Objective To investigate the stability of lumbar column after limited selective posterior rhizotomy(LSPR) and observe the healing of exscinded laminae and the deformation of lumbar column. Methods One hundred and twenty five children with spastic cerebral palsy were divided into six groups(A, B, C, D, E, F)according to the age(

Objective To detect Pneumocystis carinii (Pc) DNA by loop-mediated isothermal amplification (LAMP). Methods After injected with hydrocortisone acetate for 8 weeks, the bronchoalveolar lavage fluid (BALF) of Wistar rats were collected and a portion of BALF were examined for identifying Pc organisms using microscope. Then Pc DNA was extracted by phenol-chloroform extraction. Four primers which recognized 6 distinct regions on the mtrRNA gene of Pc were designed and used for LAMP assay. To evaluate the specificity of the assay, M. tuberculosis, M. pneumoniae, C. pneumoniae, P. gondii and rat leucocyte were used as negative controls. To compare the sensitivity of the LAMP to that of conventional PCR, Pc DNA were 10-fold serially diluted and was amplified by LAMP and PCR. LAMP results were judged by naked eye, electrophoretic analysis and restriction digestion. Results Pc organisms were detected from BALF of rats injected with hydrocortisone acetate. After LAMP reaction, positive signal was observef rats injected with hydrocortisone acetate. By contrast, no positive signal was observed for the negative controls in the study. The amplified product digested by restriction enzyme demonstrated 3 bands (82, 135, 189 lip) upon agarose gel electrophoresis, in good agreement with the predicted sizes. The detection limit of LAMP assay was 1 pg/μl of Pc DNA per reaction and that of PCR was 10 pg/μ1 of Pc DNA per reaction. Conclusion LAMP assay has usefulness for rapid detection of Pc.

Objective To detect Toxoplasma gondii DNA by loop-mediated isothermal amplification(LAMP). Methods DNA was extracted by phenol-chloroform extraction from T. gondii tachyzoites. Four primers which recognized 6 distinct regions on the B1 gene of T. gondii were designed and used for LAMP assay. To evaluate the specificity of the method, Plasmodium vivax, P. falciparum, Pneumocystis carinii, Schistosoma japonicum, and mouse leucocytes were used as controls. The parasite extract (T. gondii) was 10-fold serially diluted for evaluating the sensitivity of LAMP, and was amplified by LAMP. LAMP results were read with naked eye and analyzed by electrophoresis. Results After LAMP reaction, positive amplification was observed with T. gondii, but no positive signal was toted for the negative controls in the study. The sensitivity of LAMP assay reached up to 2-3 T. gondii tachyzoites/ml per reaction. Conclusion LAMP assay shows proper specificity and sensitivity for the detection of T. gondii.

A recombinant plasmid containing cathepsin B endopeptidase of Schistosoma japonicum(Sjcb2)was constructed,indentified by PCR,restrictive enzyme,digestion and DNA sequencing,and expressed into mammalian cells.Immunochemistry examination showed that the Sjcb2 gene can be expressed in the eukaryotic system,providing a basis for the development of schistosome DNA vaccine.

Objective To establish a method for detecting cercaria of Schistosoma japonicum.Methods The loop-mediated isothermal amplification(LAMP)was used.The cercaria DNA of Schistosoma japonicum was extracted by using GeneReleaser.Four primers which recognized 6 distinct regions on the calcium-binding protein gene of cercaria of Schistosoma japonicum were designed and used for LAMP assay and Clonorchis sinensis was used as the negative control for evaluating the specificity and 20,10,5,1 cercariae of Schistosoma japonicum were amplified by LAMP for evaluating the sensitivity.The LAMP results were judged with the naked eyes and electrophoretic analysis.Results After LAMP reaction,the positive signal was observed with cercariae of Schistosoma japonicum.By contrast,no positive signal was observed for Clonorchis sinensis.The detection limit of LAMP assay was 1 cercaria of Schistosoma japonicum per reaction.Conclusion LAMP assay has usefulness for rapid detection of cercariae of Schistosoma japonicum.

Objective To investigate the risk factors and preventive strategies of patients with diffuse axonal injury(DAI) with deep veins thrombosis in lower limbs (LDVT).Methods One hundred and thirty cases of diffuse axonal injury patients with lower limb vascular were divided into LDVT group(22 cases) and non LDVT group(108 cases) based on ultrasound.The information including long-term bed,plasma fibrinogen level,varicose veins,hypertention,sex,age,smoking,alcohol drinking,diabetes,obesity,Glasgow Coma Scale (GCS) were collected.Results There were significant different between LDVT and non-LDVT group in terms of longterm bed time,hypertension,smoking,diabetes,high plasma fibrinogen,age,low GCS score correlated with LDVT (x2 =7.08,5.99,5.17,4.70,3.55,12.72,t =27.80,P ＜ 0.05).Gender,drinking,obesity,varicose vein factors had no correlation with LDVT(P ＞ 0.05).Conclusion Diffuse axonal injury in patients with LDVT is more common in patients with older age,hypertension,low GCS score,the higher the plasma fibrinogen.

Objective To express P30 surface antigen of RH strain of Toxoplasma gondii in E.coli BL21(DE3). Methods The P30 gene from Toxoplasma gondii was cloned to the pET28b vector after PCR, and the recombinant expression plasmid pET28b-P30 was constructed. Then the recombinants were transformed into E.coli BL21(DE3) after identified by the restriction enzyme digestion, PCR and DNA sequence determination annlysis. A single colony of E.coli BL21(DE3) containing the recombinant plasmid, pET28b-P30 was inoculated in LB culture, then diluted 1∶100 into 2 ml LB culture and induced by 0.2 mmol/L IPTG, and the expression product was identified by SDS-PAGE and Western blot. Results The recombinant plasmid of pET28b-P30 was constructed. ② Plasmid pET28b-P30 could express a specific 30 kDa fusion protein in E.coli BL21(DE3). Conclusions The expression plasmid which contains the gene fragment encoding P30 surface antigen of Toxoplasma gondii has been successfully constructed and is highly expressed in E.coli BL21(DE3) as an inclusion body.

Objective To clone and sequence the partial gene of Schistosoma japonicum phosphoglycerate kinase (SjPGK). Methods A pair of primers were designed and synthesized according to the cDNA sequence of Schistosoma mansoni phosphoglycerate kinase (SmPGK) gene. The gene fragment of SjPGK was amplified and isolated from the total RNA of S.japonicum by reverse-transcription polymerase chain reaction (RT-PCR). The gene fragment was cloned into the cloning vector of pMD18-T, The positive clones were acquired and identified with restrictive enzymes and PCR amplification. After being sequenced with DNA auto-sequence analysis instrument,the cDNA sequence of SjPGK was searched for homologue identity with NCBI BLAST program. Results The gene encoding SjPGK was obtained and isolated by RT-PCR .The fragment of SjPGK was about 830 bp.The cDNA sequence of the phosphoglycerate kinase was highly homologous between Schistosoma mansoni and Schistosoma japonicum. The identity of nucleotide sequence was 85% and score 672, and the identity of amino sequence was 94% and score 473. Conclusion The partial gene of encoding SjPGK is cloned into the cloning vector of pMD18-T, which gives the basis for discovering new candidate vaccine molecular for schistosomiasis.

Adielectrophoresis-basedmicrofluidicchipappliedtocellspatterningisdesignedandfabricated, and it demonstrates non-contact and batch manipulation of cells. The microfluidic chip employs a PDMS microchannel and two ITO electrodes, which are designed as astep shape. The distribution of electric field caused by the microelectrodes is simulated by finite element simulation software, COMSOL. The position of the maximum intensity of electric field is also determined. The ITO microelectrodes and the PDMS microchannel are fabricated using MEMS fabrication process. After oxygen plasma surface treatment, the PDMS microchannel and glass substrate with the ITO microelectrodes are aligned and bonded to form experimental microfluidic chip. Through DEP experiment with the varying frequencies, DEP response of yeast cells is examined, and the electric field frequency of the both positive and negative DEP responses are confirmed. The results showed that yeast cells in solution conductivity of 60 μS/cm had negative DEP movement at the frequency of 1 kHz to 10 kHz, positive DEP movement at the 500 kHz to 10 MHz, and no DEP movement at the 50 kHz. Under the condition of the sinusoidal potential of 8Vp-p and the electric field frequency of 5 MHz, the yeast cells were aligned into chains along the step edge of microelectrodes.

AIM:To explore the effects of PPARγon the elevated level of reactive oxygen species ( ROS) in-duced by high glucose and its mechanism .METHODS:Human umbilical vein endothelial cells ( HUVECs) were cultured with DMEM containing high glucose (33 mmol/L D-glucose), and DMEM containing lower glucose (5.5 mmol/L D-glu-cose) was used as control .Superoxide anion and nitric oxide fluorescence probes were used to observe the effects of PPAR γagonist on ROS and NO productions in the HUVECs .The uncoupling protein 2 (UCP2) protein level in the HUVECs was detected by Western blotting .RESULTS:PPARγagonist pioglitazone inhibited the ROS generation and prevented the de-crease in NO level under high glucose condition , and these effects were reversed by pretreatment with PPARγantagonist GW9662.The results of Western blotting indicated that PPARγagonist pioglitazone up-regulated the UCP2 expression un-der high glucose condition , and this effect was also blocked by GW 9662.Inhibition of UCP2 by genipin attenuated the effect of pioglotazone on the ROS production .CONCLUSION: Activation of PPARγinhibits ROS generation under high glucose condition , and this effect may mediate by up-regulation of UCP2.