BACKGROUNDPatients with systemic lupus erythematosus often have various autoantibodies. The relationship between these antibodies is still poorly understood. The aim of the present study was to observe the anti-SSB antibody and anti-dsDNA antibody production profiles following immunization with synthetic SSB peptide alone, DNA alone or co-immunization with these two antigens.

METHODSSSB 214 - 225 aa peptide was synthesized by organic chemistry solid-phase peptide synthesis. Rabbits were immunized with the following antigens: synthetic SSB peptide linked with keyhole limpet hemocyanin (KLH), DNA, SSB plus dsDNA, KLH and PBS. Antibodies were measured by ELISA. Histopathology and direct immufluorescence assays were also applied.

RESULTSAnti-SSB and anti-dsDNA antibodies were produced following immunization with SSB peptide and DNA respectively. The level of SSB antibody in the co-immunization group was higher than that of the SSB peptide immunization group. The level of anti-dsDNA antibody in the co-immunization group was, however, lower than that in the DNA immunization group. Meanwhile, the level of anti-SSB antibody was higher than that of anti-DNA antibody in the co-immunization group. No morphological or immunological abnormalities were found in the heart, liver, kidney, spleen or skin tissues.

CONCLUSIONInhibition of anti-dsDNA-antibody was induced by co-immunization with synthesized SSB peptide and DNA, which might explain, at least partly, the mild disease in some LE subsets associated with SSB antibody.

Animals ; Antibodies, Antinuclear ; biosynthesis ; Autoantigens ; immunology ; DNA ; immunology ; Fluorescent Antibody Technique, Direct ; Immunization ; Rabbits ; Ribonucleoproteins ; immunology

OBJECTIVETo construct a replicative anti-tumor DNA vaccine PSCK-2PFcGB based on Semliki Forest Virus (SFV) replicon vector and observe its expression in vivo and in vitro.

METHODSThe plasmid pVAX1-2PFcGB was digested with Nhe I, and the digestion product was blunted prior to further digestion with BssH II to obtain the fragment 2PFcGB, a fusion gene containing the multitarget complex antigen 2PAG encoding both the most cytotoxic T lymphocyte epitopes of human survivin and chorionic gonadotropin β chain-CTP37 of human and monkey. The 2PFcGB fragment was inserted into the PSCK vector digested by Sma I. The products with the expected size were extracted and ligated, and the positive clones were screened by kanamycin and amplified. The recombinant PSCK-2PFcGB, following identification by colony PCR and restriction endonuclease Nde I, was transfected into 293T cells via lipofectamine 2000 and its expression was detected. The recombinant plasmid was also transfected into mouse quadriceps femoris muscle to observe its expression in vivo by immunohistochemistry.

RESULTSNde I digestion resulted in a fragment of the expected size. Transfection with the recombinant plasmid PSCK-2PFcGB resulted in successful expression of the antigen and adjuvant molecular protein in 293T cells, with the positivity rates of 5.70% and 19.75%, respectively. The fusion tumor antigen survivin and hCGβ-CTP37 were also detected in the muscular tissues of the mice.

CONCLUSIONA novel replicative anti-tumor DNA vaccine PSCK-2PFcGB has been successfully constructed and can be expressed in 293T cells and in the muscular tissues of immunized mice, which provide a basis for further studies of the antitumor activity and immunological mechanism of the DNA vaccine.

Animals ; Antibodies, Antinuclear ; immunology ; Cancer Vaccines ; biosynthesis ; immunology ; Gene Expression ; Genetic Vectors ; HEK293 Cells ; Humans ; Mice ; Muscle, Skeletal ; metabolism ; Plasmids ; Semliki forest virus ; genetics ; Vaccines, DNA ; biosynthesis ; immunology

OBJECTIVETo investigate the role of IL-17 in the overproduction of autoantibodies and IL-6 overexpression by peripheral blood mononuclear cells (PBMC) of lupus nephritis (LN) patients.

METHODSFifteen consecutively hospitalized LN patients were selected as subjects and 15 healthy adults as normal controls. PBMC were obtained by Ficoll density gradient centrifugation. IgG, anti-dsDNA antibody and IL-6 protein levels were assessed using enzyme-linked immunosorbent assays (ELISA) on the supernatant of cultured PBMC of LN patients or normal controls. IL-6 mRNA levels in PBMC were measured using reverse transcription-polymerase chain reaction (RT-PCR).

RESULTSIn medium culture, IgG, anti-dsDNA and IL-6 protein levels of the supernatant of PBMC from LN patients were significantly higher than those from normal controls (1492.1 +/- 73.2 ng/ml vs 636.7 +/- 51.9 ng/ml for IgG, 306.6 +/- 53.7 IU/ml vs 95.8 +/- 11.6 IU/ml for anti-dsDNA and 50.92 +/- 15.92 ng/ml vs 1.77 +/- 0.73 ng/ml for IL-6, all P < 0.001). In LN patients, IgG, anti-dsDNA and IL-6 protein levels were higher in the supernatants of PBMC in the IL-17-stimulated culture than the medium culture, but in normal controls, only the IL-6 protein levels were significantly higher. The increase in IgG, anti-dsDNA and IL-6 protein levels induced by IL-17 was dose-dependent and could be completely blocked by IL-17 monoclonal antibody mIgG(28) and partially blocked by dexamethasone. Similarly, IL-6 mRNA overexpression of PBMC in LN patients or normal controls induced by IL-17 was both dose- and time-dependent. During medium culture, IL-6 mRNA levels in LN patients were significantly higher than those in normal controls (1.80 +/- 0.11 vs 0.36 +/- 0.07). During stimulation with IL-17, IL-6 mRNA levels in LN patients were higher than those in normal controls (3.21 +/- 0.24 vs 1.30 +/- 0.14, P < 0.05) and also significantly higher when comparing the stimulated culture with the medium culture either in LN patients or normal control.

CONCLUSIONSIL-17 may play an important role in the pathogenesis of LN through the induction of IgG, anti-dsDNA overproduction and IL-6 overexpression of PBMC in LN patients.

Adolescent ; Adult ; Antibodies, Antinuclear ; biosynthesis ; Autoantibodies ; biosynthesis ; Female ; Humans ; Immunoglobulin G ; biosynthesis ; Interleukin-17 ; pharmacology ; Interleukin-6 ; biosynthesis ; genetics ; Leukocytes, Mononuclear ; metabolism ; Lupus Nephritis ; immunology ; Male ; RNA, Messenger ; analysis

Remitting seronegative symmetric synovitis with pitting edema (RS3 PE) syndrome is characterized by symmetrical and acute synovitis, pitting edema, the absence of rheumatoid factor, increased acute phase reactants, lack of bony erosions on radiography, and benign and short clinical course. Half of all patients with Sjogren's syndrome experience arthritis during the disease course. We here describe the first case of Sjogren's syndrome presenting as RS3PE. She had swelling in knees, ankles, and wrists. After then the swelling spread to her lower legs, feet, face, and both hands. She was admitted to another hospital and was suspected of lupus or rheumatoid arthritis. Three months later, she had dry mouth and had lower lip biopsy. She was admitted to this hospital due to development of swelling in face and lower legs for 3 days. On physical examination, she had pitting edema in both hands and feet dorsum. Laboratory test showed elevated erythrocyte sedimentation rate, positivity of rheumatoid factor, anti-nuclear antibody, and anti-Ro antibody. There was no erosion in the hands radiography. Schirmer's test and lip biopsy was compatible with Sjogren's syndrome. She was diagnosed RS3 PE and Sjogren's syndrome. She was begun with prednisolone and her symptoms improved gradually.
Adult ; Antibodies, Antinuclear/biosynthesis ; Arthritis/complications ; Biopsy ; Blood Sedimentation ; Diagnosis, Differential ; Edema/*diagnosis ; Female ; Human ; Lymphocytes/pathology ; Prednisolone/therapeutic use ; Rheumatoid Factor/biosynthesis ; Salivary Glands/pathology ; Sjogren's Syndrome/*diagnosis ; Syndrome ; Synovitis/*diagnosis

OBJECTIVETo prokaryoticly express and purify HuD protein and its RNA recognition motifs.

METHODSHuD protein was prokaryoticly expressed and purified by molecular cloning technology. Its biologic activity was testified by Western Blot.

RESULTSPurified HuD protein and its RNA recognized motifs were observed.

CONCLUSIONSThe result might aid for basic research and clinical application.

Antibodies, Antinuclear ; biosynthesis ; genetics ; isolation & purification ; Carcinoma, Small Cell ; genetics ; immunology ; metabolism ; Cloning, Molecular ; DNA, Complementary ; genetics ; ELAV Proteins ; ELAV-Like Protein 4 ; Humans ; Lung Neoplasms ; genetics ; immunology ; metabolism ; Nerve Tissue Proteins ; biosynthesis ; genetics ; isolation & purification ; Neurons ; immunology ; Paraneoplastic Syndromes, Nervous System ; genetics ; immunology ; metabolism ; RNA-Binding Proteins ; biosynthesis ; genetics ; isolation & purification

We concentrated ourselves to evaluate the prognostic significance of the p53 gene mutations, its protein expression and MIB-1 index as a proliferative marker in canine mammary tumors. In the present study, a total of 20 cases were examined, among which there were 5 malignant mixed tumors, 4 mammary gland adenocarcinomas, 1 papillary adenocarcinoma, 8 benign mixed tumors and 2 mammary gland adenomas. Positive immunostaining for p53 with PAb240 antibody was found in 2 benign (20%) and 3 malignant (30%) tumors. However, PAb421 antibody did not give positive result at all. In Western blot analysis, the p53 expression in benign and malignant tumors was detected in 4 and 3 cases, respectively. p53 mutations were found in 6 cases out of the cases with detected p53 protein expression. The MIB-1 index in benign and malignant tumors were 17.6+/-20.8% and 29.0+/-27.2%, respectively and there was no significant difference between tumor types. There was a significant correlation between p53 mutations and p53 overexpression (correlation coefficient = 0.5, p < 0.05). In Kaplan-Meier survival analysis, the p53 index was associated with significantly shortened survival time (p < 0.01). In multivariate analysis, p53 overexpression was only an independent factor for indicator of worse prognosis in canine mammary tumors (p = 0.01). These results demonstrated that p53 gene mutations and protein overexpression using the PAb240 anti-p53 antibody were useful predictors of increased malignant potential and poor prognosis in canine mammary tumors.
Animals ; Antibodies, Antinuclear/metabolism ; Antibodies, Monoclonal/metabolism ; Blotting, Western/veterinary ; Dog Diseases/genetics/*metabolism ; Dogs ; Female ; Genes, p53/*genetics ; Immunohistochemistry/veterinary ; Ki-67 Antigen/metabolism ; Mammary Neoplasms, Animal/genetics/*metabolism ; *Mutation ; Predictive Value of Tests ; Proportional Hazards Models ; Tumor Suppressor Protein p53/*biosynthesis/genetics