Effects of ischemic preconditioning on vascular reactivity and calcium sensitivity after hemorrhagic shock and their relationship to the RhoA–Rho-kinase pathway in rats

We investigated the effect of ischemic preconditioning (IPC) on vascular reactivity and calcium sensitivity after hemorrhagic shock and its relationship to the RhoA–Rho-kinase pathway. Using hemorrhagic-shock rats (40 mm Hg for 3 hours) and isolated rings of the superior mesenteric artery (SMA), the effects of IPC (abdominal aorta occlusion applied 2 hours before shock) on the pressor effect of norepinephrine (3 μg/kg), vascular reactivity and calcium sensitivity of SMA, and the activity and role of RhoA and Rho-kinase were investigated. IPC with 1-minute occlusion plus 5-minute loosening of aneurysm clips thrice significantly increased survival time and prevalence of survival at 24 hours of hemorrhagic-shock rats. This IPC condition also significantly increased the pressor response of norepinephrine and significantly improved the vascular reactivity and calcium sensitivity of the SMA. The activity of Rho-kinase and RhoA in the SMA decreased after hemorrhagic shock, but increased after IPC. Y-27632 (Rho-kinase antagonist) and C3 Transferase (RhoA antagonist) significantly decreased IPC-induced increase in vascular reactivity and calcium sensitivity. These results suggested that IPC can improve shock-induced vascular hyporeactivity and calcium desensitization. The RhoA–Rho-kinase pathway played an important part in this process. These findings suggested that the RhoA–Rho-kinase pathway may be a potential target to treat vascular hyporeactivity in severe trauma, shock, or multiple-organ dysfunction syndrome.     

精氨酸加压素对失血性休克大鼠心肌收缩力的影响及与Rho激酶的关系

目的 探讨精氨酸加压素(AVP)对失血性休克大鼠心肌收缩力的影响及与Rho激酶的关系。方法 失血性休克大鼠的离体乳头肌分别用AVP 0.1 μmol·L^-1 和 Y-27632 10 μmol·L^-1预孵育,或Y-27632+AVP合用时,先用Y-27632 孵育10 min后,再加入AVP作用10 min。平衡30 min后依次用含异丙肾上腺素（Iso）1和10 μmol·L^-1, 0.1, 1, 10 mmol·L^-1的H-K液灌流乳头肌,观察加入Iso前后收缩力变化。制备离体心脏,稳定后,进行相应再灌流30 min,检测左心室收缩压（LVSP）和左心室压力上升或下降的最大速率（±dp/dt_max）。结果 失血性休克2 h,经Iso 0.1,1 及10 mmol·L^-1灌流后,离体乳头肌收缩力显著低于假手术组（P<0.05）,AVP 0.1 μmol·L^-1预孵育可明显升高乳头肌对心肌的收缩力;与休克模型组相比,在Iso 1和10 mmol·L^-1灌流后,乳头肌收缩力明显升高(P<0.05);Y-27632 10 μmol·L^-1离体乳头肌收缩力无明显影响,但Y-27632+AVP可明显降低由AVP引起的乳头肌收缩力的增加。失血性休克2 h后,离体心脏血流动力学指标明显降低,AVP可明显改善休克模型大鼠离体心脏的血流动力学指标,同时Y-27632可明显拮抗由AVP引起休克大鼠离体心脏血流动力学指标的增加。结论 AVP可改善失血性休克所致的心肌收缩力的降低,其机制与其激活Rho 激酶有关。     

精氨酸加压素对失血性休克大鼠血管反应性的影响

目的探讨精氨酸加压素(AVP)对失血性休克大鼠血管反应性的影响,并初步探讨其与Rho激酶的关系。方法在体实验,观察大鼠失血性休克后AVP对去甲肾上腺素(NE)升压反应的影响;离体实验,测定失血性休克大鼠肠系膜上动脉(SMA)环对NE的收缩反应和去极化状态下(120mmol·L-1K+)对Ca2+的收缩反应,反映其对缩血管物质和钙的反应性。结果失血性休克(4kPa,2h)后大鼠对NE的升压反应显著下降。AVP0.1和0.4U·kg-1,iv,随后再将AVP溶于3倍失血量的复方氯化钠溶液分别以0.01和0.04U·kg-1·min-1的速度于30min内用输液泵输注,3～4h后可使NE的升压反应恢复至正常组水平。失血性休克后SMA对NE和钙的反应性显著下降,对NE和Ca2+的收缩反应量效曲线明显右移,最大反应(Emax)降低;加入NE和Ca2+前分别用0.5和5nmol·L-1AVP孵育10min可使NE和Ca2+的收缩反应量效曲线明显左移,Emax显著增高。Rho激酶拮抗剂HA1077预处理可部分取消AVP所致的Ca2+Emax变化,使Emax回降。结论AVP能升高失血性休克大鼠血管对NE的敏感性及反应效能和血管平滑肌对钙的反应效能。     

Effects of MCI-154 on vascular reactivity and its mechanisms after hemorrhagic shock in rats

The objectives of this study were to investigate the effects of 6-[4-(4'-pyridylamino)phenyl]-4,5-dihydro-3(2H)-pyridazinone hydrochloride trihydrate (MCI-154), a newly developed cardiotonic agent, on vascular reactivity and contractile responses to extracellular Ca2+ ([Ca2+]o) after hemorrhagic shock and primarily explore its mechanism. In vivo, the effects of MCI-154 (0.1, 0.5, 1.0, and 2.0 mg/kg) on the pressor effect of norepinephrine (NE) in rats subjected to hemorrhagic shock (30 mm Hg for 2 h) were observed and in vitro, the effects of MCI-154 (10(-7), 10(-6), 10(-5), 10(-4) mol/L) on vascular reactivity and contractile responses to [Ca2+]o of superior mesenteric artery (SMA) from hemorrhagic shock rats and its relationship to Rho-kinase, protein kinase C (PKC), and protein kinase G (PKG) were observed. The results showed that the NE-induced pressor response after hemorrhagic shock was significantly decreased (P<0.01), and MCI-154 made it decrease further. In vitro, MCI-154 further decreased the contractile responses of SMA to NE and Ca2+ after hemorrhagic shock as compared with untreated hemorrhagic shock group (P<0.01). Angiotensin II (Ang II), with Rho-kinase stimulating action, and PMA, a PKC agonist increased the contractile responses to [Ca2+]o of SMA after hemorrhagic shock. MCI-154 (10(-5) mol/L) partly inhibited Ang II and PMA-induced increase of the contractile responses to [Ca2+]o of SMA (P<0.01). KT-5823, the PKG antagonist, antagonized MCI-154-induced decrease of the contractile responses to [Ca2+]o. Taken together, these results suggested that the vascular reactivity and contractile responses to [Ca2+]o of vascular smooth muscle after hemorrhagic shock were significantly decreased. MCI-154 worsened hemorrhagic shock-induced vascular hyporeactivity and the decrease of contractile responses to [Ca2+]o. These effects were possibly regulated by Rho-kinase, PKC, and PKG, but this needs further confirmation.     

Rac对失血性休克大鼠血管反应性的调节作用

目的探讨Rac在失血性休克大鼠血管反应性调节中的作用。方法采用SD大鼠复制休克模型,取离体血管环,观察休克早期和晚期血管反应性的变化以及Rac激动剂和特异性抑制剂对休克早期和晚期血管反应性的影响;通过酶消化法培养原代血管平滑肌细胞（vascular smoot hmuscle cell,VSMC）,采用双室培养方式分别观察VSMC缺氧10min和90min后VSMC对去甲肾上腺素（norepinephrine,NE）的收缩反应性变化,同时观察Rac活性调节剂对缺氧后VSMC收缩反应性的影响。结果在休克早期和短暂缺氧后,离体血管环和VSMC对NE收缩反应性均有所升高,Rac的激动剂血小板衍生生长因子（platelet derived growth factor,PDGF）可部分降低休克早期或短暂缺氧后血管反应性,Rac特异性抑制剂NSC23766可拮抗由PDGF所引起的血管反应性的变化,而在休克晚期或长时间缺氧后,离体血管环和VSMC对NE收缩反应性明显降低,NSC23766对休克晚期或长时间缺氧所致血管反应性降低有升高作用。结论休克后血管反应性呈双相变化,休克早期升高,休克晚期降低,Rac参与了休克血管反应性的调节。     

三羟异黄酮对失血性休克大鼠肠系膜上动脉收缩反应性的影响

目的研究三羟异黄酮(genistein,GST)对失血性休克大鼠肠系膜上动脉收缩反应性的影响及其可能机制。方法建立大鼠失血性休克(3.9kPa,2h)模型。采用离体血管环张力测定实验,观察三羟异黄酮及酪氨酸蛋白磷酸酶抑制剂钒酸钠(sodiumorthovannadate,Na3VO4)对休克大鼠肠系膜上动脉血管收缩反应性的影响;采用细胞贴附式膜片钳记录技术,观察三羟异黄酮及钒酸钠对休克大鼠肠系膜上动脉平滑肌细胞大电导钙激活钾通道(largeconductancecalciumactivatedpotassiumchannel,BKCa)活动的影响。结果失血性休克导致大鼠肠系膜上动脉对去甲肾上腺素(noradrenine,NE)的收缩反应性降低,三羟异黄酮可在一定的剂量范围内明显改善失血性休克引起的血管低反应性;钒酸钠则可引起休克血管收缩反应性的进一步降低,且该作用可被0.1mmol·L-1TEA部分阻断;进一步的研究显示,失血性休克可引起大鼠肠系膜上动脉血管平滑肌细胞BKCa通道活动的增强,三羟异黄酮可抑制休克血管平滑肌细胞BKCa通道活动,且该作用可被钒酸钠逆转。结论三羟异黄酮可通过干预由PTK介导的酪氨酸蛋白磷酸化,防止失血性休克血管平滑肌细胞BKCa通道活动的增强,从而有效恢复失血性休克大鼠肠系膜上动脉对NE的收缩反应性。     

Pinacidil pretreatment improves vascular reactivity after shock through PKCα and PKCε in rats

We investigated the beneficial effect of pinacidil pretreatment on vascular reactivity, calcium sensitivity, and animal survival after hemorrhagic shock, its relationship to protein kinase Cα (PKCα), protein kinase Cε (PKCε), and adenosine. Using hemorrhagic shock rats, the protective effects of different extents of pinacidil pretreatment on vascular reactivity and in which the roles of PKCα, PKCε, and adenosine were observed. Pinacidil pretreatment significantly improved shock-induced decrease of vascular reactivity of superior mesenteric artery, which was antagonized by the PKCα antagonist G?-6976 (5 × 10 mole/L) and PKCε pseudosubstrate inhibitory peptide (1 × 10 mole/L). Pinacidil pretreatment induced the translocation of PKCα and PKCε from the cytoplasm to the membrane. This translocation of PKCα and PKCε was eliminated by adenosine A1 receptor antagonist DPCPX (1 × 10 mole/L). As compared with simple fluid resuscitation, combination with pinacidil pretreatment significantly improved the vascular reactivity and survival rate of hemorrhagic-shocked rats. These results suggested that pinacidil pretreatment could induce good protective effects on vascular reactivity and calcium sensitivity after hemorrhagic shock mainly through the activation of PKCα and PKCε via adenosine A1 receptor, and this protective effect made an important contribution to the overall outcome of shock therapy.     

Regulatory effect of Rac1 on vascular reactivity after hemorrhagic shock in rats

We used isolated superior mesenteric arteries (SMAs) from hemorrhagic-shock rats and hypoxia-treated vascular smooth muscle cells (VSMCs; mimicking the shock state) to observe the effects of platelet-derived growth factor (PDGF; Rac1 stimulator) and NSC23766 (Rac1 antagonist) on vascular reactivity and the relationship with the Rho kinase-myosin light-chain phosphatase (MLCP) and p21-activated kinase (PAK)-myosin light-chain kinase (MLCK) signal pathway. The results indicated that the contractile responses of the SMAs and VSMCs were significantly increased at early shock or after transient hypoxia. NSC23766 (Rac1 antagonist) further increased, whereas PDGF (Rac1 stimulator) decreased the contractile responses of SMAs and VSMCs. In the late period of shock or prolonged hypoxia, the contractile responses of SMAs and VSMCs were significantly decreased; NSC23766 increased (whereas PDGF further decreased) the contractile response of the SMAs and VSMCs. Activation of Rac1 with PDGF significantly increased the activity of PAK and MLCP, and decreased Rho kinase and MLCK activity and 20-kDa myosin light-chain phosphorylation in VSMCs. The PAK inhibitor PAK-18 significantly antagonized the PDGF-induced decrease in MLCK activity, whereas the Rho kinase antagonist Y-27632 further enforced the PDGF-induced increase in MLCP activity. Simple fluid resuscitation did not improve but in combination with NSC23766 significantly improved vascular reactivity and animal survival at 24 hours. This suggested that Rac1 has an inhibitory effect on vasoreactivity after shock. Rac1-mediated regulation of vascular reactivity is mainly through activation of PAK, inhibition of MLCK and inhibition of Rho kinase, unpack the inhibition of Rho kinase to MLCP. Rac1 may be a potential target to treat vascular hyporeactivity in many critical conditions.     

Effects of Y-27632, a Rho/Rho kinase inhibitor, on leukotriene D(4)- and histamine-induced airflow obstruction and airway microvascular leakage in guinea pigs in vivo

Recent in vitro studies have shown that the Rho/Rho kinase pathway is involved in the mechanism of not only airway smooth muscle contraction but also vascular endothelial permeability caused by certain stimuli. This suggests that Rho/Rho kinase inhibitors may become useful agents against asthma via reduction of increased airway microvascular leakage, one of the main features of this disease. Thus, we wanted to know the in vivo effect of Y-27632, a selective Rho kinase inhibitor, on airway microvascular leakage caused by leukotriene D(4) (LTD(4)) and histamine, potent mediators of allergic airway inflammation, by comparing its effect against airflow obstruction. For comparison, the effects of procaterol, a beta(2)-adrenoceptor agonist, on these responses were also studied. Tracheostomized guinea pigs were given either aerosolized Y-27632 (3 or 15 mmol/l), procaterol (6 micromol/l) or vehicle (0.9% NaCl) for 5 min under spontaneous breathing. After being mechanically ventilated, the animals were given intravenous Evans blue dye 15 min after the end of inhalation. One minute later, either 2 nmol/kg LTD(4), 300 nmol/kg histamine or vehicle was administered intravenously. After measurements of lung resistance (R(L)) for 6 min, the lungs of animals were taken out, and the amount of extravasated Evans blue dye was examined as an index of leakage. Inhaled Y-27632 dose-dependently attenuated increases in R(L) caused by LTD(4) and histamine. The degree of inhibition was almost similar between 15 mmol/l Y-27632 and 6 micromol/l procaterol. By contrast, only 15 mmol/l, but not 3 mmol/l, Y-27632 partially reduced LTD(4)-induced leakage. Histamine-induced Evans blue dye extravasation was not inhibited by 15 mmol/l Y-27632. Procaterol significantly inhibited the dye extravasation caused by either LTD(4) or histamine. These results suggest that Y-27632 is not a useful agent in attenuating airway microvascular leakage which is seen in asthma, although it is potent in inhibiting airflow obstruction.     

Rho kinase contributes to androgen amplification of renal vasoconstrictor responses in the spontaneously hypertensive rat

Androgens modulate vascular tone and hypertension development. Rho kinase contributes to norepinephrine- (NE) and vasopressin- (AVP) induced vasoconstriction. This study tested the hypothesis that Rho kinase contributes to androgen amplification of renal vasoconstrictor responses to NE or AVP in isolated perfused kidney of spontaneously hypertensive rats (SHRs).SHRs (5 weeks) underwent sham operation, castration, or castration with testosterone replacement. At 16-17 weeks, mean arterial pressure and heart rate were measured in conscious SHRs. Renal vascular reactivity to NE (10 to 10 mol) and to AVP (10 to 10 mol) was assessed in an isolated perfused kidney preparation before and after Rho kinase inhibitor treatment (fasudil; 15 microM). Castration reduced mean arterial pressure, whereas testosterone treatment of castrated SHRs increased mean arterial pressure significantly. The dose-response curves to NE and AVP obtained in isolated perfused kidneys from castrated SHRs were displaced to the right of those obtained in sham-operated and castrated + testosterone-treated SHRs. Fasudil treatment produced a rightward shift in the dose-response curves for each agonist in all of the groups and greatly attenuated the differences in renal vascular reactivity to NE and AVP among the 3 groups of SHRs.Collectively, these findings indicate that androgen modulation of hypertension development in the SHR involves a fasudil-sensitive pathway and suggest that further study is warranted in this area.     

肌内皮缝隙连接在失血性休克大鼠内皮依赖和非内皮依赖的血管舒/缩功能调节中的作用

目的 研究肌内皮缝隙连接(myo-endothelial gap junction,MEGJ)通道在失血性休克大鼠肠系膜上动脉血管(SMA)的内皮依赖和非内皮依赖的血管收缩/舒张功能调节中的作用.方法 利用在体血管管径测定技术,观察MEGJ的阻断剂18α-甘草次酸(18α-GA)对非内皮依赖的血管收缩剂去甲肾上腺素(N...     

Differential inhibition by the Rho kinase inhibitor Y-27632 of the increases in contractility and Ca<Superscript>2+</Superscript> transients induced by endothelin-1 in rabbit ventricular myocytes

The role of Rho kinase activation in the regulation of cardiac contractility and Ca²⁺ signaling remains unclear, whereas its role in smooth muscle regulation has been well documented. To study the potential role of Rho kinase in the regulation of cardiac contractility and Ca²⁺ transients induced by endothelin-1 (ET-1) and isoproterenol, we used the Rho kinase inhibitor Y-27632 in rabbit ventricular myocardium and myocytes loaded with indo-1/AM. Y-27632 (3–30 M) inhibited significantly the baseline contractility and Ca²⁺ transients. Furthermore, Y-27632 suppressed the increase in contractility and Ca²⁺ transients induced by ET-1 in a concentration-dependent manner, when it was used in a concentration at which it did not affect the effects of isoproterenol via -adrenoceptors. In the presence of Y-27632, ET-1 increased cell shortening in the absence of an increase in Ca²⁺ transients. This is an indication that the increase in myofilament Ca²⁺ sensitivity induced by ET-1 is less susceptible to the inhibitory action of Y-27632. These findings imply that the Rho kinase activation may partially contribute to the ET-1-induced regulation of contractility, primarily due to an ET-1-induced increase in Ca²⁺ transients in rabbit ventricular myocardium.     

Protein kinases participate in the contraction in response to levobupivacaine in the rat aorta

Levobupivacaine is a long-acting amide local anesthetic that intrinsically produces vasoconstriction both in vivo and in vitro. Levobupivacaine increases intracellular calcium concentrations ([Ca(2+)](i)) in vascular smooth muscle cells. The goals of this in vitro study were to investigate whether levobupivacaine-induced contraction is associated with increased Ca(2+) sensitivity and to identify the protein kinases involved in mediating contraction in response to levobupivacaine in isolated rat aortic smooth muscle. The effect of levobupivacaine and potassium chloride (KCl) on the [Ca(2+)](i) and tension was measured simultaneously with acetoxymethyl ester of fura-2-loaded aortic strips. Cumulative levobupivacaine concentration-response curves were generated in the presence or absence of the following antagonists: GF 109203X; Y-27632; genistein; SP600125; PD 98059; and SB 203580. Levobupivacaine-induced protein kinase C (PKC), extracellular signal-regulated kinase (ERK), and c-Jun NH(2)-terminal kinase (JNK) phosphorylation and Rho-kinase (ROCK-2) membrane translocation were detected in rat aortic vascular smooth muscle cells using Western blotting. The slope of the [Ca(2+)](i)-tension curve for levobupivacaine was higher than that for KCl. Y-27632, GF 109203X, and SP600125 attenuated levobupivacaine-induced contraction in a concentration-dependent manner. Genistein, PD 98059, and SB 203580 attenuated levobupivacaine-induced contraction. Pretreatment with GF 109203X and Y-27632 inhibited levobupivacaine-induced PKC phosphorylation and Rho-kinase (ROCK-2) membrane translocation, respectively. Pretreatment with SP600125 or PD 98059 attenuated the levobupivacaine-induced phosphorylation of JNK and ERK, respectively. These results indicate that levobupivacaine-induced contraction involving an increase in myofilament Ca(2+) sensitivity involves the primary activation of Rho-kinase-, PKC-, and JNK-mediated pathways of rat aortic smooth muscle.     

缝隙连接及其连接蛋白对血管加压素诱导的失血性休克大鼠血管收缩的作用研究

摘要：目的 观察缝隙连接（GJ）及其组成亚单位连接蛋白（Cx）在血管加压素（AVP）诱导失血性休克大鼠血管收缩中的作用。方法 采用失血性休克大鼠模型和缺氧培养血管平滑肌细胞（VSMC）,观察GJ阻断剂CBX和octanol、以及各Cx亚型反义寡核苷酸（AODN）对AVP诱导的血管收缩反应的影响,随后进一步观察参与AVP作用的Cx37和Cx43对AVP调节休克血管钙敏感性和缺氧VSMC内钙离子浓度的影响。结果 GJ阻断剂CBX和octanol明显抑制了AVP诱导的休克血管的收缩反应。在所有血管中表达的连接蛋白中,Cx37AODN和Cx43AODN明显抑制了AVP的血管收缩作用。进一步结果显示,Cx43AODN、而不是Cx37AODN,可拮抗AVP升高休克血管钙敏感性的作用。此外, AVP处理和干扰Cx37及Cx43对缺氧VSMC内钙离子浓度无明显影响。结论 缝隙连接在休克后AVP介导的血管收缩调节中有重要作用,Cx37和Cx43参与了这一过程,其中Cx43可能通过影响AVP介导的血管钙敏感性调节途径来发挥作用,而Cx37可能通过其它机制来参与AVP的血管调节作用。     

Enhanced relaxation to the rho-kinase inhibitor Y-27632 in mesenteric arteries from mineralocorticoid hypertensive rats

Increased vasoconstriction is characteristic of hypertension. In this study, we tested the hypothesis that changes in vascular responses during mineralocorticoid hypertension may be due to increased activation of the Rho/Rho-kinase pathway. To test this, relaxation responses to the Rho-kinase inhibitor Y-27632 were determined by measuring isometric force in deendothelialized mesenteric arteries from mineralocorticoid-hypertensive rats and sham-operated controls. Following agonist-induced contraction by serotonin (5-HT, 5-hydroxytryptamine), arteries from hypertensive rats demonstrated a greater relaxation to the Rho-kinase inhibitor Y-27632 (65 +/- 5% vs. 28 +/- 10%). Treatment with an EC50 concentration of Y-27632 following a KCl-induced contraction caused minimal relaxation of arteries in both groups of animals. These findings suggest that augmented Rho-kinase activity in the vasculature of mineralocorticoid hypertensive rats may contribute to the enhanced vascular reactivity of agonist-mediated stimuli characteristic of this model of hypertension.     

Tie-2介导Ang-1、Ang-2调节大鼠失血性休克血管反应性双相变化的作用研究

目的 观察Tie-2在血管生成素-1(angiopoietin-1, Ang-1)、血管生成素-2(angiopoietin-2, Ang-2)调节失血性休克血管反应性双相变化中的作用。方法 采用离体微血管环张力测定技术和western blot技术,观察失血性休克后不同时间点肠系膜上动脉(superior mesenteric artery,SMA)中Tie-2蛋白表达和磷酸化变化、抑制Tie-2对Ang-1和Ang-2调节缺氧早期和晚期血管反应性作用的影响,以及给予Ang-1和Ang-2后缺氧的血管内皮细胞(vascular endothelial cell, VEC)和血管平滑肌细胞(vascular smooth muscle cell, VSMC)混合细胞中Tie-2蛋白表达和磷酸化变化,并观察抑制Tie-2对缺氧早期和晚期的混合细胞NO含量的影响。结果 (1) 肠系膜上动脉Tie-2蛋白表达和酪氨酸磷酸化在正常时很低,失血性休克后逐渐增高,在休克早期(休克10min),Tie-2蛋白表达变化不大,但酪氨酸磷酸化已明显增高(P<0.01);随着休克时间延长,Tie-2蛋白表达和酪氨酸磷酸化均进一步显著增高(P<0.01)。(2)Tie-2抑制剂可降低缺氧10min的血管高反应性(NE的Emax由13.479mN降低至10.122mN,P<0.05),并显著抑制Ang-1对缺氧10min血管反应性的维持作用(Emax由15.283mN降低至10.253mN,P<0.01);Tie-2抑制剂可改善缺氧4h的血管低反应性(NE的Emax由5.875mN增高至8.003mN,P<0.05),并显著拮抗Ang-2进一步降低缺氧4h血管反应性的作用(Emax由3.444mN增高至7.643mN,P<0.01)。(3)缺氧10min时,降低血管高反应性的Ang-2可降低Tie-2磷酸化,使其由0.0403降低至0.0123(P<0.01);缺氧4h时,恢复血管低反应性的Ang-1可降低Tie-2蛋白表达,使其由0.2276降低至0.0851 (P<0.01),也可以降低Tie-2磷酸化,使其由0.1437降低至0.0359 (P<0.01)。(4) NO含量在缺氧早期显著增高,Ang-2和Tie-2抑制剂显著抑制其增高(P<0.01);缺氧晚期NO含量较正常对照组增高得更为显著,Ang-1和Tie-2抑制剂可抑制其增高(P<0.01)。结论 Ang-1、Ang-2通过Tie-2受体调节大鼠失血性休克血管反应性双相变化。     

Partial protective effect of Y-27632, a Rho kinase inhibitor, against hepatic ischemia-reperfusion injury in rats

(+)-(R)-trans-4-(1-aminoethyl)-N-(4-pyridyl)-cyclohexanecarboxamide dihydrochloride (Y-27632), a Rho kinase inhibitor, has a suppressive effect on the functions of polymorphonuclear leukocytes. In this study, the influence of Y-27632 on ischemia-reperfusion injury of the liver was examined in rats. Y-27632 (3 mg/kg) or vehicle alone was intravenously injected into rats 60 min before occlusion. Blood samples were obtained for 48 h after reperfusion. At the end of the experiment, the hepatic content of myeloperoxidase, which reflects the number of polymorphonuclear leukocytes in liver tissues, was determined. The increases in serum hepatic aminotransferases and inflammatory cytokines [tumor necrosis factor-alpha (TNF-alpha), interleukin (IL)-6] after reperfusion were partially, but significantly, inhibited by Y-27632. The increased hepatic myeloperoxidase content was significantly lowered by Y-27632. These results suggest that Y-27632 has a partial protective effect against hepatic ischemia-reperfusion injury through the suppression of polymorphonuclear leukocytes and inflammatory cytokines.     

Chronic fluvastatin treatment alters vascular contraction by inhibiting the Rho/Rho-kinase pathway

1. In the present study, we investigated the effects of chronic treatment of stroke-prone spontaneously hypertensive rats (SHRSP) with the statin fluvastatin on vascular Rho/Rho-kinase pathway mediated contraction, which has been shown to be upregulated in hypertension. 2. Contribution of the Rho/Rho-kinase pathway to noradrenaline-induced contraction of arteries from SHRSP was assessed by the inhibitory effect of Y-27632, a Rho/Rho-kinase inhibitor. Stroke-prone spontaneously hypertensive rats were treated with fluvastatin (10 mg/kg per day) for 1 month. 3. Treatment with fluvastatin tended to attenuate the contraction to noradrenaline and significantly decreased the Y-27632-sensitive component of the contraction in controls compared with fluvastatin-treated rats. 4. RhoA, as assessed by western blotting, was also reduced by fluvastatin treatment. 5. These findings suggest that chronic treatment with fluvastatin reduces the contractile response associated with Rho/Rho-kinase in arteries of hypertensive rats.     

鱼腥草注射液过敏及类过敏实验研究

目的评价鱼腥草注射液及其辅料吐温80（Tween-80）静脉给药的致敏性。方法采用Beagle犬类过敏及过敏实验,观察给药后动物行为变化和检测血浆中组胺、IgE、IgM、IgG的含量。结果含有Tween-80的鱼腥草注射液及Tween-80组动物给药后出现显著的行为异常,血浆组胺升高,而IgE变化不规律,IgG、IgM则无明显改变。结论鱼腥草注射液中导致犬严重类过敏反应与Tween-80有关。建议Beagle犬的过敏及类过敏试验,应作为中药注射液致敏试验的必做实验,行为异常及血浆中组胺为主要的判定指标,IgE作为辅助判定指标。     

Antiepileptic effects of two Rho-kinase inhibitors, Y-27632 and fasudil, in mice

BACKGROUND AND PURPOSE: Rho/Rho-kinase signalling is involved in many cellular events, including some in the CNS. However, the role of this pathway in epilepsy has not yet been assessed. Therefore, we determined the effects of two Rho-kinase inhibitors, Y-27632 and fasudil, on seizures induced by pentylenetetrazole (PTZ) or maximal electroconvulsive shock (MES). EXPERIMENTAL APPROACH: Effects of Y-27632 (5-10 mg kg(-1)) and fasudil (5-25 mg kg(-1)) on duration of myoclonic jerks, clonic and tonic convulsions, tonic hindlimb extensions and percentage of tonic convulsion index, as well as recovery latency for righting reflex were investigated in mice stimulated with PTZ (65 mg kg(-1)) or MES (50 Hz, 50 mA and 0.4 s). These inhibitors were also tested on a model of kindling induced by PTZ (35 mg kg(-1), for 11 days). Membrane and cytosolic levels of RhoA protein were measured in brain homogenates from kindled mice. KEY RESULTS: Y-27632 and fasudil diminished onset of myoclonic jerks, clonic convulsions and tonic hindlimb extensions in mice given PTZ. These inhibitors suppressed the percentage of tonic convulsion index and recovery latency for righting reflex in the mice excited with MES. Western blotting demonstrated that Rho translocation to plasma membrane increased in the brain homogenates obtained from PTZ-kindled mice. However, the Rho-kinase inhibitors at the given doses did not change motor coordination of the mice. CONCLUSIONS AND IMPLICATIONS: Rho/Rho-kinase signalling may play a role in epilepsy induced by PTZ and MES. Furthermore, Rho-kinase inhibitors could be novel important antiepileptic agents.