The effects of the interaction of vascular endothelial cells and vascular smooth muscle cells on PCSK9 and NF-kB protein expression in vascular smooth muscle cells

Year 2023, Volume: 4 Issue: 3, 133 - 141, 08.08.2023

Ece Yurtseven Kemal Baysal Hülya Yılmaz Aydoğan Oğuz Öztürk

Abstract

Introduction:

Atherosclerosis is the leading cause of death worldwide. Atherosclerosis is an inflammatory disease which causes significant structural and functional changes in vascular wall cells. It is known that the main cells of the vascular wall, endothelial cells, and smooth muscle cells communicate during atherosclerosis, but the mechanisms of this communication are not fully understood. PCSK9, on the other hand, is a molecule involved in the etiology of hyperlipidemia, and its levels increase in vascular wall cells during atherosclerosis, exerting direct atherosclerotic effects. In our study, we investigated the changes induced by endothelial cells exposed to atherogenic factors in smooth muscle cells and their effects on NF-kB expression, the most important molecule in inflammation, and on PCSK9 protein expression, which has recently been discovered to have direct atherosclerotic effects.

Methods:

Cells isolated from human umbilical veins (HUVEC) were incubated for 24 hours with 300 µM hydrogen peroxide (H2O2) or 40 µg/ml oxidized LDL (Ox-LDL) or 5% cigarette smoke extract (CSE). Following 24 hours, human aortic smooth muscle cells (HASMCs) were incubated with the supernatants of HUVECs which were treated with these substances after appropriate processing. After 48 hours, protein extraction was performed from smooth muscle cells and analyzed using the Western Blot method.

Results:

We observed no changes in the viability of HUVECs exposed to atherogenic substances. There was significant morphological changes in HASMCs incubated with endothelial cell supernatants exposed to 5% CSE and 300 µM H2O2. Increased expression of NF-kB (p=0.024, p=0.043) and PCSK9 protein (p=0.034, p=0.0137) was found in HASMCs incubated with supernatants exposed to 5% CSE and 300 µM H2O2, respectively. In HASMCs incubated with endothelial cell supernatant exposed to Ox-LDL, an increase in PCSK9 expression was observed (p=0.037), while no significant increase in NF-kB expression was observed (p=0.89).

Conclusion:

In our study, it was observed that endothelial cells exposed to atherogenic factors induced changes in vascular smooth muscle cells and increased PCSK9 and NF-kB expression in these cells.

Keywords

Atherosclerosis, Vascular smooth muscle cell, Vascular endothelial cell, Proprotein convertase subtilisin/kexin type 9 (PCSK9), Nuclear factor kappa B (NF-kB)

Supporting Institution

İstanbul Ünivertsitesi Bilimsel Araştırma Projeleri Birimi

Project Number

36268

Thanks

I would like to thank the Istanbul University Institute of Health Sciences and Istanbul University for providing me with the opportunity to conduct this study. This study was supported by the Scientific Research Projects Coordination Unit of Istanbul University. Project no: 36268

Vasküler endotel hücreleri ile vasküler düz kas hücrelerinin etkileşiminin, vasküler düz kas hücrelerinde PCSK9 ve NF-kB protein ekspresyonu üzerindeki etkileri.

Abstract

Giriş:

Ateroskleroz dünyada görülen en sık ölüm sebebidir. Ateroskleroz inflamatuar bir hastalık olup, damar duvarı hücrelerinde belirgin şekil ve fonksiyon değişikliklerine neden olur. Damar duvarındaki ana hücreler olan endotel hücreleri ile düz kas hücrelerinin ateroskleroz sırasında iletişim halinde oldukları bilinen bir gerçek olmakla beraber, bu iletişimin mekanizmaları net değildir. PCSK9 ise hiperlipidemi etyolojisinde yer alan bir molekül olmakla birlikte, damar duvar hücrelerinde ateroksleroz sırasında artar ve damar duvar hücrelerine direk aterosklerotik etkileri vardır. Çalışmamızda aterojenik etkenlere maruz kalan endotel hücrelerinin, düz kas hücrelerinde yaptığı değişiklikleri ve inflamasyonun en önemli molekülü olan NF-kB ve direct aterosklerotik etkileri son zamanlarda keşfedilmiş olan PCSK9 protein ekspresyonlarına olan etkilerini araştırdık.

Metod:
İnsan umblikal veninden izole edilen hücreler (HUVEC) ayrı ayrı 24 saat boyunca 300 µM hidrojen peroksit (H2O2), 40 µg/ml okside LDL (Ox-LDL) %5 sigara duman ekstraktı (SDE) ile inkübe edildi, 24 saat sonra aterojenik etkenlere maruz kalan endotel hücrelerinin süpernatantları uygun işlemlerden sonra insan aortic düz kas hücrelerinin (HAVSMC) inkübe edilmesi için kullanıldı. Düz kas hücrelerinden 48 saat sonra protein ekstraksiyonu yapılarak Western Blot yöntemiyle analiz edildi.

Bulgular:
Aterojenik maddelere maruz kalan HUVEC’lerde canlılığın korunduğu saptandı. Bu maddelere maruz kalan endotel hücre süpernatantları ile inkübe edilen HASMC’de belirgin bir morfolojik değişiklik saptandı. %5 SDE ve 300 µM H2O2’ye maruz kalan endotel hücre süpernatantları ile inkübe edilen HASMC’lerde NF-kB (sırasıyla p=0.024,p=0.043) ve PCSK9 protein (p=0.034, p=0.0137) ekspresyonları artmış bulundu. Ox-LDL’ye maruz kalan endotel hücre süpernatantı ile inkübe edilen HASMC’lerde ise PCSK9 ekspresyonu artmış olup (p=0.037), NF-kB ekspresyonunda anlamlı bir artış izlenmemiştir. (p=0.89)

Sonuç:

Çalışmamızın sonucunda aterojenik etkenlere maruz kalan endotel hücrelerinin, damar düz kas üzerinde değişikliklere neden olduğu,damar düz kas hücrelerindeki PCSK9 ve NF-kB ekspresyonunu arttırdığı gözlenmiştir.

Keywords

Ateroskleroz, Vasküler düz kas hücresi, Vasküler endotel hücresi, Proprotein konvertaz subtilisin/keksin tip 9 (PCSK9), Nüleer Faktör Kappa B (NF-kB)

Project Number

36268

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