1dea4aad2aeb73a5579f00683355ffac pmc_33028031.pdf 5e41d394fa0505b711dded5c65dc2bebe88878c9 pmc_33028031.pdf 07715f6b5b188c0aa26cad33c42d53b92ad65d6f613595821c45b4fcdf61bb53 pmc_33028031.pdf Title: Role of the Scavenger Receptor CD36 in Accelerated Diabetic Atherosclerosis Subject: Diabetes mellitus entails increased atherosclerotic burden and medial arterial calcification, but the precise mechanisms are not fully elucidated. We aimed to investigate the implication of CD36 in inflammation and calcification processes orchestrated by vascular smooth muscle cells (VSMCs) under hyperglycemic and atherogenic conditions. We examined the expression of CD36, pro-inflammatory cytokines, endoplasmic reticulum (ER) stress markers, and mineralization-regulating enzymes by RT-PCR in human VSMCs, cultured in a medium containing normal (5 mM) or high glucose (22 mM) for 72 h with or without oxidized low-density lipoprotein (oxLDL) (24 h). The uptake of 1,1'-dioctadecyl-3,3,3',3-tetramethylindocarbocyanine perchlorate-fluorescently (DiI) labeled oxLDL was quantified by flow cytometry and fluorimetry and calcification assays were performed in VSMC cultured in osteogenic medium and stained by alizarin red. We observed induction in the expression of CD36, cytokines, calcification markers, and ER stress markers under high glucose that was exacerbated by oxLDL. These results were confirmed in carotid plaques from subjects with diabetes versus non-diabetic subjects. Accordingly, the uptake of DiI-labeled oxLDL was increased after exposure to high glucose. The silencing of CD36 reduced the induction of CD36 and the expression of calcification enzymes and mineralization of VSMC. Our results indicate that CD36 signaling is partially involved in hyperglycemia and oxLDL-induced vascular calcification in diabetes. Keywords: scavenger receptor CD36; atherosclerosis; diabetes; inflammation; vascular calcification Author: Miquel Navas-Madroñal, Esmeralda Castelblanco, Mercedes Camacho, Marta Consegal, Anna Ramirez-Morros, Maria Rosa Sarrias, Paulina Perez, Nuria Alonso, María Galán and Dídac Mauricio Creator: LaTeX with hyperref package Producer: pdfTeX-1.40.18 CreationDate: Mon Oct 5 16:13:02 2020 CEST ModDate: Mon Oct 5 16:13:11 2020 CEST Custom Metadata: yes Metadata Stream: yes Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 19 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 1291559 bytes Optimized: yes PDF version: 1.5 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- SSNIUI+VnURWPalladioL-Bold Type 1 Custom yes yes no 666 0 UBVQAP+Pxsy Type 1 Builtin yes yes no 669 0 BQUNXL+URWPalladioL-Ital Type 1 Custom yes yes no 673 0 UMCILL+URWPalladioL-Bold Type 1 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application/pdf Profile: Linearized PDF PDFMetadata: Objects: 0 FreeObjects: 1 IncrementalUpdates: 0 DocumentCatalog: ViewerPreferences: HideToolbar: false HideMenubar: false HideWindowUI: false FitWindow: true CenterWindow: false DisplayDocTitle: false NonFullScreenPageMode: UseNone Direction: L2R ViewArea: CropBox ViewClip: CropBox PrintArea: CropBox PageClip: CropBox PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: Introduction Item: Title: Results Children: Item: Title: Hyperglycemic and Atherogenic Conditions Synergistically Induced the Expression of Scavenger Receptor CD36, and Exposure to High Glucose Enhances oxLDL Uptake in VSMC Item: Title: Exposure to HG Triggers Calcification in VSMC Item: Title: Silencing of CD36 Decreased ox-LDL Uptake and Limited Osteoblastic Differentiation of VSMC Item: Title: Expression of CD36 and ER Stress Indicators in Carotid Plaques from Subjects with and without Diabetes Item: Title: Discussion Item: Title: Methods Children: Item: Title: Human Samples Item: Title: Cell Culture Item: Title: Transfections with Small Interfering RNA (siRNA) Item: Title: Total mRNA and Protein Isolation from Tissue and Cells Item: Title: Quantitative Real-Time PCR Item: Title: OxLDL Uptake Item: Title: In Vitro Calcification Item: Title: Western Blot Item: Title: ELISA Item: Title: Immunostaining Item: Title: Determination of 7-ketocholesterol in Plasma Item: Title: Statistical Analysis Item: Title: Conclusions Item: Title: References Filters: FilterPipeline: FlateDecode Fonts: Type1: Font: BaseFont: BQUNXL+URWPalladioL-Ital FontSubset: true FirstChar: 2 LastChar: 121 FontDescriptor: FontName: BQUNXL+URWPalladioL-Ital Flags: Symbolic FontBBox: -170, -305, 1010, 941 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: JDWIZJ+EURM10 FontSubset: true FirstChar: 11 LastChar: 22 FontDescriptor: FontName: JDWIZJ+EURM10 Flags: Symbolic FontBBox: -32, -243, 1060, 720 FontFile: true Font: BaseFont: UMCILL+URWPalladioL-Bold 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SSNIUI+VnURWPalladioL-Bold FontSubset: true FirstChar: 161 LastChar: 189 FontDescriptor: FontName: SSNIUI+VnURWPalladioL-Bold Flags: Symbolic FontBBox: -152, -301, 1000, 935 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: UBVQAP+Pxsy FontSubset: true FirstChar: 2 LastChar: 121 FontDescriptor: FontName: UBVQAP+Pxsy Flags: Symbolic FontBBox: -21, -994, 1292, 912 FontFile: true XMP: application/pdf Miquel Navas-Madroñal, Esmeralda Castelblanco, Mercedes Camacho, Marta Consegal, Anna Ramirez-Morros, Maria Rosa Sarrias, Paulina Perez, Nuria Alonso, María Galán and Dídac Mauricio Diabetes mellitus entails increased atherosclerotic burden and medial arterial calcification, but the precise mechanisms are not fully elucidated. We aimed to investigate the implication of CD36 in inflammation and calcification processes orchestrated by vascular smooth muscle cells (VSMCs) under hyperglycemic and atherogenic conditions. We examined the expression of CD36, pro-inflammatory cytokines, endoplasmic reticulum (ER) stress markers, and mineralization-regulating enzymes by RT-PCR in human VSMCs, cultured in a medium containing normal (5 mM) or high glucose (22 mM) for 72 h with or without oxidized low-density lipoprotein (oxLDL) (24 h). The uptake of 1,1'-dioctadecyl-3,3,3',3-tetramethylindocarbocyanine perchlorate-fluorescently (DiI) labeled oxLDL was quantified by flow cytometry and fluorimetry and calcification assays were performed in VSMC cultured in osteogenic medium and stained by alizarin red. We observed induction in the expression of CD36, cytokines, calcification markers, and ER stress markers under high glucose that was exacerbated by oxLDL. These results were confirmed in carotid plaques from subjects with diabetes versus non-diabetic subjects. Accordingly, the uptake of DiI-labeled oxLDL was increased after exposure to high glucose. The silencing of CD36 reduced the induction of CD36 and the expression of calcification enzymes and mineralization of VSMC. Our results indicate that CD36 signaling is partially involved in hyperglycemia and oxLDL-induced vascular calcification in diabetes. 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