β-连环蛋白与卵巢癌转移的相关性及调节机制

目的 探讨β-连环蛋白(β-catenin)与卵巢癌转移的相关性及其可能的调节机制. 方法 采用免疫荧光法、免疫印迹法及反转录-聚合酶链反应(RT-PCR)检测人卵巢浆液性囊腺癌高、低转移细胞系(HO-8910PM及HO-8910)中β-catenin基因的表达差异.为进一步探讨β-catenin调节卵巢癌转移的可能机制,将低转移卵巢癌HO-8910细胞经肝细胞生长因子(HGF)处理,应用上述方法检测β-catenin基因表达的改变;同时经Transwell小室法和划痕实验检测细胞侵袭、迁移能力的改变. 结果 β-catenin在HO-8910细胞中为高表达,在HO-8910PM细胞中为低表达,差异具有显著性(P<0.05).HGF可明显降低HO-8910细胞中β-catenin基因的表达(P<0.05)并促进细胞侵袭(P<0.05)和迁移. 结论 卵巢癌细胞的侵袭、迁移等恶性行为可能与β-catenin基因表达下降有关.经由HGF/c-Met的信号很可能是β-catenin介导的黏附功能的重要调节者.     

高转移人卵巢癌细胞系HO-8910PM的建立及其生物学特性

我们在建立高转移人卵巢癌裸鼠皮下移植瘤模型的基础上［１］，又建立高转移人卵巢癌体外细胞系ＨＯ８９１０ＰＭ。现将建系过程及其生物学特性介绍如下。一、材料和方法１材料来源：取高转移人卵巢癌皮下移植瘤模型第７代３７０号裸鼠皮下肿块，在无菌条件下剪碎至１...     

转化生长因子β1信号通路在糖皮质激素抑制人卵巢癌细胞HO-8910增殖中的作用

目的 :转化生长因子 β1(transforminggrowthfactorβ1,TGFβ1)是一个分泌型的多肽 ,它对多种类型的肿瘤细胞均具有增殖抑制作用。TGFβ1受体分为I、II型 (TβRI、TβRII) ,属丝 /苏氨酸激酶受体家族。能与TGFβ1结合的是TβRII,TβRII与配体结合后与TβRI形成寡聚体 ,并使后者磷酸化 ,激活的TβRI通过激活的Smad蛋白 ,调节靶基因的转录 ,抑制细胞增殖。本教研室既往的研究表明 ,糖皮质激素能够抑制人卵巢癌细胞系HO - 8910细胞的增殖 ,但其机制尚不明确。本研究拟探讨TGFβ1信号通路在糖皮质激素抑制人卵巢癌细胞系HO - 8…     

氧化苦参碱对人卵巢癌HO-8910细胞的作用及其机制的初步研究

目的探讨氧化苦参碱（Oxymatrine,OM）在体外诱导人卵巢癌HO-8910细胞的凋亡作用及其可能的机制。方法以不同浓度的OM作用于人卵巢癌HO-8910细胞,应用MTT法检测细胞生长情况;荧光染色观察细胞核的形态学改变;琼脂糖凝胶电泳分析细胞DNA变化;Western blot检测细胞Bcl-2和Bax的蛋白表达情况。结果 OM能显著抑制HO-8910细胞的增殖作用,具有浓度和时间依赖性。荧光染色后可观察到典型的凋亡小体。细胞DNA电泳后呈现出凋亡细胞典型的DNA ladder。Bcl-2蛋白表达量逐渐降低,Bax蛋白表达量逐渐增加,呈明显的浓度依赖性。结论 OM能诱导体外培养的人肝癌HO-8910细胞凋亡。Bcl-2表达增多,Bax表达减少是OM诱导HO-8910细胞凋亡的可能机制之一。     

地塞米松促进卵巢癌细胞系p21/WAF1和TβR-Ⅱ的表达

探讨人工合成糖皮质激素地塞米松(Dex)抑制人卵巢癌细胞系HO-8910增殖的分子机理.采用RT-PCR测定细胞周期蛋白依赖性激酶抑制剂p21/WAF1,转化生长因子-β1(TGF-β1)及其I型受体(TβR-Ⅰ)和Ⅱ型受体(TβR-Ⅱ)的mRNA表达水平,免疫细胞化学方法分析TβR-Ⅱ的蛋白表达水平.发现Dex能够诱导HO-8910细胞p21/WAF1 mRNA的表达,10-7mol/L Dex处理细胞8 h组比对照组p21/WAF1 mRNA表达增加2.84倍(P＜0.01).并且证明Dex亦可上调TβR-Ⅱ mRNA的表达水平,10-7mol/L Dex处理8 h使TβR-Ⅱ mRNA的表达量比对照升高1.2倍(P＜0.01);Dex也引起TβR-Ⅱ蛋白表达的增加.这些效应均可被糖皮质激素受体(GR)拮抗剂RU486逆转.而TGF-β1和TβR-ⅠmRNA的表达不受Dex的影响.以上结果提示,Dex通过GR介导而促进p21/WAF1和TβR-Ⅱ的表达,可能参与其抑制人卵巢癌细胞HO-8910增殖过程.     

地塞米松促进卵巢癌细胞系p21/WAF1和TβR-II的表达

探讨人工合成糖皮质激素地塞米松 (Dex)抑制人卵巢癌细胞系HO 8910增殖的分子机理。采用RT PCR测定细胞周期蛋白依赖性激酶抑制剂p2 1 WAF1,转化生长因子 β1 (TGF β1 )及其I型受体 (TβR I)和II型受体 (TβR II)的mRNA表达水平 ,免疫细胞化学方法分析TβR II的蛋白表达水平。发现Dex能够诱导HO 8910细胞p2 1 WAF1mR NA的表达 ,10 - 7mol LDex处理细胞 8h组比对照组p2 1 WAF1mRNA表达增加 2 84倍 (P <0 0 1)。并且证明Dex亦可上调TβR IImRNA的表达水平 ,10 - 7mol LDex处理 8h使TβR IImRNA的表达量比对照升高 1 2倍 (P <0 0 1) ;Dex也引起TβR II蛋白表达的增加。这些效应均可被糖皮质激素受体 (GR)拮抗剂RU4 86逆转。而TGF β1 和TβR ImRNA的表达不受Dex的影响。以上结果提示 ,Dex通过GR介导而促进p2 1 WAF1和TβR II的表达 ,可能参与其抑制人卵巢癌细胞HO 8910增殖过程     

miR-141-3p对卵巢癌的作用及其机制

目的:探究miR-141-3p在卵巢癌中的作用及其相关的分子机制.方法:实时荧光定量PCR检测30例卵巢囊肿和30例卵巢癌组织中miR-141-3p和表皮生长因子受体(EGFR)的表达水平.将SKOV3细胞分为NC组(无转染的SKOV3细胞),miR-141-3p组(SKOV3细胞转染miR-141-3p),LV-EG...     

乳腺癌中Caveolin-1蛋白的表达及临床意义

目的 观察窖蛋白(Caveolin-1,Cav-1)在正常乳腺组织、良性增生性乳腺病及乳腺癌中的表达及其与乳腺癌临床病理特征的关系.方法 采用免疫组化SP法检测10例正常乳腺组织、15例良性增生性乳腺病和48例乳腺癌组织中Cav-1的表达,分析Cav-1在不同乳腺病变中表达的差异.结果 Cav-1在正常乳腺组织、良性增生性乳腺病及乳腺癌中的阳性率分别为40%、46%和100%,平均表达强度积分(±s)分别为0.90±0.83、0.93±1.16和6.24±2.18.其在正常乳腺组织与良性增生性乳腺病之间表达差异无统计学意义(P>0.05);在正常乳腺组织、良性增生性乳腺病与乳腺癌之间表达差异具有显著性(P<0.01);在乳腺癌不同组织学类型、病理分级、临床分期、细胞增殖指数(MIB-1/Ki-67)和有无淋巴结转移中阳性率和表达强度差异均无显著性(P>0.05).结论 Cav-1在正常乳腺组织及良性增生性乳腺病中呈低表达,在乳腺癌中呈高表达,表明Cav-1与乳腺癌的发生密切相关,但与乳腺癌的组织学类型、病理分级、临床分期、有无淋巴结转移和细胞增殖指数(MIB-1/Ki-67)无明显相关性.     

肿瘤转移相关蛋白1的表达与人直肠癌组织淋巴管转移及预后关系

目的观察肿瘤转移相关蛋白1(MTA1)与直肠癌淋巴结转移及预后的关系,探讨MTA1在直肠癌发生发展过程中的作用。方法应用免疫组化法和实时荧光定量PCR方法检测MTA1在45例人直肠癌和20例直肠息肉组织中的表达,并结合临床病理特征和生存资料进行相关分析。结果直肠癌组织中MTA1蛋白及mRNA的表达较直肠息肉组显著增加(P<0.05),MTA1表达与直肠癌淋巴结转移、Dukes临床分期密切相关(P<0.05)。MTA1阳性表达与生存率负相关(P<0.05)。结论 MTA1高表达促进直肠癌淋巴结转移,检测MTA1表达可成为直肠癌预后不良的重要指标。     

Caveolin-1在TGF-β1诱导人支气管上皮细胞间质化中的作用

目的:探究小窝蛋白1(caveolin-1)在转化生长因子β1(TGF-β1)诱导的人支气管上皮细胞(16HBE细胞)上皮-间充质转化(EMT)中的作用。方法:以16HBE细胞株为研究对象,免疫荧光、RT-q PCR和Western blot实验检测16HBE细胞EMT过程中caveolin-1的mRNA和蛋白表达;Western blot检测siRNA干扰caveolin-1对16HBE细胞EMT的影响。结果:Caveolin-1广泛存在于16HBE细胞膜上,TGF-β1刺激后,caveolin-1的mRNA和蛋白表达减少(P0.05)。与TGF-β1组比较,caveolin-1 siRNA和TGF-β1共同作用促进了细胞形态的转化,抑制了E-钙黏蛋白的蛋白表达而促进了α-SMA的蛋白表达(P0.05)。TGF-β1刺激16HBE细胞后,AKT和Smad3在30 min磷酸化水平最高,与0 min对照组比较显著增加(P0.05);用siRNA干扰caveolin-1基因后再用TGF-β1刺激16HBE细胞30 min,下游信号蛋白分子AKT和Smad3的磷酸化水平增高,与TGF-β1组比较显著增加(P0.05)。结论:TGF-β1能下调16HBE细胞的caveolin-1表达水平;caveolin-1可能参与了TGF-β1诱导的16HBE细胞EMT过程中TGF-β1/Smad通路和PI3K-AKT通路的活化。     

人卵巢浆液性囊腺癌高低转移细胞株E-cadherin差异表达

目的探讨上皮性细胞粘附分子(E-cadherin,E-cad)在人卵巢浆液性囊腺癌高低转移细胞株HO-8910PM及HO-8910中的表达及生物学行为的差异。方法采用免疫荧光法及W estern b lotting检测细胞中E-cad蛋白表达的差异,用粘附实验检测细胞与细胞外基质的粘附能力,用Transwell小室法检测细胞侵袭能力及迁移能力。结果E-cad在HO-8910细胞中为高表达,在HO-8910PM细胞中为低表达,HO-8910PM细胞与细胞外基质的粘附能力、侵袭能力及迁移能力均明显高于HO-8910细胞(P<0.05,P<0.01)。结论人卵巢浆液性曩腺癌HO-8910PM细胞的浸润、转移等恶性行为可能与E-cad的表达下降有关。     

非小细胞肺癌患者胸腔积液中EGFR基因突变的检测及临床意义

目的:探讨采用非小细胞肺癌患者胸腔积液标本肿瘤细胞进行表皮生长因子(epidermal growth factor receptor,EGFR)基因突变检测的可行性及其临床意义.方法:采用Sanger测序法检测17例非小细胞肺癌患者胸腔积液及对应的17例手术或肺部穿刺组织标本EGFR基因18～21外显子基因突变,并进行统计分析.结果:胸腔积液标本17例共检出5例突变,检出率29.41％.手术或穿刺组织标本17例共检出7例突变,检出率41.18％.胸腔积液标本EGFR基因突变检出率略低于手术或穿刺组织标本.结论:采用Sanger测序法进行非小细胞肺癌患者胸腔积液中EGFR基因突变的检测,方法可行,尤其适用于无法获取手术或肺部穿刺组织标本的患者.     

Mechanism and management of ovulatory failure in women with polycystic ovary syndrome

Polycystic ovary syndrome is the most important cause of chronicanovulation. In women who fail to respond to clomiphene, low-doseFSH given in a step-wise fashion can induce normal folliculargrowth and ovulation. The failure of the action of endogenousFSH in these women may be related to reduced biological activityof circulating FSH, but may also involve inhibition of its actionat follicular level by polypeptide growth factors such as EGF.     

Effect of an angiotensin II type 1 receptor blocker on caveolin-1 expression in prostate cancer cells

Introduction

Caveolin-1, the major structural protein of caveolae, interacts directly with the AT1 receptor. The biological functions of caveolin-1 in cancer are compound, multifaceted, and depend on cell type, tumour grade and cancer stage. The AT1-R-caveolin complex in caveolae may coordinate angiotensin II (Ang II) induced signalling. The aim of this study was to determine the effect of the angiotensin II receptor type 1 blocker candesartan on caveolin expression in human metastatic prostate adenocarcinoma cells PC-3.

Material and methods

WST-1 and BrdU assays were used as indicators of cell viability and proliferation after angiotensin II and/or candesartan stimulation. Real-time RT–PCR and western blot were used to study the effect of Ang II and/or candesartan on the expression of Cav-1 and AT1-R in PC-3 cells

Results

We found that the expression of caveolin-1 mRNA in the PC-3 cells treated with CV was significantly decreased in comparison with the control (2.9 ±0.17, 4.7 ±0.6, p < 0.05), whereas a higher caveolin-1 mRNA expression was observed in those after Ang II treatment (6.0 ±0.43, 4.7 ±0.6, p < 0.05). Protein analysis indicate that the expression of caveolin-1 protein in the PC-3 cells treated with candesartan was significantly decreased when compared with the control (0.69 ±0.05, 1.6 ±0.12, p < 0.05), whereas higher caveolin-1 protein expression was observed after Ang II treatment (2.5 ±0.20, 1.6 ±0.12, p < 0.05).

Conclusions

These results provide new information on the action of candesartan and may improve the knowledge about AT1 receptor inhibitors, which can be potentially useful in prostate cancer therapy.     

乳腺癌易感基因1的表达与乳腺癌亚型关系的实验研究

目的 探讨乳腺癌易基因1（BRCA1）的表达与乳腺癌分子亚型的关系.方法 运用免疫组化法检测乳腺癌组织中BRCA1的表达与雌二醇受体（ER）和人表皮生长因子受体-2（Her-2）的关系,分析BRCA1的表达在乳腺癌各亚型中的意义.结果 ER阳性和阴性时,BRCA1的阳性率分别为40.2％和60.0％,表明BRCA1表达与ER有关（P＜0.05）,与Her-2无关（P＞0.05）.在乳腺癌各分子亚型中,Luminal B型和Her-2过表达型的BRCA1阳性率（66.7％和70.0％）与Basal-like型的BRCA1阳性率（75.0％）差别无统计学意义（P＞0.05）;Luminal A型的BRCA1阳性率（23.7％）低于Basal-like型的BRCA1阳性率（75.0％）,差别具有统计学意义（P＜0.05）.结论 BRCA1在Luminal B型、Her-2过表达型和Basal-like型乳腺癌中表达较高,在Luminal A型乳腺癌中表达不高,提示BRCA1可成为治疗乳腺癌的重要靶点.     

Prognostic and predictive values of SPP1, PAI and caveolin-1 in patients with oral squamous cell carcinoma

SPP1, PAI and caveolin-1 are known to be closely associated with tumor progression in several kinds of human tumors. This study aimed to investigate the expression of SPP1, PAI and caveolin-1 in oral squamous cell carcinoma (OSCC), and to evaluate their association with the prognosis in oral carcinoma. Immunohistochemical staining was used to examine the expression of SPP1, PAI and caveolin-1 in 17 normal oral mucosa, 6 oral epithelial dysplasia and 43 OSCC specimens by tissue microarrays. High expression of SPP1, PAI and caveolin-1 was found in OSCC patients, and SPP1 and PAI expression were significantly higher in OSCC than in normal oral mucosa. No significant correlations were found between SPP1, PAI and caveolin-1 expression and clinicopathological factors. Expression of SPP1, PAI and caveolin-1 was also not associated with overall survival. Moreover, SPP1 was closely correlated with PAI, caveolin-1 and Keap1, and PAI had significant correlations with caveolin-1, Keap1 and Nrf2, and caveolin-1 was associated with Keap1 by using the Pearson correlation coefficient test. Our findings suggest that overexpressed SPP1, PAI and caveolin-1 were linked to carcinogenesis and progression, and thus they may serve as potential prognostic factors in OSCC.     

Epidermal growth factor and insulin short‐term increase hPepT1‐mediated glycylsarcosine uptake in Caco‐2 cells

Aims: Little is known about the physiological regulation of the human intestinal di/tri‐peptide transporter, hPepT1. In the present study we evaluated the effects of epidermal growth factor (EGF) and insulin on hPepT1‐mediated dipeptide uptake in the intestinal cell line Caco‐2. Methods: Caco‐2 cells were grown on filters for 23–27 days. Apical dipeptide uptake was measured using [¹⁴C]glycylsarcosine([¹⁴C]Gly‐Sar). HPepT1 mRNA levels were investigated using RT‐PCR, cytosolic pH was determined using the pH‐sensitive fluorescent probe BCECF. Results: Basolateral application of EGF increased [¹⁴C]Gly‐Sar uptake with an ED₅₀ value of 0.77 ± 0.25 ng mL^?1 (n = 3?6) and a maximal stimulation of 33 ± 2% (n = 3?6). Insulin stimulated [¹⁴C]Gly‐Sar uptake with an ED₅₀ value of 3.5 ± 2.0 ng mL^?1 (n = 3?6) and a maximal stimulation of approximately 18% (n = 3?6). Gly‐Sar uptake followed simple Michaelis‐Menten kinetics. K_m in control cells was 0.98 ± 0.11 mM (n = 8) and V_max was 1.86 ± 0.07 nmol cm^?2 min^?1 (n = 8). In monolayers treated with 200 ng mL^?1 of EGF, K_m was 1.11 ± 0.05 mM (n = 5) and V_max was 2.79 ± 0.05 nmol cm^?2 min^?1 (n = 5). In monolayers treated with 50 ng mL^?1 insulin, K_m was 1.03 ± 0.08 mM and V_max was 2.19 ± 0.06 nmol cm^?2 min^?1 (n = 5). Kinetic data thus indicates an increase in the number of active transporters, following stimulation. The incrased Gly‐Sar uptake was not accompanied by changes in hPepT1 mRNA, nor by measurable changes in cytosolic pH. Conclusions: Short‐term stimulation with EGF and insulin caused an increase in hPepT1‐mediated uptake of Gly‐Sar in Caco‐2 cell monolayers, which could not be accounted for by changes in hPepT1 mRNA or proton‐motive driving force.     

HLA-G在卵巢癌组织中的表达及对NK细胞杀伤活性的影响

通过免疫组织化学法(IHC)检测卵巢癌组织中HLA-G的表达;克隆编码全长HLA-G1cDNA,将该基因通过真核表达载体pVITRO2-mcs转染至HLA-G阴性的HO-8910、OVCAR-3细胞株,采用RT-PCR、流式细胞术(FACS)及Westernblot鉴定、分析转染细胞中HLA-G的mRNA及蛋白质表达;LDH释放法检测卵巢癌细胞表达HLA-G后对NK细胞杀伤功能的影响。结果显示,66.7%(22/33)卵巢癌组织表达HLA-G分子,而正常组织未见其表达;基因转染后,HLA-G在HO-8910及OVCAR-3细胞表面获得稳定表达。细胞毒实验结果显示,HLA-G能显著抑制NK-92对卵巢癌细胞的杀伤活性(P<0.01);HLA-G特异性抗体87G阻断后,能明显恢复NK-92细胞对HO-8910-G、OVCAR-3-G的杀伤功能。本研究结果提示,卵巢癌细胞通过表达HLA-G分子抑制NK细胞的杀伤活性,在卵巢癌细胞逃避宿主的免疫监视中起重要作用。     

Serum levels of insulin-like growth factor-1, IGF binding protein-1 and insulin and the response to human menopausal gonadotrophins in women with polycystic ovary syndrome

In order to determine which factors influence the large variationsin sensitivity to gonadotrophins witnessed in women with polycysticovary syndrome (PCOS), a prospective study was conducted ofthe correlation between basal clinical and endocrinologicalfeatures and gonadotrophin requirements of 20 women with clomiphene-resistantPCOS undergoing ovulation induction. Baseline evaluation ofserum concentrations of luteinizing hormone (LH), follicle stimulatinghormone (FSH), testosterone, fasting insulin, insulin-like growthfactor-1 (IGF-1), IGF binding protein-1 (IGFBP-1) and sex hormone-bindingglobulin (SHBG) were performed before administering gonadotrophin-releasinghormone agonist (GnRHa). Two weeks later, human menopausal gonadotrophin(HMG) was given in a standard individualized protocol accordingto ovarian response, until human chorionic gonadotrophin (HCG)was given. Serum concentrations of insulin, IGF-1, and IGFBP-1were unaffected by GnRHa. The BMI correlated positively withinsulin and inversely with IGFBP-1 serum concentrations andinsulin and IGFBP-1 were inversely correlated. The amount ofHMG required correlated positively with BMI and insulin concentrationsand inversely with IGFBP-1 in the whole group and these correlationswere maintained in the sub-group of lean women. No correlationwas observed between HMG requirements and IGF-1 or other hormones.Womenwith hyperinsulinaemia and low IGFBP-1 concentrations requiredsignificantly more HMG. Multiple regression analysis revealedthat insulin concentration is the most significant determinantof HMG requirement even when dissociated from BMI. We concludedthat requirement of HMG in PCOS is not merely determined byobesity but by a cardinal role of insulin concentrations which,when high, induce, hypothetically, a hyperandrogenic intrafollicularmilieu.