Objective To investigate the changes in cathepsin B (CatB) expression in acutely photodamaged human dermal fibroblasts (HDFs) and their significance.Methods HDFs were isolated from the foreskin of children,and subjected to primary culture and subculture.The fourth-to eighth-passage HDFs were used in the following experiment.HDFs were divided into two groups to receive irradiation with different doses of ultraviolet A (UVA) for different durations (acutely photodamaged group) or remain unirradiated (control group).Cell counting kit-8 (CCK8) assay was conducted to evaluate the proliferative activity of HDFs after irradiation with UVA at 5,10,15,20 and 25 J/cm2 respectively.Western blot and quantitative real-time reverse transcription PCR were performed to measure the protein and mRNA expressions of CatB respectively in HDFs at 24,48 and 72 hours after exposure to UVA at 10 J/cm2,and at 48 hours after exposure to UVA at 10,15,20 and 25 J/cm2.Statistical analysis was carried out by analysis of variance and least significant difference (LSD) test using the SPSS 13.0 software.Results UVA radiation induced a decrease in the proliferative activity of HDFs.When the dose of UVA was ≤ 10 J/cm2,the survival rate of HDFs maintained higher than 85％,and significant differences were observed in cell survival rate between unirradiated and irradiated HDFs at 24,48 and 72 hours (all P ＜ 0.05).Western blot showed that the gray value of CatB protein in the acutely photodamaged group irradiated with 10 J/cm2 UVA was significantly higher than that in the control group at 24 hours (0.76 ± 0.14 vs.0.35 ± 0.01,P ＜ 0.05),48 hours (1.34 ± 0.38 vs.0.45 ± 0.12,P＜ 0.05) and 72 hours (0.82 ± 0.09 vs.0.61 ± 0.06,P＜ 0.05).Increased mRNA expressions of CatB were also observed in the acutely photodamaged group compared with the control group at 24 hours (0.149 ± 0.009 vs.0.089 ± 0.015,P ＜ 0.05),48 hous (0.173 ± 0.009 vs.0.091 ± 0.010,P ＜ 0.05) and 72 hours (0.185 ± 0.158 vs.0.111 ± 0.017,P ＜ 0.05) after UVA radiation at 10 J/cm2.The gray value of CatB protein was 0.99 ± 0.07,1.49 ± 0.14,1.89 ± 0.08,2.07 ± 0.06 in HDFs at 48 hours after exposure to UVA of 10,15,20 and 25 J/cm2,respectively,significantly higher than that in the control group (0.60 ± 0.05,all P ＜ 0.05).Similarly,the mRNA expression of CatB was up-regulated in HDFs at 48 hours after UVA radiation at 10,15,20 and 25 J/cm2 compared with the unirradiated HDFs.Conclusion The protein and mRNA expressions of CatB are up-regulated in acutely photodamaged HDFs induced by UVA radiation.

Objective:To study the antiviral effect of ISG20 on HCV replicon.Methods:Wild type ISG20/mutated ISG20 cDNAs were obtained by RT-PCR/two step-PCR directed mutagenesis, and wild type ISG20 and dominant negative mutated ISG20 mammal expression vectors were consuructed. The constructed pISG20wt and pISG20m expressing vectors were transfected into Huh7 cells or Huh7 cells containing HCV replicon to investigate its effects on HCV replicon replication.Results:The ISG20wt/ISG20m expression vectors were constructed and the expressions of these two vectors were confirmed at both mRNA and protein levels. The effects of ISG20wt on HCV replicon replication were evaluated by Northern blot and Western blot. The results showed that expression of ISG20wt had significant inhibitory effect on HCV RNA replication.Conclusion:ISG20 participates in the anti-HCV action of IFN-? on HCV replicon system.

[Objective] To study the effect of electroacupuncture on NO, NOS and ET - 1 levels in rats with focal cerebral ischemia (FCI). [Methods] Rat models with FCI were established by occlusion of unilateral middle cerebral artery. Twenty - two rats were allocated to three groups: Group A (mimic operation group), Group B (model group) and Group C (model rats treated with electroacupuncture). NO, NOS and ET - 1 levels in brain homogenate were measured by spectrophotometry and radioimmunoassay method respectively. [Results] Sixty minutes after modeling, NO, NOS and ET- 1 levels were increased and then decreased 10 minutes after acupuncturing Baihui (GV20) and Dazhui (GV14). [Conclusion] Electroacupuncture of Baihui (GV20) and Dazhui (GV14) can protect neuron from ischemic injury by decreasing the levels of NO and ET - 1 and inhibiting the activity of NOS.

Objective To evaluate tension-free repair in the management of recurrent inguinal hernia. Methods From 1/1993 to 12/2002, 163 patients with recurrent inguinal hernia underwent reoperation, clinical data were reviewed. Results The male: female ratio was 138: 25, age from 34 to 76 years. The primary surgery had been traditional hemiorrhaphy without prothesis in 142 cases (87. 1 % , including 65 cases of Bassini ,35 cases of McWay and 42 cases of Shouldice) , a tension-free procedure in 12 cases(7. 4% , including 3 cases of laparoscopic mesh repair) and unknown techniques in 9 cases(5. 5% ). The average interval from the initial surgery to recurrence was 36?14 months (range from 3 months to 10 years). Recurrent hernias were treated with a Shouldice repair in 71 cases(43. 6% ) , a tension-free repair in 92 cases(56. 4% , including a laparoscopic mesh repair in 3 cases). All these 163 cases(100% ) were followed up for 56 ?1 months (range from 30 months to 12 years) after the second surgery. It was found that after reoperation the recurrence rate of Shouldice procedure and tension-free repair was 16. 1% (11/71) and 2. 2% (2/92) respectively (x2 = 8. 327 ,P

Objective A method was established for genotyping of hepatitis B virus (HBV A-D genotype),based on the PCR-restriction fragment length polymorphism (RFLP) created by Hinf Ⅰ,Ear Ⅰ,Apo Ⅰ action on an amplified segment of the S region.Methods Clinical diagnosis research.One hundred and twenty-eight HBV S sequences obtained from GeneBank were analyzed for restriction enzyme sites that would be genotype-specific.Restriction patterns following digestion with restriction enzymes Hif Ⅰ,Ear Ⅰ,Apo Ⅰ were determined to identify A-D HBV genotypes.The method was used to detect the HBV genotype of fifty severe hepatitis patients due to chronic hepatitis B in China.Then the detection results were confirmed by direct sequencing.Results The new genotyping method was established,named simple PCR-RFLP,which could identify HBV genotypes A to D.Genotypes B,C,B/C and A or D could be determined by a single step digestion with Hif Ⅰ.Eight patients of genotype A/B/C classified by single step digestion with Hif Ⅰ were conformed as genotype B variant by further digestion and direct sequencing.Extracted randomly and diluted into different concentration,three specimens were tested for genotype of HBV repeatedly and respectively.The results were all in accord with the originals,and the lowest detection limit of HBV DNA was 7 ～ 9 IU/ml.This was particularly useful in China where genotypes B and C were predominant.Twenty-three of genotype B and ten genotype C patients were classified from these fifty severe hepatitis B patients by a single step digestion with Hif Ⅰ through the simple PCR-RFLP method.The same results were also obtained by direct sequencing of PCR products (Kappa =1.00,P =0.001).The simple PCR-RFLP method was superior to direct sequencing in detecting HBV B/C polyinfection (9 cases and 0 case; x2 =18.00,P =0.001).Conclusions Both the sensitivity and repetitiveness of Simple PCR-RFLP method are satisfactory.It is superior to direct sequencing in detecting HBV B/C polyinfection,and simple,convenient.

Objective To evaluate the performance of a DNA microarray method for detecting HBV antiviral drug-resistant mutations. Methods Two hundred and twenty four serum samples from patients with CHB were tested in parallel by DNA microarray and direct sequencing for the mutations within the HBV reverse transcriptase (rt) region, which included rtL180, rtA181, rtM204 and rtN236. Samples with discrepant results were retested by clonal sequencing. Results Complete concordance between DNA microarray and direct sequencing results was observed in 214 out of 224 samples (95. 5% ). The presence of mixed viral populations in the other 10 samples detected by DNA microarray but not by direct sequencing was confirmed by clonal analysis. The DNA microarray could detect minor viral populations which constituted 5.0%-15. 0% of the total viral load. Conclusion DNA microarray is highly consistent with direct sequencing in detecting HBV mutations conferring drug resistance and more sensitive in detecting mixed mutant and wild-type sequences than direct sequencing, which makes it a useful tool for early detection of drug resistance early.

This paper elaborated that wheelchairs are the most important assistive devices for people with mobility impairment to maintain their health, improve their quality of life and promote their social participation. The situation of low service level was analyzed due to the lack of knowledge, organizations, professionals and products. Based on the foreign standard of practice for wheelchair service, the strategies and steps of wheelchair service provision in China were carried out to arouse the attention of relevant professionals, who could be active participants in wheelchair service practice and research

Objective To investigate the effects of advanced glycation end products(AGE)on the expressions and activity of cathepsin D(CatD)in ultraviolet A(UVA)?irradiated human dermal fibroblasts. Methods Human dermal fibroblasts were isolated and harvested from the circumcised foreskin of children, and subjected to a primary culture. CCK?8 assay was performed to screen non?cytotoxic concentrations of AGE?bovine serum albumin (BSA). Some fibroblasts were incubated with 50, 100 and 300 mg/L AGE?BSA separately for 24 hours, with untreated cells as the control group. Then, reverse transcription(RT)?PCR, Western?blot analysis and a fluorimetric assay were performed to measure the mRNA and protein expressions as well as activity of CatD, respectively. Some fibroblasts were classified into six groups: control group receiving no treatment, AGE?BSA group and BSA group treated with the highest non?cytotoxic concentration of AGE?BSA and the same concentration of BSA respectively for 24 hours, UVA group irradiated by 10 J/cm2 UVA, UVA?AGE?BSA group and UVA?BSA group treated with AGE?BSA and BSA at the above non?cytotoxic concentration respectively for 24 hours both before and after UVA radiation at 10 J/cm2. After the treatments, RT?PCR, Western?blot analysis and a fluorimetric assay were conducted to detect mRNA and protein expressions and activity of CatD respectively. Results AGE?BSA of 50- 200 mg/L exhibited no obvious influence on cellular proliferation of fibroblasts. The fibroblasts incubated with AGE?BSA of 50, 100 and 200 mg/L showed a significant increase in the mRNA expression(0.267 ± 0.007, 0.348 ± 0.007, and 0.418 ± 0.006 respectively), protein expression (1.403 ± 0.181, 2.233 ± 0.090 and 2.477 ± 0.111 respectively), and activity(1.760 ± 0.080, 2.330 ± 0.060 and 2.890 ± 0.080 respectively)of CatD compared with the control group(mRNA:0.161 ± 0.006;protein:0.903 ± 0.200;activity:1.100 ± 0.090, all P < 0.05). AGE?BSA increased CatD expressions and activity in a dose?dependent manner. The mRNA and protein expressions as well as activity of CatD were significantly higher in the UVA group than in the control group (mRNA expression: 0.480 ± 0.005 vs. 0.155 ± 0.005; protein expression: 2.583 ± 0.199 vs. 0.920 ± 0.235;activity:2.970 ± 0.110 vs. 1.110 ± 0.040, all P<0.05), but significantly lower in the UVA?AGE?BSA group than in the UVA group(mRNA expression:0.394 ± 0.008 vs. 0.480 ± 0.005;protein expression:2.070 ± 0.125 vs. 2.583 ± 0.199;activity: 2.560 ± 0.060 vs. 2.970 ± 0.110, all P < 0.05). Conclusion AGEs could increase CatD expressions and activity in human dermal fibroblasts not receiving UVA irradiation, but inhibit their increase in UVA?induced human dermal fibroblasts.

[Objective] To observe the protective effect of acupuncture on ischemic cerebral damage. [Methods] SD rats were randomized into 2-week sham-operation (2W-SO) group (n=6) , 5-week sham-operation (5W-SO) group (n =6) , 2-week cerebral-ischemia (2W-CI) group (n-6) , 5-week cerebral-ischemia (5W-CI) group ( n = 6), 2W-CI + electroacupuncture (EA) group (n=6) and 5W-CI + EA group (n=6) . Except the sham-operation groups, cerebral ischemia was induced by heat-occlusion of middle cerebral artery in other groups. Changes of nerve growth factor (NGF) expression were monitored in ischemic cortex of rats with 2-week and 5-week ischemia by immunohistochemical method, and the effect of electroacupuncture (EA) was observed. [Results] Few NGF-positive cells were found in the ischemic cortex of sham-operation groups and the expression of NGF were obvious in cerebral ischemia groups as compared with the sham-operation groups ( P