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研究生：	許羽婷 Hsu, Yu-Ting
論文名稱：	MK53胜肽在代謝相關脂肪性肝炎實驗模型的應用 Application of MK53 peptide in experimental models of metabolic-associated steatohepatitis
指導教授：	孫宏羽 Sun, Hung-Yu
學位類別：	碩士 Master
系所名稱：	醫學院 - 生理學研究所 Department of Physiology
論文出版年：	2026
畢業學年度：	114
語文別：	英文
論文頁數：	65
中文關鍵詞：	MK53胜肽、代謝相關脂肪性肝性炎、膽汁酸、脂質代謝
外文關鍵詞：	MK53 peptide, MASH, Bile acid, Lipid metabolism
相關次數：	點閱：97 下載：0
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代謝相關脂肪性肝炎（metabolic-associated steatohepatitis, MASH）為代謝異常相關脂肪性肝病 (metabolic dysfunction-associated steatotic liver disease, MASLD) 的進展期病程，影響超過三分之一的患者，並且經常伴隨著肝纖維化的發生。此疾病進程可能進一步發展成肝硬化、肝癌，甚至導致死亡。儘管在臨床上對MASH的治療具有高度的迫切需求，但目前對於有效的治療選擇仍然相當有限。實驗室先前已開發一種新型的治療型胜肽「MK53胜肽」，其作為載脂蛋白J (Apolipoprotein J, ApoJ) 的拮抗肽，能夠重新平衡肝臟脂質的恆定。過去研究已經證實，MK53胜肽可降低細胞內脂質累積，並恢復代謝相關脂肪性肝病 (metabolic-associated fatty liver disease, MAFLD) 中的葡萄糖與脂質代謝的恆定。本研究利用GAN diet飲食誘導的MASH小鼠模型，進一步的探討MK53胜肽對疾病的影響，且主要關注於膽汁酸的恆定。結果顯示，MK53胜肽的治療可以改善MASH小鼠模型中的脂質代謝異常特徵。MK53胜肽可以恢復肝臟膽汁酸的合成能力，並降低血清中牛磺酸結合型膽汁酸 (taurine-conjugated bile acids) 的含量。此外，膽汁酸組成的正常化也與肝臟發炎程度下降呈現相關性。p53能夠作為調控膽汁酸代謝的轉錄因子，被發現可以與ApoJ產生交互作用。在營養過載的肝細胞中，p53入核的程度會增加，促進小異二聚體伴侶 (small heterodimer partner, SHP）的表現。在MASH小鼠中，給予MK53胜肽後可降低p53和SHP的表現量，並使CYP7A1/CYP7B1的表現正常化。綜合上述結果，本研究表明MK53胜肽可以透過調控膽汁酸合成與恆定，作為MASH潛在治療方式的可能性。

Metabolic-associated steatohepatitis (MASH) represents a progressive disease stage of metabolic dysfunction-associated steatotic liver disease (MASLD), affects more than one-third of patients and is frequently accompanied by liver fibrosis. These conditions can lead to cirrhosis, hepatocellular carcinoma, or death. Despite the urgent need, therapeutic options for MASH remain limited. Our group has previously developed a novel therapeutic peptide, MK53, which antagonizes Apolipoprotein J (ApoJ) to rebalance hepatic lipid homeostasis. MK53 peptide was demonstrated to reduce intracellular lipid accumulation and to restore glucose and lipid metabolic homeostasis in metabolic-associated fatty liver disease (MAFLD). This study further investigated the effects of MK53 peptide on bile acid homeostasis using a GAN diet-induced mouse model of MASH. The results showed that treatment with MK53 improved lipid metabolic outcomes in MASH mouse model. MK53 treatment restored the hepatic bile acid synthesis and reduced serum levels of taurine-conjugated bile acids. The normalization of the bile acid profiles correlated with a decrease in hepatic inflammation. p53, a transcription factor regulating bile acid metabolism, was found to interact with ApoJ. In hepatic cells with nutrient overload, p53 was translocated to nucleus to promote expression of small heterodimer partner (SHP). In mice with MASH, administration of MK53 peptide reduced p53 and SHP levels and normalized the expression of CYP7A1/CYP7B1. Taken together, our result support the therapeutic potential of MK53 peptide in MASH through the regulation of bile acid synthesis and homeostasis.

碩士論文合格證明書 i
中文摘要 ii
Abstract iii
Acknowledgments iv
List of Abbreviations vi
Table of Contents viii
Introduction 1
1. MASLD 1
2. The treatment of MASH 1
3. Bile acid homeostasis and lipid metabolism in MASH 2
3.1 Bile acid synthesis pathway 2
3.2 Bile acid signaling pathway 3
3.3 Role of SHP in regulation 4
3.4 p53-mediated regulation 4
3.5 Bile acid homeostasis in MASH 5
4. Diet-induced animal model for MASH 5
4.1 High-fat diet (HFD) 5
4.2 Western Diet (WD) 6
4.3 Methionine and Choline Deficient diet (MCD diet) 6
4.4 Gubra-Amylin NASH diet (GAN diet) 6
5. ApoJ and its regulation of hepatic metabolism 7
5.1 Therapeutics against ApoJ 8
6. MK53 and its function in MAFLD 9
Study Goal 10
Materials and Methods 11
1. Materials 11
1.1 Kits list 11
1.2 Reagents 11
1.3 Antibody list 11
1.4 Primer list (5' to 3') 12
2. Experimental models 12
2.1 In vivo- mouse model for MASH 13
2.1.1 Experimental design 13
2.1.2 Aspartate aminotransferase (AST) and alanine aminotransferase (ALT) quantification 13
2.1.3 Glucose tolerance test (GTT) and Insulin tolerance test (ITT) analysis 14
2.1.4 Bile acid quantification 14
2.1.5 Liver sample preparation of mice 14
2.1.6 TC and FC quantification 14
2.1.7 Quantitative real-time PCR 15
2.2 In vitro- cell model for nutrient overload 15
2.2.1 Cell experimental design 15
2.2.2 Immunoprecipitation assay 16
2.2.3 Western blotting 16
2.2.4 Nuclear and cytoplasmic extract 16
2.3 Statistical analysis 17
Results 18
MK53 peptide attenuates hepatic inflammation in mouse model of MASH 18
MK53 peptide treatment improved the physiological outcomes in mouse model of MASH 18
MK53 peptide enhanced insulin sensitivity in mice with MASH 19
MK53 peptide treatment ameliorates bile acid metabolism alterations in MASH 19
Bile acid concentrations correlate with hepatic inflammation and lipid metabolism 20
MK53 treatment trends toward recovery of bile acid metabolism-related gene expression 20
p53-ApoJ-SHP axis is associated with impaired bile acid synthesis under nutrient overload 21
MK53 targets p53-ApoJ-SHP axis to restore bile acid homeostasis in MASH mouse model 22
Conclusion 23
Discussion 24
References 27
Figures 33
Figure 1. MK53 treatment ameliorates hepatic inflammation in MASH. 33
Figure 2. MK53 treatment reduces body size and body weight in mice of MASH. 34
Figure 3. MK53 improves liver and adipose tissue weights in MASH. 35
Figure 5. MK53 enhances insulin sensitivity in mice of MASH. 37
Figure 6. MK53 treatment restores bile acids in bile. 38
Figure 7. MK53 peptide reduces taurine-conjugated bile acids. 39
Figure 8. Primary bile acids are negatively correlated with liver inflammation marker AST. 40
Figure 9. Primary bile acids are negatively correlated with liver inflammation marker ALT. 41
Figure 10. Taurine-conjugated bile acids are positively correlated with liver inflammation marker AST. 42
Figure 11. Taurine-conjugated bile acids are positively correlated with liver inflammation marker ALT. 43
Figure 12. MK53 peptide reduces hepatic FC in GAN diet feeding resulted. 44
Figure 13. The primary bile acids are negatively correlated trend with hepatic TC. 45
Figure 14. The primary bile acids are negatively correlated with hepatic FC. 46
Figure 15. The mRNA level in the liver of mice. 47
Figure 16. ApoJ interaction with p53 and SHP. 48
Figure 17. Nutrient overload induces the upregulation of ApoJ, p53, and SHP expression. 49
Figure 18. Nutrient overload stimulation increased the nuclear translocation of p53 and SHP. 50
Figure 19. MK53 peptide treatment reduces bile acid synthesis. 51
Tables 52
Table 1. The bile acid levels in the MASH model. 52
Table 2. Correlation analysis of bile acids, liver function, and hepatic lipid contents. 53
                                    

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