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The Effects of Mechanical Ventilation on the Expression of Muc5ac in Airway Epithelial Cells and Its Signal Mechanism

Author: ZhongTian
Tutor: ZhouXiangDong
School: Chongqing Medical University
Course: Internal Medicine
Keywords: Mechanical Stretch The tension sensitivity cation channel Free calcium Mitogen-activated protein kinase Mucin
CLC: R363.1
Type: PhD thesis
Year: 2011
Downloads: 27
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Abstract


Objective observation ventilator mechanical ventilation, mechanical stretch Rabbit airway mucosa and airway mucosal epithelial cells (HBE16) mucin (MUC) 5AC expression and its signaling pathway mechanisms discussed. Methods Japanese rabbits were randomly divided into a control group, mechanical ventilation, 1, 3, 6, 12 and 24h group and 2,5 10cmH2O end-expiratory pressure (PEEP) group. Take bronchial alveolar lavage fluid (BALF) was detected by ELISA and RT-PCR method BALF tumor necrosis factor (TNF)-α, interleukin (IL) -8, MUC5AC protein and mRNA levels, nuclear grain count in BALF The cells, substrate detection method for the determination of the activity of a neutrophil elastase (NE). Airway mucosal epithelial cells (HBE16) cultured in vitro, using a small biological impact machines give mechanical stretch stimulation, cultured cells in each group according to impose conditions and divided into control, stretch, stretch gadolinium, stretch nifedipine Distraction independent of the low-molecular-weight heparin, the distraction domain of MARCKS effect (ED) locked nucleic acid (LNA), and stretch MARCKS ED control the LNA sequence of seven groups, using flow cytometry stretch before and after intracellular free of Ca in < / sup> changes in fluorescence intensity, laser confocal microscope cell intracellular free Ca 2 distribution, respectively, by reverse transcription polymerase chain reaction (RT-PCR) and immunofluorescence staining and extracellular spit synaptic membrane protein SNAP23 as well as the sticky protein (MUC) 5AC mRNA and protein expression, enzyme-linked immunosorbent assay (ELISA) was used to detect the cell culture supernatant MUC5AC secretion. Then built airway mucosal epithelial cell stretch model, were given the mitogen-activated protein kinase (MAPK) signaling pathway in three major pathway inhibitors: JNK inhibitor (SP600125), p38 protein kinase inhibitor (SB203580) and ERK1 / 2 inhibitor (U0126) (concentration was 30μmol / L), and set no mechanical stretch for the control group and the group of purely mechanical stretch. MUC5AC mRNA expression and protein secretion was detected by RT-PCR and ELISA, respectively. The results of mechanical ventilation can significantly enhance MUC5AC secretion (P lt; 0.05). Ventilation 3h TNF-α expression to a peak, and then decreased (P lt; 0.05). Ventilation 6h IL-8 expression to the peak, followed by a gradual decline (P lt; 0.05). Ventilation 12h nuclear granulocyte count and granulocyte elastase activity was significantly increased (P lt; 0.05) 24 hours after falling back. With the the PEEP increase of TNF-alpha, IL-8, MUC5ACmRNA expression and protein content, the number of polymorphonuclear granulocyte and NE activity with an increasing (P lt; 0.05). Mechanical stretch significantly increased airway mucosal epithelial cells intracellular Ca 2 concentration (P lt; 0.01); while significantly higher airway mucosal epithelial cells the SNAP23 and MUC5AC expression, significantly enhance cell culture supernatant MUC5AC secretion (P lt; 0.05); gadolinium, nifedipine, MARCKS ED-LNA inhibited mechanical stretch SNAP23 expression and MUC5AC expression, to enhance the role of the secretion (P lt; 0.05); while Low-molecular-weight heparin did not significantly reduce the SNAP23 expression and MUC5AC expression and secretion (P gt; 0.05). U0126 and SB203580 inhibited stretch-induced increase in MUC5AC mRNA and protein expression (both P lt; 0.01). The conclusions mechanical ventilation and inlet mucosal epithelial cells MUC5AC mRNA expression and secretion of MUC5AC distraction can promote the inflammatory response, the mechanism may be triggered by mechanical ventilation and stretch, tension-sensitive cation channels, Ca 2

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