水蛭素对博莱霉素致特发性肺间质纤维化大鼠肺组织中AKT、p-AKT蛋白表达的影响
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摘要
目的:本文以水蛭素为研究药物,通过Western blot法测定蛋白激酶B(AKT)、p-AKT蛋白在肺中的表达,分析水蛭素对特发性肺间质纤维化(Idiopathic Pulmonary Fibrosis,IPF)的干预效果及作用机制,为水蛭素的临床运用奠定基础,并拓宽IPF的治疗途径。方法:选取SPF级Wistar大鼠180只,随机选18只为空白组(K组),剩余大鼠统一造模(气管内注射博莱霉素),选取108只造模成功大鼠,随机分为模型组(M组)、激素组(J组)、低剂量组(D组)、高剂量组(G组)、中剂量组(Z组)、中剂量联合激素组(ZJ组),每组18只,初次给药为造模后第2天,在给药的第7 d、14 d、28 d分批处死取材,检测AKT、p-AKT蛋白的表达。结果:AKT蛋白表达量:K组在各时间点AKT蛋白表达量均最低,M组在7 d、28 d含量明显高于J组(P <0.01);7 d时G组低于J组(P <0.01),14、28 d时G、ZJ组均低于J组(P <0.01);7 d、28 d时D、Z组高于J组(P <0.01)。p-AKT蛋白表达量:各时间点K组最低,M组最高;J组低于M组(P <0.01);ZJ组低于J组(P <0.01);7 d、14 d时G组低于J组(P <0.01);28 d时G组高于J组,但P> 0.05。结论:IPF发病过程中,PI3K/AKT信号通路中AKT、p-AKT蛋白表达明显升高,说明该信号通路与IPF发病过程有明显相关性;水蛭素能降低AKT、p-AKT蛋白的表达量,减缓IPF病程发展;水蛭素极有可能通过PI3K/AKT通路作用于IPF。
Objective: Hirudin as a research drug, the expression of AKT and p-AKT protein in the lung were determined by Western blot. The intervention effect and mechanism of hirudin on IPF were analyzed, which laid a foundation for the clinical application of hirudin and broadened the therapeutic approach of IPF. Methods: 180 SPF-level Wistar rats were selected. 18 rats were randomly selected as the blank group(K group). The remaining rats were uniformly modeled(intratracheal injection of bleomycin); 108 rats were randomly divided into the model group(M group), hormone group(J group), hirudin low dose group(D group), high dose group(G group), medium dose group(Z group), and middle dose plus hormone group(ZJ group), 18 rats in each group. On the 2 days after modeling, rats were given first dose medicine. The samples were killed in batches on the 7 th, 14 th, and 28 th day after administration, and the expression of AKT and p-AKT protein were detected. Results: The expression of AKT protein was the lowest in K group at each time point; at 7 days and 28 days,that in M group was significantly higher than J group(P<0.01); at 7 days, that in G group was lower than J group(P<0.01); at the 14 th and28 th day, that in G group and ZJ group was lower than J group(P<0.01); at 7 days and 28 days, that in the D group and Z group was higher than J group(P<0.01). The p-AKT protein expression was the lowest in K group and the highest in M group at each time point; that in J group was lower than M group(P<0.01); that in ZJ group was lower than J group(P<0.01); that in G group was lower than J group at 7 day and 14 day(P<0.01); at 28 days, that in G group was higher than J group, P>0.05. Conclusion: During the pathogenesis of IPF, the expression of AKT and p-AKT protein in PI3 K/AKT signaling pathway was significantly increased, indicating that the signaling pathway is significantly correlated with the pathogenesis of IPF. Hirudin can reduce the expression of AKT and p-AKT protein and slow down The progression of IPF disease; hirudin is highly likely to act on IPF through the PI3 K/AKT pathway.
Objective: Hirudin as a research drug, the expression of AKT and p-AKT protein in the lung were determined by Western blot. The intervention effect and mechanism of hirudin on IPF were analyzed, which laid a foundation for the clinical application of hirudin and broadened the therapeutic approach of IPF. Methods: 180 SPF-level Wistar rats were selected. 18 rats were randomly selected as the blank group(K group). The remaining rats were uniformly modeled(intratracheal injection of bleomycin); 108 rats were randomly divided into the model group(M group), hormone group(J group), hirudin low dose group(D group), high dose group(G group), medium dose group(Z group), and middle dose plus hormone group(ZJ group), 18 rats in each group. On the 2 days after modeling, rats were given first dose medicine. The samples were killed in batches on the 7 th, 14 th, and 28 th day after administration, and the expression of AKT and p-AKT protein were detected. Results: The expression of AKT protein was the lowest in K group at each time point; at 7 days and 28 days,that in M group was significantly higher than J group(P<0.01); at 7 days, that in G group was lower than J group(P<0.01); at the 14 th and28 th day, that in G group and ZJ group was lower than J group(P<0.01); at 7 days and 28 days, that in the D group and Z group was higher than J group(P<0.01). The p-AKT protein expression was the lowest in K group and the highest in M group at each time point; that in J group was lower than M group(P<0.01); that in ZJ group was lower than J group(P<0.01); that in G group was lower than J group at 7 day and 14 day(P<0.01); at 28 days, that in G group was higher than J group, P>0.05. Conclusion: During the pathogenesis of IPF, the expression of AKT and p-AKT protein in PI3 K/AKT signaling pathway was significantly increased, indicating that the signaling pathway is significantly correlated with the pathogenesis of IPF. Hirudin can reduce the expression of AKT and p-AKT protein and slow down The progression of IPF disease; hirudin is highly likely to act on IPF through the PI3 K/AKT pathway.
引文
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[10]李晓娟,张骞云,蔡志刚,等.水蛭对肺纤维化大鼠羟脯氨酸、转化生长因子-β1及纤溶酶原激活物抑制因子-1的影响[J].安徽中医药大学学报,2013,32(03):57-61.
[11]李晓娟,张骞云,蔡志刚,等.水蛭对肺纤维化大鼠PAI-1的影响[J].贵州医科大学学报,2012,37(02):170-173.
[12]Yutaro N,Takafumi S.Idiopathic Pulmonary Fibrosis:Diagnosis and Clinical Manifestations[J].Clin Med Insights Circ Respir&Pulm Med,2015,9(Suppl 1):163-171.
[13]Raghu G,Collard HR,Egan JJ,et al.An Official ATS/ERS/JRS/ALAT Statement:Idiopathic Pulmonary Fibrosis:Evidence-based Guidelines for Diagnosis and Management[J].Am J Respir Crit Care Med,2011,183(06):788-824.
[14]Antoniou KM,Margaritopoulos GA,Soufla G,et al.Expression analysis of Akt and MAPK signaling pathways in lung tissue of patients with idiopathic pulmonary fibrosis(IPF)[J].Journal of Receptor&Signal Transduction Research,2010,30(04):262-269.
[15]Hardie W D,Hagood J S,Dave V,et al.Signaling pathways in the epithelial origins of pulmonary fibrosis[J].Cell Cycle,2010,9(14):2769.
[16]杜义安,余鹏飞.TOR信号通路与胃癌研究进展[J].中国肿瘤杂志,2014,23(09):761-764.
[17]陈德兴,张玉萍,徐丽莉,等.神农本草经[M].福州:福建科学技术出版社,2012:108.
[18]刘玉梅,章军,匙峰,等.水蛭化学成分研究进展[J].中国中医药信息杂志,2011,18(12):108-110.
[19]周乐,赵文静,常惟智.水蛭的药理作用及临床应用研究进展[J].中医药信息,2012,29(01):132-133.
[20]李晶.水蛭的药理研究进展[J].科学技术创新,2012,16(30):33.
[21]李晓娟,张骞云,马洪霞,等.水蛭对肺纤维化大鼠的影响及机制探讨[J].中药药理与临床,2015,31(02):155-156.
[22]李校天,杨书良,王军民,等.水蛭对Ang-Ⅱ刺激鼠肝星状细胞活化Ca2+效应的抑制作用[J]中国全科医学,2006,9(06):472-474.
[23]杨馨,刘娜,张佩青.水蛭对FSGS大鼠肾脏ECM积聚影响的实验研究[J].中国中西医结合肾病杂志,2017,18(10):891-893,943-944.
[24]郭睿,农晓琳,邓凌,等.水蛭素对皮肤增生性瘢痕成纤维细胞bFGF及TGFβ1表达的影响[J].中国美容医学,2011,20(04):614-617.
[25]郑燕林,王毅,武文忠,等.水蛭素对血小板源性生长因子及受体的影响[J].中华实验眼科杂志,2003,21(05):476-478.
[26]郑怡,禤坤,南澜,等.水蛭素对牙龈成纤维细胞碱性成纤维细胞生长因子和转化生长因子-β1表达的影响[J].华西口腔医学杂志,2015,33(01):6-10.
[27]梁桂文,姜敏辉.水蛭素对动脉内皮保护作用的实验研究[J].天津医药,2012,40(12):1234-1236.
[2]Raghu G,Collard H R,Egan J J,et al.An Official ATS/ERS/JRS/ALAT Statement:Idiopathic Pulmonary Fibrosis:Evidence-based Guidelines for Diagnosis and Management[J].Am J Respir Crit Care Med,2011,183(06):788-824.
[3]李宏林.TMEM16A在小鼠肺水清除、肺组织凋亡中的作用及PI3K/Akt通路表达调控的研究[D].石家庄:河北医科大学,2015.
[4]樊茂蓉,张燕萍,苗青,等.肺间质纤维化的中医发病机制与治则探讨[J].中医杂志,2012,53(03):203-204.
[5]闫蔷薇,王至婉.活血化瘀方治疗特发性肺纤维化的有效性和安全性Meta分析[J].中医研究,2017,30(09):51-55.
[6]李亚,胡金亮,李素云,等.基于数据挖掘的弥漫性肺间质疾病中医证候诊断模型建立研究[J].辽宁中医杂志,2010,37(12):2333-2335.
[7]任玉娇,朱雪,张伟.益气温阳活血化瘀法论治特发性肺纤维化合并肺动脉高压[J].中医杂志,2017,58(14):1186-1188.
[8]王海霞,李红,谢海彬,等.特发性肺间质纤维化的研究进展[J].中医临床研究,2017,9(30):9-12.
[9]李晓娟.水蛭对大鼠肺纤维化模型干预作用及机制的研究[D].石家庄:河北医科大学,2007.
[10]李晓娟,张骞云,蔡志刚,等.水蛭对肺纤维化大鼠羟脯氨酸、转化生长因子-β1及纤溶酶原激活物抑制因子-1的影响[J].安徽中医药大学学报,2013,32(03):57-61.
[11]李晓娟,张骞云,蔡志刚,等.水蛭对肺纤维化大鼠PAI-1的影响[J].贵州医科大学学报,2012,37(02):170-173.
[12]Yutaro N,Takafumi S.Idiopathic Pulmonary Fibrosis:Diagnosis and Clinical Manifestations[J].Clin Med Insights Circ Respir&Pulm Med,2015,9(Suppl 1):163-171.
[13]Raghu G,Collard HR,Egan JJ,et al.An Official ATS/ERS/JRS/ALAT Statement:Idiopathic Pulmonary Fibrosis:Evidence-based Guidelines for Diagnosis and Management[J].Am J Respir Crit Care Med,2011,183(06):788-824.
[14]Antoniou KM,Margaritopoulos GA,Soufla G,et al.Expression analysis of Akt and MAPK signaling pathways in lung tissue of patients with idiopathic pulmonary fibrosis(IPF)[J].Journal of Receptor&Signal Transduction Research,2010,30(04):262-269.
[15]Hardie W D,Hagood J S,Dave V,et al.Signaling pathways in the epithelial origins of pulmonary fibrosis[J].Cell Cycle,2010,9(14):2769.
[16]杜义安,余鹏飞.TOR信号通路与胃癌研究进展[J].中国肿瘤杂志,2014,23(09):761-764.
[17]陈德兴,张玉萍,徐丽莉,等.神农本草经[M].福州:福建科学技术出版社,2012:108.
[18]刘玉梅,章军,匙峰,等.水蛭化学成分研究进展[J].中国中医药信息杂志,2011,18(12):108-110.
[19]周乐,赵文静,常惟智.水蛭的药理作用及临床应用研究进展[J].中医药信息,2012,29(01):132-133.
[20]李晶.水蛭的药理研究进展[J].科学技术创新,2012,16(30):33.
[21]李晓娟,张骞云,马洪霞,等.水蛭对肺纤维化大鼠的影响及机制探讨[J].中药药理与临床,2015,31(02):155-156.
[22]李校天,杨书良,王军民,等.水蛭对Ang-Ⅱ刺激鼠肝星状细胞活化Ca2+效应的抑制作用[J]中国全科医学,2006,9(06):472-474.
[23]杨馨,刘娜,张佩青.水蛭对FSGS大鼠肾脏ECM积聚影响的实验研究[J].中国中西医结合肾病杂志,2017,18(10):891-893,943-944.
[24]郭睿,农晓琳,邓凌,等.水蛭素对皮肤增生性瘢痕成纤维细胞bFGF及TGFβ1表达的影响[J].中国美容医学,2011,20(04):614-617.
[25]郑燕林,王毅,武文忠,等.水蛭素对血小板源性生长因子及受体的影响[J].中华实验眼科杂志,2003,21(05):476-478.
[26]郑怡,禤坤,南澜,等.水蛭素对牙龈成纤维细胞碱性成纤维细胞生长因子和转化生长因子-β1表达的影响[J].华西口腔医学杂志,2015,33(01):6-10.
[27]梁桂文,姜敏辉.水蛭素对动脉内皮保护作用的实验研究[J].天津医药,2012,40(12):1234-1236.