Sains Malaysiana 51(4)(2022): 1181-1196

http://doi.org/10.17576/jsm-2022-5104-19

Dehydroabietic Acid Attenuates Atherosclerosis in Apolipoprotein E-Deficient Mice and VCAM-1 Expressions in-vitro

(Asid Dehidroabietik Melemahkan Aterosklerosis dalam Tikus Apolipoprotein E-Kurang dan Pengekspresan VCAM-1 in-vitro)

XIAO LIANG & YAN WU^*

Department of General Medicine, The First People's Hospital of Lianyungang, Lianyungang, Jiangsu, China, 222000

Received: 21 July 2021/ Accepted: 30 August 2021

Abstract

Dehydroabietic acid (DHA) is an analog of abietic acid (AA) for cardiovascular disease prevention, known to act pharmacologically against aging, inflammation, bacterial infections and cancer. The current research investigated the molecular mechanisms of DHA on the adhesiveness of endothelial leukocytes and activation of NF-κB in TNF-α treated HAEC and ApoE−/− mice in experimental atherosclerosis. The HAECs were tested for toxicity using MTT assay at various DHA concentrations. The cell-surface expression of CAM against endothelial leukocyte adhesion molecule-1, ICAM -1, or human VCAM-1 was determined by the ELISA test, followed by western blot analysis. Endothelial cell-leukocyte adhesion assay involving U937 cells was carried out followed by the determination of NF-κB p65 expression. ApoE−/− mice were fed with a high cholesterol diet every day followed by oral administration of DHA (10 and 20 mg/kg/day). The DHA (5 and 10 mM) substantially reduced (p < 0.05) human (U937) cell lines binding to TNF-α activated human endothelial aortic cells. The assays involving the 32P-labelled NF-kB as a probe demonstrated that DHA pre-treatment decreased the shifted bands density following the stimulation of TNF-α. Nuclear extracts immune blot assessment and immune fluorescence staining showed a significant decrease (p < 0.05) in the NF-κB p65 concentration in the nuclei with DHA treated human endothelial aortic cells. Together these findings indicate that DHA inhibits nuclear trans localization of TNF-α-induced NF-κB p65, and thus considerably suppresses (p < 0.05) the VCAM-1 expression, which contributes to lower leukocyte adherence suggesting that DHA helps in preventing inflammatory atherosclerosis in vivo.

Keywords: ApoE−/−; atherosclerosis; dehydroabietic acid; TNF-α

Abstrak

Asid dehidroabietik (DHA) ialah analog asid abietik (AA) untuk pencegahan penyakit kardiovaskular yang diketahui bertindak secara farmakologi terhadap penuaan, keradangan, jangkitan bakteria dan kanser. Penyelidikan semasa mengkaji mekanisme molekul DHA pada kelekatan leukosit endotelium dan pengaktifan NF-κB dalam TNF-α yang dirawat HAEC dan ApoE-/- tikus dalam aterosklerosis uji kaji. HAEC telah diuji untuk ketoksikan menggunakan ujian MTT pada pelbagai kepekatan DHA. Pengekspresan permukaan sel CAM terhadap molekul lekatan leukosit endotelium-1, ICAM -1, atau VCAM-1 manusia ditentukan melalui ujian ELISA, diikuti oleh analisis pemblotan western. Ujian lekatan sel-leukosit endotelium yang melibatkan sel U937 telah dijalankan diikuti dengan penentuan pengekspresan NF-κB p65. ApoE−/− tikus diberi makan dengan diet kolesterol tinggi setiap hari diikuti dengan pemberian oral DHA (10 dan 20 mg/kg/hari). DHA (5 dan 10 mM) mengurangkan dengan ketara (p <0.05) garisan sel manusia (U937) yang mengikat kepada sel aorta endotelium manusia yang diaktifkan TNF-α. Ujian yang melibatkan NF-kB berlabel 32P sebagai prob menunjukkan bahawa pra-rawatan DHA mengurangkan ketumpatan jalur beralih berikutan rangsangan TNF-α. Ekstrak nuklear penilaian pemblotan imun dan pewarnaan pendarfluor imun menunjukkan penurunan ketara (p <0.05) dalam kepekatan NF-κB p65 dalam nukleus dengan sel aorta endotelium manusia yang dirawat DHA. Bersama-sama penemuan ini menunjukkan bahawa DHA menghalang penyetempatan trans nuklear NF-κB p65 yang disebabkan oleh TNF-α dan dengan itu menekan dengan ketara (p <0.05) pengekspresan VCAM-1, yang menyumbang kepada pematuhan leukosit yang lebih rendah yang menunjukkan bahawa DHA membantu dalam mencegah aterosklerosis radang secara in vivo.

Kata kunci: ApoE−/−; asid dehidroabietik; aterosklerosis; TNF-α

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^*Corresponding author; email: tezhenwuyan@163.com