丹参素定向诱导骨髓间质干细胞分化为神经元样细胞的研究

目的 研究丹参素体外定向诱导骨髓间质干细胞(mesenchymal stem cells,MSC)分化为神经元样细胞。方法 利用Ficoll-Paque液将骨髓细胞进行密度梯度离心和贴壁筛选分离出MSC,体外扩增,流式细胞仪检测MSC表面抗原表达。采用含丹参素的无血清L-DMEM诱导MSC分化为神经元样细胞,并且探讨丹参素不同浓度及不同作用时间对MSC诱导分化为神经元样细胞的影响。以免疫组化方法检测神经元特异性烯醇化酶(NSE)、神经丝蛋白(NF-M)、巢蛋白(nestin)、胶质纤维酸性蛋白(GFAP)的表达、结果 MSC在体外传代扩增后,流式细胞仪检测结果显示CD29、CD44、CD166表达阳性,CDl4、CD34、CD45、HLA-DR为阴性。丹参素诱导后,MSC胞体收缩,突起伸出,呈典型的核周体形态,类似神经元;免疫组化显示诱导出的神经元样细胞NSE、NF-M、nestin表达阳性,GFAP阴性。结论 丹参素可以在体外诱导MSC分化为神经元样细胞,其诱导转化率与丹参素浓度和作用时间有关。     

高效液相色谱法测定复方丹参注射液中参素的含量

目的：考察不同厂家复方丹参注射液中丹参素含量。方法：选择复方丹参注射液中活性成份丹参素作为指标,用反相高效液相色谱法,以甲醇－０．４％醋酸（５：９５）作流动相,检测波长为２８１ｎｍ。结果与结论：样品平均加样加嘏率为９９．７１％,ＲＳＤ为０．９６％,测定结果不同厂家产品差异很大,为保证临床用药安全有效,应对其有效成份建立控制标准。     

丹参素对大鼠急性心肌缺血的保护作用

目的研究丹参素对大鼠急性心肌缺血的保护作用。方法采用垂体后叶素、异丙肾上腺素和冠脉结扎造成大鼠急性心肌缺血模型,以心电图T波变化,血清CK、LDH为指标,研究丹参素对大鼠急性心肌缺血的保护作用。结果丹参素（6、12、24mg／kg）剂量组均能对抗垂体后叶素及异丙肾上腺素引起的大鼠急性心肌缺血T波偏移,并且能够保护心肌细胞,显著降低冠脉结扎造成急性心肌缺血大鼠血清中CK和LDH溢出。结论丹参素对大鼠急性心肌缺血具有显著的保护作用。     

RP-HPLC测定石见穿中丹参素的含量

目的:建立石见穿中丹参素的含量测定方法。方法:采用反相高效液相法,以甲醇-0.1%冰醋酸(20∶80)为流动相,柱温为30℃,检测波长为280nm。结果:丹参素进样量在1.9248～5.7744μg具有良好的线性关系(r=0.9999),平均回收率为100.6%,RSD=1.2%。结论:该法灵敏度高、重现性好,可用于石见穿的质量控制。     

丹参素对PDGF-BB刺激下肝星状细胞的抑制作用

目的观察丹参素对PDGF-BB刺激下大鼠肝星状细胞活化增殖,Ⅰ、Ⅲ型胶原合成与分泌,ERK1/2和PI3K通道蛋白表达的影响,探讨丹参素抑制肝纤维化的分子机制。方法用不同浓度丹参素(0.250、0.125、0.062 mmol/L)、ERK通道特异性抑制剂UO126(20 nmol/L)和PI3K通道特异性抑制剂LY294002(20 nmol/L)分别作用于经血小板衍生生长因子PDGF-BB(10 ng/ml)处理的大鼠肝星状细胞(HSC)48 h,CCK8法检测HSC增殖活性,免疫化学染色法测定Ⅰ、Ⅲ型胶原合成,酶联免疫吸附法观察Ⅰ、Ⅲ型胶原分泌,Western blot测定ERK1/2、p-ERK1/2、AKT和p-AKT蛋白表达。结果 PDGF-BB 10 ng/ml处理后,HSC增殖明显增加(0.139±0.009),丹参素能抑制PDGF-BB刺激大鼠HSC增殖的作用,并随丹参素浓度增加(0.062、0.125、0.250 mmol/L)细胞增殖受到的抑制作用越强(0.102±0.008、0.083±0.007、0.066±0.014,P<0.05),Ⅰ、Ⅲ型胶原合成及分泌均显著减少(P<0.05),p-ERK1/2和p-AKT蛋白表达均显著降低(P<0.05)。结论丹参素可通过抑制PDGF-BB诱导的HSC增殖和活化,同时抑制与肝纤维化密切相关的MAPK、PI3K途径中ERK、AKT蛋白的磷酸化,负性调控肝纤维化过程,其机制可能与抑制ERK1/2和PI3K信号转导通路有关。     

RP—HPLC测定石见穿中丹参素的含量

目的：建立石见穿中丹参素的含量测定方法。方法：采用反相高效液相法,以甲醇-0．1％冰醋酸（20：80）为流动相,柱温为30℃,检测波长为280nm。结果：丹参素进样量在1．9248～5．7744肛g具有良好的线性关系（r=0．9999）,平均回收率为100．6％,RSD=1．2％。结论：该法灵敏度高、重现性好,可用于石见穿的质量控制。     

Danshensu Ameliorates Cardiac Ischaemia Reperfusion Injury through Activating Sirt1/FoxO1/Rab7 Signal Pathway

Objective: To explore the specific molecular mechanisms of Danshensu(DSS) in the treatment of ischemia reperfusion injury(IRI). Methods: IRI model was established with isolated rat hearts by performing global ischaemia for 30 min, and then followed by 60 min reperfusion. Also, H9 C2 cells were subjected to 4-h hypoxia followed by 3-h reoxygenation. Then 10 μmol/L DSS were added in the reperfusion/reoxygenation step to intervene IRI. Cardiac function, structural change and apoptosis were respectively tested by Langendorff System, hematoxylin and eosin(HE) and terminal-deoxynucleotidyl transferase mediated nick endabeling(TUNEL) stainings. Then lactate dehydrogenase(LDH), cardiac troponin T(c Tn T), reactive oxygen species(ROS), superoxide dismutase(SOD) and glutathione peroxidase(GSH-PX) were detected by enzyme-linked immunosorbent assay(ELISA). Sirt1/Fox O1/Rab7 Signal Pathway was monitored at both protein and m RNA levels. Results: The results showed that IRI not only greatly attenuated cardiac function(LVDP and ±dp/dtmax, P0.01, P0.05) and increased the level of the marker enzymes(c Tn T, LDH, P0.01) from the coronary effluents, but also markedly induced changes in the structure of cardiomyocytes and contributed to apoptosis, which were mediated by boosted endogenous ROS. However, after treatment with DSS all above indexes were improved, which was related to activating Sirt1/Fox O1/Rab7 signal pathway accompanied with the enhancement of antioxidant defense system, such as SOD and GSH-PX. Conclusion: DSS is able to protect hearts from IRI, which may be attributable to inhibiting excessive ROS through Sirt1/Fox O1/Rab7 signaling.     

近红外光谱结合不同偏最小二乘法测定乳块消片醇沉液中丹参素和橙皮苷含量

目的：应用近红外分析技术结合化学计量学方法建立中药乳块消片醇沉液中丹参素和橙皮苷含量测定的新方法。方法：采用Sample set Partitioning based on joint x-y distance（SPXY）法对训练集样本和预测集样本进行划分，应用不同的偏最小二乘方法进行有效波段范围选择以及建立定量校正模型，分别比较了间隔偏最小二乘算法(interval partial least squares, iPLS)，组合间隔偏最小二乘算法(Synergy interval partial least squares，SiPLS)，向后间隔偏最小二乘算法（backward interval partial least squares，BiPLS），窗口移动偏最小二乘算法（moving window partial least squares ,MWPLS）。结果：丹参素采用SiPLS三个区间组合、橙皮苷采用SiPLS四个区间组合建立的回归模型性能最好，预测相关系数(R)分别为0.9956和0.9940，交互验证误差均方根(RMSECV) 分别为0.0096和0.0083，预测误差均方根(RMSEP)为0.0062和0.0074。结论：该近红外光谱法对丹参素和橙皮苷含量预测结果较好，且方便快捷、无前期预处理和无污染，为中药生产过程的在线检测提供了依据。     

近红外光谱结合不同偏最小二乘法测定乳块消片醇沉液中丹参素和橙皮苷含量

目的：应用近红外分析技术结合化学计量学方法建立中药乳块消片醇沉液中丹参素和橙皮苷含量测定的新方法。方法：采用Sample set Partitioning based on joint x—y distance（SPXY）法对训练集样本和预测集样本进行划分，应用不同的偏最小二乘方法进行有效波段范围选择以及建立定量校正模型，分别比较了间隔偏最小二乘算法（interval partial least squares，iPLS），组合间隔偏最小二乘算法（Synergy interval partial least squares，SiPLS），向后间隔偏最小二乘算法（backward interval partial least squares，BiPLS），窗口移动偏最小二乘算法（movingwindowpartialleastsquares，MWPLS）。结果：丹参素采用SiPLS三个区间组合、橙皮苷采用SiPLS四个区间组合建立的回归模型性能最好，预测相关系数（R）分别为0．9956和0．9940，交互验证误差均方根（RMSECV）分别为0．0096和0．0083，预测误差均方根（RMSEP）为0．0062和0．0074。结论：该近红外光谱法对丹参素和橙皮苷含量预测结果较好，且方便快捷、无前期预处理和无污染，为中药生产过程的在线检测提供了依据。     

Favorable Maternal and Fetal Effects of Danshensu in an Experimental Mice Model of Preeclampsia

Background. Preeclampsia is a serious pregnancy-specific complication that results in high maternal and neonatal mortality and morbidity worldwide. Till date, there is no satisfactory pharmacotherapeutic treatment, except for aspirin and heparin, to stop the preeclampsia progression. Although the mechanism of preeclampsia is poorly understood, it has proved to be associated with coagulation activation. Researches on prophylactic and remedial application of anticoagulants maybe benefit the clinical aspects of preeclampsia individuals. Methods. Sixty-six preeclampsia-like pregnant mice, induced by phosphatidyleserine/phosphatidylcholine (PS/PC) microvesicle administration, were randomly divided into six groups as follows: control group (group C), preeclampsia model group (group PE), group treated with heparin (group H), group treated with aspirin (group A), group treated with low-dose danshensu (group LD), and group treated with high-dose danshensu (group HD). Systolic blood pressure (SBP), proteinuria, mean platelet counts, plasma antithrombin III activity (AT III), D-dimmer levels, thrombin time (TT), fibrin deposition with phosphotungstic acid hematoxylin (PTAH) staining, and thrombomodulin (TM) expression with immunohistochemistry staining in placentas were examined as indices for maternal syndrome. Meanwhile, the number of potentially viable fetuses, weight of fetuses and placentas, weight of fetal brains, nose-breech length, ponderal index (PI), and neurons with hematoxylin-eosin (H/E) and toluidine blue-eosin (Nissl's) staining were all evaluated as indices for fetal syndrome. Results. Heparin presents significant effects on maternal syndrome of preeclampsia such as hypertension and proteinuria, and different dose danshensu also presents the certain effects. High-dose danshensu and aspirin all process better effects than low-dose danshensu on decreasing blood pressure to normal level, whereas high-dose danshensu process better effects than aspirin and low-dose danshensu on decreasing proteinuria to normal level. As to danshensu's effects on hemostatic function, high- and low-dose danshensu's marked effects on increasing the plasma AT III activity are same as that of aspirin and inferior to heparin. High-dose danshensu's better effect on elevating the platelet counts is superior to low-dose danshensu and aspirin. Low-dose danshensu's obvious effect on decreasing D-dimmer levels is close to heparin and superior to high-dose danshensu and aspirin. High- and low-dose danshensu's significant effects on reduced TT level are same to that of heparin. Different anticoagulants all have the improvement roles on placental fibrin depositions, but heparin and high-dose danshensu's roles on lowering thrombomodulin expression in placentas are superior to low-dose danshensu and aspirin. But anticoagulant function of high-dose danshensu is still inferior to heparin. Furthermore, we found the following changes: increasing fetal body weight and length in every group, obvious overall improvement in group H, greater amelioration equaling to that in heparin group on maternal body weight, fetal nose-breech length and fetal brain weight in group HD, better changes on survival fetal number in group LD than in other groups, and more corrected brain development in group HD than in group A. We found long-term use of heparin and aspirin, in spite of low-dose administration, can raise the risk of bleeding such as placental abruption and intestinal hemorrhage. But no side effect was observed in mice treated with different dose of danshensu in our study. Conclusions. Danshensu has proven effective in ameliorating the prognosis of maternal syndrome and fetal syndrome in the PE mouse model. We suggest long-term provision of low-dose danshensu in pregnancy, leading to an improvement of preeclampsia syndrome with considerable maternal safety.