Gestational diabetes mellitus (GDM), which constitutes an estimated 86.4% of hyperglycemic pregnancies, is characterized by the onset of glucose intolerance with identified or diagnosed for the first time during pregnancy (1, 2). As the predominant form of hyperglycemia complicating pregnancies, GDM potentially results in a series of short-term and long-term complications for both maternal and fetal health (3–5). Consequently, clinical practice guidelines underscore the necessity for stringent glycemic control as a paramount objective throughout the whole pregnancy (6, 7). However, an oral glucose tolerance test (OGTT) for screening glucose metabolism during pregnancy is administered in middle and late pregnancy, conventionally between 24 to 28 weeks of gestation, or occasionally delayed until 32 weeks. It remains less time for pregnant women at risk of GDM to optimize glycemic control through lifestyle management or medication in order to reduce the duration of elevated blood glucose exposure. Therefore, early identification and diagnosis base on some simple serological indicators, and timely and effective intervention for GDM to achieve glycemic benchmarks and create a conducive intrauterine environment is of great significance to the mother and fetus.

Glypican-4, a novel adipokine sensitive to insulin and produced by adipose tissue, plays a pivotal role in insulin signaling by binding directly to the insulin receptor (8). Gesta and colleagues firstly discovered that the expression of glypican-4 varied between the visceral and subcutaneous adipose tissues of rodents and humans (9). Intriguingly, this expression pattern significantly correlated with several measures of obesity, including body fat content, body mass index (BMI) and waist-to-hip ratio. Additionally, it showed an independent association with insulin resistance (IR), as measured by euglycemic hyperinsulinemic clamps. Of greater significance, serum levels of glypican-4 have been found to be linked to key metabolic indicators such as the ratio of visceral to subcutaneous fat area, fasting plasma insulin (FINS) and the homeostasis model assessment of insulin resistance (HOMA-IR), highlighting its potential as a biomarker for metabolic disorders associated with obesity and IR (10, 11).

However, no longitudinal studies have documented the serum glypican-4 trajectories in GDM patients, particularly in the context of insulin therapy or postpartum glucose metabolism. In the present study, we address this gap by examining a cohort of pregnant women from Northeast China, to elucidate the association between glypican-4 and the risk of developing GDM, as well as its potential effects on insulin therapy and postpartum glucose metabolism in GDM patients for the first time.

The latest IDF data have revealed that the prevalence of GDM in pregnancies worldwide ranges from 7.1% to 27.6% (with a global prevalence of 14.0%), attributed to the increasing maternal age and obesity rates (1). Therefore, early identification of hyperglycemia in pregnancy is crucial for preventing adverse events. Currently, the precise etiological mechanism underlying GDM remains unclear; however, a large number of studies have indicated that IR is considered the hallmark of GDM pathogenesis (17, 18). It is widely acknowledged that insulin, a key hypoglycemic hormone, must bind with the insulin receptor to exert its biological effects through a series of intricate enzymatic reactions. Any abnormality in the involved signal cascade reaction could lead to IR (19, 20). IR can reduce the responsiveness of target organs or peripheral tissues to insulin, thereby affecting glucose output, uptake and utilization, as well as fat and protein synthesis and energy metabolism (20, 21). Under normal pregnancy, insulin sensitivity in target tissues such as fat tissue and skeletal muscle, and glucose handling capacity by insulin are lower compared with the pre-pregnancy status; however, the compensatory increase in insulin secretion could maintain normal blood glucose levels. By contrast, the degree of IR in individuals with GDM is significantly higher than that of normal healthy pregnant women, and the pancreatic β-cell dysfunction and the progressive decrease in insulin secretion cannot compensate for the increase in IR (17, 18).

Glypicans, a class of heparan sulfate proteoglycans, are anchored to the extracellular surface of the plasma membrane via glycosyl-phosphatidylinositol (GPI) linkages and have played a pivotal role in modulating signaling pathways, including those of Wnt, Hedgehog, fibroblast growth factors, and bone morphogenetic proteins (22). Glypican-4, a member of this GPI-anchored proteoglycan family, is synthesized and secreted by adipose tissue and has been shown to interact directly with the insulin receptor, modulating its activation and the subsequent signaling cascade (22). The regulatory mechanism involving glypican-4 and insulin sensitivity is linked to GPI-specific phospholipase D (GPLD), an enzyme implicated in the release of glypican-4 from the cell surface into circulation, with its activity being subject to regulation by insulin (23). Increased insulin levels have been demonstrated to upregulate GPLD-1 activity, leading to enhanced glypican-4 shedding from the cell surface and consequently, elevated levels of circulating glypican-4, which has been correlated with IR (22).

Gesta et al. reported elevated serum glypican-4 levels in insulin-resistant obese subjects, with a twofold increase compared to those with insulin sensitivity, matched for age, sex, and BMI, and noted a positive association with both FINS and HOMA-IR (9). Zhu et al. also observed that obese individuals with IR have higher serum glypican-4 levels, and glypican-4 was closely related to a variety of metabolic-related parameters, including BMI, systolic blood pressure, alanine aminotransferase, aspartate aminotransferase, FINS and HOMA-IR (10). A recent adult study demonstrated that individuals with metabolic syndrome (MetS) exhibited significantly elevated serum glypican-4 levels when contrasted with those of healthy and non-MetS counterparts (11). An elevated glypican-4 level could be an independent risk for MetS in the Chinese Han population, indicating a heightened likelihood of MetS development among individuals with increased levels (11). In concordance with prior research, serum glypican-4 levels were significantly elevated in individuals with impaired glucose tolerance (24). Conversely, serum glypican-4 levels were paradoxically lower in newly diagnosed T2DM patients compared to those with normal pregnant women, and reduced glypican-4 levels also observed in an ob/ob mice model with elevated blood glucose and hyperinsulinemia, suggesting a complex role for glypican-4 in glucose metabolism dysregulation (8, 24). Yoo et al. further delineated sex-specific disparities in serum glypican-4 levels within Asian cohorts and a significant correlation with cardiometabolic risk factors, such as body fat distribution, arterial stiffness, and IR (25). Elevated serum glypican-4 levels have also been observed in women with nonalcoholic fatty liver disease, indicative of its hepatic involvement in MetS (25). Additionally, in women with PCOS presenting metabolic risk factors, higher glypican-4 levels were associated with established cardiovascular disease predictors, including IR, hyperglycemia, hypertension, and dyslipidemia, with a notable emphasis on adipose tissue distribution (26). Serum glypican-4 levels have also been reported to escalate with the severity of obesity in pediatric populations aged 6 to 18 years (27). Collectively, these findings underscore the important role of glypican-4 in obesity and IR, and its utility as a metabolic disorder biomarker. However, no existing evidence has reported the longitudinal data on glypican-4 trajectories in GDM, particularly in the context of insulin therapy or postpartum. This study pioneers the examination of the association between maternal glypican-4 levels and GDM risk, as well as its predictive value for insulin therapy requirements and postpartum glycemic outcomes.

Our study showed that serum glypican-4 levels in GDM patients were higher than healthy pregnant women in different trimesters throughout the whole pregnancy, with significant differences observed in middle and late pregnancy. The study by Deischinger et al. also reported that glypican-4 levels in GDM patients during pregnancy increased progressively with advancing gestational weeks, but no differences were noted in comparison to healthy pregnant women (28). The study was limited by a smaller number of participants, all of whom were obese with higher BMI, differing significantly from the demographic in our study, which may account for the lack of observed differences between the groups. It is widely recognized that maternal IR progressively deteriorates throughout pregnancy, becoming more pronounced during the mid-to-late pregnancy and reaching a peak at approximately 32 weeks of gestation. Compared to healthy pregnant women, patients with GDM experience more severe IR. Our study found that serum glypican-4 levels were significantly elevated in GDM patients during the second and third trimesters of pregnancy, which was consistent with the characteristics of changes in IR during pregnancy, and exhibited the potential of glypican-4 as a serological marker associated with IR. Moreover, maternal glypican-4 levels during pregnancy demonstrated a significant positive correlation with key diagnostic indicators of glucose metabolism at the time of GDM diagnosis in our study. An elevation in glypican-4 levels was associated with an increased risk of GDM, with a statistically higher risk observed in pregnant women with glypican-4 levels exceeding 0.40 ng/mL in the first trimester (5-12 weeks of gestation) or 0.79 ng/mL in the second trimester (13-23 weeks of gestation). These results suggest that glypican-4 during pregnancy was closely associated with glucose metabolism and highlighted the potential of glypican-4 as a predictive serum marker for the identification and clinical diagnosis of GDM.

Medical nutrition therapy is a fundamental intervention for managing hyperglycemia in pregnancy, which followed individualization principle (6, 13). The intervention principles should ensure appropriate nutrition and calories intake of pregnant women, normal growth and development of the fetus, and better blood glucose maintenance to avoid ketosis (6, 13). The majority of patients with GDM can achieve optimal glycemic control through evidence-based dietary and exercise interventions; however, for those who fail to effectively control blood glucose, healthcare providers encounter a significant challenge in the early identification of high-risk cohorts and the strategic initiation of insulin therapy.

In our current study, 136 of the 373 patients with GDM necessitated supplementary insulin therapy in conjunction with ongoing lifestyle interventions to attain the predefined glycemic control targets. We found that serum glypican-4 levels in GDM with requirement for insulin therapy were higher than them with medical nutrition therapy only. Glypican-4 was a potential predictor of insulin therapy during pregnancy, a significantly increased insulin requirement with increased glypican-4 levels. It was more necessary for GDM with glypican-4 exceeding 0.87 ng/mL to require insulin therapy during pregnancy to manage blood glucose.

To the best of our knowledge, women with a history of GDM are considered to be at a significantly elevated risk for the development of diabetes, with a nearly 10 times than healthy women (29, 30). However, currently only a few patients with GDM are screened for diabetes after delivery. It is largely attributed to the insufficient understanding of diabetes and the perceived complexity of OGTT, leading to a reduced rate for early detection of postpartum glucose disorders and a subsequent increased incidence of long-term diabetes. A large retrospective study had demonstrated a significant escalation in the cumulative incidence of diabetes, rising from 2.6% at 6 weeks to over 70% within 28 years postpartum, with a particularly pronounced increase within the first five years postpartum, a period recognized as a critical window for the development of diabetes (31). Furthermore, GDM patients also confront an approximately fourfold higher risk of prediabetes, encompassing IFG and/or IGT, compared to the general female population (32). Therefore, our study determined the relationship between glypican-4 level during pregnancy and subsequent postpartum glucose metabolism in patients with GDM. A cohort of 158 GDM participants were consecutively enrolled to undergo a 75-g OGTT as part of their postpartum evaluation, of which 18 had pre-diabetes and 2 had T2DM. Participants with abnormal glucose tolerance test results had higher glypican-4 level than healthy controls throughout the whole pregnancy; however, the observed difference failed to reach statistical significance, potentially attributable to the smaller postpartum follow-up sample size and the inadequate follow-up time for OGTT assessment of glucose metabolism.

This study has several advantages. First, the trajectories of maternal serum glypican-4 concentrations throughout the whole pregnancy are first measured in this longitudinal study. Second, our study presents the first comprehensive examination of the association between serum glypican-4 levels and the risk of GDM development. Simultaneously, it elucidates the biomarker’s predictive utility in ascertaining the requirement for insulin therapy and profiling the postpartum glycemic status. However, this study also has certain limitations. Our investigation has a constrained sample size screening for glucose metabolism after delivery and lacks postpartum data on glypican-4 level. Consequently, it is limited in fully exploring the relationship between glypican-4 and the long-term risk of developing metabolic disorders such as diabetes.

In conclusion, our study introduces novel findings that establish a significant association between maternal glypican-4 levels during pregnancy and the risk of GDM development. We observe a substantial increase in GDM risk with elevated glypican-4 levels, particularly when glypican-4 levels exceeding 0.40 ng/mL in the first trimester (5-12 weeks of gestation) or 0.79 ng/mL in the second trimester (13-23 weeks of gestation). Importantly, a glypican-4 level above 0.87 ng/mL is identified as a critical threshold for GDM patients necessitating insulin therapy to maintain glycemic control during pregnancy. These insights hold profound clinical implications, enabling healthcare providers to swiftly and accurately identify pregnant women at high risk of GDM. Early and accurate diagnosis, coupled with timely intervention, is essential for the management of blood glucose levels and the safeguarding of both maternal and fetal health. It is imperative to enhance awareness among these women regarding the importance of regular glucose metabolism screening and adherence to lifestyle modifications throughout their lives, as preventive measures against the development of T2DM. Furthermore, as a potential serological indicator, further research into glypican-4 is conducive to comprehensively understand the pathophysiological mechanisms of GDM.