Objective To explore the electrophysiological characteristics and the expression of mRNA and protein of L-type calcium channel in rat bone mesenchymal stem cells(MSCs). Methods MSCs were isolated,cultured and purified.RT-PCR was used to detect the mRNA expression of ?1C,?1D,?1H,?1S.The protein expression of L-type calcium channel(?1C) was testified by immunohistochamical.Ion currents were recorded in MSCs using whole-cell patch clamp technques.Results CD29,CD44,CD106 expressed in about 93% MSCs and CD14,CD34,CD45 expressed negatively.A high expression of mRNA in ?1C was detected by RT-PCR but no expressions were observed in ?1D,?1G,?1H,?1S.Immunofluorescent double labeling showed an expression of ?1C subunits in MSCs.Moreover,inward currents were recorded in 16 of 36 cells using whole-cell patch clamp techniques.The currents were activated around-30?mV and peaked at 0 to 10?mV and were blocked by nifedipine(10??mol/L).These cells had larger currents with Ba~(2+)(10?mmol/L) in bath solution than with Ca~(2+)(2?mmol/L).Conclusion The results indicated that adult rat MSCs expresse functional L-type calcium channels.It is possible that this channel plays a role on the proliferation and differentiation of MSCs.

Objective To study the changes of antioxidant on ZCLA Mongolian gerbils' sera within different growth period.Methods The gerbils at the age of 1 month,3 months and 24 months were used in the experiment,with each 16(half male and half female),and the sera were collected for determining the MDA,SOD and GSH-Px.Results At the age of 3 months,MDA was the lowest,but GSH-Px was the highest,while the SOD was ascending with growing.Conclusion The changes of GSH-Px showed the direct relation to MDA,while the changes of the SOD was irrespective to the changes of MDA.

Objective To observe Mongolian gerbil's reproducing performance and growth indexes of biology for guidance of development and use of laboratory anima1. Methods 200 gerbils produced by seed-parents with good reproductive capacity and body constitution were selected (females and males were 100 each).When 2.5~3.0 months old,one male and one female were cohabited randomly for a long period of time.Results Every embryo gave birth to 14 young gerbils at best and 1 gerbil at least.The total embryos was 455 and the total young gerbils was 3191. The average young gerbils of every embryo were 7.01,and every embryo gave mostly birth to 5~9 young gerbils,accounting for 79.34 percent of all the embryos. The shortest time between two embryos was 20 days and the longest time was 127 days,and mostly were 20~60 days,accounting for 72.16 percent of total. Conclusion The body weight,body length and tail length of Mongolian gerbils grow rapidly from birth to 4 months old. The differences between female and male was not significantly before delectation,and the in the growing time after delectation,the body weight,body length and tail length of male gerbils were all greater than those of female gerbils.

Objective To observe the toxic reaction caused by short time contact with some common disinfectants such as 0.2% sodium hypochlorite,2%acetic acid and 2% glutaraldehyde and0.2%acetic acid,through wounded skin of laboratory animals and laborers.Methods Toxicity experiments of these disinfectants by artificial skin wound were carried on in guinea pigs,and the poisoning performance and the death situation of guinea pigs were observed.Results Light edema was caused by using 2% acetic acid,and the symptom disappeared after 3 days;it appeared the symptom of red bump and hemorrhagic spots after use 2% glutaraldehyde,and ecdysis occured after 4days,the red bump disappeared after 5 days with a brownish discoloration left;it didn't appear any toxic reaction by using 0.2% sodium hypochlorite and 0.2%acetic acid.Conclusion There was toxic effect of 2% acetic acid and 2% glutaraldehyde to animals and laborers.

Objective To observe the lumbosacral plexus nerves by diffusion tensor tractography (DTT) and quantitatively evaluate them by using diffusion tensor imaging (DTI) in healthy volunteers.Methods A total of 60 healthy volunteers (30 males and 30 females) underwent DTI scanning.Mean FA values of the lumbosacral plexus nerves (both sides of lumbar roots L3 to S1,proximal and distal to the lumbar foraminal zone) were quantified.Differences among various segments of lumbar nerve roots were compared with ANOVA test and SNK test.Differences between two sides of the lumbar nerve roots at the same lumbar segment were compared with paired-samples t test.Differences between the proximal and the distal nerve to the the lumbar foraminal zone at the same lumbar segment were compared with paired-samples t test.The lumbosacral plexus nerve was visualized with tractography.Results (1) The lumbosacral plexus nerve was clearly visualized with tractography.(2) Mean FA values of the lumbar nerve roots L3 to S1 were as followings:proximal to the left lumbar foraminal zone 0.202 ± 0.021,0.201 ± 0.026,0.201 ± 0.027,0.191 ±0.016,distal to the left lumbar foraminal zone 0.222 ± 0.034,0.250 ± 0.028,0.203 ± 0.026,0.183 ± 0.020,proximal to the right lumbar foraminal zone 0.200 ± 0.023,0.202 ± 0.023,0.205 ± 0.027,0.191 ±0.017,distal to the right lumbar foraminal zone 0.225 ±0.032,0.247 ±0.027,0.205 ± 0.033,0.183 ±0.021.Mean FA values were significantly different between the proximal nerve to the distal nerve in lumbar nerve roots L3,L4,S1 (t =-9.114-2.366,P ＜ 0.05),but not significantly different in L5 (P ＞ 0.05).Differences were not found between the right and left side nerves at the same lumbar segment (P ＞ 0.05).(3) The whole length of the lumbar roots nerve L3 to S1 can be visualized clearly by using DTT.Conclusions Diffusion tensor imaging and tractography can show and provide quantitative information of human lumbosacral plexus nerves.DTI is a potential tool for the diagnosis of lumbosacral plexus nerve disease.

The influence of intravesica1 pressure on urine for-mation was studied in dogs and rabbits prepared with ureteral fistulae and in man following ureteral cathe-terization. Reduction of urinary output following distension of the bladder occurred in all except two dogs. The mean rate of reduction in sixteen dogs was 37.3 +/- 4.9 per cent. The response was not blocked by tetracaine applied to the bladder mucosa or by systemic hexamethonium. The renal blood flow showed a significant reduction following distension of the bladder. After denervation or celiac ganglionectomy, the reduction of urinary output or of renal blood flow was prevented in the ipsilateral kidney. Coloring of the renal cortex by intravenously injected indigo carmine does not occur in animals with distended bladders. Adrenaline and serotonin produced and enhanced the effect on the urinary response of the distended bladder. The injection of a small amount of blood or urine from animals with distended bladders into undistended animals produced a significant decrease in urine formation in the recipients. In five human subjects, a marked reduction of urine flow was noted following bladder distension. We conchlde that the intravesical pressure may regulate the formation of urine through a short vesico-renal reflex mediated by the celiac ganglion and through a long vesico-hypothalamic reflex which releases the antidiuretic hormone.
Animals ; Denervation ; Dogs ; Epinephrine ; Fistula ; Ganglia, Sympathetic ; Ganglionectomy ; Hexamethonium ; Humans ; Indigo Carmine ; Kidney ; Mucous Membrane ; Rabbits ; Reflex ; Renal Circulation ; Serotonin ; Tetracaine ; Ureter ; Urinary Bladder

AIM: To investigate the changes of expression of calcium-sensing receptor(CaSR) in the rat cardiac tissue at different age and its relation with anoxia-reperfusion(A/R) injury.METHODS: RT-PCR was used to detect the mRNA expression of CaSR.The A/R injury was remodeled in vitro,and the ultrastructure of cardiac tissue was observed under transmission electron microscope.RESULTS: The expression of CaSR mRNA in the rat cardiac tissue increased gradually after birth,and reached to its maximum in the 1st month.In the 2nd to 3rd month,the expression were almost equal to that at birth and then increased again to a higher level.The expression level of CaSR mRNA increased more significantly than those in control groups after 40 min of anoxia and reperfusion of 1 h and 2 h, and decreased after 3 h and 4 h.Moreover,the longer the reperfusion time lasted,the more serious the changes of ultrastructure were observed.CONCLUSION: Our results demonstrate that the CaSR is expressed in the rat cardiac tissue.The mRNA expression of CaSR changes with age and is involved in the anoxia-reperfusion injury.

Objective To explore the effects and possible mechanism of calcium-sensing receptor(CaSR) in rat myocardial H9c2 cells hypertrophy model using angiotensin Ⅱ (Ang Ⅱ).Methods Cardiac hypertrophy model was established by treating cultured H9c2 cells with Ang Ⅱ in vitro.Hypertrophic H9c2 cells were treated with gadolinium chloride (GdCl3,a specific agonist of CaSR) and/or with Calhex231 (a specific inhibitor of CaSR) and 3-methyladenine (3-MA,a specific inhibitor of autophagy) to divided into 5 groups (six in each group):control,Ang Ⅱ,GdCl3 + Ang lⅡ,GdCl3 + Calhex231 + Ang Ⅱ,GdCl3 + 3-MA + Ang Ⅱ groups.To evaluate the status of H9c2 cells hypertrophy,protein content was determined through a coomassie brilliant blue protein kit and the expression of Ca2+/calmodulin-dependent protein kinase Ⅱ (CaMK Ⅱ) and the phosphorylation form (pCaMK Ⅱ/CaMK Ⅱ) was analyzed by Western blotting.The protein expression of CaSR,autophagy maker [Beclin-1,micmtubule-associated protein 1 light chain 3(LC3)Ⅱ/LC3 Ⅰ,P62] and Ca2+/calmodulin-dependent-protein kinase-kinase-β (CaMKKβ)-activated protein kinase (AMPK)-mammalian target of rapamycin (mTOR) pathway was analyzed by Western blotting.Results ①GdCl3 further increased H9c2 cells protein content [control group:(2.52 ± 0.84) g/L,Ang Ⅱ group:(8.72 ± 3.60) g/L GdCl3 + Ang Ⅱ group:(14.17 ± 4.49) g/L,all P ＜ 0.05] and the expression of CaSR (control group:0.22 ± 0.04,Ang Ⅱ group:0.43 ± 0.02,GdCl3 + Ang Ⅱ group:0.63 ± 0.08,all P ＜ 0.05) and pCaMK Ⅱ/CaMKⅡ (control group:0.25 ± 0.05,AngⅡ group:0.51 ± 0.03,GdCl3 + AngⅡ group:0.77 ± 0.06,all P＜ 0.05) induced by Ang Ⅱ.Calhex231 suppressed the increasing of hypertrophy indicators induced by GdCl3 [GdCl3 + Calhex231 + AngⅡ group,CaSR:0.41 ± 0.16,protein content:(9.92 ± 2.54) g/L,pCaMK Ⅱ/CaMKⅡ:0.58 ± 0.08,all P ＜ 0.05].②GdCl3 promoted the effect of Ang Ⅱ in regulation of autophagy such as Beclin-1 protein increased (control group:0.31 ± 0.06,AngⅡ group:0.55 ± 0.09,GdCl3 + AngⅡ group:0.74 ± 0.08,all P ＜ 0.05),LC3 Ⅱ/LC3 Ⅰ increased (control group:0.28 ± 0.06,Ang Ⅱ group:0.56 ± 0.10,GdCl3 + Ang Ⅱ group:1.00 ± 0.15,all P ＜ 0.05) and P62 protein decreased (control group:0.54 ± 0.03,AngⅡ group:0.34 ± 0.02,GdCl3 + AngⅡ group:0.15 ± 0.03,all P ＜ 0.05).Moreover,Calhex231 suppressed autophagy induced by GdCl3 (GdCl3 + Calhex231 + Ang Ⅱ group,Beclin-1:0.53 ± 0.14,LC3 Ⅱ/LC3 Ⅰ:0.57 ± 0.12,P62:0.28 ± 0.05,all P ＜ 0.05).③GdCl3 increased pCaMKKβ/CaMKKβ (control group:0.43 ± 0.09,AngⅡ group:0.76 ± 0.12,GdCl3 + AngⅡ group:1.19 ± 0.21,all P ＜ 0.05),pAMPK/AMPK (control group:0.38 ± 0.11,AngⅡ group:0.68 ± 0.08,GdCl3 + AngⅡ group:1.18 ± 0.08,all P ＜ 0.05) and decreased pmTOR/mTOR (control group:0.90 ± 0.10,Ang Ⅱ group:0.54 ± 0.04,GdCl3 + AngⅡ group:0.29 ± 0.09,all P ＜ 0.05).Furthermore,Calhex231 blocked the effect of GdCl3 on the above-mentioned proteins changes (GdCl3 + Calhex231 + Ang Ⅱ group,pCaMKKβ/CaMKKβ:0.75 ± 0.06,pAMPK/AMPK:0.57 ± 0.05,pmTOR/mTOR:0.51 ± 0.08,all P ＜ 0.05).Conclusion Inhibiting calcium-sensing receptor expression has reversed H9c2 cell hypertrophy induced by Ang Ⅱ,which may be related to suppressing autophagy and suppressing CaMKKβ-AMPK-mTOR pathway.

Objective To investigate the method to isolate and culture hepatic stellate cells ( HSCs) for studying the cellular mechanisms of hepatic frbrosis.Methods HSCs were isolated by nycodenz density gradient centrifugation after the hepatocytes obtained from adult male gebils were digested with pronase, collagenase and DNase, infused via portal vein.The cell viability was determined by trypan blue exclusion test.The purity of HSCs was identified by detectingα-SMA, desmin immunohistochemical staining.Results The yield rate of HSCs was 0.5~1 ×107 per gerbil liver, and the cell viability was more than 90%.The percentage ofα-SMA-positive cells was more than 75%after 3 days primary culture and almost 100% cells were α-SMA and desmin positive in passage culture.Conclusion The successful protocol of primary culture of Mongolian gerbil HSC provide a technical support for research of relevant liver diseases and drug development in the future.

OBJECTIVETo study the effects of low concentration dopamine(DA) on hydrogen peroxide-induced apoptosis in cultured rat cardiomyocytes as well as the possible molecular mechanisms.

METHODSCultured neonatal rat cardiomyocytes were randomly divided into the following groups: control group (control), hydrogen peroxide group (H2O2), pretreated with low concentration dopamine ( DA + H2O2), dopamine receptor l(DR1) antagonist group (DR1 + DA + H2O2), dopamine receptor 2(DR2) antagonist group (DR2 + DA + H2O2). The cell apoptosis was then assessed by MTT and flow cytometry. The cellular ultrastructure changes were observed by transmission electron micro- scope. The activity of lactate dehydrogenase(LDH )and superoxide dismutase (SOD) in cell medium was analyzed by colorimetry. The protein expressions of Cytochrone c, Caspase 3 and Caspase 9 were obtained by Western blot.

RESULTSCompared with hydrogen peroxide group, low concentration dopamine(10 µmol/L) decreased the apoptosis rate and the expression of protein of apoptosis related protein, enhanced SOD activity, decreased LDH activity. DR1 antagonist SCH-23390 treatment inhibited dopamine induced cardiac protective effect. DR2 antagonist haloperido treatment had no changes compared with dopamine group.

CONCLUSIONAbove findings indicate that low concentration dopanine inhibits apoptosis induced by hydrogen peroxide in neonatal rat cardiomyocytes, which is partly associated with the activation of DR1.

Animals ; Apoptosis ; Benzazepines ; Caspase 3 ; metabolism ; Caspase 9 ; metabolism ; Cells, Cultured ; Dopamine ; pharmacology ; Hydrogen Peroxide ; L-Lactate Dehydrogenase ; metabolism ; Myocytes, Cardiac ; drug effects ; Rats ; Rats, Wistar ; Receptors, Dopamine D1 ; metabolism ; Superoxide Dismutase ; metabolism