HO-1和HO-2在老年SD大鼠慢性腦血流低灌注狀態(tài)下肝臟中的動態(tài)變化及意義
發(fā)布時間：2018-08-26 17:58
【摘要】：目的:長期的腦血流灌注不足,會導(dǎo)致體內(nèi)多臟器內(nèi)過度的氧化應(yīng)激的產(chǎn)生。血紅素加氧酶(heme oxygenase,HO)是血紅素代謝分解過程中的關(guān)鍵酶和限速酶,也是體內(nèi)重要的內(nèi)源性抗氧化體系,具有顯著的抗氧化應(yīng)激能力。本課題通過建立老年SD大鼠腦血流低灌注模型,研究HO家族中重要成員HO-1和HO-2在肝臟中的表達(dá),并探討其在腦血流灌注不足引起的損傷時對肝臟的作用及意義。方法:采用持久性雙側(cè)頸總動脈結(jié)扎法(2VO)致老年SD大鼠腦血流灌注不足,首先檢測腦皮質(zhì)血流的變化情況;采用蘇木素-伊紅染色觀察大鼠肝細(xì)胞形態(tài)學(xué)改變;采集下腔靜脈的靜脈血檢測血清中丙氨酸氨基轉(zhuǎn)移酶(ALT)和天門冬氨酸氨基轉(zhuǎn)移酶(AST)的活性;采用ELISA法檢測肝臟組織中HO的活性變化;采用免疫組織化學(xué)法檢測肝臟組織中HO-1和HO-2的表達(dá)情況;并分別通過RT-PCR和Western blot法檢測肝臟組織中HO-1和HO-2在mRNA和蛋白水平的表達(dá)。結(jié)果:術(shù)后第7天,大鼠皮質(zhì)血流的面積比值明顯降低,達(dá)到最低值,而術(shù)后14、21、28天,大鼠皮質(zhì)腦血流的面積比值逐漸升高,到28天基本恢復(fù)的正常水平。HE染色結(jié)果表明:與假手術(shù)組比,在術(shù)后第1天,肝細(xì)胞因缺血出現(xiàn)水樣變性,即細(xì)胞體積明顯增大,胞質(zhì)變得透明,第7天其水樣變性最為顯著;在術(shù)后14、21、28天,肝細(xì)胞的水樣變性有所改善,但仍未恢復(fù)到正常水平。肝功能檢測結(jié)果顯示:ALT和AST在2VO術(shù)后7 d明顯增高,差異有統(tǒng)計學(xué)意義(P0.01),但增高范圍仍屬于正常,無臨床意義;隨著時間的延長,ALT和AST的含量逐漸降低,到28 d,數(shù)值基本恢復(fù)至假手術(shù)組的水平。免疫組化、RT-PCR和Western blot顯示:與假手術(shù)組比較,HO-1在術(shù)后1d即有增加,并在第7d達(dá)到高峰(p0.05),這樣的變化與HO酶活性變化一致;而HO-2在慢性腦血流灌注不足的肝臟中卻沒有顯著變化。結(jié)論:慢性腦血流灌注不足時,可導(dǎo)致肝細(xì)胞的形態(tài)結(jié)構(gòu)有輕微的損傷,但肝功能基本正常,這與HO-1和血紅素加氧酶活性的動態(tài)增加密切相關(guān),這提示HO-1在慢性腦血流灌注不足的肝臟中起著保護(hù)作用。
[Abstract]:Objective: chronic cerebral blood perfusion deficiency may lead to excessive oxidative stress in multiple organs in vivo. Heme oxygenase (heme oxygenase,HO) is a key enzyme and rate-limiting enzyme in heme catabolism and an important endogenous antioxidant system in vivo. The purpose of this study was to study the expression of HO-1 and HO-2 in the liver of aged SD rats by establishing a model of cerebral blood flow hypoperfusion, and to explore the role and significance of HO-1 and HO-2 in liver injury caused by cerebral perfusion insufficiency. Methods: chronic bilateral common carotid artery ligation (2VO) was used to induce cerebral blood perfusion insufficiency in aged SD rats. The changes of cerebral cortical blood flow were detected by hematoxylin and eosin staining, and the morphological changes of hepatic cells were observed by hematoxylin eosin staining. The activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in serum were detected by collecting venous blood from inferior vena cava, and the changes of HO activity in liver tissue were detected by ELISA method. The expression of HO-1 and HO-2 in liver tissue was detected by immunohistochemical method, and the expression of HO-1 and HO-2 in liver tissue was detected by RT-PCR and Western blot methods respectively. Results: on the 7th day after operation, the area ratio of cortical blood flow decreased significantly to the lowest value in rats, while the area ratio of cerebral blood flow increased gradually on day 14, 21 and 28 after operation. The results of HE staining showed that, compared with the sham operation group, on the first day after operation, the hepatocytes showed water-like degeneration due to ischemia, that is, the cell volume was obviously increased, the cytoplasm became transparent, and the water-like degeneration was the most obvious on the 7th day. On the 14th day, 21 ~ (th) day after operation, the degeneration of hepatocytes was improved, but it did not return to normal level. The results of liver function test showed that the levels of alt and AST increased significantly 7 days after 2VO, the difference was statistically significant (P0.01), but the range of increase was normal and had no clinical significance, and the contents of alt and AST decreased gradually with the prolongation of time. By 28 days, the value basically returned to the level of the sham operation group. Immunohistochemistry RT-PCR and Western blot showed that compared with sham-operated group, the expression of ro-HO-1 increased on the 1st day after operation and reached its peak on the 7th day (p0.05), which was consistent with the change of HO enzyme activity, but HO-2 did not change significantly in the liver of chronic cerebral blood perfusion deficiency. Conclusion: chronic cerebral blood perfusion deficiency can cause slight damage to the morphology and structure of hepatocytes, but the liver function is basically normal, which is closely related to the dynamic increase of HO-1 and heme oxygenase activities. This suggests that HO-1 plays a protective role in the liver with chronic cerebral blood perfusion deficiency.
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R743
本文編號：2205713
[Abstract]:Objective: chronic cerebral blood perfusion deficiency may lead to excessive oxidative stress in multiple organs in vivo. Heme oxygenase (heme oxygenase,HO) is a key enzyme and rate-limiting enzyme in heme catabolism and an important endogenous antioxidant system in vivo. The purpose of this study was to study the expression of HO-1 and HO-2 in the liver of aged SD rats by establishing a model of cerebral blood flow hypoperfusion, and to explore the role and significance of HO-1 and HO-2 in liver injury caused by cerebral perfusion insufficiency. Methods: chronic bilateral common carotid artery ligation (2VO) was used to induce cerebral blood perfusion insufficiency in aged SD rats. The changes of cerebral cortical blood flow were detected by hematoxylin and eosin staining, and the morphological changes of hepatic cells were observed by hematoxylin eosin staining. The activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in serum were detected by collecting venous blood from inferior vena cava, and the changes of HO activity in liver tissue were detected by ELISA method. The expression of HO-1 and HO-2 in liver tissue was detected by immunohistochemical method, and the expression of HO-1 and HO-2 in liver tissue was detected by RT-PCR and Western blot methods respectively. Results: on the 7th day after operation, the area ratio of cortical blood flow decreased significantly to the lowest value in rats, while the area ratio of cerebral blood flow increased gradually on day 14, 21 and 28 after operation. The results of HE staining showed that, compared with the sham operation group, on the first day after operation, the hepatocytes showed water-like degeneration due to ischemia, that is, the cell volume was obviously increased, the cytoplasm became transparent, and the water-like degeneration was the most obvious on the 7th day. On the 14th day, 21 ~ (th) day after operation, the degeneration of hepatocytes was improved, but it did not return to normal level. The results of liver function test showed that the levels of alt and AST increased significantly 7 days after 2VO, the difference was statistically significant (P0.01), but the range of increase was normal and had no clinical significance, and the contents of alt and AST decreased gradually with the prolongation of time. By 28 days, the value basically returned to the level of the sham operation group. Immunohistochemistry RT-PCR and Western blot showed that compared with sham-operated group, the expression of ro-HO-1 increased on the 1st day after operation and reached its peak on the 7th day (p0.05), which was consistent with the change of HO enzyme activity, but HO-2 did not change significantly in the liver of chronic cerebral blood perfusion deficiency. Conclusion: chronic cerebral blood perfusion deficiency can cause slight damage to the morphology and structure of hepatocytes, but the liver function is basically normal, which is closely related to the dynamic increase of HO-1 and heme oxygenase activities. This suggests that HO-1 plays a protective role in the liver with chronic cerebral blood perfusion deficiency.
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R743
【參考文獻(xiàn)】
相關(guān)期刊論文 前1條
1 Gi-Sang Bae;Min-Sun Kim;Kyoung-Chel Park;Bon Soon Koo;Il-Joo Jo;Sun Bok Choi;Dong-Sung Lee;Youn-Chul Kim;Tae-Hyeon Kim;Sang-Wan Seo;Yong Kook Shin;Ho-Joon Song;Sung-Joo Park;;Effect of biologically active fraction of Nardostachys jatamansi on cerulein-induced acute pancreatitis[J];World Journal of Gastroenterology;2012年25期
，本文編號：2205713
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