Objective Conduct safety assessments of the established human skin fibroblast line. Methods A series of safety assessments were performed including cell morphology,chromosome karyotyping,soft-agar test,nude mice carcinogenic test,endotoxin test,mycoplasma determination,detection of viral agent,bacteria,fungi sterile tests and abnormal toxicity test.Results No abnormal changes were observed.Conclusion The fibroblast obtained from skin is a kind of safety and reliable target cell for gene therapy.

Objective To construct the recombinant human metapneumovirus(hMPV) (defined as rhMPV NL/1/00 GFP) in vitro by reverse genetics technique. Methods BSR-T7 cells were transfected using LipofectAMINE 2000 with the full-length cDNA plasmid, and four major protein expressing plasmids, pCITE-N, pCITE-P, pCITE-M2.1 and pCITE-L. After 3 d, cells were subjected to one -80℃ freeze-thaw cycle to prepare lysates. The supernatant of lysate was used to inoculate Vero-E6 cells. After 1-4 d, cells were found for the obvious development of cytopathic effects under light microscope and green fluoroscopic signals under fluorescence microscope, and were observed up to 10 d. The supernatant were collected to de-tect virus titer. Viral RNA was extracted from the supernatant and reverse transcriptase polymerase chain re-action (RT-PCR) was used to amplify N, F and G genes of rescued virus. Results Cytopathic effects and green fluoroscopic signals was readily and obviously observed after 1-4 d post-inoculation in Vero-E6 cells, then cytopathic effects got worse and green fluoroscopic signals became stronger gradually up to 10 d. The ti-ters of the 1st, 5th, 10th,15th and 20th generation virus ranged from 105.0 to 106.5 TCID50/ml. Amplicons with size of 910 bp (N), 450 bp (F) and 980 bp (G) by RT-PCR were accordant with expectant. Nucleotide sequence analysis of above cDNA fragments showed 100% similarity with reported sequence of hMPV NI/1/00 strain. The recombinant virus was genetically constant and GFP-labeled after 20 passages in Vero-E6 cells. Conclusion Recombined hMPV was successfully rescued by reverse genetics technique. This study lays ground for exploring pathogenesis of hMPV infection and development of hMPV attenuated vac-cines.

To study the status and patterns of the drug utilization in the diseases of lower respiratory tract, the drug utilization for 290 inpatients with respiratory tract diseases in a Shanghai hospital druing 1993-1997 was analyzed by using Acute Physiology, Age, Chronic Health Evaluation(APACHE) and index of defined daily dose numbers(DDDs). The relativity between APACHE and DDDs was studied. It was found that most common drugs was anti-infection agents and expectorants, accounting for 39.06% and 38.38 %, respectively. The quantitative relationship between drug consumption and disease severity was not observed. It is concluded that the status of the drug utilization can't be demonstrated by analyzing the frequency of drug use by using DDDs.

Objective: To study the expression and significance of VEGF-C, D2-40, and E-Cad in Human breast carcinoma. Methods: S-P immunohistochemical method was used to assess the expression of VEGF-C in 88 cases of infiltrating ductal carcinoma of the breast (with intact clinicopathologic and follow-up in-formation) and 54 cases of different intraductal hyperplastic lesions. The relationship of VEGF-C expression with D2-40 and E-Cadherin in infiltrating ductal carcinoma of the breast was analyzed. Results: VEGF-C ex-pression was higher in the invasive group than that in the dysplasia group and benign lesions. VEGF-C ex-pression in the metastatic group was significantly higher than that in the group without lymph node metasta-sis, indicating that VEGF-C plays an important role in the growth and metastasis of breast cancer. VEGF ex-pression in breast cancer group was higher than that in the dysplasia group and benign lesions. LVD count in the group with lymph node metastasis was significantly higher than that in the group without lymph node me-tastasis. The positive rate of E-Cad expression in the invasive group was significantly lower than that in the dysplasia group and benign lesions. VEGF-C expression was positively correlated with D2-40 expression. VEGF-C expression was negatively correlated with E-Cad expression. Conclusion: VEGF-C and D2-40 could be used for the detection of early lymphatic metastasis of breast cancer. VEGF-C and E-Cad can be used as important indices for the evaluation of the prognosis of breast cancer.

Background: Integrative traditional Chinese and western medicine may be a new approach to improve the eradication rate of Helicobacter pylori (Hp).Aims: To compare the efficacy and safety of Weifuchun tablet versus bismuth combined with standard triple regimen as the first-line therapy of Hp infection.Methods: A total of 141 patients with Hp infection and na(i)ve to treatment were randomly assigned into 3 groups receiving a 14-day eradication therapy.In standard triple therapy group, esomeprazole, amoxicillin and clarithromycin were given twice a day;while in Weifuchun group and bismuth group, Weifuchun tablet and bismuth potassium citrate were added, respectively, to the standard triple therapy.Hp eradication was assessed by 13C-urea breath test at least 6 weeks after the end of treatment.Hp isolates were tested for resistance to antibiotics.Results: One hundred and twenty-eight patients completed the study.Hp eradication rates in Weifuchun group, bismuth group and standard triple therapy group were 83.7%, 91.8% and 79.1%, respectively by ITT analysis and 88.4%, 97.8% and 84.6%, respectively by PP analysis.The eradication rate of Weifuchun group was lower than that of bismuth group and higher than that of standard triple therapy group, but the differences were not statistically significant (P>0.05).Only PP eradication rate of bismuth group was significantly higher than that of standard triple therapy group (P<0.05).The resistant rates of Hp to clarithromycin, amoxicillin and metronidazole were 33.3%, 2.9% and 70.5%, respectively.For eradication of clarithromycin resistant strains, bismuth group was superior to Weifuchun group and standard triple therapy group (100% vs.60.0% and 66.7%, P all <0.05).All three eradication regimens showed good compliance, and no significant difference in incidence of adverse events was found between the three regimens (P>0.05).Conclusions: Weifuchun tablet combined with standard triple regimen is safe and effective for use as first-line treatment for Hp infection, however, the eradication rate is relatively low in cases infected with clarithromycin resistant strains.Bismuth combined with standard triple regimen is a good alternative in areas with high clarithromycin resistance and regions where tetracycline is unavailable.

Objective To investigate the effect of Shenlongtang decoction on learning and memory ability and expression of Bcl-2 and Bax on rats hippocampus with cerebral ischemia model. Methods The cerebral ischemia animal model was established by two-vessel occlusion. Fifty rats were randomly divided into control group, model group, pisacetam group, Shenlongtang groups (low and high dosage). After modeling, rats were administrated with corresponding drugs for 28 days, the learning and memory ability was tested with Morris water maze, the number of positive cells and the integral optical density of the immunostaining on Bcl-2 and Bax protein expression in the hippocampus was detected. Results With Morris water maze test, the latency time increased significantly and times of searching submerged platform decreased significantly in the model group, the difference was significant compared with control group (P<0.01). Compared with model group, the latency time of Shenlongtang high dosage group decreased significantly and times of searching submerged platform increased significantly (P<0.05). Compared with model group, the expression of positive cells number of Bax protein expression in the hippocampus area was significantly increased (P<0.05), the Bcl-2 protein expression in the hippocampus region was significantly decreased (P<0.01). Conclusion Shenlongtang could significantly improve the learning and memory ability of cerebral ischemia rats, the mechanism may be related to regulating the expression of Bcl-2 and Bax.

Objective To compare the differences between measured resting energy expenditure calculated by the indirect calorimetry with the resting energy expenditure calculated by the Harris-Benedict formula and weight formula in the mechanically ventilated surgical critically ill patients in SICU.Methods Patients mechanically ventilated in SICU of Xuanwu Hospital,from April 2014 to April 2015 were measured resting energy expenditure by the indirect calorimetry with the resting energy expenditure calculated by the Harris-Benedict formula and weight formula in the 1st,the 3rd and the 5th day.There were twenty-nine patients enrolled,thirteen males and sixteen females,measured the resting energy expenditure 188 times.The distribution of metabolism level was studied,and the resting energy expenditure measured by three methods were calculated and evaluated by paired sample t test.Results There were 177 times(62.24％)of low metabolism level,59 times(31.38％)of normal metabolism level,and 12 times(6.38％)of high metabolism level.Eighteen patients used these three methods to calculate the energy expenditure on 1st,3rd and 5th day:indirect calorimetry (1 627.11 ± 323.63) kcal,(1 614.67± 308.93) kcal,(1 576.11 ± 263.96) kcal;Weight formula (1 479.44 ± 200.24) kcal,(1 488.40 ± 227.72) kcal,(1434.14 ± 216.56) kcal;Harris-Benedict formula (1 777.43 ± 253.00) kcal,(1 730.08 ± 265.18) kcal,(1 689.33 ± 236.69) kcal.The results calculated from Harris-Benedict formula and the weight formula were significantly different fiom calculated from indirect calorimetry (P ＜ 0.05).Resting energy expenditure by Harris-Benedict formula was significantly higher than calculated from indirect calorimetry (All P ＜ 0.05).Resting energy expenditure by weight formula was significantly lower than calculated from indirect calorimetry(All P ＜ 0.05).Conclusions Although Harris-Benedict formula and weight formula is convenient in clinical use,while the results calculated by them is significant different from the results calculated by indirect calorimetry.So clinical nutrition support should rely on indirect calorimetry as far as possible.

Objective To investigate the effect of Shuyusan on the behavior, content of tumor necrosisfactor-α(TNF-α) in cortex of cerebral and neuropeptide expression in hippocampus of rats with chronic mild unpredictable stress (CMUS) depression. Methods Fifty mice were randomly divided into 5 groups:normal group, model group, fluoxetine group, and high-, low-dose group of Shuyusan. Except normal group, the other groups were given CMUS. Tail suspend test (TST) and sucrose preference test (SPT) were used to estimate animals’ behavior. The contents of TNF-αwas determined by enzyme linked immunosorbent assay (ELISA). The immunocytochemical technique was used to evaluate hippocampus neuropeptide immunoreactive expression. Results The fluoxetine group and high dose of Shuyusan group showed significant increase in the sucrose consumption and decrease in the immobility time in TST, there was significant difference compared with CMUS model group (P<0.05). The TNF-αlevel was significantly lower in fluoxetine group and high dose of Shuyusan group, there difference was significant compared with model group (P<0.05). The neuropeptide immunoreactive neurons in hippocampus area was decreased by a marked number in fluoxetine group and high dose of Shuyusan group, the average optical density decreased significantly, compared with the model group, there were significant differences (P<0.05). Conclusion Shuyusan has anti-depression likely effect, with dose-effect relationship.

ObjectiveTo investigate the effects of Jiuqiang Naoliqing(JNQ) on the morphology of the vital organs(the brain,kidney and heart) in acute hypertension rats.MethodsThe model was established by L-NG-nitro arginine(L-NNA),which can inhibit the synthesis of nitric oxide(NO).Affected animals were administrated with high,medium or low dose of JNQ while nifedipine was used as the positive control.Effects on cardiac function and morphology of the vital organs were investigated.Results2 weeks after the establishment of the model,the affected rats had a 36% higher blood pressure compared with the control group.The acute hypertension rats showed significant pathologic changed in the brain,kidney and heart,while there was no obvious difference in the heart rate,electrocardiaogram,and blood pressure between the administrated groups.After treatment with JNQ,the contraction force and the oxygen extraction of the myocardiocytes were significantly reduced and the increasing velocity of the left ventricular pressure was decreased.The brain,kidney and heart showed improvement in pathology analysis to different levels.ConclusionAlthough it has no obvious effects on the heart rate and blood pressure in acute hypertension rats,JNQ can decrease the oxygen extraction in myocardiocytes and can obviously alleviate brains,kidneys and hearts pathologic changes.

Objective To investigate the effect of Shenlongtang decoction (SLT) on the learning and memory ability and serum level of neuropeptide Y (NPY) and expression of NPY in hippocampus of rats with cerebral ischemia/reperfusion. Methods 50 rats were randomly divided into 5 groups: control group with sham operation; model group with cerebral ischemia/reperfusion; pisacetam group as positive control;and SLT groups (low and high dosages). The rats' model was established with two-vessel occlusion. After modeling, the rats were administrating with SLT or pisacetam for 28 d. Learning and memory ability was tested with Morris water maze, the level of NPY was detected with radioimmunoassay, and the number of positive cells of NPY in hippocampus was detected. Results Compared with the control group, the latency time increased significantly and frequence of searching submerged platform decreased significantly in the model group (P<0.01); and compared with the model group, the latency time decreased significantly and frequence of searching submerged platform increased significantly in SLT high dosage group (P<0.05). Compared with the control group, the serum levels of NPY increased significantly in the model group, and the number of positive cell of NPY in hippocampus area significantly increased (P<0.01); and compared with the model group, the serum levels of NPY decreased significantly, and the number of positive cell of NPY in hippocampus area significantly decreased in the SLT high dosage group and pisacetam group (P<0.05). Conclusion SLT could improve the learning and memory function.The mechanism may be related to regulate the serum level and expression of NPY in hippocampus area of rats with cerebral ischemia/reperfusion.