in the control group (2.10 ± 0.42) mmol/L, and the differences in the levels of TG and HDL-C of pregnant women between the two groups were statistically signi cant (P < 0.05). The level of TC (6.18 ± 1.20) mmol/L and the level of LDL-C (3.30 ± 1.01) mmol/L in the research group were slightly higher than those in the control group (5.82 ± 1.14) mmol/L and (3.14 ± 0.81) mmol/L, respectively, But the difference between the two groups was not statistically signi cant (P > 0.05) (Fig. 1). The weight of newborn was positively correlated with the TG level of pregnant women in the research group, and negatively correlated with the HDL-C level. However, no correlation was found with TC and LDL-C levels ( Table 2).

Comparison in the blood glucose levels of pregnant women between the two groups
The FPG level (4.82 ± 1.54) mmol/L, HbA1c level (7.54 ± 1.83) % and GA level (3.52 ± 1.05) of pregnant women in the research group were signi cantly higher than those in the control group (4.16 ± 1.10) mmol/L, (4.95 ± 2.17)% and (2.14 ± 0.97) mmol/L, and the differences were statistically signi cant (P < 0.05) (Fig. 2). The weight of newborns was positively correlated with the levels of FPG, HbA1c and GA in the cord blood of pregnant women in the research group (Table 3).

Comparison in the TLR4 levels of pregnant women between the two groups
The level of TLR4 (4193.02 ± 799.53) pg/mL in the umbilical cord blood of newborn in the research group was signi cantly higher than that in the control group (3669.26 ± 769.62) pg/mL. The relative expression of TLR4 protein and TLR4mRNA in the placenta of the pregnant women in the research group were respectively signi cant higher than those in the control group, and the differences were statistically signi cant (P < 0.05) (Fig. 3). The weight of the newborn was positively correlated with the level of TLR4 in the cord blood of the pregnant women, the relative expression of TLR4 protein in the placenta and TLR4mRNA (Table 4) in the research group.

Comparison in the situation of newborns between the two groups
The birth weight of newborns was (3646.05 ± 391.97) g and the incidence of macrosomia was 18.00% in the research group, which were both signi cantly greater than those in the control group (The birth weight of newborns was 3394.06 ± 388.26 and the incidence of macrosomia was 7.27%), and the difference was statistically signi cant (P < 0.05). The scores of newborns at 1 min and 5 min in the research group were lower than those in the control group, and the difference was statistically signi cant (P < 0.05) (Fig. 4).

Discussion
Under the combined action of a variety of hormones during pregnancy, the blood lipid level of pregnant women will remain in a high balance during the entire pregnancy, which can not only meet the fetus's needs for nutrients, but also store raw materials for postpartum breastfeeding [18]. The level of TG starts to increase at the 9th week of pregnancy, while TC and LDL-C decrease. At the 12th week of pregnancy, HDL-C and PL continue to increase, especially for pregnant women with GDM [19]. At present, domestic and foreign studies have con rmed that the blood glucose level of pregnant women with GDM will cause the newborn to grow too quickly in the uterus, and even large babies may appear [20]. Research by Hashimoto et al. also showed that there is a linear relationship between GA and adverse pregnancy outcomes [21]. Although the blood glucose level of pregnant women with GDM can be controlled at a reasonable level through drugs, the incidence of macrosomia has not been reduced, which proves that the incidence of macrosomia is related to the blood lipid level [22].
One of the recognized pathogenic factors of GDM is insulin resistance, and chronic in ammation can aggravate insulin resistance [23][24]. Domestic and foreign studies have shown that the expression levels of TLR4, NF-κ B and in ammatory factors (such as TNF-α, IL-6 and CRP) in placental tissue or peripheral blood of GDM pregnant women are higher than those in normal pregnant women [25][26]. Giannoukakis et al. used IκB to inhibit the expression of NF-κB and reduce the damage of in ammatory factors to pancreatic β cells [27]. These studies prove that TLR4, NF-κB, and in ammatory factors are closely related to GDM.
In this study, the blood lipid levels (TC, TG, HDL-C, LDL-C), the blood glucose levels (FPG, HbAlc, GA), and in ammatory factor TLR4 expression levels in two groups were monitored. The results showed that the levels of TG, FPG, HbA1c, GA in pregnant women, TLR4 in cord blood of newborn, and the relative expression of TLR4 protein and TLR4mRNA in the placenta in the research group were signi cantly higher than those in the control group (P<0.05), and the level of HDL-C was signi cantly lower than that in the control group (P<0.05). The weight of newborns was positively correlated with the levels of TG, FPG, HbA1c, and GA of pregnant women in the research group, the levels of TLR4 in the umbilical cord blood of pregnant women, and the relative expression of TLR4 protein and TLR4mRNA in the placenta, and negatively correlated with the levels of HDL-C, but there is no correlation with TC and LDL-C was found. The reason may be that the high levels of blood glucose and blood lipid in pregnant women with GDM will directly enter the newborn's body through the umbilical cord, causing the newborn's blood glucose to rise. The stimulation of pancreatic islet cells causes the newborn's hyperinsulinemia. When glucose and fat continue to accumulate in the newborn's body, excessive growth even macrosomia phenomenon will be easy to occur.

Conclusions
In summary, changes in the levels of blood lipids (TG and HDL-C), blood glucose (FPG, HbA1c and GA) of pregnant women with diabetes during pregnancy will affect the weight of newborns. Therefore, the monitoring of blood lipids and blood glucose of pregnant women with GDM should be strengthened to reduce the risk of mother and child. The innovation of this study is to establish the correlation between the levels of blood glucose, blood lipids and TLR4 of pregnant women with GDM and the weight of newborns, expecting to provide a theoretical basis for reducing the adverse pregnancy outcomes of pregnant women with GDM and reasonably controlling the blood glucose and blood lipids during pregnancy. The de ciencies of this study are as follows: (1) The sample included is small, which makes the research results lack of universality. (2) The research is only conducted on the expression level of TLR4, and the results obtained are biased. In view of the above shortcomings, the next step is to expand the sample of GDM pregnant women and reduce the analysis error caused by the limited sample, expecting that more representative and scienti c conclusions can be obtained, and reference for improving the pregnancy outcome of GDM pregnant women can be provided. Availability of data and materials Data are available upon reasonable request from correspondence.

Competing interests
All the authors declare that they have no con ict of interest.

Funding
Authors received no research funding for this study.
Author contributions CH and LW conceived, designed and supervised the study, interpreted the data, prepared the gure with the related legend, and wrote the manuscript; JF analyzed the data; CH and JF performed the literature search. All authors contributed to editorial changes in the manuscript. All authors read and approved the nal manuscript.   Analysis of the TLR4 levels of pregnant women in the two groups. A) the levels of TLR4 in the umbilical cord blood of the pregnant women in research group and control group, B) the relative expression of TLR4 protein in the placenta of the pregnant women in research group and control group, and C) TLR4 mRNA in the placenta in the research group and control group, Data are expressed as mean ± SD, where "*" is (P<0.05).