Monocrotaline pneumotoxicity in mice

Lung injury induced in rats by the pyrrolizidine alkaloid monocrotaline is a well-documented model of pulmonary hypertension. To our knowledge, however, monocrotaline-induced cardiopulmonary injury has rarely been described and has never been quantitated in mice. In the present study, adult male mice received 2.4, 4.8, or 24.0 mg monocrotaline/kg body weight/day in the drinking water continuously for 6 weeks. These doses represent 1, 2, and 10 times the severely pneumotoxic regimen in rats. Pulmonary endothelial function was monitored by right lung angiotensin converting enzyme (ACE) activity, plasminogen activator (PLA) activity, and prostacyclin (PGI2) and thromboxane (TXA2) production. Light and electron microscopy were performed on the left lungs. Cardiac right ventricular hypertrophy was evaluated by the right ventricle to left ventricle plus septum weight ratio (RV/LV + S). Monocrotaline-treated mice exhibited a dose-dependent decrease in lung ACE and PLA activities and an increase in PGI2 and TXA2 production, indicative of endothelial dysfunction. However, these responses were significant only after the highest monocrotaline dose. Light and electron microscopy revealed dose-dependent pulmonary inflammatory and exudative reactions. Unlike previous studies in rats, however, monocrotaline-treated mice developed relatively little lung fibrosis, cardiomegaly, or right ventricular hypertrophy, and no occlusive medial thickening of the pulmonary arteries, even at the highest dose level. These and previous data indicate that there are quantitative biochemical and qualitative morphological differences between mice and rats with respect to monocrotaline pneumotoxicity. Furthermore, in monocrotaline-treated mice (but not in rats) there appears to be a dissociation between lung endothelial dysfunction and inflammation on the one hand, and pulmonary hypertension and fibrosis on the other.     

Monocrotaline-induced cardiopulmonary damage in rats: amelioration by the angiotensin-converting enzyme inhibitor CL242817

Pulmonary injury induced by the plant alkaloid monocrotaline is partially prevented by the angiotensin-converting enzyme (ACE) inhibitor captopril. CL242817 [(S-[R*,S*])-1-([3-acetylthio]-3-benzoyl-2-methyl-propionyl)- L-proline] is a new orally active ACE inhibitor under evaluation as an antihypertensive agent. To determine whether CL242817 also can modify monocrotaline-induced pulmonary injury, male rats were divided into four groups: control; CL242817 (60 mg/kg/day, po); monocrotaline (2.4 mg/kg/day, po); or monocrotaline plus CL242817, and were sacrificed after 6 weeks of continuous treatment. Rats receiving monocrotaline alone exhibited occlusive medial thickening of the pulmonary arteries, cardiomegaly, and right ventricular hypertrophy. Electron micrographs of monocrotaline-treated lung revealed degeneration of both endothelial and Type I epithelial cells, as well as marked interstitial hypercellularity and fibrosis. Hydroxyproline (collagen) content of monocrotaline-treated lung also increased significantly, confirming the fibrosis observed in the electron micrographs. These structural changes were accompanied by decreased lung ACE and plasminogen activator (PLA) activities, indicative of pulmonary endothelial dysfunction. Concomitant CL242817 treatment ameliorated all anatomic manifestations of monocrotaline injury, particularly the right ventricular hypertrophy, pulmonary arterial occlusion, epithelial degeneration, and interstitial fibrosis. CL242817 also significantly prevented the monocrotaline-induced increase in lung hydroxyproline content. In contrast, concomitant CL242817 did not significantly influence the suppressed lung ACE and PLA activities in monocrotaline-treated rats. CL242817 alone produced retarded weight gain, decreased heart weight relative to body weight, decreased lung hydroxyproline content and ACE activity, and increased serum ACE activity and plasma AII concentration. Thus CL242817 resembles captopril, both in its ability to ameliorate monocrotaline-induced pulmonary injury in rats, and in many of its side effects.     

Monocrotaline-induced pulmonary fibrosis in rats: amelioration by captopril and penicillamine

The purpose of this study was to determine whether Captopril (an angiotensin converting enzyme inhibitor) or D-penicillamine (an inhibitor of collagen crosslinking) can ameliorate pulmonary fibrosis induced by the plant alkaloid monocrotaline. Rats were randomly assigned to one of six treatment groups: (1) control; (2) Captopril, 60 mg/kg/day, p.o.; (3) D-penicillamine, 30 mg/kg/day, p.o.; (4) monocrotaline, 2.4 mg/kg/day, p.o.; (5) monocrotaline plus Captopril, as above; (6) monocrotaline plus penicillamine, as above; and were killed after 6 weeks of continuous drug administration. Monocrotaline-treated rats exhibited several anatomic correlates of pulmonary hypertension, including cardiomegaly, right heart enlargement, and muscularization of the pulmonary arteries and arterioles. These monocrotaline reactions were accompanied by decreased lung activities of angiotensin converting enzyme (ACE) and plasminogen activator (PLA), indicative of endothelial dysfunction; and by increased lung hydroxyproline concentration, indicative of interstitial fibrosis. The presence of interstitial fibrosis was confirmed by electron microscopy. When given concomitantly with monocrotaline, both Captopril and penicillamine partially prevented the cardiomegaly, right heart enlargement, and vascular muscularization. Both agents also diminished the decreased lung PLA activity and increased hydroxyproline concentration observed in monocrotaline-treated animals. Neither modifying agent influenced the monocrotaline-induced decrease in lung ACE activity. Compared with control rats, the rats receiving Captopril alone exhibited decreased heart weight and increased serum ACE activity, and animals receiving penicillamine alone did not differ significantly from control animals for any of the endpoints studied. These data demonstrate that Captopril and penicillamine ameliorate monocrotaline-induced pulmonary fibrosis in rats. Penicillamine, known to inhibit radiation-induced lung injury, thus is shown to be effective in a second model of pulmonary fibrosis. Perhaps more importantly, the hydroxyproline data demonstrate that the ACE inhibitor Captropril exhibits antifibrotic activity in monocrotaline-treated rat lung.     

Intratracheal mesenchymal stem cell administration attenuates monocrotaline-induced pulmonary hypertension and endothelial dysfunction

The administration of mesenchymal stem cells (MSCs) has been proposed for the treatment of pulmonary hypertension. However, the effect of intratracheally administered MSCs on the pulmonary vascular bed in monocrotaline-treated rats has not been determined. In the present study, the effect of intratracheal administration of rat MSCs (rMSCs) on monocrotaline-induced pulmonary hypertension and impaired endothelium-dependent responses were investigated in the rat. Intravenous injection of monocrotaline increased pulmonary arterial pressure and vascular resistance and decreased pulmonary vascular responses to acetylcholine without altering responses to sodium nitroprusside and without altering systemic responses to the vasodilator agents when responses were evaluated at 5 wk. The intratracheal injection of 3 x 10(6) rMSCs 2 wk after administration of monocrotaline attenuated the rise in pulmonary arterial pressure and pulmonary vascular resistance and restored pulmonary responses to acetylcholine toward values measured in control rats. Treatment with rMSCs decreased the right ventricular hypertrophy induced by monocrotaline. Immunohistochemical studies showed widespread distribution of lacZ-labeled rMSCs in lung parenchyma surrounding airways in monocrotaline-treated rats. Immunofluorescence studies revealed that transplanted rMSCs retained expression of von Willebrand factor and smooth muscle actin markers specific for endothelial and smooth muscle phenotypes. However, immunolabeled cells were not detected in the wall of pulmonary vessels. These data suggest that the decrease in pulmonary vascular resistance and improvement in response to acetylcholine an endothelium-dependent vasodilator in monocrotaline-treated rats may result from a paracrine effect of the transplanted rMSCs in lung parenchyma, which improves vascular endothelial function in the monocrotaline-injured lung.     

Pulmonary endothelial dysfunction induced by unilateral as compared to bilateral thoracic irradiation in rats

Rats were sacrificed 2 months after a single dose of 10-30 Gy of 60Co gamma rays delivered to either a right unilateral or a bilateral thoracic port. Four indices of lung endothelial function were measured: the activities of angiotensin-converting enzyme (ACE) and plasminogen activator (PLA) and the production of prostacyclin (PGI2) and thromboxane (TXA2). The number of macrophages recovered by bronchoalveolar lavage (BAL) and the degree of right ventricular hypertrophy (an index of pulmonary hypertension) also were determined. Right lung ACE and PLA activity decreased linearly, and PGI2 and TXA2 production increased linearly with increasing radiation dose. The response curves for right unilateral and bilateral thoracic irradiation were not significantly different. In contrast, bilateral irradiation was more toxic than unilateral, since rats exposed to the former exhibited decreased body weight, an increased incidence of pleural effusions, an increase in the number of macrophages recovered by BAL, and right ventricular hypertrophy. These data demonstrate that pulmonary endothelial dysfunction induced by hemithorax irradiation represents a direct response of the endothelium to radiation injury and is not secondary to other phenomena such as shunting of function to the shielded lung.     

Radiation-induced pulmonary endothelial dysfunction and hydroxyproline accumulation in four strains of mice

C57BL mice exposed to 14 Gy of whole-thorax irradiation develop significant histologic lung fibrosis within 52 weeks, whereas CBA and C3H mice do not exhibit substantial fibrosis during this time. The purpose of the present study was to determine whether this strain-dependent difference in radiation histopathology is associated with genetic differences in pulmonary endothelial metabolic activity or in endothelial radioresponsiveness. C57BL/6J, C57BL/10J, CBA/J, and C3H/HeJ mice were sacrificed 12 weeks after exposure to 0 or 14 Gy of 300-kV X rays to the whole thorax. Lung angiotensin converting enzyme (ACE) activity and plasminogen activator (PLA) activity were measured as indices of pulmonary endothelial function; and lung hydroxyproline (HP) content served as an index of pulmonary fibrosis. Lung ACE and PLA activities in sham-irradiated C57BL/6J and CB57BL/10J mice were only half as high as those in sham-irradiated CBA/J and C3H/HeJ mice. Exposure to 14 Gy of X rays produced a slight but nonsignificant reduction in lung ACE and PLA activity in the C57BL strains, and a significant reduction in the CBA/J and C3H/HeJ mice. Even after 14 Gy, however, lung ACE and PLA activities in CBA/J and C3H/HeJ mice were higher than those in sham-irradiated C57BL/6J and C57BL/10J mice. Lung HP content in all four strains increased significantly after irradiation, but this increase was accompanied by an increase in lung wet weight. As a result, HP concentration (per milligram wet weight) remained constant or increased slightly in both C57BL strains and actually decreased in the CBA/J and C3H/HeJ mice. These data demonstrate significant genetic differences in both intrinsic pulmonary endothelial enzyme activity and endothelial radioresponsiveness among the four strains of mice. Specifically, strains prone to radiation-induced pulmonary fibrosis (C57BL/6J, C57BL/10J) exhibit only half as much lung ACE and PLA activity as do strains resistant to fibrosis (CBA and C3H).     

Radiation-induced endothelial dysfunction and fibrosis in rat lung: modification by the angiotensin converting enzyme inhibitor CL242817

The purpose of this study was to evaluate the angiotensin converting enzyme (ACE) inhibitor CL242817 as a modifier of radiation-induced pulmonary endothelial dysfunction and pulmonary fibrosis in rats sacrificed 2 months after a single dose of 60Co gamma rays (0-30 Gy) to the right hemithorax. CL242817 was administered in the feed continuously after irradiation at a regimen of 60 mg/kg/day. Pulmonary endothelial function was monitored by lung ACE activity, plasminogen activator (PLA) activity, and prostacyclin (PGI2) and thromboxane (TXA2) production. Pulmonary fibrosis was evaluated by lung hydroxyproline (HP) content. Lung ACE and PLA activities decreased with increasing radiation dose, and cotreatment with CL242817 significantly ameliorated both responses. CL242817 dose-reduction factors (DRF) were 1.3-1.5 for ACE and PLA activity. Lung PGI2 and TXA2 production increased with increasing radiation dose, and CL242817 almost completely prevented both radiation responses. The slope of the radiation dose-response curves in the CL242817-treated rats was essentially zero, precluding calculation of DRF values for PGI2 and TXA2 production. Lung HP content also increased with increasing radiation dose, and CL242817 significantly attenuated this response (DRF = 1.5). These data suggest that the ability of ACE inhibitors to ameliorate radiation-induced pulmonary endothelial dysfunction is not unique to captopril [Ward et al., Int. J. Radiat. Oncol. Biol. Phys. 15, 135-140 (1988)], rather it is a therapeutic action shared by other members of this class of compounds. These data also provide the first evidence that ACE inhibitors exhibit antifibrotic activity in irradiated rat lung.     

Radiation injury in rat lung. IV. Modification by D-penicillamine

To determine whether D-penicillamine, known to reduce fibrosis in irradiated rat lung (W. F. Ward, A. Shih - Hoellwarth , and R. D. Tuttle , Radiology 146, 533-537, 1983), also ameliorates radiation injury in the pulmonary endothelium, we measured angiotensin-converting enzyme (ACE) activity, plasminogen activator (PLA) activity, and prostacyclin (PGI2) production in the lungs of penicillamine-treated (10 mg/day, po, continuous after irradiation) and untreated rats from 2 weeks to 6 months after a single dose of 25 Gy of 60Co gamma rays to the right hemithorax. Both ACE and PLA activity in the irradiated right lung of untreated rats decreased dramatically between the 1st and 2nd months after exposure, then reached a plateau through 6 months at approximately 25 and 50% of the normal level, respectively. For the first 2 months after irradiation, penicillamine-treated animals exhibited significantly (P less than 0.05) higher activities of both ACE and PLA than did untreated rats. From 3 to 6 months after irradiation, however, the only significant drug effect on these enzymes was a 25% increase in PLA activity at 6 months. PGI2 production by the irradiated lung of untreated rats increased continuously, and at 6 months was approximately 10 times higher than normal. Penicillamine significantly (P less than 0.05) reduced this hypersecretion, and at 6 months after irradiation, PGI2 production by the lungs of drug-treated rats was only half that of untreated animals. In contrast, the drug had no significant effect on enzyme activities in the lungs of sham-irradiated rats. Thus the antifibrotic agent D-penicillamine delays the onset of radiation-induced enzyme dysfunction in the pulmonary endothelium. In addition at 6 months after irradiation, the lungs of penicillamine-treated rats exhibit 25% more PLA activity and only half as severe a hypersecretion of PGI2 as do the lungs of untreated animals. The drug is most effective in ameliorating endothelial damage during the first 2 months after irradiation, preceding the development of interstitial fibrosis. However, the effect of this penicillamine regimen on pulmonary endothelial function is not as large as its effect on collagen accumulation in irradiated rat lung.     

Prostacyclin and thromboxane A2 release in isolated rat lungs

The influence of platelets and platelet membranes on the generation of prostacyclin (PGI₂) and thromboxane A₂(TXA₂) by isolated rat lung and porcine aortic endothelial cell, as measured by RIA of their stable end-producs, 6-oxo-PGF_1α and TXB₂ respectively, was studied. After introduction of either aspirin-treated platelets or membranes from aspirin-treated platelets to the perfusate, 1 5-fold increase in the amount of 6-oxo-PGF_1α and TXB₂ in the perfusate was observed. Treatment of the lung with aspirin produced a 50% reduction in the platelet-stimulated release of PGI₂ and TXA₂. Treatment of the lung with the phospholipase inhibitor, mepacrine, significantly reduced the platelet-stimulated release of PGI₂ and TXA₂. Incubation of endothelial cells with untreated platelet membranes did not alter the generation of PGI₂. These results suggest that platelet-stimulated release of PGI₂ and TXA₂ occurs via mechanical stimulation of phospholipase A₂, liberating arachidonic acid.     

Neither anticoagulant nor nonanticoagulant heparin affects monocrotaline lung injury

The administration of monocrotaline to rats causes pulmonary vascular leak within 1 wk followed in 2-3 wk by perivascular proliferation and fatal pulmonary hypertension. Possibly blocking the proliferation might block the pulmonary hypertension, providing insight into its mechanism. Because heparin, given as an antiproliferative agent, reduced hypoxic pulmonary hypertension in mice, it might also block monocrotaline-induced pulmonary hypertension. Alternatively, anticoagulation could worsen the lung injury. We found that heparin (300 and 600 U/kg sc twice daily) inhibited clotting in rats given monocrotaline but did not change the vascular leak, the right ventricular pressure, the right ventricular hypertrophy, the increased medial thickness of the pulmonary arterioles, or the production of a slow-reacting substance of anaphylaxis-like material by the lungs. A nonanticoagulant heparin fragment (2 mg/kg sc twice daily), given to avoid anticoagulation also did not influence the monocrotaline injury. Thus neither anticoagulant nor nonanticoagulant heparin either attenuated or worsened the measured effects of monocrotaline.     

Pentoxifylline does not spare acute radiation reactions in rat lung and skin.

Male rats were exposed to single doses (0-30 Gy) of 60Co gamma rays to the right hemithorax. Half of each dose group consumed only control powdered chow after irradiation, and half consumed feed containing 0.10% (w/w) pentoxifylline (50 mg/kg/day). The severity of epilation and desquamation in the field of the radiation port was scored weekly. Two months after irradiation the animals were killed, and pulmonary endothelial function was monitored by the activity of lung angiotensin converting enzyme (ACE) and plasminogen activator (PLA), and by production of prostacyclin (PGI2) and thromboxane (TXA2). The amount of hydroxyproline (HP) in the lung served as an index of pulmonary fibrosis. Radiation produced a dose-dependent decrease in ACE and PLA activity in the right lung and an increase in the production of PGI2 and TXA2. This endothelial dysfunction was accompanied by an increase in wet weight and in protein and HP content in the irradiated lung. Pentoxifylline spared only the increase in lung wet weight and protein content, and actually elevated the radiation-induced hyperproduction of PGI2 and TXA2. The severity of the epilation and desquamation reactions increased with increasing radiation dose and time but was independent of diet. These data indicate that pentoxifylline, despite some promising pharmacological actions, has no beneficial effect on acute radiation reactions in rat lung and skin.     

Influence of chronic antigen exposure on the metabolism of PGH2 by microsomal preparations of airway and parenchymal tissues in the sheep

The effects of repeated antigen exposure on the synthesis of mediators by lung tissues are not well understood. To investigate the influence of antigen challenge on the synthesis of prostaglandins by central airway and peripheral lung tissues, fourteen sensitive sheep underwent biweekly exposure to aerosolized $\text{Ascaris suu}$ antigen (7) or saline (7). Following the fifth exposure, microsomal and high speed supernatant fractions were prepared from trachealis muscle and lung parenchyma. Synthesis of thromboxane (TX) A₂, prostaglandin (PG) D₂ and PGI₂ from the PG endoperoxide intermediate, PGH₂, was assayed over a range of substrate concentrations from 3–200 uM. Synthesis of PGI₂ by trachealis microsomes was approximately 5-fold greater than that of TXA₂. PGI₂ and TXA₂ production was identical in tracheal preparations from Ascaris- and saline-exposed animals. In parenchymal tissues, where TXA₂ production predominated over PGI₂ by 9-fold, preparations from Ascaris- exposed animals synthesized 50% more TXA₂ than controls at PGH₂ concentrations of 25 uM and above, whereas synthesis of PGI₂ and PGD₂ were similar in preparations from both groups of animals. The density of pulmonary mast cells was decreased by 21% in the Ascaris group, whereas polymorphonuclear leukocyte density was unchanged. These results demonstrate the differential synthesis of TXA₂ and PGI₂ in central airways and peripheral lung regions of the sheep. They further indicate that repeated exposure of the airways to antigen selectively enhances TXA₂ synthesis in the lung periphery of sensitized animals. The site of this increased enzymatic activity, whether in resident cells or newly-infiltrated cells, has not been determined.     

Myocardial prostacyclin and thromboxane A2 synthetase activities during ischemia and reperfusion in isolated rabbit heart

The aim of the study was to determine the prostacyclin (PGI₂) and thromboxane A₂ (TXA₂) synthetase activities of myocardial tissue and their variation during ischemia and reperfusion. Regional ischemia was induced by 10 min occlusion of the left anterior descending coronary artery in isolated Langendorff rabbit hearts. Biosynthesis of PGI₂ and TXA₂ were carried out by using arachidonic acid as substrate and left ventricle microsomes (LVM) from ischemic and non-ischemic areas as sources of PGI₂ and TXA₂ synthetase. 6-keto-PGF_1α and TXB₂, stable metabolites of PGI₂ and TXA₂ respectively, were determined by radioimmunoassay. Experiments carried out under the adopted conditions showed that LVM were able to synthetise PGI₂ as well as TXA₂ from arachidonic acid. On the other hand, ischemia depressed both PGI₂ and TXA₂ synthetase activities of cardiac tissue: the depression was more pronounced on TXA₂ synthetase than on PGI₂ synthetase with no significant difference between ischemic and non-ischemic regions. Moreover, ischemia increased the ratio indicating therefore that it can facilitate the formation of PGI₂. The post ischemic reperfusion of the heart counteracted the decrease in PGI₂ synthetase induced by ischemia which returned to the normal level: reperfusion also slightly reversed the decrease in TXA₂ synthetase. However, the diminution in TXA₂ synthetase of non-ischemic myocardium was attenuated but it remained lower than the normal level. These results suggested that the whole left ventricle is affected by regional ischemia. Furthermore it appears that myocardial TXA₂ synthetase is more vulnerable than PGI₂ synthetase to a lack of oxygen and nutrients.     

Fasudil dichloroacetate (FDCA), an orally available agent with potent therapeutic efficiency on monocrotaline-induced pulmonary arterial hypertension rats

Given the therapeutic efficacy of fasudil hydrochloride (F) and dichloroacetate (DCA) on pulmonary arterial hypertension (PAH), a new salt fasudil dichloroacetate (FDCA) was designed, synthesized and biologically evaluated. FDCA exhibited comparable ROCK II inhibitory activity relative to fasudil hydrochloride, and suppressed the expression of TNF-α and IL-6 in both PDGF-BB and hypoxia-treated pulmonary arterial smooth muscle cells (PASMCs) and endothelial cells (PAECs). Significantly, FDCA lowered mean pulmonary artery pressure (mPAP) and right ventricular systolic pressure (RVSP), and decreased right ventricular hypertrophy (RVH) in monocrotaline (MCT)-induced PAH rats. Meanwhile, FDCA remarkably decreased pulmonary artery medial thickness (PAMT) and hyperplasia, restoring the elasticity of elastic fiber, reduced cardiac hypertrophy, and attenuated fibrosis of heart and lung. Collectively, FDCA exhibited triple activities of pulmonary vasodilation, vascular remodeling inhibition and RVH inhibition, suggesting that it may be a promising agent for PAH intervention.     

Longitudinal in vivo SPECT/CT imaging reveals morphological changes and cardiopulmonary apoptosis in a rodent model of pulmonary arterial hypertension

Pulmonary arterial hypertension (PAH) has a complex pathogenesis involving both heart and lungs. Animal models can reflect aspects of the human pathology and provide insights into the development and underlying mechanisms of disease. Because of the variability of most animal models of PAH, serial in vivo measurements of cardiopulmonary function, morphology, and markers of pathology can enhance the value of such studies. Therefore, quantitative in vivo SPECT/CT imaging was performed to assess cardiac function, morphology and cardiac perfusion utilizing ²⁰¹Thallium (²⁰¹Tl) in control and monocrotaline-treated rats. In addition, lung and heart apoptosis was examined with ^99mTc-Annexin V (^99mTc-Annexin) in these cohorts. Following baseline imaging, rats were injected with saline or monocrotaline (50 mg/kg, i.p.) and imaged weekly for 6 weeks. To assess a therapeutic response in an established pulmonary hypertensive state, a cohort of rats received resveratrol in drinking water (3 mg/kg/day) on days 28–42 post-monocrotaline injection to monitor regression of cardiopulmonary apoptosis. PAH in monocrotaline-treated rats was verified by conventional hemodynamic techniques on day 42 (right ventricular systolic pressure (RSVP) = 66.2 mmHg in monocrotaline vs 28.8 mmHg in controls) and in terms of right ventricular hypertrophy (RV/LVS = 0.70 in monocrotaline vs 0.32 in controls). Resveratrol partially reversed both RVSP (41.4 mmHg) and RV/LVS (0.46), as well as lung edema and RV contractility +dP/dt_max. Serial ^99mTc-Annexin V imaging showed clear increases in pulmonary and cardiac apoptosis when compared to baseline, which regressed following resveratrol treatment. Monocrotaline induced modest changes in whole-heart perfusion as assessed by ²⁰¹TI imaging and cardiac morphological changes consistent with septal deviation and enlarged RV. This study demonstrates the utility of functional in vivo SPECT/CT imaging in rodent models of PAH and further confirms the efficacy of resveratrol in reversing established monocrotaline-induced PAH presumably by attenuation of cardiopulmonary apoptosis.     

Effects of nitrogen dioxide exposure on prostacyclin synthesis in lung and thromboxane A2 synthesis in platelets in rats

Effects of nitrogen dioxide (NO₂) exposure on prostacyclin (PGIP₂) synthesis in the rat lung and thromboxane A₂ (TXA₂) synthesis in the platelets were studied. Male Wistar rats were exposed to 10 ppm NO₂ for 1, 3, 5, 7 and 14 days. PGI₂ synthesizing activity of homogenized lung decreased. The damage of PGI₂ synthesizing activity reaches its maximum at 3 days. At 14 days, PGI₂ synthesizing activity returned to the normal level. The activity of PGI₂ synthetase decreased significantly. The formation of lipid peroxides due to NO₂ exposure may cause the depression of PGI₂ synthesizing activity of lung. On the other hand, platelet TXA₂ synthesizing activity increased. This increased TXA₂ synthesizing activity lasted at least till 3 days. Then, it returned to the normal level. The counts of platelet were decreased significantly by 1, 3, 5 and 7 days NO₂ exposure. Then the decreased counts of platelet returned to the normal level at 14 days NO₂ exposure. These results indicate that the depression of PGI₂ synthesizing activity lung by NO₂ exposure cause an increase in TXA₂ synthesizing activity of platelets. It may contribute to induce platelet aggregation and to the observed decrease in the number of platelets during NO₂ exposure.     

Early treatment with fumagillin, an inhibitor of methionine aminopeptidase-2, prevents Pulmonary Hypertension in monocrotaline-injured rats

Pulmonary Hypertension (PH) is a pathophysiologic condition characterized by hypoxemia and right ventricular strain. Proliferation of fibroblasts, smooth muscle cells, and endothelial cells is central to the pathology of PH in animal models and in humans. Methionine aminopeptidase-2 (MetAP2) regulates proliferation in a variety of cell types including endothelial cells, smooth muscle cells, and fibroblasts. MetAP2 is inhibited irreversibly by the angiogenesis inhibitor fumagillin. We have previously found that inhibition of MetAP2 with fumagillin in bleomycin-injured mice decreased pulmonary fibrosis by selectively decreasing the proliferation of lung myofibroblasts. In this study, we investigated the role of fumagillin as a potential therapy in experimental PH. In vivo, treatment of rats with fumagillin early after monocrotaline injury prevented PH and right ventricular remodeling by decreasing the thickness of the medial layer of the pulmonary arteries. Treatment with fumagillin beginning two weeks after monocrotaline injury did not prevent PH but was associated with decreased right ventricular mass and decreased cardiomyocyte hypertrophy, suggesting a direct effect of fumagillin on right ventricular remodeling. Incubation of rat pulmonary artery smooth muscle cells (RPASMC) with fumagillin and MetAP2-targeting siRNA inhibited proliferation of RPASMC in vitro. Platelet-derived growth factor, a growth factor that is important in the pathogenesis of PH and stimulates proliferation of fibroblasts and smooth muscle cells, strongly increased expression of MetP2. By immunohistochemistry, we found that MetAP2 was expressed in the lesions of human pulmonary arterial hypertension. We propose that fumagillin may be an effective adjunctive therapy for treating PH in patients.     

Consequences of long-term selenium-deficient diet on the prostacyclin and thromboxane release from rat aorta

It is known that peroxides, which are increased during Se deficiency because of reduced glutathione peroxidase (GSH-Px) activity, can influence the prostacyclin I₂/thromboxane A₂ (PGI₂/TXA₂) ratio. In this study we analyzed the PGI₂ and TXA₂ formation of aortas of long-term Se-deficient rats. Despite low GSH-Px activity in the Se-deficient group, the basal PGI₂ and TXA₂ formation was not different versus control animals (PGI₂: 2295 ± 1134 pg/mg vs 2940 ± 1134 pg/mg; TXA₂: 3.83 ± 1.06 pg/mg vs 5.67 ± 2.99 pg/mg). However, we checked the capacity of the aortas of Se-deficient rats to compensate for a suddenly increased peroxide concentration. After peroxide stimulation, the PGI₂ release was significantly lower in the Se-deficient group compared to the control group (PGI₂: 3507 ± 1829 pg/mg vs 7986 ± 2636 pg/mg). Again, the TXA₂ release did not show any differences. The release ratio of PGI₂/TXA₂ decreased under peroxide stress in Se-deficient animals. Although long-term Se deficiency showed a relatively well-balanced metabolism under resting conditions, sudden stress, accompanied by an excessive radical production, cannot be compensated.     

Characterization of a murine model of monocrotaline pyrrole-induced acute lung injury

Background

New animal models of chronic pulmonary hypertension in mice are needed. The injection of monocrotaline is an established model of pulmonary hypertension in rats. The aim of this study was to establish a murine model of pulmonary hypertension by injection of the active metabolite, monocrotaline pyrrole.

Methods

Survival studies, computed tomographic scanning, histology, bronchoalveolar lavage were performed, and arterial blood gases and hemodynamics were measured in animals which received an intravenous injection of different doses of monocrotaline pyrrole.

Results

Monocrotaline pyrrole induced pulmonary hypertension in Sprague Dawley rats. When injected into mice, monocrotaline pyrrole induced dose-dependant mortality in C57Bl6/N and BALB/c mice (dose range 6–15 mg/kg bodyweight). At a dose of 10 mg/kg bodyweight, mice developed a typical early-phase acute lung injury, characterized by lung edema, neutrophil influx, hypoxemia and reduced lung compliance. In the late phase, monocrotaline pyrrole injection resulted in limited lung fibrosis and no obvious pulmonary hypertension.

Conclusion

Monocrotaline and monocrotaline pyrrole pneumotoxicity substantially differs between the animal species.     

Protection of oral hydrogen water as an antioxidant on pulmonary hypertension

This study aimed to explore the protective effect of hydrogen as an antioxidant on monocrotaline (MCT)-induced pulmonary hypertension (PH). Forty-eight SD rats were equally randomized into four groups: SHAM group, MCT group, MCT+Oral-H₂ group and MCT+Inj-H₂ group. The results showed that the mean pulmonary arterial pressure, right ventricle weight and right ventricular hypertrophy index in MCT group were significant higher than those in SHAM group; pulmonary inflammatory response, atrial natriuretic factor, 3-nitrityrosine and intercellular adhesion molecule-1 were also increased significantly in MCT group. These indexes were decreased significantly in both MCT+Oral-H₂ group and MCT+Inj-H₂ group, which indicate Oral-H₂ and Inj-H₂ have similar effects of preventing the development of PH and mitigating RV hypertrophy. The protective effect of hydrogen is associated with its antioxidative ability and action of reducing pulmonary inflammatory response. While Oral-H₂ is more convenient than Inj-H₂, Oral-H₂ may be ideal for clinical use in future.