糖尿病大鼠冠状动脉平滑肌细胞体外培养方法的探讨

目的探讨糖尿病大鼠冠状动脉平滑肌细胞体外培养方法,建立糖尿病大鼠冠状动脉平滑肌细胞模型,为糖尿病性冠心病的研究奠定基础。方法建立糖尿病大鼠模型,用酶消化法体外培养冠状动脉血管平滑肌细胞。结果糖尿病大鼠造模成功后分离冠状动脉,用酶消化法培养糖尿病大鼠血管平滑肌细胞,24h更换培养液液,培养7~10d细胞重叠生长达多层,高低起伏呈“峰—谷”状。细胞α-actin免疫组织化学染色鉴定为平滑肌细胞。结论糖尿病大鼠血管平滑肌细胞增殖速度快,培养条件要求严格,在形态学上与正常大鼠平滑肌细胞相同。     

原花青素对人冠状动脉平滑肌细胞增殖和凋亡的影响

目的研究原花青素对人冠状动脉平滑肌细胞增殖和凋亡的影响。方法采用MTT实验检测原花青素对体外培养的人冠状动脉平滑肌细胞增殖的影响,运用流式细胞术、基因组DNA电泳观察凋亡特征性＂梯状＂条带检测细胞凋亡,采用Caspase活性定量检测试剂盒分析Caspase-9、Caspase-3的活性。结果原花青素对人冠状动脉平滑肌细胞具有明显的抑制作用,且其作用有剂量依赖性。流式细胞仪检测显示原花青素可以显著诱导人冠状动脉平滑肌细胞凋亡,DNA电泳显示原花青素作用于人冠状动脉平滑肌细胞后出现凋亡细胞特有的DNA阶梯状条带。原花青素作用后人冠状动脉平滑肌细胞Caspase-9、Caspase-3活性明显升高。结论原花青素可能通过线粒体信号通路抑制人冠状动脉平滑肌细胞的生长并诱导其凋亡。     

大鼠肺动脉平滑肌细胞培养及对不同类型激动剂刺激产生的反应

目的分离和培养SD大鼠肺动脉平滑肌细胞（PASMCc）,并检测其功能状态。方法显微分离肺内小动脉,并在含胶原酶（1750 U·mL^-1）和木瓜蛋白酶（9．5U·mL^-1）的低钙HBSS溶液中酶解和培养PASMCs,采用动态细胞荧光成像技术检测PASMCs胞浆游离Ca^2＋浓度（[Ca^2＋]i）的变化。结果在18～24h内可获得PASMCs,并可观察到环匹阿尼酸和5-HT可引起PASMCs[Ca^2＋]i的升高效应。结论大鼠PASMCs一步酶消化法,方法简便实用。所分离的PASMCs细胞形态和功能正常,适用于动态细胞荧光成像技术检测实验及PASMCs信号转导功能的研究。     

Direct activation of endothelial NO pathway by Ba2+ in canine coronary artery

1. We have reported that Ba²⁺ causes endothelium-dependent relaxation of canine coronary arteries through NO synthesis in Ca²⁺-free and depolarizing solution. To determine the cellular mechanisms by which the endothelium-dependent relaxation occurs, we used fura-2 fluorometry (F₃₅₀ and F₃₉₀; excitation wavelengths, 350 and 390 nm, respectively) and estimated the intracellular Ba²⁺ concentration in endothelial and vascular smooth muscle cells.
2. Ba²⁺ (10⁻³ M) increased the fura-2 ratio (F₃₅₀/F₃₉₀) recorded from a combined preparation of smooth muscle and endothelium (0.445±0.073, n=4) and contracted the arteries in the presence of 80 mM K⁺ (0.22±0.06 g, n=4).
3. Diltiazem (3×10⁻⁶ M) blocks Ba²⁺ entry into vascular smooth muscle cells via L-type Ca²⁺ channels. In this condition, Ba²⁺ increased the fura-2 ratio in endothelial cells (0.141±0.014, n=5) and relaxed the underlying smooth muscle (0.08±0.01 g, n=5) by a mechanism which was sensitive to 10⁻⁴ M N^G-methyl-L-arginine (L-NMMA).
4. Ba²⁺-induced relaxation was not attenuated with repeated application and was elicited even after endothelium-dependent relaxations in response to 10⁻⁶ M bradykinin were abolished due to tachyphylaxis. Neither 10⁻² M caffeine nor 10⁻⁶ M thapsigargin had effect upon Ba²⁺-induced relaxation.
5. To further rule out changes in intracellular Ca²⁺ as a mechanism of Ba²⁺-induced relaxation, fura-2 fluorescence was measured at the isosbestic wavelengths for Ca²⁺ (360 nm) and Ba²⁺ (370 nm) in endothelium-intact arteries. Ba²⁺ altered F₃₆₀, but not F₃₇₀, suggesting little or no contribution of intracellular Ca²⁺ to the phenomenon of Ba²⁺-induced relaxation.
6. These results suggest that the Ba²⁺-induced relaxation is due to its direct activation of endothelial NO synthesis without mobilization of intracellular Ca²⁺.

     

粉防己碱诱发大鼠尾动脉收缩的新特性

AIM: In an attempt to pharmacologically characterize the Chinese antihypertensive drug, tetrandrine, we observed in rat-tail arteries, an unusual contraction in tissues that were stimulated with high [KCI] and not those stimulated with phenylephrine. The characteristics of this contraction were studied. METHODS: Segments of perfused ventral rat-tail arteries (RTA) were contracted with a depolarizing concentration (120 mmol/L) of KCI or with phenylephrine (3.0 umol/L). At peak contraction, they were exposed to tetrandrine (40 umol/L), which caused marked relaxation in each case. Washing the RTA led to an unusual, slowly-declining contraction, hereafter referred to as tetrandrine-induced contraction (TIC) which was also observed when the tissues were exposed to 80 umol/L, but not 10 umol/L or 20 umol/L of tetrandrine. RESULTS: Pretreatment with phentolamine (non-selective a-adrenoceptor antagonist), prazosin (selective aradrenoceptor antagonist) or 6-hydroxydopamine (for denervation), but not rauwolscine     

Reduced store-operated Ca2+ entry impairs mesenteric artery function in response to high external glucose in type 2 diabetic ZDF rats

Diabetes is a major risk factor for cardiovascular disease, affecting both endothelial and smooth muscle cells. Store-operated Ca²⁺ channels (SOCCs) have been implicated in many diabetic complications. Vascular dysfunction is common in patients with diabetes, but the role of SOCCs in diabetic vasculopathy is still unclear. Our research aimed to investigate the effects of high glucose (HG) on store-operated Ca²⁺ entry (SOCE) in small arteries. Small mesenteric arteries from type 2 diabetic Zucker fatty rats (ZDF) versus their non-diabetic controls (Zucker lean, ZL) were examined in a pressurized myograph. Vascular smooth muscle cells (VSMC) were isolated and intracellular Ca²⁺ was measured (Fura 2-AM). A specific protocol to deplete intracellular Ca²⁺ stores and thereby open SOCCs, as well as pharmacological SOCE inhibitors (SKF-96365, BTP-2), were used to artificially activate and inhibit SOCE, respectively. High glucose (40 mmol/L) relaxed arteries in a SKF-sensitive manner. Diabetic arteries exhibited reduced HG-induced relaxation, as well as reduced contraction after Ca²⁺ replenishment. Further, the rise in intracellular Ca²⁺ on account of SOCE is diminished in diabetic versus non-diabetic VSMCs and was insensitive to HG in diabetic VSMCs. The expression of SOCC proteins was measured, detecting a downregulation of Orai1 in diabetes. In conclusion, diabetes leads to a reduction of SOCE and SOCE-induced contraction, which is unresponsive to HG-mediated inhibition. The reduced expression of Orai1 in diabetic arteries could account for the observed reduction in SOCE.     

Effect of calcium on the vascular contraction induced by cobra venom cardiotoxin

1. The cytotoxic effects of cardiotoxin (CTX) purified from Cobra venom were tested in endothelium-denuded rat aortic ring preparations in tissue organ baths and the effect of extracellular Ca2+ on the cytotoxic effect of CTX was investigated using a digital dynamic calcium imaging technique. 2. At 10 micromol/L, CTX induced a slowly developing and sustained contraction that amounted to approximately 50% of the maximal contraction induced by 80 mmol/L KCl. At high concentrations (> 15 micromol/L), CTX caused irreversible damage to the smooth muscle contractile function. However, washout of CTX at its peak contraction did not affect the subsequent contraction to either KCl or phenylephrine. 3. Contraction induced by CTX was dependent on the Ca2+ concentration in the external solution. A maximal contractile response to CTX was obtained in medium containing 1-2.5 mmol/L Ca2+. This contractile response induced by CTX decreased with higher Ca2+ concentrations and was completely diminished when 7 mmol/L Ca2+, 3 mmol/L Ni2+ or 30 micromol/L tetrandrine (a non-selective calcium channel blocker) was present in the external solution before addition of CTX to the bath. 4. The above observations were supported by the calcium imaging work performed with cultured aortic smooth muscle cells from Wistar-Kyoto rats, in which CTX was shown to induce the elevation of cytosolic Ca2+ in the presence, but not in the absence, of 2.5 mmol/L extracellular Ca2+. Increasing the extracellular Ca2+ concentration to 7 mmol/L, the addition of 3 mmol/L Ni2+ or inclusion of 30 micro mol/L tetrandrine inhibited the elevation of cytosolic Ca2+ induced by CTX. 5. These results suggest that: (i) a CTX-sensitive internal calcium store does not exist in rat aortic smooth muscle; (ii) the contractile effect CTX is associated with a Ca2+ influx process; and (iii) CTX interacts extracellularly with the plasma membrane at the level of the calcium channels, as well as anionic sites to which Ca2+ and other inorganic cations bind.     

Oral salmon calcitonin attenuates hyperglycaemia and preserves pancreatic beta-cell area and function in Zucker diabetic fatty rats

BACKGROUND AND PURPOSE

Oral salmon calcitonin (sCT), a dual-action amylin and calcitonin receptor agonist, improved glucose homeostasis in diet-induced obese rats. Here, we have evaluated the anti-diabetic efficacy of oral sCT using parameters of glycaemic control and beta-cell morphology in male Zucker diabetic fatty (ZDF) rats, a model of type 2 diabetes.

EXPERIMENTAL APPROACH

Male ZDF rats were treated with oral sCT (0.5, 1.0 or 2 mg·kg⁻¹) or oral vehicle twice daily from age 8 to 18 weeks. Zucker lean rats served as control group. Fasting and non-fasted blood glucose, glycosylated haemoglobin (HbA1c) and levels of pancreas and incretin hormones were determined. Oral glucose tolerance test and i.p. glucose tolerance test were compared, and beta-cell area and function were evaluated.

KEY RESULTS

Oral sCT treatment dose-dependently attenuated fasting and non-fasted hyperglycaemia during the intervention period. At the end of the study period, oral sCT treatment by dose decreased diabetic hyperglycaemia by ∼9 mM and reduced HbA1c levels by 1.7%. Furthermore, a pronounced reduction in glucose excursions was dose-dependently observed for oral sCT treatment during oral glucose tolerance test. In addition, oral sCT treatment sustained hyperinsulinaemia and attenuated hyperglucagonaemia and hypersecretion of total glucagon-like peptide-1 predominantly in the basal state. Lastly, oral sCT treatment dose-dependently improved pancreatic beta-cell function and beta-cell area at study end.

CONCLUSIONS AND IMPLICATIONS

Oral sCT attenuated diabetic hyperglycaemia in male ZDF rats by improving postprandial glycaemic control, exerting an insulinotropic and glucagonostatic action in the basal state and by preserving pancreatic beta-cell function and beta-cell area.     

Effect of tetramethylpyrazine on potassium channels to lower calcium concentration in cultured aortic smooth muscle cells

1. Tetramethylpyrazine (TMP) is one of the active principles contained in Ligusticum chuanxiong Hort. (Umbelliferae), a herb that has been used widely in China to treat vascular disorders. 2. In an attempt to elucidate the possible mechanisms of action of TMP, the effect of TMP on intracellular calcium concentrations ([Ca2+]i) was investigated in cultured vascular smooth muscle (A7r5) cells using the Ca(2+)-sensitive dye Fura-2 as an indicator. 3. The increase in [Ca2+]i in A7r5 cells produced by vasopressin (1 micromol/L) or phenylephrine (1 micromol/L) was attenuated by TMP in a concentration-dependent manner. Only inhibitors specific to ATP-sensitive potassium (KATP) channels or small conductance calcium-activated potassium (SKCa) channels attenuated the action of TMP (10 micromol/L) on [Ca2+]i. However, blockers of other K+ channels failed to modify the inhibitory action of TMP (10 micromol/L) on [Ca2+]i. 4. The action of TMP on membrane potential in A7r5 cells was monitored by the fluorescence of bisoxonol. Tetramethylpyrazine caused a concentration-dependent inhibition of changes in membrane potential elicited by KCl (20 mmol/L) or phenylephrine (1 micro mol/L), an effect that was totally reversed by glibenclamide (100 micromol/L) and apamin (100 nmol/L) in combination. 5. The results obtained indicate that the decrease in [Ca2+]i in A7r5 cells produced by TMP is mediated mainly by opening of KATP and/or SKCa channels.     

氨茶碱调节大鼠支气管平滑肌L型钙通道的活动性

目的 探讨氨茶碱对大鼠支气管平滑肌细胞L型钙通道(L-L_(Ca))的作用。方法 将20只大鼠随机分为2组,A组为正常对照组(n=10),B组为氨茶碱组(n=10),观测大鼠支气管平滑肌细胞L-L_(Ca)在正常时及给予氨茶碱治疗后的变化。结果 氨茶碱治疗后大鼠的L-L_(Ca)与正常对照组大鼠通道开放概率相比差异有显著性(P<0.05)。氨茶碱组通道开放时间常数(τ_(O1)和τ_(O2))不变,关闭时间常数(T_(c1)和T_(c2))延长,与正常对照组相比差异有显著性(P<0.05)。大鼠L-钙通道的活动经氨茶碱治疗后活动减弱,表现为通道开放时间常数不变,关闭时间常数延长。结论 氨茶碱通过缩短L-L_(Ca)的开放时间来抑制支气管平滑肌细胞的收缩,可显著抑制大鼠L-L_(Ca)的活动,促进大鼠支气管平滑肌细胞的舒张。     

粉防己碱对大鼠肺动脉平滑肌细胞小电导钙激活钾通道的作用

目的　研究粉防己碱 (Tet)对大鼠肺动脉平滑肌细胞小电导钙激活钾通道 (SKCa)的作用。方法　内面朝外膜片箝单通道记录法。结果　Tet 7 5 μmol·L-1对电导值为 10pS的SKCa无明显影响 ,15 μmol·L-1可增加SKCa开放的概率 ,改变通道的开放和关闭模式 ,30 μmol·L-1降低通道的开放概率 ,通道开放以短暂簇状为主。结论　Tet对肺动脉平滑肌SKCa的作用与Tet的浓度有关 ,合适浓度下可增加通道的开放 ,K+ 外流增多 ,与Tet降低肺动脉张力有关。     

Mechanism of asynchronous Ca2+ waves underlying agonist-induced contraction in the rat basilar artery

Background and purpose:

Uridine 5''-triphosphate (UTP) is a potent vasoconstrictor of cerebral arteries and induces Ca²⁺ waves in vascular smooth muscle cells (VSMCs). This study aimed to determine the mechanisms underlying UTP-induced Ca²⁺ waves in VSMCs of the rat basilar artery.

Experimental approach:

Isometric force and intracellular Ca²⁺ ([Ca²⁺]_i) were measured in endothelium-denuded rat basilar artery using wire myography and confocal microscopy respectively.

Key results:

Uridine 5''-triphosphate (0.1–1000 µmol·L⁻¹) concentration-dependently induced tonic contraction (pEC₅₀ = 4.34 ± 0.13), associated with sustained repetitive oscillations in [Ca²⁺]_i propagating along the length of the VSMCs as asynchronized Ca²⁺ waves. Inhibition of Ca²⁺ reuptake in sarcoplasmic reticulum (SR) by cyclopiazonic acid abolished the Ca²⁺ waves and resulted in a dramatic drop in tonic contraction. Nifedipine reduced the frequency of Ca²⁺ waves by 40% and tonic contraction by 52%, and the nifedipine-insensitive component was abolished by SKF-96365, an inhibitor of receptor- and store-operated channels, and KB-R7943, an inhibitor of reverse-mode Na⁺/Ca²⁺ exchange. Ongoing Ca²⁺ waves and tonic contraction were also abolished after blockade of inositol-1,4,5-triphosphate-sensitive receptors by 2-aminoethoxydiphenylborate, but not by high concentrations of ryanodine or tetracaine. However, depletion of ryanodine-sensitive SR Ca²⁺ stores prior to UTP stimulation prevented Ca²⁺ waves.

Conclusions and implications:

Uridine 5''-triphosphate-induced Ca²⁺ waves may underlie tonic contraction and appear to be produced by repetitive cycles of regenerative Ca²⁺ release from the SR through inositol-1,4,5-triphosphate-sensitive receptors. Maintenance of Ca²⁺ waves requires SR Ca²⁺ reuptake from Ca²⁺ entry across the plasma membrane via L-type Ca²⁺ channels, receptor- and store-operated channels, and reverse-mode Na⁺/Ca²⁺ exchange.     

Increased inward rectifier K+ current of coronary artery smooth muscle cells in spontaneously hypertensive rats; partial compensation of the attenuated endothelium-dependent relaxation via Ca2+-activated K+ channels

Endothelium-dependent vasorelaxation is partly mediated by small-conductance (SK3) and intermediate-conductance Ca²⁺-activated K⁺ channels (SK4) in the endothelium that results in endothelium-dependent hyperpolarization (EDH). Apart from the electrical propagation through myoendothelial gap junctions, the K⁺ released from the endothelium facilitates EDH by increasing inward rectifier K⁺ channel (Kir) conductance in smooth muscle cells. The EDH-dependent relaxation of coronary artery (CA) and Kir current in smooth muscle cells (CASMCs) of hypertensive animals are poorly understood despite the critical role of coronary flow in the hypertrophic heart. In spontaneously hypertensive (SHR) and control (WKY) rats, we found attenuation of the CA relaxation by activators of SK3 and SK4 (NS309 and 1-EBIO) in SHR. In isolated CASMCs, whole-cell patch-clamp study revealed larger I_Kir in SHR than WKY, whereas the myocytes of skeletal and cerebral arteries showed smaller I_Kir in SHR than WKY. While the treatment with I_Kir inhibitor (0.1 mmol/L Ba²⁺) alone did not affect the WKY-CA, the SHR-CA showed significant contractile response, suggesting relaxing influence of the higher I_Kir in the CASMCs of SHR. Furthermore, the attenuation of NS309-induced relaxation of CA by the combined treatment with 0.1 mmol/L Ba²⁺ was more prominent in SHR than WKY. Our study firstly shows a distinct increase of I_Kir in the CASMCs of SHR, which could partly compensate for the attenuated relaxation via endothelial SK3 and SK4.     

Store-operated calcium entry in vascular smooth muscle

In non-excitable cells, activation of G-protein-coupled phospholipase C (PLC)-linked receptors causes the release of Ca(2+) from intracellular stores, which is followed by transmembrane Ca(2+) entry. This Ca(2+) entry underlies a small and sustained phase of the cellular [Ca(2+)](i) increases and is important for several cellular functions including gene expression, secretion and cell proliferation. This form of transmembrane Ca(2+) entry is supported by agonist-activated Ca(2+)-permeable ion channels that are activated by store depletion and is referred to as store-operated Ca(2+) entry (SOCE) and represents a major pathway for agonist-induced Ca(2+) entry. In excitable cells such as smooth muscle cells, Ca(2+) entry mechanisms responsible for sustained cellular activation are normally considered to be mediated via either voltage-operated or receptor-operated Ca(2+) channels. Although SOCE occurs following agonist activation of smooth muscle, this was thought to be more important in replenishing Ca(2+) stores rather than acting as a source of activator Ca(2+) for the contractile process. This review summarizes our current knowledge of SOCE as a regulator of vascular smooth muscle tone and discusses its possible role in the cardiovascular function and disease. We propose a possible hypothesis for its activation and suggest that SOCE may represent a novel target for pharmacological therapeutic intervention.     

舒马曲坦对大鼠肺动脉平滑肌细胞的促有丝分裂作用

目的　观察 5 HT1B/1D受体激动剂舒马曲坦对肺动脉平滑肌细胞 (PASMC)的促有丝分裂作用 ,探讨其作用机制和诱发肺血管构型重建的可能性。方法　分离培养大鼠PASMC ,用噻唑蓝 (MTT)法检测细胞增殖 ,用流式细胞术法分析细胞周期和DNA合成。结果　MTT法检测 5 羟色胺 (5 HT) 0 .0 1,0 .1,1.0 μmol·L- 1可促进PASMC增殖 ,增殖率分别增加2 0 1% ,2 2 8%和 2 5 6 %。舒马曲坦 0 .0 1,0 .1,1.0μmol·L- 1可促进PASMC增殖 ,增殖率分别增加199% ,2 2 0 %和 2 4 5 %。流式细胞术分析 5 HT 0 .1和1.0 μmol·L- 1组的细胞增殖指数分别为 2 2 .2 %和2 5 .9% ,S期细胞分数分别为 7.2 %和 9.8%。舒马曲坦 0 .1和 1.0 μmol·L- 1组的增殖指数分别为2 1.2 %和 2 3.9% ,S期细胞分数分别为 6 .6 %和8.8% ,均较对照组明显增高 (P <0 .0 5 )。 5 HT和舒马曲坦能够促进PASMC从G0 /G1期进入S期 ,5 HT的促有丝分裂作用可能有 5 HT1B/1D受体的参与。结论　舒马曲坦能促进PASMC的增殖。 5 HT1B/1D 受体在PASMC增殖和肺血管构型重建中有重要作用。舒马曲坦有致肺动脉高压的风险     

钾通道开放剂降低血管平滑肌细胞内游离钙浓度(英文)

目的:研究PCO—Pin,Nic,Lem及RP对VMSC内[Ca~(2 )]_i的改变及其可能机制。方法:VSMC加入Fura-2 AM 2.5μmol·L~(-1)37℃下孵育50min,[Ca~(2 )]_i用荧光分光光度计检测。结果:4种PCO能较弱地抑制K~ 30 mmol·L~(-1)诱导的[Ca~(2 )]_i增加,但明显抑制ATP 0.1mmol·L~(-1)诱导的[Ca~(2 )]_i峰相及持续相增加,且呈剂量依赖性。格列苯脲完全阻断Pin,Lem及RP的作用,只部分抑制Nic的作用。无钙液中先给4种PCO,能显著抑制ATP诱导的[Ca~(2 )]_i峰相增加。结论:4种PCO均抑制ATP诱导的[Ca~(2 )]_i增加,此作用与减少细胞外钙内流及细胞内钙释放有关。     

阿托品对大鼠肠系膜动脉的舒张作用及机制

目的研究阿托品的扩血管作用及机制。方法以大鼠肠系膜动脉为标本，考察阿托品对去甲肾上腺素(NE)预收缩血管的舒张作用以及血管内皮细胞、血管平滑肌在该效应中的作用。结果阿托品能显著舒张NE预收缩的完整内皮血管，去内皮后该作用明显降低。L-N^ω-硝基精氨酸甲酯、吲哚美辛、普萘洛尔及格列本脲对阿托品的舒张作用无明显影响。阿托品对KCl的量效曲线及咖啡因缩血管作用均无明显影响，但能浓度依赖性地抑制NE诱导的内钙释放以及经受体操纵性钙通道的外钙内流。结论阿托品有明显的扩血管作用，其通过抑制受体介导的外钙内流和内钙释放而舒张血管。     

Oxidative stress and COX cause hyper-responsiveness in vascular smooth muscle of the femoral artery from diabetic rats

BACKGROUND AND PURPOSE: To investigate the dysfunction of vascular smooth muscle in streptozotocin-induced diabetic rats. EXPERIMENTAL APPROACH: Rings without endothelium of femoral arteries were suspended in organ chambers for isometric tension recording. The production of oxygen-derived free radicals was measured with 2',7'-dichlorodihydrofluorescein diacetate using confocal microscopy. The protein expressions were measured by western blotting. KEY RESULTS: The concentration-response curves to U46619 and phenylephrine, but not that to KCl, were shifted to the left, suggesting a hypersensitivity of cell membrane receptors in diabetes. Exogenous oxygen-derived free radicals induced greater vasoconstrictions in the femoral artery from diabetic rats. Chronic treatment with apocynin (inhibitor of NADPH oxidase) and acute exposure to MnTMPyP (SOD/catalase mimetic) normalized the response. The catalase activity and the total glutathione level were reduced in arteries from streptozotocin-treated rats, confirming a redox abnormality. The basal oxidative state was higher in arteries from streptozotocin-treated rats and reduced in arteries from apocynin- and streptozotocin-treated rats, suggesting that the functional changes in diabetes are due to a chronic increase in oxidative stress. In the arteries of streptozotocin-treated rats, inhibitors of COX-1 and/or COX-2 prevented the hypersensitivity and reduced the increase in oxidative stress caused by phenylephrine and U46619, suggesting that both isoforms contribute to the smooth muscle dysfunction. The expression of proteins for COX-1 and COX-2 was increased in arteries of streptozotocin-treated rats and reduced in preparations of apocynin- and streptozotocin-treated rats. CONCLUSIONS AND IMPLICATIONS: Chronic diabetes and the resulting increased oxidative stress activate the production of COX-derived vasoconstrictor prostanoids causing hypersensitivity of vascular smooth muscle.     

左旋千金藤定碱对培养牛主动脉血管平滑肌细胞胞内游离Ca~(2+)的影响

目的：研究左旋千金藤定碱（ｌ－ｓｔｅｐｈｏｌｉｄｉｎｅ，ＳＰＤ）对血管平滑肌的作用。方法：采用Ｆｕｒａ－２和ＡＲ－ＣＭ－ＭＩＣ阳离子测定系统测定培养牛主动脉血管平滑肌细胞内游离钙。结果：ＳＰＤ１～１００μｍｏｌ·Ｌ－１不影响静息［Ｃａ２＋］ｉ，但可剂量依赖地抑制高Ｋ＋引起的［Ｃａ２＋］ｉ增高，其ＩＣ５０为３９．６（９５％可信限２３．４～６７．１）μｍｏｌ·Ｌ－１，但其作用弱于尼群地平；ＳＰＤ１～１００μｍｏｌ·Ｌ－１对去甲肾上腺素、血管紧张素Ⅱ、５－ＨＴ、ＡＴＰ引起的［Ｃａ２＋］ｉ增高也有明显的抑制作用；高浓度ＳＰＤ对无外钙时去甲肾上腺素引起的［Ｃａ２＋］ｉ增高也有一定的抑制作用。结论：左旋千金藤定碱对培养血管平滑肌细胞电压依赖性钙通道和受体调控性钙通道均有抑制作用；其对电压依赖性钙通道的抑制作用弱于尼群地平。     

细胞内游离钙在辛伐他汀诱导大鼠血管平滑肌细胞凋亡中的作用

目的　研究 3 羟 3 甲戊二酰辅酶A (HMG CoA)还原酶抑制剂辛伐他汀诱导血管平滑肌细胞(VSMC)凋亡的机制。方法　以荧光染料Fura 2 /AM负载后荧光分光光度计法检测细胞内游离钙浓度 ,以DNA琼脂糖凝胶电泳、流式细胞仪PI/膜联蛋白 (an nexin)V染色及半胱天冬酶 3激活来检测细胞凋亡。结果　辛伐他汀 30 μmol·L- 1孵育VSMC后 ,细胞内游离钙浓度显著升高 ,6h时达对照的 3倍以上 (P <0 .0 1) ,维拉帕米 80 μmol·L- 1与辛伐他汀 30 μmol·L- 1共同孵育VSMC 6h后细胞内游离钙浓度为 (14 4± 34)nmol·L- 1(P <0 .0 1)。辛伐他汀可诱导细胞凋亡率增高、“DNA梯状”样改变及半胱天冬酶 3的激活 ,这些变化均可被维拉帕米所逆转。结论辛伐他汀通过使细胞外钙大量内流而诱导VSMC凋亡。