The role of γδ T cells in priming macrophages to produce tumor necrosis factor-α

The secretion of tumor necrosis factor (TNF)-α from macrophages is regulated by both priming and triggering signals. We found that macrophages from mice lacking γδ T cells [T cell receptor (TCR) δ^?/- mice], which lack the gene encoding the δ chain, produced only small amounts of TNF-α in response to lipopolysaccharide (LPS) and showed a reduced level of expression of CD14. Pre-incubation of macrophages from TCR δ-/- mice with γδ T cells from their TCR δ^+/- littermates restored their capacity to produce TNF-α in response to LPS. The priming activity of γδ T cells was in part inhibited by neutralizing anti-interferon (IFN)-γ monoclonal antibodies. Collectively, these results suggest that γδ T cells play a role in priming macrophages to a steady state of activation via IFN-γ secretion, which allows them to produce TNF-α when exposed to LPS.     

An inhibitor of cytotoxic functions produced by CD8+CD57+ T lymphocytes from patients suffering from AIDS and immunosuppressed bone marrow recipients

An inhibitor of the cytotoxic functions (ICF) mediated by human immunodeficiency virus (HIV)- or HLA-specific cytotoxic T lymphocytes, natural killer and lymphokine-activated killer (LAK) cells is secreted by CD8⁺CD57^? T lymphocytes, a subset expanded during infection with HIV and after bone marrow transplantation. We previously showed an apparent molecular mass of 20–30 kDa for this soluble glycosylated concanavalin A-binding inhibitor which is distinct from known cytokines. Here, we report a characterization of the mechanism of action of this CD8⁺CD57⁺ ICF. We show that the ICF-induced inhibition of LAK cell cytolytic activity is transient, with a spontaneous recovery of cytolytic potential after 18 h. When testing interactions of ICF with a large set of cytokines we found that the ICF-mediated inhibition of cytotoxic functions is antagonized by two cytokines: recombinant interleukin (rIL)-4 and recombinant interferon (rIFN)-γ. Finally, we show that ICF acts at the level of cytolytic effector cells, where it induces a significant increase of cyclic AMP (cAMP) level. In contrast, no modification of either cell surface antigen expression or of target/effector cell conjugate formation could be evidenced. Addition of rIL-4 and rIFN-γ reverses such an increase of cAMP levels and in parallel restores the cytolytic activity. Altogether, these data demonstrate that the glycoprotein ICF produced by CD8⁺CD57⁺ cells (1) inhibits cell-mediated cytotoxicity by sensitizing cytolytic effector cells to the cAMP pathway, and (2) is part of a cytokine network controlling cell-mediated cytotoxic functions.     

Transcriptomic and Immunophenotypic Characterization of Tumor Immune Microenvironment in Squamous Cell Carcinoma of the Oral Tongue

The tumor immune microenvironment of oral tongue squamous cell carcinoma may be accountable for differences in clinical behavior, particularly between different age groups. We performed RNA expression profiling and evaluated tumor infiltrating lymphocytes (TILs) and their T-cell subsets in order to assess the functional status of oral tongue squamous cell carcinoma tumor microenvironment and detect potentially clinically useful associations. Archival surgical pathology material from sixteen oral tongue squamous cell carcinoma patients was microscopically evaluated for TIL densities. RNA was extracted from macrodissected whole tumor sections and normal controls and RNA expression profiling was performed by the NanoString PanCancer IO 360 Gene Expression Panel. Immunostains for CD4, CD8 and FOXP3 were evaluated manually and by digital image analysis. Oral tongue squamous cell carcinomas had increased TIL densities, numerically dominated by CD4 + T cells, followed by CD8 + and FOXP3 + T cells. RNA expression profiling of tumors versus normal controls showed tumor signature upregulation in inhibitory immune signaling (CTLA4, TIGIT and PD-L2), followed by inhibitory tumor mechanisms (IDO1, TGF-β, B7-H3 and PD-L1). Patients older than 44 years showed a tumor microenvironment with increased Tregs and CTLA4 expression. Immunohistochemically assessed CD8% correlated well with molecular signatures related to CD8 + cytotoxic T-cell functions. FOXP3% correlated significantly with CTLA4 upregulation. CTLA4 molecular signature could be predicted by FOXP3% assessed by immunohistochemistry (R² = 0.619, p = 0.026). Oral tongue squamous cell carcinoma hosts a complex inhibitory immune microenvironment, partially reflected in immunohistochemically quantified CD8 + and FOXP3 + T-cell subsets. Immunohistochemistry can be a useful screening tool for detecting tumors with upregulated expression of the targetable molecule CTLA4.Electronic supplementary materialThe online version of this article (10.1007/s12105-020-01229-w) contains supplementary material, which is available to authorized users.     

Binding of lipopeptide to CD14 induces physical proximity of CD14, TLR2 and TLR1

Lipoproteins or lipopeptides (LP) are bacterial cell wall components detected by the innate immune system. For LP, it has been shown that TLR2 is the essential receptor in cellular activation. However, molecular mechanisms of LP recognition are not yet clear. We used a FLAG-labeled derivative of the synthetic lipopeptide N-palmitoyl-S-[2,3-bis(palmitoyloxy)-(2R,S)-propyl]-(R)-cysteinyl-seryl-(lysyl)(3)-lysine (Pam(3)CSK(4)) to study the roles of CD14, TLR2 and TLR1 in binding and signaling of LP and their molecular interactions in human cells. The activity of Pam(3)CSK(4)-FLAG was TLR2 dependent, whereas the binding was enabled by CD14, as evaluated by flow cytometry and confocal microscopy. Using FRET and FRAP imaging techniques to study molecular associations, we could show that after Pam(3)CSK(4)-FLAG binding, CD14 and Pam(3)CSK(4)-FLAG associate with TLR2 and TLR1, and TLR2 is targeted to a low-mobility complex. Thus, LP binding to CD14 is the first step in the LP recognition, inducing physical proximity of CD14 and LP with TLR2/TLR1 and formation of the TLR2 signaling complex.     

Human monocyte-derived hemangioma-like endothelial cells: evidence from an in vitro study

BACKGROUNDS: Hemangiomas are highly prevalent in newborns and infants and can lead to severe complications. However, the pathogenesis of hemangiomas is still unknown. This study was designed to examine the potential of human monocytes to differentiate into hemangioma endothelial cells. METHODS: Purified monocytes from adult human peripheral blood were cultured under a conditional culture environment supplemented with basic fibroblast growth factor and vascular endothelial growth factor. Cells cultured for 2 weeks were subjected to histological and immunochemical examinations in order to determine the expression of specific markers for hemangioma endothelial cells. RESULTS: Monocytes cultured for 2 weeks in angiogenic medium expressed human erythrocyte-type glucose transporter protein, FcgammaRII, and several other endothelial markers, all of which are deemed specific markers for hemangioma endothelial cells. However, neither CD133 nor alpha smooth muscle actin was detected in our monocyte culture. CONCLUSION: Our data suggested that monocytes are capable of differentiating into hemangioma endothelial cells under the angiogenic stimulation from microenvironment of proliferative hemangioma.     

胃肠道间质瘤的免疫组化诊断及P53和P21表达

目的：探讨胃肠道间质瘤（GIST）的诊断及鉴别诊断。方法：GIST患50例,进行常规检查及P^53/P^21,HHF－35,vimentin,CD34免疫组化检测。结果：本组50例GIST,恶性34例,交界性10例,良性6例,以胃及小肠为多发（68％）。免疫组化以平滑肌细胞分化为主型占46％,神经源分化型占18％,平滑肌和神经双向分化型18％,未分化型24％。肌肉共同蛋白HHF－35为平滑肌细胞分化最敏感的标志物。未分化型间质瘤的免疫表型为vimentin和CD34阳性。P^53蛋白和P^21蛋白对GIST的良恶性可资鉴别。结论：尽管胃肠道间质瘤光镜下形态相似,但其免疫表型呈异源性,需要做免疫组化准确定性。     

Decreased frequency of a novel T-lymphocyte subset,CD3+CD4−CD7+CD57− T cells,in hepatitis B virus-related end-stage liver disease might contribute to disease progression

CD7 and CD57 are related to the differentiation and functional stages of CD8⁺ T cells. However, the role of their combined presence in CD8⁺ T cells in patients with chronic hepatitis B virus (HBV) infection, especially those with end-stage liver disease, remains unclear. Blood samples from healthy volunteers and patients with chronic hepatitis B were analyzed via Luminex assay and ELISA to measure plasma cytokine levels. Further, recombinant IL-22 was used to stimulate peripheral blood mononuclear cells from healthy volunteers, and the frequency of CD3⁺CD4⁻CD7⁺CD57⁻ T cells and apoptosis rates were investigated via flow cytometry. Patients with end-stage liver disease, particularly those with acute to chronic liver failure, showed decreased CD3⁺CD4⁻CD7⁺CD57⁻ T cell frequency. Furthermore, the prevalence of CD3⁺CD4⁻CD7⁺CD57⁻ T cells was negatively correlated with disease severity, prognosis, and complications (ascites). We also observed that IL-22 promoted apoptosis and brought about a decrease in the number of CD3⁺CD4⁻CD7⁺CD57⁻ T cells in a dose-dependent manner. CD3⁺CD4⁻CD7⁺CD57⁻ T cells displayed a B and T lymphocyte attenuator (BTLA)^highCD25^highCD127^high immunosuppressive phenotype and showed low interferon-γ, tumor necrosis factor-α, granzyme A, and perforin expression levels. The present findings will elucidate the pathogenesis of HBV-related end-stage liver disease and aid the identification of novel drug targets.     

CD44v6、E-Cadherin表达与CNE-2Z-F1裸鼠移植瘤转移的关系

目的　探讨CD44v6、E -Cadherin表达与人鼻咽癌裸鼠移植瘤转移的关系。方法　分别将人鼻咽癌细胞克隆株F1在体外与鼠肺块共同孵育及胸内移植 ,然后将带瘤细胞肺块 (Ⅰ组 )和胸内瘤组织块 (Ⅱ组 )各行裸鼠皮下移植 ,并与瘤细胞悬液皮下移植瘤 (Ⅲ组 )比较 ,观察各组移植瘤转移特点。采用免疫组织化学技术检测各组移植瘤回复培养细胞CD44v6和E -Cadherin表达。结果　Ⅰ组和Ⅱ组皮下移植瘤的总转移率和淋巴结转移率均高于Ⅲ组 (其中ⅠvsⅢ ,P <0 .0 5) ;肺转移仅发生在Ⅰ组。这两组移植瘤细胞CD44v6表达水平明显增高 ,其阳性细胞数分别为 (79.7± 5.7) %、(74.1± 3.1 ) % ,第 3组为 (65.6± 4.31 ) %移植瘤转移率与CD44v6高表达密切相关 ;E -Cadherin阳性细胞分别为 (41 .7± 4.9) %、(43.8± 6.4) %和 (2 7.4± 4.9) % ,Ⅰ组、Ⅱ组与Ⅲ组之间比较 ,有显著差异 (P <0 .0 5)。结论　CD44v6的过表达和E -Cadherin功能障碍 ,可能在鼻咽癌侵袭转移中起一定作用。     

CD98 regulates the phosphorylation of HER2 and a bispecific anti-HER2/CD98 antibody inhibits the growth signal of human breast cancer cells

Human epidermal growth factor receptor (HER) family proteins are currently major targets of therapeutic monoclonal antibodies against various epithelial cancers. However, the resistance of cancer cells to HER family-targeted therapies, which may be caused by cancer heterogeneity and persistent HER phosphorylation, often reduces overall therapeutic effects. We herein showed that a newly discovered molecular complex between CD98 and HER2 affected HER function and cancer cell growth. The immunoprecipitation of the HER2 or HER3 protein from lysates of SKBR3 breast cancer (BrCa) cells revealed the HER2-CD98 or HER3-CD98 complex. The knockdown of CD98 by small interfering RNAs inhibited the phosphorylation of HER2 in SKBR3 cells. A bispecific antibody (BsAb) that recognized the HER2 and CD98 proteins was constructed from a humanized anti-HER2 (SER4) IgG and an anti-CD98 (HBJ127) single chain variable fragment, and this BsAb significantly inhibited the cell growth of SKBR3 cells. Prior to the inhibition of AKT phosphorylation, BsAb inhibited the phosphorylation of HER2, however, significant inhibition of HER2 phosphorylation was not observed in anti-HER2 pertuzumab, trastuzumab, SER4 or anti-CD98 HBJ127 in SKBR3 cells. The dual targeting of HER2 and CD98 has potential as a new therapeutic strategy for BrCa.     

Stress-induced reduction in the rat mixed lymphocyte reaction is due to macrophages and not to changes in T cell phenotypes

Exposure to aversive events or Stressors modulates various aspects of immune function. We have previously reported that exposure to an acute Stressor, inescapable tail shock (IS), resulted in a shift in T cell subpopulations in rat mesenteric lymph nodes but not in cervical lymph nodes (Fleshner et al. (1992) J. Neuroimmunol. 41, 131–142). The mesenteric ratio was increased immediately after exposure to IS and was due primarily to an increase in the percent of CD4⁺ cells. The present experiments were designed to determine the relationship between the IS-associated phenotypic shift and its significance in the function of CD4⁺ T cells. The function assessed was the in vitro proliferative response to alloantigens coded for by the Major Histocompatibility Complex (MHC). Using the mixed lymphocyte reaction (MLR), we report that exposure to IS resulted in a decrease in the MLR response of cells from both cervical and mesenteric lymph nodes. Depletion of macrophages (nylon wool adherent cells) eliminated the IS-induced reduction and co-culture of macrophages (irradiation-insensitive cells) from shocked rats produced the suppression. One interpretation of these data is that exposure to IS resulted in the activation of macrophages and the release of a suppressive factor which reduced the MLR response of peripheral lymph node lymphocytes.