AIM:To supply the theoretic evidences of elongating the lifetime of neuron-like cells differentiated from adult rat mesenchymal stem cells, we investigated the relationship between the differentiation and apoptosis in the process of induction. METHODS: The mesenchymal stem cells(MSCs) were isolated primarily from rat bone marrow, and purified by passage culture. The 5th passage of MSCs was induced by ?-mercaptoethanol and all-trans-retinoic acid (ATRA). After 1 h, 3 h and 5 h of induction, the cells were stained immunocytochemically with anti-MAP-2 and anti-GFAP antibodies, respectively. In addition to counting the ratio of neuron-like cells in MSCs, DAPI staining was employed to identify whether the differentiated cells have an apoptotic morphological changes. The ratio of apoptotic cells at 1 h, 3 h and 5 h after induction were detected by flow cytometry (FCM). RESULTS:Neuron-like cells appeared among MSCs and could be identified by MAP-2 staining 30 min after MSCs being treated by ?-mercaptoethanol and ATRA. However, 5 h after induction, some of these neuron-like cells began to float and die. With DAPI staining, the nuclei of some dying cells were shown to be condensed and later fragmented. The ratios of cells undergoing apoptosis at 1 h,3 h and 5 h after induction by ?-mercaptoethanol and ATRA were 1.5%?6.1%?21.5% and 1.5%?1.7%?12.3%, respectively . CONCLUSIONS:1. ?-mercaptoethanol and ATRA had the different ability that induced MSCs to differentiate to neuron-like cells. 2. Apoptosis was also initiated in the process of differentiation, and there is positive correlation between the ratio of differentiation and apoptosis.

Objective In previous experiment we proved that rat marrow stromal cells(MSCs) could be induced to differentiate to neuron\|like cells by ? mercaptoethanol and all\|trans\|retinoic acid(ATRA), but the neuron\|like cells showed apoptosis sooner after the differentiation.In this study, we used several sets of combinations of anti\|apoptotic reagents and inductive reagents to induce the differentiation of MSCs,to prevent the differentiated cells from apoptosis and to elongate the lifetime of the neuron\|like cells which might be use in clinic. Methods The MSCs were isolated primarily from bone marrow of adult rat,and purified by passage culture.The 5th passage of MSCs was induced by single or combinative induction,which included RA,Melatonin(10 -9 mol/L), bFGF,bFGF+RA,Melatonin+RA,Melatonin+bFGF.At 1?h,5?h and 24?h of induction, the ratio of neuron\|like cells and the numbers of survived neuron\|like cells among 6 groups were compared.In addition,immunocytochemical stainings with anti\|MAP\|2 and GFAP were also performed on cells of 5?h of induction. Results Neuron\|like cells were all MAP\|2 positive,but GFAP negative.The ratios of neuron\|like cells in MSCs at 1?h after induced by six combinations of reagents were 0 24,0 28,0,0,0 44,0 64;at 5?h were 0 56,0 28,0,0,0 76,0 83 and at 24?h were 0 10,0 27,0,0 14,0 35,0 59 respectively.The ratios of survived neuron\|like cells of differentiated cells at 24?h after induction by RA,Melatonin,Melatonin+RA and Melatonin+bFGF were 0 08,0 37,0 05,0 51 respectively.Conclusions\ 1.bFGF can't induce MSCs into neuron\|like cells independently.Moreover,it suppressed the differentiation effects of RA,while enhancing the effects of MT.2.MT can induce MSCs differentiating into neuron\|like cells and has the anti\|apoptotic effects on the induced neuron\|like cells.3.When MT and bFGF were added together,not only the ratio of neuron\|like cell in MSCs was increased, the lifetime of neuron\|like cells was also elongated.

Objective To investigate the expression of nuclear factor erythroid-2 related factor 2 (Nrf2) gene in chronic myeloid leukemia (CML) and explore its relationship with clinical characteristics and thioredoxin reductase (TrxR).Methods The expressions of Nrf2 and TrxR genes in bone marrow cells and K562 cells were analyzed in 30 bone marrow preparations of CML patients in different phases,including 20 in chronic phase,3 in accelerated phase,7 in blastic phase by real-time quantitative polymerase chain reaction.Ten health subjects were served as normal controls.Results The relative quantitation expression of Nrf2 and TrxR mRNA were 5.601±1.069 and 9.017±2.398 in chronic phase,1.698±0.349 and 5.590±1.015 in accelerated phase,1.252±0.807 and 5.050±1.469 in blastic phase,1.377± 0.344 and 1.867±0.977 in normal controls.The expressions of both Nrf2 and TrxR mRNA in CML had significant differences from those of the normal controls (x2 =14.680,P =0.002,x2 =8.271,P =0.041).The expression of Nrf2 mRNA in accelerated phase,blastic phase group showed no significant difference (Z =0.011,P =0.496),but lower than that in chronic phase group (Z =2.155,P =0.016,Z =2.534,P =0.006).The difference between the first visit and post-treated group was significant (Z =2.015,P =0.022).The expression in K562 cells and normal controls had significant difference (Z =1.898,P =0.029).In CML patients,the expression of Nrf2 was positively correlated with that of TrxR (r =0.738,P =0.037).Conclusion The expression of Nrf2 gene is higher in the first visit group of CML than that in the other groups,and is decreased after therapy,which may be the molecular marker predicting the progress of CML.Nrf2 mRNA expression level is correlated with TrxR.

Objective It is important to precisely determinate the single nucleotide polymorphisms (SNPs) in many genes in‐cluding genes related with base excision repair (BER) pathway .This research is conducted to evaluate the role of polymerase chain reaction with confronting two‐pair primers (PCR‐CTPP) in analyzing the SNPs of BER pathway .Methods Four common SNPs of BER pathway (OGG1 Ser326Cys ,XRCC1 Arg399Gln ,APE1 Asp148Glu and‐141T/G in the promoter region) was detected with PCR‐CTPP .10 of the products were sent for genotype sequencing .Compare the results of PCR and sequencing to evaluate the accu‐racy of PCR‐CTPP .Results The genotypes were exactly the same as the sequencing .Conclusion The PCR‐CTPP was a reliable and rapid detective technology for SNPs genotyping .Its broadest application would be great help for gene variant analysis .

AIM:To explore the production and cytotoxicity of the reactive oxygen species(ROS)induced by diallyl trisulfide(DATS)in HL-60 cells.METHODS:HL-60 cells were either treated with various doses of DATS alone,or DATS combination with apocynin,a specific NADPH oxidase inhibitor,or with antioxidant N-acetyl-L-cysteine(NAC)for 0,1,3,6,12 and 24 h,respectively.The intracellular ROS level was measured by flow cytometry.The activity of NADPH oxidase was evaluated by NBT reduction experiment.The content of both malondialdehyde(MDA)and the protein carbonyl were analyzed by spectrophotometer.RESULTS:The results from flow cytometry indicated that DATS significantly increased the intracellular ROS level in HL-60 cells(P

Objective To detect the expression of apurinic/apyrimidinic endonuclease 1 (APEI) and explore its correlation with the expression of mutant p53 in hepatocellular carcinoma (HCC). Methods The expression of APE1 and mutant p53 was detected by SP immunohistochemical method in 10 specimens of normal liver tissue, 40 specimens of liver cirrhosis tissue and 103 specimens of HCC tissue which were collected at the Department of Pathology of Daping Hospital from 1991 to 2004. All data were analyzed by chi-square test, correla-tion analysis and K Independent-Samples Tests. Results The expression rate of APE1 in HCC was 100.0%, which was significantly higher than that in normal liver tissue (40.0%) and liver cirrhosis tissue (82.5%) (χ~2= 47.852, P < 0.01). The expression of APE1 was only detected in the nucleus in normal liver tissue. Ectopic expression of APE1 in cytoplasm was detected in liver cirrhosis tissue and HCC tissue, with the rate of 20.0% and 53.4%, respectively (χ~2=20.757, P <0.01). There was statistical difference in clinical staging and pathological grading of HCC with different combinations of APE1 expression (intranuclear or ectopic expression) and mutant p53 expression (positive or negative expression) (χ~2=12.910, 14.481, P < 0.01), and HCC with ectopic expression of APE1 and positive expression of p53 had high malignant degree. Conclusion Overexpression and ectopic expression of APE1 in cytoplasm may play important roles in the genesis and progression of HCC, and the ectopic expression of APE1 and p53 mutation may have synergistic effect.

Objective:To investigate the association between age and cardiac tamponade after radiofrequency ablation of atrial fibrillation(RAAF).Methods:Clinical data of patients undergone de novo AF ablation procedures at Beijing Anzhen Hospital from January 2013 to December 2016 were retrospectively collected.Patients were divided into an elderly group(age ≥60 years)and a non-elderly group(age <60 years). Logistic regression analyses were used to evaluate the association between old age and the risk of cardiac tamponade complicating RAAF.Results:A total of 5 313 patients were involved in this study, including 41 patients(0.77%)with cardiac tamponade.The proportion of cardiac tamponade was higher in the elderly group than in the non-elderly group(1.1% or 32/2 950 vs.0.4% or 9/2 363, χ2=8.489, P=0.004). One patient with cardiac tamponade in the elderly group required immediate surgical repair whereas none in the non-elderly group did.No patient died in hospital.Multivariate Logistic regression analysis showed that the risk of cardiac tamponade increased in the elderly group, compared with the non-elderly group( OR=2.570, 95% CI: 1.190-5.570, P=0.017). Stratified analysis revealed that among females and patients with oral anticoagulants, left atrium dimension < 40 mm or procedure duration≥ 120 min in the elderly group carried a higher risk of cardiac tamponade than those in the non-elderly group( OR=1.011, 2.914, 3.922 and 3.244, P<0.05). Conclusions:Old age(age ≥60 years)is an independent risk factor for cardiac tamponade complicating RAAF.

AIM:To investigate high-density lipoprotein ( HDL) subclass distribution and to analyze the rela-tionship between HDL subclasses with plasma glucose and lipids in metabolic syndrome ( MS) .METHODS:Apolipopro-tein A-I ( apoA-I) contents of plasma HDL subclasses were determined by two-dimensional gel electrophoresis associated with immunodetection .The concentrations of lipids and apolipoproteins in the plasma were measured by an automated bio -chemical analyzer .RESULTS:Compared with the controls , the levels of fasting plasma glucose ( FPG) , total cholesterol (TC), triglyceride ( TG), low-density lipoprotein cholesterol ( LDL-C), LDL-C/high-density lipoprotein cholesterol (HDL-C), apolipoprotein B100(apoB100), apoB100/apoA-I, systolic blood pressure (SBP), body mass index (BMI) and HDL3b were increased in the MS patients (P<0.05).Meanwhile, HDL-C, apoA-I and preβ2-HDL, HDL2a and HDL2b were decreased in the MS patients (P<0.01).With the increase in the plasma glucose level , the contents of HDL2a and HDL2b were decreased in the MS patients (P<0.05), while preβ1-HDL was increased (P<0.05).With the decrease in the HDL-C level, the content of HDL2b was decreased in the MS patients (P<0.01), while preβ1-HDL was increased (P<0.01).With the increase in the TG level and the decrease in the HDL-C level, the content of HDL2b had a decrea-sing trend and the content of small-particle preβ1-HDL had an increasing trend , indicating that HDL maturation metabolism was disrupted.The correlation analysis showed that FPG was negatively correlated with the levels of HDL 2a and HDL2b, HDL-C was negatively correlated with the level of preβ1-HDL and positively correlated with the level of HDL 2b , and TG was positively correlated with the levels of preβ1-HDL and HDL3b .CONCLUSION:With the increases in the plasma glucose and TG, and the decrease in HDL-C in the MS patients, HDL particles have minifying tendency , and the maturation me-tabolism of HDL particles is disrupted .

Objective Our purpose was to investigate the pathogenic gene mutation of a Han Chinese family with vitreous amyloidosis.Methods The 9 individuals(proband,1 affected member and 7 unaffected members) of the family were selected and their DNA was extracted from peripheral blood.The 4 exons of transthyretin(TTR) gene were amplified by polymerase chain reaction(PCR) technique.The amplified products of TTR gene were sequencing by Sanger technique.We also selected 100 unrelated healthy individual as the control group.Results By DNA sequencing,a heterozygous mutation was found in 4 of the 9 subjects from the family.The transition of adenine to cytosine(AAG ＞ ACG) was detectable in exon 2 of TTR,which changed the amino acid composition at codon 35 (Lys35Thr).This mutation did not presented in control group.Conclusion The heterozygosis mutation of TTR gene Lys35Thr should be a pathogenic mutation for the family with vitreous amyloidosis.

Objective To discuss the methods, advantages and indications of multipoint puncturing in performing endovascular therapy for complex lower extremity arterial occlusive diseases. Methods During the period from Oct. 2011 to Oct. 2013, a total of 46 patients with complex lower extremity arterial occlusive diseases were treated with endovascular therapy by using multipoint puncturing technique. The puncturing type, the advantages of multipoint puncturing technique and the success rate of this technique were analyzed. Results The multipoint puncturing was divided into three types: type Ⅰ : puncturing from the opposite direction to deal with the same target vessel; type Ⅱ: direct puncturing of the target vessel; and type Ⅲ:using the same puncturing direction to deal with different target vessels. The success rate of endovascular treatment was 80.4%. Conclusion Multipoint puncturing technique helps improve endovascular treatment success rate for lower extremity arterial occlusive diseases. Full understanding of the advantages of multipoint puncturing technique, perfect preoperative planning, precise puncturing technique and proper interventional equipments are helpful to ensure a successful treatment.