Histocompatibility Complex Class I-related Chain Research Articles

A novel strategy for evasion of NK cell immunity by tumours expressing core2 O-glycans.

The O-glycan branching enzyme, core2 β-1,6-N-acetylglucosaminyltransferase (C2GnT), forms O-glycans containing an N-acetylglucosamine branch connected to N-acetylgalactosamine (core2 O-glycans) on cell-surface glycoproteins. Here, we report that upregulation of C2GnT is closely correlated with progression of bladder tumours and that C2GnT-expressing bladder tumours use a novel strategy to increase their metastatic potential. Our results showed that C2GnT-expressing bladder tumour cells are highly metastatic due to their high ability to evade NK cell immunity and revealed the molecular mechanism of the immune evasion by C2GnT expression. Engagement of an NK-activating receptor, NKG2D, by its tumour-associated ligand, Major histocompatibility complex class I-related chain A (MICA), is critical to tumour rejection by NK cells. In C2GnT-expressing bladder tumour cells, poly-N-acetyllactosamine was present on core2 O-glycans on MICA, and galectin-3 bound the NKG2D-binding site of MICA through this poly-N-acetyllactosamine. Galectin-3 reduced the affinity of MICA for NKG2D, thereby severely impairing NK cell activation and silencing the NK cells. This new mode of NK cell silencing promotes immune evasion of C2GnT-expressing bladder tumour cells, resulting in tumour metastasis.

Major histocompatibility complex class I-related chain A/B (MICA/B) expression in tumor tissue and serum of pancreatic cancer: Role of uric acid accumulation in gemcitabine-induced MICA/B expression

BackgroundMajor histocompatibility complex class I-related chain A and B (MICA/B) are two stress-inducible ligands that bind the immunoreceptor NKG2D and play an important role in mediating the cyotoxicity of NK and T cells. In this study, we sought to study MICA/B expression in pancreatic cancer and to determine whether and how genotoxic drugs such as gemcitabine can affect MICA/B expression and natural killer cytotoxity.MethodsSeven pancreatic cancer cell lines were analyzed for MICA/B expression by flow cytometry and for their sensitivity to NK-92 cell killing by a 51Cr release assay. MICA/B expression in tumor tissues and sera of pancreatic cancer was analyzed by immunohistochemical staining (IHC) and ELISA, respectively.ResultsTwo MICA/B-positive cell lines were sensitive to the cytotoxic activity of NK-92 cells. Other two MICA/B-positive cell lines and three MICA/B-negative cell lines were resistant to NK-92 cell killing. MICA/B expression was positive in 17 of 25 (68%) pancreatic ductal adenocarcinomas but not in normal pancreatic ductal epithelial cells. Serum MICA/B levels were significantly elevated in patients with pancreatic adenocarcinomas but did not correlate with the stage of pancreatic cancer and patient survival. Gemcitabine therapy led to increased serum MICA levels in 6 of 10 patients with detectable serum MICA. Allopurinol, an inhibitor of xanthine oxidoreductase that converts xanthine to uric acid, blocked uric acid production, MICA/B expression, and sensitivity to NK-92 cell killing toward a PANC-1 cancer cell line exposed to radiation and two genotoxic drugs, gemcitabine and 5-fluorouracil.ConclusionsThe levels of MICA/B expression in serum and tissue of pancreatic cancer are elevated. DNA damage-induced MICA/B expression is mediated through increased uric acid production.

Fibroblast growth factor‐2 enhances NK sensitivity of hepatocellular carcinoma cells

The roles of fibroblast growth factor-2 (FGF-2) in the hepatocellular carcinoma (HCC) development are still controversial. In this study, we investigated the expression of FGF-2 in chronic hepatitis (CH) type C patients with or without HCC and the immunoregulation of FGF-2 in NK sensitivity of HCC cells. The FGF-2 expressions were detected in the liver tissues of patients, but not in normal liver. The serum FGF-2 levels of the patients with CH, liver cirrhosis (LC) or HCC were significantly higher than those of healthy volunteers. The serum FGF-2 levels of patients decreased with the progression of chronic liver disease. HCC occurrence of LC patients with high levels of serum FGF-2 was significantly lower than that with low levels of serum FGF-2. Proinflammatory cytokines, such as IL-1β and IL-6, induced FGF-2 expressions in HCC cells and normal hepatocytes. FGF-2 stimulation resulted in increasing the expression of the membrane-bound major histocompatibility complex class I-related chain A (MICA), an NK activating molecule, and decreasing that of human leukocyte antigen (HLA) class I, an NK inhibitory molecule, on HCC cells. This did not occur with normal hepatocytes. Adding anti-FGF receptor-2 neutralizing antibody resulted in inhibiting the change of MICA and HLA class I expressions on FGF-2 stimulated HCC cells. FGF-2 stimulation on HCC cells resulted in increasing NK sensitivity against HCC cells. These findings indicate that FGF-2 produced by HCC cells or normal hepatocytes of chronic liver disease may play critical roles in eliminating HCC cells by innate immunity.

The liposome‐incorporating cell wall skeleton of Mycobacterium bovis bacillus Calmette‐Guéin can directly enhance the susceptibility of cancer cells to lymphokine‐activated killer cells through up‐regulation of natural‐killer group 2, member D ligands

• To conduct a preclinical evaluation of the ability of natural killer cells to cytolyze bladder cancer cells that were modified to show enhanced expression of natural-killer group 2, member D (NKG2D) ligands by R8-liposome-bacillus Calmette-Guéin (BCG)-cell wall skeleton (CWS) treatment. • The T24 cells and RT-112 cells were co-cultured with R8-liposome-BCG-CWS and BCG for 2, 4, or 6 h, and then the surface expression of NKG2D ligands was analyzed using TaqMan real-time quantitative RT-PCR. • Peripheral blood mononuclear cells were obtained with a conventional preparation kit, and then lymphokine-activated killer (LAK) cells were generated from these purified peripheral blood mononuclear cells via interleukin-2 stimulation. • The anti-tumour effect of LAK cells against untreated and R8-liposome-BCG-CWS co-cultured with cells of the human bladder cancer cell lines T24 and RT-112 was analyzed using the cytotoxic WST-8 assay method at 4 h of culture at various effector/target (E : T) ratios. • Major histocompatibility complex class I-related chain B (MICB) expression was increased ≈1.5-fold on T24 cells and RT-112 cells with BCG. • UL-16-binding protein (ULBP) 1 expression was also increased ≈1.5-fold on T24 cells and RT-112 cells with BCG. R8-liposome-BCG-CWS increased the surface expression of MICB 2.2-fold on T24 cells but did not increase it significantly on RT-112 cells. • ULBP1 expression was increased ≈2.2-fold on RT-112 cells, although no differences were observed between the expression of ULBP2 and 3 with R8-liposome-BCG-CWS. • T24 cells that were co-cultured with R8-liposome-BCG-CWS showed an ≈1.3-fold increase in sensitivity to cytolysis by LAK cells at an E : T ratio of 4 and RT-112 cells showed an ≈1.4-fold increase at an E : T ratio of 2. • In the present study, the induction of surface NKG2D ligands by R8-liposome-BCG-CWS rendered cancer cells more susceptible to cytolysis by LAK cells. • T24 cells and RT-112 cells, even when cultured singly in the absence of immune cells, can directly respond to R8-liposome-BCG-CWS. • The results obtained in the present study may therefore indicate a novel adoptive immunotherapy against bladder cancers.

The role of HIF-1 in up-regulating MICA expression on human renal proximal tubular epithelial cells during hypoxia/reoxygenation

BackgroundHuman major histocompatibility complex class I-related chain A (MICA) plays a dual role in adaptive and innate immune responses. Increasing evidence demonstrates that MICA is closely correlated with acute and chronic kidney allograft rejection. Therefore, understanding the activation mechanisms of MICA is important in kidney transplantation. We previously demonstrated that ischemia/reperfusion injury (IRI) could up-regulate MICA expression on mouse kidney allografts. Since hypoxia-inducible factor-1 (HIF-1) is the master regulator of cellular adaptive responses to hypoxia during IRI, here we investigate whether HIF-1 could up-regulate MICA expression and its influence on NK cell cytotoxicity.ResultsWe find that HIF-1alpha plays an important role in up-regulating MICA expression, inducing IFNgamma secretion and NK cell cytotoxicity during hypoxia/reoxygenation. First, we generated a HIF-1alphaDELTAODD-expressing adenovirus to stably and functionally express HIF-1alpha in human renal proximal tubular epithelial (HK-2) cells under normoxia conditions. HIF-1alpha over-expression in HK-2 cells induces MICA expression and enhances NK cell cytotoxic activity towards cells that express HIF-1alpha. Second, we used a hypoxia/reoxygenation cell model to simulate IRI in vitro and found that the suppression of HIF-1alpha by RNAi induces down-regulation of MICA expression and inhibits NK cytotoxicity. In antibody blocking experiments, an anti-MICA mAb was able to down-regulate NK cell cytotoxic activity towards HK-2 cells that over-expressed HIF-1alpha. Moreover, when NK cells were co-cultured with the HK-2 cells expressing MICA, which was up-regulated by over-expression of HIF-1alpha, there was a significant increase in the secretion of IFNgamma. In the presence of the blocking MICA mAb, IFNgamma secretion was significantly decreased.ConclusionsThese results demonstrate that hypoxia/reoxygenation-promoted MICA expression on HK-2 cells is through a HIF-1 pathway. The increased IFNgamma secretion and enhanced NK cell cytotoxicity was mainly due to the surface expression of MICA induced by over-expression of HIF-1alpha. This study enhances our understanding of MICA activation mechanisms during kidney transplantation and provides insights into how IRI can influence transplant outcome. Moreover, these findings might be also important for developing strategies to reduce the effect of MICA in kidney transplant outcomes in the future.

Expression and clinical significance of CD8+natural killer T cell stimulatory receptor NKG2D in peripheral blood of lung cancer patients

背景与目的NKT细胞活化性受体NKG2D及sMICA是近来肿瘤免疫研究领域的热点之一。本研究旨在观察肺癌患者外周血中CD8+NKT细胞受体NKG2D表达水平的变化，并对NKG2D及sMICA进行相关性分析，探讨它们在肺癌免疫监视中的作用及临床意义。方法选择82例初发未治疗的肺癌患者，采用流式细胞术检测外周血CD8+NKT细胞活化性受体NKG2D的表达，并以45例健康人作对照，采用酶联免疫吸附法检测肺癌患者血清中sMICA的表达，分析NKG2D与肺癌临床生物学特征的关系。结果肺癌患者外周血中CD8+NKT细胞表面活化性受体NKG2D水平均低于对照组，差异有统计学意义（P < 0.001）。随TNM分期的增加，NKG2D的表达率逐渐降低。其中Ⅳ期肺癌患者NKG2D的表达明显低于Ⅰ期-Ⅱ期及Ⅲ期患者该受体的表达，差异有统计学意义（P < 0.001）。肺Ca患者中吸烟人群外周血中CD8+NKT细胞受体NKG2D的表达较非吸烟者低，差异有统计学意义（P < 0.05）。CD8+NKT细胞受体NKG2D与sMICA呈负相关（r=-0.598, P < 0.001）。结论肺癌患者CD8+NKT细胞表面受体NKG2D在外周血中低表达，且与肺癌的分期有关；该受体的下调通过血清中sMICA上调的机制参与了肺癌以肿瘤为中心抑制免疫网络的形成；监测NKG2D及sMICA有助于了解患者的免疫功能，可为临床肿瘤的综合治疗提供依据。

Major histocompatibility complex class I-related chain A antigens stimulate endothelial cells proliferation and cause changes of their secreting functions

Objective To investigate the impact of major histocompatibility complex class Ⅰ -related chain A (MICA) antigen against endothelial cells and changes of secreting functions. Methods The endothelial cells were divided into three experimental groups (A5, A10, A25), which were stimulated with exogenous recombinant MICA antigen at 5, 10 and 25 μg/L respectively. Group A0 was added equivalent volume of PBS as control. The proliferation of endothelial cells was measured by MTF assay. The levels of metabolic product of PGI2 (6-keto-PGF1α), endothelin (ET-1), tissue plasminogen activator (t-PA) and its inhibitor (PAI) and soluble MICA (sMICA) in the supernatant of all groups were determined. Results A570 values in all experimental groups were higher than in control group. A570 values in both As and A10 groups were significantly higher than in control group and A25 group ( P ＜ 0. 05 ), but there were no significant differences between A5 and A10 groups (P ＞ 0. 05 ). The proliferation rate of 48 h was the highest in identical dosage. A570 of 48 h was 0. 458, 0. 446 and 0. 389 in A5, A10, A25 respectively. The proliferation rate at 72 h and 96 h was persistently decreased over time. There were significant differences (P ＜0. 05).The level of 6-keto-PGF1α in groups A10, A25 was 144. 6 and 132. 8 ng/L, which was lower than in group A0 (226. 5 ng/L) and A5 (232. 6 ng/L). The level of ET-1 in groups A10, A25 was 23. 6 and 25.8 ng/L,and that in group A0, A5 was 11. 8 and 10. 4 ng/L, respectively. The difference was statistically significant (P ＜ 0. 05). The level of t-PA in experimental groups were 161.2, 154. 2 and 157. 8 μg/L, and the level of PAI was 221.6, 248.5 and 252. 4 μg/L, respectively. sMICA in three experimental groups was 35.8,27. 4 and 21.8 ng/L respectively, which was lower than that in control group (352. 5 ng/L). There were significant differences between experimental groups and control group ( P ＜ 0. 05 ). Conclusion The small dosage of MICA antigen can stimulate endothelial cell persistent proliferation. Meanwhile, MICA antigen can cause the injury of endothelial cells and induce the enhanced coagulation function, the declined fibrinolytic function and the decreased level of sMICA. Key words: Major histocompatibility complex class; Endothelial cell; Proliferation; Prostacyclin; Endothelin

Quercetin Enhances Susceptibility to NK Cell-mediated Lysis of Tumor Cells Through Induction of NKG2D Ligands and Suppression of HSP70

It is known that treatments with heat shock, some anticancer drugs, and ionizing radiation increase the expression of heat-shock proteins (HSPs) and natural killer group 2D (NKG2D) ligands in tumor cells. The increased HSPs may make the tumor cells resistant to apoptosis and reduction of HSPs may make the tumor cells more susceptible to natural killer (NK)-cell mediated lysis of tumor cells. In this study, we investigated whether quercetin which has inhibitory activities against heat-shock factor, protein kinase C, nuclear factor-kappaB, and phosphatidyl inositol 3-kinase, can modulate the expression of NKG2D ligands and suppress the HSPs in tumor cells. The results of this study showed that quercetin significantly induced the expression of several NKG2D ligands including major histocompatibility complex class I-related chain B, UL16-binding protein 1, and UL16-binding protein 2 in K562, SNU1, and SNU-C4 cells. The quercetin-treated K562, SNU1, and SNU-C4 cells showed an enhanced susceptibility to NK-92 cells through induction of NKG2D ligands. This increased expression of NKG2D ligands seemed to be due to the inhibition of the nuclear factor-kappaB and phosphatidyl inositol 3-kinase pathways. The findings of this study suggest that the induced NKG2D ligands with the decrease of HSP70 protein by quercetin may provide an attractive strategy to improve the effectiveness of NK cell-based cancer immunotherapy.

The prognostic significance of soluble NKG2D ligands in B-cell chronic lymphocytic leukemia

Soluble or membrane-anchored ligands of NKG2D and their receptor have a critical role in the elimination of tumor cells and disease progression. Plasma samples of 98 patients with B-cell chronic lymphocytic leukemia (CLL) were analyzed with specific ELISA systems for soluble major histocompatibility complex class I-related chains (sMICA and sMICB) and UL-16-binding proteins (ULBP1, 2, and 3). The flow cytometric analysis of MICA on CLL cells and natural killer group 2 member D (NKG2D) receptors on NK cells was performed after thawing of frozen peripheral blood lymphocytes of CLL patients (N=51). Levels of sMICA, sMICB, and sULBP2 were significantly increased (P<0.001) compared with 48 controls, whereas sULBP1 3 were not detectable in patients and controls. Levels of sMICA>990 pg/ml (P=0.014), sMICB>200 pg/ml (P=0.0001), and sULBP2>105 pg/ml (P<0.0001) were associated with poor treatment-free survival (TFS). Neither MICA nor NKG2D expression could be related to clinical parameters. In multivariate analysis Binet stage (P=0.002), sULBP2 (P=0.002) and ZAP-70 (P=0.002) were independent predictive factors for TFS. In patients with Binet stage A, sULBP2 levels>105 pg/ml were strongly associated (P=0.0025) with poor TFS. Our data show that soluble but not membrane-anchored NKG2D ligands or receptors are of prognostic significance in CLL. Moreover, sULBP2 seems to be useful to identify early-stage patients with risk of disease progression.

Heat shock protein expression is low in intestinal mucosa in juvenile idiopathic arthritis: a defect in immunoregulation?

Objectives: Heat shock proteins (HSPs) are involved in the regulation of inflammation and in the maintenance of mucosal integrity. Their altered expression may be a marker of mucosal inflammation and also contribute to tissue injury. The small intestinal mucosa in children with juvenile idiopathic arthritis (JIA) shows signs of intestinal immune activation, such as increased intraepithelial cytotoxic lymphocyte counts. To further evaluate the characteristics of this immune activation in JIA, we have studied the expression of several HSPs, major histocompatibility complex (MHC) class I-related chain A (MICA), and the heat shock transcription factor 1 (HSF1) in intestinal biopsies from children with JIA.Methods: We studied 15 patients with JIA. Controls included 13 children without JIA, studied for various gastrointestinal (GI) symptoms, but eventually shown not to have any GI disease. The subjects were examined by endoscopy. The expression of HSP60, HSP70, MICA, and HSF1 was analysed in ileal and duodenal biopsies by using immunohistochemistry.Results: The expression levels of HSP60, MICA, and HSF1 were significantly lower in the duodenal epithelium in the JIA patients compared to the controls. MICA and HSF1 also showed lower expression in the ileal epithelium. The expression of HSP70 did not differ between the groups.Conclusions: The downregulation of HSP60, MICA, and HSF1 in small intestinal mucosa may indicate that intestinal epithelial cells show immune aberration in JIA. We speculate that the low heat shock response may play a role in the pathogenesis of JIA, interfering with mucosal integrity and local intestinal immunoregulation.

The soluble major histocompatibility complex class I-related chain A protein reduced NKG2D expression on natural killer and T cells from patients with prolactinoma and non-secreting pituitary adenoma

This study analyses aspects of the immune system in prolactinoma and non-secreting pituitary adenoma. Serum soluble major histocompatibility complex class I-related chain A protein (sMICA) concentrations were measured by enzyme-linked immunosorbent assay. NKG2D-expressing natural killer and T cells were analyzed by flow cytometry. A correlation analysis was also performed to associate sMICA levels with NKG2D expression. The expression of MICA was examined in specific tissues by use of the reverse transcription-polymerase chain reaction. A significant amount of sMICA was detected in the serum of nearly all patients. We found decreased percentage and mean fluorescence intensity of NKG2D-expressing natural killer and T cells from patients with prolactinoma and non-secreting pituitary adenoma compared to those from healthy donors. Pearson analysis showed a negative correlation between sMICA and NKG2D-expressing cells. The immune-escape of pituitary adenoma is related to the down-regulation of NKG2D and the up-regulation of its ligand MICA.

Major histocompatibility complex class I-related chains A and B (MIC A/B): A novel role in nonalcoholic steatohepatitis

Stress-induced soluble major histocompatibility complex class I-related chains A/B (MIC A/B) are increased in chronic liver diseases and hepatocellular malignancy. We investigated the impact of these molecules on liver injury, apoptosis, and fibrosis in nonalcoholic steatohepatitis (NASH). Blood and liver tissue were obtained from 40 patients with NASH undergoing bariatric surgery for obesity. The control group consisted of 10 healthy individuals. We also investigated 10 patients with nonalcoholic fatty liver (NAFL). Polymerase chain reaction was used to measure messenger RNA (mRNA) transcripts of MIC A/B, natural killer cell receptor G2D (NKG2D), CD95/Fas, and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-death receptor 5 (DR5). Apoptosis was quantified by way of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) (intrahepatic) and M30/M65 (systemic). Liver injury was assessed histopathologically and serologically (alanine aminotransferase/aspartate aminotransferase). Fibrosis was identified by Sirius red staining, quantitative morphometry, and alpha-smooth muscle actin and collagen 1alpha transcripts. Compared with controls, patients with NASH revealed significant increases in (1) NKG2D mRNA (13.1-fold) and MIC A/B mRNA (3.6-fold and 15.8-fold, respectively); (2) TRAIL-DR5 and CD95/Fas mRNA (2.7-fold and 3.6-fold, respectively); (3) TUNEL-positive hepatocytes (4.0-fold); and (4) M30 and M65 levels (4.6-fold and 3.4-fold, respectively). We found relevant correlations between MIC protein expression rates and NAS and fibrosis stages. In contrast, NKG2D and MIC A/B transcripts were attenuated in patients with NAFL compared with NASH. Histopathologically, NASH patients revealed increased NAS scores, an accumulation of natural killer cells, and 2.7-fold increased hepatic fibrosis by quantitative morphometry. Our findings suggest an important role for MIC A/B in liver injury. Therapeutic intervention aimed at reducing MIC A/B levels may beneficially affect the progression of NASH.

Sorafenib inhibits the shedding of major histocompatibility complex class I-related chain A on hepatocellular carcinoma cells by down-regulating a disintegrin and metalloproteinase 9

The ectodomain of major histocompatibility complex class I-related chain A (MICA) is shed from tumor cells, and may be an important means of evading antitumor immunity. This study investigated the roles of a disintegrin and metalloproteinase 9 (ADAM9) in the shedding of MICA in human hepatocellular carcinoma (HCC). Small interfering RNA-mediated knockdown (KD) of ADAM9 resulted in up-regulation of membrane-bound MICA expression on the HepG2 and PLC/PRF/5 cellular surfaces and down-regulation of soluble MICA levels in their culture supernatant. ADAM9 was cleaved at a site between Gln347 and Val348 of MICA in vitro. We constructed a plasmid of the MICA gene with mutation or deletion of the ADAM9 cleavage site to examine the detailed mechanism of MICA shedding by ADAM9 protease. The results suggested that MICA might be cleaved at the intracellular ADAM9-recognized cleavage site and was further cleaved at the extracellular ADAM9-independent cleavage site in HCC cells, resulting in the production of soluble MICA. Immunohistochemical analysis revealed that ADAM9 was overexpressed in human HCC compared to normal liver tissues. The cytolytic activity of natural killer (NK) cells against ADAM9KD-HCC cells was higher than that against control cells, and the enhancement of this cytotoxicity depended on the MICA/B and NK group 2, member D pathway. Sorafenib treatment resulted in decreased expression of ADAM9, increased expression of membrane-bound MICA expression, and decreased levels of soluble MICA in HCC cells. Adding sorafenib enhanced the NK sensitivity of HCC cells via increased expression of membrane-bound MICA. ADAM9 is involved in MICA ectodomain shedding in HCC cells, and sorafenib can modulate ADAM9 expression. Sorafenib therapy may have a previously unrecognized effect on antitumor immunity in patients with HCC.

Correlation between major histocompatibility complex class I-related chain A gene alleles and graft rejection in small intestine, liver and kidney transplantation

To investigate the correlation between major histocompatibility complex (MHC) class I chain-related gene A (MICA) gene alleles matching rates and graft rejection in small intestine, liver and kidney transplantation. Genome DNA were extracted from blood samples or pathological sections collected from donors and recipients of living-related transplantation, included 4 cases of small bowel transplantation, 5 cases of liver transplantation and 6 cases of kidney transplantation. The correlation between MICA alleles matching rates and acute graft rejection was analyzed following 13 MICA alleles determination by polymerase chain reaction based on sequence-specific primers (PCR-SSP). HLA zygosity of all donors and recipients was confirmed to be half-matching. The recipients displaying higher matching rates of MICA alleles with donors showed lighter clinical and pathological rejection and longer survival time. On the contrary, recipients with lower matching rates of MICA alleles with donors showed severer clinical and pathological rejection and shorter survival time relatively. Matching rates of MICA alleles has negative relevance to acute rejection, and positive relevance to survival time of recipients in small bowel, liver, and kidney transplantation.

MHC class I chain‐related gene A transmembrane polymorphism in Spanish women with breast cancer

The NKG2D-major histocompatibility complex class I-related chain A (MICA) system plays a key role in the antitumoral immune response. We studied five alleles of a microsatellite in the MICA transmembrane region; one of which (MICA-A5.1) gives rise to a truncated protein. The MICA-A5 allele was reduced in breast cancer patients compared with healthy controls (P = 0.04). Given the association between the HLA-B7 allele and the susceptibility to breast cancer in our area, we also analyzed the distribution of the frequency of the MICA alleles in human leukocyte antigen (HLA)-B7 patients compared with patients with the other alleles. The MICA-A5.1 allele was increased in HLA-B7 patients (P = 0.0003). These results suggest that the MICA-A5 allele appears to confer protection against human breast cancer and that the MICA-A5.1 appears to increase the susceptibility to breast cancer in HLA-B7 patients in our area.

Detection of the NK cell stimulatory receptor NKG2D and its soluble ligand MICA in patients with esophageal carcinoma.

Objective To investigate the expression of NKG2D peripheral blood of patients with esophageal carcinoma and expression of the soluble form of major histocompatibility complex class Ⅰ-related chain A (MICA). To analyze and discuss the function of NKG2D in anti-esophageal carcinoma mechanism and its relation with escape of immunosurveillance of cancer. Methods Flow cytometry analysis was used to detect NKG2D in the peripheral blood of 50 patients with esophageal carcinoma and 25 normal control individuals. Enzyme-linked immunoabsorbent assay was used to measure serum levels of soluble MICA. Results The expression of NKG2D were(87.25±3.06), (88.38±4.24), (92.46±1.46) respectively. Compared with the normal control individual, the expression of NKG2D in esophageal carcinoma group was significantly low; Among patients categorized according to most disease parameters tested (tumor size, grade of differentiation, regional lymph node status, disease stage), soluble MICA levels in sera did not statistically differ from those in normal control individuals. Patients with stage Ⅳ disease and/or regional lymph node metastasis, exhibited significantly higher serum levels of soluble MICA than control individuals. Conclusion The activity of NK cell and the anti-cancer cellular immunity level reduce in patients with esophageal carcinoma. The decrease of the receptor NKG2D is a reason for the descend of the the activity of NK cells. The escape of immunosurveiilance with esophagal carcinoma may correlate with down-regulate of NKG2D expression and up-regulate of sMICA. Key words: Esophageal neoplasms; Killer cells, natural; Receptors, neurokinin-2; Genes, MHCclass Ⅰ

N-Glycan Moieties in Neonatal Fc Receptor Determine Steady-state Membrane Distribution and Directional Transport of IgG

The neonatal Fc receptor (FcRn) is a major histocompatibility complex class I-related molecule known to protect IgG and albumin from catabolism and transport IgG across polarized epithelial cells in a bidirectional manner. Previous studies have shown species-specific differences in ligand binding, IgG transport direction, and steady-state membrane distribution when expressed in polarized epithelial cells. We hypothesized that these differences may be due to the additional N-glycans expressed on the rat FcRn, because N-glycans have been proposed to function as apical targeting signals, and that two of the N-glycan moieties have been shown to contribute to the IgG binding of rat FcRn. A panel of mutant human FcRn variants was generated to resemble the N-glycan expression of rat FcRn in various combinations and subsequently transfected into Madin-Darby canine kidney II cells together with human beta2-microglobulin. Mutant human FcRn clones that contained additional N-glycan side-chain modifications, including that which was fully rodentized, still exhibited specificity for human IgG and failed to bind to mouse IgG. At steady state, the mutant human FcRn with additional N-glycans redistributed to the apical cell surface similar to that of rat FcRn. Furthermore, the rodentized human FcRn exhibited a reversal of IgG transport with predominant transcytosis from an apical-to-basolateral direction, which resembled that of the rat FcRn isoform. These studies show that the N-glycans in FcRn contribute significantly to the steady-state membrane distribution and direction of IgG transport in polarized epithelia.

Role of NKG2D-Expressing NK Cells and sMICA in Immune Surveillance of Advanced Lung Cancer.

NKG2D-expressing NK cells and soluble major histocompatibility complex class I-related chain A (sMICA) is one of aroused general interests in tumor research area recently. The aim of the study is to investigate the levels of NKG2D-expressing NK cells and sMICA in peripheral blood of advanced lung cancer which are remarkably related to clinical significance and analyse the role of NKG2D-expressing NK cells and sMICA in immune surveillance. Flow cytometry was used to determine the percentage of NKG2D-expressing NK cells, T cell subsets, NK cells, and ELISA was used to mesure the levels of sMICA in peripheral blood of 115 advanced lung cancer patients and 50 healthy controls. Compared with control group, the levels of sMICA, CD8+T cells, NK cells increased, while the levels of NKG2D-expressing NK cells, CD3+ T cells, CD4+ T cells, CD4+ T/CD8+ T in experimental group decreased. NKG2D-expressing NK cells had a perfect negative correlation with sMICA (r =-0.319, P <0.05). NKG2D-expressing NK cells had positive correlation with CD4+ T cells, CD4+ T/CD8+ T and negative correlation with CD8+ T cells (P <0.05), sMICA had negative correlation with CD4+ T cells, CD4+ T/CD8+ T and positive correlation with CD8+ T cells (P <0.05), they had no significant correlation with CD3+ T cells, NK cells respectively (P <0.05). Accumulation of sMICA in serum may lead to the down-modulation of NKG2D-expressing NK which has been proposed to be a novel mechanism used by cancer cells to evade the tumor immunosurveillance. They may be potential indicators investigating immune functions and helpful in the evaluation of their happening and proceeding.

Major Histocompatibility Complex Class I-related Chains A and B (MIC A/B): A Role in the Pathogenesis of Non-alcoholic Steatohepatitis

Hepatocellular apoptosis is a key mechanism of liver injury in the pathogenesis of non-alcoholic steatohepatitis (NASH). Recently, soluble forms of major histocompatibility complex (MHC) class I-related chains A and B (MIC A/B) were reported to be increased in the sera of patients with chronic liver disease and hepatocellular malignancy. However, the role of MIC A and B in the pathogenesis of NASH has not been explored. We thus aimed to investigate the impact of these stress-induced ligands on liver injury, apoptosis and hepatic fibrosis in patients with NASH. Methodology: Blood and liver tissue were obtained from 40 patients with NASH undergoing gastric banding for obesity. The control group consisted of 10 healthy volunteers. Real-time PCR was used to measure mRNA transcripts of MIC A/B and the death receptors CD95/Fas, TRAIL-DR4 and DR5. Intrahepatic liver cell apoptosis was quantified by TUNEL assay, and on the systemic level, apoptosis was determined by the caspase-cleaved biomarker M30. Liver injury was assessed by histopathology using the NAFLD activity score (NAS) and serum ALT/AST determinations. Hepatic fibrosis was assessed by Sirius-Red staining, and mRNA transcript for α-SMA. Results: Our preliminary data from patients with NASH following gastric banding revealed significant increases (p&lt;0.05) (i) in hepatic MIC A/B mRNA by &gt;2.8; (ii) in CD95/Fas, DR4 and DR5 mRNAs; (iii) in TUNEL+ hepatocytes by &gt;4.1; and (iv) in systemic M30 levels. Consistent with this data, histological examination of livers from NASH patients revealed relevantly increased NAS scores. Serum ALT and AST values were significantly higher in NASH patients. Hepatic fibrosis was &gt;2.6 increased in patients with NASH. Conclusion: In patients with NASH, hepatocyte apoptosis and hepatic fibrosis are increased. Our findings suggest a role for MIC A/B-mediated liver injury. Thus, therapeutic intervention aimed at reducing MIC A/B may have a beneficial effect on the progression of NASH.

Two novel MICA alleles, MICA*054 and MICA*056

We report the identification of two novel major histocompatibility complex (MHC) class I-related chain A (MICA) alleles. MICA*054 has a nucleotide substitution of A to G at position 871 (codon 268), encoding an amino acid change of serine to glycine in the alpha-3 domain. MICA*056 has a nucleotide substitution at position 758 of G to C resulting in the substitution of tryptophan for serine at codon 230, also in the alpha-3 domain.