Schistosomiasis japonica is an endemic, zoonotic disease of major public health importance in China. Vaccination is needed as a complementary approach to the ongoing control programs. In the present study, we determined if the efficacies of DNA vaccine encoding the SjGST and Sj32 asparaginyl endopeptidase protein could be enhanced by boosting with SjGST-32 protein vaccines. Mice were inoculated with a VR1012-SjGST-32 DNA vaccine followed by boosting with rSjGST-32 at 0, 14 and 28 d. Two weeks after the final boost, mice were challenged percutaneously with cercariae. On day 45 following the challenge, all mice were sacrificed and the numbers of recovered worms and hepatic eggs were counted. Moreover, we analyzed the immune response among various vaccination groups. The results showed that DNA vaccine efficacy was enhanced when mice were boosted with protein vaccine. Adult worm and liver egg burdens were reduced 42.3% and 59.6%, respectively. We further found that DNA vaccine followed by boosting with protein significantly increased the IgG titer and T cell proliferation over those seen in mice vaccinated solely with DNA vaccines. Furthermore, the higher level of IFN-gamma expression in the splenetic CD4+ T cell showed that DNA prime-Protein boosting vaccine induced CD4+ Th1-type responses. Thus, DNA vaccine efficacy was significantly enhanced via boosting protein vaccine which might provide a basis for rational application of the Schistosoma vaccine.
Animals ; Antigens, Helminth ; immunology ; Female ; Glutathione Transferase ; administration & dosage ; immunology ; Helminth Proteins ; immunology ; Immunization, Secondary ; methods ; Mice ; Mice, Inbred C57BL ; Recombinant Fusion Proteins ; administration & dosage ; immunology ; Schistosoma japonicum ; Schistosomiasis japonica ; prevention & control ; Vaccination ; methods ; Vaccines, DNA ; administration & dosage ; immunology

This paper is aimed to investigate the transcription and expression of BCR-ABL-pIRES-SEA fusion gene vaccines in vivo in mice. The reconstructed plasmids (BCR-ABL-pIRES-SEA) which were developed previously in our laboratory were injected into the skeletal muscles of BALB/c mice at 14d intervals for three cycles. The transcription and expression of BCR-ABL and staphylococcal enterotoxin A (SEA) in injection site were detected using RT-PCR and immunohistological methods. The BCR-ABL/SEA mRNA and protein could be identified in the injection site of BCR-ABL-pIRES-SEA vaccinated mice. The reconstructed BCR-ABL-pIRES-SEA plasmids can effectively express gene production in the skeletal muscles of mice and have the common features of DNA vaccine.
Animals ; Enterotoxins ; genetics ; immunology ; metabolism ; Fusion Proteins, bcr-abl ; genetics ; immunology ; metabolism ; Male ; Mice ; Mice, Inbred BALB C ; Muscle, Skeletal ; metabolism ; Plasmids ; immunology ; RNA, Messenger ; genetics ; metabolism ; Recombinant Fusion Proteins ; genetics ; immunology ; metabolism ; Vaccines, DNA ; administration & dosage ; immunology

The delivery of transgenes to the central nervous system (CNS) can be a valuable tool to treat CNS diseases. Various systems for the delivery to the CNS have been developed; vascular delivery of viral vectors being most recent. Here, we investigated gene transfer to the CNS by intravenous injection of recombinant adenoviral vectors, containing green fluorescence protein (GFP) as a reporter gene. Expression of GFP was first observed 6 days after the gene transfer, peaked at 14 days, and almost diminished after 28 days. The observed expression of GFP in the CNS was highly localized to hippocampal CA regions of cerebral neocortex, inferior colliculus of midbrain, and granular cell and Purkinje cell layers of cerebellum. It is concluded that intravenous delivery of adenoviral vectors can be used for gene delivery to the CNS, and hence the technique could be beneficial to gene therapy.
Adenoviruses, Human/isolation & purification* ; Animals ; Blood-Brain Barrier ; Brain/virology* ; Cerebellum/cytology ; Cerebellum/virology ; Comparative Study ; Female ; Genes, Reporter ; Genetic Vectors/administration & dosage ; Genetic Vectors/isolation & purification ; Genetic Vectors/pharmacokinetics* ; Hippocampus/virology ; Inferior Colliculus/virology ; Injections, Intravenous ; Luminescent Proteins/analysis ; Luminescent Proteins/biosynthesis ; Luminescent Proteins/genetics ; Mice ; Mice, Inbred BALB C ; Neuroglia/virology ; Neurons/virology* ; Purkinje Cells/virology ; Pyramidal Cells/virology ; Recombinant Fusion Proteins/analysis ; Recombinant Fusion Proteins/biosynthesis ; Recombinant Fusion Proteins/genetics ; Tail/blood supply ; Tissue Distribution

OBJECTIVETo detect the immunoreactivity of targeted fusion anti-caries DNA vaccine pGJA-P in vitro, and the ability to enhance the immune responses compared with the non-targeted fusion anti-caries DNA vaccine pGLUA-P.

METHODSThe CHO cells were transfected with pGJA-P and the expression of recombinant protein in cultured supernatants were detected using Western blotting. 5 to 6-month-old female Japanese rabbits were immunized with either pGJA-P or pGLUA-P via either intramuscular injection (i.m.) or intranasal route (i.n.). The sera and saliva were collected and the antibody responses were checked by ELISA. The effect of immune sera on the synthesis of water-insoluble glucan by glucosyltransferase of S. mutans was examined.

RESULTSThe expressed protein could response to specific anti-GTF antibody. The antibody responses in serum generated by pGJA-P via i.m. were significantly higher than those generated by pGLUA-P (P < 0.01). The antibody responses in saliva generated by pGJA-P via i.n. were significantly higher than those generated by pGLUA-P (P < 0.01). The higher mucosal antibody response induced by pGJA-P via i.m. compared with pGLUA-P (P < 0.01) was detected. The immune sera of rabbits immunized by pGJA-P via i.m. most significantly inhibited the synthesis of water-insoluble glucan by glucosyltransferase.

CONCLUSIONSThe recombinant protein expressed by pGJA-P had the immunoreactivity to anti-GTF antibody. pGJA-P could induce faster and higher specific mucosal SIgA antibody responses via i.n. or serum IgG antibody responses via i.m. compared with non-targeted DNA vaccine, pGLUA-P. High titres of specific mucosal antibodies were found in rabbits immunized with pGJA-P via i.m. The immune sera of rabbits immunized by pGJA-P via i.m. displayed the ability of inhibiting the synthesis of water-insoluble glucan by glucosyltransferase.

Animals ; Bacterial Proteins ; immunology ; CHO Cells ; Cricetinae ; Dental Caries ; prevention & control ; Female ; Glucosyltransferases ; immunology ; metabolism ; Immunoglobulin G ; blood ; Membrane Glycoproteins ; immunology ; Rabbits ; Recombinant Fusion Proteins ; administration & dosage ; immunology ; Streptococcus mutans ; immunology ; Transfection ; Vaccines, DNA ; administration & dosage ; immunology

PURPOSE: To compare the efficacy and safety of recombinant human bone morphogenetic protein (rhBMP) and iliac crest autograft in the fusion treatment of lumbar spondylolisthesis. METHODS: The studies using randomized controlled trials to compare the rhBMP with iliac crest autograft in the treatment of lumbar spondylolisthesis were retrieved from Embase, Pubmed, ProQuest dissertations & theses (PQDT), China national knowledge infrastructure (CNKI), Chinese Biomedical Database, Wanfang Data, Cochrane Library (from March 1998 to March 2018). Postoperative fusion rate, clinical success rate, postoperative intervertebral height, complications, operation time, blood loss and duration of hospitalization were chosen as the outcome indicators. Methodological quality of the trials was critically assessed, and relevant data were extracted. Statistical software Revman 5.3 was used for data-analysis. RESULTS: Eleven articles were included in the meta-analysis. The results showed that, comparing the efficacy of rhBMP with iliac crest autograft, statistical significance was found in the 24-month fusion rate post operation [95% CI (1.38, 24.70), p = 0.02] and operation time [95% CI (-14.22, -2.08), p = 0.008]. There is not sufficient evidence for statistical differences in the remaining indicators. CONCLUSION The current literature shows rhBMP is a safe and effective grafting material in the treatment of lumbar spondylolisthesis. Further evidence is dependent on the emergence of more randomized controlled trials with higher quality and larger sample sizes in the future.
Autografts ; Bone Morphogenetic Proteins ; administration & dosage ; Databases, Bibliographic ; Humans ; Ilium ; transplantation ; Lumbar Vertebrae ; surgery ; Randomized Controlled Trials as Topic ; Recombinant Proteins ; administration & dosage ; Spinal Fusion ; methods ; Spondylolisthesis ; surgery ; Time Factors ; Treatment Outcome

OBJECTIVETo study the effects of immunization with the fusion protein CAC (a product of prokaryotic expression of recombinant HBcAg and β-amyloid peptide fusion gene) against the toxicity induced by intrahippocampal injection of aggregated β-amyloid peptide (Aβ) in rats.

METHODSSD rats were immunized intraperitoneally with the fusion protein CAC, and the titer of anti-Aβ antibody was evaluated by ELISA. When the titers of the anti-Aβ antibody reached 1:3 000, aggregated Aβ was injected into the CA1 region of the rat hippocampus. Two weeks after Aβ injection, the rats underwent morris water maze test before sacrificed to prepare the brain slices with Congo red and haematoxylin staining.

RESULTSThe titer of anti-Aβ antibody reached 1:3 000 after 5 immunizations with the fusion protein. After Aβ injection, the saline-immunized rats showed a reduced cognitive behavior in the Morris water maze test compared to the CAC-immunized rats. In the saline-immunized rats, the neurons around the site of Aβ injection exhibited obvious cell damages with Aβ deposits and glial infiltration, whereas in CAC-immunized rats, Aβ deposits were significantly reduced or even absent.

CONCLUSIONImmunization with the fusion protein CAC can inhibit the toxicity induced by intrahippocampal aggregated Aβ injection.

Amyloid beta-Peptides ; administration & dosage ; biosynthesis ; genetics ; immunology ; Animals ; Antibodies ; blood ; Hepatitis B Core Antigens ; biosynthesis ; genetics ; Hippocampus ; metabolism ; Immunization ; Injections ; Male ; Peptide Fragments ; administration & dosage ; immunology ; Rats ; Rats, Sprague-Dawley ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; immunology

OBJECTIVETo investigate the effect of immunization with prokaryotically expressed recombinant fusion protein of extracellular near-transmembrane domain of Tibet minipig leptin receptor (OBR) on fat deposition in SD rats.

METHODSA pair of specific primers containing BamHI and HindIII restriction enzyme sites was designed to amplify the extracellular near-transmembrane domain (1705-2364 bp) of Tibet minipig OBR gene. After digestion, the amplified fragment was inserted into the plasmid pRSETA between BamHI and HindIII sites. The recombinant plasmid was transformed and expressed in E.coli BL21(DE3) and the product was analyzed by SDS-PAGE and Western blotting. SD rats were immunized with the fusion protein, and the changes in body weight, feed intake, body length, Lee's index, percentage of abdominal fat, liver fat deposition and subcutaneous fat deposition were assessed.

RESULTSThe recombinant fusion protein obtained (about 27.6 kD) was expressed in E.coli induced by IPTG and identified by SDS-PAGE and Western blotting. The rats immunized with the fusion protein showed no significant changes in body weight, body length, Lee's index, percentage of abdominal fat or liver fat deposition as compared with the control rats. Nevertheless, the immunization caused significantly increased feed intake and significantly decreased volume of subcutaneous fat cells.

CONCLUSIONImmunization with the fusion protein of extracellular near-transmembrane domain of Tibet minipig OBR can promote feed intake and suppress subcutaneous fat deposition in SD rats.

Adiposity ; drug effects ; Animals ; Base Sequence ; Female ; Gene Expression ; Genetic Vectors ; Obesity ; Plasmids ; Rats ; Rats, Sprague-Dawley ; Receptors, Leptin ; administration & dosage ; genetics ; Recombinant Fusion Proteins ; administration & dosage ; genetics ; Subcutaneous Fat ; physiology ; Swine ; Swine, Miniature

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

OBJECTIVETo construct, express and purify human fusion proteins composed of a single-chain antibody fragment scFv that recognizes the prostate specific membrane antigen (PSMA) protein, Fdt, HA2 and tp, and to analyze the binding activity of the expressed fusion proteins.

METHODSThe fusion protein genes scFv, scFv-tp, and scFv-Fdt-HA2-tp were amplified by PCR, and the genes obtained were then cloned into the expression vector pET28 and expressed in E. coli BL21. The expressed products were identified by SDS-PAGE and Western blot and purified with Ni(2+)-NTA chelating agarose. The antigen-binding activity of the fusion proteins was determined by ELISA.

RESULTSThe human anti-PSMA fusion gene was successfully constructed and expressed in M15 as the inclusion body after induced with IPTG. All the target proteins expressed could bind the PSMA antigen.

CONCLUSIONFusion proteins can specifically bind the PSMA antigen. This finding contributes to the study of the targeted delivery of siRNA.

Antigens, Surface ; immunology ; Cloning, Molecular ; Electrophoresis, Polyacrylamide Gel ; Escherichia coli ; genetics ; immunology ; Glutamate Carboxypeptidase II ; immunology ; Humans ; Male ; Polymerase Chain Reaction ; RNA, Small Interfering ; administration & dosage ; immunology ; Recombinant Fusion Proteins ; genetics ; immunology ; Single-Chain Antibodies ; genetics ; immunology

BACKGROUNDMany epidemiological and experimental evidences prove that cervical cancers are strongly associated with genital high-risk types of human papillomavirus (HPV). HPV16 is present in 50% of the tumor specimens. Thus, it is important to develop vaccines against HPV16 and cervical cancer. The authors studied the expression of the HPV16 L1DeltaCE7N fusion protein in E. coli and observed its immunogenicity.

METHODSThe fragment of HPV16 L1DeltaC gene and the E7N gene were amplified by PCR separately; the fusion gene named L1DeltaCE7N was generated by fusing E7N to the C terminal of L1DeltaC then the chimeric gene was cloned into prokaryotic expression vector pGEX-2T and expressed in E. coli strain JM109. The L1DeltaCE7N protein expressed were detected by Western blot. Finally its immunogenicity was characterized in immunized mice.

RESULTSIt was proved that the sequence and open reading frame of fusion gene L1DeltaE7N was correct by sequencing; SDA-PAGE gel analysis showed that HPV16 L1/E7 fusion protein was highly expressed in E. coli; the protein was expressed as soluble form and the molecular weight was about 85 x 10(3). The fusion protein could be purified by affinity chromatography and gel filtration. The ELISA result indicated that L1/E7 could elicit specific antibodies against L1 and E7 in immunized mice. In vivo tumor protection test indicated that tumor formation was retarded or prevented in the mice after vaccination with L1/E7, when C57 BL/6 mice were challenged by syngeneic HVP16E6 and E7 transformed tumor cells.

CONCLUSIONHPV16L1/E7 fusion protein was expressed in E. coli, it can be a candidate for prophylactic and therapeutic vaccine for HPV16-associated infection and tumors.

Animals ; Blotting, Western ; Cell Line, Tumor ; Escherichia coli ; genetics ; Female ; Humans ; Immunization ; methods ; Mice ; Mice, Inbred C57BL ; Neoplasm Transplantation ; Neoplasms, Experimental ; immunology ; pathology ; prevention & control ; Oncogene Proteins, Fusion ; genetics ; immunology ; metabolism ; Oncogene Proteins, Viral ; genetics ; immunology ; metabolism ; Papillomaviridae ; genetics ; immunology ; metabolism ; Papillomavirus Infections ; immunology ; pathology ; prevention & control ; Papillomavirus Vaccines ; administration & dosage ; immunology ; Recombinant Fusion Proteins ; immunology ; metabolism ; ultrastructure