Expressions of programmed death 1 and its ligand in acute myeloid leukemia patients and their effect on anti-tumor effect of programmed death receptor 1-positive natural killer cells in vitro / 白血病·淋巴瘤

Objective:To investigate the expression of programmed death 1 (PD-1)/programmed death ligand 1 (PD-L1) pathway in patients with acute myeloid leukemia (AML) and its relationship with clinical features and prognosis, and to examine its effect on PD-1-positive natural killer (NK) cells against AML cells in vitro.Methods:The bone marrow samples of 65 AML patients and the peripheral blood of 32 AML patients diagnosed in Affiliated Cancer Hospital of Zhengzhou University from July 2019 to December 2020 were prospectively collected, and the peripheral blood of 24 healthy people was taken as healthy control. The expression level of PD-L1 in bone marrow tumor cells and expression level of PD-1 in peripheral blood NK cells were detected by flow cytometry. The correlations of PD-1 expression in bone marrow tumor cells and PD-1 expression in NK cells with the clinicopathological features, curative effect and prognosis of patients were analyzed. Flow cytometry was used to detect the expression level of PD-L1 in AML cell line THP-1 (target cells) and the expression level of PD-L1 in NK cell line NKL (effector cells). THP-1 cells treated with and without 25 μmol/L of PD-L1 inhibitor fraxinellone were used as experimental group and control group, and co-cultured with NKL cells at different effector-to-target ratios. The apoptosis of THP-1 cells and the expression of NKG2D in NKL cells were detected by flow cytometry, the cell proliferation status was detected by CCK-8 and the cell proliferation inhibition rate was calculated; the levels of interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) in the supernatant of co-culture system were detected by enzyme-linked immunosorbent assay (ELISA).Results:The proportion of AML patients with PD-L1-positive expression in bone marrow tumor cells was higher than that in the healthy control group [38.5% (25/65) vs. 8.3% (2/24), P = 0.029]. The proportion of AML patients with PD-1-positive expression in peripheral blood NK cells was higher than that in the healthy control group [40.6% (13/32) vs. 12.5% (3/24), P = 0.035]. There were no statistical differences in sex, age, hemogram, proportion of primordial cells, risk stratification, chromosomal karyotype, gene mutation (except NPM1 gene), fusion gene and French-American-British cooperative group (FAB) typing between patients with PD-L1 positive and negative in bone marrow tumor cells and between patients with PD-1 positive and negative in peripheral blood NK cells (all P > 0.05). In relapsed/refractory patients, the proportion of patients with PD-L1-positive expression in bone marrow tumor cells was higher than that in newly treated patients [58.8% (10/17) vs. 31.2% (15/48), P = 0.045]. There was no significant difference in the proportion of patients with PD-1-positive expression in peripheral blood NK cells between relapsed/refractory patients and newly treated patients [(38.5% (5/13) vs. 42.1% (8/19), P = 0.837]. There was no statistical difference in complete remission (CR) rate between PD-L1 positive and negative patients [69.6% (16/23) vs. 74.3% (26/35), P > 0.05]. There was no statistical difference in CR rate between PD-1 positive and negative patients [66.7% (8/12) vs. 70.6% (12/17), P > 0.05]. There was no statistical difference in recurrence rate after CR between PD-L1 positive and negative patients [12.5% (2/16) vs. 19.2% (5/26), P > 0.05]. There was no statistical difference in recurrence rate after CR between PD-1 positive and negative patients [25.0% (2/8) vs. 16.7% (2/12), P > 0.05]. Flow cytometry showed that the positive rate of PD-1 in NKL cells was (67±6)% and the positive rate of PD-L1 in THP-1 cells was (85±5)%. After co-culture with NKL cells, the apoptotic rate and proliferation inhibition rate of THP-1 cells were higher in the experimental group compared with the control group, the expression of NKG2D on the surface of NKL cells was elevated, and the levels of IFN-γ and TNF-α in the co-culture supernatant were increased. Conclusions:In AML patients, the expression of PD-L1 in bone marrow tumor cells is high, and the expression of PD-1 in peripheral blood NK cells is also high. The expression of PD-L1 in bone marrow tumor cells of relapsed/refractory AML patients is higher than that of newly treated patients. Inhibition of PD-L1 expression in THP-1 cells can enhance the tumor killing activity of NKL cells in vitro. The mechanism may be that inhibition of PD-L1 expression in THP-1 cells up-regulates the expression of NKL cell activated receptor NKG2D and promotes the secretion of IFN- γ and TNF- α.

Recurrent miscarriage in North Indian population: a study of association of polymorphisms in genes coding for the natural killer: cell receptor natural killer group 2, member D and its ligand MHC class I chain-related protein A

Background: The objective of this present study was to investigate the possible association of natural killer group (NKG) receptors gene polymorphisms and MHC class I chain-related protein A (MICA) gene polymorphism with recurrent spontaneous abortion (RSA).Methods: Three single-nucleotide polymorphism (SNPs) in NKG2D gene (rs2255336, rs2617160 and rs2617170) and one SNP in MICA gene (MICA129) rs1051792 were assessed in 100 controls and 100 patients employing polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and agarose gel electrophoresis.Results: NKG2D (rs2617160) and MICA 129 (rs1051792) variants are associated with RSA risk in North Indian women.Conclusions: The NKG2D and MICA129 gene polymorphisms may influence the success of pregnancy in North Indian women population.

Expresión génica de ligandos mica, micb y ulbp (1-6) del receptor NKG2D de células natural killer y metaloproteinasas adam10, adam17 y mmp14 en lineas celulares de cáncer de cervical / Gene expression of mica, micb and ulbp (1-6) ligands of the NKG2D receptor of natural killer cells and metalloproteinases adam10, adam17 and mmp14 in cells lines of cervical cancer

RESUMEN El CCU es la segunda causa de muerte en mujeres de nuestro país. Dentro de los primeros mecanismos de defensa del hospedero se encuentra la respuesta inmune de las células NK y su función lítica a expensas de su receptor activador NKG2D, el cual posee como ligandos mica, micb y ulbp (1-6), los cuales se expresan en células transformadas y/o infectadas por virus. Uno de los mecanismos de evasión por parte de la célula tumoral es el clivaje de estas proteínas a través de metaloproteinasas como adam10, adam17 y mmp14. Se analizó la expresión de estos ligandos y metaloproteinasas mediante PCR tiempo real, en lineas celulares de referencia para cáncer cervical como HeLa (positiva para VPH-18) y C33A (negativa para VPH). Se obtuvieron valores representativos de expresion relativa genica con diferencias significativas asi: mmp14 en linea HeLa (p= 0.006); y mica y ulbp-3 en la linea C33A (p= 0.020 y p=0.003 respectivamente). Por lo tanto, se podría sugerir que la expresión de mmp14 se encuentra posiblemente involucrados con la presencia de VPH causante del cancer cervical y la respuesta inmunne innata desarrollada.

ABSTRACT Cervical cancer is the second leading cause of death in women in our country. Within the first host defense mechanisms is the immune response of NK cells and their lytic function at the expense of its NKG2D receptor activator which has as ligands mica, micb and ulbp (1-6), which are expressed in transformed cells and / or virally infected. One of the mechanisms of evasion by the tumor cell is the cleavage of these proteins through metalloproteinases as adam10, adam17 and mmp14. We analyzed the expression of these ligands and metalloproteinases by real time PCR, in reference to cell lines HeLa cervical cancer (positive for HPV-18) and C33A (negative for HPV). We obtained representing relative gene expression with significant differences from the other lines of study as follows: mmp14 in HeLa (p = 0.006); and mica and ulbp-3 in C33A (p = 0.020 and p = 0.003 respectively). Thus one might suggest that the expression of mmp14 is possible involved with HPV presence causing high risk of cervical cancer and innate inmunne response developed.

The effect of entinostat on NK killing effect on non-small cell lung cancer in vitro / 中国肿瘤临床

Objective: To investigate the Effects of entinostat on the expression of NKG2D ligands in the non-small cell lung cancer (NSCLC) cell lines, A549 and HCC-827, and to detect the effect of entinostat-mediated NK cell killing of A549 and HCC-827 cells. Meth-ods: The effect of entinostat on A549 and HCC-827 cell proliferation was measured by MTT assay. Flow cytometry was used to detect the expression of NKG2D ligands. mRNA levels of the ligands were detected by RT-PCR . The level of soluble MICA in cell culture super-natant was evaluated by ELISA. The cytotoxicity of NK cells against A549 and HCC-827 cell lines (treated with entinostat) was assessed using lactate dehydrogenase release assay. Results: Entinostat showed a time-and dose-dependent inhibition effect on the prolifera-tion of A549 and HCC-827 cell lines. The expression of NKG2D ligands and mRNA transcription levels of MICA and MICB were en-hanced after treatment with 0.5, 1μmol/L entinostat for 48 h. The soluble MICA level in A549 cell culture supernatant was increased by 1μmol/L entinostat. The sensitivity of HCC-827 cells to NK cells was enhanced upon treatment with 0.5, 1μmol/L entinostat. Con-clusions: entinostat enhanced the killing effect of NK cells on non-small cell lung cancer cells by up-regulating the expression of NKG2D ligands. This provides a new method and theory for the treatment of NSCLC.

Immunoreceptor NKG2D and its ligand-mediated cellular immunity and its relationship with diseases / 国际儿科学杂志

Natural killer (NK) cells play an important role in host defense.NKG2D is expressed as a homodimer on the surface of NK cells and is an important lymphocyte activating receptor.NKG2D ligands are negatively or poorly expressed on the surface of normal cells,but when the cells are in a state of oxidative stress,malignant transformation,autoimmune or inflammatory stimulation,these ligands will be induced on the cell surface.The NKG2D receptor and its ligand-mediated cellular immune response play an important role in immune surveillance of tumor cells,and at the same time it is also closely related to certain autoimmune diseases.

Advances in antitumor research of nk cells based on drug therapy / 中国药学杂志

NK (natural killer) cells are a kind of cytotoxic lymphocytes which have difference from T and B cells. Many NK cell related antitumor immunotherapies have been applied in clinical treatment, such as transfusion of adoptive NK cells, NK cell stimulating cytokines, chimeric antigen receptor-NK (CAR-NK) cells and NK cell inhibitory receptor inhibitors. In addition, the therapy of using drugs to strengthen the anti-tumor effect of NK cells also has a good prospect. There has been many reports on the drug promoting NK cell recognition and killing of tumor cells. The mechanisms are mainly related with modulating the expression of natural-killer group 2 member D (NKG2D) ligands, natural cytotoxic receptor (NCR) ligand, NKG2D receptor and so on, and increasing the release of NK cell stimulating cytokines. In this paper, recent advances in drug therapy based NK cell anti-tumor researches are reviewed, to provide references for the future researches.

Serum thymidine kinase 1 and soluble NKG2D ligand predict prognosis of colorectal cancer patients after operation / 中华普通外科杂志

Objective To investigate the values of serum thymidine kinase 1 (TK1) and soluble NKG2D (natural killer cell group 2D) ligand (soluble major histocompatibility complex class Ⅰ-related chain A,sMICA) in predicting the prognosis of colorectal cancer patients undergoing radical resection.Methods 45 patients and 45 healthy subjects were included.Perioperative serum TK1 and NKG2D ligand levels were measured in 45 patient and 45 healthy controls.Patients were divided into high TK1 group and low TK1 group,and high sMICA group and low sMICA group according to the ROC.Results Perioperative TK1 were (4.42 ± 1.42) and (2.98 ± 0.54) pmol/L,sMICA were (135 ± 79) and (100 ± 81)pg/ml,which were significantly higher than those in healthy controls (P =0.000).The postoperative TKI and sMICA levels decreased significandy (P =0.000 and 0.042).The 3 and 5 years cumulative survival rates in the high TK1 group were 84％ and 34％,compared with that of 90％ and 75％in the low TK1 group (P =0.023).The 3 year and 5 year cumulative survival rates in high sMICA group were 61％ and 31％,compared with 71％ and 52％ in low sMICA group (P =0.148).Conclusion Patients serum thymidine kinase levels were negatively corelated with the prognosis of colorectal cancer after radical resection.

Antígenos MICA y trasplante / MICA antigens and transplantation

Los antígenos leucocitarios humanos (HLA, del inglés human leukocyte antigens), codificados por los genes del complejo principal de histocompatibilidad (MHC, del inglés m ajor histocompatibility complex), actúan como inductores de las respuestas inmunitarias en el trasplante; sin embargo, los productos de los genes relacionados a cadenas MHC clase I (MIC, del inglés MHC class I chain-related genes), constituyen también uno de los blancos del rechazo. La familia de los genes MIC consta de siete miembros, de los cuales solo MICA y MICB son funcionales. Los transcriptos son glicoproteínas de superficie celular de 62 kDA que presentan homología en su secuencia con las moléculas HLA clase I y cuya función está relacionada con la inmunidad innata. En los órganos trasplantados ocurre un incremento en la expresión de los antígenos MICA como una señal temprana de "peligro" debido al trauma quirúrgico y la isquemia. Esta sobrexpresión antigénica puede llevar al rechazo mediado por anticuerpos anti-MICA que activan el complemento y por un incremento de la citotoxicidad debido a la estimulación en los linfocitos citolíticos naturales (NK, del inglés natural killer) y los linfocitos CD8+ &+ αß y γδ, del receptor conocido como NKG2D (NK grupo 2 miembro D(AU)

Human leukocyte antigens (HLA), encoded by major histocompatibility complex (MHC) genes, act as inducers of immune responses in transplantation. However, the products of the genes related to MHC class I chains (MIC) are also one of the targets of rejection. The family of MIC genes consists of seven members, of which only MICA and MICB are functional. Transcripts are cell surface glycoproteins of 62 kDa that exhibit homology in sequence with HLA class I molecules and whose function is related to innate immunity. In transplanted organs an increase in the expression of MICA antigens occurs as an early sign of "danger" due to surgical trauma and ischemia. This antigenic overexpression can lead to rejection mediated by complement-activating anti-MICA antibodies and by increased cytotoxicity due to stimulation in natural killer (NK) lymphocytes and CD8 + + αß and γδ lymphocytes. Receptor known as NKG2D (NK group 2 member D)(AU)

Role of TRAIL pathway and NKG2D pathway in double negative T cell cell anti-tumor / 国际外科学杂志

Double negative T cell is a rare type of cells that regulates immune responses in a way that is unique.In the process of adoptive immunotherapy,it gradually becomes a hot spot,because of its obvious anti-tumor effect.However,the mechanism by which DNT cell kills tumor cell is not clear.Tumor necrosis factor-related apoptosis-inducing ligand(TRAIL) and natural-killer group 2,member D (NKG2D) pathway is a more important part of its killing mechanism,and NKG2D receptor can also regulate the production of soluble TRAIL,making the two channels to establish a certain link.In this paper,the effect of Double negative T cell and the mechanism of TRAIL and NKG2D pathway in DNT cell's anticancer were described.

Effect of ionizing radiation on the NKG2D ligand expression on the tumor cell SCC25 surface and the killing effect on tumor cells / 国际肿瘤学杂志

Objective To investigate the effect of ionizing radiation on the expression of NKG2D ligand on the surface of oral squamous cell carcinoma cell line SCC25 and its cytotoxicity to tumor cells.Methods When SCC25 cells were cultured into logarithmic growth phase,they were randomly designed as control (without treatment) and experimental group (2 Gy ionizing radiation treatment) by drawing lots.Flow cytometry was used to detect the expressions of NKG2D ligands major histocompatibility complex class Ⅰ chainrelated molecule (MIC)A,MICB,UL16 binding protein (ULBP)1 on the surface of SCC25 in the control group and the experimental group cultured for 24 h.Real-time fluorescence quantification polymerase chain reaction (RT-PCR) was used to detect the changes of NKG2D ligand mRNA expression on SCC25 cell surface after 24 h culture in the experimental group and the control group.The cells were prepared and divided into blank control group (NC),2 Gy ionizing radiation group (R),NK1 group (target ratio was 5 ∶ 1),NK2 group (target ratio was 20 ∶ 1),NK1 + R group (target ratio was 5 ∶ 1,2 Gy ionizing radiation),NK2 + R group (target ratio was 20 ∶ 1,2 Gy ionizing radiation).After each group was cultured for 24 h,the killing abilities of ionizing radiation and natural killer (NK) cells to oral squamous cell carcinoma SCC25 cells were detected by CCK8.Results Flow cytometry experiment showed that,among the NKG2D ligands,the MICA fluorescence values of experimental group and control group were respectively 21.04 ± 0.39,22.90 ± 0.40 (t =2.465,P =0.069),MICB fluorescence values were 27.58 ± 0.50,29.83 ± 1.05 (t =1.936,P =0.125),and ULBP1 fluorescence values were 21.04 ± 0.40,21.78 ± 0.50 (t =1.154,P =0.313).This indicated that after ionizing radiation on SCC25,the NKG2D ligand MICA,MICB,ULBP1 expression increased slightly,but the differences were not statistically significant.RT-PCR indicated that mRNA expressions of MICB,ULBP1 were significantly different between the control group and the experimental group (t =18.334,P =0.000;t =6.381,P =0.008).The expressions of the experimental group were respectively 6.49,1.64 times as those of the control group.The results of CCK8 showed that,there was a significant difference in cell killing ability among NK1 group,NK2 group and NC group (F =344.600,P =0.000),suggesting that NK cells could kill tumor cells,and the higher ratio of NK cells and SCC25,the stronger killing effect.The comparison between R group and NC group showed that the difference in cell killing ability was not statistically significant (P =0.567).NK1 + R group and NK1 group were compared and the difference was not statistically significant (P =0.915).There was no significant difference between NK2 + R group and NK2 group (P =0.678).This showed that the killing effect of ionizing radiation was weak.Conclusion Ionizing radiation can increase the mRNA expression of NKG2D ligands MICB and ULBP1.This may provide a new way for tumor immunotherapy.The killing effect of ionizing radiation on cells is not obvious.It may be related to low radiation dose and only 24 h for cell culture.

Effect of ionizing radiation on the NKG2D ligand expression on the tumor cell SCC25 surface and the killing effect on tumor cells / 国际肿瘤学杂志

Objective To investigate the effect of ionizing radiation on the expression of NKG2D ligand on the surface of oral squamous cell carcinoma cell line SCC25 and its cytotoxicity to tumor cells.Methods When SCC25 cells were cultured into logarithmic growth phase,they were randomly designed as control (without treatment) and experimental group (2 Gy ionizing radiation treatment) by drawing lots.Flow cytometry was used to detect the expressions of NKG2D ligands major histocompatibility complex class Ⅰ chainrelated molecule (MIC)A,MICB,UL16 binding protein (ULBP)1 on the surface of SCC25 in the control group and the experimental group cultured for 24 h.Real-time fluorescence quantification polymerase chain reaction (RT-PCR) was used to detect the changes of NKG2D ligand mRNA expression on SCC25 cell surface after 24 h culture in the experimental group and the control group.The cells were prepared and divided into blank control group (NC),2 Gy ionizing radiation group (R),NK1 group (target ratio was 5 ∶ 1),NK2 group (target ratio was 20 ∶ 1),NK1 + R group (target ratio was 5 ∶ 1,2 Gy ionizing radiation),NK2 + R group (target ratio was 20 ∶ 1,2 Gy ionizing radiation).After each group was cultured for 24 h,the killing abilities of ionizing radiation and natural killer (NK) cells to oral squamous cell carcinoma SCC25 cells were detected by CCK8.Results Flow cytometry experiment showed that,among the NKG2D ligands,the MICA fluorescence values of experimental group and control group were respectively 21.04 ± 0.39,22.90 ± 0.40 (t =2.465,P =0.069),MICB fluorescence values were 27.58 ± 0.50,29.83 ± 1.05 (t =1.936,P =0.125),and ULBP1 fluorescence values were 21.04 ± 0.40,21.78 ± 0.50 (t =1.154,P =0.313).This indicated that after ionizing radiation on SCC25,the NKG2D ligand MICA,MICB,ULBP1 expression increased slightly,but the differences were not statistically significant.RT-PCR indicated that mRNA expressions of MICB,ULBP1 were significantly different between the control group and the experimental group (t =18.334,P =0.000;t =6.381,P =0.008).The expressions of the experimental group were respectively 6.49,1.64 times as those of the control group.The results of CCK8 showed that,there was a significant difference in cell killing ability among NK1 group,NK2 group and NC group (F =344.600,P =0.000),suggesting that NK cells could kill tumor cells,and the higher ratio of NK cells and SCC25,the stronger killing effect.The comparison between R group and NC group showed that the difference in cell killing ability was not statistically significant (P =0.567).NK1 + R group and NK1 group were compared and the difference was not statistically significant (P =0.915).There was no significant difference between NK2 + R group and NK2 group (P =0.678).This showed that the killing effect of ionizing radiation was weak.Conclusion Ionizing radiation can increase the mRNA expression of NKG2D ligands MICB and ULBP1.This may provide a new way for tumor immunotherapy.The killing effect of ionizing radiation on cells is not obvious.It may be related to low radiation dose and only 24 h for cell culture.

Significance of natural killer cell G2D expression and activation in patients with different immune status of chronic hepatitis B virus infection / 中华传染病杂志

Objective To investigate the differences of expression and activation of natural killer (NK) cell G2D (NKG2D) in patients with different immune status of chronic hepatitis B virus (HBV) infection, and to explore the significance of NKG2D-mediated immune injury in HBV infection.Methods Fifteen chronic HBV carriers (immune tolerance),15 chronic hepatitis B (CHB, immune activation) patients, 15 HBV-related acute/subacute-on-chronic liver failure (HBV-ACLF, immune over-activation) patients were enro1led in this study from January 2010 to December 2011 in the Third Hospital of Hebei Medical University.The frequencies of NK cells and NKG2D+ NK cells in peripheral blood mononuclear cells (PBMC) were detected by flow cytometry.The NKG2D mRNA expressions were measured by real-time fluorescent quantitative polymerase chain reaction.Localization and hemi-quantitative analysis of NKG2D+ cells in liver tissue were performed by immunohistochemistry staining.Concentrations of serum interferon(IFN)-γ, tumor necrosis factor(TNF)-α, perforin and granzyme B were quantified by enzyme 1inked immunosorbent assay (ELISA).Normally distributed continuous variables were analyzed using one-way analysis of variance (ANOVA), followed by Student-Newman-Keuls q test for evaluating variances between each two groups.For non-normally distributed data or heterogeneity of variance, differences between groups were analyzed using nonparametric Kruskal-Wallis H test, followed by Nemenyi test for pairwise comparisons.Pearson chi-square test was used to analyze categorical variables.Results The percentages of NK cells in PBMC were (13.58±3.24)% in healthy controls, (5.42±2.18)% in chronic HBV carriers, (7.92±2.85)% in HBV-ACLF group and (8.43±2.92)% in CHB group.The percentage of NK cells in PBMCs was lower in each chronic HBV-infected group compared with healthy controls (F=22.04, P<0.05).The frequency of NKG2D+ NK cells in HBV-ACLF group (18.92±5.85)% was the highest, followed by CHB group (12.85±3.39)%, healthy controls (8.45±2.86)%, and chronic HBV carriers (3.36±1.05%), with the statistically significant differences between each two groups (H=46.09, P<0.01).Intrahepatic NKG2D mRNA expression and NKG2D+ cells density were highest in HBV-ACLF group (6.58±1.86 and 30.69±6.67, respectively), followed by CHB group (3.25±0.95 and 17.36±4.13, respectively) and chronic HBV carriers (0.69±0.20 and 3.16±1.24, respectively), with the statistically significant differences between each two groups (H=52.10 and 52.73 respectively, both P<0.01).The similar patterns were observed in serum IFN-γ, TNF-α, perforin and granzyme B concentrations.Conclusions NKG2D expresses variously in patients with different immune status of chronic HBV infection.Activation of NKG2D may take part in the immune pathogenesis of chronic HBV infection.

Baicalin promotes the expression of NKG2D ligands on BCG-infected macrophages and enhances the cytotoxic activity of NK cells / 中华微生物学和免疫学杂志

Objective To investigate the effects of BCG ( Bacillus Calmette-Guerin) infection on NKG2D (natural killer group 2, member D) ligands (MICA, MICB, ULBP1 and ULBP2) expressed on macrophages and to further analyze the effects of baicalin on these NKG2D ligands and the cytotoxic activity of NK cells. Methods PMA ( phorbol 12-myristate 13-acetate) was used to induce the differentiation of THP-1 cells into macrophages. The THP-1-derived macrophages were infected with BCG and then treated with baicalin. The expression of MICA, MICB, ULBP1 and ULBP2 at mRNA and protein levels were meas-ured by real-time PCR and Western blot assay. The BCG-infected macrophages were co-cultured with NK cells derived from human PBMC for 4 h. Real-Time Cell Analyzer ( RTCA DP) was used to evaluate the cy-totoxic activity of NK cells. Results The expression of MICA, MICB, ULBP1 and ULBP2 at mRNA level and the expression of MICA and ULBP1 at protein level were upregulated after infecting the macrophages with BCG. The expression of MICA and ULBP1 at mRNA and protein levels and the killing activity of NK cells were significantly enhanced after treating the BCG-infected macrophages with baicalin (1 mg/L) for 72 h. Conclusion BCG infection could induce the expression of NKG2D ligands on human macrophages, but could not effectively active the NK cells. Baicalin could enhance the cytotoxic activity of NK cells by further up-regulating the expression of NKG2D ligands on BCG-infected macrophages.

Effect of cisplatin and paclitaxel on the cytotoxicity of cytokine-in-duced killer cells on esophagus carcinoma and its molecular mecha-nisms / 中国肿瘤临床

Objective:To explore the effect of paclitaxel (PTX) and cisplatin (DDP) on the expression of NKG2D ligands of hu-man esophagus carcinoma cell EC9706 and on the cytotoxicity of cytokine-induced killer (CIK) cells, as well as to discuss its molecu-lar mechanisms. Methods: The half maximal inhibitory concentration (IC50) values of PTX and DDP against EC9706 cells for 24 h were measured by MTT assay. The expression levels of NKG2D ligands (MICA, MICB, ULBP1, ULBP2, and ULBP3) on the EC9706 cell surface before and after 24 h culture with 1/2 IC50 of PTX or DDP were assayed by flow cytometry. Cytotoxicity of CIK cells against EC9706 cells before and after 24 h culture with 1/2 IC50 PTX or DDP was analyzed by lactate dehydrogenase release assay at an effector to target cell ratio (E:T) of 20:1 and 30:1, respectively. The expression levels of DNA damage repair genes (ATM, ATR, CHK1, CHK2, and p53) of EC9706 cells before and after 24 h incubation with 1/2 IC50 PTX or DDP were detected by quantitative fluorescent PCR. Results:The IC50 values of PTX and DDP were 10 and 5μg/mL, respectively. MICB, ULBP2, and ULBP3 on EC9706 cells were upregulated after 24 h culture with 1/2 IC50 PTX (P0.05), whereas ATM, ATR, CHK1, and CHK2 were over-expressed after 24 h treatment with 1/2 IC50 DDP (P<0.05). Conclusion:PTX or DDP can enhance the susceptibility of EC9706 cells to CIK cell-mediated lysis by upregulating the expression of NKG2D ligands through activating DNA damage repair genes.

Analysis of correlation between NKG2D receptor pathway and the mechanism of DNT cell' killing effect / 国际外科学杂志

NKG2D is an activating receptor expressed on various immune cells' surfaces.Currently,eight ligands have been found,they can be divided into two kinds:MHC Ⅰ related protein and ULBP molecules.NKG2D receptors and their respective ligands differentially exist in normal and cancerous tissues.So far,many researchers have found that a lot of immune cells' anti-tumor mechanism is mediated by NKG2D receptor and it can't be replaced in the resistance to the development of carcinoma.adoptive immunotherapy is a new method of treating cancer that propose to find tumor-specific cytotoxicity immune cells and enlarge the amount of it,then injected into human body to resist tumor.Recently,many researchers have found that DNT cells (CD4-CD8-T 细胞)have the function of anti-tumor;sadly its mechanisms are unclear.After the summary of NKG2D receptor-mediated pathway of destruction,this paper try to analyze its relevance to DNT cells' killing effect,on the purpose of finding the Killing mechanism of DNT cells that may existed.

Advances in double negative T cells in tumor immunotherapy / 国际外科学杂志

Double negative T cells,namely DNT cells,are specific regulatory cells with TCR+ CD3 + CD4-CD8-in cell phenotype,divide into αβ T cell and γδ T cell,play a very important role in regulating immunoreaction.Previous research restricted to the treatment of autoimmune disease and organ transplantation immunity.Recently dnt cells adoptive immunotherapy,as a new anti-tumor method,gradually come into public view.This review mainly focuses on the source of DNT cells,their anti-tumor effect,as well as the possible mechanism.

Natural killer cell receptors and human cytomegalovirus infection / 国际儿科学杂志

Human cytomegalovirus(HCMV)infection is quite prevalent in population. HCMV triggers important disorders in pregnant women,immune-compromised individuals and organ-transplant patients. For dec-ades,more and more scholars believe that NK cells are important immune cells against HCMV. The activity of NK cells largely depends on the balance between the signals transducted by inhibitory receptors and activatory re-ceptors,therefore the study of NK receptors is of great importance. It may be helpful to the basic research and clinical treatment of HCMV infection. Here,this paper reviews the changes and the molecular mechanism of NK receptors in the control of HCMV infection.

Effect of inhibition of TGF-β1 receptors on lymphocytes during infection of Echi-nococcus granulosus / 中国免疫学杂志

Objective:To investigate the effects of TGF-β1 on T lymphocytes of BALB/c mice infected with Echinococcus granulosus( E.granulosus ) in vitro.Methods: The inhibitor group:the spleen cells of BALB/c mouse were co-cultured with E.granulosus and SB525334.The control group:the spleen cells of BALB/c mouse were co-cultured with E.granulosus and PBS.The blank group:the spleen cells of BALB/c mouse were co-cultured with RPMI-1640 medium and SB525334.The lymphocytes were collected at 48 h post-infection.The T lymphocyte subsets, the number of CD4+CD25+T cells, the number of NK cells, and the expression of NKG2D receptor were detected by flow cytometry.The NK cell activity was determined with the lactate dehydrogenase leakage assay(LDH).Results:The inhibition the TGF-β1 receptors resulted in the increase of in the number of CD4+T cells,the decrease in the number of CD8+T cells,the increase of in the ratio CD4+/CD8+T cells,the decrease of in the number of CD4+CD25+T cells,the increase in the expression of the NKG2D receptors,the increase in the lysis rate of Yac-1 cells by NK cells,and a positive cor-relation between the expression of activity receptor NKG2D and killing activity of NK, which were mediated by E.granulosus.Conclusion: The inhibition of TGF-β1 receptors can enhance the immune response of T lymphocytes against E.granulosus infection in vitro.

Combination of Jiehe pellet and the standardized anti-tuberculosis therapeutic regimen for pulmonary tuberculosis complicating cervical lymph node tuberculosis in the aged / 国际中医中药杂志

Objective To assess the efficacy of combination of Jiehe pellet and the standardized anti-tuberculosis therapeutic regimen for pulmonary tuberculosis complicating cervical lymph node tuberculosis in the aged. Methods A total of 103 aged patients with pulmonary tuberculosis complicating cervical lymph node tuberculosis were enrolled and randomly allocated to either a standardized anti-tuberculosis therapeutic regimen group (control group with 51 patients) or a standardized anti-tuberculosis therapeutic regimen plus Jiehe pellet group (treatment group with 52 patients). The patients in the control group and the treatment group received the treatment with 2HRZE/4HR and 2HRZE/4HR plus Jiehe pellet for 6 months, respectively. The abscessed lymph nodes were treated by either total excision or incision and drainage after 4 weeks of medicine treatment in both groups. Sputum smear was examined for acid-fast bacilli. The CD8 cells expressing natural killer T cells receptors NKG2A, NKG2D in peripheral blood were detected by flow cytometry. The treatment outcome was measured at the end of treatment. Results The rates of lesion resolution (78.85%vs. 58.82%;χ2=4.439, P<0.05) and cavity closure (62.86% vs. 35.48%;χ2=3.893, P<0.05) in the treatment group were significantly higher than those in the control group. In the end of 2, 4 and 6 months of treatment, cumulative rates of sputum conversion from positive to negative in the treatment group were significantly higher than those in the control group (χ2 were 5.343, 5.067 and 4.118,all P<0.05). The CD8 cells expressing NKG2A after treatment in the treatment group were significantly lower than those before treatment in the treatment group (t=9.510, P<0.01) and after treatment in the control group (t=9.832, P<0.01);the CD8 cells expressing NKG2D after treatment in the treatment group were significantly higher than those before treatment in the treatment group (t=10.622, P<0.01) and after treatment in the control group (t=10.433, P<0.01). The serum levels of IL-6 and TNF-αafter treatment were significantly lower than those before treatment in both groups (t were 17.344 and 21.142 in the treatment group, 10.984 and 12.203 in the control group;all P<0.01 );the serum levels of IL-6 and TNF-α after treatment in the treatment group were significantly lower than those after treatment in the control group (t were 7.832 and 5.478,all P<0.01). The serum IL-10 levels after treatment were significantly higher than those before treatment in both groups (t were 12.454 in the treatment group, 7.934 in the control group; all P<0.01 ); and the serum IL-10 level after treatment in the treatment group was significantly higher than that after treatment in the control group (t=4.720, P<0.01). The effective rate for cervical lymph node tuberculosis in the treatment was significantly higher than that in the control group (88.5%vs. 64.7%;χ2=6.855, P<0.01). Conclusion Combination of Jiehe pellet and the standardized anti-tuberculosis therapeutic regimen may improve immune function, increase the rate of sputum conversion from positive to negative, and facilitate lesion resolution in aged patients with pulmonary tuberculosis complicating cervical lymph node tuberculosis.

El receptor NKG2D en la frontera de la inmunovigilancia y la carcinogénesis / The NKG2D receptor in the border of immune surveillance and carcinogenesis

El sistema inmune es capaz de realizar la detección y eliminación de células transformadas por un mecanismo fisiológico conocido como inmunovigilancia. En este proceso participa el receptor activador NKG2D presente en linfocitos T y células NK, ambos de suma relevancia en la inmunovigilancia contra el cáncer. Al reconocer el receptor NKG2D a sus ligandos (NKG2DLs) en las células que experimentan neotransformación se desencadena la respuesta lítica específica de las células linfoides citotóxicas. Asimismo, se ha descrito en diversos tipos de cáncer formas solubles de NKG2DLs que se ha demostrado son utilizadas para la evasión tumoral al saturar los receptores NKG2D presentes en las células efectoras linfoides evitando de esta manera ser reconocidas y eliminadas y, con ello escapando de la inmunovigilancia. Aunque este fenómeno de evasión inmune, donde participan algunos NKG2DLs, ha sido ya descrito y corroborado clínicamente no se ha estudiado si el receptor NKG2D está presente en las células tumorales per se ya que también podría estar implicado en subvertir la inmunovigilancia. En este trabajo se analizan evidencias recientes de que la expresión del receptor NKG2D no es exclusiva de linfocitos T y NK ya es expresado por células epiteliales tumorales tanto in vitro como in vivo. Las consecuencias de esta anómala expresión en células no linfoides tiene amplias implicaciones en la carcinogénesis que serán revisadas. También se analizan estudios clínicos recientes donde se comprueba la participación del receptor NKG2D en diferentes patologías tumorales.

The immune system is able to perform the detection and elimination of transformed cells by a mechanism known as physiological immune surveillance. This process involves the NKG2D receptor activator present in T lymphocytes and NK cells, both of paramount importance in the immune surveillance against cancer. To recognize the receptor NKG2D ligands (NKG2DLs) in cells that experience retransformation triggers the specific lithic response of the cytotoxic lymphoid cells. Also, soluble forms of NKG2DLs have been described in various types of cancer that have proven to be used for tumor evasion by saturating the NKG2D receptors present in the effector lymphoid cells thus avoiding their recognition and elimination, which makes them escape immune surveillance. Although this phenomenon of immune evasion, where some NKG2DLs participate, has already been described and corroborated, clinically, it has not been studied whether the receptor NKG2DL is present in the tumor cells per se because it could also be involved in reversing immune surveillance. This paper analyzes recent evidence that the expression of the NKG2D receptor is not lymphocyte T and NK exclusive it is already expressed by tumor epithelial cells in vitro and in vivo. Consequences of this anomalous expression in non-lymphoid cells have widespread implications in carcinogenesis, which will be revised. Recent clinical studies to prove the participation of NKG2D receptor in several tumor pathologies are analyzed.