Growth and differentiation factor-11 is developmentally regulated in skeletal muscle and inhibits myoblast differentiation

Growth and differentiation factor-11 (GDF-11) is a secreted protein that is closely related to myostatin, a known inhibitor of skeletal muscle development. The role of GDF-11 in regulating skeletal muscle growth remains unclear and the pattern of expression during post-natal growth has not been reported. Therefore, we sought to determine the expression of GDF-11 during post-natal growth and its effect on myoblast proliferation and differentiation. We collected gastrocnemius muscles from male and female mice at 2, 3, 4, 6, 12, 20 and 32 weeks of age (n = 6 per sex and age). In addition, gastrocnemius muscles were col- lected from male wild-type and myostatin knockout mice at 4, 6, 12 and 20 weeks of age (n = 6 per age and genotype). RNA was extracted and reverse tran- scribed. Northern analysis identified an expected 4.4 kb mRNA transcript for GDF-11 in gastrocnemius muscles of mice. The concentration of GDF-11 mRNA, as determined by quantitative PCR, was increased in gastrocnemius muscles from 2 to 6 weeks—a period of rapid postnatal muscle growth—and remained higher in male than female mice from 4 to 20 weeks of age (P gastrocnemius muscles of myostatin knockout compared with wild-type mice (P < 0.05), which may suggest a compensatory mecha- nism for the lack of myostatin. In support, recombi- nant GDF-11 inhibited differentiation of C2C12 mur- ine myoblasts and those isolated from myostatin knockout and wild-type mice (P < 0.05). Inhibited dif-ferentiation of C2C12 myoblasts was associated with decreased mRNA expression of early and late mo- lecular markers of differentiation (MyoD, myogenin, IGF-II, desmin and MyHC, P < 0.05). Our results suggest that GDF-11 regulates growth of skeletal muscles by inhibiting myoblast differentiation in an autocrine/paracrine manner and, perhaps, also plays a role in regulating sexually dimorphic growth.

Correlation between growth differentiation factor-15 and collagen metabolism indicators in patients with myocardial infarction and heart failure

BackgroundGrowth differentiation factor （GDF）-15, a divergent member of the transforming growth factor beta super-family does appear to be up-regulated in response to experimental pressure overload and progression of heart failure （HF）. HF frequently develops after myocardial infarction （MI）, contributing to worse outcome. The aim of this study is to assess the correlation between GDF-15 levels and markers related to collagen turnover in different stages of HF.MethodsThe study consists of a cohort of 179 patients, including stable angina pectoris patients （AP group,n= 50）, old MI patients without HF （OMI group,n = 56）, old MI patients with HF （OMI-HF group,n= 38） and normal Control group （n = 35）. Both indicators reflecting the synthesis and degradation rates of collagen including precollagen I N-terminal peptide （PINP）, type I collagen carboxy-terminal peptide （ICTP）, precollagen III N-terminal peptide （PIIINP） and GDF-15 were measured using an enzyme-linked inmunosorbent assay.ResultsThe plasma GDF-15 level was higher in OMI-HF group （1373.4 ± 275.4 ng/L） than OMI group （1036.1 ± 248.6 ng/L）, AP group （784.6 ± 222.4 ng/L） and Control group （483.8 ± 186.4 ng/L） （P〈 0.001）. The indi-cators of collagen turnover （ICTP, PINP, PIIINP） all increased in the OMI-HF group compared with Control group （3.03 ± 1.02μg/Lvs. 2.08 ± 0.95μg/L, 22.2 ± 6.6μg/Lvs. 16.7 ± 5.1μg/L and 13.2 ± 7.9μg/Lvs. 6.4 ± 2.1μg/L, respectively;P〈 0.01）. GDF-15 positively cor-related with ICTP and PIIINP （r = 0.302,P〈 0.001 andr= 0.206,P= 0.006, respectively）. GDF-15 positively correlated to the echocardio-graphic diastolic indicators E/Em and left atrial pressure （r= 0.349 and r= 0.358, respectively;P〈 0.01）, and inversely correlated to the systolic indicators left ventricular ejection fraction and the average of peak systolic myocardial velocities （Sm） （r=-0.623 and r=-0.365, respectively;P〈 0.01）.ConclusionPlasma GDF-15 is associated with the indicators of type I and III collagen turnover.

Growth differentiation factor-15 is a prognostic marker in patients with intermediate coronary artery disease

Background Growth differentiation factor-15(GDF-15)is involved in multiple processes that are associated with coronary artery disease(CAD).However,little is known about the association between GDF-15 and the future ischemic events in patients with intermediate CAD.This study was conducted to investigate whether plasma GDF-15 constituted risk biomarkers for future cardiovascular events in patients with intermediate CAD.Methods A prospective study was performed based on 541 patients with intermediate CAD(20%–70%).GDF-15 of each patient was determined in a blinded manner.The primary endpoint was major adverse cardiac event(MACE),which was defined as a composite of all-cause death,nonfatal myocardial infarction,revascularization and readmission due to angina pectoris.Results After a median follow-up of 64 months,504 patients(93.2%)completed the follow-up.Overall,the combined endpoint of MACE appeared in 134 patients(26.6%)in the overall population:26 patients died,11 patients suffered a nonfatal myocardial infarction,51 patients underwent revascularization,and 46 patients were readmitted for angina pectoris.The plasma levels of GDF-15(median:1172.02 vs.965.25 pg/m L,P=0.014)were higher in patients with ischemic events than those without events.After adjusting for traditional risk factors,higher GDF-15 levels were significantly associated with higher incidence of the composite endpoint of MACE(HR=1.244,95%CI:1.048–1.478,Quartile 4 vs.Quartile 1,P=0.013).Conclusions The higher level of GDF-15 was an independent predictor of long-term adverse cardiovascular events in patients with intermediate CAD.

GROWTH DIFFERENTIATION FACTOR-5 STIMULATES THE GROWTH AND ANABOLIC METABOLISM OF ARTICULAR CHONDROCYTES

Objective To observe the effect of growth differentiation factor-5 (GDF-5) on the growth and anabolic metabolism of articular chondrocytes. Methods The articular chondrocytes isolated from rats were treated with various concentrations of rmGDF-5, and the growth of chondrocytes measured by MTT assay, the cellular cartilage matrices formation detected sulfated glycosaminoglycan by Alcian blue staining and type Ⅱcollagen by RT-PCR. Results After 7 days culture, MTT assay showed that GDF-5 enhanced the growth of chondrocytes in a dose-dependent manner, RT-PCR showed that GDF-5 clearly induced the synthesis of type Ⅱ collagen because of the col2a1 mRNA band more and more strong in a dose-dependent. Chondrocytes were cultured with GDF-5 for 14 days, the intensity of Alcian blue staining was greatly enhanced, especially, at a high concentration of 1000ng/mL, and GDF-5 enhanced the accumulation of the Alcian blue-stainable material in a concentration-dependent manner and in a does-dependent manner. Conclusion GDF-5 enhanced the growth of mature articular chondrocytes, and stimulated the cellular cartilage matrices formation in mono-layer culture.

Growth differentiation factor 11 promotes macrophage polarization towards M2 to attenuate myocardial infarction via inhibiting Notch1 signaling pathway

Background:Myocardial infarctions(MI)is a major threat to human health especially in people exposed to cold environment.The polarization of macrophages towards different functional phenotypes(M1 macrophages and M2 macrophages)is closely related to MI repairment.The growth differentiation factor 11(GDF11)has been reported to play a momentous role in inflammatory associated diseases.In this study,we examined the regulatory role of GDF11 in macrophage polarization and elucidated the underlying mechanisms in MI.Methods:In vivo,the mice model of MI was induced by permanent ligation of the left anterior descending coronary artery(LAD),and mice were randomly divided into the sham group,MI group,and MI+GDF11 group.The protective effect of GDF11 on myocardial infarction and its effect on macrophage polarization were verified by echocardiography,triphenyl tetrazolium chloride staining and immunofluorescence staining of heart tissue.In vitro,based on the RAW264.7 cell line,the effect of GDF11 in promoting macrophage polarization toward the M2 type by inhibiting the Notch1 Signaling pathway was validated by qRT-PCR,Western blot,and flow cytometry.Results:We found that GDF11 was significantly downregulated in the cardiac tissue of MI mice.And GDF11 supplementation can improve the cardiac function.Moreover,GDF11 could reduce the proportion of M1 macrophages and increase the accumulation of M2 macrophages in the heart tissue of MI mice.Furthermore,the cardioprotective effect of GDF11 on MI mice was weakened after macrophage clearance.At the cellular level,application of GDF11 could inhibit the expression of M1 macrophage(classically activated macrophage)markers iNOS,interleukin(IL)-1β,and IL-6 in a dose-dependent manner.In contrast,GDF11 significantly increased the level of M2 macrophage markers including IL-10,CD206,arginase 1(Arg1),and vascular endothelial growth factor(VEGF).Interestingly,GDF11 could promote M1 macrophages polarizing to M2 macrophages.At the molecular level,GDF11 significantly down-regulated the Notch1 signaling pathway,the activation of which has been demonstrated to promote M1 polarization in macrophages.Conclusions:GDF11 promoted macrophage polarization towards M2 to attenuate myocardial infarction via inhibiting Notch1 signaling pathway.

Significance of 125I radioactive seed implantation on growth differentiation factor and programmed death receptor-1 during treatment of oral cancer

BACKGROUND Oral cancer(OC)is the most common malignant tumor in the oral cavity,and is mainly seen in middle-aged and elderly men.At present,OC is mainly treated clinically by surgery or combined with radiotherapy and chemotherapy;but recently,more and more studies have shown that the stress trauma caused by surgery and the side effects of radiotherapy and chemotherapy seriously affect the prognosis of patients.AIM To determine the significance of 125I radioactive seed implantation on growth differentiation factor 11(GDF11)and programmed death receptor-1(PD-1)during treatment of OC.METHODS A total of 184 OC patients admitted to The Second Affiliated Hospital of Jiamusi University from May 2015 to May 2017 were selected as the research subjects for prospective analysis.Of these patients,89 who received 125I radioactive seed implantation therapy were regarded as the research group(RG)and 95 patients who received surgical treatment were regarded as the control group(CG).The clinical efficacy,incidence of adverse reactions and changes in GDF11 and PD-1 before treatment(T0),2 wk after treatment(T1),4 wk after treatment(T2)and 6 wk after treatment(T3)were compared between the two groups.RESULTS The efficacy and recurrence rate in the RG were better than those in the CG(P<0.05),while the incidence of adverse reactions and survival rate were not different.There was no difference in GDF11 and PD-1 between the two groups at T0 and T1,but these factors were lower in the RG than in the CG at T2 and T3(P<0.05).Using receiver operating characteristic(ROC)curve analysis,GDF11 and PD-1 had good predictive value for efficacy and recurrence(P<0.001).CONCLUSION 125I radioactive seed implantation has clinical efficacy and can reduce the recurrence rate in patients with OC.This therapy has marked potential in clinical application.The detection of GDF11 and PD-1 in patients during treatment showed good predictive value for treatment efficacy and recurrence in OC patients,and may be potential targets for future OC treatment.

Glycosylation-independent binding to extracellular domains 11-13 of mannose-6-phosphate/insulin-like growth factor-2 receptor mediates the effects of soluble CREG on the phenotypic proliferation of vascular smooth muscle cells

Background The present study aimed to investigate the detailed mode and specific sites for their binding as well as the functional relevance of this binding in the phenotypic proliferation of vascular smooth muscle cells(SMCs). Methods CREG knocked-down SMCs were employed to evaluate the biological activity of wtCREG and mCREG.Expressions of SMC differentiation markers SM myosin heavy chain(SM-MHC),SM-actin,heavy caldesmon and myocardin were determined by Western blotting using specific antibodies. Cellular growth of SMCs was assessed by bromide dewuridine (BrdU) incorporation and cell cycle analysis on fluorescence-activated cell sorting(FACS).A solid-phase binding assay was used to study the binding of CREG to extracellular domains of M6P/IGF2R.The cellular co-localization of the two recombinant CREGs with M6P/IGF2R was detected on SMC surface by immunoprecipitation and immunofluorescence analysis.Results The molecular weight of wtCREG was around 30 kD while that of the mCREG was～25 kD.Treatment of wtCREG with PNGase F reduced its molecular weight from～30 kD to～25 kD,whereas PNGase F treatment had no effect on the molecular weight of mCREG.Both wtCREG and mCREG proteins enhanced SMC differentiation,inhibited BrdU incorporation,and arrested cell cycle progression when added to the culture medium.In CREG knocked-down SMCs,the amount of CREG detected by immunoblotting in M6P/IGF2R immunoprecipitates was significantly reduced when compared to normal cells.Both recombinant CREGs co-immunoprecipitated with M6P/IGF2R, although slightly reduced amount of the mutant CREG was detected in M6P/IGF2R immunoprecipitates.Immunostaining revealed that His-tagged CREGs co-localized with IGF2R on the cell surface in a glycosylation-independent manner.In vitro binding assay showed that CREGs bound to M6P/ IGF2R extracellular domains 7-10 and 11-13 in a glycosylation -dependent and -independent manner,respectively.Further blocking experiments using soluble M6P/IGF2R fragments and M6P/IGF2R neutralizing antibody indicated that the biological activities of recombinant CREGs in SMC growth and the up-regulation of SMC differentiation markers were all abolished by treatment with the M6P/IGF2R neutralizing antibody. However,although the growth inhibitory effect of wtCREG was nearly abolished by D7-10 or D11-13,the effect of mCREG was only reversed by Dll-13,indicating that the binding to domains 11-13 is required for CREG to modulate the proliferation of SMCs.Conclusions These data suggest that solubleCREG proteins can exert their biological function via binding to the extracellular domains 7-10 and 11-13 of cell surface M6P/IGF2R in both a glycosylation-dependent and -independent manner.

生长分化因子11激活PI3K/Akt信号途径促进糖尿病小鼠ADSCs体外矿化的研究

目的探讨生长分化因子11(GDF11)对糖尿病小鼠来源的脂肪间充质干细胞(ADSCs)成骨分化的影响及信号机制。方法分离、培养糖尿病小鼠ADSCs。流式细胞术检测GDF11对ADSCs增殖的影响。成骨诱导培养后,检测碱性磷酸酶(ALP)活性,实时荧光定量PCR(qPCR)检测成骨相关基因的表达,Western blot检测PI3K、Akt的磷酸化水平。结果GDF11不影响ADSCs增殖,呈浓度依赖性地增强ALP活性,且具有饱和性;显著上调Runx2[(2.41±0.19)vs.(1.00±0.11)]、Osx[(1.41±0.21)vs.(1.00±0.10)]和ALP[(1.42±0.20)vs.(1.00±0.09)]的表达(P<0.05);明显提高PI3K[(2.84±0.19)vs.(1.00±0.11)]和Akt[(4.58±0.20)vs.(1.00±0.19)]的磷酸化水平(P<0.05)。而且,GDF11促进糖尿病小鼠ADSC s成骨分化的作用被PI3K抑制剂LY294002部分减弱。结论GDF11促进糖尿病小鼠ADSCs骨向分化,其作用机制可能与激活PI3K/Akt信号通路有关。

外源性GDF11通过调控LOX-1介导的内质网应激途径改善高血压大鼠血管重构

[目的]探讨外源性生长分化因子11(GDF11)通过调控凝集素样氧化型低密度脂蛋白受体1(LOX-1)介导的内质网应激途径对高血压大鼠血管重构的影响。[方法]将大鼠随机分为对照组、模型组(AngⅡ诱导的高血压大鼠模型组)、r-GDF11组、Ab-LOX-1组和r-GDF11+r-LOX-1组,每组10只。采用动物无创血压仪检测大鼠尾动脉血压变化;HE染色评估主动脉血管形态;Masson染色评估主动脉组织胶原沉积情况;Western blot检测主动脉组织中GDF11、LOX-1及内质网应激相关蛋白表达。[结果]与对照组比较,模型组大鼠血压升高,主动脉组织中膜厚度(MT)、MT/管腔内径(LD)比值增大,LD减小(均P<0.05);主动脉组织胶原纤维容积分数(CVF)值、葡萄糖调节蛋白78(GRP78)和转录激活因子6(ATF6)蛋白表达、磷酸化PKR样内质网调节激酶(p-PERK)/PERK和磷酸化肌醇需求酶1α(p-IRE1α)/IRE1α比值均升高(均P<0.05)。与模型组比较,r-GDF11组和Ab-LOX-1组大鼠血压降低,主动脉组织MT、MT/LD均减小,LD增大(均P<0.05);主动脉组织CVF值、GRP78和ATF6蛋白表达、p-PERK/PERK和p-IRE1α/IRE1α比值均显著降低(均P<0.05)。与r-GDF11组比较,r-GDF11+r-LOX-1组大鼠血压升高,主动脉组织MT、MT/LD增大,LD减小(均P<0.05);主动脉组织CVF值、GRP78和ATF6蛋白表达、p-PERK/PERK和p-IRE1α/IRE1α比值显著升高(均P<0.05)。[结论]外源性GDF11通过抑制LOX-1介导的内质网应激途径改善高血压大鼠血管重构。

糖尿病肾病患者血清GDF-11、S100A4水平与病情、疾病转归的关系分析

目的探讨血清生长分化因子-11(GDF-11)、S100钙结合蛋白A4(S100A4)水平与糖尿病肾病(DN)患者病情严重程度、疾病转归的关系。方法选取2021年5月至2023年1月该院收治的95例DN患者作为研究组,另选取110例体检健康者作为健康组。将DN患者根据病情严重程度分为轻度组(n=66)、重度组(n=29),患者出院后连续随访半年,根据预后情况分为预后良好组(n=64)和预后不良组(n=31)。采用酶联免疫吸附试验检测血清GDF-11、S100A4水平,采用Spearman等级相关分析探讨血清GDF-11、S100A4水平与病情严重程度的关系,采用受试者工作特征(ROC)曲线评估血清GDF-11、S100A4对DN患者疾病转归的预测价值,采用多因素Logistic回归分析DN患者疾病转归的影响因素。结果轻度组、重度组血清GDF-11、S100A4水平高于健康组,且重度组高于轻度组(P<0.05)。DN患者血清GDF-11、S100A4水平与病情严重程度均呈正相关(P<0.05)。预后良好组血清GDF-11、S100A4水平低于预后不良组(P<0.05)。血清GDF-11、S100A4预测DN患者疾病转归的曲线下面积(AUC)分别为0.785、0.839,二者联合预测的AUC为0.902。预后不良组2型糖尿病(T2DM)病程、肾小球分级为Ⅲ~Ⅳ级、间质炎症评分为2分、间质性纤维化和小管萎缩(IFTA)评分为2~3分、估算肾小球滤过率<60 mL/(min·1.73 m^(2))占比及总胆固醇、甘油三酯、低密度脂蛋白胆固醇、24 h尿蛋白定量、糖化血红蛋白、C肽、血细胞比容、红细胞沉降率水平高于预后良好组(P<0.05)。多因素Logistic回归分析显示,T2DM病程≥12.0年、IFTA评分≥2分、GDF-11≥700.82 ng/mL、S100A4≥211.53 ng/L是DN患者预后不良的危险因素(P<0.05)。结论血清S100A4、GDF-11水平在DN患者中呈高表达,且与病情严重程度及疾病转归相关,有望作为评估DN病情及预后的潜在标志物。

Identification of cytokines involved in hepatic differentiation of mBM-MSCs under liver-injury conditions

AIM: To identify the key cytokines involved in hepatic differentiation of mouse bone marrow mesenchymal stem cells (mBM-MSCs) under liver-injury conditions. METHODS: Abdominal injection of CCl4 was adopted to duplicate a mouse acute liver injury model. Global gene expression analysis was performed to evaluate the potential genes involved in hepatic commitment under liver-injury conditions. The cytokines involved in hepatic differentiation of mBM-MSCs was function-ally examined by depletion experiment using specifi c antibodies, followed by rescue experiment and direct inducing assay. The hepatic differentiation was characterized by the expression of hepatic lineage genes and proteins, as well as functional features. RESULTS: Cytokines potentially participating in hepatic fate commitment under liver-injury conditions were initially measured by microarray. Among the up-regulated genes determined, 18 cytokines known to closely relate to liver growth, repair and development, were selected for further identif ication. The f ibroblast growth factor-4 (FGF-4), hepatocyte growth factor (HGF) and oncostatin M (OSM) were fi nally found to be involved in hepatic differentiation of mBM-MSCs under liver-injury conditions. Hepatic differentiation could be dramatically decreased after removing FGF-4, HGF and OSM from the liver-injury conditioned medium, and could be rescued by supplementing these cytokines. The FGF-4, HGF and OSM play different roles in the hepatic differentiation of mBM-MSCs, in which FGF-4 and HGF are essential for the initiation of hepatic differentiation, while OSM is critical for the maturation of hepatocytes. CONCLUSION: FGF-4, HGF and OSM are the key cytokines involved in the liver-injury conditioned medium for the hepatic differentiation of mBM-MSCs.

BMP-4 induced proliferation and oriented differentiation of rat hepatic oval cells into hepatocytes

Objective:To explore the role of bone morphogenetic protein 4(BMP-4) in hepatic progenitor cells(HPCs).Methods:The effect of BMP-4 on rat hepatic oval cells was examined by using the WB-F344 rat hepatocytic epithelial stem-cell-like cell line.This hepatocytic cell line could exert various hepatocytc functions including the secretion of albumin and urea.Immunohistochemistry was used to examine the effects of BMP-4 and its antagonist,Noggin,on the proliferation and differentiation of these cells,cellular uptake and excretion of indocyanine green,the periodic acid-schiff(PAS) assay for glycogen storage and the expression of hepatic markers.Results:Our results showed for the first time that BMP-4 may acted as a potential inducer of hepatic differentiation in rat hepatic oval cells.Conclusions:This cell source offers a much-needed attractive and expandable source for future investigations of drug screening,stem cell technologies and cellular transplantation,in a society with increasing levels of liver disease and damage.

慢性心力衰竭患者氧化应激及血清GDF11、HO-1水平与心功能指标的相关性分析

目的探讨慢性心力衰竭(CHF)患者生长分化因子11(GDF11)、血红素加氧酶-1(HO-1)、氧化应激水平与心功能指标的相关性。方法选取2021年2月—2023年2月川北医学院附属医院收治的134例CHF患者为研究组,并根据纽约心脏协会心功能分级将其分为3个亚组[Ⅱ级组(47例)、Ⅲ级组(61例)、Ⅳ级组(26例)]。另选取同期于该院体检的70例健康者为对照组。比较研究组与对照组、不同病情严重程度CHF患者的血清GDF11、HO-1、氧化应激指标[谷胱甘肽过氧化物酶3(GSH-Px3)、总抗氧化能力(T-AOC)、晚期蛋白氧化产物(AOPP)、过氧化脂质(LPO)]及心功能指标[脑钠肽(BNP)、心肌肌钙蛋白I(cTnI)、左室舒张末期内径(LVEDD)、左心室质量指数(LVMI)、左心室射血分数(LVEF)];采用Pearson法分析GDF11、HO-1、氧化应激水平与心功能指标的相关性。结果研究组GDF11、HO-1水平高于对照组(P<0.05)。研究组GSH-Px3、T-AOC水平低于对照组,AOPP、LPO水平高于对照组(P<0.05)。研究组BNP、cTnI、LVEDD、LVMI水平高于对照组,LVEF水平低于对照组(P<0.05)。Ⅱ级组GDF11、HO-1、AOPP、LPO、BNP、cTnI、LVEDD、LVMI水平低于Ⅲ级组与Ⅳ级组(P<0.05),Ⅲ级组GDF11、HO-1、AOPP、LPO、BNP、cTnI、LVEDD、LVMI水平低于Ⅳ级组(P<0.05),Ⅱ级组GSH-Px3、T-AOC、LVEF水平高于Ⅲ级组与Ⅳ级组(P<0.05),Ⅲ级组GSH-Px3、T-AOC、LVEF水平高于Ⅳ级组(P<0.05)。Pearson相关性分析显示,GDF11、HO-1、AOPP、LPO与BNP、cTnI、LVEDD、LVMI呈正相关,与LVEF呈负相关(P<0.05);GSH-Px3、T-AOC与BNP、cTnI、LVEDD、LVMI呈负相关,与LVEF呈正相关(P<0.05)。结论CHF患者GDF11、HO-1、氧化应激水平、心功能均处于异常状态,且GDF11、HO-1、氧化应激水平与心功能之间存在密切联系。

GDF11调控NF-κB通路影响巨噬细胞M1极化介导的炎症反应

目的探讨生长转化因子11(GDF11)对巨噬细胞M1极化介导的炎症反应的影响及其作用机制。方法体外培养RAW264.7巨噬细胞,100 ng/mL LPS+30 ng/mL IFN-γ诱导巨噬细胞M1极化,并将细胞分成对照组、LPS+IFN组、GDF11组及LPS+IFN+GDF11组。流式细胞术检测巨噬细胞的极化,实时PCR检测IL-6、TNF-α、NF-κB及GDF11 mRNA的表达水平,Western blotting检测细胞GDF11、NF-κB及P-NF-κB蛋白表达水平,免疫荧光检测细胞GDF11、NF-κB的蛋白表达和分布。结果与对照组相比,LPS+IFN组巨噬细胞M1极化比例增加,IL-6、TNF-α及NF-κB mRNA表达升高,GDF11 mRNA及蛋白表达降低,NF-κB、P-NF-κB蛋白表达升高,P-NF-κB/NF-κB比值升高,NF-κB入核明显(P<0.05);与对照组相比,GDF11组巨噬细胞M1极化比例、IL-6、TNF-α、NF-κB mRNA表达及P-NF-κB/NF-κB比值无明显变化(P>0.05);与LPS+IFN组相比,LPS+IFN+GDF11组RAW264.7巨噬细胞M1极化比例显著降低,IL-6、TNF-α、NF-κB mRNA表达降低,NF-κB、P-NF-κB蛋白表达降低,P-NF-κB/NF-κB比值降低(P<0.05)。结论GDF11可减轻巨噬细胞M1极化介导的炎症反应,其机制可能与调节NF-κB通路有关。

Effect of the gap junction blocker 1-heptanol on chondrogenic differentiation of mouse bone marrow mesenchymal stem cells in vitro

Objective：To investigate the effect of the gap junction blocker 1-heptanol on the in vitro chondrogenic differentiation of mouse bone marrow mesenchymal stem cells（MSCs） following induction by GDF-5. Methods：MSCs were isolated from mouse bone marrow and cultured in vitro. After 3 passages cells were induced to undergo chondrogenic differentiation with recombinant human GDF-5（100 ng/ml）, with or without 1-heptanol（2.5 la mol/L）. The effect of 1-heptanol on MSCs proliferation was investigated using the MTT assay. Type II collagen mRNA and protein were examined by RT-PCR and immunocytochemistry respectively, and the sulfate glycosaminoglycan was assessed by Alcian blue dye staining. Connexin43（Cx43） protein was examined by western blotting. Results：GDF-5 induced proliferation and chondrogenic differentiation of MSCs. While 1-heptanol treatment had no effect on this proliferation, it inhibited the expression of both type II collagen mRNA and protein. The Alcian blue staining revealed that 1-heptanol also inhibited the deposition of the typical cartilage extracellular matrix promoted by recombinant GDF-5. Western blotting demonstrated that 1-heptanol had no effect on the expression of Cx43. Conclusion：These results suggest that mouse bone marrow MSCs can be differentiated into a chondrogenic phenotype by GDF- 5 administration in vitro. While the gap junction blocker, 1-heptanol, did not reduce gap junction Cx43, these intercellular communication pathways clearly played an important functional role in GDF-5-induced cartilage differentiation.

血清GDF11、Copeptin水平对充血性心力衰竭病人新活素治疗疗效及死亡风险的预测作用

目的:探讨血清生长分化因子11(growth differentiation factor 11,GDF11)、羧基端糖基化肽(Copeptin)水平对充血性心力衰竭(CHF)病人新活素治疗疗效及死亡风险的预测作用。方法:选取156例CHF病人,均给予新活素治疗,比较受试者治疗前后NYHA心功能分级、心电图QRS宽度、左心室舒张末期内径(LVEDD)、左室射血分数(LVEF)、血清GDF11和Copeptin的水平。随访至2021年10月,根据病人预后情况,对2组病人基线资料采用单因素和多因素logistic回归分析,并应用受试者工作特征曲线(ROC)分析血清GDF11、Copeptin预测死亡风险的价值。结果:治疗前NYHA心功能分级中,Ⅱ级为48例,Ⅲ级为36例,Ⅳ级为72例,治疗后NYHA心功能分级Ⅰ级为29例,Ⅱ级为67例,Ⅲ级为43例,Ⅳ级为17例,心功能明显改善(P<0.01);治疗后QRS宽度和LVEDD数值明显低于治疗前(P<0.01),LVEF数值明显高于治疗前(P<0.01),GDF11和Copeptin数值明显低于治疗前(P<0.01);Pearson相关分析显示,血清GDF11和Copeptin均与LVEDD(r=0.291和0.268)和QRS宽度(r=0.247和0.222)成正相关关系(P<0.01),而与LVEF(r=-0.310和-0.261)成负相关关系(P<0.01);随访至2021年10月,失访5例,中位随访时间为(29.23±3.00)个月。死亡组病人的hs-CRP、TNF-α、GDF11和Copeptin指标均高于生存组(P<0.01)。logistic回归分析结果表明血清GDF11(OR=1.702,95%CI:1.348~2.150)和血清Copeptin(OR=2.166,95%CI:1.458~3.219)是CHF病人死亡的影响因素(P<0.01)。根据ROC曲线可得,血清GDF11诊断的临界值为765.44 ng/mL,其对应的敏感度为70.37%,特异性为70.16%,AUC为0.785(95%CI:0.730~0.839);血清Copeptin诊断的临界值为26.29 pmol/L,其对应的敏感度为59.26%,特异性为59.68%,AUC为0.635(95%CI:0.571~0.700)。两者联合诊断的敏感度为88.89%,特异性为78.23%,AUC为0.878(95%CI:0.831~0.908),敏感度和AUC明显高于GDF11和Copeptin单独预测(P<0.05)。结论:应用新活素治疗CHF效果确切,可有效改善心功能,且降低血清GDF11和Copeptin。血清GDF11和Copeptin是CHF死亡的独立影响因素,且可通过联合诊断提高预测CHF病人死亡的诊断效能。

生长分化因子11对股动脉介入损伤术后小鼠血管的修复作用

目的 探讨生长分化因子11(growth differentiation factor 11,GDF11)对股动脉介入损伤术后小鼠血管的修复作用。方法 利用导丝损伤的方法构建C57BL/6小鼠股动脉损伤模型,将30只小鼠随机分为对照组、模型组和重组人GDF11蛋白处理组(GDF11组),每组10只。干预14天时,采用苏木精-伊红(hematoxylin-eosin,HE)染色检测各组小鼠股动脉内膜增生情况,同时采用免疫荧光检测各组小鼠损伤段血管CD31表达水平,利用酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)检测各组小鼠外周血血清中血管内皮生长因子(vascular endothelial growth factor,VEGF)、表皮生长因子(epidermal growth factor,EGF)、基质细胞衍生因子1(stromal-derived factor-1,SDF-1)、碱性成纤维细胞生长因子(basic fibroblast growth factor,bFGF)的含量变化。结果 与对照组比较,模型组股动脉损伤小鼠外周血中促血管修复因子VEGF、SDF-1、bFGF、EGF的含量明显升高(P均<0.01);损伤血管增生内膜与中膜面积比显著升高(P<0.01);模型组小鼠CD31表达明显降低(P<0.01)。与模型组相比,GDF11组小鼠促血管修复因子VEGF、SDF-1、bFGF、EGF的含量明显升高(P均<0.01);损伤段血管增生内膜与中膜面积比下降(P<0.01);CD31表达明显增多(P<0.01)。结论 GDF11能够抑制股动脉损伤小鼠损伤血管的新生内膜过度增生,促进再内皮化,从而对损伤血管发挥修复作用。

血清GDF-11、S100A4对冠状动脉造影术后对比剂肾病的预测效能

目的探讨血清生长分化因子11(GDF-11)、钙卫蛋白A4(S100A4)对冠状动脉造影(CAG)术后对比剂肾病(CIN)的预测效能。方法选取2020年12月至2023年11月于西安交通大学第一附属医院行CAG术的528例患者为研究对象,根据患者是否发生CIN将其分为非肾病组(472例)与肾病组(56例)。采用酶联免疫吸附试验检测血清GDF-11、S100A4水平,采用多因素Logistic回归分析探讨CAG术后CIN的影响因素,采用受试者工作特征(ROC)曲线评估血清GDF-11、S100A4对CAG术后CIN的预测价值。结果肾病组血清GDF-11、S100A4水平高于非肾病组(P<0.05)。肾病组对比剂剂量、术后血肌酐水平高于非肾病组(P<0.05)。多因素Logistic回归分析显示,对比剂剂量≥131.84 mL(OR=2.158,95%CI 1.284~3.627)、术后血肌酐≥87.57μmol/L(OR=2.445,95%CI 1.533~3.898)、GDF-11≥450.84 ng/mL(OR=3.043,95%CI 1.789~5.177)是CAG术后发生CIN的影响因素(P<0.05)。血清GDF-11、S100A4预测CAG术后CIN的曲线下面积(95%CI)分别为0.861(95%CI 0.810~0.912)、0.798(95%CI 0.747~0.849),最佳临界值分别为450.84 ng/mL、86.98 pg/mL,特异度分别为65.89%、57.62%,灵敏度分别为94.74%、94.74%,二者联合诊断的曲线下面积为0.906(95%CI 0.856~0.957),特异度为87.09%,灵敏度为84.26%。结论血清GDF-11、S100A4水平升高与CAG术患者术后发生CIN密切相关,可作为评估CAG术患者术后发生CIN的生物学指标,且二者联合预测的效能更高。

老年维持性血液透析患者血清GDF11和Cdc42变化与心血管事件的相关性研究

目的探究老年维持性血液透析患者血清生长分化因子11(growth differentiation factor 11,GDF11)、细胞分裂周期蛋白42(cell division cycle 42,Cdc42)水平变化与患者发生主要不良心血管事件(major adverse cardiac events,MACE)的相关性。方法选择四川省南充市中心医院2019年6月至2021年6月收治的164例老年维持性血液透析患者作为观察对象(血液透析组),同时选取同时期未进行血液透析的老年终末期肾病患者150例作为未血液透析组,在本院体检的健康老年人员148名作为对照组,根据2年内是否发生MACE,将患者分为非MACE组92例和MACE组72例,采用酶联免疫吸附法测定血清GDF11、Cdc42水平,多因素Logistic回归分析影响老年维持性血液透析患者2年发生MACE的影响因素,受试者工作特征曲线分析GDF11、Cdc42水平对老年维持性血液透析患者发生MACE的预测价值。结果对照组、未血液透析组及血液透析组之间比较,血液透析组患者血清中GDF11水平[(640.38±137.55)μg/L比(851.56±215.61)μg/L,(785.27±174.62)μg/L]显著降低(t=10.414,P<0.001),Cdc42水平[(57.22±11.34)μg/L比(35.61±10.25)μg/L,(43.58±10.72)μg/L]显著升高(t=17.589,P<0.001);MACE组Cdc42水平显著高于非MACE组[(65.31±21.22)μg/L比(50.89±15.56)μg/L],GDF11水平显著低于非MACE组[(576.56±125.71)μg/L比(690.33±159.84)μg/L],差异具有统计学意义(均P<0.05)。GDF11、Cdc42水平和二者联合预测发生MACE的曲线下面积分别为0.762、0.794、0.868,二者联合预测的价值优于单独预测(Z=3.183、3.047;P=0.002、0.002)。结论老年维持性血液透析患者血清中GDF11水平降低,Cdc42水平升高,二者均与患者MACE发生密切相关,可能作为老年终末期肾病血液透析患者病情诊断、治疗和预后评估的标志物。

血清FGFR-4、CCL11、CA125检测对分化型甲状腺癌的诊断价值

目的探究血清成纤维生长因子受体4(FGFR-4)、嗜酸性粒细胞趋化因子(CCL11)、糖类抗原(CA125)检测对分化型甲状腺癌的诊断价值。方法选取2019年2月至2023年2月来我院治疗的133例确诊为分化型甲状腺癌的患者为观察组,选同期来我院治疗的108例甲状腺良性病变患者为良性病变组,另选取同期来我院体检的120例健康者为对照组。采用酶联免疫吸附测定法(ELISA)测定血清FGFR-4、CCL11水平,采用电化学发光法测定CA125水平;受试者工作特征(ROC)曲线分析血清FGFR-4、CCL11、CA125水平对分化型甲状腺癌的诊断价值。结果甲状腺癌组和良性病变组患者血清FGFR-4、CCL11、CA125水平与对照组相比均显著升高,且甲状腺癌组与良性病变组相比均显著升高(P<0.05);甲状腺癌组患者手术后血清FGFR-4、CCL11、CA125水平均显著下降(P<0.05);血清FGFR-4、CCL11、CA125水平在低分化、有淋巴结转移、III+IV期患者中的水平显著升高(P<0.05);血清FGFR-4、CCL11、CA125诊断分化型甲状腺癌的曲线下面积(AUC)分别为0.793、0.808、0.647,3项联合诊断的AUC为0.867,3项联合诊断优于各个指标单独诊断(Z_(二者联合vs FGFR-4)=2.334、Z_(二者联合vs CCL11)=1.966、Z_(二者联合vs CA125)=5.132,P=0.020、0.048、0.000)。结论分化型甲状腺癌患者血清FGFR-4、CCL11、CA125水平均上调,且血清FGFR-4、CCL11、CA125水平与分化程度、淋巴结转移情况以及TNM分期有关,3项联合诊断分化型甲状腺癌具有更高的诊断价值。