To introduce the implication,research overview and appraisal method of patient reported outcome being used in the clinical research of stroke.The basic process and principle of stroke-associated scale research was described in this article,the author think that PRO can be the important supplement of clinical therapeutic e ect evaluation of stroke.The study method of PRO is worth to extension and application in clinical research of other chronic disease.

BACKGROUND:Human mesenchymal stem cells (hMSCs) become aging and even die after several passages. Some investigations have shown that telomere has a close correlation with life span of the cells. Whether the ectopic expression of human telomerase reverse transcriptase (hTERT) could induce the activity of the telomerase, maintain the length of telomere, and finally prolong the life cycle of MSCs without losing their multipotent differentiation capacity is still uncertain.OBJECTIVE: To observe the influence of the ectopic expression of hTERT on the telomerase activity and cell life cycles of hMSCs.DESIGN: Repetitive measurement trails.SETTING: Research Center of Stem Cell, Zhengzhou University Medical College.MATERIALS: The experiment was conducted in the Research Center of Stem Cell, Zhengzhou University Medical College from October 2003 to December 2005. hMSCs were obtained from 20 healthy donators from the Department of Pediatric Surgery and Outpatient, the Third and First Affiliated Hospitals of Zhengzhou University. Enhanced green fluorescent protein plasmid (pEGFP-C1) and pEGFP-hTERT were provided by Dr. Chantal Autexier of Canada. DH5α strain provided by Dr. Hou Wei-hong, the Key Molecular Medical Laboratory of Zhengzhou University Medical College.METHODS.: Under sterile condition, 2 mL bone marrow of sternum of healthy donors were harvested, and prepared after centrifugalization,dilution and passage.① Transfection of pEGFP-hTERT into hMSCs and the screening and amplification of resistance cloning:The 5th passage cells were seeded in a 24-well plate,and transfected by pEGFP-hTERT with lipofectamine method.The cells were divided into four groups including untransfected group,lipofectamine group,pEGFP-C1 group and pEGFP-hTERT group. Resistance cloning screen and amplification was performed by G418. ②hTERT mRNA expression and detection of telomerase activity:RT-PCR and PCR-ELISA were used to detect the hTERT mRNA expressions of the fifth passage hMSCs transfected with pEGFP-hTERT, and pEGFP-C1, the untransfected tenth passage hMSCs and K562 cells (positive control), and the telomerase activity of the fifth and thirtieth passage hMSCs transfected with pEGFP-hTERT,the fifth pEGFP-C1-transfected cells and the tenth passage untransfected cells. ③Karyotype analysis of hTERT-transfected MSCs: Chromosome analysis was performed by conventional Giemsa staining.④Inducement of differentiation from telomerase-positive MSCs into neuron-like cells and RE-PCR identification:The transfected MSCs were cultured in a medium containing epidermal growth factor and basic fibroblast growth factor, which could induce the cells differentiate into neuron-like cells. The culture solution was changed every 3 days, and the changes in cell growth condition and morphologic characteristics were observed under an inverted microscope. The microtubule associate protein (MAP2) and neurofliament subunit M (NF-M) were identified by RT-PCR.MAIN OUTCOME MEASURES:①hMSCs transfection with different kathion liposomes and the screening and amplification of resistance cloning; ②hTERT mRNA expressions of each group and detection of telomerase activity; ③Karyotype analysis of pEGFP-hTERT-transfected MSCs; ④ Induction of differentiation from telomerase-positive MSCs into neuron-like cells and RE-PCR identification.RESULTS: ①With the decrease of G418 concentration, the cells in the untransfected and lipofectamine groups died, and stably EGFP expressed MSCs were obtained; after G418 screening, there was a cell clone undergone 35 passages and continued to proliferate, whose appearance and growth characteristics were similar to the untransfected MSCs observed under inverted microscope. ②The fifth passage pEGFP-C1-transfected hMSCs and tenth passage untransfected hMSCs remained telomerase-negative, but the K562 and fifth passage hTERT-transfected cells showed positive telomerase activity. ③The telomerase activity of the fifth and thirtieth passage hTERT-transfected cells was positive. ④The hTERT-MSCs at passage 10, 20 and 30 had 23 pairs of chromosomes, and two X chromosomes. So they were still normal diploid with normal chromosome appearance and number. ⑤Many hTERT-transfected MSCs had the typical appearance of neuron-like cells. RT-PCR analysis showed that th expressions of MAP2 and NF-M were increased.CONCLUSION:Ectopic expression of the hTERT gene is found in hMSCs,and can induce the telomerase activity of hMSCs.The ectopic expression of the hTERT gene in hMSCs could extend the life spans of cells and maintain their multipotent differentiation capacity.

Aims To study single dose toxicity of poly-phenols effective parts from Punica granatum,to eval-uate their safety,and thus to provide a theoretical basis for drug development and clinical use.To observe their protective effect on ethanol-induced gastric ulcer in rats.Methods 50 healthy Kunming mice were ran-domly divided into five groups and given different doses of polyphenols’effective parts from Punica granatum via intragastric administration.Toxicity and death in each group of mice were observed and recorded after administration for 14 d.The median lethal dose was calculated by Bliss method.70 rats were randomly di-vided into normal group,model group(constant volume of normal saline),sanjiuweitai particles(1 850 mg· kg-1 )group,colloidal bismuth subcitrate (33 mg · kg-1 )group and polyphenols effective parts from Puni-ca granatum low-dose,medium-dose,high-dose(430, 852,1 704 mg·kg-1 )groups.On the 9th day of 10 days’gavage,all except the normal group were fed ethanol (1.5 mL/only)to induce gastric mucosal inju-ry in rats with acute gastric ulcer.Gastric ulcer index, the rate of ulcer inhibition were calculated for each group.The morphological changes of gastric mucosa were observed.The gastric mucosa levels of PGE2 , NO,SOD and MDA were determined.Results The LD50 and 95%confidence limit of the polyphenols’ef-fective parts from Punica granatum were 8 520.9 mg· kg-1 and 7 291.2 ~9 914.4 mg·kg-1,respectively. Pathology showed that the organs receiving dose of 16 000 mg · kg-1 had different degrees of damage . Compared with the model group,the extract from Puni-ca granatum significantly repaired the gastric mucosa, and significantly increased the gastric mucosa levels of NO and reduced MDA content,and improved SOD content and the levels of PGE2 .Conclousion The dose of 5 063 mg · kg-1 of polyphenols effective part from Punica granatum showed no death.The dose of 16 000 mg · kg-1 of polyphenols effective parts from Punica granatum could cause varying degrees of dam-age in heart,liver,lung,kidney or the death of mice. The LD50 and 95% confidence limit of the polyphenols effective parts from Punica granatum were 8 520.9 mg ·kg-1 and 7 291.2 ~9 914.4 mg·kg-1,respective-ly.The extract from Punica granatum plays a protective role against gastric mucosa damage induced by absolute ethanol,and the mechanism may be related to promo-ting ulcer epithelial cells synthesis,enhancing mucosal regeneration function,regulating NO content and en-hancing antioxidant capacity.

Objective To develop a method for determination of the plasma protein binding rate of bisoprolol in human plasma by high performance liquid chromatography ( HPLC) combined with hollow fiber-based liquid-phase microextraction ( HF-LPME) . Methods Method of liquid phase microextraction was optimized. The concentration of bisoprolol in the reconstitute solution was analyzed by HPLC. The mobile phase consisted of water-methanol-acetonitrile-0. 1% phosphoric acid (50:34:6:10). The excitation wavelength was 232 nm and emission wavelength was 300 nm. Through the linear regression equations, the total and free concentrations were obtained, and then the protein binding rate was calculated. Results At low, middle, and high concentration, the protein binding rate of bisoprolol was 31. 2%, 32. 0% and 31. 8%, respectively. Conclusion The proposed method is proven to be simple, fast and reproducible, and is feasible for the determination of plasma protein binding rate of bisoprolol. Bisoprolol moderately binds with plasma protein independent of concentration.

Objective To investigate the protective mechanism of Interleukin 33 (IL-33) in preventing myocardial ischemia and reperfusion injury in rats. Methods A rat model with myocardial I/R with 32the adult male SD rats , which were randomly divided into 4 groups: SO group , I/R group , IL-33 + I/R group , SB230580 + IL-33 + I/R group. The levels of LDH, CK, TNF-α, IL-6, HMGB1, Bcl-2, total caspase-3, cleaved caspase-3 and P-P38 were detected. Results After reperfusion, IL-33 significantly decreased the levels of serum LDH and CK and the expression of TNF-α, IL-6 , cleaved caspase-3 , but significantly increased the expressions of Bcl-2, p-P38 (P < 0.05). SB230580 attenuated the protective role of IL-33 on myocardial I/R in a certain degree. Conclusions IL-33 may prevent myocardial I/R injury via inhibiting inflammation and cardiocyteapoptosis by way of P38 MAPK signaling pathway.

Objective To observe the alteration of capillary density and apelin/APJ expression in soleus muscles of high-fat diet rats.Methods Male 5-week-old Sprague-Dawley rats were randomly divided into a control group and a high-fat diet group.After 12 weeks of high-fat diet,16 rats were se lected and randomly divided into a sedentary group and a treadmill running group.The exercise rats underwent 60-minute treadmill running at 26 m/min 5 days a week for 10 weeks.The body weight,body fat and blood lipid level were measured for all rats.The protein expression of Soleus CD31 and apelin was determined using immunohistochemical staining,soleus apelin content was determined using the radioimmunoassay,and the mRNA expression of apelin/APJ was detected using real-time PCR.Results Compared with the control group,significant increase was observed in the body weight,body fat and the level of total triglyceride,total cholesterol and low density lipoprotein cholesterin,but significant decrease was found in the high density lipoprotein cholesterin in the high-fat diet group.There were no significant differences in the capillary density and mRNA levels of apelin/APJ between the two groups.Compared with the sedentary high-fat diet group,significant improvement was observed in the body weight and blood lipid level of the treadmill running group.Moreover,significant increase was also observed in the capillary density,the expression of apelin/APJ mRNA,as well as that of apelin protein in the treadmill running group (P＜0.05).Conclusion The treadmill running can significantly increase capillary density of obese rats,as it may activate the expression of Apelin/APJ.

Objective To detect the PML-RARα fusion gene expression of APL patients with FISH and fluorescent quantitative PCR and discuss the sensitivity and specificity of two techniques. Methods The detection of the PML-RARα fusion gene expression of 75 APL patients with FISH and fluorescent quantitative PCR were carried out simultaneously. Results Eighty eight patients of primary and relapse or remission phases were examinated and total conformity rate was 96.59 %. Fourteen primary patients were detected and conformity rate was 100 %. Seventy four relapse or remission patients were detected and conformity rate was 95.95 %. Stem cell essays were detected for six times and conformity rate was 100 %. Conclusion The sensitivity of FISH and fluorescent quantitative PCR is identical for primary APL patients and FISH is more sensitive. But the sensitivity of FISH is weaker than that of fluorescent quantitative PCR for detection of residue disease.

Objective To analyze the risk factors of fevers after percutaneous nephrostolithotomy (PCNL) and to determine a more effective prophylaxis method.Methods A retrospective analysis of 320cases who underwent PCNL for renal calculi from 2008 to 2011 (men 233,women 87,age between 22 years to 72 years) was made.The average age of the patients and the average diameter of the stones were 42 years and 3cm(0.8 -6 cm),respectively.We analyzed factors such as age ( ＞60 years and ≤60 years),stone size ( ＞2.0 cm and ≤2.0 cm),operative time ( ＞60 min and ≤60 min),irrigation pump pressure ( ＞120 mm Hg and ≤ 120 mm Hg),obstructive conditions,preoperative urinary tract infection and fever.Wethen compared the fever rate with each of the risk factors.Results There were 59 cases with fever after PCNL; 18.4％ of the cases had a temperature over 38 ℃.There were two cases of pyemia.The patients whose stone diameter ＞ 2 cm had a post-operative fever rate of 22.4％.The fever rate in patients whose stone diameter ≤2.0 cm was 10.4％.The fever rate in patients with an operative time ＞60 min and ≤60 min was 27.2％ and 10.4％,respectively.Patients with irrigation pump pressure ＞ 120 mm Hg and ≤ 120mm Hg had post-operative fever rates of 28.3％ and 11.0％,respectively.There was statistical significance between each post-operative fever risk factor group.Conclusions The post PCNL fever risk factors are stone diameter ＞ 2.0 cm,operation time ≤60 min and irrigation pump pressure ＞ 120 mm Hg.The effective prophylaxis policy of post-operative fever are the pre-operative using of broad-spectrum antibacterial agents,shorter operative time and lower irrigation pump pressure.A two-stage operation procedure will also reduce the post-operative fever.

Objective To delineate the potency of YC-1 on the proliferation,apoptosis,cell cycle and the protein expression of Caspase-3,-8,-9 in U937 and THP-1 leukemia cell lines.Methods MTT assay was performed to detect proliferation.Flow cytometry was used to measure the apoptosis and cell cycle.The expression of Caspase-3,-8 and-9 were detected by Western blot.Results The MTT assay showed that cell proliferation was inhibited in a concentration-dependent manner in 1.0,3.0,10.0 μmol/L YC-l-treated U937 and THP-1 cells.The survival rates for YC-1 after 24 h in U937 cells were (76.5±4.4) ％,(68.7±6.8) ％,(60.9±13.2) ％ respectively and (94.1±1.4) ％,(81.4±2.0) ％,(72.7±3.0) ％ respectively in THP-1 cell,compared with the control group (100 ％),there were significant differences (F =15.870,126.629,P ＜ 0.01).The apoptosis rates for 1.0,3.0,10.0 μmol/L YC-1 after 24 h were (40.7±1.0) ％,(55.6±2.3) ％,(71.8±1.5) ％respectively in U937 cells and (34.6±2.0) ％,(50.3±3.5) ％,(59.6±4.6) ％ respectively in THP-1 cells.With the control group (4.7±1.4) ％,(1.8±1.0) ％,there were significant difference (F =937.229,200.447,P ＜ 0.01).However,there was no significant difference for cell cycle.In addition,Cleaved Caspase-8 and Cleaved Caspase-3 expression after 1.0,3.0,10.0 μmol/L YC-1 treated for 24 h were significantly higher than control,but the expression of Caspase-9 did not appear significant change in U937 cells.As the same concentration and time point,Cleaved Caspase-3 expression increased with no change of Caspase-9 or Caspase-8 in THP-1 cells.Conclusion YC-1 effectively suppress the proliferation with little effect on cell cycle,but induce the apoptosis,have no effect on cell cycle,and the mechanism of apoptosis may be related to the Caspase activation in U937 and THP-1 cell lines.

Objectives To provide prenatal diagnosis and genetic counseling for four athigh-risk pregnant women with a suspected family or personal history of fragile X syndrome (FXS) by genetic screening of fragile X mental retardation (FMR1) gene.Methods This study was conducted on four pregnant women (No.l to 4) who received outpatient treatment in Peking University First Hospital from August 2014 to June 2016.Genomic DNA was extracted from peripheral blood samples of the pregnant women and six of their family members,four of which were suspected or confirmed FXS and the other two were FMR1 gene carriers.Amplide X kits were used to detect CGG repeat size in FMR1 gene.Two amniocytes and one chorionic villi samples were collected from three pregnant women to extract DNAs for FMR1 gene and karyotyping analyses.Results There were patients diagnosed with FXS in all the families by detecting CGG repeat numbers in FMR1 gene.The pregnant woman No.1 was a permutation carrier;No.2 carried normal FMR1 alleles while her brother had a mutation with over 20 CGG repeats in FMRI gene at chromosome X.No.3 and 4 were full mutation carriers with over 200 CGG repeats in FMR1 gene.After genetic counseling,No.3 decided to terminate the pregnancy due to abnormal fetal karyotype (47,XY,+21) and full mutation of FMR1 alleles.No.1 and 4 continued to pregnancy as their fetuses were normal in FMR1 alleles and karyotype.No.2 continued to pregnancy as her fetus was free of FXS risk.Conclusions Prenatal diagnosis and genetic counseling should be conducted on women at highrisk for FXS to avoid birth defects.People with a family history of FXS should be tested for FMR1 gene carrier status.