OBJECTIVETo investigate the effects of adipose-derived stem cells (ADSC) and non-methylated CpG-oligodeoxynucleotides (CpG-ODN) on the expression of peripheral blood CD4CD25regulatory T (Treg) cells in young mice with food allergy, as well as their immune intervention effects.

METHODSA total of 40 female BALB/c mice were randomly divided into control group, allergic group, ADSC treatment group, and CpG-ODN treatment group, with 10 mice in each group. A mouse model of food allergy was established by intraperitoneal injection and intragastric administration of ovalbumin (OVA) for sensitization and challenge. The mice in the control group were treated with normal saline at the same dose; the mice in the ADSC treatment group were given intraperitoneal injection of ADSC (1×10cells for each mouse) before and after OVA challenge, and those in the CpG-ODN treatment group were given intraperitoneal injection of non-methylated CpG-ODN solution (40 μg for each mouse) at 1 hour before challenge by gavage. The allergic symptom scores were determined for each group after model establishment. ELISA was used to measure the serum level of OVA-IgE. Flow cytometry was used to measure the percentage of peripheral blood CD4CD25Treg cells. Hematoxylin and eosin staining was used for the pathological analysis of the jejunum.

RESULTSThe allergic group had significantly higher allergic symptom scores and serum level of OVA-IgE than the control group (P<0.05). There were no significant differences in the allergic symptom score and the serum level of OVA-IgE between the ADSC treatment group and the CpG-ODN treatment group (P>0.05), but these two groups had significantly lower allergic symptom scores and serum level of OVA-IgE than the allergic group and significantly higher allergic symptom scores and serum level of OVA-IgE than the control group (P<0.01). The allergic group had a significantly lower percentage of peripheral blood CD4CD25Treg cells than the control group (P<0.05). The ADSC treatment group and the CpG-ODN treatment group had a significantly higher percentage of peripheral blood CD4CD25Treg cells than the allergic group (P<0.05); there were no significant differences between these two groups or between them and the control group (P>0.05). Pathological results showed structural damage and edema in the jejunal villi, a large number of eosinophils, and lymphocyte infiltration in the allergic group, while the ADSC treatment group and the CpG-ODN treatment group had less structural damage and edema in the jejunal villi, a lower number of eosinophils, and less lymphocyte infiltration.

CONCLUSIONSADSC and non-methylated CpG-ODN have a certain effect in the treatment of food allergy and can increase the percentage of peripheral blood CD4CD25Treg cells and reduce the level of OVA-IgE. They may be associated with the induction of immune tolerance and these two treatment have comparable effects. Detailed mechanisms of action still need further investigation.

Adipose Tissue ; cytology ; Adjuvants, Immunologic ; pharmacology ; Animals ; Female ; Food Hypersensitivity ; immunology ; therapy ; Immunoglobulin E ; blood ; Mice ; Mice, Inbred BALB C ; Oligodeoxyribonucleotides ; pharmacology ; Ovalbumin ; immunology ; Stem Cell Transplantation ; T-Lymphocytes, Regulatory ; drug effects ; immunology

OBJECTIVETo explore the effect of non-methylated cytosine-phosphate-guanine oligodeoxynucleotides (CpG-ODN) on serum transforming growth factor (TGF)-β and immune regulation in ovalbumin (OVA)-sensitized young mice.

METHODSThirty female BALB/c mice (2-3 weeks old) were randomly divided into control, model, and CpG-ODN intervention groups. A young mouse model of food allergy was established by OVA sensitization. Normal saline of the same volume was used for replacement in the control group. The mice in the intervention group were intraperitoneally injected with CpG-ODN solution 1 hour before every OVA sensitization. Allergic symptoms were observed and scored for each group. The jejunal tissue was histopathologically examined with hematoxylin-eosin staining. Serum OVA-IgE level was measured using ELISA. Serum concentrations of interleukin (IL)-4, interferon (IFN)-γ, and TGF-β were determined by CBA.

RESULTSAllergic symptoms were observed in the model group and the jejunal tissue showed the pathological characteristics of type I allergic reaction. The allergic symptom scores in the model and CpG-ODN intervention groups were significantly higher than in the control group (P<0.01). The serum levels of OVA-IgE, IL-4, and TGF-β were significantly higher in the model group than in the control and CpG-ODN intervention groups (P<0.05). The CpG-ODN intervention group had significantly higher serum levels of OVA-IgE, IL-4, and TGF-β than the control group (P<0.05). Compared with the control and CpG-ODN intervention groups, the model group had a significantly reduced IFN-γ level (P<0.05).

CONCLUSIONSThe serum TGF-β level is increased in the young mouse model of OVA-sensitized food allergy and is involved in the allergy mechanism. Non-methylated CpG-ODN can reduce the serum TGF-β level in sensitized young mice and play an immunoregulatory role in food allergy.

Aging ; Animals ; DNA Methylation ; Female ; Food Hypersensitivity ; drug therapy ; immunology ; Immunoglobulin E ; blood ; Interleukin-4 ; blood ; Mice ; Mice, Inbred BALB C ; Oligodeoxyribonucleotides ; pharmacology ; Ovalbumin ; immunology ; Transforming Growth Factor beta ; blood

OBJECTIVEThis aimed to investigate the effect of specific sequence oligodeoxynucleotide MT01 on the biological properties of osteoblasts invaded by Porphyromonas gingivalis (P. gingivalis ) by evaluating proliferation, cell cycle, and apoptosis.

METHODSMG63 osteoblasts were recovered and incubated with MT01, CpG ODN, metronidazole (MNZ), and gentamicin (GEN) for 3 h. P. gingivalis (the multiplicity of infection was 100:1) was added subsequently and cocultured for another 24 and 48 h. Cells with PBS comprised the blank group, whereas cells with P. gingivalis comprised the negative controls. Six experimental groups were established: PBS group, P. gingivalis group, MT01+P. gingivalis group, CpG ODN+ P. gingivalis group, MNZ+P. gingivalis group, and GEN+P. gingivalis group. The proliferative ability was measured by methyl thiazolyl tetrazolium assay, and the percentages of apoptosis and cell cycle were examined by flow cytometry.

RESULTSCompared with the blank group, proliferation increased significantly in the MT01+P. gingivalis group (P < 0.05). The ratio of cells was lower at the G₁ phase and higher at the S phase in the MT01+P. gingivalis group compared with the results in the P. gingivalis group (P < 0.05). Early cell apoptosis in the MT01+P. gingivalis group was significantly lower than that in the P. gingivalis group (P < 0.05).

CONCLUSIONMT01 can promote the proliferation, reduce the ratio of the G₁phase, increase the ratio of the S phase, and inhibit the early apoptosis of osteoblasts invaded by P. gingivalis.

Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Division ; Cell Proliferation ; drug effects ; Flow Cytometry ; Gentamicins ; pharmacology ; Humans ; Metronidazole ; pharmacology ; Oligodeoxyribonucleotides ; pharmacology ; Osteoblasts ; cytology ; drug effects ; Porphyromonas gingivalis ; pathogenicity

Aptamers are capable of binding a wide range of biomolecular targets with high affinity and specificity. It has been widely developed for diagnostic and therapeutic purposes. Because of unique three dimensional structures and cell-membrane penetration, aptamers inhibit virus infection not only through binding specific target, such as the viral envelope, genomic site, enzyme, or other viral components, but also can be connected to each other or with siRNA jointly achieve antiviral activity. Taking human immunodeficiency virus and hepatitis C virus as examples, this paper reviewed the effects and mechanisms of aptamers on disturbing viral infection and replication steps. It may provide an insight to the development of aptamer-based new antiviral drugs.
Antiviral Agents ; pharmacology ; Aptamers, Nucleotide ; pharmacology ; therapeutic use ; Genome, Viral ; drug effects ; HIV ; drug effects ; HIV Reverse Transcriptase ; metabolism ; Hepacivirus ; drug effects ; genetics ; Humans ; Macular Degeneration ; drug therapy ; Neoplasms ; drug therapy ; Oligodeoxyribonucleotides ; therapeutic use ; RNA, Small Interfering ; pharmacology ; SELEX Aptamer Technique ; Viral Envelope Proteins ; metabolism ; Virus Replication ; drug effects

OBJECTIVETo investigate the protective effect of suppressive oligodeoxynucleotides (Sup ODN) on interferon-γ (IFN-γ) and signal transducers and activators of transcription (pSTAT4) expression of Silica-induced pulmonary inflammation in Mice.

METHODSSixty Balb/c mice were randomly divided into 4 groups, normal control group, silicious group, suppressive oligodeoxynucleotides (Sup ODN) group, control oligodeoxynucleotides (Con ODN) group. Except the normal control group injected normal saline, the rest groups were induced by the intratracheal instillation of 0.1 ml (5 g/L) of sterilized silica suspension. Sup ODN group and Con ODN group were treated by i.p. injection of 0.3 ml (1mg/mL) of suppressive or control ODN 3 h before silica administration. After 7 days, the animals were killed and levels of IFN-γ were detected by ELISA. The pathologic changes in lung tissues of mice were observed with HE staining. Expressions of IFN-γ and pSTAT4 in lung tissue were detected with immunohistochemistry and quantified by Image-Pro Plus 7.0.

RESULTSHE staining showed that the lung tissue of silicious group were damaged seriously than Sup ODN group. Compared with the normal control group (serum: (280.1±41.3) pg/ml, lung tissue: (0.249±0.373), IFN-γ increased in silicious group (serum: (886.3±81.7) pg/ml, lung tissue: (0.270±0.300) (P < 0.05). Compared with the normal control group and Con ODN group [(894.5±91.6) pg/ml], IFN-γ in the serum of Sup ODN group decreased significantly (P < 0.01). Compared with the silicious group , IFN-γ in lung tissue decreased in Sup ODN group (0.241±0.250) (P < 0.05). Compared with the normal control group (0.279±0.353), pSTAT4 in lung tissue increased significantly in silicious group (0.313±0.231) (P < 0.01). Compared with the silicious group, pSTAT4 in lung tissue decreased significantly in Sup ODN group (0.269±0.523) (P < 0.01).

CONCLUSIONSup ODN attained protective effect on Silica treated mice by suppressing expression of IFN-γ and pSTAT4.

Animals ; Female ; Inflammation ; metabolism ; Interferon-gamma ; metabolism ; Lung ; drug effects ; metabolism ; pathology ; Mice ; Mice, Inbred BALB C ; Oligodeoxyribonucleotides ; pharmacology ; Phosphorylation ; STAT4 Transcription Factor ; metabolism ; Silicon Dioxide ; toxicity

Animals ; Inflammation ; chemically induced ; Lung ; drug effects ; Oligodeoxyribonucleotides ; pharmacology ; Pulmonary Alveoli ; drug effects ; pathology ; Silicon Dioxide ; adverse effects

OBJECTIVETo investigate the high expression of HPV16L2N120E7E6 fusion protein by prokaryotic expression system, and evaluate its immunogenicity and antitumor efficacy in vaccinated mice.

METHODSThe HPV16L2N120E7E6 fusion gene, its codons were optimized to increase the expression of the protein, was constructed by overlap extension PCR and inserted into prokaryotic expression vector pET9a. Then the fusion protein was expressed by inducing with IPTG in E. coli strain BL21 (DE3) harboring with plasmid pETL2N120E7E6, and further detected by SDS-PAGE and Western-blot. Finally, the humoral and cellular immune responses were measured by ELISA and ELISPOT, respectively, in vaccinated mice with the purified HPV16L2N120E7E6 fusion protein, and the antitumor efficacy was assessed in mice using the TC-1 tumor challenge model.

RESULTSThe codon-optimized HPV16L2N120E7E6 fusion gene was highly expressed in E. coli strain BL21 (DE3) harboring with plasmid pETL2N120E7E6, and the amount of fusion protein was nearly 48.6% of the total bacterial protein. The purified fusion protein could induce high titer of specific antibody against L2, E7 and E6 in vaccinated mice. When accompanied with the adjuvant CpG, the fusion protein was able to elicit strong and moderate cellular immune responses in vaccinated mice against peptide HPV16E7(49-57) and peptide pools of HPV16E6, respectively. Furthermore, the tumor therapeutic experiment showed that HPV16L2N120E7E6 + CpG could prevent the tumor formation in 80.0% (8/10) vaccinated mice.

CONCLUSIONSThe data of this study suggest that HPV16L2N120E7E6 fusion protein could be a promising candidate vaccine for treatment of chronic HPV16 infection and post-operative adjuvant therapy for cervical cancer.

Adjuvants, Immunologic ; pharmacology ; Animals ; Cancer Vaccines ; immunology ; therapeutic use ; Capsid Proteins ; genetics ; immunology ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; Codon ; Escherichia coli ; immunology ; metabolism ; Female ; Humans ; Immunization ; methods ; Immunotherapy ; methods ; Mice ; Mice, Inbred C57BL ; Neoplasm Transplantation ; Oligodeoxyribonucleotides ; immunology ; Oncogene Proteins, Viral ; genetics ; immunology ; metabolism ; Papillomavirus E7 Proteins ; genetics ; immunology ; metabolism ; Papillomavirus Vaccines ; immunology ; therapeutic use ; Plasmids ; Recombinant Fusion Proteins ; genetics ; immunology ; metabolism ; Repressor Proteins ; genetics ; immunology ; metabolism

We have investigated the effect of various forms of phosphodiester cytidine-phosphate-guanosine oligodeoxynucleotides (CpG ODNs) on the production of pro-inflammatory cytokines and related genes in RAW 264.7 macrophages. Treatment with the CpG ODNs increased the expression of tumor necrosis factor alpha (TNF-alpha), IL-6, and inducible nitric oxide synthase but not interleukin-1beta (IL-1beta). We also investigated the effect of CpG ODNs on the expression of ATP-binding cassette transporter A1 (ABCA1) and G1 (ABCG1) genes which are known to facilitate cholesterol efflux from macrophages for anti-atherosclerosis. CpG 2006 significantly reduced the levels of ABCG1 mRNA as determined by real-time polymerase chain reaction, whereas ABCA1 mRNA level was not changed. Western blot analysis further confirmed the reduction of ABCG1 protein expression by CpG 2006. In addition, we also determined the protein level of peroxisome proliferator activated receptor gamma (PPARgamma), which is recognized as a transcriptional activator of ABC transporters, was also reduced by CpG 2006. Thus, these results suggest that ABCG1 is specifically down-regulated by CpG 2006 in a PPARgamma-dependent manner in macrophages.
ATP-Binding Cassette Transporters/drug effects/genetics/*metabolism ; Animals ; Atherosclerosis/metabolism ; Cholesterol/metabolism ; Cytokines/drug effects/metabolism ; Gene Expression Regulation ; Inflammation/*metabolism ; Interleukin-1beta/drug effects/metabolism ; Interleukin-6/metabolism ; Lipoproteins/drug effects/genetics/*metabolism ; Macrophages/*cytology/*metabolism ; Mice ; Nitric Oxide Synthase/drug effects/metabolism ; Oligodeoxyribonucleotides/*pharmacology ; PPAR gamma/genetics/*metabolism ; Tumor Necrosis Factor-alpha/drug effects/metabolism

CD8-Positive T-Lymphocytes ; metabolism ; Case-Control Studies ; Hepatitis B Surface Antigens ; immunology ; Hepatitis B, Chronic ; immunology ; Humans ; Interferon-gamma ; biosynthesis ; Lymphocytes ; immunology ; Oligodeoxyribonucleotides ; pharmacology

This study focused on prevention and treatment of acute and chronic asthma by oligonucleotides containing unmethylated CpG motifs (CpG-ODNs). Acute and chronic asthma models of mice were made by sensitizing and inhaling ovalbumin (OVA); The number of white blood cells (WBC) and eosnophils (EOS) in bronchoalveolar lavage fluid (BALF) was counted and the concentration of cytokines and vascular endothelial growth factor (VEGF) was examined in BALF by ELISA kit. After that, TLR-9 mRNA was detected in mice spleen cells by reverse transcription polymerase chain reaction (RT-PCR) and TLR-9 protein was determined in mice lung tissues by Western blotting. Compared with acute asthma models of mice, WBC in BALF decreased obviously in the groups of Bordetella pertussis, CpG-ODNs and seq A to seq I which were administrated by both of intragastric (ig) and intraperitoneal (ip) injection group, EOS decreased obviously in Bordetella pertussis, CpG+ and seq A to seq D ig groups, and in all ip administrating groups, although it was not effective in the groups of seq E to seq I. In chronic asthma models of mice, IFN-gamma increased ((1) control: 176.45 +/- 23.46 pg x mL(-1); (2) model: 174.11 +/- 22.71 pg x mL(-1); (3) CpG+ ip: 220.56 +/- 15.42 pg x mL(-1); (4) seq A ip: 225.23 +/- 21.60 pg x mL(-1)) and IL-4 decreased obviously (1) control: 66.91 +/- 5.81 pg x mL(-1); (2) model: 81.02 +/- 11.24 pg x mL(-1); (3) CpG+ ip: 63.99 +/- 6.09 pg x mL(-1); (4) seq A ip: 62.75 +/- 10.03 pg x mL(-1)) in the BALF of CpG+ and seq A ip group, although VEGF was not changed in this research. And also, TLR-9 mRNA in spleen cells (TLR-9/GAPDH: (1) control: 0.62 +/- 0.13; (2) model: 0.66 +/- 0.17; (3) CpG+ ip: 1.46 +/- 0.26; (4) seq A ip: 1.42 +/- 0.34) and TLR-9 protein in lung tissues (TLR-9/beta-actin: (1) control: 0.63 +/- 0.16; (2) model: 0.61 +/- 0.07; (3) CpG+ ip: 1.15 +/- 0.25; (4) seq A ip: 1.03 +/- 0.29) both increased in ip groups, but the change was not significant in ig group. The study confirms that CpG-ODNs and seq A could inhibit airway inflammation remarkably, this mechanism might be related with regulating Th1/Th2 balance and controlling the expression of TLR-9.
Adjuvants, Immunologic ; isolation & purification ; pharmacology ; Animals ; Asthma ; chemically induced ; metabolism ; pathology ; Bordetella pertussis ; Bronchoalveolar Lavage Fluid ; Eosinophils ; drug effects ; Female ; Interferon-gamma ; metabolism ; Interleukin-4 ; metabolism ; Leukocyte Count ; Leukocytes ; drug effects ; Lung ; metabolism ; Mice ; Mice, Inbred BALB C ; Mice, Inbred ICR ; Oligodeoxyribonucleotides ; isolation & purification ; pharmacology ; Ovalbumin ; RNA, Messenger ; metabolism ; Random Allocation ; Signal Transduction ; drug effects ; Spleen ; metabolism ; Th1-Th2 Balance ; Toll-Like Receptor 9 ; genetics ; metabolism ; Vascular Endothelial Growth Factor A ; metabolism