In order to explore the available sero-diagnotic method for surveillance in basically controlled and controlled areas of schistosomiasis japonica, a 3 years longitudinal investigation of children 6~14 years old with COPT in 5 endemic areas was carried out after being basically controlled for 8~16 years. The results showed that the examination of children with COPT is one of the available serodiagnostic methods of schistosomiasis for surveillance. In all five areas the positive rate of children in this age group was less than 3%, and the circumoval precipitin rate among the positive cases was less than 5%. Therefore, it is indicated that the above mentioned two indexes can be used for sero-surveillance of schistosomiasis japonica.

Objective To construct and evaluate a novel PAMAM dendrimer vector-DNA vaccine for schistosomiasis japonica.Methods Lysine was used to modify 4.0G PAMAM.and the modified product PAMAM-Lys was synthesized.Agarose gel electrophoresis was used to confirm the composite ratio of plasmid DNA and dendrimer.Micrestructure of the compound was observed by using transmission electronic microscopy,and the stability was analyzed by using electrophoresis.The viability of the cells transfected with dendrimers was evaluated by using a MTT technique in vitro.Fiftyty mice were immunized with purified plasmid pJW4303,pJW4303-Sj23 dendrimer PAMAM-Lys and compound PAMAM-Lys/pJW4303-Sj23,respectively.The specific antibodies of the mice in each group were detected to access the immunoreactivity.Results The agarese gel electrophoresis showed that when the charge ratio of the dendrimer vector and DNA was between 2 and 4,the positive and negative charges could be counteraeted completely,and the compound was blocked completely by DNA electrophoresis.The obscrvation results with transmission electronic microscopy showed that the composition of dendrimer vector and DNA caused shrink of DNA structure.Dendrimer-DNA compound had a good stability.MTT showed the modified dendrimer vector and DNA compound system produced a lower cell toxicity on 293T cell than the unmodified Ones.Thk levels of specific antibodies of the mice immunized with PAMAM-Lys/pJW4303Sj23 were significantly higher than those of the mice immunized with naked DNA vaccine(P<0.05).Conclusions The lysinemodified PAMAM-lys is an excellent vector,and has an appropriate biocompatibility.Lysine-modification can reduce the cell toxicity of PAMAM dendrimer significantly.PAMAM-Lys can enhance the immunoreactivity of DNA vacmine which merits further application in schistosomiasis DNA vaccine.

Objective To enhance the protective immunity effects against Schistosoma japonicum infection by priming with cocktail DNA vaccines and boosting with cocktail protein vaccines in infected BALB/c mice.Methods Plasmids and proteins for immunization were prepared and diluted in no bacterial saline solution to final concentration of 1.5 mg/ml,and mixed with pcDNA3.1-SjC23,pcDNA3.1-SjCTPI,pcDNA3.1-(CDR3)6 plasmid DNAs by equal volume to form the cocktail DNA vaccine,and also mixed with recombinant proteins SjC23-HD,SjCTPI,and NP30 by equal volume to form the cocktail protein vaccine.Seventy female BALB/c mice of 4-5 weeks old were randomly divided into 5 groups(A,B,C,D,E).In Group A(control group),each mouse was immunized with 100 ?l saline solution by intramuscular(i.m.);in Group B(pcDNA3.1 control group),each mouse was immunized(i.m.)with 100 ?l pcDNA3.1 for three times at week 0,3,6;in Group C(pcDNA3.1 and cocktail protein group),each mouse was immunized(i.m.)with 100 ?l pcDNA3.1 for three times at week 0,3,6 and immunized with 100 ?l mixed protein vaccines plus 100 ?l FCA by subcutaneous at week 9;in Group D(cocktail DNA vaccines group),each mouse was immunized(i.m.)with 100 ?l mixed DNA vaccines for three times at week 0,3,6;in Group E(cocktail DNA vaccines plus cocktail proteins),each mouse was immunized(i.m.)with 100 ?l mixed DNA vaccines for three times at week 0,3,6 and immunized with 100 ?l mixed protein vaccines plus 100 ?l FCA by subcutaneous at week 9.Four weeks after the last DNA immunization or two weeks after protein boosting,all the mice were challenged with(40?1)cercariae of Schistosoma japonicum by abdominal skin penetration at the same time.Forty-two days post-challenge,the mice were sacrificed and perfused,and the numbers of recovered worms and eggs in liver were counted.The blood was collected from the tail veins of all the mice two days before the first immunization and challenge,respectively,the serum was prepared for detection of IgG,IgG1 and IgG2a.Two days before the challenge,the spleen cells of two mice from each group were cultured and stimulated with ConA and soluble egg antigen(SEA),and the supernatant was collected for detection of IL-2,IL-4 and IFN-?.Results The worm reduction rates in Group C,D and E were 17.70%,32.88% and 45.35%,respectively,compared with the control group.The worm reduction rates in Group D and E were significantly higher than that in Group C(P

Objective To construct, express and purify human scFv antibody (S1) against the recombinant SAG1 of Toxoplasma fused to magainin, and observe its protective effect against Toxoplasma in infected mice. Methods The S1 scFv antibody gene amplified from phagmid S1/pIT-2 fused to magainin was cloned into procaryotic expression vector pET-32c. The recombinant plasmid S1M/pET-32c proved by DNA sequencing was transformed into E.coli BL21, and induced for fusion expression of S1M with IPTG. The expressed S1M was purified with Ni 2+ chelating HiTrap HP column and detected with SDS-PAGE. The effect of reduction of infection of Toxoplasma was observed through in vivo and in vitro experiments in mice. Results The fused gene of S1 and magainin was successfully cloned into procaryotic expression vector pET-32c proved by DNA sequencing. The recombinant S1M protein about 43 kDa was expressed in E.coli as inclusion body, and prepared with Ni 2+ column purification. Tachyzoite of Toxoplasma preincubated with S1M showed decreased infectivity in mice, the result of in vivo experiments showed that mice treated with S1M hadlonger survival time than the mice untreated. Conclusion The purified targeting antimicrobial peptide S1M could reduce the infectivity of tachyzoites of Toxoplasma in a certain extent and has a potential value for biological therapy of toxoplasmosis; otherwise, the constructed targeting antimic robial peptide S1M also provides a new model for biological therapy of toxoplasmosis.

Objective To study the protective effect of codon optimized TPI DNA vaccine against Schistosoma japonicum infection.Methods Sixty female BALB/c mice were randomly divided into 5 groups.The mice were injected through musculus quadriceps fexoris with 100 ?g pcDNA 3.1 control(Group A), pcDNA3.1-TPI(Group B), pcDNA 3.1-TPI-mHSP70(Group C), pcDNA3.1-TPI.opt(Group D), and pcDNA3.1-TPI.opt-mHSP70(Group E) respectively.All mice were immunized for three times with an interval of two weeks.The mice were challenged with(40?1) cercariae of S.japonicum per mouse by abdominal skin penetration 4 weeks after the last immunization, and sacrificed at 42 days post-challenge, the number of worms or hepatic eggs was counted.Blood was taken for the detection of IgG, IgG1, and IgG2a 2 days before immunization and before challenge, respectively.Spleen cells of 2 mice from each group were cultured and stimulated with ConA and rSjCTPI peptide, and the supernatant was collected for detection of IL-2, IL-4, IL-5, IFN-?, and TNF by flow cytometry.Results ELISA showed that the mice in groups B, C, D, and E produced specific IgG and IgG1, IgG2a antibody isotypes, and the ratio of IgG2a/IgG1 was 1.73, 2.06, 2.44, and 3.09, respectively.The levels of IL-2, IFN-? and TNF in groups D and E were higher than that of groups B and C.The worm reduction rate and hepatic egg reduction rate in groups D(36.03%, 41.7%) and E(39.03%, 46.85%) were higher than those of groups B(26.28%, 28.35%) and C(28.38%, 31.39%)(P

Objective To enhance the protective immunity effects of nucleic acid vaccines against Schistosoma japonicum infection by electroporation(EP)in vivo in infected BALB/c mice.Methods Plasmids and proteins for immunization were prepared and diluted in no bacterial saline solution to final concentration of 1.5 mg/ml.pcDNA3.1-SjC23,pcDNA3.1-SjCTPI,pcDNA3.1-(CDR3)6 plasmid DNAs were mixed by equal volume to form the cocktail DNA vaccine,and also mixed with recombinant proteins SjC23-HD,SjCTPI,and NP30 by equal volume to form the cocktail protein vaccine.Seventy female BALB/c mice of 4-5 weeks old were randomly divided into 5 groups(A,B,C,D,E).In Group A(control group),each mouse was immunized with 100 ?l saline solution by intramuscular(i.m.);in Group B(pcDNA3.1/EP control group),each mouse was immunized(i.m.)with 100 ?l pcDNA3.1 followed by EP in vivo for three times at week 0,3,6;in Group C(pcDNA3.1/EP plus cocktail protein vaccine group),each mouse was immunized(i.m.)with 100 ?l pcDNA3.1 followed by EP for three times at week 0,3,6 and boosted with 100 ?l cocktail protein vaccine plus 100 ?l FCA by subcutaneous at week 9;in Group D(cocktail DNA vaccine/EP group),each mouse was immunized(i.m.)with 100 ?l cocktail DNA vaccine followed by EP for three times at week 0,3,6;in Group E(cocktail DNA vaccine/EP plus cocktail protein vaccine group),each mouse was immunized(i.m.)with 100 ?l cocktail DNA vaccine followed by EP for three times at week 0,3,6 and boosted with 100 ?l cocktail protein vaccine plus 100 ?l FCA by subcutaneous at week 9.Four weeks after the last DNA immunization or two weeks after protein boosting,all the mice were challenged with(40?1)cercariae of Schistosoma japonicum by abdominal skin penetration.Forty-two days post-challenge,the mice were sacrificed and perfused,and the numbers of recovered worms and eggs in liver were counted.The blood was collected from the tail veins of all the mice two days before the first immunization and challenge,respectively,the serum was prepared for detection of IgG,IgG1 and IgG2a.Two days before the challenge,the spleen cells of two mice from each group were cultured and stimulated with ConA and soluble egg antigen(SEA),and the supernatant was collected for detection of IL-2,IL-4 and IFN-? by flow cytometre.Results The worm reduction rates in Group C,D and E were 18.09%,45.00% and 57.09%,respectively,compared with the control group.The worm reduction rates in Group D and E were significantly higher than that in Group C(P

Objective To predict B cell epitopes in Sj22, Sj23, Sj14-3-3, Sj26 of Schistosoma japonicum with bioinformatics, and evaluate the antigenicity of these epitope proteins. Methods The complete DNA sequences of S.japonicum were predicted by BioSun system, the target B cell epitope genes were selected, cloned and expressed. The expressed fusion proteins were detected with the sera of schistosomiasis patients and health people for evaluation of their antigenicity. Results Eight B cell epitopes from four molecules of S.japonicum were predicted. The B cell epitopes of Sj22 probably located in 56-62 and 127-133 amino acids. The B cell epitopes of Sj23 probably located in 149-156 and 160-167 amino acids. The B cell epitopes of S14-3-3 probably located in 118-125 and 130-137 amino acids. The B cell epitopes of Sj26 probably located in 143-149 and 191-197 amino acids. The predicted epitope genes were cloned into pET-32c plasmid and expressed. Three of eight expressed fusion proteins of epitopes were reacted with the sera of schistosomiasis patients but not with health people. Conclusion Three epitope antigens with potential diagnosis value are determined.

Objective To understand the potential impact of south-north water transfer project on transmission and distribution of Schistosomiasis japonica, and to put forward the countermea-sures of prevention of the disease transferring into other places. Methods The information on the progress of south-north water transfer project and factors related to the distribution of Schistosoma juponicum were collected, and the suggestions on improving the countermeasures were obtained through the group discussions and field visits. Results The potential impact of the project on the disease transferring is existed, mainly the disease transferring will be through the Lixia River basin in Jiangsu Province, and Chaohu areas of Anhui Province in the east route, and Sihu areas of Hubei Province in the middle route. The snail transferring northward will be affected both by the project and global warming, as a result, the endemic areas of Schistosomiasis will probably transfer into the Hongzehu and Chaohu areas in the future. Conclusion In the east route of the project, if the project is not combined with Schistosomiasis control, the endemic areas of Schistosomiasis will extend into other regions, the loss in the society and economy will be very large.

Paget's disease of bone (PDB) is a common metabolic bone disease that is characterized by aberrant focal bone remodeling, which is caused by excessive osteoclastic bone resorption followed by disorganized osteoblastic bone formation. Genetic factors are a critical determinant of PDB pathogenesis, and several susceptibility genes and loci have been reported, including SQSTM1, TNFSF11A, TNFRSF11B, VCP, OPTN, CSF1 and DCSTAMP. Herein, we report a case of Chinese familial PDB without mutations in known genes and identify a novel c.163G>C (p.Val55Leu) mutation in FKBP5 (encodes FK506-binding protein 51, FKBP51) associated with PDB using whole-exome sequencing. Mutant FKBP51 enhanced the Akt phosphorylation and kinase activity in cells. A study of osteoclast function using FKBP51V55L KI transgenic mice proved that osteoclast precursors from FKBP51V55L mice were hyperresponsive to RANKL, and osteoclasts derived from FKBP51V55L mice displayed more intensive bone resorbing activity than did FKBP51WT controls. The osteoclast-specific molecules tartrate-resistant acid phosphatase, osteoclast-associated receptor and transcription factor NFATC1 were increased in bone marrow-derived monocyte/macrophage cells (BMMs) from FKBP51V55L mice during osteoclast differentiation. However, c-fos expression showed no significant difference in the wild-type and mutant groups. Akt phosphorylation in FKBP51V55L BMMs was elevated in response to RANKL. In contrast, IκB degradation, ERK phosphorylation and LC3II expression showed no difference in wild-type and mutant BMMs. Micro-CT analysis revealed an intensive trabecular bone resorption pattern in FKBP51V55L mice, and suspicious osteolytic bone lesions were noted in three-dimensional reconstruction of distal femurs from mutant mice. These results demonstrate that the mutant FKBP51V55L promotes osteoclastogenesis and function, which could subsequently participate in PDB development.
Acid Phosphatase ; Animals ; Asian Continental Ancestry Group ; Bone Diseases, Metabolic ; Bone Remodeling ; Bone Resorption ; Femur ; Humans ; Mice ; Mice, Transgenic ; Osteitis Deformans* ; Osteoblasts ; Osteoclasts ; Osteogenesis ; Phosphorylation ; Phosphotransferases ; Tacrolimus Binding Proteins ; Transcription Factors