大鼠肝损伤过程中肝组织神经营养因子神经突蛋白水平变化及其意义研究
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摘要
目的探讨大鼠肝损伤过程中肝组织神经营养因子神经突蛋白(Neuritin)水平变化及其意义。方法2015年11月—2016年11月,采用随机数字表法将清洁级SD大鼠126只分为空白组(42只)、非肝损伤组(42只)、肝损伤组(42只)。肝损伤组采用自制改良型BIM-IV生物撞击机建立肝损伤模型,非肝损伤组大鼠于同一部位给予同一力度致其股骨骨折;空白组不给予任何处置。分别于造模后6 h、12 h、1 d、2 d、3 d、5 d采用随机数字表法在各组选取7只大鼠,采集肝组织标本及血清,对肝组织进行HE染色,检测血清丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST)水平及肝组织Neuritin水平。结果肝损伤组造模后6 h时可见肝细胞排列不规则、胞质疏松化及气球样变;造模后12 h、1 d、2 d时可见肝细胞大片坏死甚至出现细胞核脱失;造模后3 d、5 d时可见肝细胞排列、胞质、细胞核大小形态接近正常。肝损伤组造模后6 h、12 h、1 d、2 d血清ALT、AST水平高于空白组、非肝损伤组(P<0.05)。肝损伤组造模后12 h血清ALT、AST水平高于造模后6 h,造模后2 d、3 d、5 d血清ALT、AST水平低于造模后6 h、12 h,造模后2 d、3 d、5 d血清ALT、AST水平低于造模后1 d,造模后3 d、5 d血清ALT、AST水平低于造模后2 d(P<0.05)。肝损伤组造模后6 h、12 h、1 d、2 d、3 d肝组织Neuritin水平高于空白组、非肝损伤组(P<0.05)。肝损伤组造模后12 h、1 d肝组织Neuritin水平高于造模后6 h,造模后3 d、5 d肝组织Neuritin水平低于造模后6 h,造模后1 d、2 d、3 d、5 d肝组织Neuritin水平低于造模后12 h,造模后2 d、3 d、5 d肝组织Neuritin水平低于造模后1 d,造模后3 d、5 d肝组织Neuritin水平低于造模后2 d,造模后5 d肝组织Neuritin水平低于造模后3 d(P<0.05)。肝损伤组肝组织Neuritin水平与血清ALT、AST水平均呈正相关(r=0.796 2,P<0.001;r=0.769 1,P<0.001)。结论大鼠肝损伤后,肝组织中神经营养因子Neuritin水平先升高再降低,与血清中ALT、AST水平变化一致,其可能在肝损伤后肝细胞的再生或增殖过程中发挥一定的作用。
Objective To investigate the changes in the expression level of neuritin(a neurotrophic factor) expressed in the liver during liver injury in rats and its significance. Methods From November 2015 to November 2016, a total of 126 healthy Sprague-Dawley rats were randomly divided into 3 groups, blank group, non-liver injury group and liver injury group with 42 in each. The liver injury model was established by a self-made modified BIM-IV biological impact machine in liver injury group, while rats of the non-liver injury group were hit with a uniform strength at the same part to cause a femoral fracture. Rats of the blank group did not receive any interventions. Seven rats were randomly selected from each group at 6 h, 12 h, 1 d, 2 d, 3 d and 5 d after the establishment of liver injury model to collect the liver specimens and serum samples. HE staining was used to detect the pathological changes in the liver tissue. Serum alanine aminotransferase(ALT) and aspartate aminotransferase(AST) were measured. Expression levels of Neuritin expressed in the liver tissue were measured with immunohistochemical staining.Results In the liver injury group, irregular arrangement of hepatocytes, loose and balloon-shaped cytoplasms were observed at 6 h after modeling; 12 h, 1 d, and 2 d after modeling, large areas of hepatocellular necrosis and even nuclear lysis were observed; while 3, 5 days after the modeling, it was found that the arrangement of hepatocytes, morphology and size of cytoplasm and nucleus were nearly became normal. At 6 h, 12 h, 1 d, 2 d after injury, liver injury group demonstrated higher serum ALT and AST levels compared with other groups(P<0.05). In the liver injury group, the levels of ALT and AST at 12 h after modeling were significantly higher compared with 6 h after modeling(P<0.05), but both of them were higher than the following measurement levels(at 2 d, 3 d and 5 d after modeling)(P<0.05), moreover, the third measurement levels were higher than the following three measurement levels(P<0.05), and the fourth measurement levels were higher than the latter two measurement levels(P<0.05). At 6 h, 12 h, 1 d, 2 d, 3 d after modeling, liver injury group demonstrated higher Neuritin expression levels compared with other groups(P<0.05). In the liver injury group, compared with at 6 h after modeling, Neuritin expression levels were higher at 12 h, 1 d after modeling, but lower at 3 d and 5 d after modeling(P<0.05); Neuritin expression level at 12 h after modeling was higher than that measured at each of the following four times(at 1 d, 2 d, 3 d and 5 d after modeling)(P<0.05), and it was higher at 1 d after modeling than that measured at each of the following three times(at 2 d, 3 d and 5 d after modeling)(P<0.05), likewise, it was higher at 2 d after modeling than that measured at each of the following two times(at 3 d and 5 d after modeling)(P<0.05), and higher at 3 d after modeling than that of the final measurement(P<0.05). Overall,the expression level of Neuritin in liver injury group was positively correlated with serum ALT(r=0.796 2, P<0.001) and AST(r=0.769 1, P<0.001). Conclusion After liver injury, the expression level of Neuritin in rats first increased then decreased, which changed with the same tendency as ALT and AST in serum. So, Neuritin may play a role in the regeneration or proliferation of hepatic cells after liver injury.
Objective To investigate the changes in the expression level of neuritin(a neurotrophic factor) expressed in the liver during liver injury in rats and its significance. Methods From November 2015 to November 2016, a total of 126 healthy Sprague-Dawley rats were randomly divided into 3 groups, blank group, non-liver injury group and liver injury group with 42 in each. The liver injury model was established by a self-made modified BIM-IV biological impact machine in liver injury group, while rats of the non-liver injury group were hit with a uniform strength at the same part to cause a femoral fracture. Rats of the blank group did not receive any interventions. Seven rats were randomly selected from each group at 6 h, 12 h, 1 d, 2 d, 3 d and 5 d after the establishment of liver injury model to collect the liver specimens and serum samples. HE staining was used to detect the pathological changes in the liver tissue. Serum alanine aminotransferase(ALT) and aspartate aminotransferase(AST) were measured. Expression levels of Neuritin expressed in the liver tissue were measured with immunohistochemical staining.Results In the liver injury group, irregular arrangement of hepatocytes, loose and balloon-shaped cytoplasms were observed at 6 h after modeling; 12 h, 1 d, and 2 d after modeling, large areas of hepatocellular necrosis and even nuclear lysis were observed; while 3, 5 days after the modeling, it was found that the arrangement of hepatocytes, morphology and size of cytoplasm and nucleus were nearly became normal. At 6 h, 12 h, 1 d, 2 d after injury, liver injury group demonstrated higher serum ALT and AST levels compared with other groups(P<0.05). In the liver injury group, the levels of ALT and AST at 12 h after modeling were significantly higher compared with 6 h after modeling(P<0.05), but both of them were higher than the following measurement levels(at 2 d, 3 d and 5 d after modeling)(P<0.05), moreover, the third measurement levels were higher than the following three measurement levels(P<0.05), and the fourth measurement levels were higher than the latter two measurement levels(P<0.05). At 6 h, 12 h, 1 d, 2 d, 3 d after modeling, liver injury group demonstrated higher Neuritin expression levels compared with other groups(P<0.05). In the liver injury group, compared with at 6 h after modeling, Neuritin expression levels were higher at 12 h, 1 d after modeling, but lower at 3 d and 5 d after modeling(P<0.05); Neuritin expression level at 12 h after modeling was higher than that measured at each of the following four times(at 1 d, 2 d, 3 d and 5 d after modeling)(P<0.05), and it was higher at 1 d after modeling than that measured at each of the following three times(at 2 d, 3 d and 5 d after modeling)(P<0.05), likewise, it was higher at 2 d after modeling than that measured at each of the following two times(at 3 d and 5 d after modeling)(P<0.05), and higher at 3 d after modeling than that of the final measurement(P<0.05). Overall,the expression level of Neuritin in liver injury group was positively correlated with serum ALT(r=0.796 2, P<0.001) and AST(r=0.769 1, P<0.001). Conclusion After liver injury, the expression level of Neuritin in rats first increased then decreased, which changed with the same tendency as ALT and AST in serum. So, Neuritin may play a role in the regeneration or proliferation of hepatic cells after liver injury.
引文
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[12]KOJIMA N,SHIOJIRI N,SAKAI Y,et al.Expression of neuritin during liver maturation and regeneration[J].FEBS Lett,2005,579(21):4562-4566.DOI:10.1016/j.febslet.2005.07.015.
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[14]RAGGO C,RUHL R,MCALLISTER S,et al.Novel cellular genes essential for transformation of endothelial cells by Kaposi's sarcomaassociated herpesvirus[J].Cancer Res,2005,65(12):5084-5095.DOI:10.1158/0008-5472.CAN-04-2822.
[15]雷箴,温韬.松果菊苷对刀豆蛋白A所致急性肝损伤小鼠的保护作用及对细胞外组蛋白的影响[J].解放军医学杂志,2016,41(2):97-102.DOI:10.11855/j.issn.0577-7402.2016.02.03.LEI Z,WEN T.Effect of echinacoside on the protection of acute liver injury induced by concanavalin A in mice and its effect on extracellular histones[J].Medical Journal of Chinese People's Liberation Army,2016,41(2):97-102.DOI:10.11855/j.issn.0577-7402.2016.02.03.
[16]吴思荣,王之敏,李向东,等.caspase-3在创伤性脑损伤诱导神经细胞凋亡中的作用[J].苏州大学学报(医学版),2007,27(1):49-51.DOI:10.3969/j.issn.1673-0399.2007.01.016.WU S R,WANG Z M,LI X D,et al.Effects of caspase-3 during traumatic brain injury for inducing neuronal apoptosis in rats[J].Journal of Soochow University(Medical Science Edition),2007,27(1):49-51.DOI:10.3969/j.issn.1673-0399.2007.01.016.
[17]KARAMOYSOYLI E,BURNAND R C,TOMLINSON DR,et al.Neuritin mediates nerve growth factor-induced axonal regeneration and is deficient in experimental diabetic neuropathy[J].Diabetes,2008,57(1):181-189.DOI:10.2337/db07-0895.
[18]ALME M N,WIBRAND K,DAGESTAD G,et al.Chronic fluoxetine treatment induces brain region-specific upregulation of genes associated with BDNF-induced long-term potentiation[J].Neural Plast,2007,2007:26496.DOI:10.1155/2007/26496.
[2]陈伯棠,刘臻玉,黄擎雄,等.90例肝外伤手术预后的临床分析[J].中国临床实用医学,2010,4(1):84-85.DOI:10.3760/cma.j.issn.1673-8799.2010.01.46.CHEN B T,LIU Z Y,HUANG Q X,et al.The analysis about the clinical prognosis of 90 patients with blunt hepatic injuries after surgieal management[J].China Clinical Practical Medicine,2010,4(1):84-85.DOI:10.3760/cma.j.issn.1673-8799.2010.01.46.
[3]MOORE E E,SHACKFORD S R,PACHTER H L,et al.Organ injury scaling:spleen,liver,and kidney[J].J Trauma,1989,29(12):1664-1666.
[4]杨龙,张雅敏,崔子林,等.肝细胞生长因子激活抑制因子在肝硬化大鼠部分肝切除后肝再生过程中的表达[J].中华肝胆外科杂志,2015,21(5):324-327.DOI:10.3760/cma.j.issn.1007-8118.2015.05.011.YANG L,ZHANG Y M,CUI Z L,et al.HGFA and its inhibitors manifested differential expressions during liver regeneration after partial Hepatectomy in cirrhotic rat model[J].Chinese Journal of Hepatobiliary Surgery,2015,21(5):324-327.DOI:10.3760/cma.j.issn.1007-8118.2015.05.011.
[5]FRIEDMAN S L.Seminars in medicine of the Beth Israel Hospital,Boston.The cellular basis of hepatic fibrosis.Mechanisms and treatment strategies[J].N Engl J Med,1993,328(25):1828-1835.DOI:10.1056/NEJM199306243282508.
[6]WIBRAND K,MESSAOUDI E,H?VIK B,et al.Identification of genes co-upregulated with Arc during BDNF-induced long-term potentiation in adult rat dentate gyrus in vivo[J].Eur J Neurosci,2006,23(6):1501-1511.DOI:10.1111/j.1460-9568.2006.04687.x.
[7]NEDIVI E,HEVRONI D,NAOT D,et al.Numerous candidate plasticity-related genes revealed by differential c DNA cloning[J].Nature,1993,363(6431):718-722.DOI:10.1038/363718a0.
[8]NEDIVI E,FIELDUST S,THEILL L E,et al.A set of genes expressed in response to light in the adult cerebral cortex and regulated during development[J].Proc Natl Acad Sci U S A,1996,93(5):2048-2053.
[9]李晓天,赵冬,代林志,等.Neuritin对大鼠蛛网膜下腔出血后早期脑损伤影响的研究[J].中国全科医学,2016,19(24):2908-2915.DOI:10.3969/j.issn.1007-9572.2016.24.008.LI X T,ZHAO D,DAI L Z,et al.Effects of Neuritin in early brain injury after subarachnoid hemorrhage of rats[J].Chinese General Practice,2016,19(24):2908-2915.DOI:10.3969/j.issn.1007-9572.2016.24.008.
[10]李维嘉,钟晨,王锦国,等.Neuritin在胃癌中的表达水平及意义[J].重庆医学,2014,43(22):2845-2847.DOI:10.3969/j.issn.1671-8348.2014.22.005.LI W J,ZHONG C,WANG J G,et al.Expression of Neuritin and its clinical significance in gastric cancer[J].Chongqing Medicine,2014,43(22):2845-2847.DOI:10.3969/j.issn.1671-8348.2014.22.005.
[11]刘京敏,董金波,王维山,等.不同途径给予Neuritin对大鼠骨折愈合的影响[J].重庆医学,2013,42(10):1128-1131,1133.DOI:10.3969/j.issn.1671-8348.2013.10.018.LIU J M,DONG J B,WANG W S,et al.Effect of Neuritin given in different ways on promoting rats frcture healing[J].Chongqing Medicine,2013,42(10):1128-1131,1133.DOI:10.3969/j.issn.1671-8348.2013.10.018.
[12]KOJIMA N,SHIOJIRI N,SAKAI Y,et al.Expression of neuritin during liver maturation and regeneration[J].FEBS Lett,2005,579(21):4562-4566.DOI:10.1016/j.febslet.2005.07.015.
[13]NEDIVI E,WU G Y,CLINE H T.Promotion of dendritic growth by CPG15,an activity-induced signaling molecule[J].Science,1998,281(5384):1863-1866.
[14]RAGGO C,RUHL R,MCALLISTER S,et al.Novel cellular genes essential for transformation of endothelial cells by Kaposi's sarcomaassociated herpesvirus[J].Cancer Res,2005,65(12):5084-5095.DOI:10.1158/0008-5472.CAN-04-2822.
[15]雷箴,温韬.松果菊苷对刀豆蛋白A所致急性肝损伤小鼠的保护作用及对细胞外组蛋白的影响[J].解放军医学杂志,2016,41(2):97-102.DOI:10.11855/j.issn.0577-7402.2016.02.03.LEI Z,WEN T.Effect of echinacoside on the protection of acute liver injury induced by concanavalin A in mice and its effect on extracellular histones[J].Medical Journal of Chinese People's Liberation Army,2016,41(2):97-102.DOI:10.11855/j.issn.0577-7402.2016.02.03.
[16]吴思荣,王之敏,李向东,等.caspase-3在创伤性脑损伤诱导神经细胞凋亡中的作用[J].苏州大学学报(医学版),2007,27(1):49-51.DOI:10.3969/j.issn.1673-0399.2007.01.016.WU S R,WANG Z M,LI X D,et al.Effects of caspase-3 during traumatic brain injury for inducing neuronal apoptosis in rats[J].Journal of Soochow University(Medical Science Edition),2007,27(1):49-51.DOI:10.3969/j.issn.1673-0399.2007.01.016.
[17]KARAMOYSOYLI E,BURNAND R C,TOMLINSON DR,et al.Neuritin mediates nerve growth factor-induced axonal regeneration and is deficient in experimental diabetic neuropathy[J].Diabetes,2008,57(1):181-189.DOI:10.2337/db07-0895.
[18]ALME M N,WIBRAND K,DAGESTAD G,et al.Chronic fluoxetine treatment induces brain region-specific upregulation of genes associated with BDNF-induced long-term potentiation[J].Neural Plast,2007,2007:26496.DOI:10.1155/2007/26496.