| Part ?.Protective effects of tanshinone ?A sodium sulfonate on LPS induced acute lung injury in miceObjective:To investigate the protective effects of tanshinone ?A sodium sulfonate on lipopolysaccharide induced acute lung injury in mice.Methods:108 male C57/black mice,weight 25 to 30g,were randomly divided into three groups:(1)Control group(n=36),mice received equal volume saline intraperitoneally;(2)LPS group(n=36),mice were administrated with LPS 10 mg/kg intraperitoneally;(3)LPS+TSS group(n=36),mice were pretreated with TSS 30mg/kg,and were administrated with LPS 10 mg/kg intraperitoneally 30 minutes later.Mice were sacrificed by neck broken at 1h,6h,12h after NS or LPS injection.At each time point,12 mice were sacrificed,and lung tissues were sampled.The wet-to-dry ratio(W/D)of lung tissue was measured,and lung injury and inflammatory cell infiltration were evaluated after HE staining.Results:The alveolar structure of the control group was intact and the alveolar septum was uniform.In LPS group,pulmonary tissue surface was congested and edema,alveolar structure was damaged,alveolar space was widened,and a part of alveolar cavities exudate and bleed.Compared with the control group,the wet-dry ratio(W/D)of lung tissue in the LPS group was significantly higher at 1h,6h and 12h(P<0.05),while that in the LPS+TSS group was significantly higher at 1h and 6h(P<0.05).Tanshinone ?A sulfonate pretreatment can significantly reduce the pulmonary edema at 1h,6h,12h.There was a significant difference in the number of inflammatory cell infiltration at 1h and 6h after LPS stimulation compared with the control group(P<0.05),and there was no difference at 12h(P>0.05).Tanshinone ?A sodium sulfonate pretreatment significantly reduce the number of inflammatory cells infiltration at 1 h,6 h(P<0.05).Conclusions:LPS can dramatically induce acute lung injury in mice,when compared with control mice.TSS pretrement can protective alveolar structure,and significantly reduce pulmonary edema,inflammatory cells infiltration.Part ?.Effect of tanshinone ?A sodium sulfonate on HIF-1? and inflammatory factors in LPS induced acute lung injury in miceObjective:To investigated the effect of tanshinone ?A sodium sulfonate on HIF-1?and inflammatory factors in LPS induced acute lung injury in mice.Methods:108 male C57/black mice,weight 25 to 30g,were randomly divided into three groups:(1)Control group(n=36),mice received equal volume intraperitoneally;(2)LPS group(n=36),mice were administrated with LPS 10 mg/kg intraperitoneally;(3)LPS+TSS group(n=36),mice were pretreated with TSS 30mg/kg,and were administrated with LPS 10 mg/kg intraperitoneally 30 minutes later.Mice were sacrificed by neck broken at 1h,6h,12h after LPS injection.At each time point,12 mice were sacrificed,and lung tissues were sampled.The expression of HIF-1? in lung tissue was detected by RT-PCR and Western blot.Proinflammatory cytokines(TNF-?,IL-1?,and IL-6)and anti-inflammatory cytokines(IL-10)in the lung tissue were examined by enzyme linked immunosorbent assay kit(ELISA)and Real-time PCR(RT-PCR).Results:Compared with the control group,both of mRNA and protein expressions of HIF-1?,proinflammatory cytokine(TNF-?,IL-1? and IL-6)and anti-inflammatory cytokine(IL-10)were markedly increased at all observed time points after LPS stimulation(P<0.05).After LPS stimulation,HIF-1? mRNA and protein levels in the lung group decreased after peaking at 6h.Both of mRNA and protein expressions of TNF-?,IL-1?,mRNA of IL-6 and IL-10 were significantly increased at 1h and decreased at 6h after LPS stimulation.Compared with LPS group,sodium tanshinone ?A sulfonate significantly decreased the expression of HIF-1? mRNA and protein at 6h and 12h after LPS stimulation.Tanshinone ?A sodium sulfonate can reduce mRNA of TNF-?,IL-1?,and both of mRNA and protein of IL-6 at lh,6h,12h,meanwhile the expression of IL-10 mRNA and protein were up-regulated at 12h.Conclusions:The protective effect of TSS on LPS-induced lung injury may be attributed to the inhibition of HIF-1?,proinflammatory cytokines(TNF-?,IL-1?,and IL-6)and the up regulation of anti-inflammatory cytokines(IL-10).Part ?.Effect of HIF-1? on inflammatory factors in lipopolysaccharide induced acute lung injury in miceObjective:To investigate the effect of HIF-1? on inflammatory factors in lipopolysaccharide induced acute lung injury in mice.Methods:36 male C57/black mice,weight 25 to 30g,were randomly divided into three groups:(1)Control group(n=12),mice received equal volume intraperitoneally;(2)LPS group(n=12),mice were administrated with LPS 10 mg/kg intraperitoneally;(3)LPS+2ME2 group(n=12),mice were pretreated with 2ME2 20mg/kg,and were administrated with LPS 10 mg/kg intraperitoneally 30 minutes later.Mice were sacrificed by neck broken at 6h after NS or LPS injection,and lung tissue was taken for examination.The wet-to-dry ratio(W/D)of lung tissue was measured,and lung injury was evaluated after HE staining.ELISA and RT-PCR were used to detect tumor necrosis factor-?(TNF-?),interleukin-1?(IL-1?)and interleukin-6(interleukin-6)and interleukin-10(IL-10)in lung tissue.Results:Compared with the control group,the mRNA and protein levels of proinflammatory cytokines TNF-?,IL-1?,IL-6 and anti-inflammatory cytokines IL-10 were significantly up-regulated in LPS-induced ALI mice(P<0.05).Compared with the LPS group,the lung tissue damage and pulmonary edema were reduced in the LPS+2ME2 group,and the pro-inflammatory factors TNF-?,IL-1?,IL-6 mRNA and protein levels were significantly down-regulated(P<0.05),but the anti-inflammatory factors IL-10 mRNA and protein levels did not change significantly(P>0.05).Conclusions:Inhibition of HIF-1? expression can attenuate LPS-induced ALI pulmonary edema and the expression of pro-inflammatory factors TNF-?,IL-1? and IL-6,and play a protective role in the lung.Part ?.Tanshinone ?A sodium sulfonate induced attenuation of LPS induced acute lung injury is associated with reduction of hypoxia-inducible factor l? expression via PI3K/Akt Signaling pathwayObjective:To investigated the mechanism of tanshinone ?A sodium sulfonate on HIF-1? in LPS induced acute lung injury in mice.Methods:1.Effect of TSS on PI3K/AKT signaling pathway in LPS induced acute lung injury in mice36 male C57/black mice,weight 25 to 30g,were randomly divided into 3 groups:(1)Control group(n=12),mice received equal volume saline intraperitoneally;(2)LPS group(n=12),mice were administrated with LPS 10 mg/kg intraperitoneally;(3)LPS+TSS group(n=12),mice were pretreated with TSS 30mg/kg,and were administrated with LPS 10 mg/kg intraperitoneally 30 minutes later.Mice were sacrificed by neck broken at 6h after NS or LPS injection,and lung tissues were sampled.Akt phosphorylation and PI3K were examined by Western blot.2.Effect of PI3K inhibitor(LY294002)on HIF-1? in LPS induced acute lung injury in mice36 male C57/black mice,weight 25 to 30g,were randomly divided into 3 groups:(1)Control group(n=12),mice received equal volume saline intraperitoneally;(2)LPS group(n=12),mice were administrated with LPS 10 mg/kg intraperitoneally;(3)LPS+LY294002 group(n=12),mice were pretreated with LY294002 30mg/kg,and were administrated with LPS 10 mg/kg intraperitoneally 30 minutes later.Mice were sacrificed by neck broken at 6h after NS or LPS injection,and lung tissues were sampled.The expression of HIF-1? in lung tissue was detected by RT-PCR and Western blot.Results:PI3K and p-Akt/Akt were expressed at low levels in the control group.Compared with the control group,PI3K and p-Akt/Akt were significantly elevated in the LPS group.Compared with the LPS group,sodium tanshinone ?A sulfonate can significantly inhibit the expression of PI3K protein and phosphorylation of Akt in lung tissue.The PI3K inhibitor(LY294002)significantly inhibited the expression of HIF-1?mRNA and protein in lung tissue of ALI.Conclusions:Tanshinone II A sulfonate inhibits the expression of HIF-la through the PI3K/Akt pathway and plays a protective role in the lung. |
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