Objective To investigate the effect ofvalproate acid(VAP) on the activity ofneutrophils in children with epilepsy and its clinical significance.Methods A total of 34 cases of children with epilepsy(epilepsy group) were enrolled in this study.Another 30 healthy children were as control group.The activation rate of neutrophils was detected by flow cytometry.The levels of serum high sensitivity-C reactive protein(hs-CRP) were measured by latex enhanced immunoturbidimetric analysis.The levels of serum tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) were measured by enzyme linked immunosorbent assay analysis.Results The activation rate of neutrophils had no significant difference between epilepsy group before treatment and control group (P ＞ 0.05).The activation rate of neutrophils after treatment of 6,12 months in epilepsy group was higher than that in control group [(12.36 ± 4.72)％,(15.87 ± 5.68)％ vs.(5.32 ± 1.41)％,P ＜ 0.01].The activation rate of neutrophils after treatment of 12 months was higher than that after treatment of 6 months in epilepsy group (P ＜ 0.05).There was no significant difference in the levels of serum hs-CRP,TNF-α and IL-6 between epilepsy group before treatment and control group (P ＞ 0.05).The levels of serum hs-CRP,TNF-α and IL-6 after treatment of 6,12 months in epilepsy group were higher than those in control group [(3.64 ± 1.22),(6.96 ± 2.64) mg/L vs.(1.46 ± 0.27) mg/L,(74.72 ± 22.58),(96.67 ± 30.25) ng/L vs.(31.72 ± 12.16) ng/L,(32.59 ± 8.45),(46.74 ± 12.16) ng/L vs.(15.36 ± 4.45)ng/L,P ＜ 0.01],and those after treatment of 12 months were higher than those after treatment of 6 months (P ＜ 0.05).The activation rate of neutrophils was positively related with the serum levels of hs-CRP,TNF-αand IL-6 (r =0.328,0.402,0.344,P ＜ 0.05).Conclusions The high activation rate of neutrophils is found in children with epilepsy.The abnormal activation of neutrophils makes the body undergo high oxidative stress status.

Objective To detect the expression of serum creatinine (SCr),urine output,and urinary chalone C (CysC) in children with acute kidney injury (AKI) and to explore its clinical significance.Methods A total of 34 children infected by H1N1 or EV71 virus were enrolled in this study.Thirty-four cases of children were divided into AKI group(12 cases) and non-AKI group(22 cases).The level of SCr,urine output and CysC in two groups was compared on admission and 1,3 d after admission.Results The incidence rate of AKI in virus infected children was 35.3％ (12/34).There was no significant difference in SCr and urine output between two groups on admission (P ＞ 0.05).But the CysC in AKI group was higher than that in non-AKI group [(1.64 ± 0.23) mg/L vs.(0.85 ± 0.16) mg/L] (P ＜ 0.05).There was no significant difference of SCr,urine output and CysC in non-AKI group (P ＞ 0.05),but the level of SCr and CysC in AKI group after admission was significantly higher than that on admission [(38.25 ±7.18),(40.54 ± 7.62) μ mol/L vs.(25.26 ± 5.42) μ mol/L; (2.04 ± 0.33),(2.56 ± 0.41) mg/L vs.(1.64 ± 0.23) mg/L] (P ＜ 0.05 or ＜ 0.01),and urine output was significantly lower[(0.37 ± 0.12),(0.41 ± 0.14) ml/(kg·h) vs.(0.54 ±0.20) ml/ (kg·h)](P＜ 0.05).There was no significant difference in AKI group in SCr and urine between 1 day and 3 day after admission,while the CysC was statistically significant (P ＜ 0.01).Correlation analysis showed that the expression of serum CysC and SCr had positively significant correlation (r =0.412,P ＜0.05),and the urine volume had significantly negative correlation (r =-0.364,P ＜ 0.05).Conclusions High incidence of AKI occurs in children of virus infection.CysC value in early diagnosis of AKI is significantly superior to SCr and urine volume,and is expected to be an index in early prevention and treatment of children with AKI.

Objective: To analyze the published domestic documents of the clinical trial for treatment of anal fissure in past 10 years, to evaluate the quality of literatures about anal fissure randomized controlled clinical trials, and to provide references of clinical studies for the treatments of anal fissure in Traditional Chinese Medicine (TCM). Methods: In term of principles of evidence-based medicine, we evaluate the published domestic literatures of the randomized controlled clinical trial for treatments of anal f issure in past 10 years. Results: We analyze 52 literatures which are in the method of randomized controlled clinical studies. 28 literatures (accounting for 53.85%) record diagnostic criteria, 38 literatures (accounting for 73.08%) make sure standards of evaluating the effect and 18 literatures (accounting for 34.62%) mention comparability between groups without describing the baseline statistics. Only 5 are marked as a funded research projects. Conclusion: in current published literatures of randomized controlled study for treatments of anal fi ssure, the problems are focused on: ① Experimental idea is unclear and design is unreasonable, ② The description of implemented process in randomized control is unspecif ic, ③ Signifi cant parameters in statistical data are missed and with less credibility, ④ Criteria of diagnosis and effi cacy are different, ⑤ Evaluation indicators are fuzzy. All above clue to the quality of clinical research needs to be improved rapidly, and standardized evaluation of eff icacy also needs to be created urgently.

Objective To construct of shRNA lentiviral expression vector targeting APRIL (aproliferation-inducing ligand) gene in CFPAC-1 cell of human pancreatic cancer. Methods We used gene engineering to screen RNA interference targeting sequence of APRIL gene. The complementary DNA containing both sense and antisense Oligo DNA of the targeting sequence was designed, synthesized and cloned into the pGCL-GFP vector. The resulting lentiviral vector containing shAPRIL were named LV-shAPRIL. Then it was conformed by PCR and DNA sequencing identification. 293T cells were eotransfected with LV-shAPRIL,pHelper 1.0 and pHelper 2.0 to product ientivirus. The titer of virus was tested according to the expression level of GFP in the 293T cells. After recombinant lentivirus infection into CFPAC-1 cells, we used real-time RT-PCR and Western blotting to examine APRIL mRNA and protein expression at different cell culture period.Results PCR analysis and DNA sequencing conformed that shAPRIL DNA was successfully inserted into the lentiviral vector. The titer of concentrated virus were 5 × 107 TU/ml. APRIL expression in CFPAC-1 cells were inhibited significantly at both mRNA and protein level. APRIL mRNA expression were decreased 73%, 70%and 71% , respectively, after the infection of 4 days, 4 weeks and 8 weeks by LV-shAPRIL. APRIL protein expression were decreased 66%, 63% and 62%, respectively , after the infection of 4 days , 4 weeks and 8weeks by LV-shAPRIL. Conclusions ShRNA lentiviral expression vector targeting APRIL gene has been successully constructed, and it can effectively inhibit the expression of APRIL gene in CFPAC-1 ceils. This study lays a foundatin for in vivo research APRIL gene scilence in pancreatic cancer cell using the model of nude mice.

Objectives To establish real-time fluorenscence quantitative polymerase chain reaction ( RFQ-PCR ) for measurement of the expression level of guanylyl cyclase-C(GC-C) in the Peripheral blood mononuclear cell(PBMC)in 30 blood donors, 10 cases colorectal cancer tissue and 1 case T84 human colon cancer cell line. Methods Specific primers and TaqMan probe have been designed,and fluorenscence of the PCR product was detected continuously during amplification. According to the standard curve created by plasmid DNA, the expression level of GC-C in clinical samples has been determined using software, and the results were presented as the ratios of GC-C mRNA to?2-microgluobulin(?2M)mRNA. Results The detection range of the assay was from 101 pg/ml to109pg/ml,the coefficient of variation values of both intra-experimental and inter- experimental reproducibility were 6. 87% to 11. 12% and 8. 86% to 15. 19% . None of 30 blood donors and 11 benign intestinal patients expressed GC-C mRNA,it was expressed in 31/37 colorectal cancer patients. The expression level of GC-C mRNA in colorectal cancer patients was 0. 88?0.06,and the expression level of its in colorectal carcinoma tissue and T84 cells were 0. 86?0.07/ug tissue and 0.0082/per cell. Conclusions This assay had high sensitivity,specificity and reproducibility.

Objective To investigate the inhibitive effect of shRNA (short hairpin RNA) targeting APRIL gene on the pancreatic cancer cells in vitro and in vivo, in order to explore the feasibility of gene therapy for pancreatic cancer. Methods The LV-shAPRIL targeting APRIL gene had been constructed before, and was used to infect the CFPAC-1 cells. Cell proliferation and apoptosis were examined by MTT and flow cytometry. Then CFPAC-1 cells were used to construct the model of transplantation tumor into the nude mice, the tumor growth was assessed after LV-shAPRIL treatment. Results 96 hours after the LV-shAPRIL infection into CFPAC-1 cells, the cell proliferation was significantly inhibited when compared with control group and lentivirus infection group (P＜0.05 ). Flow cytometry showed the apoptosis ratio of the CFPAC-1 cells was (17.35±0.96)% in LV-shAPRIL group, which was higher than that in control group and lentivirus infection group (P＜0.05 ). After LV-shAPRIL injection into the model of nude mice, the tumor growth was slower than that in the two control groups. The tumor's volume of the LV-shAPRIL group was(821.8±123.3) mm3 and the mass was (2.16±0.18)g at 27 day, and were obviously depressed, when compared with two control groups (P±0.05). Conclusions LV-shAPRIL targeting APRIL gene can inhibit the growth of the CFPAC-1 cells in vitro and vivo. This may provide a new gene therapy approach for pancreatic cancer.

Objective To construct small hairpin RNA(shRNA) lentiviral vectors targeting human a proliferation-inducing ligand(APRIL) gene and detect the titer of virus and suitable multiplicity of infection (MOI) after 293T cells were infected by the lentival vectors. Methods Three RNA interference targeting sequences of APRIL gene were screened including shAPRIL1210, shAPRIL1754 and shAPRIL1604. Both sense and antisense Oligo DNA of the targeting sequences were synthesized and cloned into the pGCL-GFP vector, respectively. The resulting lentiviral vectors containing shAPRIL were named LV-shAPRIL1210, LV-shAPRIL1754, LV-shAPRILI604. Then they were confirmed by PCR and DNA sequencing. 293T cells were co-transfected with LV-shAPRIL, pHelper 1.0 and pHelper 2. 0 to product lentivirus, respectively. The titer of virus and suitable MOI were tested according to the expression level of GFP in the 293T cells. Results PCR analysis and DNA sequencing confirmed that three shAPRIL DNA were successfully inserted into the lentiviral vectors. The titers of concentrated virus were 5 × 107, 6 × 107 and 4 × 107(transduction units )TU/ml, respectively, and the suitable MOI was 5. Conclusions Three shRNA lentiviral vectors targeting human APRIL gene have been successfully constructed, which lays a foundation for future studying APRIL's gene silencing in related target cells.

0.05 ), those in the malignant group were 53%(18/34) and 94%(32/34) with a distinguished improvement after enhancement(P

Objective To evaluate the frequency of mutations that occur in PHEX,FGF - 23 and DMP - I genes associated with familial hypophosphatemic vitamin D resistant rickets among 6 patients from 4 families in China. Methods The peripheral blood samples from 4 families were collected and other 10 persons from different families were selected as normal controls,and then the total gene DNA was extracted from the whole blood. Using polymerase chain reaction(PCR)amplication,sequences of the exons and flanking zones in PHEX,FGF - 23 and DMP - I genes were sequenced by direct DNA sequencing and TA cloning,and then the mutations found were analyzed. Results In exon 6 of DMP - I gene,c1218 C ﹥ T and c1230 G ﹥ A mutations were detected in lineage 1,as same sense mutation (propositus and its sister:homozygous mutation;mother:heterozygous mutation);c1333 - 1334 GC ﹥ TT mutation,as missense mutation,was found in exon 12 of PHEX gene on the propositus of lineage 2,determined as heterozygous muta-tion,but the same mutation was not found from their parents. In exon 3 of FGF - 23 gene,c716 C ﹥ T,p. T239M hetero-zygous mutation was found on the propositus and its mother. In exon 6 of the DMP - I gene,c205 A ﹥ T homozygous mutation was detected in lineage 3. In lineage 3,c716 C ﹥ T mutation of the FGF - 23 gene was detected,and the pro-positus and their father had the same mutation. No disease causing mutations of the PHEX,FGF - 23 and DMP - I genes were detected in the family members of lineage 1,3 and 4. Conclusions The mutation c1333 - 1334 GC ﹥ TT detected in exon 12 of PHEX gene might be the cause of disease for the propositus of lineage 2,as missense mutation, which needs further verification;c716 C ﹥ T,p. T239M mutation of the FGF - 23 gene detected in lineage 2 and 3 might not be the causes of the hypophosphatemic rickets and abnormal phenotype.