Jeremy P. T. Ward

Hypoxic Pulmonary Vasoconstriction

It has been known for more than 60 years, and suspected for over 100, that alveolar hypoxia causes pulmonary vasoconstriction by means of mechanisms local to the lung. For the last 20 years, it has been clear that the essential sensor, transduction, and effector mechanisms responsible for hypoxic pulmonary vasoconstriction (HPV) reside in the pulmonary arterial smooth muscle cell. The main focus of this review is the cellular and molecular work performed to clarify these intrinsic mechanisms and to determine how they are facilitated and inhibited by the extrinsic influences of other cells. Because the interaction of intrinsic and extrinsic mechanisms is likely to shape expression of HPV in vivo, we relate results obtained in cells to HPV in more intact preparations, such as intact and isolated lungs and isolated pulmonary vessels. Finally, we evaluate evidence regarding the contribution of HPV to the physiological and pathophysiological processes involved in the transition from fetal to neonatal life, pulmonary gas exchange, high-altitude pulmonary edema, and pulmonary hypertension. Although understanding of HPV has advanced significantly, major areas of ignorance and uncertainty await resolution.

Communication and information technology in medical education

The past few years have seen rapid advances in communication and information technology (C&IT), and the pervasion of the worldwide web into everyday life has important implications for education. Most medical schools provide extensive computer networks for their students, and these are increasingly becoming a central component of the learning and teaching environment. Such advances bring new opportunities and challenges to medical education, and are having an impact on the way that we teach and on the way that students learn, and on the very design and delivery of the curriculum. The plethora of information available on the web is overwhelming, and both students and staff need to be taught how to manage it effectively. Medical schools must develop clear strategies to address the issues raised by these technologies. We describe how medical schools are rising to this challenge, look at some of the ways in which communication and information technology can be used to enhance the learning and teaching environment, and discuss the potential impact of future developments on medical education.

Voltage-independent calcium entry in hypoxic pulmonary vasoconstriction of intrapulmonary arteries of the rat

1 It has been proposed that hypoxic pulmonary vasoconstriction (HPV) is mediated via K+ channel inhibition and Ca2+ influx through voltage‐gated channels. HPV depends strongly on the degree of preconstriction, and we therefore examined the effect of Ca2+ channel blockade on tension and intracellular [Ca2+] ([Ca2+]i) during HPV in rat intrapulmonary arteries (IPAs), whilst maintaining preconstriction constant. We also investigated the role of intracellular Ca2+ stores. 2 HPV demonstrated a transient constriction (phase I) superimposed on a sustained constriction (phase II). Nifedipine (1 μm) partially inhibited phase I, but did not affect phase II. In arteries exposed to 80 mm K+ and nifedipine or diltiazem the rises in tension and [Ca2+]i were blunted during phase I, but were unaffected during phase II. 3 At low concentrations (< 3 μm), La3+ almost abolished the phase I constriction and rise in [Ca2+]i, but had no effect on phase II, or constriction in response to 80 mm K+. Phase II was inhibited by higher concentrations of La3+ (IC50∼50 μm). 4 IPA treated with thapsigargin (1 μm) in Ca2+‐free solution to deplete Ca2+ stores showed sustained constriction upon re‐exposure to Ca2+ and an increase in the rate of Mn2+ influx, suggesting capacitative Ca2+ entry. The concentration dependency of the block of constriction by La3+ was similar to that for phase I of HPV. Pretreatment of IPA with 30 μm CPA reduced phase I by > 80%, but had no significant effect on phase II. 5 We conclude that depolarization‐mediated Ca2+ influx plays at best a minor role in the transient phase I constriction of HPV, and is not involved in the sustained phase II constriction. Instead, phase I appears to be mainly dependent on capacitative Ca2+ entry related to release of thapsigargin‐sensitive Ca2+ stores, whereas phase II is supported by Ca2+ entry via a separate voltage‐independent pathway.

Dietary soy isoflavone-induced increases in antioxidant and eNOS gene expression lead to improved endothelial function and reduced blood pressure in vivo

Epidemiological evidence suggests that populations consuming large amounts of soy protein have a reduced incidence of coronary heart disease (1–5). The cardiovascular risks associated with conventional hormone replacement therapy in postmenopausal women (5–7) have precipitated a search for alternative estrogen receptor modulators. Here we report that long‐term feeding of rats with a soy protein‐rich (SP) diet during gestation and adult life results in decreased oxidative stress, improved endothelial function, and reduced blood pressure in vivo measured by radiotelemetry in aged male offspring. Improved vascular reactivity in animals fed an SP diet was paralleled by increased mitochondrial glutathione and mRNA levels for endothelial nitric oxide synthase (eNOS) and the antioxidant enzymes manganese superoxide dismutase and cytochrome c oxidase. Reduced eNOS and antioxidant gene expression, impaired endothelial function, and elevated blood pressure in animals fed a soy‐deficient diet was reversed after refeeding them an SP diet for 6 months. Our findings suggest that an SP diet increases eNOS and antioxidant gene expression in the vasculature and other tissues, resulting in reduced oxidative stress and increased NO bioavailability. The improvement in endothelial function, increased gene expression, and reduced blood pressure by soy isoflavones have implications for alternative therapy for postmenopausal women and patients at risk of coronary heart disease.

Inhibition of sustained hypoxic vasoconstriction by Y-27632 in isolated intrapulmonary arteries and perfused lung of the rat

We have examined the effects of Y‐27632, a specific inhibitor of Rho‐activated kinases (ROCK I and ROCK II) upon sustained hypoxic pulmonary vasoconstriction (HPV) in both rat isolated small intrapulmonary arteries (IPA) and perfused rat lungs in situ. Y‐27632 (100 nM–3 μM) was found to cause a concentration‐dependent inhibition of acute sustained HPV in rat IPA. Application of Y‐27632 (10–600 nM) in perfused rat lungs caused no change in basal perfusion pressure, but was found to inhibit HPV in a concentration‐dependent manner, resulting in complete ablation of the pressor response to hypoxia at a concentration of 600 nM. Furthermore, addition of Y‐27632 at any point during hypoxia caused a reversal of HPV in perfused rat lungs. These results suggest that activation of Rho‐associated kinase may be a pivotal step in the generation of sustained HPV.

Divergent roles of glycolysis and the mitochondrial electron transport chain in hypoxic pulmonary vasoconstriction of the rat: identity of the hypoxic sensor

1 The mechanisms responsible for sensing hypoxia and initiating hypoxic pulmonary vasoconstriction (HPV) are unclear. We therefore examined the roles of the mitochondrial electron transport chain (ETC) and glycolysis in HPV of rat small intrapulmonary arteries (IPAs). 2 HPV demonstrated a transient constriction (phase 1) superimposed on a sustained constriction (phase 2). Inhibition of complex I of the ETC with rotenone (100 nm) or complex III with myxothiazol (100 nm) did not cause vasoconstriction in normoxia, but abolished both phases of HPV. Rotenone inhibited the hypoxia‐induced rise in intracellular Ca2+ ([Ca2+]i). Succinate (5 mm), a substrate for complex II, reversed the effects of rotenone but not myxothiazol on HPV, but did not affect the rise in NAD(P)H fluorescence induced by hypoxia or rotenone. Inhibition of cytochrome oxidase with cyanide (100 μm) potentiated phase 2 constriction. 3 Phase 2 of HPV, but not phase 1, was highly correlated with glucose concentration, being potentiated by 15 mm but abolished in its absence, or following inhibition of glycolysis by iodoacetate or 2‐deoxyglucose. Glucose concentration did not affect the rise in [Ca2+]i during HPV. 4 Depolarisation‐induced constriction was unaffected by hypoxia except in the absence of glucose, when it was depressed by ∼50 %. Depolarisation‐induced constriction was depressed by rotenone during hypoxia by 23 ± 4 %; cyanide was without effect. 5 Hypoxia increased 2‐deoxy‐[3H]glucose uptake in endothelium‐denuded IPAs by 235 ± 32 %, and in mesenteric arteries by 218 ± 38 %. 6 We conclude that complex III of the mitochondrial ETC acts as the hypoxic sensor in HPV, and initiates the rise in smooth muscle [Ca2+]i by a mechanism unrelated to changes in cytosolic redox state per se, but more probably by increased production of superoxide. Additionally, glucose and glycolysis are essential for development of the sustained phase 2 of HPV, and support an endothelium‐dependent Ca2+‐sensitisation pathway rather than the rise in [Ca2+]i.

Activation of nitric oxide synthase by β2‐adrenoceptors in human umbilical vein endothelium in vitro

Some animal studies suggest that β‐adrenoceptor‐mediated vasorelaxation is in part mediated through nitric oxide (NO) release. Furthermore, in humans, we have recently shown that forearm blood flow is increased by infusion of β2‐adrenergic agonists into the brachial artery, and the nitric oxide synthase (NOS) inhibitor NG‐monomethyl‐L‐arginine (L‐NMMA) inhibits this response. The purpose of the present study was to determine whether stimulation of human umbilical vein endothelial β‐adrenoceptors causes vasorelaxation and nitric oxide generation, and whether this might be mediated by cyclic adenosine‐3′,5′‐monophosphate (cyclic AMP). Vasorelaxant responses were determined in umbilical vein rings to the nonselective β‐adrenergic agonist isoprenaline and to the cyclic AMP analogue dibutyryl cyclic AMP, following precontraction with prostaglandin F2α. NOS activity was measured in cultured human umbilical vein endothelial cells (HUVEC) by the conversion of [3H]‐L‐arginine to [3H]‐L‐citrulline, and adenylyl cyclase activity by the conversion of [α‐32P]‐ATP to [32P]‐cyclic AMP. Isoprenaline relaxed umbilical vein rings, and this vasorelaxation was abolished by β2‐ (but not β1‐) adrenergic blockage, and by endothelium removal or 1 mM L‐NMMA. In addition, vasorelaxant responses to dibutyryl cyclic AMP were inhibited by 1 mM L‐NMMA, with a reduction in Emax from 90.0±9.3% to 50.5±9.9% (P<0.05). Isoprenaline 1 μM increased NOS activity in HUVEC (34.0±5.9% above basal, P<0.001). Furthermore, isoprenaline increased adenylyl cyclase activity in a concentration‐dependent manner; this response was inhibited by β2 (but not β1‐) adrenergic blockade. Forskolin 1 μM and dibutyryl cyclic AMP 1 mM each increased NOS activity in HUVEC, to a degree similar to isoprenaline 1 μM. The increase in L‐arginine to L‐citrulline conversion observed with each agent was abolished by co‐incubation with NOS inhibitors. These results indicate that endothelial β2‐adrenergic stimulation and cyclic AMP elevation activate the L‐arginine/NO system, and give rise to vasorelaxation, in human umbilical vein.

Hypoxic pulmonary vasoconstriction: mechanisms and controversies

The pulmonary circulation differs from the systemic in several important aspects, the most important being that pulmonary arteries constrict to moderate physiological (∼20–60 mmHg PO2) hypoxia, whereas systemic arteries vasodilate. This phenomenon is called hypoxic pulmonary vasoconstriction (HPV), and is responsible for maintaining the ventilation–perfusion ratio during localized alveolar hypoxia. In disease, however, global hypoxia results in a detrimental increase in total pulmonary vascular resistance, and increased load on the right heart. Despite many years of study, the precise mechanisms underlying HPV remain unresolved. However, as we argue below, there is now overwhelming evidence that hypoxia can stimulate several pathways leading to a rise in the intracellular Ca2+ concentration ([Ca2+]i) in pulmonary artery smooth muscle cells (PASMC). This rise in [Ca2+]i is consistently found to be relatively small, and HPV seems also to require rho kinase‐mediated Ca2+ sensitization. There is good evidence that HPV also has an as yet unexplained endothelium dependency. In this brief review, we highlight selected recent findings and ongoing controversies which continue to animate the study of this remarkable and unique response of the pulmonary vasculature to hypoxia.

Mechanisms of hypoxic pulmonary vasoconstriction: can anyone be right?

Despite extensive studies over many years. there is still no real consensus regarding the mechanisms responsible for hypoxic pulmonary vasoconstriction (HPV). This is partially related to extensive variation between preparations, species, and the length of the hypoxic challenge, but also to an apparent abundance of potential mechanisms. Whereas there is good evidence that hypoxia causes inhibition of K channels in pulmonary artery smooth muscle, with subsequent depolarisation and Ca2+ influx through voltage-activated Ca2+ channels, there is also strong support for a critical role for Ca2+ release from intracellular stores. Moreover other studies suggest that the endothelium provides an essential component of the overall response. We suggest in this review that sustained HPV, as seen in the intact animal, is multi-factorial in origin and requires activation of more than one process for the full response to develop. Fundamental issues that remain unresolved are outlined.

Mechanisms of hypoxic pulmonary vasoconstriction and their roles in pulmonary hypertension: new findings for an old problem

Hypoxic pulmonary vasoconstriction (HPV) normally optimises ventilation-perfusion matching in the lung, but leads to pulmonary hypertension (PH) under conditions of global hypoxia. The past few years have provided some major advances in our understanding of this complex phenomenon, but significant controversy remains concerning many of the key underlying mechanisms. On balance, recent evidence is most consistent with an elevation in mitochondria-derived reactive oxygen species as a key event for initiation of HPV, with consequent Ca2+ release from intracellular ryanodine-sensitive stores, although the activation pathways and molecular identity of the associated Ca2+ entry pathways remain unclear. Recent studies have also raised our perception of the critical role played by Rho kinase (ROCK) in both sustained HPV and the development of PH, further promoting ROCK and the pathways regulating its activity and expression as important therapeutic targets.