Byakangelicin inhibits IL-1β–induced mouse chondrocyte inflammation in vitro and ameliorates murine osteoarthritis in vivo

doi:10.1016/j.intimp.2020.106605

International Immunopharmacology

Volume 85, August 2020, 106605

https://doi.org/10.1016/j.intimp.2020.106605 Get rights and content

Highlights

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BYA inhibited the IL-1β-induced expression of iNOS, COX-2, TNF-α, and IL-6 in vitro.
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BYA altered the expression of various proteins, leading to ECM degradation.
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BYA suppressed the NF-κB signaling in the IL-1β-induced chondrocytes.
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BYA exerted protective effects in OA development in an in vivo DMM model.
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In conclusion, BYA may be a potential therapeutic approach for OA.

Abstract

Osteoarthritis (OA) is a chronic musculoskeletal degeneration disease, resulting in severe consequences such as chronic pain and functional disability. Owing to the complex pathology, there are currently available preventative clinical therapies for OA. Several studies have reported the potential anti-inflammatory effects of byakangelicin (BYA), a component of the Angelica dahurica root extract; however, the effects of BYA in OA remain unknown. In this study, we investigated the protective effects of BYA in interleukin (IL)-1β–induced mouse chondrocytes in vitro and on surgical destabilization in a medial meniscus (DMM) mouse OA model in vivo. In vitro, BYA treatment inhibited IL-1β–mediated inducible nitric oxide synthase, cyclooxygenase-2, tumor necrosis factor-alpha, and IL-6 expression. Moreover, BYA promoted the expression of type two collagen and aggrecan but inhibited the expression of thrombospondin motifs 5 and matrix metalloproteinases, leading to degradation of the extracellular matrix. In addition, BYA mechanistically suppressed nuclear factor-kappa B signaling in the IL-1β–induced chondrocytes. The protective effects of BYA in OA development were also observed in vivo using the DMM model. In conclusion, our results highlight BYA as a candidate for OA treatment and prevention.

Introduction

Osteoarthritis (OA) is a painful degenerative joint disease characterized by complicated mechanisms involving synovial inflammation, articular cartilage degradation, subchondral bone remodeling, and osteophyte formation [1], [2]. These pathological events cause irreversible disabilities that affect the quality of life of millions of people worldwide, particularly the elderly [3], [4]. It is widely accepted that chondrocytes, the only cell type in the articular cartilage, are closely related to OA progression; therefore, investigating the molecular mechanisms of chondrocytes can help to improve our understanding of cartilage degradation and aid in the development of effective therapeutic approaches for OA.

Until recently, chondrocytes in the articular cartilage were considered to mainly function in maintaining the balance between anabolic metabolism and catabolic metabolism in the extracellular matrix (ECM). However, accumulating evidence has demonstrated that pro-inflammatory cytokines such as interleukin (IL)-1β and tumor necrosis factor alpha (TNF-α), regulate the release of proteolytic enzymes, including matrix metalloproteinases (MMPs) and A disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS), which can accelerate the anabolic activity of chondrocytes via ECM degradation [5], [6]. In particular, IL-1β exerts inflammatory effects by significantly increasing the secretion of inflammatory cytokines such as IL-6, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2), and by inhibiting the synthesis of anabolic macromolecules such as type II collagen and aggrecan [7], [8], [9], leading to the initiation and progression of OA along with chondrocyte dysfunction. Therefore, inhibiting the IL-1β–induced inflammatory reaction can be a promising target for OA therapy [10], [11].

Byakangelicin (7H-furo[3,2-g][1]benzopyran-7-one,9-(2,3-dihydroxy-3-methylbutoxy)-4-methoxy-; BYA), identified in Angelica dahurica root extracts, has been shown to have a wide range of biological activities [12], [13], [14]. In China, BYA is used as a traditional medicine to treat colds, headache, and toothaches [15], [16], and has also been reported to increase the expression of all pregnane X receptor target genes in human hepatocytes, thereby inducing a wide range of inter-drug interactions [13]. Importantly, BYA shows high anti-inflammatory potential, which has been confirmed in some disease models. One study revealed that BYA could greatly reduce lipopolysaccharide-induced inflammation to improve the accumulation of diverse active compounds in vivo [17]. Kang et al. [18] reported that BYA significantly improved cellular reactive oxygen species (ROS) levels in RAW264.7 cells. Furthermore, BYA can inhibit the gene expression of inflammatory cytokines such as TNF-α and IL‐1β by inhibiting NF-κB activation [19], [20]. Despite its potential role as a treatment agent indicated by these preclinical studies, the effects of BYA in OA remain unclear.

Therefore, in this study, we investigated the protective effects of BYA on IL-1β–induced inflammation in chondrocytes in vitro and its therapeutic effect in a surgical destabilization in the medial meniscus (DMM) mouse model of OA in vivo.

Section snippets

Media and reagents

BYA (purity ≥ 95%) and carboxymethylcellulose (CMC) were obtained from Sigma-Aldrich (St. Louis, MO, USA). Primary antibodies against MMP3 (Rabbit, #14351), p65 (Rabbit, #8242), p-p65 (Rabbit, #3033), IκBα (Rabbit, #4812), Lamin B1 (Rabbit, #13435), and GADPH (Rabbit, #5174) were acquired from Cell Signaling Technology (Danvers, MA, USA). Primary antibodies against MMP13 (Rabbit, ab219620), COX-2 (Rabbit, ab15191), iNOS (Rabbit, ab15323), ADAMTS5 (Rabbit, ab41037), aggrecan (Rabbit, ab3778),

Phenotypic determination of mouse primary chondrocytes

First, we examined the morphology of the primary chondrocytes (Fig. 1A), and their phenotype was determined by type II collagen immunofluorescent staining (Fig. 1B).

Effects of BYA on the viability of mouse chondrocytes

The chemical structure of BYA is shown in Fig. 2A. To assess its cytotoxicity, the mouse chondrocytes were exposed to varying doses of BYA (0–10 µM) for 24 h or 48 h. The chondrocyte viability was unaffected by BYA doses up to 2.5 µM (Fig. 2B, C); therefore, we used BYA doses of 0.625 µM, 1.25 µM, and 2.5 μM in the subsequent

Discussion

OA is one of the most common chronic diseases characterized by synovial inflammation, articular cartilage degradation, subchondral bone remodeling, and osteophyte formation [25], [26]; however, current therapeutic strategies such as non-steroidal anti-inflammatory drugs only help to alleviate pain and are associated with serious side effects, including gastrointestinal bleeding [27]. High morbidity and a lack of therapeutic strategies for OA necessitate the urgent development of new therapeutic

Funding sources

This study was funded by the National Natural Science Foundation of China (grant no. 81871801), Natural Science Foundation of Zhejiang Province (grant no. LY15H060005), Zhejiang Basic Public Welfare Research Project (grant no. LGF20H060011), Projects of Medical and Health Technology Development Program in Zhejiang Province (grant no. 2018KY824), and Natural Science Foundation of Zhejiang Province (grant no. LQ19H060001).

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

We would also like to thank Dr. An Qin from the Department of Orthopedics, Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine for technical help and advice.

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¹: These authors contributed equally to this work and should be considered as co-first authors.

View full text

Byakangelicin inhibits IL-1β–induced mouse chondrocyte inflammation in vitro and ameliorates murine osteoarthritis in vivo

Highlights

Abstract

Introduction

Section snippets

Media and reagents

Phenotypic determination of mouse primary chondrocytes

Effects of BYA on the viability of mouse chondrocytes

Discussion

Funding sources

Declaration of Competing Interest

Acknowledgements

Osteoarthr. Cartil.

Bone

Cell. Signal.

Phytomedicine

Phytomedicine

J. Pharmacol. Sci.

Cancer Lett.

Osteoarthr. Cartil.

Phytomedicine: Int. J. Phytother. Phytopharmacol.

Osteoarthr. Cartil.

Osteoarthr. Cartil.

Bone

J. Oral Maxillofac. Surg.

Trends Cell Biol.

J. Biol. Chem.

Cytokine

Osteoarthr. Cartil.

Int. J. Biochem. Cell Biol.

Mutat. Res./Fundam. Mol. Mech. Mutagen.

Exp. Cell Res.

Radiocarbon dating reveals minimal collagen turnover in both healthy and osteoarthritic human cartilage

Sci. Transl. Med.

Systemic inhibition of IL-6/Stat3 signalling protects against experimental osteoarthritis

Ann. Rheum. Dis.

Clinical Perspective on Pain and Pain Phenotypes in Osteoarthritis

Curr. Rheumatol. Rep.

Role of proinflammatory cytokines in the pathophysiology of osteoarthritis

Nat. Rev. Rheumatol.

Shifting HIFs in osteoarthritis

Nat. Med.