519c88d8e2bb107eeda2192886757519 284461.pdf 527ffd224d6a7ff67b31bfdf20b530530460ccc4 284461.pdf edf095b8ce21720f6b3654b5cf1c554f07a0d6d4a8cf31f4485520c00fcc3873 284461.pdf Title: Involvement of the PI3K/AKT Intracellular Signaling Pathway in the AntiCancer Activity of Hydroxytyrosol, a Polyphenol from Olea europaea, in Hematological Cells and Implication of HSP60 Levels in Its Anti-Inflammatory Activity "2279 Subject: Hydroxytyrosol (HT), the main representative of polyphenols of olive oil, has been described as one of the most powerful natural antioxidants, also showing anti-inflammatory, antimicrobial, cardioprotective and anticancer activity in different type of cancers, but has been little studied in hematological neoplasms. The objective of this work was to evaluate the anticancer potential of HT in acute human leukemia T cells (Jurkat and HL60) and the anti-inflammatory potential in murine macrophages (Raw264.7). For this, cytotoxicity tests were performed for HT, showing IC50 values, at 24 h, for Jurkat, HL60 and Raw264.7 cells, of 27.3 gmL-1, 109.8 gmL-1 and 45.7 gmL-1, respectively. At the same time, HT caused cell arrest in G0/G1 phase in both Jurkat and HL60 cells by increasing G0/G1 phase and significantly decreasing S phase. Apoptosis and cell cycle assays revealed an antiproliferative effect of HT, decreasing the percentage of dividing cells and increasing apoptosis. Furthermore, HT inhibited the PI3K signaling pathway and, consequently, the MAPK pathway was activated. Inflammation tests revealed that HT acts as an anti-inflammatory agent, reducing NO levels in Raw264.7 cells previously stimulated by lipopolysaccharide (LPS). These processes were confirmed by the changes in the expression of the main markers of inflammation and cancer. In conclusion, HT has an anticancer and anti-inflammatory effect in the cell lines studied, which were Raw264.7, Jurkat, and HL60, and could be used as a natural drug in the treatment of liquid cancers, leukemias, myelomas and lymphomas. Keywords: apoptosis; cell cycle arrest; HSP60; hydroxytyrosol; inflammation; leukemia cells; PI3K/AKT signaling pathway; ROS level Author: Alberto M. Parra-Perez, Amalia Pérez-Jiménez, Isabel Gris-Cárdenas, Gloria C. Bonel-Pérez, Luis M. Carrasco-Díaz, Khalida Mokhtari, Leticia García-Salguero, José A. Lupiáñez and Eva E. Rufino-Palomares Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Tue Jul 5 10:45:57 2022 CEST ModDate: Tue Jul 5 10:50:07 2022 CEST Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 21 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 2981371 bytes Optimized: no PDF version: 1.7 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- IOJCNK+VnURWPalladioL-Bold Type 1 Custom yes yes no 10 0 JDWIZJ+EURM10 Type 1 Builtin yes yes no 15 0 XPHJAJ+URWPalladioL-Bold Type 1 Custom yes yes no 19 0 ONBVPV+URWPalladioL-Roma Type 1 Custom yes yes no 24 0 QZBRZC+URWPalladioL-Ital Type 1 Custom yes yes no 28 0 PGGYTP+URWPalladioL-BoldItal Type 1 Custom yes yes no 32 0 OJYQXC+CMSY10 Type 1 Builtin yes yes no 36 0 MEJHZL+VnURWPalladioL Type 1 Custom yes yes no 40 0 IRWHZX+TimesNewRomanPSMT Type 1C WinAnsi yes yes no 58 0 CKUPDE+TimesNewRomanPS-BoldItalicMT 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0 FNTSBS+PalatinoLinotype-Roman CID TrueType Identity-H yes yes yes 313 0 FNTSBS+PalatinoLinotype-Italic CID TrueType Identity-H yes yes yes 319 0 FNTSBS+SymbolMT CID TrueType Identity-H yes yes yes 325 0 FNTSBS+PalatinoLinotype-Bold CID TrueType Identity-H yes yes yes 339 0 FNTSBS+PalatinoLinotype-Roman CID TrueType Identity-H yes yes yes 345 0 FNTSBS+PalatinoLinotype-Italic CID TrueType Identity-H yes yes yes 351 0 PYTLFY+EURB10 Type 1 Builtin yes yes no 384 0 Jhove (Rel. 1.28.0, 2023-05-18) Date: 2024-02-22 02:33:45 CET RepresentationInformation: 284461.pdf ReportingModule: PDF-hul, Rel. 1.12.4 (2023-03-16) LastModified: 2024-02-21 12:41:39 CET Size: 2981371 Format: PDF Version: 1.7 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 644 FreeObjects: 1 IncrementalUpdates: 0 DocumentCatalog: PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: Introduction Destination: section.1 Item: Title: Results Destination: section.2 Children: Item: Title: Hydroxytyrosol Has a Cytotoxic Effect on Different Blood Cell Lines Destination: subsection.2.1 Item: Title: Hydroxytyrosol Induces G0/G1 Arrest in Jurkat and HL60 Cells Destination: subsection.2.2 Item: Title: Hydroxytyrosol Enhances Apoptosis in Jurkat and HL60 Cells Destination: subsection.2.3 Item: Title: Hydroxytyrosol Increases ROS Production in Jurkat and HL60 Cells Destination: subsection.2.4 Item: Title: Hydroxytyrosol Produces up and Down-Regulation of Target Proteins of Apoptosis and Proliferation Processes Destination: subsection.2.5 Item: Title: Hydroxytyrosol Modulates Dual PI3K/MAPK Signaling Pathway in Jurkat Cells Destination: subsection.2.6 Item: Title: Hydroxytyrosol Reduces Inflammation Levels in Raw264.7 Cells Destination: subsection.2.7 Item: Title: Discussion Destination: section.3 Item: Title: Materials and Methods Destination: section.4 Children: Item: Title: Cell Culture Destination: subsection.4.1 Item: Title: Natural Extract Destination: subsection.4.2 Item: Title: Cytotoxicity Assays Destination: subsection.4.3 Item: Title: Cell Cycle Analysis Destination: subsection.4.4 Item: Title: Apoptosis Assay Destination: subsection.4.5 Item: Title: Measurement of ROS Production Destination: subsection.4.6 Item: Title: Preparation of Cytosolic Extracts Destination: subsection.4.7 Item: Title: Western Blotting Destination: subsection.4.8 Item: Title: PI3K/MAPK Dual Pathway Activation Assay Destination: subsection.4.9 Item: Title: Study of Inflammation Using the Griess Method Destination: subsection.4.10 Item: Title: Statistical Analysis Destination: subsection.4.11 Item: Title: Conclusions Destination: section.5 Item: Title: References Destination: section.5 Info: Title: Involvement of the PI3K/AKT Intracellular Signaling Pathway in the AntiCancer Activity of Hydroxytyrosol, a Polyphenol from Olea europaea, in Hematological Cells and Implication of HSP60 Levels in Its Anti-Inflammatory Activity "2279 Author: Alberto M. Parra-Perez, Amalia Pérez-Jiménez, Isabel Gris-Cárdenas, Gloria C. Bonel-Pérez, Luis M. Carrasco-Díaz, Khalida Mokhtari, Leticia García-Salguero, José A. Lupiáñez and Eva E. Rufino-Palomares Subject: Hydroxytyrosol (HT), the main representative of polyphenols of olive oil, has been described as one of the most powerful natural antioxidants, also showing anti-inflammatory, antimicrobial, cardioprotective and anticancer activity in different type of cancers, but has been little studied in hematological neoplasms. The objective of this work was to evaluate the anticancer potential of HT in acute human leukemia T cells (Jurkat and HL60) and the anti-inflammatory potential in murine macrophages (Raw264.7). For this, cytotoxicity tests were performed for HT, showing IC50 values, at 24 h, for Jurkat, HL60 and Raw264.7 cells, of 27.3 gmL-1, 109.8 gmL-1 and 45.7 gmL-1, respectively. At the same time, HT caused cell arrest in G0/G1 phase in both Jurkat and HL60 cells by increasing G0/G1 phase and significantly decreasing S phase. Apoptosis and cell cycle assays revealed an antiproliferative effect of HT, decreasing the percentage of dividing cells and increasing apoptosis. Furthermore, HT inhibited the PI3K signaling pathway and, consequently, the MAPK pathway was activated. Inflammation tests revealed that HT acts as an anti-inflammatory agent, reducing NO levels in Raw264.7 cells previously stimulated by lipopolysaccharide (LPS). These processes were confirmed by the changes in the expression of the main markers of inflammation and cancer. In conclusion, HT has an anticancer and anti-inflammatory effect in the cell lines studied, which were Raw264.7, Jurkat, and HL60, and could be used as a natural drug in the treatment of liquid cancers, leukemias, myelomas and lymphomas. 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