Objective To investigate the role of lysine-specific demethylase 1 （LSD1） in the process of THP-1 monocyte-to-macrophage differentiation. Methods Quantitative reverse transcription-polymerase chain reaction （qRT-PCR） and Western blotting were performed to analyze the expression of LSD1 and interleukin-6 （IL-6） in THP-1 monocytes and THP-l-derived macrophages. Chromatin immunoprecipitation （ChiP） assay was applied to detect the occupancy of LSD1 and H3K4 methylation at IL-6 promoter during THP-1 monocyte-to-macrophage differentiation. IL-6 mRNA level and H3K4 methylation at IL-6 promoter were analyzed using qRT-PCR and ChiP assay in LSD 1 -knockdown THP- 1 cells treated with 12-O-tetradecanoylphorbol- 13-acetate （TPA） for 0 4, 8, 12, and 24 hours. Fluorescence activated flow cytometry was performed to reveal the percentage of macrophages differentiated from THP- 1 monocytes. Results The expression of LSD1 reduced during THP-1 monocyte-to-macrophage differentiation （P〈0.01）. LSD1 occupancy decreased and H3K4 methylation increased at IL-6 promoter during the differentiation. With knockdown of LSD1, H3K4 methylation at IL-6 promoter was found increased after TPA treatment at different times points （all P〈0.05, except 24 hours）. The percentage of macrophages increased significantly in theTHP-I cells with LSD1 knockdown （P〈0.05）. Conclusions LSD1 is repressed during the monocyte-to-macrophage differentiation of THP-1 cells. Suppression of LSD 1-mediated H3K4 demethylation may be required for THP-1 monocyte-to-macrophage differentiation.
Rui-feng YangGuo-wei ZhaoShu-ting LiangHou-zao ChenDe-pei Liu
SIRT1,a mammalian ortholog of yeast silent information regulator 2(Sir2),is an NAD+-dependent protein deacetylase that plays a critical role in the regulation of vascular function.The current study aims to investigate the functional significance of deacetylase activity of SIRT1 in heart.Here we show that the early postnatal hearts expressed the highest level of SIRT1deacetylase activity compared to adult and aged hearts.We generated transgenic mice with cardiac-specific expression of a dominant-negative form of the human SIRT1(SIRT1H363Y),which represses endogenous SIRT1 activity.The transgenic mice displayed dilated atrial and ventricular chambers,and died early in the postnatal period.Pathological,echocardiographic and molecular phenotype confirmed the presence of dilated cardiomyopathy.Terminal deoxynucleotidyl transferase-mediated dUTP nick-end-labeling analysis revealed a greater abundance of apoptotic nuclei in the hearts of transgenic mice.Furthermore,we show that cardiomyocyte apoptosis caused by suppression of SIRT1 activity is,at least in part,due to increased p53acetylation and upregulated Bax expression.These results indicate that dominant negative form of SIRT1(SIRT1H363Y)overexpression in mouse hearts causes cardiomyocyte apoptosis and early-onset heart failure,suggesting a critical role of SIRT1 in preserving normal cardiac development during the early postnatal period.
