BACKGROUND:Previous experiments reported that basic fibroblast growth factor(bFGF) can change the shape of corneal endothelial cells,accelerate cell division and chemotaxis,and repair corneal endothelia cells injury in vivo and in vitro. OBJECTIVE:To investigate the effect of bFGF on cat corneal endothelial cells proliferation. DESIGN,TIME AND SETTING:The cell morphological observation was performed at the Central Laboratory of the Affiliated Hospital of Medical College of Qingdao University between January 2005 and December 2006. MATERIALS:Domestic cats were purchased from Experimental Animal Center of the Affiliated Hospital of Medical College of Qingdao University. METHODS:The corneal endothelial cells of cats were primarily cultured,and neurone specific enolase(NSE) immunocytochemistry staining was performed. After cell passage,bFGF(1,10,100 ?g/L) was added,continuously hatched for 1-5 days. MAIN OUTCOME MEASURES:MTT method was used to determine the proliferation of corneal endothelial cells;meantime,inverted phase contrast microscope and transmission electron microscope were used to observe the ultrastructure of cells. RESULTS:At days 1,3 and 5 after bFGF was added,the absorbance value in 490 nm was significantly higher than that of control group,especially in 10 ?g/L group. CONCLUSION:The bFGF can promote proliferation of cat corneal endothelial cells with effective concentration of 10 ?g/L .

BACKGROUND:Taurine is an important non-enzymatic system antioxidant in the lens.The mechanism of its anti-oxidative effect is mainly to protect lens from oxidative injury by anti-lipid peroxidation.OBJECTIVE:This study was to observe the effect of exogenous taurine on lens epithefial cell apoptosis-induced by H2O2 in vitro.DESIGN:A randomized controlled animal experiment.SETTING:Department of Ophthalmology,Affiliated Hospital of Qingdao University Medical College.MATERIALS:Seventy-five adult New Zealand standard tabbits,of either gender,weighing 1.5-2.5 kg,were provided by Qingdao Laboratory Animal Center.Reagent kit for in situ detecting cell apoptosis(Sigma Company,USA),taurine and H2O2(Shanghai Guangda Chemical Reagent Factory,China)were included in this study.METHODS:This study was performed in the Department of Ophthalmology and Central Laboratory.Affiliated Hospital of Qingdao University Medical College from Mav 2005 to June 2007.Rabbit lenses were harvested and randomly divided into 3 groups:control group,in which,clear lenses were incubated in non-serum and non-phenolsulfonphthalein MEM medium which was renewed every 24 hours,H2O2 group,in which,clear lenses were incubated in non-serum and non-phenolsulfonphthalein MEM medium containing 1 mmol/L H2O2 with addition of 62 μL H202(30 g/L)every 6 hours,and H2O2+taurine group,in which,clear lenses were incubated in non-serum and non-phenolsulfonphthalein MEM medium containing 1 mmol/L H2O2 and 10 g/L taurine.which was renewed every 6 hours.The protocol was conducted in accordance with ethical guidelines for the use and care of animals.MAIN OUTCOME MEASURES:Observation of lens opacity 6.12,24.48 and 72 hours after culture;Lens epithelial cell apoptosis determined by DNA in situ end labeling and DNA fragment analysis.RESULTS:Lens opacity:The lens opacity in the H2O2 group was aggregrated gradually along with the time of oxidative injury.The lens opacity in the H2O2 group was severer than that in the H2O2+taurine group.Lens epithelial cell apoptosis:There were no apoptotic cells in the control group within 72 hours.The number of apoptotic cells in the H2O2 group was increased gradually with the prolonged time of oxidative injury.Till the 72nd hour,the cells were all tamed into apoptotic cells.A few aopototic celIs were found in the H2O2+taurine group since hour 24,and then were more and more,and the number of apoptotic cells accounted for about 30%at hour 72.The apoptotic rate in the H202+taurine group was significantly lower than that in the H2O group at each time point(q=8.6845,P＜0.01).There was no significant difference in apoptotic rate between the H202+taurine group and the control group (P＞0.05). Findings of DNA fragmentation assay:The DNA"ladder"was found in the H2O2 group at hours 24,36,48 and 72,while no DNA"ladder"but only normal electrophoresis straps were present in the other two groups 24 hours after culture.CONCLUSION:Taurine can inhibit the oxidative injury-induced apoptosis of rabbit lens epithelial cells,and alleviate lens opacity.

BACKGROUND: Oxidation injury can lead to the apoptosis of lens epithelial cells. Some researches have found that body supplement of vitamin C is beneficial to inhibit oxidation injury by increasing the level of vitamin C in aqueous humor.OBJECTIVE: To observe the inhibitive effect of vitamin C on H2O2 induced apoptosis of lens epithelial cell.DESIGN: Randomized controlled trial taking animal's lens as subject.SETTING: Department of Ophthalmology, Medical College of Qingdao University.MATERIALS: 120 adult New Zealand rabbits of both genders, weighing 3-5 kg, were selected. Vitamin C and 300 g/L H2O2 were purchased from Shanghai Guangda Chemical Reagent Factory.METHODS: The experiment was carried out in the Central Laboratory, the Affiliated Hospital of Medical College of Qingdao University from January 2002 to January 2004. ①Culture of lens: 240 lenses were isolat ed from 120 rabbits after being killed. Among them, 192 clear lenses were selected and divided randomly to three groups with 64 lenses in each group: control group, H2O2 group and H2O2+vitamin C group. Lenses in the control group were incubated in non-serum and non- phenolsulfonphthalein MEM medium, those in the H2O2 group incubated in non serum and non- phenolsulfonphthalein MEM medium containing 1 mmol/L H2O2 (62 μL of 30 g/L H2O2 was added into the medium every 6 hours to keep H2O2 level maintain 1 mmol/L in the medium), and those in the H2O2+vitamin C group incubated in non-serum and non- phenolsulfonphthalein MEM medium containing 1 mmol/L H2O2 and 1 mmol/L vitamin C. Under the same culture condition, the lenses in each group were detected at hours 6, 12, 18, 24, 36, 48, 72 and 108 after culture, respectively. ②Measurement of the opacity of lens: Under the white background, two black lines 0.5 mm in width, which were mutually vertical with an interval of 10 mm. The lenses were placed above the crossing to measure the opacity. ③Detecting the apoptosis of lens epithelial cells:The apoptosis of lens epithelial cells was examined using TUNEL method and DNA fragmentation assay under light microscope to calculate the apoptotic rate of epithelial cells.MAIN OUTCOME MEASURES: ①Opacity of the lenses in each group;②apoptotic rate of lens epithelial cells in each group.RESULTS: ①After 108 hours of culture, the opacity of the lenses in the H2O2+vitamin C group was obviously lower than that in the H2O2 group. ②The apoptotic rates in the control group were lower that those in the H2O2group at different time point and those in the H2O2+vitamin C groupat hours 24, 48 and 72 after culture(P＜0.05-0.01). The apoptotic rates in the H2O2+vitamin C group were significantly lower than those in the H2O2group at different time points after culture (P＜0.01). ③The findings of DNA fragment analysis showed that the DNA "ladder" was found in the H2O2 group during 24-72 hours, while no DNA "ladder" but only normal electrophoresis strap were present in the other two groups 24 hours after culture.CONCLUSION: Lens can take refuge from the oxidative injury of H2O2with the addition of vitamin C. As a result, the apoptosis rate of lens epithelial cells and further, cataract formation can be restrained.

Objective:To observe influence of enhanced external counterpulsation (EECP) on plasma levels of lipo‐protein-associated phospholipase A2 (Lp‐PLA2) and hsCRP in patients with coronary heart disease (CHD) ,and preliminarily explore anti -inflammatory mechanism of EECP in prevention and treatment of CHD .Methods :A to‐tal of 85 CHD patients were selected from our hospital ,randomly divided into routine treatment group (n=42) and EECP group [n=43 ,received EECP based on routine treatment ,once/d ,60min each time ,continuous five weeks were regarded as one course (35 times)] .Plasma concentrations of Lp‐PLA2 and hsCRP were measured in all sub‐jects at the enrollment day and after five -week treatment .Results:Compared with before treatment ,after five-week treatment ,there were significant reductions in plasma levels of Lp‐PLA2 [(58.46 ± 40.04)μg/L vs .(33.94 ± 23.22)μg/L] and hsCRP [ (3.54 ± 2.22)μg/ml vs .(2.19 ± 1.16)μg/ml] in EECP group (P<0.01 both) ,and there were no significant difference in routine treatment group between before and after treatment .Compared with routine treatment group ,after five-week treatment ,there were significant reductions in plasma levels of Lp‐PLA2 [ (56.87 ± 33.69)μg/L vs .(33.94 ± 23.22)μg/L] and hsCRP [ (3.63 ± 1.60)μg/ml vs .(2.19 ± 1.16)μg/ml] in EECP group , P<0.01 both .Conclusion:1. EECP of one course can significantly reduce plasma concentrations of Lp‐PLA2 and hsCRP in CHD patients , indicating that anti - inflammatory may be one of its mechanism in preven‐tion and treatment of CHD .

Objective:To explore the relationship among plasma level of lipoprotein‐associated phospholipase A2 (Lp‐PLA2) ,ankle brachial index (ABI) ,brachial‐ankle pulse wave velocity (baPWV) and coronary heart disease (CHD) .Methods :According to results of coronary angiography ,a total of 120 patients with suspected or diagnosed CHD were divided into CHD group (n=90) and non‐CHD group (n=30) .CHD group was further divided into sin‐gle vessel coronary disease group (single vessel group ,n=30) ,double‐vessel coronary disease group (double‐vessel group ,n=30) and multi‐vessel coronary disease group (multi‐vessel group ,n= 30) .Plasma Lp‐PLA2 level ,ABI and baPWV were measured and compared among all groups .Results:Compared with non‐CHD group ,there were significant rise in plasma Lp‐PLA2 level [ (23.60 ± 13.33)μg/L vs .(36.65 ± 17.24)μg/L] and baPWV [ (1244.27 ± 127.85) cm/s vs .(1753.08 ± 284.32) cm/s] in CHD group ,P<0.01 both .In CHD group ,compared with single vessel group ,plasma Lp‐PLA2 level significantly rose [ (25.81 ± 8.97)μg/L vs .(35.03 ± 9.80)μg/L vs .(49.13 ± 21.22)μg/L] in double‐vessel group and multi‐vessel group ,and that of multi‐vessel group was significantly higher than that of double‐vessel group ( P<0.05 or <0.01 );baPWV significantly rose [ (1579.77 ± 178.05 ) cm/s vs . (1808.07 ± 272.11) cm/s ,(1871.40 ± 306.03) cm/s] in double‐vessel group and multi‐vessel group , P<0.01 both . ABI of single vessel group and double‐vessel group were significantly higher than that of multi‐vessle group [ (1.19 ± 0.08) ,(1.17 ± 0.07) vs .(1.11 ± 0.15)] ,P<0.01 or <0.05 .Conclusion:Plasma Lp‐PLA2 level ,ABI and baP‐WV are related to CHD and its lesion degree ,combined measurement of these three indexes can predict CHD and its severity more accurately ,then preventing and treating CHD should be more effective .

Inflammation plays an important role in occurrence and progression of atherosclerosis.Among numerous studies on inflammatory markers, the relationship between high sensitive C-reactive protein (hsCRP) and coronary heart disease (CHD) has been widely studied.The present article made a review about the relationship between hsCRP and CHD.

PurposeTo explore the value of transesophageal echocardiography (TEE) monitoring mini-incision transthoracic occlusion of ventricular septal defect (VSD) with asymmetric ventricular defects occlude in preoperative selection of patients, intraoperative guidance monitoring and postoperative evaluation.Materials and MethodsForty-five cases of VSD undertook mini-incision transthoracic occlusion with asymmetric ventricular defects occlude were enrolled, their clinical data was analyzed retrospectively, preoperative transthoracic echocardiography (TTE) was applied for choosing appropriate cases. During surgery, TEE was employed for evaluating the VSD and in which perimembranous VSD was found in 21 cases, intracristal VSD in 15 cases and subpulmonic VSD in 9 cases, suitable eccentric type occluders were chosen, guide occluder was placed, and the occlusion effect was evaluated right after operation.ResultsThe procedures were completed successfully in 42 cases, with a successful rate of 93.3%, of which 21 cases had perimembranous VSD, 15 cases had intracristal VSD, and 6 cases had subpulmonic VSD. The diameter of the VSD ranged from 3 to 7 mm, averaging (4.5±0.7) mm, the diameter of occluders ranged from 4 to 8 mm, averaging (5.7±1.2) mm, there was a good positive correlation between size of VSD and occlude (r=0.87,P<0.05). All patients received follow-ups from 3 months to 24 months after operation, all the occluders located normally, with no more than mildly residual shunt, valve regurgitation or severe arrhythmia discovered.ConclusionMini-incision transthoracic occlusion of ventricular septal defect (VSD) with asymmetric ventricular defects occlude has high success rates, minimal injury, and lower complication rate. TEE can play a vital role by improving the success rate and safety of surgery.

Objective To investigate the effect of mitogen-activated protein kinase(MAPK)pathway on the transcriptional expression of mdr1 gene induced by doxorubicin ( DOX)and study the transcription regulation of mdr1 gene.Methods K562 cells were treated with DOX(0.01 μg/ml)with the initial concentration of 0.01 μg/ml for 24 hours,then change the culture media without DOX.K562 cells were cultured until the its status wag recovered.Subsequently the cells were treated with DOX(0.02μg/ml)for 24 hours again.The concentration of DOX was increaged until 0.05 μg/ml by following the protocol above.K562 cells were collected at the concentration of 0.01 μg/ml,0.03μg/ml and 0.05μS/ml DOX.Expression of mdr1 gene were examined by reverse transcription-polymerase chain reaction(RT-PCR).Pglycoprotein(P-gP)wag detected by flow cytometry.Western blot wag performed to detect ERK and P-ERk.K562 cells were pretreated with MAPK inhibitor PD98059 for 1 hour.and then DOX was added.RT-PCR and FCM were used to detect the expression of mdr1 mRNA and P-gp.Results When K562 cells were exposured to DOX.the phosphorylation of ERK wag increaged.the mdr1 gene wag highly expressed as well as its corresponding protein P-gp.When the concentration of DOX was 0.05μg/ml,the expression of mdr1 gene and P-gp were increased over 5 fold.When K562 cells were pretreated with MAPK inhibitor PD98059,the expression of mdr1 gene induced by DOX(the concentration was 0.03 μg/ml and 0.05 μg/m1)was effectively inhibited by(74.1±0.11)%and(70.2±0.14)%respectively.Conclusions DOX could induce the expression of mdr1 gene in K562 cells accompanied by the activation of MAPK/ERK pathway.The block of activation of ERK could inhibit the induced expression of mdr1 gene.

Objective To investigate the correlation between the mutation of pncA gene and the susceptibility to pyrazinamide ( PZA) in Mycobacterium tuberculosis complex ( MTBC) strains and to analyze the mutation of panD and rpsA genes in wild type isolates without pncA gene mutation.Methods The sus-ceptibilities of 108 MTBC strains to first-line drugs including PZA were detected by using the MGIT 960 TB system.PCR was performed to amplify the 16S rDNA and pncA, panD and rpsA genes.The PCR products were analyzed by DNA sequencing analysis .Results Among the 78 multidrug-resistant MTBC strains , 47 isolates (60%) were resistant to PZA.Four out of 30 (13%) strains that were sensitive to ethambutol , iso-niazid, rifampicin and streptomycin (EIRS) were resistant to PZA.The drug-resistant MTBC strains showed higher resistance rate to PZA than that of the EIRS sensitive strains .There were 49 ( 96%) PZA-resistant isolates and 4 (7%) PZA-sensitive isolates occurred pncA gene mutation.Most of the pncA gene mutations in the genomes of PZA-resistant strains were base substitution mutation , especially the His57Asp substitu-tion.The pncA gene mutations centralized in the regions of 160-169, 203-289, 309-396 and 413-467.Seven novel mutation sites of pncA gene were observed including T175C, C188A, G insertion at 68, AGC insertion at 235, C insertion at 339, CC insertion at 392 and GT deletion at 395.The mutation sites founded in the genomes of PZA-sensitive strains were different from those of the PZA-resistant strains .No mutation of the pncA gene and the upstream regulatory sequence was found in two PZA-resistant strains , NJ44 and NJ108 . The sequence analysis of panD and rpsA gene showed that the NJ 108 strain had panD gene mutation at G419A, but no mutation was detected in the NJ 44 strain.Conclusion The multidrug-resistant MTBC strains showed higher resistance rate to PZA .The pncA gene mutation was common in PZA-resistant MTB strains and the panD gene mutation was also worthy of attention .

Objective To evaluate the effects of interleukin-22(IL-22)on the expression of tazarotene-induced gene 3(TIG3)in HaCaT cells. Methods Cultured HaCaT cells were randomly divided into several groups to be treated with different concentrations (12.5, 25, 50, 100 μg/L)of IL-22 alone, or the combination of 50 μg/L IL-22 with the MAPK-ERK1/2 inhibitor PD98059 or the JAK/STAT inhibitor AG490 for 24 hours. Those HaCaT cells treated with phosphate buffered saline served as the control group. Subsequently, total proteins and mRNAs were extracted from the HaCaT cells. An immunofluorescence assay, Western blot and enzyme-linked immunosorbent assay (ELISA) were performed to determine the protein expression level of TIG3, and real-time fluorescence-based quantitative PCR to quantify the mRNA expression of TIG3 in HaCaT cells. Results The immunofluorescence assay showed that TIG3 protein was mainly expressed in the cytoplasm of HaCaT cells. As Western blot revealed, the protein expression level of TIG3 was 0.743 ± 0.035, 0.678 ± 0.040, 0.582 ± 0.041 and 0.328 ± 0.032 in HaCaT cells treated with IL-22 of 12.5, 25, 50 and 100μg/L, respectively, significantly lower than that in the control group (0.839 ± 0.045, all P<0.05). ELISA also showed a decrease in the protein expression of TIG3 in IL-22-treated HaCaT cells, which was consistent with Western blot results. Further more, the mRNA expression level (2-△△Ct)of TIG3 was significantly weaker in HaCaT cells treated with IL-22 of 12.5, 25, 50 and 100μg/L than in the control group (0.838 ± 0.036, 0.686 ± 0.061, 0.565 ± 0.047 and 0.457 ± 0.033 vs. 1.000, all P< 0.05). The decrease in TIG3 mRNA and protein expressions was significantly attenuated in HaCaT cells treated with the combination of 50 μg/L IL-22 with PD98059 or AG490 compared with those treated with 50 μg/L IL-22 alone. Conclusion IL-22 can dose-dependently inhibit the expression of TIG3 in HaCaT cells, likely through the MAPK-ERK1/2 and JAK2/STAT3 signaling pathways.