HSP70多肽复合物修饰DCs疫苗抗胰腺癌荷瘤小鼠的实验研究

目的:探讨负载热休克蛋白70多肽复合物(HSP70-PC)抗原的树突状细胞(DCs)疫苗对胰腺癌荷瘤小鼠的免疫治疗作用.方法:采用低渗裂解、ConA-Sepharose亲和层析及ADP-Agarose亲和层析法,从小鼠胰腺癌(MPC83)肿瘤组织中纯化HSP70-PC,修饰小鼠骨髓来源的树突状细胞(DCs),制备树突状细胞HSP70多肽肿瘤疫苗,用MTT法检测混合淋巴细胞反应(MLR)中HSP70-PC致敏DC对CTL的增殖及活化效果;建立MPC83荷瘤小鼠模型,观察树突状细胞HSP70多肽肿瘤疫苗对荷瘤小鼠治疗的效果和小鼠存活期.结果:经上述方法分离、纯化获得了较高纯度的HSP70-PC蛋白质;HSP70-PC在1.5～2.0 μg/ml范围内可达到刺激树突状细胞最强效果,用HSP70-PC修饰的DCs能增强T细胞的增殖和活化能力;应用树突状细胞HSP70多肽肿瘤疫苗治疗荷瘤小鼠能显著抑制荷瘤小鼠肿瘤的生长,延长荷瘤小鼠存活期.结论:采用低压亲和层析柱可从胰腺癌瘤块中获得较高纯度HSP70多肽复合物,其修饰的DCs疫苗用于荷瘤小鼠免疫治疗有显著疗效,为临床胰腺癌生物免疫治疗奠定基础.     

Hsp70-肿瘤抗原肽复合物修饰的DC疫苗体内外特异性抗瘤作用

目的 :探讨树突状细胞 (DC)及Hsp70 肿瘤抗原肽复合物修饰后的体内外特异性抗瘤作用。方法 :采用一定的生化技术 ,从H2 2肝癌细胞中提取肿瘤抗原肽 ,并在体外与Hsp70进行结合 ;采用细胞培养技术 ,培养rmGM CSF、rmIL 4诱导的小鼠骨髓细胞 ,体外获取大量的DC ,后者经Hsp70 肿瘤抗原肽复合物修饰后刺激小鼠脾淋巴细胞 ,通过MTT法进行检测淋巴细胞的激活 ;收集上述刺激传代培养的淋巴细胞 ,检测其对H2 2瘤细胞和艾氏腹水癌细胞的杀伤功能 ;采用H2 2瘤细胞肌肉接种和H2 2瘤细胞、艾氏腹水癌细胞腹腔接种 ,对接种小鼠给予经体外修饰的DC回输 ,观察其抑制肿瘤的效果。结果 :Hsp70 肿瘤抗原肽复合物可使DC成熟 ,大量分泌IL 12、TNF α、IL 1β等细胞因子 ,并能够使DC激活小鼠脾淋巴细胞 ;激活后传代培养的淋巴细胞对H2 2瘤细胞能够特异性地杀伤 ,而对艾氏腹水癌细胞无效 ;经Hsp 70 肿瘤抗原肽复合物修饰后的DC可作为一种有效的瘤苗 ,体内能特异性地抑制小鼠H2 2肿瘤生长。结论 :Hsp70 肿瘤抗原肽复合物能够很好地修饰体外诱导获取的DC ,使后者成为一种有效的瘤苗 ,体外能够特异性地激活淋巴细胞 ,体内有效地抑制肿瘤生长     

负载不同形式肝癌抗原的树突状细胞抑瘤功能的比较

目的：探讨不同形式的肝癌抗原修饰的树突状细胞(DC)的抑瘤功能。方法：分别用肝癌H22冻融抗原、H22小分子抗原肽和Hsp70-H22抗原肽复合物修饰DC；用MTT比色法分析DC激活的CTL对H22细胞的杀伤能力，并用RT-PCR法测定脾脏T细胞中IFN-γ mRNA的表达水平；用不同修饰的DC免疫小鼠，观察其对H22肝癌的生长抑制作用。结果：单独的H22肝癌抗原肽修饰的DC不能激活CTL。Hsp70-H22肽复合物修饰的DC激活CTL的能力强于H22肝癌冻融抗原修饰的DC，对H22细胞的杀伤率分别为47．3％和18．3％。各组T细胞中IFN-γ表达水平的变化与杀伤率的变化相一致。用H22肝癌冻融抗原和Hsp70-H22肽复合物修饰的DC免疫小鼠后，均可抑制H22细胞生长，但后者的抑制作用更强，成瘤率仅40％。其他各组的成瘤率均为100％。结论：Hsp70-H22肽复合物是一种DC的强致敏物．可通过激活CTL、诱导CD4^ T细胞分化成Th1型细胞而参与肝癌的免疫排斥。     

体外构建的HSP70-肝癌抗原肽诱导抗原肽特异性免疫反应1

目的研究体外构建的HSP70-肝癌抗原肽复合物诱导针对肝癌的特异性免疫反应能力,为该复合物的临床应用奠定基础.方法在体外构建HSP70-肝癌抗原肽复合物,联合应用粒/巨细胞集落刺激因子(GM-CSF)及白介素-4(IL-4)直接从志愿者外周血中培养出DC;以HSP70、HSP70-肝癌抗原肽、抗原肽分别刺激DC,DC激活同源的T淋巴细胞产生细胞毒性T淋巴细胞(CTL),检测其杀伤T2细胞和肝癌细胞系的能力.结果HSP70-抗原肽、抗原肽均可诱导CD8+的抗原肽特异性CTL,而前者的诱导效果更强.结论体外构建的HSP70-抗原肽复合物具有免疫原性,HSP70可以增强抗原肽诱导特异性免疫反应的能力,HSP70-抗原肽复合物有可能作为肽疫苗用于临床肿瘤免疫治疗.     

体外构建的HSP70—肝癌抗原肽诱导抗原肽特异性免疫反应

目的：研究体外构建的HSP70-肝癌抗原肽复合物诱导针对肝癌的特异性免疫反应能力,为该复合物的临床应用奠定基础。方法：在体外构建HSP70-肝癌抗原肽复合物,联合应用粒/巨细胞集落刺激因子（GM-CSF）及白介素-4-（IL-4）直接从志愿者外周血中培养出DC;以HSP70、HSP70-肝癌抗原肽、抗原肽分别刺激DC,DC激活同源的T淋巴细胞产生细胞毒性T淋巴细胞（CTL）,检测其杀伤72细胞和肝癌细胞系的能力。结果：HSP70-肝癌抗、抗原肽均可诱导CD8^ 的抗原肽特异性CTL,而前者的诱导效果更强。结论：体外构建的HSP70-抗原肽复合物具有免疫原性,HSP70可以增强抗原肽诱导特异性免疫反应的能力,HSP70-抗原肽复合物有可能作为肽疫苗用于临床肿瘤免疫治疗。     

HSP70-肿瘤肽的纯化及其抗肿瘤免疫效应

目的：探讨热休克蛋白70（HSP70）－肿瘤肽分离、纯化方法，并观察其抗肿瘤免疫保护效应。方法：采用低渗均浆、超速离心、ConA-Sepharose亲和层析，ADP－Agarose亲和层析和DEAE离子交换的亲和层析，从热处理的小鼠肝癌（HCaF)细胞中分离、纯化HSP70－肿瘤肽，并通过主动免疫保护试验观察其抗肿瘤免疫效应。结果：蛋白得率为每g湿重瘤细胞可纯化50－100μg HSP70－肿瘤肽；经SDS－PAGE和Western blot检测，纯化的HSP70－肿瘤肽具有很高的纯度和特异性；HSP70－肿瘤肽主动免疫的小鼠可抵抗HCaF细胞的攻击，存活率达75％，结论：采用低压亲和层析可获得高纯度的HSP70－肿瘤肽，且可诱导明显的抗肿瘤免疫保护效应。     

异种HSP70及固相抗体法体外扩增人γδT细胞的比较

目的：比较异种HSP70及固相抗体法体外扩增人γδT细胞。方法：分别用固相抗体法、HSP70BCG及小鼠榄香烯复合瘤苗热休克蛋白70-肽复合物(HSP70m-肽复合物)体外扩增人PBMCs的γδT细胞，流式细胞仪检测细胞表型及亚型分布，RT-PCR检测各亚型基因的取用。MTT检测γδT细胞的细胞毒作用。结果：培养14天后，抗体法扩增的PMBCs总数增加30～40倍，御细胞的比率由3.2％提高到86．7％；HSP70BCG组PBMCs总数增加7—8倍，7趼细胞的比率由3．2％提高到71．32％；HSP70m-肽复合物组PBMCs总数增加4～5倍，γδT细胞的比率由3.2％提高到27.26％。抗体及HSP70BCG扩增的γδT细胞以表达Vγ9/Vδ2亚型为主，且Vδ1、Vδ2、Vδ3三种基因均得到取用，对Daudi具较高的杀伤活性。结论：抗体及HSP70BCG均能扩增出高纯度的γδT细胞，且具有完整的受体库，扩增的γδT细胞具细胞毒活性。     

树突状细胞激活的肿瘤浸润淋巴细胞抗小鼠乳腺癌研究

目的 探讨C127细胞全细胞性抗原致敏的DC激活的TIL体外抗小鼠乳腺癌活性,并将C127细胞全细胞性抗原致敏的DC激活的TIL(C127-DC-TIL)过继免疫荷瘤小鼠,研究其对C127荷瘤小鼠免疫功能的影响及抑瘤作用.方法 从小鼠四肢长骨骨髓中获取DC,应用粒,巨噬细胞集落刺激因子(GM-CSF)、白介素-4(IL-4)和肿瘤全细胞性抗原致敏DC,然后用DC激活TIL,观察TIL在体外对C127细胞、MA782细胞和B16细胞的杀伤活性;检测应用C127-DC-TIL后荷瘤小鼠的脾淋巴细胞的NK、LAK、CTL活性、血清TNF活性、抑瘤作用以及瘤体病理改变,并与对照组相比较.结果 ①C127-DC-TIL具有很强的对C127细胞杀伤活性[杀伤率为(70.21±2.86)%],明显高于其对MA782和B16细胞的杀伤活性[杀伤率分别为(51.31±3.25)%,(31.41±2.65)%],也明显高于未经DC激活的TIL、C127-DC-脾淋巴细胞和未经DC激活的脾淋巴细胞对C127细胞杀伤活性[杀伤率分别为(48.30±2.97)%,(47.76±3.43)%和(17.23±2.56)%]和对MA782细胞杀伤活性[杀伤率分别为(38.52±2.87)%,(36.62±2.75)%和(18.07±2.40)%]以及对B16细胞杀伤活性[杀伤率分别为(25.38±2.63)%,(24.82±2.81)%和(17.34±2.81)%],同时B16细胞全细胞性抗原致敏的DC激活的TIL(B16-DC-TIL,TIL来源于C127瘤体)也可诱导相对较低的对B16细胞的特异性细胞杀伤活性.②C127-DC-TIL可明显诱导提高荷瘤小鼠脾淋巴细胞NK、LAK和CTL活性[活性分别为(32.21±1.24)%、(30.35±1.72)%和(37.43±1.54)%],并可检测到血清TNF水平明显上升[血清TNF水平为(38.41±1.77)U/ml],它们均达正常对照组水平,与未经DC激活的TIL组、C127-DC-脾淋巴细胞组、未经DC激活的脾淋巴细胞组、生理盐水组分别对应比较,差异均有显著性(P＜0.01).该组瘤体内淋巴细胞浸润程度也高于对照组,其瘤体生长明显受到抑制.结论 ①C127-DC-TIL可产生很强的体外针对C127细胞的特异性杀伤活性.②C127-DC-TIL具有很强的特异性抗小鼠乳腺癌作用.     

HSP_(70BCG)-Daudi肿瘤肽复合物对γδT细胞的活化作用

用固相抗体法[1] 体外扩增人PBMC的γδT细胞 ,静息化处理后用HSP70BCG Daudi肿瘤肽复合物再次刺激研究HSP70BCG Daudi肿瘤肽复合物对γδT细胞的活化作用。MTT检测γδT细胞的细胞毒作用及TNF α生物学活性。3 H TdR掺入法测定细胞增殖。HSP70BCG Daudi肿瘤肽复合物活化的γδT细胞在效靶比为 10∶1及 5∶1时对靶细胞Daudi的杀伤活性分别为 88 36 %±8 2 1%及 75 2 3%± 16 36 % ,与对照组有显著性差别。 2 4及 4 8h增殖结果各组间无差别 ;未检测出各刺激组的上清中TNF α生物学活性有何差别。结论表明HSP70BCG Daudi肿瘤肽复合物活化的γδT细胞对Daudi有较高的杀伤活性     

TLR4在HSP70_(EC-TCV)冲激的DC成熟过程中的作用

观察TLR4在Hca-F榄香烯复合瘤苗来源的HSP70(HSP70EC-TCV)冲激处理的小鼠骨髓来源的DC成熟过程中的作用。用rmGM-CSF和rmIL-4诱导小鼠骨髓来源的DC,以HSP70EC-TCV或加入抗TLR4抗体30 min后再用HSP70EC-TCV冲激处理DC,流式细胞仪检测DC的CD40和CD86表达,ELISA法检测DC上清中IL-12和T细胞上清中IL-2浓度,MTT法检测T细胞对Hca-F细胞的杀伤率。结果表明,抗TLR4抗体对DC的CD40和CD86表达无明显影响,但对HSP70EC-TCV诱导DC分泌IL-12和进而致敏的T细胞分泌IL-2及产生杀瘤功能有明显的抑制作用。TLR4信号通路参与HSP70EC-TCV诱导DC成熟过程。     

Silencing LINC01234 represses pancreatic cancer progression by inhibiting the malignant phenotypes of pancreatic cancer cells

ObjectivePrevious works have outlined the pivotal involvement of long intergenic non-coding RNA (lincRNA) in cancer progression, while the efficiency of LINC01234 in pancreatic cancer remained obscure. The purpose of this research is to unravel the regulatory mechanism of LINC01234 in pancreatic cancer via modulating microRNA (miR)-513a-3p and hexose 6-phosphate dehydrogenase (H6PD).MethodsPancreatic cancer cells were cultured and clinical tissue specimens were collected. LINC01234, miR-513a-3p and H6PD levels in pancreatic cancer cells and tissues were examined. Plasmids altering LINC01234, miR-513a-3p and H6PD expression were transfected into pancreatic cancer cells to assess the change in biological behaviors of pancreatic cancer cells. The targeting relations among LINC01234, miR-513a-3p and H6PD were validated.ResultsLINC01234 and H6PD levels were elevated while miR-513a-3p level was reduced in pancreatic cancer cells and tissues. LINC01234 deficiency hindered the malignant biological activities of pancreatic cancer cells. MiR-513a-3p depletion or H6PD elevation could abrogate the inhibitory effects of LINC01234 silencing on pancreatic cancer cells. LINC01234 sponged miR-513a-3p that targeted H6PD.ConclusionThe reduced LINC01234 exerts inhibitory impacts on pancreatic cancer cells via targeting miR-513a-3p to restrain H6PD level. The current study broadens the understanding of LINC01234 function and affords novel therapeutic targets for pancreatic cancer treatment.     

Role of pancreatic stellate cells in pancreatic cancer metastasis

Pancreatic stellate cells (PSCs) produce the stromal reaction in pancreatic cancer (PC), and their interaction with cancer cells facilitates cancer progression. This study investigated the role of human PSCs (hPSCs) in the metastatic process and tumor angiogenesis using both in vivo (orthotopic model) and in vitro (cultured PSC and PC cells) approaches. A sex mismatch study (injection of male hPSCs plus female PC cells into the pancreas of female mice) was conducted to determine whether hPSCs accompany cancer cells to metastatic sites. Metastatic nodules were examined by fluorescent in situ hybridization for the presence of the Y chromosome. Angiogenesis was assessed by i) immunostaining tumors for CD31, an endothelial cell marker; and ii) quantifying human microvascular endothelial cell (HMEC-1) tube formation in vitro on exposure to conditioned media from hPSCs. Transendothelial migration was assessed in vitro by examining the movement of fluorescently labeled hPSCs through an endothelial cell monolayer. Human PSCs i) were found in multiple metastatic sites in each mouse injected with male hPSCs plus female PC cells; ii) increased CD31 expression in primary tumors from mice injected with MiaPaCa-2 and hPSCs and stimulated tube formation by HMEC-1 in vitro; and iii) exhibited transendothelial migration that was stimulated by cancer cells. Human PSCs accompany cancer cells to metastatic sites, stimulate angiogenesis, and are able to intravasate/extravasate to and from blood vessels.     

大黄素通过上调Smad4蛋白表达抑制胰腺癌细胞

目的以Jab1为作用靶点探讨大黄素抗胰腺癌作用及可能的机制。方法 293T细胞随机分为对照组(不处理)、大黄素组(20μmol/L大黄素处理)、Jab1组(转染HA-Jab1质粒),用免疫印迹实验(Western blot)法测定胰腺癌PANC-1和As PC-1细胞系中Jab1与Smad4的蛋白表达。用免疫共沉淀(IP)测定大黄素组及Jab1对β-Tr CP1与Smad4结合的影响。采用Western blot法检测对照组及大黄素干预组PANC-1和As PC-1细胞系中Smad4蛋白表达。结果胰腺癌PANC-1细胞和As PC-1细胞中Jab1呈高表达,Smad4呈低表达,Jab1与Smad4呈负相关关系(P0.01)。293T细胞中Jab1能促使β-Tr CP1与Smad4结合,促进Smad4蛋白降解;而大黄素通过抑制β-Tr CP1与Smad4结合,增加Smad4的表达水平(均P0.05)。结论与Jab1的作用相反,大黄素可能通过抑制泛素化酶途径,抑制Smad4的降解,从而起到抗肿瘤作用。     

Immunoglobulin E antibodies from pancreatic cancer patients mediate antibody-dependent cell-mediated cytotoxicity against pancreatic cancer cells

In addition to allergy and parasitic infections, immunoglobulin E (IgE) has been shown recently to possess anti-viral and anti-cancer effects. We investigated serum levels of IgE, its low-affinity receptor, soluble CD23 (sCD23) in patients with pancreatic cancer and the effect of IgE against pancreatic cancer cells. Twelve patients were evaluated for pancreatic cancer by imaging and confirmed by biopsy. Fifteen healthy volunteers served as controls. Serum Igs (IgG, IgM, IgA, IgE) and sCD23 levels were determined (enzyme-linked immunosorbent assay, nephelometry) and the presence of cancer-specific IgE was assessed (fluorescence microscopy, Western blot). IgE anti-cancer activity was determined by antibody-dependent cell-mediated cytotoxicity (ADCC). Serum levels of IgE and sCD23 were elevated significantly in patients with pancreatic cancer versus controls, whereas no differences were observed in other Ig isotypes (IgG, IgM, IgA). Flow cytometry and immunofluorescence microscopy demonstrated similar presence of IgG and IgE pancreatic cancer Igs. However, Western blot analysis indicated differences in IgG and IgE antigen-specific antibodies; IgE antibody recognized a 50 kD protein. ADCC studies demonstrated that serum and purified IgE-mediated cytotoxicity against pancreatic cancer cells, effects which were reversed with anti-IgE neutralizing antibody and IgE depletion (immunoaffinity); greater cytotoxicity was observed in patient serum when compared with healthy controls. These data suggest that IgE and sCD23 may serve as useful biomarkers for patients with pancreatic cancer and may be important in the immune response to this disease in that IgE-directed therapy may help to direct treatment.     

Benzyl isothiocyanate sensitizes human pancreatic cancer cells to radiation by inducing apoptosis

Isothiocyanates are a class of naturally occurring chemopreventive agents known to suppress proliferation of cancer cells in culture. The present study was undertaken in order to examine the effects of benzyl isothiocyanate (BITC), one of the common dietary isothiocyanates, on the radiosensitivity of human pancreatic cancer cells and to gain insights into the underlying molecular mechanism of BITC-induced radiosensitization. Two human pancreatic cancer cell lines, PANC-1 and MIAPaCa-2, were treated with BITC and irradiated with X-rays. Radiation sensitivity, apoptosis, and protein levels were determined by a clonogenic assay, fluorescence microscopic analysis with DAPI staining and Western blotting, respectively. MIAPaCa-2 cells were relatively more sensitive to BITC treatment compared with PANC-1 cells. Radiosensitization was observed in both PANC-1 and MIAPaCa-2 cells incubated with BITC at 5 to 10 μM and 2.5 to 5 μM for 24 h, respectively. The combination treatments with BITC and X-rays also revealed an increased percentage of apoptotic cells. In addition, treatment with BITC and X-rays resulted in a decrease in the protein levels of the X-linked inhibitor of apoptosis (XIAP), inhibitor of apoptosis (IAP) family protein, and in a marked increase in the apoptosis protease activating factor-1 (Apaf-1), essential for activation of caspase-9 in stress-induced apoptosis. BITC may be a useful radiosensitizer for radiotherapy of pancreatic cancers.     

吉西他滨抑制胰腺癌细胞糖酵解

目的研究吉西他滨对miR-1208在胰腺癌细胞中表达的影响及其抑制胰腺癌细胞糖酵解的分子机制。方法用real-time PCR法检测胰腺癌组织及细胞系中miR-1208以及糖酵解关键基因的表达;在胰腺癌细胞系Bx PC-3与PANC-1分别转染miR-1208模拟物与阴性对照,利用CCK-8试剂盒、乳酸以及葡萄糖检测试剂盒,研究细胞增殖、乳酸分泌以及葡萄糖利用情况;设计拯救实验研究吉西他滨、miR-1208与胰腺癌细胞代谢的关系。结果 miR-1208在胰腺癌组织中表达下调(60%,12/20)(P0.05);miR-1208过表达明显抑制胰腺癌细胞增殖、乳酸分泌以及葡萄糖的消耗(P0.05),miR-1208导致LDH-A与LDH-D的内源性表达水平下调;经吉西他滨处理的胰腺癌细胞系Bx PC-3与PANC-1,其内源性miR-1208表达水平明显上调(P0.01),而其LDH-A、LDH-D的表达水平明显下调(P0.01)。在胰腺癌细胞中,敲低miR-1208表达抑制吉西他滨诱导的细胞代谢方式转换。LDH-A是miR-1208在胰腺癌细胞中的功能靶基因。结论吉西他滨通过调控miR-1208介导的LDH-A通路发挥抑制胰腺癌细胞糖酵解的功能。     

重组腺病毒介导的野生型p53对人胰腺癌细胞凋亡的诱导作用

目的研究恢复外源性野生型（ｗｔｐ）ｐ５３的表达对人胰腺癌细胞生长和凋亡的作用。方法构建了一个复制缺陷型５型腺病毒ｗｔｐ５３表达载体Ａｄ５ＣＭＶｗｔｐ５３，含有人ＣＭＶ启动子，人野生型ｐ５３和ＳＶ４０ｐｏｌｙＡ信号，经腺病毒包装细胞２９３细胞包装、扩增后，转染胰腺癌ＰＣ２细胞。ＰＣＲ和免疫沉淀技术证实转染的细胞有外源ｗｔｐ５３基因的存在和表达。结果转染的ＰＣ２细胞的生长率和３Ｈ掺入率降低。原位凋亡检测、流式细胞术和ＤＮＡ凝胶电泳显示转染细胞凋亡明显增多。而ＰＣ２细胞和用Ａｄ５ｐＸＪ转染的细胞没有这些改变。结论复制缺陷型腺病毒是一种安全高效的基因转移载体，恢复ｗｔｐ５３的表达可以有效地诱导凋亡和抑制胰腺癌细胞生长     

Establishment of pancreatic cancer stem cells by flow cytometry and their biological characteristics

To investigate the method of separating human pancreatic cancer stem cells by Hoechst 33342 labeled flow cytometry and to analyze the biological properties of pancreatic cancer stem cells. The human pancreatic cancer cell line PC-3 was divided into SP and non-SP cells by flow cytometry. The number of two cell clone spheres and nude mice tumor formation rates were compared by cultivating in serum-free medium; The expression of CD133, Nestin mRNA and protein was analyzed by real-time fluorescence quantitative PCR and Western blot; The expression of two cell drug resistance genes (MDR1, ABCG2, ABCA2 and MRP1) was analyzed by real time fluorescent quantitative PCR. The number of the cloned spheres in SP cells in serum-free medium was significantly higher than that of non-SP cells (P<0.05). The incidence of SP cells in the tumor of immunodeficiency nude mice was significantly higher than that of non-SP cells, and the difference was statistically significant (P<0.05). Real-time fluorescence quantitative PCR analysis showed that the expression of CD133 and Nestin mRNA in SP cells was significantly higher than those of non-SP cells, and the expression of CD133 and Nestin protein in SP cells was also significantly higher than those of non-SP cells (P<0.05). In conclusion, SP side population pancreatic cancer cells by Hoechst 33342 separation have the stem cell characteristics, higher tumor formation rate and higher drug resistance, which may be related to chemotherapy resistance.     

Rab5在胰腺癌细胞BxPC3中对Notch1活性的调节

目的 本研究拟阐明内吞调节蛋白Rab5在Notch活化中的作用.方法 用RNA干扰的方法抑制BxPC3细胞中Rab5蛋白的表达,用Western blot测定Notch1活性型Notch ICD的表达.用Wortmannin和LY294002抑制PI3激酶的活性,观察Notch活性的变化.结果 抑制Rab5蛋白的表达,或者抑制PI3激酶的活性后,Notch1的活性型Notch ICD的表达量均显著下降,同时BxPC3细胞的生长受到抑制.结论 在胰腺癌细胞BxPC3中内吞调节蛋白质Bab5和PI3激酶在Notch活化中起着关键的作用,提示Notch的活化依赖于内吞.