The Role Of Hydrogen Sulfide On Apoptosis In Rats Of Acute Lung Injury Induced By Agkistrodon Venom

Objective: To study the role of hydrogen sulfide on apoptosis in rats model ofacute lung injury induced by Agkistrodon Venom and explore the probablemechanism.Methods: Thirty six male SD rats were randomly divided into six groups(n=6):control group (group N,injection saline2ml/kg),venom group (group C, venom1.50mg/kg), venom+low-dose NaHS group (group L,venom1.50mg/kg+NaHS0.78mg/kg), venom+middle-dose NaHS group (group M, venom1.50mg/kg+NaHS1.56mg/kg),venom+high-dose NaHS group(group H, venom1.50mg/kg+NaHS3.12mg/kg), venom+L-propargylglycine(PPG) group (group P, venom1.50mg/kg+PPG30mg/kg). Saline or venom were administrated with intraperitonealinjection in groups according to the experimental grouping above.NaHS or PPG wererespectively administrated in L,M,H and P group at three hour right after the firstinjection of saline or venom.The rats were sacrificed at six hour after the firstinjection of saline or venom. The histopathological changes were measured by lightmicroscope and a semi-quantitative histological index of quantitative assessment afterHE staining.The apoptosis rate of lung cells was analysised by flow cytometry(FCM).The distribution and expression of Bcl-2and Bax protein were detected byimmunohistochemistry(IHC). The Caspase-3and Caspase-9protein expressions weretested by western blot.Results:1. Changes in histopathology of lung tussue(1) General view:The surface of lung in N group was pale pink and smoothwithout hemorrhagic foci and edema. The appearance of C group was significantlycongestion and edema, red with dark red hemorrhagic foci on surface. The damages tothe lung tissue of the L group, M group and H group were mitigated sequentially versus the C group. Compared with the C group, the appearances of the P group wereserious red of the lung tissue, large tracts of hemorrhagic foci, severe congestion andedema in the lung tissue.(2).Endoscopic view:The normal structure of pulmonary alveolar was damagedin S group versus the N group, such as the dilation of the capillary in the interstitial oflung tussue, the thickened alveolar septum associated with the interstitial edema,theincreased capillary permeability, the infiltration of inflammatory cells and RBC intoalveoli, and large tracts of pulmonary hyaline membrane either. Furthermore,thealveolar collapse and atelectasis also can be observed,and the scores of IQA(index ofquantitative assessment) increased evidently(P<0.01or P<0.05). Compared with thatof C group, the grade of IQA was significantly decreased (P<0.01) and the lung injurywere markedly mitigated in the M group and H group.Otherwise, the degree of lunginjury of P group was much severer than the other groups such as the extensivehyaline membrane and severe atelectasis were detected, and the scores of IQA weresignificantly increased versus those of other groups(P<0.05).2.Changes of apoptosis rateCompared with the N group,the apoptosis rate of lung cells were significantlyincreased in the rest of the groups(P<0.01).The rate of apoptosis was evidentlydecreased in L group,M group and H group versus C group(P<0.01)while that of the Pgroup was significantly increased (P<0.01).3.The distribution and expression of Bax and Bcl-2The positive expression of Bax and Bcl-2were stained buffy,and weredistributed in alveolar epithelial cells,vascular endothelial cells and infiltratedinflammatory cells.In the N group,there were trace positive expression of both Baxand Bcl-2in the lung tussue.In the C group,the expression of Bcl-2and Bax wasmarkedly increased while the ratio of Bcl-2/Bax were evidently decreased versus theN group(P<0.01).Compared with the C group,the expression of Bcl-2was obviouslyincreased in the L group,M group and H group(P<0.01),and the expression of Baxwas significantly decreased in the M group and H group(P<0.01).The changes of the expression of Bcl-2and Bax maked the significant increase of the ratio of Bcl-2/Baxin the L group,M group and H group(P<0.01or P<0.05).In the P group,otherwise,theexpression of Bax was markedly increased while Bcl-2and the ratio of Bcl-2/Baxwere significantly decreased compared with the C group(P<0.05).4. The expression of Caspase-9and Caspase-3The expression of Caspase-9and Caspase-3were significantly increased in the Cgroup versus those of the N group(P<0.01).Compared with the C group,theexpression of Caspase-9and Caspase-3were markedly dereased in the M group andH group while those of the P group were evidently increased(P<0.01or P<0.05).Conclusions1. The acute lung injury can be induced by the Agkistrodon Venom in SD rats.2. The apoptosis rate of lung tissue was significantly increased in the acute lung injuryassociated with the Agkistrodon Venom in rats, which illustrated that the apoptosisplays a core role in the acute lung injury induced by venom.3. The hydrogen sulfide can inhibit the apoptosis of lung tissue in SD rats with acutelung injury induced by Agkistrodon Venom, and thus to mitigate the damage of lungtissue from acute lung injury.The probable mechanism is that the hydrogen sulfidemay upregulated the expression of Bcl-2and reduced the expression of Bax,Caspase-9and Caspase-3.