肺癌患者肺泡巨噬细胞抗肿瘤免疫功能研究

用镀铜镉还原法、放射免疫法、比色法、酶标双抗体夹心法及逆转录－聚合酶链式反应（ＲＴ－ＰＣＲ）研究肺癌患者肺泡巨噬细胞（ＡＭ）抗肿瘤免疫功能状态。结果显示肺癌患者荷瘤侧肺ＢＡＬＦ中ＮＯ，ＡＭ培养上清中ＮＯ和ＴＮＦα活性低于非荷瘤侧肺与对照组，ｓＩＬ－２Ｒ活性高于对照组；经ＧＭ－ＣＳＦ刺激后ＡＭ培养上清中ＮＯ，ＴＮＦα及ＳＯＤ活性增强，ｓＩＬ－２Ｒ水平降低；两组ＡＭｉＮＯＳｍＲＮＡ表达强度明显弱于对照组。表明肺癌患者机体免疫功能及肿瘤局部ＡＭ抗肿瘤功能均存在不均一性缺陷。     

肺癌患者肺泡巨噬细胞一氧化氮活性研究

用镀铜镉还原－Ｃｒｉｅｓｓ法检测肺癌患者ＢＡＬＦ及肺泡巨噬细胞（ＡＭ）培养上清液中ＮＯ水平；ＲＴ－ＰＣＲ检测ＡＭ ｉＮＯＳ ｍＲＮＡ表达。结果显示，肺癌组ＢＡＬＦ及ＡＭ培养上清液中ＮＯ水平均明显低于对照组。两组ＡＭ ｉＮＯＳ ｍＲＮＡ表达阳性率分别为６９％和９１％（Ｐ〉０．０５），但表达强度肺癌组明显弱于对照组（Ｐ〈０．０１）。经ＧＭ－ＣＳＦ刺激后，ＡＭ ｉＮＯＳ表达强度及培养上清液中ＮＯ水平均显     

肺癌患者肺泡巨噬细胞一氧化氮活性研究

用镀铜镉还原－Ｇｒｉｅｓｓ法检测肺癌患者ＢＡＬＦ及肺泡巨噬细胞（ＡＭ）培养上清液中ＮＯ水平；ＲＴ－ＰＣＲ检测ＡＭｉＮＯＳｍＲＮＡ表达。结果显示，肺癌组ＢＡＬＦ及ＡＭ培养上清液中ＮＯ水平均明显低于对照组。两组ＡＭｉＮＯＳｍＲＮＡ表达阳性率分别为６９％和９１％（Ｐ＞０．０５），但表达强度肺癌组明显弱于对照组（Ｐ＜０．０１）。经ＧＭ－ＣＳＦ刺激后，ＡＭｉＮＯＳ表达强度及培养上清液中ＮＯ水平均显著提高。提示肺癌局部ＡＭ抗肿瘤功能可能存在某些缺陷；ＧＭ－ＣＳＦ可刺激ＡＭ使其功能增强     

健择对非小细胞肺癌患者肺泡巨噬细胞免疫功能影响的研究

目的探讨健择对非小细胞肺癌患者肺泡巨噬细胞(AM)凋亡及其分泌细胞毒效应分子功能的影响。方法ELISA法、酶法分别检测35例非小细胞肺癌患者支气管肺泡灌洗液中及其AM培养上清夜中TNF-α和NO水平,并分别用IFN-α、健择和两药联合干预AM培养,检测其培养上清液中TNF-α、NO变化水平。流式细胞仪检测健择对AM凋亡的影响。结果所有非小细胞肺癌患者其肺泡巨噬细胞均可自发地产生一定量的TNF-α和NO。非小细胞肺癌患者荷瘤侧肺NO和TNF-α活性无论是整体状态,还是细胞水平均明显低于非荷瘤侧肺。健择单独干预组肺泡巨噬细胞培养上清液中TNF-α和NO的浓度无明显变化(P>0.05);经IFN-α刺激肺泡巨噬细胞培养上清液中TNF-α和NO的浓度明显升高,与对照组比较差异呈显著性(P<0.01);健择和IFN-α联合干预下肺泡巨噬细胞培养上清液中TNF-α和NO水平较对照组、IFN-α干预组明显升高,差异呈显著性(P<0.05)。不同时间不同浓度健择对肺泡AM凋亡率无明显差异(P>0.05)。结论健择单独应用对肺泡巨噬细胞免疫功能无明显影响,联合干扰素时能提高非小细胞肺癌患者肺泡巨噬细胞抗肿瘤免疫功能。其机制有待进一步研究。     

肺癌患者肺泡巨噬细胞精氨酸酶活性研究

检测肺泡巨噬细胞（ＡＭ）培养液中精氨酸酶活性。结果表明，肺癌患者荷瘤侧肺ＡＭ较非荷瘤侧肺和非癌性肺病患者ＡＭ产生精氨酸酶活性低；ＡＭ经卡介菌（ＢＣＧ），干扰素－α（ＩＦＮ－α）刺激后产生精氨酸酶活性明显增强。提示肺癌患者肿瘤局部ＡＭ抗肿瘤活性存在缺陷；ＢＣＧ，ＩＦＮ－α诱导ＡＭ产生精氨酸酶可能为其抗肿瘤、抗感染的机制之一。     

肺癌患者肺泡巨噬细胞表现HLA-DR的表达

目的 了解肺癌患者肺泡巨噬细胞表面人白细胞抗原-DR(HLA-DR)的表达。方法 经支气管肺泡灌洗获肺泡巨噬细胞，贴壁分离及培养，免疫组化方法检测HLA-DR表达的阳性细胞百分率。结果 ① 肺癌及良性肺病患者，其肺泡巨噬细胞未经刺激，均已有部分表达HLA-DR，但前者明显高于后者；② 干扰素-α(IFN-α)或脂多糖(LPS)刺激后，HLA-DR的表达均增加，且两者联合刺激优于单独刺激；③ 无论刺激与否，肺癌组荷瘤侧肺与非荷瘤侧肺比较，肺泡巨噬细胞表达HLA-DR的阳性细胞率明显增高。结论 肺癌患者，尤其是肿瘤局部，肺泡巨噬细胞可能通过表达HLA-DR提高抗原递呈功能。IFN-α、LPS均能明显增强肺癌患者肺泡巨噬细胞的抗原提呈功能，且两者有协同作用。     

肺疾患肺泡巨噬细胞功能的变化

用伊红－美蓝染色法和中性红染料摄入法检测了４１例ＢＡＬＦ中人肺泡巨噬细胞的百分率和吞噬能力。结果显示：肺癌患者肺泡巨噬细胞百分率与对照组相比较无显著性差异；但吞噬能力低于对照组；肺间质性病变患者肺泡巨噬细胞百分率和吞噬能力低于对照组。     

肺癌患者肺泡巨噬细胞表现HLA—DR的表达

目的 了解肺癌患肺泡巨噬细胞表面人白细胞原－ＤＲ（ＨＬＡ－ＤＲ）的表达。方法 经支气管肺泡灌洗获肺泡巨噬细胞,贴壁分离及培养,免疫组化方法检测ＨＬＡ－ＤＲ表达的阳性细胞百分率。结果 ①肺癌及良性肺病患,其肺泡巨噬细胞未经刺激,均忆有部分表达ＨＬＡ－ＤＲ,但前明显高于后;②干扰素α（ＩＦＮ－α）或脂多糖（ＬＰＳ）刺激后,ＨＬＡ－ＤＲ的表达均增加,且两联合刺激优于单独刺激;③无论刺激与否,肺     

肺癌患者肺泡巨噬细胞精氨酸酶活性研究

检测肺泡巨噬细胞培养液中中精氨酸活性。结果表明，肺癌患者荷瘤侧肺ＡＭ较非荷瘤侧肺和非癌性肺癌患者ＡＭ产生精氨酸活性低；ＡＭ经卡介菌，干扰素－α刺激后产生精氨酸活性明显增强。     

Immunological status of alveolar macrophages in patients with lung cancer and benign pulmonary diseases]

This is a report of the study on the immunological status of alveolar macrophages (aM phi) in patients with lung cancer (LC, n = 27) and benign pulmonary diseases (BD, n = 26). Patients were undergone bronchoalveolar lavage by fiberoptic bronchoscopy. aM phi in the lavage fluid isolated by adherence on plastic surface were examined in vitro for their cytostatic and cytolytic activities against tumor target cells, secretion of interleukin 1 (IL-1) and tumor necrosis factor (TNF), intracellular IL-1 activity and mRNA expression of IL-1 beta and TNF-alpha. aM phi, both non-activated and activated, were shown to be highly cytostatic against P815 cells by 3H-TdR post-labelling assay. There was no statistical difference between the LC and BD group. As shown by isotope release assay, regardless of being activated or not, aM phi were not cytolytic against P815 and NS-1 cells in both groups of patients. TNF activity could be demonstrated in the culture supernatants of aM stimulated with LPS. Statistically, the TNF activity was not different in the two groups of patients. Spontaneous release of TNF activity was occasionally detected in unstimulated aM phi. While both intracellular and extracellular IL-1 activity of unstimulated aM phi was demonstrated in the two patient groups, the former activity was 1 to 5 times as high as the latter. When stimulated with LPS, there was some increase in extracellular but not intracellular IL-1 activity. mRNA expression of IL-1 beta and TNF-alpha by dot blot hybridization was demonstrable in aM phi from both patient groups irrespective of activation. These results indicate that the immune status of aM phi in lung cancer patients examined does not differ from that in patients with benign pulmonary diseases.     

肺癌患者与吸烟及非吸烟者支气管肺泡灌洗液T淋巴细胞...

The measurements of T-lymphocyte subsets (as expressed by, CD3 CD4 and CD8) in the peripheral blood and bronchial alveolar lavage fluid (BALF) on 30 cases of lung cancer, 26 cases of smoker and 25 cases of nonsmoker have been done. The results showed that there existed no differences in the peripheral blood of the above said subsets of lymphocyte. In BALF the percentage of CD3 in all the lymphocytes did not show any significant difference among the three tested groups, either (P > 0.05). But the percentage of CD4 and the ratio of CD4/CD8 manifested the following peculiarity: patients with lung cancers < smokers < nonsmokers (P < 0.01). On the contrary, the percentage of CD8 showed just the opposite: patients with lung cancer > smokers nonsmokers (P < 0.01). It was suggested that the cellular immunity as shown in the peripheral blood did not correspond with the findings in the BALF. The cellular immunity of the lungs was decreased both in patients with lung cancer and smokers, but more severely in the group of lung cancer patients. Smokers who have had changes of above said subsets of lymphocytes in the lungs may develop lung cancer. As the lung cancer advances, it may suppress body immunity and in turn enhance the cancer growth. Both the cause and the effect are interrelated. Therefore, the provocative agents for cellular immunity may act as one of the adjunctive therapies for patients with lung cancer. Giving up smoking should play an important role in recovering or promoting the immunity of the lungs and decreasing the incidence as well as improving the prognosis in patients with lung cancer.     

人参单体皂甙Rh2对非小细胞肺癌患者肺泡巨噬细胞免疫活性的影响

目的:探讨人参单体皂甙Rh2(G-Rh2)对非小细胞肺癌患者肺泡巨噬细胞(AM)分泌细胞毒效应分子的影响.方法:采用ELISA法和酶法分别检测35例非小细胞肺癌患者支气管肺泡灌洗液及其AM培养上清液中肿瘤坏死因子(TNF-α)和NO浓度,并分别用干扰素-α(IFN-α)、G-Rh2以及两药联合干预AM培养,检测其上清液中TNF-α和NO变化情况.结果:所有非小细胞肺癌患者其AM均可产生一定量的TNF-α和NO;非小细胞肺癌患者荷瘤侧肺NO和TNF-α活性在肺泡灌洗液及AM培养上清液中均明显低于非荷瘤侧肺.G-Rh2干预下AM培养上清液中TNF-α和NO的浓度较对照组明显升高(P＜0.05);与TNF-α干预AM培养上清液中TNF-α和NO的浓度比较差异无统计学意义(P＞0.05).G-Rh2和TNF-α联合干预时AM培养上清液中TNF-α和NO的浓度明显大于TNF-α或G-Rh2单独干预(P＜0.01).结论:G-Rh2能促进非小细胞肺癌患者肺泡巨噬细胞分泌细胞毒效应分子,联合干扰素时其作用更显著,两者具有协同性.     

肺泡巨噬细胞来源的微囊泡对肺损伤的作用

目的探究肺泡巨噬细胞来源微囊泡(microvesicles,MVs)对肺损伤的作用。方法培养小鼠肺泡巨噬细胞,通过梯度离心法获取磷酸缓冲盐溶液(phosphate buffer saline,PBS)或脂多糖刺激的肺泡巨噬细胞来源的MVs,并经透射电镜、纳米粒径追踪技术及蛋白免疫印迹进行鉴定。经气管内给予肺泡巨噬细胞来源的MVs,观察小鼠肺组织变化情况,并经蛋白免疫印迹探究肺组织中紧密连接相关蛋白的表达变化。结果透射电镜发现MVs呈现sharp结构,纳米粒径追踪显示MVs粒径集中于100~200 nm,并且其表达CD63、CD68及ALIX标志物。苏木精-伊红染色发现脂多糖刺激的肺泡巨噬细胞来源的MVs可刺激小鼠发生肺损伤,并降低紧密连接相关蛋白闭合小环蛋白,咬合蛋白及闭合蛋白的表达。结论炎性肺泡巨噬细胞来源的MVs可导致肺损伤并降低肺部紧密连接相关蛋白的表达,肺泡巨噬细胞来源的MVs参与肺损伤的发生发展。     

内毒素血症时小鼠肺泡巨噬细胞CD14和清道夫受体的表达

目的 探讨内毒素肺损伤时,肺泡巨噬细胞逐步由免疫防御型转变为效应型的分子机制。方法 建立内毒素肺损伤动物模型,免疫组化法观察肺泡巨噬细胞ＣＤ１４及清道夫受体（ＳＲ）表达的变化,并辅以计算机图像分析;同时检测肺体指数,肺组织的病理变化及动物存活率。结果 内毒素对ＣＤ１４的表达呈时间依赖性升高,但加大剂量并未使其进一步增加;ＳＲ呈时间及剂量依赖性降低。肺泡巨噬细胞ＳＲ、ＣＤ１４的表达变化与小鼠肺损伤程     

受体相互作用蛋白140在脓毒症急性肺损伤大鼠肺泡巨噬细胞中的表达及其作用

目的 探讨受体相互作用蛋白140 (receptor interacting protein140,RIP140)在脓毒症急性肺损伤(acute lung injury,ALI)大鼠肺泡巨噬细胞中的表达及其在炎症反应过程中的作用.方法 36只雄性SD大鼠按随机数字表法分为正常对照组、假手术组和盲肠结扎穿孔术(CLP)致脓毒症肺损伤3、6、12、24 h组.HE染色观察肺组织病理变化;免疫组化检测RIP140在肺组织的表达分布;Western blot及免疫荧光观察肺泡巨噬细胞RIP140的表达变化及定位;RT-qPCR测定肺泡巨噬细胞IL-1β、TNF-α、IL-6 mRNA水平变化.结果 CLP组可见明显肺损伤病理改变且随时间进展逐渐加重.RIP140在脓毒症肺损伤大鼠肺组织中主要表达于炎性渗出细胞,且随时间进展,表达RIP140的细胞数量逐渐增多.CLP术后6h大鼠肺巨噬细胞RIP140蛋白表达水平较正常对照组开始升高(P＜0.05),至12～24 h维持一较高水平,且主要表达于细胞核.CLP术后3h大鼠肺巨噬细胞IL-1β、TNF-α、IL-6 mRNA水平较正常对照组开始升高(P＜0.05),至12～24 h维持一较高水平,下调肺泡巨噬细胞RIP140的表达可明显抑制IL-1 β、TNF-α、IL-6 mRNA的表达.结论 脓毒症肺损伤大鼠肺泡巨噬细胞可通过上调RIP140表达增强肺组织炎症反应,从而参与早期急性肺损伤的炎症应答.     

硝酸甘油、地塞米松对哮喘肺泡巨噬细胞源性一氧化氮、内皮素的影响

研究哮喘患者肺泡巨噬细胞(AM)源性一氧化氮(NO)、内皮素(ET)的变化及硝酸甘油(NTC)、地塞米松(DXM)对两者的影响及机制。对15例轻、中度过敏性支气管哮喘发作期患者的AM(分为未干预组、DXM干预组、NTG干预组),7名健康自愿受试者的AM(未干预组)培养48h,用镀铜镉还原法、放射免疫法和原位杂交法分别测定AM培养上清液中NO,ET水平和iNOS-mRNA,ET-mRNA的表达。结果发现,哮喘AM iNOS-mRNA,ET-mRNA表达增强,分别导致NO,ET水平升高;NTG以直接作用的方式促进AM源性NO的产生,反馈抑制iNOS-mRNA表达并明显抑制ET mRNA的表达,降低ET的水平;DXM降低哮喘AM源性NO,ET水平及iNOS-mRNA,ET-mRNA的表达,尤以抑制iNOS-mRNA表达和降低NO水平为甚,使NO,ET处于低水平的异常状态。