ABSTRACT
<p><b>BACKGROUND</b>Allergen-specific immunotherapy can induce immune tolerance to specific allergens by regulating immune status of individuals. However, its clinical application is limited due to individual differences in efficacy among patients and un-confirmed safety. 1,25 Dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) has been shown to be involved in a variety of physiological processes, including immune response regulation. In the present study we explored the role of 1,25(OH)(2)D(3) pretreatment for immunotherapy.</p><p><b>METHODS</b>Seventy-five BALB/c mice were randomly divided into five groups (15 mice per group). The mouse allergic asthma model was established by intra-peritoneal injection of ovalbumin (OVA, 10 µg) and aluminium hydroxide (2 mg) as an adjuvant. Intra-peritoneal injection of 50 ng of 1,25(OH)(2)D(3) served as a pretreatment, subcutaneous injection of OVA (100 µg) as an immunotherapy, and 1% OVA inhalation as a challenge. Histopathological analysis was performed on four mice per group. The number of cells and their classification in bronchoalvolar lavage (BAL) fluid were assayed. Levels of serum OVA-specific immunoglobulin E (sIgE) and IFN-γ, IL-4, IL-5 and IL-10 in BAL fluid were measured by ELISA.</p><p><b>RESULTS</b>After 1,25(OH)(2)D(3) pretreatment, immunotherapy could significantly inhibit the infiltration of inflammatory cells into lung tissues and BAL fluid of mice with allergic asthma when compared with un-treated animals (eosinophils: (7.46 ± 1.34) × 10(4)/ml vs. (13.41 ± 1.67) × 10(4)/ml, P < 0.05). In addition, levels of IL-4 ((36.91 ± 7.87) pg/ml vs. (43.70 ± 6.42) pg/ml, P > 0.05) and IL-5 ((41.97 ± 7.93) pg/ml vs. (60.14 ± 8.35) pg/ml, P < 0.05) in BAL fluid and serum sIgE ((0.42 ± 0.05) vs. (0.75 ± 0.06) OD units, P < 0.05) were profoundly reduced. However, the IL-10 level in BAL fluid was significantly increased ((67.74 ± 6.57) pg/ml vs. (44.62 ± 8.81) pg/ml, P < 0.05).</p><p><b>CONCLUSIONS</b>These results indicated that 1,25(OH)(2)D(3) pretreatment enhanced the inhibitory effects of immunotherapy on allergic airway inflammation. In the treatment of allergic diseases, 1,25(OH)(2)D(3) pretreatment may be beneficial for improving the efficacy of immunotherapy.</p>
Subject(s)
Animals , Female , Mice , Asthma , Allergy and Immunology , Pathology , Therapeutics , Bronchoalveolar Lavage Fluid , Allergy and Immunology , Calcitriol , Therapeutic Uses , Cytokines , Desensitization, Immunologic , Disease Models, Animal , Immunoglobulin E , Blood , Mice, Inbred BALB C , Ovalbumin , Allergy and ImmunologyABSTRACT
<p><b>OBJECTIVE</b>To investigate whether insertion of TC motif into hepatitis B virus (HBV) core protein c/e1 site affects the expression of S and e antigen.</p><p><b>METHODS</b>Different oligonucleotides encoding TC motif were inserted into the c/e1 site of the core gene of a 1.3 copy wild-type HBV genome vector. HepG2 cells were divided into several groups of cells to transiently transfect with the wild-type and mutant HBV vectors, respectively. In each group, the expression level of core protein inside cells was detected by western blotting, and the levels of S and e antigen in culture medium were analyzed by ELISA assay.</p><p><b>RESULTS</b>Western blotting showed that these TC-tagged core proteins were expressed at similar level of wild-type one. ELISA assay indicated that the level of S and e antigen in culture medium of different groups were not significantly different.</p><p><b>CONCLUSION</b>Insertion of TC motif into HBV core protein c/e1 site does not interference with the expression of viral protein encoded by HBV genome.</p>
Subject(s)
Humans , Amino Acid Sequence , Blotting, Western , Enzyme-Linked Immunosorbent Assay , Genetic Vectors , Genetics , Metabolism , Hep G2 Cells , Hepatitis B Core Antigens , Genetics , Metabolism , Hepatitis B Surface Antigens , Metabolism , Hepatitis B e Antigens , Metabolism , Hepatitis B virus , Genetics , Mutagenesis, Insertional , Recombinant Proteins , Genetics , Metabolism , Transfection , Viral Core Proteins , Genetics , Metabolism , Viral Proteins , Genetics , Metabolism , Virus ReplicationABSTRACT
<p><b>OBJECTIVE</b>To explore the in vitro immune response to taxol resistance associated antigen 3 (TRAG-3)-derived cytotoxic T lymphocyte (CTL) epitope-pulsed dendritic cell (DC).</p><p><b>METHODS</b>The HLA-A2.1 restricted CTL epitope of TRAG-3 was previously identified to be a nonameric peptide sequence from 58 to 66 amino acid residues (TRAG-3(58-66)). The peptide was synthesized according to Fmos procedure, purified and molecular weight determined. Peripheral blood mononuclear cells (PBMC) of normal HLA-A2.1(+) individuals were used to isolate DC and DCs' phenotype identified by flow cytometry. Induction of CTL was achieved by in vitro stimulation of HLA-A2.1(+) PBMC with peptide-pulsed DC. CTL activity against HLA-A2.1(+)/TRAG-3(+) melanoma cell line LB373-MEL was assessed by (51)Cr release assay and IFN-gamma released by ELISA.</p><p><b>RESULTS</b>The synthetic nonameric peptide was above 90% pure and the determined values of molecular weight were conformed to their theoretical values. The phenotype of the isolate DCs was characteristic for mature ones. PBMC stimulated in vitro by TRAG-3-derived epitope-pulsed DCs for three times could specifically kill the target cells and secreted high concentration of IFN-gamma.</p><p><b>CONCLUSION</b>TRAG-3-derived epitope-pulsed DC can elicit specific anti-tumor immune response in vitro. Clinical trials using it as tumor vaccine may be feasible as specific immunotherapy for patients with TRAG-3 expressing cancers.</p>