Effect of Jiere Xingshen Injection on cAMP,IL-1β content in hypothalamus of ET febrile rabbits

AIM: The effects of Jiere Xingshen(JRXS) Injection on cAMP, IL-1_β content in hypothalamus (HP) of endotoxin(ET)-induced feverish rabbits were studied. METHODS: The ET-induced fever model was established in rabbits and the cAMP content in hypothalamus (HP) and csf, IL-1_β content in HP were determined by radioimmunoassay following intravenous infusion of JRXS. RESULTS: In ET group, the ΔT[(0.40±0.11)℃], TRI₁(1.78±0.79), cAMP content in HP[(2.90±0.40)nmol/g], cAMP content in csf[(0.40±0.11)nmol/L)], IL-1_β content in HP[(6.08±0.79)ng/g] were higher than that of NS and JRXS+ET group (P＜0.01). In JRXS+ET group, the ΔT[(0.10±0.10)℃], TRI₁(0.36±0.64), cAMP content in HP[(1.37±0.27)nmol/g], cAMP content in csf[(14.4±3.69)nmol/L)], IL-1_β content in HP[(2.90±0.37)ng/g] were very close to that of NS group but lower than that of the ET group (P＜0.01);The cAMP content in HP and csf, IL-1_β content in HP paralleled with the fluctuation of temperature. CONCLUTION: JRXS Injection has significant inhibitory effect on ET-induced fever by inhibiting cAMP and IL-1_β production in hypothalamus.     

Transfer of siRNA against CD73 suppresses human breast cancer cell line MB-MDA-231 adhesion to extracellular matrix

AIM:To study the effect of CD73 on breast cancer cell line MB-MDA-231 adhesion to extracellular matrix.METHODS: ① CD73 siRNA plasmid was constructed and transfected into MB-MDA-231 cells by lipofectamine 2000.② The transfection efficiency was analyzed by flow cytometry using eGFP as a marker gene.Stable transfected MB-MDA-231 cells were selected using G418.③ RT-PCR and Western blotting analysis of CD73 expression in MB-MDA-231 cells was performed.④ The effects of CD73 on MB-MDA-231 cells adhesion to extrace...     

北冬虫夏草发酵液中虫草素和腺苷含量的HPLC分析

试用高效液相色谱法同时检测了5株北冬虫夏草菌株发酵液中虫草素和腺苷的含量,并系统地研究了各种发酵条件对虫草素和腺苷含量的影响.结果表明,虫草素和腺苷在所设色谱条件下得到了很好的分离,二组分的线性关系良好,回收率满意,其中虫草素峰保留时间为11.443min,腺苷峰保留时间为9.054min;虫草素的回收率为103.22%,RSD=4.4%,腺苷的回收率为98.42%,RSD=3.2%;虫草素和腺苷的线性范围分别为0.49～98μg/mL,0.46～92μg/mL.5株待测菌株发酵液中的菌丝干重第5天都达到了最大值,其中Cordycepsmilitaris G5124菌株的虫草素含量第9天达最大值,腺苷含量第1天达最大值.试验结果还表明,不同菌株发酵液中的虫草素和腺苷含量具有明显差异,这可能与菌株本身的遗传特性有关.其中C.militaris G（04）5f菌株的虫草素含量最高,可用作发酵生产虫草素的高功能菌株,进一步试验表明,当起始pH值为9时该菌株的虫草素含量达到最高.     

AMP对铝胁迫诱导丹波黑大豆柠檬酸分泌及其铝耐受性的影响

为了考察AMP(Adenosine 5’-monophosphate)对铝耐受型丹波黑大豆(RB)柠檬酸的分泌及铝抗性的影响,在50 mmol·L-1铝溶液中添加100 mmol·L-1AMP共处理RB,结果表明RB根的相对生长率显著小于单独用50 mmol·L-1铝胁迫处理的植株。免疫共沉淀分析显示铝胁迫下AMP的存在降低RB根中14-3-3蛋白与磷酸化质膜H+-ATP酶的相互作用,使根尖质膜H+-ATP酶活性降低约1倍,氢泵活性和H+分泌能力显著降低,根柠檬酸的分泌量减少约1倍。说明铝胁迫下AMP通过抑制14-3-3蛋白和质膜H+-ATP酶的互作来减少RB根尖的氢泵活性和柠檬酸分泌作用,从而降低RB根的相对生长率及其对铝胁迫的耐受性。     

体外25℃家蚕卵酯酶A4的ATP酶活性变化及其与滞育性的关系

从家蚕卵纯化出EA4酶蛋白 ,调查了温度对EA4的ATP酶最高活性和其出现时间的影响以及与蚕卵滞育性的关系。EA4的酶活性反应温度为 2 5℃时比 5℃时提高约 1 0倍。从滞育性卵和盐酸处理卵纯化EA4 ,或者用EA4的活性抑制多肽 (PIN)体外处理酶蛋白质 ,2 5℃中EA4的最高活性出现时间比 5℃中显著提早。随纯化时卵龄或PIN处理时间的改变 ,2 5、5℃中EA4的最高活性出现时间以及与蚕卵 5℃冷藏时滞育发育时间有相同趋势的变化。体外 2 5℃和体外 5℃一样 ,EA4的最高活性出现时间与蚕卵的滞育发育有密切的关系     

Study of platelet aggregations in patients with coronary heart disease and the effect of ticlopidine treatment

AIM: To investigate the platelet aggregations in patients with coronary heart disease(CHD) and the effect of ticlopidine treatment. METHODS: Platelet aggregations induced by adenosine diphosphate(ADP), epinephrine(EPI), collagen(Coll), arachidonic acid(ACA) in CHD group before and after ticlopidine treatment were measured by turbidity assay. RESULTS: Maximum ratios of platelet aggregations (max%) induced by ADP, EPI in CHD group (0.78±0.23, 0.86±0.25) were significantly higher than that of control group (0.65±0.19, 0.73±0.21, P＜0.05). After the treatment with ticlopidine, they were lowered obviously (0.68±0.18, P＜0.05;0.75±0.20, P＜0.05). There were no difference in max% induced by Coll and ACA between two groups and there were no significantly changes of max% induced by Coll and ACA by ticlopidine in CHD group. CONCLUSION: The platelet aggregations were increased in patients with coronary heart disease and could be inhibited by ticlopidine.     

蚕蛹虫草中虫草素和腺苷含量影响因素研究

以双蒸水为溶剂,用反相高效液相色谱-紫外检测法测定蚕蛹虫草样品中虫草素和腺苷含量,并用该方法测定了19-3、17-3、MS、1-1以及1-Y等12种不同虫草菌株栽培的蚕蛹虫草、不同品质的蚕蛹虫草以及蚕蛹虫草不同组织中的虫草素和腺苷的含量。结果表明:1-Y菌株的虫草素和腺苷含量均最高,质量分数达15.45 mg/g和4.40 mg/g;不同品质的蚕蛹虫草中虫草素则以感染而未出草的僵蚕最高;蚕蛹虫草的僵蚕体对虫草素的富集能力高于子座,蚕蛹虫草的子座对腺苷的富集能力高于僵蚕体。该结果可对探求高虫草素含量和高腺苷含量的虫草材料提供理论依据。     

SKE法在ATP生物发光快速检测中的应用

通过研究ATP缓冲液种类和浓度,ATP提取时间等实验因素,优化SKE法中的细菌ATP提取条件,并对优化后的SKE法和TCA法进行对比,得出采用0.2mol/L Tis-乙酸作为ATP缓冲液,提取时间为1min的SKE法作为食品样品前处理的最佳条件。     

Regulation of AMPK/PPARα/SCAD signal pathway on cardiac hypertrophy

AIM：To study the effect of short-chain acyl-coenzyme A dehydrogenase (SCAD）on cardiac hypertrophy and to explore the role of adenosine monophosphate-activated protein kinase (AMPK)/peroxisome proliferator-activated receptor α (PPARα） signal pathway in the regulation of SCAD during the development of cardiac hypertrophy. METHODS：The optimal sequence of SCAD interference was chosen by Western blotting and real-time PCR. The cardiomyocytes were treated with fenofibrate (10 μmol/L) for 24 h and subsequently stimulated with the optimal sequence of SCAD interference. The changes of SCAD expression at mRNA and protein levels, the enzyme activity of SCAD, the cardiomyocyte surface area and free fatty acids were determined. Using real-time PCR for analyzing the markers of cardiac hypertrophy, the mRNA expression of atrial natriuretic factor (ANF） and brain natriuretic peptide (BNP) was detected to judge the development of cardiac hypertrophy. The cardiomyocytes were treated with fenofibrate (10 μmol/L) or AMPK activator 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR, 0.5 mmol/L) for 30 min and subsequently stimulated with phenylephrine (PE, 20 μmol/L) for 24 h. The changes of cardiomyocyte surface area, free fatty acids, and the expression of SCAD, PPARα and p-AMPKα (T172) at mRNA and protein levels were observed. RESULTS：The effect of optimal sequence siRNA-1186 and PE on the cardiomyocytes was the same. Compared with control group, the expression of ANF and BNP at mRNA level, the cardiomyocyte surface area and free fatty acids were increased obviously in siRNA-1186 group. After pretreated with fenofibrate (10 μmol/L), the expression of PPARα and SCAD, and the enzyme activity of SCAD were significantly increased, while the free fatty acids were decreased, indicating that fenofibrate prevented the development of cardiac hypertrophy induced by knockdown of SCAD. Compared with control group, the expression of SCAD, PPARα and p-AMPKα (T172) at mRNA and protein levels was significantly down-regulated, and the enzyme activity of SCAD was obviously decreased in PE group. Compared with PE group, the expression of SCAD, PPARα and p-AMPKα (T172) was significantly up-regulated, and the cardiomyocyte surface area and the content of free fatty acids were obviously decreased in the cardiomyocytes pretreated with fenofibrate or AICAR for 30 min. CONCLUSION：Down-regulation of SCAD is related to the cardiac hypertrophy and energy metabolism. AMPK/PPARα/SCAD signaling pathway may regulate cardiac hypertrophy directly.     

Exogenous ATP induces formation of membrane pore in PC12 cells

AIM：To investigate the formation of membrane pore in PC12 cells induced by exogenous adenosine triphosphate (ATP) and to identify the key molecular targets. METHODS：PC12 cells were treated with different concentrations of ATP to establish the injury model. The morphological change was observed under an inverted phase-contrast microscope. The viability of the PC12 cells was measured by CCK-8 assay. Fluorescent dye YO-PRO-1 was used to detect the membrane permeability. The expression of P2X7 receptor and pannexin 1 (Panx1) at mRNA and protein levels was assessed by real-time PCR and Western blotting. RESULTS：After exposed to ATP (1 mmol/L, 3 mmol/L and 5 mmol/L) for 3 h, the PC12 cells became edematous, and the number of adherent cells decreased gradually in a dose-dependent manner. The cell viabilities in 3 mmol/L ATP group and 5 mmol/L ATP group were significantly decreased compared with control group (P<0.05). YO-PRO-1 uptake in the PC12 cells exposed to ATP (0, 1, 3 and 5 mmol/L) for 15 min, 30 min and 60 min increased in a dose-dependent and time-dependent manner. The cell viability increased and the intracellular fluorescence intensity induced by ATP were significantly antagonized in brilliant blue G (a P2X7 receptor inhibitor) pretreatment group (P<0.05), whereas it did not change in carbenoxolone (a Panx1 inhibitor) pretreatment group (P>0.05). The expression of P2X7 receptor at mRNA and protein levels was significantly increased (P<0.05), but the expression of Panx1 was not changed (P>0.05) when PC12 cells were exposed to ATP for 3 h. CONCLUSION：Extracellular ATP at high concentration may induce membrane pore formation with the expression and activation of P2X7 receptor in PC12 cells.