The response of cultured human kidney capillary endothelium to immunologic stimuli

We have developed in culture for the first time one of the actual target cells in kidney rejection by establishing long-term cultures of human kidney capillary endothelium and have begun to explore their response to immunologic stimuli. Microvascular endothelial cells were obtained from the human kidney cortex by mechanical separation, enzyme digestion, and density gradient centrifugation. The cells were exposed to 1% PHA, a PHA-stimulated PBL supernatant, human and mouse IL-1 and human gamma-interferon. They were stained with FITC-conjugated monoclonal antibodies specific for beta 2-microglobulin, HLA-DR, and DQ and analyzed by flow cytometry (FACS-II). HKCE cells express HLA-ABC antigens during growth and confluent phases but do not express DR or DQ antigens. Only the PHA-stimulated PBL supernatant and gamma-interferon induced class II antigen expression. A change in cellular morphology was noted after 72 hr of incubation in the presence of the lectin-stimulated PBL supernatant. Our findings show that microvascular endothelial cells express selected Ia antigens only after immunologic induction. This is in agreement with studies using umbilical and aortic endothelial cells. These data suggest that a dynamic interaction takes place between activated lymphocytes whose products induce expression of class II antigens and human kidney endothelial cells.     

NK cell dysfunction with down‐regulated CD16 and up‐regulated CD56 molecules in patients with esophageal squamous cell carcinoma

NK cells can be divided into two subsets, CD56^dim and CD56^bright NK cells, based on their expression of CD56 and CD16. In the present study, we analyzed NK cell dysfunction in patients with esophageal squamous cell carcinoma (ESCC), with a particular focus on the expression of CD16 and CD56 molecules. Expression of CD16 and CD56, and the distribution of CD56^dim or CD56^bright NK cells gated on CD56(+)CD3(–) NK cells were compared between ESCC patients (n= 40) and healthy donors (n= 38). Purified NK cells were evaluated for Cetuximab‐mediated antibody‐dependent cellular cytotoxicity (ADCC) against epidermal growth factor receptor (EGFR)‐expressing ESCC cell lines. Although there were no significant differences in the distribution of CD56^dim and CD56^bright NK cells between ESCC patients and healthy donors, down‐regulated CD16 and up‐regulated CD56 were significantly observed on NK cells of ESCC patients, paralleling the impairment of Cetuximab‐mediated ADCC, in comparison with healthy donors. After patients received curative resections of ESCC, the down‐regulated CD16 and up‐regulated CD56 were significantly restored to the levels of healthy donors. Moreover, TGF‐beta1 partially contributed to down‐regulation of CD16 on NK cells. Down‐regulated CD16 and up‐regulated CD56 molecules on NK cells were observed in ESCC patients, resulting in NK cell dysfunction.     

Antibody-dependent in-vitro cytotoxicity of newborn Trichinella spiralis larvae: nature of the cells involved

The cytotoxic reaction of normal peritoneal mouse cells, containing less than 3% eosinophils, against newborn Trichinella spiralis larvae, in the presence of antibody, was studied using newborn worms less than 2 h of age, or newborn worms that had been maintained in culture at 37 degrees C for 20 h. Newborn worms 2 h old were killed, whereas 20 h newborn worms were not. The cells that initially adhered to the larvae were examined by electron microscopy. Only eosinophils adhered to 2 h newborn worms and only macrophages to 20 h ones. The attached eosinophils degranulated and died after a few hours. The macrophages that adhered to, but did not kill the 20 h newborn worms were morphologically in good state and no lysis of the larvae was observed. These results suggest that different antibody classes are involved in eosinophil and macrophage adherence, and strongly support the hypothesis that eosinophils mediate larval destruction. They also show that rapid changes are taking place after birth in the structure of the larval cuticle and that the age of Trichinella newborn worms is a major factor in the antibody-dependent cellular cytotoxicity reaction.     

实验性哮喘小鼠免疫细胞的ADCC作用

目的 :探讨支气管哮喘的发病机制。方法 :在小鼠 OVA支气管哮喘模型的基础上 ,分离哮喘鼠和正常鼠脾脏各种免疫细胞 ,包括单核 /巨噬细胞、树突状细胞、B细胞、CD3+ T细胞、CD4+ T细胞和 CD8+ T细胞 ,并以细胞增殖法和改良的 MTT比色法检测上述各种细胞的抗体依赖的细胞介导细胞毒效应 ( antibody dependentcell- mediated cytotoxicity,ADCC)。结果 :支气管哮喘小鼠上述各种细胞自然状态下的 ADCC作用均较健康对照组减弱 ( P<0 .0 5 )。结论 :免疫细胞自然状态下 ADCC作用减弱可能作为支气管哮喘免疫功能紊乱的一部分参与了哮喘的发病     

靶向抗PD-1/CD19双特异性抗体的表达与活性鉴定

目的:重组表达和纯化靶向抗PD-1/CD19双特异性抗体(bispecific antibody, BsAb)并验证其活性.方法:以pCAR1为载体,利用分子克隆技术构建抗PD-1/CD19 BsAb真核表达载体,通过PEI试剂转染哺乳动物细胞株CHO-S瞬时表达抗体.利用亲和层析法对BsAb进行纯化,用SDS-PAG...     

K562工程细胞联合IL-2扩增方案对人NK细胞扩增和活化的效果

目的：评价新建立的K562工程细胞联合IL-2扩增方案对人NK细胞扩增和活化的效果。 方法：采集健康志愿者和肿瘤患者的外周血PBMC并分离NK细胞,采用前期构建的K562工程细胞(将IL-15、4-1BBL和IL-18在白血病K562细胞上进行跨膜表达获取）联合IL-2培养方案对NK细胞进行扩增和活化,以流式细胞术检测NK细胞的扩增效果和NK细胞表面受体表达水平,CCK-8法检测扩增后NK细胞对肿瘤细胞的杀伤活性和ADCC活性,CCK-8法检测在培养方案扩增末期加入TKD多肽对NK细胞的活化效果。结果：对于健康志愿者的NK细胞,新建立扩增培养方案可使NK细胞在PBMC中的比例提高至（93±3）%;使NK细胞中活化性受体NKG2D、CD94、NKp30、NKp44和NKp46的比例分别提高60%、40%、20%、40%和63%,而抑制性受体的表达变化不大;扩增后NK细胞对白血病细胞K562、肺癌细胞A549、肝癌细胞SMMU-7721和乳腺癌细胞MCF-7的杀伤活性分别提高了19%、29%、26%和28%,其ADCC活性从（33±5.6）%上升至（65±12）%;方案中增加TKD可使NK细胞的杀伤活性从（86±4）%提高至（96±2）%。对于肿瘤患者的NK细胞,新扩增方案使其在PBMC的比例提高至（90.0±8.0）%,其对K562细胞的杀伤活性提高了17%左右。 结论： K562工程细胞联合IL-2扩增方案可高效扩增NK细胞,明显激活其杀伤活性,扩增和活化的NK细胞可满足临床治疗的需要。