The Cap gene of antisense strand of circovirus has the most variation of the genome, and encodes a capsid protein which has the main immunogenicity. The N-terminal of capsid protein makes up of nuclear localization signal which is involved with virus location. This review summarizes the research advance of Cap gene of circovirus in the sequence characteristics, its encoding capsid protein, basic functions of the capsid protein and its interaction with MKRN1 protein, Hsp40 protein, receptor protein gClqR and complement factor C1qB protein. This paper lays a theory foundation for the further study of the capsid protein in the aspects of viral attachment, replication and transportation.
Animals ; Capsid Proteins ; genetics ; immunology ; metabolism ; Circoviridae Infections ; veterinary ; virology ; Circovirus ; genetics ; immunology ; Genetic Variation ; Genome, Viral ; genetics ; Nuclear Localization Signals ; Protein Binding ; Virus Replication

OBJECTIVETo study the Ag-NORs expressive character of T lymphocyte in patients with oral and maxillofacial tumor and evaluate its diagnostic value for the patients.

METHODSNucleolar organizer regions(NORs) of T lymphocyte from 86 normal adults, 102 patients with oral and maxillofacial benign tumors and 87 patients with oral and maxillofacial malignant tumors were analyzed through KL imaging system.

RESULTSThere was significant difference among normal adults, benign and malignant patients (P < 0.01). Their average I.S% values were 7.87 +/- 0.12, 5.72 +/- 0.14, 4.19 +/- 0.13, respectively.

CONCLUSIONThe expression of Ag-NORs of T lymphocyte has definite relation with oral and maxillofacial tumors; detection of Ag-NORs might play valuable in diagnosis of oral and maxillofacial tumors.

Adolescent ; Adult ; Antigens, Nuclear ; Diagnosis, Differential ; Female ; Humans ; Male ; Maxillary Neoplasms ; diagnosis ; immunology ; metabolism ; Middle Aged ; Mouth Neoplasms ; diagnosis ; immunology ; metabolism ; Nuclear Proteins ; biosynthesis ; genetics ; metabolism ; Nucleolus Organizer Region ; metabolism ; Silver Staining ; T-Lymphocytes ; diagnostic imaging ; metabolism ; Ultrasonography

OBJECTIVETo identify and select smooth muscle progenitor cells from mouse bone marrow mesenchyme stem cell population and to characterize smooth muscle progenitor cells in peripheral blood.

METHODSRecombinant expression vector with the promoter of sm22alpha was constructed to have an enhancement type green fluorescent protein expression plasmid (EGFP-1). The construct was transfected into mouse bone marrow mesenchyme stem cells using Lipofectamine 2000. Morphological assessment was performed and the expressions of myocardin at protein and mRNA levels by fluorescence microscope and RT-PCR were evaluated at 3, 5, 7, and 10 d targeting CD34 positive bone mesenchyme stem cells.

RESULTSThe transfection efficiency of the positive control group was 70% +/- 1.5% (P > 0.05). Expected green fluorescent proteins expressed at 3rd day. The numbers of green fluorescent cells in experimental groups increased with the time and reached the peak at the 7th day, and declined thereafter. The shapes of the green fluorescent cells were also different from each others. The positive ratios of green fluorescent cells at different time points: 3 d: 7% +/- 0.13%, 5 d: 10% +/- 0.32%, 7 d: 20% +/- 0.26%, 10 d: 12% +/- 0.18%, P < 0.05. Myocardin mRNA expression roughly correlated with green fluorescent expressions. CD34 was expressed on the 5th day in transfected bone mesenchyme stem cells. The CD34 positive ratio was 5.2% +/- 0.21% (P > 0.05).

CONCLUSIONSThere are smooth muscle progenitor cells among mouse bone marrow mesenchyme stem cell population. Smooth muscle progenitor cells can be selected using a Psm22alpha-EGFP-1 recombinant expression approach.

Animals ; Antigens, CD34 ; immunology ; Bone Marrow Cells ; cytology ; immunology ; Cell Separation ; methods ; Cell Shape ; Green Fluorescent Proteins ; Mesenchymal Stromal Cells ; cytology ; immunology ; Mice ; Microfilament Proteins ; genetics ; Microscopy, Fluorescence ; Muscle Proteins ; genetics ; Myocytes, Smooth Muscle ; cytology ; Nuclear Proteins ; genetics ; metabolism ; Promoter Regions, Genetic ; RNA, Messenger ; genetics ; Recombinant Proteins ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Trans-Activators ; genetics ; metabolism ; Transfection

OBJECTIVETo produce BMI1 polyclonal antibody, mouse Bmi1 cDNA was cloned from mouse testis and expressed in E. coli BL21.

METHODSBmi1 gene was amplified from mouse testis by RT-PCR and inserted into the prokaryotic expression vector pET-28c(+). Subsequently the recombined vector was transformed and expressed in E. coli BL21 (DE3) and the immunogenicity of recombined protein BMI1 (rBMI1) was tested by Western blot.

RESULTSMouse Bmi1 cDNA of 975 bp was successfully cloned and recombined. E. coli BL21 strains expressed rBMI1 were screened. The expression protein amounted to 12% of the total bacterial protein after induced with IPTG, which included inclusion body and soluble protein. Inclusion body was the major pattern of the expression that amounted to 71% of the insoluble protein. Western blot analysis showed that rBMI1 could be specially recognized by mouse monoclonal IgG1 anti-BMI1 and His-tag antibody.

CONCLUSIONThere was expression of Bmi1 gene in mouse testis. Mouse Bmi1 cDNA was successfully cloned and expressed prokaryoticly.

Animals ; Antibodies, Monoclonal ; immunology ; Cloning, Molecular ; DNA, Complementary ; genetics ; Escherichia coli ; genetics ; Gene Expression ; Male ; Mice ; Nuclear Proteins ; biosynthesis ; genetics ; immunology ; Polycomb Repressive Complex 1 ; Proto-Oncogene Proteins ; biosynthesis ; genetics ; immunology ; Recombinant Proteins ; biosynthesis ; immunology ; Repressor Proteins ; biosynthesis ; genetics ; immunology ; Testis ; metabolism

Exosomes are small membrane vesicles secreted from various types of cells. Tumor-derived exosomes contain MHC class I molecules and tumor-specific antigens, receiving attention as a potential cancer vaccine. For induction of efficient anti-tumor immunity, CD4+ helper T cells are required, which recognize appropriate MHC class II-peptide complexes. In this study, we have established an MHC class II molecule-expressing B16F1 murine melanoma cell line (B16F1-CIITA) by transduction of the CIITA (Class II transactivator) gene. Exosomes from B16-CII cells (CIITA-Exo) contained a high amount of MHC class II as well as a tumor antigen TRP2. When loaded on dendritic cells (DCs), CIITA-Exo induced the increased expression of MHC class II molecules and CD86 than the exosomes from the parental cells (Exo). In vitro assays using co-culture of immunized splenocytes and exosome-loaded DCs demonstrated that CIITA-Exo enhanced the splenocyte proliferation and IL-2 secretion. Consistently, compared to B16-Exo, CIITA-Exo induced the increased mRNA levels of inflammatory cytokines such as TNF-alpha, chemokine receptor CCR7 and the production of Th1-polarizing cytokine IL-12. A tumor preventive model showed that CIITA-Exo significantly inhibited tumor growth in a dose-dependent manner. Ex vivo assays using immunized mice demonstrated that CIITA-Exo induced a higher amount of Th1-polarized immune responses such as Th1-type IgG2a antibodies and IFN-gamma cytokine as well as TRP2-specific CD8+ T cells. A tumor therapeutic model delayed effects of tumor growth by CIITA-Exo. These findings indicate that CIITA-Exo are more efficient as compared to parental Exo to induce anti-tumor immune responses, suggesting a potential role of MHC class II-containing tumor exosomes as an efficient cancer vaccine.
Animals ; Cancer Vaccines/genetics/immunology ; Cell Line, Tumor ; Cell Proliferation ; Dendritic Cells/immunology ; Exosomes/genetics/*metabolism ; Gene Expression Regulation ; Gene Transfer Techniques ; Immunity, Cellular/immunology ; Immunity, Humoral/immunology ; Immunotherapy ; Lymphocyte Activation/immunology ; Melanoma, Experimental/mortality/pathology/*physiopathology ; Mice ; Mice, Inbred C57BL ; Nuclear Proteins/*genetics/*metabolism ; Survival Analysis ; T-Lymphocytes/immunology/metabolism ; Trans-Activators/*genetics/*metabolism ; Transduction, Genetic

We examined the effect of class II transactivator (CIITA) down-modulation on allograft rejection. To inhibit the function of CIITA, we constructed a series of CIITA mutants and found one exhibiting the dominant-negative effect on the regulation of major histocompatibility complex (MHC) class II expression. To test whether the CIITA dominant-negative mutant reduces immunogenecity, CIITA-transfected melanoma cells were injected into allogeneic host and assessed for immune evading activity against host immune cells. We demonstrated that the CIITA dominant-negative mutant allowed tumor nodules to develop earlier in the lung than control by this tumor challenge study. Furthermore, skin grafts deficient for CIITA also survived longer than wild-type in allogeneic hosts. Both the tumor challenge and skin graft studies suggest the inhibition of CIITA molecules in donor tissue would be beneficial to the control of allo-response.
Transplantation, Homologous ; Transfection ; Trans-Activators/genetics/*immunology/metabolism ; Trans-Activation (Genetics)/genetics/immunology ; Skin Transplantation ; Nuclear Proteins/genetics/*immunology/metabolism ; Mutation ; Mice, Transgenic ; Mice, Knockout ; Mice, Inbred C57BL ; Mice, Inbred BALB C ; Mice ; Melanoma, Experimental/genetics/immunology/pathology ; Male ; Interferon Type II/pharmacology ; Humans ; Histocompatibility Antigens Class II/genetics/*immunology/metabolism ; Graft Survival/genetics/immunology ; Graft Rejection/genetics/*immunology ; Genes, MHC Class II/genetics/immunology ; Flow Cytometry ; DNA, Complementary/genetics ; Cell Proliferation/drug effects ; Cell Line, Tumor ; Animals

Tumor hypoxia contributes to the progression of a malignant phenotype and resistance to ionizing radiation and anticancer drug therapy. Many of these effects in hypoxic tumor cells are mediated by expression of specific set of genes whose relation to therapy resistance is poorly understood. In this study, we revealed that DNA-dependent protein kinase (DNA-PK), which plays a crucial role in DNA double strand break repair, would be involved in regulation of hypoxia inducible factor-1 (HIF-1). HIF-1beta-deficient cells showed constitutively reduced expression and DNA-binding activity of Ku, the regulatory subunit of DNA-PK. Under hypoxic condition, the expression and activity of DNA- PK were markedly induced with a concurrent increase in HIF-1alpha expression. Our result also demonstrated that DNA-PK could directly interact with HIF- and especially DNA-PKcs, the catalytic subunit of DNA-PK, could be involved in phosphorylation of HIF-1alpha, suggesting the possibility that the enhanced expression of DNA- PK under hypoxic condition might attribute to modulate HIF-1alpha stabilization. Thus, the correlated regulation of DNA-PK with HIF-1 could contribute to therapy resistance in hypoxic tumor cells, and it provides new evidence for developing therapeutic strategies enhancing the efficacy of cancer therapy in hypoxic tumor cells.
Antibodies/immunology ; Cell Hypoxia ; Cell Line, Tumor ; DNA Helicases/immunology/metabolism ; DNA-Binding Proteins/genetics/*metabolism/*physiology ; Deferoxamine/pharmacology ; Drug Resistance, Neoplasm/*physiology ; Humans ; Immunoprecipitation ; Neoplasms/enzymology/*metabolism ; Nuclear Proteins/genetics/*metabolism/physiology ; Phosphorylation ; Protein-Serine-Threonine Kinases/metabolism/*physiology ; Research Support, Non-U.S. Gov't ; Transcription Factors/genetics/*metabolism/physiology ; Up-Regulation

OBJECTIVETo compare the immunophenotypic and clinical characteristics between NPM1 mutated acute myeloid leukemia (AML) (NPM1m(+)AML) and unmutated AML(NPM1m(-)AML) not otherwise characterized (NOS) under similar FAB subtypes constituent ratio.

METHODSImmunophenotyping and NPM1 gene mutation type-A, B and D and other leukemic related fusion genes were detected by multiparameter flow cytometry and real time RT-PCR or PCR, respectively. 104 AML patients with NPM1m(+)AML and performed immunophenotyping assay were included, 97 with NPM1m(-)AML.

RESULTSThere were significant difference between the two groups at presentation in terms of sex, white blood count(WBC), platelet counts (PLT), blast ratio, normal karyotype ratio, WT1 expression level, FLT3-ITD mutation positive rate and remission rate of first course of induction therapy (P < 0.05). On the immunophenotype, the expression of early differentiation antigens (CD34, HLA-DR, CD117, CD38), lymphocytic antigens (CD7, CD4, CD19, CD2), myeloid and monocytic differentiation-associated antigens (CD13, CD14, CD15) were lower, and that of CD33 as well as CD123 were higher in NPM1m(+)AML patients. Among them, only CD34, HLA-DR, CD7, and CD4 positive cases were significantly lower in NPM1m(+)AML group than in NPM1m(-)AML group (P < 0.05), the rest of them had significant difference in the number of positive cells (P < 0.05). Above features were further analyzed between the M1/M2 and M4/M5 subgroups. M1/M2 cases retained the women prominent and had a higher WT1 expression level (P < 0.05). The expression of monocytic differentiation-associated antigens including HLA-DR and lymphocytic antigens were higher and that of CD117 were lower in M4/M5 subtype (P < 0.05). Among them, the positive rates of HLA-DR, CD64, CD11b, CD10, CD15, and CD4 were significantly higher in M4/M5 than in M1/M2 in NPM1m(+)AML group (P < 0.05).

CONCLUSIONThe most clinical characteristics in NPM1m(+)AML patients are consistent with reports, but some immunophenotype are different to the previous reports under similar FAB subtypes constituent ratio. The major immunophenotypic features of NPM1m(+)AML patients are lower expression of progenitor, myeloid and lymphoid lineage antigens. Monocytic differentiation-associated antigens are only higher expression in M4/M5 cases when comparison with M1/M2 cases within NPM1m(+)AML group.

Adolescent ; Adult ; Aged ; Antigens, CD ; metabolism ; Child ; Child, Preschool ; Female ; HLA-DR Antigens ; immunology ; Humans ; Immunophenotyping ; Leukemia, Myeloid, Acute ; diagnosis ; genetics ; immunology ; Male ; Middle Aged ; Mutation ; Nuclear Proteins ; genetics ; Young Adult

OBJECTIVETo study Twist expression in thyroid papillary carcinoma (PTC) by immunohistochemistry and to assess its usefulness as marker in the differential diagnosis of PTC, follicular adenomas (FA) and benign papillary lesions (BPL).

METHODSFifty cases of PTC, 48 cases of FA and 47 cases of BPL were evaluated using manual tissue chip and SP immunohistochemical stain to detect the expression of Twist and HBME-1, and comparing the staining to that of cytokeratin 19 (CK19).

RESULTSIn PTC, positive rates of Twist, HBME-1 and CK19 were 100% (48/48), 94.0% (47/50) and 78.0% (39/ 50) respectively; in FA, positive rates were 0, 6.7% (3/45) and 0 respectively; in BPL, positive rates were 7.0% (3/34), 2.1% (1/47) and 0, respectively. The differences between PTC and FA and between PTC and BPL were both statistically significant (P = 0. 000). The sensitivity of Twist, HBME-1 and CK19 was 100%, 94.0% and 78.0%; the specifity was 96.4%, 95.7% and 100%; overall accurary was 97.7%, 95.1% and 91.9%, respectively.

CONCLUSIONSPositive rates of Twist is higher than the other markers in PTC. Immunohistochemical staining of Twist has important significance in the differential diagnosis of thyroid lesions. Twist immunohistochemistry maybe helpful in diagnosis and differential diagnosis of PTC.

Adenocarcinoma, Follicular ; metabolism ; Adenocarcinoma, Papillary ; pathology ; Adenoma ; diagnosis ; metabolism ; Biomarkers, Tumor ; immunology ; Carcinoma, Papillary ; diagnosis ; metabolism ; pathology ; Carcinoma, Papillary, Follicular ; metabolism ; Diagnosis, Differential ; Galectin 3 ; genetics ; metabolism ; Immunohistochemistry ; Keratin-19 ; genetics ; Keratins ; genetics ; metabolism ; Nuclear Proteins ; genetics ; metabolism ; Thyroid Neoplasms ; diagnosis ; metabolism ; pathology ; Thyroid Nodule ; pathology ; Twist-Related Protein 1 ; genetics ; metabolism

MHC class II expression is controlled mainly at transcriptional level by class II transactivator (CIITA), which is a non-DNA binding coactivator and serves as a master control factor for MHC class II genes expression. Here, we describe the function of a novel splice-isoform of CIITA, DC-expressed caspase inhibitory isoform of CIITA (or DC-CASPIC), and we show that the expression of DCCASPIC in DC is upregulated upon lipopolysaccharides (LPS) induction. DC-CASPIC localizes to mitochondria, and protein-protein interaction study demonstrates that DC-CASPIC interacts with caspases and inhibits its activity in DC. Consistently, DC-CASPIC suppresses caspases-induced degradation of nitric oxide synthase-2 (NOS2) and subsequently promotes the synthesis of nitric oxide (NO). NO is an essential regulatory molecule that modulates the capability of DC in stimulating T cell proliferation/activation in vitro; hence, overexpression of DC-CASPIC in DC enhances this stimulation. Collectively, our findings reveal that DC-CASPIC is a key molecule that regulates caspases activity and NO synthesis in DC.
Alternative Splicing ; Amino Acid Sequence ; Animals ; Base Sequence ; CARD Signaling Adaptor Proteins ; genetics ; metabolism ; Cell Line ; Dendritic Cells ; drug effects ; immunology ; metabolism ; Humans ; In Vitro Techniques ; Lipopolysaccharides ; pharmacology ; Lymphocyte Activation ; Mice ; Mice, Inbred C57BL ; Mitochondria ; metabolism ; Molecular Sequence Data ; Nitric Oxide ; biosynthesis ; Nitric Oxide Synthase Type II ; metabolism ; Nuclear Proteins ; genetics ; metabolism ; Protein Isoforms ; genetics ; metabolism ; RNA, Messenger ; genetics ; metabolism ; T-Lymphocytes ; immunology ; metabolism ; Trans-Activators ; genetics ; metabolism ; Up-Regulation ; drug effects