Effects of aminoguanidine on endotoxin-induced acute lung injury in rabbits

AIM: To observe the effect of aminoguanidine (AG) on hemodynamics and lung capillary permeability in acute lung injury (ALI) in rabbits. METHODS:24 rabbits were equally divided into four groups: saline group, endotoxin group, AG group and AG plus endotoxin group. In AG plus endotoxin group, endotoxin was injected to animals to make an ALI model, 25mg/kg AG was injected following that and let this sustain 3 hours. Meanwhile, mean arterial pressure (MAP), mean pulmonary arterial pressures (MPAP) and blood gas analyses were observed during this period. At the end of the experiment, broncho-alveolus lavage was performed, pathologic samples were treated routinely and lung wet weight/dry weight ratio was calculated. RESULTS:After endotoxin injection, MAP and arterial oxygen pressure (PaO₂) decreased, and MPAP increased significantly. The injection of AG had little effect on MAP, but AG could markedly decrease MPAP and increase PaO₂. Cell count in broncho-alveolus lavage fluid (BALF) was less in AG plus endotoxin group than in endotoxin group. Although AG did not affect total protein in BALF, low molecular weight proteins decreased in AG plus endotoxin group by the assay of electrophoresis. Tissue wet weight/dry weight ratio also decreased in this group. Pathologic study showed that there were fewer inflammatory cells and less lung edema in AG plus endotoxin group. CONCLUSION:AG could improve hemodynamics status and attenuate acute lung injury induced by endotoxin in rabbits.     

Levels of CGRP and ET-1 in plasma of pulmonary artery and thoracic aorta and in the extractives of lung and ventricular tissues of the chronic hypoxic rats

AIM AND METHODS: To explore the effects calcitonin gene-related peptide (CGRP) and endothelin-1(ET-1) on the mechanisms of hypoxic pulmonary hypertension (HPH), the contents of CGRP and ET-1 in plasma of pulmonary artery and thoracic aorta and in extractives of lung and ventricular tissues of the chronic hypoxic rats were determined by radioimmunoassay. The changes of their hemodynamic indices and right heart hypertrophy index were monitored simultaneously. RESULTS: The level of pulmonary artery plasma CGRP was significantly higher than that of thoracic aorta plasma, but just the reverse was ET-1 or the ratio of ET-1 and CGRP in control rats( P<0.01). Compared with controls, the level of pulmonary artery plasma CGRP was gradually reduced in all hypoxic rats, but ET-1 was enhanced after 7 and 14 days of hypoxia and was decreased after 21 days of hypoxia. With prolonging time exposed to hypoxia, the level of thoracic aorta plasma CGRP was markedly increased in hypoxic animals compared with controls ( P<0.05), the positive correlation significantly with increased pulmonary arterial pressure( r = -0.896, P =0.05), but the lower level of thoracic aorta plasma ET-1 showed or negative correlation with pulmonary arterial pressure. CONCLUSIONS: These data suggest that the unbalance of effects of CGRP and ET may plays an important role in regulating the resistance of pulmonary circulation and has close relation with the formation of HPH.     

Effect of inhaled nitric oxide on aquaporin expression in newborn rat lung in acute hyperoxic lung injury

AIM:To investigate the effect of inhaled nitric oxide on aquaporin expression and alveolar epithelial fluid transport in newborn rats with acute hyperoxic lung injury. METHODS:32 newborn SD rats were randomized to breathe for 48 h room air (C), >95%O₂ (O), >95%O₂+10^-5 NO (NO only in the first 24 h, ONO), room air + NO (CN). Then, the rats were killed, the lung wet-to-dry weight ratio (Q_W/Q_D), the histology, and AQP1, AQP5, α₁-NKA, α-ENaC mRNA expressions in the lungs were measured. RESULTS:Compared with C group, the Q_W/Q_D in O group significantly increased (P<0.01), and AQP1 mRNA expression decreased significantly (P<0.01). Compared with O group, ONO group had a lower level of Q_W/Q_D (P<0.05), and AQP1 mRNA expression increased (P<0.05). AQP5 mRNA expression in all groups remained unchanged. CONCLUSION:In newborn rats with acute hyperoxic lung injury, inhaled 10-5 nitric oxide for 24 h may attenuate lung edema and increase AQP1 mRNA expression, suggesting that inhaled 10^-5 nitric oxide for 24 h may promote the AQP1 expression in lung in this model of acute lung injury.     

Effect of NG-nitro-L-arginine on mitochondria in acute lung injury induced by LPS in rats

AIM: To examine the effect of nonselective nitric oxide synthase inhibitor, NG-nitro-L-arginine (L-NA), on mitochondria from acute lung injury induced by lipopolysaccharides(LPS) in rats. METHODS: The rats were randomly divided into control group, LPS injury group and L-NA treatment group. The model of acute lung injury was prepared with injection of LPS in rats. L-NA was respectively administrated through intraperitoneal injection at 3 h after injury induced by LPS. The rats were killed and the mitochondria in lung tissues were isolated by differential centrifugation. The activities of T-NOS, iNOS, ATPase, SOD and GSH-Px, and the contents of NO and MDA from mitochondria were respectively measured. The changes of ultrastructure in lung mitochondria were examined by electronic microscope after injury and L-NA treatment. RESULTS: The activities of T-NOS and iNOS were significantly increased, the activities of ATPase, SOD and GSH-Px were significantly decreased, the contents of NO and MDA were increased after acute lung injury. L-NA significantly enhanced the activities of ATPase, SOD and GSH-Px, and markedly decreased the contents of NO and MDA and the activities of T-NOS and iNOS. CONCLUSION: L-NA inhibits the activity of NOS in mitochondria, decreases the production of NO, improves mitochondria energy pump, ameliorates oxidative injury, and effectively protects lung tissue against acute lung injury induced by LPS.     

Studies on the role of ET-1 and CGRP in the pathophysiology of ankylosing spondylitis and rheumatoid arthritis

AIM: To clarify the role of endothelin-1 (ET-1) and calcitonin gene related peptide(CGRP) in the pathophysiology of ankylosing spondylitis and rheumatoid arthritis. METHOD: Plasma/synovial fluid ET-1 and CGRP were measured by radioimmunoassay in patients with ankylosing spondylitis (AS) or rheumatoid arthritis (RA) and healthy control. RESULTS: ET-1 level in plasma of patients with AS and RA were significantly higher than that of healthy controls (P<0.01). No difference was found in plasma CGRP level between AS or RA and healthy control (P>0.05). CGRP level in synovial fluid was significantly higher than that in plasma (P<0.01), but ET-1 level was significantly lower than in plasma (P<0.01). CONCLUSION: These results suggest that ET-1 and CGRP play a pathogenic role in AS and RA.     

Protective effects of hypercapnia on acute lung injury and it's mechanisms

AIM: To investigate whether hypercapnia is protective against acute lung injury (ALI) in a rabbit model, and study it's potential mechanisms. METHODS: Twenty-two healthy New Zealand white rabbits were involved in this study, and randomly allocated to control group (group C), normocapnic group (group N) and hypercapnic group (group H). Oleic acid (0.1 mL/kg) was injected intravenously to establish ALI model. Lung mechanics, hemodynamics, blood-gas analysis, the content of malondialdehyde (MDA) and superoxide dismutase (SOD) activity in lung tissue were measured. Apoptosis was analyzed after 3h mechanical ventilation. RESULTS: (1) Peak airway pressure in group H was significantly lower than that in group N (P<0.05) and the dynamic lung compliance was significantly higher than that in group N (P<0.05). (2) PaO₂ in group H was significantly higher than that in group N(P<0.05). (3) The content of MDA was significantly lower but the activity of SOD was significantly higher in group H than that in group N (P<0.05). (4) Apoptosis index in group H was significantly lower than that in group N (P<0.01). (5) Histologic damage was significantly severer in group N than group H. (6) PaCO₂ was correlated with pH, PaO₂, dynamic lung compliance, peak airway pressure and pulmonary permeability index (r=－0.928, P<0.01; r=0.511, 0.526, -0.506, -0.556, P<0.05, respectively). CONCLUSION: Hypercapnia protects lung from oleic acid-induced injury in rabbits. The mechanisms of protection might be associated with improvement of oxidation/anti-oxidation imbalance and inhibition of apoptosis.     

Role of selectin in lung injury in rabbits with diabetic ketoacidosis

AIM: To study the effect of selectin on lung injury in rabbits with diabetic ketoacidosis(DKA). METHODS: 12 male New Zealand rabbits were divided into experimental group (DKgroups, intravenous injection of streptozocin and alloxan monohydroate, n =6) and normal control group (received same dose of saline, n=6). After 72 hours, all the animals were killed and blood and lung tissue were collected. Blood gases, expressions of selectin-Pand selectin-Lwere detected. RESULTS: The expression of selectin in DKgroup were higher than that in NSgroup. CONCLUSION: These results indicate that selectin expression may play an important role in lung injury during diabetic ketoacidosis.     

Effects of rat mesenchymal stem cells modified by CGRP on proliferation and phenotype transformation of vascular smooth muscle cells in vitro

AIM：To explore the effects of rat mesenchymal stem cells (MSCs) modified by calcitonin gene-related peptide (CGRP) on the proliferation and phenotype transformation of rat vascular smooth muscle cells (VSMCs) in vitro. METHODS：Rat MSCs and VSMCs were isolated, cultured and identified. The MSCs were infected by lentivirus which carried genes encoding enhanced green fluorescent protein (EGFP) and CGRP. The expression levels of CGRP in CGRP-modified MSCs were detected using real-time PCR and enzyme-linked immunosorbent assay (ELISA). The prolife-ration and migratory abilities of VSMCs were evaluated by MTT assay, Trypan blue staining and scratch test. The expression levels of α-smooth muscle actin (α-SMA) and osteopontin (OPN) were assessed by Western blotting. RESULTS：Compared with MSCs and MSCs-EGFP groups, the expression levels of CGRP in MSCs-CGRP group were markedly increased (P<0.01). The results of MTT assay and scratch test demonstrated that the proliferation and migratory abilities of VSMCs in MSCs-CGRP group were significantly inhibited compared with MSCs and MSCs-EGFP groups (P<0.05). Furthermore, cell viability was >90% shown by Typan blue staining. Western blotting showed that the expression of α-SMA was increased and that of OPN was decreased in MSCs-CGRP group compared with MSCs and MSCs-EGFP groups (P<005). CONCLUSION：CGRP-modified MSCs could secrete CGRP protein and inhibit the proliferation and migration of VSMCs, which may be associated with deterring the phenotype transformation from contractile type to synthetic type. These results lay a foundation for gene therapy in vivo.     

Paracrine effect of stem cells on treatment of acute lung injury/acute respiratory distress syndrome

Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) represent the leading cause of morbidity and mortality in critical patients. Despite improvements in supportive care and mechanical ventilation, recent data indicate that the mortality of ALI/ARDS is still high. Cell-based therapy with stem cells is an attractive new therapeutic approach. Stem cells have the capacity to secrete multiple paracrine factors that can regulate inflammation, improve alveolar fluid clearance, regulate endothelial and epithelial permeability, enhance tissue repair via chemotaxis and inhibit bacterial growth. This review focuses on the recent studies, which support the potential therapeutic use of stem cells in ALI/ARDS with an emphasis on the role of paracrine soluble factors.     

NO isn’t involved in the effect of TMP on vasomotion of arterioles in the rabbit mesentery

AIM: To elucidate the role of NO in the effect of tetramethylpyrazine (TMP) on vasomotion of arterioles induced by noradrenalin(NA).METHODS: Albino rabbits were anesthetized,respiration were maintained,the arterial pressure were measured with a pressure transducer attached to a polyethylene canula in the left common carotid artery,the intestinal loop was mounted on the stage of an inverted microscope and bathed in balanced solution.The inside diameter of the arterial was manually measured on a TV monitor using the TV camera mounted on the microscope.RESULTS: Topical applicated NA to measured field inducd spontaneous vasomotion which has frequency and amplitude.TMP inhibited vasomotion in vasodilation. Treatment with L-NMMA did not affect the effect of TMP.CONCLUSION: L-NMMA can not inhibit action of TMP on vasomotion,TMP may act on smooth muscle directly, not through endogenous NO.     

Effect of ghrelin on iNOS expression in alveolar macrophages and lung tissues in rats with sepsis-associated lung injury

AIM: To investigate the effect of ghrelin on inducible nitric oxide synthase (iNOS) expression in alveolar macrophages and lung tissues in sepsis-induced acute lung injury (ALI) rats. METHODS: The septic rat model was established by cecal ligation and puncture (CLP). Male SD rats were divided into sham group, CLP group and CLP+ghrelin group. The rats in the former 2 groups were further divided into 3 subgroups, which were 6 h, 12 h and 20 h post-operation groups. Ghrelin was administered by intraperitoneal injection at 3 h and 15 h after operation in ghrelin group. The samples were harvested 20 h after operation. The mRNA expression of iNOS in alveolar macrophages collected from bronchoalveolar lavage was detected by RT-PCR. The protein levels of lung iNOS were measured by Western blotting. The lung pathological examination was performed 20 h after operation. RESULTS: In CLP group, the mRNA expression levels of iNOS in the alveolar macrophages were 1.33±0.05, 1.44±0.08, 1.57±0.11 at 6 h, 12 h and 20 h after CLP, respectively, which were higher than that in sham group, but did not show time correlation. However, it was lower in CLP group than that in CLP+ghrelin group at 20 h after CLP (2.27±0.37, P<0.05). At 20 h after CLP, the protein level of lung iNOS was decreased in CLP+ghrelin group (0.87± 0.03) as compared with CLP group (1.08±0.05). Compared with sham group, the histopathological score was increased in both CLP group and CLP+ghrelin group, but it was lower in CLP+ghrelin group (5.83±0.477) than that in CLP group (7.83±0.75). CONCLUSION: Ghrelin treatment improves the degree of ALI. During 6 h to 20 h after CLP, the mRNA expression of iNOS in alveolar macrophages was elevated, but the difference was not seen as the time went on. Ghrelin up-regulates the mRNA expression of iNOS in alveolar macrophages and inhibits iNOS expression in lungs of septic rats.     

Upregulative effect of CGRP on expression of CREB and phospho-CREB protein during focal cerebral ischemia/reperfusion in rat parietal cortex

AIM: To investigate the effect of calcitonin gene related peptide (CGRP) on the expression of cyclic AMP response element binding protein (CREB) and phosphorylated-CREB in rat parietal cortex during focal cerebral ischemia/reperfusion (I/R).METHODS: Focal cerebral ischemia/reperfusion model was induced by occlusion of the right middle cerebral artery using the intraluminal suture method. The expressions of CREB and phospho-CREB in the parietal cortex in different groups (sham group, focal cerebral ischemia/reperfusion group and CGRP group) were detected with immunohistochemistry and Western blotting, and the positive products were analyzed by image analysis system.RESULTS: There was definite expression of CREB in right parietal cortex in sham group, while it was lesser in I/R group than that in sham group, but it became more in CGRP group than that in I/R group (P<0.05). Phospho-CREB was barely detected in right parietal cortex in sham group and it became more in I/R group than that in sham group. The expression of phospho-CREB increased in CGRP group than that in I/R group of the right parietal cortex (P<0.05).CONCLUSION: CGRP upregulates the expression of CREB and phospho-CREB in the ischemic neurons of the parietal cortex during focal cerebral ischemia/reperfusion and CREB probably involves in the mechanism of protective role of CGRP to ischemic neurons.     

Roles of endogenous nitric oxide in acute mountain sickness

Acute mountain sickness (AMS) is the greatest challenge of military medicine and wilderness medicine, as the increasing populations of entering Tibet and others who rapidly ascend to high altitude. AMS is considered as a kind of vascular dysfunction and disequilibrium of body fluid diseases induced by environmental stress. Studies reveal that endogenous nitric oxide (NO), a predominant endogenous vessel dilator, plays a crucial role in the pathogenesis of AMS. The decreased production and blocked delivery of NO cause dysfunction of the vascular system in hypoxic situations. Subsequently, AMS develops under the facilitations of alterations in brain and pulmonary blood vessels. NO or L-arginine (the precursor of NO) can also be used to prevent and treat AMS. This review will summarize the progress on the roles of NO in the pathogenesis, pathophysiology, treatment and prophylaxis of AMS.     

Effects of partial liquid ventilation on acute lung injury in piglets

AIM: To observe the effect of partial liquid ventilation with perfluorocarbon on gas exchange, hemodynamics and lung histological change in the piglets with surfactant depletion-induced acute lung injury. METHODS: 12 piglets were treated by lung lavage to a partial pressure of oxygen in arterial blood (PaO₂) below 100 mmHg for one hour and randomly divided into gas ventilation group (control group) and partial liquid ventilation (PLV) group, in PLV group, piglets received PFC (FC3280) intratracheally at doses of 15 mL/kg. The parameters of gas exchange and hemodynamics were measured before lung lavage, after lung lavage when the acute lung injury (ALI) was established, and 1 hour, 2 hours after ALI. Animals underwent euthanasia at the end of the study, lung histologic analysis followed. RESULTS: Surfactant depletion by lung lavage induced a stable acute lung injury.Gas exchange increased markedly in the animals that underwent PLV, less hemodynamic damage was observed in PLV group compared with the animals in GV group. Lung histologic analysis demonstrated a less lung damage, including atelectasis, neutrophil excudation, intra-alveolar hemorrhage and interstitial edema in PLV group compared with control group. CONCLUSION: In piglets with surfactant depletion-induced acute lung injury, partial liquid ventilation with perfluorocarbon can improve the gas exchange with less adverse hemodynamic effect and less lung injury compared with conventional gas ventilation.     

Effect of the combination of positive end expiratory pressure and nitric oxide inhalation on oleic acid-induced acute lung injury in canine

AIM:To observe the effects of the combination of positive end expiratory pressure (PEEP) and 80×10^-6 nitric oxide (NO) inhalation on oleic acid induced acute lung injury (ALI) in canine.METHEDS:30 dogs were divided into 6 groups. Oleic acid was injected through Swan-Ganz catheter to induced ALI. Pulmonary and systemic hemodynamics,blood gas were measured in dogs before and after injection of oleic acid and the period of inhaled NO for 1-6 h. The methemoglobin(MHb) concentrations were measured. Histology and ultrastructure of the lung tissue were observed. RESULTS:(1) The combination of PEEP and 80×10^-6 NO inhalation rapidly reduced mean pulmonary arterial pressure (MPAP) and pulmonary vascular resistance (PVR), increased PaO₂/FiO₂, reduced A-aDO₂ without inducing significant change on systemic hemodynamics. Arterial blood levels of MHb did not change significantly. (2)The combination group was showed the lightest ALI change by HE stain and electron microscopy. CONCLUSION:The combination of PEEP and inhalation of 80×10^-6 NO significantly and rapidly increased PaO₂/FiO₂ without producing lung injury induced by high FiO₂ and high PEEP.     

Effect of endogenous H2S on pulmonary hypertension during acute lung injury induced by LPS and its interaction with NO

AIM: To investigate the effect of H2S on pulmonary artery hypertension during acute lung injury induced by LPS and the interaction between the systems of hydrogen sulfide (H2S)/cystathionine-β-lyase (CSE) and nitric oxide (NO)/nitric oxide synthase (NOS) in this process. METHODS: Seventy-two adult male rats were randomly divided into four groups: control group, LPS group, LPS+L-NAME group and LPS+propargylglycine (PPG) group. Mean pulmonary artery pressure (mPAP) of each rat was examined at 2 h, 4 h, 6 h and 8 h after treatment. H2S and NO contents in plasma, NO content, iNOS, cNOS and CSE activity in lung were measured at 4 h or 8 h after treatment, respectively. Expression of iNOS in lung tissue was also detected by immunohistochemistry technique, and the injury of lung was evaluated with morphological changes under microscope. RESULTS: LPS could induce severe lung injury, and mPAP, NO content, iNOS activity and its protein expression in LPS group significantly increased, but cNOS activity, H2S content and CSE activity decreased compared with those of control group. Administration of L-NAME before LPS could attenuate the changes induced by LPS. Pre-administration of PPG, a CSE inhibitor, exacerbated the injury by LPS, but there was no prominent variation in cNOS activity. CONCLUSION: Reduced endogenous H2S could increase pulmonary artery hypertension during acute lung injury induced by LPS. There is a negative effect between H2S/CSE system and NO/NOS system in this process.     

Sphingosine-1-phosphate receptor-2 attenuates lipopolysaccharide-induced acute lung injury

AIM: To investigate the effects and mechanisms of sphingosine-1-phosphate receptor-2 (S1P2R)on lipopolysaccharide (LPS)-induced acute lung injury (ALI). METHODS: ALI model was induced by intratracheal administration of LPS in both wild-type mice and S1P2R -deficient mice. The pathological changes in the lung tissues were observed, and the protein concentration, total cell number, neutrophil ratio, TNF-α level and IL-6 level were determined in the bronchoalveolar lavage fluid (BALF) 24 h after LPS injection. In order to investigate the mechanisms of S1P2R in LPS-induced ALI, 10 min before LPS injection, both wild-type mice and S1P2R -deficient mice were injected with nitric oxide synthase inhibitor by tail vein injection, the pathological changes of the lung tissues were observed, and the protein concentration and total cell number in BALF were determined 12 h after LPS injection. RESULTS: Compared with wild-type mice, S1P2R -deficient mice showed more severe LPS-induced ALI, and the protein concentration, neutrophils and inflammatory cytokines in BALF were significantly increased in S1P2R -deficient mice. Administration of nitric oxide synthase inhibitor N^ω-L-nitro-arginine methyl ester protected S1P2R -deficient mice from aggravation of ALI. CONCLUSION: S1P2R mediates the protection from LPS-induced ALI possibly through inhibiting nitric oxide synthase.     

Shenfu injection prevents kidney from acute ischemia-reperfusion injury in rabbits

AIM:To further investigate preventive effects of Shenfu(SF) injection, a Chinese herb drug, on acute renal ischemic reperfusion injury (IRI).METHODS:After SF or normal saline was administered intravenously one t ime a day for four days,the renal ichemia-reperfusion(I-R)model was established by occlusion of right renal artery and vein for an hour and reperfusion for three hours after left nephroectomy.The activity of superoxide dismutase (SOD),content of malondialdehyde(MDA)in serum and renal tissue,and content of nitric oxide(NO),concentrat ions of Na⁺ and Ca²⁺ ,numbers of WBC adhesion in renal t issue were detected and light and electronic microscopy were used or the detection of the renal histological changes. RESULTS:SF lowered significantly MDA content in either renal tissues or serum , concentration of Na⁺, the number of WBC adhesion, and scores of tubules in renal tissue after renal I-R, and the SOD activity in renal tissues and serum and NO content in renal tissue were obviously increased by SF.In addition, renal histomorphological damage in either light or electronic microscope were lightened by SF. But Ca²⁺concentration in renal tissue appeared to be only mildly affected. CONCLUSION:The mechanisms that SF protects renal structure and function against acute renal IRI may be involved in increasing SOD activity, scavenging directly oxygen free redicals(OFR), raising NO content, inhibiting WBC adhesion and recruiment, preventing Na⁺ influx to form Na⁺ overload.     

Effect of calcitonin gene-related peptide on myocardin expression and phenotypic switch in vascular smooth muscle cells

AIM: To investigate the effects of calcitonin gene-related peptide (CGRP) on myocardin expression and phenotypic switch in vascular smooth muscle cells (VSMCs). METHODS: VSMCs were obtained by aortic tissue adherent culture and treated with angiotensin Ⅱ (AngⅡ), AngⅡ + CGRP or AngⅡ + CGRP + CGRP_8-37. The protein expression of myocardin and the phenotypic proteins of the VSMCs was detected by Western blot. RESULTS: The expression of myocardin in cultured VSMCs showed downregulation along with time expansion. The protein level of myocardin was higher at 48 h and 72 h than that at baseline in the cultured VSMCs (P<0.05). However, the myocardin was lower at 48 h and 72 h than that at baseline after treatment with CGRP in cultured VSMCs (P<0.05). Furthermore, at 48 h in cultured VSMCs, the myocardin decreased along with α-smooth muscle actin (α-SMA) (P<0.05), and osteopontin (OPN) increased (P<0.05) in AngⅡ group compared with control group. After treatment with CGRP, the levels of myocardin and α-SMA become higher (P<0.05) but OPN was lower (P<0.05) in CGRP group than those in AngⅡ group. CGRP_8-37 abrogated CGRP-induced increase in myocardin and α-SMA and decrease in OPN in CGRP_8-37 group compared with CGRP group. CONCLUSION: CGRP may regulate the phenotypic switch of the VSMCs and maintain the cells in contractile phenotype through the upregulation of myocardin protein, which may be accomplished by the combination of CGRP and its receptor.