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Published ahead of print on December 30, 2003, doi:10.1164/rccm.200308-1154OC
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American Journal of Respiratory and Critical Care Medicine Vol 169. pp. 829-835, (2004)
© 2004 American Thoracic Society

Original Article

Altered Pulmonary Vascular Reactivity in Mice with Excessive Erythrocytosis

Jo Hasegawa, Klaus F. Wagner, Dörte Karp, Dechun Li, Junpei Shibata, Matthias Heringlake, Ludger Bahlmann, Reinhard Depping, Joachim Fandrey, Peter Schmucker and Stefan Uhlig

Department of Anesthesiology and Critical Care Medicine; Department of Physiology, University of Lübeck, Lübeck; Department of Physiology, University of Essen, Essen; and Division of Pulmonary Pharmacology, Research Center Borstel, Borstel, Germany

Correspondence and requests for reprints should be addressed to Stefan Uhlig, Ph.D., Division of Pulmonary Pharmacology, Research Center Borstel, Center for Medicine and Biosciences, Parkallee 22, D-23845 Borstel, Germany. E-mail: suhlig{at}fz-borstel.de


   ABSTRACT
TOP
 ABSTRACT
METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Pulmonary vascular remodeling during chronic hypoxia may be the result of either oxygen deprivation or erythrocytosis. To separate experimentally the effects of hypoxia and erythrocytosis, we analyzed transgenic mice that constitutively overexpress the human erythropoietin gene in an oxygen-independent manner. These mice are characterized by polycythemia but have normal blood pressure, heart rate, and cardiac output. In transgenic mice, pulmonary artery pressure (PAP) was increased in vivo but was reduced in blood-free perfused lungs. The thromboxane receptor agonist U46619 caused a smaller rise in PAP in isolated transgenic lungs than in lungs from wild-type mice. The transgenic pulmonary vasculature was characterized by elevated prostacyclin production, stronger endothelial nitric oxide synthase expression, and reduced pulmonary vascular smooth muscle thickness. The fact that transgenic polycythemic mice have marked pulmonary hypertension in vivo but not in vitro suggests that their pulmonary hypertension is due to the increased blood viscosity, thus supporting an independent role of polycythemia in the development of pulmonary hypertension. In addition, our findings indicate that the lungs of transgenic animals adapt to the high PAP by elevated synthesis of vasodilators and reduced vascular smooth muscle thickness that tend to reduce vascular tone and vascular responsiveness.

Key Words: pulmonary hypertension • erythropoietin • isolated mouse lung • vascular remodeling

Patients with chronic pulmonary disease bear an increased perioperative risk. Apart from the inherent oxygenation problems and the circulatory fragility, the intraoperative management of these patients is complicated by the more profound anesthesia-associated increase in ventilation-perfusion mismatch (1). In addition, the vasoconstrictor response to acute hypoxia is decreased (2).

In hypoxic chronic pulmonary disease, pulmonary hypertension frequently develops. The hallmarks of pulmonary hypertension under these conditions are pulmonary vascular remodeling with media hypertrophy of arteries, polycythemia, altered reactivity of the pulmonary vasculature, and subsequently right ventricular hypertrophy (3). The exact mechanisms of the pulmonary hypertension are incompletely understood, and a variety of factors, including polycythemia, hypoxia, vascular remodeling, and nitric oxide (NO), have been implicated in the disease process. Because in chronic pulmonary disease both hypoxemia and polycythemia are simultaneously present, their individual effects on the pulmonary circulation have been difficult to define.

Recently, a mouse model of an isolated erythrocytosis was generated by establishing a transgenic mouse line constitutively overexpressing the human erythropoietin gene in an oxygen-independent manner (4). These mice reach a hematocrit plateau of 0.80–0.85 within the first 2 months without altering their blood pressure, heart rate, and cardiac output (5). The transgenic mice show heart abnormalities and have dramatically reduced exercise performance and a significantly reduced life span of only 7.5 months. However, blood pressure and cardiac output in 4-month-old transgenic animals are normal, probably as result of enhanced NO production by the endothelial NO synthase (eNOS). Increased eNOS expression was noted in the endothelium of transgenic aorta and pulmonary arteries, and feeding transgenic mice an NO synthase inhibitor in their drinking water resulted in deaths within 3 days (4).

Other than the increased expression of eNOS in the pulmonary artery, little is known about the effect of erythrocytosis on the pulmonary circulation and the lungs in these transgenic mice. Such information would extend our understanding of the pulmonary changes occurring in hypoxic chronic obstructive pulmonary disease and in high-altitude disease. Under these conditions, hypoxia and erythrocytosis are always linked, making it difficult to identify which of these two is causing the pulmonary artery remodeling. The transgenic mice offer the unique opportunity to investigate the pulmonary consequences of erythrocytosis under normoxic conditions. Therefore, in this study, we histologically and functionally characterized the pulmonary circulation of 4-month-old mice with excessive erythrocytosis. Some of the results of these studies have been previously reported in the form of an abstract (6).


   METHODS
TOP
 ABSTRACT
 METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Animals
Four-month-old mice were studied (mouse line tg6) that overexpressed the human erythropoietin gene (4), which resulted in a 10-fold increased plasma erythropoietin concentration (data not shown). For organ weight determination, the animals were perfused free of blood with a total of 50 ml of 0.9% saline via an aortic and a vena cava cannula; the organs were excised, and the weight was measured immediately (wet weight) and after heat drying at 60°C for 5 days (dry weight).

In Vivo Pulmonary Artery Pressure Measurement
Animals were tracheotomized and ventilated with oxygen-enriched air (Minivent Type 845; Hugo Sachs Elektronik, March, Germany) without positive end-expiratory airway pressure (130 and 180 breaths/minute; VT, 180–230 µl). After median sternotomy, the pulmonary artery was punctured with a with a 22-gauge needle to measure the pulmonary artery pressure (PAP).

Isolated Perfused Mouse Lung Preparation
The isolated mouse lungs were ventilated by negative pressure (-3 to -9 cm H2O, 90 breaths · minute-1, VT, 200 µl) and perfused in a nonrecirculating fashion with RPMIRoswell Park Memorial Institute medium containing 4% albumin at a constant flow of 1 ml · min-1, resulting in 0–5 cm H2O PAP (7, 8). Although for technical reasons (minimize edema formation, reduce costs of perfusate) the chosen perfusate flow rate was only approximately a 10th of the physiologic flow rate, relative comparisons between different mouse strains are still feasible under these conditions (9).

Histology
Lungs were fixed by perfusion with 4% neutral-buffered formalin, processed routinely, and embedded in paraffin. Serial sections (2 µm) were stained with hematoxylin-eosin, Masson's trichrome, and van Gieson's elastica stain (10).

Immunohistochemistry
Two-micrometer lung sections were deparaffinized with xylene, progressively rehydrated in decreasing percentages of ethanol, and immersed in 0.3% hydrogen peroxide in methanol. Antigen sites were retrieved by heating the sections on slides in 0.01-M sodium citrate in a microwave oven and cooling for 30 minutes to room temperature. eNOS protein was immunolocalized with a rabbit polyclonal primary antibody and visualized with the PAP technique (11). For morphometric analysis of the muscularization of the pulmonary vessels, sections were immunostained for smooth muscle {alpha}-actin and examined by digital imaging analysis (12).

Morphometric Analysis
The morphometric analysis was performed in smooth muscle {alpha}-actin immunostained slides with the investigator blinded for the genotype of the material measured. The media thickness of 50 fully muscularized small pulmonary vessels (70–100 µm) per lung was measured, and the mean value of the total of 5 x 50 single-thickness measurements was calculated. Fully muscularized vessels had a continuous smooth muscle layer. Media thickness was measured perpendicular to the circumference of the vessel as the diameter of the smooth muscle layer. The thickness of the pulmonary artery wall was measured from the endothelium to the outer edge of the smooth muscle layer. It was measured at five different regions of the circumference of the main pulmonary artery.

Statistical Analysis
Single time point data were tested for significance of differences using the unpaired t test. Data for multiple comparisons were analyzed by one-way analysis of variance followed by Bonferroni's post hoc test. Differences were considered significant at p < 0.05. All data are shown as means ± SD unless otherwise specified.


   RESULTS
TOP
 ABSTRACT
 METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
Histology
On inspection, the lungs of transgenic mice showed a prominent purple color, which was in sharp contrast to the normal light pink of the lungs of wild-type mice. Subsequently, microscopy was employed for morphologic assessment. In transgenic mice, the alveolar capillaries, arterioles, and arteries were hyperemic and congested with erythrocytes (Figures 1B and 1D), and alveolar septi appeared thickened. In line with these findings, the lung wet weight of transgenic animals was increased (wild type 168.4 ± 15.7 mg vs. transgenic 209.4 ± 20.8 mg, p = 0.001, n = 7). This significant difference persisted when lung dry weight was measured (Figure 1E), indicating that lung mass and not tissue fluid content was increased.



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  		Figure 1. A plethora of erythrocytes within the vasculature is the histologic hallmark of the lung morphology in erythropoietin transgenic mice with excessive erythrocytosis (B and D). For comparison, the corresponding wild-type sections are shown in A and C. Lung dry weight (E) was significantly increased in transgenic mice. In vivo measurement of pulmonary artery pressure (PAP) (F) demonstrated pulmonary hypertension in transgenic mice with excessive erythrocytosis. Microscopic slides are stained with Masson's trichrome. (A and B) x100, scale 50 µm. (C and D) x400, scale 10 µm; wt = wild type (n = 5–8); tg = transgenic (n = 5–8). *p < 0.05 and #p < 0.001 compared with wild-type control subjects.

 
We further analyzed the thickness of the pulmonary arterial vessels. For small pulmonary arteries (70–100 µm), the media thickness was significantly reduced in transgenic mice (Figure 2C). In addition, the wall of the main pulmonary artery in erythropoietin transgenic mice was significantly thinner compared with wild-type control subjects (Figure 2D).



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  		Figure 2. Erythropoietin transgenic mice with excessive erythrocytosis showed a significant reduction of media thickness in small pulmonary arteries (A, wild type; B, transgenic). The mean media thickness of small pulmonary arteries (C) (n = 250 determinations, lungs from five wild-type and five transgenic animals) as well as the mean wall thickness of the main pulmonary artery (D) (n = 25 determinations, lungs from five wild-type and five transgenic animals) was significantly reduced (p < 0.001) in transgenic animals compared with wild-type control subjects. (A and B) Immunohistochemistry for {alpha} smooth muscle actin, chromogen fast-red, x400, scale 10 µm. #p < 0.001 compared with wild-type control subject.

 
Investigating the pulmonary expression of NO synthase immunohistochemistry revealed an increased eNOS protein expression in endothelial cells of pulmonary arteries in erythropoietin transgenic mice (Figure 3).



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  		Figure 3. Expression of endothelial NO synthase (eNOS) in pulmonary vessels. Immunostaining revealed an increased expression of eNOS within the vascular endothelium of pulmonary vessels of erythropoietin transgenic mice compared with wild-type mice. (A) Wild type. (B) Transgenic, chromogen DAB, x100, scale 50 µm.

 
PAP In Vivo
Transgenic mice had a significantly increased hematocrit (wild type 0.45 ± 0.01 vs. transgenic 0.86 ± 0.03, p < 0.001). Because hematocrit is a major determinant of blood viscosity, it was assumed that pulmonary vascular resistance is increased in transgenic mice. In keeping with this, PAP was markedly increased in transgenic mice in vivo (Figure 1F).

Isolated Perfused Lungs
To investigate whether the increased pressure was a result of the higher blood viscosity or due to vascular remodeling, we measured PAP in isolated lungs perfused with blood-free salt buffer at a low flow rate of 1 ml/minute. In contrast to the in vivo situation, PAP was significantly lower in transgenic mice than in control animals (Table 1), indicating a lower tone in pulmonary resistance vessels. Note that in our model perfusion pressure may become negative with respect to the ambient pressure because the lungs are ventilated by negative pressure, which is added to the pressure of the venous effluent buffer. Such a setup is recommended to provide physiologic transmural pressures in the lungs (13–15). Perfusate levels of 6-keto-PGF1{alpha}, the stable metabolite of the vasodilator prostacyclin, were higher in mice from transgenic lungs (Table 1). Unfortunately, NO could not be measured in our samples because of the low sensitivity of currently available NO assays. Baseline pulmonary compliance and airway resistance did not differ between the two strains (Table 1).


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  		TABLE 1. Isolated perfused lung baseline data of wild-type and transgenic mice

 
Responsiveness to the Thromboxane Mimetic U46619
We next compared the responsiveness of airways and pulmonary vessels between transgenic and control mice. Because baseline PAP differed between the two strains (Table 1), the data are expressed as the relative increase in perfusion pressure. Injection of the thromboxane receptor agonist U46619 (10 nM) increased PAP and airway resistance in lungs from both strains. The increase in PAP in control mice was two to three times stronger than in transgenic mice (Figure 4A). There was no difference in the airway responses of both strains (Figure 4B).



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  		Figure 4. Time course of vascular (upper panel) and airway responses (lower panel) to 10 nM U46619. Lungs were perfused and ventilated for 30 minutes without any treatment to obtain a baseline. After 30 minutes, lungs were perfused with 10 nM U46619 for another 30 minutes. Shown are the changes in PAP (upper panel) and in airway resistance (lower panel). (Circles) Wild-type animals (n = 6). (Squares) Transgenic animals (n = 5). Data are mean ± SEM. Based on the measurement after 30 minutes of treatment with U46619, the increase in PAP in wild-type lungs was significantly (p < 0.001) higher than in transgenic lungs.

 
To investigate the hypothesis that increased production of prostacyclin and NO in transgenic mice masks the U46619-induced vasoconstriction, lungs were pretreated with indomethacin to block cyclooxygenase, with N(G)-nitro-L-arginine methyl ester (L-NAME) to block NO synthase, or with both antagonists simultaneously. The addition of indomethacin and/or L-NAME did not affect the baseline PAP (data not shown). In transgenic mice, pretreatment with indomethacin had a small effect on the U46619-induced vasoconstriction (Figure 5). In contrast, in the presence of L-NAME or L-NAME/indomethacin, the vascular response in the transgenic mice to U46619 was markedly enhanced (Figure 5). In another set of experiments, the simultaneous pretreatment with L-NAME/indomethacin increased the vascular response to U46619 by nearly the same degree as it did in the transgenic animals (Figure 6).



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  		Figure 5. Time course of vascular responses in transgenic animals to 10 nM U46619. Lungs were perfused and ventilated for 20 minutes without any treatment to obtain a baseline. N(G)-nitro-L-arginine methyl ester (L-NAME) (100 µM) or indomethacin (10 µM) was added after 20 minutes of perfusion and 10 nM U46619 after 30 minutes. (Open squares) U46619 alone (n = 5). (Closed circles) Indomethacin/U46619 (n = 3). (Closed squares) L-NAME/U46619 (n = 3). (Closed diamonds) Indomethacin/L-NAME/U46619 (n = 3). Data are mean ± SEM. Based on the measurement after 30 minutes of treatment with U46619, the increase in PAP in lungs pretreated with L-NAME or indomethacin/L-NAME was significantly (p < 0.01) higher than in lungs perfused with indomethacin or without any inhibitor.

 


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  		Figure 6. Time course of vascular responses in wild-type animals to 10 nM U46619. Lungs were perfused and ventilated for 20 minutes without any treatment to obtain a baseline. L-NAME (100 µM) and indomethacin (10 µM) were added after 20 minutes of perfusion and 10 nM U46619 after 30 minutes. (Open circles) U46619 alone (n = 6). (Closed triangles) Indomethacin/L-NAME/U46619 (n = 3). Data are mean ± SEM. Based on the measurement after 30 minutes of treatment with U46619, the increase in PAP in lungs pretreated with indomethacin/L-NAME was significantly (p < 0.001) higher than in lungs perfused with U46619 alone.

 

   DISCUSSION
TOP
 ABSTRACT
 METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
We investigated the properties of the pulmonary vasculature in mice overexpressing the human erythropoietin gene. Our data suggest that in these mice the increased hematocrit and hence blood viscosity lead to the pulmonary hypertension in vivo. At the same time, however, these animals are characterized by a reduced thickness of the pulmonary artery wall and enhanced production of vasodilators such as NO and prostacyclin. These alterations may be interpreted as a countermeasure of the lungs to lower the elevated PAP. And in fact, in the absence of blood the vascular tone was lower in transgenic animals as compared with control mice. The functional consequences of the reduced thickness of the pulmonary artery became also evident by the observation that the vessels of transgenic animals responded weaker to the vasopressor action of U46619, a thromboxane receptor agonist. Thus, the lungs appear to adapt to chronically increased pulmonary hypertension in at least three ways: reduced vascular wall thickness, increased production of vasodilators, and attenuated responses to vasoconstrictors.

Altered vascular responses in the lungs of polycythemic individuals have been noted before, such as a reduced hypoxic pulmonary vasoconstriction (16) or impaired vasodilator responses to acetylcholine (17). Among other things, the transgenic mice are characterized by a decrease in pulmonary vascular smooth muscle thickness (Figure 2) and an increase in NO synthase protein expression (Figure 3), NO production (4, 5, 18), and blood viscosity (19). Because increased blood viscosity elevates vascular shear stress, which is known to stimulate the production of endothelial vasodilators such as prostacyclin and NO, it is tempting to hypothesize that the reduced basal pulmonary vascular tone in transgenic mice is a consequence of the elevated blood viscosity. On the other side, in cell culture experiments, a stimulatory effect of erythropoietin on NO production has been shown (20, 21); therefore, a direct action of erythropoietin as cause for the reduced basal pulmonary vascular tone cannot be excluded. However, cell culture studies have also demonstrated a mitogenic effect of erythropoietin on vascular smooth muscle cells (22), which is in contrast to the diminished pulmonary artery smooth muscle in transgenic mice. In addition, the fact that the changes in transgenic mice were confined to the vasculature lends further support to our conclusion that increased blood viscosity rather than direct effects of erythropoietin caused the vascular alterations because otherwise similar changes might be expected in the airways' tone and responsiveness. Our proposition that the vascular changes are a consequence of polycythemia rather than of erythropoietin itself are further supported by the observation that administration of erythropoietin to hypoxic polycythemic rats did not worsen the pulmonary hypertension (16).

In perfused lungs of transgenic mice, the responses to thromboxane were reduced, and this hyporesponsiveness was not altered by simultaneous blockade of prostacyclin and NO synthesis. Thus, although these inhibitors exaggerated the response to U46619, this effect was not stronger in transgenic animals than in control mice. In addition, these inhibitors had no effect on the basal vascular tone. These findings suggest that the major determinants of the reduced vascular tone and responsiveness are not vasodilators such as NO or prostacyclin, but rather the diminution of the pulmonary artery smooth muscle. However, we cannot exclude the possibility that the effect of these mediators is more pronounced at higher perfusate flow rates. It also appears possible that the observed anatomic changes are related to mediators such as prostacyclin or NO. Interestingly, with regard to the physiologic role of NO under these conditions, opposing results were obtained when NO inhibitors were administered to erythropoietin (EPO) transgenic mice or to chronic hypoxic rats: In the former case, NO synthase inhibition caused death in the transgenic animals within three days (4), whereas in the hypoxic animals, NO inhibition ameliorated the pulmonary changes (23).

Our study resembles in some aspects that by Walker and colleagues, who investigated the effect of a 2-week treatment with EPO in rats (24). Also in that study, a lesser vasoconstrictor response to U46619 was observed, although this was not as pronounced as reported here. Otherwise, however, the results from that study substantially differ from the results reported here. The 2-week treatment of rats with EPO resulted in a hematocrit of approximately 65 compared with a hematocrit of approximately 80 in the transgenic mice. Thus, in their model, erythrocytosis was less pronounced and present for a shorter period of time. Under these conditions, expression of eNOS, NO production, and PAP were all normal, and right ventricular hypertrophy was not observed. Interestingly, however, systemic blood pressure was increased. These findings are in contrast to the observations made in the transgenic mice and thus suggest that the changes in eNOS and PAP require elevated hematocrit levels for a longer time.

The decreased thickness of the pulmonary artery wall does well explain the decreased tone and responsiveness of the pulmonary vessels of transgenic animals. However, these findings should also be discussed against the background of studies showing increased media thickness of muscular pulmonary arteries in chronic hypoxic rats (25, 26). As media thickening was not observed in the transgenic mice, one obvious explanation for the discrepancy to hypoxic animals would be that the increased media thickness results from the hypoxia and not from the polycythemia; yet media thickening appears unlikely as a determinant of the increased pulmonary vascular pressure in hypoxic animals, as the angiotensin-converting enzyme inhibitor cilazapril completely prevented the thickening of the media of pulmonary arteries in chronic hypoxic rats without decreasing PAP or right ventricular weight (27). More recent studies even suggest that media thickening may not be a prominent feature of hypoxia after all. A careful analysis of the media of the pulmonary artery of hypoxic rats revealed only a nonsignificant trend of media thickening and explained previous stronger findings by an artifact caused by active hypoxic pulmonary vasoconstriction (28). In addition, in rats exposed to low oxygen levels corresponding to 5,500 m sea level, the external diameter of various arteries was reduced compared with rats exposed to hypoxia corresponding to only 4,000 m (29).

The EPO transgenic mice (4, 5, 30) allowed us to study the effect of isolated polycythemia on the pulmonary circulation independent from coexisting hypoxia. The observed pathologic alterations, in particular the increased PAP in conjunction with the diminished pulmonary artery thickness and reduced vascular responsiveness, may be of relevance for a number of clinical conditions where elevated hematocrit values ranging from 0.65 to 0.91 have been reported, such as postrenal transplant polycythemia (31), polycythemia of the newborn (32), congenital cyanotic heart disease (33), and chronic mountain sickness (34, 35), not to mention the etiologically distinct polycythemia vera (36). Secondary polycythemia results from tissue hypoxia, and under these conditions, it is difficult to define which cardiopulmonary consequences are caused by chronic polycythemia and which by hypoxia. The coexistence of polycythemia in patients with chronic pulmonary disease has been reported to increase mortality significantly (37). In patients with chronic hypoxemic lung disease, polycythemia has been shown to impair the vasodilator response to acetylcholine (17). Therefore, polycythemia appears to add significantly to the overall morbidity and mortality of the underlying chronic pulmonary disease. Therapeutic effects that could be explained by a reduction in polycythemia are the successful treatment of excessive erythrocytosis (1) in chronic mountain sickness by phlebotomy or relocation to sea level (38) and (2) after renal transplantation or exposure to high altitude with angiotensin-converting enzyme inhibitors that reduce hematocrit, blood pressure, and proteinuria (39, 40).

Taken together, transgenic polycythemic mice show marked pulmonary hypertension in vivo but pulmonary hypotension in vitro. These findings suggest that the increased pulmonary hypertension is due to the increased blood viscosity, thus supporting an independent role of polycythemia in the development of pulmonary hypertension. The clinical relevance of our further observations in transgenic mice, in particular the diminished pulmonary artery smooth muscle and the attenuated vascular responsiveness, is unknown at present and deserves further study.


   Acknowledgments
 
The perfect technical assistance of Ann-Katrin Hellberg is gratefully acknowledged. The authors wish to thank M. Gassmann for the generous gift of the tg6 mouse line and W. Jelkmann, M. Gassmann, and R. Wenger for helpful discussions.


   FOOTNOTES
 
Supported by the Department of Anesthesiology and Resuscitation, Shinshu University, and in part by the Deutsche Forschungsgemeinschaft grant Uh 88/2–4 (S.U.) and a grant from the Max Kade Foundation (K.F.W.).

J.H. and K.W. contributed equally to this study.

This article has an online supplement, which is accessible from this issue's table of contents online at www.atsjournals.org

Conflict of Interest Statement: J.H. has no declared conflict of interest; K.F.W. has no declared conflict of interest; D.K. has no declared conflict of interest; D.L. has no declared conflict of interest; J.S. has no declared conflict of interest; M.H. has no declared conflict of interest; L.B. has no declared conflict of interest; R.D. has no declared conflict of interest; J.F. has no declared conflict of interest; P.S. has no declared conflict of interest; S.U. has no declared conflict of interest.

Received in original form August 19, 2003; accepted in final form December 27, 2003


   REFERENCES
TOP
 ABSTRACT
 METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 

  1. Seigne PW, Hartigan PM, Body SC. Anesthetic considerations for patients with severe emphysematous lung disease. Int Anesthesiol Clin 2000;38:1–23.
  2. Peinado VI, Santos S, Ramirez J, Roca J, Rodriguez-Roisin R, Barbera JA. Response to hypoxia of pulmonary arteries in chronic obstructive pulmonary disease: an in vitro study. Eur Respir J 2002;20:332–338.[Abstract/Free Full Text]
  3. Meyrick B. The pathology of pulmonary artery hypertension. Clin Chest Med 2001;22:393–404.[CrossRef][Medline]
  4. Ruschitzka FT, Wenger RH, Stallmach T, Quaschning T, de Wit C, Wagner K, Labugger R, Kelm M, Noll G, Rulicke T, et al. Nitric oxide prevents cardiovascular disease and determines survival in polyglobulic mice overexpressing erythropoietin. Proc Natl Acad Sci USA 2000;97:11609–11613.[Abstract/Free Full Text]
  5. Wagner KF, Katschinski DM, Hasegawa J, Schumacher D, Meller B, Gembruch U, Schramm U, Jelkmann W, Gassmann M, Fandrey J. Chronic inborn erythrocytosis leads to cardiac dysfunction and premature death in mice overexpressing erythropoietin. Blood 2001;97:536–542.[Abstract/Free Full Text]
  6. Wagner KF, Hasegawa J, Shibata J, Li D, Heringlake M, Karp D, Uhlig S. Altered pulmonary vascular reactivity in transgenic mice with excessive erythrocytosis [abstract]. Am J Respir Crit Care Med 2003;167:A698.
  7. von Bethmann AN, Brasch F, Nüsing R, Vogt K, Volk HD, Müller KM, Wendel A, Uhlig S. Hyperventilation induces release of cytokines from perfused mouse lung. Am J Respir Crit Care Med 1998;157:263–272.
  8. Held HD, Martin C, Uhlig S. Characterization of airway and vascular responses in murine lungs. Br J Pharmacol 1999;126:1191–1199.[CrossRef][Medline]
  9. Held HD, Uhlig S. Basal lung mechanics and airway and pulmonary vascular responsiveness in different inbred mouse strains. J Appl Physiol 2000;88:2192–2198.[Abstract/Free Full Text]
  10. Romeis B. Mikroskopische Technik, 17th ed. Munich: Urban & Schwarzenberg; 1989.
  11. Bancroft JD, Stevens A. Theory and practice of histological techniques, 3rd ed. Edinburgh, London, Melbourne, New York: Churchill Livingstone; 1990.
  12. Quinlan TR, Li D, Laubach VE, Shesely EG, Zhou N, Johns RA. eNOS-deficient mice show reduced pulmonary vascular proliferation and remodeling to chronic hypoxia. Am J Physiol Lung Cell Mol Physiol 2000;279:L641–L650.[Abstract/Free Full Text]
  13. Permutt B. Mechanical influences on water accumulation in the lung. In: Fishman A, Renkin EMJ, editors. Pulmonary edema. Bethesda, MD: American Physiological Society; 1979. p. 175–194.
  14. Culver BH, Butler J. Mechanical influences on the pulmonary microcirculation. Annu Rev Physiol 1980;42:187–198.[CrossRef][Medline]
  15. Dritsas KG, Brown D, Couves CM. Effect of alveolar pleural pressures on interstitial pressures in isolated dog lungs. J Appl Physiol 1969;65:611–616.
  16. Petit RD, Warburton RR, Ou LC, Hill NS. Pulmonary vascular adaptations to augmented polycythemia during chronic hypoxia. J Appl Physiol 1995;79:229–235.[Abstract/Free Full Text]
  17. Defouilloy C, Teiger E, Sediame S, Andrivet P, Roudot-Thoraval F, Chouaid C, Housset B, Adnot S. Polycythemia impairs vasodilator response to acetylcholine in patients with chronic hypoxemic lung disease. Am J Respir Crit Care Med 1998;157:1452–1460.
  18. Shibata J, Hasegawa J, Siemens HJ, Wolber E, Dibbelt L, Li D, Katschinski DM, Fandrey J, Jelkmann W, Gassmann M, et al. Hemostasis and coagulation at a hematocrit of 0.85: functional consequences of erythrocytosis. Blood 2003;101:4416–4422.[Abstract/Free Full Text]
  19. Vogel J, Kießling I, Heinicke K, Stallmach T, Ossent P, Vogel O, Aulmann M, Frietsch T, Schmid-Schonbein H, Kuschinsky W, et al. Transgenic mice overexpressing erythropoietin adapt to excessive erythrocytosis by regulating blood viscosity. Blood 2003;102:2278–2284.[Abstract/Free Full Text]
  20. Wilcox CS, Deng X, Doll AH, Snellen H, Welch WJ. Nitric oxide mediates renal vasodilation during erythropoietin-induced polycythemia. Kidney Int 1993;44:430–435.[Medline]
  21. Banerjee D, Rodriguez M, Nag M, Adamson JW. Exposure of endothelial cells to recombinant human erythropoietin induces nitric oxide synthase activity. Kidney Int 2000;57:1895–1904.[CrossRef][Medline]
  22. Ito C, Kusano E, Furukawa Y, Yamamoto H, Takeda S, Akimoto T, Rimura O, Ando Y, Asano Y. Modulation of the erythropoietin-induced proliferative pathway by cAMP in vascular smooth muscle cells. Am J Physiol Cell Physiol 2002;283:C1715–C1721.[Abstract/Free Full Text]
  23. Mitani Y, Maruyama K, Sakurai M. Prolonged administration of L-arginine ameliorates chronic pulmonary hypertension and pulmonary vascular remodeling in rats. Circulation 1997;96:689–697.[Abstract/Free Full Text]
  24. Walker BR, Resta TC, Nelin LD. Nitric oxide-dependent pulmonary vasodilation in polycythemic rats. Am J Physiol Heart Circ Physiol 2000;279:H2382–H2389.[Abstract/Free Full Text]
  25. Rabinovitch M, Gamble W, Nadas AS, Miettinen OS, Reid L. Rat pulmonary circulation after chronic hypoxia: hemodynamic and structural features. Am J Physiol 1979;236:H818–H827.
  26. Meyrick B, Reid L. Hypoxia-induced structural changes in the media and adventitia of rat hilar pulmonary artery and their regression. Am J Pathol 1980;100:151–178.[Abstract]
  27. Clozel JP, Saunier C, Hartemann D, Fischli W. Effects of cilazapril, a novel angiotensin converting enzyme inhibitor, on the structure of pulmonary arteries of rats exposed to chronic hypoxia. J Cardiovasc Pharmacol 1991;17:36–40.[CrossRef][Medline]
  28. van Suylen RJ, Smits JF, Daemen MJ. Pulmonary artery remodeling differs in hypoxia- and monocrotaline-induced pulmonary hypertension. Am J Respir Crit Care Med 1998;157:1423–1428.
  29. Nakanishi K, Tajima F, Osada H, Nakamura A, Yagura S, Kawai T, Suzuki M, Torikata C. Pulmonary, vascular responses in rats exposed to chronic hypobaric hypoxia at two different altitude levels. Pathol Res Pract 1996;192:1057–1067.[Medline]
  30. Wiessner C, Allegrini PR, Ekatodramis D, Jewell UR, Stallmach T, Gassmann M. Increased cerebral infarct volumes in polyglobulic mice overexpressing erythropoietin. J Cereb Blood Flow Metab 2001;21:857–864.[Medline]
  31. Swales JD, Evans DB. Erythraemia in renal transplantation. BMJ 1969;2:80–83.
  32. Swetnam SM, Yabek SM, Alverson DC. Hemodynamic consequences of neonatal polycythemia. J Pediatr 1987;110:443–447.[CrossRef][Medline]
  33. Pearson TC. Rheology of the absolute polycythaemias. Baillieres Clin Haematol 1987;1:637–664.[CrossRef][Medline]
  34. Jefferson JA, Escudero E, Hurtado ME, Pando J, Tapia R, Swenson ER, Prchal J, Schreiner GF, Schoene RB, Hurtado A, et al. Excessive erythrocytosis, chronic mountain sickness, and serum cobalt levels. Lancet 2002;359:407–408.[CrossRef][Medline]
  35. Winslow R. High-altitude polycythemia. In: West JB, Lahiri S, editors. High altitude and man. Bethesda, MD: American Physiological Society; 1984.
  36. Murphy S. Polycythemia vera. Dis Mon 1992;38:153–212.[Medline]
  37. Renzetti AD Jr, McClement JH, Litt BD. The Veterans Administration cooperative study of pulmonary function: 3: mortality in relation to respiratory function in chronic obstructive pulmonary disease. Am J Med 1966;41:115–129.[CrossRef][Medline]
  38. Jefferson JA, Escudero E, Hurtado ME, Pando J, Tapia R, Swenson ER, Prchal J, Schreiner GF, Schoene RB, Hurtado A, et al. Excessive erythrocytosis, chronic mountain sickness, and serum cobalt levels. Lancet 2002;359:407–408.
  39. Remuzzi G, Perico N. Routine renin-angiotensin system blockade in renal transplantation? Curr Opin Nephrol Hypertens 2002;11:1–10.[CrossRef][Medline]
  40. Plata R, Cornejo A, Arratia C, Anabaya A, Perna A, Dimitrov BD, Remuzzi G, Ruggenenti P. Angiotensin-converting-enzyme inhibition therapy in altitude polycythemia: a prospective randomised trial. Lancet 2002;359:663–666.[CrossRef][Medline]



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