Sains Malaysiana 48(8)(2019): 1737–1744

http://dx.doi.org/10.17576/jsm-2019-4808-20

Inhibitory Effects of Gynura procumbens Ethanolic Extract on Nitric Oxide Production and Inducible Nitric Oxide Synthase (iNOS) Protein Expression in Macrophages

(Kesan Perencatan Ekstrak Etanol Gynura procumbens terhadap Penghasilan Nitrik Oksida dan Ekspresi Protein Sintase Nitrik Oksida Teraruh (iNOS) di dalam Makrofaj)

TAN JIAH NING¹, SYARATUL DALINA YUSOFF¹, ZAKIAH JUBRI², FHATAHEYA BUANG¹, TAN ZE SONG¹, AMEERAH BUDIONO¹, IBRAHIM JANTAN³, ROZA DIANITA⁴, ENDANG KUMOLOSASI¹, NORAZRINA AZMI¹ & NORSYAHIDA MOHD FAUZI¹*

¹Drug and Herbal Research Centre, Faculty of Pharmacy, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur, Malaysia

²Department of Biochemistry, Faculty of Medicine, Universiti Kebangsaan Malaysia, Jalan Yaacob Latiff, Bandar Tun Razak, 56000 Cheras, Kuala Lumpur, Federal Territory, Malaysia

³School of Pharmacy, Taylor's University, Lakeside Campus, 47500 Subang Jaya, Selangor Darul Ehsan, Malaysia

⁴School of Pharmaceutical Sciences, Universiti Sains Malaysia, 11800 USM Pulau Pinang, Malaysia

Received: 14 September 2018/Accepted: 31 May 2019

ABSTRACT

Nitric oxide (NO) overproduction by inducible nitric oxide synthase (iNOS) may be associated with acute and chronic inflammations. Macrophages as important cells in the innate immune system are able to be stimulated and can lead to iNOS activation and excessive NO production. Gynura procumbens is a medicinal plant traditionally used in treating various ailments including inflammation but the mechanism of anti-inflammatory activity of this plant is still elusive. This study was carried out to investigate the anti-inflammatory therapeutic effects of Gynura procumbens ethanolic extract on NO production and iNOS protein expression in RAW 264.7 macrophages stimulated with lipopolysaccharide (LPS). Cell viability of RAW 264.7 macrophages treated with Gynura procumbens ethanolic extract was determined by MTT assay. NO production was determined by Griess assay following Gynura procumbens ethanolic extract treatment alone or in combination with LPS stimulation. Protein expression of iNOS was determined by western blot. RAW 264.7 macrophages viability of more than 90% was observed after 24 h treatment with Gynura procumbens ethanolic extract concentration range of 3.9 μg/mL to 500 μg/mL. Significant inhibition of NO production level has been identified in LPS-stimulated RAW 264.7 cells pre-treated with 250 μg/mL Gynura procumbens ethanolic extract (p<0.05) while all selected concentrations of Gynura procumbens ethanolic extract showed no significant alteration of NO production in the absence of LPS stimulation. Pre-treatment of 250 μg/mL Gynura procumbens ethanolic extract also demonstrated significant suppression of iNOS protein expression in LPS-stimulated RAW 264.7 cells (p<0.05). In conclusion, this study demonstrates that Gynura procumbens ethanolic extract exhibits anti-inflammatory potential through inhibition of NO production and iNOS protein expression in LPS-stimulated macrophages, suggesting that this plant could be further researched for its beneficial use in inflammatory disorders.

Keywords: Gynura procumbens; inducible nitric oxide synthase; inflammation; lipopolysaccharide; macrophage; nitric oxide

ABSTRAK

Penghasilan nitrik oksida (NO) yang berlebihan oleh sintase nitrik oksida teraruh (iNOS) mungkin boleh dikaitkan dengan radang akut dan kronik. Makrofaj sebagai sel yang penting dalam sistem keimunan inat berupaya dirangsang dan menyebabkan aktivasi iNOS dan penghasilan NO yang berlebihan. Gynura procumbens ialah tumbuhan perubatan tempatan yang digunakan secara tradisi untuk merawat pelbagai jenis penyakit termasuk radang namun mekanisme aktiviti anti-radang oleh tumbuhan ini masih sukar difahami. Kajian ini dijalankan untuk mengkaji kesan terapeutik anti-radang oleh ekstrak etanol Gynura procumbens terhadap penghasilan NO dan ekspresi protein iNOS dalam makrofaj RAW 264.7 yang dirangsang oleh lipopolisakarida (LPS). Kebolehhidupan sel makrofaj RAW 264.7 oleh rawatan ekstrak etanol Gynura procumbens ditentukan dengan asai MTT. Reagen Griess digunakan untuk menentukan penghasilan NO diikuti rawatan ekstrak etanol Gynura procumbens atau dengan gabungan rangsangan LPS. Ekspresi protein iNOS dikaji dengan pemblotan western. Di bawah rawatan ekstrak etanol Gynura procumbens selama 24 jam menggunakan kepekatan 3.9 μg/mL hingga 500 μg/mL, makrofaj RAW 264.7 menunjukkan kebolehhidupan sel lebih daripada 90%. Perencatan paras penghasilan NO yang signifikan telah dikenal pasti dalam sel RAW 264.7 yang dirangsang oleh LPS bersama pra-rawatan 250 μg/mL ekstrak etanol Gynura procumbens (p<0.05) manakala semua kepekatan ekstrak etanol Gynura procumbens yang dipilih menunjukkan tiada perubahan yang signifikan dalam penghasilan NO tanpa perangsangan LPS. Pra-rawatan 250 μg/mL ekstrak etanol Gynura procumbens juga mendemonstrasikan perencatan ekspresi protein iNOS yang signifikan dalam sel RAW 264.7 yang dirangsang oleh LPS (p<0.05). Kesimpulannya, kajian ini menunjukkan bahawa ekstrak etanol Gynura procumbens mempamerkan potensi anti-radang melalui perencatan penghasilan NO dan ekspresi protein iNOS dalam makrofaj yang dirangsang oleh LPS, menunjukkan bahawa tumbuhan ini boleh dikaji secara lebih lanjut tentang kegunaannya yang bermanfaat dalam gangguan keradangan.

Kata kunci: Gynura procumbens; lipopolisakarida; makrofaj; nitrik oksida; radang; sintase nitrik oksida teraruh

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*Corresponding author; email: drnorsyahida@ukm.edu.my