Cryptotanshinone protects against IL-1β-induced inflammation in human osteoarthritis chondrocytes and ameliorates the progression of osteoarthritis in mice

Osteoarthritis (OA) is a common degenerative disease characterized by progressive erosion of articular cartilage, subchondral bone sclerosis and synovitis. Cryptotanshinone (CTS), an active component extracted from the root of Salvia miltiorrhiza Bunge, has been shown to have potent anti-inflammatory effects. However, its effects on OA have not been clearly elucidated. This study aimed to assess the effect of CTS on human OA chondrocytes and mice OA models. Human OA chondrocytes were pretreated with CTS (5, 10 and 20 μM) for 2 h and subsequently stimulated with IL-1β for 24 h. Production of NO, PGE2, IL-6, TNF-α was evaluated by the Griess reaction and ELISA. The protein expression of COX-2, iNOs, MMP-3, MMP13, COX-2, ADAMTS-5, JNK, p-JNK, ERK, p-ERK, p38, p-p38, p-IKKα/β, p65, p-p65, IκB-α, and p-IκB-α was tested by Western blot. In vivo, the severity of OA was determined by histological analysis. We found that CTS significantly inhibited the IL-1β-induced production of NO and PGE2; expression of COX-2, iNOS, MMP-3, MMP-13, and ADAMTS-5. Furthermore, CTS in dramatically suppressed IL-1β-stimulated NF-κB and MAPK activation. Immunofluorescence staining demonstrated that CTS could suppress IL-1β-induced phosphorylation of p65 nuclear translocation. In vivo, treatment of CTS prevented the destruction of cartilage and the thickening of subchondral bone in mice OA models. These results indicate that the therapeutic effect of CTS on OA is accomplished through the inhibition of both NF-κB and MAPK signaling pathways. Our findings provide the evidence to develop CTS as a potential therapeutic agent f or patients with OA.     

Fisetin inhibits IL-1β-induced inflammatory response in human osteoarthritis chondrocytes through activating SIRT1 and attenuates the progression of osteoarthritis in mice

Osteoarthritis (OA) is a degenerative joint disease characterized by cartilage degradation and inflammation. Fisetin, a polyphenol extracted from fruits and vegetables, has been reported to have anti-inflammatory effects. Our study aimed to investigate the effect of fisetin on OA both in vitro and in vivo. In vitro, chondrocytes were pretreated with fisetin alone or fisetin combined with sirtinol (an inhibitor of SIRT1) for 2 h before IL-1β stimulation. Production of NO, PGE2, TNF-α and IL-6 were evaluated by the Griess reaction and ELISAs. The mRNA (COX-2, iNOS, MMP-3, MMP-13, ADAMTS-5, Sox-9, aggrecan and collagen-II) and protein expression (COX-2, iNOS, MMP-3, MMP-13, ADAMTS-5 and SIRT1) were measured by qRT-PCR and Western blot respectively. Immunofluorescence was used to assess the expression of collagen-II and SIRT1. SIRT1 activity was quantified with SIRT1 fluorometric assay kit. The in vivo effect of fisetin was evaluated by gavage in mice OA models induced by destabilization of the medial meniscus (DMM). We found that fisetin inhibited IL-1β-induced expression of NO, PGE2, TNF-α, IL-6, COX-2, iNOS, MMP-3, MMP-13, ADAMTS-5. Besides, fisetin remarkably decreased IL-1β-induced degradation of Sox-9, aggrecan and collagen-II. Furthermore, fisetin significantly inhibited IL-1β-induced SIRT1 decrease and inactivation. However, the inhibitory effect of fisetin was obvious abolished by sirtinol, suggesting that fisetin exerts anti-inflammatory effects through activating SIRT1. In vivo, fisetin-treated mice exhibited less cartilage destruction and lower OARSI scores. Moreover, fisetin reduced subchondral bone plate thickness and alleviated synovitis. Taken together, these findings indicate that fisetin may be a potential agent in the treatment of OA.     

UFL1 attenuates IL-1β-induced inflammatory response in human osteoarthritis chondrocytes

Osteoarthritis (OA) is a chronic inflammatory joint disease characterized by degradation of articular cartilage. Ubiquitin-fold modifier 1 (UFM1)-specific ligase 1 (UFL1) is an UFM1 E3 ligase that has been identified as a regulator of inflammatory response. However, the role of UFL1 in OA remains unknown. The aim of the present study was to explore the function of UFL1 in an in vitro OA system in chondrocytes. Our results showed that UFL1 was lowly expressed in both OA articular tissues and chondrocytes with IL-1β induction. Ectopic expression of UFL1 improved cell viability of IL-1β-induced chondrocytes. UFL1 suppressed the production of NO and PGE2, as well the expression levels of iNOS and COX-2 in IL-1β-induced chondrocytes. The IL-1β-induced increases in TNF-α and IL-6 levels were attenuated by UFL1. Ectopic expression of UFL1 inhibited the production of extracellular matrix (ECM) degrading enzymes including matrix metalloproteinase 3 (MMP-3), MMP-13, ADAMTS-4 and ADAMTS-5 in chondrocytes with IL-1β induction. Additionally, UFL1 suppressed IL-1β-induced activation of NF-κB signaling pathway in chondrocytes. In conclusion, these findings indicated that UFL1 exerted protective effect on IL-1β-induced chondrocytes. Thus, UFL1 might be a potential target for the treatment of OA.     

Geraniol-mediated osteoarthritis improvement by down-regulating PI3K/Akt/NF-κB and MAPK signals: In vivo and in vitro studies

Osteoarthritis (OA) is a degenerative disease that has received increasing attention among the elderly. Its clinical manifestation is primarily long-term joint pain. Evidence for the pharmacological effects of geraniol in various diseases is accumulating. However, whether geraniol has a therapeutic effect against OA remains to be determined. In this study, we discussed the anti-inflammatory effects of geraniol in IL-1β-induced chondrocytes and the anti-cartilage degradation effects in a mouse model of destabilization of the medial meniscus (DMM). In cell experiments, we found that the treatment of geraniol inhibited the expression of IL-1β-induced PGE2, NO, COX-2, iNOS, TNF-α and IL-6 by western blot, qRT-PCR and immunofluorescence staining. Besides, geraniol inhibited the expression of MMP-9 and ADAMTS-5, and reversed the degradation of aggrecan and type II collagen. Mechanistically, we revealed that geraniol suppressed IL-1β-stimulated PI3K/Akt/NF-κB and MAPK activation. Importantly, we have found in animal experiments that oral treatment of geraniol was beneficial in protecting articular cartilage from degradation. Overall, these data indicated that geraniol may have the potential to be developed as an effective treatment for OA.     

The protective effect of myricitrin in osteoarthritis: An in vitro and in vivo study

Osteoarthritis (OA) is a long-term, chronic, progressive joint condition caused by a pathology characterized by the deterioration of joint cartilage and proliferation of subchondral bone. Myricitrin (Myr) is a flavonoid compound extracted from myrica rubra with potent anti-inflammatory properties, as demonstrated in various studies. However, the mechanisms by which Myr plays a protective role in OA are not completely understood. In this study, the anti-inflammatory properties and potential mechanisms of Myr on mouse chondrocytes treated with interleukin (IL) −1beta (β) were explored in vitro and the role of Myr in a mouse model of OA in vivo. The production of pro-inflammatory factors, such as IL-6, tumor necrosis factor alpha (TNF-α), prostaglandin E2 (PGE2) and nitric oxide (NO) were assessed by enzyme linked immunosorbent assay (ELISA) and the Griess reaction. Protein expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), Collagen-II, matrix metalloproteinase(MMP)-13, MMP-3, thrombospondin motifs 5(ADAMTS5), inhibitor of nuclear factor kappa-B (IκB), p-IκB, p65, p-p65, c-jun-terminal kinase (JNK), p-JNK, extracellular regulated protein kinases (ERK), p-ERK, p38 and p-p38 were quantified using Western blot analysis. In the present study, we found that Myr inhibited IL-1β-induced production of NO and PGE2, expression of MMP-13, MMP-3 and ADAMTS5 and degradation of collagen-II in mouse chondrocytes. Mechanistically, Myr inhibited the activation of mitogen-activated protein kinase (MAPK) and nuclear factor kappa-B (NF-κB) treated with IL-1β in mouse chondrocytes. In vivo, Myr decreased OA Research Society International (OARSI) scores in a surgically-induced mouse model of OA. These data suggest that Myr could be developed as a potential therapy for OA.     

Protectin DX attenuates IL-1β-induced inflammation via the AMPK/NF-κB pathway in chondrocytes and ameliorates osteoarthritis progression in a rat model

Protectin DX (PDX) has been reported to have extensive anti-inflammatory effects. However, it is unknown whether PDX acts as an anti-inflammatory agent in the context of osteoarthritis (OA). This study aimed to evaluate the anti-inflammatory activity of PDX in vitro and in vivo in a model of OA. Primary rat chondrocytes were preincubated with PDX 1 h prior to IL-1β treatment for 24 h. We found that PDX was nontoxic, and pretreatment with PDX increased cell viability in IL-1β-induced chondrocytes. Preincubation with PDX also efficiently inhibited the degradation of type II collagen dose-dependently. Additionally, the expression of MMP-3, MMP-13, ADAMTS4, iNOS, COX-2, NO, and PGE2 decreased after IL-1β stimulation when cells were preincubated with PDX. Moreover, PDX inhibited the increase in phosphorylated NF-κB p65 and IκBα upon IL-1β stimulation, and the negative effects of IL-1β on chondrocytes were partially blocked by treatment with pyrrolidine dithiocarbamate (PDTC), a selective NF-κB inhibitor. In addition, we found that PDX increased AMPK phosphorylation in IL-1β-mediated chondrocytes. The phosphorylation of AMPK could be inhibited by compound C, a classic AMPK inhibitor. Compound C also remarkably reversed the decrease in p65 phosphorylation and MMP-13 expression caused by PDX. Furthermore, nuclear translocation of NF-κB was visible by immunofluorescence after PDX-induced AMPK activation. Additionally, we verified that PDX ameliorated cartilage degradation in monosodium iodoacetate (MIA)-induced OA rats through histological evaluation and ELISA of TNF-α in the serum and intra-articular lavage fluid. In conclusion, we have shown that PDX suppresses inflammation in chondrocytes in vitro and in vivo, likely through the AMPK/NF-κB signaling pathway. Our results suggest that PDX could be a useful novel therapeutic agent for OA treatment.     

Baicalin suppresses IL-1β-induced expression of inflammatory cytokines via blocking NF-κB in human osteoarthritis chondrocytes and shows protective effect in mice osteoarthritis models

Osteoarthritis (OA) is a degenerative joint disease with an inflammatory component that drives the degradation of cartilage extracellular matrix. Baicalin, a predominant flavonoid isolated from the dry root of Scutellaria baicalensis Georgi, has been reported to have anti-inflammatory effects. However, the anti-inflammatory effects of baicalin on OA have not been reported. Our study aimed to investigate the effect of baicalin on OA both in vitro and in vivo. In vitro, human OA chondrocytes were pretreated with baicalin (10, 50, 100 μM) for 2 h and subsequently stimulated with IL-1β for 24 h. Production of NO and PGE2 were evaluated by the Griess reaction and ELISAs. The mRNA expression of COX-2, iNOS, MMP-3, MMP-13, ADAMTS-5, aggrecan and collagen-II were measured by real-time PCR. The protein expression of COX-2, iNOS, MMP-3, MMP-13, ADAMTS-5, p65, p-p65, IκBα and p-IκBα was detected by Western blot. The protein expression of collagen-II was evaluated by immunofluorescence. Luciferase activity assay was used to assess the relative activity of NF-kB. In vivo, the severity of OA was determined by histological analysis. We found that baicalin significantly inhibited the IL-1β-induced production of NO and PGE2, expression of COX-2, iNOS, MMP-3, MMP-13 and ADAMTS-5 and degradation of aggrecan and collagen-II. Furthermore, baicalin dramatically suppressed IL-1β-stimulated NF-κB activation. In vivo, treatment of baicalin not only prevented the destruction of cartilage but also relieved synovitis in mice OA models. Taken together, these results suggest that baicalin may be a potential agent in the treatment of OA.     

Alpha-Mangostin protects rat articular chondrocytes against IL-1β-induced inflammation and slows the progression of osteoarthritis in a rat model

Osteoarthritis (OA) is a joint disease characterized by inflammation and cartilage degradation. α-Mangostin (α-MG), which can be isolated from the fruit of the tropical evergreen tree Garcinia mangostana-L, is known to have anti-inflammatory properties. The aim of the study was to investigate the use of α-MG in the treatment of OA, using both rat chondrocytes and an OA rat model induced by destabilization of the medial meniscus (DMM). Rat chondrocytes were pretreated with α-MG (0, 1.25, 2.5, and 5.0 μg/ml for 24 h) prior to stimulation with interleukin-1β (IL-1β) (10 ng/ml for 24 h). Nitric oxide (NO) production was determined using the Griess method and prostaglandin E₂ (PGE₂) was assessed using an enzyme-linked immunosorbent assay (ELISA). The expression of inducible nitric oxide synthase (INOS), cyclooxygenase-2 (COX-2), matrix metalloproteinase-3, -9, and -13 (MMP-3, MMP-9, and MMP-13), Collagen II, and Aggrecan were detected by both quantitative real-time PCR (qRT-PCR) and a western blot analysis. Nuclear factor-κB (NF-κB) signaling molecules were detected by western blot analysis. Detection of p65 nuclear translocation of NF-κB was examined using immunofluorescence staining. The OA rats received intraperitoneal injections of α-MG (10 mg/kg) or saline every other day. Hematoxylin and eosin and Safranin-O-Fast green staining were used to evaluate the severity of cartilage lesions up to 8 weeks following surgery. α-MG inhibited the production of NO and PGE₂. The elevated expression of INOS, COX-2, MMP-3, MMP-9, and MMP-13, and the degradation of Collagen II and Aggrecan, were reversed by α-MG in IL-1β-stimulated chondrocytes. In addition, IL-1β induced considerable phosphorylation of the NF-kB signaling pathway, which was inhibited by α-MG. Furthermore, the immunofluorescence staining demonstrated that α-MG could suppress IL-1β-induced p65 nuclear translocation. In vivo, cartilage treated with α-MG showed attenuated degeneration and had low Osteoarthritis Research Society International (OARSI) scores compared with the control group. Taken together, these results show that α-MG has potential therapeutic value in the treatment of OA.     

Theobromine mitigates IL-1β-induced oxidative stress,inflammatory response,and degradation of type II collagen in human chondrocytes

Osteoarthritis is one of the major causes of disability in elderly adults. Chondrocytes are responsible for the formation and remodeling of articular cartilage in joint tissue. The dysfunction of chondrocytes is a significant factor in the development of osteoarthritis. In the current study, we found that theobromine, a constituent of the cacao plant, possesses a preventive effect against interleukin (IL)-1β-induced chondrocyte dysfunction. Theobromine ameliorates IL-1β-induced production of cellular reactive oxygen species (ROS) and inflammatory mediators including cyclooxygenase-2 (COX-2) and prostaglandin E₂ (PGE₂). The presence of theobromine suppresses IL-1β-induced inducible nitro oxide synthase (iNOS) expression and cellular nitro oxide (NO) production. Theobromine also suppresses IL-1β-induced production of the pro-inflammatory cytokines tumor necrosis factor-α (TNF-α) and monocyte chemoattractant protein-1 (MCP-1), as well as matrix metalloproteinases (MMP)-3 and MMP-13. Additionally, theobromine mitigates IL-1β-induced type II collagen degradation. Mechanistically, we show that theobromine inhibits IL-1β-induced IκBα activation, nuclear factor-κB (NF-κB) protein p65 accumulation, and transfected NF-κB promoter activity, indicating that theobromine suppresses the NF-κB pathway in chondrocytes. Collectively, our study demonstrates that the natural molecule theobromine has a protective effect to counter cytokine-induced chondrocyte dysfunction, implying its beneficial effect in the prevention of osteoarthritis.     

Anti-inflammatory effects of aromatic-turmerone through blocking of NF-κB, JNK, and p38 MAPK signaling pathways in amyloid β-stimulated microglia

Amyloid β (Aβ) induces the production of neuroinflammatory molecules, which may contribute to the pathogenesis of numerous neurodegenerative diseases. Therefore, suppression of neuroinflammatory molecules could be developed as a therapeutic method. Aromatic (ar)-turmerone, turmeric oil isolated from Curcuma longa, has long been used in Southeast Asia as both a remedy and a food. In this study, we investigated the anti-inflammatory effects of ar-turmerone in BV2 microglial cells. Aβ-stimulated microglial cells were tested for the expression and activation of MMP-9, iNOS, and COX-2, the production of proinflammatory cytokines, chemokines, and ROS, as well as the underlying signaling pathways. Ar-turmerone significantly suppressed Aβ-induced expression and activation of MMP-9, iNOS, and COX-2, but not MMP-2. Ar-turmerone also reduced TNF-α, IL-1β, IL-6, and MCP-1 production in Aβ-stimulated microglial cells. Further, ar-turmerone markedly inhibited the production of ROS. Impaired translocation and activation of NF-κB were observed in Aβ-stimulated microglial cells exposed to ar-turmerone. Furthermore, ar-turmerone inhibited the phosphorylation and degradation of IκB-α as well as the phosphorylation of JNK and p38 MAPK. These results suggest that ar-turmerone impaired the Aβ-induced inflammatory response of microglial cells by inhibiting the NF-κB, JNK, and p38 MAPK signaling pathways. Lastly, ar-turmerone protected hippocampal HT-22 cells from indirect neuronal toxicity induced by activated microglial cells. These novel findings provide new insights into the development of ar-turmerone as a therapeutic agent for the treatment of neurodegenerative disorders.     

Protective effects of alogliptin against TNF-α-induced degradation of extracellular matrix in human chondrocytes

Osteoarthritis (OA) is a common debilitating disease most prevalent among the elderly population worldwide. Excessive degradation of the articular extracellular matrix is a pivotal event in the development of OA. Preventative treatments against the destruction of type II collagen and aggrecan, the two main components of the articular extracellular matrix, may serve as a novel therapy against the progression of OA. In the current study, we investigated whether the DPP-4 inhibitor alogliptin could prevent degradation of the articular extracellular matrix in human primary chondrocytes. Pretreatment with alogliptin successfully prevented degradation of type II collagen and aggrecan in a dose-dependent manner by reducing increased expression of MMP-1, -3, and -13 as well as ADAMTS-4 and -5 induced by treatment with TNF-α. Furthermore, pretreatment with alogliptin also reduced TNF-α-induced expression of IKKα/β, IκBα and NF-κB in human primary chondrocytes. This suggests that DPP-4 inhibitors such as alogliptin may be used as an effective preventative therapy against continued destruction of the articular extracellular matrix in OA.     

Morin inhibits interleukin-1β-induced nitric oxide and prostaglandin E2 production in human chondrocytes

It is well known that the inflammatory cytokines play important roles in osteoarthritis (OA). In the present study, we investigated the anti-inflammatory properties of morin in chondrocytes. The nitric oxide (NO) production was determined by Griess method, the prostaglandin E2 (PGE(2)) production was detected by Enzyme-linked immunosorbent assay (ELISA). The expression of inducible NO synthase (iNOS) and cyclooxygenase (COX)-2 were investigated by quantitative real-time PCR and western blot. In addition, western blotting and immunofluorescence staining were performed to investigate the protein level of inhibitor of nuclear factor-κB (IκB-α) and the translocation of nuclear factor kappa B (NF-κB). For the in vivo study, morin was administered by intra-articular injection in rats, and the gene expression of iNOS and COX-2 was assessed. We showed that morin inhibited the production of NO and PGE(2) as well as the expression of iNOS and COX-2 in interleukin-1-beta (IL-1β)-induced chondrocytes. In addition, morin suppressed the degradation of IκB-α as well as the translocation of NF-κB. In vivo study, morin exerted anti-inflammatory properties in an IL-1β-induced rat OA model. Our data suggest that morin possess potential value in the treatment of OA.     

Aucubin prevents interleukin-1 beta induced inflammation and cartilage matrix degradation via inhibition of NF-κB signaling pathway in rat articular chondrocytes

Proinflammatory cytokine interleukin-1β (IL-1β) plays a crucial role in the pathogenesis of Osteoarthritis (OA) by stimulating several mediators contributed to cartilage degradation. Aucubin, a natural compound derived from plants which has been shown to possess diverse biological activities including anti-inflammatory property, may benefit the IL-1β stimulated chondrocytes. The present study was aimed to investigate the effects of Aucubin on IL-1β stimulated rat chondrocytes. Rat chondrocytes were cultured and pretreated with Aucubin (1, 10, 20, 50 μM), and then stimulated with or without IL-1β (10 ng/ml). Gene and protein expression of MMP-3, MMP-9, MMP-13, cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS) was determined by real-time PCR and Western blotting respectively. Nitric oxide (NO) production was quantified by Griess reagent. Phosphorylation and nuclear translocation of p65 were detected by western blotting and immunofluorescence, respectively. We found that Aucubin significantly reversed the elevated gene and protein expression of MMP-3, MMP-9, MMP-13, iNOS, COX-2 and the production of NO induced by IL-1β challenge in rat chondrocytes. Furthermore, Aucubin was able to suppress the IL-1β-mediated phosphorylation and nuclear translocation of p65, indicating Aucubin may possibly act via the NF-κB signaling pathway. The present study proposes that Aucubin may be a potential therapeutic choice in the treatment of OA due to its anti-inflammatory and chondroprotective features.     

The protective effect of sophocarpine in osteoarthritis: An in vitro and in vivo study

BackgroundOsteoarthritis (OA) is a type of degenerative joint disease affecting millions of individuals worldwide. However, there are currently no great inflammatory treatments available for it. Sophocarpine (SPC), one of the key bioactive compounds derived from Sophora flavescens, has shown remarkable anti-inflammatory effects.MethodsIn this study, we evaluated the effect of SPC on preventing the progression of OA and investigated its molecular target involved. In brief, rat chondrocytes were pretreated with SPC and subsequently stimulated with IL-1β. We found that SPC reduced the production of pro-inflammatory cytokines, such as nitric oxide (NO), prostaglandin E2 (PGE2), tumor necrosis factor alpha (TNF-α), and interleukin-6 (IL-6). SPC also inhibited the expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) at both the gene and protein level. Moreover, SPC promoted the expression of anabolic factors Sox-9 and aggrecan, while inhibiting the expression of catabolic factors, such as matrix metalloproteinases 13 (MMP-13) and thrombospondin motifs 5 (ADAMTS-5) in rat chondrocytes. Mechanistically, we found that SPC inhibited nuclear factor kappa B (NF-κB) via the phosphatidylinositol 3 kinase (PI3K)/AKT pathway. The beneficial effects of SPC were also observed in vivo using a rat OA model.ConclusionsOur findings indicate that SPC may be a potential novel therapeutic in the treatment of OA.     

Downregulating Akt/NF-κB signaling and its antioxidant activity with Loureirin A for alleviating the progression of osteoarthritis: In vitro and vivo studies

Osteoarthritis(OA) is one of the most common diseases in orthopedics. It is characterized by degeneration of articular cartilage and chronic inflammation. In this study, we aim to elucidate the mechanism of Loureirin A’s therapeutic effect in OA progression. In vitro, Loureirin A pretreatment can significantly inhibit production of NO, PGE2, COX-2, TNF-α, iNOS andIL-6 induced by IL-1β in mouse articular chondrocytes. Moreover, Loureirin A suppressed the expression of matrix metalloproteinase-9(MMP-9), which leads to degradation of the extracellular matrix. The degradation of aggrecan and type II collagen protein in the extracellular matrix (ECM) stimulated by IL-1β was reversed. For signal pathway research, Loureirin A dramatically inhibited the phosphorylation of AKT and subsequent NF-κB entering into the nucleus caused by IL-1β in chondrocytes. Besides, a number of related indicators suggested that Loureirin A has a strong antioxidant activity in the treatment of osteoarthritis via increasing content of SOD2 and suppressing MDA and ROS. In addition, in vivo study demonstrated that Loureirin A could ameliorated the progression of OA in mice DMM model In conclusion, all results showed that Loureirin A may be a potential therapeutic candidate for the OA.     

龙胆苦苷对慢性前列腺炎的作用及对NF-κB和MAPKs信号通路活化的研究

目的探讨龙胆苦苷对慢性前列腺炎大鼠的作用及相关分子机制。方法采用完全弗氏佐剂诱导自身免疫性慢性前列腺炎大鼠模型,评估龙胆苦苷对大鼠前列腺器官指数、组织病理形态,以及大鼠前列腺组织中炎症因子IL-6、IL-1β、TNF-α和炎症介质COX-2、PGE2的表达水平及iNOS酶活性的影响。采用Western blot法检测大鼠前列腺组织中NF-κB和MAPKs通路相关分子蛋白表达水平。结果龙胆苦苷对慢性前列腺炎有保护作用,其可以降低前列腺器官指数,缓解前列腺组织病理损伤,此外,龙胆苦苷下调大鼠前列腺组织中炎症因子IL-6、IL-1β、TNF-α和炎症介质COX-2、PGE2的表达水平及iNOS酶活性,下调p-p65、p-ERK/ERK、p-JNK/JNK、p-p38/p38的表达水平。结论龙胆苦苷对慢性前列腺炎具有明显的保护作用,且这种作用可能与抑制NF-κB和MAPKs信号通路活化有关。     

狐臭柴茎挥发油体外抗炎活性及基于NF-κB和MAPKs信号通路作用机制的研究

目的:探究狐臭柴茎挥发油的体外抗炎活性及其作用机制。方法:通过水蒸气蒸馏法制备狐臭柴茎挥发油,采用GC-MS对其化学成分进行分析。通过Griess法和ELISA法测定挥发油对LPS诱导RAW264.7细胞上清液中一氧化氮(NO)、肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)水平的影响;qRT-PCR检测iNOS、COX-2、TNF-α、IL-6、IL-1β mRNA的表达;Western blot测定挥发油对iNOS、COX-2、NF-κB和MAPKs信号通路蛋白的影响。结果:从挥发油中共鉴定出74种化学成分,其中主要的化学成分是茅术醇(13.50%)。挥发油显著抑制NO、TNF-α和IL-6的分泌(P＜0.01),同时降低了促炎因子(TNF-α、IL-6和IL-1β)和促炎酶(iNOS和COX-2)的mRNA表达水平(P＜0.01)。Western blot研究表明挥发油能下调NF-κB信号通路细胞核p65蛋白表达、p65和IκBα磷酸化(P＜0.05或P＜0.01)。此外,还降低了MAPKs信号通路p38、JNK和ERK蛋白的磷酸化(P＜0.01)。结论:狐臭柴茎挥发油对LPS诱导RAW264.7细胞炎症模型具有较好的抗炎效果,其内在的分子机制与下调NF-κB和MAPKs信号通路有关。     

Salvianolic acid B inhibits IL-1β-induced inflammatory cytokine production in human osteoarthritis chondrocytes and has a protective effect in a mouse osteoarthritis model

Osteoarthritis (OA) is a chronic progressive disease that has complicated mechanisms that involve inflammation and cartilage degradation. In this study, we investigated the anti-inflammatory action of Salvianolic acid B (Sal B) in both human OA chondrocytes and a mouse OA model that was induced by destabilization of the medial meniscus. In vitro, chondrocytes were pretreated with Sal B (0, 25, 50, 100 μM) for 2 h, then incubated with IL-1β (10 ng/mL) for 24 h. NO production was determined by Griess method and PGE2 was assessed by ELISA. The expression of INOS, COX-2, MMP-13, ADAMTS-5 and NF-κB-related signaling molecules were tested by Western blotting. Immunofluorescence staining was used to detect P65 nuclear translocation. In vivo, the mouse OA model received intraperitoneal-injection of either Sal B (25 mg/kg) or saline every other day. Hematoxylin and Eosin, as well as Safranin-O-Fast green staining, were utilized to evaluate the severity of cartilage lesions up to 8 weeks following the surgery. Sal B inhibited the over-production of NO and PGE2, while the elevated expression of INOS, COX-2, MMP-13 and ADAMTS-5 were reversed by Sal B in IL-1β-induced chondrocytes. In addition, IL-1β significantly induced phosphorylation of NF-κB signaling, and this phosphorylation response was blocked by Sal B. Immunofluorescence staining demonstrated that Sal B could suppress IL-1β-induced p65 nuclear translocation. In vivo, the cartilage in Sal B-treated mice exhibited less cartilage degradation and lower OARSI scores. Taken together, Sal B possesses great potential value as a therapeutic agent for OA treatment.     

Anti-arthritic effects of chlorogenic acid in interleukin-1β-induced rabbit chondrocytes and a rabbit osteoarthritis model

Cartilage degradation is one of the pathological changes of osteoarthritis (OA), and accumulating evidence suggests an excess of matrix metalloproteinases (MMPs) plays a role in this cartilage breakdown. Here, we investigated the effects of chlorogenic acid (CGA) on the mRNA and protein expression of MMPs in interleukin (IL)-1β-induced rabbit chondrocytes and evaluated the in vivo effects of CGA in experimental OA induced by anterior cruciate ligament transection (ACLT) in rabbits. Using quantitative real-time PCR and ELISA to investigate the expression levels of MMP-1, MMP-3, MMP-13, and tissue inhibitors of metalloproteinase-1(TIMP-1) in IL-1β-induced rabbit chondrocytes, we showed that CGA inhibits the expression of these MMPs while increasing TIMP-1 expression, at both the mRNA and protein levels. In addition, IL-1β-induced activation of nuclear factor kappa B (NF-κB) and the degradation of inhibitor of κB (IκB)-α were suppressed by CGA. In rabbits, CGA decreased cartilage degradation as assessed by morphological and histological analyses. The down-regulation of MMP-1, MMP-3, and MMP-13 expression and up-regulation of TIMP-1 expression were also detected in CGA-treated cartilage compared with vehicle-treated cartilage, confirming these findings in an in vivo model. Taken together, these findings indicate that CGA may be considered as a possible candidate agent in the treatment of OA.