The Mediating Effect of Insulin on the Influence of 25(OH)D in Recurrent Pregnancy Loss: A Case–Control Study

Abstract

Background:

Vitamin D deficiency (VDD) and insulin resistance (IR) are well-known risk factors for recurrent pregnancy loss (RPL). Since VDD may contribute to the development of IR, this study aimed to investigate the role of fasting insulin (FINS) levels and the homeostasis model assessment of insulin resistance (HOMA-IR) in the association between vitamin D and RPL.

Methods:

A total of 934 women were retrospectively analyzed between 2019 and 2022, including patients with RPL and age-matched controls. Clinical and biochemical data were collected, including serum 25(OH)D, FINS, fasting blood glucose (FBG), HOMA-IR, and sex hormone levels. Correlation, multivariate logistic regression, restricted cubic spline (RCS), and mediation analyses were conducted.

Results:

Compared to controls, the RPL group exhibited lower levels of 25(OH)D and higher levels of FINS and HOMA-IR. In the RPL group, 25(OH)D was negatively correlated with FINS and HOMA-IR. Higher levels of 25(OH)D were associated with reduced RPL, whereas elevated FINS and HOMA-IR levels were linked to an increased risk. A mediation analysis confirmed that FINS and HOMA-IR partially mediated the relationship between vitamin D and RPL, accounting for 10.9% and 10.7% of the total effects, respectively.

Conclusions:

VDD is closely associated with increased RPL risk, potentially through impaired glucose metabolism. Therefore, improving vitamin D status and insulin sensitivity may help in reducing pregnancy loss and enhancing reproductive outcomes.

Graphical abstract

Keywords

vitamin D / recurrent pregnancy loss / fasting insulin / insulin resistance / mediating effect

1. Introduction

Vitamin D is a fat-soluble vitamin that plays an essential role in maintaining the calcium and phosphorus balance, promoting bone health. In recent years, it has been recognized for its essential biological functions in immune regulation, inflammation inhibition, cell proliferation, and differentiation [1, 2, 3]. Recurrent pregnancy loss (RPL) is a common pregnancy complication that has generated significant concern [4]. Although the exact etiology remains unclear, more and more evidence shows that vitamin D deficiency (VDD) may be closely related to the occurrence and development of RPL [5, 6, 7]. Previous studies have found that with the increase of vitamin D level, the clinical pregnancy rate (CPR) also increases, while the miscarriage rate (MR) decreases [6, 8]. These findings support the crucial role of vitamin D in pregnancy, especially in avoiding RPL.

Glucose metabolism plays a vital role during pregnancy, providing necessary energy and nutrition for the normal development of embryos [9, 10]. Some studies have found a complex interaction between vitamin D and glucose metabolism [11, 12, 13, 14]. Vitamin D receptor (VDR) is widely expressed in islet cells, suggesting that vitamin D may affect glucose metabolism through insulin secretion and resistance [15, 16]. Therefore, exploring the relationship between VDD and RPL, as well as its potential mechanistic involvement in glucose metabolism, is essential for understanding the etiology of RPL and finding preventative and therapeutic strategies.

This study explores the potential role of vitamin D in the occurrence and development of RPL and whether there is a mechanism related to glucose metabolism disorder. The result of the study will help provide a theoretical basis for clinical intervention research and individualized treatment in the future.

2. Materials and Methods

2.1 Study Population and Grouping

This retrospective study analyzed 934 patients who visited the Reproductive Medicine Center of the Second Hospital of Lanzhou University between September 2019 to February 2022. This study was approved by the hospital’s ethics committee. Inclusion criteria were as follows: ① Age between 18 and 40 years; ② Natural conception and no use of assisted reproductive technologies; ③ For the RPL group: history of two or more consecutive spontaneous or induced abortions before 14 weeks of gestation, consistent with the clinical diagnostic criteria for RPL; ④ For the control group: age-matched women without a diagnosis of RPL. Women with no history of pregnancy loss or with one or two isolated (non-recurrent) early miscarriages were eligible for inclusion in the control group, provided they had at least one successful pregnancy. Exclusion criteria included: ① Use of assisted reproductive technology (e.g., in vitro fertilization, intracytoplasmic sperm injection); ② Known chromosomal abnormalities; ③ Chronic metabolic or endocrine diseases (e.g., diabetes, thyroid dysfunction, cardiovascular diseases); ④ Use of medications known to affect glucose or lipid metabolism; ⑤ Vitamin D supplementation within 3 months before or during pregnancy; ⑥ More than four previous pregnancies; ⑦ Severe pregnancy complications such as gestational hypertension, early fetal demise or placental insufficiency.

The included population was divided into two groups according to their history of pregnancy: the RPL group and the normal control group. Then, according to the serum 25(OH)D level, the patients in the RPL group were divided into the VDD group [defined as 25(OH)D 30 nmol/L], the vitamin D insufficiency group [defined as 30 nmol/L \le 25(OH)D 50 nmol/L], and the vitamin D sufficiency group [defined as 50 nmol/L \le 25(OH)D] [17].

2.2 Research Indicators and Outcomes

Maternal baseline data were obtained from the hospital’s electronic medical record system. Collected variables included sociodemographic characteristics (e.g., maternal age, education), reproductive history (e.g., gravidity, parity, pregnancy loss), lifestyle factors (e.g., smoking, alcohol consumption, manual labor), and clinical measurements (e.g., blood pressure, lipid profile). Pregnancy outcomes were obtained through a review of hospital records and telephone follow-ups.

The primary outcomes of this study were the occurrence of RPL, serum 25(OH)D level, fasting insulin (FINS) level, and the homeostasis model assessment of insulin resistance (HOMA-IR), to investigate the effect of vitamin D on RPL and the potential mediating role of FINS and HOMA-IR. Secondary outcomes included CPR (defined as the presence of an intrauterine gestational sac and yolk sac), clinical live birth rate (LBR) (defined as the delivery of a live fetus after 28 weeks of gestation), MR (defined as pregnancy losses before 20 gestational weeks, including spontaneous abortion and termination due to intrauterine fetal death) and sex hormone levels [18, 19, 20].

Serum biochemical parameters were measured using standardized protocols and were measured during the early follicular phase (days 2–5 of the menstrual cycle) during routine preconception or reproductive evaluations. All participants were in a non-pregnant state at the time of blood collection. The measured parameters include 25(OH)D, FINS, estradiol (E2), luteinizing hormone (LH), and follicle-stimulating hormone (FSH) were measured using an electrochemiluminescence immunoassay on the cobas® 8000 analyzer (Roche Diagnostics, e801, Mannheim, Germany), in accordance with the manufacturer’s instructions. Lipid profiles, including total cholesterol (TC), triglycerides (TG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C), were determined by enzymatic colorimetric methods using the cobas® 8000 analyzer (Roche Diagnostics, c702, Mannheim, Germany). Fasting blood glucose (FBG) was measured using the glucose oxidase-peroxidase method on the cobas® 8000 fully automated biochemical analyzer (Roche Diagnostics, c702, Mannheim, Germany). HOMA-IR was calculated as [FINS (mIU/L) \times FBG (mmol/L)]/22.5 [21].

2.3 Statistical Analysis

Statistical analyses were conducted using SPSS version 26.0 (IBM Corp., Armonk, NY, USA) and R version 4.2.2 (R Foundation for Statistical Computing, Vienna, Austria). Graphs were created using the ggplot2 (https://ggplot2.tidyverse.org) and plotRCS (https://cran.r-project.org/src/contrib/Archive/plotRCS/) packages. Missing data were handled using multiple imputations with chained equations, assuming data were missing at random. A two-sided p-value 0.05 was considered statistically significant throughout the analysis. The Kolmogorov-Smirnov test was used to assess normality. Normally distributed variables were reported as mean \pm standard deviation and compared by one-way analysis of variance. Non-normally distributed variables were expressed as median (P25, P75) and compared by the Mann-Whitney U test. Categorical variables were expressed as frequencies (ratios) and compared using chi-square or Fisher’s exact test, as appropriate. Spearman correlation was employed to evaluate associations between serum 25(OH)D and FINS or HOMA-IR. In the RPL group, multiple linear regression was conducted to examine the relationship between 25(OH)D and FINS or HOMA-IR. Multivariate logistic regression models were used to assess the associations between 25(OH)D, FINS, HOMA-IR, and the risk of RPL. Odds ratio (OR) and 95% confidence interval (CI) were reported. To evaluate potential nonlinear relationships, restricted cubic spline (RCS) logistic models were fitted using the rms package (v8.0-0, https://hbiostat.org/R/rms/), with 4 knots at default percentiles. Nonlinear and total effects were assessed, and results were visualized via spline curves. Mediation analysis was performed using the PROCESS macro in SPSS (v4.1, https://processmacro.org/download.html) with 5000 bootstrap samples to estimate indirect, direct, and total effects and their 95% CI. The mediation proportion was calculated as the ratio of the indirect to total effect. Statistical significance was defined as a 95% CI that excludes zero.

3. Results

3.1 Baseline Characteristics of the Study Population

Table 1 summarizes the baseline characteristics of women in the RPL group and the control group. Compared with the control group, women in the RPL group had a significantly higher cumulative number of pregnancies (3.00 vs. 1.00) and prior pregnancy losses (2.00 vs. 1.00), but fewer previous live births (0.00 vs. 1.00). The RPL group also exhibited lower serum 25(OH)D levels (31.16 nmol/L vs. 35.05 nmol/L) and a higher proportion of VDD (46.10% vs. 34.90%). Additionally, FINS levels (9.77 mIU/L vs. 7.97 mIU/L) and HOMA-IR (2.17 vs. 1.73) were elevated in the RPL group. No significant differences were observed in maternal age, height, weight, body mass index (BMI), education level, manual labor proportion, smoking proportion, drinking proportion, age at menarche or first pregnancy, FBG, or lipid parameters.

3.2 Correlation Between Serum 25(OH)D and FINS or HOMA-IR

Fig. 1 shows the correlations between 25(OH)D and FINS or HOMA-IR. In the RPL group, Spearman analysis revealed significant negative correlations between 25(OH)D and FINS (r = –0.407, p 0.001), and between 25(OH)D and HOMA-IR (r = –0.373, p 0.001). No significant correlations were found in the control group. Multiple linear regression confirmed these findings after adjusted for confounding factors including maternal age, BMI, reproductive history, lifestyle factors (such as smoking and alcohol use, manual labor), blood pressure, and lipid profiles (non-standardized coefficient \beta = –0.006, 95% CI = –0.008 to –0.005, p 0.001).

3.3 Association of 25(OH)D, FINS, and HOMA-IR With RPL Risk

After adjusting for potential confounding factors, multivariate logistic regression analysis (Table 2) demonstrated that higher serum 25(OH)D levels were significantly associated with a decreased risk of RPL [adjusted odds ratio (aOR): 0.98, 95% CI: 0.97–0.99, p 0.01]. In contrast, elevated FINS levels (aOR: 1.06, 95% CI: 1.03–1.09, p 0.01) and higher HOMA-IR (aOR: 1.27, 95% CI: 1.23–1.44, p 0.01) were significantly linked with an increased likelihood of RPL. When vitamin D status was analyzed as a categorical variable, women with VDD had a significantly higher risk of RPL compared to those with sufficient vitamin D levels.

To further investigate the potential dose-response relationship and assess the linearity of these associations, we performed RCS analyses within logistic regression models adjusted for the same confounders. The results showed significant overall associations for 25(OH)D (p = 0.0019), FINS (p = 0.00006), and HOMA-IR (p = 0.0002) with RPL. However, none of the nonlinear components were statistically significant (all p > 0.05), indicating that the relationships were approximately linear. The dose-response trends are visualized in Fig. 2.

3.4 Mediating Effect of FINS and HOMA-IR

Mediation analysis revealed that both FINS and HOMA-IR partially mediated the association between 25(OH)D and RPL (Table 3). The indirect effects were statistically significant: FINS (OR = 0.9962, 95% CI: 0.9929–0.9986) and HOMA-IR (OR = 0.9963, 95% CI: 0.9929–0.9986), explaining 10.9% and 10.7% of the total effect, respectively.

3.5 Subgroup Analysis Based on Vitamin D Status in the RPL Group

Patients in the RPL group were further stratified into VDD, insufficiency, and sufficiency subgroups. Compared to patients with sufficient vitamin D, those with deficiency had significantly elevated FINS and HOMA-IR levels (p 0.01). Although not statistically significant, the VDD subgroup showed lower E2 levels, CPR, and LBR compared to the other subgroups (Table 4).

4. Discussion

RPL is a common adverse outcome in pregnancy with a multifactorial etiology. Although the exact mechanisms remain unclear, increasing evidence suggests that VDD may play a crucial role in the onset and progression of RPL [22]. In this study, we observed lower serum 25(OH)D levels and a higher prevalence of VDD among women with RPL compared to control subjects. Concurrently, FINS levels and HOMA-IR were elevated in the RPL group and negatively correlated with serum 25(OH)D concentrations, suggesting a potential link between vitamin D status and glucose metabolism in the pathogenesis of RPL.

Previous studies have reported a high prevalence of VDD among women with RPL [23]. A meta-analysis demonstrated that VDD is associated with an increased risk of miscarriage, with women receiving vitamin D supplementation having significantly lower odds of miscarriage (OR: 1.94, 95% CI: 1.25–3.02) [8]. In our study, patients in the RPL group exhibited lower serum 25(OH)D levels compared to controls (31.16 nmol/L vs. 35.05 nmol/L), along with a higher prevalence of VDD (46.10% vs. 34.90%). These findings align with previous research and highlight the potential role of vitamin D in maintaining pregnancy health. Furthermore, multivariate logistic regression analysis revealed that increasing serum 25(OH)D levels were significantly associated with a decreased risk of RPL (aOR: 0.98, 95% CI: 0.97–0.99), suggesting that VDD may act as a risk factor for RPL.

However, the underlying mechanisms linking low vitamin D levels to increased miscarriage risk remain unclear. One widely accepted hypothesis is that vitamin D exerts immunomodulatory effects that are critical for successful pregnancy. Ota et al. [24] reported that 1,25(OH)₂D₃ significantly inhibits the cytotoxicity of natural killer (NK) cells, downregulates pro-inflammatory cytokines such as interferon-\gamma and tumour necrosis factor-\alpha, and upregulates anti-inflammatory and implantation-supportive cytokines, including interleukin (IL)-10, IL-1\beta, vascular endothelial growth factor, and granulocyte colony-stimulating factor. This promotes a shift from a type I to a type II immune response conducive to maintaining pregnancy [24]. Consistently, previous research has demonstrated a heightened type I immune response in women with RPL [22], implying that VDD may exacerbate NK cell activity and contribute to pregnancy failure. Although the immunomodulatory role of vitamin D is widely recognized and likely relevant to RPL, our study did not measure immune-related indicators, which represents a limitation. Future research, including immune biomarkers, is necessary to clarify the potential influence of vitamin D on immune regulation in RPL.

In addition to peripheral immune modulation, vitamin D is also thought to influence the maternal-fetal interface. A study has found significantly reduced levels of 25(OH)D, transforming growth factor-beta, and VDR in the decidual tissue of RPL patients, alongside elevated levels of IL-17 and IL-23 [25]. Given that 25(OH)D negatively regulates the IL-23/IL-17 axis, these findings suggest that insufficient vitamin D may impair local immune tolerance at the maternal-fetal interface, further increasing the risk of pregnancy loss [26].

Furthermore, glucose metabolism disturbances may represent another pathway linking VDD and RPL. Previous research has identified elevated insulin levels and HOMA-IR in women with RPL, both in natural conception and in those who used assisted reproduction [27, 28, 29]. In our cohort, RPL patients had higher FINS and HOMA-IR, and their increase was associated with greater RPL risk. Previous literature suggests that insulin resistance may impair endometrial vascularization, embryo development, and implantation, or lead to hypercoagulability and androgen excess, all of which may contribute to adverse pregnancy outcomes [30, 31, 32].

Importantly, our mediation analysis revealed that FINS and HOMA-IR partially mediated the association between 25(OH)D levels and RPL risk, accounting for 10.9% and 10.7% of the total effect, respectively. These findings suggest that glucose metabolism may serve as a mechanistic bridge between VDD and RPL. One potential mechanism is that VDD leads to reduced expression of insulin receptors on insulin-responsive cells, thereby decreasing insulin sensitivity and resulting in elevated circulating insulin levels [33]. Elevated insulin, in turn, may reduce the expression of glycoproteins at the implantation site and increase homocysteine levels, both of which could impair endometrial receptivity [34]. These alterations may disrupt endometrial blood flow and vascular integrity, ultimately compromising successful embryo implantation. Although subgroup analysis indicated that patients with VDD had higher FINS levels and HOMA-IR, and slightly worse reproductive outcomes, these trends did not reach statistical significance, likely due to limited sample size. Larger studies are needed to validate these associations.

Generally speaking, the results of this study emphasize the relationship between low levels of vitamin D and RPL and its potential regulation mechanism of glucose metabolism. However, it should be pointed out that this study has some limitations, including the relatively small sample size and the cross-sectional characteristics of the research design. Although this study is cross-sectional in design, all serum 25(OH)D measurements were conducted before pregnancy, and pregnancy outcomes were collected retrospectively or through follow-up. This temporal sequence helps mitigate, although not eliminate, the concern of reverse causality. Nevertheless, prospective studies are still warranted to clarify the causal direction of the association between vitamin D status and RPL. Therefore, further large-scale research and clinical trials are necessary to deeply understand the role of vitamin D in RPL and provide more powerful theoretical support for future intervention and treatment strategies. To sum up, vitamin D may be a potential therapeutic target, which is expected to play an essential role in preventing and managing RPL.

5. Conclusions

In summary, our study supports the hypothesis that VDD is associated with an increased risk of RPL and may act, at least in part, through dysregulation of glucose metabolism. FINS and HOMA-IR partially mediated the association between 25(OH)D and RPL, suggesting a novel mechanistic link. While these findings are promising, they are observational and cross-sectional. Future large-scale, prospective, and mechanistic studies are needed to confirm causality and elucidate the precise role of vitamin D and insulin pathways in the pathogenesis of RPL. Vitamin D may serve as a potential target for preventive or therapeutic interventions aimed at improving reproductive outcomes in women at risk of RPL.

References

[1] Lopez AG, Kerlan V, Desailloud R. Non-classical effects of vitamin D: Non-bone effects of vitamin D. Annales D’endocrinologie. 2021; 82: 43–51. https://doi.org/10.1016/j.ando.2020.12.002.

[2] Bartley J. Vitamin D: emerging roles in infection and immunity. Expert Review of Anti-infective Therapy. 2010; 8: 1359–1369. https://doi.org/10.1586/eri.10.102.

[3] Bhan I. Vitamin d binding protein and bone health. International Journal of Endocrinology. 2014; 2014: 561214. https://doi.org/10.1155/2014/561214.

[4] Potdar N, Iyasere C. Early pregnancy complications including recurrent pregnancy loss and obesity. Best Practice & Research. Clinical Obstetrics & Gynaecology. 2023; 90: 102372. https://doi.org/10.1016/j.bpobgyn.2023.102372.

[5] Mousavi Salehi A, Ghafourian M, Amari A, Zargar M. Evaluation of CD3+ T Cell Percentage, Function and its Relationship with Serum Vitamin D Levels in Women with Recurrent Spontaneous Abortion and Recurrent Implantation Failure. Iranian Journal of Immunology. 2022; 19: 369–377. https://doi.org/10.22034/IJI.2022.91464.2083.

[6] Zhao H, Wei X, Yang X. A novel update on vitamin D in recurrent pregnancy loss (Review). Molecular Medicine Reports. 2021; 23: 382. https://doi.org/10.3892/mmr.2021.12021.

[7] Sharif K, Sharif Y, Watad A, Yavne Y, Lichtbroun B, Bragazzi NL, et al. Vitamin D, autoimmunity and recurrent pregnancy loss: More than an association. American Journal of Reproductive Immunology (New York, N.Y.: 1989). 2018; 80: e12991. https://doi.org/10.1111/aji.12991.

[8] Tamblyn JA, Pilarski NSP, Markland AD, Marson EJ, Devall A, Hewison M, et al. Vitamin D and miscarriage: a systematic review and meta-analysis. Fertility and Sterility. 2022; 118: 111–122. https://doi.org/10.1016/j.fertnstert.2022.04.017.

[9] Ornoy A, Becker M, Weinstein-Fudim L, Ergaz Z. Diabetes during Pregnancy: A Maternal Disease Complicating the Course of Pregnancy with Long-Term Deleterious Effects on the Offspring. A Clinical Review. International Journal of Molecular Sciences. 2021; 22: 2965. https://doi.org/10.3390/ijms22062965.

[10] Yessoufou A, Moutairou K. Maternal diabetes in pregnancy: early and long-term outcomes on the offspring and the concept of “metabolic memory”. Experimental Diabetes Research. 2011; 2011: 218598. https://doi.org/10.1155/2011/218598.

[11] Sacerdote A, Dave P, Lokshin V, Bahtiyar G. Type 2 Diabetes Mellitus, Insulin Resistance, and Vitamin D. Current Diabetes Reports. 2019; 19: 101. https://doi.org/10.1007/s11892-019-1201-y.

[12] Niroomand M, Fotouhi A, Irannejad N, Hosseinpanah F. Does high-dose vitamin D supplementation impact insulin resistance and risk of development of diabetes in patients with pre-diabetes? A double-blind randomized clinical trial. Diabetes Research and Clinical Practice. 2019; 148: 1–9. https://doi.org/10.1016/j.diabres.2018.12.008.

[13] Berridge MJ. Vitamin D deficiency and diabetes. The Biochemical Journal. 2017; 474: 1321–1332. https://doi.org/10.1042/BCJ20170042.

[14] Contreras-Bolívar V, García-Fontana B, García-Fontana C, Muñoz-Torres M. Mechanisms Involved in the Relationship between Vitamin D and Insulin Resistance: Impact on Clinical Practice. Nutrients. 2021; 13: 3491. https://doi.org/10.3390/nu13103491.

[15] Morró M, Vilà L, Franckhauser S, Mallol C, Elias G, Ferré T, et al. Vitamin D Receptor Overexpression in β-Cells Ameliorates Diabetes in Mice. Diabetes. 2020; 69: 927–939. https://doi.org/10.2337/db19-0757.

[16] Neelankal John A, Jiang FX. An overview of type 2 diabetes and importance of vitamin D3-vitamin D receptor interaction in pancreatic β-cells. Journal of Diabetes and its Complications. 2018; 32: 429–443. https://doi.org/10.1016/j.jdiacomp.2017.12.002.

[17] Ross AC, Manson JE, Abrams SA, Aloia JF, Brannon PM, Clinton SK, et al. The 2011 report on dietary reference intakes for calcium and vitamin D from the Institute of Medicine: what clinicians need to know. The Journal of Clinical Endocrinology and Metabolism. 2011; 96: 53–58. https://doi.org/10.1210/jc.2010-2704.

[18] Xu J, Yin MN, Chen ZH, Yang L, Ye DS, Sun L. Embryo retention significantly decreases clinical pregnancy rate and live birth rate: a matched retrospective cohort study. Fertility and Sterility. 2020; 114: 787–791. https://doi.org/10.1016/j.fertnstert.2020.04.043.

[19] Yang X, Bu Z, Hu L. Live Birth Rate of Frozen-Thawed Single Blastocyst Transfer After 6 or 7 Days of Progesterone Administration in Hormone Replacement Therapy Cycles: A Propensity Score-Matched Cohort Study. Frontiers in Endocrinology. 2021; 12: 706427. https://doi.org/10.3389/fendo.2021.706427.

[20] Glujovsky D, Pesce R, Sueldo C, Quinteiro Retamar AM, Hart RJ, Ciapponi A. Endometrial preparation for women undergoing embryo transfer with frozen embryos or embryos derived from donor oocytes. The Cochrane Database of Systematic Reviews. 2020; 10: CD006359. https://doi.org/10.1002/14651858.CD006359.pub3.

[21] Ma C, Cheng B, Zhou L, Cai S, Qin B, Sun J, et al. Association between insulin resistance and vascular damage in an adult population in China: a cross-sectional study. Scientific Reports. 2024; 14: 18472. https://doi.org/10.1038/s41598-024-69338-y.

[22] Gonçalves DR, Braga A, Braga J, Marinho A. Recurrent pregnancy loss and vitamin D: A review of the literature. American Journal of Reproductive Immunology (New York, N.Y.: 1989). 2018; 80: e13022. https://doi.org/10.1111/aji.13022.

[23] Chen X, Yin B, Lian RC, Zhang T, Zhang HZ, Diao LH, et al. Modulatory effects of vitamin D on peripheral cellular immunity in patients with recurrent miscarriage. American Journal of Reproductive Immunology (New York, N.Y.: 1989). 2016; 76: 432–438. https://doi.org/10.1111/aji.12585.

[24] Ota K, Dambaeva S, Han AR, Beaman K, Gilman-Sachs A, Kwak-Kim J. Vitamin D deficiency may be a risk factor for recurrent pregnancy losses by increasing cellular immunity and autoimmunity. Human Reproduction (Oxford, England). 2014; 29: 208–219. https://doi.org/10.1093/humrep/det424.

[25] Li N, Wu HM, Hang F, Zhang YS, Li MJ. Women with recurrent spontaneous abortion have decreased 25(OH) vitamin D and VDR at the fetal-maternal interface. Brazilian Journal of Medical and Biological Research = Revista Brasileira De Pesquisas Medicas E Biologica. 2017; 50: e6527. https://doi.org/10.1590/1414-431X20176527.

[26] Wu Z, Ma B, Xiao M, Ren Q, Shen Y, Zhou Z. Vitamin D Modified DSS-Induced Colitis in Mice via STING Signaling Pathway. Biology. 2025; 14: 715. https://doi.org/10.3390/biology14060715.

[27] Tian L, Shen H, Lu Q, Norman RJ, Wang J. Insulin resistance increases the risk of spontaneous abortion after assisted reproduction technology treatment. The Journal of Clinical Endocrinology and Metabolism. 2007; 92: 1430–1433. https://doi.org/10.1210/jc.2006-1123.

[28] Baban RS, Ali NM, Al-Moayed HA. Serum leptin and insulin hormone level in recurrent pregnancy loss. Oman Medical Journal. 2010; 25: 203–207. https://doi.org/10.5001/omj.2010.57.

[29] Suzuki K, Sata F, Yamada H, Saijo Y, Tsuruga N, Minakami H, et al. Pregnancy-associated plasma protein-A polymorphism and the risk of recurrent pregnancy loss. Journal of Reproductive Immunology. 2006; 70: 99–108. https://doi.org/10.1016/j.jri.2005.11.004.

[30] Cai WY, Luo X, Lv HY, Fu KY, Xu J. Insulin resistance in women with recurrent miscarriage: a systematic review and meta-analysis. BMC Pregnancy and Childbirth. 2022; 22: 916. https://doi.org/10.1186/s12884-022-05256-z.

[31] Glueck CJ, Wang P, Fontaine RN, Sieve-Smith L, Tracy T, Moore SK. Plasminogen activator inhibitor activity: an independent risk factor for the high miscarriage rate during pregnancy in women with polycystic ovary syndrome. Metabolism: Clinical and Experimental. 1999; 48: 1589–1595. https://doi.org/10.1016/s0026-0495(99)90250-0.

[32] Ispasoiu CA, Chicea R, Stamatian FV, Ispasoiu F. High fasting insulin levels and insulin resistance may be linked to idiopathic recurrent pregnancy loss: a case-control study. International Journal of Endocrinology. 2013; 2013: 576926. https://doi.org/10.1155/2013/576926.

[33] Park CY, Shin S, Han SN. Multifaceted Roles of Vitamin D for Diabetes: From Immunomodulatory Functions to Metabolic Regulations. Nutrients. 2024; 16: 3185. https://doi.org/10.3390/nu16183185.

[34] Alghamdi AA, Alotaibi AS. High Insulin Resistance in Saudi Women with Unexplained Recurrent Pregnancy Loss: A Case-control Study. Saudi Journal of Medicine & Medical Sciences. 2023; 11: 314–318. https://doi.org/10.4103/sjmms.sjmms_82_23.

Funding

Medical Innovation and Development Project of Lanzhou University(lzuyxcx-2022-137)

The Science Foundation of Lanzhou University(054000229)

A Real-World Study on RPL in China(YJS-BD-19)