柴芩承气汤对急性胰腺炎大鼠腺泡细胞外分泌功能的影响

目的研究柴芩承气汤(CQCQ-D)对急性胰腺炎(AP)大鼠胰腺腺泡细胞外分泌功能和腺泡细胞内钙离子浓度([Ca2 ]i)变化的影响。方法SD大鼠灌喂CQCQ-D以制备柴芩承气汤含药血清(CQCQ-S);SD大鼠分为AP组和假手术组,分离胰腺腺泡细胞并与CQCQ-S共同孵育,观测腺泡细胞淀粉酶分泌水平和[Ca2 ]i变化。结果AP大鼠腺泡细胞淀粉酶分泌水平较假手术组降低(P<0.01),CQCQ-S使AP大鼠腺泡细胞淀粉酶分泌水平进一步降低(P<0.05);[Ca2 ]i随AP病程延长而升高(P<0.05),CQCQ-S可抑制AP大鼠腺泡细胞内[Ca2 ]i升高(P<0.05)。结论CQCQ-D对AP大鼠的胰腺腺泡细胞的外分泌功能和腺泡细胞内钙超载有抑制作用。     

质子泵抑制剂对离体胰腺腺泡淀粉酶分泌的影响

目的探讨质子泵抑制剂(PPI)对胰腺外分泌是否有直接影响。方法首先,参照Williams方法制备离体胰腺腺泡悬液,加入蛙皮素(10-10M)刺激。①加入不同浓度的奥美拉唑(1×10-6M,2×10-6M,4×10-6M,8×10-6M)(10-8M~10-5M),取上清液以碘-淀粉比色法测量淀粉酶量;②将相同浓度的奥美拉唑(10-6M)加入蛙皮素刺激后的胰腺腺泡中作用30,60,90,120,150min后,测定其胰腺腺泡分泌率;③加入不同浓度善宁(10-9M~10-6M),取上清液测定其淀粉酶分泌程度。结果不同浓度善宁作用后的淀粉酶分泌率明显降低,有显著性差异(P<0.05);不同浓度奥美拉唑作用后的淀粉酶分泌率间无显著性差异(P>0.05);奥美拉唑(10-6M)作用30,60,90,120,150min后胰腺腺泡分泌率无显著性差异(P>0.05)。结论奥美拉唑对胰腺腺泡外分泌功能无直接抑制作用。     

CCK-8对LPS诱导大鼠TASMCsCuZn—SOD基因表达影响的受体机制

目的 探讨八肽胆囊收缩素(CCK-8)对体外脂多糖(LPS)诱导大鼠胸主动脉平滑肌细胞(TASMCs) CuZn-SOD基因表达影响的受体机制.方法 用贴块法培养大鼠TASMCs,经CCK-8、LPS、CR-1409、CR-2945、CCK+LPS、CR-1409+LPS、CR-2945+LPS、CR-1409+CCK-8+LPS、CR-2945+CCK-8+LPS及溶剂单独或共同孵育细胞,用RT-PCR技术检测CuZn-SOD mRNA表达.结果 对照组TASMCs CuZn-SOD mRNA低水平表达,LPS 诱导TASMCs CuZn-SOD mRNA表达上调(P<0.05),CCK-8预处理后,LPS对TASMCs CuZn-SOD mRNA表达的诱导作用可部分受抑,预先用CR-1409(1×10-5 mol/L)、CR-2945(1×10-5 mol/L)处理后,再分别加入LPS和CCK-8,CR-1409+CCK-8+LPS、CR-2945+CCK-8+LPS两组CuZn- SOD mRNA表达均高于CCK-8+LPS组(P<0.01),但仍低于LPS 组(P<0.05),其中CR-2945的拮抗作用比CR-1409稍明显; CR-1409、2945单独作用与对照组相比差异无显著性(P>0.05).结论 CCK-8通过其受体下调LPS 对TASMCs CuZn-SOD基因表达的诱导作用,其中CCK-BR较CCK-AR作用稍大.     

亚低温对雨蛙肽致大鼠胰腺腺泡细胞损伤的保护作用

目的观察亚低温对雨蛙肽刺激大鼠胰腺腺泡细胞的保护作用,并探讨其机制。方法取大鼠胰腺组织块,分为常温对照组、亚低温对照组、常温雨蛙肽组和亚低温雨蛙肽组。分别测定不同时间点(15、30、60、90min)上清液中淀粉酶、脂肪酶和组织匀浆液中胰蛋白酶活性、乳酸脱氢酶(LDH)漏出率及胰腺腺泡细胞内钙离子([Ca~(2 )]i)浓度,并测定1、3、5 h的胰腺细胞存活率。结果与常温雨蛙肽组比较,30、60和90 min时亚低温雨蛙肽组的淀粉酶和脂肪酶均显著降低(P值分别<0.05、0.01),15和30 min时胰蛋白酶活性显著降低(P值分别<0.05、0.01),60和90 min时LDH漏出率显著降低(P值分别<0.05、0.01),30、60和90 min时胰腺腺泡细胞内[Ca~(2 )]i浓度显著降低(P值分别<0.05、0.01),3和5 h时胰腺细胞存活率显著升高(P值均<0.05)。结论亚低温对雨蛙肽刺激的胰腺腺泡细胞具有保护作用,其机制可能与抑制胰蛋白酶原激活、减轻细胞内钙超载有关。     

高压氧对急性坏死性胰腺炎胰腺腺泡细胞NF-κB活性与血液炎症细胞因子影响的研究

目的探讨高压氧对ANP胰腺腺泡细胞核转录因子NF-κB活性的影响。方法采用5%牛磺胆酸钠胰管逆行注射制备ANP动物模型。SD雄性大鼠40只,按随机分配原则分为ANP高压氧治疗组和ANP空白对照组。采用EMSA法检测胰腺腺泡细胞胞核NF-κB活性、Western-blot法检测胰腺腺泡细胞胞质IκBa活性及ELISA法检测血液IL-2、IL-6、IL-10、TNF-a及ICAM-1含量。结果高压氧在1、3、5及7h均可显著抑制ANP胰腺腺泡细胞胞核NF-κB活性[(24.47±5.39)μmol/Lvs(31.36±5.72)μmol/L、(28.58±4.51)μmol/Lvs(39.44±6.31)μmol/L、(26.47±4.13)μmol/Lvs(33.80±5.96)μmol/L及(20.38±3.79)μmol/Lvs(25.69±4.91)μmol/L](P<0.01);显著增强ANP胰腺腺泡细胞胞质IκBa活性[(8.94±1.43)μmol/Lvs(6.37±1.19)μmol/L、(7.83±1.72)μmol/Lvs(5.91±1.65)μmol/L、(8.31±1.54)μmol/Lvs(6.85±1.37)μmol/L及(8.62±1.12)μmol/Lvs(6.97±0.86)μmol/L](P<0.01);显著抑制血液中炎症细胞因子IL-2、IL-6、IL-10、TNF-a及ICAM-1活性(P<0.05)。结论高压氧可通过显著抑制ANP胰腺腺泡细胞NF-κB活性,增加胰腺腺泡细胞胞质IκBa活性的能力,减少血液中炎症细胞因子活性,从而抑制ANP引起的过度炎症反应。     

银杏黄酮苷元通过调控DUSP1表达对急性胰腺炎大鼠腺泡细胞炎症因子分泌和凋亡的影响

目的:探讨银杏黄酮苷元(GA)对雨蛙素诱导的急性胰腺炎(AP)大鼠腺泡细胞炎症因子分泌和凋亡的影响.方法:采用雨蛙素诱导大鼠胰腺腺泡细胞株AR42J建立AP腺泡细胞模型.①正常培养的AR42J细胞作为空白对照组,AP模型细胞为AP组,以用15、30、60 mg/L GA处理的AP模型细胞作为GA低、中、高浓度组.②将p...     

硫化氢对离体大鼠胰腺腺泡细胞炎症因子表达的影响

目的 以雨蛙素刺激的大鼠胰腺腺泡细胞为体外急性胰腺炎(acute pancreatitis,AP)模型,研究硫化氢(H2S)对AP时胰腺腺泡细胞炎症因子表达的影响及机制.方法 分离25只SD大鼠胰腺腺泡细胞,用雨蛙素(10-7 mol/L)诱导体外AP模型,用不同浓度NaHS(5、10、20、40 μmol/L)及不同时间(10、30、60 min) NaHS(5μmol/L)处理,Real-time PCR检测炎症因子mRNA水平.再用CSE抑制剂PAG处理,Real-time PCR检测炎症因子mRNA水平及CSE mRNA水平.进一步用NF-κB抑制剂BAY 11-7082处理后,Real-time PCR检测炎症因子mRNA水平,Western blot检测浆内IκBα及核内NF-κBp65的蛋白表达.结果 当5μmol/L NaHS预处理60 min时,TNF-α水平(14.70±1.10)、IL-1β水平(14.42±0.83)显著低于雨蛙素组(P＜0.05),IL-10水平(12.20 ±0.55)显著高于雨蛙素组(P＜0.05);CSE抑制剂PAG预处理60 min后,TNF-α水平(37.38 ±0.89)、IL-1β水平(24.45 ±0.76)显著高于雨蛙素组(P＜0.05),CSE水平(0.14±0.03)、IL-10水平(0.14 ±0.03)显著低于雨蛙素组(P＜0.05);NF-κB抑制剂BAY 11-7082预处理30 min,TNF-α水平(9.17±0.96)、IL-1β水平(2.90 ±0.17)显著低于雨蛙素组(P＜0.05),IL-10水平(40.00±1.00)显著高于雨蛙素组(P＜0.05),并且胞浆内IκBα(0.90 ±0.01)的降解及细胞核内NF-κBp65 (0.70 ±0.04)的蛋白表达明显低于雨蛙素组(P＜0.05).结论 小剂量H2S供体NaHS在雨蛙素孵育的胰腺腺泡细胞中具有一定抗炎作用,且该作用是通过抑制NF-κB通路从而抑制腺泡细胞表达促炎因子实现的.     

外源性八肽胆囊收缩素对吗啡所致SH-SY5Y和PC12细胞毒的保护作用

目的 观察外源性八肽胆囊收缩素(cholecystokinin octapeptide,CCK-8)对吗啡处理SH-SY5Y及PC12细胞毒性作用的影响.方法 应用二甲基噻唑二苯基四噻唑盐[3-(4,5)-dimethylthiazo (-2-yl)-3,5-di-phenytetrazoliumromide,MTT]法观察吗啡对SH-SY5Y及PC12细胞毒性作用的剂量和时间依赖关系,并且观测外源性CCK-8对吗啡细胞毒性作用的影响.结果 吗啡在100~10 000μmol/L浓度范围内显著降低SH-SY5Y和PC12细胞的生存率,半数抑制率分别为2 520μmol/L和680μmol/L;上述作用在孵育48h明显,随时间延长逐渐增强;CCK-8(10-6和10-8mol/L)可明显抑制3 000μmol/L吗啡作用48h后SH-SY5Y细胞生存率的下降,CCK-8(10-6、10-8和10-10mol/L)可明显抑制1 000μmol/L吗啡作用48h后PC12细胞生存率的下降;3 000μmol/L和1 000μmol/L 吗啡分别处理SH-SY5Y细胞和PC12细胞48h后,可见细胞胞体变圆,突触变短或消失,失去原有的细胞形态.CCK-8(10-6和10-8mol/L)可明显改善吗啡作用后SH-SY5Y和PC12细胞的形态改变.结论 外源性CCK-8可浓度依赖地对抗吗啡产生的细胞毒性作用.     

CRH对腹腔巨噬细胞分泌IL-10的影响及其信号机制

目的 研究促肾上腺皮质激素释放激素(corticotrophin-releasing hormone,CRH)对大鼠腹腔巨噬细胞分泌白介素-10(interleukin-10,IL-10)的影响及其信号转导机制.方法 将原代培养的大鼠腹腔巨噬细胞分为正常对照组、CRH组、CP+CRH组、HL+CRH组、H89+CRH组以及PKAi+CRH组.其中正常对照组仅加入新鲜RPMI1640培养液2 ml;CRH处理组,加入含CRH终浓度为10~(-9) mol/L的培养液2 ml;后4组在分别用CRHRI选择性阻断剂CP-154526(5×10~(-4) mol/L)、CRHR1/2非选择性阻断剂α-helical CRH9-41(10~(-6)mol/L)、Ac抑制剂H89(10~(-6)mol/L)和PKA抑制剂PKAi(6-22)amide(10~(-4)mol/L)预孵育2 h后,加入含终浓度为10~(-9)mol/L的CRH培养液,再分别加入CRH(终浓度为10~(-9)mol/L)刺激.各组分别于CRH刺激后0.5、1、2、4、8 h收集培养皿中的上清液,应用ELISA方法 检测IL-10的含量.结果 正常对照组巨噬细胞分泌IL-10的含量为(29.41±3.10)μg/g.CRH组IL-10含量呈现双峰,在1、4 h达到高峰,分别为(53.15±7.98)、(46.01±1.70)μg/g,与CRH组0.5 h相比差异均非常显著(P<0.01),接近正常对照组的2倍(P<0.01).CP+CRH组与HL+CRH组IL-10含量在1、4 h均明显低于CRH组(P<0.01),并接近正常对照组水平.但在8 h时明显增加,显著高于正常对照组(P<0.05)和CRH组(P<0.05).且两组之间IL-10含量在0.5～4 h差异不显著,但在8 h时HL+CRH组显著低于CP+CRH组.H89预孵育后细胞IL-10含量在1、4、8 h显著下降,与CRH组比较差异显著(P<0.01);在4、8 h低于正常对照组水平(P<0.05).PKAi(6-22)amide预孵育细胞后,IL-10含量在0.5～8 h下降,与正常对照组以及CRH组比较差异非常显著(P<0.01).结论 CRH可通过CRHRI促进巨噬细胞分泌产生抗炎因子IL-10,其与cAMP/PKA信号通路密切相关.     

白细胞介素-1受体拮抗剂在急性胰腺炎中的作用及机制探讨

目的从胰腺腺泡细胞凋亡的角度探讨白细胞介素-1受体拮抗剂(IL-1Ra)在急性胰腺炎中的作用及机制。方法用牛磺胆酸钠建立大鼠急性坏死性胰腺炎模型,IL-1Ra腹腔注射诱导胰腺腺泡细胞凋亡,并应用细胞凋亡原位标记检测(TUNEL)染色、免疫组化等技术检测胰腺细胞凋亡及Fas/FasL的蛋白表达。结果干预组1,2,6,12 h胰腺细胞凋亡指数显著高于胰腺炎组相应时间点(均P<0.01);对照组见较弱的Fas的表达,干预组各时间点胰腺细胞Fas染色阳性率明显高于胰腺炎组(均P<0.01);胰腺炎组、干预组中均没有见到胰腺腺泡细胞中FasL表达,但是在间质中均可见浸润的炎性细胞FasL免疫组化染色阳性。结论IL-1Ra减轻急性胰腺炎的机制可能不仅限于对炎症反应的抑制,它可能是通过上调凋亡调控基因Fas/FasL系统的表达,诱导胰腺腺泡细胞的凋亡从而减轻胰腺炎的严重程度。     

Relationship between carbachol hyperstimulation-induced pancreatic acinar cellular injury and trypsinogen or NF-kappaB activation in rats in vitro.

The relationship between M3 cholinergic receptor agonist (carbachol) hyperstimulation-induced pancreatic acinar cellular injury and trypsinogen activation or NF-kappaB activation in rats was studied in vitro. Rat pancreatic acinar cells were isolated, cultured and treated with carbachol, the active protease inhibitor (pefabloc), and NF-kappaB inhibitor (PDTC) in vitro. Intracellular trypsin activity was measured by using a fluorogenic substrate. The cellular injury was evaluated by measuring the leakage of LDH from pancreatic acinar cells. The results showed that as compared with control group, 10(-3) mol/L carbachol induced a significant increase of the intracellular trypsin activity and the leakage of LDH from pancreatic acinar cells. Pretreatment with 2 mmol/L pefabloc could significantly decrease the activity of trypsin and the leakage of LDH from pancreatic acinar cells (P < 0.01) following the treatment with a high concentration of carbachol (10(-3) mol/L) in vitro. The addition of 10(-2) mol/L PDTC didn't result in a significant decrease in the activity of trypsin and the leakage of LDH from pancreatic acinar cells treated with a high concentration of carbachol (10(-3) mol/L) in vitro (P > 0.05). It was concluded that intracellular trypsinogen activation is likely involved in pancreatic acinar cellular injury induced by carbachol hyperstimulation in vitro. NF-kappaB activation may not be involved in pancreatic acinar cellular injury induced by carbachol hyperstimulation in vitro.     

Relationship between Carbachol Hyperstimulation-Induced Pancreatic Acinar Cellular Injury and Trypsinogen or NF-κB Activation in Rats in vitro

The relationship between M3 cholinergic receptor agonist (carbachol) hyperstimulationinduced pancreatic acinar cellular injury and trypsinogen activation or NF-κB activation in rats was studied in vitro. Rat pancreatic acinar cells were isolated, cultured and treated with carbachol, the active protease inhibitor (pefabloc), and NF-κB inhibitor (PDTC) in vitro. Intracellular trypsin activity was measured by using a fluorogenic substrate. The cellular injury was evaluated by measuring the leakage of LDH from pancreatic acinar cells. The results showed that as compared with control group, 10-3 mol/L carbachol induced a significant increase of the intracellular trypsin activity and the leakage of LDH from pancreatic acinar cells. Pretreatment with 2 mmol/L pefabloc could significantly decrease the activity of trypsin and the leakage of LDH from pancreatic acinar cells (P ＜0.01) following the treatment with a high concentration of carbachol (10-3 mol/L) in vitro. The addition of 10-2 mol/L PDTC didn't result in a significant decrease in the activity of trypsin and the leakage of LDH from pancreatic acinar cells treated with a high concentration of carbachol (10-3 mol/L) in vitro (P＞0.05). It was concluded that intracellular trypsinogen activation is likely involved in pancreatic acinar cellular injury induced by carbachol hyperstimulation in vitro. NF-κB activation may not be involved in pancreatic acinar cellular injury induced by carbachol hyperstimulation in vitro.     

Relationship between Carbachol Hyperstimulation-induced Pancreatic Intracelluar Trypsinogen and NF-кB Activation in Rats in vitro

The relationship between intracelluar trypsinogen activation and NF-r,B activation in rat pancreatic acinar cells induced by M3 cholinergic receptor agonist (carbachoi) hyperstimulation was studied. Rat pancreatic acinar cells were isolated, cultured and treated with carbachol, the active pro- tease inhibitor (pefabloc) and NF-кB inhibitor (PDTC) in vitro. Intracelluar trypsin activity was measured by using a fluorogenie substrate. The activity of NF-кB was monitored by using electro- phoretic mobility shift assay. The results showed that after pretreatment with 2 mmol/L pefabloc, the activities of trypsin and NF-кB in pancreatic acinar cells treated with high concertrations of carbachol (10-3 mol/L) in vitro was significantly decreased as compared with control group (P＜0.01). The addi- tion of 10-2mol/L PDTC resulted in a significant decrease of NF-кB activities in pancreatic acinar cells after treated with high concertrations of carbachol (10-3 mol/L) in vitro, but the intracelluar trypsinogen activity was not obviously inhibited (P＞0.05). It was concluded that intracelluar trypsi- nogen activation is likely involved in the regulation of high concertrations of carbachol-induced NF-кB activation in pancreatic acinar cells in vitro. NF-кB activation is likely not necessary for high concertrations of carbachol-induced trypsinogen activation in pancreatic acinar cells in vitro.     

Relationship between carbachol hyperstimulation-induced pancreatic acinar cellular injury and trypsinogen or NF-κB activation in ratsin vitro

Summary The relationship between M3 cholinergic receptor agonist (carbachol) hyperstimulation-induced pancreatic acinar cellular injury and trypsinogen activation or NF-κB activation in rats was studiedin vitro. Rat pancreatic acinar cells were isolated, cultured and treated with carbachol, the active protease inhibitor (pefabloc), and NF-κB inhibitor (PDTC)in vitro. Intracellular trypsin activity was measured by using a fluorogenic substrate. The cellular injury was evaluated by measuring the leakage of LDH from pancreatic acinar cells. The results showed that as compared with control group, 10⁻³ mol/L carbachol induced a significant increase of the intracellular trypsin activity and the leakage of LDH from pancreatic acinar cells. Pretreatment with 2 mmol/L pefabloc could significantly decrease the activity of trypsin and the leakage of LDH from pancreatic acinar cells (P<0.01) following the treatment with a high concentration of carbachol (10⁻³ mol/L)in vitro. The addition of 10⁻² mol/L PDTC didn't result in a significant decrease in the activity of trypsin and the leakage of LDH from pancreatic acinar cells treated with a high concentration of carbachol (10⁻³ mol/L)in vitro (P>0.05). It was concluded that intracellular trypsinogen activation is likely involved in pancreatic acinar cellular injury induced by carbachol hyperstimulationin vitro. NF-κB activation may not be involved in pancreatic acinar cellular injury induced by carbachol hyperstimulationin vitro. Zheng Hai, male, born in 1972, Doctor in Charge, M. D., Ph.D.     

胆囊收缩素对豚鼠结肠平滑肌及其细胞膜L-型钙电流和膜电位的影响

目的 探讨胆囊收缩素8肽(CCK-8S)对豚鼠近端结肠平滑肌及细胞膜L-型钙电流和膜电位的电生理作用及其机制.方法 (1)采用RM6240多道生理信号采集处理系统记录CCK-8S对豚鼠近端结肠平滑肌肌条收缩的影响;(2)检测Fluo-2/AM标记的近端结肠平滑肌细胞膜内钙离子浓度([Ca2+]i)变化;(3)用EPC-10膜片钳放大器测量全细胞模式下CCK-8S对近端结肠平滑肌细胞膜静息电位(RP)、动作电位(AP)和L-型钙通道电流(ICa-L)的影响.结果 CCK-8S(10-1mol/L)作用后:(1)豚鼠近端结肠平滑肌肌条收缩幅值和频率分别为对照组的(149±12)%和(132±13)%(均P＜0.05);(2)近端结肠平滑肌细胞膜的[Ca2+]i为对照组的(738±24)%(P＜0.05);(3)RP、AP峰值及AP快速复极时间分别为对照组的(52±9)%、(140±4)%和(61±13)%(均P＜0.05),该效应可被预先加入的CCK1受体拮抗剂地伐西匹和(或)L-型钙通道阻断剂尼非地平阻断(n=8,均P＜0.05),而预先向灌流液中加入CCK2受体拮抗剂CI 988后,CCK-8S对RP、AP峰值及AP快速复极时间的作用仍存在;(4)从-60 mV去极化至+10 mV,ICa-L为对照组的(138±7)%(P＜0.05),但分别可被相应拮抗剂抑制;当向电极内液加入肝素(10-6 mol/L)或向细胞外液加入staurosporine(10-6 mol/L)后,CCK-8S对Ica-L均没有影响(2.9%±2.6%和1.9%±2.3%,均P＞0.05).结论 CCK-8S通过CCK1受体促进近端结肠平滑肌自发性收缩频率和强度;其机制与激活1,4,5-三磷酸肌醇介导的蛋白激酶C途径进而促进L-型钙通道开放有关.

Abstract：

Objective To investigate the effects and mechanism of cholecystokinin octapeptide (CCK-8S) on the contractile activity of smooth muscles,L-type calcium current and membrane potentials of proximal colon myocytes in guinea pig.Methods ( 1 ) Strips of proximal colon were obtained from adult guinea pigs.The contraction of these stripes was measured by a RM6240 multi-channel physiological signal system.(2) Suspension of single smooth muscle cells (SMCs) were obtained from proximal colon and isolated by enzymatic digestion.The effect of CCK-8S on intracellular calcium concentration ( [Ca2+] i) of SMCs was examined by fura-2-1oaded miscrofluorimetric measurement.(3) Resting potential ( RP),action potential (AP) and L-type calcium current (ICa-L ) were recorded by patch-clamp technique.Results ( 1 )The contractile amplitude and frequency of muscle stripes enhanced by CCK-8S ( 10 -7 mol/L) were ( 149 ±12)% and (132 ± 13 )% respectively of those of control group (all P ＜ 0.05 ).They were significantly attenuated by pretreating strips with CCK1 receptor antagonist devazepide ( 10-7 mol/L),L-type calcium channel blocker nifedipine ( 10 -5 mol/L),Ca2+ -ATPase inhibitor TG (thapsigargin) ( 10-5 mol/L) and BA (boric acid) (10-5 mol/L) respectively.(2) [Ca2+]i of SMCs intensified by CCK-8S was (738 ±24)% of that of control group.And it was inhibited by pretreating SMCs with devazepide(all P ＜0.05).(3) After the superfusion of CCK-8S,RP depolarized to (52 ±9)%,the exogenously stimulated peak values of AP rose to (140±4)% and fast repolarization time of AP decreased to (61 ± 13)% (all P ＜0.05).They were significantly inhibited when these cells were pretreated with devazepide and/or nifedipine (n = 8,P ＜0.05 for each group) whereas CI 988 had little effect.(4) The CCK-8S-evoked ICa-L of SMCs at the voltage of + 10 mV was boosted to ( 138 ± 7 )%.Such an effect was suppressed by a pretreatment with nifedipine,devazepide,TG and BA respectively.In the presence of an inhibitor of inositol 1 ,4,5-trisphosphate (IP3 ) receptors,heparin (10-6 mol/L) and an protein kinase C (PKC) inhibitor,saurosporine (10-6 mol/L),CCK-8S did not significantly intensify ICa-L (all P ＞ 0.05 ).Conclusion CCK-8S promotes proximal colon contraction by CCK1 receptors through the activation of IP3-mediated PKC pathway.     

阿托伐他汀对骨髓基质细胞增殖及向成骨细胞分化的影响

目的比较不同浓度的阿托伐他汀对小鼠骨髓基质细胞向成骨细胞分化、矿化成骨的影响。方法分别将10^-8、10^-7、10^-6mol／L的阿托伐他汀和空白药物加入传代培养的小鼠骨髓基质细胞，通过细胞形态学观察、细胞增殖率检测、碱性磷酸酶（ALP）检测和钙结节染色，比较药物在成骨细胞分化过程中对细胞增殖和分化的影响。结果阿托伐他汀可以剂量依赖性方式抑制骨髓基质细胞的增殖，10^-7mol／L组和10^-6mol／L组较明显（均P〈0．05）。阿托伐他汀可以提高骨髓基质细胞ALP活性，培养10～22d，10^-7mol／L组和10^-6mol／L组ALP活性高于对照组和10^-8mol／L组（均P〈0．05）。矿化成骨染色显示在培养26d后，10^-6mol／L组染色明显深于其它各组。结论阿托伐他汀可促进骨髓基质细胞来源的成骨细胞的分化，抑制了细胞增殖，促进骨结节钙化。     

尼尔雌醇和左炔诺孕酮对人成骨肉瘤 MG-63细胞株雌激素受体亚型的作用

目的：观察不同浓度的尼尔雌醇(NYL)和左炔诺孕酮(LNG)对人成骨肉瘤MG-63细胞株ERα和ERβ表达的作用，探讨旁分泌效应对ERα和ERβ基因表达的影响。方法：人成骨肉瘤MG-63细胞株分为（1）空白对照组：用含1% BSA无血清MEM培养液培养48 h；（2）阳性对照组：培养液中加入10-8 mol/L的17β-estradiol (E2) 培养48 h；（3）药物处理组：培养液中分别加入10-10，10-8，10-6 mol/L的NYL或LNG培养48 h；（4）药物处理+换液组：培养液中分别加入10-10 mol/L的NYL或10-8 mol/L LNG培养48 h，每12 h更换新的含有相应药物的培养液。用半定量RT-PCR检测各组细胞ERα和ERβmRNA的表达。结果：NYL及LNG均能诱导ERα和ERβ亚型mRNA表达上调，诱导ERαmRNA表达的最强作用浓度均为10-6 mol/L。NYL，LNG对ERβ mRNA表达最强作用浓度分别为10-10，10-8 mol/L。每12 h更换干预培养液对ERα表达无影响，但可抑制ERβ的表达。结论：NYL和LNG均可诱导MG-63细胞株ERα和ERβ mRNA表达上调，且2种亚型的表达存在相互制约关系；在NYL和LNG诱导ER亚型表达上调过程中ERβ的表达可能受到旁分泌作用的影响。     

Relationship between Carbachol Hyperstimulation-induced Pancreatic Intracelluar Trypsinogen and NF-κB Activation in Rats in vitro

The relationship between intracelluar trypsinogen activation and NF-κB activation in rat pancreatic acinar cells induced by M3 cholinergic receptor agonist （carbachol） hyperstimulation was studied. Rat pancreatic acinar cells were isolated, cultured and treated with carbachol, the active protease inhibitor （pefabloc） and NF-κB inhibitor （PDTC） in vitro. Intracelluar trypsin activity was measured by using a fluorogenic substrate. The activity of NF-κB was monitored by using electrophoretic mobility shift assay. The results showed that after pretreatment with 2 mmol/L pefabloc, the activities of trypsin and NF-κB in pancreatic acinar cells treated with high concertrations of carbachol （10^-3 mol/L） in vitro was significantly decreased as compared with control group （P〈0.01 ）. The addition of 10^-2 mol/L PDTC resulted in a significant decrease of NF-κB activities in pancreatic acinar cells after treated with high concertrations of carbachol （10^-3 mol/L） in vitro, but the intracelluar trypsinogen activity was not obviously inhibited （P〉0.05）. It was concluded that intracelluar trypsinogen activation is likely involved in the regulation of high concertrations of carbachol-induced NF-κB activation in pancreatic acinar cells in vitro. NF-κB activation is likely not necessary for high concertrations of carbachol-induced trypsinogen activation in pancreatic acinar cells in vitro.     

辛伐他汀对大鼠磨牙牙根根端复合体细胞矿化的影响

目的研究辛伐他汀对大鼠磨牙牙根根端复合体（DAC）细胞矿化作用的影响。方法将3代大鼠磨牙牙根根端复合体细胞在条件矿化液中诱导培养,同时加入四种不同浓度的辛伐他汀（1×10^-9、1×10^-8、1×10^-7、1×10^-6mol/L）,检测DAC组织碱性磷酸酶（ALP）活性的表达并对形成的矿化结节进行计数。结果四种不同浓度的辛伐他汀均能促进大鼠磨牙牙根根端复合体细胞的矿化,其中1×10^-8、1×10^-7、1×10^-6mol/L与对照组比较,差异有统计学意义（P〈0.01）,1×10^-7mol/L的辛伐他汀组促进细胞矿化作用的影响最明显。结论适宜浓度的辛伐他汀可有效促进大鼠磨牙牙根根端复合体细胞的矿化。     

塞来昔布对乳腺癌阿霉素耐药细胞株MCF-7/ADM的增殖抑制作用

目的： 通过研究选择性环氯化酶2(COX-2)抑制剂塞来昔布联合阿霉素(ADM)对乳腺癌MCF-7/ADM细胞株生长的作用,探讨塞来昔布的抗肿瘤作用。方法：MCF-7/ADM细胞加入不同浓度等倍稀释的ADM（0.05、0.10、0.20、0.40、0.80和1.60 mg/L）为对照组,MCF-7/ADM细胞加入无细胞的完全培养基为阴性对照组,在对照组的基础上联合应用10、20 μmol/L塞来昔布为实验组,各组细胞于24、48、72 h后采用CCK-8检测细胞生长抑制率。MCF-7/ADM细胞单加0.05 mg?L-1ADM及联合塞来昔布(10、20 μmol/L)处理后3 h收集细胞,采用流式细胞术检测细胞内ADM水平。结果：不同浓度ADM单药及联合塞来昔布（10和20 μmol/L）　作用于MCF-7/ADM 24、48和72 h后细胞生长受到抑制,实验组不同时间细胞生长抑制率均高于对照组(P<0.05),且20 μmol/L塞来昔布实验组细胞生长抑制率高于10 μmol?L-1塞来昔布实验组,差异具有统计学意义（P<0.05）。对照组24、48和72 h的细胞未见明显变化,但实验组细胞作用48和72 h后出现细胞改变及死亡,呈塞来昔布剂量依赖性。与对照组比较,塞来昔布实验组（10和20 μmol/L）细胞内ADM浓度升高（P<0.05）;20 μmol/L塞来昔布实验组细胞内ADM浓度高于 10 μmol/L塞来昔布实验组(P<0.05)。结论：选择性COX-2抑制剂塞来昔布联合ADM可有效逆转乳腺癌ADM细胞株耐药。
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