The Effect Of Suberoylanilide Hydroxamic Acid On Claudins In Lung In Rats Suffering Serious Hemorrhagic Shock On Plateau

Objective To investigate the effect of Suberoylanilide hydroxamic acid on claudins in lung in rats suffering serious hemorrhagic shock on plateau,and the lung protection effect of SAHA and its possible mechanism.Methods Eighty-four Wistar rats were completely randomized into 7 groups(n = 12/group):Sham group,shock without resuscitation group(NR group),Lactated Ringer’s solution group(LR group),Lactated Ringer’s solution combined with suberoylanilide hydroxamic acid group(LR+SAHAgroup),Suberoylanilide hydroxamic acid group,hypertonic sodium chloride hydroxyethyl starch 40 solution group(HSH group)and hypertonic sodium chloride hydroxyethyl starch 40 solution combined with suberoylanilide hydroxamic acid group(HSH+SAHA group).Serious hemorrhagic shock model of Wistar rats was reproduced by Wigger’s method in Maxianshan(Gansu,3780m above sea level).The rats were only anesthetized,no shock,no resuscitation,and were sacrificed after 3h in the Sham group.The NR group rats were only bled and not resuscitated,and then were sacrificed at 90min after shock,specimens were taken immediately after death to be tested in the NR group.The other groups’ rats were bled to remain a mean arterial pressure of(35±5)mmHg for 90min.Then different resuscitation programes were implemented:LR group and HSH group received 1.5 times of blood loss volume of LR and 4ml/kg HSH within 20 minute and 5 minute respectively.While 7.5mg/kg SAHA were added in LR+SAHA group and HSH+SAHA group in the above mentioned basis.SAHA group was resuscitated with 7.5mg/kg SAHA dissolved in 0.25ml Saline merely,which was constantly infused within 5 minutes.Animals were sacrificed at 3h after resuscitation.The damage degree of lung was observed with Optical microscope.Tight junction in lung Epithelial cells,cell apoptosis and ultrastructural were observed with transmission electronic microscope.Wet to dry weight ratio(W/D),lung permeability index(LPI),myeloperoxidase activity(MPO)and malondialdehyde(MDA)in lung were measured.Enzyme-linked immunosorbent assay was used to detect the TNF-a in lung tissue.The expression and distribution of claudin-3 and claudin-4 in lung tissue were verified by Immunohistochemistry method.The level of claudin-3,claudin-4,P-JNK,P-P38 and NF-κB in lung was measured by Western blot..Results Compared with Sham group,W/D,lung injury scores,lung permeability index,pulmonary respiration index,PaO2/FiO2 and claudin-4 were significantly decreased(P<0.05).But MPO,claudin-4 were obviously increased(P<0.05)in NR group,LR group,LR+SAHA group and HSH + SAHA group.Western blot demonstrated that claudin-3 and claudin-4 were significantly reduced.The expression of P-JNK,P-P38 and NF-κB were significantly increased(P<0.05).W/D,lung injury scores,lung permeability index,pulmonary respiration index,claudin-3 and claudin-4 in HSH+SAHA group were increased(P<0.05).MPO was deincreased(P<0.05),But the expression of claudin-3,claudin-4 and P-P38 in lung of Western blot were cut down.P-JNK and NF-κB were increased(P<0.05).Lung MDA in NR group and LR group was rised(P<0.05),While there was no significant difference among LR+SAHA group,HSH group and HSH+SAHA group(P>0.05).Compared with NR group,W/D,claudin-3 and claudin-4 in LR group were obviously increased.While lung MPO,MDA and TNF-α were distinctly reduced(P<0.05).Western blot demonstrated that the expression of claudin-3 was rised.But NF-κB,P-JNK and P-P38 were deincreased(P<0.05).Compared with NR group W/D,lung injury scores,lung permeability index,pulmonary respiration index,MPO,MDA and TNF-α were significantly reduced in LR+SAHA group,SAHA group,HSH group and HSH+SAHA group(P<0.05).PaO2/FiO2,claudin-3 and claudin-4 were increased(P<0.05).The expression of claudin-3,claudin-4 in lung of Western blot were increased.P-JNK,P-P38 and NF-κB were droped(P<0.05).These change in HSH+SAHA group were the greatest.Compared with SAHA group,lung injury scores,W/D,lung permeability index,claudin-3 and claudin-4 were reduced(P<0.05).Pulmonary respiration index and PaO2/FiO2 appear no difference(P>0.05).MPO,MDA and TNF-a were significantly increased(P<0.05).The expression of lung claudin-3 and claudin-4 in Western blot were reduced.NF-κB,P-JNK and P-P38 were clearly increased(P<0.05).W/D,MPO,MDA and TNF-α in LR group were reduced,But claudin-3 and claudin-4 were increased(P<0.05),There was no difference among the other index(P>0.05),Western blot demonstrated that lung claudin-3 and claudin-4 in LR group were reduced,While NF-κB,P-JNK and P-P38 were distinctly gone up(P<0.05).W/D,MPO,MDA and TNF-α in HSH group were reduced(P<0.05),but claudin-3 and claudin-4 were rised(P<0.05),There was no difference among the other index(P>0.05).The expression of claudin-3 and claudin-4 in HSH group of Western blot were increased,yet NF-κB,P-JNK and P-P38 were reduced(P<0.05).Lung injury scores,W/D,lung permeability index,MPO,MDA and TNF-α were obviously reduced in LR+SAHA group and HSH+SAHA group,Pulmonary respiration index,PaO2/FiO2,claudin-3 and claudin-4 appeared increased(P<0.05),The expression of lung claudin-3,claudin-4,NF-κB,P-JNK and P-P38 in Western blot appeared evidently difference(P<0.05).Compared with LR group,Lung injury scores,W/D,lung permeability index,MPO,MDA and TNF-a in HSH group were obviously reduced,claudin-3 and claudin-4 were increased(P<0.05),There was no difference between pulmonary respiration index and PaO2/FiO2(P>0.05),Western blot demonstrated that the expression of claudin-3 and claudin-4 were rised,P-JNK,P-P38 and NF-κB were deincreased(P<0.05).There was no difference in above index between HSH group and LR+SAHA group(P>0.05).Conclusion Hemorrhagic shock on plateau can activate the expression of MPO,TNF-a and MDA in lung firstly,which subsequently resulting in the loss of claudin-3 and claudin-4 in lung epithelial cells,The barrier function of lung epithelial is disturbed,Eventually evolved into ALI.Different resuscitation programs can display different lung protection to a certain extent,with SAHA combined with hypertonic sodium chloride hydroxyethyl starch 40 solution being the best among the liquids under examination.The mechanism is associated with the suppression of TLR4-MAPK and NF-κB signal transduction pathways.