The Protective Mechanism Of Recombinant Mouse UBC13 On LPS-induced Acute Lung Injury In Mice

Objective: To investigate the protective effect of recombinant mouse UBC13 on lipopolysaccharide(LPS)-induced acute lung injury(Acute Lung Injury,ALI)and its molecular mechanism.Methods:(1)The effect of recombinant mouse UBC13 on normal mice was analyzed: 60 SPF female Kunming mice were randomly divided into 4 groups: PBS control group,UBC13 groups(200 μg,100 μg,50μg).The mice were sacrificed by subcutaneous injection for 1 day,7 days and 13 days.The changes of body weight,organ index,pathological sections and serum inflammatory factors were detected.(2)To analyze the role of recombinant mouse UBC13 in the LPS model: 80 SPF female Kunming mice were randomly divided into 8 groups,which were set as control group,PBS control group,LPS group(20 mg/kg),UBC13 high dose group(100 μg),UBC13 medium dose group(50 μg),UBC13 low dose group(25 μg)dexamethasone positive control group(0.2 mg/kg)and CST11 negative control group(100 μg).The pathological tissue sections were used to observe the pathological changes of the lungs.The expression of cytokines was detected by real-time PCR and ELISA.The expression of key proteins in NF-κB signaling pathway was detected by Western Blot.The expression of CD4 and CD8 was detected by immunohistochemistry.Results:(1)The test results showed that injection UBC13 protein in mice of indicators compared witn PBS control group,no significant differences(P>0.05).(2)Histopathological observation showed that obvious lesions in the lung tissue of LPS group and CST negative control group,UBC13 treatment group and dexamethasone positive control group were alleviated;The results of ELISA,NO and MPO showed that the levels of IL-1β,TNF-α,NO and MPO in the LPS group were significantly higher than those in the control group(P<0.01).Compared with the LPS group,the UBC13 treatment group and the positive group were significantly lower(P<0.05),and the negative group no significantly different(P>0.05);The results of real-time PCR showed that the expression of IL-1β,TNF-α,IL-6,CXCL-1 and VCAM-1 m RNA in LPS group was significantly increased compared with the control group(P<0.01).Compared with the LPS group,the expression levels of IL-1β,TNF-α,IL-6 and VCAM-1 m RNA in UBC13 treatment group andpositive control group were significantly decreased(P<0.05),and there was no significant difference in CXCL-1 m RNA expression(P<0.05).The results of Wstern Blot showed that the expression levels of p-IκBα and p65 in LPS group was significantly increased compared with the control group(P<0.01),and the expression levels of IκBα and cytosolic p65 were significantly decreased(P<0.01).Compared with the LPS group,the expression levels of p-IκBα and nuclear p65 in the UBC13-treated group and the positive control group were significantly decreased(P<0.05),the expression levels of IκBα and cytosolic p65 were significantly increased(P<0.05),and there was no significant difference in the negative control group(P>0.05).The results of immunohistochemistry showed that the expression of CD4 and CD8 in the LPS group was significantly higher than that in the control group(P<0.01).Compared with the LPS group,the UBC13 treatment group and the positive control group were significantly lower(P<0.05),there was no significant difference in the negative control group(P>0.05).In the acute lung injury model,UBC13 treatment group can inhibit the activation of NF-κB signaling pathway,reduce the expression of cytokines and NO content,reduce the activity of MPO and the expression of CD4 and CD8 of which high dose of UBC13 group work well.Conclusion: The acute lung injury(ALI)model was successfully established,and the recombinant mouse UBC13 protein can inhibit the LPS-induced inflammatory response through NF-κB signaling pathway,which has a certain protective effect on acute lung injury.