Diagnostic significance of serum levels of serum amyloid A,procalcitonin,and high-mobility group box 1 in identifying necrotising enterocolitis in newborns

BACKGROUND Necrotising enterocolitis(NEC)is a critical gastrointestinal emergency affecting premature and low-birth-weight neonates.Serum amyloid A(SAA),procalcitonin(PCT),and high-mobility group box 1(HMGB1)have emerged as potential biomarkers for NEC due to their roles in inflammatory response,tissue damage,and immune regulation.AIM To evaluate the diagnostic value of SAA,PCT,and HMGB1 in the context of NEC in newborns.METHODS The study retrospectively analysed the clinical data of 48 newborns diagnosed with NEC and 50 healthy newborns admitted to the hospital.Clinical,radiological,and laboratory findings,including serum SAA,PCT,and HMGB1 Levels,were collected,and specific detection methods were used.The diagnostic value of the biomarkers was evaluated through statistical analysis,which was performed using chi-square test,t-test,correlation analysis,and receiver operating characteristic(ROC)analysis.RESULTS The study demonstrated significantly elevated levels of serum SAA,PCT,and HMGB1 Levels in newborns diagnosed with NEC compared with healthy controls.The correlation analysis indicated strong positive correlations among serum SAA,PCT,and HMGB1 Levels and the presence of NEC.ROC analysis revealed promising sensitivity and specificity for serum SAA,PCT,and HMGB1 Levels as potential diagnostic markers.The combined model of the three biomarkers demonstrating an extremely high area under the curve(0.908).CONCLUSION The diagnostic value of serum SAA,PCT,and HMGB1 Levels in NEC was highlighted.These biomarkers potentially improve the early detection,risk stratification,and clinical management of critical conditions.The findings suggest that these biomarkers may aid in timely intervention and the enhancement of outcomes for neonates affected by NEC.

Inhibition of high-mobility group box 1 expression by siRNA in rat hepatic stellate cells

AIM:To explore the role of high-mobility group box 1 (HMGB1) protein during liver fibrogenesis and investigate the functional effects of HMGB1 gene silencing in hepatic stellate cells (HSCs) using siRNA.METHODS:Hepatic fibrosis in rats was induced through serial subcutaneous injections of dimethylnitrosamine,and expression of HMGB1 was detected by immunohistochemistry.HMGB1 siRNAs were developed and transiently transfected into HSC-T6 cells using Lipofectamine 2000.HMGB1 expression was evaluated by real-time polymerase chain reaction (PCR) and Western blotting analysis.Expression of α-smooth muscle actin (α-SMA) and collagen typesⅠand Ⅲ was evaluated by real-time PCR.Cell proliferation and the cell cycle were determined using the methyl thiazolyl tetrazolium method.Finally,collagen content in HSC supernatant was evaluated by an enzyme-linked immunosorbent assay.RESULTS:The results showed that HMGB1 was upregulated during liver fibrosis and that its expression was closely correlated with the deposition of collagen.siRNA molecules were successfully transfected into HSCs and induced inhibition of HMGB1 expression in a time-dependent manner.Moreover,HMGB1 siRNA treatment inhibited synthesis of α-SMA and collagen types Ⅰ and Ⅲ in transfected HSCs.CONCLUSION:This study suggests a significant functional role for HMGB1 in the development of liver fibrosis.It also demonstrates that downregulation of HMGB1 expression might be a potential strategy to treat liver fibrosis.

Inhibition of LOX-1 alleviates the proinflammatory effects of high-mobility group box 1 in Aspergillus fumigatus keratitis

AIM: To investigate the inflammatory amplification effect of high-mobility group box 1(HMGB1) in Aspergillus fumigatus(A. fumigatus) keratitis and the relationship between lectin-like oxidized low-density lipoprotein receptor 1(LOX-1) and HMGB1 in keratitis immune responses.METHODS: Phosphate buffer saline(PBS), and Boxb were injected into BALB/c mice subconjunctivally before the corneas were infected with A. fumigatus. RAW264.7 macrophages and neutrophils were pretreated with PBS and Boxb to determine the HMGB1 inflammatory amplification effects. Abdominal cavity extracted macrophages were pretreated with Boxb and Poly(I)(a LOX-1 inhibitor) before A. fumigatus hyphae stimulation to prove the the relationship between the two molecules. LOX-1, interleukin-1β(IL-1β), tumor necrosis factor-α(TNF-α), macrophage inflammatory protein-2(MIP-2) and IL-10 were assessed by polymerase chain reaction and Western blot.RESULTS: Pretreatment with Boxb exacerbated corneal inflammation. In macrophages and neutrophils, A. fumigatus induced LOX-1, IL-1β, TNF-α and MIP-2 expression in Boxb group was higher than those in PBS group. Poly(I) treatments before infection alleviated the proinflammatory effects of Boxb in abdominal cavity extracted macrophages. Pretreatment with Boxb did not influence Dectin-1 mRNA levels in macrophages and neutrophils.CONCLUSION: In fungal keratitis, HMGB1 is a proinflammatory factor in the first line of immune response. HMGB1 mainly stimulates neutrophils and macrophages to produce inflammatory cytokines and chemokines during the immune response. LOX-1 participates in HMGB1 induced inflammatory exacerbation in A. fumigatus keratitis.

Scolopendra subspinipes mutilans protected the ceruleininduced acute pancreatitis by inhibiting high-mobility group box protein-1

AIM:To evaluate the inhibitory effects of Scolopendra subspinipes mutilans(SSM) on cerulein-induced acute pancreatitis(AP) in a mouse model.METHODS:SSM water extract(0.1,0.5,or 1 g/kg) was administrated intraperitoneally 1 h prior to the first injection of cerulein.Once AP developed,the stable cholecystokinin analogue,cerulein was injected hourly,over a 6 h period.Blood samples were taken 6 h later to determine serum amylase,lipase,and cytokine levels.The pancreas and lungs were rapidly removed for morphological examination,myeloperoxidase assay,and real-time reverse transcription polymerase chain reaction.To specify the role of SSM in pancreatitis,the pancreatic acinar cells were isolated using collagenase method.Then the cells were pre-treated with SSM,then stimulated with cerulein.The cell viability,cytokine productions and high-mobility group box protein-1(HMGB-1) were measured.Furthermore,the regulating mechanisms of SSM action were evaluated.RESULTS:The administration of SSM significantly attenuated the severity of pancreatitis and pancreatitis associated lung injury,as was shown by the reduction in pancreatic edema,neutrophil infiltration,vacuolization and necrosis.SSM treatment also reduced pancreatic weight/body weight ratio,serum amylase,lipase and cytokine levels,and mRNA expression of multiple inflammatory mediators such as tumor necrosis factor-α and interleukin-1β.In addition,treatment with SSM inhibited HMGB-1 expression in the pancreas during AP.In accordance with in vivo data,SSM inhibited the cerulein-induced acinar cell death,cytokine,and HMGB-1 release.SSM also inhibited the activation of c-Jun NH2-terminal kinase,p38 and nuclear factor(NF)-κB.CONCLUSION:These results suggest that SSM plays a protective role during the development of AP and pancreatitis associated lung injury via deactivating c-Jun NH2-terminal kinase,p38 and NF-κB.

Clinical signification of high-mobility group box 1protein(HMGB1) expression in infiltrating ductalcarcinoma breast tissue

Objective: Exploring the clinical signification of high-mobility group box 1 protein(HMGB1) expression in infiltrating ductal carcinoma(IDC) breast tissue. Methods: The expression of HMGB1 protein in IDC breast tissue was detected by immunohistochemistry, and the relations among size of tumour, lymph node metastasis, clinical staging, estrogen receptor(ER), progesterone receptor(PR) and human epidermal growth factor receptor 2(HER-2) were also analyzed. Results: Fortysix cases out of 60 cases of IDC breast tissue showed positive or strong positive HMGB1 expression(76.67%), statistical significance was observed between HMGB1 expression with clinical staging(P < 0.01), lymph node metastasis(P < 0.01), breast cancer ER(P < 0.05) and HER-2(P < 0.05), however same conclusion can not be drawn between HMGB1 with either size of tumour or PR expression(P > 0.05) in IDC breast tissue. Spearman analysis showed negative correlation between HMGB1 expression and ER, and positive correlation between HMGB1 expression and clinical staging, lymph node metastasis together with HER-2. Conclusion: It's promising that HMGB1 expression in IDC tissue can be one of biological indicators of poor prognosis.

Relationship of plasma homocysteine, soluble intercellular adhesion molecule-1, high mobility group box 1 protein with carotid intima-media thickness in elderly patients with type 2 diabetes mellitus

Objective:To explore the relationship of plasma homocysteine(Hcy),soluble intercellular adhesion molecule-1(sICAM-1)and high mobility group box 1 protein(HMGB1)with carotid intima-media thickness(c-IMT)in elderly patients with type 2 diabetes mellitus.Methods:A total of 100 elderly patients who were diagnosed as type 2 diabetes mellitus in Baogang Hospital of Inner Mongolia from June 2017 to May 2020 were chosen as research objects.According to c-IMT,they were divided into the normal group(n=35),the mild to moderate group(n=41)and the severe group(n=24).The expression levels of plasma Hcy,sICAM-1 and HMGB1 were compared between groups respectively.Pearson’s correlation coefficient was used to analyze the relationship of plasma Hcy,sICAM-1,HMGB1 with c-IMT.Results:The comparison in plasma Hcy,sICAM-1,HMGB1 and c-IMT among the three groups of patients was of statistical significance(p<.05).The results of correlation analysis showed that the expression levels of plasma Hcy,sICAM-1 and HMGB1 were positively correlated with c-IMT in elderly patients with type 2 diabetes mellitus(r=.627,.598,.614;p<.05).Conclusions:The expression levels of plasma Hcy,sICAM-1 and HMGB1 are abnormally increased in elderly patients with type 2 diabetes mellitus,and related to c-IMT,which can provide a strong evidence for clinical diagnosis and treatment by detecting their levels in clinical practice.

下调HMGB2表达对肝癌LM3细胞上皮-间质转化的抑制作用及其AKT/mTOR信号通路机制

目的:探讨下调肝癌细胞中高迁移率族框蛋白2 (HMGB2)表达对肝癌细胞生物学行为及上皮-间质转化(EMT)进程的影响,并阐明其作用机制。方法:对数生长期的人肝癌LM3细胞分为阴性对照组和HMGB2 RNA干扰组(HMGB2 siRNA组),分别以Lipofectamin 2000为载体转染无关序列的RNA寡核苷酸(RNA oligo)和敲除HMGB2序列的RNA oligo。采用实时荧光定量PCR(RT-qPCR)法和Western blotting法检测2组细胞中HMGB2 mRNA和蛋白表达水平,分别采用细胞划痕实验和Transwell小室实验检测2组细胞的迁移和侵袭能力,采用Western blotting法检测2组细胞中E-钙黏蛋白(E-cadherin)、 N-钙黏蛋白(N-cadherin)、波形蛋白(Vimentin)和蛋白激酶B(AKT)/哺乳动物雷帕霉素靶蛋白(mTOR)通路相关蛋白表达水平。结果:与阴性对照组比较,HMGB2 siRNA组细胞中HMGB2 mRNA和蛋白表达水平均明显降低(P<0.05),HMGB2 siRNA组细胞划痕愈合率明显降低(P<0.01),侵袭细胞数明显减少(P<0.01),细胞中E-cadherin蛋白表达水平明显升高(P<0.01),N-cadherin、Vimentin、mTOR、AKT和磷酸化AKT (p-AKT)蛋白表达水平明显降低(P<0.05或P<0.01)。结论:下调HMGB2的表达可降低肝癌LM3细胞迁移和侵袭能力并抑制EMT,其作用机制可能与参与调节AKT/mTOR通路相关蛋白表达有关。

血清Ang-2和HMGB1水平与脓毒症患者病情及预后相关性的研究

目的 探究血清血管生成素-2(Ang-2)和高迁移率族蛋白B1(HMGB1)水平与脓毒症患者病情及28天预后的相关性。方法 回顾性分析41例脓毒症患者资料,根据患者28天的预后情况分为存活组和死亡组;根据患者病情严重程度将2组患者分别分为脓毒症组和脓毒性休克组。记录所有入组患者的一般资料并进行第1天、第3天、第5天及第7天的APACHEⅡ评分,测定Ang-2和HMGB1水平。结果 存活组与死亡组患者的一般资料比较差异无统计学意义。死亡组脓毒症休克患者占比及入科APACHEⅡ评分为(85.71%)和(29.29±5.99)分,均明显高于存活组(51.84%)和(24.96±5.84)分。2组患者血清Ang-2和HMGB1水平比较差异无统计学意义。第7天的HMGB1水平与脓毒症患者预后显著相关,而第1天、第3天、第5天的HMGB1水平及上述各时间点的Ang-2水平与患者预后均无相关性。各时间点血清Ang-2水平的算术平均值与APACHEⅡ评分的算术平均值呈正相关,HMGB1水平与APACHEⅡ评分并无相关性。联合第7天的APACHEⅡ评分、Ang-2和HMGB1水平的曲线下面积最大为0.78,灵敏度为92.9%,特异度为59.3%。联合第7天的Ang-2和HMGB1水平预测预后不良的曲线下面积为0.71,灵敏度为92.9%,特异度为55.6%。结论 脓毒症患者第1天、第3天、第5天及第7天的平均血清Ang-2水平与病情严重程度呈正相关;联合脓毒症患者第7天的血清Ang-2和HMGB1水平对其预后进行评估灵敏度最高,明显高于单独某项指标的评估价值。

miR-let-7c-5p对膀胱癌细胞恶性行为、血管管腔形成及HMGB2的调控作用

目的探讨miR-let-7c-5p对膀胱癌细胞恶性行为、血管管腔形成及高迁移率族蛋白(HMGB)2的调控作用。方法将人膀胱癌UM-UC-3细胞分别进行常规培养(空白组)、转染mimics-NC(mimics-NC组)、转染miR-let-7c-5p-mimics(miR-let-7c-5p-mimics组)、转染inhibitor-NC(inhibitor-NC组)及转染miR-let-7c-5p-inhibitor(miR-let-7c-5p-inhibitor组)。采用实时荧光定量聚合酶链反应检测各组细胞miR-let-7c-5p转染成功率;采用Transwell法检测各组细胞侵袭能力;采用划痕试验检测各组细胞迁移能力;采用Matrigel胶小管形成试验检测各组细胞血管生成;采用免疫印迹法检测各组细胞HMGB2、血管内皮生长因子(VEGF)及葡萄糖转运蛋白-1(GLUT-1)蛋白表达水平;采用荧光素酶试验验证miR-let-7c-5p对HMGB2的调控作用。结果空白组、mimics-NC组及inhibitor-NC组miR-let-7c-5p mRNA表达水平、侵袭数目、划痕愈合率、血管管腔数目,HMGB2、VEGF及GLUT-1蛋白表达水平比较,差异均无统计学意义(P>0.05);与空白组、mimics-NC组及inhibitor-NC组比较,miR-let-7c-5p-mimics组miR-let-7c-5p mRNA表达水平升高,侵袭数目、划痕愈合率、血管管腔数目,HMGB2、VEGF及GLUT-1蛋白表达水平均降低,miR-let-7c-5p-inhibitor组miR-let-7c-5p mRNA表达水平降低,侵袭数目、划痕愈合率、血管管腔数目,HMGB2、VEGF及GLUT-1蛋白表达水平均升高,差异均有统计学意义(P<0.05)。荧光素酶试验结果显示,与mimics-NC组比较,miR-let-7c-5p-mimics组HMGB2-Wt水平降低,差异有统计学意义(P<0.05)。结论上调miR-let-7c-5p可明显抑制膀胱癌细胞侵袭、转移及血管生成,其机制与抑制HMGB2蛋白表达相关。

儿童肺炎支原体肺炎肺泡灌洗液CARDS TX、HMGB1、TLR2表达及临床意义

目的研究肺炎支原体肺炎(MPP)患儿肺泡灌洗液中社区获得性呼吸窘迫综合征毒素(CARDS TX)、高迁移率族蛋白1(HMGB1)、Toll样受体2(TLR2)表达,探讨其在MPP发病中的临床意义。方法回顾性分析55例MPP病例组及20例对照组临床资料,同时检测两组支气管肺泡灌洗液(BALF)中CARDS TX、HMGB1、TLR2、TLR4、晚期糖基化终末产物受体(RAGE)及髓样分化因子88(MyD88)表达,体外检测不同浓度CARDS TX刺激下HMGB1、TLR2的表达并进行比较。结果与对照组相比,MPP组LYM绝对值计数、PA、CD3^(+)、CD3^(+)CD4^(+)、CD3^(+)CD8^(+)、CD3-CD19^(+)、NK cell、CD19^(+)CD23^(+)明显下降,CRP、LDH、D-二聚体、IgA、IgG、IgM、CARDS TX、HMGB1、TLR2、MyD88均升高,差异有统计学意义(P<0.05或0.001)。CARDS TX刺激后HMGB1、TLR2的表达升高,且呈正相关,差异有统计学意义(P<0.001)。结论CARDS TX可能通过HMGB1-TLR2-MyD88途径参与肺炎支原体(MP)的致病。

慢性宫颈炎合并HPV感染140例血清SOX2和PD-L1表达与临床病理特征相关性分析

目的探究慢性宫颈炎合并人乳头状瘤病毒(HPV)感染病人血清性别决定区Y框蛋白2(SOX2)和程序性死亡配体1(PD-L1)表达与临床病理特征相关性。方法选取2020年10月至2022年10月在重庆市江津区中医院就诊治疗的未合并HPV感染慢性宫颈炎病人160例作为慢性宫颈炎组,慢性宫颈炎合并HPV感染病人140例作为合并HPV组,另选取在该院体检健康女性110例作为对照组。采用酶联免疫吸附测定(ELISA)检测研究对象血清SOX2水平,实时荧光定量PCR(qRT-PCR)测定血清PD-L1 mRNA水平,二代杂交捕获实验技术(HC2)检测HPV-DNA。结果对照组、慢性宫颈炎组、合并HPV组血清SOX2、PD-L1mRNA水平呈现逐渐升高的趋势(P<0.05);低危型HPV组、中危型HPV组、高危型HPV组血清SOX2、PD-L1 mRNA水平呈现逐渐升高的趋势(P<0.05);慢性宫颈炎合并HPV病人中单一感染率为45.71%(64/140),多重感染率为54.29%(76/140)(P>0.05),其中慢性宫颈炎合并HPV病人中低危型单一感染与多重感染病例数差异有统计学意义(P<0.05);高水平SOX2与低水平SOX2以及高水平PD-L1 mRNA与低水平PD-L1 mRNA慢性宫颈炎病人合并HPV率差异有统计学意义(P<0.05)。结论慢性宫颈炎合并HPV感染病人血清SOX2、PD-L1 mRNA水平均显著升高,与HPV分型密切相关,且病人血清SOX2、PD-L1 mRNA水平与HPV感染均呈正相关。

颅脑外伤患者血清IL-33、sST2、TNF-α、HMGB1水平与病情严重程度及短期预后的关系

目的探讨颅脑外伤患者血清白细胞介素(IL)-33、可溶性生长刺激表达基因2蛋白(sST2)、肿瘤坏死因子-α(TNF-α)、高迁移率族蛋白B1(HMGB1)与病情严重程度及短期预后的关系。方法回顾性分析2022年1月至2024年1月渭南市中心医院收治的142例颅脑外伤患者的临床资料。根据格拉斯哥昏迷量表(GCS)评估病情严重程度分为轻度组(13~15分)(n=47)、中度组(9~12分)(n=59)、重度(3~8分)(n=36)。根据格拉斯哥预后量表(GOS)评估28 d预后分为预后良好组(4~5分)(n=115)、预后不良组(1~3分)(n=27)。收集患者资料,包括性别、年龄、体重指数、吸烟、饮酒、高血压、糖尿病、外伤原因、IL-33、sST2、TNF-α、HMGB1。比较不同病情严重程度、不同预后患者的临床资料;采用Spearman秩相关分析检验颅脑外伤患者病情严重程度与IL-33、sST2、TNF-α、HMGB1水平的相关性;通过受试者操作特征(ROC)分析IL-33、sST2、TNF-α、HMGB1预测颅脑外伤患者预后不良的价值。结果重度组IL-33、sST2、TNF-α、HMGB1水平分别为(40.69±9.64)pg/mL、(7.75±2.14)ng/mL、(252.43±41.37)ng/L、(11.42±3.27)μg/L,中度组IL-33、sST2、TNF-α、HMGB1水平分别为(30.47±7.22)pg/mL、(6.45±1.37)ng/mL、(181.29±22.75)ng/L、(8.05±2.13)μg/L,均高于轻度组[(21.53±5.84)pg/mL、(5.61±1.05)ng/mL、(140.36±16.41)ng/L、(4.87±1.42)μg/L],差异均有统计学意义(P<0.05)。IL-33、sST2、TNF-α、HMGB1水平与颅脑外伤患者病情严重程度呈正相关(r=0.561、0.518、0.593、0.537,P<0.05)。预后不良组IL-33、sST2、TNF-α、HMGB1水平分别为(74.08±20.37)pg/mL、(10.56±2.54)ng/mL、(286.31±42.81)ng/L、(13.45±3.07)μg/L,均高于预后良好组[(22.64±6.19)pg/mL、(6.95±1.73)ng/mL、(153.76±31.45)ng/L、(5.86±1.49)μg/L],差异均有统计学意义(P<0.05)。ROC曲线分析结果显示,IL-33、sST2、TNF-α、HMGB1预测颅脑外伤患者预后不良的曲线下面积为0.891、0.846、0.905、0.928,均有P<0.05。结论IL-33、sST2、TNF-α、HMGB1表达水平随颅脑外伤患者病情严重程度升高而升高,且与其短期不良预后有关。

轻中型颅脑创伤继发认知功能障碍患者血清HMGB1和TLR2的表达

目的观察轻中型颅脑损伤继发轻度认知功能障碍(MCI)后血清中高迁移率族蛋白B1(highmobility group box-1,HMGB1)和Toll样受体2(toll like receptor-2,TLR2)的表达情况并探讨其临床意义。方法选择郑州大学第五附属医院神经外科自2016年1月至2017年12月收治的格拉斯哥昏迷评分(GCS)9~15分的轻中型颅脑损伤患者115例为颅脑损伤组,同期于门诊体检的20例符合条件的健康者作为健康组。应用双抗体夹心酶联免疫吸附实验(ELISA)测定两组血清中HMGB1、TLR2水平;分析各组血清中HMGB1和TLR2的表达。结果 3个月后,轻型颅脑损伤组和中型颅脑损伤组与健康组比较,其血清HMGB1、TLR2明显增高(q=42.442,24.264;均P<0.05);中型颅脑损伤组血清HMGB1水平比轻型颅脑损伤组比明显增高(q=17.122,P<0.05),TLR2水平无明显差异(q=2.416,P>0.05)。多组间血清中HMGB1和TLR2水平比较有统计学意义(F=520.008,169.249;均P<0.05);进一步分析,轻型颅脑损伤不伴MCI组和中型颅脑损伤不伴MCI组比健康组血清中HMGB1和TLR2水平高(q=33.252,24.126;均P<0.05),轻型颅脑损伤不伴MCI组和中型颅脑损伤不伴MCI组之间HMGB1和TLR2水平无明显差异(q=2.422,P>0.05);轻型颅脑损伤伴MCI组和中型颅脑损伤伴MCI组比健康组血清中HMGB1和TLR2水平高(q=48.374,44.522;均P<0.05),轻型颅脑损伤伴MCI组和中型颅脑损伤伴MCI组比轻型颅脑损伤不伴MCI组血清中HMGB1和TLR2水平高(q=28.674,21.351;均P<0.05),轻型颅脑损伤伴MCI组和中型颅脑损伤伴MCI组比中型颅脑损伤不伴MCI组血清中HMGB1和TLR2水平高(q=19.974,16.465;均P<0.05),轻型颅脑损伤伴MCI组和中型颅脑损伤伴MCI组之间HMGB1和TLR2水平无明显差异(q=3.584,P>0.05);颅脑损伤伴MCI患者血清中TLR2和HMGB1的水平呈正相关[Y=0.372X-0.408(r=0.874,P<0.01)];颅脑损伤不伴MCI患者血清中TLR2和HMGB1无明显相关性[Y=0.285X+0.038(r=0.459,P=0.064)]。结论轻中型颅脑损伤患者血清中高表达的HMGB1/TLR2与颅脑损伤合并MCI密切相关,有可能为预防和治疗颅脑损伤继发MCI提供新思路。

HMGB2沉默对乳腺癌MDA-MB-231细胞增殖、凋亡及侵袭迁移的影响

目的探讨HMGB2沉默对乳腺癌MDA-MB-231细胞增殖、凋亡及侵袭迁移的影响。方法采用将靶向HMGB2的寡核苷酸克隆至p LKO.1 puro载体质粒构建靶向沉默HMGB2表达的慢病毒shRNA。将MDA-MB-231细胞分为对照组、空转染组及HMGB2干扰组,其中空转染组及HMGB2干扰组分别转染包含随机序列shRNA和靶向HMGB2 shRNA的病毒液,对照组仅行常规培养而不行任何转染。采用Western blotting检测各组转染48 h后的HMGB2蛋白水平以评价转染效率,采用MTT法计算各组细胞的存活率,流式细胞术检测转染48 h后的细胞凋亡率,Transwell法检测转染48 h后的细胞侵袭和转移水平。结果 HMGB2干扰组的HMGB2蛋白水平低于对照组和空转染组,差异有统计学意义(P<0.05),提示构建的载体沉默HMGB2的效果显著。与对照组和空转染组比较,HMGB2干扰组的细胞存活率降低,且随着转染时间的延长而呈降低趋势,差异均有统计学意义(P<0.05);HMGB2干扰组细胞的早期、晚期及总凋亡率均高于其余两组,且侵袭细胞数目和迁移细胞数目均低于其余两组,以上差异均有统计学意义(P<0.05)。结论 shRNA靶向沉默MDA-MB-231细胞中HMGB2表达效果显著,可抑制细胞增殖及侵袭迁移并诱导细胞凋亡,为乳腺癌靶向治疗提供一定参考。

Sox2通过Wnt信号通路对宫颈癌侵袭及迁移能力的影响

目的探讨干细胞转录因子Sox2对宫颈鳞癌SiHa细胞侵袭及迁移能力的影响及其机制。方法采用RTPCR及Western blot方法分别检测本实验前期构建的稳定高表达Sox2的SiHa-Sox2细胞及对照组SiHa-EGFP细胞中Sox2的表达情况;细胞划痕实验及Transwell小室实验观察Sox2对SiHa细胞侵袭及迁移能力的影响;Western blot检测两组细胞中Wnt信号通路关键基因β-catenin的表达情况。结果稳定表达Sox2的SiHa-Sox2细胞中Sox2的mRNA及蛋白表达均明显增加;SiHa-Sox2组细胞侵袭及迁移能力增强;Western blot检测结果显示β-catenin表达上调。结论 Sox2通过上调β-catenin的表达激活Wnt信号通路,使宫颈鳞癌SiHa细胞的侵袭及迁移能力增强,从而促进宫颈鳞癌的发生发展。

HMGB1对淋巴细胞增殖、凋亡及Th1/Th2、Tc1/Tc2漂移的影响

目的：探索HMGB1是否参与淋巴细胞免疫功能的调节。方法：系列浓度HMGB1单独或与ConA联合刺激培养小鼠脾淋巴细胞。MTT法检测细胞增殖,流式细胞术（FCM）检测细胞凋亡、细胞表面CD3、CD8及细胞内IL-4、IFN-γ表达。结果：（1）HMGB1时间-剂量依赖性调节淋巴细胞增殖,而不影响其凋亡。（2）不同HMGB1浓度和刺激时间不影响淋巴细胞Th1、Th2及Th1/Th2变化,但10μg/L和100μg/L HMGB1刺激12-24h,Th1亚群占优势;培养12-24h,淋巴细胞Tc1亚群明显减少,Tc2无变化。Tc1/Tc2变化显示,1μg/L和10μg/L HMGB1刺激,Tc1亚群占优势。（3）培养12-24h,上清中IL-2增加,sIL-2R减少,IL-2/sIL-2R比例升高20-50倍,尤以10μg/L HMGB1刺激明显。结论：低剂量HMGB1可增强淋巴细胞免疫功能。

血清HMGB1、esRAGE水平与2型糖尿病颈动脉粥样硬化的相关性

目的 探讨血清高迁移率蛋白B1（HMGB1）、内源性分泌型糖基化终末产物受体（esRAGE）与2型糖尿病颈动脉粥样硬化的关系。方法 选取198例住院患者作为病例组，根据是否存在2型糖尿病及颈动脉彩超检测结果，将病例组分为3组，分别为单纯颈动脉粥样硬化组（AS）72例；单纯2型糖尿病组（T2DM）56例；2型糖尿病合并颈动脉粥样硬化组（T2DM＋AS）70例。同时选取46例健康体检者作为正常对照组。应用酶联免疫吸附试验（ELISA）测定血清HMGB1、esRAGE。结果 T2DM＋AS组患者血清HMGB1较其他组明显升高（P〈0.05），esRAGE显著降低（P〈0.01）；T2DM＋AS组血清HMGB1与esRAGE呈负相关（r=-0.317，P=0.002）；Logistic回归分析结果显示，HMGB1、esRAGE与2型糖尿病合并颈动脉粥样硬化独立相关（OR=1.202，P=0.021, 95% CI 1.028～1.406；OR=0.301, P=0.046, 95% CI 0.004～0.848）。结论 外周循环中高水平的HMGB1及低水平的esRAGE可能参与2型糖尿病合并颈动脉粥样硬化的发生发展。

脂多糖诱导小鼠腹腔巨噬细胞炎性反应中前列腺素E2/前列腺素E受体4调控高迁移率族蛋白1的机制

目的:探讨前列腺素E2(prostaglandin E2,PGE2)对脂多糖(lipopolysaccharide,LPS)诱导的小鼠腹腔巨噬细胞高迁移率族蛋白1(high mobility group box-1 protein,HMGB1)表达的影响及其可能的机制。方法:运用常规方法提取、培养小鼠腹腔巨噬细胞;采用LPS诱导小鼠腹腔巨噬细胞构建炎症模型;使用PGE2和前列腺素E受体(prostaglandin E receptor,EP)激动剂、RNAi技术下调巨噬细胞EP4受体表达、抑制性表达DN.CREB质粒处理LPS诱导的小鼠腹腔巨噬细胞,并通过Western印迹测定HMGB1表达水平。结果:与单纯应用LPS处理巨噬细胞比较,PGE2联合LPS共孵育小鼠腹腔巨噬细胞24 h后,HMGB1蛋白的表达水平明显上升,且EP4受体激动剂联合LPS处理小鼠腹腔巨噬细胞24 h后HMGB1蛋白的表达亦增加(均P<0.05);与PGE2+LPS联合处理比较,应用RNAi技术可下调小鼠腹腔巨噬细胞EP4受体表达,再给予外源性PGE2加LPS处理,HMGB1水平下降(P<0.05);使用DN.CREB质粒抑制c AMP结合元件结合蛋白(c AMP respondse element bound protein,CREB)后再加上EP4受体激动剂联合LPS处理,与EP4受体激动剂+LPS处理比较,HMGB1水平差异无统计学意义(P>0.05);与单纯应用LPS比较,EP4受体激动剂联合LPS处理小鼠腹腔巨噬细胞可上调表皮生长因子受体(epidermal growth factor receptor,EGFR)及蛋白激酶B(protein kinase B,PKB,亦称Akt)thr308磷酸化水平(均P<0.05);使用EGFR抑制剂预处理细胞后,Akt thr308磷酸化水平及HMGB1表达均降低(均P<0.05);采用Akt特异性抑制剂预处理细胞后,HMGB1表达降低(P<0.05)。结论:PGE2可以通过EP4受体上调LPS诱导的小鼠腹腔巨噬细胞HMGB1的表达,且这一作用与激活EGFR/磷脂酰肌醇3激酶(PI3K)/Akt信号通路有关。

新诊断2型糖尿病患者血清高迁移率族蛋白1和Toll样受体4水平变化及其与胰岛素抵抗的关系研究

目的探讨新诊断2型糖尿病(T2DM)患者血清高迁移率族蛋白1(HMGB1)和Toll样受体4(TLR4)表达水平及其与胰岛素抵抗的关系。方法选取2011年6—11月在我院内分泌科就诊的新诊断T2DM患者60例(T2DM组)及同期在我院体检的健康者30例(NC组)。观察两组受检者的空腹血清HMGB1和TLR4水平,并计算其胰岛素抵抗指数(HOMA-IR)、胰岛β细胞功能指数(HOMA-β)、胰岛素敏感性指数(ISI),并将HMGB1、TLR4与其他观察指标(如血糖、血脂、肝功能、胰岛素)进行单因素相关分析,然后进行多元逐步回归分析。结果(1)T2DM组患者血清HMGB1水平为(89.8±13.4)μg/L,TLR4水平为(35.8±3.1)μg/L,NC组分别为(58.8±10.4)μg/L和(21.6±3.6)μg/L,两组HMGB1、TLR4水平比较差异均有统计学意义(t=11.059,19.373;P<0.01)。(2)单因素相关分析显示,血清HMGB1及TLR4均与体质指数(BMI)、三酰甘油(TG)、空腹血糖(FPG)、胰岛素抵抗指数(HOMA-IR)呈正相关,而与HOMA-β、ISI呈负相关,且HMGB1与TLR4亦呈正相关(P均<0.05)。(3)多元逐步回归分析发现,TLR4为影响HMGB1的最显著因素(β=0.281,P=0.000);HMGB1、HOMA-β、ISI为影响TLR4最显著的因素(β值分别为-4.146、-4.325和-5.061;P<0.05)。结论 T2DM患者血清HMGB1、TLR4均显著升高;且血清HMGB1与TLR4呈正相关;血清HMGB1与TLR4水平升高可能参与了胰岛素抵抗及T2DM的发生、发展。

卵巢上皮性癌YAP1和SOX2的表达及其临床意义

目的:检测卵巢上皮性癌中Yes相关蛋白1(Yes-associated protein1,YAP1)和Y染色体性别决定区相关的高迁移率族盒蛋白2(sex-determing region of Y chromosome related high mobility group box 2,SOX2)的表达及意义。方法:选取86例卵巢上皮性癌组织和20例行卵巢活检的正常卵巢组织(少数局灶伴良性上皮性囊肿),采用免疫组织化学法检测YAP1和SOX2的表达;记录并统计86例卵巢癌患者的生存时间、肿瘤相关的临床病理参数,分析其与YAP1高表达的关系;建立SOX2稳定转染的卵巢上皮癌细胞株,免疫印迹法检测SOX2及YAP1的表达,划痕实验检测SOX2过表达细胞的迁移能力。结果:卵巢上皮性癌中YAP1和SOX2的高表达率明显高于正常卵巢组织(P均<0.01);YAP1高表达患者生存时间明显短于低表达者(P<0.01);卵巢上皮性癌组织中YAP1高表达与年龄无关,但与化疗耐药、肿瘤转移、FIGO分期、组织分化相关。YAP1和SOX2表达与卵巢上皮性癌相关,且两者表达呈正相关(r=0.635,P<0.01)。与对照组比较,SOX2过表达细胞中YAP1蛋白表达明显增加(t=6.304,P<0.01),细胞迁移能力明显提高(t=11.77,P<0.01)。结论:YAP1和SOX2在卵巢上皮性癌中呈高表达,且与卵巢上皮性癌发生发展相关。