β-Adrenoceptor activates endothelium-dependent release of nitric oxide in rat aorta

AIM:To examine the possible role of agents elevating cAMP to release NO from aortic en-dothelial cells. METHODS:NG-nitro-L-arg inine methylester (L-NAME) , an inhibitor of NO synthase, partially inhibited endothelium-dependent relaxation evoked in phenylephrine-precontracted rings by isoproterenol and abolished relaxation mediated by forskolin 0. 2 umol L-1.RESULTS: In rings without en-dothelium, isoproterenol and forskolin were less effective relaxants and L-NAME had no effect on the responses. In methylene blue-treated rings isoproterenol- and forskolin-induced relaxation were prevented in both en-dothelium-intact and -denuded rings, but the inhibitory effects of methylene blue were significantly more in rings with endothelium than in those without. On the other hand, relaxation induced by sodium nitroprusside was not inhibited by L-NAME, but was inhibited by methylene blue in both the endothelium-intact and -denuded rings. The concentration relaxation curves to sodium nitroprusside after methylene bl     

Deacetylation of 4-(5-acetylamino-2-furyl)thiazole and formation of 1-(4-thiazolyl)-3-cyano-1-propanone by rat liver tissues

The reductive metabolism of the rat carcinogen 4-(5-nitro-2furyl)thiazole (NFT) to 1-4-thiazolyl)-3-cyano-1-propanone (TCP) is reported. Formation of TCP from NFT involved furan ring fission. This could have occurred through involvement of either aminofuran or N-hydroxylaminofuran as precursors. To examine if 4-(5-amino-2-furyl)thiazole is a precursor for TCP, a stable model compound, 4-(5-acetylamino-2-furyl)thiazole (AAFT), was prepared and subjected to enzymatic deacetylation, using rat liver tissue homogenates. AAFT was synthesized by catalytic hydrogenation of NFT with 5% palladium on activated carbon, followed by acetylation with acetic anhydride. AAFT, a white crystalline powder, melted at 168–170°, had an extinction coefficient of 17.9 mM^?1 cm^?1 at 293 nm in ethyl acetate, and exhibited spectroscopic and mass spectral characteristics consistent with the assigned structure. Incubation with rat liver 10,000 g supernatant preparations resulted in the biotransformation of AAFT as evidenced by a decrease in absorption at 290 nm. Incubation of ¹⁴C-labeled AAFT followed by extraction with chloroform-diethyl ether (1:1) resulted in the recovery of a major portion (56%) of the radioactivity in the organic phase when the label was at the 2-position of the thiazole ring, while the major amount (82%) of radioactivity was recovered in the aqueous phase when the 1-¹⁴C-acetyl group was labeled. The radioactivity from the aqueous phase was extractable into the organic phase following acidification to pH 1, an observation consistent with deacetylation. Furthermore, the deacetylation product exhibited a mass spectrum, and retention times in gas and high pressure liquid chromatography, similar to those of synthetic TCP. These data establish 4-(5-amino-2-furyl)thiazole, derived from AAFT by deacetylation, as a precursor for TCP.     

Niflumic acid inhibits chloride conductance of rat skeletal muscle by directly inhibiting the CLC-1 channel and by increasing intracellular calcium

BACKGROUND AND PURPOSE: Given the crucial role of the skeletal muscle chloride conductance (gCl), supported by the voltage-gated chloride channel CLC-1, in controlling muscle excitability, the availability of ligands modulating CLC-1 are of potential medical as well as toxicological importance. Here, we focused our attention on niflumic acid (NFA), a molecule belonging to the fenamates group of non-steroidal anti-inflammatory drugs (NSAID). EXPERIMENTAL APPROACH: Rat muscle Cl(-) conductance (gCl) and heterologously expressed CLC-1 currents were evaluated by means of current-clamp (using two-microelectrodes) and patch-clamp techniques, respectively. Fura-2 fluorescence was used to determine intracellular calcium concentration, [Ca(2+)](i), in native muscle fibres. KEY RESULTS: NFA inhibited native gCl with an IC(50) of 42 muM and blocked CLC-1 by interacting with an intracellular binding site. Additionally, NFA increased basal [Ca(2+)](i) in myofibres by promoting a mitochondrial calcium efflux that was not dependent on cyclooxygenase or CLC-1. A structure-activity study revealed that the molecular conditions that mediate the two effects are different. Pretreatment with the Ca-dependent protein kinase C (PKC) inhibitor chelerythrine partially inhibited the NFA effect. Therefore, in addition to direct channel block, NFA also inhibits gCl indirectly by promoting PKC activation. CONCLUSIONS AND IMPLICATIONS: These cellular effects of NFA on skeletal muscle demonstrate that it is possible to modify CLC-1 and consequently gCl directly by interacting with channel proteins and indirectly by interfering with the calcium-dependent regulation of the channel. The effect of NFA on mitochondrial calcium stores suggests that NSAIDs, widely used drugs, could have potentially dangerous side-effects.     

General anesthetics selectively modulate glutamatergic and dopaminergic signaling via site-specific phosphorylation in vivo

Isoflurane, propofol and ketamine are representative general anesthetics with distinct molecular mechanisms of action that have neuroprotective properties in models of excitotoxic ischemic damage. We characterized the effects of these agents on neuronal glutamate and dopamine signaling by profiling drug-induced changes in brain intracellular protein phosphorylation in vivo to test the hypothesis that they affect common downstream effectors. Anesthetic-treated and control mice were killed instantly by focused microwave irradiation, frontal cortex and striatum were removed, and the phosphorylation profile of specific neuronal signaling proteins was analyzed by immunoblotting with a panel of phospho-specific antibodies. At anesthetic doses that produced loss of righting reflex, isoflurane, propofol, and ketamine all reduced phosphorylation of the activating residue T183 of ERK2 (but not of ERK1); S897 of the NR1 NMDA receptor subunit; and S831 (but not S845) of the GluR1 AMPA receptor subunit in cerebral cortex. At sub-anesthetic doses, these drugs only reduced phosphorylation of ERK2. Isoflurane and ketamine also reduced phosphorylation of spinophilin at S94, but oppositely regulated phosphorylation of presynaptic (tyrosine hydroxylase) and postsynaptic (DARPP-32) markers of dopaminergic neurotransmission in striatum. These data reveal both shared and agent-specific actions of CNS depressant drugs on critical intracellular protein phosphorylation signaling pathways that integrate multiple second messenger systems. Reduced phosphorylation of ionotropic glutamate receptors by all three anesthetics indicates depression of normal glutamatergic synaptic transmission and reduced potential excitotoxicity. This novel approach indicates a role for phosphorylation-mediated down-regulation of glutamatergic synaptic transmission by general anesthetics and identifies specific in vivo targets for focused evaluation of anesthetic mechanisms.     

利拉鲁肽治疗超重的2型糖尿病24例临床观察

目的观察胰高糖素样激动剂1（GLP-1）的类似物利拉鲁肽治疗超重或肥胖的2型糖尿病的有效性及安全性。方法选择于2011—01—12～2013—01—20在本院接受治疗的超重2型糖尿病患者24例,所有患者在二甲双胍及吡咯列酮治疗的基础上加用利拉鲁肽,最初1周给予患者利拉鲁肽0．6mg晚上睡前皮下注射,第2周加量至1．6mg。观察患者的临床疗效,每2周随访1次,观察治疗12周后患者的FPG、2h-PPG、HbAlc、体重、收缩压及舒张压水平变化和不良反应情况。结果治疗12周后患者的FPG、2h-PPG、HbAIc、体重、收缩压及舒张压与治疗前比较,均有明显的改善,差异有统计学意义（P〈0．05）。无严重的不良反应。结论利拉鲁肽治疗超重或肥胖的2型糖尿病疗效肯定,安全,不增加体重,无明显不良反应的另一类糖尿病合适选择的治疗方法。     

Calcineurin-NFAT signaling is involved in phenylephrine-induced vascular smooth muscle cell proliferation

Aim： Catecholamine-induced vascular smooth muscle cell （VSMC） proliferation is one of the major events in the pathogenesis of atherosclerosis and vascular remodeling. The calcineurin-NFAT pathway plays a role in regulating growth and differentiation in various cell types. We investigated whether the calcineurin-NFAT pathway was involved in the regulation of phenylephrine-induced VSMC proliferation. Methods： Proliferation of VSMC was measured using an MTT assay and cell counts. Localization of NFATcl was detected by immunofluorescence staining. NFATcl-DNA binding was determined by EMSA and luciferase activity analyses. NFATcl and calcineurin levels were assayed by immunoprecipitation. Results： Phenylephrine （PE, an α1-adrenoceptor agonist） increased VSMC proliferation and cell number. Prazosin （an α1-adrenoceptor antagonist）, cyclosporin A （CsA, an inhibitor of calcineurin） and chelerythrine （an inhibitor of PKC） decreased PE-induced proliferation and cell number. Additional treatment of VSMC with CsA or chelerythrine further inhibited proliferation and cell number in the chelerythrine-pretreatment group and the CsA-pretreatment group. CsA and chelerythrine alone had no effect on either absorbance or cell number. CsA decreased PE-induced calcineurin levels and activity. NFATcl was translocated from the cytoplasm to the nucleus upon treatment with PE. This translocation was reversed by CsA. CsA decreased the PE-induced NFATcl level in the nucleus. PE increased NFAT＇s DNA binding activity and NFAT-dependent reporter gene expression. CsA blocked these effects. Conclusion： CsA partially suppresses PE-induced VSMC proliferation by inhibiting calcineurin activity and NFATcl nuclear translocation. The calcineurin-NFATcl pathway is involved in the hyperplastic growth of VSMC induced by phenylephrine.     

调肝与健脾对节育器致人子宫异常出血症子宫内膜组织ERK1/2、PI_3K信号通路的影响差异

目的:探讨柴胡止血液对置器后子宫异常出血患者的影响机制。方法:原代培养置器后子宫异常出血患者的宫内膜组织,给含药血清后,分别采用western blot和ELISA的方法测定ERK1/2和PI3K磷酸化的水平。结果:4号组织的归脾汤组升高了PI3K磷酸化水平,P<0.05;1号组织的柴胡止血液组有升高ERK1/2磷酸化水平的趋势;3号组织的柴胡止血液组有升高ERK1/2磷酸化水平的趋势;5号组织的柴胡止血液组有升高PI3K磷酸化水平的趋势;2号组织的归脾汤组有升高ERK1/2磷酸化水平的趋势;4号组织的归脾汤组有升高ERK1/2和PI3K磷酸化水平的趋势。结论:对于出血量大的肝旺患者,柴胡止血液可能通过激活ERK1/2通路来增强血管的收缩从而达到止血的目的;对于出血量较小的肝旺患者证,柴止血液可能通过激活PI3K通路来调节组织抗炎作用,从而达到治疗效果。     

化痰降气胶囊对COPD模型大鼠支气管上皮细胞中多药耐药相关蛋白1功能和表达的影响

目的 通过建立慢性阻塞性肺疾病（COPD)大鼠模型,观察化痰降气胶囊对支气管上皮细胞多药耐药相关蛋白1（MRP1）功能和表达的影响,探讨化痰降气胶囊治疗COPD的作用机制。方法 雄性Wistar大鼠24只随机分为正常对照组、模型组和化痰降气胶囊组（简称化痰降气组)。除正常对照组外,其余各组采用烟、二氧化硫定量熏吸并复合木瓜蛋白酶雾化吸入法,建立COPD大鼠模型;观察化痰降气胶囊治疗后肺功能指标、支气管肺泡灌洗液细胞计数与分类、大鼠的肺组织病理学改变,并采用ELISA方法测定肺组织中白三烯C4 （LTC4）的浓度及免疫组化测定肺支气管上皮MRP1的表达。结果 (1)模型组大鼠的肺顺应性（CL）、第0.3秒用力呼气容积（FEV0.3%)/用力肺活量(FVC)、呼气流量峰值、最大呼气中段流量较正常对照组显著降低（P<0.05)。化痰降气组上述肺功能指标较模型组明显升高(P<0.05)。(2)模型组气道炎症加重, 且出现明显肺气肿;化痰降气组也出现了炎症和肺气肿, 但程度较轻。(3)与正常对照组比较,模型组及化痰降气组肺组织中的LTC4均显著升高(P<0.01)。与模型组比较,化痰降气组中肺组织的LTC4显著下降(P<0.05)。(4)与正常对照组比较,模型组肺支气管上皮MRP1蛋白表达显著降低(P<0.01)。与模型组比较,化痰降气组肺支气管上皮MRP1蛋白表达显著增强(P<0.05)。结论 化痰降气胶囊可能通过影响慢性阻塞性肺疾病状态下MRP1功能和表达,从而达到有效减轻COPD大鼠的气道炎症, 延缓肺功能恶化等COPD发生和发展的目的。     

川芎嗪干预BMNCs移植对MCAO小鼠血清TGF—β1的影响

目的观察川芎嗪干预BMNCs移植对MCAO小鼠血清TGF-β1的影响。方法采用线栓法制作小鼠MCAO缺血再灌注模型,分离同种系小鼠的骨髓单个核细胞,尾静脉注射BMNCs于造模成功的MCAO小鼠体内,腹腔注射川芎嗪注射液进行干预,以ELISA法测定血清TGF-β1水平。结果川芎嗪干预的BMNCs移植能够使血清TGF-β1值降低更为显著（P〈0．05）。结论川芎嗪干预的BMNCs移植更能促进TGF-β1向脑缝缸局部转移,改善小鼠脑缺血症状。     

黄芪沙参煎剂对博莱霉素诱导的大鼠肺纤维化模型的干预作用

目的：观察中药黄芪沙参煎剂对肺纤维化大鼠病理、肺泡灌洗液中细胞因子（TGF-β1）表达的影响,探讨黄芪沙参煎剂作用机制,为其临床治疗肺纤维化提供实验依据。方法：取体重190-210g的健康雌性SD大鼠90只,采用随机数字表随机分为6组,每组15只。博莱霉素（B组）、黄芪沙参煎剂高剂量组（H组）、中剂量组（M组）、低剂量组（L组）、强的松组（P组）、生理盐水组（N组）,各组大鼠于造模后第7、14和28天分别随机处死5只。行HE染色、Masson三色染色;右肺支气管肺泡灌洗,收集灌洗液（BALF）,用ELISA法测BALF上清液细胞因子TGF-β1的表达。结果：H组、M组、P组与B组早期肺泡炎和晚期肺纤维化程度比较,差异有统计学意义（P〈0.01）;第7、14、28天P组、H组、M组的BALF中TGF-β1水平与Β组相比差异有显著统计学意义（Ρ〈0.01）。结论：黄芪沙参煎剂具有与强的松相似的疗效,但副反应小、服用方便,可以明显减少肺泡炎症及肺纤维化形成。黄芪沙参煎剂治疗肺纤维化机制可能与早期抑制炎症细胞浸润,减少炎症细胞分泌释放细胞因子TGF-β1,从而抑制成纤维细胞产生胶原和ECM积聚,起到对肺纤维化的治疗作用。