TABLE 1.
In vitro studies outcomes.
| No | References | Aim | Cells and treatment | Tests | Findings | Signaling molecules/Pathways | Conclusion/correlation with IH |
|---|---|---|---|---|---|---|---|
| 1 | Nakbi et al. (2011) | To investigate the potential of HT and T on oxidative damage caused by ROS production and MMP-9 expression in PMA induced THP-1 |
Cells
THP-1 Treatment HT (1, 5, 10 and 50 μM) and T (0.05, 0.15, 0.5 and 2 mM) for 4, 15 and 24 h followed by addition of PMA (0.1 μM) |
1. Superoxide anion production 2. MMP-9 expression |
1. HT and T reduced superoxide release | ROS | HT reduced MMP-9 production that could prevent the migration of smooth muscle cell |
| 2 | Scoditti et al. (2012) | To study polyphenols effect on COX-2 and MMP-9 expression induced by pro-angiogenic factor PMA |
Cells
1. HUVEC 2. HMEC-1 Treatment HT (0.1–50 µMmol/L) |
1. Cell cytotoxicity 2. MMP-9 release 3. MMP-9 gelatinolytic activity 4. PGE2 production 5. COX-2, COX-1, b-actin, and p65 NF-kB expression 6. ROS production |
1. HT prevented inflammatory tube formation and cell migration 2. HT inhibited MMP-9 expression 3. HT inhibited COX-2 activity 4. HT decreased ROS level 5. HT suppressed translocation and transactivation of p65 NF-κB |
NF-κB | HT suppressed the ROS level and NF-κB activation that regulates the proliferation of endothelial and smooth muscle cells |
| 3 | Lamy et al. (2014) | To investigate effect of phenolic compounds toward endothelial cell angiogenesis |
Cells
1. HUVECs (HMVECs-d-Ad) Treatment 50 μM olive oil compounds for 18 followed by addition of 1 μg/ml VEGF |
1. Tube formation 2. Cell proliferation 3. Cell migration 4. VEGFR-2 phosphorylation study |
1. HT suppressed VEGF-induced tube formation 2. HT inhibited cell proliferation 3. HT inhibited phosphorylation of VEGFR-2 4. HT suppressed phosphorylation of ERK-1/2 and SAPK/JNK |
1. VEGF 2 2. ERK-1/2 3. SAPK/JNK |
HT potently suppressed ERK-1/2, SAPK and JNK pathways involved in endothelial apoptosis |
| 4 | Scoditti et al. (2014) | To study the HT effect on MMP-9 expression involved in COX-2/PGE2 pathway in PMA stimulated human monocytes stimulated |
Cells
1. PBMC 2. U937 Treatment HT (1–10 μmol/L) for 1 h followed by stimulation with 30 nmol/L PMA for 0–24 h |
1. MMP-9 and TIMP-1 secretion 2. PGE2 production 3. COX-2, COX-1, PKCa, PKCb1, NF-kB expression 4. MMP-9, COX-2, MCP-1, ICAM-1, IL-1b, TNF-a gene expression 5. NF-kB activation 6. PKC translocation |
1. HT suppressed MMP-9 secretion 2. HT reduced MMP-9 mRNA levels 3. HT suppressed PGE2 production 4. HT inactivated NF-kB 5. HT decreased MCP-1, ICAM-1, IL-1b, and TNF-α mRNA level 6. HT inactivated PKCα and PKCβ1 |
1. PGE2 2. NF-kB |
HT exhibits protection against vascular endothelial inflammation by suppressing inflammatory cytokines and activating COX-2 and PGE2 pathway |
| 5 | Zrelli et al. (2011b) | To study the potential of HT on ROS reduction by enhancing catalase activity through AMPK-FOXO3a pathway |
Cells
PPAECs Treatment HT (10, 30 and 50 μM) |
1. ROS production 2. Catalase mRNA level 3. Phosphorylation of AMPKα and AMPKβ1 4. Protein level of catalase, FOXO3a and AMPK |
1. HT reduced ROS 2. HT increased catalase expression 3. HT upregulated FOXO3a expression and mediated nuclear translocation 4. HT activated AMPK phosphorylation |
AMPK–FOXO3 | HT positively regulated endothelial oxidative defense while prevents endothelial dysfunction and apoptosis by activating AMPK-FOXO3 pathways |
| 6 | Zrelli et al. (2013) | To study the effect of hydroxytyrosol with carbon monoxide-releasing Molecule-2 in prevention of endothelial dysfunction through NO production and NFκB inactivation |
Cells
PAECs Treatment HT (1, 10, or 100) μmol/L |
1. eNOS,NFκBp65, IκBα, cleaved 2. caspase-3 expression 3. NO production 4. Cell cytotoxicity 5. Cell morphology 6. NFκB activation |
1. HT inhibited cytotoxicity 2. HT suppressed cellular damage 3. HT inhibited apoptotic morphology changes and apoptotic cell death 4. HT alone and HT + CORM-2 reduced NFκBp65 protein level 5. HT + CORM-2 increased Enos phosphorylation 6. HT + CORM-2 increased NO release 7. HT + CORM-2 blocked activation of caspase-3 8. HT alone inhibited NFκBp65 phosphorylation while CORM-2 enhanced it 9. HT + CORM-2 inactivates NFκB |
NFκB | HT + CORM-2 potentially inhibited endothelial apoptosis by inhibiting caspase 3 and NFκB pathway while supported vascular healing through NO production |
| 7 | Abe et al. (2012) | To examine the potential of olive oil phenols in inhibition of smooth muscle cell proliferation through a G1/S cell cycle block regulated by ERK1/2 |
Cells
BVSMCs Treatment HT (1, 10, or 100 μmol/L) |
1. Cell proliferation 2. Cell cycle 3. (ERK)1/2 phosphorylation |
1. HT inhibited cell proliferation 2. HT disrupted cell cycle and controlled over proliferation 3. HT inhibited ERK1/2 phosphorylation |
ERK1/2 | HT has potential to inhibit intimal hyperplasia by reducing migration and proliferation of SMC via blocking cell cycle regulated by ERK1/2 phosphorylation |
| 8 | Torul et al. (2020) | To evaluate phenolic compounds of olive extract on endothelial toxicity induced by hydrogen peroxide |
Cells
HUVECs Treatment HT (1.0–10.0 μmol/L |
1. Determination of phenolic compounds 2. Induction of ROS 3. Cell cytotoxicity |
1. HT suppressed cell toxicity 2. HT decreased ROS production |
ROS | HT shown to decrease ROS generation in endothelial which could promote vascular healing |
| 9 | Fortes et al. (2012) | To investigate effect of hydroxytyrosol and tyrosol in preventing inflammatory angiogenesis |
Cells
1. HUVECs 2. HMECs 3. BAECs Treatment HT 10 mg/ml |
1. Cell cytotoxicity 2. Cell migration 3. Tube formation 4. Cell cycle analysis 5. MMP-2 production |
1. HT inhibited cell proliferation 2. HT inhibited cell migration 3. HT suppressed tube formation 4. HT enhances apoptosis 5. HT regulated cell cycle 6. HT inhibited MMP-2 activity |
HT regulated endothelial cell cycle while decreased production of MMP-2 that possibly could prevent smooth muscle cells migration | |
| 10 | Abate et al. (2020) | To investigate the effect of HT in endothelial vascularization |
Cells
1. HUVECs 2. HVECs Treatment (0–160 µM) for 24 and 48 h |
1. Cell viability 2. Cell proliferation 3. Wound healing 4. Cell migration 5. Tube formation 6. Angiogenesis protein expression |
1. HT safe for cells up to 160 µM 2. HT enhanced wound healing process 3. HT stimulated HUVEC migration 4. HT upregulated migration and adhesion related protein expression such as ROCK, MMP-2, Phospho-Src, Src, Phospho Erk1/2, Erk1/2, RhoA, Rac1 and Ras 5. HT enhanced tube formation 6. HT upregulated VEGF) receptor 2 7. eNOS, PI3-Kinase, m-TOR, AMPK and Akt |
1. PI3K/AKT/mTor 2. Erk1/2 |
HT positively regulated vascular remodeling by promoting reendothelization and wound healing by activating PI3K/AKT/mTor pathways |
| 11 | Wang et al. (2018) | To assess the effect of HT on autophagosis of VAFs and its related signaling pathways |
Cells
VAFs Treatment HT (12.5, 25, 50, 100, 200 and 400 µM) for 1 h followed by induction of TNF-α (5 ng/ml) for 24 h |
1. Cell viability 2. SIRT1 siRNA level 3. Autophagy related protein level 4. Inflammatory cytokines level |
1. HT was shown no cytotoxicity up to 100 µM 2. HT upregulated conversion of LC3 I to LC3 II and the expression of LC3 mRNA in VAFs stimulated with TNF-α 3. HT increased protein level and mRNA expression of Beclin1 4. HT regulated the expression of SIRT1 5. HT and SIRT1 shown compatibility in molecular docking 6. HT activated Akt/mTOR signaling pathway 7. HT decreased TNF-α induced inflammatory cytokine IL-1β |
1. SIRT1 2. Akt/mTOR |
Hydroxytyrosol promoted autophagy of VAFs via SIRT1- signaling pathway and inhibited inflammatory cytokines in vascular inflammation pathophysiology |
| 12 | Cheng et al. (2017) | To study the potential of HT together with PEMFs on HUVECs proliferation |
Cells
HUVECs Treatment PEMFs at days 0, 1, 2, 3 or 4, or treated with HTY (0, 10, 30, 50, 100, 150 µM) at day 2, or treated with a combination on days 0, 1, 2 or 4 |
1. Cell viability 2. Cell migration 3. Cell apoptosis |
1. HTY + PEMF increases cell proliferation 2. HTY + PEMF enhanced cell migration 3. HTY + PEMFs prevented apoptosis 4. HTY increases mRNA and protein level of Akt, mTOR and TGF-β, but not p53 |
1. Akt 2. mTOR 3. TGF-β |
PEMFs and HTY enhanced endothelial migration and proliferation that could promote reendothelization in vascular remodeling |
| 13 | Kouka et al. (2017) | To examine antioxidant property of pure HT from EVOO phenolic fraction |
Cells
1. EA. hy926 2. C2C12 Treatment HT (0–40 μg/ml) |
1. Extraction of TPF from EVOO 2. Purification of HT from TPF 3. radical scavenging assay 4. Cell viability 5. Assessment of GSH and ROS levels |
1. HT exhibited highest antioxidant DPPH 2. HT reduced ROS 3. HT increased GSH |
HT found to have decreased ROS and increased GSH which possibly enhance endothelial proliferation and functioning | |
| 14 | Kitsati et al. (2016) | To assess the potential of HT in rescuing cells from oxidative stress induced by H2O2 |
Cells
Jurkat cells Treatment HT (0.05 and 0.1 mM) for 30 min |
1. Comet assay 2. Labile iron level 3. H2O2 generation |
1. HT inhibited H2O2 induced labile iron level 2. Hydroxytyrosol inhibits H2O2-induced and mitochondrial-mediated apoptosis 3. Hydroxytyrosol inhibits H2O2-induced apoptosis 4. inhibits H2O 5. HT inhibited phosphorylation and activation of the JNK and p38 MAPKs |
1. JNK 2. p38 MAPKs |
HT prevented cellular apoptosis by inactivating JNK and p38 MAPKs pathway |
| 15 | Zrelli et al. (2015) | To examine the action of hydroxytyrosol in the vascular wound healing mechanism |
Cells
PPAECs Treatment HT (10–100 μM) 0–24 h |
1. Expression of HO-1 and Nrf2 2. Wound healing |
1. HT inclined HO-1 mRNA and protein level 2. HT induced HO-1 expression supported by PI3K/Akt and ERK1/2 3. HT mediated Nrf2 expression and nuclear localization |
1. PI3K/Akt 2. ERK1/2 3. Nrf2 |
HT enhanced wound healing process in endothelial through activating expression of HO-1 and Nrf2 |
| 16 | Zrelli et al. (2011a) | To study the effect of HT in vascular smooth muscle cell VSMCs proliferation |
Cells
RVSMCs Treatment HT (10, 30, and 100 µM) with and without 20 ng/mL of PDGF |
1. Cell migration 2. Cell viability 3. NO production 4. Akt phosphorylation |
1. HT decreased the number of viable VSMCs either in the presence or not of PDGF 2. HT promotes VSMCs apoptosis 3. HT increased NO production 4. HT increased iNOS protein expression 5. HT dephosphorylate Akt 6. PP2A mediated HT induced Akt phosphorylation |
1. Akt 2. PPA |
HT prevents VSMCs apoptosis through NO production and Akt dephosphorylation via activation of PP2A |
| 17 | Zrelli et al. (2011b) | To assess the proliferation and protective effect of HT on oxidative injury induced VECs injury |
Cells
PPAECs Treatment HT (10–100 µM) for 24 h followed by 0–700 3M) of H2O2 for 24 h |
1. Cell viability 2. Wound healing 3. HO-1 mRNA expression 4. phosphorylation of Akt, p38 MAPK, and ERK1/2 5. ROS production |
1. HT enhanced cell proliferation 2. HT repaired wound healing 3. HT prevented H2O2-Induced cytotoxicity 4. HT-induced phosphorylation of Akt, p38 MAPK, and ERK1/2 5. HT accumulates Nrf2 in nucleus 6. HT reduced ROS generation 7. HT increased mRNA and protein level of HO-1 |
1. Akt 2. MAPK 3. ERK1/2 4. Nrf2 |
HT protects VECs from oxidative damage through activation of the PI3K/Akt and ERK1/2 pathways |
| 18 | Catalan et al. (2015) | To evaluate the effect of hydroxytyrosol and its plasma metabolites toward endothelial protection |
Cells
HAEC Treatment HT (1, 2, 5, and 10 µM) co-incubated with TNF -α (10 ng/ml) for 18 and 24 h |
1. HT metabolites production 2. Adhesion molecules production 3. Chemokine protein production 4. Cytotoxicity |
1. HT and HT metabolites reduced E-selectin, P-selectin, VCAM-1, and ICAM-1 2. HT metabolites only reduced MCP-1 |
HT and HT metabolites exhibited vascular protection by reducing endothelial inflammation cytokines | |
| 19 | Terzuoli et al. (2020) | To investigate the HT-3Os effects on endothelial-to-mesenchymal transition (EndMT) in the inflamed endothelium | Cells 1. EC 2. HUVEC 3. HREC Treatment 1. IL-1β (10 ng/ml) with or without HT-3Os (10 μM, every 24 h for 7 days |
1. Morphology evaluation 2. Immunomarkers detection 3. Cytoplasmic and nuclear protein detection 4. miRNA expression analysis 5. Cytotoxicity |
1. HT-3Os reverses EndMT-phenotypic changes induced by IL-1β 2. HT-3Os restores let-7 miRNA expression and inhibits TGF-β signaling 3. HT-3Os upregulated CD31 in IL-1β induced HUVEC and HREC 4. HT-3Os decreased fibroblast markers as FN1 and VIM or SMCin IL-1β induced HUVEC and HREC) 5. HT-3Os upregulated NOTCH3 and MMP2 and MMP9 |
1. let-7 miRNA 2. MMP 2 3. MMP 9 |
HT-3Os halts EndMT process in inflamed EC, by increasing let-7 miRNA expression and preventing activation of TGF-β signaling |