phenanthrenes and Arthritis--Rheumatoid

Celastrol and triptolide, chemical compounds isolated from Tripterygium wilfordii hook (also known as thunder god vine), are effective against rheumatoid arthritis (RA). Celastrol targets numerous signaling pathways involving NF‑κB, endoplasmic reticulum Ca2+‑ATPase, myeloid differentiation factor 2, toll‑like receptor 4, pro‑inflammatory chemokines, DNA damage, cell cycle arrest and apoptosis. Triptolide, inhibits NF‑κB, the receptor activator of NF‑κB (RANK)/RANK ligand/osteoprotegerin signaling pathway, cyclooxygenase‑2, matrix metalloproteases and cytokines. The present review examined the chemistry and bioavailability of celastrol and triptolide, and their molecular targets in treating RA. Clinical studies have demonstrated that T. wilfordii has several promising bioactivities, but its multi‑target toxicity has restricted its application. Thus, dosage control and structural modification of T. wilfordii are required to reduce the toxicity. In this review, future directions for research into these promising natural products are discussed.

Topics: Arthritis, Rheumatoid; Chemokines; Cyclooxygenase 2; Diterpenes; Epoxy Compounds; Humans; NF-kappa B; Pentacyclic Triterpenes; Phenanthrenes; Signal Transduction; Toll-Like Receptor 4; Tripterygium; Triterpenes

Triptolide (TPL), the active component of Tripterygium wilfordii Hook F (Twhf) has been used to treat cancer and bone loss conditions for over two hundred years in traditional Chinese medicine (TCM). In this paper, we reviewed the specific molecular mechanisms in the treatment of cancer, bone loss and cardiovascular disease. In addition, we analyze the toxicity of TPL and collect some optimized derivatives extracted from TPL. Although positive results were obtained in most cell culture and animal studies, further studies are needed to substantiate the beneficial effects of TPL.

Topics: Animals; Anti-Inflammatory Agents; Apoptosis; Arthritis, Rheumatoid; Autophagy; Cardiovascular Diseases; Diterpenes; Epoxy Compounds; Humans; Medicine, Chinese Traditional; Neoplasms; Osteoporosis; Phenanthrenes; Plant Extracts; Signal Transduction; Tripterygium

Triptolide (TP), a major extract of the herb

Topics: Animals; Antirheumatic Agents; Arthritis, Rheumatoid; Diterpenes; Drug Evaluation, Preclinical; Epoxy Compounds; Humans; Phenanthrenes

Extracts of Tripterygium wilfordii are effective in traditional Chinese medicine for treatment of rheumatoid arthritis (RA). Triptolide, a diterpenoid triepoxide purified from TWHF, has been identified as the major component of TWHF and might account for its therapeutic effects. To make for the clinical reasonable application and further development of triptolide, in this review was introduced the recent ten-years progress in mechanisms of anti-RA of it, including immunosuppression, anti-inflammation, inducing cell apoptosis, inhibiting vascular proliferation, protecting article cartilage and gene regulation. Triptolide is a potent immunosuppressant.

Topics: Animals; Apoptosis; Arthritis, Rheumatoid; Cell Proliferation; Cytokines; Dinoprostone; Diterpenes; Epoxy Compounds; Humans; Immunosuppressive Agents; Neovascularization, Pathologic; NF-kappa B; Phenanthrenes; Plants, Medicinal; T-Lymphocytes; Tripterygium

Glucocorticoids have anti-inflammatory, transrepression-mediated effects, although adverse events (AEs; transactivation-mediated effects) limit long-term use in patients with rheumatoid arthritis (RA). We evaluated the efficacy and safety of fosdagrocorat (PF-04171327), a dissociated agonist of the glucocorticoid receptor, versus prednisone or placebo.. In this 12-week, phase II, randomised controlled trial, 323 patients with moderate to severe RA were randomised 1:1:1:1:1:1:1 to fosdagrocorat (1 mg, 5 mg, 10 mg or 15 mg), prednisone (5 mg or 10 mg) or placebo, once daily. The primary endpoints (week 8) were American College of Rheumatology 20% improvement criteria (ACR20) responses, and percentage changes from baseline in biomarkers of bone formation (procollagen type 1 N-terminal peptide [P1NP]) and resorption (urinary N-telopeptide to urinary creatinine ratio [uNTx:uCr]). Safety was assessed.. ACR20 responses with fosdagrocorat 10 mg and 15 mg were superior to placebo, and fosdagrocorat 15 mg was non-inferior to prednisone 10 mg (week 8 model-predicted ACR20 responses: 47%, 61%, 69% and 73% vs 51%, 71% and 37% with fosdagrocorat 1 mg, 5 mg, 10 mg and 15 mg vs prednisone 5 mg, 10 mg and placebo, respectively). Percentage changes from baseline in P1NP with fosdagrocorat 1 mg, 5 mg and 10 mg met non-inferiority criteria to prednisone 5 mg. Corresponding changes in uNTx:uCr varied considerably. All fosdagrocorat doses reduced glycosylated haemoglobin levels. AEs were similar between groups; 63 (19.5%) patients reported treatment-related AEs; 9 (2.8%) patients reported serious AEs. No patients had adrenal insufficiency, treatment-related significant infections or laboratory abnormalities. No deaths were reported.. In patients with RA, fosdagrocorat 10 mg and 15 mg demonstrated efficacy similar to prednisone 10 mg and safety similar to prednisone 5 mg.. NCT01393639.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Antirheumatic Agents; Arthritis, Rheumatoid; Biomarkers; Drug Therapy, Combination; Female; Humans; Male; Middle Aged; Organophosphates; Phenanthrenes; Prednisone; Treatment Outcome; Young Adult

Dissociated agonists of the glucocorticoid receptor (DAGRs) show similar antiinflammatory effects but improved tolerability compared with standard glucocorticoid receptor (GR) agonists. The prodrug fosdagrocorat (PF-04171327), with active DAGR metabolite PF-00251802 (Metabolite-1), is postulated to show superior efficacy over placebo and prednisone in patients with moderate to severe rheumatoid arthritis (RA). We investigated the population pharmacokinetics of active Metabolite-1 and its active metabolite PF-04015475 (Metabolite-2) in patients with moderate to severe RA enrolled in a 12-week, phase II, randomized, double-blind study (NCT01393639). A simultaneous fit of a two-compartment model for Metabolite-1 and a one-compartment model for Metabolite-2 provided an adequate fit to the data. Significant covariates included weight, with an additional female effect on clearance of Metabolite-1 (∼26%) and Metabolite-2 (∼33%) compared with males. Age influenced clearance of Metabolite-1. In combination, age, weight, and sex predicted >twofold differences in area under the concentration-time curve of Metabolite-1 at the extremes.

Topics: Administration, Oral; Adult; Age Factors; Aged; Aged, 80 and over; Area Under Curve; Arthritis, Rheumatoid; Biological Variation, Population; Body Weight; Dose-Response Relationship, Drug; Double-Blind Method; Female; Glucocorticoids; Humans; Male; Middle Aged; Models, Biological; Organophosphates; Phenanthrenes; Prednisone; Prodrugs; Receptors, Glucocorticoid; Severity of Illness Index; Sex Factors; Young Adult

To assess efficacy and safety of fosdagrocorat (PF-04171327), a potential dissociated agonist of the glucocorticoid receptor, in rheumatoid arthritis (RA) patients.. This multicenter, double-blind, parallel-group, active- and placebo-controlled Phase 2 study (NCT00938587) randomized 86 patients (1 : 1 : 1 : 1) to receive fosdagrocorat 10 mg, fosdagrocorat 25 mg, prednisone 5 mg or placebo, all with stable background methotrexate therapy. The primary outcome was change from baseline in Disease Activity Score of 28 joints (DAS28-4[C-reactive protein (CRP)]) after 2 weeks of treatment. Secondary outcomes included American College of Rheumatology (ACR) response rates, change from baseline in ACR core components and Health Assessment Questionnaire Disability Index.. At week 2, improvements from baseline in DAS28-4(CRP) with fosdagrocorat 10 and 25 mg, prednisone 5 mg and placebo were -1.69, -2.22, -1.17 and -0.96, respectively, and were statistically significantly greater for both fosdagrocorat doses versus placebo (P < 0.05) and for fosdagrocorat 25 mg versus prednisone 5 mg (P < 0.001). The effects of fosdagrocorat on secondary outcomes were generally consistent with those observed for the primary outcome. Adverse events (AEs) were reported for eight (38%), three (14%), four (19%) and 12 (55%) patients treated with fosdagrocorat 10 and 25 mg, prednisone 5 mg and placebo, respectively. Most AEs were mild in severity. Four patients discontinued treatment due to AEs (fosdagrocorat 10 mg, n = 2; placebo, n = 2). There were no serious AEs.. Fosdagrocorat 10 and 25 mg demonstrated efficacy in improving signs and symptoms in RA patients, with manageable AEs. Additional studies are needed to assess the longer-term safety and efficacy of fosdagrocorat.

Topics: Adult; Aged; Antirheumatic Agents; Arthritis, Rheumatoid; Biomarkers; C-Reactive Protein; Disability Evaluation; Double-Blind Method; Drug Partial Agonism; Drug Therapy, Combination; Female; Glucocorticoids; Humans; Hydrocortisone; Joints; Male; Methotrexate; Middle Aged; Organophosphates; Phenanthrenes; Prednisone; Receptors, Glucocorticoid; Recovery of Function; Severity of Illness Index; Surveys and Questionnaires; Time Factors; Treatment Outcome

Fosdagrocorat (PF-04171327), a dissociated agonist of the glucocorticoid receptor, has potent anti-inflammatory activity in patients with rheumatoid arthritis with reduced adverse effects on bone health. To identify fosdagrocorat doses with bone formation marker changes similar to prednisone 5 mg, we characterized treatment-related changes in amino-terminal propeptide of type I collagen (P1NP) and osteocalcin (OC) with fosdagrocorat (1, 5, 10, or 15 mg) and prednisone (5 or 10 mg) in a phase II randomized trial (N = 323). The time course of markers utilized a mixed-effects longitudinal kinetic-pharmacodynamic model. Median predicted changes from baseline at week 8 with fosdagrocorat 5, 10, and 15 mg were -18, -22, and -22% (P1NP), and -7, -13, and -17% (OC), respectively. Changes with prednisone 5 and 10 mg were -15% and -18% (P1NP) and -10% and -17% (OC). The probability of fosdagrocorat doses up to 15 mg being noninferior to prednisone 5 mg for P1NP and OC changes was >90%.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Anti-Inflammatory Agents; Arthritis, Rheumatoid; Biomarkers; Double-Blind Method; Female; Humans; Male; Middle Aged; Models, Biological; Organophosphates; Osteocalcin; Peptide Fragments; Phenanthrenes; Prednisone; Procollagen; Receptors, Glucocorticoid; Young Adult

PF-04171327 is a dissociated agonist of the glucocorticoid receptor (DAGR) being developed to retain anti-inflammatory efficacy while reducing unwanted effects. Our aim was to conduct a longitudinal dose-response analysis to identify the DAGR doses with efficacy similar to or greater than prednisone 10 mg once daily (QD). The data included were from a Phase 2, randomized, double-blind, parallel-group study in 323 subjects with active rheumatoid arthritis on a background of methotrexate. Subjects received DAGR 1, 5, 10 or 15 mg, prednisone 5 or 10 mg, or placebo QD for 8 weeks. The Disease Activity Score 28-4 calculated using C-Reactive Protein (DAS28-4 CRP) was the efficacy endpoint utilized in this dose-response model. For DAGR, the maximum effect (Emax) on DAS28-4 CRP was estimated to be -1.2 points (95 % CI -1.7, -0.84), and the evaluated dose range provided 31-87 % of the Emax; for prednisone 5 and 10 mg, the estimated effects were -0.27 (95 % CI -0.55, 0.006) and -0.94 point (95 % CI -1.3, -0.59), respectively. Stochastic simulations indicated that the DAGR 1, 5, 10 and 15 mg have probabilities of 0.9, 29, 54 and 62 %, respectively, to achieve efficacy greater than prednisone 10 mg at week 8. DAGR 9 mg estimated probability was 50 % suggesting that DAGR ≥9 mg QD has an effect on DAS28-4 CRP comparable to or greater than prednisone 10 mg QD. This work informs dose selection for late-stage confirmatory trials.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Anti-Inflammatory Agents; Arthritis, Rheumatoid; C-Reactive Protein; Computer Simulation; Dose-Response Relationship, Drug; Double-Blind Method; Female; Humans; Longitudinal Studies; Male; Middle Aged; Models, Biological; Organophosphates; Phenanthrenes; Probability; Receptors, Glucocorticoid; Severity of Illness Index; Stochastic Processes; Treatment Outcome; Young Adult

To explore the correlations between diagnostic information and therapeutic efficacy in rheumatoid arthritis (RA) with decision tree model analysis.. Three hundred and ninety seven patients came from 9 clinical centers were randomly divided into the Western medicine (WM) group (n=194) treated with non-steroidal anti-inflammatory drugs and slow-acting antirheumatic drug and the Chinese medicine (CM) group (n=203) with basic therapy and syndrome-differentiation dependant TCM treatment. TCM and WM diagnostic information were collected. The ACR 20 was used for efficacy evaluation and the information of patients before treatment was analyzed by SAS 8.2 statistical package. Through single-factor exploratory analysis, odds ratio of efficacy and variable was calculated taken P < 0.2 as the including criteria for data mining analysis with decision tree model. All data were classified into the training set (75%) and verifying set (25%) with efficacy as the variable for layering to make further verification of the data-mining analysis.. Twenty variables were included in the CM group and 26 in the WM group in the data-mining model. In the former, 9 variables were positively correlated to the efficacy, including degree of arthralgia, tenderness and morning stiffness, number of swollen joint, and joint with tenderness, levels of IgM, rheumatoid factor (RF), C-reactive protein (CRP), and total assessment from doctor; and disease duration and degree of nocturnal polyuria were negatively correlated to that. While in the latter, 8 were positively correlated to the efficacy, including erythrocyte sedimentation rate (ESR), sour and weak waist and knees, white fur in tongue, joint ache and stiffness, swollen joint, and total assessment from doctor and patient, and red tongue with yellow fur and leucocyte count negatively correlated to it. Data mining with decision tree analysis revealed that different combinations of morning stiffness, slight red tongue, joint tenderness and nocturnal polyuria in the CM group, and those of white fur in tongue, CRP level, leucocyte count and morning stiffness in the WM group showed different efficacy, which were also verified in the randomly chosen verifying set.. To analyze the correlations between diagnostic information and therapeutic efficacy with decision tree analysis is conformed to the theory of TCM in applying treatment according to syndrome differentiation individually, thus it would contribute to elevate the accuracy of therapy.

Topics: Adolescent; Adult; Aged; Anti-Inflammatory Agents, Non-Steroidal; Arthritis, Rheumatoid; Decision Trees; Diterpenes; Drugs, Chinese Herbal; Epoxy Compounds; Female; Humans; Male; Medicine, Chinese Traditional; Middle Aged; Phenanthrenes; Phytotherapy; Treatment Outcome

Rheumatoid arthritis (RA) is an autoimmune disease characterized by synovial joint hyperplasia, joint inflammation, cartilage erosion and bone destruction. Macrophages play an essential role in the pathogenesis of RA, and folate receptor β (FR-β) is highly expressed on the surface of activated synovial macrophages in RA patients. Triptolide (TP) has anti-inflammatory properties, and it can protect the cartilage matrix, but its clinical application has been limited due to poor solubility, low bioavailability and systemic toxicity. Therefore, we constructed folate-modified triptolide liposomes (FA-TP-Lips) to target macrophages, thereby treating RA in a safe and effective way. The experiments indicated that FA-TP-Lips had properties of small particle size, uniform particle size distribution, high drug encapsulation and long circulation. Furthermore, FA-TP-Lips showed reduced cytotoxicity, increased cellular uptake and significant anti-inflammatory effects

Topics: Arthritis, Rheumatoid; Diterpenes; Epoxy Compounds; Folic Acid; Humans; Liposomes; Macrophages; Phenanthrenes

Triptolide (TPL) is an active component derived from Tripterygium wilfordii Hook F (TwHF) with therapeutic potential for rheumatoid arthritis (RA). However, the underlying mechanism of TPL is remains under-studied. Competing endogenous RNA (ceRNA) networks may participate in the response to TPL in RA. Herein, we sought to identify a TPL response-related ceRNA axis. A circular RNA (circRNA)-microRNA (miRNA)-mRNA ceRNA axis associated with the TPL response was constructed according to our previous study. Modulatory mechanisms of the ceRNA axis were ascertained through a series of experimentations. The clinical relevance of the ceRNA axis was also determined using computational models. Here, we found that TPL had excellent clinical effect on RA and promising therapeutic efficacy in experimental animals. The ceRNA axis of hsa-circ-0003353 (circ0003353), miR-31-5p, and CDK1 was identified as a candidate biomarker for the response of RA patients to TPL. TPL inhibited the viability, proliferation, and cell cycle entry of RA-fibroblast-like synoviocytes (FLSs), as well as the production of cytokines. Overexpression of circ0003353 abolished the inhibitory effects of TPL on RA-FLSs. Mechanistically, circ0003353 sponged miR-31-5p that inversely targeted CDK1 and manipulated the p21/Cyclin B axis. Additionally, consecutive rescue experiments indicated that the inhibitory impacts of TPL on RA-FLSs were dependent on the circ0003353/miR-31-5p/CDK1 axis. Molecular docking was also applied to predict the specific binding sites and binding capacity of TPL to related targets. In conclusion, the present study demonstrated that TPL repressed the cell growth and inflammatory response of RA-FLSs by mediating the expression of the circ0003353/miR-31-5p/CDK1 axis. This novel ceRNA axis may serve as a biomarker for screening RA patients who respond to TPL treatment, which holds potential applications in the diagnosis and therapy of RA.

Topics: Animals; Arthritis, Rheumatoid; CDC2 Protein Kinase; Cell Cycle; Cell Proliferation; Cells, Cultured; Diterpenes; Epoxy Compounds; Fibroblasts; Humans; MicroRNAs; Molecular Docking Simulation; Phenanthrenes; RNA, Circular; Synoviocytes

The abundant M1 macrophages in the joint synovium were the main factors that exacerbate rheumatoid arthritis (RA) by secreting various types of inflammatory cytokines. Here, we note that cGAS-STING, an important pro-inflammatory pathway, was significantly up-regulated in RA, enabling it be the potential target for RA therapy. Therefore, in this work, we developed M1 macrophages targeted micelles capable of cGAS-STING pathway inhibition for the smart treatment of RA. The folic acid (FA) and lauric acid (LA) were modified on dextran to obtain an amphiphilic polymer (FDL). Then, FDL was subsequently applied to encapsulate triptolide (TP) to form FDL@TP nanomicelles. The FDL@TP could target the joint and enhance the cell uptake of TP by M1 macrophages (overexpressing folate receptor-β), which also reduced the side effects of TP on normal tissues. In M1 macrophages, the released TP, acted as an anti-inflammatory and immunosuppressant, obviously down-regulated the expressions of cGAS and STING protein, and thus reduced the secretion of TNF-α, IL-1β and IL-6. Importantly, compared with the same dose of free TP, FDL@TP could significantly enhance the anti-inflammatory effect. Therefore, FDL@TP nanomicelles were believed to be superior candidates for the clinical treatment of RA.

Topics: Anti-Inflammatory Agents; Arthritis, Rheumatoid; Cytokines; Dextrans; Diterpenes; Epoxy Compounds; Folic Acid; Humans; Immunosuppressive Agents; Interleukin-6; Macrophages; Micelles; Nucleotidyltransferases; Phenanthrenes; Tumor Necrosis Factor-alpha

Triptolide (TP) has its unique curative effect in the treatment of rheumatoid arthritis (RA), but its application is limited by the poor water solubility and multi-organ toxicity. We herein developed a novel nanoparticle platform composed of L-ascorbate palmitate (VP, vitamin C derivative) that can deliver TP to synergistically treat arthritis and inhibit the occurrence of oxidative stress. The TP-loaded nanoparticles (termed TP-VP NPs) showed the suitable particle size (about 145 nm) and good physical stability. TP-VP NPs effectively down-regulated IL-1β, IL-6 and TNF-α levels to inhibit the erosion of synovitis and bone tissue, and alleviate the swelling and deformation of CIA mice's feet. Compared to the TP, TP-VP NPs could inhibit effectively the oxidative stress in liver, and alleviate significantly the triptolide-induced toxicity injury in liver, kidney and testicle. The results demonstrated that TP-VP NPs is a promising triptolide delivery system for the treatment of RA, which enhances the water solubility of TP and reduces the toxicity of TP

Topics: Animals; Arthritis, Rheumatoid; Ascorbic Acid; Diterpenes; Epoxy Compounds; Mice; Micelles; Palmitates; Phenanthrenes; Water

Rheumatoid arthritis (RA), the most common type of inflammatory arthritis, can cause bone damage and disability. Triptolide, a prominent treatment for RA, has satisfactory anti-inflammatory effects. However, the mechanism of action of triptolide in RA remains unknown.. This study aimed to explore the molecular mechanisms underlying triptolide-mediated improvements in RA and identify the miRNA pathway responsible for these effects.. We identified various dysregulated miRNAs associated with RA by mining previously described microarray data and verified and screened these candidates using RT-qPCR. Hematoxylin-eosin staining was then applied to identify pathological changes in the affected joints, and cell counting kit-8 analysis and flow cytometry were employed to examine cell proliferation and apoptosis, respectively. Extracted exosomes were verified using transmission electron microscopy.. Our results revealed that the legs of rats with collagen-induced arthritis presented with obvious swelling and bone damage, a high degree of inflammatory cell infiltration into the synovium, and structural changes to the cartilage. Data mining identified 39 dysregulated miRNAs in these tissues, and RT-qPCR further refined these observations to highlight miR-221 as a potential RA biomarker. Subsequent evaluations revealed that fibroblast-like synovial (FLS) cells secrete Exs carrying dysregulated miR-221 in vitro. These Exs mediate miR-221 levels, inflammation, and TLR4/MyD88 signaling via their fusion with chondrocytes, leading to changes in chondrocyte growth and metabolic factor levels. Additionally, the addition of triptolide impaired miR-221 expression, cell proliferation, inflammatory factors, and the protein levels of TLR4/MyD88 in RA-FLS and promoted the apoptosis of FLS. The therapeutic effect of triptolide on miR-221 Exs was reversed by miR-221 inhibitor in both normal and RA FLS.. Our research shows that effective treatment with triptolide is mediated by its regulation of growth and secretory functions of chondrocytes via the inhibition of miR-221 secretion by FLS, providing a new target and natural medicinal candidate for future RA treatments.

Topics: Animals; Anti-Inflammatory Agents; Apoptosis; Arthritis, Rheumatoid; Cell Proliferation; Cells, Cultured; Chondrocytes; Diterpenes; Down-Regulation; Eosine Yellowish-(YS); Epoxy Compounds; Exosomes; Fibroblasts; Hematoxylin; MicroRNAs; Myeloid Differentiation Factor 88; Phenanthrenes; Rats; Synovial Membrane; Toll-Like Receptor 4

The polarization of macrophages plays a critical role in the physiological and pathological progression of rheumatoid arthritis (RA). Activated M1 macrophages overexpress folate receptors in arthritic joints. Hence, we developed folic acid (FA)-modified liposomes (FA-Lips) to encapsulate triptolide (TP) (FA-Lips/TP) for the targeted therapy of RA. FA-Lips exhibited significantly higher internalization efficiency in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells than liposomes (Lips) in the absence of folate. Next, an adjuvant-induced arthritis (AIA) rat model was established to explore the biodistribution profiles of FA-Lips which showed markedly selective accumulation in inflammatory paws. Moreover, FA-Lips/TP exhibited greatly improved therapeutic efficacy and low toxicity in AIA rats by targeting M1 macrophages and repolarizing macrophages from M1 to M2 subtypes. Overall, a safe FA-modified liposomal delivery system encapsulating TP was shown to achieve inflammation-targeted therapy against RA via macrophage repolarization.

Topics: Animals; Arthritis, Experimental; Arthritis, Rheumatoid; Chemistry, Pharmaceutical; Cytokines; Diterpenes; Drug Carriers; Drug Liberation; Epoxy Compounds; Folic Acid; Inflammation Mediators; Lipopolysaccharides; Liposomes; Macrophages; Male; Mice; Phenanthrenes; Rats; Rats, Sprague-Dawley; RAW 264.7 Cells

Attenuating inflammatory response and relieving pain are two therapeutic therapeutical goals for rheumatoid arthritis (RA). Anti-inflammatory and analgesic drugs are often associated with many adverse effects due to nonspecific distribution. New drug delivery systems with practical targeting ability and other complementary strategies urgently need to be explored. To achieve this goal, an acupoint drug delivery system that can target deliver anti-inflammatory drugs and simulate acupuncture in relieving pain was constructed, which can co-deliver triptolide (TP) and 2-chloro-N (6)-cyclopentyl adenosine (CCPA).. We have successfully demonstrated that acupoint nanocomposite hydrogel composed of TP-Human serum album nanoparticles (TP@HSA NPs) and CCPA could effectively treat RA. The result shows that CCPA-Gel can enhance analgesic effects specifically at the acupoint, while the mechanical and thermal pain threshold was 4.9 and 1.6 times compared with non-acupoint, respectively, and the nanocomposite gel further enhanced. Otherwise, the combination of acupoint and nanocomposite hydrogel exerted synergetic improvement of inflammation, bone erosion, and reduction of systemic toxicity. Furthermore, it could regulate inflammatory factors and restore the balance of Th17/Treg cells, which provided a novel and effective treatment strategy for RA. Interestingly, acupoint administration could improve the accumulation of the designed nanomedicine in arthritic paws (13.5% higher than those in non-acupoint at 48 h), which may explain the better therapeutic efficiency and low toxicity.. This novel therapeutic approach-acupoint nanocomposite hydrogel, builds a bridge between acupuncture and drugs which sheds light on the combination of traditional and modern medicine.

Topics: Acupuncture Points; Acupuncture Therapy; Animals; Anti-Inflammatory Agents; Arthritis, Rheumatoid; Behavior, Animal; Delayed-Action Preparations; Diterpenes; Drug Delivery Systems; Epoxy Compounds; Humans; Male; Nanogels; Nanomedicine; Phenanthrenes; Rats; Rats, Sprague-Dawley

Rheumatoid arthritis (RA) is a chronicautoimmune disease, marked by joint swelling and pain, articular synovial hyperplasia, as well as cartilage and bone destruction. Triptolide (TP) is an anti-inflammatory molecule but its use to treat RA is limited due to poor solubility and extremely high toxicity. In this study, by encapsulating TP into a star-shaped amphiphilic block copolymer, POSS-PCL-b-PDMAEMA, we engineered a pH-sensitive TP-loaded nanomedicine (TP@NPs) to simultaneously reduce the toxicity of TP and improve its therapeutic efficacy. TP@NPs shows a uniform spherical structure with a hydrodynamic diameter of ~92 nm and notable pH-responsiveness. In vitro TP@NPs showed reduced cytotoxicity and cell apoptosis of treated RAW264.7 cells compared to free TP. And in vivo intravenous injection of indocyanine green-labeled NPs into a collagen-induced arthritis model in mice showed that the engineered compound had potent pharmacokinetic and pharmacodynamic profiles, while exhibiting significant cartilage-protective and anti-inflammatory effects with a better efficacy and neglible systemic toxicity even at an ultralow dose compared to free TP. These results suggest that TP@NPs may be a safe and effective therapy for RA and other autoimmune diseases.

Topics: Animals; Arthritis, Rheumatoid; Diterpenes; Epoxy Compounds; Hydrogen-Ion Concentration; Mice; Nanomedicine; Phenanthrenes

The UPLC-MS/MS method was established with good precision, accuracy and stability to determine the concentrations of TPL in biological samples, such as heart, liver, spleen, lung, kidney, plasma and joint.After being made into microspheres, TPL can stay in the joint tissue for a long time, further reducing the number of times joint cavity administration, and its sustained release effect was significantly improved compared with the solution dosage form.The pharmacokinetic parameters, such as AUC

Topics: Animals; Arthritis, Rheumatoid; Chromatography, Liquid; Diterpenes; Epoxy Compounds; Injections, Intra-Articular; Microspheres; Phenanthrenes; Rats; Tandem Mass Spectrometry

Rheumatoid arthritis (RA), recognized as a common chronic autoimmune disease, is characterized by the excessive proliferation and inflammatory infiltration of fibroblast-like synoviocytes (FLS). In this study, our purpose is to elucidate the mechanisms of triptolide (TPL) in the treatment of RA by regulating the long non-coding RNA (lncRNA) ENST00000619282, which promoted apoptosis and reduced inflammatory infiltration of FLS in RA (RA-FLS). RA-FLS was treated with different concentrations of TPL at different time points. CCK-8 assay, ELISA, RT-qPCR, immunofluorescence, TUNEL assay, and the transmission electron microscopy were used to measure the changes of cell viability, apoptosis, and the release of inflammatory cytokines. Next, the involvement of ENST00000619282 in TPL-mediated protection against RA was explored. ENST00000619282 expression was significantly increased in the peripheral blood mononuclear cells (PBMCs) of RA patients. ENST0000061928 expression in RA PBMCs was positively associated with ESR, RF, CCP, and DAS28, while TPL treatment led to a downregulation of ENST00000619282. In addition, ENST00000619282 was significantly increased in RA-FLS. Furthermore, overexpression of ENST00000619282 elevated the levels of pro-apoptotic and pro-inflammatory factors, while reduced the levels of anti-apoptotic proteins and antiinflammatory factors. Besides, TPL treatment could reverse these effects by ENST00000619282 overexpression. The anti-RA potential of TPL might be achieved by downregulating ENST00000619282, thereby promoting apoptosis, and reducing the inflammatory response in RA.

Topics: Apoptosis; Arthritis, Rheumatoid; Cell Proliferation; Cells, Cultured; Diterpenes; Epoxy Compounds; Fibroblasts; Humans; Inflammation; Leukocytes, Mononuclear; Phenanthrenes; RNA, Long Noncoding; Synoviocytes

Our previous study observed that long non-coding RNA (lncRNA) RP11-83J16.1 promoted rheumatoid arthritis (RA)-fibroblast-like synoviocyte (RA-FLS) proliferation, invasion and inflammation, which was downregulated by triptolide treatment. Therefore, the present study aimed to further investigate the mechanism and interaction between triptolide and lncRNA RP11-83J16.1 in RA treatment in vitro and in vivo.. RA-FLS was isolated and treated by different concentration of triptolide and lncRNA RP11-83J16.1 overexpression plasmid. Furthermore, collagen-induced arthritis (CIA) rat model was constructed followed by triptolide and lncRNA RP11-83J16.1 overexpression plasmid treatment.. Triptolide inhibited RA-FLS viability and lncRNA RP11-83J16.1 expression in a dose-dependent manner. Afterward, triptolide treatment inhibited RA-FLS proliferation, invasion, levels of inflammatory markers (TNF-α, IL-1β, IL-6, MMP-3, and MMP-9), inactivated lncRNA RP11-83J16.1, URI1 and β-catenin signaling, but promoted apoptosis. However, lncRNA RP11-83J16.1 overexpression weakened the effects of triptolide on regulating RA-FLS cell behaviors, URI1 signaling and β-catenin signaling. In CIA model, triptolide decreased arthritis score, hyperproliferation of synovial cells, inflammation infiltration of synovial tissue, inflammatory markers (TNF-α, IL-1β, IL-6, MMP-3, and MMP-9), inactivated lncRNA RP11-83J16.1, URI1 and β-catenin signaling, but increased cell apoptosis rate of synovial tissue. Nevertheless, lncRNA RP11-83J16.1 curtailed the treatment effect of triptolide in CIA model.. Triptolide decreases RA-FLS proliferation, invasion, inflammation and presents a therapeutic effect in CIA model via inactivating lncRNA RP11-83J16.1 mediated URI1 and β-catenin signaling.

Topics: Animals; Arthritis, Experimental; Arthritis, Rheumatoid; beta Catenin; Cell Movement; Cell Proliferation; Cells, Cultured; Diterpenes; Epoxy Compounds; Fibroblasts; Humans; Male; Phenanthrenes; Primary Cell Culture; Rats; Repressor Proteins; RNA, Long Noncoding; Signal Transduction; Synovial Membrane; Synoviocytes

Interruption of the Warburg effect - the observation that un-stimulated macrophages reprogram their core metabolism from oxidative phosphorylation toward aerobic glycolysis to become pro-inflammatory M1 macrophages upon stimulation - is an emerging strategy for the treatment of cancer and anti-inflammatory diseases such as rheumatoid arthritis. We studied this process with view to the discovery of novel therapeutics, and found that tylophorine-based compounds targeted a ribonucleoprotein complex containing caprin-1 and mRNAs of c-Myc and HIF-1α in LPS/IFN-γ stimulated Raw264.7 cells, diminished the protein levels of c-Myc and HIF-1α, and consequently downregulated their targeted genes that are associated with the Warburg effect, as well as the pro-inflammatory iNOS and COX2. The tylophorine-based compound DBQ 33b significantly meliorated the severity and incidence of type II collagen-monoclonal antibody-induced rheumatoid arthritis and diminished gene expressions of c-Myc, HIF-1α, iNOS, COX2, TNFα, and IL-17A in vivo. Moreover, pharmacological inhibition of either c-Myc or HIF-1α exhibited similar effects as the tylophorine-based compound DBQ 33b, even though inhibition of c-Myc reversed the induction of iNOS and COX2 in LPS/IFN-γ stimulated Raw264.7 cells to a lesser degree. Therefore, simultaneous inhibition of both c-Myc and HIF-1α is efficacious for anti-inflammation in vitro and in vivo and merits further study.

Topics: Alkaloids; Animals; Anti-Inflammatory Agents; Arthritis, Rheumatoid; Cell Cycle Proteins; Cyclooxygenase 2; Edema; Female; Hypoxia-Inducible Factor 1, alpha Subunit; Indolizines; Interferon-gamma; Lipopolysaccharides; Mice; Mice, Inbred BALB C; Nitric Oxide Synthase Type II; Phenanthrenes; Proto-Oncogene Proteins c-myc; Rats, Sprague-Dawley; RAW 264.7 Cells; RNA, Messenger; Tumor Necrosis Factor-alpha

Network pharmacology is a novel approach that uses bioinformatics to predict and identify multiple drug targets and interactions in disease. Here, we used network pharmacology to investigate the mechanism by which triptolide acts in rheumatoid arthritis (RA). We first searched public databases for genes and proteins known to be associated with RA, as well as those predicted to be targets of triptolide, and then used Ingenuity Pathway Analysis (IPA) to identify enriched gene pathways and networks. Networks and pathways that overlapped between RA-associated proteins and triptolide target proteins were then used to predict candidate protein targets of triptolide in RA. The following proteins were found to occur in both RA-associated networks and triptolide target networks: CD274, RELA, MCL1, MAPK8, CXCL8, STAT1, STAT3, c-JUN, JNK, c-Fos, NF-κB, and TNF-α. Docking studies suggested that triptolide can fit in the binding pocket of the six top candidate triptolide target proteins (CD274, RELA, MCL1, MAPK8, CXCL8 and STAT1). The overlapping pathways were activation of Th1 and Th2 cells, macrophages, fibroblasts and endothelial cells in RA, while the overlapping networks were involved in cellular movement, hematological system development and function, immune cell trafficking, cell-to-cell signaling and interaction, inflammatory response, cellular function and maintenance, and cell death and survival. These results show that network pharmacology can be used to generate hypotheses about how triptolide exerts therapeutic effects in RA. Network pharmacology may be a useful method for characterizing multi-target drugs in complex diseases.

Topics: Antirheumatic Agents; Arthritis, Rheumatoid; Binding Sites; Computational Biology; Diterpenes; Drug Development; Epoxy Compounds; Feasibility Studies; Humans; Molecular Docking Simulation; Phenanthrenes; Protein Interaction Maps; Signal Transduction

Tripterygium wilfordii Hook. F. (TwHF), a traditional Chinese Medicine, is effective in treating rheumatoid arthritis (RA), but its severe nephrotoxicity limits its extensive application. The nephrotoxic mechanism of Triptolide (TP), the main pharmacological and toxic component of TwHF, has not been fully revealed. This study was designed to explore the nephrotoxicity of TP in the RA state and the potential molecular mechanism. A rat collagen-induced arthritis (CIA) model was constructed and administered with TP for 28 days in vivo. Results showed that the kidney injury induced by TP was aggravated in the CIA state, the concentration of TP in the renal cortex was higher than that of the medulla after TP administration in the CIA rats, and the expression of organic cation transporter 2 (Oct2) in kidney was up-regulated under CIA condition. Besides, rat kidney slice study demonstrated that TP was transported by Oct2 and this was confirmed by transient silencing and overexpression of OCT2 in HEK-293T cells. Furthermore, cytoinflammatory models on HK-2 and HEK-293T cell lines were constructed by exposure of TNF-α or IL-1β to further explore the TP's renal toxicity. Results suggested that TNF-α exposure aggravated TP's toxicity and up-regulated the protein expression of OCT2 in both cell lines. TNF-α treatment also increased the function of OCT2 and finally OCT2 silencing confirmed OCT2 mediated nephrotoxicity of TP in HEK-293T cells. In summary, the exposure of TNF-α in RA state induced the expression of OCT2, which transported more TP into kidney cortex, subsequently exacerbated the kidney injury.

Topics: Animals; Arthritis, Experimental; Arthritis, Rheumatoid; Cell Line; Cytokines; Diterpenes; Epoxy Compounds; Female; HEK293 Cells; Humans; Kidney; Medicine, Chinese Traditional; Organic Cation Transporter 2; Phenanthrenes; Rats; Rats, Wistar; Renal Insufficiency; Tripterygium; Up-Regulation

This study aims to discuss the effect of triptolide (TPL) on rheumatoid arthritis (RA) and the mechanism related to osteoclast precursor (OCP) and osteoclast (OC). TNF-transgenic RA mice were treated with different doses of TPL by gavage. After the administration was finished, the curative effects were evaluated and compared, and the OCP apoptosis rates, the OC number, and the OC differentiation ability in vitro were detected. Finally, splenocytes of wild-type mice were cultured in vitro and induced to differentiate into OCP, and the cell apoptosis rate, cIAP2, and apoptotic effectors expression level were detected after cIAP2 overexpression and TPL administration. After TPL administration, the RA symptoms in the TPL groups were all better, the apoptosis rate of OCP was higher, and the amount of OC in vitro were lower than that in the control group (all P < 0.05), and all of the changes in the high-dose group were more obvious than the low-dose group. In splenocytes cells cultured in vitro, cIAP2 overexpression could decrease the apoptosis rate of OCPs and increase the OC number, and TPL treatment could down-regulate the cIAP2 and promote OCP apoptosis and OC reduction. In conclusion, TPL could induce OCP apoptosis and inhibit OC formation to effectively treat RA by mediating cIAP2 degradation.

Topics: Adult Stem Cells; Animals; Apoptosis; Arthritis, Rheumatoid; Baculoviral IAP Repeat-Containing 3 Protein; Diterpenes; Epoxy Compounds; HEK293 Cells; Humans; Mice; Mice, Transgenic; Osteoclasts; Phenanthrenes; Proteolysis; Tumor Necrosis Factor-alpha

The objective of this present study was to develop and evaluate the triptolide-loaded nanostructured lipid carriers (TPL-NLCs) for transdermal drug delivery system (TDDS). TPL-NLCs was prepared with emulsification technique and optimized by central composite design of a response surface methodology (CCD-RSM). The optimized TPL-NLCs were spherical and physically stable with the average size of 139.6.0 ± 2.53 nm and Zeta potential of -36.03 ± 2.41 mV. The encapsulation efficiency and drug loading were 97.15% ± 9.46 and 10.35% ± 1.12, respectively. Moreover, the in vitro release study showed that TPL-NLCs had a sustained release profiles and the in vitro penetration study indicated that TPL-NLCs could effectively penetrate into skin. Besides, the methodology of skin-blood synchronous microdialysis was established to evaluate the pharmacokinetics of TPL-NLCs in vivo and the results displayed that the TPL concentration in skin was higher than that in blood. And TPL-NLCs presented a remarkable effect of decreasing knee edema, inhibiting inflammation by regulating the levels of TNF-α, IL-1β and IL-6, which indicated that TPL-NLCs was a promising topical administration in treatment of edema and inflammation associated with rheumatoid arthritis (RA).

Topics: Administration, Cutaneous; Animals; Arthritis, Rheumatoid; Delayed-Action Preparations; Disease Models, Animal; Diterpenes; Drug Carriers; Drug Delivery Systems; Drug Liberation; Edema; Epoxy Compounds; Immunosuppressive Agents; Lipids; Male; Microdialysis; Nanostructures; Particle Size; Phenanthrenes; Rats; Rats, Sprague-Dawley; Skin Absorption

The present study investigated the mechanisms of apoptosis induced by cryptotanshinone (CT) in human rheumatoid arthritis fibroblast‑like synoviocytes (RA‑FLSs). Cell Counting kit‑8 assay was performed to determine the cytotoxic effects of CT in human RA‑FLSs, including primary RA‑FLS, HFLS‑RA and MH7A cells, and in HFLS cells derived from normal synovial tissue. Annexin V‑FITC/PI staining was used to detect the apoptotic effects of CT in HFLS‑RA and MH7A cells. Flow cytometry was performed to detect the apoptotic and reactive oxygen species (ROS) levels induced by CT in HFLS‑RA cells. Western blotting was used to assess the expression levels of proteins associated with apoptosis and with the mitogen‑activated protein kinase (MAPK), protein kinase B (Akt), and signal transducer and activator of transcription‑3 (STAT3) signaling pathways. The results demonstrated that CT treatment significantly suppressed HFLS‑RA and MH7A cell growth, whereas no clear inhibitory effect was observed in normal HFLS cells. CT exposure downregulated the expression levels of B‑cell lymphoma 2 (Bcl‑2), p‑Akt, p‑extracellular signal‑related kinase and p‑STAT3, while it upregulated the expression levels of Bcl‑2‑associated death promoter (Bad), caspase‑3, poly (ADP‑ribose) polymerase (PARP), p‑p38 and p‑c‑Jun N‑terminal kinase. Following ROS scavenging, the CT‑induced apoptosis and altered expression levels of Bcl‑2, Bad, cleaved caspase‑3 and cleaved PARP were restored. Furthermore, the Akt, MAPK and STAT3 signaling pathways were regulated by intracellular ROS. These results suggest that ROS‑mediated Akt, MAPK and STAT3 signaling pathways serve important roles in the CT‑induced apoptosis of RA‑FLSs.

Topics: Apoptosis; Arthritis, Rheumatoid; Biomarkers; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cell Survival; Fibroblasts; Humans; Mitochondria; Phenanthrenes; Proto-Oncogene Proteins c-akt; Reactive Oxygen Species; STAT3 Transcription Factor; Synoviocytes

Topics: Anti-Inflammatory Agents; Antioxidants; Arthritis, Rheumatoid; Autophagy; Cell Culture Techniques; Cell Line; Cell Movement; Cell Proliferation; Diterpenes; Epoxy Compounds; Humans; Oxidation-Reduction; Oxidative Stress; Phenanthrenes; Synoviocytes

The purpose of this study was to develop and evaluate triptolide-loaded cubic and hexagonal liquid crystals for transdermal drug delivery systems (TDDSs). We prepared and characterized triptolide-loaded lyotropic liquid crystals and evaluated for their percutaneous permeation properties in vitro and in vivo. We then used the adjuvant arthritic rat model and HaCaT cells to analyze the pharmacodynamics and conduct cell-stimulating studies of these liquid crystals. The optimized preparations were identified as cubic and hexagonal phase structures, respectively. Moreover, the in vitro percutaneous penetration studies demonstrated that compared to the homemade triptolide gel, cubic and hexagonal liquid crystals could significantly increase the percutaneous cumulative penetration of drugs within 48 h. Besides, the results of skin-blood synchronous microdialysis showed that the triptolide concentration in skin was higher than that in blood, and the cubic and hexagonal liquid crystals significantly increased the bioavailability of triptolide. Triptolide-loaded cubic and hexagonal liquid crystals presented excellent anti-arthritic effects, alleviating paw swelling and inhibiting inflammation by downregulating the levels of TNF-α and IL-1β. In vitro cell-stimulating studies displayed that triptolide-loaded cubic and hexagonal liquid crystals exhibited no obvious toxicity, which exhibited that triptolide-loaded cubic and hexagonal liquid crystals were remarkable biocompatibility. Collectively, triptolide-loaded cubic and hexagonal liquid crystals represented a promising candidate for rheumatoid arthritis therapy.

Topics: Administration, Cutaneous; Animals; Anti-Inflammatory Agents; Arthritis, Experimental; Arthritis, Rheumatoid; Cell Line; Diterpenes; Drug Carriers; Epidermal Cells; Epoxy Compounds; Humans; Interleukin-1beta; Liquid Crystals; Phenanthrenes; Rats; Rats, Sprague-Dawley; Skin; Skin Absorption; Surface Properties; Tumor Necrosis Factor-alpha

Our study aimed to determine the effects of sodium tanshinone IIA sulfonate (STS) on proliferation, migration, invasion, and inflammation in rheumatoid arthritis human fibroblast-like synoviocytes (RA-HFLSs). Firstly, results demonstrated STS reduced proliferation, migration, invasion in HFLSs. Also, we found that STS could alleviate the reorganizations of F-actin cytoskeleton in TNF-α-treated HFLSs. In addition, STS decreased the production of IL-1β, IL-6, MMP-1, and MMP-3 in TNF-α-treated RA-HFLSs. Further study showed that STS blocked MAPK/NF-κB activations in TNF-α-stimulated RA-HFLSs. Moreover, we illustrated that STS could alleviate rheumatoid arthritis progression and prevent inflammation damage in joint tissues of collagen-induced arthritis (CIA) mice. Taken together, this study suggested that STS inhibited proliferation, migration, invasion, and inflammation of RA-HFLSs by blocking MAPK/NF-κB pathways.

Topics: Animals; Anti-Inflammatory Agents; Arthritis, Experimental; Arthritis, Rheumatoid; Cell Line; Cell Movement; Cell Proliferation; Humans; Interleukin-1beta; Interleukin-6; Male; Matrix Metalloproteinase 1; Matrix Metalloproteinase 3; Mice, Inbred DBA; Phenanthrenes; Synoviocytes; Tumor Necrosis Factor-alpha

To discuss the curative effect and security of triptolide (TPL) on TNF transgenic (TNF-Tg) mice with rheumatoid arthritis (RA), and to explore the mechanism primarily.. 40 TNF-Tg RA mice were randomlydivided into five groups averagely: the control group, low-dose group (3.3 μg/kg/d TPL), middle-dose group (10 μg/kg/d TPL), high-dose group (33 μg/kg/d TPL) and MTX group (0.1 mg/kg/d MTX). Mice were administrated five days a week for six weeks. The arthritis deformation index, arthritis detumescencepercentage and the level of inflammatory factor in each group were recorded during theadministration. After administration, body weight, liver and renal function indexes, the apoptosis rates of osteoclast precursors (OCP), T and B lymphocytes in the peripheral blood and the number of osteoclast (OC) were detected and compared. μCT scanning and HE staining methods were taken to observethebone histomorphometry and bony erosion.. After administration, the arthritis deformation indexes were lower and arthritis detumescence percentageswere higher in TPL groups thanthe control group (P < 0.05), and the arthritis detumescence percentage in the high-dose group was higher than the MTX group (P < 0.05). The liver function index ALT increased after administrationin the high-dose group but was lower than that in the MTX group (P < 0.05). The level of IL-1α, IL-1β, and TNF-α decreased in the TPL groups and MTX group after administration;The apoptosis rates of OCP and T lymphocytes in middle and high dose TPL groups and MTX group were higher than other groups, and that in the high-dose group was higher than the MTX group (P < 0.05). Compared with the other groups, the bony erosion degree was lower and the number of OC was less and the parameters of bone histomorphometry were better in the high-dose group.. TPL could improvearthritic of TNF-Tg mice by decreasing the levels of pro-inflammatory cytokines, promoting the apoptosis of OCP, inhibiting the generation of OC and bone resorption. There was some toxic and side effect on liver for high-dose TPL which was weaker than the MTX.

Topics: Animals; Antirheumatic Agents; Apoptosis; Arthritis, Rheumatoid; Biomarkers; Bone Resorption; Chemical and Drug Induced Liver Injury; Cytokines; Disease Models, Animal; Diterpenes; Dose-Response Relationship, Drug; Epoxy Compounds; Inflammation Mediators; Joints; Lymphocytes; Mice, Inbred C57BL; Mice, Inbred CBA; Mice, Transgenic; Osteoclasts; Phenanthrenes; Risk Assessment; Time Factors; Tumor Necrosis Factor-alpha

Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by cellular infiltration into the joints and cartilage destruction. Neutrophils play a crucial role in the pathogenesis of RA. Triptolide (TP) is a bioactive compound derived from Tripterygium wilfordii Hook F, which has been used in folk medicine as a treatment for a variety of inflammatory disorders, including RA, for many centuries. Previous studies have shown that TP possesses anti-arthritic activity. However, the anti-arthritic mechanism of TP remains to be fully defined. In the present study, we used the adjuvant-induced arthritis (AA) murine model of RA to investigate the impact of TP on RA and neutrophil function. TP alleviated AA by reducing neutrophil recruitment and suppressing the expression of interleukin-6 and tumour necrosis factor-α in vivo. TP also suppressed the expression of pro-inflammatory cytokines in neutrophils, promoted neutrophil apoptosis and inhibited the migration, NETosis and autophagy of neutrophils in vitro. Based on our findings, TP effectively ameliorates RA by down-regulating neutrophil inflammatory functions, indicating that TP represents a potential therapeutic agent for RA.

Topics: Animals; Apoptosis; Arthritis, Experimental; Arthritis, Rheumatoid; Autophagy; Chronic Disease; Cytokines; Diterpenes; Epoxy Compounds; Extracellular Traps; Inflammation; Leukocyte Elastase; Lipopolysaccharides; Male; Mice, Inbred C57BL; Neutrophil Infiltration; Neutrophils; Peroxidase; Phenanthrenes

The balance between T helper 17 (Th17) cells and regulatory T (Treg) cells, plays a critical role in rheumatoid arthritis (RA). The differentiation of Th17 cells requires the activation of STAT3, which determines the balance of Th17/Treg. Here, we investigated the therapeutic effect of Cryptotanshinone (CTS) on collagen induced mouse arthritis and explored the underlying mechanisms.. Our study suggested that the anti-arthritis effects of CTS were attained through suppression of p300-mediated STAT3 acetylation. Our data suggest that CTS might be a potential immune modulator for RA treatment.

Topics: Acetylation; Animals; Antirheumatic Agents; Arthritis, Experimental; Arthritis, Rheumatoid; Cells, Cultured; Dose-Response Relationship, Drug; Drugs, Chinese Herbal; Female; HEK293 Cells; Humans; Male; Mice, Inbred DBA; Mutation; p300-CBP Transcription Factors; Phenanthrenes; Protein Processing, Post-Translational; Random Allocation; Rats, Wistar; Recombinant Proteins; RNA Interference; Specific Pathogen-Free Organisms; STAT3 Transcription Factor; Synoviocytes

High-resolution atomic force microscopy (AFM) was used for the in situ evaluation of the anti-inflammatory effects of triptolide on rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS) to understand the anti-RA effects of triptolide, based on the morphological and biophysical changes observed in RA-FLS. RA-FLS have been reported to play a primary role in inflammatory bone destruction during the development of RA and thus are regarded as an important target for RA treatment. Triptolide pretreatment significantly inhibited tumor necrosis factor-α-induced expression of the interleukin (IL)-1β, IL-6, and IL-8 genes in MH7A cells. Using AFM, we showed that triptolide-induced morphological damage in MH7A cells by inducing significant ultrastructure changes in the membrane, which were closely related to triptolide-induced apoptosis in MH7A cells. Using force measurements determined with AFM, triptolide was shown to increase the stiffness of MH7A cells. These findings not only revealed the strong anti-inflammatory effects of triptolide on RA-FLS, highlighting triptolide as a potential anti-RA agent, but also revealed the possible use of AFM for studying anti-inflammatory responses in RA-FLS, which we expect to be developed into a potential tool for anti-RA drug studies in RA-FLS.

Topics: Anti-Inflammatory Agents; Apoptosis; Arthritis, Rheumatoid; Cell Line; Cell Membrane; Cell Proliferation; Diterpenes; Epoxy Compounds; Humans; Interleukin-1beta; Interleukin-6; Interleukin-8; Microscopy, Atomic Force; Phenanthrenes; Synoviocytes; Tumor Necrosis Factor-alpha

Triptolide (TP), an active component isolated from Tripterygiumwilfordii Hook F, has therapeutic potential against rheumatoid arthritis (RA). However, the underlying molecular mechanism has not been fully elucidated. The aim of this study is to investigate the mechanisms of TP acting on RA by combining bioinformatics analysis with experiment validation. The human protein targets of TP and the human genes of RA were found in the PubChem database and NCBI, respectively. These two dataset were then imported into Ingenuity Pathway Analysis (IPA) software online, and then the molecular network of TP on RA could be set up and analyzed. After that, both in vitro and in vivo experiments were done to further verify the prediction. The results indicated that the main canonical signal pathways of TP protein targets networks were mainly centered on cytokine and cellular immune signaling, and triggering receptors expressed on myeloid cells (TREM)-1 signaling was searched to be the top one shared signaling pathway and involved in the cytokine and cellular immune signaling. Further in vitro experiments indicated that TP not only remarkably lowered the levels of TREM-1 and DNAX-associated protein (DAP)12, but also significantly suppressed the activation of janus activating kinase (JAK)2 and signal transducers and activators of transcription (STAT)3. The expression of tumor necrosis factor (TNF)-α, interleukin (IL)-1β and IL-6 in lipopolysaccharides (LPS)-stimulated U937 cells also decreased after treatment with TP. Furthermore, TREM-1 knockdown was able to interfere with the inhibition effects of TP on these cytokines production. In vivo experiments showed that TP not only significantly inhibited the TREM-1 mRNA and DAP12 mRNA expression, and activation of JAK2 and STAT3 in ankle of rats with collagen-induced arthritis (CIA), but also remarkably decreased production of TNF-α, IL-1β and IL-6 in serum and joint. These findings demonstrated that TP could modulate the TREM1 signal pathway to inhibit the inflammatory response in RA.

Topics: Animals; Anti-Inflammatory Agents; Antineoplastic Agents, Phytogenic; Arthritis, Rheumatoid; Cell Line, Tumor; Cytokines; Diterpenes; Epoxy Compounds; Humans; Male; Membrane Glycoproteins; Phenanthrenes; Rats, Sprague-Dawley; Receptors, Immunologic; Signal Transduction; Triggering Receptor Expressed on Myeloid Cells-1; Tripterygium

Triptolide, a primary active ingredient extracted from a traditional Chinese herb, Tripterygium wilfordii Hook F, has been demonstrated to have a positive therapeutic effect on patients with rheumatoid arthritis (RA); however, its mechanism of action against RA is not well established. Therefore, in the present study, we observed the effect of triptolide on the aggressive behavior of RA fibroblast-like synoviocytes (RA FLSs), and we explored its underlying signal mechanisms. We found that triptolide treatment significantly reduced the migratory and invasive capacities of RA FLSs in vitro. We also demonstrated that the invasion of RA FLSs into the cartilage, evaluated in the severe combined immunodeficiency (SCID) mouse co-implantation model, was attenuated by treatment with triptolide in vivo. Additionally, the immunofluorescence results showed that triptolide treatment decreased the polymerization of F-actin and the activation of matrix metalloproteinase 9 (MMP-9). To gain insight into the molecular signal mechanisms, we determined the effect of triptolide on the activation of MAPK signal pathways. Our results indicate that triptolide treatment reduced the TNF-α-induced expression of phosphorylated JNK, but did not affect the expression of phosphorylated p38 and ERK. A JNK-specific inhibitor decreased the migration of RA FLSs. We also observed that triptolide administration improved clinical arthritic conditions and joint destruction in mice with collagen-induced arthritis (CIA). Thus, our findings suggest that the therapeutic effects of triptolide on RA might be, in part, due to its contribution to the aggressive behavior of RA FLSs.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Arthritis, Experimental; Arthritis, Rheumatoid; Cell Movement; Cells, Cultured; Diterpenes; Epoxy Compounds; Female; Humans; Joints; MAP Kinase Signaling System; Mice, Inbred NOD; Mice, SCID; Phenanthrenes; Phosphorylation; Synoviocytes; Tumor Necrosis Factor-alpha

To explore the anti-inflammatory effect of triptolide (TPT) by regulating miR-155 in monocytes pre-stimulated by lipopolysaccharide (LPS) from rheumatoid arthritis (RA) patients.. Monocytes were isolated by CD14⁺ magnetic beads from peripheral blood mononuclear cells (PBMCs) of RA and stimulated by LPS for 24 hours. The levels of tumor-necrosis factor-α (TNF-α) and interleukin 6 (IL-6) in monocytes were detected by ELISA and the expression of miR-155 was measured by real-time quantitative PCR (qRT-PCR) in monocytes before and after the treatment of TPT at different concentrations. MiR-155 mimic and negative control were respectively transfected into the LPS-stimulated monocytes by Lipofectamine(TM)2000. Twenty-four hours later, the monocytes were treated with or without TPT for another 24 hours. TNF-α and IL-6 expressions in the cell culture supernatants were detected by ELISA and the expressions of suppressor of cytokine signaling-1 (SOCS1) and Src homology 2 domain-containing inositol 5-phosphatase 1 (SHIP-1) were tested by Western blotting.. TPT suppressed the expressions of TNF-α, IL-6 and miR-155 in LPS-stimulated peripheral blood monocytes from RA patients. Over-expression of miR-155 significantly reversed the down-regulation of TNF-α and IL-6 by TPT in monocytes. TPT up-regulated the expressions of SOCS1 and SHIP-1 in monocytes, but over-expressed miR-155 antagonized the effect of TPT on SHIP-1 while the expression of SOCS1 was not affected.. TPT suppressed the expression of miR-155 and up-regulated the release of SHIP-1, thus inhibiting the inflammatory response in the LPS-stimulated monocytes of RA patients.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Arthritis, Rheumatoid; Blotting, Western; Cells, Cultured; Diterpenes; Enzyme-Linked Immunosorbent Assay; Epoxy Compounds; Gene Expression Regulation; Humans; Inositol Polyphosphate 5-Phosphatases; Interleukin-6; Lipopolysaccharides; MicroRNAs; Monocytes; Phenanthrenes; Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases; Phosphoric Monoester Hydrolases; Reverse Transcriptase Polymerase Chain Reaction; Suppressor of Cytokine Signaling 1 Protein; Suppressor of Cytokine Signaling Proteins; Tripterygium; Tumor Necrosis Factor-alpha

Triptolide (TPT), an active compound extracted from Chinese herb Tripterygium wilfordii , has been used in therapy of rheumatoid arthritis (RA). In this study, after synovial fibroblasts from rheumatoid arthritis (RASFs) were treated with TPT, we investigated its effect on the differentiation of Th17 cells. Firstly, the mRNA level of cyclooxygenase (COX) wad detected by qRT-PCR and the protein level of prostaglandin E2 (PGE2) was tested by ELISA in RASFs treated with different concentrations (0, 10, 50, 100 nmol L-1 ) of TPT. Then after TPT pre-treated RASFs and RA CD4 + T cells wer e co-cultured for 3 days in the presence or absence of PGE2, IL-17 and IFN-gamma production in CD4 T cell subsets were detected by flow cytometry. The results showed TPT decreased the mRNA experssion of COX2 and the secretion of PGE2 in RASFs in a dose-dependent manner(P <0. 05). We further found that differentiation of Thl7 cells was downregulated in a dose-dependent manner, and exogenous PGE2 could reverse the inhibition of Th17 cell differentiation(P <0. 05). Taken together, our results demonstrated that TPT inhibited the mRNA level of COX2 and the secretion of PGE2 in RASFs, which partly led to impaired Th17 cell differentiation in vitro.

Topics: Arthritis, Rheumatoid; Cell Differentiation; Cell Line; Cyclooxygenase 2; Dinoprostone; Diterpenes; Epoxy Compounds; Fibroblasts; Gene Expression Regulation, Enzymologic; Humans; Middle Aged; Phenanthrenes; Synovial Fluid; Th17 Cells

Rheumatoid arthritis (RA) is characterized by a pre-vascular seriously inflammatory phase, followed by a vascular phase with high increase in vessel growth. Since angiogenesis has been considered as an essential event in perpetuating inflammatory and immune responses, as well as supporting pannus growth and development of RA, inhibition of angiogenesis has been proposed as a novel therapeutic strategy for RA. Triptolide, a diterpenoid triepoxide from Tripterygium wilfordii Hook F, has been extensively used in treatment of RA patients. It also acts as a small molecule inhibitor of tumor angiogenesis in several cancer types. However, it is unclear whether triptolide possesses an anti-angiogenic effect in RA. To address this problem, we constructed collagen-induced arthritis (CIA) model using DA rats by the injection of bovine type II collagen. Then, CIA rats were treated with triptolide (11-45 µg/kg/day) starting on the day 1 after first immunization. The arthritis scores (P<0.05) and the arthritis incidence (P<0.05) of inflamed joints were both significantly decreased in triptolide-treated CIA rats compared to vehicle CIA rats. More interestingly, doses of 11~45 µg/kg triptolide could markedly reduce the capillaries, small, medium and large vessel density in synovial membrane tissues of inflamed joints (all P<0.05). Moreover, triptolide inhibited matrigel-induced cell adhesion of HFLS-RA and HUVEC. It also disrupted tube formation of HUVEC on matrigel and suppressed the VEGF-induced chemotactic migration of HFLS-RA and HUVEC, respectively. Furthermore, triptolide significantly reduced the expression of angiogenic activators including TNF-α, IL-17, VEGF, VEGFR, Ang-1, Ang-2 and Tie2, as well as suppressed the IL1-β-induced phosphorylated of ERK, p38 and JNK at protein levels. In conclusion, our data suggest for the first time that triptolide may possess anti-angiogenic effect in RA both in vivo and in vitro assay systems by downregulating the angiogenic activators and inhibiting the activation of mitogen-activated protein kinase downstream signal pathway.

Topics: Angiogenesis Modulating Agents; Animals; Arthritis, Experimental; Arthritis, Rheumatoid; Cell Adhesion; Cell Movement; Cells, Cultured; Collagen; Diterpenes; Drug Combinations; Epoxy Compounds; Human Umbilical Vein Endothelial Cells; Humans; Laminin; Male; Mitogen-Activated Protein Kinases; Neovascularization, Pathologic; Phenanthrenes; Proteoglycans; Rats; Synovial Membrane; Vascular Endothelial Growth Factor A

To date, Kaposi sarcoma has not been mentioned among the adverse effects of triptolide/tripdiolide, ethyl acetate extracts or polyglycosides of the Chinese herbal remedy Tripterygium wilfordii Hook F.. A patient was diagnosed with rheumatoid arthritis at the age of 29 years. She underwent treatment with corticosteroids, methotrexate and gold sodium thiosulfate, and was chronically taking ketoprofen. At the age of 59 years she started to take a powder (≈2 g/day) from a Chinese physician for treatment of rheumatoid arthritis. This powder was supplied to her regularly for 10 years. At the age of 69 years, multiple soft, violaceous to dark-red patches, plaques, nodules and blisters of varying sizes appeared on a background of severely edematous skin on her legs, and later on her arms. Biopsy specimens of the leg lesions were diagnostic for human herpesvirus 8-associated Kaposi sarcoma. Triptolide (235 µg/1 g) and tripdiolide were found in the Chinese powder by the use of Liquid Chromatography Electrospray Ionization Mass Spectrometry. Administration of the powder was stopped and medication with paclitaxel was introduced. General condition of the patient improved and skin lesions diminished significantly.. This case indicates a possible association between triptolide/tripdiolide chronic intake and development of human herpesvirus 8-associated Kaposi sarcoma. Triptolide/tripdiolide could contribute to development of Kaposi sarcoma by reactivation of latent human herpesvirus 8, permitted by immunosuppression induced by triptolide.

Topics: Adult; Aged; Arthritis, Rheumatoid; Diterpenes; Edema; Epoxy Compounds; Female; Humans; Immunosuppressive Agents; Leg; Middle Aged; Phenanthrenes; Powders; Sarcoma, Kaposi; Skin Neoplasms

To study the effect of programmed cell death 5 (PDCD5) on apoptosis of rheumatoid arthritis fibroblast-like synoviocyte (RAFLS) induced by triptolide.. Cultured synovial cells in vitro from RA patients were transfected with Ad-PDCD5. In protein level, expression of PDCD5 protein in Ad-PDCD5 transfected RAFLS was detected by Western blot. RAFLS transfected with Ad-PDCD5 were cultured in presence or absence of triptolide and RAFLs apoptosis was determined by flow cytometry.. Transfection of RAFLS with increasing concentration of Ad-PDCD5 (50-300 MOI) resulted in dose-dependent increase of PDCD5 production. Apoptotic cells percentage of no transfection group, Ad-null group and Ad-PDCD5 group were, respectively, (22.41 +/- 3.87)%, (28.77 +/- 12.97)%, and (48.87 +/- 12.69)%. Alternatively, transfection without triptolide stimuli had no effect. The data showed that gene transfection of PDCD5 alone without triptolide was not sufficient to activate RAFLS apoptosis; PDCD5 acted as an enhancer rather than inductor of apoptosis.. Overexpression of PDCD5 could enhance apoptosis of RA FLS induced by triptolide; PDCD5 may be a potential therapeutic target to RA.

Topics: Adenoviridae; Apoptosis; Apoptosis Regulatory Proteins; Arthritis, Rheumatoid; Cell Separation; Cells, Cultured; Cloning, Molecular; Diterpenes; Epoxy Compounds; Fibroblasts; Flow Cytometry; Genetic Vectors; Humans; Neoplasm Proteins; Phenanthrenes; Synovial Membrane; Transfection

To determine the effects of triptolide (TP) on the expression of interleukin-18 (IL-18) and its receptor in phorbol 12-myristate 13-acetate (PMA)-stimulated rheumatoid arthritis synovial fibroblasts (RASF).. RASF were pretreated with TP (0-100 microg/L) for 2 h before stimulation with PMA (50 microg/L). The bioactivity of IL-18 in the supernatant was detected based on IFN-gamma secretion from IL-18-responding human myelomonocytic KG-1 cells. IL-18 level was analyzed by ELISA. To estimate the protein and mRNA expression of IL-18 and IL-18Ralpha in RASF, Western blot and quantitative RT-PCR were performed. Nuclear factor-kappaB (NF-kappaB) activity in the whole-cell extract of treated RASF was also measured using an ELISA-based method.. TP effectively inhibited the bioactivity of IL-18 in PMA-stimulated RASF. The expression of IL-18 and IL-18R at protein and gene levels was reduced by TP. NF-kappaB activity in PMA-stimulated RASF was profoundly suppressed by TP. These effects showed a high correlation with TP concentration (0-100 microg/L).. TP effectively inhibited the expression of IL-18 and its receptor in PMA-stimulated RASF. These results suggest a mechanism of TP in RA therapy.

Topics: Arthritis, Rheumatoid; Diterpenes; Epoxy Compounds; Fibroblasts; Humans; Immunosuppressive Agents; Interleukin-18; NF-kappa B; Phenanthrenes; Receptors, Interleukin-18; Synovial Membrane

To determine the effects of triptolide (TP) on the expression of interleukin-18 (IL-18) and its receptor in phorbol 12-myristate 13-acetate (PMA)-stimulated rheumatoid arthritis synovial fibroblasts (RASF).. RASF were obtained from the synovial tissue of patients with RA. RASF were pretreated with TP (0~100 ng/ml) for 2 h before stimulation with PMA (50 ng/ml). The bioactivity of IL-18 in the supernatant was detected based on IFN-gamma secretion from IL-18-responding human myelomonocytic KG-1 cells. IL-18 level was analyzed by ELISA. In situ expression of IL-18Ralpha was determined by immunofluorescence assay. To estimate the protein and mRNA expression of IL-18 and IL-18Ralpha in RASF, western blot and quantitative RT-PCR were performed. Nuclear factor-kappaB (NF-kappaB) activity in the whole-cell extract of treated RASF was also measured using an ELISA-based method.. TP effectively inhibited the bioactivity of IL-18 in PMA-stimulated RASF. The expression of IL-18 and IL-18R at protein and gene levels was reduced by TP. NF-kappaB activity in PMA-stimulated RASF was profoundly suppressed by TP. These effects showed a high correlation with TP concentration (0~100 ng/ml).. TP effectively inhibited the expression of IL-18 and its receptor in PMA-stimulated RASF. These results suggest a mechanism of TP in RA therapy.

Topics: Animals; Arthritis, Rheumatoid; Cells, Cultured; Diterpenes; Epoxy Compounds; Fibroblasts; Humans; Immunosuppressive Agents; Interleukin-18; Phenanthrenes; Receptors, Interleukin-18; Synovial Membrane; Tetradecanoylphorbol Acetate; Transcription Factor RelA

To explore the effect of programmed cell death 5(PDCD5) on apoptosis of rheumatoid arthritis fibroblast-like synoviocytes(RA FLS) induced by triptolide.. Cultured synovial cells in vitro from RA patients were transfected with Ad-PDCD5.At protein level, expression of PDCD5 in RA FLS infected with Ad-PDCD5 was detected by Western blot.RA FLS infected with Ad-PDCD5 were cultured in presence or absence of triptolide and apoptosis of RA FLS was determined by flow cytometry.. Infection of RA FLS with increasing concentrations of Ad-PDCD5(50-300 MOI) resulted in a does-dependent increase in the production of PDCD5. Apoptotic cells percentage for noinfection group, Ad-null group and Ad-PDCD5 group were(22.41 +/- 3.87)%, (28.77 +/- 12.97)% and (48.87 +/- 12.69)%, respectively. Alternatively, infection without addition of triptolide stimuli had no effect. The data showed that gene transfection of PDCD5 alone without addition of triptolide was not sufficient to activate RA FLS apoptosis, PDCD5 acted as an enhancer rather than inductor of apoptosis.. Overexpression of PDCD5 could enhance apoptosis of RA FLS induced by triptolide, PDCD5 may be a potential therapeutic target to RA.

Topics: Adenoviridae; Apoptosis; Apoptosis Regulatory Proteins; Arthritis, Rheumatoid; Cells, Cultured; Diterpenes; Epoxy Compounds; Fibroblasts; Humans; Neoplasm Proteins; Phenanthrenes; Synovial Membrane; Transfection

Apoptosis is reduced in the synovial tissue of patients with rheumatoid arthritis (RA), possibly due to decreased expression of pro-apoptotic genes. Programmed Cell Death 5 (PDCD5) has been recently identified as a protein that mediates apoptosis. Although PDCD5 is down-regulated in many human tumors, the role of PDCD5 in RA has not been investigated. Here we report that reduced levels of PDCD5 mRNA and protein are detected in RA synovial tissue (ST) and fibroblast-like synoviocytes (FLS) than in tissue and cells from patients with osteoarthritis (OA). We also report differences in the PDCD5 expression pattern in tissues from patients with these two types of arthritis. PDCD5 showed a scattered pattern in rheumatoid synovium compared with OA, in which the protein labeling was stronger in the synovial lining layer than in the sublining. We also observed increased expression and nuclear translocation of PDCD5 in RA patient-derived FLS undergoing apoptosis. Finally, overexpression of PDCD5 led to enhanced apoptosis and activation of caspase-3 in triptolide-treated FLS. We propose that PDCD5 may be involved in the pathogenesis of RA. These data also suggest that PDCD5 may serve as a therapeutic target to enhance sensitivity to antirheumatic drug-induced apoptosis in RA.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Apoptosis; Apoptosis Regulatory Proteins; Arthritis, Rheumatoid; Caspase 3; Cells, Cultured; Diterpenes; Epoxy Compounds; Fibroblasts; Gene Expression Regulation; Humans; Neoplasm Proteins; Phenanthrenes; Synovial Membrane; Tissue Distribution; Transfection

To test the effects of a novel tylophorine analog, DCB 3503, on the prevention and treatment of collagen-induced arthritis (CIA) and to elucidate its underlying mechanisms.. DBA/1J mice were immunized with type II collagen, and in some cases, lipopolysaccharide (LPS) was used to boost the development of arthritis. DCB 3503 was injected intraperitoneally before or after the onset of CIA. Mice were monitored to assess the effects of DCB 3503 on the clinical severity of the disease, and pathologic changes in the joints were examined histologically. Levels of tumor necrosis factor alpha (TNFalpha) and interleukin-1beta (IL-1beta) in serum and joint tissues were measured by enzyme-linked immunosorbent assay and by cytometric bead array analysis. The effect of DCB 3503 on LPS-induced proinflammatory cytokines from bone marrow-derived dendritic cells was determined by flow cytometry.. DCB 3503 significantly suppressed the development and progression of CIA. Moreover, DCB 3503 completely blocked the LPS-triggered acceleration of joint inflammation and destruction. Consistent with its effects in vivo, DCB 3503 significantly suppressed the synthesis of proinflammatory cytokines in inflamed joints as well as cytokine synthesis by macrophages examined ex vivo. Treatment also reduced the levels of inflammatory cytokines (IL-6, IL-12, TNFalpha, and monocyte chemotactic protein 1) produced by bone marrow-derived dendritic cells in vitro. However, DCB 3503 showed no direct effects on T cell proliferation and B cell antibody response.. Because of its ability to specifically suppress innate immune responses, DCB 3503 may be a novel therapeutic agent for inflammatory arthritis in humans.

Topics: Alkaloids; Animals; Arthritis, Experimental; Arthritis, Rheumatoid; Chemokine CCL2; Collagen Type II; Dendritic Cells; Enzyme-Linked Immunosorbent Assay; Indolizines; Interleukin-1; Interleukin-12; Male; Mice; Mice, Inbred DBA; Phenanthrenes; Tumor Necrosis Factor-alpha

To explore the effects of FK506 on the inhibition of cell proliferation and the expression of cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS) and their products PGE subset2 and NO in TNF-alpha-stimulated human rheumatoid arthritis synovial fibroblasts (RASF) treated with triptolide (TP), and to study the mechanisms involved when combining FK506 and TP in RA therapy.. RASF used in the experiments were obtained from the synovial tissue of patients with RA before being cultured. RASF were pretreated with FK506 (10 approximately 1000 nM) for 2 hours before being stimulated with TNF-alpha (20 ng/ml) in the presence or absence of TP (10 ng/ml) . RASF proliferation was determined by [3 supersetH]-TdR incorporation. Production of PGE subset2 and NO in culture supernatants of RASF was detected by competitive ELISA and enzymatic reduction of nitrate, respectively. Expressions of COX-2 and iNOS mRNA in RASF were analyzed by semi-quantitative RT-PCR. Expressions of COX-2 and iNOS protein were estimated by Western-blot and a cellular enzyme immunoassay. NFkappaB activity in whole-cell extract of treated RASF was also measured using an ELISA-based method.. Neither FK506 nor TP at a lower concentration (10 ng/ml) affected TNF-alpha-induced COX-2 and iNOS expressions or PGE subset2 and NO productions in synovial cells. Combined treatment of FK506 and a lower concentration of TP (10 ng/ml) reduced both COX-2 and iNOS mRNA and protein expression, and correspondingly reduced PGE subset2 and NO produced by synovial fibroblasts. This effect was highly correlated with FK506 concentration (10 approximately 1000 nM). NFkappaB activity in TNF-alpha-stimulated synovial cells was suppressed more profoundly by FK506 plus TP (10 ng/ml) than by TP (10 ng/ml) alone. However, no change was observed regarding the inhibition of synovial cell proliferation after combined treatment of FK506 and TP.. FK506 enhanced TP-mediated down-regulation of COX-2 and iNOS as well as their products PGE subset2 and NO in human TNF-alpha-stimulated RASF by more profoundly suppressing the activity of NFkappaB.

Topics: Arthritis, Rheumatoid; Base Sequence; Cell Line; Cyclooxygenase 2; Dinoprostone; Diterpenes; DNA Primers; Down-Regulation; Drug Synergism; Epoxy Compounds; Fibroblasts; Humans; Immunosuppressive Agents; Membrane Proteins; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Phenanthrenes; Prostaglandin-Endoperoxide Synthases; Reverse Transcriptase Polymerase Chain Reaction; Synovial Membrane; Tacrolimus; Tumor Necrosis Factor-alpha

The expression and activity of NF-kappaB in the synovium of collagen-induced arthritis (CIA) rats was detected in order to investigate the possible therapeutic effects of triptolide on rheumatoid arthritis (RA). The experimental Wistar rat model of CIA was set up by intradermal injection of emulsion of bovine collagen II and the successful rate of setting-up models was evaluated by arthritis index (AI). Rats were grouped randomly into three groups: normal, model and treatment group. The expression of TNF-alpha and IL-6 in synovial fluid was detected by ELISA, and the expression and activity of NF-kappaB in synovium by immunohistochemistry method and by electrophoretic mobility shift assay (EMSA) respectively. As compared with normal group, the expression of TNF-alpha and IL-6 in synovia (P < 0.05), and the expression and activity of NF-kappaB (P < 0.05) in synovium were increased in model group. There was statistical difference in above-mentioned indexes between model group and treatment group. Triptolide may play a protective role in RA via downregulating the expression and activity of NF-kappaB in synovium.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Arthritis, Experimental; Arthritis, Rheumatoid; Cattle; Diterpenes; Epoxy Compounds; Interleukin-6; Male; NF-kappa B; Phenanthrenes; Random Allocation; Rats; Rats, Wistar; Synovial Membrane; Tumor Necrosis Factor-alpha

Extracts of Tripterygium wilfordii Hook F (TWHF), a traditional Chinese herb, have been reported to show efficacy in patients with rheumatoid arthritis (RA). Since RA is not only characterized by inflammation but also by synovial proliferation in the joints, we examined whether triptolide (a constituent of TWHF) could influence the proliferation of rheumatoid synovial fibroblasts (RSF) by induction of apoptosis.. RSF were obtained from RA patients during surgery and were treated with triptolide under various conditions. The viability and proliferation of RSF were measured by the 4-[3-(4-iodophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]-1,3-benzene disulfonate (WST-1) assay and by 5-bromo-2'-deoxyuridine incorporation, respectively. Apoptosis was identified by detection of DNA fragmentation using an enzyme-linked immunosorbent assay and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL). The role of caspases in apoptosis of RSF was analyzed by measuring caspase-3 activity. Activation of the peroxisome proliferator-activated receptor (PPAR) gamma was assessed by a luciferase reporter gene assay using RSF transfected with a plasmid containing the peroxisome proliferator response element. Triptolide decreased viability, inhibited proliferation, and induced apoptosis of RSF in a concentration-dependent manner at very low (nM) concentrations. Caspase-3 activity was increased by treatment with triptolide and was suppressed by caspase inhibitors. Although PPARgamma activation was induced by 15-deoxy-Delta12,14-prostaglandin J2, triptolide did not induce it under the same experimental conditions. An extract of TWHF also induced DNA fragmentation in RSF.. The mechanism of action remains to be studied; however, triptolide may possibly have a disease-modifying effect in patients with RA.

Topics: Apoptosis; Arthritis, Rheumatoid; Caspases; Cell Division; Cell Survival; Cells, Cultured; Diterpenes; Drugs, Chinese Herbal; Epoxy Compounds; Fibroblasts; Humans; Phenanthrenes; Synovial Membrane; Tripterygium

To explore the effects of Triptolide (TP) on TNFalpha-induced cell proliferation and expressions of cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS) and their inducing products PGE2, NO in human rheumatoid arthritis synovial fibroblasts (RASF).. Fibroblasts (RASF) were obtained from synovial tissue of patients with RA and were cultured in vitro. RASF were stimulated with TNFalpha(20 microg/L) in the presence or absence of TP(0 - 100 microg/L) for 20 h. The RASF proliferation was determined by (3)H-TdR incorporation, and the productions of PGE2 and NO in culture supernatants of RASF were detected with competitive ELISA and enzyme reduction of nitrate. Expressions of COX-2 and iNOS mRNA in RASF were analyzed by semi-quantitative RT-PCR. Expressions of COX-2 and iNOS protein were estimated by Western-blot method and cellular enzyme immunoassay in synovial fibroblasts. NF-kappaB activity in whole-cell extract of RASF was also measured by an ELISA-based method.. TP (>20 microg/L) down-regulated markedly TNFalpha-induced COX-2 and iNOS mRNA and protein expression, and their inducing products PGE2 and NO of synovial fibroblasts. This effect was positively correlated with TP concentrations. NF-kappaB activity in TNFalpha-stimulated synovial cells was suppressed profoundly by TP treatment (IC(50) approximately 35microg/L). The activity of NF-kappaB was correlated with the levels of COX-2 and iNOS expression in TNFalpha-stimulated RASF. No change was observed in proliferation of synovial cells after treatment of TP.. TP could significantly down-regulate TNFalpha-induced COX-2, iNOS expression and production of PGE2, NO in human RASF, which is associated with the suppression of NF-kappaB activity.

Topics: Arthritis, Rheumatoid; Cyclooxygenase 2; Diterpenes; Epoxy Compounds; Fibroblasts; Gene Expression Regulation; Humans; Isoenzymes; Membrane Proteins; NF-kappa B; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Phenanthrenes; Prostaglandin-Endoperoxide Synthases; RNA, Messenger; Synovial Membrane; Tumor Necrosis Factor-alpha

To explore the effects of FK506 on the inhibition by triptolide (TP) of cell proliferation and expressions of cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS) and their inducing products PGE2, NO in human rheumatoid arthritis synovial fibroblasts (RASF), and to study the mechanisms of combination of FK506 and TP in RA therapy, RASF used in the experiments were obtained from synovial tissue of patients with RA and were cultured. RASF were pretreated with FK506(10-1000 nmol/L)for 2 h, then the cells were stimulated with TNF alpha(20 microg/L) in the presence or absence of TP (10 microg/L). The RASF proliferation was determined by [(3)H]-TdR incorporation, and the productions of PGE2 and NO in culture supernatants of RASF were detected with competitive ELISA and enzyme reduction of nitrate. Expression of COX-2 and iNOS mRNA in RASF were analyzed by semi quantitative RT-PCR. Expressions of COX-2 and iNOS protein were estimated by Western blot method and cellular enzyme immunoassay in synovial fibroblasts. NF-kappa B activity in whole-cell extract of RASF was also measured by an ELISA-based method. Results showed that neither FK506 nor TP at lower concentration (10 microg/L) alone affected TNF alpha-induced COX-2, iNOS expression and production of PGE2, NO in synovial cells. Combined treatment of FK506 and a lower concentration of TP (10 microg/L) down-regulated COX-2 and iNOS mRNA and protein expression, and their inducing products PGE2 and NO of synovial fibroblasts. This effect was positively correlated with FK506 concentrations (10-1000 nmol/L). NF-kappa B activity in TNF alpha-stimulated synovial cells was suppressed more profoundly by FK506 plus TP (10 microg/L) treatment than those with TP (10 microg/L) alone. No change was observed in inhibition of proliferation of synovial cells after combined treatment of FK506 and TP. In conclusion, FK506 enhanced TP-mediated down-regulation of COX-2, iNOS and their inducing products PGE2, NO in human RASF by suppressing the activity of NF-kappa B.

Topics: Arthritis, Rheumatoid; Blotting, Western; Cell Division; Cells, Cultured; Cyclooxygenase 2; Dinoprost; Diterpenes; Enzyme-Linked Immunosorbent Assay; Epoxy Compounds; Fibroblasts; Gene Expression Regulation, Enzymologic; Humans; Immunosuppressive Agents; Isoenzymes; Membrane Proteins; NF-kappa B; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Phenanthrenes; Prostaglandin-Endoperoxide Synthases; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Synovial Membrane; Tacrolimus; Tumor Necrosis Factor-alpha

Tripterygium Wilfordii Hook multi-glycosides T2 has been wildly used in China in treatment of RA. T4 was abstracted from T2 and was reported much more efficient in anti-inflammatory and immune suppression than T2. This study was to investigate the effect Tripterygium Wilfordii Hook T4 monomer on proliferation and interleukin-6 production of synovial fibroblasts of patients with rheumatoid arthritis.. Synovium was obtained from patients with rheumatoid arthritis undergoing synovectomies or joint replacement. Cultures of synovial fibroblasts were established. After 3 generations, cultured synovial fibroblasts were stimulated with IL-1. Then 1.5 ng/ml, 5 ng/ml and 15 ng/ml T4 were added, and synovial fibroblasts were cultured in the presence of T4 for 48 hours. Cell proliferation was assayed using MTT method. IL-6 level of supernatant was measured by ELISA.. Proliferation of synovial fibroblasts was inhibited by T4. The proliferation inhibition effect of T4 was dose dependent and inhibition rate was 5.18%, 10.95% and 21.37%, respectively. And T4 had no effect on IL-6 production by IL-1 stimulated synovial fibroblasts.. T4 might control the disease activity of RA by inhibiting the proliferation of synoviocyte. And T4 might not influence the concentration of IL-6 in synovial fluid, as a central effect, since IL-6 has protective effect on articular cartilage.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Arthritis, Rheumatoid; Cell Division; Cells, Cultured; Diterpenes; Drug Antagonism; Female; Fibroblasts; Humans; Interleukin-1; Interleukin-6; Male; Middle Aged; Phenanthrenes; Synovial Membrane; Tripterygium

Tripchlorolide(T4) is an active ingredient recently isolated from Tripterygium Wilfordii Hook. The in vitro effects of T4 on the peripheral blood mononuclear cells(PBMC) of healthy persons(n = 10), the digested single synovium cells(DSSC) (n = 3) and PBMC(n = 6) from RA patients on production of immunoglobulins (Ig) were studied using Elisa method. The results showed that T4 at concentrations from 5 ng.ml-1 to 35 ng.ml-1 significantly reduced the production of Ig by PWM-stimulated PBMC of healthy persons in a dose dependent manner. At concentration of 25 ng.ml-1, T4 also reduced the Ig secretion of RA-PBMC and DSSC. It is hoped that T4 would be an encouraging new drug of herbal nature for the treatment of RA.

Topics: Adult; Arthritis, Rheumatoid; Cells, Cultured; Diterpenes; Dose-Response Relationship, Drug; Drugs, Chinese Herbal; Female; Humans; Immunoglobulin G; Immunoglobulin M; Immunosuppressive Agents; Leukocytes, Mononuclear; Male; Phenanthrenes; Synovial Membrane

Encouraged by good therapeutic results of the semipurified preparations of Tripterygium Wilfordii Hook (TWH) (code name T4) a single-active ingredient of TWH with the code name of T4 was obtained. The in vitro effects of T4 on the bioactivity of TNF produced by PBMC of normal subjects (n = 10) and RA patients (n = 6) or the RA digested synovium single cells (DSSC) were studied. The results showed that under pretreatment of PBMC of healthy persons with T4 (35 ng/ml) for two hours, TNF were reduced to 36.13 +/- 1.75% in comparison with the control of 63.33 +/- 2.51% (P < 0.001), with an inhibition rate of 40.31%. Similarly, the inhibition rate for the PBMC of RA patients (P < 0.001), was 25.54%. The fact that coculture of PBMC and T4 for 6 hours instead of treatment with T4 for 2 hours caused an inhibition rate of 40.51% (P < 0.01), might indicate that time of interaction was also a factor of importance. T4 also reduced TNF secretion by RA-DSSC from 72.1 +/- 6.1% to 51.6 +/- 2.0% with an inhibition rate of 28.4%. The aformentioned indicates that T4 appears to possess suppressive actions on production of TNF by PBMC and DSSC of RA patients. Therefore, it is possible that T4 may be used clinically for RA patients.

Topics: Adult; Anti-Inflammatory Agents, Non-Steroidal; Arthritis, Rheumatoid; Cells, Cultured; Diterpenes; Drugs, Chinese Herbal; Female; Humans; Immunosuppressive Agents; Leukocytes, Mononuclear; Male; Phenanthrenes; Synovial Membrane; Tripterygium; Tumor Necrosis Factor-alpha

Tripchlorolide (T4) is a single active ingredient recently isolated from Tripterygium wilfordii Hook. The effects of T4 on the proliferation of peripheral blood mononuclear cells (PBMC) of rheumatoid arthritis (RA) patients and normal subjects were studied, RA patients and sex- and age-matched normal subjects (each 8 cases) were selected. PBMC were incubated with T4 of various doses (10, 20, 30 and 40 ng/ml) in the presence or absence of PHA for 72 hours. The results were as follows: T4 remarkably inhibited the proliferation of both PHA-stimulated and unstimulated PBMC of normal subjects and PHA-stimulated PBMC of RA patients as measured by MTT colormetric method. However, T4 showed a biphasic reaction in unstimulated PBMC of 3 RA patients. These results indicates that T4 would be useful in the therapy of RA. The significance of biphasic reaction occurring in RA patients needs to be further investigated.

Topics: Adult; Anti-Inflammatory Agents, Non-Steroidal; Arthritis, Rheumatoid; Cell Division; Diterpenes; Female; Humans; Leukocytes, Mononuclear; Male; Phenanthrenes

The effects of tripchlorolide, an active ingredient (T4) of Tripterygium wilfordii Hook on the production of prostaglandin E2(PGE2) by synovial cells of rheumatoid arthritis (RA) patients were investigated. Six cases of definite RA (female 5, male 1, mean age 45 with an average course of disease of 9 years) were selected. Surgically obtained synovium specimen were dissociated into digested synovial single cells (DSSC). The cells were incubated with various concentrations of T4 for 48 hours. Using radioimmunoassay T4 was found to significantly inhibit the production of PGE2 (control 6.10 +/- 2.30 vs T4 treated 0.58 +/- 0.47 x 10(-5) mol.L-1) by short-term cultured DSSC of RA patients. The results of this study suggests that T4 may be useful in the forthcoming treatment of RA due to its inhibition of production of PGE2 by synovial cells.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Arthritis, Rheumatoid; Cells, Cultured; Dinoprostone; Diterpenes; Drugs, Chinese Herbal; Female; Humans; Male; Middle Aged; Phenanthrenes; Synovial Membrane; Tripterygium

It was satisfactory using the ethyl acetate extract of Tripterygium wilfordii (TW) to treat rheumatoid arthritis (RA). The results showed that the ethyl acetate extract of TW was the effective component part of TW. Although the therapeutic effect of 15 cases with RA cured by triptolide was as effective as the ethyl acetate extract of TW, triptolide could impair some patients' hearts. This suggested that triptolide was one of the main effective elements of TW, but it was also one of the main toxic elements. Clinical research indicated that: (1) the effect of TW in treating RA was the synergistic action of elements with triptolide as the main; (2) triptolide may act as a major standard of controlling the quality of the preparation of TW and assure clinical use safely.

Topics: Adolescent; Adult; Arthritis, Rheumatoid; Diterpenes; Drugs, Chinese Herbal; Epoxy Compounds; Female; Humans; Male; Middle Aged; Phenanthrenes; Tripterygium

Topics: Animals; Anti-Inflammatory Agents; Arthritis, Rheumatoid; Glycyrrhiza; Granuloma; Humans; Phenanthrenes; Plants, Medicinal; Rats; Steroids