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淋巴细胞瘤

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To date,it has been documented after transfusion of unirradiated blood components to at least 87 patients:in patients with severe combined immunodeficiency,thymic hypoplasia,and Wiskott Aldrich syndrome premature newborns and those with erythroblastosis fetalis; patients with hematologic malignancies including Hodgkin\'s and non-Hodgkin\'s lymphomas,acute myelocytic and lymphoblastic leukemias,chronic lymphocytic leukemia,and aplastic patients with solid tumors including neuroblastomas, glioblastoma,rhabdomyosarcoma,cervical carcinoma,small cell lung cancer,and germ cell tumor;patients after cardiac surgery and cholecystectomp-60;and in an apparently healthy 22-year-old woman.

目前为止,输注未辐照血液成分后,至少引起了87例TA-GVHD的发生,主要见于以下病人:严重的免疫缺陷病人、胸腺发育不全、Wiskott Aldrich综合症、早产儿、胎儿红细胞增多症、何杰金与非何杰金氏淋巴瘤等恶性血液病人、急性粒细胞性和淋巴细胞性白血病、慢性淋巴细胞性白血病、成神经细胞瘤等实体瘤病人、横纹肌赘瘤、宫劲癌、小细胞肺癌、微细胞瘤、心脏和胆囊手术病人。

Results: Among 38 transgenic mice surviving beyond one year.12(9 myc and 3 myc+ ras)mice developed multiple neoplasms.Most tumors were observed as lymphoblastoma and fibrosarcoma in histological types.

结果:在存活一年以上的38只转基因小鼠中有12只(9myc和3myc+ras)发生多种肿瘤,其组织类型以成淋巴细胞瘤和纤维肉瘤居多。

DCs acquired by our reformed methods express both CD83 and CD 14 molecules highly, and have a higher density than other domestic reports. The higher TNF in DCs culture medium of HC patient suggests DCs in patient still have antigen presenting ability and by optimization the culture medium would improve its presenting ability and have a potential value in design and application individual vaccine. Although antigens pulsed DCs have a decrescent antigen presenting ability but BCG HSP70 could induce its mature and improve its presenting ability. Suggests BCG HSP70 would be a useful mature inducer. Lymphocytes primed by DCs based HC vaccine have the specific cytotoxicity against HCC lines. The CTL after freezing and anabiotic could prophylaxis and therapy HC xenograft on nude mouse. The results also suggests that CD4〓 lymphocytes play a important role in HC with a good differentiation and would be useful in treatment this kind of HC. After being activated by Peptide LLNQHACAV of hAFP and apoptotic HCCs pulsed DCs respectively, the culture medium of activated lymphocytes both contains a high level Th1 cytokines IL-12 and TNF. Primed lymphocytes appeared a characteristic of NK cells. DCs not only inhibited the growth of human HCC and other cancer cells in vitro but also prevented the growth of HC xenograft on nude mouse in vivo. There are at least three kinds of mechanism playing important role in DC based vaccine ,there are inhibition of DCs, HC specific CTL and cytokines pathway.

诱导出的DC共同表达CD83和CD14分子,CD83分子表达明显高于国内报道;肝癌患者DC培养上清中TNF水平高于健康人,提示肝癌患者DC仍具一定的抗原呈递能力,适当调控可使其行使APC功能,以期在肝癌个体化疫苗中发挥作用;DC负载肝癌可溶性抗原后,抗原呈递能力降低,BCG HSP70可促进DC成熟,增加其抗原呈递能力,预示BCG HSP70有可能成为促进DC活化和成熟的另一重要分子;肝癌DC疫苗在体外诱导肝癌特异性淋巴细胞,活化的淋巴细胞在体外对肝癌细胞的杀伤以特异性CTL为主,同时分泌较高水平Th1型细胞因子IL-12和TNF,并抑制4种肝癌细胞生长;冷冻复苏后的肝癌特异性淋巴细胞可以预防和抑制人肝癌裸鼠皮下移植瘤,提示DC负载肝癌可溶性抗原后诱发的MHC-Ⅱ类限制性CD4〓T细胞有可能在分化程度高的肝癌治疗中发挥作用;用DC和HLA-A2〓DC分别负载凋亡肝癌细胞和hAFP218-226位LLNQHACAV HLA-A2限制性九肽,在体外诱导肝癌特异性淋巴细胞,活化后的CTL细胞分泌较高水平的Th1型细胞因子IL-12和TNF,并具较强杀伤活性,此CTL同时具备NK细胞特征;DC对肿瘤细胞的抑制作用可能是通过吞噬实现的,Fas-L在DC抑制中也起一定作用;DC对人肝癌裸鼠皮下移植瘤的抑制率为97%;在肝癌DC疫苗的作用中,至少联合3种以上机制,即通过DC的直接作用、肝癌特异性CTL和细胞因子途径直接或间接地杀伤和抑制肝癌细胞。

Get high purity DCs by Cultured plastic-adherent monocytes isolated from healthy human peripheral blood with GM-CSF and IL-4 for 7 days. To observe the morphology of DCs by inverted phase contrast microscope ,electron microscope and laser confocal microscope. Analyse phenotype of DCs with flow cytometry. Investigate the endocytosis ability of DCs as a group by Horseradish peroxidase endocytosis assay. To appraise allogeneic mixed lymphocytes reaction of DCs by MTT reduction assay. Analyse the levels of IL-12 and TNF in liquids of cultured medium by ELISA and MTT reduction assay respectively. Soluble antigens of HCCs was obtained by 3 freeze-and-thaw cycles. Biological characteristics of HC soluble antigens pulsed DCs were monitored by flow cytometry. According to MTT reduction assay estimated the cell proliferation of self lymphocytes activated by HC antigens pulsed DCs. Get high purity BCG HSP 70 protein by SDS-PAGE electrophoresis and determined its biological activity with ELISA. Analyse phenotype of antigen pulsed DCs primed by BCG HSP70 with flow cytometry. By MTT reduction assay estimated the cell proliferation of self lymphocytes and the MLR of DC based vaccine. Analyse expression of HLA-DR molecule on surface of HCC lines. The IFN-γ mRNA in lymphocytes after actived by DC vaccine and the Fas-L expression on DC and DC vaccine primed lymphocytes were detected by in situ hybridization and flow cytometry respectively. Specific cytotoxity lysis of T lymphocytes and nonspecific inhibition of liquids in culture medium against HCC lines were also tested. Detect expression of hAFP on four HCC lines with Cell-ELISA. Induce apoptosis of HCCs with actinomycin-D. Interaction of DCs and apoptotic cells was observed under transmission electron microscope. Growth inhibition test of DC against HCC lines was also performed. Establish the nude mouse model bearing human HC xenografts and indentify the characteristic of tumour by histochemistry and immunohistochemistry techniques. Prevent and treat transplanted human HC on nude mouse with Freezing and anabiotic HC specific lymphocytes.

用GM-CSF和IL-4从健康人外周血诱导DC;分别用倒置相差显微镜、电子显微镜及激光共聚焦显微镜观察DC形态;流式细胞术检测DC表型;HRP吞噬实验测定DC的群体内吞能力;MTT法检测同种异体混合淋巴细胞反应;ELISA法和MTT法分别测定DC培养上清液中IL-12和TNF水平;冻融法制备肝癌细胞可溶性抗原;流式细胞术检测负载肝癌可溶性抗原后DC的生物学特性;MTT法检测DC负载肝癌抗原后对自身淋巴细胞增殖的影响;SDS-PAGE制备电泳纯化BCG HSP70并鉴定纯度,ELISA测定活性;流式细胞术检测负载抗原DC经BCGHSP 70活化后的表型;MTT法检测肝癌DC疫苗对自身淋巴细胞增殖的影响和混合淋巴细胞反应;流式细胞术检测肝癌细胞表面HLA-DR表达;MTT法检测肝癌DC疫苗对自身淋巴细胞的活化;原位杂交法检测肝癌DC疫苗活化后的淋巴细胞IFN-γmRNA表达;流式细胞术检测DC和肝癌DC疫苗活化后淋巴细胞表面Fas-L;MTT法分别检测肝癌DC疫苗活化的淋巴细胞和其培养上清对肝癌细胞的特异性杀伤和非特异性抑制作用;Cell-ELISA检测人肝癌细胞hAFP表达;MTT法检测负载AFP表位肽和凋亡肝癌细胞DC对自身淋巴细胞增殖的影响;ELISA法和MTT法分别测定活化后淋巴细胞培养上清中TNF和IL-12水平;肝癌细胞凋亡的诱导和检测;DC吞噬凋亡肝癌细胞后的电子显微镜观察;DC对肝癌细胞的生长抑制试验;人肝癌裸鼠皮下移植瘤动物模型的建立及其组织学和免疫组织化学鉴定;DC及肝癌特异性淋巴细胞预防和治疗人肝癌裸鼠皮下移植瘤;冻存和复苏后的肝癌特异性淋巴细胞预防和治疗人肝癌裸鼠皮下移植瘤。

Methods By establishing the apoptosis model of primary culture of hippocampal neurons induced by hyperthermia, the dynamic change of concentration Ca(superscript 2+)in hippocampal neurons and the influence to Bcl-2 expression were observed after using chelerythrine chloride which was the special inhibitor of PKC.

通过建立体外高温诱导海马神经细胞原代培养的凋亡模型,应用PKC特异性抑制剂氯化百屈菜红碱,观察海马神经元胞内Ca(上标 2+)浓度的动态变化以及对B淋巴细胞瘤基因2蛋白(Bcl-2)表达的影响。

Methods Granulocyte/macrophage colony stimulating factors and interleukin 4 were used to cultivate DC from peripheral blood of hepatocellular carcinoma patients.Tumor associated antigen extracted from human liver cancer HepG2 cell line was used to activate the DC,which will initiate homogenic T lymphocyte to form cytotoxic T lymphocyte.HepG2 cells were transplanted to the nude mice and were treated with CTL.Expression of PCNA in tumor cells was evaluated.

联合应用粒/巨细胞集落刺激因子及白介素-4(IL-4)直接从肝癌患者外周血中培养出DC;以源于人肝癌细胞系HepG2细胞的肿瘤抗原粗提物刺激DC;DC激活同源的T淋巴细胞产生细胞毒性T淋巴细胞;建立人肝癌细胞系HepG2裸鼠移植瘤模型;CTL治疗人肝癌细胞系HepG2裸鼠移植瘤;检测移植瘤标本肿瘤细胞PCNA表达水平。

The list of possible causes of diabetes insipidus in this case and many others is extensive and includes infection (tuberculous pachymeningitis and Whipple's disease), malignant neoplasms (germ-cell tumor and lymphoma), infiltrative processes (hemochromatosis and amyloidosis), autoimmune diseases (Wegener's granulomatosis, lymphocytic infundibulitis, and sarcoidosis), and histiocytosis (Langerhans'-cell histiocytosis and non-Langerhans'-cell histiocytosis).

这例病例其他许多病例尿崩症的可能病因广泛,包括感染(结核性脑脊髓膜炎和Whipple病),恶性肿瘤,变性疾病,自身免疫病(Wegener肉芽肿,垂体茎淋巴细胞瘤和粒细胞肉瘤)和组织细胞增生症(Langerhans细胞和非Langerhans细胞组织细胞增生症

Methods DCs were prepared from peripherad blood mononuclear induce d with gra-nulocyt e/macrophage colony-stimulating factor and interleukin-4. Apoptosis of glioma cells were induced with γ-radiation. We design these experiment groups including (1) coculture of DCs and apoptotic glioma cells and T cells,(2) coculture of DCs and U937 cells and T cells,(3) coculture of DCs and cultured glioma cell and T cells,(4) coculture of Des and T cell.

用粒-巨噬细胞集落刺激因子加白介素-4(IL-4)从人外周血分化、诱导DCs、γ-射线在体外诱导培养的人脑胶质瘤细胞凋亡,将DCs、T淋巴细胞和凋亡胶质瘤细胞共培养,同时设计不同类型肿瘤细胞(U937及培养胶质瘤细胞)作对照,分离、富集DCs、T淋巴细胞进行免疫应答及肿瘤细胞杀伤试验。

Methods:Human CD80 cDNA was transfected into B16 melanoma cells by lipofectin-mediated gene transfer before the expansion of tumor cells were tested by MTT method in vitro;C57BL/6 mice were inculated subcutaneously either mock-transfected B16 cells (B16-neo)or CD80-transfected B16 cells (B16-CD80),then the latency,survival times and tumor mass growth were investigated.The lymphocytes were examined for both proliferation indices and specific cytotoxic activity by MTT method and improved LDH-releasing method,after syngenic Mixed Lymphocyte tumor cultures.

通过脂质体介导将CD80基因导入小鼠黑色素瘤B16细胞后,利用SABC法检测CD80的表达;MTT法测定肿瘤细胞体外增殖能力;将肿瘤细胞接种至同源小鼠皮下后观察成瘤期、荷瘤小鼠存活期及肿瘤结节生长速度;在同源淋巴细胞肿瘤细胞混合培养后,通过测定淋巴细胞增殖指数和CTL杀伤活性检测肿瘤细胞的免疫原性。

Result: With a high cancer-inhibiting rate on the animal model having been transplanted the cancer, this medicine can remarkably restrain the growth of cancer cells cultivated in vitro and improve the life quality of mouse model, and also shows obvious promotion and enhancement function on the DTH ear-swelling of mouse with cancer, the proliferation of mouse spleen lymphocyte induced by ConA and the cytokine (TNF-α、IFN-γ) of mouse with cancer respectively. It can obviously decrease the adverse effects like weight reduction, damage of viscera and degradation of life quality caused by the chemotherapeutic medicines, and has a remarkable stimulation on the lymphocyte proliferation induced by ConA. It is also able to improve the ratio of SMMC-7721 cells in G0-G1 period and reduce the cell amount in G2/M period to cut down the possibility of stepping into the division period; additionally, it can obviously inhibit some cancer genes of the cancer cells and meanwhile up regulate the cancer-inhibiting gene.

结果:本品能明显抑制体外培养肿瘤细胞的生长,对在体肿瘤移植模型动物具有较高的抑瘤率,能明显提高模型小鼠的生存质量;对荷瘤小鼠DTH耳肿胀、ConA诱导的小鼠脾淋巴细胞增殖、荷瘤小鼠细胞因子(TNF-α、IFN-γ)呈现明显的促进或提高作用;能显著减少化疗药所引起的小鼠体重降低、脏器损害以及生存质量下降等不良作用;本品含药血清对ConA诱导的淋巴细胞增殖具有明显的促进作用;能提高SMMC-7721细胞G0-G1期比例,降低G2/M期细胞数目,而减少其进入分裂期;对肿瘤细胞的多个癌基因具有明显的抑制作用,同时上调抑癌基因等。

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