The inhibitory NK cell receptor CD94/NKG2A and the activating receptor CD94/NKG2C bind the top of HLA-E through mostly shared but partly distinct sets of HLA-E residues

The human non-classical MHC class I molecule HLA-E is a ligand for both an inhibitory NK cell receptor (CD94/NKG2A) and an activating receptor (CD94/NKG2C). To identify HLA-E surface recognized by both receptors, especially to determine if both receptors recognize the same epitope, we made a series of individually Ala-substituted HLA-E proteins and analyzed their binding to CD94/NKG2A orCD94/NKG2C. Eight HLA-E mutations that significantly impaired HLA-E binding to CD94/NKG2A are all found in the top of alpha1/alpha2 domain of HLA-E. These results suggest that CD94/NKG2A binds a HLA-E surface equivalent to a NKG2D binding site on MICA. Of the eight mutations that impaired HLA-E binding to CD94/NKG2A, six significantly impaired HLA-E binding to CD94/NKG2C suggesting that CD94/NKG2C also binds a similar surface of HLA-E. Unexpectedly, the two HLA-E mutations (D69A and H155A) selectively abrogated HLA-E binding to CD94/NKG2A, not largely affected CD94/NKG2C. These results indicate that a mostly shared, but partly distinct set of HLA-E residues is discriminated by the two receptors.     

Aggregation of MHC class I molecules on a CD8+ alphabeta T cell clone specifically inhibits non-antigen-specific lysis of target cells

MHC class I molecules are target molecules recognized by TCR or NK receptors encoded in the NK gene cluster or leukocyte receptor cluster. We show that aggregation of MHC class I molecules by specific monoclonal antibodies on cytotoxic T cells, inhibits the anti-CD94 redirected lysis of P815. This inhibition is not the consequence of apoptosis or anergy of the cytotoxic T lymphocytes. In contrast, aggregation of MHC class I molecules does not inhibit either the anti-CD3 redirected cytotoxicity or the CD94-triggered up-regulation of CD25 molecules of the same T cell clone. MHC class I ligand molecules expressed by antigen presenting cells and/or T lymphocytes could therefore be able to modulate nonspecific cytotoxicity upon interaction with MHC class I molecules expressed by effector cytotoxic T lymphocytes.     

Human CD8+ intraepithelial lymphocytes: a unique model to study the regulation of effector cytotoxic T lymphocytes in tissue

Summary:  The epithelium of the human small intestine contains a large population of intraepithelial cytolytic αβ T-cell receptor (TCR) CD8αβ T lymphocytes (IE-CTLs), whose main role is to sustain epithelial integrity by rapidly eliminating infected and damaged cells. In mouse, the recognition of inducible/modified self-molecules, i.e. non-classical major histocompatibility complex (MHC) class I molecules, is mediated by the TCR and natural killer receptors (NKRs) co-expressed on the cell surface of a non-conventional autoreactive CD8αααβTCR cell subset. In contrast, in humans, the recognition of non-classical MHC class I molecules induced by stress and inflammation on intestinal epithelial cells (IECs) is principally mediated by NKRs expressed on conventional CD8αβαβTCR cells. By sensing microenvironmental signals of inflammation and stress through NKRs, IE-CTLs fine tune their TCR activation threshold. Furthermore, IE-CTLs under particular conditions, involving interleukin-15 upregulation, acquire the capacity to kill distressed intestinal epithelial cells in an antigen non-specific manner. Adaptive IE-CTLs appear hence to have autoreactive properties and modulate their immune response based on innate signals, reflecting the fitness of the tissue.     

MS患者CD94/NKG2A-bright和CD94/NKG2A-dim NK细胞亚群对IFN-β的差异性反应

目的： 分析多发性硬化（MS）进展型患者不同表型NK细胞亚群对临床主要治疗方法的反应性差异.方法： 分离患者外周血中的NK细胞, 以流式细胞术根据表面抑制性受体CD94/NKG2A表达情况分为两个亚群CD94/NKG2A-bright和CD94/NKG2A-dim.分别加入IFN-β, 测定两个亚群表面CD94/NKG2A变化及细胞增殖, 同时检测两种亚群分泌IL-10和TGF-β情况.结果： CD94/NKG2A阳性表达的NK细胞占25.5%, 其中CD94/NKG2A-bright和CD94/NKG2A-dim分别占其中的23.6%和76.4%.加入IFN-β, CD94/NKG2A-bright组增殖率明显低于CD94/NKG2A-dim组, CD94/NKG2A表达峰度变化不大.CD94/NKG2A-dim组中CD94/NKG2A表达显著增加.两个亚群分泌的IL-10和TGF-β与未刺激组相比, 均有明显差异.CD94/NKG2A-bright和CD94/NKG2A-dim组间亦有明显差异.结论： IFN-β通过诱导NK细胞CD94/NKG2A表达在非特异免疫系统中抑制NK细胞; 同时刺激IL-10 和TGF-β分泌进一步发挥对免疫系统的抑制.CD94/NKG2A-bright和CD94/NKG2A-dim对IFN-β反应有差异性.     

Expression of autophagy and ER stress-related proteins in primary salivary adenoid cystic carcinoma

Autophagy is the endogenous cellular pathway that facilitates cellular survival by maintaining energy homeostasis and macromolecular synthesis during cellular stress and nutrient deprivation. Endoplasmic reticulum (ER) stress is the process in which disruption of these physiological functions leads to an accumulation of unfolded proteins and induces the unfolded protein response (UPR). ER stress and autophagy are involved in human cancer. We investigated the expression of autophagic proteins (LC3 and beclin 1) and ER stress-related protein (GRP78) in head and neck adenoid cystic carcinoma tissue. Tissue samples from 79 cases of head and neck adenoid cystic carcinoma tissue were utilized for immunohistochemistry. LC3 expression was significantly correlated with lymph node involvement (P = .016) and TNM (P = .021). Beclin 1 expression was significantly correlated with the histological growth pattern (P = .002), the histological grade (P = .000), and longer survival (P = .000). GRP78 expression was significantly correlated with the histological growth pattern (P = .019), the histological grade (P = .019), and longer survival (P = .001). LC3 expression was positively correlated with beclin 1 expression (P = .000); LC3 and beclin 1 expressions were positively correlated with GRP78 expression respectively (P = .035) (P = .008). Our study describes the expression of LC3, beclin 1, and GRP78 in adenoid cystic carcinoma and its relationship with clinicopathologic factors and overall survival. These results suggest that LC3, beclin 1, and GRP78 may play an important role in the tumorigenesis of adenoid cystic carcinoma, and that beclin 1 and GRP78 may serve as new prognostic indicators for the outcome of patients with adenoid cystic carcinoma.     

葡萄糖调节蛋白94在人胚胎发育中的表达

目的 为探讨葡萄糖调节蛋白９４（ＧＲＰ９４）在胚胎发育中的作用,研究了１０－３１周人胎组织ＧＲＰ９４的定位及表达。方法 用ＳＰ免疫组织化学方法。结果 结肠粘膜上皮,肾小管上皮及肝细胞的胞质内,在１０周已有ＧＲＰ９４的表达,１８－２０周时表达增多,一直维持较高水平。胃粘膜上皮及心肌细胞也有较强的表达。     

内质网应激诱导人乳腺癌MDA-MB-231细胞系凋亡

目的探讨内质网应激(ERS)致雌激素受体阴性(ER-)人乳腺癌MDA-MB-231细胞系凋亡及钙激活中性蛋白酶2(calpain2)在凋亡效应中的作用。方法实验设空白对照组(DMEM)、对照组(DMEM+DMSO)和实验组,用不同浓度衣霉素(TM)诱导乳腺癌细胞不同时间,MTT法和流式细胞术检测细胞增殖抑制率及凋亡率;Western blot法检测葡萄糖调节蛋白78(GRP78)表达量,以GRP78表达最高点确定ERS模型建立,检测凋亡相关蛋白caspase4和CHOP表达以及calpain及其内源性抑制酶calpastatin的表达,用ERS抑制剂(4-PBA)和calpain抑制剂(calpeptin)分别预处理细胞2 h,观察对上述效应的影响。结果 9、12和18μmol/L TM诱导ERS对该细胞增殖有明显抑制效应,抑制率分别为33.88%±1.32%、51.51%±8.85%和55.77%±2.61%,细胞凋亡率分别为9μmol/L TM组16.70%±0.46%和12μmol/L TM组28.1%±1.09%,与对照组相比有明显差异(P0.05);9μmol/L TM诱导细胞24 h的GRP78表达上调最高(P0.01);ERS还可明显上调caspase4、CHOP和calpain表达,下调calpastatin表达(P0.01或P0.05),上述效应均能被4-PBA和calpeptin减弱或阻断(P0.05)。结论 ERS可诱导MDA-MB-231细胞凋亡,calpain2参与调控凋亡发生。     

CRIPTO promotes an aggressive tumour phenotype and resistance to treatment in hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is the third leading cause of cancer‐related death worldwide. Despite increasing treatment options for this disease, prognosis remains poor. CRIPTO (TDGF1) protein is expressed at high levels in several human tumours and promotes oncogenic phenotype. Its expression has been correlated to poor prognosis in HCC. In this study, we aimed to elucidate the basis for the effects of CRIPTO in HCC. We investigated CRIPTO expression levels in three cohorts of clinical cirrhotic and HCC specimens. We addressed the role of CRIPTO in hepatic tumourigenesis using Cre‐loxP‐controlled lentiviral vectors expressing CRIPTO in cell line‐derived xenografts. Responses to standard treatments (sorafenib, doxorubicin) were assessed directly on xenograft‐derived ex vivo tumour slices. CRIPTO‐overexpressing patient‐derived xenografts were established and used for ex vivo drug response assays. The effects of sorafenib and doxorubicin treatment in combination with a CRIPTO pathway inhibitor were tested in ex vivo cultures of xenograft models and 3D cultures. CRIPTO protein was found highly expressed in human cirrhosis and hepatocellular carcinoma specimens but not in those of healthy participants. Stable overexpression of CRIPTO in human HepG2 cells caused epithelial‐to‐mesenchymal transition, increased expression of cancer stem cell markers, and enhanced cell proliferation and migration. HepG2‐CRIPTO cells formed tumours when injected into immune‐compromised mice, whereas HepG2 cells lacking stable CRIPTO overexpression did not. High‐level CRIPTO expression in xenograft models was associated with resistance to sorafenib, which could be modulated using a CRIPTO pathway inhibitor in ex vivo tumour slices. Our data suggest that a subgroup of CRIPTO‐expressing HCC patients may benefit from a combinatorial treatment scheme and that sorafenib resistance may be circumvented by inhibition of the CRIPTO pathway. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Adaptive reconfiguration of the human NK‐cell compartment in response to cytomegalovirus: A different perspective of the host‐pathogen interaction

As discussed in this review, human cytomegalovirus (HCMV) infection in healthy individuals is associated with a variable and persistent increase of NK cells expressing the CD94/NKG2C activating receptor. The expansion of NKG2C⁺ NK cells reported in other infectious diseases is systematically associated with HCMV co‐infection. The functionally mature NKG2C^bright NK‐cell subset expanding in HCMV⁺ individuals displays inhibitory Ig‐like receptors (KIR and LILRB1) specific for self HLA class I, and low levels of NKp46 and NKp30 activating receptors. Such reconfiguration of the NK‐cell compartment appears particularly marked in immunocompromised patients and in children with symptomatic congenital infection, thus suggesting that its magnitude may be inversely related with the efficiency of the T‐cell‐mediated response. This effect of HCMV infection is reminiscent of the pattern of response of murine Ly49H⁺ NK cells against murine CMV (MCMV), and it has been hypothesized that a cognate interaction of the CD94/NKG2C receptor with HCMV‐infected cells may drive the expansion of the corresponding NK‐cell subset. Yet, the precise role of NKG2C⁺ cells in the control of HCMV infection, the molecular mechanisms underlying the NK‐cell compartment redistribution, as well as its putative influence in the response to other pathogens and tumors remain open issues.