The inositol trisphosphate pathway mediates platelet-activating-factor-induced pulmonary oedema

R. Göggel, S. Uhlig
European Respiratory Journal 2005 25: 849-857; DOI: 10.1183/09031936.05.00069804

Abstract

Platelet-activating factor (PAF) is a pro-inflammatory lipid mediator that increases vascular permeability by simultaneous activation of two pathways, one dependent on the cyclooxygenase metabolite prostaglandin E2 and the other on the sphingomyelinase metabolite ceramide. The hypothesis that part of the PAF-induced oedema is mediated via the inositol 1,4,5-trisphosphate (IP3) pathway or Rho kinase pathway was investigated.

Oedema formation was induced in isolated perfused rat lungs by injection of 5 nmol PAF into the pulmonary artery. Lungs were pre-treated with specific inhibitors: edelfosine (L108) to block phosphatidyl-inositol-specific phospholipase C, xestospongin to block the IP3 receptor, 5-iodonaphthalene-1-sulphonyl-homopiperazine (ML-7) to block myosin light chain kinase, and (+)-R-trans-4-(aminoethyl)-N-(4-pyridyl)cyclohexanecarboxamide (Y27632) to block Rho-associated protein kinase.

Pre-treatment with L108 or xestospongin reduced PAF-induced oedema formation by 58 and 56%, respectively. The effect of L108 was additive to that of the cyclooxygenase inhibitor acetyl salicylic acid (88% oedema reduction). PAF-induced oedema formation was also reduced if extracellular calcium concentrations were lowered. Furthermore, treatment with ML-7 reduced oedema formation by 54%, whereas Y27632 was without effect.

It is concluded that platelet-activating-factor-triggered oedema is mediated by activation of the inositol 1,4,5-trisphosphate pathway, influx of extracellular calcium and subsequent activation of a myosin light chain kinase-dependent and Rho-associated-protein-kinase-independent mechanism.

Platelet-activating factor (PAF) affects a variety of different lung functions, such as airway tone, vascular tone, ciliary beating frequency and endothelial permeability. Accordingly, PAF has been implicated in lung diseases such as acute respiratory distress syndrome 1 and asthma 2. Among these actions of PAF, the mechanisms of the pressor responses are known to depend on thromboxane and peptidoleukotrienes 3, mainly via activation of the Rho kinase pathway 4. The molecular mechanisms of PAF-induced oedema formation have only recently been studied in greater detail.

PAF induces oedema by activation of two lipid-modifying enzymes, cyclooxygenase (COX) and sphingomyelinase, which give rise to the formation of prostaglandin E2 and ceramide 5, 6. The signalling pathways through which these lipid mediators alter vascular permeability are largely unknown. The actions of prostaglandin E2 depend on voltage-gated potassium channels in a poorly understood manner 5, whereas the non-COX-dependent pathway is blocked by quinolines such as quinine, quinidine and chloroquine 7. Among the known molecular targets of quinolines are the inositol 1,4,5-trisphosphate (IP3) receptors 8, phospholipase A2 and phospholipase C (PLC) isoenzymes 9–11. A particular group of currently 11 PLCs, called phosphoinositide-specific PLC (PI-PLC) isozymes, triggers the rapid hydrolysis of membrane phosphatidylinositol 4,5-bisphosphate to generate two intracellular messengers, diacylglycerol and IP3 12, 13. While diacylglycerol activates protein kinase C (PKC), IP3 rapidly releases Ca2+ from intracellular Ca2+ pools within the endoplasmic reticulum and other cellular membranes by binding to the IP3 receptor (IP3R). Increased cytosolic Ca2+ levels regulate a plethora of intracellular processes, among them smooth muscle contraction 14 and vascular permeability 15. The Ca2+ signals are sensed by calcium-binding proteins, of which calmodulin is the most important. Many G-protein-coupled receptors are known to activate PI-PLC, among them PAF. Since, in mesangial cells, PAF has been shown to raise intracellular Ca2+ concentrations through the IP3 pathway 16 and because Ca2+ has frequently been implicated in oedema formation 17, it was speculated that this pathway might be involved in PAF-induced oedema formation in the lung.

Current models of the development of increased vascular permeability suggest that elevated intracellular Ca2+ concentrations activate myosin light chain kinase (MLCK), which, in turn, results in endothelial cell contraction and thus enhanced permeability 17, 18. In addition, a possible contribution of the small guanosine triphosphate (GTP)-binding protein Rho and Rho-associated protein kinase (ROCK) to the loss of barrier function has been described 19. However, the majority of the current knowledge regarding the role of Ca2+, IP3, MLCK and ROCK in vascular permeability was derived from studies using endothelial cells in culture, mostly with thrombin as a model agent 20. To date, these mechanisms have not been investigated in the whole intact organ, with the notable exception of a recent study showing reduced oedema formation in response to lipopolysaccharide and overventilation in mice that lacked endothelial MLCK 21. In addition, the involvement of the IP3 pathway, MLCK and ROCK in PAF-induced oedema has not yet been examined in the whole intact organ.

In vivo, besides bronchoconstriction 22, PAF produces many effects that may contribute to oedema formation, for example, hydrostatic mechanisms, such as activation of neutrophils 23, 24, pulmonary vasoconstriction 25, 26 and platelet aggregation 24. Although these factors may aggravate pulmonary oedema, the major factor in PAF-induced oedema formation appears to be increased vascular permeability. It is important to note that the PAF-induced alterations in vascular permeability are independent of neutrophils 27 and platelets 28. Here, the mechanisms of PAF-induced changes in vascular permeability are investigated using the model of the isolated blood-free perfused rat lung. Compared to the in vivo situation, this model permits the exclusion of oedema formation due to increased hydrostatic pressure by the use of constant pressure perfusion; accordingly, under these conditions, PAF does not alter capillary pressure 3, 5. Conversely, PAF increases the capillary filtration coefficient and vascular permeability 5, 29. Thus, in the present model, PAF-induced pressor responses and oedema formation can be completely separated; the pressor responses but not oedema formation are prevented by thromboxane and leukotriene antagonists 7, whereas, conversely, treatment with ceramide antibodies attenuates oedema formation without affecting the pressor responses 6. In summary, constant-pressure perfused isolated lungs are ideally suited to investigation of the mechanisms of increased vascular permeability in the whole intact organ. Here, this model has been used to investigate the role of the IP3 pathway, extracellular Ca2+, MLCK and ROCK in PAF-induced oedema formation.

MATERIALS AND METHODS

Study design

Lungs were always perfused for 10 min under control conditions (tidal volume 1.8–2 mL, pulmonary artery pressure 10 cmH2O and perfusate flow rate 30 mL·min−1) before capillary filtration coefficient (Kf,c) measurement, for which the arterial and venous reservoir were raised by 5 cm for 10 min. The Kf,c was also measured 30 min after addition of PAF (Sigma, Deisenhofen, Germany), which was always injected as a 5-nmol bolus directly into the perfusate (which contained fraction V bovine albumin (Serva, Heidelberg, Germany) to maintain oncotic pressure) after 30 min of perfusion. All other agents were added to the buffer reservoir. Acetylsalicylic acid (ASA) (Sigma) was made up in bicarbonate solution and added 10 min before PAF. The phospholipase inhibitor edelfosine (L108) (Biomol, Hamburg, Germany; median inhibitory concentration (IC50) 10 μM 30, used at 30 μM) and the membrane-permeable IP3R antagonist xestospongin C (Biomol; IC50 0.4 μM 31, used at 1 μM) were prepared as stock solutions in ethanol, the MLCK inhibitor 5-iodonaphthalene-1-sulphonyl-homopiperazine (ML-7) (Biomol; IC50 0.3 μM 31, used at 35 μM) in 50% aqueous ethanol, and the ROCK inhibitor (+)-R-trans-4-(aminoethyl)-N-(4-pyridyl)cyclohexanecarboxamide (Y27632) (Tocris, Avonmouth, UK; IC50 0.8 μM 32, 33, used at 10 μM) in dimethyl sulphoxide; aliquots from these stock solutions were added 10 min before PAF. The calmodulin-dependent protein kinase II (CaMKII) inhibitor K252a (Biomol; IC50 2 nM 34, used at 10 nM (IC50 for inhibition of protein kinase A (PKA), PKC and MLCK 20 nM 35, 36)) was used to inhibit CaMKII. For the experiments on the role of extracellular Ca2+, the lungs were perfused with Krebs-Henseleit buffer containing different Ca2+ concentrations from the beginning of the preparation. None of the solvents alone had any affect on PAF-induced weight gain (data not shown).

The data on airway and vascular resistance, for most of the experiments shown in the present study, have previously been reported 4, and are therefore not repeated here. However, this information is not pertinent to the present study since, in the present model, oedema formation occurs completely independently of the haemodynamic changes (see introduction).

Isolated perfused rat lung preparation

The isolated perfused rat lung set-up has previously been described in detail 37, 38. Briefly, female Wistar rats (Harlan Winkelmann, Borchen, Germany; mean±sd 220±20 g body weight) were anaesthetised by intraperitoneal injection of 16 mg·kg body weight−1 pentobarbital sodium (Nembutal®; Wirtschaftsgenossenschaft Deutscher Tierärzte, Hanover, Germany). They were intubated with a tracheal cannula and ventilated at 80 breaths·min−1 on room air with a tidal volume of 1.8–2.1 mL. Every 5 min, a deep breath (hyperinflation at -16 cmH2O) was initiated in order to prevent atelectasis. After laparotomy, the diaphragm was removed. The animals were ex-sanguinated and the chest was opened. A ligature was placed around the pulmonary artery and aorta. An arterial cannula was inserted into the pulmonary artery and fixed by the ligature. Then, the apex of the heart was cut off, the venous cannula inserted into the left atrium and fixed by a ligature around the heart. The excised lungs were perfused at constant hydrostatic pressure, which resulted in a flow rate of ∼30 mL·min−1 through the pulmonary artery with Krebs–Henseleit buffer containing 2% albumin, 0.1% glucose and 0.3% hydroxyethyl piperazine ethanesulphonic acid (HEPES). The total recirculating volume of buffer was 100 mL. The lungs were suspended by the trachea and ventilated by negative pressure ventilation in an artificial thorax chamber. Thorax chamber pressure was measured using a differential pressure transducer, and air flow velocity was measured via a pneumotachograph tube connected to a differential pressure transducer. Lung weight was followed with a specifically designed weight transducer 29. The Kf,c was measured by fitting the weight gain to a bi-exponential equation, as described previously 39. Perfusate flow, buffer pH, and arterial and venous pressure were continuously monitored. The pH of the perfusate before entering the lung was kept at 7.35 by automatic bubbling of carbon dioxide into the buffer.

The data for the control groups (PAF and control) are from experiments performed over recent years, in which it was checked, at regular intervals, that PAF and perfusion without any treatment give the expected responses. These data show that the response to PAF is highly reproducible over time. During studies on the mechanisms of PAF-induced oedema, various hypotheses were followed simultaneously; therefore, the data from the control groups have also been used in previous studies performed at the same time 5, 6.

Statistical analysis

The data in the tables are presented as mean±sd, and data in the graphs as mean±sem. In case of heteroskedasticity, data were logarithmically transformed prior to analysis. Data were analysed using an unpaired t-test, one-sided or two-sided as indicated. The false discovery rate due to multiple comparisons was controlled by the method of Benjamini and Hochberg 40. This procedure controls the false discovery rate, i.e. the expected proportion of false discoveries amongst the rejected hypotheses, and is a less stringent condition than the family-wise error rate, making this method more powerful. This method has recently been recommended “as the best practical solution to the problems of multiple comparisons that exist within science” 41. A p-value of <0.05 was considered significant.

RESULTS

Perfusing lungs with PAF increased lung weight as shown before 5, 29. The contribution of the IP3 pathway to PAF-induced oedema was investigated using L108, an inhibitor of PI-PLC, and xestospongin C, a sponge toxin that interferes with the IP3R.

Pre-treating the lungs with 30 μM L108 attenuated the PAF-induced increase in lung weight (fig. 1⇓, table 1⇓) and Kf,c (table 1⇓). As mentioned above, PAF induces oedema via two separate mechanisms that can be blocked by the COX inhibitor ASA and quinine 7. When the lungs were pre-treated with ASA, oedema formation was reduced by ∼34%, and when ASA was co-perfused with L108, the effect of PAF on lung weight gain was almost completely inhibited (fig. 1⇓, table 1⇓). Pre-treatment with xestospongin C (1 μM) attenuated the PAF-induced weight gain and the increase in Kf,c to the same extent as did L108 (table 1⇓). Furthermore, heparin, another agent reported to inhibit IP3Rs 42, 43, had beneficial effects on PAF-induced oedema formation (table 1⇓). However, heparin may bind directly to PAF 44. Since, in the present experiments, heparin was effective against both oedema formation and pressor responses (data not shown) the possibility that the beneficial effects of heparin were simply due to binding of PAF cannot be excluded.

Fig. 1—
Fig. 1—

Effect of acetylsalicylic acid (ASA) and edelfosine (L108) on platelet-activating factor (PAF)-induced oedema formation (•: PAF (n = 42); ▴: ASA+PAF (n = 9); ▪: L108+PAF (n = 4); ♦: ASA/L108+PAF (n = 4); ○: control (n = 12)). PAF was given as a 5-nmol bolus injection 30 min after beginning the experiment. ASA and L108 were given 10 min before injection of PAF. Data are presented as mean±sem; for statistics, see table 1⇓.

View this table:
Table. 1—

-activating factor (PAF)-induced oedema formation: inhibition of the inositol 1,4,5-trisphosphate pathway

Activation of the IP3 pathway and subsequent emptying of intracellular Ca2+ stores may lead to activation of store-operated calcium channels (SOCs) 45, 46. In order to examine the importance of extracellular Ca2+, lungs were perfused with different Ca2+ concentrations. These experiments were difficult to carry out, since reduced extracellular Ca2+ itself may enhance permeability in perfused lungs 47, 48. The lowest Ca2+ concentration tolerated was 250 μM. Reduction of extracellular Ca2+ concentration reduced PAF-induced weight gain in a concentration-dependent manner (fig. 2a⇓). PAF-induced weight gain was also attenuated by the unspecific calcium channel inhibitor lanthanum chloride (fig. 2b⇓).

Fig. 2—
Fig. 2—

Effect on platelet-activating factor (PAF)-induced oedema formation of a) reduced Ca2+ concentration (normal concentration 2.5 mM) and b) lanthanum chloride (2.5 mM Ca2+). PAF was given as a 5-nmol bolus injection 30 min after beginning the experiment (▓), and buffer containing different Ca2+ concentrations was used from the beginning of the experiment (□: no PAF). The unspecific Ca2+ channel inhibitor lanthanum chloride was added 20 min before injection of PAF (n = 5). Data are presented as mean±sem (no PAF (2.5 mM Ca2+): n = 12; 0.25 mM Ca2+: n = 7; 1.25 mM Ca2+: n = 3; 2.5 mM Ca2+: n = 42; 5.0 mM Ca2+: n = 3). *: p<0.05 versus PAF alone (2.5 mM Ca2+).

One mechanistic explanation for the increased vascular permeability of endothelial cells is endothelial cell contraction and gap formation. This phenomenon is assumed to be at least partly dependent on activation of MLCK 49. ML-7 is a relatively specific, albeit rather weak, inhibitor of MLCK 50. Inhibition of MLCK with ML-7 (35 μM) reduced PAF-induced oedema formation (fig. 3⇓, table 2⇓) and attenuated the PAF-induced increase in Kf,c (table 2⇓). In contrast, inhibition of CaMKII with K252a (10 nM) resulted in enhanced oedema formation (table 2⇓), an observation which might be explained by the observation that, under certain conditions, CaMKII counter-regulates the action of MLCK 51. Further experiments with calmidazolium chloride (10 μM), an inhibitor of Ca2+/calmodulin that, amongst other things, also induces MLCK activation, resulted in strong oedema formation even in the absence of PAF (data not shown).

Fig. 3—
Fig. 3—

Inhibition of myosin light chain kinase (MLCK) reduces platelet-activating factor (PAF)-induced oedema formation (•: PAF (n = 42); ▪: 5-iodonaphthalene-1-sulphonyl-homopiperazine (ML-7)+PAF (n = 4); ○: control (n = 12)). PAF was given as a 5-nmol bolus injection 30 min after beginning the experiment. The specific MLCK inhibitor ML-7 was administered 10 min before injection of PAF. Data are expressed as mean±sem. Oedema formation in lungs pre-treated with ML-7 and control lungs was significantly reduced compared to PAF alone.

View this table:
Table. 2—

-activating factor (PAF)-induced oedema formation: inhibition of myosin light chain kinase and Rho-associated protein kinase

It was recently shown that thrombin-induced endothelial cell monolayer hyperpermeability results from increased F-actin stress-fibre-related contractile tension, a process regulated by the small GTP-binding protein Rho and ROCK 19. Y27632 is a highly specific ROCK inhibitor that inhibits other kinases such as PKA, PKC and MLCK, but only at concentrations of >25 μM 30, 31. Treatment with Y27632 influenced neither PAF-induced weight gain nor the increase in Kf,c (table 2⇑).

DISCUSSION

PAF is an important mediator of oedema formation in many different models of acute lung injury, including endotoxic shock 5, 52, intestinal ischemia–reperfusion 53 and acid instillation 54. PAF fulfils many of the criteria of a terminal executor of increased vascular permeability: it is formed during inflammation, it increases vascular permeability, and it does so in <10 min 6. The intracellular signalling pathways of PAF-induced oedema formation are only poorly defined. The present findings suggest that part of the PAF-induced pulmonary oedema is mediated via Ca2+-dependent activation of MLCK, the Ca2+ originating from intracellular IP3-sensitive stores and from the extracellular space.

Given the importance of PAF in many models of acute lung injury, surprisingly little is known about the underlying mechanisms. One probable reason for the lack of mechanistic studies is the lack of effect of PAF on endothelial cell permeability in vitro 55–57, although such an effect has been reported once 58. Therefore, nearly all of the studies on this subject have been performed in isolated organs or whole animals. However, results from whole-animal studies are hard to interpret, partly because the current methods for measuring oedema formation in vivo are not very sensitive, and partly because the haemodynamic changes evoked by PAF make it very difficult to distinguish between permeability and hydrostatic types of oedema. Therefore, many authors have used isolated perfused lungs to study the mechanisms of PAF-induced oedema formation. Such studies have identified several agents that attenuate PAF-induced oedema formation, such as COX inhibitors 5, steroids 6, 59, vitamin D3 60, agents increasing cyclic adenosine monophosphate levels 61, tricyclodecan-9-yl xanthogenate (D609) and ceramide-specific antibodies 6, but these studies have not yet led to the identification of a specific intracellular signalling mechanism.

At present, only one PAF receptor is known, a typical seven-transmembrane domain receptor coupled to G proteins 62. In mesangial cells, it was shown that activation of this receptor activates PI-PLC to produce IP3 that, upon binding to its receptor, releases Ca2+ from intracellular stores 16, 45. The present findings in the intact organ are in line with this sequence of events (fig. 4⇓). Inhibition of PI-PLC with L108 and blockade of IP3Rs with xestospongin C attenuated the PAF-induced oedema formation and increase in Kf,c. L108 and xestospongin C were selected because they are cell permeable and can therefore, be used in studies with intact organs. L108 is also known as ET-18-OCH3 and acts as a proapoptotic agent 63. In kidney cells, it has been suggested that L108 may activate PAF receptors 63, but since, in the present experiments, L108 had no effect when given alone (it caused neither oedema formation nor contraction of airway or vascular smooth muscle), the present authors do not believe that this happens in the current model. Xestospongin C (20 μM) has been shown to discriminate between IP3-sensitive and ryanodine-sensitive Ca2+ stores 64; however, at least in some tissues, xestospongin C may also block voltage-dependent potassium and calcium channels 65. Reasonable specificity of xestospongin C and L108 in the present study is suggested by the fact that both inhibitors were used at concentrations only three times greater than their IC50, the observation that both inhibitors failed to affect other responses to PAF (in particular PAF-induced bronchoconstriction) 4, and the fact that the only known common denominator of both inhibitors is inhibition of the IP3 pathway. In addition, both PLC and IP3Rs have been described as targets for quinine, which also reduces PAF-induced oedema formation 7. Alternative to an increase in Ca2+ concentration caused by response to IP3, intracellular Ca2+ levels might also be increased by ceramide 66, which was recently identified as a mediator of PAF 6. This interesting possibility needs to be explored further.

Fig. 4—
Fig. 4—

Summary of intracellular signalling pathways of rat lungs in response to platelet-activating factor (PAF). The left-hand side shows the effect of PAF on what are presumably endothelial cells as inferred from the present study (for further details, see Discussion section). The right-hand side shows the effect of PAF on what are presumably vascular and airway smooth muscle cells, which is largely mediated by thromboxane. Here, the major pathway is activation of Rho and Rho-associated protein kinases (p160 ROCK), which attenuate the activity of myosin light chain phosphatase (MLCP) with consequent increases in myosin light chain phosphorylation and, hence, contraction 4. PLC: phospholipase C; IP3: inositol 1,4,5-trisphosphate; IP3R: IP3 receptor; MLCK: myosin light chain kinase; ML-7: 5-iodonaphthalene-1-sulphonyl-homopiperazine; L108: edelfosine. –––∣: inhibition; ↑: increase or activation.

The intracellular Ca2+ response to PAF has been shown to be biphasic 67, 68. The first phase is mediated by IP3Rs which are regulated by Ca2+/calmodulin, whereas the second phase occurs due to transmembrane Ca2+ movement. The increase in intracellular Ca2+ concentrations during phase one activates SOCs 43–46. Alternatively, activation of the PAF receptor may directly activate other calcium channels in the plasma membrane 69. Manipulation of extracellular Ca2+ content or unspecific inhibition of calcium channels attenuated PAF-induced pulmonary oedema formation (fig. 2⇑). This argues for a role of a capacitive Ca2+ entry through SOCs or receptor-coupled activation of calcium channels in PAF-induced oedema formation. This interpretation is supported by the finding that thapsigargin, an inhibitor of the endoplasmic reticular Ca2+-adenosine triphosphatase, which leads to emptied intracellular Ca2+ stores and activation of SOCs, induces pulmonary oedema formation by a mechanism that requires extracellular Ca2+ (data not shown) 70.

Increased intracellular Ca2+ concentration activates MLCK, an effect mediated by Ca2+/calmodulin 71. In line with this, it was shown that the enhanced permeability induced by PAF was attenuated by pre-treatment with an MLCK inhibitor, indicating that myosin phosphorylation plays an important role in PAF-induced oedema formation. Recently, the significance of the endothelial MLCK in endotoxin-induced pulmonary oedema formation was demonstrated 21. Since, in this model, PAF is critical to oedema formation 5, 52, the blockade of MLCK in that study may also be explained by protection against the effects of PAF.

With respect to myosin light chain phosphorylation, a Rho-kinase-dependent pathway of enhanced permeability 19 and cytoskeletal reorganisation of nonmuscle cells 72 has recently been suggested. Carbajal et al. 19 reported that thrombin-induced monolayer hyperpermeability was partially mediated via ROCK. However, in the present study, the specific ROCK inhibitor Y27632 failed to affect PAF-induced weight gain, indicating that Rho kinase does not mediate PAF-induced oedema. The lack of effect of Y27632 on oedema formation is also supported by another recent study showing that, at relevant concentrations, Y27632 does not attenuate airway microvascular leakage evoked by histamine or leukotriene 73. In both perfused rat lungs and guinea pigs, however, Y27632 was effective in inhibiting airflow obstruction 4, 73. Taken together, these data suggest that, in the intact organ, the Rho kinase pathway contributes to smooth muscle contraction, but has no role in permeability oedema formation (fig. 4⇑).

Unfortunately, the role of calmodulin could not be explored, because inhibition of Ca2+-binding to calmodulin with calmidazolium caused severe pulmonary oedema of itself. Nevertheless, this is an interesting finding, suggesting that calmodulin is essential for the maintenance of the normal vascular barrier function. CaMKII was reported to counter-regulate the action of MLCK in smooth muscle cells. In these experiments, phosphorylation of MLCK decreased the Ca2+ sensitivity of myosin light chain phosphorylation 51. Therefore, inhibition of CaMKII should lead to enhanced MLCK activity. In line with this, blocking CaMKII with K252a resulted in increased oedema formation in the presence of PAF. It is of note that these results were obtained using 10 nM K252a, a concentration at which this inhibitor should not block other enzymes, such as PKA, MLCK and PKC 35, 36. Again, the present findings are in contrast to observations in endothelial monolayers, in which inhibition of CaMKII reduced thrombin-induced hyperpermeability 74. Whether these differences are to be attributed to the cell culture model (where PAF does not act) or to differences between thrombin and PAF remains to be elucidated.

Taken together, the present findings suggest that platelet-activating factor activates phosphoinositide-specific phospholipase C, which, in turn, produces inositol 1,4,5-trisphosphate. This increase in cytosolic calcium concentration then leads to activation of calcium-gated or receptor-activated calcium channels, leading to a secondary calcium influx from the extracellular space and subsequent activation of myosin light chain kinase.

Acknowledgments

The perfect technical assistance of F. Seel is gratefully acknowledged.

  • Received June 11, 2004.
  • Accepted December 27, 2004.

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The inositol trisphosphate pathway mediates platelet-activating-factor-induced pulmonary oedema
R. Göggel, S. Uhlig
European Respiratory Journal May 2005, 25 (5) 849-857; DOI: 10.1183/09031936.05.00069804
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