背景、目的: 動物體血糖之恆定需依靠胰島β細胞分泌胰島素調控血糖。胰島之β細胞數目於一生中維持著一種動態平衡狀態以恆定血糖。於出生前後時期，β細胞會短暫快速的增生，以增加β細胞團以因應出生後體內之血糖恆定。在成年期，如體內產生胰島素阻抗之病理狀態時，β細胞具有細胞生長與增生的能力以代償性增加β細胞團以分泌足夠的胰島素因應。轉譯調控腫瘤蛋白(TCTP)是一種演化上高度保留之蛋白質，已被證實與細胞生長及增生有關。近期的體外細胞研究證實其在胰島β細胞內扮演著一種可受葡萄糖調控及支持細胞存活之蛋白。但TCTP於活體動物β細胞內之角色仍未明。本研究之目的在探討於正常生理發育階段與產生病理胰島素阻抗時期，TCTP在活體動物胰島β細胞扮演之角色為何?
實驗方法: 進行小鼠胚胎時期到成鼠階段TCTP於胰島β細胞表達量之評估後，利用選擇性基因剔除小鼠模式，將TCTPfl/fl小鼠與RIP-cre小鼠交配以產生β細胞專一性剔除TCTP基因之基因剔除鼠。進行血糖代謝相關測試與胰島組織病理切片以分析TCTP於小鼠胰島β細胞剔除後之影響。並利用免疫螢光染色法、西方墨點法與即時定量反轉錄聚合脢連鎖反應來分析胰島細胞內之蛋白質與mRNA之改變。
結果:剛出生前後之β細胞高度增生期及餵食高脂肪飲食後產生胰島素阻抗期為TCTP高度表達於β細胞之兩時期。將TCTP於小鼠胰島β細胞剔除後會導致胰島β細胞內總FoxO1、核內FoxO1及腫瘤抑制蛋白P53表達量增加，以及降低磷酸化態p70S6激酶、cyclin D2及CDK2之表達。進而降低β細胞生長與增生的能力並減少胰島β細胞團，使得胰島素分泌不足而產生高血糖現象。
結論: TCTP對於生長發育階段與胰島素阻抗階段之β細胞團建立與增生扮演重要之角色。在未來，或許可藉由調控TCTP促進β細胞團增生達到治療糖尿病的目的。

Aim: Pancreatic β-cells undergo dynamic remodeling for glucose homeostasis throughout life. In perinatal stage, they are characterized by a transient burst in proliferation to increase β-cell mass in response to the need for glucose homeostasis throughout life. In adulthood, the ability of β-cells to grow, proliferate, and expand their mass is also characteristic of pathological states of insulin resistance. Translationally controlled tumor-associated protein (TCTP), an evolutionarily highly conserved protein that is implicated in cell growth and proliferation, has been identified as a novel glucose-regulated survival-supporting protein in pancreatic β-cells in vitro. The aim of the present study is to define the in vivo role of TCTP in pancreatic β-cells during development and in insulin resistant states in mice.
Materials and methods: The expression of TCTP was assessed in pancreatic β-cells from embryo to adult mice. Mice with deletions of TCTP specific in β-cells were generated by mating TCTPfl/fl mice to rat insulin II promoter Cre recombinase mice (RIP-cre mice). Metabolic and islet morphological studies were performed to determine the effects of deleting this gene on β-cell mass and glucose homeostasis. Changes in islet proteins and mRNA were investigated by immunofluorescence, immunoblotting and quantitative RT-PCR.
Results: The expression of TCTP in pancreatic β-cells paralleled the enhanced β-cell proliferation during perinatal islet development and in insulin-resistant states of high-fat feeding. Specific knockout of TCTP in β-cells led to increased expression of total and nuclear FoxO1 and tumor suppressor P53, and decreased expression of p70S6 kinase phosphorylation and cyclin D2 and CDK2. These changes resulted in decreased β-cell proliferation and growth, reduced β-cell mass and insulin secretion; together these effects led to hyperglycemia.
Conclusions: TCTP is essential for β-cell mass expansion during development and β-cell adaptation in response to insulin resistance. TCTP may be a potential therapeutic target that could be harnessed to promote β-cell mass expansion in diabetes in the future.

目錄 1
中文摘要 4
Abstract 6
Abbreviations 8
Introduction 9
The primary cause of diabetes – the pancreatic β-cells defect 9
The pancreatic β-cell differentiation and β-cell mass establishment 10
The translationally controlled tumor-associated protein (TCTP) 11
The TCTP and β-cells mass establishment. 12
Materials and Methods 14
Production, breeding, and genotyping of conditional knockout mice 14
Metabolic studies and hormone measurement 15
Immunofluorescence and immunohistochemical staining 16
Analyses of β-cell mass, size, proliferation, apoptosis; and determinations of nuclear FoxO1+ cells percentage and α-cell/β-cell ratio 16
Islet isolation 17
Western blot analysis 18
RNA isolation and real-time reverse transcriptase polymerase chain reaction (RT-PCR) 19
Statistical analysis 20
Results 21
TCTP is transiently expressed in mouse pancreatic β-cells during embryonic and neonatal periods 21
Production of pancreatic β-cell-specific TCTP-deficient mice 21
Ablation of TCTP in pancreatic β-cells leads to progressive hyperglycemia and glucose intolerance 22
Ablation of TCTP in β-cells leads to decreased β-cell mass caused by impaired cell proliferation 23
Ablation of TCTP in pancreatic β-cell leads to diabetes after HFD feeding 24
TCTP is required for β-cell adaptation in response to insulin resistance 25
Ablation of TCTP changes FoxO1 cellular distribution and decreases p70S6K phosphorylation 26
Ablation of TCTP induces P53 activation and decreases cyclin D2, CDK2 expression 27
Discussion 29
Conclusion 33
Acknowledgment 34
References 35
Figure 1 43
Figure 2 45
Figure 3 46
Figure 4 47
Figure 5 49
Figure 6 50
Figure 7 52
Figure 8 54
Figure 9 56
Figure 10 58
Figure 11 60
Figure 12 62
Figure 13 64
Figure 14 66
Figure 15 68
Figure 16 70
Figure 17 72
Figure 18 73
Figure 19 75
Figure 20 77
Figure 21 79
Figure 22 80
Figure 23 82
Figure 24 83
Figure 25 85
Figure 26 87
Figure 27 89
Appendix 91

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