Journal: Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease

Article Title: Coronary Microvascular Dysfunction Is Associated With Augmented Lysosomal Signaling in Hypercholesterolemic Mice

doi: 10.1161/JAHA.124.037460

Figure Lengend Snippet: MCECs were cultured in low glucose DMEM with 5% FBS and treated with 7‐ketocholesterol for the indicated times. A and D , Representative immunofluorescence images and quantification show mitochondrial superoxide levels. Representative immunofluorescence images and quantification show the expression of proinflammatory proteins VCAM‐1 ( B and E ) and CCL2 ( C and F ). G , Representative immunofluorescence images and quantification show the nuclear TFEB positive percentage. Nuclei were stained with DAPI. H , Representative immunoblots and summarized data show the effects of 7‐ketocholesterol on the protein expression levels of microtubule‐associated proteins light chain 3‐II. I , Real‐time reverse transcription polymerase chain reaction analyses of TFEB, LAMP‐1, LAMP‐2A, beclin‐1, microtubule‐associated proteins light chain 3, and p62/SQSTM1 mRNA levels after treatment with 0 or 40 μM 7‐ketocholesterol for 24 hour. Scale bar=20 μm. * vs 0, P <0.05 (n=4–5). CCL2 indicates C‐C motif chemokine ligand 2; LAMP‐1, lysosomal‐associated membrane protein; MCECs, mouse cardiac endothelial cells; TFEB, transcriptional factor EB; and VCAM‐1, vascular cell adhesion molecule 1.

Article Snippet: MCECs (Cedarlane, CLU510) were cultured in low glucose DMEM with 5% FBS, 1% penicillin/streptomycin, and 1 mmol/L of HEPES at 37 °C with 5% CO 2 .

Techniques: Cell Culture, Immunofluorescence, Expressing, Staining, Western Blot, Reverse Transcription, Polymerase Chain Reaction, Membrane

Journal: Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease

Article Title: Coronary Microvascular Dysfunction Is Associated With Augmented Lysosomal Signaling in Hypercholesterolemic Mice

doi: 10.1161/JAHA.124.037460

Figure Lengend Snippet: MCECs were cultured and treated in low glucose DMEM with 5% FBS, pretreated with or without 50 nM BAF for 1 hour, and then cotreated with or without 40 μM of 7‐ketocholesterol for 6 hour. A , Representative immunofluorescence images and quantification show the nuclear TFEB‐positive percentage. Nuclei were stained with DAPI. B through G , MCECs are treated in low‐glucose DMEM with 1% FBS for 2 hour before pretreatment with or without 50 nM of BAF for 1 hour, and then the cells are cotreated with or without 40 μM of 7‐ketocholesterol for 24 hour. B , Representative images of VCAM‐1 and summarized data. C through G , Representative images of FLICA/PI staining and summarized data. H , Cell numbers were detected by using CCK8 kit. Scale bar=20 μm. * vs 0, # vs BAF or 7‐ketocholesterol, P <0.05 (n=4). BAF indicates bafilomycin A1; CCK8, Cell‐Counting Kit 8; MCECs, mouse cardiac endothelial cells; PI, propidium iodide; TFEB, transcriptional factor EB; and VCAM‐1, vascular cell adhesion molecule 1.

Article Snippet: MCECs (Cedarlane, CLU510) were cultured in low glucose DMEM with 5% FBS, 1% penicillin/streptomycin, and 1 mmol/L of HEPES at 37 °C with 5% CO 2 .

Techniques: Cell Culture, Immunofluorescence, Staining, Cell Counting

Journal: Journal of the American Heart Association: Cardiovascular and Cerebrovascular Disease

Article Title: Coronary Microvascular Dysfunction Is Associated With Augmented Lysosomal Signaling in Hypercholesterolemic Mice

doi: 10.1161/JAHA.124.037460

Figure Lengend Snippet: MCECs were cultured in low glucose DMEM with 5% FBS, then treated with EZE with or without 7‐ketocholesterol for the indicated time. A , Representative immunofluorescence images and quantification show the effect of ezetimibe on nuclear TFEB positive percentage. B , Representative immunofluorescence images and quantification show the effect of ezetimibe and 7‐ketocholesterol on nuclear TFEB positive percentage. Representative immunofluorescence images and quantification of mitochondrial superoxide ( C ), proinflammatory proteins vascular cell adhesion molecule 1 (VCAM‐1) ( D ) and CCL2 ( E ), and monocyte adhesion ( F ). Scale bar=20 μm. * vs 0, # vs 7‐keto, P <0.05 (n=4). CCL2 indicates C‐C motif chemokine ligand 2; EZE, ezetimibe; MCECs, mouse cardiac endothelial cells; TFEB, transcriptional factor EB; and VCAM‐1, vascular cell adhesion molecule 1.

Article Snippet: MCECs (Cedarlane, CLU510) were cultured in low glucose DMEM with 5% FBS, 1% penicillin/streptomycin, and 1 mmol/L of HEPES at 37 °C with 5% CO 2 .

Techniques: Cell Culture, Immunofluorescence