Accessory cell function of a human colonic epithelial cell line HT-29 for bacterial superantigens

The expression and up-regulation of cell adhesion molecules on a human colonic epithelial cell line HT-29, and the peripheral blood T lymphocyte proliferation responses to bacterial superantigens presented by this cell line were investigated, compared with peripheral blood monocytes. In HT-29 cells, there was constitutive expression of intercellular adhesion molecule-1 (ICAM-1) and lymphocyte function-associated antigen-3 (LFA-3) at a low level, but no constitutive expression of HLA-DR, LFA-1, B7-1 and B7-2 molecules. After stimulation with the supernatants of staphylococcal enterotoxin B (SEB)-stimulated peripheral blood mononuclear cells for 48 h, there was significant up-regulation of HLA-DR and ICAM-1 molecules (both >90% positive). However, this stimulation had no effect on the expression of LFA-1, B7-1, B7-2 and LFA-3 molecules. In the presence of all tested superantigens SEB, toxic shock syndrome toxin-1, and streptococcal pyogenic exotoxin A, stimulated HT-29 cells caused significant T cell proliferation. When monocytes were used as antigen-presenting cells (APC), the MoAbs against HLA-DR, B7-2 and LFA-3 showed a significant inhibition of SEB-induced T cell proliferation. Anti-ICAM-1 MoAb had no effect on this response. On the other hand, when stimulated HT-29 cells were used as APC, the MoAbs against HLA-DR and ICAM-1 significantly inhibited SEB-induced T cell proliferation. In contrast to monocytes, anti-B7-2 and anti-LFA-3 had no effect on this response. SEB could not induce HT-29 cells to produce IL-8 directly; however, SEB significantly induced the stimulated HT-29 cells to produce IL-8 in the presence of T cells. Thus these data demonstrate that the products of superantigen-stimulated T cell activation can increase the expression of HLA-DR and ICAM-1 molecules on HT-29 cells significantly. Stimulated HT-29 cells can serve as APC to bacterial superantigens. This response is an HLA-DR- and ICAM-1-dependent, but B7-2- and LFA-3-independent process, which was different from professional APC monocytes.     

Localization of antigen-presenting cells in Helicobacter pylori-infected gastric mucosa

Helicobacter pylori (HP) infection is known to induce the specific immune response in the gastric mucosa. The immune response is triggered by presentation of antigen peptides on the major histocompatibility assembly of the antigen-presenting cells (APC) with the assistance of costimulatory molecules such as B7-1 (CD80) and B7-2 (CD86). Their counter-receptors or ligands on T cells are CD28 or cytotoxic lymphocyte-associated molecule-4. The aim of the present study was to clarify the localization of APC and their relation with T cells in HP-infected human gastric mucosa. Our findings suggest that the macrophages in the lamina propria may mainly act as APC in the HP-infected gastric mucosa, and the triggered immune response might be involved in the mucosal immune response in the inflamed gastric mucosa to invasive antigens related to HP organisms.     

Antithetical effects of hemicellulase-treated Agaricus blazei on the maturation of murine bone-marrow-derived dendritic cells

We report the effects of hemicellulase-treated Agaricus blazei (ABH) on the maturation of bone-marrow-derived dendritic cells (BMDCs). ABH activated immature BMDCs, inducing up-regulation of surface molecules, such as CD40, CD80 and major histocompatibility complex class I antigens, as well as inducing allogeneic T-cell proliferation and T helper type 1 cell development. However, unlike lipopolysaccharide (LPS), ABH did not stimulate the BMDCs to produce proinflammatory cytokines, such as interleukin-12 (IL-12) p40, tumour necrosis factor-alpha, or IL-1beta. In addition, ABH suppressed LPS-induced DC responses. Pretreatment of DCs with ABH markedly reduced the levels of LPS-induced cytokine secretion, while only slightly decreasing up-regulation of the surface molecules involved in maturation. ABH also had a significant impact on peptidoglycan-induced or CpG oligodeoxynucleotide-induced IL-12p40 production in DCs. The inhibition of LPS-induced responses was not associated with a cytotoxic effect of ABH nor with an anti-inflammatory effect of IL-10. However, ABH decreased NF-kappaB-induced reporter gene expression in LPS-stimulated J774.1 cells. Interestingly, DCs preincubated with ABH and then stimulated with LPS augmented T helper type 1 responses in culture with allogeneic T cells as compared to LPS-stimulated but non-ABH-pretreated DCs. These observations suggest that ABH regulates DC-mediated responses.     

ICOS/GL50信号在T细胞依赖性体液免疫应答中的作用

研究ICOS/GL5 0信号在T细胞体外增殖、细胞因子分泌以及B细胞抗体分泌中的作用 ,进一步探讨该信号途径在机体体液免疫应答中的作用机制。采用3 H TdR检测GL5 0 L92 9转染细胞和特异性鼠抗人GL5 0单抗对T细胞体外增殖的作用 ;ELISA法检测GL5 0 L92 9转染细胞和抗人GL5 0单抗对T细胞分泌IL 2、IL 10以及ICOS L92 9转染细胞和特异性鼠抗人GL5 0单抗对PWM介导的B细胞分泌抗体的效应。结果提示GL5 0 L92 9转染细胞能够促进经抗CD3单抗活化的T细胞增值和分泌IL 10 ,而抗GL5 0单抗可以阻断这些效应。ICOS分子与B细胞上GL5 0分子的结合上调PWM介导的B细胞分泌抗体而抗GL5 0单抗则下调这一效应。这些表明 ,ICOS/GL5 0信号途径在机体T细胞依赖的体液免疫应答反应中发挥着重要的调节作用     

人OX40转基因细胞株的建立及其对树突状细胞的促分化作用

应用RT PCR方法 ,从PHA活化的扁桃体T细胞的mRNA中 ,扩增获得编码人OX4 0分子的cDNA ,并将其克隆到pMD18 T载体 ,经PCR、酶切和测序鉴定确证。进而构建pcDNA3 1 OX4 0重组真核表达载体 ,脂质体法转染L92 9细胞 ,经G4 18筛选 ,获得能稳定高表达OX4 0分子的细胞株。经OX4 0 FITC单抗标记和FCM检测 ,阳性表达率为 89 1%。转基因细胞在可溶性CD4 0L共存条件下 ,与未成熟DC共育 ,能促进DC的进一步分化成熟 ,使DC膜分子CD4 0、CD86和CD83呈上调性表达。     

Differential expression of costimulatory molecules in chronic inflammatory periodontal disease tissue

Although B cell activation and subsequent immunoglobulin production are the immunopathological features of chronic inflammatory periodontal disease, in situ expression of costimulatory molecules in humoral immunity has not been investigated. In the present study we examined the expression of CD40, CD40 ligand (CD40L), CD80, CD86, CD28 and cytolytic T lymphocyte-associated antigen-4 (CTLA-4) on lymphocytes immunohistochemically. Cryostat sections were prepared from the gingival tissue samples of 14 patients with moderate to advanced adult periodontitis. In vitro kinetics of the expression of CD40L and CTLA-4 by peripheral blood T cells and that of CD80 and CD86 by peripheral blood B cells were also investigated by flow cytometry. Positive percentage expression of CD40L, CD28 and CTLA-4, and CD40, CD80 and CD86 was calculated for the number of CD3⁺ and CD19⁺ cells, respectively. Flow cytometric analysis demonstrated that the expression of CD40L and CTLA-4 on T cells, and CD80 and CD86 on B cells of peripheral blood was up-regulated upon activation. While most T cells and B cells expressed CD28, and CD80 and CD86, respectively, in gingival tissues, the expression of CD40L and CTLA-4 was lower but highly variable between specimens. Furthermore, these two molecules seemed to be expressed reciprocally in the lesion. As both CD40L and CTLA-4 expression are induced transiently by stimulation, variability in the expression of the molecules may reflect immunological activities and participation in the regulation of B cell activation of the lesion.     

穿梭质粒pCEPmB7的构建及其对小鼠宫颈癌U14细胞体内生长的影

目的：构建可用于肿瘤基因治疗研究的基因表达载体ｐＣＥＰｍＢ７,并研究该质粒转染小鼠宫颈癌Ｕ１４细胞后的体内致瘤性变化。方法：（１）用限制性核酸内切酶ＨｉｎｄⅢ和ＸｈｏⅠ分别对质粒ｐＣＥＰ４和ｐＬＸＳＮｍＢ７进行双酶切,分别回收１０．４ｋｂ的线性ｐＣＥＰ４     

Clinicopathological significance of CD28 overexpression in adult T‐cell leukemia/lymphoma

CD28, one of the costimulatory molecules, has a pivotal role in T‐cell activation, and its expression is strictly regulated in normal T cells. Gain‐of‐function genetic alterations involving CD28 have been frequently observed in adult T‐cell leukemia/lymphoma (ATLL). These abnormalities, such as CD28 fusions and copy number variations, may not only confer continuous, prolonged, and enhanced CD28 signaling to downstream pathways but also induce overexpression of the CD28 protein. In this study, 120 ATLL cases were examined by immunohistochemistry for CD28 and its ligands CD80 and CD86, and their expression on tumor cells was semiquantitatively evaluated. CD28 was overexpressed in 55 (46%) cases, and CD80 or CD86 (CD80/CD86) was infrequently overexpressed in 12 (11%). Compared with non‐overexpressers, CD28 overexpressers showed a higher frequency of CD28 genetic alterations and had an increased number of CD80/CD86‐positive non‐neoplastic cells infiltrating tumor microenvironment. In the entire ATLL patient cohort, CD28 overexpressers showed a significantly poorer overall survival (OS) compared with non‐overexpressers (P = .001). The same was true for a subgroup who were treated with multidrug regimens with or without mogamulizumab. CD28 overexpression had no prognostic impact in the group who received allogeneic hematopoietic stem cell transplantation. In the multivariate analysis for OS, CD28 overexpression was selected as an independent risk factor. These results suggest ATLL patients with CD28 overexpression have more aggressive clinical course and are more refractory to treatment with multidrug chemotherapy. CD28 overexpression appears to be a novel unfavorable prognostic marker in ATLL patients, and further prospective studies are warranted to establish its prognostic significance.     

The modulation of B7.2 and B7.1 on B cells by immunosuppressive agents.

Several recent studies demonstrate that B7.2, but not B7.1, play an important role in allergic inflammation and IgE production. Agents that down-regulate B7.2 may therefore be of benefit for the treatment of Th2-driven allergic diseases. Our current study was carried out to investigate the effect of immunosuppressive agents, cyclosporin A (CsA) and dexamethasone, on B7.2 and B7.1 expression on B cells stimulated with the superantigen, toxic shock syndrome toxin-1 (TSST-1). The analysis of B7.2 and B7.1 on the same cells by flow cytometry demonstrated that TSST-1 up-regulated B7.2+B7.1- but not B7.1+B7.2- on B cells in a dose-dependent fashion. CsA and dexamethasone significantly down-regulated B7.2+B7.1- but up-regulated B7.2-B7.1+ B cells in the presence or absence of TSST-1 (100 ng/ml). Interestingly, the combination of CsA and dexamethasone was much more potent in the inhibition of B7.2 expression than either of these agents alone. As CD40 is known to up-regulate B7.2 expression on B cells, the mechanism of B7.2 down-regulation by CsA and dexamethasone was further studied by investigating the effect of these agents on CD40 expression on B cells. TSST-1 significantly increased CD40 expression on B cells. However, the addition of CsA or dexamethasone significantly down-regulated CD40 expression. Anti-CD40 MoAb significantly reversed the effects of CsA or dexamethasone on B7.2 and B7.1 expression, suggesting that T cell engagement of CD40 plays a role in the mechanisms by which CsA and dexamethasone acts on B cells. These data demonstrate the modulatory effect of CsA and dexamethasone on B7.2 and B7.1 expression on B cells and the potential role of CD40 in mediating this effect.     

Treatment of acute lymphoblastic leukaemia with the second generation of CD19 CAR‐T containing either CD28 or 4‐1BB

T cells modified with anti‐CD19 chimeric antigen receptor (CAR) containing either CD28 or 4‐1BB (also termed TNFRSF9, CD137) costimulatory signalling have shown great potential in the treatment of acute lymphoblastic leukaemia (ALL). However, the difference between CD28 and 4‐1BB costimulatory signalling in CAR‐T treatment has not been well elucidated in clinical trials. In this study, we treated 10 relapsed or refractory ALL patients with the second generation CD19 CAR‐T. The first 5 patients were treated with CD28‐CAR and the other 5 patients were treated with 4‐1BB CAR‐T. All the 10 patients were response‐evaluable. Three patients achieved complete remission and 1 patient with extramedullary disease achieved partial response after CD28‐CAR‐T treatment. In the 4‐1BB CAR‐T treatment group, 3 patients achieved complete remission. Furthermore, FLT‐3 ligand (FLT3LG) was highly correlated with response time and may serve as a prognosis factor. No severe adverse events were observed in these 10 treated patients. Our study showed that both CD28 CAR‐T and 4‐1BB CAR‐T both worked for response but they differed in response pattern (peak reaction time, reaction lasting time and reaction degree), adverse events, cytokine secretion and immune‐suppressive factor level.