高原肺水肿患者血流动力学变化及对吸氧反应的测定

目的:探讨高原肺水肿的发病机理。方法:采用右心漂浮导管检测法, 对9例高原肺水肿患者及9例同海拔高原健康人的血流动力学指标进行了检测, 同时也观察了吸入纯氧对高原肺水肿患者血流动力学的影响。结果:高原肺水肿患者发病时, 肺动脉平均压、肺血管阻力、心脏指数均明显高于同海拔高度健康人, 而患者肺动脉楔压, 右心房压力同对照组相比, 未见显著差异;吸氧后, 高原肺水肿患者心率、肺动脉平均压力, 肺血管阻力及心脏指数均较吸氧前明显下降, 特别是肺动脉平均压及肺血管阻力下降尤为明显, 肺动脉平均压力在吸氧1min后即明显下降, 吸氧5min后, 下降至最低值, 但吸氧20min后仍未达对照组水平。结论:高原肺水肿是非心源性肺水肿, 肺动脉高压在其发病中起重要作用。     

Role of O(2)-sensitive K(+) and Ca(2+) channels in the regulation of the pulmonary circulation: potential role of caveolae and implications for high altitude pulmonary edema

High altitude pulmonary edema (HAPE) is a potentially fatal complication in response to exposure to low O(2) at high altitudes. Hypoxia, by causing pulmonary vasoconstriction, increases pulmonary vascular resistance and pulmonary arterial pressure, both of which are features in the pathogenesis of HAPE. Uneven hypoxic pulmonary vasoconstriction is thought to be responsible for increased capillary pressure and leakage, resulting in edema. O(2)-sensitive ion channels are known to play pivotal roles in determining vascular tone in response to hypoxia. K(+), Ca(2+) and Na(+) channels are ubiquitously expressed in both endothelial and smooth muscle cells of the pulmonary microvasculature, subfamilies of which are regulated by local changes in P(O(2)). Hypoxia reduces activity of voltage-gated K(+) channels and down-regulates their expression leading to membrane depolarization, Ca(2+) influx in pulmonary artery smooth muscle cells (by activating voltage-dependent Ca(2+) channels) and vasoconstriction. Hypoxia up-regulates transient receptor potential channels (TRPC) leading to enhanced Ca(2+) entry through receptor- and store-operated Ca(2+) channels. Altered enrichment of ion channels in membrane microdomains, in particular in caveolae, may play a role in excitation-contraction coupling and perhaps in O(2)-sensing in the pulmonary circulation and thereby may contribute to the development of HAPE. We review the role of ion channels, in particular those outlined above, in response to low O(2) on vascular tone and pulmonary edema. Advances in the understanding of ion channels involved in the physiological response to hypoxia should lead to a greater understanding of the pathogenesis of HAPE and perhaps in the identification of new therapies.     

High altitude pulmonary edema: a pressure-induced leak

High altitude pulmonary edema (HAPE) is a non-cardiogenic pulmonary edema that can occur in healthy individuals who ascend rapidly to altitudes above 3000-4000m. Excessive pulmonary artery pressure (PAP) is crucial for the development of HAPE, since lowering pulmonary artery pressure by nifedipine or tadalafil (phosphodiesterase-5-inhibitor) will in most cases prevent HAPE. Recent studies using microspheres in swine and magnetic resonance imaging in humans strongly support the concept and primacy of nonuniform hypoxic arteriolar vasoconstriction to explain how hypoxic pulmonary vasoconstriction occurring predominantly at the arteriolar level can cause leakage. Evidence is accumulating that the excessive PAP response in HAPE-susceptible individuals is due to a reduced NO bioavailability. HAPE-susceptible individuals show an endothelial dysfunction in the systemic circulation in hypoxia. Lower levels of exhaled NO in hypoxia before and during HAPE suggest that this abnormality also occurs in the lungs and polymorphisms of the eNOS gene are associated with susceptibility to HAPE in the Indian and Japanese population.     

急性高原病患者支气管肺泡灌洗前后肺功能及血气的改变

目的:探讨急性高原反应(HAAR)及高原肺水肿(HAPE)的发病机理。方法:对10例HAAR患者及6例HAPE患者灌洗前和灌洗后进行肺功能和动脉血气检测, 并与10例高原健康者进行对比。结果:HAAR患者及HAPE患者灌洗前动脉血氧分压明显低于对照组, HAAR同HAPE均存在弥散功能障碍;HAPE肺弥散功能(DLCO%)由灌洗前的(76.01±6.29)%, 上升到灌洗后的(103.31±9.23)%;气体转化因子(DLCO/VA%)由灌洗前的(150.30±15.20)%, 上升到灌洗后的(176.04±16.10)%;动脉血氧分压(PaO₂)由(31.73±3.01)mmHg上升到(45.31±3.56)mmHg。而HAAR及对照组灌洗后上述指标差异不显著。结论:HAPE患者肺泡内大量的液体渗出是HAPE病情恶化的主要原因之一。HAAR属HAPE发展的初级阶段, 存在着间质性肺水肿。     

［Ca2+］i对大鼠肺动脉平滑肌细胞膜钙激活氯离子通道的调节作用

目的： 探讨细胞浆内游离钙离子浓度（［Ca²⁺］_i）在常氧、急性和慢性低氧条件下对大鼠肺动脉平滑肌细胞(PASMCs)膜钙激活氯离子通道（Cl_Ca）的调节作用。 方法: 常规离体血管灌流法检测急性低氧时肺动脉环张力变化；钙荧光探针(Fura-2/AM)负载培养PASMCs，观察常氧和慢性低氧条件下［Ca²⁺］_i的变化并由此对Cl_Ca的影响；同时用四唑盐(MTT)比色法观察当［Ca²⁺］_i变化时Cl_Ca对PASMCs增殖的影响。 结果: (1) Cl_Ca阻断剂尼氟灭酸（NFA）和indaryloxyacetic acid (IAA-94)可以舒张急性低氧引起的肺动脉环收缩。(2) 慢性低氧时［Ca²⁺］_i升高：常氧状态下, PASMCs ［Ca²⁺］_i为(123.63±18.98)nmol/L，低氧时为(281.75±16.48)nmol/L (P<0.01)。(3) 常氧时，NFA和IAA-94对［Ca²⁺］_i 无明显影响(P>0.05)。(4) 慢性低氧时, NFA和IAA-94使PASMCs ［Ca²⁺］_i由(281.75±16.48)nmol/L降低到(117.66±15.36)nmol/L (P<0.01)。(5)MTT比色法中，慢性低氧状态下NFA和IAA-94引起升高的吸光度(A)值降低，由0.459±0.058到0.224±0.025 (P<0.01)。 结论: 低氧引起［Ca²⁺］_i升高，这可能激活Cl_Ca，对［Ca²⁺］_i起正反馈作用，Cl_Ca可能在低氧肺动脉高压中起作用；慢性低氧条件下Cl_Ca可能参与促进大鼠PASMCs的增殖。     

NADH and NAD modulates Ca2+-activated K+ channels in small pulmonary arterial smooth muscle cells of the rabbit

We have investigated the effect of NADH and NAD on the gating of large conductance Ca²⁺-activated K(K_Ca) channels in arterial smooth muscle cells isolated from small pulmonary artery(outer diameter <300μm) and ear artery, using the patch clamp technique. In the inside-out configuration, intracellularly applied 2 mM NADH inhibited the activity of K_Ca, channels in pulmonary arterial smooth muscle cells, while it had no significant effect on ear arterial smooth muscle cells. On the other hand, 2 mM NAD increased the opening of K_Ca, channels in pulmonary arterial smooth muscle cells. The effects of another intracellular redox couple, glutathione(GSH) and glutathione disulfide(GSSG) were also dependent on their redox potentials. GSH(5 mM) inhibited K_Ca. channels activity, while GSSG(5 mM) increased the activity of pulmonary arterial smooth muscle cells. It could be concluded that the modulation of K_Ca channels by intracellular redox state contributes, at least in part, to the hypoxic suppression of outward current in pulmonary arterial smooth muscle cells.     

钾通道对大鼠肺动脉平滑肌细胞[Ca2+]i的调节

目的:探讨在常氧、低氧条件下钾通道对大鼠肺动脉平滑肌细胞(PASMCs)[Ca²⁺]_i的调节。方法:采用钙荧光探针(Fura-2/AM)负载培养的大鼠PASMCs,观察常氧、低氧培养后3种钾通道抑制剂(4AP,TEA、Glib)对PASMCs[Ca²⁺]_i的调节,同时用四唑盐(MTT)比色法比较4AP、TEA、Glib对大鼠PASMCs增殖的影响。结果:(1)常氧状态下,PASMCs[Ca²⁺]_i为(156.91±8.60)nmol/L,低氧时为(294.01±16.81)nmol/L(P＜0.01)。(2)常氧状态下,4AP可引起PASMCs[Ca²⁺]_i升高,达(280.52±23.21)nmol/L(P＜0.01),而TEA、Glib无此作用。(3)低氧时,4AP和TEA都可引起PASMCs[Ca²⁺]_i的升高,分别为(422.41±24.28)nmol/L、(380.84±11.02)nmol/L(P<0.01),Glib无作用。(4)MTT比色法中,常氧和低氧状态下4AP均引起吸光度(A)值升高,分别是0.582±0.062,0.873±0.043(P＜0.01)。TEA仅在低氧时A值升高(0.729±0.041,P＜0.05),而Glib无论常氧还是低氧均无影响。结论:无论常氧还是低氧条件下,电压依赖性钾通道(K_V)对PASMCs[Ca²⁺]_i及其增殖起主要作用。钙激活的钾通道(K_Ca)在常氧条件下对[Ca²⁺]_i不起调节作用,而在低氧下使[Ca²⁺]_i降低,反应性地调节PASMCs增殖。ATP敏感性钾通道(K_ATP)无论在常氧还是低氧情况下对[Ca²⁺]_i的调节不起作用。     

K+通道在正常与慢性低氧大鼠低氧性肺血管收缩反应中的作用

目的:探讨K⁺通道在慢性低氧致低氧性肺血管收缩反应降低中的作用。方法:采用离体肺灌流实验,研究4-AP(4-aminopyridine,电压依赖性K⁺通道-Kv阻滞剂)、TEA(tetraethylamonium,Ca²⁺激活性K⁺通道-K_Ca阻滞剂)、GLIB(glibenclamide,ATP敏感性K⁺通道-K_ATP阻滞剂)对正常与慢性低氧大鼠肺血管低氧反应的影响。结果:4-AP、TEA均可使正常大鼠肺动脉基础压上升,且使其肺血管低氧反应明显增强;对于慢性低氧大鼠,其肺血管对低氧反应明显低下,4-AP、TEA升肺动脉基础压的作用明显低于对照鼠肺,GLIB也呈现升高肺动脉基础压力作用,4-AP、TEA、GLIB均可使肺血管低氧反应大大增强,增强的比例明显大于正常对照组。结论:在离体灌流鼠肺HPV中,Kv、K_Ca的开放起调节作用,大鼠经慢性低氧后,肺血管反应性明显降低,可能与Kv、K_Ca、K_ATP在HPV中的调节作用相对增强有关。     

K channels in O2 sensing and postnatal development of carotid body glomus cell response to hypoxia

The sensitivity of carotid body chemoreceptors to hypoxia is low just after birth and increases over the first few weeks of the postnatal period. At present, it is believed that the hypoxia-induced excitation of carotid body glomus cells begins with the inhibition of the outward K⁺ current via one or more O₂ sensors. Although the nature of the O₂ sensors and their signals that inhibit the K⁺ current are not well defined, studies suggest that the postnatal maturation of the glomus cell response to hypoxia is largely due to the increased sensitivity of K⁺ channels to hypoxia. As K_V, BK and TASK channels that are O₂-sensitive contribute to the K⁺ current, it is important to identify the O₂ sensor and the signaling molecule for each of these K⁺ channels. Various O₂ sensors (mitochondrial hemeprotein, hemeoxygenase-2, NADPH oxidase) and associated signals have been proposed to mediate the inhibition of K⁺ channels by hypoxia. Studies suggest that a mitochondrial hemeprotein is likely to serve as an O₂ sensor for K⁺ channels, particularly for TASK, and that multiple signals may be involved. Thus, changes in the sensitivity of the mitochondrial O₂ sensor to hypoxia, the sensitivity of K⁺ channels to signals generated by mitochondria, and/or the expression levels of K⁺ channels are likely to account for the postnatal maturation of O₂ sensing by glomus cells.     

Effect of phospholipase A2 on development of pulmonary edema and on the pulmonary hemodynamics in intact and vagotomized animals

Laboratory of Pathophysiology of Respiration and Bioengineering Laboratory, Research Institute of General Pathology and Pathological Physiology, Academy of Medical Sciences of the USSR, Moscow. Department of Pathophysiology, Yaroslavl' Medical Institute. (Presented by Academician of the Academy of Medical Sciences of the USSR B. I. Tkachenko.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 110, No. 9, pp. 254–257 September, 1990.     

Biophysical properties of Ca2+- and Mg-ATP-activated K+ channels in pulmonary arterial smooth muscle cells isolated from the rat

A novel class of Ca²⁺-activated K⁺ channel, also activated by Mg-ATP, exists in the main pulmonary artery of the rat. In view of the sensitivity of these K_Ca,ATP channels to such charged intermediates it is possible that they may be involved in regulating cellular responses to hypoxia. However, their electrophysiological profile is at present unknown. We have therefore characterised the sensitivity of K_Ca,ATP channels to voltage, intracellular Ca²⁺ ([Ca²⁺]_i) and Mg-ATP. They have a conductance of 245 pS in symmetrical K⁺ and are approximately 20 times more selective for K⁺ ions than Na⁺ ions, with a K⁺ permeability (P _K) of 4.6×10^–13cm s^–1. Ca²⁺ ions applied to the intracellular membrane surface of K_Ca,ATP channels causes a marked enhancement of their activity. This activation is probably the result of simultaneous binding of at least two Ca²⁺ ions, determined using Hill analysis, to the channel or some closely associated protein. This results in a shift of the voltage activation threshold to more hyperpolarized membrane potentials. The activation of K_Ca,ATP channels by Mg-ATP has an EC₅₀ of approximately 50 M. Although the EC₅₀ is unaffected by [Ca²⁺]_i, channel activation by Mg-ATP is enhanced by increasing [Ca²⁺]_i. One possible interpretation of these data is that Mg-ATP increases the sensitivity of K_Ca,ATP channels to Ca²⁺. It is therefore possible that under hypoxic conditions, where lower levels of Mg-ATP may be encountered, the sensitivity of K_Ca,ATP channels to Ca²⁺ and therefore voltage is reduced. This would tend to induce a depolarising influence, which would favour the influx of Ca²⁺ through voltage-activated Ca²⁺ channels, ultimately leading to increased vascular tone.     

Role of calcium-dependent K+ channels in the regulation of arterial and venous tone by nitric oxide in pigs

Effects of inhibition of calcium-dependent potassium channels (K⁺ _Ca channels) on the regulation of arterial and venous tone by nitric oxide (NO) were studied in anaesthetized pigs following vagotomy and blockade of autonomic ganglia. Selective inhibition of K⁺ _Ca channels by charybdotoxin (CTX, 2 μg/kg iv) or iberiotoxin (IbTX, 1 μg/kg) significantly augmented mean total peripheral resistance (TPR) to levels 30–60% above control. Venous and pulmonary vascular tone were assessed by changes in effective compliances of the venous (EVC) and pulmonary (EPC) vascular beds as calculated from changes in central venous and diastolic pulmonary arterial blood pressure during haemorrhagia (−5 ml/kg) and hypervolaemia (+5 ml/kg). Blockade of K⁺ _Ca channels significantly decreased both EVC (−20 to −30%) and EPC (−30 to −50%). Both CTX and IbTX significantly diminished the vasodilation caused by the NO-donor S-nitroso-N-acetylpenicillamine (SNAP) both during control conditions and following experimental vasoconstriction induced by systemic inhibition of NO-synthesis by NG-nitro-L-arginine methyl ester (L-NAME) or infusion of vasoconstrictor agonists. Dilator effects of the adenosine 3′,5′-cyclic monophosphate (cAMP)-dependent agonist adenosine were only slightly reduced. However, blockade of K⁺ _Ca channels did not increase vasoconstriction induced by L-NAME significantly. These results suggest that activation of vascular K⁺ _Ca channels is an important mechanism by which NO attenuates the constrictor tone of resistance and capacitance vessels in vivo. + _Ca channel blockade during vasoconstriction by agonists The effects of CTX on the haemodynamic responses to infusions of AII, AVP, NA and ET-1, at doses producing similar increases in MAP of about 50 mmHg, are listed in Table 1. The increase in TPR caused by NA and ET-1 was significantly smaller after CTX, whereas the responses to AII and AVP were similar both before and after CTX. To characterize this effect of K⁺ _Ca channel blockade further, we constructed dose-response curves for AII and NA with and without pretreatment with IbTX. The results for TPR are shown in Fig. 5. The constrictor responses to the two lower doses of NA were significantly reduced by IbTX. Received: 12 February 1996 / Accepted: 31 March 1996     

前列腺素、一氧化氮和钾通道在鼠兔缺氧肺血管反应钝化中的作用

目的:探索前列腺素、一氧化氮和电压依赖性钾通道在高原动物鼠兔的缺氧性肺血管收缩反应钝化中的作用。方法:用鼠兔肺组织条替代肺血管进行实验,并以Wistar鼠肺组织条作为对照组。分别观察环加氧酶(cyclooxygenase,COX)抑制剂吲哚美辛(indomethacin)、一氧化氮合酶(nitricoxidesynthase,NOS)阻断剂L-NAME、电压门控的钾通道的阻断剂4-AP,对缺氧性肺血管收缩反应的影响。结果:(1)吲哚美辛组:吲哚美辛使鼠兔肺组织条的缺氧张力升高平均值比用吲哚美辛前增加64.7%;Wistar鼠肺组织条仅增加19.7%;两组差异显著,P＜0.05;(2)L-NAME组:L-NAME使鼠兔肺组织条缺氧性张力升高平均值增加102.7%;Wistar鼠肺组织条仅增加60.7%;两组差异显著,P＜0.05;(3)4-AP组:4-AP使鼠兔肺组织条缺氧张力平均值降低43.8%;Wistar鼠肺组织条降低28.53%;两组无显著差异,P＞0.05。结论:(1)NO和前列腺素在肺血管缺氧收缩反应中可能起调节作用,而电压门控的钾通道可能起介导作用;(2)鼠兔肺血管对缺氧收缩反应性钝化的机制中NO和前列腺素可能起更重要的介导作用,而电压门控的钾通道可能不起重要作用。     

Influence of age on L-type Ca2+ channels in the pulmonary artery and vein of spontaneously hypertensive rats

The influence of age on the density and localization of L-type Ca²⁺ channels was studied during development of hypertension in the pulmonary artery and vein of spontaneously hypertensive rats (SHR) and age-matched normotensive Wistar–Kyoto (WKY) rats by radioligand binding assay and light microscope autoradiography. SHR were examined at 6 weeks (juvenile, pre-hypertensive stage), 12 weeks (young, developing hypertension) and 24 weeks (mature, established hypertension). The dihydropyridine-type Ca²⁺ antagonist [³H]nicardipine was used as a radioligand. It was bound specifically to sections of rat pulmonary artery and vein. Dissociation constant (K_d) values were similar in WKY rats and SHR, whereas maximum density of binding sites (B_max) values increased in SHR in comparison with WKY rats. This increase was noticeable from the pre-hypertensive phase. The pharmacological profile of [³H]nicardipine binding was similar in different age groups of either normotensive and hypertensive rats. Quantitative analysis of autoradiographs from SHR revealed a progressive increase of silver grains in smooth muscle of tunica media and to a lesser extent in the adventitia of pulmonary artery but not of pulmonary vein from pre-hypertensive stage to developing hypertension. No further changes were observed in established hypertension. The above data indicate that the density of L-type Ca²⁺ channels of pulmonary arteries is increased in SHR. This augmentation after the pre-hypertensive phase suggests the occurrence of dysregulation of Ca²⁺ handling in the pulmonary vasculature of developing SHR.     

ATP敏感性钾通道的开放通过抑制TLR4/NF-κB通路对抗高糖引起的H9c2心肌细胞损伤和炎症

目的:探讨开放ATP敏感性钾通道(K_(ATP)通道)能否抑制Toll样受体4(TLR4)/核因子-κB(NF-κB)通路对抗高糖(HG)引起的H9c2心肌细胞损伤和炎症。方法:应用Western blot测定TLR4和NF-κB p65的蛋白水平;应用ELISA法检测细胞培养液中白细胞介素-1β(IL-1β)和肿瘤坏死因子-α(TNF-α)的水平;采用细胞计数试剂盒8(CCK-8)测定心肌细胞存活率;罗丹明123染色荧光显微镜照相法测定线粒体膜电位(MMP);双氯荧光素染色荧光显微镜照相法测定细胞内活性氧簇(ROS)水平;Hoechst 33258核染色荧光显微镜照相法检测凋亡细胞数量。结果:H9c2心肌细胞经HG(35 mmol/L葡萄糖)处理24 h,胞内TLR4和磷酸化NF-κB p65(p-NF-κB p65)的蛋白水平明显增加,100μmol/L K_(ATP)通道开放剂二氮嗪(DZ)预处理30 min可抑制HG对TLR4和p-NF-κB p65蛋白水平的上调作用;此外,30μmol/L TAK-242(TLR4抑制剂)和HG共处理心肌细胞24 h也可减轻HG对p-NF-κB p65的上调作用。另一方面,100μmol/L DZ预处理有明确的心肌保护作用,可抑制HG引起的细胞毒性、炎症反应、线粒体损伤、氧化应激和细胞凋亡,使细胞存活率升高,并减少IL-1β和TNF-α分泌水平、MMP丢失、ROS生成及凋亡细胞数量;而30μmol/L TAK-242或100μmol/L PDTC(NF-κB抑制剂)共处理心肌细胞24 h也可发挥和DZ相类似的作用,能抑制HG引起的上述损伤和炎症反应。结论:开放K_(ATP)通道可通过抑制TLR4/NF-κB通路对抗HG引起的H9c2肌细胞损伤和炎症。     

Urinary Ca2+ and the regulation of K+ secretion in toad bladder by neurohypophyseal hormones

We measured net K⁺ fluxes in the isolated toad urinary bladder to determine whether neurohypophyseal hormones control K⁺ secretion in this tissue. To determine if the pathway involved in K⁺ secretion is similar to a Ca²⁺-blockable alkali cation channel in the apical membrane of toad bladder, previously described in electrophysiological studies, we measured K⁺ fluxes in either the presence or absence of Ca²⁺ in the mucosal bathing solution. Oxytocin enhanced K⁺ secretion in both cases; however, the enhancement was markedly greater in the absence of mucosal Ca²⁺. In other experiments the transepithelial voltage was held constant at a value of 120 mV (mucosa negative) to find whether hyperpolarization would enhance K⁺ secretion to the levels seen in Ca²⁺-free solutions. The response to oxytocin was markedly greater in the absence of mucosal Ca²⁺ even when the transepithelial voltage was continuously hyperpolarized. These observations suggest that the properties of the activated pathway are akin to those of the previously described Ca²⁺-blockable alkali cation channel. We also found that toad bladder urine often contained extremely low levels of Ca²⁺; therefore neurohypophyseal hormonal control of K⁺ transport across the bladder may play an important role in amphibian K⁺ balance.     

Whole-cell and unitary Ca channel currents in mammalian intestinal smooth muscle cells: evidence for the existence of two types of Ca channels

Whole-cell and single Ca channel currents were recorded in smooth muscle cells isolated from guinea-pig taenia coli to examine whether multiple types of Ca channels exist. Two different types of voltage-dependent Ca channels with different conductances and inactivation kinetics were identified from cell-attached patch clamp recordings using 50 mM Ba in the patch pipettes. One type of channel, with a large conductance of 25 pS, had a threshold of activation near –40 mV and the mean current reconstructed by averaging individual current responses inactivated slowly. A second type of channel, with a small conductance of 12 pS, had a similar threshold value to that of the 25 pS channel, but the averaged current inactivated rapidly. The steady state inactivation of the 12 pS channel was complete at a holding potential of –40 mV. We concluded that both channel types represent fast and slow inactivating voltage-dependent Ca channels which have been found in many excitable cells.