CEA mRNA转染的成熟树突状细胞体外诱导特异性抗肿瘤作用

目的:将体外转录的癌胚抗原(carcinoembryonic antigen,CEA)mRNA转染入成熟树突状细胞(dendritic cell,DCs),观察其体外诱导的特异性抗肿瘤作用。方法:GM-CSF、IL-4、TNF-α体外诱导成熟DCs并用流式鉴定。构建pcDNA3.1-CEA载体,体外转录为CEA mRNA,电穿孔法将CEA mRNA转染入DCs。流式细胞术检测转染后DCs中CEA蛋白的表达;MTT法检测DCs刺激T细胞增殖能力;LDH法检测DCs体外诱导的CTL的特异性抗肿瘤作用;ELISA法检测诱导的CTL上清中IFN-γ的水平。结果:CEA mRNA转染后DCs细胞内CEA蛋白显著高于对照组(83.32%vs3.34%,P<0.01)。CEA mRNA转染组DCs在效靶比为1∶10时,刺激T细胞增殖作用最强,明显高于未转染组[(19.11±1.89)%vs(15.59±0.70)%,P<0.05]。CEAmRNA转染组DCs能产生CEA特异性的CTL效应,在效靶比为5∶1、10∶1、20∶1和40∶1时杀伤率分别为[(5.42±0.87)%、(14.09±1.13)%、(27.16±0.72)%、(32.49±0.84)%,P<0.01],而未转染组和对照靶细胞均无杀伤作用。转染组DCs诱导的CTL上清中IFN-γ分泌量显著高于未转染组[(141.73±28.61)vs(9.45±4.63)pg/ml,P<0.01]。结论:CEAmRNA转染的成熟DCs体外能产生特异性抗肿瘤作用,为研制CEA RNA-DCs疫苗提供了实验依据。     

人肺癌组织总RNA转染树突状细胞方法的优化

背景与目的树突状细胞(dendriticcell,DCs)作为体内惟一能活化初始T细胞的抗原递呈细胞,是激发机体特异性免疫反应的关键,以DCs为基础构建的疫苗广泛应用于各种肿瘤免疫治疗中。其中RNA转染DCs构建DCs疫苗的方法,因避免了抗原的筛选和鉴定,所需肿瘤量小等优点,特别适于像肺癌这样的缺乏特异性肿瘤抗原、异质性大、抗原性弱的肿瘤。然而,目前缺乏RNA转染DCs的最佳时间和方法的研究。本研究以人肺癌组织总RNA体外转染DCs,试图摸索转染DCs的最佳条件。方法用免疫组化的方法选择肿瘤组织癌胚抗原CEA、粘蛋白MUC1双阳性的10例肺癌患者,一步法提取肿瘤组织总RNA,采集和分离外周血单核细胞,体外培养DCs和T细胞,将RNA于不同时间和不同方法转染DCs,用流式细胞仪检测CEA和MUC1的表达,混合淋巴细胞反应测定DCs刺激自体T细胞增殖能力。结果未成熟期DCs转染人肺癌总RNA后CEA和MUC1表达量(11.33±2.64和39.68±7.25)明显高于成熟期DCs(5.46±1.63和27.17±4.16),且刺激自身T细胞增殖反应能力更强。电穿孔法转染人肺癌总RNA后DCs表达CEA和MUC1的量(20.53±3.64和65.39±9.33)明显多于脂质体转染(11.33±2.64和39.68±7.25)和直接混合法(0.91±0.27和18.53±3.26),且刺激自身T细胞增殖反应能力更强。结论采用电穿孔法将肺癌总RNA体外转染未成熟DCs可以使DCs表达肿瘤抗原量更高,刺激T细胞增殖的活性更强。     

肿瘤源性血管内皮细胞RNA转染树突细胞体外诱导特异性CTL的研究

目的:探讨转染肿瘤血管内皮细胞RNA的树突细胞(DC)能否诱导特异性的细胞毒性T淋巴细胞(CTL)。方法:采用肿瘤细胞的培养上清诱导人脐静脉内皮细胞(HUVEC)增殖,从而制备肿瘤源性血管内皮细胞(Td-EC)。用RT-PCR检测肿瘤内皮标志1(TEM1)在Td-EC上的表达。制备Td-EC的总RNA,转染从人外周血中扩增的未成熟DC,用流式细胞仪检测DC表型的变化,用LDH法检测DC诱导的CTL对Td-EC、HUVEC和人脐静脉内皮细胞株CRL-1730的杀伤率。结果:Td-EC特异性表达TEM1。转染RNA的DC组CD83、CD86表达率分别为(85.22±7.73)%和(88.38±8.02)%,而未成熟DC组CD83、CD86表达率分别为(51.58±9.49)%和(61.77±2.91)%,两组差异有统计学意义,P<0.05。在效靶比为20∶1、10∶1和5∶1时,CTL对Td-EC的杀伤率分别为(34.15±1.14)%、(27.02±1.31)%和(13.12±1.63)%;CTL对HUVEC杀伤率分别为(11.35±0.97)%、(8.24±0.37)%和(5.30±0.73)%;CTL对CRL-1730杀伤率分别为(13.24±1.76)%、(8.41±0.91)%和(5.31±0.94)%;CTL对Td-EC杀伤率均显著高于对HUVEC和CRL-1730的杀伤率,P<0.05。结论:转染Td-EC RNA的DC可在体外诱导CTL特异性地杀伤Td-EC。     

GPC3基因转染的树突状细胞对人肝癌HepG2细胞杀伤作用的观察

目的:以GPC3基因转染人外周血单个核细胞来源的树突状细胞(DCs),诱导CTL产生特异性抗肝癌免疫.方法:从HLA-A2正常健康成人外周血分离单个核细胞,经GM-CSF和IL-4诱导产生DCs;通过脂质体转染法将携带人GPC3基因的质粒pEF-hGPC3转染DCs,蛋白质印迹法检测磷脂酰肌醇蛋白聚糖-3表达,流式细胞仪检测DCs表型的变化;MTT法检测DC诱导CTL对人肝癌细胞HepG2的特异性杀伤.结果:经细胞因子诱导产生了具有典型形态学的DCs,经肿瘤坏死因子-α(TNF-α)刺激成熟后可高表达CD80、CD86及CD83,而转染GPC3基因对协同刺激因子在DCs中的表达的影响并不显著.转染GPC3基因后,DCs可促进CTL特异性杀伤HepG2细胞,在不同效靶比的杀伤率:100∶1为(38.90±0.95)%,50∶1为(30.83±1.24)%,10∶1为(20.84±0.65)%,明显高于空载体组和对照组,P＜0.05.结论:GPC3基因转染人外周血单个核细胞来源的DCs可诱导和促进CTL特异性杀伤肝癌细胞HepG2作用,为治疗肝癌提供了新的免疫靶点.     

体外诱导尤文肉瘤DC疫苗的特异性抗肿瘤免疫鉴定及初步临床应用报告

目的：探讨尤文肉瘤细胞总RNA转染的DC疫苗体外诱导特异性抗肿瘤免疫的能力及临床初步应用的效果。方法：分离尤文肉瘤患者外周血单核细胞体外诱导为DC细胞。Trizol法提取患者尤文肉瘤细胞总RNA,总RNA转染DC并于体外诱导特异性CTL克隆扩增。RT-PCR方法检测肿瘤总RNA加载的DC细胞内EWS-FLI1mRNA的表达。MTT法检测淋巴细胞的增殖和CTL的杀伤活性。在试验成功基础上,将成熟DC疫苗接种患者并测定免疫学OKT值。结果：RT-PCR测定显示尤文肉瘤细胞总RNA转染的DC细胞可以提呈肿瘤特异性抗原,并特异性地表达其特有序列EWS-FLI1。经总RNA转染的DC特异性表面标志及功能相关分子表达均上调,转染后的DC可显著刺激自体T淋巴细胞增殖。诱导的特异性CTL对携带EWS-FLI1抗原的靶细胞的杀伤率显著高于LAK细胞和未经转染的DC。经疫苗免疫的患者血清CD8明显升高（P〈0.05）。结论：尤文肉瘤细胞总RNA转染的DC疫苗可在体外诱导出特异性抗肿瘤免疫,并在临床初步证实可提高尤文肉瘤患者的特异性抗肿瘤免疫力。     

Caveolin1对KDR信号通路的作用

目的:研究以小鼠结肠癌细胞CT-26RNA作为抗原体外转染经mIL-12基因修饰的树突状细胞(dendriticcells,DC),观察其诱导特异性抗肿瘤的效应。方法:小鼠骨髓细胞体外以rmGM-CSF、rmIL-4诱导培养获取树突状细胞,流式细胞术检测纯度;293细胞扩增携带mIL-12基因的重组腺病毒,体外转染树突状细胞;Trizol法提取CT-26细胞总RNA,应用Trans-Messenger体外转染mIL-12基因修饰的树突状细胞,免疫接种小鼠。ELISA法检测细胞上清及小鼠血液中mIL-12水平,LDH释放法检测小鼠体内细胞毒性T淋巴细胞(CTL)杀伤活性。结果:小鼠骨髓细胞经诱导培养后,获得大量高纯度的树突状细胞,流式细胞术检测CD11c 的树突状细胞>90%;提取的CT-26细胞总RNA体外经TransMessenger介导,转染mIL-12基因修饰的树突状细胞后,回输小鼠,可以诱导体内生成较高水平的特异性CTL活性,亲本肿瘤接种后小鼠100%长期存活,而以该RNA转染Ad-LacZ修饰DC后的对照组及RNA转染DC的对照组,诱导机体生成的特异性CTL活性显著低于实验组(P<0.01),亲本肿瘤接种后小鼠60%长期存活,DC、PBS对照组则均未诱导机体生成特异性CTL活性,小鼠无长期存活。结论:树突状细胞经小鼠结肠癌CT26细胞RNA转染和mIL-12基因修饰后免疫接种小鼠,可在体内有效提呈肿瘤抗原,诱导机体产生高水平的CTL,更有效地诱发特异性抗肿瘤效应。     

黏蛋白1基因转染DC对乳腺癌细胞MCF-7裸鼠移植瘤的免疫抑制作用

目的:探讨黏蛋白1 (mucin 1,MUC1基因转染DC对人乳腺癌MCF-7细胞裸鼠移植瘤的抑制作用.方法:体外诱导培养健康成人DC,应用脂质体转染法将pcDNA3.1-MUC1转染DC,ELISA法检测转染后DC分泌细胞因子IL-12和YNF-α的能力,LDH释放法检测基因转染后DC诱导特异性CTL对乳腺癌MCF-7细胞的杀伤活性.应用MU C1基因转染DC、空质粒转染DC、及生理盐水皮下注射治疗人乳腺癌MCF-7细胞裸鼠移植瘤,观测其对肿瘤生长的抑制作用.结果:转染pcDNA3.1-MUC1的DC分泌IL-12、TNF-α的能力较转染空质粒DC明显增强[IL-12:(202.52±29.61)vs(10.83±1.02)pg/ml;TNF-α:(349.07±79.42)vs(9.26±1.52)pg/ml,均P＜0.01];转染pcDNA3.1-MUC1的DC诱导产生特异性CTL,对人乳腺癌MCF-7细胞具有更明显的杀伤活性,效靶比为10∶1、5∶1和2.5∶1时的杀伤率分别达到56.2％、38.9％和25.8％,显著高于对照组CTL(均P＜0.01).MUC1基因转染DC对乳腺癌MCF-7裸鼠移植瘤生长抑制作用明显强于空质粒转染DC组(P＜0.05).结论:MUC1基因转染DC可以诱导特异性CTL,对乳腺癌MCF-7细胞具有更强的抗肿瘤免疫效应.     

Hep-2细胞总RNA转染树突状细胞诱发抗喉癌特异性免疫效应的研究

目的:研究转染喉鳞状细胞癌总RNA对树突状细胞(DC)免疫功能的影响,观察肿瘤总RNA修饰后的DC在体外诱导高效而特异的抗喉癌免疫效应.方法:采用分离脐血单核细胞体外诱导DC,Trizol法提取喉鳞状细胞癌细胞株Hep-2总RNA后转染入DC,流式细胞仪分析DC表面分子CD40、CD80、CD83、CD86的表达,LDH法评估转染肿瘤总RNA的DC瘤苗的细胞毒性T淋巴细胞反应.结果:转染后的DC表面分子表达CD40(60.6±0.15)%,CD80(60.2±0.21)%,CD83(85.1±0.06)%,和CD86(91.9%±0.09)%,而在未转染的未成熟DC中低表达;转染RNA的DC瘤苗对于Hep-2细胞株有特异性杀伤效应,在E/T为40:1时,杀伤效率达45.6%,而对对照组靶细胞杀伤效率较低.结论: 喉鳞状细胞癌总RNA转染修饰的DC能特异性诱导细胞毒性T淋巴细胞,显著提高DC的抗原提呈功能,在体外能诱导高效而特异的抗喉癌免疫效应.     

PD-L1分子在肺癌细胞株上的表达及其生物学意义

背景与目的目前研究表明,多种人类肿瘤大量表达的PD-L1分子参与了肿瘤免疫逃逸,其机制主要在于肿瘤细胞通过高表达PD-L1分子,与T细胞上的受体PD-1的结合,传递负性调控信号,导致肿瘤抗原特异性T细胞的凋亡和免疫无能。本文着重探讨PD-L1分子在肺癌细胞株上的表达及其对T细胞杀伤效应的调节作用。方法采用常规方法从健康人外周血单个核细胞诱导DCs,并经凋亡肿瘤细胞和激发型CD40单克隆抗体刺激获得成熟DCs,与自体T细胞共育后获得肿瘤特异性CTL细胞;流式细胞术检测肺癌细胞上PD-L1分子的表达;JAM法和单抗阻断实验分析CTL细胞对肺癌细胞株的杀伤效应,ELISA法检测细胞培养上清中IFN-γ的含量。结果经凋亡肿瘤细胞负载的成熟DCs可诱导自体T细胞分化为肿瘤特异性CTL;H1299高表达PD-L1分子(90.3±4.2)%,而A549低表达PD-L1分子(19.4±5.2)%;CTL对A549具有高效特异的杀伤力,而对H1299不能高效杀伤;联合应用PD-L1单抗可促进CTL对H1299的杀伤作用和IFN-γ的分泌(P<0.05)。结论在肺癌细胞株上表达的PD-L1分子,可降低CTL对肺癌细胞的杀伤效应。     

CIK、DC-CIK细胞对神经母细胞瘤细胞杀伤作用的研究

目的：研究细胞因子诱导的杀伤细胞（ CIK）与树突状细胞（ DC）共培养后对神经母细胞瘤（ neuro-blastoma,NB）细胞株的杀伤作用。方法：取健康人和肿瘤患者外周血单个核细胞（ PBMC）,加入不同的细胞因子分别诱导出DC和CIK细胞,用流式细胞术测定诱导培养前后DC和CIK细胞的表型,MTT法测定不同组CIK细胞对NB细胞株的杀伤活性。结果：流式细胞仪检测健康人PBMC培养后CD3＋CD56＋淋巴细胞百分比以及对NB细胞株的杀伤活性均显著高于肿瘤患者（ P〈0.05）。此外,与单纯CIK细胞相比,DC-CIK细胞具有更强的杀伤NB细胞株的活性（ P〈0.05）。结论：DC-CIK细胞是一种细胞毒作用高于单纯CIK细胞的免疫活性细胞。健康人和肿瘤患者的PBMC经诱导培养获得的CIK细胞有显著差别,为临床进一步提高CIK细胞的治疗效果提供了实验依据。     

Differences in inducibility of morphological differentiation of parental cells, selected variant cells and cells from spontaneous metastases of mouse neuroblastoma cells

The in vitro sensitivities to differentiating agents of a murine neuroblastoma cell line (N18) and a selected variant cell line (N18-LM5) were examined. In addition, the sensitivities to differentiating agents of cells from spontaneous metastases produced by N18 cells were examined. When N18 cells (1 X 10(5) cells/mouse) were injected into the lateral tail vein of syngeneic A/J mice, only a few metastatic nodules formed in the liver and lung, while similar injection of N18-LM5 cells produced larger numbers of metastatic nodules. Exposure of N18 cells to differentiating agents, such as dibutyryl cyclic 3':5'-AMP (db-cAMP), prostaglandin E1, and dexamethasone, resulted in induction of differentiation in terms of neurite extension. N18-LM5 cells responded to differentiating agents to a greater extent than N18 cells, and most of the cells extended neurites when they were exposed to 1 mM db-cAMP for 3 days. On the other hand, not all cell lines from spontaneous metastases produced by N18 cells responded to db-cAMP. These results suggest that the colonizing potential of neuroblastoma cells is not necessarily correlated with loss of responsiveness to differentiating agents and that various spontaneous metastases show heterogeneity in responsiveness to differentiating agents.     

Melanoma cells interfere with the interaction of dendritic cells with NK/LAK cells

Dendritic cells (DCs) and natural killer (NK) cells are key players at the interface between innate resistance and acquired immunity. NK cells can induce DC maturation, a differentiation process whereby DCs respond to a environmental stimulus and acquire the ability of eliciting adaptive immunity. Conversely, maturing DCs promote NK functions in vivo and in vitro. This interplay has important consequences on the immune response to pathogens and possibly to neoplastic cells. Here, we show that B16 melanoma cells actively modulate the interaction between DCs derived from bone marrow precursors and NK/LAK cells propagated from the spleen of C57BL/6 mice. DCs increased in a dose-dependent manner the ability of NK/LAK cells to kill melanoma cells and to produce cytokines. This activatory cross-talk entailed the production of IL-18 by DCs and of IFN-gamma by NK/LAK cells. Melanoma cells were not a passive target of NK activity; they regulated the outcome of the interaction between DCs and NK/LAK cells, inhibiting the in vitro production of cytokines as effectively as the genetic deletion of IL-18 or IFN-gamma. Interference with the NK/DC interaction possibly represents a mechanism used by growing tumors to evade the immune response.     

Immunogenic reactivity of CTLs induced by electrofusion cells of human dendritic cells and gastric cancer cells

In tumor immunotherapy, there were several reports of attempts to induce anti-tumor immunity by fusion hybrid cells generated with dendritic and tumor cells. One of them reported that vaccination of hybrid cells resulted in a remarkable reduction of tumor cells in a lab mouse experiment. In our study, fusion cells were generated successfully with human matured dendritic and human gastric cancer cells by electrofusion technique and employed to induce CTLs. The evaluated fusion rate was 47.8% by FACS analysis. We tried to induce CTLs by co culture of effector and stimulator cells in the presence of IL-2, IL-7 and IL-12 for 4 weeks. Although it was not statistically significant in tumor cytotoxic assay, effector cells induced by the fusion cells as stimulator cells showed a few cytotoxic responses in an immunological tumor specific manner. Our data suggest that fusion hybrid cells may facilitate stimulation and expansion of tumor-specific T cells, but further investigation is required for clinical application of fusion cells in adoptive immunotherapy.     

Marker profile of mesothelial cells versus ovarian carcinoma cells

We investigated the marker profile of human ascitic and cultured mesothelial cells, and compared it to that of ovarian carcinoma cells which are related in terms of their histogenesis, unrelated colon carcinomas being used as reference. Mesothelial and ovarian carcinoma cells could not be distinguished by (intermediate) filament typing, using monoclonal antibodies (MAbs) to keratins, vimentins and desmins. Colon carcinomas differed from mesothelial cells and ovarian carcinomas by the absence of keratin-7 filaments. The epithelial marker BW 495/36 was completely negative on mesothelial cells and positive on all ovarian and colon carcinoma cells. While CEA was found on about 85% of all colon carcinomas, CEA expression on mesothelial cells and ovarian carcinoma cells was below 20%. The ovarian carcinoma markers (OV-TL 3, OV-TL 10, OC 125, MOV 18) were strongly positive on ovarian carcinomas and negative on colon carcinomas (or limited to traces of immunofluorescence on some samples). Although the mesothelial cells showed weak or negative reactivity with these markers, OC 125 antigen was found by immunoelectron microscopy on the surface of cultured mesothelial cells, and was shed in the culture supernatant at concentrations of 50, 28, and 25 CA 125 U/ml/10(4) positive cells. This suggests that mesothelial cells may be responsible for the synthesis of CA 125 in ascitic fluid. The data indicate that ovarian carcinomas, mesothelial cells and colon carcinomas can be distinguished using a combination of anti-keratin antibodies with BW 495/36 and anti-ovarian carcinoma markers.     

Radial glia cells are candidate stem cells of ependymoma

Tumors of the same histologic type often comprise clinically and molecularly distinct subgroups; however, the etiology of these subgroups is unknown. Here, we report that histologically identical, but genetically distinct, ependymomas exhibit patterns of gene expression that recapitulate those of radial glia cells in the corresponding region of the central nervous system. Cancer stem cells isolated from ependymomas displayed a radial glia phenotype and formed tumors when orthotopically transplanted in mice. These findings identify restricted populations of radial glia cells as candidate stem cells of the different subgroups of ependymoma, and they support a general hypothesis that subgroups of the same histologic tumor type are generated by different populations of progenitor cells in the tissues of origin.     

Can cancer cells transform normal host cells into malignant cells?

A human prostate tumour cell line, LNCaP C4-2, when injected into athymic male nude mice, produced tumours containing: (1) only human cancer cells similar to those injected; (2) only murine stromal cells containing abnormal chromosome constitutions; or (3) both human prostate cancer cells similar to those injected and the transformed murine stromal cells with altered chromosome constitutions. Karyotypic analysis of murine metaphases from all the host-derived tumours showed mostly pseudodiploid chromosome constitutions, with multiple copies (amplification) of mouse chromosome 15 and the absence of a typical Y chromosome. Fluorescence in situ hybridization analysis of these murine cells, using a biotin-labelled total human DNA painting probe, further demonstrated the absence of human DNA and the presence of only mouse metaphase and interphase cells in these transformed stromal cells. These results suggest that cancer cells are capable of inducing neoplastic transformation in stromal cells of the host organ by some, as yet unknown, epigenetic mechanism(s).     

α-半乳糖神经酰胺负载DC促进CIK细胞对肝癌细胞的杀伤效应

目的:研究负载α-半乳糖神经酰胺(alpha-galactosylceramide,α-GalCer)的DC功能与成熟度的改变情况,探讨α-GalCer-DC与CIK共培养对CIK细胞表型、增殖活性及杀伤肝癌细胞效率的影响.方法:采用密度梯度离心法从人外周血中分离出单个核细胞,悬浮细胞诱导培养CIK细胞,贴壁细胞诱导培养DC;流式细胞仪检测α-GalCer负载DC的表型,Real-time PCR检测DC相关基因的mRNA表达改变.将α-GalCer-DC与CIK细胞共培养,流式细胞仪检测DC与CIK细胞表面标志物;锥虫蓝染色法检测CIK细胞的增殖倍数;Real-time PCR检测CIK细胞功能相关基因的表达情况;CCK-8试剂盒检测α-GalCer-DC对CIK杀伤HepG2细胞的影响.结果:经过多种细胞因子诱导,可获得CIK细胞和成熟的DC;α-GalCer负载可促进DC成熟,DC表面标志物CD80、CD86、CD83和CD11c的阳性率均升高(P＜0.05或P＜0.01),表面趋化因子受体CCR-7、IL-12、IL-10的mRNA水平升高(P＜0.05).α-GalCer-DC与CIK共培养,可显著提高CIK细胞CD3+ CD56+的表达和增殖活性(P＜0.05或P＜0.01),并显著提高INF-γ、IL-12、穿孔素和颗粒酶素B的mRNA表达水平(P＜0.05或P＜0.01);CIK、DC-CIK、α-GalCer-DC-CIK细胞对HepG2细胞的杀伤作用随效靶比的升高而增强,在同一效靶比时α-GalCer-DC-CIK细胞对靶细胞的杀伤作用最强(P＜0.05).结论:α-GalCer负载可促进DC成熟,α-GalCer-DC与CIK共培养能促进后者增殖和成熟,能显著增强其对肝癌细胞的杀伤活性,为DC-CIK在肿瘤免疫治疗中的应用提供了实验依据.     

Chemoresistant colorectal cancer cells and cancer stem cells mediate growth and survival of bystander cells

Background:

Recent studies suggest that cancer stem cells (CSCs) mediate chemoresistance, but interestingly, only a small percentage of cells in a resistant tumour are CSCs; this suggests that non-CSCs survive by other means. We hypothesised that chemoresistant colorectal cancer (CRC) cells generate soluble factors that enhance survival of chemonaive tumour cells.

Methods:

Chemoresistant CRC cells were generated by serial passage in oxaliplatin (Ox cells). Conditioned media (CM) was collected from parental and oxaliplatin-resistant (OxR) cells. CRC cells were treated with CM and growth and survival were assessed. Tumour growth rates were determined in nude mice after cells were treated with CM. Mass spectrometry (MS) identified proteins in CM. Reverse phase protein microarray assays determined signalling effects of CM in parental cells.

Results:

Oxaliplatin-resistant CM increased survival of chemo-naive cells. CSC CM also increased growth of parental cells. Parental and OxR mixed tumours grew larger than tumours composed of parental or OxR cells alone. Mass spectrometry detected unique survival-promoting factors in OxR CM compared with parental CM. Cells treated with OxR CM demonstrated early phosphorylation of EGFR and MEK1, with later upregulation of total Akt .We identified progranulin as a potential mediator of chemoresistance.

Conclusion:

Chemoresistant tumour cells and CSCs may promote resistance through soluble factors that mediate survival in otherwise chemosensitive tumour cells.