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The role of Dab2 in the skeletal muscle development and differentiation. / Dab2基因在骨骼肌發育與分化中的作用 / CUHK electronic theses & dissertations collection / Dab2 ji yin zai gu ge ji fa yu yu fen hua zhong de zuo yongJanuary 2012 (has links)
Dab2是一個細胞內接頭蛋白和腫瘤抑制因子。在小鼠胚胎中,應用免疫熒光染色技術,從E8.5-E11.0 Dab2發現表達於肌節的生皮肌節中。從E8.5 E9.5,Dab2表達於生皮肌節的中部。在E10.5,Dab2表達於生皮肌節的腹外側唇部,與肌肉發育的早期標誌基因Pax3和 Myf5共定位。從E11.5-E14.5,Dab2表達於四肢與軀體的肌肉中,Dab2在出生後小鼠肌肉中的表達逐漸減弱。此外,因為肌肉正常發育需要很多細胞信號的調節並且Dab2已經發現調節MAPK, TGF-β和 Wnt信號轉導通路。這些發現預示了Dab2在肌肉發育和分化中可能具有重要作用。 / 為了進一步研究它在肌肉發育中的作用,非洲爪蟾的胚胎和C2C12 肌原細胞在此研究中分別被用作體內和體外的研究模型。原位雜交結果揭示非洲爪蟾的Dab2基因表達於其胚胎的肌節中,並與肌肉發育的標誌基因XPax3, XMyoD, XMef2c和 XMyos共定位於此。用morpholino敲低XDab2 在非洲爪蟾胚胎中的表達,下調了許多肌肉發育標誌基因的表達,例如:XPax3, XMyf5, XMef2c, XMyoS 和XAC100。與此同時,免疫熒光技術也檢測到MHC(MF20)和12/101在肌節中的表達下調。 / 來源於小鼠肌肉衛星細胞的C2C12肌原細胞系被用作體外模型來檢測Dab2基因在骨骼肌發育和分化中的作用。在C2C12肌原細胞被誘導分化形成肌管的過程中,Dab2基因在RNA和蛋白水平的表達被誘導性的升高。Dab2基因超表達能夠加速肌原細胞的融合,從而增加肌小管的形成。利用miRNA敲低Dab2基因的表達能夠減緩肌原細胞的融合,從而減少肌小管的形成。利用慢病毒shRNA技術我們得到了2個Dab2穩定敲低細胞系,命名為克隆5-2和克隆5-7。這兩個克隆具有減少或抑制減少或抑制肌小管形成的特點。蛋白免疫印跡實驗表明,磷酸化p38 MAPK的表達在這兩個克隆中被抑制。在克隆5-2中超表達Dab2基因能夠恢復肌小管的形成。這個研究表明Dab2基因在肌小管的形成過程中具有至關重要的作用。 / 利用Affymetrix微陣列技術,我們檢測並分析了在克隆5-2和對照細胞中差異表達的基因。235個探針(155個基因)的顯示出超過2倍的差異表達。在這155個基因中,127個基因下調表達,28個基因上調表達。熒光定量PCR結果顯示出與微陣列結果相一致的結果。這些差異表達基因的功能發現與肌肉系統的發育和功能具有顯著地聯系。它影響了與肌肉收縮,橫紋肌的收縮,肌前體細胞的分化和肌肉發育相關功能的基因。基因網絡分析結果揭示,在克隆5-2中Mef2c基因的下調表達可能是一個導致肌細胞分化抑制的原因。 Mef2c基因在克隆5-2中超表達能夠拯救肌細胞的分化。 / 總括來說,體內和體外實驗共同表明Dab2基因是一個肌肉發育和分化的正調控基因。 / Dab2 is an intracellular adaptor protein and a tumor suppressor. In mouse embryos, Dab2 was found to be expressed in the dermomyotome of somites from E8.5 to E11.0 using immunofluorescence staining, with expression first detected in the medial aspect of the dermomyotome at E8.5 and then co-localized with the early muscle markers Pax3 and Myf5 at the ventrolateral lip of the dermomyotome at E10.5. From E11.5 to E14.5, Dab2 was expressed in muscle masses of limb buds and the trunk. Dab2 expression in skeletal muscles was gradually decreased after birth. These observations suggested potential roles of Dab2 in the skeletal muscle myogenesis. In addition, since the normal development of skeletal muscles requires proper signal transduction, and Dab2 has been known to be involved in the MAPK, TGF-β and Wnt signaling pathways, Dab2 may therefore be important for the muscle development. / To determine the role of Dab2 in the skeletal muscle development, Xenopus laevis embryos and C2C12 myoblasts were employed as in vivo and in vitro models, respectively. In situ hybridization results showed that XDab2 was expressed in somites of Xenopus embryos and co-localized with the muscle markers XPax3, XMyoD, XMef2c and XMyos. Knockdown of XDab2 expression with antisense morpholinos down regulated the expression of several muscle markers in somites including XPax3, XMyf5, XMef2c, XMyoS and XAC100. Down-regulation of MHC and 12/101 were also observed in whole mount preparations and transverse sections of XDab2 morpholino-injected embryos after immunohistochemical staining. / The C2C12 cell line derived from mouse muscle satellite cells was then employed as an in vitro model to determine the role of Dab2 during early muscle development. When C2C12 myoblasts were induced to differentiate into myotubes, Dab2 expression was simultaneously increased at RNA and protein levels. Dab2 over-expression after transfection with Dab2 plasmids resulted in enhanced myoblast fusion and increased numbers of myotubes. Conversely, suppression of Dab2 expression with miRNAs resulted in reduced myoblast fusion and decreased numbers of myotubes. Lentiviral shRNA-mediated Dab2 stable knockdown reduced myotube formation in 2 representative stable clones, clone 5-2 and clone 5-7. Western blot analysis showed that expression of phospho-p38 MAPK was down-regulated in clone 5-2 and 5-7. Dab2 re-expression through plasmid-mediated transient transfection in clone 5-2 could partially restore the myotube formation. These observations therefore suggested that Dab2 plays essential roles in the formation of myotubes. / Comprehensive profiling of differentially expressed genes was performed with the Affymetrix microarray analysis between the Dab2-knockdown clone 5-2 and the C2C12 parental cell line. As compared to the parental cells, the clone 5-2 showed significant changes in the expression of 235 probe sets representing 155 genes (p<0.05) with 2 folds or greater changes. Among the 155 genes, 127 were down-regulated, while 28 up-regulated. qRT-PCR results were found to be consistent with the microarray results. Functions of the differentially expressed genes were found to be significantly associated with the development and functions of the muscular system. Knockdown of Dab2 affected the genes involved in muscle contraction, the contraction of striated muscle, differentiation of muscle precursor cells, and the development of skeletal muscle fibers. A network analysis and a gene expression study revealed that Mef2c down-regulation was related to the inhibition of myogenic differentiation in the clone 5-2. Furthermore, forced expression of Mef2c in the clone 5-2 could rescue the myogenic differentiation. / In conclusion, these results indicated that Dab2 is positive regulator of the skeletal muscle development and differentiation both in vivo and in vitro. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Shang, Na. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 211-227). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese. / Abstract --- p.i / 摘要 --- p.iv / Acknowledgements --- p.vi / Table of contents --- p.vii / Abbreviation --- p.xiii / Chapter Chapter 1 --- General Introduction --- p.1 / Chapter 1.1 --- Characterizations of the Dab2 gene --- p.1 / Chapter 1.2 --- The role of Dab2 in Wnt/ beta-catenin signaling --- p.2 / Chapter 1.3 --- The role of Dab2 in TGF beta signaling --- p.3 / Chapter 1.4 --- The role of Dab2 in Ras-MAPK signaling --- p.4 / Chapter 1.5 --- The role of Dab2 in protein trafficking and endocytosis --- p.5 / Chapter 1.6 --- Dab2 expression and its functions. --- p.7 / Chapter 1.7 --- Somite and skeletal muscle development --- p.8 / Chapter 1.8 --- The formation of the somite and its structure --- p.9 / Chapter 1.9 --- The formation of dermomyotome and its function --- p.10 / Chapter 1.10 --- The formation of myotome and its function --- p.11 / Chapter 1.11 --- The formation of muscle fibers and musculatures --- p.12 / Chapter 1.12 --- The formation of satellite cells and its function in skeletal muscle differentiation --- p.12 / Chapter 1.13 --- The gene expression during skeletal muscle development and differentiation --- p.13 / Chapter 1.14 --- Dab2 genetically modified mice --- p.16 / Chapter 1.15 --- Objectives of this research --- p.17 / Chapter Figures and legends --- p.21 / Chapter Chapter 2 --- Expression of Dab2 in the mouse somites and skeletal muscles --- p.32 / Chapter 2.1 --- Introduction --- p.32 / Chapter 2.2 --- Materials and Methods --- p.34 / Chapter 2.2.1 --- Mouse embryos and tissue isolation --- p.34 / Chapter 2.2.2 --- Histological preparation of embryos and tissues --- p.34 / Chapter 2.2.3 --- Immunostaining using Tyramide signal amplification kits --- p.35 / Chapter 2.3 --- Results --- p.36 / Chapter 2.3.1 --- Dab2 expression in somites of the mouse embryos --- p.36 / Chapter 2.3.2 --- Dab2 expression in skeletal muscles of embryonic and postnatal mice --- p.36 / Chapter 2.3.3 --- Co-localization of Dab2 and Pax3 immunoreactivities with double immunofluorescence staining --- p.37 / Chapter 2.3.4 --- Co-localization of Dab2 and Myf5 immunoreactivities with double immunofluorescence staining --- p.38 / Chapter 2.3.5 --- Co-localization of Dab2 and Myogenin immunoreactivities with double immunofluorescence staining --- p.38 / Chapter 2.4 --- Discussion --- p.40 / Chapter 2.5 --- Summary --- p.42 / Chapter Table 2.1 --- p.44 / Chapter Figures and Legends --- p.45 / Chapter Chapter 3 --- Dab2 is a positive regulator of skeletal muscle development in Xenopus embryos --- p.58 / Chapter 3.1 --- Introduction --- p.58 / Chapter 3.2 --- Materials and Methods --- p.61 / Chapter 3.2.1 --- RNA extraction --- p.61 / Chapter 3.2.2 --- Reverse-transcription polymerase chain reaction (RT-PCR) --- p.61 / Chapter 3.2.3 --- Gene cloning and sequencing analysis --- p.61 / Chapter 3.2.4 --- Transformation --- p.62 / Chapter 3.2.5 --- Plasmid mini and midi-preparation --- p.62 / Chapter 3.2.6 --- Frogs and embryos handling --- p.63 / Chapter 3.2.7 --- Synthesis of mRNA for microinjection --- p.64 / Chapter 3.2.8 --- Microinjection --- p.64 / Chapter 3.2.9 --- Synthesis of DIG-labeled anti-sense RNA probe --- p.65 / Chapter 3.2.10 --- Whole mount in situ hybridization (WMISH) and whole mount immunohistochemical localization --- p.65 / Chapter 3.3 --- Results --- p.67 / Chapter 3.3.1 --- Cloning of Xenopus Dab2 long isoform and the sequence analysis --- p.67 / Chapter 3.3.2 --- Phylogenetic analysis --- p.67 / Chapter 3.3.3 --- RT-PCR analysis of Xenopus Dab2 (XDab2) expression --- p.68 / Chapter 3.3.4 --- Xenopus Dab2 spatial and temporal expression examined by WMISH analysis --- p.68 / Chapter 3.3.5 --- Dab2 expression in somites and its colocalization with myogenic transcription factors --- p.69 / Chapter 3.3.6 --- XDab2 knockdown led to down-regulation of myogenic transcription factors and muscle markers at the RNA level --- p.70 / Chapter 3.3.7 --- XDab2 knockdown led to down-regulation of muscle markers at the protein level --- p.70 / Chapter 3.3.8 --- XDab2 overexpression led to up-regulation of XPax3, XMyf5 and XMyoS --- p.71 / Chapter 3.4 --- Discussion --- p.72 / Chapter 3.5 --- Summary --- p.77 / Chapter Table 3.1 --- p.78 / Chapter Figures and Legends --- p.79 / Chapter Chapter 4 --- Potential roles of Dab2 in C2C12 myoblast differentiation --- p.99 / Chapter 4.1 --- Introduction --- p.99 / Chapter 4.2 --- Materials and Methods --- p.101 / Chapter 4.2.1 --- Cell culture and differentiation in vitro --- p.101 / Chapter 4.2.2 --- Cell sample preparation --- p.102 / Chapter 4.2.3 --- Real-time PCR --- p.102 / Chapter 4.2.4 --- SDS-PAGE --- p.103 / Chapter 4.2.5 --- Western blotting and immunodetection --- p.104 / Chapter 4.2.6 --- Plasmids used for transient over-expression --- p.105 / Chapter 4.2.7 --- Generation of miRNAs targeting at Dab2 --- p.105 / Chapter 4.2.8 --- C2C12 differentiation after transfection --- p.106 / Chapter 4.2.9 --- Immunohistochemical staining for myotubes --- p.106 / Chapter 4.2.10 --- Lentiviral shRNA mediated Dab2 stable knockdown --- p.107 / Chapter 4.2.10.1 --- shRNA Lentiviral Transduction Particles and sequence information --- p.107 / Chapter 4.2.10.2 --- Optimization of puromycin treatment on C2C12 myoblasts --- p.107 / Chapter 4.2.10.3 --- Determination of the optimal MOI for C2C12 --- p.108 / Chapter 4.2.10.4 --- Lentivirus transduction method --- p.109 / Chapter 4.2.10.5 --- Stable cell line generation --- p.109 / Chapter 4.2.11 --- Rescue experiments --- p.109 / Chapter 4.2.12 --- Serum starvation and FGF treatment --- p.110 / Chapter 4.2.13 --- Microarray and data analysis --- p.110 / Chapter 4.3 --- Results --- p.113 / Chapter 4.3.1 --- Expression of Dab2 during myogenesis --- p.113 / Chapter 4.3.2 --- Generation of miRNAs targeting at Dab2 --- p.113 / Chapter 4.3.3 --- Improvement of the transfection efficiency --- p.114 / Chapter 4.3.4 --- Knockdown efficiencies of the 4 miRNAs --- p.114 / Chapter 4.3.5 --- Down-regulation of Dab2 expression by transient transfection inhibited C2C12 differentiation --- p.115 / Chapter 4.3.6 --- Up-regulation of Dab2 expression by transient transfection enhanced myogenic differentiation --- p.116 / Chapter 4.3.7 --- Lentivirus-mediated Dab2 stable knockdown inhibited myotube formation --- p.117 / Chapter 4.3.8 --- Re-expression of Dab2 partially restored myogenic differentiation in the clone 5-2 --- p.120 / Chapter 4.3.9 --- Dab2 knockdown affected the MAPK signaling pathway --- p.122 / Chapter 4.3.10 --- Transcriptome and network analysis revealed changes of gene expression patterns in the C2C12 cell line after Dab2 knockdown --- p.123 / Chapter 4.3.11 --- Mef2c down-regulation was related to the inhibition of the myotube formation in the clone 5-2 --- p.126 / Chapter 4.4 --- Discussion --- p.128 / Chapter 4.4.1 --- Dab2 expression was found to be induced upon differentiation and down-regulated after myotube formation --- p.128 / Chapter 4.4.2 --- Dab2 was found to be a positive regulator of C2C12 differentiation --- p.129 / Chapter 4.4.3 --- Dab2 knockdown affected the MAPK signaling pathway --- p.131 / Chapter 4.4.4 --- Potential roles of Dab2 in myogenic differentiation revealed by transcriptome and network analysis --- p.133 / Chapter 4.4.5 --- Mef2c down-regulation may be involved in the inhibition of myogenic differentiation after Dab2 knockdown --- p.135 / Chapter 4.5 --- Summary --- p.138 / Chapter Table 4.1 --- p.141 / Chapter Table 4.2 --- p.142 / Chapter Table 4.3 --- p.143 / Chapter Table 4.4 --- p.144 / Chapter Table 4.5 --- p.147 / Chapter Table 4.6 --- p.148 / Chapter Table 4.7 --- p.149 / Chapter Figures and Legends --- p.150 / Chapter Chapter 5 --- Conclusions and discussion --- p.192 / Chapter 5.1 --- Dab2 expression in somites and skeletal muscles of mouse embryos --- p.192 / Chapter 5.1 --- Dab2 as a positive regulator for skeletal muscle development in Xenopus embryos in vivo --- p.194 / Chapter 5.3 --- Dab2 as a positive regulator of skeletal muscle development in vitro --- p.196 / Chapter 5.3.1 --- Dab2 was found to be a positive regulator of C2C12 differentiation --- p.196 / Chapter 5.3.2 --- Dab2 knockdown affected the MAPK signaling pathway --- p.198 / Chapter 5.3.3 --- Potential functions of Dab2 revealed by transcriptomeand network analysis --- p.200 / Chapter 5.3.4 --- Mef2c down-regulation was closely related to the inhibition of myogenic differentiation upon Dab2 knockdown --- p.202 / Appendix I --- p.204 / Appendix II --- p.205 / References --- p.211
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Proteomic study of the effects of palmitic acid on skeletal muscle cell and its relation with mitochondrial function. / CUHK electronic theses & dissertations collectionJanuary 2012 (has links)
2 型糖尿病(T2D)的發展歷史悠久,但導致T2D 患者胰島素抵抗的確切病理還沒有完全理解。骨骼肌佔大多數(70-80%)的胰島素引導的葡萄糖的吸收,所以它一直是胰島素抵抗的研究焦點。許多 T2D 患者的骨骼組織顯示線粒體功能障礙,但線粒體功能障礙和胰島素抵抗之間的關係尚不清楚還在辯論中。在這個項目中,這種關係是通過研究游離脂肪酸(FFA)( 24 小時處理)對 C2C12 小鼠骨骼肌細胞的效果來闡明。 / 免疫印記法顯示FFA 誘導胰島素抵抗,結合二維電泳和質譜分析的蛋白質組學研究發現FFA 有抑制糖酵解,增加β-氧化作用,沒有改變檸檬酸循環和抑製氧化磷酸化的作用。FFA 抑制電子傳遞鏈的幾個組成部分,揭示線粒體功能障礙,背後的原因可推測為FFA 增加令β-氧化作用增加,但沒有協調改變率檸檬酸循環,導致積累不完全β-氧化的中轉體,導致線粒體過載,最終導致胰島素抵抗。 / There is a long history of Type 2 diabetes (T2D) research development, but the exact pathology leading to insulin resistance of T2D is still not fully understood. T2D is frequently characterized by tissue insulin resistance and it is often associated with an elevated concentration of palmitic acid (PA, a major kind of dietary fatty acid) in serum. Due to this correlation, much of the effort in the field had been concentrated on the effect of PA in insulin action and glucose metabolism, and how elevated PA could possibly cause insulin resistance in specific tissues. / Skeletal muscle accounts for the majority (70-80%) of insulin-mediated glucose uptake, so it has been the focus of insulin resistance studies. Many T2D patients having elevated serum free fatty acid (FFA, where PA is a kind of FFA) also show mitochondrial dysfunction in their skeletal tissue, but the relationship between mitochondrial dysfunction and insulin resistance in skeletal muscle remains unclear and under debate. In this project, the three-party relationship was elucidated by studying the effect of 24hrs of incubation of palmitic acid (PA) on skeletal muscle using C2C12 mouse skeletal cells as model. / PA-treated C2C12 cells show reduction in insulin-stimulated Akt phosphorylation when compared with untreated C2C12 cells. Comparative proteomic study for both total proteins and mitochondrial proteins with 2D gel electrophoresis and mass spectrometry unveil, when compared with untreated cells, PA-treated C2C12 cells show down-regulation in enzymes involved in glycolysis(e.g. glyceraldehyde-3-phosphate dehydrogenase, phosphoglycerate kinase, fructose-bisphosphate aldose A), up-regulation in enzymes involved in beta-oxidation(e.g. 3-ketoacyl-CoA thiolase, 3-hydroxyacyl-CoA dehydrogenase), and down-regulation in proteins involved in oxidative phosphorylation(e.g. ATP synthase subunits, NADH-ubiquinone oxidoreductase 75kDa subunit, cytochrome b-c complex subunit 1). / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Lam, Chor Kwan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 69-78). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts also in Chinese. / Thesis/Assessment Committee --- p.i / Declaration --- p.ii / Abstract (in English) --- p.iii / Abstract (in Chinese) --- p.v / Acknowledgments --- p.vi / Table of Contents --- p.vii / List of Abbreviations --- p.x / List of Figures --- p.xiii / List of Tables --- p.xiv / Chapter 1. --- Literature review --- p.1 / Chapter 1.1. --- Introduction to diabetes mellitus --- p.1 / Chapter 1.1.1. --- Definition and prevalence --- p.1 / Chapter 1.1.2. --- Diagnosis and classification --- p.2 / Chapter 1.1.3. --- Symptoms and complications --- p.4 / Chapter 1.1.4. --- Causes and risk factors --- p.5 / Chapter 1.1.5. --- Prevention and treatment --- p.6 / Chapter 1.2. --- The role of muscle tissue in pathophysiology of T2DM --- p.7 / Chapter 1.3. --- Insulin receptor substrate-1 and Fatty acids-induced insulin resistance --- p.15 / Chapter 1.4. --- Introduction of proteomics --- p.18 / Chapter 1.4.1. --- The application of proteomics in disease discovery --- p.18 / Chapter 1.4.2. --- Application of Proteomics --- p.19 / Chapter 1.4.3 --- Two-dimensional gel electrophoresis --- p.20 / Chapter 1.4.4 --- Organelles proteomics --- p.21 / Chapter 1.4.5. --- Mass spectrometry --- p.22 / Chapter 1.4.6 --- Application of proteomic technology in disease pathology --- p.24 / Chapter 1.4.7 --- Current challenges --- p.25 / Chapter 1.5 --- Objectives --- p.27 / Chapter 2 --- Materials and Methods --- p.28 / Chapter 2.1 --- Fatty acid preparation --- p.28 / Chapter 2.2 --- Cell culture --- p.28 / Chapter 2.2.1 --- Treatment of C2C12 myotubes with Palmitic acid --- p.28 / Chapter 2.2.2 --- MTT assay for viability measurement --- p.29 / Chapter 2.2.3 --- Determination of the IC₅₀ values --- p.31 / Chapter 2.3 --- Proteomic analysis of C2C12 cells with and without PA treatment --- p.32 / Chapter 2.3.1 --- Protein sample preparation from C2C12 skeletal muscle cells --- p.32 / Chapter 2.3.2 --- Protein quantitation --- p.33 / Chapter 2.3.3 --- 2D Gel electrophoresis --- p.34 / Chapter 2.3.4 --- Image analysis --- p.36 / Chapter 2.3.5 --- In gel digestion and MALDI-ToF MS --- p.37 / Chapter 2.4 --- Mitochondrial purification and protein extraction --- p.38 / Chapter 2.4.1 --- Ultracentrifugation method --- p.38 / Chapter 2.4.2 --- Mitochondrial Isolation Kit --- p.39 / Chapter 2.5 --- Western Immunoblotting --- p.40 / Chapter 2.5.1 --- Protein sample preparation --- p.40 / Chapter 2.5.2 --- SDS-PAGE --- p.40 / Chapter 2.5.3 --- Western blotting --- p.40 / Chapter 2.5.4 --- Membrane Blocking and Antibody Incubations --- p.41 / Chapter 2.5.5 --- Detection of Proteins --- p.42 / Chapter 3 --- Results --- p.43 / Chapter 3.1 --- Differentiation of C2C12 myoblast into myotubes --- p.43 / Chapter 3.2 --- The effect of Palmitic acid on C2C12 Proliferation --- p.44 / Chapter 3.3 --- Comparison of total protein profiles of palmitic acid-treated C2C12 myotubes with control myotubes --- p.45 / Chapter 3.4 --- Western blotting of Akt and Phospho-Akt in C2C12 cells treated with Palmitic acid after acute exposure to insulin --- p.50 / Chapter 3.5 --- Comparison of two mitochondria isolation methodsultracentrifugation and mitochondrial isolation kit --- p.51 / Chapter 3.5.1 --- Quantity of extracted mitochondrial protein --- p.51 / Chapter 3.5.2 --- Purity of extracted mitochondrial protein --- p.52 / Chapter 3.6 --- Comparison of mitochondrial protein profiles between palmitic acid-treated and control C2C12 myotubes --- p.53 / Chapter 3.7 --- Western blotting of insulin receptor substrate-1 and its serine phosphorylation --- p.58 / Chapter 4 --- Discussion --- p.59 / Chapter 4.1 --- Investigation of anti-proliferating effect of Palmitic acid on C2C12 using MTT assay --- p.59 / Chapter 4.2 --- Comparison of total protein profiles of palmitic C2C12 myotubes with control myotubes --- p.60 / Chapter 4.3 --- Western blotting of insulin receptor substrate-1and its serine phosphorylation --- p.62 / Chapter 4.4 --- Western blotting of Akt and Phospho-Akt in C2C12 cells treated with Palmitic acid after acute exposure to insulin --- p.63 / Chapter 4.5 --- Comparison of mitochondrial protein profiles between palmitic acid-treated and control C2C12 myotubes --- p.65 / Chapter 4.6 --- Problems faced and future prospect --- p.68 / Chapter 5 --- References --- p.69
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The extrinsic apoptotic pathway in aged skeletal muscle roles of tumor necrosis factor-[alpha] and interleukin-15 /Pistilli, Emidio E. January 2006 (has links)
Thesis (Ph. D.)--West Virginia University, 2006. / Title from document title page. Document formatted into pages; contains x, 189 p. : ill. (some col.). Includes abstract. Includes bibliographical references.
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The Effect Of Diabetes On Rat Skeletal Muscle Tissues At Molecular LevelBozkurt, Ozlem 01 September 2006 (has links) (PDF)
In the present study Fourier Transform Infrared Spectroscopy was used to examine the effects of streptozotocin-induced diabetes mellitus on the structural components of slow- and fast-twitch rat skeletal muscles, at molecular level.
Diabetes mellitus is a chronic disorder of carbohydrate, fat and protein metabolism, which is characterized by hyperglycemia caused by a defective or deficient insulin secretory response. The effect of diabetes is seen on a variety of tissues leading to important secondary complications
such as kidney failure, liver dysfunction, cardiac disorders, etc. Skeletal muscle is one of the major tissues determining carbohydrate and lipid metabolism in the body / therefore, is one of the target tissues of diabetes.
The two main types of muscle fibers are type I (slow-twitch) and type II (fast-twitch) fibers / having different structural organization and metabolic features.
The FTIR spectra revealed a considerable decrease in lipid and protein content of diabetic skeletal muscles, indicating an increased lipolysis and protein breakdown or decreased protein synthesis. Moreover changes in protein structure and conformation were observed. In diabetes, muscle membrane lipids were more ordered and the amount of unsaturated lipids
was decreased possibly due to lipid peroxidation. Diabetes caused a decrease in the content of nucleic acids, especially RNA, and hydrogen bonded phospholipids in the membrane structures of skeletal muscles.
In all of the spectral parameters investigated slow-twitch muscle was more severely affected from diabetes. Thus, FTIR spectroscopy appears to be a useful method to evaluate the effect of diabetes on skeletal muscle tissues at molecular level.
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Orchestration of skeletal myogenesis by the myogenic bHLH family of transcription factors /Bergstrom, Donald Alan, January 2000 (has links)
Thesis (Ph. D.)--University of Washington, 2000. / Vita. Includes bibliographical references (leaves 53-58).
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Vasomotor responses of rat skeletal muscle arterioles to norepinephrine and adenosineAaker, Aaron Paul, January 2001 (has links)
Thesis (Ph. D.)--University of Missouri--Columbia, 2001. / Typescript. Vita. Includes bibliographical references (leaves 122-137). Also available on the Internet.
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Effects of hindlimb unweighting on soleus muscle resistance artery endothelial function and eNOS expressionSchrage, William January 2001 (has links)
Thesis (Ph. D.)--University of Missouri--Columbia, 2001. / Typescript. Vita. Includes bibliographical references (leaves 141-150). Also available on the Internet.
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Novel pharmaceutical approaches to regulate glucose homeostasisSundbom, Maj, January 2010 (has links)
Diss. (sammanfattning) Stockholm : Karolinska institutet, 2010. / Härtill 3 uppsatser.
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Aging differences in mechanisms of human skeletal muscle hypertrophyKosek, David J. January 2007 (has links) (PDF)
Thesis (Ph.D.)--University of Alabama at Birmingham, 2007. / Title from PDF title page (viewed on Feb. 18, 2010). Includes bibliographical references.
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Nutrient and energy sensing in skeletal muscleDeshmukh, Atul S., January 2009 (has links)
Diss. (sammanfattning) Stockholm : Karolinska institutet, 2009. / Härtill 5 uppsatser.
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