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The Influence of Poly ADP-ribose Polymerase 1 (PARP-1) on HMGB1 Localization and Secretion in RAW264.7 Cell

Author: YuanWeiWei
Tutor: WuHeShui
School: Huazhong University of Science and Technology
Course: Surgery
Keywords: High mobility group box protein 1 (HMGB1) Poly ( ADP-ribose ) -based polymerase 1 (PARP-1) Lipopolysaccharide (LPS) Nuclear localization sequence (NLS) 3 - aminobenzamide (3-AB)
CLC: R576
Type: Master's thesis
Year: 2011
Downloads: 14
Quote: 0
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Abstract


Objective: To study the poly ( ADP-ribose ) - polymerase 1 (PARP-1) of HMGB1 positioning RAW264.7 cells and the secretion of HMGB1 from the nucleus to the cytoplasm shift from subcellular structures observed for the study of severe treatment of acute pancreatitis is to provide a new strategy . : 100ng/ml of LPS treatment of RAW264.7 cells 1, 2, 4, 8 and 16h , use the time pattern of the cell ELISA determination stimulated intracellular PARP-1 activity ; using LPS (100ng/ml), LPS 3 -AB (10mmol / l) RAW264.7 cells stimulated 1,2,4,8 and 16h time intracellular localization of HMGB1 variation observed under a fluorescence microscope using immunofluorescence , using Western-blot detection of HMGB1 in the level in the cell culture supernatant ; build HMGB1-EGFP (wild type, WT) of the expression plasmid and HMGB1NLS region 40,47,179 in the glutamic acid mutation to alanine that HMGB1 - EGFP ( Mutated type , MT), of the the expression plasmid, RAW264.7 cells by transient transfection , cells treated with 100ng/ml of LPS after transfection with HMGB1 in the intracellular localization was observed under a fluorescence microscope . Results: LPS stimulation RAW264.7 cells , PARP-1 activation and activity enhancement , 4 hours after the stimulation reached a peak , and then the activity decreases gradually ; HMGB1 h after stimulation by the nucleus to the cytoplasm of the shift , with the stimulation time, and gradually to the cytoplasm agglomeration; HMGB1 from the nucleus to the cytoplasm of the shift inhibition of PARP-1 activity can be reduced , inhibition of HMGB1 to extracellular secretion ; of HMGB1 NLS region glutamate mutations , not weaken HMGB1 shift from the nucleus to the cytoplasm , HMGB1 cell localization . Conclusion : PARP-1 involved in HMGB1 positioning and secretion in RAW264.7 cells , which by HMGB1 protein ADP-ribose base modification , regulation of HMGB1 in the cells the positioning and extracellular secretion . HMGB1 NLS region glutamate mutant the HMGB1 cells positioned within no effect. By inhibiting the activity of PARP-1 , 3-AB thereby inhibiting ADP - ribosylation of the HMGB1 modified , thereby inhibiting HMGB1 a shift to the cytoplasm and extracellular secretion process . PARP-1 inhibitors can inhibit HMGB1 to intracellular localization and extracellular secretion , provides a new way of thinking for the treatment of HMGB1 - induced inflammatory response .

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CLC: > Medicine, health > Internal Medicine > Digestive and abdominal diseases > Pancreatic diseases
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