Part 2:Acteoside Counteracts Interleukin-1β-Induced Catabolic Processes Through The Modulation Of Mitogen-Activated Protein Kinases And The NFκB Cellular Signaling Pathway
Mar 05, 2022
Contact: Audrey Hu Whatsapp/hp: 0086 13880143964 Email: audrey.hu@wecistanche.com
HyangI Lim ,1 Do Kyung Kim ,1 Tae-Hyeon Kim ,1 Kyeong-Rok Kang ,
Osteoarthritis (OA) is the most common degenerative joint disease with chronic joint pain caused by progressive degeneration of articular cartilage at synovial joints. Acteoside, a caffeoylphenylethanoid glycoside, has various biological activities such as antimicrobial, anti-inflammatory, anticancer, antioxidative, cytoprotective, and neuroprotective effect. Further, oral administration of acteoside at a high dosage does not cause genotoxicity. Therefore, the aim of the present study is to verify the anticatabolic effects of acteoside against osteoarthritis and its anticatabolic signaling pathway. Acteoside did not decrease the viabilities of mouse fibroblast L929 cells used as normal cells and primary rat chondrocytes. Acteoside counteracted the IL-1β-induced proteoglycan loss in the chondrocytes and articular cartilage by suppressing the expression and activation of a cartilage-degrading enzyme such as matrix metalloproteinase- (MMP-) 13, MMP-1, and MMP-3. Furthermore, acteoside suppressed the expression of inflammatory mediators such as inducible nitric oxide synthase, cyclooxygenase-2, nitric oxide, and prostaglandin E2 in the primary rat chondrocytes treated with IL-1β. Subsequently, the expression of proinflammatory cytokines was decreased by acteoside in the primary rat chondrocytes treated with IL-1β. Moreover, acteoside suppressed not only the phosphorylation of mitogen-activated protein kinases in primary rat chondrocytes treated with IL-1β but also the translocation of NFκB from the cytosol to the nucleus through suppression of its phosphorylation. Oral administration of 5 and 10mg/kg acteoside attenuated the progressive degeneration of articular cartilage in the osteoarthritic mouse model generated by destabilization of the medial meniscus. Our findings indicate that acteoside is a promising potential anticatabolic agent or supplement to attenuate or prevent progressive degeneration of articular cartilage.
4. Discussion
The synovial (diarthrosis) joint is a complex anatomical structure consisting of several different types of tissues located at the potential space between bones to permit mobility and stability at the body through counteracting the differ- ent intensities of mechanical loading and control fine movements [16]. As the elderly population is increasing worldwide, OA is emerging as a degenerative disease associated with psychological and socioeconomic problems that are to be solved urgently [17]. Unfortunately, there are still no effective medications for OA; therefore, the prevention of articular cartilage degeneration is the most important to maintain the mechanical joint functions associated with the permission of body mobility and stability.
Generally, the synovial joint is composed of two bones to provide stability and support the muscle by ligament and tendons and is surrounded by a synovial fibrous joint capsule filled with synovial fluid to reduce friction between the articular cartilages located on the articular surfaces of the joined bones [16]. Especially, the extracellular matrix (ECM) of articular cartilage is composed mainly of type II collagen and proteoglycans that are synthesized and regulated by specialized cells called chondrocytes. The homeostasis of articular cartilage is precisely balanced between anabolism (synthesis of ECM) and catabolism (degeneration of ECM) in synovial joints [18]. Generally, catabolic factors such as proinflammatory cytokines and inflammatory medi- ators induce the progressive degeneration of articular carti- lage through the expression of cartilage-degrading enzymes such as matrix metalloproteinase (MMPs) and metallopro- teinase with thrombospondin motifs (ADAMTs) from chon- drocytes [18]. Hence, recent biochemical strategies to prevent or attenuate the progressive degeneration of articular cartilage have targeted the suppression of cartilage-degrading enzymes, proinflammatory cytokines, and inflammatory mediators based on the long-term biological safeties in syno- vial joints [19, 20]. Recent studies demonstrate that natural products, originating from herbal or oriental medicine, possess long-term biological safeties, anti-inflammatory, and antioxidative properties and may promote joint health and managing OA through suppressing the release of proinflammatory cytokines [21].
cistanche extract: acteoside
Acteoside (called verbascoside; C29H36O15) is a glyco- side that is isolated from the flowers or leaves of many herbal plants such as Scrophularia ningpoensis, Cistanche deserticola, Digitalis purpurea, and Osmanthus fragrans [22, 23]. Recently, Henn et al. reported that the high concentration (100μg/mL) of acteoside isolated from the leaves of Plantago australis did not only show less cytotoxicity in V79 Chinese hamster cells used as a normal cell but also did not have mutagenic or genotoxic activities and phototoxic properties [6]. Furthermore, Perucatti et al. have reported that in vivo cytogenetic test that is feeding 5mg/kg acteoside to rabbit (Oryctolagus cuniculus) for 80 days revealed no toxicity with any other mutagenic activity, resulting in no cytotoxicity for the animals [24]. These studies suggest that acteoside is a bio-active material that can be used in both animal and human diets [6, 24]. As shown in Figure 2, similar to previous studies, 100μM (62.459μg/mL) acteoside did not affect the viabilities of mouse fibroblast cell line L929 used as a normal cell and primary rat chondrocytes in the present study. Hence, these data indicate that acteoside may have secured potential biological safety and can be used as a supplement. ECM, a large amount of up to 98% of cartilage volume, is a highly organized network of hyaluronan, proteoglycans, and type II collagen [25]. Especially, proteoglycans are proteins glycosylated with sulfated glycosaminoglycan to form an aggregating network that generates a static charge density to counteract compressive forces during the mechanical function of synovial joints [25]. Hence, the loss of proteoglycan in the articular cartilage of synovial joints leads to disability of mechanical joint function [25]. Degeneration of articular cartilage due to the loss of proteoglycan results in an imbalance between the anabolic and catabolic processes. Hence, recent bio- logical strategies related with the regeneration of articular cartilage and the prevention or attenuation of progressive articular cartilage degeneration are considering the increase of anabolic process through the synthesis of major articular cartilage component such as proteoglycan and type II collagen and the increase of anticatabolic process against catabolic factors such as proinflammatory cytokines, inflammation mediators, and catabolic growth factors. As shown in Figure 3, acteoside did not only recover the proteoglycan content through the counteraction against proinflammatory cytokine IL-1β-induced proteoglycan depletion in the primary rat chondrocytes but also suppressed the proteoglycan loss in the articular cartilage tissues treated with IL-1β for 7 days. Taken together, these data indicate consistently that acteoside may protect or attenuate the progressive degeneration of articular cartilage through counteraction to proinflammatory cytokine-induced catabolic process in the articular cartilage of the synovial joint.
Elevated cartilage-degrading enzymes including MMP-1, MMP-3, MMP-13, ADMITS-4, and ADAMTS-5 in the synovial fluid of patients with OA are the key enzymes responsible for the progressive degeneration of articular cartilage through degradation of collagen and ECM compo- nent [26, 27]. Hence, the inhibition of MMP expression and activation seems to be an attractive therapeutic strategy to prevent and attenuate the progressive degeneration of articular cartilage for maintaining the mechanical function of synovial joints [26]. In the present study, acteoside effectively suppressed the expression and activation of the cartilage-degrading enzyme in the primary rat chondrocytes treated with proinflammatory cytokine IL-1β as shown in Figure 4. These data indicate that acteoside may attenuate the progressive degeneration of articular cartilage by suppressing the expression and activation of articular cartilage in the synovial joint with catabolic conditions.
The inflammatory mediators such as iNOS, NO, COX-2, and PGE2 are integral to OA pathogenesis [28]. Especially, proinflammatory cytokines such as IL-1β and TNFα upregulate the production of NO and PGE2 through the increase of iNOS and COX2, respectively, in the synovial joint with OA [29, 30]. Upregulated NO inhibits the synthesis of ECM components such as type II collagen and proteoglycan. Besides, increased PGE2 inhibits the proliferation of chondrocytes and reduces the synthesis of ECM [28]. Hence, suppression of inflammatory mediators may attenuate the progressive degeneration of articular cartilage through the inhibition of ECM reduction in the synovial joint with OA. In the present study, acteoside effectively suppressed the upregulation of inflammatory mediators as shown in Figure 5. These data indicate consistently that acteoside may attenuate the progressive degeneration of articular cartilage through the suppression of inflammatory mediators in the synovial joint with OA.
Moreover, the overexpression of proinflammatory cytokines by the inflamed synovium and chondrocytes is a major risk pathogenic factor in OA pathogenesis. Especially, the expression of proinflammatory cytokine is thought to be generated by the synovial membrane at the stage of OA initiation. Sequentially, upregulated proinflammatory cytokines activate chondrocytes to express their own expression and to synthesize the cartilage-degrading enzymes, chemokines, and inflammatory mediators [31]. Therefore, the suppression of proinflammatory cytokines can prevent OA and may attenuate the progressive degeneration of articular cartilage through the inhibition of other proinflammatory cytokines, inflammatory mediators, and cartilage-degrading enzymes. In the present study, acteoside suppressed the production of proinflammatory cytokines such as CINC-2, CINC-3, CNTF, fractalkine, IL-1α, IL-1β, leptin, MCP-1, MIP-3α, and β-NGF in primary rat chondrocytes treated with IL-1β compared with IL-1β alone, as shown in Figure 6.
Gouze et al. reported that CINC-2 was significantly increased in chondrocytes treated with IL-1β similar with our study [32]. However, a recent study showed that spinal processing of painful inputs is closely altered during OA pathogenesis [33]. With regard to joint pain, CINC-2 and CINC-3 were significantly upregulated in the spinal dorsal horn of OA animals generated by the intra-articular injection of monosodium iodoacetate into the knee joint [34, 35]. Although the pathophysiological role of CINC-2 and CINC-3 in OA pathogenesis is still largely unknown, these studies indicate that the expression of CINC-2 and CINC-3 in the spinal dorsal horn under OA conditions may be closely associated with the development of joint pain during OA pathogenesis.
cistanche herb
CNTF, which is a pluripotent neurotropic factor and is related with the cytokine family that includes IL-6, IL-11, leu- kemia inhibitory family, and oncostatin, binds and signals to maintain the bone homeostasis through the gp130 corecep- tor subunit [36]. Although the biological function of CNTF is still largely unknown in OA, recent studies have shown that CNTF-gp130 signaling may be associated with the pathologic bone remodeling evident in rheumatoid arthritis (RA), periodontal disease, spondyloarthropathies, and OA through regulating the differentiation and activity of osteoblast, oste- oclast, and chondrocytes [36]. In addition, a recent study showed that β-NGF, a neurotrophic factor involved with the physiological regulation of neuronal cells, was upregulated in blood and synovial fluid in patients with OA [37]. However, several studies have reported that the blockade of NGF reduces OA pain [38–40]. Therefore, neurotrophic factors including CNTF and NGF not only are considered pathogenic risk factors of OA progression but also provide the neurological linkage between the progressive degeneration of articular cartilage and the development of chronic OA pain. Furthermore, it has been considered a therapeutic targeting molecule to reduce the chronic OA pain.
Fractalkine also known as chemokine CX3CL1 is exuberantly expressed in both adult human and rat articular chondrocytes treated with IL-1β [41, 42]. Recent studies have reported that fractalkine promotes the expression of MMP- 3 through the CX3CR1, c-Raf, MEK, ERK, and NFκB cellular signaling pathways in the synovial tissue obtained from the patients with OA [43]. Furthermore, the genomic-wide DNA methylation analysis in OA chondrocytes revealed that the fractalkine gene was not only hypomethylated but also constantly correlated with its mRNA expression [44]. MCP-1, a member of chemokine family to induce the inflammation, trigger the chemotaxis and transendothelial migration of monocyte to inflammatory lesion. Recently, Xu et al., have reported that MCP-1 and chemokine (C-C motif) receptor 2 axis are involved with the degradation of articular cartilage through the expression of MMP-13 and the increase of OA
chondrocyte apoptosis [45]. Furthermore, MIP-3α also called a chemokine CCL20 is abundantly expressed in the articular cartilage of patients with OA and increases the progressive degeneration of articular cartilage through the expression of cartilage-degrading enzymes such as MMP-1 and MMP-3, inflammatory mediator such as PGE2, and proinflammatory cytokine IL-6 [46]. Hence, chemokines such as fractalkine, MCP-1, and MIP-3α have been also considered a pathophysiological risk factor to initiate the progression of OA.
Leptin is a peptide hormone belonging to adipokines, which are cytokines secreted by adipose tissue [47]. Recent studies have reported that the level of leptin is not only elevated significantly in the human body with obesity but also increased in the serum and synovial fluid collected from the patients with OA that is correlated with the severity of OA [48]. Hence, recent studies have suggested that the expressions of leptin and its receptor have been considered positively as a risk factor associated with the development of OA [49–51]. [52]
IL-1 family, including IL-1α and IL-1β, is considered the most key cytokine associated with the pathogenesis of OA that induces the inflammatory catabolic process combined with other catabolic factors such as aging, obesity, and traumatic joint injury [53]. Generally, the level of IL-1 family in the synovial fluid, synovial membrane, articular cartilage, and subchondral bone is elevated in the synovial joint of patients with OA [54]. After the IL-1 family binds onto their receptors, it manifests the progressive degeneration of articular cartilage by the expression of other cytokines, chemo- kines, adhesion molecules, inflammatory mediators, and cartilage-degrading enzymes through the phosphorylation of cellular signaling transcriptional factors such as the NFκB and MAPKs [54]. As shown in Figure 7, acteoside not only reduced the phosphorylation of ERK1/2, p38, and JNK but also inhibited the phosphorylation of NFκB in the primary rat chondrocytes treated with IL-1β. Moreover, Figure 8 shows that acteoside inhibited the translocation of NFκB from the cytosol to nucleus in the primary rat chondrocytes treated with IL-1β. Therefore, our results consistently indi- cate that acteoside counteract the IL-1β-induced catabolic effects such as the expression of cartilage-degrading enzymes and the production of proinflammatory cytokines and inflammatory mediators through the inactivation of cellular signaling pathways such as MAPK and NFκB in the primary rat chondrocytes. Recently, similar to our study, Qiao et al. have reported that acteoside inhibits inflammatory response in OA-induced animals [55]. They showed the suppression of inflammatory cytokines through the inactivation of the JAK/STAT signaling pathway in the synovial tissue of DMM-induced OA animals that were administered an intraperitoneal injection of acteoside [55]. However, to estimate the effectiveness of acteoside as an OA preventive supplement, acteoside was orally administrated to DMM-induced OA animals in the present study. Thereafter, the alteration of articular cartilage was histologically assessed as shown in Figure 9. Our histological assessment showed that the oral administration of acteoside consistently prevented the progressive degeneration of articular cartilage through the inhibition of proteoglycan loss in DMM-induced OA animals.
cistanche acteoside
5. Conclusions
Our findings suggest that acteoside is capable of oral administration and may be used as an effective supplement to prevent or attenuate OA based on the biological safety and anticatabolic effects against proinflammatory cytokines.
References
[1] J. S. Di Chen, W. Zhao, T. Wang, L. Han, J. L. Hamilton, and H.-J. I'm, “Osteoarthritis: toward a comprehensive understanding of the pathological mechanism,” Bone Research, vol. 5, no. 1,2017.
[2] G. Musumeci, F. C. Aiello, M. A. Szychlinska, M. Di Rosa, P. Castrogiovanni, and A. Mobasheri, “Osteoarthritis in the XXIst century: risk factors and behaviors that influence disease onset and progression,” International Journal of Molec- ular Sciences, vol. 16, no. 12, pp. 6093–6112, 2015.
[3] A. Ghouri and P. G. Conaghan, “Prospects for therapies in osteoarthritis,” Calcified Tissue International, 2020.
[4] B. Klimek, “6′-0-Apiosyl-verbascoside in the flowers of mullein (Verbascum species),” Acta Poloniae Pharmaceutica, vol. 53, no. 2, pp. 137– 140, 1996.
[5] F. Pardo, F. Perich, L. Villarroel, and R. Torres, “Isolation of verbascoside, an antimicrobial constituent of Buddleja Globo leaves,” Journal of Ethnopharmacology, vol. 39, no. 3, pp. 221- 222, 1993.
[6] J. G. Henn, L. Steffens, N. D. de Moura Sperotto, et al., “Toxicological evaluation of a standardized hydroethanolic extract from leaves of Plantago australis and its major compound, verbascoside,” Journal of Ethnopharmacology, vol. 229, pp. 145– 156, 2019.
[7] M. Khullar, A. Sharma, A. Wani et al., “Acteoside ameliorates inflammatory responses through NFkB pathway in alcohol-induced hepatic damage,” International Immunopharmacology, vol. 69, pp. 109– 117, 2019.
[8] T. W. Hwang, D. H. Kim, D. B. Kim et al., “Synergistic anticancer effect of acteoside and temozolomide-based glioblastoma chemotherapy,” International Journal of Molecular Medicine, vol. 43, no. 3, pp. 1478– 1486, 2019.
[9] X. Li, Y. Xie, K. Li et al., “Antioxidation and cytoprotection of acteoside and its derivatives: comparison and mechanistic chemistry,” Molecules, vol. 23, no. 2, p. 498, 2018.
[10] M. Li, F. Zhou, T. Xu, H. Song, and B. Lu, “Acteoside protects against 6-OHDA-induced dopaminergic neuron damage via Nrf2-ARE signaling pathway,” Food and Chemical Toxicology, vol. 119, pp. 6– 13, 2018.
