本文關(guān)鍵詞：Hand2及Pten在第二心場增殖和發(fā)育中的功能研究，由筆耕文化傳播整理發(fā)布。

【摘要】：心血管疾病在發(fā)展中國家以及發(fā)達(dá)國家中是導(dǎo)致死亡的主要疾病。而在這類疾病當(dāng)中,先天性心臟病,由于在胚胎發(fā)育過程中心臟發(fā)育的畸形,約影響超過5%的新生兒童的健康。因而,了解早期心臟發(fā)育的調(diào)節(jié)機(jī)制是必需的。早期心臟的前體細(xì)胞來源于側(cè)板中胚層并由兩部分前體細(xì)胞群構(gòu)成,即第一心場和第二心場細(xì)胞來源。 第一心場貢獻(xiàn)于心臟左心室的結(jié)構(gòu),而第二心場貢獻(xiàn)于心臟流出道,右心室,靜脈竇以及部分左右心房的形成。在早期心臟發(fā)育過程中,FGF,BMP,Wnt等信號對于誘導(dǎo)心臟特異的轉(zhuǎn)錄因子的表達(dá)發(fā)揮著重要的作用。然而,這些信號與轉(zhuǎn)錄因子如何相互作用來調(diào)節(jié)心臟發(fā)生這一過程目前研究的依然不是很清楚。Hand2,與心臟的其他轉(zhuǎn)錄因子如工s11,Mef2c,Fgf8/10以及FoxH等一樣,對于調(diào)節(jié)心臟第二心場的發(fā)育是必需的。全身敲除Hand2基因?qū)е屡咛ブ滤啦橛行呐K環(huán)化的缺陷,因而,我們利用Mef2c-cre轉(zhuǎn)基因工具小鼠特異性的在第二心場敲除Hand2基因來研究心臟發(fā)育的過程。Hand2敲除小鼠表現(xiàn)有右心室發(fā)育不全和流出道發(fā)育缺陷等表型,并最終死于胚胎期第13.5天,提示著Hand2在這一區(qū)域中的必要性。通過免疫熒光染色實(shí)驗(yàn),我們發(fā)現(xiàn)在敲除Hand2小鼠心臟中,細(xì)胞增殖是減少的,而細(xì)胞凋亡和自噬是增加的,而細(xì)胞的分化未受到明顯的影響。此外,ERK信號通路的活性是下調(diào)的。以此同時(shí),參與直接調(diào)控ERK磷酸化活性的FGFs的表達(dá)量在Hand2敲除的心臟中也是下調(diào)的。近一步實(shí)驗(yàn)證明Fgf10是Hand2的一個(gè)直接下游靶基因。然而,我們通過刪除Pten提高ERK的活性可以明顯的恢復(fù)由于Hand2缺失而造成的心臟發(fā)育缺陷的表型,不論在分子水平還是在動(dòng)物水平上。與此同時(shí),與Hand2單敲小鼠相比較,Hand2和Pten雙敲小鼠的壽命明顯延長。在早期胚胎發(fā)育過程中,MAPK/ERK及PI3K/AKT信號通路能夠直接被FGF信號所誘導(dǎo),對調(diào)節(jié)細(xì)胞的存活,增殖和分化是必需的。而之前的研究表明Pten作為腫瘤抑制基因發(fā)揮其作用通過負(fù)調(diào)控PI3K/AKT及MAPK/ERK信號通路,而Pten對脊椎動(dòng)物發(fā)育的調(diào)節(jié)依然不是很清楚。是否Pten刪除導(dǎo)致的這兩條信號通路的失調(diào)對早期心臟形態(tài)發(fā)生過程有重要影響,我們著重于研究該基因在心腔第二心場中的作用。組織學(xué)研究表明,刪除Pten基因在該區(qū)域?qū)е滦呐K的肥大,心室間隔的發(fā)育不全,以及心臟瓣膜和心肌小梁發(fā)育的缺陷,不論是在胚胎期還是在出生后小鼠心臟均發(fā)現(xiàn)此類表型。以此同時(shí),敲除Pten基因引起第二心場祖細(xì)胞的過度增殖和提前分化。近一步的研究表明Pten通過Akt-FoxO介導(dǎo)的上調(diào)BMP信號通路來調(diào)節(jié)祖細(xì)胞的增殖。此外,我們還發(fā)現(xiàn)Pten正調(diào)控NOTCH信號通路來調(diào)節(jié)心肌小梁的發(fā)育。染色體重塑復(fù)合物BAF也參與Pten介導(dǎo)的右心室的發(fā)育,提示著Pten可能通過表觀遺傳來調(diào)節(jié)第二心場的發(fā)育。綜上所述,我們的研究結(jié)果表明Hand2及Pten在調(diào)節(jié)第二心場祖細(xì)胞的增殖和心臟發(fā)育過程中發(fā)揮著重要的作用。
【關(guān)鍵詞】：Hand2 Pten 心臟發(fā)育 心臟發(fā)育缺陷 第二心場 祖細(xì)胞增殖 提前分化
【學(xué)位授予單位】：南京大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2015
【分類號】：Q344
【目錄】：

• Abstract7-10
• 中文摘要10-12
• List of Abbreviations12-14
• Chapter 1 Introduction14-64
• 1.1. Cardiac development and signaling transduction15-33
• 1.1.1. Cardiac development from cardiac mesoderm15-17
• 1.1.2. Morphogenesis of the cardiac17-20
• 1.1.3. First heart field and second heart field20-22
• 1.1.4. Signaling pathways and transcriptional regulators in cardiac development22-33
• 1.1.4.1 Early signals in cardiac development22-26
• 1.1.4.2. Transcription factors mark cardiac progenitor cells26
• 1.1.4.3. Transcription factors in 1~(st) heart field derivatives26-28
• 1.1.4.4. Transcription factors in 2~(st) heart field derivatives28-33
• 1.2. The role of Hand transcription factors in cardiac morphogenesis33-40
• 1.2.1. Basic Helix-Loop-Helix (bHLH) Transcription Factor33-34
• 1.2.2. Hand Transcription Factors and Cardiovascular Development34-40
• 1.2.2.1. Expression of Hand transcription factors in cardiac development34-35
• 1.2.2.2. Function of Hand transcription factors in cardiac development35-38
• 1.2.2.3. Molecular and cellular mechaniams regulated by Hand238-39
• 1.2.2.4. HAND factors in human congenital heart disease (CHD)39-40
• 1.3. Characterization of Pten and its functions in development40-47
• 1.3.1. The discovery of Pten40-41
• 1.3.2. Structure, function and regulation of Pten41-45
• 1.3.2.1. Structure of Pten41-42
• 1.3.2.2 Function of Pten42
• 1.3.2.3. Regulation of Pten42-45
• 1.3.3. Function of Pten in tumorigenesis and embryonic development45-47
• 1.4. Aim and objectives47-49
• 1.5. References49-64
• Chapter 2 The HAND2-FGF-ERK Signaling Controls Proliferation of Second HeartField Progenitor64-94
• 2.1. Introduction65-67
• 2.2. Results67-85
• 2.2.1. Loss of Hand2 resulted in abnormal cardiac morphology67-69
• 2.2.1.1. Generation of Hend2 second heart field-specific knockout mice67
• 2.2.1.2. Ablation of Hand2 resulted in severe cardiac defects67-69
• 2.2.2. Cell proliferation,differentiation,apoptosis and autophagy in Hand2 mutant mice69-71
• 2.2.2.1. Ablation of Hand2 decreased the SHF progenitors and increased myocardial cells apoptosis and autophagy69-71
• 2.2.2.2. Ablation of Hand2 did not affect cell differentiation in the SHF71
• 2.2.3. Molecular studies of Hand2 mutant embryos71-75
• 2.2.3.1. Hand2 mediated activation of FGF-ERK signaling in the SHF development71-73
• 2.2.3.2. The Fgf10 promoter was activated directly by Hand273-75
• 2.2.4. Enhancement of phspho-ERK activity by loss of Pten could rescue the defects of cell proliferation in Hand2 mutant embryos75-85
• 2.2.4.1. Ablation of Pten ed to enlarged right ventricle and lengthen outflow tract75-77
• 2.2.4.2. Increased cell proliferation in Pten mulant embryos77-78
• 2.2.4.3. Ablation of Pten ed to premature differentiation of SHF progenitor cells78-80
• 2.2.4.4. Increased both of phspho-ERK and phspho-AKT activity in Pten mutant embryos80-81
• 2.2.4.5. Loss of Pten in the SHF rescued the SHF defects of Hand2 Mutants at molecular level81-83
• 2.2.4.6. Genetic ablation of Hand2 and Pten partially rescued the SHF phenotypes of Hand2 deletion mice83-85
• 2.3. Discussion85-88
• 2.4. Materials and methods88-90
• 2.5. References90-94
• Chapter 3 Pten Function in Second Heart Field is Required for Embryonic andPostnatal Heart Development94-131
• 3.1. Introduction95-96
• 3.2. Results96-118
• 3.2.1. Pten is required for embryonic cardiac development96-98
• 3.2.2. Analysis of cardiac defects in the surviving Pten~(f/f);Mef2c-Cre mice98-105
• 3.2.2.1. Loss of Pten in the second heart field results in cardiac hypertrophy and ventricular septal hyperplasia98-100
• 3.2.2.2. Loss of Pten in the second heart field results in cardiac valve and trabecular defects100-103
• 3.2.2.3. cell morphology abnormal in Pten~(f/f);Mef2c-Cre heart103-105
• 3.2.3. Pten promotes SHF cell proloferation through positvely regulating BMP signaling105-109
• 3.2.3.1. BMP signaling pathway is downregulated in Pten~(f/f);Mef2c-Cre heart105-106
• 3.2.3.2.Bmp7 promoter is directly regulated by Foxo3a in vitro106-109
• 3.2.4. Genetic ablation of Pten and Akt1 rescue RV hypertrophy and extend life span of Pten~(f/f);Mef2c-Cre mice109-110
• 3.2.5. The Combinatorial deletion of Pten and β-catenin partially rescued Pten mutant heart110-112
• 3.2.6. Analysis of Pten~(f/f);mef2c-Cre heart at late stage112-114
• 3.2.7. Chromstin-Remodeling Factors are involved in Pten~(f/f);Mef2c-Cre mice114-118
• 3.3. Discussion118-121
• 3.4. Materials and methods121-126
• 3.4.1. Animal studies121-124
• 3.4.2. Molecular studies124-126
• 3.5. References126-131
• Curriculum Vitae:Wen Luo131-133
• 致謝133-134

  本文關(guān)鍵詞：Hand2及Pten在第二心場增殖和發(fā)育中的功能研究，由筆耕文化傳播整理發(fā)布。

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本文編號：397700