蛋白激酶Cβ抑制剂Enzastaurin逆转肿瘤多药耐药机制的研究

目的探讨蛋白激酶Cβ抑制剂Enzastaurin逆转多药耐药(MDR)的作用及机制。方法 Enzastaurin处理高表达ABCB1、ABCC1及ABCG2的MDR基因细胞株后,通过MTT法测定细胞毒作用,流式细胞仪测定阿霉素及罗丹明123在细胞内累积,Western blotting测定蛋白表达,应用半定量RT-PCR测定m RNA表达水平及采用化学发光法测定ATP酶活性。结果 Enzastaurin可逆转ABCB1介导MDR,对ABCC1及ABCG2介导MDR无逆转作用;Enzastaurin抑制ABCB1药物外排功能而逆转肿瘤MDR,但与ABCB1表达及阻断Akt及Erk1/2磷酸化无关;Enzastaurin通过抑制ATP酶激活而抑制ABCB1药物外排泵功能。结论 Enzastaurin在体外可通过抑制药物外排泵ATP酶活性逆转ABCB1介导MDR作用。

米非司酮对离体大鼠垂体细胞促黄体激素分泌的影响及其机制

本实验应用垂体细胞体外培养模型 ,观察了米非司酮 (MP)对GnRH、高浓度细胞外K+([K+]e)和蛋白激酶C激活剂PMA诱导的LH分泌的影响。结果证实MP可以剂量和时间依赖方式抑制GnRH诱导的LH分泌 ,并可拮抗P调节GnRH诱导的LH分泌效应。同时发现 10 -7mol/LMP短时间处理 4h能抑制 6 0mmol/LKCl和 10 -8mol/LPMA诱导的LH分泌 ,而 10 -7mol/LP短时间处理则起促进作用。当处理时间延长为 5 2h时 ,P对 6 0mmol/LKCl和 10 -8mol/LPMA诱导的LH分泌无明显作用 ,P也仅对 6 0mmol/LKCl刺激的LH分泌起抑制作用 ,但不影响 10 -8mol/LPMA诱导的LH分泌。当P和MP同时处理时 ,则MP可逆转P对高 [K+]e和PMA诱导的LH分泌的调节作用 ,表明MP影响GnRH诱导的LH分泌的机制可能与MP影响电压依赖性钙离子通道和PKC的活性有关。

Breviscapine attenuates acute pancreatitis by inhibiting expression of PKCα and NF-κB in pancreas

AIM:To study the effect of breviscapine (Bre) on activity of protein kinase Cα (PKCα) and nuclear factor (NF)-κB in pancreas,and the mechanism of Bre attenuating acute pancreatitis (AP). METHODS:One hundred and eight rats were randomly divided into acute necrotizing pancreatitis (ANP) group,Bre group (ANP + Bre group) and sham operation (SO) group,36 rats in each group. ANP model was induced by a retrograde injection of 4% sodium deoxycholate into the bilio-pancreatic duct. Fifteen minutes after the ANP model was induced,the rats in Bre group were intraperitoneally injected with Bre (0.4 mg/100 g body weight or 0.1 mL/100 g body weight). Survival time and mortality of rats were calculated. Serum amylase and malondialdehyde levels were measured,volume of ascites was recorded and morphology of pancreas and lung was evaluated at 1,5 and 10 h,after the ANP model was induced,respectively. Expressions of PKCα and subunit p65 of NF-κB in pancreas were detected by immunohistochemistry and Western blotting. RESULTS:The life span of rats was longer and the mortality was lower in Bre group than in ANP group 13.51 ± 5.46 vs 25.36 ± 8.11 (P < 0.05). The amylase and MDA levels as well as the volume of ascites were lower and the pathological changes in pancreas and lung were less in Bre group than ANP group (P < 0.05),indicating that the pancreatitis is less severe in Bre group than ANP group. The activation of PKCα and NF-κB p65 in pancreas was induced rapidly and reached their peak at 1 h or 5 h after ANP,but their activity in Bre group was significantly inhibited. CONCLUSION:Bre exerts its therapeutic effect on AP by inhibiting the activation of PKCα and NF-κB p65 in pancreas.

天香丹胶囊通过调节G蛋白偶联雌激素受体保护心肌缺血再灌注损伤

目的探讨天香丹胶囊(TXD)对预处理去卵巢雌性合并心肌缺血再灌注(I/R)大鼠心肌损伤的保护作用与其可能机制。方法将60只SD雌性大鼠摘去卵巢后随机分为假手术组、模型组、阳性对照组、低、中、高3个剂量实验组,每组10只。阳性对照组灌胃0.8 mg·kg^(-1)雌二醇溶液,低、中、高剂量实验组分别灌胃给予0.2、0.4和0.8 g·kg^(-1)天香丹溶液,假手术组、模型组灌胃等量双蒸水。连续处理60 d。末次给药12 h后建立心脏缺血再灌注模型;其中假手术组只穿线不结扎,另取10正常SD雌性大鼠为正常组(不做任何处理)。用酶联免疫吸附试验法检测血清中大鼠心肌肌酸激酶同工酶(CK-MB)活性、三磷酸肌醇(IP3)的含量;用微量法检测血清中乳酸脱氢酶(LDH);用MTB微板法检测心肌组织中Ca^(2+)浓度;用苏木精-伊红染色检测大鼠心肌组织病理变化;用蛋白质印迹法检测心肌组织中G蛋白偶联雌激素受体(GPER)、磷脂酶C(PLC-β)、三磷酸肌醇受体(IP3R)以及肌浆网钙ATP酶(SERCA2)蛋白表达水平。结果正常组、假手术组、模型组、阳性对照组和高剂量实验组的血清中CK-MB活性表达分别为(64.88±4.70)、(65.86±4.40)、(161.40±10.97)、(78.31±6.64)和(79.62±7.43)pg·mL^(-1);Ca^(2+)浓度分别为(3.20±0.11)、(3.41±0.12)、(5.05±0.15)、(4.10±0.12)和(4.18±0.18)mmol·L^(-1);模型组与假手术组比较,高剂量实验组与模型组比较,差异均有统计学意义(均P<0.05)。正常组、假手术组、模型组、阳性对照组和高剂量实验组的GPER蛋白相对表达水平分别为0.74±0.06、0.73±0.04、0.36±0.03、0.67±0.06和0.65±0.05;模型组与假手术组比较,差异均有统计学意义(P<0.05或P<0.01);阳性对照组、高剂量实验组与模型组比较,差异均有统计学意义(P<0.05或P<0.01)。结论TXD对I/R大鼠心肌损伤有保护作用,可能与调节GPER/PLC/IP3R通路有关。

Angiotensin Ⅱ type 1 receptor modulation of L-type calcium currents in guinea-pig ventricular cells

Objective To study the cellular mechanism of the effect of Ang Ⅱ on ICa,L in single guinea-pig ventricular cells by using losartan and 1-(5-Isoquinolinylsulfonyl)-2-Methyl-Piperazine (H-7) as the Ang Ⅱ type 1 receptor (AT1) inhibitor and protein kinase C inhibitor, respectively.Methods Patch clamp techniques were used to study the cellular mechanism of the effect of Ang Ⅱ on ICa,L in single guinea-pig ventricular cells.Results In the whole cell patch clamp recording model, Ang Ⅱ stimulated ICa,L in a concentration dependent manner; the maximal effect was obtained at 100?nmol/L (n=9). At 30?nmol/L, Ang Ⅱ stimulated peak ICa,L from 11.3±0.6?pA/pF to 15.3±0.6?pA/pF (at +10?mV, n=9, P<0.05). 100?nmol/L Losartan, a specific AT1 receptor inhibitor, had no effect on ICa,L (n=9), but the effect of Ang Ⅱ on ICa,L was inhibited by 100?nmol/L Losartan. Ang Ⅱ on ICa,L was also inhibited by 20?μmol/L H-7, a specific protein kinase C inhibitor, whereas H-7 alone has no effect on ICa,L (n=9).Conclusion Ang Ⅱ stimulates ICa,L in guinea-pig ventricular cells by binding to AT1 through a transduction pathway involving protein kinase C.