Aim To investigate the role of T type calcium channel of spinal cord and supraspinal on the pain threthold of the rats following chronic constriction injury(CCI) of the sciatic nerve.Methods Intrathecal and lateral ventricle injection were employed in this study.With Von Frey hair and radiant thermal stimulator,we measured the mechanical withdrawal threshold(MWT) and thermal withdrawal latency(TWL)of the rats after injected mibefradil.Results The rats of CCI group formed steady mechanical and heat hyperaglsia from the third day after operation to the end of this study.Administered intrathecally mibefradil 50,100,200 ?g can increase the CCI rats MWT and TWL.However,mibefradil administered lateral ventricle can reduce the CCI rats MWT and TWL.Conclusion Blocking T type calcium channel of spinal can inhibit mechanical and heat hyperalgesia of the CCI rats,However,bloking the T type calcium channel of supraspinal can enhance the mechanical and heat hyperalgesia of the CCI rats.

Objective To investigate the effects of isoflurane and sevonumne on apoptosis and expression of CD44 and CD54 in human lung cancer cell line A549.Methods Human lung cancer A549 cells were obtained from Shanghai Cell Biology Medical Research Institute,Chinese Academy of Sciences,and inoculated in 24 well culture plate.After being cultured for 24 h the cells were randomly divided into 3 groups:group Ⅰ control(group C);group Ⅱ isoflurane (group Iso) and group Ⅲ sevoflurane (group Sev).A 549 cells were exposed to 1.7% isoflurane and 2.5%sevoflurane for 4 h respectively in group Iso and Sev respectively,and were then cultured for another 24 h.Apoptosis and expression of CD44 and CD54 in A549 cells were detected with flow cytometer at 0 (T0),2 h(T1) and 4 h(T2) of and 24 h after(T3) exposure to isoflurane and sevoflurane.Results The percentage of apoptotic cells wag significantly higher at T2 and T3 in group Iso than in group C.The percentage of apoptotic cells was significantly higher at T1,T2 and T3 in group Sev than in group Iso and C.The expression of CD44 and CD54 at T1,T2 and T3 was significantly decreased as compared with the baseline at T0 in group Iso and Sev and was significantly lower in group Iso and Sev than in group C.Conclusion Isoflurane and sevoflurane can induce apoptesis of human lung cancer cell line A549, and sevoflurane is more effective. Isoflurane and sevoflurane can inhibit the expression of CD44 and CD54 of human lung cancer cell line A549.

Objective To investigate the effects of different concentrations of sevoflurane on Survivin expression in human adenocarcinoma cell line A549. Methods A549 cells were obtained from Shanghai Cell Biology Medical Research Institute, Chinese Academy of Sciences and inoculated in 96 well culture plate. After being cultured for 24 h, the cells were randomly divided into 4 groups: Ⅰ , Ⅱ , Ⅲ and Ⅳ groups exposed to 95 % O2 -5 %CO2,1.7%, 3.4% and 5.1% sevoflurane respectively. A549 cells were exposed to sevoflurane for 2, 4 and 6 h respectively and then cultured for another 48 h in Ⅱ , Ⅲ and Ⅳ groups. Proliferation of A549 cells were measured by methyl thiazolyl tetrazolium (MTT) assay, and apoptosis was detected with flow cytometer at 48 h after 2, 4 and 6 h sevoflurane exposure. The expression of Survivin in A549 cells was determined by Western blot analysis at 48h after 4 h sevoflurane exposure. Results The rate of proliferation inhibition and percentage of apoptotic cells were significantly higher while the expression of Survivin was significantly lower in a concentration-dependent manner in Ⅱ , Ⅲ and Ⅳ groups as compared with group Ⅰ . Conclusion Sevoflurane can inhibit proliferation and induce apoptosis of A549 cells by inhibition of Survivin expression.

ObjectiveTo investigate the role of T-type calcium channel in the spinal neurotoxicity of intrathecal (IT) lidocaine in rats.MethodsForty-eight adult male SD rats in which IT catheter was successfully implanted,weighing 230-270 g,were randomly divided into 4 groups ( n =12 each):dimethyl sulfoxide (DMSO)group (group D),lidocaine group (group L),mibefradil + lidocaine group (group M),normal saline + lidocaine group (group N).Another 12 rats served as control group (group C).DMSO and 10％ lidocaine 20μl were injected intrathecally in groups D and L respectively.After mibefradil 200 μg/10μl and normal saline 10 μl were injected intrathecally in groups M and N respectively,10％ lidocaine 20 μl was injected intrathecally in the two groups.The mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) were measured before IT injection and at 2,4,8 and 12 h and 1,2,3,4 and 5 d after IT injection (T0-9).Four rats were sacrificed at T6 in each group and their lumbar enlargements were removed for microscopic examination.ResultsCompared with group C,no significant change in MWT and TWL was found at each time point in group D,MWT was significantly increased at T1-8 and TWL was significantly prolonged at T1-7 in groups L and N,and MWT was significantly increased at T1-6 and TWL was significantly prolonged at T1-6 in group M ( P ＜ 0.05 ).Compared with groups L and N,MWT was significantly decreased at T1-4 and TWL was significantly shortened at T1-4 in group M ( P ＜ 0.05).Pathological injury was significantly reduced in group M as compared with groups L and N.ConclusionT-type calcium channel is involved in the spinal neurotoxicity of IT lidocaine in rats.

ObjectiveTo investigate the effects of sevoflurane on inhibition of invasive activity and migration of human adenocarcinoma cell line A549 by cisplatin.MethodsThe.human lung adenocarcinoma cell line A549 was obtained from Shanghai Cell Biology Institute,Chinese Academy of Sciences and cultured in RPMI 1640 culture medium containing 10％ fetal calf serum.The cells were inoculated in culture plate and cultured for 24 h and randomly divided into 4 groups:control group; 2.5 ％ sevoflurane group ; cisplatin group and cisplatin + 2.5 ％sevoflurane group.In groups sevoflurane,cisplatin and cisplatin + sevoflurane the cells were exposed to 2.5％sevoflurane or/and cisplatin 10μmol/L for 4 h respectively.The invasive activity of the cells was evaluated by Transwell chamber assay.The migration of the cells was determined by wound healing assay.The expression of MMP-2,MMP-9,Ezrin,and Fascin in the cells was detected by Western blot.ResultsBoth 2.5％ sevoflurane and cisplatin depressed invasive activity and migration of the A549 cells and down-regulated MMP-2,MMP-9,Ezrin and Fascin expression in A549 cells.The inhibitory effects of cisplatin on the A549 cells were potentiated by 2.5 ％ sevoflurane.ConclusionSevoflurane can enhance the inhibition of invasive activity and migration of human adenocarcinoma cell line A549 by cisplatin.

Objective To evaluate the role of calcium/calmodulin-dependent protein kinase Ⅱ (CaMK Ⅱ) in the up-regulation of expression of Cav3.2 T-type calcium channels in spinal cord in a rat model of neuropathic pain (NP).Methods Forty-eight male Sprague-Dawley rats,aged 3 months,weighing 220-250 g,were randomly divided into 6 groups (n =8 each)∶ sham operation group (group S),group NP,dimethyl sulfoxide group (group D) and different concentrations of a specific CaMK Ⅱ inhibitor KN93 groups (groups K1-3).NP was produced by chronic compression of dorsal root ganglion.The rats in groups D and K1-3 received a single intrathecal injection of dimethyl sulfoxide and KN93 15,30,60 nmol/L (10 μl),respectively,on 5th day after NP.Paw withdrawal threshold to von Frey filament stimulation (MWT) and paw withdrawal latency to thermal nociceptive stimulus (TWL) were measured before NP,before intrathecal injection on 5th day after NP,and at 30 and 60 min and 3,6 and 8 h after intrathecal injection on 5th day after NP (T1-7).The rats were sacrificed after the measurement of pain threshold at T7 and their lumbar enlargements were removed to detect the expression of Cav3.2 mRNA and protein using Western blot and RT-PCR.Results Compared with group S,MWT was significantly decreased,TWL was shortened and the expression of Cav3.2 mRNA and protein was up-regulated in NP,D and K1-3 groups (P ＜ 0.05).Compared with NP group,MWT was significantly increased,TWL was prolonged and the expression of Cav3.2 mRNA and protein was down-regulated in a concentration-dependent manner in K1-3 groups (P ＜ 0.05),and no significant change was found in the parameters mentioned above in group D (P ＞ 0.05).Conclusion CaMK Ⅱ is involved in the development and maintenance of chronic NP by up-regulating the expression of Cav3.2 T-type calcium channels in rat spinal cord.

Objective To evaluate the role of calcium/calmodulin-dependent protein kinase Ⅱ (CaMK Ⅱ) in the neuronal damage induced by lidocaine.Methods SH-SY5Y cells were seeded in 96-well plates (100 μl/hole) with a density of 5 × 105/ml and randomly divided into 4 groups (n =63 each):normal culture group (C group),CaMK Ⅱ inhibitor KN93 (K group),lidocaine group (L group) and KN93 + lidocaine group (KL group).KN93 (final concentration 1 μmol/L) was added to the culture medium and the cells were then cultured for 24 h in group K.Lidocaine (final concentration 10 mmol/L) was added to the culture medium and the cells were then cultured for 24 h in group L.KN93 (final concentration 1 μmol/L) and lidocaine (final concentration 10 mmol/L) were added to the culture medium and the cells were then cultured for 24 h in group KL.The cell morphology was examined with microscope after 24 h of incubation.The viability of cells was measured by MTT assay before incubation and at 1,6,12 and 24 h of incubation.The apoptosis in the cells was assessed by flow cytometry.The apoptotic rate was calculated.Results Compared with C and K groups,the cell viability was significantly decreased and the apoptotic rate was increased in L and KL groups (P ＜ 0.05).The cell viability was significantly higher and the apoptotic rate was lower in group KL than in group L (P ＜ 0.05).There was no significant difference in the cell viability and apoptotic rate between C group and K group (P ＞ 0.05).The pathological changes were obviousin group L and significantly reduced in group KL.Conclusion CaMK Ⅱ is involved in the neuronal damage induced by lidocaine.

Objective To investigate the effects of dexmedetomidine on perioperative cellular immune function and micro-metastasis in blood circulation in patients undergoing radical operation for colon cancer.Methods Sixty ASA Ⅰ or Ⅱ patients,aged 38-69 yr,weighing 45-67 kg,undergoing radical operation for colon cancer,were randomly divided into 2 groups (n =30 each) ∶ dexmedetomidine group (D group) and control group (C group).Anesthesia was induced with midazolam,cisatracurium and sufentanil,and maintained with propofol,remifentanil and sevoflurane.After tracheal intubation,a loading dose of dexmedetomidine 1 μg/kg was injected intravenously,followed by infusion at 0.5μg· kg-1 · h-1 until the end of operation in D group.The equal volume of normal saline was administered in C group.Venous blood samples were obtained at 5 min before induction of anesthesia (T0),1 h after beginning of operation (T1),the end of operation (T2) and 24 h after the end of operation (T3) for determination of the levels of T lymphocyte subsets (CD3+,CD4+,CD8+) and NK cells by flow cytometry.CD4+/CD8+ ratio was calculated.Cytokeratin 20 (CK20) expression in circulation was detected by RT-PCR at T0 and T3 and the positive rate was calculated.Results Compared with the baseline value at T0,the levels of CD3+ and CD4+,CD4+/CD8+ ratio and level of NK cells were significantly decreased at T2 and T3 in group C,and the levels of CD3+ and CD4+ were significantly decreased at T2 and T3 in group D (P ＜ 0.05).The levels of CD3+,CD4+ and NK cells at T2 and T3 were significantly higher and positive rate at T3 was significantly lower in group D than in group C (P ＜ 0.05).Conclusion Dexmedetomidine can improve the cellular immune function and decrease the probability of micro-metastasis in blood circulation in patients undergoing radical operation for colon cancer.

Objective To observe the expression of calcium/calmodulin dependent protein kinase Ⅱ (CaMK Ⅱ) in the spinal cord of the rats followed lidocaine hydrochloride intrathecal injection.Methods 48 male SD rats weight(230 ± 20) g,after intrathecal indwelling catheter,were randomly divided into four groups (n =12,8 rats for behavioral detection and 4 rats for western blotting):normal group (C group),sham group (S group),DMSO group (D group),10％ lidocaine group (L group).Mechanical withdrawal threshold(MWT) and thermal withdrawal latency(TWL) were detected before and after 2 h,4 h,8 h,12 h,1 d,2 d,3 d,4 d and 5 d with drug treatment.Intumescentia lumbalis of the spinal cord were collected to measure the expression of CaMK Ⅱ with western blotting after drug treatment for 12 h.Results The based MWT of the rats in C,S and D group were (11.2 ± 3.1) g,(11.8 ± 2.2) g and (11.4 ± 2.4) g respectively.There were no differences among the every time points (n=8,P＞0.05).The MWT of the rats in L group significantly increased at 2 h,4 h,8 h,12 h,1 d,2 d,3 d and 4 d after treatment with lidocaine hydrochloride,and the data were (22.0 ± 6.6) g,(22.2 ± 5.3) g,(20.5 ±5.8)g,(18.5 ±4.3)g,(16.7 ±3.2)g,(15.2 ±3.1)g,(15.5 ±3.5)g,(13.7 ±2.4)g respectively (n=8,P＜0.01).TWL had no difference among the rats in C,S,and D group(n=8,P＞0.05).The TWL of the rats in L group significantly increased at 2 h,4 h,8 h,12 h,1 d,2 d and 3 d after treatment with lidocaine hydrochloride(n =8,P＜ 0.01).The expression of CaMK Ⅱ of the rats in C group,S group,D group and L group were 0.17 ± 0.03,0.16 ± 0.03,0.19 ± 0.05,0.42 ± 0.11,and significantly upregulated in L goup (n =4,P ＜ 0.01).Conclusion Lidocaine hydrochloride intrathecal injection can increase the expression of the CaMK Ⅱ in the spinal cord of the rats.Those indicate that CaMK Ⅱ may be involved with the nerve damage induced by lidocaine hydrochloride.