BACKGROUND:Fetal liver stem cel s have the potential to differentiate into hepatocytes and bile duct cel s, and participate in the repair and reconstruction of the liver, which is an important source of hepatocytes. But there are a little amount of fetal liver stem cel s in human body, and how to obtain a certain number of high-purity fetal liver stem cel s is currently a hot research. OBJECTIVE:To construct a siRNA carrier that can effectively inhibit the expression of connexin 43 (Cx43) in rat fetal liver stem cel s, and to investigate the effect of Cx43 inhibition on the proliferation and cel cycle of fetal liver stem cel s cultured in vitro. METHODS:Fetal liver stem cel s were cultured by the suspension culture in vitro, siRNA sequences targeting Cx43 (Cx43-siRNA) and negative control sequence (NC-siRNA) were designed and synthesized. Then, rat fetal liver stem cel s were transferred electrophoretical y and divided into three groups:blank group, NC-siRNA group, Cx43-siRNA group. Real-time PCR and western blot were used to assess the knockdown efficiency. Cel ular proliferation was determined by cel growth curve and cel counting kit-8 assay. The cel cycle was analyzed by flow cytometry. RESULTS AND CONCLUSION:After transfection, the Cx43 gene and protein expression levels were declined dramatical y in the Cx43-siRNA, NC-siRNA and blank groups, and the cel s grew faster. The number of cel s at G0/G1 phase decrease, but the number of cel s in S phase increased. There were significant differences between the groups (P<0.05). Electrophoretic transfer of Cx43-siRNA can promote the proliferation of cultured fetal liver stem cel s and optimize the cel culture.

The combination of teachers'leading role and the students'main role is a basic principle of teaching. This paper discusses how to play the leading role of teachers from pre-teaching preparation and classroom teaching to coaching students and answering the questions after school..

Thetherapeuticeffectsofintermitentcyclicetidronatedisodium(HEBP)ofdifferentdosesonovariectomy-inducedosteoporosiswerestudied.53femaleWistarrats,weighingwithanaver-ageof266gmwererandomlydividedinto5groups.Gourp1hadbilateralovariectomy(OVX)andwasgivenlowdoseHEBPof25mg/kgBW/day;group2hadOVXandgivenmediumdoseHEBPof36mg/kgBW/day;group3hadOVXandgivenhighdoseHEBPof51mg/kgBW/day;group4hadOVXandgroup5hadshamOVX.HEBPwasadministratedoralyduringfastingfor5daysfolowedbyarestperiodof25daysandtheregimewasrepeatedfor3times.Alanimalsweresacrificedat90thdayafterOVX.RadioimmunoassayofserumestradiolconfirmedthesuccessofOVX.TheserumCaandPcon-tent,AKPaseactivity,femoralrelativedryweight,CaandPcontentwereanalysedrespectively.Therighttibiaewereprocessedundecalcifiedforquantitativebonehistomorphometry.TheresultsshowedthatthebonecalciumcontentandtrabecularbonevolumeofOVX-ratstreatedwithHEBP,especialywiththelowdosegroupwerehigherthanthatofthesimplyOVX-ratandtheboneturnoverparameterssuchasAKPaseactivity,boneformationsurface,boneresorptionsurface,activeformationsurface,bonemineralappositionrateandboneformationratewerelowerthanthoseofthesimplyOVX-rat.TheresultsindicatetheHEBPcouldretardthedegreeofosteopososisthroughreducingtheelevatedboneresorptionandboneturnoverinducedbyOVX.Meanwhile,theresultssuggestthattheefectoflowdoseHEBP(25mg/kgBW/day)wasmoreefectivethanthatofthehighdose.

Objective To establish a polarographic method for detecting fluoroacetamide.Methods On the condition of strong basic solution with high temperature fluoroacetamide was decomposed to form a decomposed product ALC-La-F - complex.In a pH4.90buffer solution containing KNO 3 and hexamethylenetetramine,the decomposed product was determined with com-plex-adsorptive wave at-0.67V by polarography.Re sults The detection limit and linear range of the method was2.0?10 -8 mol/L and5.0?10 -8 -1.2?10 -6 mol/L respectively.The regressive equation was y=3.49x+0.042with a correlation coefficient r=0.9996.The method had been applied to the determination of fluoroacetamide in wheat flour samples satisfyingly.Conclu-sion This method was simple and suitable for the determination of fluoroacetamide in flour.

Objective To investigate the expression and regulation of ASPP family mRNA in NSCLC cell lines.and study the diagnostic and therapeutic significances of apoptosis stimulating of ps3 expression in NSCLC Patients.Methods Real-time fluorescence quantitative PCR wag established and used to measure the expression Ievels of ASPP mRNA in NSCLC cell lines A549(wild-type p53)and NCI-H157 (mutant-type p53)with p53 different genetic background,and ASPP mRNA was also detected in NSCLC tissue,peripheral blood of 37 NSCLC patients.Western Bloting was used to examine the levels of ASPP1 and ASPP2 proteins of NSCLC cell Iines.Chemosensitivity of the cell lines to CDDP was analyzed by MTT assay,and detect the change of ASPP1 and ASPP2.Compared with the ASPPs mRNA expression between NSCLC patients and healthy controls.and monitoring ASPPs mRNA expression in patients with NSCLC chemotherapy.Results The slope rotes of standard curves were-3.249,-3.358 respectively.and correlation coefficients were more than 0.98.Amplification efficiency of standard curves were 1.156.1.028 respectively.The IC_(50) values of NCI-H157 and A549 cells were 3.70.10.48 μg/ml respectively.Mean ASPP1 ASPP2 mRNA levels were a statistically significant 2.66 folds and 6.98 folds higher in NCI-H157 cell compared to A549 cell The levels of ASPP1 mRNA in tumor tissues and peripheral blood were (4.27±0.57)×10~3,(2.49±0.32)×10~3 in patients with NSCLC respectively,and the levels of ASPP2 mRNA in tumor tissues and peripheral blood were(2.34±0.75)×10~3,(7.00±1.17)×10~3 in Patients with NSCLC respectively.The levels of ASPP1 and ASPP2 mRNA in tissues and peripheral blood were(1.32±0.21)×10~4,(1.46±0.31)×10~4 and(1.38±0.19)×10~4,(1.28±0.18)×10~4 in control groups respectively.The ASPP1,ASPP2 mRNA levels in patients with NSCLC were significantly lower than those of control groups(t=2.58,3.94,3.62,3.76,P＜0.05).Before chemotherapy the levels of ASPP1 and ASPP2 mRNA in patients with NSCLc were(2.34±0.56)×10~3 and(6.64±0.72)×10~3,and after 2 periods of postoperative chemotherapy the levels were(3.66±0.64)×10~3 and(9.42±0.44)×10~3,the expression was higher than those of ehemotherapy before(t=3.02,4.50,P＜0.05).Conclusions The expression of ASPP1.ASPP2 mRNA in p53 mutant-type cell lines is higher than that in p53 wild-type cell lines.The expression of ASPP maybe has close relationship with chemotherapeutic sensitivity of NSCLC.In NSCLC patient chemotherapy process,the change of ASPP1 and ASPP2 mRNA expression may be related with chemotherapy medicine function.Enhance the expression of ASPP1 and ASPP2 will be beneficial to the tumot treatment.

Objective To explore the inhibitory effects of endogenous hydrogen sulfide, a novel and important gas-eous transmitter generated in mammalian tissues mainly by cystathionine β-synthase (CBS) or cystathionineγ-lyase (CSE) on the apoptosis of the rat hepatic BRL cell line in physiological condition. Methods BRL cells were cultured, and divid-ed randomly into several groups in different phases of the experiment, including negative-siRNA (control) group, CBS siRNA (CBS 1~3) group and CSE siRNA (CSE 1~3) group, which were used to select the most efficient sequences of siRNAs at 48 or 24-hour transfection. Solution group and (CBS+CSE) siRNA group were added to detect the variation of apoptosis. The BRL cell line was observed and evaluated at 0, 4, 8, 12, and 24 hrs after siRNA transfection. When the mechanisms of the apoptosis were detected, CBS/CSE siRNAs were transfected individually or jointly into BRL cells, and compared with nega-tive-siRNA group to examine the variation. The genic and protein expression of CBS/CSE were detected by RT-PCR and Western blot assay. After transfection of CBS/CSE siRNA, the apoptosis of BRL cells was detected by Hoechst stain and flow cytometry (FCM). The mitochondrial membrane potential (MMP) changes were observed by fluorescent staining. Western blot assay was used to examine the protein expression of intracytoplasm cytochrome C (Cyt C) and cleaved-caspase 3. Re-sults CBS and CSE were observed in BRL cells. After transfection of CBS/CSE siRNA, endogenous H2S generation de-creased and the apoptosis of BRL cells increased. Accordingly, the expression of intracytoplasm-Cyt C and cleaved-caspase 3 increased. Conclusion The inhibition of endogenous H2S synthesis induced the apoptosis of BRL cells under physiologi-cal condition, which may be involved in mitochondrial pathway of apoptosis.

Objective: To study the effects of TNFα on wound healing. Methods: The skin lesion and incisions were made on mice and rhTNFα was applied in both skin lesion and incision. Tissue strips were evaluated for tensiometry and histologic features after healing. Results: It was found that rhTNFα markedly improved wound healing both in skin lesion and incision wound model, it also markedly enhanced the breaking strength of treated incision and increased wound cellularity in rh TNF α treated groups. Conclusion: The results suggest that TNFα play an important role in regulating inflammatory reactions of local injured tissues and it may improve the functions of fibroblasts, mononuclear phagocytes and endothelial cells. rhTNFα of suitable doses can promote wound healing. This shows that TNFα, as a wound healing promoter, has the potentials for clinical application.

Objective To investigate expression of inducible nitric oxide synthase (iNOS) and production of nitric oxide(NO) in cultured vascular smooth muscle cells(SMCs)from human abdominal aortic aneurysm(AAA). Methods TypeⅠ collagenase enzyme digestion method was used to culture AAA SMC . Cell proliferation curves for AAA SMC proliferation ability were evaluated,? smooth muscle actin (? SMA) , iNOS protein expression of AAA SMC and nitrite and nitrate (NO 2 -+NO 3 -,NO X ) in cultured media were measured. Results AAA SMCs had a limited proliferation ability, with 45%?5 8% positive rate of ? SMA and 86 7%?4 6% of iNOS protein expression. High concentration of NO X existed in the cultured media. Conclusion AAA SMCs present abnormal but limited proliferation and changed phenotype; The expression of iNOS and high production of NO of AAA SMCs may play an important role in AAA pathogenesis.

Objective: To investigate the effect of N methyl D aspartate(NMDA) on cochlear potentials and to find out the possible neurotoxic effect of NMDA on cochlea function in guinea pigs. Methods: After basal compound action potentials (CAP) and cochlear microphonics (CM) were recorded by round window electrode, animals ( n =5) were treated with Hanks applied to the round window membrane (RWM) for 20 min as control. Then 100 ?mol/L NMDA was applied to the RWM for another 20 min. Results: Hanks produced no obvious changes in CAP threshold, CAP N1 and CM latency and amplitude. CAP thresholds at all frequencies of tone burst were significantly elevated by application of NMDA, threshold shifts ranged from 13 27 dB. NMDA significantly reduced the CAP N1 amplitudes at all intensities of stimulations. CAP amplitudes were suppressed by 50% 75%. NMDA also significantly increased the CAP latency, the latency of CAP evoked by 6 kHz tone burst at intensity of -90 dB(output attenuation) was (1.9?0.06) ms after Hanks treatment and (2.76?0.21) ms after NMDA treatment ( P

Objective To explore the application of autologous bone marrow stem celI transplantation in the treatment of chronic ischemic disease of lower extremity and evaluate the effect.Methods Forty patients with chronic ischemic limbs were treated from January 2008 to April 2009.The bone marrow of each patient was stimulated by an injection of the recombinant human granulocyte macrophage colony-stimulatory factor(GCSF),300?g /d,for 2-3 days.Then,200 mL of bone marrow was withdrawn from the iliac spine and the stem cells were obtained.After the bone marrow separation and purification,they were transplanted into the more severely ischemic limb.After the transplantation,they were foIlowed up by phone once a month for six months.The skin temperature,transcutaneous oxygen pressure(TcPO2),digital subtraction angiography(DSA)and ankle/brachial index(ABI)were rechecked at 6 months later,and the symptoms of pain,cold,numbness,intermittent claudication and physical signs of ulcer and gangrene were also evaluated.Results All of the 40 patients were followed-up.At 6 months post-transplantation,the various indices were compared with before transplantation.(1)The skin temperature and TcPO2 had significante improvement (P0.05).(3)DSA and all of the clinical symptoms showed general improvement in more than 60% of cases.(4)There were no serious complications.Conclusions Transplantation of stimulated autologous bone marrow stem cells is effective in the treatment of ischemic disease of lower limb,which is a simple,safe and effective treatment for ischemic disease of lower limb.