OBJECTIVE: To evaluate the effect of intranasal immunization with CTA1-DD as mucosal adjuvant combined with H3N2 split vaccine. METHODS: Mice were immunized intranasally with PBS (negative control), or H3N2 split vaccine (3 μg/mouse) alone, or CTA1-DD (5 μg/mouse) alone, or H3N2 split vaccine (3 μg/mouse) plus CTA1-DD (5 μg/mouse). Positive control mice were immunized intramuscularly with H3N2 split vaccine (3 μg/mouse) and alum adjuvant. All the mice were immunized twice, two weeks apart. Then sera and mucosal lavages were collected. The specific HI titers, IgM, IgG, IgA, and IgG subtypes were examined by ELISA. IFN-γ and IL-4 were test by ELISpot. In addition, two weeks after the last immunization, surivival after H3N2 virus lethal challenge was measured. RESULTS: H3N2 split vaccine formulated with CTA1-DD could elicit higher IgM, IgG and hemagglutination inhibition titers in sera. Furthermore, using CTA1-DD as adjuvant significantly improved mucosal secretory IgA titers in bronchoalveolar lavages and vaginal lavages. Meanwhile this mucosal adjuvant could enhance Th-1-type responses and induce protective hemagglutination inhibition titers. Notably, the addition of CTA1-DD to split vaccine provided 100% protection against lethal infection by the H3N2 virus. CONCLUSION CTA1-DD could promote mucosal, humoral and cell-mediated immune responses, which supports the further development of CTA1-DD as a mucosal adjuvant for mucosal vaccines.
Adjuvants, Immunologic ; Administration, Intranasal ; Animals ; Cholera Toxin ; Female ; Immunity, Humoral ; Influenza A Virus, H3N2 Subtype ; immunology ; Influenza Vaccines ; Mice, Inbred BALB C ; Nasal Mucosa ; immunology ; Random Allocation ; Recombinant Fusion Proteins

OBJECTIVE: This study aimed to characterize the diagnostic and vaccine potential of a novel Mycobacterium tuberculosis antigen Rv0674. METHODS: To evaluate the diagnostic potential and antigenicity of Rv0674, IgG was evaluated using ELISA and interferon (IFN)-γ was done by using ELISpot assay among TB patients and healthy donors. For immunogenicity evaluation, BALB/c mice were immunized with Rv0674. Cytokine production was determined by cytokine release assay using an ELISA kit, and the antibodies were tested using ELISA. RESULTS: The results of serum Elisa tests showed that Rv0674 specific immunoglobulin G (IgG) response was higher in TB patients than negative controls. And Rv0674 had good performance in serological test with sensitivity and specificity of 77.1% and 81.1%, respectively. While it shows poor sensitivity and specificity of 26.23% and 79.69% for IFN-γ tests. In BALB/c mice, Rv0674 adjuvant by DDA/Poly I:C could also induce a high level of IFN-γ, interleukin-2 and interleukin-6 as well as a high IgG titer in both high- and low-dose groups indicating that Rv0674 is essential in humoral and cellular immunity. Moreover, the cytokine profile and IgG isotype characterized Rv0674 as a Th1/Th2-mixed-type protective immunity with the predominance of Th1 cytokines. CONCLUSION Rv0674 may be a good potential candidate for the development of TB serological diagnosis and a new TB vaccine.
Adult ; Aged ; Animals ; Antigens, Bacterial ; immunology ; Female ; Humans ; Immunity, Cellular ; Immunity, Humoral ; Male ; Mice ; Mice, Inbred BALB C ; Middle Aged ; Tuberculosis ; diagnosis ; immunology ; Young Adult

Myasthenia gravis (MG) is a prototypical antibody-mediated neurological autoimmune disease with the involvement of humoral immune responses in its pathogenesis. T follicular helper (Tfh) cells have been implicated in many autoimmune diseases. However, whether and how Tfh cells are involved in MG remain unclear. Here, we established and studied a widely-used and approved animal model of human MG, the rat model with acetylcholine receptor alpha (AChRα) subunit (R-AChR)-induced experimental autoimmune myasthenia gravis (EAMG). This model presented mild body-weight loss 10 days after the first immunization (representing the early stage of disease) and more obvious clinical manifestations and body-weight loss 7 days after the second immunization (representing the late stage of disease). AChR-specific pre-Tfh cells and mature Tfh cells were detected in these two stages, respectively. In co-cultures of Tfh cells and B cells, the number of IgG2b-secreting B cells and the level of anti-AChR antibodies in the supernatant were higher in the cultures containing EAMG-derived Tfh cells. In immunohistochemistry and immunofluorescence assays, a substantial number of CD4/Bcl-6 T cells and a greater number of larger germinal centers were observed in lymph node tissues resected from EAMG rats. Based on these results, we hypothesize that an AChR-specific Tfh cell-mediated humoral immune response contributes to the development of EAMG.
Animals ; B-Lymphocytes ; immunology ; Disease Models, Animal ; Female ; Immunity, Humoral ; Lymph Nodes ; immunology ; Myasthenia Gravis, Autoimmune, Experimental ; immunology ; Protein Subunits ; immunology ; Proto-Oncogene Proteins c-bcl-6 ; immunology ; Rats, Inbred Lew ; Receptor Cross-Talk ; Receptors, Cholinergic ; immunology ; T-Lymphocytes, Helper-Inducer ; immunology

OBJECTIVETo eliminate the side effects of aluminum adjuvant and His-tag, we constructed chimeric VLPs displaying the epitope of EV71 (SP70) without His-tagged. Then evaluating whether the VLPs could efficiently evoke not only humoral but also cellular immune responses against EV71 without adjuvant.

METHODSThe fusion protein was constructed by inserting SP70 into the MIR of truncated HBcAg sequence, expressed in E. Coli, and purified through ion exchange chromatography and density gradient centrifugation. Mice were immunized with the VLPs and sera were collected afterwards. The specific antibody titers, IgG subtypes and neutralizing efficacy were detected by ELISA, neutralization assay, and EV71 lethal challenge. IFN-γ and IL-4 secreted by splenocytes were tested by ELISPOT assay.

RESULTSHBc-SP70 proteins can self-assemble into empty VLPs. After immunization with HBc-SP70 VLPs, the detectable anti-EV71 antibodies were effective in neutralizing EV71 and protected newborn mice from EV71 lethal challenge. There was no significant difference for the immune efficacy whether the aluminum adjuvant was added or not. The specific IgG subtypes were mainly IgG1 and IgG2b and splenocytes from the mice immunized produced high levels of IFN-γ and IL-4.

CONCLUSIONThe fusion proteins without His-tagged was expressed and purified as soluble chimeric HBc-SP70 VLPs without renaturation. In the absence of adjuvant, they were efficient to elicit high levels of Th1/Th2 mixed immune response as well as assisted by aluminum adjuvant. Furthermore, the chimeric VLPs have potential to prevent HBV and EV71 infection simultaneously.

Adjuvants, Immunologic ; Animals ; Antibodies, Neutralizing ; Antibodies, Viral ; blood ; Enterovirus A, Human ; genetics ; Enterovirus Infections ; immunology ; virology ; Epitopes ; immunology ; metabolism ; Escherichia coli ; metabolism ; Female ; Immunity, Cellular ; Immunity, Humoral ; Mice ; Recombinant Fusion Proteins ; immunology

OBJECTIVETo investigate the changes in serum YKL-40 level and humoral immune function and their significance in children with recurrent pneumonia.

METHODSBlood samples were collected from 30 children with recurrent pneumonia (recurrent pneumonia group), 30 children with acute pneumonia (acute pneumonia group), and 30 healthy children (control group). Serum YKL-40 levels were measured by enzyme-linked immunosorbent assay. The correlation between serum YKL-40 level and laboratory indices related to humoral immune function was analyzed. The receiver operating characteristic (ROC) curve was used to analyze the diagnostic value of serum YKL-40 level for recurrent pneumonia.

RESULTSThe recurrent pneumonia group had a significantly higher serum YKL-40 level than the acute pneumonia and control groups (P<0.05). The acute pneumonia group had a significantly higher serum YKL-40 level than the control group (P<0.05). Serum levels of IgG and complement 4 in the recurrent pneumonia group were significantly lower than in the acute pneumonia group (P<0.05). Serum YKL-40 level was negatively correlated with serum IgG level (r=-0.309, P=0.047) and serum complement 4 level (r=-0.324, P=0.039). The area under the ROC curve of serum YKL-40 level for diagnosing recurrent pneumonia was 0.958 (95%CI: 0.921-0.994).

CONCLUSIONSHumoral immune function is low in children with recurrent pneumonia. Serum YKL-40 may be involved in the occurrence of recurrent pneumonia and can be used as a reference index for diagnosing recurrent pneumonia.

Child, Preschool ; Chitinase-3-Like Protein 1 ; blood ; Complement C4 ; analysis ; Female ; Humans ; Immunity, Humoral ; Immunoglobulin G ; blood ; Male ; Pneumonia ; immunology ; Recurrence

To analyze the immunogenicity and protective ability of recombinant IgG-binding protein (EAG) of Streptococcus equi subspecies equi and to evaluate its value when used as equine vaccine antigen, EAG gene was amplified by PCR and inserted into pET-28a vector. The EAG recombinant proteins were expressed and purified to immune mice. The serum antibody and challenge protection were tested. The purified recombinant protein of EAG was 26 kDa, and the protein reacted specifically with positive serum of Streptococcus equi subspecies equi. The mice antibody level for EAG immunization group was 1∶8 100. The immunological protection result showed that the protection rate of the EAG recombinant protein was 90%. The results suggested that the EAG protein has good immunogenicity and immunological protection, and it can effectively increase the humoral immune response and immunological protection of mice.
Animals ; Antibodies, Bacterial ; blood ; Antigens, Bacterial ; immunology ; Bacterial Proteins ; immunology ; Bacterial Vaccines ; immunology ; Immunity, Humoral ; Immunoglobulin G ; blood ; Mice ; Polymerase Chain Reaction ; Protein Binding ; Recombinant Proteins ; immunology ; Streptococcal Infections ; prevention & control ; Streptococcus equi ; Vaccination

OBJECTIVETo construct a recombinant human adenovirus type 5 (Ad5) expressing luciferase and GFP reporter gene and detect neutralizing antibodies against adenovirus type 5 in common marmosets (Callithrix jacchus) to provide basic laboratory data for evaluating adenovirus vaccines.

METHODSLuciferase and GFP reporter genes from plasmid pHAGE-CMV-GFP were inserted into pDC315 to construct the recombinant adenovirus shutter plasmid pDC315-Luc-GFP. The shutter plasmid was co-transduced with pBHGlox(delta)E1,3Cre in 293A cell line to package the recombinant adenovirus rAd5/Luc/GFP. Three rounds of plaque formation experiment were performed to select the monoclonal adenovirus followed by purification with cesium chloride density gradient centrifugation and virus titration with TCID50 method. Chemiluminescence assay and flow cytometry were employed to detect the neutralizing antibody levels in 14 common marmosets.

RESULTSThe shuttle plasmid pDC315-Luc-GFP was successfully constructed and the recombinant adenovirus rAd5/Luc/GFP was packaged with a the titer reaching 6.9×10(11.5) PFU/mL. In the 14 marmosets, chemiluminescence assay identified 4 (28.6%) marmosets that were positive for Ad5-neutralizing antibodies, including 2 with a antibody titer of 1/16 and another 2 with a titer of 1/32; flow cytomery detected Ad5-neutralizing antibodies in 3 marmosets at the titer of 1/16.

CONCLUSIONChemiluminescence assay is a simple, sensitive, and accurate modality for detecting Ad5-neutralizing antibodies. Common marmosets have a very low positivity rate for Ad5-neutralizing antibodies and are therefore promising models for studying adenovirus-based vaccines and therapies.

Adenoviruses, Human ; immunology ; Animals ; Antibodies, Neutralizing ; blood ; Antibodies, Viral ; blood ; Callithrix ; Cell Line ; Humans ; Immunity, Humoral ; Luciferases ; Plasmids

EPSAH is an exopolysaccharide from Aphanothece halophytica GR02. The present study was designed to evaluate its toxicity and adjuvant potential in the specific cellular and humoral immune responses in ovalbumin (OVA) in mice. EPSAH did not cause any mortality and side effects when the mice were administered subcutaneously twice at the dose of 50 mg·kg(-1). Hemolytic activity in vitro indicated that EPSAH was non-hemolytic. Splenocyte proliferation in vitro was assayed with different concentrations of EPSAH. The mice were immunized subcutaneously with OVA 0.1 mg alone or with OVA 0.1 mg dissolved in saline containing Alum (0.2 mg) or EPSAH (0.2, 0.4, or 0.8 mg) on Day 1 and 15. Two weeks later, splenocyte proliferation, natural killer (NK) cell activity, production of cytokines IL-2 from splenocytes, and serum OVA-specific antibody titers were measured. Phagocytic activity, production of pro-inflammatory cytokines IL-1 and IL-12 in mice peritoneal macrophages were also determined. EPSAH showed a dose-dependent stimulating effect on mitogen-induced proliferation. The Con A-, LPS-, and OVA-induced splenocyte proliferation and the serum OVA-specific IgG, IgG1, and IgG2a antibody titers in the immunized mice were significantly enhanced. EPSAH also significantly promoted the production of Th1 cytokine IL-2. Besides, EPSAH remarkably increased the killing activities of NK cells from splenocytes in the immunized mice. In addition, EPSAH enhanced phagocytic activity and the generation of pro-inflammatory cytokines IL-1 and IL-12 in macrophages. These results indicated that EPSAH had a strong potential to increase both cellular and humoral immune responses, particularly promoting the development of Th1 polarization.
Adjuvants, Immunologic ; administration & dosage ; Animals ; Cyanobacteria ; chemistry ; Female ; Immunity, Cellular ; Immunity, Humoral ; Immunization ; Interleukin-12 ; immunology ; Interleukin-2 ; immunology ; Killer Cells, Natural ; immunology ; Mice ; Mice, Inbred ICR ; Ovalbumin ; immunology ; Polysaccharides ; administration & dosage ; immunology ; Rabbits ; Th1 Cells ; immunology ; Th2 Cells ; immunology

OBJECTIVE: To explore the effects of pure adenoidectomy,adenoidectomy with partial tonsillectomy,and adenoidectomy with total tonsillectomy on humoral and cellular immunity of children with OSAHS. METHOD: The children with OSAHS diagnosed by polysomnography were divided into pure adenoidectomy group(group A),adenoidectomy with partial tonsillectomy group(group B), and adenoidectomy with total tonsillectomy(group C), and there were 50 cases in each group. The serum IgG, IgA, IgM level and peripheral blood T cell subgroup per-centage were detected at 6 months preoperatively and postoperatively. Tonsil grading and polysomnography wereconducted, recording symptoms improvement situation at postoperative 6 months. RESULT: There was no statisticallysignificant difference compared with preoperative(P>0. 05) in humoral immunity and cellular immunity index ofpostoperative 6 months. There was no significani difference(P>C. 05) in curative effect among three groups in the 6th month post-operatively. CONCLUSION All of these three surgical procedures had no obvious effect on humoral orcellular immune function in children, and could effectively treat children OSAHS.
Adenoidectomy ; Antibodies ; blood ; Child ; Humans ; Immunity, Cellular ; Immunity, Humoral ; Palatine Tonsil ; pathology ; Polysomnography ; Postoperative Period ; Sleep Apnea, Obstructive ; immunology ; surgery ; T-Lymphocyte Subsets ; cytology ; Tonsillectomy

To enhance the immunogenicity of DNA and adenoviral vector vaccines expressing HIV-1 subtype B gp160, human interleukin 15 (hIL15) DNA adjuvant (pVR-hIL15) was constructed. BALB/c mice received DNA prime/protein boost immunization with pVR-HIVgp160/Ad5-HIVgp160 alone or combined with pVR-hIL15. Cellular and humoral immune responses were evaluated by IFN-gamma enzyme-linked immunosorbent spot assay and enzyme-linked immunosorbent assay, respectively. Compared with those immunized with vaccines alone, the mice immunized with vaccines combined with pVR-hIL15 had significantly increased specific cellular response and antibody titer (P < 0.05). It suggests that the IL15 DNA adjuvant can enhance the immune responses induced by prime-boost regimen using DNA and adenoviral vector encoding HIV-1 subtype B gp160.
Adenoviridae ; genetics ; Adjuvants, Immunologic ; Animals ; Antibodies, Viral ; immunology ; Antibody Specificity ; Female ; Genetic Vectors ; genetics ; HIV Envelope Protein gp120 ; immunology ; HIV Envelope Protein gp160 ; genetics ; immunology ; HIV Envelope Protein gp41 ; immunology ; Humans ; Immunity, Cellular ; Immunity, Humoral ; Interleukin-15 ; genetics ; Mice ; Mice, Inbred BALB C ; Vaccines, DNA ; genetics ; immunology