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Sains Malaysiana 53(3)(2024): 591-604
http://doi.org/10.17576/jsm-2024-5303-09
Thymoquinone Reverses Homocysteine-Induced Endothelial
Dysfunction via Inhibition of Endoplasmic Reticulum-Stress Induced Oxidative
Stress Pathway
(Timoquinon Membalikkan Disfungsi Endotelium Aruhan Homosistein melalui Perencatan Laluan Tekanan Oksidatif Aruhan Retikulum Endoplasma)
SITI SARAH M. SOFIULLAH1,
DHARMANI DEVI MURUGAN2, SUHAILA ABD MUID3,4, WU YUAN SENG5,6,
NOR HISAM ZAMAKSHSHARI 7, QUAN FU GAN8, MELONNEY PATRICK 3,4,
NORASIKIN AB AZIS9, SRINIVASA RAO SIRASANAGANDLA10 & CHOY KER WOON1,*
1Department of Anatomy, Faculty of Medicine, Universiti Teknologi MARA (UiTM), Sungai Buloh Campus, Jalan Hospital, 47000 Sungai Buloh, Petaling Jaya, Selangor, Malaysia Diserahkan: 31 Julai 2023/Diterima: 9 Februari 2024
Abstract
Hyperhomocysteinemia causes endoplasmic reticulum (ER)
stress, which elevates reactive oxygen species (ROS) and induces
endothelial dysfunction, the hallmark of cardiovascular diseases. Nigella sativa seeds contain thymoquinone (TQ), a cardioprotective bioactive component. Nevertheless, research on
investigating the effectiveness of TQ in preventing endothelial dysfunction
caused by homocysteine (Hcy) is scarce. Therefore,
the purpose of this work was to examine the role of TQ in restoring Hcy-induced endothelial dysfunction as well as the
mechanisms behind this role. Male Sprague-Dawley (SD) rat aortas were isolated
and then co-treated in an organ bath with Hcy and TQ, tauroursodeoxycholic acid (TUDCA), apocynin, or Tempol to examine
vascular function. Furthermore, human umbilical vein endothelial cells (HUVECs)
were treated with Hcy and TQ, Tempol, apocynin, TUDCA or H2O2 to
determine the cell viability via a phase contrast microscope and dye exclusion
test. ER stress pathway involvement, ROS and NO bioavailability were
investigated using immunoassays and fluorescence staining, respectively. The
binding affinity of TQ to GRP78 has been identified using molecular docking. According to
our findings, Hcy hindered endothelium-dependent
relaxation in an isolated aorta and caused apoptosis in HUVECs. TQ, TUDCA, Tempol, and apocynin were able to
counteract these negative effects. In HUVECs, treatment with TQ decreased ROS
levels, increased NO bioavailability, and decreased GRP78 and NOX4 protein.
According to the molecular docking study outcomes, TQ could attach to GRP78
effectively via a hydrogen bond and a hydrophobic connection to the amino acid
at GRP78 ATP binding pocket. Taken together, the findings show that TQ protected
endothelial function caused by Hcy via inhibiting ER
stress-mediated ROS and eNOS uncoupling.
Keywords: Endoplasmic reticulum stress; endothelial dysfunction; homocysteine;
oxidative stress; thymoquinone
Abstrak
Hiperhomosisteinemia meninggikan tekanan retikulum endoplasma (RE) yang boleh meningkatkan spesies oksigen reaktif (SOR), yang membawa kepada disfungsi endotelium sel dan penyakit kardiovaskular. BijiNigella sativa mengandungi timoquinon (TQ), komponen bioaktif berkardio-protektif. Walau bagaimanapun, tiada kajian yang menilai kesan TQ terhadap disfungsi endotelial yang disebabkan oleh homosistein (Hcy). Oleh itu, penyelidikan ini bertujuan untuk mengkaji kesan dan mekanisme TQ dalam menormalkan disfungsi endotelial yang disebabkan oleh Hcy. Aorta diasingkan daripada tikus Sprague-Dawley
(SD) jantan yang diinkubasi dengan Hcy dan dirawat bersama dengan atau tanpa TQ, TUDCA, apocynin atau Tempol dalam mandian organ untuk mengkaji fungsi vaskular. Di samping itu, sel endotelial vena umbilik manusia (HUVECs) diinkubasi dengan Hcy dan TQ, Tempol, apocynin, TUDCA atau H2O2 untuk menilai keviabelan sel dengan menggunakan mikroskop kontras fasa dan ujian pengecualian pewarna. Penglibatan laluan tekanan ER, ROS dan NO bioketersediaan diakses masing-masing melalui immunoasai dan pewarnaan pendarfluor. Dok molekul telah dilakukan untuk menilai pertalian mengikat TQ kepada
GRP78. Keputusan kami mendedahkan bahawa Hcy merosakkan disfungsi endotelial
dalam aorta dan apoptosis dalam HUVECs. Kesan
ini telah dinormalkan oleh TQ, TUDCA, Tempol dan apocynin. Rawatan dengan TQ
mengurangkan tahap ROS, meningkatkan bioketersediaan NO serta mengurangkan
protein GRP78 dan NOX4 dalam HUVECs. Hasil kajian dok molekul menunjukkan
bahawa TQ boleh mengikat dengan baik kepada GRP78 melalui ikatan hidrogen dan
interaksi hidrofobik dengan asid amino pada poket pengikat ATP GRP78.
Kesimpulannya, TQ membaikipulih fungsi endotelial yang dirosakkan oleh Hcy
melalui perencatan ROS pengantara tekanan ER dan meningkatkan bioketersediaan
NO.
Kata
kunci: Disfungsi endotelium; homosistein; tekanan oksidatif; tekanan retikulum endoplasma; timoquinon
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*Pengarang untuk
surat-menyurat; email: choykerwoon@uitm.edu.my
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