Interferon-gamma induces dipeptidylpeptidase IV expression in human glomerular epithelial cells.

The expression of interleukin-2 receptors (IL-2R) and proliferating cell nuclear antigens (PCNA) were compared for their usefulness as markers of lymphocyte activation. Heterologous polyclonal (anti-bovine IL-2R) and monoclonal (anti-human PCNA) antibodies were used to detect the expression of these molecules on activated deer lymphocytes. Both molecules were co-expressed on blast cells which had been activated with mitogen [concanavalin A (Con A)]. There was detectable up-regulation of IL-2R expression in response to antigen [Mycobacterium bovis-derived purified protein derivative (PPD)] stimulation while PCNA expression mimicked lymphocyte transformation (LT) reactivity. PCNA expression was found to more accurately reflect both antigen- and mitogen-activated lymphocyte activation, as estimated by LT activity. The expression of PCNA was used to identify antigen reactive cells from animals exposed to M. bovis. A very low percentage (1.1 +/- 0.4%) of peripheral blood lymphocytes from non-infected animals could be stimulated to express PCNA by in vitro culture with antigen (PPD). Within the infected group both diseased and healthy, 'in-contact', animals expressed significantly higher levels of PCNA upon antigen stimulation.     

Interferon-gamma enhances rhinovirus-induced RANTES secretion by airway epithelial cells

Respiratory viruses, including rhinoviruses, infect respiratory epithelium and induce a variety of cytokines and chemokines that can initiate an inflammatory response. Cytokines, such as interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha, could enhance epithelial cell activation by inducing virus receptors. To test this hypothesis, effects of IFN-gamma or TNF-alpha on expression of intercellular adhesion molecule (ICAM)-1, rhinovirus binding, and virus-induced chemokine secretion on A549 and human bronchial epithelial cells (HBEC) were determined. The results varied with the type of cell. IFN-gamma was a stronger inducer of ICAM-1 and viral binding on HBEC, whereas TNF-alpha had greater effects on A549 cells. In addition, IFN-gamma, but not TNF-alpha, synergistically enhanced regulated on activation, normal T cells expressed and secreted (RANTES) mRNA expression and protein secretion induced by RV16 or RV49. To determine whether IFN-gamma could enhance RANTES secretion independent of effects on ICAM-1 and RV binding, HBEC were transfected with RV16 RNA in the presence or absence of IFN-gamma. RV16 RNA alone stimulated RANTES secretion, and this effect was enhanced by IFN-gamma. These results demonstrate that IFN-gamma can enhance rhinovirus-induced RANTES secretion by increasing viral binding, and through a second receptor-independent pathway. These findings suggest that IFN-gamma, by upregulating RANTES secretion, could be an important regulator of the initial immune response to rhinovirus infections.     

Human airway trypsin-like protease increases mucin gene expression in airway epithelial cells

Human airway trypsin-like protease (HAT) is a serine protease found in sputum of patients with chronic airway diseases and is an agonist of protease-activated receptor-2 (PAR-2). Results from this study show that HAT treatment also enhances mucus production by the airway epithelial cell line NCI-H292 in vitro. Histologic examination showed that HAT enhances mucous glycoconjugate synthesis, whereas the PAR-2 agonist peptide (PAR-2 AP) has no such effect. HAT, but not PAR-2 AP, enhances MUC2 and MUC5AC gene expression 23-fold and 32-fold, respectively. The proteolytic activity of HAT is required to enhance MUC5AC gene expression; the addition of the inhibitors of trypsin-like protease activity of HAT, aprotinin and leupeptin, abolishes its enhancing effect. AG1478, anti-epidermal growth factor receptor (anti-EGFR)-neutralizing antibody, and anti-amphiregulin (AR)-neutralizing antibody all inhibited the stimulatory effect of HAT. Furthermore, HAT increases AR gene expression and subsequent AR protein release, whereas PAR-2 AP shows no such effects. These results indicate that HAT enhances mucin gene expression through an AR-EGFR pathway, and PAR-2 is not sufficient for or does not directly cause HAT-induced mucin gene expression. Thus, HAT might be a possible therapeutic target to prevent excessive mucus production in patients with chronic airway diseases.     

Aquaporin-3 expression in human fetal airway epithelial progenitor cells

Airway epithelium stem cells have not yet been prospectively identified, but it is generally assumed that both secretory and basal cells have the capacity to divide and differentiate. Previously, we developed a test for progenitor cells of the human airway epithelium, relying on the transplantation of fetal respiratory tissues into immunodeficient mice. In this study, we hypothesized that airway-repopulating epithelial progenitors can be marked with surface antigens, and we screened an array of such candidate markers, including lectin ligands, the CD44 and CD166 adhesion molecules, and the aquaporin-3 (AQP3) water channel. We observed that AQP3 is selectively expressed on the surface of basal cells, allowing the separation by flow cytometry of AQP3+ basal cells and AQP3- ciliated and secretory cells. Functional evaluation of sorted cells in vivo showed that AQP3+ cells can restore a normal pseudostratified, mucociliary epithelium as well as submucosal glands. AQP3- cells are also endowed with a similar potential, although faster engraftment suggests their inclusion of more committed progenitors. These results show that stem cell candidates in the human tracheo-bronchial mucosa can be positively selected with a novel marker but also, for the first time, that epithelial progenitors exist among both basal and suprabasal cell subsets within the human airway.     

哮喘大鼠模型信号转导子和转录激活子6在气道上皮细胞表达增强

目的探讨哮喘大鼠肺组织中信号转导子和转录激活子6(STAT6)的活化规律及其意义。方法将SD大鼠随机分为对照组和哮喘组,后者根据末次激发后处死时相不同分成0、3、8、24、72、120和144 h组。采用透射电镜、细胞计数、免疫组化和图像分析等,分别观察肺组织超微结构、支气管肺泡灌洗液(BALF)中嗜酸性粒细胞(EOS)数量及STAT6蛋白表达。结果①哮喘组肺组织EOS炎性浸润较对照组明显增加;②哮喘组STAT6蛋白主要在气道上皮细胞表达;③3 h时STAT6即开始明显升高,24 h达到峰值,其后有所下降;④STAT6蛋白表达的变化与BALF中EOS绝对值及EOS%的动态变化均显著正相关(P<0.01)。结论哮喘大鼠气道上皮细胞存在STAT6的持续活化及过度表达,并与EOS的生成密切相关,提示STAT6的活化在哮喘气道炎症调控中起重要作用。     

IL—6信号转导与转录激活子活性研究

目的　探讨 Ｉ Ｌ６ 生物效应与胞内磷酸化蛋白和转录激活子活性变化之间的关系。方法　应用原位杂交和 Ａ Ｐ Ａ Ａ Ｐ 方法检测细胞内 Ｉ Ｌ６ Ｒ ｍ Ｒ Ｎ Ａ 和蛋白的表达;采用 Ｄ Ｎ Ａ 结合蛋白凝胶阻滞电泳分析 Ｉ Ｌ６ 信号转导与 Ａ Ｐ Ｒ Ｆ 活性的相关性;用免疫沉淀和 Ｓ Ｄ Ｓ Ｐ Ａ Ｇ Ｅ 观察 Ｉ Ｌ６ 信号转导中磷酸化蛋白的变化情况。结果　（１） Ｉ Ｌ６ 受体（ Ｉ Ｌ６ Ｒ） ｍ Ｒ Ｎ Ａ 和蛋白表达阳性的人骨髓瘤细胞系（ Ｓ Ｋ Ｏ００７） 体外对 Ｉ Ｌ６ 有增殖反应;（２） 胞内转录激活子活性与 Ｉ Ｌ６ 诱导时间和剂量有相关性,抗ｇｐ１３０ 单克隆抗体和酪氨酸蛋白激酶抑制剂均可特异性抑制转录激活子活性;（３） Ｉ Ｌ６ 诱导 Ｓ Ｋ Ｏ００７细胞后,胞浆内出现一组相对分子质量为（１３０ 、９０ 、５４ 、３６） ×１０３ 磷酸化蛋白,它们的磷酸化活性与转录激活子活性一样,也具有时间和剂量相关性。结论　结果提示,转录激活子和这组相对分子质量不同的酪氨酸磷酸化蛋白参与 Ｉ Ｌ６ 信号转导调节。     

Interferon-gamma protects human thyroid epithelial cells against cell-mediated cytotoxicity

Interferons have been shown to have antagonistic effects on cell-mediated cytotoxicity. In the present study, we investigated various types of immunologically mediated cytotoxic reactions with a 51chromium release assay, using human thyroid cells as targets, which were cultured in the presence and absence of interferon-gamma (IFN-gamma). Antibody-dependent cell-mediated cytotoxicity (ADCC), natural killer (NK) cell-mediated cytotoxicity, and autologous lymphocyte-mediated cytotoxicity (ALC) were measured utilizing thyroid tissue, sera and lymphocytes from patients with autoimmune thyroid disease as well as from normal subjects. ADCC, determined with sera from patients with Hashimoto's thyroiditis, was reduced by 70% when the thyroid target cells were cultured in the presence of 1000 U human IFN-gamma/ml culture medium. Similarly, natural killer cell-mediated cytotoxicity, as well as autologous lymphocyte-mediated cytotoxicity, was significantly reduced after pretreatment of thyroid target cells with 500-1000 U IFN-gamma/ml culture medium. Although it is known that IFN-gamma renders target cells resistant to NK cell-mediated lysis, this is the first report on this effect using human epithelial cells, which may have major implications for the suggested role of IFN-gamma in the induction and perpetuation of autoimmune thyroid disease.     

Manumycin inhibits cell proliferation and the Ras signal transduction pathway in human hepatocellular carcinoma cells

Manumycin was reported to have inhibitory effect on farnesyltransferase by competing with the farnesyl pyrophosphate substrate. It exhibited different antiproliferative activity in human hepatocellular carcinoma HepG2 cells, primary cultured human cardiac muscle cells and human liver cells (CLC). HepG2 cells overexpressing ras gene were more sensitive to manumycin than the other cells. The difference might be related to Ras protein levels in these cell lines. Manumycin reduced the amount of functional ras localized at the cytoplasmic membrane, resulting in blocked C-raf-1 assocation with Ras. Manumycin inhibited ERK1/2 phosphorylation in HepG2 cells without reduced expression of ERK1/2 protein. The levels of protein MKP-1 were significantly up-regulated. Our study also demonstrated that manumycin inhibited p85/PI3K and Akt phosphorylation without reduced expression of p85/PI3K and Akt, and interfered with the association of p85/PI3K and Ras. These findings indicated that manumycin interfered with Ras membrane localization, shut down the downstream pathways of Ras and inhibited cell proliferation in HepG2 cells.