肿瘤坏死因子α诱导蛋白家族8样蛋白2在小鼠内毒素性急性肺损伤中的作用

Role of TIPE2 in endotoxin-induced acute lung injury in mice

目的评价肿瘤坏死因子α诱导蛋白家族8样蛋白2(TIPE2)在小鼠内毒素性急性肺损伤(ALI)中的作用。方法SPF级健康成年雄性BALB/c小鼠40只,6~8周龄,体重20~25 g,采用随机数字表法分为4组(n=10):空载质粒组(VP组)、空载质粒组+ALI组(VP+ALI组)、腺相关病毒过表达TIPE2组(T组)和腺相关病毒过表达TIPE2组+ALI组(T+ALI组)。VP组和VP+ALI组气管内注射空载腺相关病毒,T组和T+ALI组气管内给予携带TIPE2干扰序列的腺相关病毒,3周后制备小鼠内毒素性ALI模型。VP组和T组气管内注射等容量PBS,VP+ALI组和T+ALI组气管内注射LPS 5 mg/kg。各组于注射LPS后24 h时采集腹主动脉血样,行血气分析并计算氧合指数(OI),采用ELISA法检测血清TNF-α浓度,随后处死小鼠取肺组织,HE染色观察病理学结果并行肺损伤评分,确定肺湿重/干重(W/D)比值,测定髓过氧化物酶(MPO)活性,采用Western blot法检测TIPE2、磷酸化c-Jun氨基末端激酶(p-JNK)和NF-κB的表达。结果与VP组比较,VP+ALI组肺损伤评分、W/D比值、MPO活性和血清TNF-α浓度升高,PaO2和OI降低,肺组织TIPE2表达下调,p-JNK和NF-κB表达上调(P<0.05),T组上述指标差异无统计学意义(P>0.05);与VP+ALI组比较,T+ALI组肺损伤评分、W/D比值、MPO活性和血清TNF-α浓度降低,PaO2和OI升高,肺组织TIPE2表达上调,p-JNK和NF-κB表达下调(P<0.05)。结论TIPE2表达下调参与了小鼠内毒素性ALI的过程。

Objective To evaluate the role of tumour necrosis factor-α-induced protein 8-like 2(TIPE2)in the acute lung injury(ALI)induced by endotoxin in mice.Methods Forty SPF healthy adult male BALB/c mice,aged 6-8 weeks,weighing 20-25 g,were divided into 4 groups(n=10 each)using a random number table method:vehicle plasmid group(VP group),vehicle plasmid plus ALI group(VP+ALI group),TIPE2 adeno-associated virus overexpression group(T group)and TIPE2 adeno-associated virus overexpression plus ALI group(T+ALI group).The mice in VP and VP+ALI groups were injected with empty adeno-associated virus,while the mice in T and T+ALI groups were intratracheally given adeno-associated virus carrying TIPE interference sequence.Three weeks later,the model of endotoxin-induced ALI was established.Lipopolysaccharide(LPS)5 mg/kg was intratracheally given in VP+ALI and T+ALI groups,and the equal volume of phosphate buffered saline(PBS)was given in VP and T groups.Blood samples were obtained from the abdominal aorta at 24 h after injection of LPS for blood gas analysis,oxygenation index(OI)was calculated,and tumor necrosis factor-alpha(TNF-α)in serum were detected by enzyme-linked immunosorbent assay.The animals were then sacrificed,and lung tissues were removed for examination of pathological changes which were scored after haematoxylin and eosin staining,for calculation of the wet/dry weight ratio(W/D ratio)and for determination of myeloperoxidase(MPO)activity and the expression of TIPE2,phosphorylated c-Jun N-terminal kinase(p-JNK)and nuclear factor kappa B(NF-κB)(by Western blot).Results Compared with VP group,the lung injury score,W/D ratio,MPO activity and concentration of serum TNF-αwere significantly increased,PaO2 and OI were decreased,expression of TIPE2 was down-regulated and expression of p-JNK and NF-κB was up-regulated in VP+ALI group(P<0.05).Compared with VP+ALI group,the lung injury score,W/D ratio,MPO activity and concentration of serum TNF-αwere significantly decreased,PaO2 and OI were increased,expression of TIPE2 was up-regulated and expression of p-JNK and NF-κB was down-regulated in T+ALI group(P<0.05).Conclusion The down-regulation of TIPE2 expression is involved in the process of ALI induced by endotoxin in mice.