Aim To investigate whether Ca2+/calmodulin-dependent kinase Ⅱ(CaMKⅡ)contribute to tumor necrosis factor ?(TNF-?)-induced cardiomyocyte hypertrophy.Methods The protein content was assayed with Lowry's method.The cardiomyocytes volumes were measured by computer photograph analysis system.The protein synthesis was assayed with[3H]-lencine incorporation method.[Ca2+]i transient was measured by Till image system by cell-loading Fura-2/AM.The expression of CaMKⅡ?B was determined by Western blot.Results ① TNF-? significantly induced the increase of protein content,[3H]-leucine incorporation and cell size;These responses were significantly suppressed by KN93,a selective CaMKⅡ inhibitor.② TNF-? increased the amplitude of the spontaneous Ca2+ transients in cultured ventricular myocytes from the neonatal rat;CaMKⅡ inhibitor KN93 can suppress the elevation induced by TNF-?.③ TNF-? significantly increased the expression of CaMKⅡ?B.Concluslon CaMKⅡ signal pathway are involved in TNF-?-induced cardiomyocyte hypertrophy in rats.

Aim To investigate whether Ca~(2+)/calmodulin-dependent kinase Ⅱ(CaMKⅡ)contribute to tumor necrosis factor α(TNF-α)-induced cardiomyocyte hypertrophy.Methods The protein content was assayed with Lowry's method.The cardiomyocytes volumes were measured by computer photograph analysis system.The protein synthesis was assayed with[~3H]-lencine incorporation method.[Ca~(2+)]_i transient was measured by Till image system by cell-loading Fura-2/AM.The expression of CaMKⅡδ_B was determined by Western blot.Results ① TNF-α significantly induced the increase of protein content, [~3H]-leucine incorporation and cell size;These responses were significantly suppressed by KN93, a selective CaMKⅡ inhibitor.② TNF-α increased the amplitude of the spontaneous Ca~(2+) transients in cultured ventricular myocytes from the neonatal rat;CaMKⅡ inhibitor KN93 can suppress the elevation induced by TNF-α.③ TNF-α significantly increased the expression of CaMKⅡδ_B.Concluslon CaMKⅡ signal pathway are involved in TNF-α-induced cardiomyocyte hypertrophy in rats.

Objective To investigate the dose-respeuse relationship of levobupivacaine for spinal anesthesia in patients undergoing cesarean section.Methods Sixty ASA Ⅰ or Ⅱ patients aged 25-35 yr weighing 60-90 kg scheduled for elective cesarean section under spinal anesthesia were enrolled in this study.Spinal puncture was performed at the L3,4 interspace.A 25 G spinal needle was inserted into the subarachnoid space cephalad.The patients were randomized to receive levobupivacaine 7.5 mg (group L1 ),10 mg (group L2),12.5 mg (group L3) or 15 mg (group L4) diluted to 3 ml with 5% glucose over 30 s (n = 15 each).The degree of pain was evaluated at the time of skin incision using visual analogue scales (VAS) (0 = no pain and 10 = worse pain ever).Effective analgesia was defined as VAS = 0.Spinal analgesia was inadequate when VAS≥1 and then 1% lidocaine 5-10 ml was given epidurally.The doses of levobupivacaine for effective analgesia in 50% and 95% of patients ( ED50 and ED95 ) and 95 % confidence interval were calculated by Probit method.Results The ED50 of levobupivacaine was 9.0 mg (95% confidence interval 7.8-9.9 mg) and ED95 13.2 mg (95% confidence interval 11.6-17.6 mg) for spinal anesthesia.Conclusion The ED50 and ED95 of levobupivacaine for spinal anesthesia in patients undergoing cesarean section are 9.0 mg and 13.2 mg respectively.

Background It is estimated that discoidin domain receptor 2 (DDR2) and matrix metalloproteinase-13 (MMP-13) play an important role in the development of tumor angiogenesis.However,their effects on choroidal neovascularizaiton (CNV) have not been clarified yet.Objective This study was to observe the expression pattern of DDR2 and MMP-13 in CNV and to further investigate the regulation role of MEK/ERK and PI3K/Akt pathways to the expression of DDR2 and MMP-13 in CNV.Methods CNV models were established in 78 Brown Norway (BN) rats by retinal photocoagulation with 532 nm laser.Then the animals were randomly divided into the normal control group (n =7),the model control group (n =39),PD98059 (MEK1 inhibitor) group (n =16) and LY294002 (PI3K inhibitor) group (n =16),and 5 mmol/L PD98059 or 5 mmol/L LY294002 3 μl was intravitreally injected 1 day and 7 days after photocoagulation in the PD98059 group or LY294002 group.The expression of DDR2 and MMP-13 mRNA and proteins in the CNV area were detected by using reverse transcription PCR (RT-PCR),and the expression levels of p-ERK/ERK and p-Akt/Akt protein were detected by Western blot assay.CNV thickness was measured by pathological examination 14 days after photocoagulation,and the changes of CNV thickness,the expression levels of DDR2 and MMP-13 in CNV were compared among the model control group,PD98059 group and LY294002 group.Results Three days after photocoagulation,the cells within the lasered lesions proliferated,then CNV formed 7 days after photocoagulation and became stable 14 days after photocoagulation.Immunohistochemistry staining indicated that DDR2 was weakly expressed in the cells of ganglion cell layer,inner nuclear layer and vascular endothelial cells;while MMP-13 was strongly expressed in the cells of the inner limiting membrane layer,photoreceptor layer and sclera layer.Both DDR2 and MMP-13 were strongly expressed in CNV area.Double immunofluorescence staining revealed that MMP-13 and DDR2 co-expression in CNV area.RT-PCR revealed that the relative DDR2 mRNA levels at 1 day,3 days,7 days and 14 days after photocoagulation were 55.22±4.03,47.74±2.23,14.82±4.56 and 5.59±2.47 respectively,while the relative MMP-13 mRNA levels were 25.54±3.83,43.51±4.36,10.90±4.00 and 5.23±3.23 respectively.Compared with the normal control group,the expression of DDR2 and MMP-13 were significantly increased (all at P＜0.05).Immunofluorescence staining showed that the relative fluorescence unit (RFU) values at 1 day,3 days,7 days and 14 days after photocoagulation were 2.73±0.53,5.21±0.31 and 3.22±0.33 for DDR2 and 1.66±0.17,3.57±0.44,2.67±0.21 for MMP-13,respectively.The RFU values in the PD98059 group and LY294002 group 14 days after photocoagulation were 1.14±0.19,1.03±0.14 for DDR2 and 1.37±0.25,1.24±0.20 for MMP-13,respectively.Compared with the model control group,the differences were statistically significant (both at P＜0.05).Western blot results showed that,compared to the normal control group,the expression levels of p-ERK and p-Akt pretein increased at day 7 after photocoagulation (both at P＜0.05),and returned back to the baseline at day 14 after photocoagulation (both at P＞0.05).Both PD98059 and LY294002 treatment were able to attenuate the thickness of CNV to 57.21％ and 50.34％ at day 14 after photocoagulation and further decrease the expression levels of DDR2 and MMP-13 in CNV (all at P＜0.05).Conclusions The expressions of DDR2 and MMP-13 up-regulate in laser-induced CNV.MEK/ERK and PI3K/Akt pathways suppress the development of CNV by regulating the expression of DDR2 and MMP-13.

Objective To evaluate the effect of dexmedetomidine on the efficacy of patient-controlled intravenous analgesia (PCIA) with morphine after elective radical gastrectomy. Methods One hundred and twenty ASA Ⅰ or Ⅱ patients aged 41-64 yr weighing 50-80 kg undergoing elective radical gastrectomy were randomly divided into 2 groups of 60 patients, according to the composition of PCIA solution:group I morphine (group M)and group Ⅱ morphine + dexmedetomidine (group MD). In group M the PCIA solution contained morphine 100 mg in 200 ml of normal saline (NS), while in group MD the PCIA solution contained morphine 100 mg+dexmedetomidine 200 μg in NS 200 ml. PCIA was started immediately after operation. A loading dose of 6 ml was given iv at the end of operation. PCIA setting was as follows:background infusion 1 ml/h, bolus dose 3 ml and lockout interval 10 min. VAS score was maintained at ≤4 and Ramsay score at 2-3. The total amount of morphine consumed, the number of attempts and successfully delivered doses within 24 and 48 h after operation were recorded. Postoperative complications including nausea, vomiting, bradycardia, hypotension, oversedation and respiratory depression were recorded. Results The total amount of morphine consumed, the number of attempts and successfully delivered doses within 24 and 48 h after operation were significantly smaller and the incidence of nausea and vomiting and pruritus was significantly lower in group MD than in Sroup M. No bradycardia,hypotension, oversedation or respiratory depression was observed in either group. Conclusion Dexmedetomidine added to intravenous morphine PCA can improve the analgesic efficacy after radical gastrectomy with less adverse effects.

Objective To determine the dose-response relationship of 0.2％ ropivacaine for ultrasoundguided stellate ganglion block (SGB).Methods Seventy-five ASA physical status [or Ⅱ patients with migraine,aged 23-55 yr,with body mass index of 22-28 kg/m2,scheduled for elective ultrasound-guided SGB,were randomly divided into R1-5 groups (n =15 each) using a random number table.In R1,R2,R3,R4 and R5 groups,the patients underwent ultrasound-guided SGB with 0.2％ ropivacaine 1,2,3,4 and 5 ml,respectively.A successful SGB block was confirmed by the onset of ptosis (Horner syndrome) on the injected side.Probit analysis was used to calculate the effective dose of 0.2 ％ ropivacaine in 50 ％ and 95 ％ of the patients (ED50 and ED95) and 95％ confidence interval (95％ CI).Results The ED50 of 0.2％ ropivacaine for ultrasound-guided SGB was 2.2 ml (95％CI 1.9-2.5 ml) and ED95 was 3.2 ml (95％CI 2.8-4.1 ml).Conclusion The ED50 and ED95 of 0.2％ ropivacaine for ultrasound-guided SGB are 2.2 and 3.2 ml,respectively.

Background Our previous study demonstrated that hyperglycemia aggravate the choroidal neovascularization (CNV) by promoting the chemotaxis process of bone marrow-derived cells (BMCs).Bioluminescence imaging (BLI) can dynamically monitor CNV in vivo.However,how diabetes mellitus (DM)participate in CNV is still in research.Objective This study was to dynamically observe the influence of BMCs to CNV under hyperglycaemia by using BLI combined with histopathology.Methods BMCs from luciferase-green fluorescent protein (Fluc-GFP) double transgenic mice were injected to adult wild type C57BL/6J mice (nine mice per group) via caudal vein to create the chimera models with a chimerism degree higher than 85％,and the chimeric mice were randomized into the control group and DM group based on randomized number table.Streptozotocin [60 mg/(kg · d)] was intraperitoneally injected daily for 5 days to establish the DM models in the chimeric mice of the DM group.CNV was induced in the chimeric mice of both control group and DM group with 532 nm laser photocoagulation.BLI signal of BMCsFluc+GFP+ was in vivo examined by IVIS Kinetics system 1,3,5,7,14,21 and 28 days after CNV modeling.At the seventh day after laser,part of mice were sacrificed,and choroidal and retinal sections were prepared for histopathological examination.The length and thickness of CNV were compared between the control group and DM group.The use and care of experimental followed Statement of ARVO.Results The chimerism degree of the chimeric mice was (88.85 ± 2.46) ％ 28 days after BMCs transplantation,and the blood glucose concentration in the DM group was (17.88±0.86)mmol/L.Histopathological examination revealed that CNV broke through the Bruch membrane toward subretinas.The length of the CNV was (338.67±33.17) μm in the DM group and (180.33±24.68)μm in the control group,showing a significant difference between the two groups (t =8.943,P＜0.05).However,no significant difference was seen in the CNV thickness between the two groups (t =1.790,P＞0.05).Light signals appeared 1 day and reach strongest 7 days after CNV modeling in both groups.The Light signals were stronger in the DM group than those in the control group on 5,7,14 and 21 days after CNV modeling (t =3.411,5.594,5.067,2.663,all at P＜0.05).Conclusions Hyperglycemia can promote more BMCs to participate in the pathogenesis and aggravation of CNV.The behavior of BMCs in CNV can be evaluated using BLI in vivo.

Objective To investigate the effect of hydrogen-rich saline postconditioning on the expression of vascular endothelial growth factor receptor-1 (VEGFR1) during myocardial ischemiareperfusion (I/R) injury in rats.Methods Thirty-six adult male Sprague-Dawley rats,weighing 250-280 g,were randomly divided into 3 groups (n=12 each) using a random number table:sham operation group (group S),group I/R and hydrogen-rich saline group (group H2).Myocardial I/R was induced by 30 min ligation of anterior descending branch of the left coronary artery followed by 2 h reperfusion.In group H2,hydrogen-rich saline 5 ml/kg was injected intravenously at 5 min before reperfusion,while the equal volume of normal saline was given in group I/R.Left ventricular systolic pressure (LVSP),left ventricular enddiastolic pressure (LVEDP) and ± dp/dtmax were measured and recorded during reperfusion and at 120 min of reperfusion.The rats were sacrificed at 120 min of reperfusion,and myocardial specimens were obtained for determination of myocardial infarct size,contents of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) (using ELISA),and expression of VEGFR1 (by Western blot).At 120 min of reperfusion,blood samples were collected from the common carotid artery to measure cardiac troponin I (cTnI) concentrations in serum.Results Compared with group S,LVSP and ± dp/dtmax were significantly decreased,LVEDP was increased,the myocardial infarct size was enlarged,cTnI concentrations in serum and contents of IL-6 and TNF-α were increased,and the expression of VEGFR1 was down-regulated at 120 min of reperfusion in H2 and I/R groups.Compared with group I/R,LVSP and ± dp/dtmax were significantly increased,LVEDP and myocardial infarct size were decreased,cTnI concentrations in serum and contents of IL-6 and TNF-α were decreased,and the expression of VEGFR1 was up-regulated at 120 min of reperfusion in group H2.Conclusion Hydrogen-rich saline postconditioning can reduce myocardial I/R injury possibly by upregulating myocardial VEGFR1 expression and inhibiting inflammatory responses in the myocardium of rats.

Objective To evaluate the effects of impaired glucose tolerance (IGT) on ventricular remodeling. Methods Parameters of every subject including left ventricular mass ( LVM), left ventricular mass index (LVMI), E/A ratio, 75 g oral glucose tolerance test (OGTT), ambulatory blood pressure monitoring(ABPM) data including 24-hour mean systolic blood pressure(mSBP) and 24-hour mean diastolic blood pressure(mDBP) were collected. Then the relationship of IGT and myocardial remodeling related parameters were analyzed. Results The rate of diastolic dysfunction was higher in the IGT combined with hypertensive group(74% ) compared with the hypertensive group( 39% )( x2 = 6. 5, P ＜ 0. 05 ). The rate of diastolic dysfunction was higher in the IGT group( 34% ) compared with the normal group( 10% ) (x2 = 5.2,P ＜0. 05). The rate of Left Ventricular Hypertrophy (LVH)in the IGT combined with hypertensive group (24%) was higher than the other three groups (Hypertension group 7%, IGT group 0, Normal group 0) (x2 =4.561,P ＜0.05), and there was no significance between the rest three groups (P ＞0.05).Stepwise multiple regression showed age and 2 Hours' Postprandial Blood Glucose were independent risk factors of E/A ratio. Conclusions These results suggested that IGT is a possible contributor to left ventricular hypertrophy and diastolic dysfunction, and is one of the histopathology of left ventricular remodeling.

OBJECTIVETo determine if locally administered bone morphogenetic protein-2 (BMP-2) and osteoprotegerin (OPG) improved osteogenesis and new bone formation by trans-sutural distraction osteogenesis.

METHODSTwenty four dogs were divided into three groups randomly and received new internal trans-sutural distraction osteogenesis treatment. Five days after operation, infusion apparatus with double-tube was inserted to submucosa near the distracted zone to deliver controlled release agent of recombinant human bone morphogenetic protein-2/poly (lactic-co-glycolic acid)/fibrin sealant (rhBMP-2/PLGA/FS) in group A and group C. Recombinant human osteoprotegerin/fibrin sealant (rhOPG/ FS) was injected three weeks later in group B and group C. Histology staining and bone histomorphometry were used to measure the changes of maxillary bone sutura after distraction for 1, 2, 4 and 6 weeks.

RESULTSNew bone formation observed in distracted zone showed a significant increase in group A and C. Transmission electron microscope showed the osteoblast and osteocyte were active with dilated rough endoplasmic reticulum and a large number of chondriosomes and Golgi complex. After distraction for 6 weeks, indexes of osteoblast of group A, B, and C were 38.5 +/- 7.7, 35.7 +/- 6.5, and 41.7 +/- 11.0, indexes of osteoclast (Ioc) were 5.9 +/- 1.0, 1.2 +/- 0.3, and 2.8 +/- 0.4, bone trabecula thicknesses were (38.36 +/- 13.28), (66.20 +/- 9.16), and (51.85 +/- 9.92) microm respectively. Increased bone density and decreased Ioc were found in group B and C.

CONCLUSIONThe new elastic distractor is effective in inducing new bone formation. BMP-2 and OPG combination acts synergistically, and leads to significant enhancement of bone formation and remodeling.

Animals ; Bone Density ; Bone Morphogenetic Protein 2 ; Dogs ; Humans ; Lactic Acid ; Maxilla ; Osteoblasts ; Osteogenesis ; Osteogenesis, Distraction ; Osteoprotegerin ; Polyesters ; Polyglycolic Acid ; Polymers ; Recombinant Proteins ; Transforming Growth Factor beta