Developmental expression of vasoactive and growth factors in human lung. Role in pulmonary vascular resistance adaptation at birth.

The factors that mediate the postnatal fall in pulmonary vascular resistance, which is crucial for normal gas exchange, are not fully understood. The endothelium has been implicated in this phenomenon, through the release of vasorelaxant factors such as nitric oxide (NO). Human pulmonary expression of endothelial NO synthase increases up to 31 wk of gestation, together with vascular endothelial growth factor (VEGF), and both factors potently mediate pulmonary angiogenesis and vasorelaxation. During the perinatal period, when pulmonary vasodilatation is maximal, endothelial NO synthase and VEGF are weakly expressed. This raises the involvement of vasorelaxant factors other than NO at birth. One candidate is endothelial-derived hyperpolarizing factor, which induces smooth muscle cell hyperpolarization by activating K(ATP) channels. The marked vasorelaxation induced by activation of these channels in newborn animals, and their strong perinatal expression in the human lung, suggest their involvement during this phase. Another candidate is endothelin (ET)-1, together with its receptors ET-A and ET-B. ET-A receptors are located exclusively on smooth muscle cells and mediate vasoconstriction, whereas ET-B receptors mediate vasoconstriction when located on smooth muscle cells and vasodilatation when located on endothelial cells. ET-B receptors, which are strongly expressed in the human fetal lung both at the end of gestation and after birth, may be involved in perinatal pulmonary vasodilatation. Thus, in human fetal lung, K(ATP) channels and ET-B receptors could be important in mediating the perinatal pulmonary vasodilatation crucial for adapting the pulmonary circulation to extrauterine life.

Noninvasive assessment of fetal pulmonary blood flow in experimental pulmonary hypertension in the fetal lamb.

The aim of this study was to assess pulmonary arterial blood flow changes induced by the creation of a systemic arteriovenous fistula (120 d gestation) in the fetal lamb using Doppler technique. Doppler echocardiographic assessment of the pulmonary artery blood flow performed 1, 6, and 14 d after surgery showed that mean pulmonary arterial blood flow in the left or right pulmonary artery was 224 +/- 58 mL/min at day 1 in the fistula group, significantly higher than in the control group (113 +/- 22 mL/min; p < 0.01, ANOVA test) whether no difference was found at days 6 and 14. The mean inner diameter of the left pulmonary artery measured on postmortem lung arteriograms compared favorably to the one measured on day 14 at the same level on ultrasound. The mean left pulmonary arterial blood flow, measured at birth on day 14 after surgery, using ultrasonic flow transducer, was not statistically different from the one measured by Doppler on day 14. Our data demonstrate that echocardiography allows accurate assessment of pulmonary arterial blood flow in utero, providing evidence suggesting transient high pulmonary blood flow that did not last >6 d after the creation of a systemic fistula.

Systemic arteriovenous fistula leads to pulmonary artery remodeling and abnormal vasoreactivity in the fetal lamb.

Several cases of systemic arteriovenous fistula diagnosed in the human fetus have been associated with the postnatal development of persistent pulmonary hypertension. The aim of this study was to determine the effects of a prenatally created systemic arteriovenous fistula on the structure and reactivity of the pulmonary circulation in the fetal lamb. A fistula between the jugular vein and carotid artery was created in fetal lambs at 119-124 days of gestation. At delivery (134-139 days), left pulmonary artery (LPA) pressure was increased in the fistula group (n = 12) compared with controls (n = 11, P < 0.01). The pulmonary vascular resistance was significantly higher in the fistula group (P < 0.05), whereas mean LPA blood flow was not statistically different between the two groups. Morphometric analysis of the pulmonary vascular bed revealed an increase in the number of peripheral muscular arteries, together with an increase in pulmonary arterial medial thickness in the fistula group. There was no difference in the relative number or size of intraacinar arteries. In vitro organ bath studies on pulmonary arterial rings showed impaired endothelium-dependent relaxation in the fistula group compared with controls. However, endothelial nitric oxide synthase protein expression was similar in both groups, whereas endothelium-independent relaxation to sodium nitroprusside was greater in the fistula group compared with controls. A systemic arteriovenous fistula leads to both structural and functional alteration of the pulmonary vasculature, which might lead to the development of persistent pulmonary hypertension after birth.

Involvement of Ca2+/calmodulin-dependent protein kinase II in endothelial NO production and endothelium-dependent relaxation.

Nitric oxide (NO) is synthesized from l-arginine by the Ca(2+)/calmodulin-sensitive endothelial NO synthase (NOS) isoform (eNOS). The present study assesses the role of Ca(2+)/calmodulin-dependent protein kinase II (CaMK II) in endothelium-dependent relaxation and NO synthesis. The effects of three CaMK II inhibitors were investigated in endothelium-intact aortic rings of normotensive rats. NO synthesis was assessed by a NO sensor and chemiluminescence in culture medium of cultured porcine aortic endothelial cells stimulated with the Ca(2+) ionophore A23187 and thapsigargin. Rat aortic endothelial NOS activity was measured by the conversion of l-[(3)H]arginine to l-[(3)H]citrulline. Three CaMK II inhibitors, polypeptide 281-302, KN-93, and lavendustin C, attenuated the endothelium-dependent relaxation of endothelium-intact rat aortic rings in response to acetylcholine, A23187, and thapsigargin. None of the CaMK II inhibitors affected the relaxation induced by NO donors. In a porcine aortic endothelial cell line, KN-93 decreased NO synthesis and caused a rightward shift of the concentration-response curves to A23187 and thapsigargin. In rat aortic endothelial cells, KN-93 significantly decreased bradykinin-induced eNOS activity. These results suggest that CaMK II was involved in NO synthesis as a result of Ca(2+)-dependent activation of eNOS.

Altered guanylyl-cyclase activity in vitro of pulmonary arteries from fetal lambs with congenital diaphragmatic hernia.

Nitric oxide (NO) plays a major role in the modulation of perinatal pulmonary vascular tone. Congenital diaphragmatic hernia (CDH), a major cause of severe persistent pulmonary hypertension of the newborn (PPHN), is often refractory to inhaled NO. Alterations in NO/cyclic guanosine 3',5' monophosphate (cGMP)-mediated pulmonary vasodilatation may contribute to PPHN in CDH. We assessed NO/cGMP-mediated pulmonary vasorelaxation in vitro in 140-d gestational lamb fetuses with surgically created left CDH (term = 147 d) to age-matched controls. Relaxation of fourth generation intralobar pulmonary artery rings in response to the endothelium-dependent vasodilator, acetylcholine (ACh), and to the specific inhibitor of cGMP-phosphodiesterase (PDE), zaprinast, did not differ between the two groups. By contrast, relaxation in response to the calcium ionophore A23187 was impaired in CDH as compared with control animals. Relaxation in response to the NO donor sodium nitroprusside (SNP) (a direct activator of soluble guanylyl cyclase [sGC]) was also impaired in CDH animals as compared with controls. Repeating the challenge increased vasorelaxation in response to SNP in CDH as compared with control animals. Immunohistochemistry revealed the presence of endothelial NO-synthase in the endothelium of pulmonary arteries from both control and CDH animals. We conclude that endothelium-dependent vasodilatation in response to ACh and A23187 was differently affected in the fetal surgical CDH-lamb model. Furthermore, activity of sGC but not that of PDE was impaired in CDH animals. PPHN and decreased inhaled NO responsiveness in CDH may involve decreased sGC activity.

Involvement of Na(+)/Ca(2+) exchanger in endothelial NO production and endothelium-dependent relaxation.

Endothelial nitric oxide (NO) synthase (eNOS) is controlled by Ca(2+)/calmodulin and caveolin-1 in caveolae. It has been recently suggested that Na(+)/Ca(2+) exchanger (NCX), also expressed in endothelial caveolae, is involved in eNOS activation. To investigate the role played by NCX in NO synthesis, we assessed the effects of Na(+) loading (induced by monensin) on rat aortic rings and cultured porcine aortic endothelial cells. Effect of monensin was evaluated by endothelium-dependent relaxation of rat aortic rings in response to acetylcholine and by real-time measurement of NO release from cultured endothelial cells stimulated by A-23187 and bradykinin. Na(+) loading shifted the acetylcholine concentration-response curve to the left. These effects were prevented by pretreatment with the NCX inhibitors benzamil and KB-R7943. Monensin potentiated Ca(2+)-dependent NO release in cultured cells, whereas benzamil and KB-R7943 totally blocked Na(+) loading-induced NO release. These findings confirm the key role of NCX in reverse mode on Ca(2+)-dependent NO production and endothelium-dependent relaxation.

Successful cardiopulmonary resuscitation after cardiac arrest as a "sepsis-like" syndrome.

BACKGROUND: We investigated the immunoinflammatory profile of patients successfully resuscitated after cardiac arrest, representing a model of whole-body ischemia/reperfusion syndrome. METHODS AND RESULTS: Plasma cytokine, endotoxin, and ex vivo cytokine production in whole-blood assays was assessed in 61, 35, and 11 patients, respectively. On admission, high levels of plasma interleukin (IL)-6, IL-8, IL-10, and soluble tumor necrosis factor (TNF) receptor type II could discriminate between survivors and nonsurvivors. Among nonsurvivors, the initial need for a vasopressor agent was associated with higher levels of IL-1 receptor antagonist, IL-10, and IL-6 on day 1. Plasma endotoxin was detected in 46% of the analyzed patients within the 2 first days. Endotoxin-induced TNF and IL-6 productions were dramatically impaired in these patients compared with healthy control subjects, whereas an unaltered production was observed with heat-killed Staphylococcus aureus. In contrast, IL-1 receptor antagonist productions were enhanced in these patients compared with healthy control subjects. The productions of T-cell-derived IL-10 and interferon-gamma were also impaired in these patients. Finally, using in vitro plasma exchange between healthy control subjects and patients, we demonstrated that the endotoxin-dependent hyporeactivity was an intrinsic property of patients' leukocytes and that an immunosuppressive activity was also present in their plasma. CONCLUSIONS: Altogether, the high levels of circulating cytokines, the presence of endotoxin in plasma, and the dysregulated production of cytokines found in these patients recall the immunological profile found in patients with sepsis.