Objective To study the roles of regulated upon activation normal T cell expressed and secreted(RANTES) and monocyte cbemoattractant protein-1 (MCP-1) in human immunodeficien- cy virus-1(HIV 1) infected patients.Methods RANTES and MCP-1 in HIV-1 infected patients, including treated and untreated groups,and healthy control group were determined by enzyme-linked immunosorbent assay(ELISA).The recombinant plasmids,hMCP-pcDNA3.1,bRANTES- peDNA3.1 and hMCP/bRANTES-pcDNA3.1,were constructed and transfected into CHO cells to overexpress the corresponding recombinant proteins,whose chemoattract function was then studied. Results The level of RANTES was (164.3?21.3) pg/mL in healthy control group,(1 224.1?62.0) pg/mL in untreated group and (475.3?36.2) pg/mL in treated group.The level of MCP-1 was (90.6?28.5) pg/mL in healthy control group,(335.0?30.3) pg/mL in untreated group and (807.2?62.6) pg/mL in treated group.In HIV-1 infected patients,the levels of RANTES and MCP-1 were significantly increased.After highly active antiretroviral therapy (HAART),the level of RANTES declined,but MCP-1 increased further.Western blot assay revealed that the three recombinant proteins could be recognized by monoclonal antibodies respectively.All of them could chemoattract human peripheral blood mononuclear cells(PBMCs).And the chemoattractant potency of MCP/RANTES fusion protein was stronger.When the recombinant proteins were used with con- centrations as 50,200,400 and 800 pg/mL,respectively,the number of PBMCs chemoattracted by MCP/RANTES fusion protein was 52?10~4/mL,102?10~4/mL,132?10~4/mL and 184?10~4/mL; the number of PBMCs chemoattracted by RANTES was 27?10~4/mL,51?10~4/mL,65?10~4/mL and 96?10~4/mL;the number of PBMCs chemoattracted by MCP-1 was 18?10~4/mL,44?10~4/mL, 54?10~4/mL and 74?10~4/mL.Conclusion RANTES and MCP-1 may both be involved in the HIV infection process and host immunological reaction against HIV.

OBJECTIVETo study the role of interleukin(IL)18 and interferon gamma (IFN-Gamma) in the pathogenesis of condyloma acuminatum(CA).

METHODSDouble antibody sandwich ELISA was used to measure the levels of IL-18 and IFN-Gamma in serum of 30 cases of CA.

RESULTThe serum levels of IL-18 and IFN-Gamma of the CA patients were significantly higher than those of normal controls IL-18 132.00 +/-52.92 microg/L compared with 50.79 +/- 23.48 microg/L, P<0.01 IFN-Gamma:80.83 +/-20.46 microg/L compared with 20.71 +/-11.18 microg/L P <0.01) and IL-18 levels were positively correlated with IFN-Gamma concentrations (r=0.754, t=6.081, P <0.01).

CONCLUSIONThese data suggest that during infection of human papilloma viruses, anti-infective immune responses are activated.

Adult ; Condylomata Acuminata ; immunology ; Female ; Humans ; Interferon-gamma ; blood ; Interleukin-18 ; blood ; Male ; Middle Aged

20(Z= -2.117, P=0.034), and between the patients with OD and without OD (Z=-3.089, P=0.002), but PCT was not so between the non-surviror and survivor (Z=-1.307, P=0.191). The serum PCT level correlated with the incidence of organ dysfunction (x~2=14.82, P=0.033) and APACHEII (x~2=12.83, P

OBJECTIVETo investigate the role of cyclin D1 and p27(kip1) in the occurrence and development of conventional renal cell carcinoma(CRCC).

METHODSRT-PCR and Western-blot were used to detect mRNA and protein contents of cyclin D1 and p27(kip1) in 25 CRCCs and 10 normal renal tissue distant to tumor. Immunohistochemistry was used to investigate the expression of cyclin D1 and p27(kip1) in pathological tissue sections of 76 CRCCs. The relationship between those index and clinicopathological parameters was analyzed statistically.

RESULTIn CRCC, the expression of cyclin D1 was higher than that of the control group. The higher cyclin D1 content was related to big tumor size (P<0.05); The expression of p27(kip1) was lower than that of the control group, and the lower p27(kip1) was related to higher nuclear grade and TNM stage (P<0.01). The 5-year cumulative survival rate of the p27(kip1) high expression group is longer than that of the low group (P<0.01).

CONCLUSIONThe excessive expression of cyclin D1 and lower expression of p27(kip1) play an important role in the carcinogenesis of CRCC. The lower expression of p27(kip1) may affect the progression of the tumors. The detection of p27(kip1) may be as a reference marker in the prognosis of CRCC.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Blotting, Western ; Carcinoma, Renal Cell ; genetics ; metabolism ; pathology ; Cyclin D1 ; biosynthesis ; genetics ; Cyclin-Dependent Kinase Inhibitor p27 ; biosynthesis ; genetics ; Female ; Gene Expression Regulation, Neoplastic ; Humans ; Kidney Neoplasms ; genetics ; metabolism ; pathology ; Male ; Middle Aged ; RNA, Messenger ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVE: To investigate the effects of IL-18 gene-modified fetal hepatocytes (AdmIL-18/MNL.CL2) intrasplenic transplantation on mouse immune function. METHODS: Forty mice were evenly divided into 4 groups of 10. Each group received an intrasplenic transplantation one of the following: AdmIL-18/BNL.CL2, Ad-LacZ/BNL.CL2 (virus control), BNL.CL2 (cell control) and PBS (blank control). After two weeks, the mice were sacrificed. Serum cytokine levels, Mpsi and splenic cell culture supernatant and liver tissue extracts supernatants were measured using ELISA. Hepatic cytokines mRNA expression were determined by RT-PCR. THe cytotoxicity of peritoneal Mpsi and NK activity of spienocytes were detected by LDH release assay. The proliferation of splenic lymphocytes was determined by MTT assay. RESULTS: The IL-18, IL-2,IFN-gamma, TNF-alpha levels of serum, Mpsi and splenocyte culture supernatant, liver tissue extracts supernatants in mice transplanted with AdmIL-18/BNL.CL2 were higher and the IL-4, IL-10 levels were lower compared to their levels in other 3 groups. The highest IL-18, IL-2, IFN-gamma, TNF-alpha and the lowest IL-4, IL-10 mRNA expressions in the liver were observed in mice transplanted with AdmIL-18/BNL.CL2. The mice transplanted with AdmIL-18/BNL.Cl2 showed significantly increases cytotoxicity of Mpsi, NK activity and splenic cell proliferation compared with the other 3 groups. CONCLUSION: AdmIL-18 can be effectively transfected into mice fetal heptocytes which subsequently IL-18. Intransplenic transplantation of IL-18 gene-modified fetal hepatocytes may augment mouse immune function and provide an useful basis for targeted gene therapy of liver disease.

OBJECTIVETo explore the effects of beta-elemene combined with aclarubicin on the induction of HL-60 cell apoptosis and its mechanisms in antileukemia therapy.

METHODSHL-60 cells were treated for 20 hours with different dose of aclarubicin (0.05, 0.10, 0.25 microg/ml) or with different concentrations of beta-elemene (10, 20, 40 microg/ml) in the presence or absence of aclarubicin (0.10 microg/m). The apoptotic rate was analyzed by flow cytometry (FCM), the productions of PGE2 in culture supernatants was detected by competitive ELISA and the expressions of COX-2 and NF-kappaB activity in HL-60 cells by Western blot.

RESULTSLower concentration of aclarubicin (0.05, 0.10 microg/ml) didn't affect apoptotic rate, and COX-2, NF-kappa B and PGE2 expression on HL-60 cells. Combined treatment of beta-elemene and aclarubicin (0.10 microg/ml) enhanced the apoptotic effect and down-regulated COX-2, NF-kappaB and PGE2 expressions. There was a positive correlation between the effects and beta-elemene concentrations.

CONCLUSIONbeta-elemene enhances aclarubicin-mediated apoptotic effect, down-regulation of COX-2 and their inducing products PGE2 in HL-60 cells by suppressing activitation of NF-kappaB.

Aclarubicin ; Apoptosis ; drug effects ; Cell Line, Tumor ; Down-Regulation ; HL-60 Cells ; Humans ; NF-kappa B ; metabolism

OBJECTIVETo prepare a monoclonal antibody (mAb) against extracellular domain of B7-H4 and to investigate the expression of B7-H4 in pancreatic cancer tissue with the prepared mAb.

METHODSBalb/c mice were immunized with 3T3-B7-H4 cells and the splenic cells of the immunized mice were fused with Sp2/0 myeloma cells by conventional hybridoma techniques. An indirect ELISA method using 3T3-B7-H4 lysate as antigen was established to screen antibody-producing hybridoma cell lines. Western blott, immunoprecipitation (IP), and immunohistochemistry (IHC) were applied to characterize the mAb. Immunohistochemical staining was used to detect the expression of B7-H4 in human pancreatic cancer tissue. The correlation of B3-H4 expressions and pathological features of pancreatic cancer was analyzed.

RESULTSA hybridoma cell line secreting mAb against B7-H4 was obtained. The subclass of this mAb was IgM, and the light chain was Kappa. Western blot and IP showed that the mAb specifically recognized B7-H4. IHC staining revealed that the mAb stained in a predominantly diffuse plasmalemmal or cytoplasmic pattern when applied to certain tumor tissues. The B7-H4 was diffusely expressed in the cytoplasma and/or membrane of pancreatic cancer tissue, which was much higher than that expressed in normal pancreatic tissue (4.00 ± 1.44 compared with 1.12 ± 0.78, P ± 0.01). The expression of B7-H4 was higher in pancreatic cancer tissues with higher pathological grade or with lymph node metastasis as compared with that in pancreatic cancer tissues with lower grade or with no lymph mode metastasis (6.10 ± 0.72 compared with 3.55 ± 1.12,P<0.01: 6.14 ± 0.66 compared with 3.70 ± 1.25,P<0.01). The expression level of B7-H4 was not related to patients'age and gender.

CONCLUSIONMonoclonal antibody against B7-H4 with high activity and specificity has been prepared successfully. The expression of B7-H4 in pancreatic cancer tissue is up-regulated,which is closely related to the tumor grade and lymph node metastasis in pancreatic cancer.

3T3 Cells ; Animals ; Antibodies, Monoclonal ; biosynthesis ; Cell Line, Tumor ; Humans ; Immunohistochemistry ; Lymphatic Metastasis ; Mice ; Mice, Inbred BALB C ; Neoplasm Grading ; Pancreatic Neoplasms ; metabolism ; pathology ; V-Set Domain-Containing T-Cell Activation Inhibitor 1 ; metabolism

OBJECTIVE: To observe whether angiotensin II (AngII) influences expression of lectin-like oxidized low density lipoprotein receptor-1 (LOX-1) in cultured human umbilical vein endothlial cells(HUVECs). METHODS: Cultured HUVECs were incubated with 10(-5)mol/L approximate, equals 10(-10) mol/L AngII for 24 h or with 10(-6) mol/L AngII for various time up to 48 h. Then HUVECs LOX-1 protein expression was measured by endothlial cell enzyme linked immunosorbent assay, and mRNA level of LOX-1 was detected by quantitative competitive reverse transcription-polymerase chain reaction. RESULTS: Incubation of HUVECs with AngII for 24 h significantly increased LOX-1 protein expression (P<0.001). The increase was dependent on AngII concentration (10(-5)mol/L approximate, equals 10(-9)mol/L). LOX-1mRNA expression was also induced by AngII, and after 24 h incubation of AngII(10(-5)mol/L approximate, equals 10(-8)mol/L), LOX-1mRNA expression increased 4.43, 4.25, 2.71, 1.84 times, respectively. After treatment with 10(-6) mol/L AngII for 3 h, LOX-1 expression (protein and mRNA) was elevated (P<0.001) in HUVECs, reaching its maxium at 24 approximate, equals 36 h. CONCLUSION: AngII upregulates LOX-1 expression concentration-dependently and time-dependently in HUVECs.

Fibroblast growth factor-21 (FGF-21) is an important metabolism regulator, however, whether FGF-21 has effects on cardiovascular remains unclear. In this study, H2O2-induced injury in H9c2 cells was used as a cell model, the anti-apoptosis potential and mechanism of FGF-21 against oxidative injury were evaluated by MTT assay, flow cytometry assay and real-time PCR. The results showed that FGF-21 could increase the cell survival of H2O2-induced injury in H9c2 cells and prevent H9c2 cells from oxidative stress-induced apoptosis. Furthermore, FGF-21 can elevate SOD activity and regulate Bcl-2/Bax expression in H9c2 cells. The results suggest that FGF-21 have protective effect against the H2O2-induced apoptosis in H9c2 cells.
Animals ; Apoptosis ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Fibroblast Growth Factors ; pharmacology ; Hydrogen Peroxide ; toxicity ; Malondialdehyde ; metabolism ; Myocytes, Cardiac ; cytology ; drug effects ; metabolism ; Oxidative Stress ; drug effects ; Protective Agents ; pharmacology ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; metabolism ; RNA, Messenger ; metabolism ; Rats ; Reactive Oxygen Species ; metabolism ; Superoxide Dismutase ; metabolism ; bcl-2-Associated X Protein ; genetics ; metabolism

OBJECTIVETo construct vectors stably expressing bioactive VEGF165 and VEGF121 in MC3T3-E1 cells.

METHODSVEGF165 and VEGF121 genes were obtained by RT-PCR from HL-60 cells and inserted into eukaryotic expression plasmid pcDNA3.1(+). The constructed plasmids containing right sequence of target genes were transfected into MC3T3-E1 cells by lipofectin. The cells were limitedly diluted twice and selected by G418, the expression of VEGF was tested by ELISA, Western-blot and immunofluorescence essay. The bioactivity of expressed products was proved by MTT method.

RESULTDNA sequencing indicated the sequences of the obtained VEGF165 and VEGF121 were identical to the those reported in Genebank, and the newly-constructed plasmids were VEGF165/pcDNA3.1(+) and VEGF121/pcDNA3.1(+). The results of ELISA, Western-blot and immunofluorescence showed that the transfected cells could express VEGF165 and VEGF121 proteins. And MTT proved the proteins were bioactive.

CONCLUSIONThe transfected MC3T3-E1 cells could stably express high levels of bioactive VEGF165 and VEGF121.

3T3 Cells ; Animals ; HL-60 Cells ; Humans ; Mice ; Plasmids ; genetics ; Recombinant Proteins ; biosynthesis ; genetics ; Transfection ; Vascular Endothelial Growth Factor A ; biosynthesis ; genetics