Protective role of endogenous hydrogen sulfide in cholecystokinin octapeptid against acute lung injury induced by lipopolysaccharide

AIM: To explore the role of endogenous hydrogen sulfide (H₂S) in the mechanism of cholecystokinin octapeptide (CCK-8) to alleviate acute lung injury (ALI) induced by lipopolysaccharide (LPS). METHODS: Eighty-four Sprague-Dawley rats were randomly divided into seven groups: control, LPS (instilled intratracheally to reproduce the model of ALI), NaHS (H₂S donor) +LPS, propargylglycine ［inhibitor of cysathionine-γ-lyase (CSE), PPG］+LPS, CCK-8+LPS, PPG+CCK-8+LPS and CCK-8 group. Animals were sacrificed at 4 h and 8 h after agent instillation. The wet and dry ratio (W/D) of the lung weight was measured and calculated. Morphological changes of lung tissues were observed. H₂S concentration in plasma, malondialdehyde (MDA) content, myeloperoxidase (MPO) and CSE activities in the lung were determined. Furthermore, the level of P-selectin of lung tissue was measured by radioimmunoassay, the CSE mRNA expression in the lung was detected by RT-PCR, and the protein content in bronchoalveolar lavage fluid (BALF) was detected. RESULTS: Compared with control, severe injury of lung tissues and increase in W/D, protein content in BALF, MDA content, MPO activity and P-selectin level in the lung were observed in rats treated with LPS. LPS also lead to a drop in plasma H₂S concentration, lung CSE activity and CSE mRNA expression. Administration of NaHS before LPS could attenuate the changes induced by LPS, while H₂S concentration, CSE activity and CSE mRNA expression were higher than those in LPS group. However, pre-treatment with PPG exacerbated the lung injury induced by LPS, H₂S concentration, CSE activity and CSE mRNA expression were lower than those in LPS and CCK-8 +LPS group, respectively. CONCLUSION: CCK-8 attenuates LPS-induced acute lung injury by means of anti-oxidation and inhibition of PMN adhesion and aggregation, both of which are mediated by endogenous H₂S.     

Effects of ginsenoside Rh1 on expression of inflammatory factors in mouse asthma model

AIM: To observe the effects of ginsenoside Rh1 on the levels of inflammatory factors in serum and bronchoalveolar lavage fluid (BALF), and the pathological changes of the lung tissues in an experimentally induced mouse asthma model. METHODS: Male BALB/c mice (n=40) were divided into 4 groups:normal control group, asthma mo-del group, and low-dose (40 mg·kg^-1·d^-1) and high-dose (80 mg·kg^-1·d^-1) ginsenoside Rh1 groups. The bronchial asthma mouse model was established by the method of ovalbumin induction and excitation, and during the excitation period, the mice were daily treated with ginsenoside Rh1 for 2 weeks. At 24 h after the final dose of ginsenoside Rh1, the mice were sacrificed. The number of eosinophils (EOS) and the concentrations of interleukin (IL)-4, IL-5 and interferon (IFN)-γ in BALF were determined. The levels of IgG and IgE in serum were measured, and the expression of transforming growth factor (TGF)-β1 and the pathological changes in lung tissues were evaluated. RESULTS: Ginsenoside Rh1 inhibited the increases in the number of EOS and the concentrations of IL-4, IL-5, IFN-γ and IgE, reversed the increased expression of TGF-β1, and improved the pathological changes of the lung tissues in asthmatic mice. CONCLUSION: Ginsenoside Rh1 improves the immuno-inflammatory profile and pathological changes in the experimentally induced mouse asthma model, implying its potential therapeutic effect on asthma.     

Protective effects and mechanism of astragalus injection on asthmatic rats

AIM: To explore the protective effects and mechanism of astragalus injection on asthmatic rats.METHODS: OVA was injected intraperitoneally and inhaled to produce the asthmatic model.Forty rats were randomly divided into five groups: control group,asthma group and astragalus groups of high,medium and low dose.The concentrations of IL-4,IFN-γ in BALF,the expression of IL-4 mRNA,IFN-γ mRNA and phospho-p38 MAPK in lung tissues were respectively measured by ELISA,RT-PCR and Western blotting.The number of inflammatory cells in BALF and histropathology changes were observed.RESULTS: In asthmatic group,the number of inflammatory cells and the concentrations of IL-4 in BALF and the expression of IL-4 mRNA,phospho-p38 MAPK in lung tissue were higher,but IFN-γ and IFN-γ mRNA were lower than those in normal control rats (P<0.01).In astragalus group,the number of inflammatory cells,the concentrations of IL-4 in BALF and the expression of IL-4 mRNA,phospho-p38 MAPK in lung tissue were lower,but IFN-γ and IFN-γ mRNA were higher than those in normal control rats (P<0.01),and histropathology damage was alleviated significantly.The efficacies in the astragalus groups of high,medium and low dose were similar,which no significant difference was observed among them.There were positive correlations between the expression of 〖JP3〗phospho-p38 MAPK and the number of eosinophil,the concentration of IL-4,IL-4 mRNA (r=0.63,r=0.69,r=0.71,〖JP〗 P<0.01),and negative correlations between the expression of phospho-p38 MAPK and IFN-γ and IFN-γ mRNA (r=-0.65,r=-0.68,P<0.01).CONCLUSION: p38 MAPK may play a role in pathological process of asthma.Astragalus effectively treats asthma by inhibiting the expression of phospho-p38 MAPK,correcting the inbalance of IFN-γ/ IL-4 and decreasing the number of inflammatory cells.     

Endogenous hydrogen sulfide regulates oxidant/antioxidant balance in rat with chronic obstructive pulmonary disease

AIM: To investigate the role of hydrogen sulfide (H₂S) in the protection against oxidative stress in rats with chronic obstructive pulmonary disease (COPD).METHODS: The rat model of COPD was established by cigarette smoking (CS) combined with lipopolysaccharide (LPS) instillation. Thirty-two healthy male Sprague-Dawley rats were randomly divided into 4 groups: control group, CS+LPS group, CS+LPS+NaHS (H₂S donor) group and CS+LPS+PPG (DL-propargylglycine, an inhibitor of cystathionine-γ-lyase) group. After 30 days, the lung functions of the rats were measured, the histological changes of lungs were observed under light microscope and the pathological scores were calculated. The H₂S level in plasma and the protein expression of cystathionine-γ-lyase (CSE) in the lung tissues were measured. The content of malondialdehyde (MDA), the activity of superoxide dismutase (SOD) and catalase (CAT) were detected to reflect oxidative stress.RESULTS: Compared with control group, the peak expiratory flow (PEF) decreased by 24% and intra-pressure (IP) increased by 66% in CS+LPS group. The pathological scores of the lung tissues also increased. Compared with CS+LPS group, no change in the lung function was observed after given NaHS or PPG, but the pathological scores decreased in CS+ LPS+ NaHS group. Compared with control group, the content of H₂S in plasma was increased by 26% on day 16. Compared with CS+LPS group, the content of H₂S in plasma of CS+LPS+PPG group was decreased by 22% after 30 days. Compared with control group, the protein expression of CSE increased, and no statistical difference among CS+LPS group, CS+ LPS+ NaHS group and CS+LPS+PPG group was observed. Compared with control group, MDA content in the lung tissues was increased by 24% in CS+LPS group, the activity of SOD was increased by 47% and the activity of CAT was increased by 52%. Compared with CS+LPS group, the MDA content in CS+LPS+NaHS group was decreased by 21%, and no statistical difference in the activity of SOD and CAT was observed. The activity of SOD decreased by 33% after given PPG.CONCLUSION: H₂S plays a role as anti-oxidant in the rats with COPD. The CSE/ H₂S pathway may be involved in the development of COPD.     

H2S inhibits neuron apoptosis induced by anoxia-reoxygenation through cAMP-mediated PI3-K/Akt/P70S6K kinase cell-survival signaling pathways

AIM: To investigate the effect of hydrogen sulfide on neuron apoptosis through PI₃-K/Akt/P70S6K cell-survival signal transduction pathways after neuron anoxia-reoxygenation.METHODS: Newborn (24-48 h) Wistar rats were decapitated.The hippocampus tissue was dissected and cells were suspended.Cells were plated at 1.0×10⁸ cells/L on poly-dlysine-treated 96-well (100 μL/well) plates and 6-well (2 mL/well) plates.Cells were used after 7 days.For anoxia-reoxygenation (oxygen glucose deprivation,OGD) experiments,cells were washed three times in a glucose-free balanced salt solution (BSS).They were then placed in deoxygenated glucose-free medium and cultured under 95% N₂,5% CO₂ in an anaerobic chamber equilibrated to 37 ℃ and 100% humidity for 45 min.OGD was terminated by replacement of stored medium and by returning the cultures to a standard incubator maintained at 37 ℃ in 95% air,5% CO₂.In experimental group,cells were respectively carried out OGD,OGD+150 μmol/L NaHS,OGD+150 μmol/L NaHS+10 μmol/L triciribin,OGD+150 μmol/L NaHS+10 nmol/L rapamycin and OGD+150 μmol/L NaHS+10 μmol/L triciribin+10 nmol/L rapamycin.Control cells were cultured normally.24 h later,neuron viability and apoptosis were measured.The level of cAMP and protein expression of PI₃-K,Akt and P70S6K were detected.RESULTS: NaHS enhanced concentration of cAMP and expression of PI₃-K,Akt and P70S6K.Meanwhile,increased neuron viability and decreased neuron apoptosis (P<0.01 vs group C or group I/R) were observed.Triciribin inhibited Akt and P70S6K,as well as increased neuron apoptosis and decreased neuron viability (P<0.05,P<0.01 vs group NaHS).Rapamycin inhibited P70S6K,as well as increased neuron apoptosis and decreased neuron viability (P<0.05,P<0.01 vs group NaHS).CONCLUSION: H₂S inhibits hippocampus neuron apoptosis and protects neuron from anoxia-reoxygenation injury through cAMP-mediated PI₃-K/Akt/P70S6K kinase cell-survival signaling pathways.     

Effect of hydrogen sulfide on airway inflammation induced by ozone in mice

AIM: To investigate the effect of hydrogen sulfide (H₂S) on airway inflammation induced by ozone (O₃) exposure and its mechanisms.METHODS: C57BL/6 mice (n=32) were randomly divided into control group, O₃ group, NaHS+O₃ group and NaHS group. The mice in O₃ group and O₃+NaHS group were exposed to 2.14 mg/m³ O₃ for 3 h on days 1, 3 and 5, while the mice in control group and NaHS group were exposed to filtered air. NaHS (14 μmol/kg) was administered intraperitoneally to the mice in NaHS group and O₃+NaHS group 30 min before each exposure. After the last exposure for 24 h, the airway responsiveness was determined, and bronchoalveolar lavage fluid (BALF) was collected for counting inflammatory cells and measuring total protein concentration. The lung tissues were collected for observing the morphological changes with HE staining. The levels of interleukin-6 (IL-6), interleukin-8 (IL-8), malondialdehyde (MDA) and NF-κB p65 protein in the lungs were determined.RESULTS: Compared with control group, the airway responsiveness, inflammatory cells, protein concentration, inflammation score, levels of IL-6, IL-8, MDA and NF-κB p65 in O₃ group increased significantly, but these in NaHS+O₃ group decreased compared with O₃  group.CONCLUSION: The present findings suggest that H₂S attenuates O₃ induced airway inflammation by inhibiting NF-κB expression and preventing lipid peroxidation.     

Effect of hydroxylamine on pulmonary arterial hypertension induced by chronic hypoxic hypercapnia in rats

AIM: To explore the effects of hydroxylamine on the pulmonary arterial pressure in chronic hypoxic hypercapnic rats. METHODS: Twenty-four male Sprague-Dawley rats were randomly divided into 3 groups (8 rats in each group): the normal control group (NC), hypoxic hypercapnia+normal saline group (NS), hypoxic hypercapnia+hydroxylamine group (HA). The animals in NS and HA groups were kept in the O₂ (9%-11%) and CO₂ (5%-6%) cabin, 8 h a day and 6 days a week for 4 weeks. Before entering the cabin, the rats in HA group were administered with 1 mL hydroxylamine (12.5 mg/kg) by intraperitoneal injection, while the rats in NS group were given intraperitoneal injection of 1 mL saline solution. The mean pulmonary arterial pressure (mPAP) was measured by external jugular vein cannulation. The heart was removed, and the right ventricle (RV) and the left ventricle plus the septum (LV+S) were dissected. The ratio of the wet weight of the RV to that of the LV+S was calculated. The changes of the pulmonary vascular construction were observed under optical microscope. The concentration of H₂S in the plasma was measured with a spectrometer. The expression of cystathionine-γ-lyase (CSE) in the pulmonary arterioles and bronchi was measured by immunohistochemistry and RT-PCR. RESULTS: The values of mPAP, RV/(LV+S),vessel wall area/total area (WA/TA) and media thickness of pulmonary arterioles (PAMT) in NS group and HA group were significantly higher than those in NC group (P<0.05). The level of H₂S in the plasma, the content of CSE protein and the expression of CSE mRNA in NC group were significantly lower than those in NS group (P<0.05). The values of mPAP, RV/(LV+S), WA/TA and PAMT in HA group were significantly lower than those in NS group (P<0.05). The level of H₂S in the plasma, the content of CSE protein and the expression of CSE mRNA in HA group were significantly higher than those in NS group (P<0.05). CONCLUSION: Hydroxylamine may decrease the pulmonary arterial hypertension induced by chronic hypoxic hypercapnia in rats by increasing the level of H₂S in the plasma, the content of CSE protein and the mRNA expression of CSE, thus improving the pulmonary vascular structural remodeling.     

Role of hydrogen sulfide in acute lung injury induced by ischemia-reperfusion of hind limbs in rats

AIM: To explore the role of endogenous and exogenous hydrogen sulfide (H₂S) in acute lung injury (ALI) induced by ischmia-reperfusion (IR) of hind limbs in rats.METHODS: A Sprague-Dawley rat model of acute lung injury was induced by ischemia of the hind limbs for 4 h and reperfusion for another 4 h. The rats (n=120) were randomly divided into 4 groups: control, IR, NaHS (H₂S donor)+IR, and propargylglycine +IR. The animals were sacrificed after reperfusion. Lung weight/body weight ratio (LW/BW) was measured and calculated. Morphological changes of the lung tissues were observed. The concentrations of H₂S, nitric oxide (NO) and carbon monoxide (CO) in plasma were tested. The content of malondialdehyde (MDA), the activity of CSE, inducible nitric oxide synthase (iNOS) and hemeoxygenase (HO) in the lungs were determined. The polymorpho-nuclear neutrophils(PMN) and protein content in bronchoalveolar lavage fluid(BALF) were also measured. The correlation of H₂S content with the above indices was analyzed.RESULTS: Compared with control group, severe injuries of the lung tissues, raised LW/BW, MDA concentration, PMN and protein contents in BALF were observed in IR group. Limb IR also made a drop in the concentration of plasma H₂S and the activity of lung CSE, while the activity of iNOS and HO in the lung tissues and the levels of plasma NO and CO increased. Administration of NaHS before IR attenuated the changes induced by IR, while pre-administration of PPG exacerbated the IR injuries and increased the plasma NO level and lung iNOS activity. The H₂S content was positively correlated with CSE activity, CO content and HO-1 activity (P<0.01), and negatively correlated with the other indices (P<0.01).CONCLUSION: Down-regulation of H₂S/CSE is involved in the pathogenesis of acute lung injury induced by IR. Endogenous and exogenous H₂S protects against lung injuries. The anti-injury effects of H₂S are related with its anti-oxidative activity to attenuate the inflammatory over-reactions in the lung induced by PMN. Down-regulation of NO/iNOS system and up-regulation of CO/HO-1 system by H₂S are also involved in the process of anti-injury to ALI.     

Adipolin/CTRP12 protects against LPS-induced ARDS by up-regulating alveolar epithelial sodium channel in mice

AIM: To investigate the effect of adipolin/CTRP12 in LPS-induced acute respiratory distress syndrome(ARDS) and its potential regulation on alveolar epithelial sodium channel(ENaC) in mice. METHODS: C57BL/6J mice(n=40) were randomly divided into control group, LPS group, adipolin group and wortmannin(PI3K inhibitor) group with 10 mice in each group using random number table. The pathological changes of the lung tissues were evaluated by HE staining. The alveolar fluid clearance(AFC) was measured by Evans blue-marked albumin, and the concentrations of total protein in bronchoalveolar lavage fluid(BALF) were assessed by bicinchoninic acid(BCA) method. In BALF, the levels of IL-1β and TNF-α were determined by ELISA, and the activity of myeloperoxidase(MPO) was detected by an MPO assay kit. The total cell counts and polymorphonuclear neutrophil(PMN) counts in the BALF were analyzed by Giemsa staining. The mRNA levels of α-ENaC were assessed by qPCR, while the protein levels of α-ENaC and p-Akt were determined by Western blot. RESULTS: Compared with control group, the classic ARDS pathological changes were observed in the mice in LPS group, manifesting by severe pathological lung injury(P<0.05), increases in W/D weight ratio, total protein levels, cell counts, MPO activitiy, and IL-1β and TNF-α levels in the BALF, and decrease in AFC(P<0.05), accompanied by down-regulated levels of α-ENaC and p-Akt in the lung tissues(P<0.05). The deteriorating effects triggered by LPS were significantly reversed by administration of adipolin. However, PI3K inhibitor wortmannin canceled the beneficial effects of adipolin on LPS-induced ARDS, as evidenced by aggravated lung injury, increased levels of W/D weight ratio, protein levels, cell counts, MPO activity, and IL-1β and TNF-α levels in the BALF(P<0.05), and decreased levels of AFC, α-ENaC and p-Akt in the lung tissues. CONCLUSION: Adipolin protects against LPS-induced ARDS in the mice by up-regulating α-ENaC and enhancing AFC via PI3K/Akt signal pathway.     

Changes in endothelin-1 and atrial natriuretic factor and the effect of atrial natriuretic factor on the change of endothelin-1 in asthmatic guinea pigs

AIM: To investigate the possible role of endothelin(ET-1) in asthma pathogenesis and the effect of atrial natriuretic factor (ANF) on changes of ET-1. METHODS: Measuring the contents of endothelin-1(ET-1),atrial natriuretic factor(ANF),cGMP in plasma and bronchoalveolar lavage fluid (BALF) of guinea pigs. RESULTS: The contents of ET-1, ANF and cGMP in asthma group were higher than that of the control group; There was a significant negative correlation between ET-1 and ANF( r=-0.638,P <0.05) in plasma of the asthma group, and a significantly negative correlation between ET-1 and ANF( r=-0.921,P <0.01) in BALF of the asthma group. There was a significantly positive correlation between ANF and cGMP( r=0.848,P <0.01) in plasma of the asthma group,and a significantly positive correlation between ANF and cGMP ( r=0.831,P <0.01) in BALF of the asthma group. The levels ET-1 in the asthma+rat ANF(rANF) group were lower than those in the asthma group,and the levels of cGMP in the asthma+rat ANF(rANF) group were higher than those in the asthma group after ceasing to infuse rANF for guinea pigs for 30 minutes.CONCLUSION: ET-1 might play an important role in the pathogenesis of asthma.ANF might inhibit production of ET-1.     

CB2 receptor agonist JWH133 exerts protective effects on rat model of paraquat-induced acute lung injury

AIM: To study the protective effects of cannabinoid CB2 receptor agonist JWH133 on rat acute lung injury induced by paraquat (PQ).METHODS: Male Sprague-Dawley rats (n=72) were randomly divided into 4 groups. PQ group: PQ was administered intraperitoneally at the dose of 20 mg/kg; Low-dose JWH133 pretreatment group (L-JWH133 group): JWH133 (5 mg/kg, ip) was administered 1 h before PQ exposure; high-dose JWH133 pretreatment group (H-JWH133 group): JWH133 (20 mg/kg, ip) was administered 1 h before PQ exposure; control group: 1 mL saline was administered intraperitoneally. Arterial blood, bronchoalveolar lavage fluid (BALF) and lung tissue samples were collected at 8 h, 1 d and 3 d after PQ exposure. PaO2 and the levels of TNF-α and IL-1β in BALF were measured via blood gas analyzer and ELISA, respectively. The pathological changes and lung injury scores were assessed at 3 d after PQ exposure. NF-κB and AP-1 protein levels were also determined by Western blotting.RESULTS: The decrease in PaO2, structural injury of the lung tissues, interstitial pulmonary edema, and the increase in IL-1β and TNF-α in BALF were observed in PQ-treated rats compared with control group. JWH133 pretreatment reduced the degree of lung tissue injury, decreased the levels of IL-1β and TNF-α in BALF and the NF-κB and AP-1 protein expression in the lung tissue compared with PQ group, especially in H-JWH133 group. CONCLUSION: CB2 receptor agonist JWH133 inhibits NF-κB and AP-1 protein expression in the lung tissues, and reduces the secretion of IL-1β and TNF-α in BALF after paraquat exposure, thus attenuating paraquat-induced acute lung injury.     

Expression of IFN-γ, IL-4 and IL-17A in asthmatic mice vaccinated with BCG and HepB in neonatal period

AIM: To investigate the expression of IFN-γ, IL-4 and IL-17A in asthmatic mice vaccinated with bacillus Calmette-Guérin (BCG) and hepatitis B (HepB) in the neonatal period. METHODS: BALB/c mice were randomly divided into BGG+HepB+ovalbumin (OVA) group (B/H/O group), B/O group, H/O group, B/H group, OVA group, BCG group, HepB group and normal saline (NS) group (n=6). The mice in B/H/O group and B/H group at 0, 7 and 14 d received subcutaneous injection of 1×10⁵ CFU BCG for 3 times, while at 0 and 28 d received intramuscular injection of 1.5 μg HepB on the hindlimb twice. The mice in other groups were individually vaccinated with BCG or HepB. OVA sensitization and aerosol inhalation were performed to establish the asthma model. The lung tissues were collected for HE staining. Bronchoalveolar lavage fluid (BALF) and peripheral blood (PB) were collected, and the number of eosinophils (EOS) in BALF was counted. The serum levels of IFN-γ and IL-4, and the level of IL-17A in lung tissue homogenate were measured by ELISA. RESULTS: The pathological changes of the lung in OVA group, B/O group, B/H/O group and H/O group were observed. There were extensive inflammatory cell infiltration around the bronchus, and epithe-lial cell hypertrophy. Those in B/H/O group and H/O group were worse than those in OVA group, while those in B/O group was better than those in OVA group. Total BALF cell counts in B/H/O group, B/O group and H/O group were decreased (P<0.05) as compared with OVA group. The BALF EOS count in B/H/O group was higher than that in B/H group, that in B/O group was higher than that in BCG group, and that in H/O group was higher than that in HepB groups (P<0.05). Compared with H/O group, OVA group and NS group, the serum IFN-γ/IL-4 ratio in HepB group was increased (P<0.05), and compared with B/H/O group, B/O group, OVA group and NS group, that in B/H group was also increased (P<0.05). Compared with OVA group, the level of IL-17A in the lung tissues of B/H/O group and B/O group was decreased (P<0.05), and compared with B/O group, that in B/H/O group was further decreased (P<0.05). CONCLUSION: Combined vaccination of BCG and HepB reduces the inflammotory responses in the lung tissues of asthmatic mice. The mechanism may be related with the decrease in the release of IL-4, the increase in IFN-γ/IL-4, and the inhibition of IL-17A expression.     

Protective effect of Auricularia auricular-judae polysaccharide on pulmonary tissues of rats with LPS-induced acute lung injury

AIM: To investigate the effects of Auricularia auricular-judae polysaccharide(AAP) on pulmonary tissues of rats with LPS-induced acute lung injury(ALI) and its mechanisms.METHODS: Adult Sprague-Dawley rats were randomly divided into control group, LPS group,low-dose AAP group, middle-dose AAP group, high-dose APP group, and dexamethasone group. The rats were injected with LPS(8 mg/kg, ip) to induce ALI. The rats in the AAP groups were treated with AAP for 7 d before the induction of ALI. The protein concentration in the bronchoalveolar lavage fluid(BALF) was measured. The lung edema degree was measured by detecting the wet/dry weight ratio. The myeloper-oxidase(MPO), total antioxidant capacity(T-AOC), total superoxide dismutase(T-SOD), nitric oxide synthase(NOS) and malondialdehyde(MDA) levels were determined. The pathological changes of the lung tissues were evaluated by HE staining.RESULTS: Treatment with AAP significantly improved LPS-induced lung pathological changes, attenuated the protein concentration in the BALF and wet/dry weight ratio, inhibited the activities of MPO and NOS, reduced MDA level and increased the activities of T-AOC and T-SOD.CONCLUSION: AAP protects against LPS-induced acute lung injury in rats.     

Effects of M. vaccae on eosinophil apoptosis and Bcl-2 protein expression in lung tissues of asthmatic guinea pigs

AIM: To study the effects of M. vaccae on eosinophil apoptosis and Bcl-2 protein expression in lung tissues of asthmatic guinea pigs. METHODS: 30 guinea pigs were divided into normal saline (NS) group, asthma group and M. vaccae treatment group at random, every group included 10 guinea pigs. Guinea pigs in M. vaccae treatment group were injected intramuscularly with 22.5 μg M. vaccae 10 days before OVA immunization. TdT-mediated dUTP nick end labeling (TUNEL) technique was used to investigate the apoptosis of eosinophils and immunohistochemistry method was used to study the expression of Bcl-2 protein in lung tissues. RESULTS: The apoptosis index (AI) of eosinophils in lung tissues in M. vaccae treatment group was significant higher than that in asthma group ［(23.78±5.42)% vs (4.56±0.68)%, P<0.01］. The mean optical density value of Bcl-2 protein in lung tissues of M. vaccae treatment group was significant lower than that of asthma group ［(1 556.3±492.4) vs (2 321.9±751.2), P<0.05］. CONCLUSION: The apoptosis of eosinophils induced by M. vaccae in lung tissues of asthmatic guinea pigs may be due to the inhibition of Bcl-2 protein expression.     

Meconium induces formation of nitrotyrosine and expression of inducible nitric oxide synthase in the rat lung

AIM: To evaluate the contribution of inducible nitric oxide synthase (iNOS) and nitrotyrosine to acute lung injury (ALI) in rats with meconium aspiration. METHODS: 16 health male Sprage-Dawley rats were randomized to control group and meconium group, followed by intratracheally administration of 1 mL/kg saline or 1 mL/kg 20% human newborn meconium suspension. The animals were killed after 24 h of treatment. The measurements included bronchoalveolar lavage fluid (BALF) cell count, pulmonary myoloperoxidase (MPO) activity and nitric oxide (NO) level. Western bloting was used to determine the expression of pulmonary nitrotyrosine-a specific “footprint” of peroxynitrite and iNOS. RESULTS: Compared to control group, the rats in the meconium group had increased BALF cell counts ［(4.04±1.01)×109cells/L vs （0.53±0.19)×109cells/L］, pulmonary MPO activity ［(1.49±0.22）U/g wet lung tissue vs (0.62±0.16) U/g wet lung tissue］, NO level ［(12.77±5.00） mmol/g protein vs （4.89±1.32) mmol/g protein］, increased expression of nitrotyrosine and iNOS (0.46±0.19 and 1.49±0.60 vs 0.15±0.04 and 0.09±0.04, respectively), all P<0.01. CONCLUSIONS: Meconium results in an increase in expression of pulmonary iNOS, leading to over production of NO and nitrotyrosine, which may be of pathogenic importance in the ALI with meconium aspiration.     

Inhalation of inactivated Mycobacterium phlei down-regulates expression of nuclear factor-kappa B and intercellular adhesion molecule-1 in asthmatic mice

AIM:To investigate the effect of inhalation of inactivated Mycobacte-rium phlei on the expression of nuclear factor-kappa B (NF-κB), intercellular adhesion molecule (ICAM)-1 and vascular cell adhesion molecule (VCAM)-1 in the lung tissues of asthmatic mice. METHODS:Male BALB/c mice (n=24) were randomly divided into normal control group (A), asthmatic model group (B), and inactivated Mycobacterium phlei inhalation group (C). Asthmatic model was made by inhalation of chicken ovalbumin. The mice in group C were treated with inactivated Mycobacterium phlei for 5 d. The lung tissues and bronchoalveolar lavage fluid (BALF) were harvested. HE and AB-PAS staining were used to measure the lung inflammation and mucus production. The inflammatory cells in the BALF were counted. The mRNA expression of NF-κB, ICAM-1 and VCAM-1 was detected by real-time fluorescence quantitative PCR. RESULTS:Mycobacterium phlei treatment alleviated lung inflammation, attenuated mucus production, and reduced the percentage of eosinophils in the BALF. The mRNA levels of NF-κB and ICAM-1 were significantly decreased after treated with Mycobacterium phlei. However, no significant difference of VCAM-1 mRNA expression was found before and after treatment. The correlation between NF-κB mRNA and ICAM-1 mRNA, and between NF-κB mRNA and VCAM-1 mRNA was not found. CONCLUSION:Inhalation of inactivated Mycobacterium phlei attenuates asthmatic airway inflammation. NF-κB participates in the pathogenesis of asthma. NF-κB signal pathway may be associated with the therapeutic mechanism. Another important mechanism is the reduction of adhesion molecule expression.     

Bone marrow-derived mesenchymal stem cell-conditioned medium atte-nuates LPS-induced acute lung injury in mice

AIM:To explore the effects of bone marrow-derived mesenchymal stem cells-conditioned medium (MSCs CdM) on lipopolysaccharide (LPS)-induced acute lung injury. METHODS:Lung injury was induced in mice by intraperitoneal injection of LPS. The mice were given a tail vein injection of MSCs CdM or normal saline 1 h after LPS administration. The mice were killed by an intraperitoneal injection of pentobarbital 6 h after LPS injection for either bronchoalveolar lavage fluid (BALF) and serum collection or lung histological analysis. RESULTS:Compared with control group, the BALF levels of protein, interleukin-10 (IL-10) and keratinocyte growth factor (KGF), the serum levels of tumor necrosis factor α (TNF-α) and IL-6, and the myeloperoxidase (MOP) activity in the lung tissues were significantly higher in LPS group, and severe pathological damages in the lung tissues were also observed. Treatment with MSCs CdM significantly reduced the BALF prtein level, the seum TNF-α and IL-6　levels and the lung MPO activity, and attenuated the lung pathological damages, but further increased the levels of IL-10 and KGF in the BALF. CONCLUSION:Treatment with MSCs CdM attenuates the lung injuries induced by LPS, which may be via regulating the expression of TNF-α, IL-6, IL-10 and KGF.     

Effects of zanthoxylum seed oilA2 on NF-кB signaling pathway and inflammatory factor in lung tissue of asthmatic mice

AIM: To investigate the dynamic influence of zanthoxylum seed oilA2 (ZSOA2) on NF-кB signaling pathway and inflammatory factor in pulmonary tissue of asthmatic mice. METHODS: The suspensoid (0.2 mL containing 20% albumin hydroxide and 10% ovalbumin) was administered by intraperitoneal injection to sensitize the BALB/c mice on day 1, then 0.4% ovalbumin solution (50 μL in phosphate buffer fluid) was dripped into the respiratory tract through nasal cavity to establish the asthmatic mouse model. After dripped ovalbumin for 24 h, 48 h, 3 d, 7 d and 14 d, the mice were killed at specified time points. The contents of interleukin-4 (IL-4), interleukin-5 (IL-5) and interferon-γ (IFN-γ) in bronchoalveolar lavage fluid (BALF) were determined by ELISA. The pathological changes of the lung tissues were observed with HE staining. The inflammatory cell counts were conducted by Eosin staining. The protein levels of adhesion molecule and the molecules of NF-κB signaling pathway in lung tissues were determined by Western blotting. RESULTS: In ZSOA2 treated mice, the pathological injury of the lung was significantly attenuated as compared to the model mice, the counts of eosinophils and lymphocytes were reduced obviously in lung bronchial area of asthmatic mice at all observed time points (P<0.05). The levels of IL-5 and IL-4 decreased and IFN-γ increased in BALF. The results of Western blotting showed that ZSOA2 down-regulated the protein levels of intercellular adhesion molecule-1, vascular cell adhesion molecule-1, I kappa B kinase alpha-α and phosphorylation inhibitory-κB. ZSOA2 also up-regulated the protein levels of tumor necrosis factor receptor 1 and phosphorylation nuclear factor-kappaB in lung tissue at all observed time points. CONCLUSION: ZSOA2 has therapeutic effect on asthma by down-regulating the protein expression of IκB-β and p-IκB, inhibiting the releases of cytokines and chemotactic factors, and attenuating the infiltration of inflammatory cells in the lungs of ovalbumin challenged asthma mice.