声明
摘要
ABSTRACT
CONTENTS
ABBREVIATIONS
1．Introduction
1．1．Epidemiological investigation
1．2 The pathogenesis of DGP
1．2．1 Autonomic nerve damage
1．2．2 ICC and neuronal nitric oxide synthase(nNOS)
1．2．3 Hyperglycemia
1．2．4 The gastrointestinal hormones
1．2．5 Microcirculatory disturbances
1．2．6 Helicobacter pylori(H．pylori)infection
1．3 Diagnosis and treatment
1．4 Study design
Part ⅠChanges in AMPK activity，apoptosis and energy metabolism in gastric smooth muscle in a rat model of diabetic gastroparesis
2．1 Materials
2．2 Experimental Methods
2．2．1 Diabetic rat model establishment and grouping
2．2．2 Preparation of diabetic gastroparesis model
2．2．3 Isolated muscle strip experiments
2．2．4 Immunohistochemieal staining
2．2．5 Flow eytometry detection
2．2．6 Enzyme-linked immunosorbent assay(Elisa)
2．2．7 Western blot analysis
2．3 Statistical analysis
2．4 Results
2．4．1 Comparison of general conditions，body weight and blood glucose concentrations between groups
2．4．2 Comparison of gastric residual pigment ratio，intestinal transit rate and intestinal propulsion rate between groups
2．4．3 Changes in spontaneous contraction of gastric smooth muscle in DGP rats
2．4．4 Identification of gastric smooth muscle cell
2．4．5 Comparison of gastric smooth muscle cell apoptosis rates
2．4．6 Comparison of ADP，AMP and ATP contents and the energy charge in rat gastric smooth muscle tissues between groups
2．4．7 Relative expression of phosphor-AMPK and AMPK in rat gastric smooth muscle tissues of each group
2．4．8 Ratios of phospho-LKBl／LKBl and phospho-TAKl／TAKl，and relative expression of CaMKKp in rat gastric smooth muscle tissues of each group
2．5 Discussion
Part ⅡEffect of AMPK on apoptosis and energy metabolism of rat gastric smooth muscle cells under high glucose condition
3．1 Materials
3．2 Experimental Methods
3．2．1 Primary culture of rat gastric smooth muscle cells and grouping
3．2．2 Western blot analysis
3．2．3 Confocal laser scanning microscopy detection
3．2．4 Flow cytometry detection
3．2．5 Detection of energy metabolism of rat gastric smooth muscle cells by using Seahorse XFe analyzer
3．3 Statistical analysis
3．4 Results
3．4．2 Changes in intracellular Ca2+ concentration in rat gastric smooth muscle cells under high glucose condition
3．4．3 Effect of AMPK inhibitor Compound C and agonist 2-DG on gastric smooth muscle cell apoptosis in each group under high glucose condition
3．4．4 Effects of AMPK inhibitor and agonist on mitochondrial metabolism pathway in rat gastric smooth muscle cells in each group under high glucose condition
3．4．5 Effects of AMPK inhibitor and agonist on glycolysis pathway in rat gastric smooth muscle cells in each group
3．5 Discussion
Part ⅢMechanism of action of AMPK on influencing apoptosis and energy metabolism of rat gastric smooth muscle cells under high glucose condition
4．1 Materials
4．2 Experimental Methods
4．2．2 Silencing of AMPK with RNA interference
4．2．3 Western blot analysis
4．2．4 Enzyme-linked immunosorbent assay(Eiisa)
4．3 Statistical analysis
4．4 Results
4．4．1 Phosphorylation changes of AMPK signal pathway in rat gastric smooth muscle tissues of DGP
4．4．2 The conseq uent of Silencing of AMPK with RNA interference
4．4．3 Relative expression of Akt，Phospho-AktSer473，p70S6K，Phospho-p70S6KThr389 and p53
4．4．4 Relative expression of Bcl-2，BAX and Caspase-3
4．4．5 Relative expression of 4E-BP1 and PLC-β3
4．4．6 Relative expression of CaMK Ⅱ，Phospho-CaMK Ⅱ Thr305，CaMKⅣ and Phospho-CaMKⅣThr196/200
4．4．7 PKA activity as detected by Eiisa
4．4．8 Relative expression of ACC1 and eEF2
4．5 Discussion
4．5．1 The mechanisms of effects of AMPK activation on gastric smooth muscle cell apoptosis
4．5．2 Mechanism of the effect of AMPK activation on energy metabolism in rat gastric smooth muscle cells
CONCLUTION
REFERENCES
致谢
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