MN9202手性对映体对心肌细胞作用的差异性及其对内毒素致心肌损伤的保护作用与机制
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摘要
目的:①建立MN9202手性固定相HPLC拆分方法,制备单一对映体,比较MN9202对映体对心肌细胞收缩、舒张功能及钙瞬变的影响,及其对离子通道的立体选择性;②探讨MN9202对心肌的保护作用及其机制。为正确评价MN9202消旋体及其对映体作用的差异性,为二氢吡啶类钙通道拮抗剂的临床应用提供科学依据。
方法:①采用Daicel OJ-H手性固定相色谱柱,通过对流动相中正己烷与乙醇的比例、添加三乙胺的量对MN9202对映体的保留时间及分离度的影响进行考察,优化色谱条件,拆分MN9202消旋体。②采用IonOptix单细胞动缘探测系统同步检测MN9202消旋体、S-(-)-MN9202和R-(+)-MN9202作用后心肌细胞收缩、舒张和钙瞬变的变化。③采用全细胞膜片钳方法,比较MN9202消旋体、S-(-)-MN9202和R-(+)-MN9202对心肌细胞L型钙通道电流密度的影响。进而探讨各药物对L型钙通道稳态激活动力学和稳态失活动力学特性的影响。④通过股静脉注射LPS建立感染性休克致动物心肌损伤模型,研究MN9202对心肌的保护作用;RM-6200型四道生理记录仪实时监测血压等参数;用ELISA法测定血浆TNF-α水平;取心肌组织样本,进行HE染色,比较心肌形态学改变。⑤原代培养新生大鼠心肌细胞,用LPS和不同浓度的MN9202消旋体、对映体及氨氯地平作用,采用ELISA法测定培养上清液中TNF-α含量,RT-PCR及免疫荧光法分别测定心肌细胞中TNF-αmRNA和蛋白的表达。⑥RT-PCR和western blot法分别测定LPS刺激不同时间及DHPs作用后心肌细胞iNOS、COX-2 mRNA和蛋白的表达。Western blot法测定磷酸化Akt和总Akt的表达。
结果:①本研究建立的MN9202 HPLC手性拆分条件为:Daicel OJ-H手性分析柱,正己烷-乙醇-三乙胺(93︰7︰0.5,V︰V︰V)为流动相,流速为0.5 ml/min,检测波长为237 nm。用此法MN9202对映异构体可达到基线分离,并可少量制备S-(-)-MN9202和R-(+)-MN9202。②不同浓度的MN9202消旋体和S-(-)-MN9202可剂量依赖性抑制大鼠单个心肌细胞的收缩:在1×10-5 mol/l浓度时,百分峰值收缩幅度(PTA %baseline)与对照组相比分别降低了73.8 %和92.7 %;而R-(+)-MN9202则呈剂量依赖性增强心肌细胞的收缩,1×10-5 mol/l时, PTA %baseline升高了18.2 %。MN9202消旋体和S-(-)-MN9202剂量依赖性抑制单个心肌细胞钙瞬变幅度(△FFI)。与对照组相比,1×10-5 mol/l MN9202消旋体和S-(-)-MN9202,细胞钙瞬变分别由0.45±0.04下降到0.07±0.02,及由0.45±0.04下降到0.05±0.01(P<0.01)。而R-(+)-MN9202则增加了钙瞬变幅度:在最大浓度1×10-5 mol/l时,细胞钙瞬变由0.45±0.01升高到0.58±0.05(P<0.01)。③MN9202消旋体剂量依赖性的降低了L型钙通道电流密度。检测电压为0 mV,MN9202浓度为0.03、0.1、0.3、1和3μmol/l时,CaL电流峰值的密度较对照组分别减少了6.7 %、15.0 %(P<0.01)、27.5 %(P<0.01)、50.7 %(P<0.01)和69.5 %(P<0.01);但药物并未改变L型钙通道激活的阈电位(-40 mV)和峰值电流电位(0 mV)。S-(-)-MN9202也抑制了L型钙通道电流密度,检测电压为0 mV时,1μmol/l S-(-)-MN9202使CaL电流峰值的密度从对照组的-9.8±1.0减少到-3.2±0.9(减少了67 %)(P<0.01)。而1μmol/l R-(+)-MN9202则引起了L型钙通道电流密度增加:检测电压为0 mV时,使CaL电流峰值的密度从对照组的-9.3±0.7增加到-10.3±0.6(P<0.01)。MN9202消旋体,S-(-)-MN9202和R-(+)-MN9202均未明显改变L型钙通道的激活特性。但各药物均对通道稳态失活有一定的影响:MN9202消旋体和S-(-)-MN9202均使稳态失活曲线左移,而R-(+)-MN9202使稳态失活曲线轻微右移。④LPS在30 min内引起大鼠平均动脉压的快速下降,从119±4降至85±2 mmHg(P<0.05);60 min后,大鼠平均动脉压仍持续下降,在240 min时为81±3 mmHg。10及30μg/kg MN9202消旋体给药后,延缓了LPS对大鼠平均动脉压的下降。但MN9202的应用并未改变LPS对心率的影响。给予LPS使血浆中TNF-α的水平发生了变化,先升高,至1小时达到峰值,而后逐渐降低。MN9202给药后可使LPS引起的大鼠血浆TNF-α升高的峰值显著降低(P<0.05)。⑤正常心肌细胞几乎不表达TNF-α,但细胞培养液中TNF-α的含量可随LPS浓度的增加而提高;TNF-α的释放与mRNA表达也随着LPS刺激时间的延长而显著升高,在刺激后12小时达高峰。0.1 ~ 10μmol/l的氨氯地平、MN9202消旋体、R-(+)-MN9202对映体可浓度依赖性的抑制LPS刺激的TNF-α的释放和mRNA的表达,S-(-)-MN9202对映体对TNF-α的释放及mRNA的表达无明显影响。1.0μmol/l的氨氯地平、MN9202消旋体、R-(+)-MN9202、S-(-)-MN9202对TNF-α释放的抑制率分别为52.41、31.62、49.81和2.25 %。⑥MN9202消旋体可浓度依赖性地抑制LPS刺激所致iNOS和COX-2转录和翻译水平的表达;1.0μmol/l的MN9202消旋体对COX-2的抑制作用与氨氯地平相当,对iNOS的抑制作用强于氨氯地平;R-(+)-MN9202明显抑制iNOS和COX-2的表达(P<0.05),而S-(-)-MN9202对iNOS和COX-2的表达均无影响。⑦LPS刺激的同时,用MN9202消旋体、R-(+)-MN9202对映体或氨氯地平处理心肌细胞,均可明显增加pAkt的表达(P<0.05);但S-(-)-MN9202对Akt的活化无明显影响。用PI3K抑制剂wortmannin或LY294002预处理2 h后,LPS和DHPs(MN9202消旋体、R-(+)-MN9202对映体和氨氯地平)共处理心肌细胞,各DHPs处理组Akt的活化均受到抑制;3种DHPs类化合物对TNF-α释放的抑制作用均可被wortmannin或LY294002不同程度的降低;Cox-2和iNOS蛋白的表达与无PI3K抑制剂预处理组相比无显著性差异。
结论:①本研究建立的手性固定相拆分条件为MN9202单一对映异构体的分离、制备及含量测定等提供了简单、快捷的方法。②两种MN9202对映体对心肌细胞收缩与舒张特性、钙瞬变和L型钙通道均具有立体选择性。S-(-)-MN9202表现出了拮抗特性,而R-(+)-MN9202则具有一定的激动活性。③MN9202对LPS致大鼠心肌损伤有保护作用,此作用与其抑制LPS引起的TNF-α、iNOS和COX-2表达升高密切相关,其中R-(+)-MN9202是抑制炎症相关基因表达的优映体。④MN9202通过激活PI3K,引起下游Akt的活化,参与了TNF-α的调节过程,但对Cox-2和iNOS的调节作用可能依赖于其它信号转导通路。
Aim In this study, we attempt to set up a resolution approach to get single enantiomer of MN9202 on a chiral stationary phase column; then, to determine the stereoselectivity effects of the optical enantiomers of MN9202 on cell shortening and relengthening, calcium transient and L-type calcium channels in rat ventricular myocytes; in addition, to pursue the effects and mechanisms of the optical enantiomers of MN9202 on myocardium injury induced by LPS. From above, we want to get a scientific evaluation for MN9202, which will be helpful for enlarging our understanding of clinic applications of dihydropyridine-type drugs.
Methods①The enantiomers of MN9202 were separated on Daicel OJ-H column by optimizing conditions of chromatogram, such as the proportion of hexane or ethanol in the mobile phase.②Mechanical contraction properties and calcium transient were recorded in myocytes and assessed by video-edge detection system in absence or presence of racemic MN9202, S-(-)-MN9202 and R-(+)-MN9202.③To investigate the stereoselectivity of the optical enantiomers of MN9202 to ion channels, the actions (including ICa,L, steady-state activation and steady-state inactivation curves) of the optical enantiomers of MN9202 and racemic MN9202 were studied on L-type calciun channels in single isolateded adult rat ventricular myocytes, using the whole-cell configuration of the patch clamp technique.④Sepsis model was set up by LPS (5 mg/kg, i.v.) in anesthetized rats. The right carotid artery was connected to a pressure transducer (RM-6200) for the measurement of mean arterial blood pressure (MAP) and heart rate (HR). After recorded baseline hemodynamic parameters, animals received vehicle or LPS were monitored for 240 min. 0.5 ml blood was taken to measure the plasma levels of TNF-αby ELISA. Finally, myocardial samples were taken to stain with hematoxylin and eosin.⑤The primary cultured neonatal rat cardiomyocytes were treated with LPS, alone or in the presence of various concentrations of racemic MN9202, MN9202 enantiomers or amlodipine. The TNF-αrelease in the culture medium was measured by ELISA. TNF-αmRNA and proteins in myocytes were measured by RT-PCR and immunofluorescent staining analysis, respectively.⑥The iNOS and COX-2 mRNA or protein levels in LPS-treated cardiomyocytes at different times or cotreated with DHPs were assessed by RT-PCR and western blot assays, respectively. Regulation of phospho-Akt (pAkt) and Akt was assessed by western blot assays.
Results①The chromatographic separations were performed using a OJ-H column and using a mobile phase, hexane: ethanol: triethylamine (93︰7︰0.5, V ︰V︰V). The flow-rate of the mobile phase was 0.5 ml/min, and the samples were monitored with UV detection at 237 nm. Under the optimized conditions, the enantiomers of MN9202 were well separated from the baseline and several milligrams two enantiomers, S-(-)-MN9202 and R-(+)-MN9202 were obtained.②The exposure of racemic MN9202 and S-(-)-MN9202 (1×10-8 mol/l ~ 1×10-5 mol/l) caused concentration-dependent inhibition of PTA and ?FFI in single rat ventricular myocytes. The extent of maximal percentage inhibition of PTA %baseline was significantly reduced by 73.8 % and 92.7 %, respectively. ?FFI was reduced from 0.45±0.04 to 0.07±0.02 and from 0.45±0.04 to 0.05±0.01 in presence of 1×10-5 mol/l racemic MN9202 and S-(-)-MN9202, respectively. However, R-(+)-MN9202 induced concentration-dependent increases of PTA and ?FFI in single rat ventricular myocytes. The extent of maximal percentage inhibition of PTA %baseline was significantly increased by 18.2 %, and ?FFI was increased from 0.45±0.01 to 0.58±0.05 in presence of 1×10-5 mol/l R-(+)-MN9202 (P<0.01, compared with control group).③In response to depolarizations positive to 0 mV, ICa,L density was reduced by 6.7 %,15.0 % (P<0.01), 27.5 % (P<0.01), 50.7 % (P<0.01) and 69.5 % (P<0.01), respectively in 0.03, 0.1, 0.3, 1 and 3μmol/l racemic MN9202 groups. S-(-)-MN9202 (10-6 mol/l) reduced the ICa,L density from -9.8±1.0 to -3.2±0.9 (P<0.01, 67% decrease) in response to depolarizations positive to 0 mV, exhibiting more effective antagonist activity than racemic MN9202. However, R-(+)-MN9202 (10-6 mol/l) increased the ICa,L density from -9.3±0.7 to -10.3±0.6 (P<0.01) in response to depolarizations positive to 0 mV. In ventricular myocytes, racemic MN9202, S-(-)-MN9202 and R-(+)-MN9202 didn’t affect the steady-state activation curve of ICa, L. However, a shift of the midpoint of the steady-state inactivation curve was produced in the hyperpolarizing direction from ?28.8±0.2 mV to ?35.4±0.7 mV in the presence of 1μmol/l racemic MN9202 and from?28.8±0.2 mV to ?37.7±0.6 mV in the presence of 1μmol/l S-(-)-MN9202. But, in contrast, R-(+)-MN9202 at the same concentration did cause only a small shift of the midpoint of the steady-state inactivation curve (to the right) in the depolarizing direction from ?28.8±0.2 mV to ?27.1±0.6.④Administration of LPS caused a rapid fall in MAP from 119±4 to 85±2 mmHg (P<0.05)within 30 min. After 60 min, there was a continuous further fall in MAP. 10 and 30μg/kg MN9202 prevented the delayed fall in MAP observed in LPS groups. Administration of LPS resulted in an increase of heart rate, and in the MN9202 group, there was no significant difference of heart rate compared with LPS group. The injection of LPS resulted in bell-shape changes in the plasma levels of TNF-αwhich reached a peak at 60 min after LPS injection and subsequently decreased slowly (P<0.05). Treatment of 10μg/kg and 30μg/kg MN9202 decreased the TNF-αlevel in plasma. In LPS group, increase and oedema of interstitial cell, vacuolar degeneration of cardiomyocytes, inflammatory cell infiltrate have been significantly found, compared with control group. Administration of 10μg/kg and 30μg/kg MN9202 mitigated these above symptoms.⑤Neonatal rat cardiomyocytes did not release detectable amounts of TNF-αin the absence of LPS. However, in the presence of LPS, TNF-αrelease in the culture medium increased in a concentration-depended manner and mRNA levels also increased rapidly, reaching a peak at 12 h. Amlodipine, racemic MN9202 and R-(+)-MN9202, in concentrations ranging from 0. 1 to 10μmol/l, could concentration-dependently inhibit the release and mRNA expression of TNF-αin LPS-stimulated cardiomyocytes, but S-(-)-MN9202 had no obvious effect on the TNF-αrelease and mRNA expression. The inhibition rates of TNF-αrelease from LPS-stimulated neonatal rat cardiomyocytes in 1.0μmol/l amlodipine, racemic MN9202, R-(+)-MN9202 and S-(-)-MN9202 groups were 52.41, 31.62, 49.81 and 2.25 %, respectively.⑥Racemic MN9202 significantly attenuated the iNOS and TNF-αmRNA and protein levels in cardiomyocytes stimulated with LPS in a concentration-dependent manner. While the inhibition rate of 1.0μmol/l MN9202 on COX-2 expression was almost as same as the one of amlodipine, MN9202 showed more effective regulation on the iNOS levels in same concentration. Furthermore, R-MN9202 significantly depressed the levels of iNOS and COX-2, but S-MN9202 could not significantly attenuate these inflammation-relevant genes expression.⑦Compared with LPS-stimulated cells, when cardiomyocytes were cotreated with LPS and racemic MN9202, R-(+)-MN9202 or amlodipine, the phosphorylation levels of Akt were markedly increased (P<0.05), but S-(-)-MN9202 had no effect on the pAkt levels. Pre-treatment with selective inhibitors of the PI3K/Akt pathway, wortmannin or LY294002, in LPS and DHPs (racemic MN9202, R-(+)-MN9202 and amlodipine)-treated cells, Akt phosphorylation was partially inhibited. To elucidate the contribution of PI3K/Akt pathway to the regulation of TNF-α, Cox-2 and iNOS protein levels, cardiomyocytes were pretreated with wortmannin or LY294002 for 2 h and then cotreated with LPS and DHPs. The inhibition of PI3K with wortmannin or LY294002 partly attenuated the depression effects of DHPs on TNF-αprotein expression, but there was no significiant difference on the Cox-2 and iNOS expression between the groups which pre-treated with the PI3K/Akt inhibitors or not.
Conclusion①The proposed chromatographic method is easy and accurate. It is reliable in the separation, preparation and quantification of MN9202.②S-(-)-MN9202 and R-(+)-MN9202 produce differential stereoselectivity effects in mechanical contraction properties (shortening and relengthening of individual myocytes), myocyte Ca2+ transients and L-type calcium channels. S-(-)-MN9202 displayed an antagonist activation, whereas R-(+)-MN9202 exerted an agonist effect.③MN9202 protected the impaired myocardium induced by LPS through inhibiting the LPS-induced expression of TNF-α, iNOS and COX-2. R-(+)-MN9202 has more markedly inhibitory effects on these LPS-induced inflammation-relevant genes expression.④The anti-TNF-αeffects of MN9202 were partly mediated by activation of Akt, downstream of the PI3K signal cascade, but anti-iNOS and COX-2 effects might be mediated by other signal pathway.
方法:①采用Daicel OJ-H手性固定相色谱柱,通过对流动相中正己烷与乙醇的比例、添加三乙胺的量对MN9202对映体的保留时间及分离度的影响进行考察,优化色谱条件,拆分MN9202消旋体。②采用IonOptix单细胞动缘探测系统同步检测MN9202消旋体、S-(-)-MN9202和R-(+)-MN9202作用后心肌细胞收缩、舒张和钙瞬变的变化。③采用全细胞膜片钳方法,比较MN9202消旋体、S-(-)-MN9202和R-(+)-MN9202对心肌细胞L型钙通道电流密度的影响。进而探讨各药物对L型钙通道稳态激活动力学和稳态失活动力学特性的影响。④通过股静脉注射LPS建立感染性休克致动物心肌损伤模型,研究MN9202对心肌的保护作用;RM-6200型四道生理记录仪实时监测血压等参数;用ELISA法测定血浆TNF-α水平;取心肌组织样本,进行HE染色,比较心肌形态学改变。⑤原代培养新生大鼠心肌细胞,用LPS和不同浓度的MN9202消旋体、对映体及氨氯地平作用,采用ELISA法测定培养上清液中TNF-α含量,RT-PCR及免疫荧光法分别测定心肌细胞中TNF-αmRNA和蛋白的表达。⑥RT-PCR和western blot法分别测定LPS刺激不同时间及DHPs作用后心肌细胞iNOS、COX-2 mRNA和蛋白的表达。Western blot法测定磷酸化Akt和总Akt的表达。
结果:①本研究建立的MN9202 HPLC手性拆分条件为:Daicel OJ-H手性分析柱,正己烷-乙醇-三乙胺(93︰7︰0.5,V︰V︰V)为流动相,流速为0.5 ml/min,检测波长为237 nm。用此法MN9202对映异构体可达到基线分离,并可少量制备S-(-)-MN9202和R-(+)-MN9202。②不同浓度的MN9202消旋体和S-(-)-MN9202可剂量依赖性抑制大鼠单个心肌细胞的收缩:在1×10-5 mol/l浓度时,百分峰值收缩幅度(PTA %baseline)与对照组相比分别降低了73.8 %和92.7 %;而R-(+)-MN9202则呈剂量依赖性增强心肌细胞的收缩,1×10-5 mol/l时, PTA %baseline升高了18.2 %。MN9202消旋体和S-(-)-MN9202剂量依赖性抑制单个心肌细胞钙瞬变幅度(△FFI)。与对照组相比,1×10-5 mol/l MN9202消旋体和S-(-)-MN9202,细胞钙瞬变分别由0.45±0.04下降到0.07±0.02,及由0.45±0.04下降到0.05±0.01(P<0.01)。而R-(+)-MN9202则增加了钙瞬变幅度:在最大浓度1×10-5 mol/l时,细胞钙瞬变由0.45±0.01升高到0.58±0.05(P<0.01)。③MN9202消旋体剂量依赖性的降低了L型钙通道电流密度。检测电压为0 mV,MN9202浓度为0.03、0.1、0.3、1和3μmol/l时,CaL电流峰值的密度较对照组分别减少了6.7 %、15.0 %(P<0.01)、27.5 %(P<0.01)、50.7 %(P<0.01)和69.5 %(P<0.01);但药物并未改变L型钙通道激活的阈电位(-40 mV)和峰值电流电位(0 mV)。S-(-)-MN9202也抑制了L型钙通道电流密度,检测电压为0 mV时,1μmol/l S-(-)-MN9202使CaL电流峰值的密度从对照组的-9.8±1.0减少到-3.2±0.9(减少了67 %)(P<0.01)。而1μmol/l R-(+)-MN9202则引起了L型钙通道电流密度增加:检测电压为0 mV时,使CaL电流峰值的密度从对照组的-9.3±0.7增加到-10.3±0.6(P<0.01)。MN9202消旋体,S-(-)-MN9202和R-(+)-MN9202均未明显改变L型钙通道的激活特性。但各药物均对通道稳态失活有一定的影响:MN9202消旋体和S-(-)-MN9202均使稳态失活曲线左移,而R-(+)-MN9202使稳态失活曲线轻微右移。④LPS在30 min内引起大鼠平均动脉压的快速下降,从119±4降至85±2 mmHg(P<0.05);60 min后,大鼠平均动脉压仍持续下降,在240 min时为81±3 mmHg。10及30μg/kg MN9202消旋体给药后,延缓了LPS对大鼠平均动脉压的下降。但MN9202的应用并未改变LPS对心率的影响。给予LPS使血浆中TNF-α的水平发生了变化,先升高,至1小时达到峰值,而后逐渐降低。MN9202给药后可使LPS引起的大鼠血浆TNF-α升高的峰值显著降低(P<0.05)。⑤正常心肌细胞几乎不表达TNF-α,但细胞培养液中TNF-α的含量可随LPS浓度的增加而提高;TNF-α的释放与mRNA表达也随着LPS刺激时间的延长而显著升高,在刺激后12小时达高峰。0.1 ~ 10μmol/l的氨氯地平、MN9202消旋体、R-(+)-MN9202对映体可浓度依赖性的抑制LPS刺激的TNF-α的释放和mRNA的表达,S-(-)-MN9202对映体对TNF-α的释放及mRNA的表达无明显影响。1.0μmol/l的氨氯地平、MN9202消旋体、R-(+)-MN9202、S-(-)-MN9202对TNF-α释放的抑制率分别为52.41、31.62、49.81和2.25 %。⑥MN9202消旋体可浓度依赖性地抑制LPS刺激所致iNOS和COX-2转录和翻译水平的表达;1.0μmol/l的MN9202消旋体对COX-2的抑制作用与氨氯地平相当,对iNOS的抑制作用强于氨氯地平;R-(+)-MN9202明显抑制iNOS和COX-2的表达(P<0.05),而S-(-)-MN9202对iNOS和COX-2的表达均无影响。⑦LPS刺激的同时,用MN9202消旋体、R-(+)-MN9202对映体或氨氯地平处理心肌细胞,均可明显增加pAkt的表达(P<0.05);但S-(-)-MN9202对Akt的活化无明显影响。用PI3K抑制剂wortmannin或LY294002预处理2 h后,LPS和DHPs(MN9202消旋体、R-(+)-MN9202对映体和氨氯地平)共处理心肌细胞,各DHPs处理组Akt的活化均受到抑制;3种DHPs类化合物对TNF-α释放的抑制作用均可被wortmannin或LY294002不同程度的降低;Cox-2和iNOS蛋白的表达与无PI3K抑制剂预处理组相比无显著性差异。
结论:①本研究建立的手性固定相拆分条件为MN9202单一对映异构体的分离、制备及含量测定等提供了简单、快捷的方法。②两种MN9202对映体对心肌细胞收缩与舒张特性、钙瞬变和L型钙通道均具有立体选择性。S-(-)-MN9202表现出了拮抗特性,而R-(+)-MN9202则具有一定的激动活性。③MN9202对LPS致大鼠心肌损伤有保护作用,此作用与其抑制LPS引起的TNF-α、iNOS和COX-2表达升高密切相关,其中R-(+)-MN9202是抑制炎症相关基因表达的优映体。④MN9202通过激活PI3K,引起下游Akt的活化,参与了TNF-α的调节过程,但对Cox-2和iNOS的调节作用可能依赖于其它信号转导通路。
Aim In this study, we attempt to set up a resolution approach to get single enantiomer of MN9202 on a chiral stationary phase column; then, to determine the stereoselectivity effects of the optical enantiomers of MN9202 on cell shortening and relengthening, calcium transient and L-type calcium channels in rat ventricular myocytes; in addition, to pursue the effects and mechanisms of the optical enantiomers of MN9202 on myocardium injury induced by LPS. From above, we want to get a scientific evaluation for MN9202, which will be helpful for enlarging our understanding of clinic applications of dihydropyridine-type drugs.
Methods①The enantiomers of MN9202 were separated on Daicel OJ-H column by optimizing conditions of chromatogram, such as the proportion of hexane or ethanol in the mobile phase.②Mechanical contraction properties and calcium transient were recorded in myocytes and assessed by video-edge detection system in absence or presence of racemic MN9202, S-(-)-MN9202 and R-(+)-MN9202.③To investigate the stereoselectivity of the optical enantiomers of MN9202 to ion channels, the actions (including ICa,L, steady-state activation and steady-state inactivation curves) of the optical enantiomers of MN9202 and racemic MN9202 were studied on L-type calciun channels in single isolateded adult rat ventricular myocytes, using the whole-cell configuration of the patch clamp technique.④Sepsis model was set up by LPS (5 mg/kg, i.v.) in anesthetized rats. The right carotid artery was connected to a pressure transducer (RM-6200) for the measurement of mean arterial blood pressure (MAP) and heart rate (HR). After recorded baseline hemodynamic parameters, animals received vehicle or LPS were monitored for 240 min. 0.5 ml blood was taken to measure the plasma levels of TNF-αby ELISA. Finally, myocardial samples were taken to stain with hematoxylin and eosin.⑤The primary cultured neonatal rat cardiomyocytes were treated with LPS, alone or in the presence of various concentrations of racemic MN9202, MN9202 enantiomers or amlodipine. The TNF-αrelease in the culture medium was measured by ELISA. TNF-αmRNA and proteins in myocytes were measured by RT-PCR and immunofluorescent staining analysis, respectively.⑥The iNOS and COX-2 mRNA or protein levels in LPS-treated cardiomyocytes at different times or cotreated with DHPs were assessed by RT-PCR and western blot assays, respectively. Regulation of phospho-Akt (pAkt) and Akt was assessed by western blot assays.
Results①The chromatographic separations were performed using a OJ-H column and using a mobile phase, hexane: ethanol: triethylamine (93︰7︰0.5, V ︰V︰V). The flow-rate of the mobile phase was 0.5 ml/min, and the samples were monitored with UV detection at 237 nm. Under the optimized conditions, the enantiomers of MN9202 were well separated from the baseline and several milligrams two enantiomers, S-(-)-MN9202 and R-(+)-MN9202 were obtained.②The exposure of racemic MN9202 and S-(-)-MN9202 (1×10-8 mol/l ~ 1×10-5 mol/l) caused concentration-dependent inhibition of PTA and ?FFI in single rat ventricular myocytes. The extent of maximal percentage inhibition of PTA %baseline was significantly reduced by 73.8 % and 92.7 %, respectively. ?FFI was reduced from 0.45±0.04 to 0.07±0.02 and from 0.45±0.04 to 0.05±0.01 in presence of 1×10-5 mol/l racemic MN9202 and S-(-)-MN9202, respectively. However, R-(+)-MN9202 induced concentration-dependent increases of PTA and ?FFI in single rat ventricular myocytes. The extent of maximal percentage inhibition of PTA %baseline was significantly increased by 18.2 %, and ?FFI was increased from 0.45±0.01 to 0.58±0.05 in presence of 1×10-5 mol/l R-(+)-MN9202 (P<0.01, compared with control group).③In response to depolarizations positive to 0 mV, ICa,L density was reduced by 6.7 %,15.0 % (P<0.01), 27.5 % (P<0.01), 50.7 % (P<0.01) and 69.5 % (P<0.01), respectively in 0.03, 0.1, 0.3, 1 and 3μmol/l racemic MN9202 groups. S-(-)-MN9202 (10-6 mol/l) reduced the ICa,L density from -9.8±1.0 to -3.2±0.9 (P<0.01, 67% decrease) in response to depolarizations positive to 0 mV, exhibiting more effective antagonist activity than racemic MN9202. However, R-(+)-MN9202 (10-6 mol/l) increased the ICa,L density from -9.3±0.7 to -10.3±0.6 (P<0.01) in response to depolarizations positive to 0 mV. In ventricular myocytes, racemic MN9202, S-(-)-MN9202 and R-(+)-MN9202 didn’t affect the steady-state activation curve of ICa, L. However, a shift of the midpoint of the steady-state inactivation curve was produced in the hyperpolarizing direction from ?28.8±0.2 mV to ?35.4±0.7 mV in the presence of 1μmol/l racemic MN9202 and from?28.8±0.2 mV to ?37.7±0.6 mV in the presence of 1μmol/l S-(-)-MN9202. But, in contrast, R-(+)-MN9202 at the same concentration did cause only a small shift of the midpoint of the steady-state inactivation curve (to the right) in the depolarizing direction from ?28.8±0.2 mV to ?27.1±0.6.④Administration of LPS caused a rapid fall in MAP from 119±4 to 85±2 mmHg (P<0.05)within 30 min. After 60 min, there was a continuous further fall in MAP. 10 and 30μg/kg MN9202 prevented the delayed fall in MAP observed in LPS groups. Administration of LPS resulted in an increase of heart rate, and in the MN9202 group, there was no significant difference of heart rate compared with LPS group. The injection of LPS resulted in bell-shape changes in the plasma levels of TNF-αwhich reached a peak at 60 min after LPS injection and subsequently decreased slowly (P<0.05). Treatment of 10μg/kg and 30μg/kg MN9202 decreased the TNF-αlevel in plasma. In LPS group, increase and oedema of interstitial cell, vacuolar degeneration of cardiomyocytes, inflammatory cell infiltrate have been significantly found, compared with control group. Administration of 10μg/kg and 30μg/kg MN9202 mitigated these above symptoms.⑤Neonatal rat cardiomyocytes did not release detectable amounts of TNF-αin the absence of LPS. However, in the presence of LPS, TNF-αrelease in the culture medium increased in a concentration-depended manner and mRNA levels also increased rapidly, reaching a peak at 12 h. Amlodipine, racemic MN9202 and R-(+)-MN9202, in concentrations ranging from 0. 1 to 10μmol/l, could concentration-dependently inhibit the release and mRNA expression of TNF-αin LPS-stimulated cardiomyocytes, but S-(-)-MN9202 had no obvious effect on the TNF-αrelease and mRNA expression. The inhibition rates of TNF-αrelease from LPS-stimulated neonatal rat cardiomyocytes in 1.0μmol/l amlodipine, racemic MN9202, R-(+)-MN9202 and S-(-)-MN9202 groups were 52.41, 31.62, 49.81 and 2.25 %, respectively.⑥Racemic MN9202 significantly attenuated the iNOS and TNF-αmRNA and protein levels in cardiomyocytes stimulated with LPS in a concentration-dependent manner. While the inhibition rate of 1.0μmol/l MN9202 on COX-2 expression was almost as same as the one of amlodipine, MN9202 showed more effective regulation on the iNOS levels in same concentration. Furthermore, R-MN9202 significantly depressed the levels of iNOS and COX-2, but S-MN9202 could not significantly attenuate these inflammation-relevant genes expression.⑦Compared with LPS-stimulated cells, when cardiomyocytes were cotreated with LPS and racemic MN9202, R-(+)-MN9202 or amlodipine, the phosphorylation levels of Akt were markedly increased (P<0.05), but S-(-)-MN9202 had no effect on the pAkt levels. Pre-treatment with selective inhibitors of the PI3K/Akt pathway, wortmannin or LY294002, in LPS and DHPs (racemic MN9202, R-(+)-MN9202 and amlodipine)-treated cells, Akt phosphorylation was partially inhibited. To elucidate the contribution of PI3K/Akt pathway to the regulation of TNF-α, Cox-2 and iNOS protein levels, cardiomyocytes were pretreated with wortmannin or LY294002 for 2 h and then cotreated with LPS and DHPs. The inhibition of PI3K with wortmannin or LY294002 partly attenuated the depression effects of DHPs on TNF-αprotein expression, but there was no significiant difference on the Cox-2 and iNOS expression between the groups which pre-treated with the PI3K/Akt inhibitors or not.
Conclusion①The proposed chromatographic method is easy and accurate. It is reliable in the separation, preparation and quantification of MN9202.②S-(-)-MN9202 and R-(+)-MN9202 produce differential stereoselectivity effects in mechanical contraction properties (shortening and relengthening of individual myocytes), myocyte Ca2+ transients and L-type calcium channels. S-(-)-MN9202 displayed an antagonist activation, whereas R-(+)-MN9202 exerted an agonist effect.③MN9202 protected the impaired myocardium induced by LPS through inhibiting the LPS-induced expression of TNF-α, iNOS and COX-2. R-(+)-MN9202 has more markedly inhibitory effects on these LPS-induced inflammation-relevant genes expression.④The anti-TNF-αeffects of MN9202 were partly mediated by activation of Akt, downstream of the PI3K signal cascade, but anti-iNOS and COX-2 effects might be mediated by other signal pathway.
引文
[1] 朱肖星, 梅其炳, 刘莉, 等. MN9202 对心肌细胞的收缩和舒张功能的影响. 中华老年多器官疾病杂志. 2005, 4, 50-53.
[2] 王剑波, 郭炜, 梅其炳, 等. 钙拮抗剂 MN9202 对心肌肥厚大鼠全血和心肌中5-HT 的影响. 第四军医大学学报. 2003, 24, 1688-90.
[3] 王莉莉, 梅其炳, 赵德化. 2,6-二甲基-4-(3-硝基苯基)-1,4-二氢-3,5-吡啶二羧酸-3-甲酯-5-正戊酯(MN9202)抗实验性血栓形成作用. 中国药学杂志. 1998, 33, 282-95.
[4] 姚秀娟, 甘洪泉, 贾敏, 等. 钙拮抗剂 MN9202 对高脂血症大鼠的影响. 第四军医大学学报. 2000, 21, 311-3.
[5] 甘戈, 梅其炳, 赵德化. MN9202 对兔小肠缺血再灌注损伤时血管内皮细胞功能的保护作用. 第四军医大学学报. 1998, 19, 571-3.
[6] 李新立. 钙拮抗剂的分类及其在高血压治疗中的选用. 高血压杂志. 1999, 7, 72-5.
[7] De Camp, W.H. Chiral drugs: the FDA perspective on manufacturing and control. J Pharm Biomed Anal. 1993, 11, 1167-72.
[8] 王普善, 王宇梅. 手性药物开发战略的再认识. 精细与专用化学品. 2004, 12, 4-8.
[9] 王丹, 李亚, 何浪. 手性药物及其开发与应用. 现代医药卫生. 2007, 23, 837-8.
[10] Assmann, I., Dittrich, P., Schwela, H. and Fiehring, H. [Clinico-pharmacological studies with Nifedipine--a new coronary substance]. Z Gesamte Inn Med. 1975, 30, 575-9.
[11] Triggle, D.J. 1,4-Dihydropyridines as calcium channel ligands and privileged structures. Cell Mol Neurobiol. 2003, 23, 293-303.
[12] Triggle, D.J. Calcium antagonists. History and perspective. Stroke. 1990, 21,IV49-58.
[13] Triggle, D.J. Calcium channel antagonists: Clinical uses-Past, present and future. Biochem Pharmacol. 2007 Jan 13; [Epub ahead of print]
[14] Grossman, E. and Messerli, F.H. Calcium antagonists. Prog Cardiovasc Dis. 2004, 47, 34-57.
[15] Leenen, F.H. Clinical relevance of 24 h blood pressure control by 1,4-dihydropyridines. Am J Hypertens. 1996, 9, 97S-104S; discussion 108S-109S.
[16] Wenzel, R.R., Allegranza, G., Binggeli, C., Shaw, S., Weidmann, P., Luscher, T.F. and Noll, G. Differential activation of cardiac and peripheral sympathetic nervous system by nifedipine: role of pharmacokinetics. J Am Coll Cardiol. 1997, 29, 1607-14.
[17] Haria, M. and Wagstaff, A.J. Amlodipine. A reappraisal of its pharmacological properties and therapeutic use in cardiovascular disease. Drugs. 1995, 50, 560-86.
[18] Motomura, S., Wu, Z.J. and Hashimoto, K. Lacidipine, a new long-acting dihydropyridine calcium antagonist, has high vascular selectivity against all intracardiac variables. Heart Vessels. 1993, 8, 16-22.
[19] Shan, R., Velazquez, C. and Knaus, E.E. Syntheses, calcium channel agonist-antagonist modulation activities, and nitric oxide release studies of nitrooxyalkyl 1,4-dihydro-2,6-dimethyl-3-nitro-4-(2,1,3-benzoxadiazol-4-yl)pyridine-5-ca rboxylate racemates, enantiomers, and diastereomers. J Med Chem. 2004, 47, 254-61.
[20] Ariens, E.J. Stereochemistry: a source of problems in medicinal chemistry. Med Res Rev. 1986, 6, 451-66.
[21] Inotsume, N. and Nakano, M. Stereoselective determination and pharmacokinetics of dihydropyridines: an updated review. J Biochem Biophys Methods. 2002, 54, 255-74.
[22] Millership, J.S. and Fitzpatrick, A. Commonly used chiral drugs: a survey. Chirality. 1993, 5, 573-6.
[23] Nerurkar, S.G., Dighe, S.V. and Williams, R.L. Bioequivalence of racemic drugs. J Clin Pharmacol. 1992, 32, 935-43.
[24] Bischer, A., Zia-Amirhosseini, P., Iwaki, M., McDonagh, A.F. and Benet, L.Z. Stereoselective binding properties of naproxen glucuronide diastereomers to proteins. J Pharmacokinet Biopharm. 1995, 23, 379-95.
[25] 曾苏. 手性药物与手性药理学. 浙江大学出版社. 2002
[26] Carter, R.B. and Dykstra, L.A. Effects of picenadol (LY150720) and its stereoisomers on electric shock titration in the squirrel monkey. J Pharmacol Exp Ther. 1985, 234, 299-306.
[27] Middlefell, V.C. and Price, T.L. 5-HT3 receptor agonism may be responsible for the emetic effects of zacopride in the ferret. Br J Pharmacol. 1991, 103, 1011-2.
[28] Tabi, T., Magyar, K. and Szoko, E. Chiral characterization of deprenyl-N-oxide and other deprenyl metabolites by capillary electrophoresis using a dual cyclodextrin system in rat urine. Electrophoresis. 2003, 24, 2665-73.
[29] Kean, W.F., Lock, C.J. and Howard-Lock, H.E. Chirality in antirheumatic drugs. Lancet. 1991, 338, 1565-8.
[30] Jamali, F., Mehvar, R. and Pasutto, F.M. Enantioselective aspects of drug action and disposition: therapeutic pitfalls. J Pharm Sci. 1989, 78, 695-715.
[31] 余细勇, 林曙光. 手性药物对映体的药动学与药效学立体选择性差异研究进展. 广州医药. 1997, 28, 1-3.
[32] Lane, R.M. and Baker, G.B. Chirality and drugs used in psychiatry: nice to know or need to know? Cell Mol Neurobiol. 1999, 19, 355-72.
[33] 李高. 手性药物的临床药物动力学立体选择性. 中国医院药学杂志. 1999, 19, 429-31.
[34] Bode, W., Toet, A.E., Stolker, A.A., van Ginkel, L.A., Groen, K., Wemer, J. and de Wildt, D.J. Toxicokinetics of a single intravenous dose of rac-propranolol versus optically pure propranolol in the rat. Chirality. 1995, 7, 626-31.
[35] Caldwell, J., Hutt, A.J. and Fournel-Gigleux, S. The metabolic chiral inversion and dispositional enantioselectivity of the 2-arylpropionic acids and their biological consequences. Biochem Pharmacol. 1988, 37, 105-14.
[36] Lee, E.J. and Williams, K.M. Chirality. Clinical pharmacokinetic and pharmacodynamic considerations. Clin Pharmacokinet. 1990, 18, 339-45.
[37] 章立, 姚彤炜, 曾苏. 手性药物对映体的药物代谢动力学. 中国现代应用药学杂志. 1999, 16, 4-7.
[38] Schramm, M., Thomas, G., Towart, R. and Franckowiak, G. Novel dihydropyridines with positive inotropic action through activation of Ca2+ channels. Nature. 1983, 303, 535-7.
[39] Muto, K., Kuroda, T., Kawato, H., Karasawa, A., Kubo, K. and Nakamizo, N. Synthesis and pharmacological activity of stereoisomers of 1,4-dihydro-2,6-dimethyl-4-(3-nitrophenyl)-3,5-pyridine-dicarboxylic acid methyl 1-(phenylmethyl)-3-piperidinyl ester. Arzneimittelforschung. 1988, 38, 1662-5.
[40] Zhang, X.P., Loke, K.E., Mital, S., Chahwala, S. and Hintze, T.H. Paradoxical release of nitric oxide by an L-type calcium channel antagonist, the R+ enantiomer of amlodipine. J Cardiovasc Pharmacol. 2002, 39, 208-14.
[41] Berjukow, S. and Hering, S. Voltage-dependent acceleration of Ca(v)1.2 channel current decay by (+)- and (-)-isradipine. Br J Pharmacol. 2001, 133, 959-66.
[42] Furukawa, T., Miura, R., Honda, M., Kamiya, N., Mori, Y., Takeshita, S., Isshiki, T. and Nukada, T. Identification of R(-)-isomer of efonidipine as a selective blocker of T-type Ca2+ channels. Br J Pharmacol. 2004, 143, 1050-7.
[43] Jabor, V.A., Coelho, E.B. and Lanchote, V.L. Enantioselective pharmacokinetics of lercanidipine in healthy volunteers. J Chromatogr B Analyt Technol Biomed Life Sci. 2004, 813, 343-6.
[44] Hirano, T. et al. Differential properties of the optical-isomers of pranidipine, a 1,4-dihydropyridine calcium channel modulator. Fundam Clin Pharmacol. 1999, 13, 650-5.
[45] Okumura, K., Ichihara, K., Nagasaka, M., Oda, N. and Tajima, K. Calcium entry blocking activities of MPC-1304 and of its enantiomers and metabolites. Eur J Pharmacol. 1993, 235, 69-74.
[46] Wang, S., He, L. and Yun, B. The difference between nicardipine and its enantiomers on inhibiting vasoconstriction of isolated rabbit thoracic artery. Arch Pharm Res. 2005, 28, 319-24.
[47] Vo, D., Matowe, W.C., Ramesh, M., Iqbal, N., Wolowyk, M.W., Howlett, S.E. and Knaus, E.E. Syntheses, calcium channel agonist-antagonist modulation activities, and voltage-clamp studies of isopropyl 1,4-dihydro-2,6-dimethyl-3-nitro-4-pyridinylpyrid- ine-5-carboxylate racemates and enantiomers. J Med Chem. 1995, 38, 2851-9.
[48] Hof, R.P., Ruegg, U.T., Hof, A. and Vogel, A. Stereoselectivity at the calcium channel: opposite action of the enantiomers of a 1,4-dihydropyridine. J Cardiovasc Pharmacol. 1985, 7, 689-93.
[49] Tokuma, Y., Fujiwara, T. and Noguchi, H. Determination of (+)- and (-)-nilvadipinein human plasma using chiral stationary-phase liquid chromatography and gas chromatography-mass spectrometry, and a preliminary pharmacokinetic study in humans. J Pharm Sci. 1987, 76, 310-3.
[50] Uno, T., Ohkubo, T., Motomura, S. and Sugawara, K. Effect of grapefruit juice on the disposition of manidipine enantiomers in healthy subjects. Br J Clin Pharmacol. 2006, 61, 533-7.
[51] Marques, M.P., Santos, N.A., Coelho, E.B., Bonato, P.S. and Lanchote, V.L. Enantioselective assay of nisoldipine in human plasma by chiral high-performance liquid chromatography combined with gas chromatographic-mass spectrometry: applications to pharmacokinetics. J Chromatogr B Biomed Sci Appl. 2001, 762, 87-95.
[52] He, L.C. and Wang, S.C. Studies on differences of pharmacokinetic behavior and tissue distribution of nimodipine and its two enantiomers in rats using achiral and chiral liquid chromatography. Yao Xue Xue Bao. 2003, 38, 603-8.
[53] Fischer, C., Schonberger, F., Muck, W., Heuck, K. and Eichelbaum, M. Simultaneous assessment of the intravenous and oral disposition of the enantiomers of racemic nimodipine by chiral stationary-phase high-performance liquid chromatography and gas chromatography/mass spectroscopy combined with a stable isotope technique. J Pharm Sci. 1993, 82, 244-50.
[54] Mast, V., Fischer, C., Mikus, G. and Eichelbaum, M. Use of pseudoracemic nitrendipine to elucidate the metabolic steps responsible for stereoselective disposition of nitrendipine enantiomers. Br J Clin Pharmacol. 1992, 33, 51-9.
[55] Kobayashi, H. and Kobayashi, S. Enantiomer-enantiomer interaction of a calcium channel antagonist, benidipine hydrochloride, during liver metabolism in the rat. EurJ Drug Metab Pharmacokinet. 1999, 24, 121-6.
[56] Tokuma, Y., Fujiwara, T., Niwa, T., Hashimoto, T. and Noguchi, H. Stereoselective disposition of nilvadipine, a new dihydropyridine calcium antagonist, in the rat and dog. Res Commun Chem Pathol Pharmacol. 1989, 63, 249-62.
[57] 李雷鸣, 高连勋. 对映体制备性分离方法的进展. 化学通报. 1997, 2, 17-21.
[58] Kiddle, J.J. Quebrachitol: A Versatile Building Block in the Construction of Naturally Occurring Bioactive Materials. Chem. Rev. 1995, 95, 2189-202.
[59] Taylor, M.S. and Jacobsen, E.N. Asymmetric catalysis in complex target synthesis. Proc Natl Acad Sci U S A. 2004, 101, 5368-73.
[60] Gawronski, J. Asymmetric syntheses and transformations--tools for chirality multiplication in drug synthesis. Acta Pol Pharm. 2006, 63, 333-51.
[61] Yun, O. Profile of William S. Knowles. Proc Natl Acad Sci U S A. 2005, 102, 16913-5.
[62] 刘二畅, 杨定乔, 董建霞. 现代手性药物的合成与发展. 广东医学院学报. 2004, 22, 665-8.
[63] Joguet, L., Sondi, I. and Matijevic, E. Preparation of nanosized drug particles by the coating of inorganic cores: naproxen and ketoprofen on alumina. J Colloid Interface Sci. 2002, 251, 284-7.
[64] Archelas, A. and Furstoss, R. Synthesis of enantiopure epoxides through biocatalytic approaches. Annu Rev Microbiol. 1997, 51, 491-525.
[65] Chirumamilla, R.R., Marchant, R. and Nigam, P. Captopril and its synthesis from chiral intermediates Journal of Chemical Technology & Biotechnology 2001, 76, 123 - 7.
[66] 胡健, 张乐之, 罗平, 李福涛, 周红. 手性药物消旋体拆分的研究进展. 西南国防医药. 2005, 15, 105-7.
[67] 尤田耙. 手性化合物的现代研究方法. 中国科技大学出版社. 1993. 北京.
[68] Xin, J.Y., Li, S.B., Xu, Y. and Wang, L.L. Enzymatic resolution of (S)-(+)-naproxen in a trapped aqueous-organic solvent biphase continuous reactor. Biotechnol Bioeng. 2000, 68, 78-83.
[69] Ahmar, M., Girard, C. and Block, R. Enzymatic resolution of methyl 2-alkyl-2-arylacetates. Tetrahedron letters. 1989, 30, 7053-6.
[70] 陈雄裕, 郭华云. 手性药物的液相色谱法分离. 海峡药学. 2000, 12, 105.
[71] 罗维早, 李章万, 张志荣. 手性药物的色谱分离进展. 华西药学杂志. 2000, 15, 295-8.
[72] 彭清涛, 胡文样, 谭生键. 药物对映体 HPLC 分离测定研究新进展. 药学学报. 1998, 33, 793-800.
[73] 侯经国, 周志强, 陈立仁. 高效液相色谱手性拆分中的“刷型”手性固定相. 色谱. 1997, 15, 206-11.
[74] 李坤, 王海荣, 王海, 杜玉民. 高效液相色谱法拆分 1,4-二氢-2,6-二甲基-4-(3-硝基苯基)-3,5-吡啶二羧酸单甲酯对映体. 色谱. 2006, 24, 318.
[75] Ranade, V.V. and Somberg, J.C. Chiral cardiovascular drugs: an overview. Am J Ther. 2005, 12, 439-59.
[76] 赵卫光, 陈寒松, 王素华, 李正名. 2,2,6-三甲基-4H-1,3-二噁烷-4-酮在有机合成中的应用. 精细化工原料及中间体. 2004, 11, 12-5.
[77] Satoh, Y., Okumura, K. and Shirokawa, Y. Studies on nilvadipine IV-synthesis of and deuterated and optically active isopropyl-2 cyano-3-methoxycarbaonyl-4-(3-nitrophe- nyl)-6-methyl 1,4 dihydroxy pyridine 5-carbox-ylate (Nilvadipine). Chem Pharm Bull (Tokyo). 1994, 42, 950–2.
[78] Scharpf, F., Riedel, K.-D., Laufen, H. and Leitold, M. Enantioselective gas chromatographic assay with electroncapture detection for amlodipine in biological samples. J Chromatogr 1994, 655, 225–33.
[79] Owens, P.K., Fell, A.F., Coleman, M.W. and Berridge, J.C. Method development in liquid chromatography with a charged cyclodextrin additive for chiral resolution of rac-amlodipine utilising a central composite design. Chirality. 1996, 8, 466-76.
[80] Uno, T., Ohkubo, T. and Sugawara, K. Enantioselective high-performance liquid chromatographic determination of nicardipine in human plasma. J Chromatogr. 1997, 698, 181–6.
[81] Yamaguchi, M., Yamashita, K., Aoki, I., Tabata, T., Hirai, S.-I. and Yashiki, T. Determination of manidipine enantiomers in human serum using chiral chromatography and column-switching liquid chromatography. J Chromatogr. 1992, 575, 123- 9.
[82] Fu, Q., Matsunaga, H. and Haginaka, J. Resolution of dihydropyridine calcium antagonist enantiomers using HPLC with ovoglycoprotein as a chiral stationary phase. Anal Sci. 2001, 17, 897-900.
[83] Boatto, G., Nieddu, M., Faedda, M.V. and de Caprariis, P. Enantiomeric separation by HPLC of 1,4-dihydropyridines with vancomycin as chiral selector. Chirality. 2003, 15, 494-7.
[84] 傅颖珺, 谢勇. 内毒素血症与 Toll 受体. 江西医学院学报. 2006, 46, 178-80.
[85] Cohen, J. The immunopathogenesis of sepsis. Nature. 2002, 420, 885-91.
[86] Thiru, Y., Pathan, N., Bignall, S., Habibi, P. and Levin, M. A myocardial cytotoxic process is involved in the cardiac dysfunction of meningococcal septic shock. Crit Care Med. 2000, 28, 2979-83.
[87] Turner, A., Tsamitros, M. and Bellomo, R. Myocardial cell injury in septic shock. Crit Care Med. 1999, 27, 1775-80.
[88] 许夕海, 施光峰. 感染性休克中脂多糖的信号传导机制研究进展. 中国抗感染化疗杂志. 2003, 3, 47-50.
[89] Tobias, P.S., Soldau, K. and Ulevitch, R.J. Isolation of a lipopolysaccharide-binding acute phase reactant from rabbit serum. J Exp Med. 1986, 164, 777-93.
[90] 赵虎, 周庭银, 孔宪涛. LPS 结合蛋白的结构和功能. 细胞与分子免疫学杂志. 2000, 16, 91-2.
[91] Wright, S.D., Ramos, R.A., Tobias, P.S., Ulevitch, R.J. and Mathison, J.C. CD14, a receptor for complexes of lipopolysaccharide (LPS) and LPS binding protein. Science. 1990, 249, 1431-3.
[92] Gupta, D., Kirkland, T.N., Viriyakosol, S. and Dziarski, R. CD14 is a cell-activating receptor for bacterial peptidoglycan. J Biol Chem. 1996, 271, 23310-6.
[93] Katz, S.S., Chen, K., Chen, S., Doerfler, M.E., Elsbach, P. and Weiss, J. Potent CD14-mediated signalling of human leukocytes by Escherichia coli can be mediated by interaction of whole bacteria and host cells without extensive prior release of endotoxin. Infect Immun. 1996, 64, 3592-600.
[94] 杨一新, 李桂源. LPS 所介导的信号转导通路研究进展. 中南大学学报(医学版). 2006, 31, 141-5.
[95] Hamann, L., Schumann, R.R., Flad, H.D., Brade, L., Rietschel, E.T. and Ulmer, A.J. Binding of lipopolysaccharide (LPS) to CHO cells does not correlate with LPS-induced NF-kappaB activation. Eur J Immunol. 2000, 30, 211-6.
[96] Medzhitov, R., Preston-Hurlburt, P. and Janeway, C.A., Jr. A human homologue of the Drosophila Toll protein signals activation of adaptive immunity. Nature. 1997,388, 394-7.
[97] Underhill, D.M., Ozinsky, A., Hajjar, A.M., Stevens, A., Wilson, C.B., Bassetti, M. and Aderem, A. The Toll-like receptor 2 is recruited to macrophage phagosomes and discriminates between pathogens. Nature. 1999, 401, 811-5.
[98] Underhill, D.M. and Ozinsky, A. Toll-like receptors: key mediators of microbe detection. Curr Opin Immunol. 2002, 14, 103-10.
[99] Guillot, L., Medjane, S., Le-Barillec, K., Balloy, V., Danel, C., Chignard, M. and Si-Tahar, M. Response of human pulmonary epithelial cells to lipopolysaccharide involves Toll-like receptor 4 (TLR4)-dependent signaling pathways: evidence for an intracellular compartmentalization of TLR4. J Biol Chem. 2004, 279, 2712-8.
[100] Monick, M.M., Yarovinsky, T.O., Powers, L.S., Butler, N.S., Carter, A.B., Gudmundsson, G. and Hunninghake, G.W. Respiratory syncytial virus up-regulates TLR4 and sensitizes airway epithelial cells to endotoxin. J Biol Chem. 2003, 278, 53035-44.
[101] Abreu, M.T., Vora, P., Faure, E., Thomas, L.S., Arnold, E.T. and Arditi, M. Decreased expression of Toll-like receptor-4 and MD-2 correlates with intestinal epithelial cell protection against dysregulated proinflammatory gene expression in response to bacterial lipopolysaccharide. J Immunol. 2001, 167, 1609-16.
[102] Dziarski, R., Wang, Q., Miyake, K., Kirschning, C.J. and Gupta, D. MD-2 enables Toll-like receptor 2 (TLR2)-mediated responses to lipopolysaccharide and enhances TLR2-mediated responses to Gram-positive and Gram-negative bacteria and their cell wall components. J Immunol. 2001, 166, 1938-44.
[103] Pfeiffer, A. et al. Lipopolysaccharide and ceramide docking to CD14 provokes ligand-specific receptor clustering in rafts. Eur J Immunol. 2001, 31, 3153-64.
[104] Fitzgerald, K.A. et al. Mal (MyD88-adapter-like) is required for Toll-like receptor-4 signal transduction. Nature. 2001, 413, 78-83.
[105] Muzio, M., Natoli, G., Saccani, S., Levrero, M. and Mantovani, A. The human toll signaling pathway: divergence of nuclear factor kappaB and JNK/SAPK activation upstream of tumor necrosis factor receptor-associated factor 6 (TRAF6). J Exp Med. 1998, 187, 2097-101.
[106] Horng, T., Barton, G.M. and Medzhitov, R. TIRAP: an adapter molecule in the Toll signaling pathway. Nat Immunol. 2001, 2, 835-41.
[107] Singh, H., Sen, R., Baltimore, D. and Sharp, P.A. A nuclear factor that binds to a conserved sequence motif in transcriptional control elements of immunoglobulin genes. Nature. 1986, 319, 154-8.
[108] Hayden, M.S. and Ghosh, S. Signaling to NF-kappaB. Genes Dev. 2004, 18, 2195-224.
[109] Sharif, O., Bolshakov, V.N., Raines, S., Newham, P. and Perkins, N.D. Transcriptional profiling of the LPS induced NF-kappaB response in macrophages. BMC Immunol. 2007, 8, 1.
[110] Dunn, K.L., Espino, P.S., Drobic, B., He, S. and Davie, J.R. The Ras-MAPK signal transduction pathway, cancer and chromatin remodeling. Biochem Cell Biol. 2005, 83, 1-14.
[111] Arndt, P.G., Suzuki, N., Avdi, N.J., Malcolm, K.C. and Worthen, G.S. Lipopolysaccharide-induced c-Jun NH2-terminal kinase activation in human neutrophils: role of phosphatidylinositol 3-Kinase and Syk-mediated pathways. J Biol Chem. 2004, 279, 10883-91.
[112] Schabbauer, G., Tencati, M., Pedersen, B., Pawlinski, R. and Mackman, N. PI3K-Aktpathway suppresses coagulation and inflammation in endotoxemic mice. Arterioscler Thromb Vasc Biol. 2004, 24, 1963-9.
[113] 吴文溪, 吴肇光. 外科基本问题 休克:低血容量性休克和感染性休克. 外科理论与实践. 2001, 6, 47-51.
[114] Karima, R., Matsumoto, S., Higashi, H. and Matsushima, K. The molecular pathogenesis of endotoxic shock and organ failure. Mol Med Today. 1999, 5, 123-32.
[115] Kumar, A., Thota, V., Dee, L., Olson, J., Uretz, E. and Parrillo, J.E. Tumor necrosis factor alpha and interleukin 1beta are responsible for in vitro myocardial cell depression induced by human septic shock serum. J Exp Med. 1996, 183, 949-58.
[116] Joe, E.K., Schussheim, A.E., Longrois, D., Maki, T., Kelly, R.A., Smith, T.W. and Balligand, J.L. Regulation of cardiac myocyte contractile function by inducible nitric oxide synthase (iNOS): mechanisms of contractile depression by nitric oxide. J Mol Cell Cardiol. 1998, 30, 303-15.
[117] Wang, X. and Evans, R.D. Effect of endotoxin and platelet-activating factor on lipid oxidation in the rat heart. J Mol Cell Cardiol. 1997, 29, 1915-26.
[118] Devaux, Y. et al. Lipopolysaccharide-induced increase of prostaglandin E(2) is mediated by inducible nitric oxide synthase activation of the constitutive cyclooxygenase and induction of membrane-associated prostaglandin E synthase. J Immunol. 2001, 167, 3962-71.
[119] Kapadia, S., Lee, J., Torre-Amione, G., Birdsall, H.H., Ma, T.S. and Mann, D.L. Tumor necrosis factor-alpha gene and protein expression in adult feline myocardium after endotoxin administration. J Clin Invest. 1995, 96, 1042-52.
[120] Comstock, K.L. et al. LPS-induced TNF-alpha release from and apoptosis in rat cardiomyocytes: obligatory role for CD14 in mediating the LPS response. J Mol CellCardiol. 1998, 30, 2761-75.
[121] Haudek, S.B., Spencer, E., Bryant, D.D., White, D.J., Maass, D., Horton, J.W., Chen, Z.J. and Giroir, B.P. Overexpression of cardiac I-kappaBalpha prevents endotoxin-induced myocardial dysfunction. Am J Physiol Heart Circ Physiol. 2001, 280, H962-8.
[122] Fauvel, H., Marchetti, P., Chopin, C., Formstecher, P. and Neviere, R. Differential effects of caspase inhibitors on endotoxin-induced myocardial dysfunction and heart apoptosis. Am J Physiol Heart Circ Physiol. 2001, 280, H1608-14.
[123] Stamm, C., Cowan, D.B., Friehs, I., Noria, S., del Nido, P.J. and McGowan, F.X., Jr. Rapid endotoxin-induced alterations in myocardial calcium handling: obligatory role of cardiac TNF-alpha. Anesthesiology. 2001, 95, 1396-405.
[124] Rauchhaus, M., Koloczek, V., Volk, H., Kemp, M., Niebauer, J., Francis, D.P., Coats, A.J. and Anker, S.D. Inflammatory cytokines and the possible immunological role for lipoproteins in chronic heart failure. Int J Cardiol. 2000, 76, 125-33.
[125] Oprins, J.C., Meijer, H.P. and Groot, J.A. TNF-alpha potentiates the ion secretion induced by muscarinic receptor activation in HT29cl.19A cells. Am J Physiol Cell Physiol. 2000, 278, C463-72.
[126] Dinarello, C.A. Proinflammatory cytokines. Chest. 2000, 118, 503-8.
[127] Pantano, C., Shrivastava, P., McElhinney, B. and Janssen-Heininger, Y. Hydrogen peroxide signaling through tumor necrosis factor receptor 1 leads to selective activation of c-Jun N-terminal kinase. J Biol Chem. 2003, 278, 44091-6.
[128] Pattle, S.B. and Farrell, P.J. The role of Epstein-Barr virus in cancer. Expert Opin Biol Ther. 2006, 6, 1193-205.
[129] 王永清, 樊寻梅. 脓毒症及感染性休克时心肌抑制的炎症介质机制. 小儿急救医学. 2005, 12, 151-3.
[130] Roig, E., Orus, J., Pare, C., Azqueta, M., Filella, X., Perez-Villa, F., Heras, M. and Sanz, G. Serum interleukin-6 in congestive heart failure secondary to idiopathic dilated cardiomyopathy. Am J Cardiol. 1998, 82, 688-90, A8.
[131] Schulz, R., Nava, E. and Moncada, S. Induction and potential biological relevance of a Ca2+-independent nitric oxide synthase in the myocardium. Br J Pharmacol. 1992, 105, 575-80.
[132] Gunnett, C.A., Chu, Y., Heistad, D.D., Loihl, A. and Faraci, F.M. Vascular effects of LPS in mice deficient in expression of the gene for inducible nitric oxide synthase. Am J Physiol. 1998, 275, H416-21.
[133] Wolfsgruber, W., Feil, S., Brummer, S., Kuppinger, O., Hofmann, F. and Feil, R. A proatherogenic role for cGMP-dependent protein kinase in vascular smooth muscle cells. Proc Natl Acad Sci U S A. 2003, 100, 13519-24.
[134] Kumar, A., Kosuri, R., Kandula, P., Dimou, C., Allen, J. and Parrillo, J.E. Effects of epinephrine and amrinone on contractility and cyclic adenosine monophosphate generation of tumor necrosis factor alpha-exposed cardiac myocytes. Crit Care Med. 1999, 27, 286-92.
[135] Iwase, M., Yokota, M., Kitaichi, K., Wang, L., Takagi, K., Nagasaka, T., Izawa, H. and Hasegawa, T. Cardiac functional and structural alterations induced by endotoxin in rats: importance of platelet-activating factor. Crit Care Med. 2001, 29, 609-17.
[136] Araujo, C.V., Barbosa-Filho, J.M., Cordeiro, R.S. and Tibirica, E. Protective effects of yangambin on cardiovascular hyporeactivity to catecholamines in rats with endotoxin-induced shock. Naunyn Schmiedebergs Arch Pharmacol. 2001, 363, 267-75.
[137] Elkayam, U., Amin, J., Mehra, A., Vasquez, J., Weber, L. and Rahimtoola, S.H. A prospective, randomized, double-blind, crossover study to compare the efficacy and safety of chronic nifedipine therapy with that of isosorbide dinitrate and their combination in the treatment of chronic congestive heart failure. Circulation. 1990, 82, 1954-61.
[138] Francis, G.S. Calcium channel blockers and congestive heart failure. Circulation. 1991, 83, 336-8.
[139] Packer, M. Pathophysiological mechanisms underlying the adverse effects of calcium channel-blocking drugs in patients with chronic heart failure. Circulation. 1989, 80, IV59-67.
[140] 胡大一, 刘晓惠. 评价钙拮抗剂在治疗慢性充血性心力衰竭中的地位. 高血压杂志. 1997, 5, 71-3.
[141] Packer, M. et al. Effect of amlodipine on morbidity and mortality in severe chronic heart failure. Prospective Randomized Amlodipine Survival Evaluation Study Group. N Engl J Med. 1996, 335, 1107-14.
[142] Lee, H.C. and Lum, B.K. Protective action of calcium entry blockers in endotoxin shock. Circ Shock. 1986, 18, 193-203.
[143] Veeraveedu, P.T. et al. Comparative effects of pranidipine with amlodipine in rats with heart failure. Pharmacology. 2006, 77, 1-10.
[144] Bravo, G., Kurtansky, A., López-Mu?oz, F.J., Hong, E., Rojas, G., Villalón, C.M. and Huang, F. Protective Action of Amlodipine on Cardiac Negative Inotropism Induced by Lipopolysaccharide in Rats. Basic & Clinical Pharmacology & Toxicology. 2007,
[145] Fukuzawa, M. et al. Modulation of tumor necrosis factor-alpha production with anti-hypertensive drugs. Immunopharmacology. 2000, 48, 65-74.
[146] 宋浩明, 罗明, 邓南伟, 等. 氨氯地平在心衰治疗中对肿瘤坏死因子-α、白细胞介素-1,6 的作用. 中国新药与临床杂志. 2002, 21, 661-3.
[147] 宋革, 苏海砾, 杜芳, 等. 新二氢吡啶类钙拮抗剂 MN9202 对实验性小鼠脑缺血的防护作用. 中国微循环. 2001, 5, 208-10.
[148] Yamamoto, C., Yashima, E. and Okamoto, Y. Computational studies on chiral discrimination mechanism of phenylcarbamate derivatives of cellulose. Bull. Chem.Soc. Jpn. 1999, 72, 1815-25.
[149] Zhao, M.G., Mei, Q.B., Zhang, Y.F., Xing, B., Li, X.Q. and Cui, Y. Inhibitory effects of serotonin on transient outward potassium current in rat ventricular myocytes. Acta Pharmacol Sin. 2002, 23, 617-22.
[150] Zhao, M.G., Mei, Q.B., Zhang, Y.F., Xiong, X.Y., Lu, B.H., Xing, B. and Zhao, D.H. 5-Hydroxytryptamine enhances L-type calcium current in norepinephrine-induced hypertrophic ventricular myocytes. Acta Pharmacol Sin. 2002, 23, 523-8.
[151] Ren, J. and Wold, L.E. Measurement of Cardiac Mechanical Function in Isolated Ventricular Myocytes from Rats and Mice by Computerized Video-Based Imaging. Biol Proced Online. 2001, 3, 43-53.
[152] Zhang, B., Zhang, H., Fan, Q., Ma, X. and Gao, F. Insulin improves cardiac myocytes contractile function recovery in simulated ischemia-reperfusion: key role of Akt. Chin Sci Bull 2003, 48, 1364-9.
[153] Sipido, K.R. and Callewaert, G. How to measure intracellular [Ca2+] in single cardiac cells with fura-2 or indo-1. Cardiovasc Res. 1995, 29, 717-26.
[154] Takahashi, A., Camacho, P., Lechleiter, J.D. and Herman, B. Measurement of intracellular calcium. Physiol Rev. 1999, 79, 1089-125.
[155] Molkentin, J.D. Dichotomy of Ca2+ in the heart: contraction versus intracellularsignaling. J Clin Invest. 2006, 116, 623-6.
[156] Adachi-Akahane, S. [Molecular and pharmacological bases for the gating regulation of L-type voltage-dependent Ca2+ channels]. Nippon Yakurigaku Zasshi. 2004, 123, 197-209.
[157] Takamatsu, H., Nagao, T., Ichijo, H. and Adachi-Akahane, S. L-type Ca2+ channels serve as a sensor of the SR Ca2+ for tuning the efficacy of Ca2+-induced Ca2+ release in rat ventricular myocytes. J Physiol. 2003, 552, 415-24.
[158] Xie, M.J., Zhang, L.F., Ma, J. and Cheng, H.W. Functional alterations in cerebrovascular K+ and Ca2+ channels are comparable between simulated microgravity rat and SHR. Am J Physiol Heart Circ Physiol. 2005, 289, H1265-76.
[159] 刘振伟. 实用膜片钳技术. 军事医学科学出版社. 2006.
[160] 康华光. 膜片钳技术及其应用. 科学出版社. 2003.
[161] Parrillo, J.E., Parker, M.M., Natanson, C., Suffredini, A.F., Danner, R.L., Cunnion, R.E. and Ognibene, F.P. Septic shock in humans. Advances in the understanding of pathogenesis, cardiovascular dysfunction, and therapy. Ann Intern Med. 1990, 113, 227-42.
[162] Jean-Baptiste, E. Cellular mechanisms in sepsis. J Intensive Care Med. 2007, 22, 63-72.
[163] Rice, T.W. Treatment of severe sepsis: where next? Current and future treatment approaches after the introduction of drotrecogin alfa. Vasc Health Risk Manag. 2006, 2, 3-18.
[164] Aggarwal, B.B. Signalling pathways of the TNF superfamily: a double-edged sword. Nat Rev Immunol. 2003, 3, 745-56.
[165] Ghezzi, P. and Cerami, A. Tumor necrosis factor as a pharmacological target. MolBiotechnol. 2005, 31, 239-44.
[166] Szabo, C., Mitchell, J.A., Gross, S.S., Thiemermann, C. and Vane, J.R. Nifedipine inhibits the induction of nitric oxide synthase by bacterial lipopolysaccharide. J Pharmacol Exp Ther. 1993, 265, 674-80.
[167] Haupt, W., Fritzsche, H., Hohenberger, W. and Zirngibl, H. Selective cytokine release induced by serum and separated plasma from septic patients. Eur J Surg. 1996, 162, 769-76.
[168] Arras, M., Hoche, A., Bohle, R., Eckert, P., Riedel, W. and Schaper, J. Tumor necrosis factor-alpha in macrophages of heart, liver, kidney, and in the pituitary gland. Cell Tissue Res. 1996, 285, 39-49.
[169] 荣仔萍, 常中飞, 宋欣伟. 肿瘤坏死因子与心力衰竭及抗细胞因子治疗研究进展. 医学综述. 2006, l2, 58-60.
[170] Chovolou, Y., Watjen, W., Kampkotter, A. and Kahl, R. Resistance to tumor necrosis factor-alpha (TNF-alpha)-induced apoptosis in rat hepatoma cells expressing TNF-alpha is linked to low antioxidant enzyme expression. J Biol Chem. 2003, 278, 29626-32.
[171] Kawano, H., Okada, R., Kawano, Y., Sueyoshi, N. and Shirai, T. Apoptosis in acute and chronic myocarditis. Jpn Heart J. 1994, 35, 745-50.
[172] 李世英, 倪新海, 陈曦, 等. 血管紧张素Ⅱ促进培养的新生鼠心肌细胞表达TNF-α. 高血压杂志. 2003, 11, 161-4.
[173] Carswell, E.A., Old, L.J., Kassel, R.L., Green, S., Fiore, N. and Williamson, B. An endotoxin-induced serum factor that causes necrosis of tumors. Proc Natl Acad Sci U S A. 1975, 72, 3666-70.
[174] 邱飞, 王礼琛. 肿瘤坏死因子 α 与心力衰竭的研究进展. 国外医学(药学分册).2003, 30, 326-30.
[175] Kubota, T., McTiernan, C.F., Frye, C.S., Slawson, S.E., Lemster, B.H., Koretsky, A.P., Demetris, A.J. and Feldman, A.M. Dilated cardiomyopathy in transgenic mice with cardiac-specific overexpression of tumor necrosis factor-alpha. Circ Res. 1997, 81, 627-35.
[176] Bozkurt, B. et al. Pathophysiologically relevant concentrations of tumor necrosis factor-alpha promote progressive left ventricular dysfunction and remodeling in rats. Circulation. 1998, 97, 1382-91.
[177] Kriegler, M., Perez, C., DeFay, K., Albert, I. and Lu, S.D. A novel form of TNF/cachectin is a cell surface cytotoxic transmembrane protein: ramifications for the complex physiology of TNF. Cell. 1988, 53, 45-53.
[178] 尹丙姣, 李卓娅, 余上斌, 等. 跨膜型和分泌型 TNF-α 在内毒素性休克过程中的作用. 中华微生物学和免疫学杂志. 2002, 22, 334-8.
[179] Torre-Amione, G., Kapadia, S., Lee, J., Durand, J.B., Bies, R.D., Young, J.B. and Mann, D.L. Tumor necrosis factor-alpha and tumor necrosis factor receptors in the failing human heart. Circulation. 1996, 93, 704-11.
[180] 陈齐红, 覃数. 肿瘤坏死因子-α 和充血性心力衰竭. 心血管病进展. 2004, 25, 89-91.
[181] Palladino, M.A., Bahjat, F.R., Theodorakis, E.A. and Moldawer, L.L. Anti-TNF-alpha therapies: the next generation. Nat Rev Drug Discov. 2003, 2, 736-46.
[182] Stuber, F. Effects of genomic polymorphisms on the course of sepsis: is there a concept for gene therapy? J Am Soc Nephrol. 2001, 12 Suppl 17, S60-4.
[183] Matsui, T. et al. Akt activation preserves cardiac function and prevents injury aftertransient cardiac ischemia in vivo. Circulation. 2001, 104, 330-5.
[184] Chen, K.Q., Iribarren, P., Gong, W.H. and Wang, J.M. The Essential Role of Phosphoinositide 3-Kinases (PI3Ks) in Regulating Pro-Inflammatory Responses and the Progression of Cancer. Cell Mol Immunol. 2005, 2, 241-52.
[185] Sambrook, J., Russell, D. (黄培堂等译). 分子克隆实验指南(第 3 版). 科学出版社. 2002, 北京
[186] Freyer, D. et al. Cerebral endothelial cells release TNF-alpha after stimulation with cell walls of Streptococcus pneumoniae and regulate inducible nitric oxide synthase and ICAM-1 expression via autocrine loops. J Immunol. 1999, 163, 4308-14.
[187] Fiorucci, S. et al. Relative contribution of acetylated cyclo-oxygenase (COX)-2 and 5-lipooxygenase (LOX) in regulating gastric mucosal integrity and adaptation to aspirin. Faseb J. 2003, 17, 1171-3.
[188] Knowles, R.G. and Moncada, S. Nitric oxide synthases in mammals. Biochem J. 1994, 298 ( Pt 2), 249-58.
[189] Gaston, B., Drazen, J.M., Loscalzo, J. and Stamler, J.S. The biology of nitrogen oxides in the airways. Am J Respir Crit Care Med. 1994, 149, 538-51.
[190] Forstermann, U., Closs, E.I., Pollock, J.S., Nakane, M., Schwarz, P., Gath, I. and Kleinert, H. Nitric oxide synthase isozymes. Characterization, purification, molecular cloning, and functions. Hypertension. 1994, 23, 1121-31.
[191] Hoit, B.D. Two faces of nitric oxide: lessons learned from the NOS2 knockout. Circ Res. 2001, 89, 289-91.
[192] Ullrich, R., Scherrer-Crosbie, M., Bloch, K.D., Ichinose, F., Nakajima, H., Picard, M.H., Zapol, W.M. and Quezado, Z.M. Congenital deficiency of nitric oxide synthase 2 protects against endotoxin-induced myocardial dysfunction in mice. Circulation.2000, 102, 1440-6.
[193] Goren, N., Cuenca, J., Martin-Sanz, P. and Bosca, L. Attenuation of NF-kappaB signalling in rat cardiomyocytes at birth restricts the induction of inflammatory genes. Cardiovasc Res. 2004, 64, 289-97.
[194] Schmedtje, J.F., Jr., Ji, Y.S., Liu, W.L., DuBois, R.N. and Runge, M.S. Hypoxia induces cyclooxygenase-2 via the NF-kappaB p65 transcription factor in human vascular endothelial cells. J Biol Chem. 1997, 272, 601-8.
[195] Lai, J. et al. Prostaglandin F2 alpha induces cardiac myocyte hypertrophy in vitro and cardiac growth in vivo. Am J Physiol. 1996, 271, H2197-208.
[196] Hocherl, K., Dreher, F., Kurtz, A. and Bucher, M. Cyclooxygenase-2 inhibition attenuates lipopolysaccharide-induced cardiovascular failure. Hypertension. 2002, 40, 947-53.
[197] Wong, S.C., Fukuchi, M., Melnyk, P., Rodger, I. and Giaid, A. Induction of cyclooxygenase-2 and activation of nuclear factor-kappaB in myocardium of patients with congestive heart failure. Circulation. 1998, 98, 100-3.
[198] Ejima, K. et al. Cyclooxygenase-2-deficient mice are resistant to endotoxin-induced inflammation and death. Faseb J. 2003, 17, 1325-7.
[199] Chou, T.C., Yang, S.P. and Pei, D. Amlodipine inhibits pro-inflammatory cytokines and free radical production and inducible nitric oxide synthase expression in lipopolysaccharide/interferon-gamma-stimulated cultured vascular smooth muscle cells. Jpn J Pharmacol. 2002, 89, 157-63.
[200] Sugimoto, Y., Whitman, M., Cantley, L.C. and Erikson, R.L. Evidence that the Rous sarcoma virus transforming gene product phosphorylates phosphatidylinositol and diacylglycerol. Proc Natl Acad Sci U S A. 1984, 81, 2117-21.
[201] Macara, I.G., Marinetti, G.V. and Balduzzi, P.C. Transforming protein of avian sarcoma virus UR2 is associated with phosphatidylinositol kinase activity: possible role in tumorigenesis. Proc Natl Acad Sci U S A. 1984, 81, 2728-32.
[202] Guha, M. and Mackman, N. The phosphatidylinositol 3-kinase-Akt pathway limits lipopolysaccharide activation of signaling pathways and expression of inflammatory mediators in human monocytic cells. J Biol Chem. 2002, 277, 32124-32.
[203] Fang, H., Pengal, R.A., Cao, X., Ganesan, L.P., Wewers, M.D., Marsh, C.B. and Tridandapani, S. Lipopolysaccharide-induced macrophage inflammatory response is regulated by SHIP. J Immunol. 2004, 173, 360-6.
[204] 尹华华, 夏浩, 程晓刚, 等. LPS 对小鼠肺组织及 RAW264. 7 细胞 Akt 蛋白表达及磷酸化的调控. 第三军医大学学报. 2007, 29, 188-190.
[205] Matsumori, A., Nunokawa, Y. and Sasayama, S. Nifedipine inhibits activation of transcription factor NF-kappaB. Life Sci. 2000, 67, 2655-61.
[2] 王剑波, 郭炜, 梅其炳, 等. 钙拮抗剂 MN9202 对心肌肥厚大鼠全血和心肌中5-HT 的影响. 第四军医大学学报. 2003, 24, 1688-90.
[3] 王莉莉, 梅其炳, 赵德化. 2,6-二甲基-4-(3-硝基苯基)-1,4-二氢-3,5-吡啶二羧酸-3-甲酯-5-正戊酯(MN9202)抗实验性血栓形成作用. 中国药学杂志. 1998, 33, 282-95.
[4] 姚秀娟, 甘洪泉, 贾敏, 等. 钙拮抗剂 MN9202 对高脂血症大鼠的影响. 第四军医大学学报. 2000, 21, 311-3.
[5] 甘戈, 梅其炳, 赵德化. MN9202 对兔小肠缺血再灌注损伤时血管内皮细胞功能的保护作用. 第四军医大学学报. 1998, 19, 571-3.
[6] 李新立. 钙拮抗剂的分类及其在高血压治疗中的选用. 高血压杂志. 1999, 7, 72-5.
[7] De Camp, W.H. Chiral drugs: the FDA perspective on manufacturing and control. J Pharm Biomed Anal. 1993, 11, 1167-72.
[8] 王普善, 王宇梅. 手性药物开发战略的再认识. 精细与专用化学品. 2004, 12, 4-8.
[9] 王丹, 李亚, 何浪. 手性药物及其开发与应用. 现代医药卫生. 2007, 23, 837-8.
[10] Assmann, I., Dittrich, P., Schwela, H. and Fiehring, H. [Clinico-pharmacological studies with Nifedipine--a new coronary substance]. Z Gesamte Inn Med. 1975, 30, 575-9.
[11] Triggle, D.J. 1,4-Dihydropyridines as calcium channel ligands and privileged structures. Cell Mol Neurobiol. 2003, 23, 293-303.
[12] Triggle, D.J. Calcium antagonists. History and perspective. Stroke. 1990, 21,IV49-58.
[13] Triggle, D.J. Calcium channel antagonists: Clinical uses-Past, present and future. Biochem Pharmacol. 2007 Jan 13; [Epub ahead of print]
[14] Grossman, E. and Messerli, F.H. Calcium antagonists. Prog Cardiovasc Dis. 2004, 47, 34-57.
[15] Leenen, F.H. Clinical relevance of 24 h blood pressure control by 1,4-dihydropyridines. Am J Hypertens. 1996, 9, 97S-104S; discussion 108S-109S.
[16] Wenzel, R.R., Allegranza, G., Binggeli, C., Shaw, S., Weidmann, P., Luscher, T.F. and Noll, G. Differential activation of cardiac and peripheral sympathetic nervous system by nifedipine: role of pharmacokinetics. J Am Coll Cardiol. 1997, 29, 1607-14.
[17] Haria, M. and Wagstaff, A.J. Amlodipine. A reappraisal of its pharmacological properties and therapeutic use in cardiovascular disease. Drugs. 1995, 50, 560-86.
[18] Motomura, S., Wu, Z.J. and Hashimoto, K. Lacidipine, a new long-acting dihydropyridine calcium antagonist, has high vascular selectivity against all intracardiac variables. Heart Vessels. 1993, 8, 16-22.
[19] Shan, R., Velazquez, C. and Knaus, E.E. Syntheses, calcium channel agonist-antagonist modulation activities, and nitric oxide release studies of nitrooxyalkyl 1,4-dihydro-2,6-dimethyl-3-nitro-4-(2,1,3-benzoxadiazol-4-yl)pyridine-5-ca rboxylate racemates, enantiomers, and diastereomers. J Med Chem. 2004, 47, 254-61.
[20] Ariens, E.J. Stereochemistry: a source of problems in medicinal chemistry. Med Res Rev. 1986, 6, 451-66.
[21] Inotsume, N. and Nakano, M. Stereoselective determination and pharmacokinetics of dihydropyridines: an updated review. J Biochem Biophys Methods. 2002, 54, 255-74.
[22] Millership, J.S. and Fitzpatrick, A. Commonly used chiral drugs: a survey. Chirality. 1993, 5, 573-6.
[23] Nerurkar, S.G., Dighe, S.V. and Williams, R.L. Bioequivalence of racemic drugs. J Clin Pharmacol. 1992, 32, 935-43.
[24] Bischer, A., Zia-Amirhosseini, P., Iwaki, M., McDonagh, A.F. and Benet, L.Z. Stereoselective binding properties of naproxen glucuronide diastereomers to proteins. J Pharmacokinet Biopharm. 1995, 23, 379-95.
[25] 曾苏. 手性药物与手性药理学. 浙江大学出版社. 2002
[26] Carter, R.B. and Dykstra, L.A. Effects of picenadol (LY150720) and its stereoisomers on electric shock titration in the squirrel monkey. J Pharmacol Exp Ther. 1985, 234, 299-306.
[27] Middlefell, V.C. and Price, T.L. 5-HT3 receptor agonism may be responsible for the emetic effects of zacopride in the ferret. Br J Pharmacol. 1991, 103, 1011-2.
[28] Tabi, T., Magyar, K. and Szoko, E. Chiral characterization of deprenyl-N-oxide and other deprenyl metabolites by capillary electrophoresis using a dual cyclodextrin system in rat urine. Electrophoresis. 2003, 24, 2665-73.
[29] Kean, W.F., Lock, C.J. and Howard-Lock, H.E. Chirality in antirheumatic drugs. Lancet. 1991, 338, 1565-8.
[30] Jamali, F., Mehvar, R. and Pasutto, F.M. Enantioselective aspects of drug action and disposition: therapeutic pitfalls. J Pharm Sci. 1989, 78, 695-715.
[31] 余细勇, 林曙光. 手性药物对映体的药动学与药效学立体选择性差异研究进展. 广州医药. 1997, 28, 1-3.
[32] Lane, R.M. and Baker, G.B. Chirality and drugs used in psychiatry: nice to know or need to know? Cell Mol Neurobiol. 1999, 19, 355-72.
[33] 李高. 手性药物的临床药物动力学立体选择性. 中国医院药学杂志. 1999, 19, 429-31.
[34] Bode, W., Toet, A.E., Stolker, A.A., van Ginkel, L.A., Groen, K., Wemer, J. and de Wildt, D.J. Toxicokinetics of a single intravenous dose of rac-propranolol versus optically pure propranolol in the rat. Chirality. 1995, 7, 626-31.
[35] Caldwell, J., Hutt, A.J. and Fournel-Gigleux, S. The metabolic chiral inversion and dispositional enantioselectivity of the 2-arylpropionic acids and their biological consequences. Biochem Pharmacol. 1988, 37, 105-14.
[36] Lee, E.J. and Williams, K.M. Chirality. Clinical pharmacokinetic and pharmacodynamic considerations. Clin Pharmacokinet. 1990, 18, 339-45.
[37] 章立, 姚彤炜, 曾苏. 手性药物对映体的药物代谢动力学. 中国现代应用药学杂志. 1999, 16, 4-7.
[38] Schramm, M., Thomas, G., Towart, R. and Franckowiak, G. Novel dihydropyridines with positive inotropic action through activation of Ca2+ channels. Nature. 1983, 303, 535-7.
[39] Muto, K., Kuroda, T., Kawato, H., Karasawa, A., Kubo, K. and Nakamizo, N. Synthesis and pharmacological activity of stereoisomers of 1,4-dihydro-2,6-dimethyl-4-(3-nitrophenyl)-3,5-pyridine-dicarboxylic acid methyl 1-(phenylmethyl)-3-piperidinyl ester. Arzneimittelforschung. 1988, 38, 1662-5.
[40] Zhang, X.P., Loke, K.E., Mital, S., Chahwala, S. and Hintze, T.H. Paradoxical release of nitric oxide by an L-type calcium channel antagonist, the R+ enantiomer of amlodipine. J Cardiovasc Pharmacol. 2002, 39, 208-14.
[41] Berjukow, S. and Hering, S. Voltage-dependent acceleration of Ca(v)1.2 channel current decay by (+)- and (-)-isradipine. Br J Pharmacol. 2001, 133, 959-66.
[42] Furukawa, T., Miura, R., Honda, M., Kamiya, N., Mori, Y., Takeshita, S., Isshiki, T. and Nukada, T. Identification of R(-)-isomer of efonidipine as a selective blocker of T-type Ca2+ channels. Br J Pharmacol. 2004, 143, 1050-7.
[43] Jabor, V.A., Coelho, E.B. and Lanchote, V.L. Enantioselective pharmacokinetics of lercanidipine in healthy volunteers. J Chromatogr B Analyt Technol Biomed Life Sci. 2004, 813, 343-6.
[44] Hirano, T. et al. Differential properties of the optical-isomers of pranidipine, a 1,4-dihydropyridine calcium channel modulator. Fundam Clin Pharmacol. 1999, 13, 650-5.
[45] Okumura, K., Ichihara, K., Nagasaka, M., Oda, N. and Tajima, K. Calcium entry blocking activities of MPC-1304 and of its enantiomers and metabolites. Eur J Pharmacol. 1993, 235, 69-74.
[46] Wang, S., He, L. and Yun, B. The difference between nicardipine and its enantiomers on inhibiting vasoconstriction of isolated rabbit thoracic artery. Arch Pharm Res. 2005, 28, 319-24.
[47] Vo, D., Matowe, W.C., Ramesh, M., Iqbal, N., Wolowyk, M.W., Howlett, S.E. and Knaus, E.E. Syntheses, calcium channel agonist-antagonist modulation activities, and voltage-clamp studies of isopropyl 1,4-dihydro-2,6-dimethyl-3-nitro-4-pyridinylpyrid- ine-5-carboxylate racemates and enantiomers. J Med Chem. 1995, 38, 2851-9.
[48] Hof, R.P., Ruegg, U.T., Hof, A. and Vogel, A. Stereoselectivity at the calcium channel: opposite action of the enantiomers of a 1,4-dihydropyridine. J Cardiovasc Pharmacol. 1985, 7, 689-93.
[49] Tokuma, Y., Fujiwara, T. and Noguchi, H. Determination of (+)- and (-)-nilvadipinein human plasma using chiral stationary-phase liquid chromatography and gas chromatography-mass spectrometry, and a preliminary pharmacokinetic study in humans. J Pharm Sci. 1987, 76, 310-3.
[50] Uno, T., Ohkubo, T., Motomura, S. and Sugawara, K. Effect of grapefruit juice on the disposition of manidipine enantiomers in healthy subjects. Br J Clin Pharmacol. 2006, 61, 533-7.
[51] Marques, M.P., Santos, N.A., Coelho, E.B., Bonato, P.S. and Lanchote, V.L. Enantioselective assay of nisoldipine in human plasma by chiral high-performance liquid chromatography combined with gas chromatographic-mass spectrometry: applications to pharmacokinetics. J Chromatogr B Biomed Sci Appl. 2001, 762, 87-95.
[52] He, L.C. and Wang, S.C. Studies on differences of pharmacokinetic behavior and tissue distribution of nimodipine and its two enantiomers in rats using achiral and chiral liquid chromatography. Yao Xue Xue Bao. 2003, 38, 603-8.
[53] Fischer, C., Schonberger, F., Muck, W., Heuck, K. and Eichelbaum, M. Simultaneous assessment of the intravenous and oral disposition of the enantiomers of racemic nimodipine by chiral stationary-phase high-performance liquid chromatography and gas chromatography/mass spectroscopy combined with a stable isotope technique. J Pharm Sci. 1993, 82, 244-50.
[54] Mast, V., Fischer, C., Mikus, G. and Eichelbaum, M. Use of pseudoracemic nitrendipine to elucidate the metabolic steps responsible for stereoselective disposition of nitrendipine enantiomers. Br J Clin Pharmacol. 1992, 33, 51-9.
[55] Kobayashi, H. and Kobayashi, S. Enantiomer-enantiomer interaction of a calcium channel antagonist, benidipine hydrochloride, during liver metabolism in the rat. EurJ Drug Metab Pharmacokinet. 1999, 24, 121-6.
[56] Tokuma, Y., Fujiwara, T., Niwa, T., Hashimoto, T. and Noguchi, H. Stereoselective disposition of nilvadipine, a new dihydropyridine calcium antagonist, in the rat and dog. Res Commun Chem Pathol Pharmacol. 1989, 63, 249-62.
[57] 李雷鸣, 高连勋. 对映体制备性分离方法的进展. 化学通报. 1997, 2, 17-21.
[58] Kiddle, J.J. Quebrachitol: A Versatile Building Block in the Construction of Naturally Occurring Bioactive Materials. Chem. Rev. 1995, 95, 2189-202.
[59] Taylor, M.S. and Jacobsen, E.N. Asymmetric catalysis in complex target synthesis. Proc Natl Acad Sci U S A. 2004, 101, 5368-73.
[60] Gawronski, J. Asymmetric syntheses and transformations--tools for chirality multiplication in drug synthesis. Acta Pol Pharm. 2006, 63, 333-51.
[61] Yun, O. Profile of William S. Knowles. Proc Natl Acad Sci U S A. 2005, 102, 16913-5.
[62] 刘二畅, 杨定乔, 董建霞. 现代手性药物的合成与发展. 广东医学院学报. 2004, 22, 665-8.
[63] Joguet, L., Sondi, I. and Matijevic, E. Preparation of nanosized drug particles by the coating of inorganic cores: naproxen and ketoprofen on alumina. J Colloid Interface Sci. 2002, 251, 284-7.
[64] Archelas, A. and Furstoss, R. Synthesis of enantiopure epoxides through biocatalytic approaches. Annu Rev Microbiol. 1997, 51, 491-525.
[65] Chirumamilla, R.R., Marchant, R. and Nigam, P. Captopril and its synthesis from chiral intermediates Journal of Chemical Technology & Biotechnology 2001, 76, 123 - 7.
[66] 胡健, 张乐之, 罗平, 李福涛, 周红. 手性药物消旋体拆分的研究进展. 西南国防医药. 2005, 15, 105-7.
[67] 尤田耙. 手性化合物的现代研究方法. 中国科技大学出版社. 1993. 北京.
[68] Xin, J.Y., Li, S.B., Xu, Y. and Wang, L.L. Enzymatic resolution of (S)-(+)-naproxen in a trapped aqueous-organic solvent biphase continuous reactor. Biotechnol Bioeng. 2000, 68, 78-83.
[69] Ahmar, M., Girard, C. and Block, R. Enzymatic resolution of methyl 2-alkyl-2-arylacetates. Tetrahedron letters. 1989, 30, 7053-6.
[70] 陈雄裕, 郭华云. 手性药物的液相色谱法分离. 海峡药学. 2000, 12, 105.
[71] 罗维早, 李章万, 张志荣. 手性药物的色谱分离进展. 华西药学杂志. 2000, 15, 295-8.
[72] 彭清涛, 胡文样, 谭生键. 药物对映体 HPLC 分离测定研究新进展. 药学学报. 1998, 33, 793-800.
[73] 侯经国, 周志强, 陈立仁. 高效液相色谱手性拆分中的“刷型”手性固定相. 色谱. 1997, 15, 206-11.
[74] 李坤, 王海荣, 王海, 杜玉民. 高效液相色谱法拆分 1,4-二氢-2,6-二甲基-4-(3-硝基苯基)-3,5-吡啶二羧酸单甲酯对映体. 色谱. 2006, 24, 318.
[75] Ranade, V.V. and Somberg, J.C. Chiral cardiovascular drugs: an overview. Am J Ther. 2005, 12, 439-59.
[76] 赵卫光, 陈寒松, 王素华, 李正名. 2,2,6-三甲基-4H-1,3-二噁烷-4-酮在有机合成中的应用. 精细化工原料及中间体. 2004, 11, 12-5.
[77] Satoh, Y., Okumura, K. and Shirokawa, Y. Studies on nilvadipine IV-synthesis of and deuterated and optically active isopropyl-2 cyano-3-methoxycarbaonyl-4-(3-nitrophe- nyl)-6-methyl 1,4 dihydroxy pyridine 5-carbox-ylate (Nilvadipine). Chem Pharm Bull (Tokyo). 1994, 42, 950–2.
[78] Scharpf, F., Riedel, K.-D., Laufen, H. and Leitold, M. Enantioselective gas chromatographic assay with electroncapture detection for amlodipine in biological samples. J Chromatogr 1994, 655, 225–33.
[79] Owens, P.K., Fell, A.F., Coleman, M.W. and Berridge, J.C. Method development in liquid chromatography with a charged cyclodextrin additive for chiral resolution of rac-amlodipine utilising a central composite design. Chirality. 1996, 8, 466-76.
[80] Uno, T., Ohkubo, T. and Sugawara, K. Enantioselective high-performance liquid chromatographic determination of nicardipine in human plasma. J Chromatogr. 1997, 698, 181–6.
[81] Yamaguchi, M., Yamashita, K., Aoki, I., Tabata, T., Hirai, S.-I. and Yashiki, T. Determination of manidipine enantiomers in human serum using chiral chromatography and column-switching liquid chromatography. J Chromatogr. 1992, 575, 123- 9.
[82] Fu, Q., Matsunaga, H. and Haginaka, J. Resolution of dihydropyridine calcium antagonist enantiomers using HPLC with ovoglycoprotein as a chiral stationary phase. Anal Sci. 2001, 17, 897-900.
[83] Boatto, G., Nieddu, M., Faedda, M.V. and de Caprariis, P. Enantiomeric separation by HPLC of 1,4-dihydropyridines with vancomycin as chiral selector. Chirality. 2003, 15, 494-7.
[84] 傅颖珺, 谢勇. 内毒素血症与 Toll 受体. 江西医学院学报. 2006, 46, 178-80.
[85] Cohen, J. The immunopathogenesis of sepsis. Nature. 2002, 420, 885-91.
[86] Thiru, Y., Pathan, N., Bignall, S., Habibi, P. and Levin, M. A myocardial cytotoxic process is involved in the cardiac dysfunction of meningococcal septic shock. Crit Care Med. 2000, 28, 2979-83.
[87] Turner, A., Tsamitros, M. and Bellomo, R. Myocardial cell injury in septic shock. Crit Care Med. 1999, 27, 1775-80.
[88] 许夕海, 施光峰. 感染性休克中脂多糖的信号传导机制研究进展. 中国抗感染化疗杂志. 2003, 3, 47-50.
[89] Tobias, P.S., Soldau, K. and Ulevitch, R.J. Isolation of a lipopolysaccharide-binding acute phase reactant from rabbit serum. J Exp Med. 1986, 164, 777-93.
[90] 赵虎, 周庭银, 孔宪涛. LPS 结合蛋白的结构和功能. 细胞与分子免疫学杂志. 2000, 16, 91-2.
[91] Wright, S.D., Ramos, R.A., Tobias, P.S., Ulevitch, R.J. and Mathison, J.C. CD14, a receptor for complexes of lipopolysaccharide (LPS) and LPS binding protein. Science. 1990, 249, 1431-3.
[92] Gupta, D., Kirkland, T.N., Viriyakosol, S. and Dziarski, R. CD14 is a cell-activating receptor for bacterial peptidoglycan. J Biol Chem. 1996, 271, 23310-6.
[93] Katz, S.S., Chen, K., Chen, S., Doerfler, M.E., Elsbach, P. and Weiss, J. Potent CD14-mediated signalling of human leukocytes by Escherichia coli can be mediated by interaction of whole bacteria and host cells without extensive prior release of endotoxin. Infect Immun. 1996, 64, 3592-600.
[94] 杨一新, 李桂源. LPS 所介导的信号转导通路研究进展. 中南大学学报(医学版). 2006, 31, 141-5.
[95] Hamann, L., Schumann, R.R., Flad, H.D., Brade, L., Rietschel, E.T. and Ulmer, A.J. Binding of lipopolysaccharide (LPS) to CHO cells does not correlate with LPS-induced NF-kappaB activation. Eur J Immunol. 2000, 30, 211-6.
[96] Medzhitov, R., Preston-Hurlburt, P. and Janeway, C.A., Jr. A human homologue of the Drosophila Toll protein signals activation of adaptive immunity. Nature. 1997,388, 394-7.
[97] Underhill, D.M., Ozinsky, A., Hajjar, A.M., Stevens, A., Wilson, C.B., Bassetti, M. and Aderem, A. The Toll-like receptor 2 is recruited to macrophage phagosomes and discriminates between pathogens. Nature. 1999, 401, 811-5.
[98] Underhill, D.M. and Ozinsky, A. Toll-like receptors: key mediators of microbe detection. Curr Opin Immunol. 2002, 14, 103-10.
[99] Guillot, L., Medjane, S., Le-Barillec, K., Balloy, V., Danel, C., Chignard, M. and Si-Tahar, M. Response of human pulmonary epithelial cells to lipopolysaccharide involves Toll-like receptor 4 (TLR4)-dependent signaling pathways: evidence for an intracellular compartmentalization of TLR4. J Biol Chem. 2004, 279, 2712-8.
[100] Monick, M.M., Yarovinsky, T.O., Powers, L.S., Butler, N.S., Carter, A.B., Gudmundsson, G. and Hunninghake, G.W. Respiratory syncytial virus up-regulates TLR4 and sensitizes airway epithelial cells to endotoxin. J Biol Chem. 2003, 278, 53035-44.
[101] Abreu, M.T., Vora, P., Faure, E., Thomas, L.S., Arnold, E.T. and Arditi, M. Decreased expression of Toll-like receptor-4 and MD-2 correlates with intestinal epithelial cell protection against dysregulated proinflammatory gene expression in response to bacterial lipopolysaccharide. J Immunol. 2001, 167, 1609-16.
[102] Dziarski, R., Wang, Q., Miyake, K., Kirschning, C.J. and Gupta, D. MD-2 enables Toll-like receptor 2 (TLR2)-mediated responses to lipopolysaccharide and enhances TLR2-mediated responses to Gram-positive and Gram-negative bacteria and their cell wall components. J Immunol. 2001, 166, 1938-44.
[103] Pfeiffer, A. et al. Lipopolysaccharide and ceramide docking to CD14 provokes ligand-specific receptor clustering in rafts. Eur J Immunol. 2001, 31, 3153-64.
[104] Fitzgerald, K.A. et al. Mal (MyD88-adapter-like) is required for Toll-like receptor-4 signal transduction. Nature. 2001, 413, 78-83.
[105] Muzio, M., Natoli, G., Saccani, S., Levrero, M. and Mantovani, A. The human toll signaling pathway: divergence of nuclear factor kappaB and JNK/SAPK activation upstream of tumor necrosis factor receptor-associated factor 6 (TRAF6). J Exp Med. 1998, 187, 2097-101.
[106] Horng, T., Barton, G.M. and Medzhitov, R. TIRAP: an adapter molecule in the Toll signaling pathway. Nat Immunol. 2001, 2, 835-41.
[107] Singh, H., Sen, R., Baltimore, D. and Sharp, P.A. A nuclear factor that binds to a conserved sequence motif in transcriptional control elements of immunoglobulin genes. Nature. 1986, 319, 154-8.
[108] Hayden, M.S. and Ghosh, S. Signaling to NF-kappaB. Genes Dev. 2004, 18, 2195-224.
[109] Sharif, O., Bolshakov, V.N., Raines, S., Newham, P. and Perkins, N.D. Transcriptional profiling of the LPS induced NF-kappaB response in macrophages. BMC Immunol. 2007, 8, 1.
[110] Dunn, K.L., Espino, P.S., Drobic, B., He, S. and Davie, J.R. The Ras-MAPK signal transduction pathway, cancer and chromatin remodeling. Biochem Cell Biol. 2005, 83, 1-14.
[111] Arndt, P.G., Suzuki, N., Avdi, N.J., Malcolm, K.C. and Worthen, G.S. Lipopolysaccharide-induced c-Jun NH2-terminal kinase activation in human neutrophils: role of phosphatidylinositol 3-Kinase and Syk-mediated pathways. J Biol Chem. 2004, 279, 10883-91.
[112] Schabbauer, G., Tencati, M., Pedersen, B., Pawlinski, R. and Mackman, N. PI3K-Aktpathway suppresses coagulation and inflammation in endotoxemic mice. Arterioscler Thromb Vasc Biol. 2004, 24, 1963-9.
[113] 吴文溪, 吴肇光. 外科基本问题 休克:低血容量性休克和感染性休克. 外科理论与实践. 2001, 6, 47-51.
[114] Karima, R., Matsumoto, S., Higashi, H. and Matsushima, K. The molecular pathogenesis of endotoxic shock and organ failure. Mol Med Today. 1999, 5, 123-32.
[115] Kumar, A., Thota, V., Dee, L., Olson, J., Uretz, E. and Parrillo, J.E. Tumor necrosis factor alpha and interleukin 1beta are responsible for in vitro myocardial cell depression induced by human septic shock serum. J Exp Med. 1996, 183, 949-58.
[116] Joe, E.K., Schussheim, A.E., Longrois, D., Maki, T., Kelly, R.A., Smith, T.W. and Balligand, J.L. Regulation of cardiac myocyte contractile function by inducible nitric oxide synthase (iNOS): mechanisms of contractile depression by nitric oxide. J Mol Cell Cardiol. 1998, 30, 303-15.
[117] Wang, X. and Evans, R.D. Effect of endotoxin and platelet-activating factor on lipid oxidation in the rat heart. J Mol Cell Cardiol. 1997, 29, 1915-26.
[118] Devaux, Y. et al. Lipopolysaccharide-induced increase of prostaglandin E(2) is mediated by inducible nitric oxide synthase activation of the constitutive cyclooxygenase and induction of membrane-associated prostaglandin E synthase. J Immunol. 2001, 167, 3962-71.
[119] Kapadia, S., Lee, J., Torre-Amione, G., Birdsall, H.H., Ma, T.S. and Mann, D.L. Tumor necrosis factor-alpha gene and protein expression in adult feline myocardium after endotoxin administration. J Clin Invest. 1995, 96, 1042-52.
[120] Comstock, K.L. et al. LPS-induced TNF-alpha release from and apoptosis in rat cardiomyocytes: obligatory role for CD14 in mediating the LPS response. J Mol CellCardiol. 1998, 30, 2761-75.
[121] Haudek, S.B., Spencer, E., Bryant, D.D., White, D.J., Maass, D., Horton, J.W., Chen, Z.J. and Giroir, B.P. Overexpression of cardiac I-kappaBalpha prevents endotoxin-induced myocardial dysfunction. Am J Physiol Heart Circ Physiol. 2001, 280, H962-8.
[122] Fauvel, H., Marchetti, P., Chopin, C., Formstecher, P. and Neviere, R. Differential effects of caspase inhibitors on endotoxin-induced myocardial dysfunction and heart apoptosis. Am J Physiol Heart Circ Physiol. 2001, 280, H1608-14.
[123] Stamm, C., Cowan, D.B., Friehs, I., Noria, S., del Nido, P.J. and McGowan, F.X., Jr. Rapid endotoxin-induced alterations in myocardial calcium handling: obligatory role of cardiac TNF-alpha. Anesthesiology. 2001, 95, 1396-405.
[124] Rauchhaus, M., Koloczek, V., Volk, H., Kemp, M., Niebauer, J., Francis, D.P., Coats, A.J. and Anker, S.D. Inflammatory cytokines and the possible immunological role for lipoproteins in chronic heart failure. Int J Cardiol. 2000, 76, 125-33.
[125] Oprins, J.C., Meijer, H.P. and Groot, J.A. TNF-alpha potentiates the ion secretion induced by muscarinic receptor activation in HT29cl.19A cells. Am J Physiol Cell Physiol. 2000, 278, C463-72.
[126] Dinarello, C.A. Proinflammatory cytokines. Chest. 2000, 118, 503-8.
[127] Pantano, C., Shrivastava, P., McElhinney, B. and Janssen-Heininger, Y. Hydrogen peroxide signaling through tumor necrosis factor receptor 1 leads to selective activation of c-Jun N-terminal kinase. J Biol Chem. 2003, 278, 44091-6.
[128] Pattle, S.B. and Farrell, P.J. The role of Epstein-Barr virus in cancer. Expert Opin Biol Ther. 2006, 6, 1193-205.
[129] 王永清, 樊寻梅. 脓毒症及感染性休克时心肌抑制的炎症介质机制. 小儿急救医学. 2005, 12, 151-3.
[130] Roig, E., Orus, J., Pare, C., Azqueta, M., Filella, X., Perez-Villa, F., Heras, M. and Sanz, G. Serum interleukin-6 in congestive heart failure secondary to idiopathic dilated cardiomyopathy. Am J Cardiol. 1998, 82, 688-90, A8.
[131] Schulz, R., Nava, E. and Moncada, S. Induction and potential biological relevance of a Ca2+-independent nitric oxide synthase in the myocardium. Br J Pharmacol. 1992, 105, 575-80.
[132] Gunnett, C.A., Chu, Y., Heistad, D.D., Loihl, A. and Faraci, F.M. Vascular effects of LPS in mice deficient in expression of the gene for inducible nitric oxide synthase. Am J Physiol. 1998, 275, H416-21.
[133] Wolfsgruber, W., Feil, S., Brummer, S., Kuppinger, O., Hofmann, F. and Feil, R. A proatherogenic role for cGMP-dependent protein kinase in vascular smooth muscle cells. Proc Natl Acad Sci U S A. 2003, 100, 13519-24.
[134] Kumar, A., Kosuri, R., Kandula, P., Dimou, C., Allen, J. and Parrillo, J.E. Effects of epinephrine and amrinone on contractility and cyclic adenosine monophosphate generation of tumor necrosis factor alpha-exposed cardiac myocytes. Crit Care Med. 1999, 27, 286-92.
[135] Iwase, M., Yokota, M., Kitaichi, K., Wang, L., Takagi, K., Nagasaka, T., Izawa, H. and Hasegawa, T. Cardiac functional and structural alterations induced by endotoxin in rats: importance of platelet-activating factor. Crit Care Med. 2001, 29, 609-17.
[136] Araujo, C.V., Barbosa-Filho, J.M., Cordeiro, R.S. and Tibirica, E. Protective effects of yangambin on cardiovascular hyporeactivity to catecholamines in rats with endotoxin-induced shock. Naunyn Schmiedebergs Arch Pharmacol. 2001, 363, 267-75.
[137] Elkayam, U., Amin, J., Mehra, A., Vasquez, J., Weber, L. and Rahimtoola, S.H. A prospective, randomized, double-blind, crossover study to compare the efficacy and safety of chronic nifedipine therapy with that of isosorbide dinitrate and their combination in the treatment of chronic congestive heart failure. Circulation. 1990, 82, 1954-61.
[138] Francis, G.S. Calcium channel blockers and congestive heart failure. Circulation. 1991, 83, 336-8.
[139] Packer, M. Pathophysiological mechanisms underlying the adverse effects of calcium channel-blocking drugs in patients with chronic heart failure. Circulation. 1989, 80, IV59-67.
[140] 胡大一, 刘晓惠. 评价钙拮抗剂在治疗慢性充血性心力衰竭中的地位. 高血压杂志. 1997, 5, 71-3.
[141] Packer, M. et al. Effect of amlodipine on morbidity and mortality in severe chronic heart failure. Prospective Randomized Amlodipine Survival Evaluation Study Group. N Engl J Med. 1996, 335, 1107-14.
[142] Lee, H.C. and Lum, B.K. Protective action of calcium entry blockers in endotoxin shock. Circ Shock. 1986, 18, 193-203.
[143] Veeraveedu, P.T. et al. Comparative effects of pranidipine with amlodipine in rats with heart failure. Pharmacology. 2006, 77, 1-10.
[144] Bravo, G., Kurtansky, A., López-Mu?oz, F.J., Hong, E., Rojas, G., Villalón, C.M. and Huang, F. Protective Action of Amlodipine on Cardiac Negative Inotropism Induced by Lipopolysaccharide in Rats. Basic & Clinical Pharmacology & Toxicology. 2007,
[145] Fukuzawa, M. et al. Modulation of tumor necrosis factor-alpha production with anti-hypertensive drugs. Immunopharmacology. 2000, 48, 65-74.
[146] 宋浩明, 罗明, 邓南伟, 等. 氨氯地平在心衰治疗中对肿瘤坏死因子-α、白细胞介素-1,6 的作用. 中国新药与临床杂志. 2002, 21, 661-3.
[147] 宋革, 苏海砾, 杜芳, 等. 新二氢吡啶类钙拮抗剂 MN9202 对实验性小鼠脑缺血的防护作用. 中国微循环. 2001, 5, 208-10.
[148] Yamamoto, C., Yashima, E. and Okamoto, Y. Computational studies on chiral discrimination mechanism of phenylcarbamate derivatives of cellulose. Bull. Chem.Soc. Jpn. 1999, 72, 1815-25.
[149] Zhao, M.G., Mei, Q.B., Zhang, Y.F., Xing, B., Li, X.Q. and Cui, Y. Inhibitory effects of serotonin on transient outward potassium current in rat ventricular myocytes. Acta Pharmacol Sin. 2002, 23, 617-22.
[150] Zhao, M.G., Mei, Q.B., Zhang, Y.F., Xiong, X.Y., Lu, B.H., Xing, B. and Zhao, D.H. 5-Hydroxytryptamine enhances L-type calcium current in norepinephrine-induced hypertrophic ventricular myocytes. Acta Pharmacol Sin. 2002, 23, 523-8.
[151] Ren, J. and Wold, L.E. Measurement of Cardiac Mechanical Function in Isolated Ventricular Myocytes from Rats and Mice by Computerized Video-Based Imaging. Biol Proced Online. 2001, 3, 43-53.
[152] Zhang, B., Zhang, H., Fan, Q., Ma, X. and Gao, F. Insulin improves cardiac myocytes contractile function recovery in simulated ischemia-reperfusion: key role of Akt. Chin Sci Bull 2003, 48, 1364-9.
[153] Sipido, K.R. and Callewaert, G. How to measure intracellular [Ca2+] in single cardiac cells with fura-2 or indo-1. Cardiovasc Res. 1995, 29, 717-26.
[154] Takahashi, A., Camacho, P., Lechleiter, J.D. and Herman, B. Measurement of intracellular calcium. Physiol Rev. 1999, 79, 1089-125.
[155] Molkentin, J.D. Dichotomy of Ca2+ in the heart: contraction versus intracellularsignaling. J Clin Invest. 2006, 116, 623-6.
[156] Adachi-Akahane, S. [Molecular and pharmacological bases for the gating regulation of L-type voltage-dependent Ca2+ channels]. Nippon Yakurigaku Zasshi. 2004, 123, 197-209.
[157] Takamatsu, H., Nagao, T., Ichijo, H. and Adachi-Akahane, S. L-type Ca2+ channels serve as a sensor of the SR Ca2+ for tuning the efficacy of Ca2+-induced Ca2+ release in rat ventricular myocytes. J Physiol. 2003, 552, 415-24.
[158] Xie, M.J., Zhang, L.F., Ma, J. and Cheng, H.W. Functional alterations in cerebrovascular K+ and Ca2+ channels are comparable between simulated microgravity rat and SHR. Am J Physiol Heart Circ Physiol. 2005, 289, H1265-76.
[159] 刘振伟. 实用膜片钳技术. 军事医学科学出版社. 2006.
[160] 康华光. 膜片钳技术及其应用. 科学出版社. 2003.
[161] Parrillo, J.E., Parker, M.M., Natanson, C., Suffredini, A.F., Danner, R.L., Cunnion, R.E. and Ognibene, F.P. Septic shock in humans. Advances in the understanding of pathogenesis, cardiovascular dysfunction, and therapy. Ann Intern Med. 1990, 113, 227-42.
[162] Jean-Baptiste, E. Cellular mechanisms in sepsis. J Intensive Care Med. 2007, 22, 63-72.
[163] Rice, T.W. Treatment of severe sepsis: where next? Current and future treatment approaches after the introduction of drotrecogin alfa. Vasc Health Risk Manag. 2006, 2, 3-18.
[164] Aggarwal, B.B. Signalling pathways of the TNF superfamily: a double-edged sword. Nat Rev Immunol. 2003, 3, 745-56.
[165] Ghezzi, P. and Cerami, A. Tumor necrosis factor as a pharmacological target. MolBiotechnol. 2005, 31, 239-44.
[166] Szabo, C., Mitchell, J.A., Gross, S.S., Thiemermann, C. and Vane, J.R. Nifedipine inhibits the induction of nitric oxide synthase by bacterial lipopolysaccharide. J Pharmacol Exp Ther. 1993, 265, 674-80.
[167] Haupt, W., Fritzsche, H., Hohenberger, W. and Zirngibl, H. Selective cytokine release induced by serum and separated plasma from septic patients. Eur J Surg. 1996, 162, 769-76.
[168] Arras, M., Hoche, A., Bohle, R., Eckert, P., Riedel, W. and Schaper, J. Tumor necrosis factor-alpha in macrophages of heart, liver, kidney, and in the pituitary gland. Cell Tissue Res. 1996, 285, 39-49.
[169] 荣仔萍, 常中飞, 宋欣伟. 肿瘤坏死因子与心力衰竭及抗细胞因子治疗研究进展. 医学综述. 2006, l2, 58-60.
[170] Chovolou, Y., Watjen, W., Kampkotter, A. and Kahl, R. Resistance to tumor necrosis factor-alpha (TNF-alpha)-induced apoptosis in rat hepatoma cells expressing TNF-alpha is linked to low antioxidant enzyme expression. J Biol Chem. 2003, 278, 29626-32.
[171] Kawano, H., Okada, R., Kawano, Y., Sueyoshi, N. and Shirai, T. Apoptosis in acute and chronic myocarditis. Jpn Heart J. 1994, 35, 745-50.
[172] 李世英, 倪新海, 陈曦, 等. 血管紧张素Ⅱ促进培养的新生鼠心肌细胞表达TNF-α. 高血压杂志. 2003, 11, 161-4.
[173] Carswell, E.A., Old, L.J., Kassel, R.L., Green, S., Fiore, N. and Williamson, B. An endotoxin-induced serum factor that causes necrosis of tumors. Proc Natl Acad Sci U S A. 1975, 72, 3666-70.
[174] 邱飞, 王礼琛. 肿瘤坏死因子 α 与心力衰竭的研究进展. 国外医学(药学分册).2003, 30, 326-30.
[175] Kubota, T., McTiernan, C.F., Frye, C.S., Slawson, S.E., Lemster, B.H., Koretsky, A.P., Demetris, A.J. and Feldman, A.M. Dilated cardiomyopathy in transgenic mice with cardiac-specific overexpression of tumor necrosis factor-alpha. Circ Res. 1997, 81, 627-35.
[176] Bozkurt, B. et al. Pathophysiologically relevant concentrations of tumor necrosis factor-alpha promote progressive left ventricular dysfunction and remodeling in rats. Circulation. 1998, 97, 1382-91.
[177] Kriegler, M., Perez, C., DeFay, K., Albert, I. and Lu, S.D. A novel form of TNF/cachectin is a cell surface cytotoxic transmembrane protein: ramifications for the complex physiology of TNF. Cell. 1988, 53, 45-53.
[178] 尹丙姣, 李卓娅, 余上斌, 等. 跨膜型和分泌型 TNF-α 在内毒素性休克过程中的作用. 中华微生物学和免疫学杂志. 2002, 22, 334-8.
[179] Torre-Amione, G., Kapadia, S., Lee, J., Durand, J.B., Bies, R.D., Young, J.B. and Mann, D.L. Tumor necrosis factor-alpha and tumor necrosis factor receptors in the failing human heart. Circulation. 1996, 93, 704-11.
[180] 陈齐红, 覃数. 肿瘤坏死因子-α 和充血性心力衰竭. 心血管病进展. 2004, 25, 89-91.
[181] Palladino, M.A., Bahjat, F.R., Theodorakis, E.A. and Moldawer, L.L. Anti-TNF-alpha therapies: the next generation. Nat Rev Drug Discov. 2003, 2, 736-46.
[182] Stuber, F. Effects of genomic polymorphisms on the course of sepsis: is there a concept for gene therapy? J Am Soc Nephrol. 2001, 12 Suppl 17, S60-4.
[183] Matsui, T. et al. Akt activation preserves cardiac function and prevents injury aftertransient cardiac ischemia in vivo. Circulation. 2001, 104, 330-5.
[184] Chen, K.Q., Iribarren, P., Gong, W.H. and Wang, J.M. The Essential Role of Phosphoinositide 3-Kinases (PI3Ks) in Regulating Pro-Inflammatory Responses and the Progression of Cancer. Cell Mol Immunol. 2005, 2, 241-52.
[185] Sambrook, J., Russell, D. (黄培堂等译). 分子克隆实验指南(第 3 版). 科学出版社. 2002, 北京
[186] Freyer, D. et al. Cerebral endothelial cells release TNF-alpha after stimulation with cell walls of Streptococcus pneumoniae and regulate inducible nitric oxide synthase and ICAM-1 expression via autocrine loops. J Immunol. 1999, 163, 4308-14.
[187] Fiorucci, S. et al. Relative contribution of acetylated cyclo-oxygenase (COX)-2 and 5-lipooxygenase (LOX) in regulating gastric mucosal integrity and adaptation to aspirin. Faseb J. 2003, 17, 1171-3.
[188] Knowles, R.G. and Moncada, S. Nitric oxide synthases in mammals. Biochem J. 1994, 298 ( Pt 2), 249-58.
[189] Gaston, B., Drazen, J.M., Loscalzo, J. and Stamler, J.S. The biology of nitrogen oxides in the airways. Am J Respir Crit Care Med. 1994, 149, 538-51.
[190] Forstermann, U., Closs, E.I., Pollock, J.S., Nakane, M., Schwarz, P., Gath, I. and Kleinert, H. Nitric oxide synthase isozymes. Characterization, purification, molecular cloning, and functions. Hypertension. 1994, 23, 1121-31.
[191] Hoit, B.D. Two faces of nitric oxide: lessons learned from the NOS2 knockout. Circ Res. 2001, 89, 289-91.
[192] Ullrich, R., Scherrer-Crosbie, M., Bloch, K.D., Ichinose, F., Nakajima, H., Picard, M.H., Zapol, W.M. and Quezado, Z.M. Congenital deficiency of nitric oxide synthase 2 protects against endotoxin-induced myocardial dysfunction in mice. Circulation.2000, 102, 1440-6.
[193] Goren, N., Cuenca, J., Martin-Sanz, P. and Bosca, L. Attenuation of NF-kappaB signalling in rat cardiomyocytes at birth restricts the induction of inflammatory genes. Cardiovasc Res. 2004, 64, 289-97.
[194] Schmedtje, J.F., Jr., Ji, Y.S., Liu, W.L., DuBois, R.N. and Runge, M.S. Hypoxia induces cyclooxygenase-2 via the NF-kappaB p65 transcription factor in human vascular endothelial cells. J Biol Chem. 1997, 272, 601-8.
[195] Lai, J. et al. Prostaglandin F2 alpha induces cardiac myocyte hypertrophy in vitro and cardiac growth in vivo. Am J Physiol. 1996, 271, H2197-208.
[196] Hocherl, K., Dreher, F., Kurtz, A. and Bucher, M. Cyclooxygenase-2 inhibition attenuates lipopolysaccharide-induced cardiovascular failure. Hypertension. 2002, 40, 947-53.
[197] Wong, S.C., Fukuchi, M., Melnyk, P., Rodger, I. and Giaid, A. Induction of cyclooxygenase-2 and activation of nuclear factor-kappaB in myocardium of patients with congestive heart failure. Circulation. 1998, 98, 100-3.
[198] Ejima, K. et al. Cyclooxygenase-2-deficient mice are resistant to endotoxin-induced inflammation and death. Faseb J. 2003, 17, 1325-7.
[199] Chou, T.C., Yang, S.P. and Pei, D. Amlodipine inhibits pro-inflammatory cytokines and free radical production and inducible nitric oxide synthase expression in lipopolysaccharide/interferon-gamma-stimulated cultured vascular smooth muscle cells. Jpn J Pharmacol. 2002, 89, 157-63.
[200] Sugimoto, Y., Whitman, M., Cantley, L.C. and Erikson, R.L. Evidence that the Rous sarcoma virus transforming gene product phosphorylates phosphatidylinositol and diacylglycerol. Proc Natl Acad Sci U S A. 1984, 81, 2117-21.
[201] Macara, I.G., Marinetti, G.V. and Balduzzi, P.C. Transforming protein of avian sarcoma virus UR2 is associated with phosphatidylinositol kinase activity: possible role in tumorigenesis. Proc Natl Acad Sci U S A. 1984, 81, 2728-32.
[202] Guha, M. and Mackman, N. The phosphatidylinositol 3-kinase-Akt pathway limits lipopolysaccharide activation of signaling pathways and expression of inflammatory mediators in human monocytic cells. J Biol Chem. 2002, 277, 32124-32.
[203] Fang, H., Pengal, R.A., Cao, X., Ganesan, L.P., Wewers, M.D., Marsh, C.B. and Tridandapani, S. Lipopolysaccharide-induced macrophage inflammatory response is regulated by SHIP. J Immunol. 2004, 173, 360-6.
[204] 尹华华, 夏浩, 程晓刚, 等. LPS 对小鼠肺组织及 RAW264. 7 细胞 Akt 蛋白表达及磷酸化的调控. 第三军医大学学报. 2007, 29, 188-190.
[205] Matsumori, A., Nunokawa, Y. and Sasayama, S. Nifedipine inhibits activation of transcription factor NF-kappaB. Life Sci. 2000, 67, 2655-61.