Objective To investigate the change of splenocyte subsets in Balb/c mice immunized with transgenic alfalfa(Medicago sativa)containing Eg95-EgA31 fusion gene of Echinococcus granulosus(Eg) and challenged with Eg protoscoleces.Methods Leaf protein was extracted from transgenic alfalfa containing Eg95-EgA31 fusion gene by heat-coagulation method,and concentration of 20 g/L was used in the study.Meanwhile,leaf protein extracted from the transgenic alfalfa containing blank vector(pBI121)and the normal alfalfa was served as control.Thirty-two female Balb/c mice were randomly divided into 4 groups,8 mice in each group.Oral group was immunized with the leaf protein containing Eg95-EgA31 fusion antigen intragastrically(100μl per mouse);intranasal group was immunized with the leaf protein containing Eg95-EgA31 fusion antigen intranasally(10 μl per mouse);blank vector group was vaccinated intranasally with 10μl leaf protein with blank vector(pBI121);and normal control group was given 100μl normal leaf protein intragastrically.All mice in the above mentioned groups were immunized every 3 days for 2 months.Then,the mice were challenged intraperitoneally with Eg protoscoleces(50 protoscoleces per mouse)8 weeks after last vaccination and sacrified 24 weeks pest infection to separate the splenocytes.Flow cytometry was used to measure the percentages of CD4+ and CD8+ T ceils subsets.Resuits Compared with the normal control group(0.166±0.018,0.083±0.006,2.019 ±0.369),the percentages of CD4+(0.286±0.009)and CD8+(0.102±0.004)T cell subsets and the ratio of CD4+/CD8+(2.814±0.014)in oral group increased significantly (P<0.01 or<0.05).The percentage of CD4+ subset(0.269±0.016)and the ratio of CD4+/CD8+(2.955±0.986) in intranasal group was significantly higher than that ofthe normal control group(all P<0.01).The percentage of CD4+ subset in oral group was significantly higher than that of the intranasal group(P<0.05).No significant difference was found in the percentages of CD4+ and CD8+ T cell subsets and the ratio of CD4+/CD8+ between the blank vector group(0.169±0.018,0.093±0.019,1.852±0.188)and the normal control group(all P>0.05).Conclusions CD4+ T cell may play an important role in the protection induced by transgenic alfalfa vaccine against the challenge of Eg protoscoleces.Intragastrical immunization may be a good route.

0.05) after the training,while for the students of control group,the levels of IAP and Npt increased significantly after the training(P

ObjectiveTo probe into the maternal care utilization by minority women, for the purpose of policy recommendations on better maternal care in minority areas. MethodsA combination of stratified random sampling and typical sampling was made on 445 married women of reproductive age in six counties in Yunnan, Guizhou, Qinghai and Tibet provinces, a field survey on their utilization of maternal care services. ResultsTheir average prenatal detection rate is 78.24％, a level lower than the national rural average of 93.7％ and grade-4 rural average of 81.2％ in 2008; their post partook rate is 30.7％, lower than the national rural average of 54.3％ and grade-4 rural average of 58.9％ in the same period; their average coverage rate is 52.18％, a level lower than the national rural average of 87.1％and grade-4 rural average of 64.3％ in 2008. ConclusionThe maternal care utilization is found to be low for women in minority areas. Effective solutions are expected for payment of indirect expenditure of hospital delivery; better health education for enhancing health knowledge and health awareness of minority women; effective incentive mechanism for village doctors, consolidating the base of the three-level healthcare network.

Objective: To isolate and screen active marine microorganisms from the East China Sea and to identify the phenotypes of the isolated strains. Methods: Thirty strains of bacteria isolated from the East China Sea were selected randomly, and the active strains were screened out by the anti-Escherichia coli model. The active strains were subjected to physiological and biochemical characteristics, salt-aggregation test, and 16S rDNA sequence analysis. Neighbor-joining trees were constructed by comparing the results of 16S rDNA sequences with sequences described in the BLAST server of the National Center for Biotechnology Information (NCBI); the strains were subsequently identified to genus level. Results: Strains F81612 and F201721 were screened out with the optimum salinities of 10% and 7.5%, respectively. Their morphology and biochemical characteristics were similar to those of Bacillus sp.. 16S rDNA sequence analysis showed that sequences of F81612 had a higher similarity to those of Bacillus subtilis; F201721 was similar to Bacillus amylolique faciens. Conclusion: Two Macrolactin S-producing strains have been screened out by the anti-Escherichia coli model, and they are identified as moderate halophilic Bacillus sp.

Objective: To exploit marine microorganisms and study their secondary metabolites for new drugs. Methods: An antibacterial model was used to screen for active strains. The ethyl acetate (EtOAc) extract was separated by silica chromatography, gel filtration chromatography, and high-performance liquid chromatography. The structures of the compounds were elucidated by 1 HNMR, 13CNMR and MS technologies; Kirby-Bauer Disc Diffusion method and MTT method were employed to detect the biological activities of the separated compounds. Results: Eleven compounds were separated and identified as macrolactin A (1), 3-Hydroxyl acetyl-indole (2), 3-indolethanol (3), cyclo-(Try-Pro) (4), cyclo-(Ile-Try) (5), cyclo-(Leu-Pro) (6), cyclo-(Leu-Val) (7), cyclo-(Ile-Pro) (8), cyclo-(Phe-Val) (9), N- phenethylacetamide (10), P-hydroxy benzaldehyde (PHB) (11). Conclusion: Compound 1 shows strong inhibitory activities against Pyricularia oryzae, Escherichia coli and Staphylococcus aureus (with MIC values being 3.6, 0.45 and 6.3 μg/ml, respectively), and tumor cell lines HeLa and HepG2 (with the IC50 values being 2.0 and 1.8 μg/ml, respectively).

Objective To investigate the dynamic changes of splenocyte cytokines in mice immunized with the transgenic alfalfa containing Eg95-EgA31 fusion gene of Echinococcus granulosus (Eg). Methods Eighty-eight Balb/c mice were divided into 2 groups randomly according to body weights,and immunized orally or intranasally with 100μl or 10μl extracted leaf protein from the transgenic alfalfa(20 g/L) respectively once per 3 days for 2 months. Four mice randomized from each group were killed to get splenocyte on week 0(control),2,4,6,8,10,12,14,16,18 and 20 after the last immunization. The splenocyte were cultured in medium for 48 hours with EgAg or concanavalin A (ConA) stimulation to induce the interleukin (IL)-12,interferon γ(IFN-γ) and IL-10,and cultured for 72 hours with EgAg or lipopolysaccharide (LPS) stimulus to induce the tumor necrosis factor α (TNF-α). Then the supernatant was collected to measure the level of IL-12,IFN-γ,TNF-α and IL-10 by ELISA. Results In the oral immunization group,the level of IL-12,IFN-γ,TNF-α and IL-10 increased significantly from week 4 to week 6,week 2 to week 8,week 2 to week 6 and week 4 to week 12,respectively,reaching the highest level(25.0±5.8)ng/L on week 4,(575.0±28.9)ng/L on week 2,(50.0±11.5)ng/L on week 2 and (42.5±2.9)ng/L on week 8,respectively,as compared with the values on week 0[(11.3±2.5),(125.0±28.9),(11.3±2.5),(12.5±2.9)ng/L,all P < 0.01]; in the intranasal immunization group,it was similar about the values of IL-12,IFN-γ,TNF-α and IL-10 could be seen from week 4 to week 6,week 2 to week 10,week 4 to week 10 and week 6 to week 16,respectively,reaching the highest level(25.0±5.8)ng/L on week 6,(725.0±28.9)ng/L on week 4,(27.5±2.9)ng/L on week 6 and (60.0±11.5)ng/L on week 6,respectively,as compared with the values on week 0[(11.3±2.5),(125.0±28.9),(11.3±2.5),(12.5±2.9)ng/L,all P < 0.01]. The cytokine levels in the groups with EgAg,ConA or LPS stimulus were significantly higher than those in the corresponding splenocyte suspension groups(P < 0.05 or < 0.01),and the cytokine levels in the groups with ConA or LPS stimulus were obviously higher than those in the corresponding groups with EgAg stimulation (P < 0.05 or < 0.01). Conclusion The mixed responses of Th1 and Th2 types can be induced in mice immunized with the transgenic alfalfa in the early period post immunization(2-10 weeks).

AIM: To evaluate the preservation of anterior capsule during vitrectomy and lensectomy.ment (RD) and grade C proliferative vitreoretinopathy (PVR)underwent pars plana vitrectomy (PPV) and pars plana lensectomy (PPL) with preservation and polishing of the anterior capsule. Of the 15 eyes, 4 eyes had giant tear, 3 had recurrent rhegmatogenous retinal detachment (RRD), 2 had diabetic retinopathy. Totally 6 eyes had gas and 9 had silicone oil tamponade. The surgeries were evaluated according to the visual acuity (VA) and the postoperative complications during the follow-up of at least 3 months.in all eyes, improved by 3± 3 lines overall. Eight eyes were implanted posterior chamber intraocular lens (PCIOL) successfully at 2-3 months after operation, including 6 having gas and 2 having silicone oil tamponade. No eyes had central anterior capsule opacity, corneal decompensation, puplillary block, retina redetachment or other complications.an intact anterior capsule in eyes with RD and PVR. Preserving the anterior capsule can help preventing intraoperative and postoperative complications of gas or silicone oil, simplify future PCIOL placement, and maintaining a normal iris appearance.

Objective To cultivate and identify the transgenic affalfa containing Echinococcus granulosus Eg95 gene. Methods The alfalfa plants were transformed by co-cultivating alfalfa cotyledons via recombinant Agrobacterium tumefaciens LBA4404 harboring pBI-Eg95. The transgenic alfalfa explants were selected by kanamyein after calli formation, shoots and roots regeneration in the selective medium, the seedlings of transgenic plants were obtained which were finally transplanted into pots containing nutrient soil. After 2-3 months growth, the complete transgenic alfalfa plants containing Echinococcus granulosus Eg95 gene were obtained. To identify the transgenic alfalfa plants, the total DNA, RNA and leaf protein were extracted from fresh leaf tissue of the transgenic alfalfa plants and confirmed by PCR, RT-PCR, SDS-PAGE and Western blot assay. Results A specific band around 471 bp was amplified by PCR with total DNA, and the same band was obtained by RT-PCR with total RNA, which confirmed that the Eg95 gene was stably integrated into the transformed alfalfa genome. SDS-PAGE analysis showed that the relative molecular mass(Mr) of the expressed protein was about 16.5×103, consistent with the Eg95 protein, and the level of Eg95 expression was up to 0.06% of total soluble leaf protein by Bio-Rad Quantity one assay. Western blot verified the expressed protein was reactive with the sera of mice infected with Echinococcus granulosus. Conclusion The transgenic alfalfa plants containing Echinococcus granulosus Eg95 gene are successfully cultivated.

Objective To construct a prokaryotic expression vector of cystatin C (Cys C), purify Cys C protein produced by the expression system, and prepare its antiserum. Methods Total RNA was isolated from HL-60 cells, and human Cys C gene was amplified with RT-PCR. The cDNA fragment was cloned into pMD18-T vector and which was confirmed by sequencing. The enzyme-digested target fragment was cloned into PET-32(a) expression vector and transfected into E.coli. BL 21(DE3), in which Cys C expression was induced. After the inclusion body protein was purified through Ni2+ affinity chromatography, processed by dialysis, identified by Western blotting, a rabbit was immunized with the fusion protein, and the antiserum was obtained. Results The result of DNA sequence analysis showed that the cloned Cys C gene sequence was completely corresponding to GenBank data. SDS-PAGE and Western blotting showed that the expressed Cys C fusion protein was about 35?103, mainly existing in the inclusion body of E.coli., that could be purified through Ni2+ affinity chromatography. The titer of the antiserum to the purified protein was 1∶8 000 by ELISA, and Western blotting confirmed that the antiserum reacted specifically to the Cys C protein. Conclusion A recombinant Cys C protein and the specific polyclonal antibody have been obtained, which provides a basis for establishment of immunoassays of human Cys C.

Objective To investigate the mechanism of effects of acitretin on the treatment of psoriasis. Methods The expression of vascular endothelial growth factor (VEGF) and micro-vascular density (MVD) were detected by immunohistochemical technique before and after treatment with acitretin in 32 patients with psoriasis. Serum level of VEGF was measured by double antibody sandwich enzyme-linked immunosorbent assay (ELISA). Results The expression of VEGF protein and MVD were significantly higher in psoriatic lesions before treatment with acitretin than those after treatment (P