抑制法尼基焦磷酸合成酶改善心臟重塑作用的研究
發(fā)布時(shí)間：2018-08-02 11:27
【摘要】：第一部分:法尼基焦磷酸合成酶抑制劑改善心肌肥厚并影響連接蛋白重塑研究背景:心肌肥厚是心臟疾病中的一種病理改變,其可能與機(jī)體過(guò)度激活的神經(jīng)內(nèi)分泌體系和內(nèi)環(huán)境失穩(wěn)密切相關(guān)。心肌持續(xù)而過(guò)度的肥厚反應(yīng)將最終導(dǎo)致心臟功能減退及慢性充血性心力衰竭。在心衰形成的整個(gè)過(guò)程中,心臟重構(gòu)是一個(gè)動(dòng)態(tài)發(fā)展的過(guò)程,其間涉及到細(xì)胞凋亡、纖維化、脂肪堆積、線粒體異常、離子通道改變及信號(hào)轉(zhuǎn)導(dǎo)通路的變化等。以往的研究表明,法尼基焦磷酸合成酶(famesyl pyrophosphate synthase,FPPS)是甲羥戊酸途徑中的一種關(guān)鍵酶,其催化合成下游產(chǎn)物法尼基焦磷酸(Farnesyl pyrophosphate,FPP)和r{牛兒基r{牛兒基焦磷酸(Geranylgeranylpyrophosphat,GGPP)所參與的信號(hào)轉(zhuǎn)導(dǎo)通路直接參與心肌肥厚及心臟重構(gòu)的發(fā)生。心肌細(xì)胞的纖維化是心臟重構(gòu)中及其重要的特征,但其他如肌鈣蛋白、縫隙連接蛋白等的變化也間接地反應(yīng)了心肌重構(gòu)的程度與趨勢(shì)。目的:探討法尼基焦磷酸合成酶抑制劑在腹主動(dòng)脈縮窄術(shù)所致心肌肥厚的動(dòng)物模型中心肌重構(gòu)的影響與作用。方法:1.腹主動(dòng)脈縮窄術(shù)建立心肌肥厚大鼠模型并觀察手術(shù)效果;2.將腹主動(dòng)脈縮窄術(shù)后雄性SD大鼠隨機(jī)分為3組:腹主動(dòng)脈縮窄術(shù)組、腹主動(dòng)脈縮窄術(shù)加藥物組和假手術(shù)對(duì)照組。分別給予生理鹽水10mg/kg/day或阿倫磷酸鈉10mg/kg/day 灌胃維持 3、5、8 周。3.運(yùn)用超聲、血流動(dòng)力學(xué)等手段檢測(cè)大鼠組織形態(tài)學(xué)變化;4.脛骨尺測(cè)量大鼠左側(cè)脛骨長(zhǎng)度;5.高效液相色譜法(HPLC)技術(shù)對(duì)大鼠心肌組織中FPP和GGPP含量進(jìn)行檢測(cè);6.用BNP檢測(cè)試劑盒檢測(cè)大鼠血液樣本中的BNP含量;7.對(duì)大鼠心肌組織切片染色并對(duì)I型纖維膠原蛋白(Col-I)、T型肌鈣蛋白(t-TnT)及縫隙連接蛋白(cx43)等進(jìn)行免疫組化分析。結(jié)果:1.腹主動(dòng)脈縮窄術(shù)導(dǎo)致大鼠心肌肥厚并增加心肌組織中FPP及GGPP的含量;2.FPPS抑制劑可部分改善心肌肥厚表現(xiàn),同時(shí)心肌組織中FPP及GGPP含量下降;3.心肌肥厚中大鼠脛骨長(zhǎng)度及血BNP水平無(wú)明顯變化;4.心肌重構(gòu)中心肌纖維化、肌鈣蛋白及縫隙連接蛋白等發(fā)生改變,抑制FPPS可以部分逆轉(zhuǎn)這種重構(gòu)表現(xiàn)。結(jié)論:FPPS參與了心肌肥厚及重塑的發(fā)生、發(fā)展;抑制FPPS的活性可以部分改善心肌肥厚。第二部分:法尼基焦磷酸合成酶抑制劑阿倫磷酸鈉對(duì)心肌肥厚大鼠中RhoA通路影響的研究研究背景:心臟重塑是心肌肥厚發(fā)生與發(fā)展中重要的病理過(guò)程,并以心力衰竭為最終結(jié)局。很多研究表明被激活的RhoA/ROCK通路及其相關(guān)蛋白會(huì)介導(dǎo)心肌肥厚及細(xì)胞凋亡的發(fā)生。Rho激酶屬于小G蛋白(蛋白激酶A,G,C)家族的絲氨酸/蘇氨酸激酶,可以通過(guò)其下游靶效應(yīng)分子,進(jìn)一步調(diào)節(jié)ROCK,這些蛋白的調(diào)節(jié)可以對(duì)細(xì)胞收縮、黏附、遷移和增殖等多種生物學(xué)行為產(chǎn)生影響。而已知的事實(shí)是,RhoA/Rock信號(hào)通路在心室重塑過(guò)程中起著上調(diào)炎性細(xì)胞因子,抑制心肌收縮,促進(jìn)心肌肥大與凋亡等作用。研究表明,阿倫磷酸鈉是經(jīng)典的FPPS抑制劑,其通過(guò)抑制FPP、GGPP的生成進(jìn)而影響RhoA/Rock信號(hào)通路的轉(zhuǎn)導(dǎo)達(dá)到改善心肌肥厚及纖維化等的病理過(guò)程。法尼基焦磷酸合成酶(FPPS)可能是治療心肌肥厚和改善心臟重構(gòu)的新思路。目的:探討FPPS抑制劑在改善心肌肥厚及重塑時(shí)可能的RhoA通路機(jī)制。方法:1.腹主動(dòng)脈縮窄術(shù)建立心肌肥厚大鼠模型;2.將雄性SD大鼠分為3組:腹主動(dòng)脈縮窄術(shù)組、腹主動(dòng)脈縮窄術(shù)加藥物組和假手術(shù)對(duì)照組。分別給予生理鹽水10mg/kg/day或阿倫酸納10mg/kg/day灌胃維持3、5、8 周。3.運(yùn)用超聲、血流動(dòng)力學(xué)等手段檢測(cè)大鼠的組織形態(tài)學(xué)變化;4.高效液相色譜法(HPLC)技術(shù)對(duì)大鼠心肌組織中FPP和GGPP含量進(jìn)行檢測(cè);5.RhoA試劑盒測(cè)定心肌組織中RhoA在490nm處的OD值;6.免疫蛋白印記檢測(cè)心肌組織中FDPS、RhoA及p-ERKl/2的蛋白含量。結(jié)果:1.抑制FPPS可改善心肌肥厚及心臟重塑;2.抑制FPPS可降低心肌組織中FPP、GGPP含量;3.抑制FPPS可降低心肌組織中RhoA含量;4.抑制FPPS可通過(guò)抑制RhoA相關(guān)通路的蛋白活性抑制心肌肥厚與心臟重塑。結(jié)論:抑制FPPS可通過(guò)抑制RhoA通路上蛋白的表達(dá)從而改善心肌肥厚及心臟重塑;FPPS抑制劑可能是治療心肌肥厚的新思路。
[Abstract]:The first part: the farnesyl pyrophosphate synthetase inhibitor improves the cardiac hypertrophy and affects the study of the remodeling of connexin: myocardial hypertrophy is a pathological change in the heart disease, which may be closely related to the excessive activation of the neuroendocrine system and the instability of the internal environment. The persistent and excessive hypertrophy of the heart muscle will eventually lead to the heart In the whole process of the formation of heart failure, cardiac remodeling is a process of dynamic development, which involves apoptosis, fibrosis, fat accumulation, mitochondrial abnormalities, changes in ion channels and signal transduction pathways. Previous studies showed that farnesyl pyrophosphate synthetase (famesyl Pyrophosphate synthase, FPPS) is a key enzyme in the metholate pathway, which catalyzes the synthesis of downstream product farnyl pyrophosphate (Farnesyl pyrophosphate, FPP) and r{bovine based r{bovine basic pyrophosphate (Geranylgeranylpyrophosphat, GGPP) involved in the direct involvement of the cardiac hypertrophy and cardiac remodeling. Cell fibrosis is an important feature of cardiac remodeling, but other changes such as troponin and gap connexin also indirectly reflect the degree and trend of cardiac remodeling. Objective: To investigate the effect of farnesyl pyrophosphate synthetase inhibitor on cardiac muscle remodeling in the animal model of cardiac hypertrophy induced by abdominal aortic coarctation. Methods: 1. abdominal aorta coarctation was used to establish the rat model of myocardial hypertrophy and to observe the effect of operation. 2. the male SD rats were randomly divided into 3 groups after the coarctation of the abdominal aorta: abdominal aorta coarctation group, abdominal aorta coarctation plus drug group and sham operation control group. The rats were given saline 10mg/kg/day or Allen phosphate sodium 10mg/kg/day irrigation respectively. Gastric maintenance 3,5,8 week.3. using ultrasound, hemodynamics and other means to detect the histomorphological changes in rats; 4. tibia length measurement of the length of the left tibia of rats; 5. high performance liquid chromatography (HPLC) technique to detect the content of FPP and GGPP in rat myocardial tissue; 6. BNP detection kit to detect the content of BNP in rat blood samples; 7. rats Myocardial tissue section staining and immunohistochemical analysis of I type fiber collagen (Col-I), T type troponin (t-TnT) and gap connexin (Cx43). Results: 1. abdominal aorta coarctation caused myocardial hypertrophy in rats and increased FPP and GGPP in myocardial tissue; 2.FPPS inhibitor could partly improve the performance of myocardial hypertrophy and myocardium. The content of FPP and GGPP in the tissue decreased, and there was no significant change in the length of tibia and blood BNP in 3. cardiac hypertrophy rats; 4. cardiac fibrosis, troponin and gap connexin were changed, and the inhibition of FPPS could partly reverse this reconstruction. Conclusion: FPPS is involved in the occurrence and development of myocardial hypertrophy and remodeling, and the inhibition of FPPS. Activity can partly improve myocardial hypertrophy. Second: the second part: the research background of the effect of farnesyl pyrophosphate synthase inhibitor Allen phosphate on the cardiac hypertrophy rats: cardiac remodeling is an important pathological process in the development and development of cardiac hypertrophy, and heart failure is the final outcome. Many studies have shown that activated RhoA/ The ROCK pathway and its related proteins mediate the occurrence of myocardial hypertrophy and apoptosis..Rho kinase belongs to the serine / threonine kinase in the family of small G protein (protein kinase A, G, C), which can further regulate ROCK through its downstream target effect molecules, which can be regulated by many biological behaviors such as cell contraction, adhesion, migration and proliferation. The fact is that the RhoA/Rock signaling pathway plays the role of up regulating inflammatory cytokines, inhibiting myocardial contraction, promoting myocardial hypertrophy and apoptosis during ventricular remodeling. The study shows that Allen phosphate is a classic FPPS inhibitor, and it affects the transduction of RhoA/Rock signaling pathway by inhibiting the formation of FPP and GGPP. To improve the pathological process of myocardial hypertrophy and fibrosis, farnesyl pyrophosphate synthetase (FPPS) may be a new idea for the treatment of myocardial hypertrophy and improvement of cardiac remodeling. Objective: To explore the possible RhoA pathway mechanism of FPPS inhibitors in improving myocardial hypertrophy and remodeling. Methods: 1. abdominal aorta coarctation was used to establish a rat model of myocardial hypertrophy; 2. Male SD rats were divided into 3 groups: abdominal aorta coarctation group, abdominal aorta coarctation and drug group and sham control group. The histomorphological changes of rats were measured by 10mg/kg/day or Allen acid, 10mg/kg/day and 3,5,8 weeks, respectively. The histomorphology of rats was detected by hemodynamics, and 4. high performance liquid chromatography (HPLC) technique. The content of FPP and GGPP in myocardium of rats was detected, 5.RhoA kit was used to determine the OD value of RhoA at 490nm in myocardial tissue, and the protein content of FDPS, RhoA and p-ERKl/2 in myocardial tissue was detected by 6. immunoglobulin. Results: 1. inhibition of FPPS could improve myocardial hypertrophy and cardiac remodeling; 2. inhibition of FPPS could reduce FPP, GGPP content and 3. of myocardium tissue. Inhibition of FPPS can reduce the content of RhoA in myocardial tissue; 4. inhibition of FPPS can inhibit cardiac hypertrophy and cardiac remodeling by inhibiting the protein activity of the RhoA related pathway. Conclusion: inhibition of FPPS can improve cardiac hypertrophy and cardiac remodeling by inhibiting the expression of protein on the RhoA pathway, and FPPS suppressor may be a new idea for the treatment of myocardial hypertrophy.
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2015
【分類號(hào)】：R54
本文編號(hào)：2159285
[Abstract]:The first part: the farnesyl pyrophosphate synthetase inhibitor improves the cardiac hypertrophy and affects the study of the remodeling of connexin: myocardial hypertrophy is a pathological change in the heart disease, which may be closely related to the excessive activation of the neuroendocrine system and the instability of the internal environment. The persistent and excessive hypertrophy of the heart muscle will eventually lead to the heart In the whole process of the formation of heart failure, cardiac remodeling is a process of dynamic development, which involves apoptosis, fibrosis, fat accumulation, mitochondrial abnormalities, changes in ion channels and signal transduction pathways. Previous studies showed that farnesyl pyrophosphate synthetase (famesyl Pyrophosphate synthase, FPPS) is a key enzyme in the metholate pathway, which catalyzes the synthesis of downstream product farnyl pyrophosphate (Farnesyl pyrophosphate, FPP) and r{bovine based r{bovine basic pyrophosphate (Geranylgeranylpyrophosphat, GGPP) involved in the direct involvement of the cardiac hypertrophy and cardiac remodeling. Cell fibrosis is an important feature of cardiac remodeling, but other changes such as troponin and gap connexin also indirectly reflect the degree and trend of cardiac remodeling. Objective: To investigate the effect of farnesyl pyrophosphate synthetase inhibitor on cardiac muscle remodeling in the animal model of cardiac hypertrophy induced by abdominal aortic coarctation. Methods: 1. abdominal aorta coarctation was used to establish the rat model of myocardial hypertrophy and to observe the effect of operation. 2. the male SD rats were randomly divided into 3 groups after the coarctation of the abdominal aorta: abdominal aorta coarctation group, abdominal aorta coarctation plus drug group and sham operation control group. The rats were given saline 10mg/kg/day or Allen phosphate sodium 10mg/kg/day irrigation respectively. Gastric maintenance 3,5,8 week.3. using ultrasound, hemodynamics and other means to detect the histomorphological changes in rats; 4. tibia length measurement of the length of the left tibia of rats; 5. high performance liquid chromatography (HPLC) technique to detect the content of FPP and GGPP in rat myocardial tissue; 6. BNP detection kit to detect the content of BNP in rat blood samples; 7. rats Myocardial tissue section staining and immunohistochemical analysis of I type fiber collagen (Col-I), T type troponin (t-TnT) and gap connexin (Cx43). Results: 1. abdominal aorta coarctation caused myocardial hypertrophy in rats and increased FPP and GGPP in myocardial tissue; 2.FPPS inhibitor could partly improve the performance of myocardial hypertrophy and myocardium. The content of FPP and GGPP in the tissue decreased, and there was no significant change in the length of tibia and blood BNP in 3. cardiac hypertrophy rats; 4. cardiac fibrosis, troponin and gap connexin were changed, and the inhibition of FPPS could partly reverse this reconstruction. Conclusion: FPPS is involved in the occurrence and development of myocardial hypertrophy and remodeling, and the inhibition of FPPS. Activity can partly improve myocardial hypertrophy. Second: the second part: the research background of the effect of farnesyl pyrophosphate synthase inhibitor Allen phosphate on the cardiac hypertrophy rats: cardiac remodeling is an important pathological process in the development and development of cardiac hypertrophy, and heart failure is the final outcome. Many studies have shown that activated RhoA/ The ROCK pathway and its related proteins mediate the occurrence of myocardial hypertrophy and apoptosis..Rho kinase belongs to the serine / threonine kinase in the family of small G protein (protein kinase A, G, C), which can further regulate ROCK through its downstream target effect molecules, which can be regulated by many biological behaviors such as cell contraction, adhesion, migration and proliferation. The fact is that the RhoA/Rock signaling pathway plays the role of up regulating inflammatory cytokines, inhibiting myocardial contraction, promoting myocardial hypertrophy and apoptosis during ventricular remodeling. The study shows that Allen phosphate is a classic FPPS inhibitor, and it affects the transduction of RhoA/Rock signaling pathway by inhibiting the formation of FPP and GGPP. To improve the pathological process of myocardial hypertrophy and fibrosis, farnesyl pyrophosphate synthetase (FPPS) may be a new idea for the treatment of myocardial hypertrophy and improvement of cardiac remodeling. Objective: To explore the possible RhoA pathway mechanism of FPPS inhibitors in improving myocardial hypertrophy and remodeling. Methods: 1. abdominal aorta coarctation was used to establish a rat model of myocardial hypertrophy; 2. Male SD rats were divided into 3 groups: abdominal aorta coarctation group, abdominal aorta coarctation and drug group and sham control group. The histomorphological changes of rats were measured by 10mg/kg/day or Allen acid, 10mg/kg/day and 3,5,8 weeks, respectively. The histomorphology of rats was detected by hemodynamics, and 4. high performance liquid chromatography (HPLC) technique. The content of FPP and GGPP in myocardium of rats was detected, 5.RhoA kit was used to determine the OD value of RhoA at 490nm in myocardial tissue, and the protein content of FDPS, RhoA and p-ERKl/2 in myocardial tissue was detected by 6. immunoglobulin. Results: 1. inhibition of FPPS could improve myocardial hypertrophy and cardiac remodeling; 2. inhibition of FPPS could reduce FPP, GGPP content and 3. of myocardium tissue. Inhibition of FPPS can reduce the content of RhoA in myocardial tissue; 4. inhibition of FPPS can inhibit cardiac hypertrophy and cardiac remodeling by inhibiting the protein activity of the RhoA related pathway. Conclusion: inhibition of FPPS can improve cardiac hypertrophy and cardiac remodeling by inhibiting the expression of protein on the RhoA pathway, and FPPS suppressor may be a new idea for the treatment of myocardial hypertrophy.
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2015
【分類號(hào)】：R54
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