Objective To investigate the correlation of serum levels of Spexin,FoxO1 and insulin resistance(IR)with prognosis in patients with gestational diabetes mellitus(GDM).Methods A total of 198 patients with GDM were prospectively selected as the GDM group,and 195 healthy pregnant women who underwent physical examinations during the same period were selected as the control group.The age at enrollment,gestational age,triglyceride(TG),pre-pregnancy body weight,fasting insulin(FINS),parity,fasting blood glucose(FBG),total cholesterol(TC)and homeostasis model assessment of insulin resistance index(HOMA-IR)of the subjects were collected.The serum levels of Spexin and FoxO1 in subjects were detected by enzyme-linked immunosorbent assay(ELISA).According to the pregnancy outcome,the GDM patients were divided into the adverse pregnancy outcome group(96 cases)and the good pregnancy outcome group(102 cases).The correlations between serum Spexin,FoxO1,and HOMA-IR in GDM patients were analyzed by Pearson's method.Logistic regression was used to analyze risk factors affecting the pregnancy outcome of GDM patients.The predictive value of serum Spexin and FoxO1 levels for the pregnancy outcome of GDM patients was analyzed by receiver operating characteristic(ROC)curve.Results The serum levels of FINS,TG,FBG,Spexin,FoxO1 and HOMA-IR were higher in the GDM group than those in the control group(P＜0.01).Pearson correlation analysis showed that serum levels of Spexin and FoxO1 in GDM patients were positively correlated with HOMA-IR,and serum level of Spexin was positively correlated with FoxO1(P＜0.05).The incidences of gestational hypertension,cesarean section,macrosomia,neonatal malformations,low birth weight infants and neonatal asphyxia were higher in the GDM group than those in the control group(P＜0.05).Serum levels of Spexin and FoxO1 and FINS,FBG and HOMA-IR were significantly higher in the adverse pregnancy group than those in the good pregnancy group(P＜0.01).Multivariate Logistic regression analysis showed that the increased Spexin and FoxO1 were the risk factor for adverse pregnancy outcomes in GDM patients(P＜0.01).The ROC curve analysis showed that the area under the curve(AUC)of serum Spexin and FoxO1 levels for predicting pregnancy outcomes in GDM patients was 0.887 and 0.883,respectively,and the AUC of combined prediction was 0.942.Conclusion Serum levels of Spexin and FoxO1 are elevated in GDM patients,and both are related to IR.The combination of the two can assist in judging the pregnancy outcome of GDM patients.

Intervertebral disc degeneration (IDD) is largely attributed to impaired endogenous repair. Nucleus pulposus-derived stem cells (NPSCs) senescence leads to endogenous repair failure. Small extracellular vesicles/exosomes derived from mesenchymal stem cells (mExo) have shown great therapeutic potential in IDD, while whether mExo could alleviate NPSCs senescence and its mechanisms remained unknown. We established a compression-induced NPSCs senescence model and rat IDD models to evaluate the therapeutic efficiency of mExo and investigate the mechanisms. We found that mExo significantly alleviated NPSCs senescence and promoted disc regeneration while knocking down thioredoxin (TXN) impaired the protective effects of mExo. TXN was bound to various endosomal sorting complex required for transport (ESCRT) proteins. Autocrine motility factor receptor (AMFR) mediated TXN K63 ubiquitination to promote the binding of TXN on ESCRT proteins and sorting of TXN into mExo. Knocking down exosomal TXN inhibited the transcriptional activity of nuclear factor erythroid 2-related factor 2 (NRF2) and activator protein 1 (AP-1). NRF2 and AP-1 inhibition reduced endogenous TXN production that was promoted by exosomal TXN. Inhibition of NRF2 in vivo diminished the anti-senescence and regenerative effects of mExo. Conclusively, AMFR-mediated TXN ubiquitination promoted the sorting of TXN into mExo, allowing exosomal TXN to promote endogenous TXN production in NPSCs via TXN/NRF2/AP-1 feed-forward circuit to alleviate NPSCs senescence and disc degeneration.

Objective:To investigate the expression and methylation status of the SALL4 gene in placental tissues of fetal growth restriction (FGR) and its effects on trophoblast cell proliferation, migration, and invasion. Methods:Placental tissues were collected from 20 full-term FGR patients and 20 healthy term controls who underwent regular prenatal examination and cesarean section at the Third Affiliated Hospital, Zhengzhou University between July 2023 and February 2024. SALL4 mRNA and protein expression were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot. Methylation specific polymerase china reaction (MSP) assessed promoter methylation levels. HTR8/SVneo cells were transfected with SALL4-targeting small interfering RNA (si-SALL4) or negative control small interfering RNA (si-NC). HTR8/SVneo cells were treated with the demethylating agent 5-aza-2′-deoxycytidine (5-Aza-dC) to inhibit gene methylation (5-Aza-dC group) or with 10% RPMI-1640 medium as a vehicle control. Transfection efficiency (for siRNA) and the efficacy of 5-Aza-dC-induced demethylation were assessed by qRT-PCR and Western blot. The functional effects of SALL4 knockdown and methylation inhibition on trophoblast cells were evaluated using proliferation assays, scratch wound healing assays, and Transwell invasion assays. Statistical analyses included independent t-tests and Chi-square test. Results:(1) Human tissues: FGR placentas showed lower SALL4 mRNA (0.802±0.194 vs. 1.015±0.186, t=3.55) and protein expression (0.445±0.114 vs. 0.701±0.113, t=3.19), alongside higher methylation rates of SALL4 [80% (16/20) vs. 15% (3/20), χ2=14.44] compared to controls (all P<0.05). (2) In vitro: si-SALL4 transfection reduced HTR8/SVneo proliferation (OD450 at 48 h: 0.653±0.021 vs. 0.827±0.040, t=6.60), migration [healing rate at 48 h: (24.317±2.637)% vs. (49.327±1.961)%, t=13.18], and invasion [counted invaded cells: (133.000±6.557) vs. (272.667±18.009) cells, t=12.62] versus si-NC (all P<0.05). Conversely, 5-Aza-dC treatment increased HTR8/SVneo proliferation (0.917±0.042 vs. 0.783±0.031, t=－4.47), migration [(71.097±3.354)% vs. (51.632±2.877)%, t=－7.63], and invasion [(384.000±12.166) vs. (202.833±7.095) cells, t=－13.69] versus vehicle control (all P<0.05). Conclusions:Hypermethylation of the SALL4 promoter in FGR placentas suppresses its expression, impairing trophoblast cell function. Demethylation restores SALL4 expression and enhances cellular proliferation, migration, and invasion, involving in the occurrence and development of FGR disease.

Objective To investigate the correlation of serum levels of Spexin,FoxO1 and insulin resistance(IR)with prognosis in patients with gestational diabetes mellitus(GDM).Methods A total of 198 patients with GDM were prospectively selected as the GDM group,and 195 healthy pregnant women who underwent physical examinations during the same period were selected as the control group.The age at enrollment,gestational age,triglyceride(TG),pre-pregnancy body weight,fasting insulin(FINS),parity,fasting blood glucose(FBG),total cholesterol(TC)and homeostasis model assessment of insulin resistance index(HOMA-IR)of the subjects were collected.The serum levels of Spexin and FoxO1 in subjects were detected by enzyme-linked immunosorbent assay(ELISA).According to the pregnancy outcome,the GDM patients were divided into the adverse pregnancy outcome group(96 cases)and the good pregnancy outcome group(102 cases).The correlations between serum Spexin,FoxO1,and HOMA-IR in GDM patients were analyzed by Pearson's method.Logistic regression was used to analyze risk factors affecting the pregnancy outcome of GDM patients.The predictive value of serum Spexin and FoxO1 levels for the pregnancy outcome of GDM patients was analyzed by receiver operating characteristic(ROC)curve.Results The serum levels of FINS,TG,FBG,Spexin,FoxO1 and HOMA-IR were higher in the GDM group than those in the control group(P＜0.01).Pearson correlation analysis showed that serum levels of Spexin and FoxO1 in GDM patients were positively correlated with HOMA-IR,and serum level of Spexin was positively correlated with FoxO1(P＜0.05).The incidences of gestational hypertension,cesarean section,macrosomia,neonatal malformations,low birth weight infants and neonatal asphyxia were higher in the GDM group than those in the control group(P＜0.05).Serum levels of Spexin and FoxO1 and FINS,FBG and HOMA-IR were significantly higher in the adverse pregnancy group than those in the good pregnancy group(P＜0.01).Multivariate Logistic regression analysis showed that the increased Spexin and FoxO1 were the risk factor for adverse pregnancy outcomes in GDM patients(P＜0.01).The ROC curve analysis showed that the area under the curve(AUC)of serum Spexin and FoxO1 levels for predicting pregnancy outcomes in GDM patients was 0.887 and 0.883,respectively,and the AUC of combined prediction was 0.942.Conclusion Serum levels of Spexin and FoxO1 are elevated in GDM patients,and both are related to IR.The combination of the two can assist in judging the pregnancy outcome of GDM patients.

Objective:To investigate the expression and methylation status of the SALL4 gene in placental tissues of fetal growth restriction (FGR) and its effects on trophoblast cell proliferation, migration, and invasion. Methods:Placental tissues were collected from 20 full-term FGR patients and 20 healthy term controls who underwent regular prenatal examination and cesarean section at the Third Affiliated Hospital, Zhengzhou University between July 2023 and February 2024. SALL4 mRNA and protein expression were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot. Methylation specific polymerase china reaction (MSP) assessed promoter methylation levels. HTR8/SVneo cells were transfected with SALL4-targeting small interfering RNA (si-SALL4) or negative control small interfering RNA (si-NC). HTR8/SVneo cells were treated with the demethylating agent 5-aza-2′-deoxycytidine (5-Aza-dC) to inhibit gene methylation (5-Aza-dC group) or with 10% RPMI-1640 medium as a vehicle control. Transfection efficiency (for siRNA) and the efficacy of 5-Aza-dC-induced demethylation were assessed by qRT-PCR and Western blot. The functional effects of SALL4 knockdown and methylation inhibition on trophoblast cells were evaluated using proliferation assays, scratch wound healing assays, and Transwell invasion assays. Statistical analyses included independent t-tests and Chi-square test. Results:(1) Human tissues: FGR placentas showed lower SALL4 mRNA (0.802±0.194 vs. 1.015±0.186, t=3.55) and protein expression (0.445±0.114 vs. 0.701±0.113, t=3.19), alongside higher methylation rates of SALL4 [80% (16/20) vs. 15% (3/20), χ2=14.44] compared to controls (all P<0.05). (2) In vitro: si-SALL4 transfection reduced HTR8/SVneo proliferation (OD450 at 48 h: 0.653±0.021 vs. 0.827±0.040, t=6.60), migration [healing rate at 48 h: (24.317±2.637)% vs. (49.327±1.961)%, t=13.18], and invasion [counted invaded cells: (133.000±6.557) vs. (272.667±18.009) cells, t=12.62] versus si-NC (all P<0.05). Conversely, 5-Aza-dC treatment increased HTR8/SVneo proliferation (0.917±0.042 vs. 0.783±0.031, t=－4.47), migration [(71.097±3.354)% vs. (51.632±2.877)%, t=－7.63], and invasion [(384.000±12.166) vs. (202.833±7.095) cells, t=－13.69] versus vehicle control (all P<0.05). Conclusions:Hypermethylation of the SALL4 promoter in FGR placentas suppresses its expression, impairing trophoblast cell function. Demethylation restores SALL4 expression and enhances cellular proliferation, migration, and invasion, involving in the occurrence and development of FGR disease.

Objective To investigate the expression levels of secreted frizzled-related protein 5 (sFRP5), heat shock protein 60 (HSP60) and solute carrier family 16 member 11 (SLC16A11) in serum of patients with gestational diabetes mellitus (GDM) and their relationships with insulin resistance. Methods A total of 120 GDM patients in the hospital from January 2022 to December 2023 were selected as study group, and another 120 healthy pregnant women in the same period were selected as control group. The expression levels of serum sFRP5, HSP60 and SLC16A11 were detected; the levels of insulin resistance-related indicators[fasting blood glucose (FBG), fasting insulin (FINS), and homeostasis model assessment of insulin resistance (HOMA-IR)]were measured and calculated; the Pearson's correlation method was used to analyze the correlations of serum sFRP5, HSP60 and SLC16A11 with insulin resistance; the Logistic regression andysis and receiver operating characteristic (ROC) curve analysis were used to assess the relationships of serum sFRP5, HSP60 and SLC16A11 with GDM. Results The serum sFRP5 expression in the study group was significantly lower than that in the control group, while the serum HSP60, SLC16A11, FBG, FINS and HOMA-IR levels were significantly higher than those in the control group (P < 0.05). Pearson's correlation analysis showed that serum sFRP5 was negatively correlated with HSP60, SLC16A11, FBG, FINS and HOMA-IR (P < 0.05); serum HSP60 was positively correlated with SLC16A11, FBG, FINS and HOMA-IR (P < 0.05); serum SLC16A11 was positively correlated with FBG, FINS and HOMA-IR (P < 0.05). Multivariate Logistic regression analysis revealed that HSP60, SLC16A11, FBG, FINS and HOMA-IR were risk factors for GDM in pregnant patients, while sFRP5 was a protective factor (P < 0.05). The area under the curve (AUC) for the combined diagnosis of GDM in pregnant patients by serum sFRP5, HSP60 and SLC16A11 was 0.924, indicating that the combined diagnostic value of three indicators was superior to that of each individual indicator (all P < 0.05). Conclusion GDM patients exhibit decreased serum sFRP5 level and increased HSP60 and SLC16A11 levels; three indicators are factors influencing the occurrence of GDM in pregnant patients, and are associated with insulin resistance; the combination of three indicators exhibits high diagnostic efficacy for GDM.