Impact of Lycium Barbarum Polysaccharide and Danshensu on vascular endothelial growth factor in the process of retinal neovascularization of rabbit

AIM:To discuss the impact of Lycium Barbarum Polysaccharide (LBP) and Danshensu purified from Traditional Chinese Medicine (TCM) on vascular endothelial growth factor (VEGF) of rabbits with retinal neovascularization. METHODS:Forty rabbits were divided into normal control group, model control group, LBP group and Danshensu group. Animals in the normal control group were fed in the normal oxygen environment. Animals in the other three groups were put into the environment with 70% oxygen for 5 days in order to build the model of oxygen-induced vascular proliferation retinopathy. And then different TCM extract was injected into the abdominal cavities of these annimals. After 7 days, the VEGF content of in the serum of rabbit was measured by double antibody sandwich method. RESULTS:Data analysis indicated that VEGF content was as follows:Danshensu group was lower than model control group (12.92 ±3.84ng/L vs 19.32 ±4.15ng/L, P 【 0.05); LBP group and normal control group were lower than model control group (12.92±3.84ng/L, 9.26±1.61ng/L vs 19.32±4.15ng/L, P【0.01); total blood viscosity, plasma viscosity, cholesterol content, fibrinogen content and triacylglycerol content after peritoneal injection of LBP and Danshensu were obviously lower than before injection. CONCLUSION:TCM extract-LBP and Danshensu can prominently reduce the content of VEGF in the process of vascular proliferative retinopathy of rabbit; can prevent the occurrence of retinal microvascular disease by improving partial oxygen -deficient environment or affecting all kinds of new growth factor.

Novel derivative of Danshensu acts as anti-thrombotic agent for modulation of protein disulfide isomerase family member ERp57

OBJECTIVE To identify the specific targets of a novel derivative of Danshensu ADTM as the protein disulfide isomerase(PDI)family proteins including ERp57.To further investigate the underlying mechanism of ADTM to modulate ERp57 to regulate platelet function with direct interaction withαⅡbβ3 integrin.METHODS To isolate the protein targets that bound to ADTM,a biotin-conjugated ADTM analogue(BAA)was designed and synthesized.BAA(300μmol·L-1)was incubated with rat blood platelet lysates and the BAA-protein complexes were pulled down with NeutrAvidin-agarose followed by protein profiling using LC-MS/MS.To determine platelet aggregation in vitro,rabbit platelets were incubated with the indicated concentrations of compounds and aggregation was induced by ADP(10μmol·L-1)or AA(200μmol·L-1)and measured using a platelet aggregometer.To determine platelet aggregation-induced by ADP in rat in vivo,ADTM(5-20mg·kg-1)in comparison with DSS(10mg·kg-1)and clopidogrel(18mg·kg-1)were administered daily by i.v.injection for 5d,respectively.To determine the action of ADTM on the ERp57/αⅡbβ3 interaction,it was examined by immunoprecipitation with anti-αⅡbβ3antibody,followed by detection of ERp57 immunoreactivity using immunoblotting.RESULTS BAA could bind to various proteins involved in platelet function.In particular,platelet aggregation-associated proteins were identified with>95% protein identification probability including ERp72,ERp57ERp5 and PDI,which are members of the protein disulfide isomerase(PDI)family related to platelet function and redox homeostasis.ADTM exhibited potent inhibition on the redox activity of ERp57 in a concentration-dependent manner(IC50=100 300μmol·L-1).In in vitro studies,ADTM exhibited concentration-dependent inhibition on ADP-induced and AA-induced platelet aggregation with comparable effects to aspirin and clopidogrel.In vivo study showed that ADP-induced platelet aggregation was significantly compromised(>40%reduction)in rats treated with ADTM(20mg·kg-1).Similarly,ADTM also exhibited significant anti-thrombotic effect in vivo as shown in the ferric chloride(FeCl3)-induced venous thrombosis.Immunoprecipitation with anti-αⅡbβ3antibody,followed by detection of ERp57 immunoreactivity using immunoblotting showed that ADTM disrupted the interaction of ERp57 with αⅡ bβ3.CONCLUSION These results demonstrated that ADTM exhibited broad-spectrum anti-platelet activities and ERp57 is a potential therapeutic target for anti-platelet therapy.

Cardioprotective and chemosensitizing effects of novel Danshensu derivatives

OBJECTIVE To investigate the cardioprotective effect of novel danshensu derivatives against doxorubicin(Dox)cardiotoxicity and their synergistic anti-tumor effect with Dox on breast cancer cells.METHODS Two new Danshensu derivatives were synthesized by conjugation with tetramethylpyrizine and/or4-(3-thioxo-3 H-1,2-dithiol-4-yl)-benzoic acid and tested for protective effects against Dox induced cardiotoxicity in cell and zebrafish.H9c2 cardiomyoblasts were co-treated with Dox and Danshensu derivatives for 24 h and then were measured for cell viability and cytotoxicity by MTT and LDH assays.The expression levels of mitochondrial biogenesis related proteins PGC-1α,NRF-1and Nrf2 were detected by Western blotting and qPCR.Moreover,in a Dox-induced cardiotoxicity model of zebrafish,zebrafish embryos were treated with Dox for 36 h,followed by measurement of numerous ventricular function parameters including heart rate,stroke volume,cardiac output and fractional shortening.In addition,the synergistic anti-tumor effects of the Danshensu derivatives and Dox had been studied in MCF-7 breast cancer cells.The effects of the Danshensu derivatives on the cell death and metabolism of MCF-7 cells were measured using apoptosis assay and Seahorse Metabolic Analyzers respectively.RESULTS Our results showed that the Danshensu derivatives were more potent than the parental compounds in ameliorating Dox-induced cytotoxicity in H9c2 cells and significantly preserving stroke volume of heart function in Dox-treated zebrafish.Further mechanistic studies identified that the danshensu derivatives increased mitochondrial copy numbers and protein expressions of PGC-1α,NRF-1 and Nrf2 in H9c2 cells.In addition,the Danshensu derivatives enhanced Dox-induced apoptosis,and decreased glycolysis and mitochondrial function in MCF-7 tumor cells.CONCLUSION Our results revealed that two new Danshensu derivatives displayed promising cardioprotective effects against Dox induced cardiotoxicity both in vivo and in vitro,at least partially through activating mitochondrial biogenesis.Also,the new Danshensu derivatives potentiated the anticancer effects of Dox in breast tumor cells involving induction of glycolytic inhibition and mitochondrial dysfunction.

脯氨酸丹参素冰片酯对小鼠中枢神经系统的一般药理学研究

目的 考察脯氨酸丹参素冰片酯对小鼠中枢神经系统的影响。方法 将小鼠随机分为对照组和脯氨酸丹参素冰片酯大剂量(120 mg/kg)、中剂量(60 mg/kg)、小剂量(30 mg/kg)给药组,每组10只,雌雄各半,灌胃给药后考察对小鼠自主活动、协同睡眠、协调运动能力和镇痛的作用。结果 与对照组相比,脯氨酸丹参素冰片酯对小鼠自主活、协同睡眠、协调运动能力和镇痛方面无显著影响。结论 脯氨酸丹参素冰片酯对小鼠中枢神经系统无明显影响,在药效有效剂量范围内安全性较好。

基于PPARγ/PGC-1α通路探讨丹参素对糖尿病心肌病大鼠心肌线粒体功能的影响

目的基于过氧化物酶体增殖物激活受体γ(PPARγ)/过氧化物酶体增殖物激活受体γ辅助激活因子1α(PGC-1α)通路探究丹参素对糖尿病心肌病(DCM)大鼠心肌线粒体功能的影响。方法将大鼠随机分为正常组、模型组、丹参素低剂量组(5 mg·kg^(-1))、丹参素中剂量组(10 mg·kg^(-1))、丹参素高剂量组(20 mg·kg^(-1))、二甲双胍组(140 mg·kg^(-1))、丹参素高剂量+GW9662组(20 mg·kg^(-1)丹参素+1 mg·kg^(-1)GW9662),每组12只。除正常组外,其余各组均采用高糖高脂饲料喂养联合腹腔注射链脲佐菌素的方法构建DCM大鼠模型。模型复制成功后,各组按照上述设定剂量灌胃给药,每日1次,连续6周。采用动物血糖仪检测空腹血糖值;超声心动图评估大鼠心功能,包括左室短轴缩短率(LVFS)、左室射血分数(LVEF);HE染色法观察心肌组织病理变化;透射电镜观察心肌组织线粒体超微结构;JC-1染色法检测心肌细胞线粒体膜电位;通过试剂盒检测心肌组织中三磷酸腺苷(ATP)含量及活性氧(ROS)的表达水平;Western Blot法检测心肌组织中PPARγ、PGC-1α蛋白表达水平。结果与正常组比较,模型组大鼠的空腹血糖水平明显升高(P<0.05),LVFS、LVEF明显降低(P<0.05);心肌组织明显受损,心肌线粒体肿胀;心肌细胞线粒体膜电位未降低细胞的百分比明显降低(P<0.05);心肌组织中ATP含量明显降低(P<0.05),ROS表达水平明显升高(P<0.05);心肌组织中PPARγ、PGC-1α蛋白表达明显下调(P<0.05)。与模型组比较,丹参素各剂量组及二甲双胍组大鼠空腹血糖水平明显降低(P<0.05),LVFS、LVEF明显升高(P<0.05);心肌组织损伤减轻,心肌线粒体结构损伤有所减轻;心肌细胞线粒体膜电位未降低细胞的百分比明显升高(P<0.05);心肌组织中ATP含量明显升高(P<0.05),ROS表达水平明显降低(P<0.05);心肌组织中PPARγ、PGC-1α蛋白表达明显上调(P<0.05)。与丹参素高剂量组比较,丹参素高剂量+GW9662组大鼠空腹血糖水平明显升高(P<0.05),LVFS、LVEF明显降低(P<0.05);心肌组织损伤、心肌线粒体结构损伤加重,心肌线粒体肿胀;心肌细胞线粒体膜电位未降低细胞的百分比明显降低(P<0.05);心肌组织中ATP含量明显降低(P<0.05),ROS表达水平明显升高(P<0.05);心肌组织中PPARγ、PGC-1α蛋白表达明显下调(P<0.05)。结论丹参素改善DCM大鼠线粒体功能可能与激活PPARγ/PGC-1α通路有关。

丹参素对UVB诱导的皮肤光老化小鼠的保护作用和抗氧化机制研究

研究丹参素(DSS)对紫外线B(UVB)诱导的皮肤光老化(SP)小鼠的保护作用和抗氧化机制。建立了UVB诱导的SP小鼠模型,使用3个剂量的DSS(20,40和80 mg/(kg·d))治疗小鼠8周,结束后分别测定了各组小鼠的表皮含水量。通过HE染色评价皮肤组织形态,Masson三色染色评价胶原蛋白沉积。按照试剂盒说明测定皮肤组织中氧化应激指标(SOD、CAT、GSH-Px和MDA)和炎症指标(TNF-α和IL-6)水平。通过RT-qPCT和Western blotting检测了皮肤组织中MMP-1、Collagen I、Nrf2、Keap1、HO-1、NF-κB p65和p-NF-κB p65的mRNA或蛋白水平。结果显示,DSS剂量依赖性地提高了UVB诱导的SP小鼠表皮含水量,减轻皮肤损伤,促进胶原形成(P<0.05)。DSS抑制了UVB诱导的SP小鼠皮肤组织中MMP-1的转录和表达,促进了Collagen的转录和表达(P<0.05)。DSS升高了UVB诱导的SP小鼠皮肤组织中SOD、CAT和GSH-Px的水平,降低了MDA的水平(P<0.05)。DSS降低了UVB诱导的SP小鼠皮肤组织中TNF-α和IL-6的水平(P<0.05)。DSS促进了UVB诱导的SP小鼠皮肤组织中Nrf2和HO-1的转录和表达,抑制了Keap1的转录和表达(P<0.05)。DSS抑制了UVB诱导的SP小鼠皮肤组织中p-NF-κB p65的表达(P<0.05)。本研究表明DSS可有效改善UVB诱导的小鼠SP,其机制与Nrf2和NF-κB信号通路有关。

肾茶地上部分丹参素衍生物的分离与绝对构型的确定

目的对肾茶(Clerodendranthus spicatus(Thunb)C Y Wu ex H.W.Li)地上部分的化学成分进行研究,进一步丰富其物质基础。方法采用加热回流提取法对肾茶的干燥地上部分进行提取,联合应用硅胶、ODS及高效液相色谱等多种色谱法对肾茶地上部分提取物的化学成分进行分离纯化,结合圆二色光谱、核磁、质谱等波谱技术以及化学反应的方法,鉴定化合物的结构,确定其绝对构型。结果从肾茶地上部分分离鉴定了7个丹参素衍生物,分别为(R)-迷迭香酸(1)、(R)-迷迭香酸甲酯(2)、(R)-迷迭香酸乙酯(3)、2R-羟基-3-(4-羟基苯基)丙酸甲酯(4)、2R-羟基-3-(3,4-二羟基苯基)丙酸(5)、2R-羟基-3-(3,4-二羟基苯基)丙酸甲酯(6)和2R-羟基-3-(3,4-二羟基苯基)丙酸乙酯(7)。结论化合物4为首次从肾茶属植物中分离得到。此外,发现对于α-羟基羧酸类化合物,即使是只有一个手性中心,也不能仅仅通过旋光值的比对确定其绝对构型。本文所提供的方法为该类成分构型的研究提供了借鉴。

丹参素在妊娠期高血压疾病中的作用机制研究进展

丹参素是丹参的主要水溶性活性成分,具有多种药理活性,近年来在心血管系统疾病中被广泛研究,并应用于临床治疗。妊娠期高血压的防治至今仍是尚未攻克的难题,由于其与心血管系统疾病相似的发病机制,丹参素在防治妊娠期高血压疾病中的作用也成为学者们研究的热点。研究显示,丹参素及其衍生物可以通过抗氧化应激、抗炎、抗凋亡、抗凝、保护血管内皮等途径发挥作用。本文从上述几个方面探讨其在妊娠期高血压疾病中可能的作用机制。

Effects of Danshensu on maternal syndrome in phosphatidyleserine/ phosphatidylcholine microvesicle induced-mouse model： is it a candidate for preeclampsia remedy？

Backgroud Up to date, there is few satisfactory pharmacotherapy, except for aspirin and heparin, to stop the preeclampsia progression. Although the mechanism of preeclampsia is poorly understood, it has been proven to be associated with coagulation activation. Researches on prophylactic and therapeutic application of anticoagulants may benefit the clinical aspects of preeclampsia individuals. This study aimed to evaluate the effects of Danshensu on maternal syndrome in phosphatidylserine/phosphatidylcholine （PS/PC） microvesicle induced-mouse model. Methods Sixty-six preeclampsia-like pregnant mice, induced by PS/PC microvesicle administration, were randomly divided into six groups. From days 5.5 to 16.5 of pregnancy, each group was respectively treated as follows： a） mice in group C （n=12, control group） were injected with 100 μl of filtered phosphate-buffered saline into the tail vein every day; b） group PE （n=15, preeclampsia model group） were injected in the same way with 100 μl of filtered PS/PC vesicle suspension; c） group H （n=9, group treated with heparin） were injected with 1 unit heparin together with PS/PC vesicle suspension; d） group A （n=10, group treated with aspirin） were injected with 20 μg/g aspirin-DL lysine as well; e） group LD （n=10, group treated with low-dose Danshensu） were injected with 10 μg/g Danshensu; and f） group HD （n=10, group treated with high-dose Danshensu） were injected with 30 μg/g Danshensu. Systolic blood pressure, total urinary protein levels, blood tests for some hemostatic function parameters （mean platelet counts, plasma antithrombin III activity （AT-Ⅲ）, D-D dimmer levels, and thrombin time）, fibrin deposition by phosphotungstic acid hematoxylin staining, and thrombomodulin expression by immunohistochemistry staining in placentas were examined as indices for maternal syndrome. Results Heparin showed significant effects on maternal syndrome of preeclampsia such as hypertension and proteinuria, and different doses of Danshensu also presented the certain effects. High-dose Danshensu and aspirin all demonstrated better effects than low-dose Danshensu on decreasing blood pressure to normal level, while high-dose Danshensu demonstrated 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 were the same as that of aspirin and inferior to that of heparin. High-dose Danshensu＇s better effect on elevating the platelet counts was superior to low-dose Danshensu and aspirin. Low-dose Danshensu＇s obvious effect on decreasing D-D levels was close to heparin and superior to high-dose Danshensu and aspirin. High- and low-dose Danshensu＇s significant effects on reduced thrombin time level are same to heparin. Different anticoagulants all played improvement roles in placental fibrin depositions, but heparin and high-dose Danshensu＇s roles on lowering thrombomodulin expression in placentas were superior to low-dose Danshensu and aspirin. However, anticoagulant function of high-dose Danshensu was still inferior to heparin. We found long-term use of heparin and aspirin, in spite of low-dose administration, could raise the risk of bleeding such as placental abruption and intestinal hemorrhage. But no any side effect was observed in mice treated with different doses of Danshensu in our study.Conclusions Danshensu has proven to be effective and safe in ameliorating the prognosis of maternal syndrome in a preeclampsia mouse model. We suggest long-term provision of low-dose Danshensu in pregnancy, leading to an improvement of preeclampsia syndrome with considerable maternal safety.

A lipophilic prodrug of Danshensu:preparation,characterization,and in vitro and in vivo evaluation

Danshensu [3-(3, 4-dihydroxyphenyl) lactic acid, DSS], one of the significant cardioprotective components, is extracted from the root of Salvia miltiorrhiza. In the present study, an ester prodrug of Danshensu(DSS), palmitoyl Danshensu(PDSS), was synthesized with the aim to improve its oral bioavailability and prolong its half-life. The in vitro experiments were carried out to evaluate the physicochemical properties and stability of PDSS. Although the solubility of PDSS in water was only 0.055 mg·mL^(-1), its solubility in Fa SSIF and Fe SSIF reached 4.68 and 9.08 mg·mL^(-1), respectively. Octanol-water partition coefficient(log P) was increased from-2.48 of DSS to 1.90 of PDSS. PDSS was relatively stable in the aqueous solution in pH range from 5.6 to 7.4. Furthermore, the pharmacokinetics in rats was evaluated after oral administration of PDSS and DSS. AUC and t1/2 of PDSS were enhanced up to 9.8-fold and 2.2-fold, respectively, compared to that of DSS. Cmax was 1.67 ± 0.11 μg·mL^(-1) for PDSS and 0.81 ± 0.06 μg·m L-1 for DSS. Thus, these results demonstrated that PDSS had much higher oral bioavailability and longer circulation time than its parent drug.

Danshensu Ameliorates Cardiac Ischaemia Reperfusion Injury through Activating Sirt1/Fox01/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,H9C2 cells were subjected to 4-h hypoxia followed by 3-h reoxygenation.Then 10|i mol/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(cTnT),reactive oxygen species(ROS),superoxide dismutase(SOD)and glutathione peroxidase(GSH-PX)were detected by enzyme-linked immunosorbent assay(ELISA).Sirt1/FoxO1/Rab7 Signal Pathway was monitored at both protein and mRNA levels.Results:The results showed that IRI not only greatly attenuated cardiac function(LVDP and±dp/dtmax,P<0.01,P<0.05)and increased the level of the marker enzymes(cTnT,LDH,P<0.01)from the coronary effluents,but also markedly induced changes in the structure of cardiomyocytes and contributed to apoptosis,which were mediated by boosted en doge nous ROS.However,after treatment with DSS all above indexes were improved,which was related to activating Sirt1/FoxO1/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/FoxO1/Rab7 signaling.

Sodium Danshensu promotes the healing of stage 2 pressure injury wounds in ischemia/reperfusion injury rat models:possible regulation of apoptosis and inflammatory response

OBJECTIVE:To investigate the protective effect and possible mechanism of sodium Danshensu(SDSS)against pressure injury caused by ischemia/reperfusion(I/R)injury.METHODS:Sprague-Dawley rats were randomly divided into five groups of eight rats each:control group,model group,10 mg/kg SDSS-treated group,20 mg/kg SDSS-treated group,and 40 mg/kg SDSS-treated group.We used two round ferrite magnetic plates of 15 mm diameter and 3 mm thickness to establish stage 2 pressure injury model rats.Each rat was subjected to five cycles of ischemia and reperfusion to induce pressure injury.One cycle consisted of 2 h of ischemia and 0.5 h of reperfusion,which meant that each cycle included2 h of pressure and 0.5 h of pressure relief.The outline of the wound was delineated by butter paper and marker pen,and histopathological changes were observed by hematoxylin and eosin staining.In addition,the number of apoptotic cells and the activity of caspase-3 were assessed by terminal deoxynucleotidyl transferase-mediated nick end labeling and caspase-3 assay kits,respectively.The expression of apoptosis-regulatory proteins and inflammatory mediators was investigated by enzyme-linked immunosorbent assay.RESULTS:Results showed that treatment with SDSS for 7 d after establishing the pressure injury model remarkably improved the healing rate of the wound.SDSS also inhibited the levels of tumor necrosis factor-α,myeloperoxidase,and intercellular cell adhesion molecule-1;decreased the number of apoptotic cells;increased the ratio of B-cell lymphoma-2(Bcl-2)/Bcl-2-associated X(Bax);and regulated the expression and activity of caspase-3.CONCLUSION:Our results suggest that SDSS exhibits a treatment efficacy for pressure injury caused by I/R injury possibly by inhibiting apoptosis and inflammatory response.

超滤工艺对香丹注射液有效成分的影响分析

目的 明确香丹注射液中有效成分在不同超滤条件下的差异,确定香丹注射液超滤工艺的最优条件。方法 采用正交试验设计,以超滤装置进出口压力差、药液温度、药液pH值和超滤膜的截留分子量为因素,以香丹注射液中丹参素与原儿茶醛平均透过率为指标,评估超滤工艺各试验条件下香丹注射液有效成分的差异。结果 正交试验结果显示,丹参素和原儿茶醛平均透过率最高的超滤工艺条件为超滤膜截留分子量30 KDa,超滤装置进出口压力差0.08 MPa,药液pH值9.0,药液温度20℃。丹参素及原儿茶醛平均透过率方差分析结果显示,除超滤膜截留分子量外,均P>0.05。结论 丹参素和原儿茶醛2种成分的超滤工艺影响因素基本一致,超滤膜的截留分子量对香丹注射液有效成分有显著的影响,其余因素的影响不显著。优化后超滤工艺对香丹注射液中有效成分影响较小,相对稳定,适用性强。

高效液相色谱测定复方丹参片中丹参素钠的含量

为建立质控指标,有效控制药品质量,建立测定复方丹参片中丹参素钠的含量的高效液相色谱法。采用Agilent zorbax SB-C_(18)色谱柱(4.6 mm×150 mm,5μm),以甲醇-0.05%磷酸溶液(15∶85)为流动相,流速1.0 mL/min,检测波长280 nm,柱温30℃。丹参素钠在4.979~99.590μg/mL范围内与其峰面积值呈良好的线性关系(r=0.9997),平均回收率为100.36%,RSD为1.7%。24 h内供试品溶液稳定性良好,RSD为0.37%。本方法简便易行,准确可靠,可用于复方丹参片的质量控制。

高效液相色谱法(HPLC)测定乐脉丸中丹参素钠含量

本研究采用高效液相色谱法(HPLC)测定乐脉丸中丹参素钠含量的方法。采用C18柱(4.6mm×250mm,Column),流动相以甲醇-0.2%冰醋酸(10:90),柱温30℃,检测波长280nm,流速1mL/min。结果表明,本方法具有良好的准确性、稳定性和重复性,可为乐脉丸质量控制和产品研发提供有效参考。同时对目前市面上在售的两种品牌乐脉丸中丹参素钠进行测定,通过测定结果发现丹参素钠含量差别微小,质量相当。

丹参素介导Wnt-β-catenin通路增强丙泊酚对小鼠缺血再灌注肾组织损伤的保护作用机制

本研究基于Wnt-β-catenin通路探讨丹参素增强丙泊酚对小鼠缺血再灌注肾损伤的保护机制。本研究将96只健康雄性小鼠按照随机数字表法分为空白对照组(C组)、假手术对照组(S组)、缺血再灌注组(IR组)、缺血再灌注前进行丙泊酚处理组(IRP组)、缺血再灌注前进行丹参素处理组(IRS组)、缺血再灌注前进行丙泊酚联合丹参素处理组(IRPS组),每组16只。C组正常处理,S组仅游离双侧肾脏,IR组建立小鼠缺血再灌注模型,IRP组于小鼠缺血损伤前30min给予丙泊酚处理,IRS组于小鼠缺血损伤前30min给予丹参素处理,IRPS组于小鼠缺血损伤前30 min给予丙泊酚和丹参素处理,其余处理均与IR组相同。手术2 d后,收集小鼠血清和肾组织,检测血清中肌酐(Cr)和尿氮素(BUN)含量,检测肾组织中丙二醛(MDA)含量和超氧化物歧化酶(SOD)活性;HE染色观察肾脏病理学变化;检测肾脏组织Wnt4、β-catenin、Lrp-6的mRNA和蛋白表达水平。结果表明,IR组、IRS组、IRP组、IRPS组的Cr、BUN、MDA水平显著高于C组和S组,且IRPS组的Cr、BUN、MDA水平显著低于IR组、IRP组和IRS组,差异均有显著性(P <0.05);IR组、IRS组、IRP组、IRPS组的SOD活性显著低于C组和S组,且IRPS组的SOD活性显著高于IR组、IRP组和IRS组,差异均有显著性(P <0.05);IR组、IRS组、IRP组、IRPS组的HE染色肾损害程度显著高于C组和S组,且IRPS组的肾损害程度显著低于IR组、IRP组和IRS组,差异均有显著性(P <0.05);IR组、IRS组、IRP组、IRPS组的Wnt4、β-catenin、Lrp-6的mRNA和蛋白水平显著高于C组和S组,IRP组、IRS组、IRPS组的Wnt4、β-catenin、Lrp-6的mRNA和蛋白表达水平显著高于IR组,且IRPS组的Wnt4、β-catenin、Lrp-6的mRNA和蛋白水平显著高于IRP组,差异均有显著性(P <0.05)。丹参素可增强丙泊酚对小鼠缺血再灌注肾组织损伤的保护作用,减小肾组织损伤,其机制可能是通过上调Wnt-β-catenin通路达到肾保护作用。

基于VEGF途径探究丹参素对大鼠视网膜静脉阻塞及对视网膜功能的改善作用机制

目的探讨丹参素对视网膜静脉阻塞(RVO)大鼠的改善作用及可能机制。方法随机选择8只SPF级雄性SD大鼠作为对照组,剩余大鼠左眼利用孟加拉红联合激光光动力法诱导BRVO模型,成模大鼠随机分为模型组、丹参素低(15 mg/kg)、中(30 mg/kg)、高剂量组(60 mg/kg),每组各8只,对照组及模型组大鼠每日经尾静脉注射等量生理盐水,连续21 d。荧光素眼底血管造影(FFA)技术观察视网膜静脉结构;耳静脉微循环检测静脉血液流速;全视野视网膜电图(ffERG)评估视网膜功能;光学相干断层扫描(OCT)测量视网膜层厚度;HE染色观察视网膜组织学变化;RT-qPCR技术检测VEGF mRNA相对表达量;Western blot技术检测视网膜组织中缺氧诱导因子1α(HIF1α)、细胞外信号调节激酶1/2(ERK1/2)、p-ERK1/2、黏着斑激酶(FAK)、p-FAK、VEGF及色素上皮衍生因子(PEDF)蛋白表达情况。结果给药前,与对照组比较,模型大鼠视网膜静脉血流中断、远端血管迂曲、扩张,视网膜厚度增加,ffERG a波、b波振幅降低,耳静脉血液流速降低(P<0.05);给药后,与对照组比较,模型组及丹参素三个给药组大鼠视网膜静脉阻塞处再通,视网膜厚度、ffERG a波、b波振幅及耳静脉血液流速降低(P<0.05),视网膜存在不同程度损伤,模型组大鼠视网膜结构不完整,神经节细胞层(GCL)细胞数目减少,外核层(ONL)感光细胞层几乎丧失,视网膜组织中VEGF mRNA及蛋白和HIF1α、p-ERK1/2、p-FAK蛋白相对表达量升高,PEDF蛋白相对表达量降低(P<0.05);与模型组比较,丹参素三个给药组大鼠上述指标及因子表达水平均逆转(P<0.05),视网膜病理损伤逐渐减轻;丹参素作用效果呈剂量依赖性(P<0.05)。结论丹参素可减轻RVO大鼠视网膜组织损伤,促进阻塞静脉微循环恢复并改善视网膜功能,其作用机制可能与抑制VEGF血管生成相关途径的激活有关。

丹参有效成分最佳配伍对载脂蛋白E^(-/-)小鼠血管外膜成纤维细胞增殖的影响

目的:探讨丹参水溶性有效成分最佳配伍(SABP)对载脂蛋白E^(-/-)小鼠体内和体外血管外膜成纤维细胞(VAFs)增殖的干预作用。方法:对原代培养C57BL/6J小鼠、载脂蛋白E^(-/-)小鼠VAFs,体外分别给予正常培养基、SABP、瑞舒伐他汀。将载脂蛋白E^(-/-)小鼠分为模型组、SABP组、瑞舒伐他汀组,C57BL/6J小鼠作正常对照组,原代培养给药后的各组小鼠VAFs,采用CCK-8法检测VAFs的增殖情况。结果:体外和体内结果均显示,SABP和瑞舒伐他汀均明显抑制了载脂蛋白E^(-/-)小鼠VAFs的增殖,且SABP作用更显著。结论:SABP对载脂蛋白E^(-/-)小鼠VAFs的增殖有明显的抑制作用,有望改善血管重构,成为治疗心血管疾病替代药物。

基于小鼠骨骼肌损伤模型分析丹参素对骨骼肌损伤后修复再生的促进作用

旨在探明丹参单体成分丹参素在骨骼肌损伤后修复中的作用,以期为临床治疗提供理论依据。本试验建立骨骼肌损伤模型,设置丹参素治疗组,通过HE染色和Masson染色观察肌纤维修复过程,Western blot和QPCR方法检测肌再生相关因子(α-Actinin、eMyHC、MyoD、MyoG、Pax7和MSTN),炎症相关因子(IL-6和IL-10),巨噬细胞类型相关因子(F4/80),瘢痕组织形成相关因子(CollagenⅠ和α-SMA)及肌间脂肪相关因子PPAR-γ的表达。结果显示,丹参素降低损伤组腓肠肌对Evans blue的摄取,显著增加α-Actinin和eMyHC因子的表达水平。与损伤组相比,肌肉再生相关因子MyoD、MyoG和Pax7的表达水平在丹参素治疗之后显著升高,而MSTN显著减少。丹参素显著降低损伤组F4/80的表达,加快M1型巨噬细胞向M2型巨噬细胞的转化,降低IL-6 mRNA水平,增加IL-10 mRNA水平。Masson染色结果显示,丹参素减少受损骨骼肌胶原纤维的分布,并且降低CollagenⅠ和α-SMA的表达。同时,丹参素显著增加肌间脂肪因子PPAR-γ的表达。因此,丹参素能够加快肌纤维的再生,促进肌间脂肪的形成,减少纤维化瘢痕的生成,从而加快骨骼肌修复进程,提高骨骼肌修复质量。

丹参及其提取物治疗口腔恶性肿瘤药理机制研究进展

丹参的主要提取物包括丹参酮、丹酚酸、隐丹参酮、丹参素等,其中丹参酮ⅡA可抑制肿瘤细胞有氧糖酵解进程,以阻止肿瘤细胞增殖;丹酚酸B和丹参酮ⅡA可通过抑制磷脂酰肌醇三羟基激酶/蛋白质丝氨酸苏氨酸激酶(phosphoinositide-3 kinase/serine threonine kinase, PI3K/Akt)信号通路传导,促进口腔鳞状细胞癌肿瘤细胞凋亡;丹酚酸B和丹参酮ⅡA可通过下调环氧合酶-2(cyclooxygenase 2,COX-2)/B淋巴细胞瘤-2基因(B-cell lymphoma-2,Bcl-2)的表达,有助于降低头颈部鳞状细胞癌和鼻咽癌的炎症状态,以促使细胞凋亡;丹酚酸B能通过抑制蛋白质丝氨酸苏氨酸激酶/雷帕霉素靶蛋白(serine threonine kinase/Mammalian target of rapamycin, Akt-mTOR)信号通路传导,促使口腔白斑癌变线粒体凋亡和肿瘤细胞凋亡;丹参提取物能激活并提高半胱天冬酶-3(caspase-3)的活性,促使口腔癌耐药细胞凋亡;隐丹参酮可通过抑制Janus激酶/信号传导和转录激活因子3(janus kinase/signal transducers and activators of transcription 3,JAK/STAT3)信号通路的信号传导,促使舌鳞状细胞癌肿瘤细胞凋亡;丹酚酸A和丹参素可通过抑制基质金属蛋白酶2(matrix metalloproteinase 2,MMP-2)的表达,降低肿瘤细胞的侵袭和迁移;丹酚酸B能抑制缺氧诱导因子1α的表达,降低血管内皮细胞和微血管密度水平,抑制口腔癌血管的形成。本文旨在探讨丹参及其提取物治疗口腔恶性肿瘤的药理机制,为临床研究提供参考。