OBJECTIVES: To investigate the mechanism by which transforming growth factor‑β (TGF‑β) regulates major histocompatibility complex class I (MHC-I) expression in hepatocellular carcinoma (HCC) cells and its role in immune evasion of HCC. METHODS: HCC cells treated with TGF‑β alone or in combination with SB-431542 (a TGF-β type I receptor inhibitor) were examined for changes in MHC-I expression using RT-qPCR and Western blotting. A RNA interference experiment was used to explore the role of miR-23a-3p/IRF1 signaling in TGF‑β‑mediated regulation of MHC-I. HCC cells with different treatments were co-cultured with human peripheral blood mononuclear cells (PBMCs), and the changes in HCC cell proliferation was assessed using CCK-8 and colony formation assays. T-cell cytotoxicity in the co-culture systems was assessed with lactate dehydrogenase (LDH) release and JC-1 mitochondrial membrane potential assays, and T-cell activation was evaluated by flow cytometric analysis of CD69 cells and ELISA for TNF-α secretion. RESULTS: TGF‑β treatment significantly suppressed MHC-I expression in HCC cells and reduced T-cell activation, leading to increased tumor cell proliferation and decreased HCC cell death in the co-culture systems. Mechanistically, TGF-β upregulated miR-23a-3p, which directly targeted IRF1 to inhibit MHC-I transcription. Overexpression of miR-23a-3p phenocopied TGF‑β‑induced suppression of IRF1 and MHC-I. CONCLUSIONS We reveal a novel immune escape mechanism of HCC, in which TGF‑β attenuates T cell-mediated antitumor immunity by suppressing MHC-I expression through the miR-23a-3p/IRF1 signaling axis.
Humans ; MicroRNAs/genetics* ; Carcinoma, Hepatocellular/metabolism* ; Liver Neoplasms/metabolism* ; Interferon Regulatory Factor-1/metabolism* ; Transforming Growth Factor beta/metabolism* ; Signal Transduction ; Histocompatibility Antigens Class I/metabolism* ; Cell Line, Tumor ; Tumor Escape ; Coculture Techniques

OBJECTIVE: To detect the expression level of suppressors of cytokine signaling 3 （SOCS3） in acute lymphoblastic leukemia (ALL), and to observe the effect of over-expresson of SOCS3 in Jurkat cells on the cytotoxicity of NK cells. METHODS: The expression levels of SOCS3 mRNA in peripheral blood mononuclear cells of 20 children with ALL and 20 healthy children (normal control group) were detected by RT-PCR. The peripheral blood NK cells from healthy subjects were selected by immunomagnetic technique, and the purity was detected by flow cytometry. SOCS3 was overexpressed in Jurkat cells infected with lentivirus vector, and SOCS3 mRNA expression was detected by RT-PCR after lentivirus infection. The NK cells were co-cultured with the infected Jurkat, and LDH release method was used to detect the cytotoxicity of NK cells on the infected Jurkat cells. The concentrations of TNF-α and IFN-γ were determined by ELISA. The expression of NKG2D ligands MICA and MICB on the surface of Jurkat cells were detected by flow cytometry. Western blot was used to detect the effect of SOCS3 overexpression on STAT3 phosphorylation in Jurkat cells. RESULTS: Compared with the control group, the mRNA expression of SOCS3 in the peripheral blood mononucleated cells of ALL children was significantly decreased. The purity of NK cells isolated by flow cytometry could reach more than 70%. The expression of SOCS3 mRNA in Jurkat cells increased significantly after lentivirus infection. Overexpression of SOCS3 in Jurkat cells significantly promoted the killing ability of NK cells and up-regulated the secretion of TNF-α and IFN-γ from NK cells. The results of flow cytometry showed that the expression of NKG2D ligands MICA and MICB on Jurkat cells increased significantly after SOCS3 overexpression. Western blot results showed that overexpression of SOCS3 significantly reduced the phosphorylation level of STAT3 protein in Jurkat cells. CONCLUSION SOCS3 mRNA expression was significantly decreased in ALL patients, and overexpression of SOCS3 may up-regulate the expression of MICA and MICB of NKG2D ligands on Jurkat cell surface through negative regulation of JAK/STAT signaling pathway, thereby promoting the cytotoxic function of NK cells.
Child ; Histocompatibility Antigens Class I/metabolism* ; Humans ; Killer Cells, Natural/cytology* ; Leukocytes, Mononuclear/cytology* ; Ligands ; NK Cell Lectin-Like Receptor Subfamily K/metabolism* ; Precursor Cell Lymphoblastic Leukemia-Lymphoma/genetics* ; RNA, Messenger/genetics* ; Suppressor of Cytokine Signaling 3 Protein/metabolism* ; Tumor Necrosis Factor-alpha/metabolism*

To study the relationship between acute graft-versus-host disease (aGVHD) and the SIRT1 expression in peripheral blood CD4+T cells from patients after allogeneic hematopoietic stem cell transplantation (allo-HSCT).
 Methods: We collected 40 patients who underwent allo-HSCT from human leukocyte antigen (HLA)-identical sibling donors. SIRT1 expression level in CD4+T cells was measured by real-time PCR and Western blot. Acetylation and phosphorylation of STAT3 in CD4+T cells were detected by Western blot. The binding level between SIRT1 and STAT3 in CD4+T cells was analyzed by co-immunoprecipitation and Western blot. Over-expression of SIRT1 in aGVHD CD4+T cells, as well as STAT3 acetylation and phosphorylation were measured by Western blot. The mRNA levels of RORγt, IL-17A, IL-17F related to Th17 were detected by real-time PCR.
 Results: SIRT1 expression was significantly down-regulated, while STAT3 expression, acetylation and phosphorylation levels were significantly up-regulated in patients with aGVHD compared with patients without aGVHD. The STAT3 acetylation was positively correlated with STAT3 phosphorylation (r=0.69, P<0.01). Less SIRT1-STAT3 complexes were found in CD4+T cells from patients with aGVHD compared with patients without aGVHD. After SIRT1 over-expression in aGVHD CD4+T cells, the STAT3 acetylation and phosphorylation, and the expression of RORγt, IL-17A, and IL-17F related to Th17 were significantly down-regulated (P<0.05).
 Conclusion: SIRT1 deficiency in CD4+T cells plays a crucial role in up-regulation of STAT3 acetylation and phosphorylation, the increase of Th17 related gene expression, and induction of aGVHD after allogeneic hematopoietic stem cell transplantation.
Acute Disease ; CD4-Positive T-Lymphocytes ; metabolism ; Down-Regulation ; Graft vs Host Disease ; etiology ; metabolism ; Hematopoietic Stem Cell Transplantation ; Histocompatibility Antigens Class I ; Humans ; Interleukin-17 ; metabolism ; Nuclear Receptor Subfamily 1, Group F, Member 3 ; metabolism ; STAT3 Transcription Factor ; metabolism ; Sirtuin 1 ; deficiency ; metabolism ; Transplantation, Homologous ; Up-Regulation

Animals ; Antibodies, Monoclonal ; pharmacokinetics ; therapeutic use ; Antibodies, Viral ; therapeutic use ; Hepatitis B virus ; immunology ; Histocompatibility Antigens Class I ; metabolism ; Humans ; Macaca fascicularis ; Mice ; Protein Binding ; Protein Engineering ; Receptors, Fc ; metabolism

Killer cell immunoglobulin-like receptors (KIRs) are members of the immunoglobulin superfamily expressed on natural killer (NK) cells and a subset of T cells. Given the receptor-ligand relationship between certain KIR and human leukocyte antigen (HLA) classⅠmolecules, the KIRs are involved in the regulation of NK cell activation through conveying activating or inhibitory signals, which plays an important role in immunities involved in transplantation, tumor, infection as well as autoimmune diseases. This paper has provided a review for the research on KIR gene polymorphisms and summarized the characteristics of the sequence-based typing method for KIR genes.
HLA Antigens ; genetics ; Histocompatibility Antigens Class I ; genetics ; Humans ; Killer Cells, Natural ; metabolism ; Polymorphism, Genetic ; genetics ; Receptors, KIR ; genetics

OBJECTIVE: To investigate the mRNA and protein expression of MICA in laryngeal squamous cell carcinoma tissue and the Hep-2 cells. METHOD: Reverse transcriptase-polymerase chain reaction (RT-PCR) and Western-blot were used to detect the expression of MICA mRNA and protein levels in the Hep-2 cells and laryngeal cancer tissues. RESULT: The MICA mRNA showed higher expression in Hep-2 cells by RT-PCR. Compared with the control, the mRNA expression of MICA was significantly enhanced in laryngeal cancer tissues (t = 11.878, P < 0.01). The intensity of MICA expression is not related to the clinical stage of cancer. MICA protein demonstrated higher level expression by Western blot. The intensity of MICA protein expression was decreased with increased clinical stage in laryngeal cancer tissues. CONCLUSION The MICA mRNA showed stronger expression in Hep-2 cells and laryngeal cancer tissues. The intensity of its expression is not related to clinical stage of cancer. The MICA protein expression was strong in Hep-2 cells. The intensity of MICA protein expression was decreased with increased clinical stage in laryngeal cancer tissues. MICA may play an important role in laryngeal carcinoma process.
Carcinoma, Squamous Cell ; metabolism ; pathology ; Cell Line, Tumor ; Head and Neck Neoplasms ; metabolism ; pathology ; Histocompatibility Antigens Class I ; metabolism ; Humans ; Laryngeal Neoplasms ; metabolism ; pathology ; Squamous Cell Carcinoma of Head and Neck

Leukocyte immunoglobulin-like receptors (LILRs), also called CD85s, ILTs, or LIRs, are important mediators of immune activation and tolerance that contain tandem immunoglobulin (Ig)-like folds. There are 11 (in addition to two pseudogenes) LILRs in total, two with two Ig-like domains (D1D2) and the remaining nine with four Ig-like domains (D1D2D3D4). Thus far, the structural features of the D1D2 domains of LILR proteins are well defined, but no structures for the D3D4 domains have been reported. This is a very important field to be studied as it relates to the unknown functions of the D3D4 domains, as well as their relative orientation to the D1D2 domains on the cell surface. Here, we report the crystal structures of the D3D4 domains of both LILRB1 and LILRB2. The two Ig-like domains of both LILRB1-D3D4 and LILRB2-D3D4 are arranged at an acute angle (∼60°) to form a bent structure, resembling the structures of natural killer inhibitory receptors. Based on these two D3D4 domain structures and previously reported D1D2/HLA I complex structures, two alternative models of full-length (four Ig-like domains) LILR molecules bound to HLA I are proposed.
Amino Acid Sequence ; Antigens, CD ; chemistry ; Crystallography, X-Ray ; Histocompatibility Antigens Class I ; chemistry ; Humans ; Immunoglobulins ; chemistry ; Leukocyte Immunoglobulin-like Receptor B1 ; Membrane Glycoproteins ; chemistry ; Models, Molecular ; Peptides ; chemistry ; metabolism ; Protein Binding ; Protein Structure, Tertiary ; Receptors, Immunologic ; chemistry ; Signal Transduction

The NOD like receptors (NLRs), a class of intracellular receptors that respond to pathogen attack or cellular stress, have gained increasing attention. NLRC5, the largest member of the NLR protein family, has recently been identified as a critical regulator of immune responses. While NLRC5 is constitutively and widely expressed, it can be dramatically induced by interferons during pathogen infections. Both in vitro and in vivo studies have demonstrated that NLRC5 is a specific and master regulator of major mistocompatibility complex (MHC) class I genes as well as related genes involved in MHC class I antigen presentation. The expression of MHC class I genes is regulated by NLRC5 in coordination with the RFX components through an enhanceosome-dependent manner. And the involvement of NLRC5 in MHC class I mediated CD8+ T cell activation, proliferation and cytotoxicity is proved to be critical for host defense against intracellular bacterial infections. Nevertheless, the role of NLRC5 in innate immunity remains to be further explored. Here, we review the research advances on the structure, expression regulation and function of NLRC5.
Animals ; CD8-Positive T-Lymphocytes ; immunology ; metabolism ; Histocompatibility Antigens Class I ; genetics ; metabolism ; Humans ; Immunity, Innate ; Intracellular Signaling Peptides and Proteins ; chemistry ; genetics ; metabolism

Serum levels of soluble MHC class I-related chain A (sMICA) are related with the prognosis of various types of cancer; however, few studies on the prognostic value of sMICA in hepatocellular carcinoma (HCC) have been reported. In this study, we retrospectively investigated the relationship between sMICA levels and clinical features of advanced HCC, and we assessed the prognostic value of sMICA in advanced HCC. Furthermore, the relationship of serum sMICA levels and natural killer group 2, member D (NKG2D) expression on natural killer (NK) cells was also evaluated. We detected sMICA levels in the serum of 60 advanced HCC patients using enzyme-linked immunosorbent assay (ELISA) and measured expression levels of NKG2D on NK cells using flow cytometry. We found that serum sMICA levels in HCC patients were in the range of 0.10-6.21 ng/mL. Chi-square analyses showed that sMICA level was significantly related with only tumor size. Survival analysis showed that a high sMICA level was significantly related with poor prognosis among HCC patients. Multivariate analyses indicated that sMICA was an independent prognostic factor. In addition, the levels of CD56+NKG2D+ NK cells were within the range of 11.2%-55.4%, and correlation analyses indicated that sMICA level was negatively correlated with the level of NKG2D+ NK cells. Our results suggest that serum sMICA levels may be an independent prognostic factor for advanced HCC.
Adult ; Carcinoma, Hepatocellular ; blood ; immunology ; pathology ; Female ; Histocompatibility Antigens Class I ; blood ; Humans ; Killer Cells, Natural ; immunology ; metabolism ; Liver Neoplasms ; blood ; immunology ; pathology ; Male ; Middle Aged ; Multivariate Analysis ; NK Cell Lectin-Like Receptor Subfamily K ; metabolism ; Neoplasm Staging ; Retrospective Studies ; Survival Rate ; Tumor Burden

OBJECTIVETo evaluate the association of major histocompatibility complex class I chain related gene A (MICA) antibodies with acute rejection (AR), chronic rejection (CR) and renal function after renal transplantation.

METHODSSerum MICA antibodies were detected with ELISA before and after transplantation with also examinations of panel reactive antibodies (PRA), serum creatinine, urine, graft ultrasound, lymphocyte subsets and the pathology of graft biopsy. The study was carried out in two parts to monitor MICA antibodies in acute and chronic rejections after renal transplantation.

RESULTSIn the first part of the study 18 of the 41 recipients experienced episodes of acute rejection, and the incidence rate was markedly higher in MICA(+) group than in MICA(-) group (P<0.05). Compared with the recipients with stable renal functions, the patients with acute graft rejection showed a significantly higher positivity rate of MICA antibodies. Postoperative MICA antibody monitoring showed that MICA antibody level increased gradually 2-3 days after the occurrence of acute rejection; anti-rejection treatment lowered serum creatinine to a normal level but MICA antibodies remained positive. In the second part, 21 of 40 patients had chronic graft rejection and showed significantly higher positivity rate of MICA than the patients with stable renal functions (P<0.05). In patients with chronic rejections, the serum creatinine levels were significantly higher in MICA(+) than in MICA(-) cases (P<0.05). Graft biopsy of all MICA(+) cases showed C4d deposition.

CONCLUSIONThe status of MICA antibodies can predict the occurrence and treatment outcomes of acute rejection, and also as one of the major causes of chronic graft rejection, they affect the long-term survival of the renal grafts.

Adolescent ; Adult ; Antibodies ; blood ; immunology ; Complement C4b ; metabolism ; Creatinine ; blood ; Follow-Up Studies ; Graft Rejection ; blood ; immunology ; pathology ; HLA Antigens ; immunology ; Histocompatibility Antigens Class I ; immunology ; Humans ; Kidney ; metabolism ; physiopathology ; Kidney Transplantation ; Peptide Fragments ; metabolism ; Young Adult