RESUMO
BACKGROUND: Gout is caused by monosodium urate (MSU) crystals deposition to trigger immune response. A recent study suggested that inhibition of Class I Histone deacetylases (HDACs) can significantly reduce MSU crystals-induced inflammation. However, which one of HDACs members in response to MSU crystals was still unknown. Here, we investigated the roles of HDAC3 in MSU crystals-induced gouty inflammation. METHODS: Macrophage specific HDAC3 knockout (KO) mice were used to investigate inflammatory profiles of gout in mouse models in vivo, including ankle arthritis, foot pad arthritis and subcutaneous air pouch model. In the in vitro experiments, bone marrow-derived macrophages (BMDMs) from mice were treated with MSU crystals to assess cytokines, potential target gene and protein. RESULTS: Deficiency of HDAC3 in macrophage not only reduced MSU-induced foot pad and ankle joint swelling but also decreased neutrophils trafficking and IL-1ß release in air pouch models. In addition, the levels of inflammatory genes related to TLR2/4/NF-κB/IL-6/STAT3 signaling pathway were significantly decreased in BMDMs from HDAC3 KO mice after MSU treatment. Moreover, RGFP966, selective inhibitor of HDAC3, inhibited IL-6 and TNF-α production in BMDMs treated with MSU crystals. Besides, HDAC3 deficiency shifted gene expression from pro-inflammatory macrophage (M1) to anti-inflammatory macrophage (M2) in BMDMs after MSU challenge. CONCLUSIONS: Deficiency of HDAC3 in macrophage alleviates MSU crystals-induced gouty inflammation through inhibition of TLR2/4 driven IL-6/STAT3 signaling pathway, suggesting that HDAC3 could contribute to a potential therapeutic target of gout.
Assuntos
Acrilamidas , Gota , Histona Desacetilases , Macrófagos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fenilenodiaminas , Ácido Úrico , Animais , Ácido Úrico/toxicidade , Histona Desacetilases/metabolismo , Histona Desacetilases/genética , Histona Desacetilases/deficiência , Macrófagos/metabolismo , Macrófagos/efeitos dos fármacos , Gota/metabolismo , Gota/patologia , Camundongos , Inflamação/metabolismo , Inflamação/induzido quimicamente , Masculino , Artrite Gotosa/induzido quimicamente , Artrite Gotosa/metabolismo , Artrite Gotosa/patologia , Modelos Animais de Doenças , Transdução de Sinais/efeitos dos fármacosRESUMO
BACKGROUND: Previous studies have revealed that Sirt3 deficiency is associated with several inflammatory responses. The purpose of this study is to investigate the role and potential molecular mechanisms of Sirt3 in the inflammation induced by monosodium urate (MSU) crystals. METHODS: The Sirt3 expression level in the peripheral blood mononuclear cells (PBMCs) of patients with gout was measured. Function and molecular mechanism of Sirt3 in MSU crystal-induced inflammation were investigated in bone marrow-derived macrophages (BMDMs), C57BL/6 mouse, and Sirt3-/- mouse. RESULTS: Sirt3 expression was decreased in the PBMCs of patients with gout. Sirt3 agonist (Viniferin) inhibited the acetylation levels of mitochondrial proteins including the SOD2 protein. RNA sequencing, bio-informatics analysis, RT-PCR, and Western blot demonstrated that Sirt3 could suppress the expression of Acod1 (Irg1), which plays an important role in gout. In BMDMs treated with palmitic acid (C16:0) plus MSU crystals, Acod1 knockdown repressed mitochondrial reactive oxygen species (mtROS) over-production, macrophage migration, and mitochondrial fragmentation, and Acod1 improved AMPK activity. The over-expression of Acod1 did not significantly affect the level of itaconic acid, but greatly decreased the levels of some important intermediate metabolites of the tricarboxylic acid (TCA) cycle. These data indicate that Acod1 exerts a pro-inflammatory role in MSU crystal-induced inflammation and is independent of the metabolic level of itaconic acid. Sirt3 deficiency exacerbates inflammatory response induced by MSU crystals in vitro and in vivo. CONCLUSION: The current study has shown that Sirt3 can alleviate the MSU crystal-induced inflammation by inhibiting the expression of Acod1.
Assuntos
Gota , Sirtuína 3 , Animais , Camundongos , Gota/induzido quimicamente , Gota/tratamento farmacológico , Gota/metabolismo , Inflamação/induzido quimicamente , Inflamação/metabolismo , Leucócitos Mononucleares/metabolismo , Camundongos Endogâmicos C57BL , Sirtuína 3/genética , Sirtuína 3/metabolismo , Ácido Úrico/toxicidadeRESUMO
AIMS: Polygonum cuspidatum Sieb. et Zucc is one of the commonly used herbs for the treatment of gouty arthritis, and polydatin is one of its main effective components. This study evaluated the therapeutic potential of polydatin for the treatment of gout. MAIN METHODS: The ankle joint of C57BL/6 mice were injected with MSU suspensions to simulate human gouty arthritis, and oral treatment with polydatin (25, 50, and 100 mg/kg body weight) was performed at 1 h after MSU crystal injection. The effect of polydatin on model mice was evaluated by measuring ankle swelling, gait, histopathological analysis, proinflammatory cytokine expression, as well as the contents of NO, MDA and GSH. The targets of polydatin were explored by Real-Time PCR and IHC. KEY FINDINGS: Treatment with polydatin inhibited ankle swelling, improved abnormal gait, and reduced ankle lesions dose-dependently. Moreover, polydatin decreased pro-inflammatory cytokine expression, and promoted expression of anti-inflammatory cytokine. In addition, polydatin inhibited MSU-induced oxidative stress by decreasing oxidative product (NO, MDA) generation and promote the antioxidant (GSH). Further, we found that polydatin reduced inflammation by decreasing the expression of NLRP3 inflammasome component via activating PPAR-γ. Moreover, polydatin can protect against iron overload and attenuate oxidative stress by promoting the activation of ferritin. SIGNIFICANCE: Our findings indicates that polydatin ameliorates MSU-induced inflammation and oxidative stress by regulating PPAR-γ and ferritin activation in gouty arthritis model mice, and this research result suggests that polydatin has therapeutic potential for the treatment of gout in humans through multiple targets.
Assuntos
Artrite Gotosa , Gota , Camundongos , Humanos , Animais , Artrite Gotosa/induzido quimicamente , Artrite Gotosa/tratamento farmacológico , Artrite Gotosa/metabolismo , PPAR gama/metabolismo , Ácido Úrico/toxicidade , Camundongos Endogâmicos C57BL , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Gota/tratamento farmacológico , Estresse Oxidativo , Inflamassomos/metabolismo , Citocinas/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismoRESUMO
Activation of the nod-like receptor protein 3 (NLRP3) inflammasome by monosodium urate (MSU) crystals has been identified as the molecular basis for the acute inflammatory response in gouty arthritis. However, MSU crystals alone are not sufficient to induce acute gouty arthritis (AGA). Adenosine triphosphate (ATP) is an endogenous signaling molecule involved in the NLRP3 inflammasome activation. We aimed to explore the role of ATP in MSU crystal-induced AGA development. In peripheral blood mononuclear cell-derived macrophages obtained from gout patients, we observed a synergistic effect of ATP on MSU crystal-induced IL-1ß release. Furthermore, in a rat model of spontaneous gout, we demonstrated that a synergistic effect of ATP and MSU crystals, but not MSU crystals alone, is essential for triggering AGA. Mechanistically, this synergistic effect is achieved through the purinergic receptor P2X7 (P2X7R). Blockade of P2X7R prevented AGA induction in rats after local injection of MSU crystals, and carrying the mutant hP2X7R gene contributed to the inhibition of NLRP3 inflammasome activation induced by costimulation of MSU crystals and ATP in vitro. Taken together, these results support the synergistic effect of ATP on MSU crystal-induced NLRP3 inflammasome activation facilitating inflammatory episodes in AGA. In this process, P2X7R plays a key regulatory role, suggesting targeting P2X7R to be an attractive therapeutic strategy for the treatment of AGA.
Assuntos
Artrite Gotosa , Gota , Ratos , Animais , Artrite Gotosa/induzido quimicamente , Artrite Gotosa/tratamento farmacológico , Artrite Gotosa/metabolismo , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Trifosfato de Adenosina , Leucócitos Mononucleares/metabolismo , Ácido Úrico/toxicidade , Ácido Úrico/química , Gota/metabolismoRESUMO
Bee venom (BV) acupuncture has anti-inflammatory and analgesic effects; therefore, it was used as a traditional Korean medicine for various musculoskeletal disorders, especially arthritis. In this study, we investigated the effect of BV on monosodium urate (MSU) crystal-induced acute gouty rats. An intra-articular injection of MSU crystal suspension (1.25 mg/site) was administered to the tibiotarsal joint of the hind paw of Sprague Dawley rats to induce MSU crystal-induced gouty arthritis. Colchicine (30 mg/kg) was orally administered 1 h before MSU crystal injection as a positive control, and BV (0.5 mg/kg) was injected into the tibiotarsal joint immediately after MSU crystal injection. The ankle thickness, mechanical allodynia, and expression of proinflammatory cytokines (TNF-α, IL-1ß, IL6, COX2 and iNOS) and chemokines (MIP-1α, MIP-1ß, MCP-1, GRO-α, MIP-2α) were then evaluated. BV reduced the expression of proinflammatory cytokines and chemokines, which are important mediators of MSU crystal-induced inflammatory responses. This anti-inflammatory effect was also confirmed histologically to attenuate synovitis and neutrophil infiltration. We demonstrated that BV markedly ameliorated ankle edema and mechanical allodynia in gouty rats. These results suggest that BV acupuncture is a potential clinical therapy for acute gouty management.
Assuntos
Artrite Gotosa/induzido quimicamente , Artrite Gotosa/tratamento farmacológico , Venenos de Abelha/uso terapêutico , Edema/tratamento farmacológico , Inflamação/tratamento farmacológico , Dor/tratamento farmacológico , Ácido Úrico/toxicidade , Animais , Modelos Animais de Doenças , Humanos , Masculino , Ratos , Ratos Sprague-Dawley , Resultado do TratamentoRESUMO
Mesenchymal stem cells (MSCs), due to their multi-directional differentiation, paracrine and immunomodulation potentials, and the capacity of homing to target organ, have been reported to facilitate regeneration and repair of kidney and improve kidney function in acute or chronic kidney injury. The present study was aimed to evaluate whether MSCs could have a protective effect in hyperuricemic nephropathy (HN) and the underlying mechanisms. A rat HN model was established by oral administration of a mixture of potassium oxonate (PO, 1.5 g/kg) and adenine (Ad, 50 mg/kg) daily for 4 weeks. For MSCs treatment, MSCs (3 × 106 cells/kg per week) were injected via tail vein from the 2nd week for 3 times. The results showed that along with the elevated uric acid (UA) in HN rats, creatinine (CREA), blood urea nitrogen (BUN), microalbuminuria (MAU) and 24-hour urinary protein levels were significantly increased comparing with the normal control rats, while decreased after MSCs treatment. Moreover, the mRNA levels of inflammation and fibrosis-related gene were reduced in UA + MSCs group. Consistently, hematoxylin-eosin (HE) staining results showed the destruction of kidney structure and fibrosis were significantly alleviated after MSCs administration. Similarly, in vitro, NRK-52Es cells were treated with high concentration UA (10 mg/dL) in the presence of MSCs, and we found that MSCs co-culture could inhibited UA-induced cell injury, characterized as improvement of cell viability and proliferation, inhibition of apoptosis, inflammation, and fibrosis. Collectively, MSCs treatment could effectively attenuate UA-induced renal injury, and thus it might be a potential therapy to hyperuricemia-related renal diseases.
Assuntos
Hiperuricemia/metabolismo , Células-Tronco Mesenquimais/metabolismo , Ácido Úrico/metabolismo , Animais , Proteínas Reguladoras de Apoptose/metabolismo , Técnicas de Cultura de Células , Sobrevivência Celular , Fibrose , Hiperuricemia/sangue , Hiperuricemia/induzido quimicamente , Inflamação , Rim/patologia , Masculino , Transplante de Células-Tronco Mesenquimais , Ratos , Ratos Sprague-Dawley , Ácido Úrico/sangue , Ácido Úrico/toxicidadeRESUMO
Using molecular dynamics, we address uric acid (UA) replacement by a model small-molecule inhibitor, allopurinol (AP), from its aggregated cluster in a columnar fashion. Experimentally it has been affirmed that AP is efficient in preventing UA-mediated renal stone formation. However, no study has presented the underlying mechanisms yet. Hence, a theoretical approach is presented for mapping the AP, which binds to melamine (MM) and UA clusters. In AP's presence, the higher-order cluster of UA molecules turns into a lower-order cluster, which "drags" fewer MM to them. Consequently, the MM-UA composite structure gets reduced. It is worth noting that UA-AP and AP-MM hydrogen-bonding interactions often play an essential role in reducing the UA-MM cluster size. Interestingly, an AP around UA makes a pillar-like structure, confirmed by defining the point-plane distribution function. The decomposition of the preferential interaction by Kirkwood-Buff integral into different angles like 0°-30°, 30°-60°, and 60°-90° firmly establishes the phenomenon mentioned above. However, the structural order for such π-stacking interactions between AP and UA molecules is not hierarchical but rather more spontaneous. The driving force behind UA-AP-MM composite formation is the favorable complexation energy that can be inferred by computing pairwise binding free energies for all possible combinations. Performing enhanced sampling and quantum calculations further confirms the evidence for UA degradation.
Assuntos
Alopurinol/química , Triazinas/química , Ácido Úrico/química , Ligação de Hidrogênio , Cálculos Renais/prevenção & controle , Simulação de Dinâmica Molecular , Eletricidade Estática , Termodinâmica , Triazinas/toxicidade , Ácido Úrico/toxicidadeRESUMO
Lagotis brachystachya Maxim is a herb widely used in traditional Tibetan medicine. Our previous study indicated that total extracts from Lagotis brachystachya could lower uric acid levels. This study aimed to further elucidate the active components (luteolin, luteoloside and apigenin) isolated from Lagotis brachystachya and the underlying mechanism in vitro and in vivo. The results showed that treatment with luteolin and luteoloside reversed the reduction of organic anion transporter 1 (OAT1) levels, while apigenin attenuated the elevation of urate transporter 1 (URAT1) and glucose transporter 9 (GLUT9) levels in uric acid-treated HK-2 cells, which was consistent with the finding in the kidneys of potassium oxonate (PO)-induced mice. On the other hand, hepatic xanthine oxidase activity was inhibited by the components. In addition, all of these active components improved the morphology of the kidney in hyperuricemic mice. Moreover, molecular docking showed that luteolin, luteoloside and apigenin could bind Toll-like receptor 4 (TLR4) and NLR family pyrin domain containing 3 (NLRP3). Congruently, western blot analysis showed that the components inhibited TLR4/myeloid differentiation primary response 88 (MyD88)/NLRP3 signaling. In conclusion, these results indicated that luteolin, luteoloside and apigenin could attenuate hyperuricemia by decreasing the production and increasing the excretion of uric acid, which were mediated by inhibiting inflammatory signaling pathways.
Assuntos
Medicamentos de Ervas Chinesas/farmacologia , Hiperuricemia/metabolismo , Rim/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Receptor 4 Toll-Like/metabolismo , Ácido Úrico/metabolismo , Animais , Relação Dose-Resposta a Droga , Medicamentos de Ervas Chinesas/isolamento & purificação , Medicamentos de Ervas Chinesas/uso terapêutico , Homeostase/efeitos dos fármacos , Homeostase/fisiologia , Hiperuricemia/tratamento farmacológico , Rim/efeitos dos fármacos , Masculino , Camundongos , Proteína 3 que Contém Domínio de Pirina da Família NLR/antagonistas & inibidores , Plantas Medicinais , Estrutura Secundária de Proteína , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Receptor 4 Toll-Like/antagonistas & inibidores , Ácido Úrico/toxicidadeRESUMO
BACKGROUND: Gout is initiated by the precipitation of monosodium urate (MSU) crystals within the joints and soft tissues, and it can eventually cause acute or chronic arthritis. MSU crystals trigger, amplify, and maintain a strong inflammatory response through promoting proinflammatory activity. In this study, the therapeutic effects of Stephania hainanensis (S. hainanensis) total alkaloid (SHA) were tested and evaluated on MSU-induced acute gouty arthritis in a mouse model. METHODS: After oral administration of SHA (10 or 20 mg/kg) or the antigout medicine colchicine (0.5 mg/kg) once daily for 3 consecutive days, MSU crystals suspended in saline (2.5 mg/50 µl) were intradermally injected into the right paw of the mice. Then, SHA and colchicine were administered for another 2 days. During this period, swelling of the ankle and clinical scores were measured at 12, 24, and 48 h postinjection. After the mice were euthanized, inflammatory cytokine expression and paw tissue inflammation-related gene and protein expression, and a histopathological analysis was performed. RESULTS: SHA had obvious therapeutic effects on MSU-induced acute gouty arthritis in mice. SHA alleviated ankle swelling and inhibited the production of cytokines, such as IL-1ß and TNF-α. In addition, NLRP3, Caspase-1 and IL-1ß, which are activated by MSU were also suppressed by SHA. The histological evaluation showed that SHA relieved the infiltration of inflammation around the ankle. CONCLUSIONS: These results suggest that SHA is capable of anti-inflammatory activities and may be useful for treating gouty arthritis.
Assuntos
Alcaloides/farmacologia , Anti-Inflamatórios/farmacologia , Antioxidantes/toxicidade , Artrite Gotosa/induzido quimicamente , Stephania/metabolismo , Ácido Úrico/toxicidade , Animais , Antioxidantes/farmacologia , CamundongosRESUMO
Tanshinones, the active ingredients derived from the roots of Salvia miltiorrhiza, have been widely used as traditional medicinal herbs for treating human diseases. Although tanshinones showed anti-inflammatory effects in many studies, large knowledge gaps remain regarding their underlying mechanisms. Here, we identified 15 tanshinones that suppressed the activation of NLRP3 inflammasome and studied their structure-activity relationships. Three tanshinones (tanshinone IIA, isocryptotanshinone, and dihydrotanshinone I) reduced mitochondrial reactive-oxygen species production in lipopolysaccharide (LPS)/nigericin-stimulated macrophages and correlated with altered mitochondrial membrane potentials, mitochondria complexes activities, and adenosine triphosphate and protonated-nicotinamide adenine dinucleotide production. The tanshinones may confer mitochondrial protection by promoting autophagy and the AMP-activated protein kinase pathway. Importantly, our findings demonstrate that dihydrotanshinone I improved the survival of mice with LPS shock and ameliorated inflammatory responses in septic and gouty animals. Our results suggest a potential pharmacological mechanism whereby tanshinones can effectively treat inflammatory diseases, such as septic and gouty inflammation.
Assuntos
Abietanos/farmacologia , Furanos/farmacologia , Gota/tratamento farmacológico , Inflamassomos/antagonistas & inibidores , Fenantrenos/farmacologia , Quinonas/farmacologia , Choque Séptico/tratamento farmacológico , Proteínas Quinases Ativadas por AMP/metabolismo , Abietanos/uso terapêutico , Animais , Autofagia/efeitos dos fármacos , Autofagia/imunologia , Modelos Animais de Doenças , Feminino , Furanos/uso terapêutico , Gota/induzido quimicamente , Gota/imunologia , Gota/patologia , Humanos , Inflamassomos/metabolismo , Inflamação/tratamento farmacológico , Inflamação/imunologia , Inflamação/patologia , Masculino , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/patologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/antagonistas & inibidores , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Fenantrenos/uso terapêutico , Quinonas/uso terapêutico , Ratos , Espécies Reativas de Oxigênio/metabolismo , Choque Séptico/imunologia , Choque Séptico/patologia , Ácido Úrico/administração & dosagem , Ácido Úrico/toxicidadeRESUMO
High uric acid (HUA) is associated with insulin resistance (IR) in cardiomyocytes. We investigated whether metformin protects against HUA-induced IR in cardiomyocytes. We exposed primary cardiomyocytes to HUA, and cellular glucose uptake was quantified by measuring the uptake of 2-NBDG, a fluorescent glucose analog. Western blot was used to examine the levels of signalling protein. Membrane of glucose transporter type 4 (GLUT4) was analysed by immunofluorescence. We monitored the impact of metformin on HUA-induced IR and in myocardial tissue of an acute hyperuricaemia mouse model established by potassium oxonate treatment. Treatment with metformin protected against HUA-reduced glucose uptake induced by insulin in cardiomyocytes. HUA directly inhibited the phosphorylation of Akt and the translocation of GLUT4 induced by insulin, which was blocked by metformin. Metformin promoted phosphorylation of AMP-activated protein kinase (AMPK) and restored the insulin-stimulated glucose uptake in HUA-induced IR cardiomyocytes. As a result of these effects, in a mouse model of acute hyperuricaemia, metformin improved insulin tolerance and glucose tolerance, accompanied by increased AMPK phosphorylation, Akt phosphorylation and translocation of GLUT4 in myocardial tissues. As expected, AICAR, another AMPK activator, had similar effects to metformin, demonstrating the important role of AMPK activation in protecting against IR induced by HUA in cardiomyocytes. Metformin protects against IR induced by HUA in cardiomyocytes and improves insulin tolerance and glucose tolerance in an acute hyperuricaemic mouse model, along with the activation of AMPK. Consequently, metformin may be an important potential new treatment strategy for hyperuricaemia-related cardiovascular disease.
Assuntos
Quinases Proteína-Quinases Ativadas por AMP/metabolismo , Hiperuricemia/tratamento farmacológico , Hipoglicemiantes/farmacologia , Resistência à Insulina , Metformina/farmacologia , Miócitos Cardíacos/efeitos dos fármacos , Animais , Células Cultivadas , Glucose/metabolismo , Transportador de Glucose Tipo 4/metabolismo , Hiperuricemia/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Miócitos Cardíacos/metabolismo , Transdução de Sinais , Ácido Úrico/toxicidadeRESUMO
The aim of this study was to identify the effects of melatonin on acute gouty inflammation and to investigate the underlying mechanisms. We found significantly lower serum melatonin levels in gout patients in the acute phase than in those in the remission phase or in normal individuals. The mRNA expression of melatonin receptor 2 (MT2) was also lower in gout patients than in normal individuals. To verify the in-vivo role of melatonin, a gouty arthritis model was established by intraarticular injection of monosodium urate (MSU, 1 mg) crystals into the paws of C57BL/6 mice. Joint inflammation in the mouse model was evaluated by measuring the thickness of the right paw/left paw, and the inflammation index was determined by examining infiltrating neutrophils with haematoxylin and eosin (H&E) staining. Melatonin was found to reduce both paw thickness and the inflammation index in the mouse model, and melatonin also reduced the mRNA levels of interleukin-1 beta (IL-1ß), IL-6 and NLR family pyrin domain containing 3 (NLRP3) inflammasome. To mimic gouty inflammation in vitro, mouse peritoneal macrophages were stimulated with lipopolysaccharides (LPS) plus MSU. Melatonin was revealed to reduce IL-1ß secretion by stimulated macrophages. The mRNA expression levels of IL-1ß and IL-6 were also inhibited by melatonin. Western blot analysis showed that the expression of NLRP3, caspase-1 and pro-IL-1ß was also inhibited by melatonin. In conclusion, our study demonstrated that melatonin alleviated gouty inflammation in vivo and in vitro, and the underlying mechanism may involve inhibiting the assembly of the NLRP3 inflammasome.
Assuntos
Artrite Gotosa/tratamento farmacológico , Gota/tratamento farmacológico , Inflamação/tratamento farmacológico , Melatonina/farmacologia , Receptor MT2 de Melatonina/sangue , Doença Aguda/epidemiologia , Animais , Artrite Gotosa/sangue , Artrite Gotosa/induzido quimicamente , Artrite Gotosa/genética , Modelos Animais de Doenças , Gota/metabolismo , Gota/patologia , Humanos , Inflamação/sangue , Inflamação/induzido quimicamente , Inflamação/genética , Interleucina-1beta/genética , Interleucina-6/genética , Articulações/efeitos dos fármacos , Articulações/patologia , Macrófagos/efeitos dos fármacos , Macrófagos/patologia , Melatonina/sangue , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , RNA Mensageiro/sangue , Ácido Úrico/toxicidadeRESUMO
This work aimed to investigate the effect of fucoidan (FPS) on urate transporters induced by uric acid (UA). The results showed that UA stimulated the expression of glucose transporter 9 (GLUT9) and urate transporter 1 (URAT1) in HK-2 cells, and FPS could reverse the effect. Moreover, UA could activate NF-κB, JNK and PI3K/Akt pathways, but both pathway inhibitors and FPS inhibited the UA-induced activation of these three pathways. These data suggested that FPS effectively inhibited the expression induction of reabsorption transporters URAT1 and GLUT9 by UA, through repressing the activation of NF-κB, JNK and PI3K/Akt signal pathways in HK-2 cells. The in vitro research findings support the in vivo results that FPS reduces serum uric acid content in hyperuricemia mice and rats through inhibiting the expression of URAT1 and GLUT9 in renal tubular epithelial cells. This study provides a theoretical basis for the application of FPS in the treatment of hyperuricemia.
Assuntos
Proteínas Facilitadoras de Transporte de Glucose/metabolismo , Supressores da Gota/farmacologia , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Túbulos Renais Proximais/efeitos dos fármacos , Laminaria , NF-kappa B/metabolismo , Transportadores de Ânions Orgânicos/metabolismo , Proteínas de Transporte de Cátions Orgânicos/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Polissacarídeos/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Linhagem Celular , Supressores da Gota/isolamento & purificação , Humanos , Túbulos Renais Proximais/enzimologia , Laminaria/química , Polissacarídeos/isolamento & purificação , Transdução de Sinais , Ácido Úrico/toxicidadeRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Simiao Wan (SMW) is a classical traditional Chinese medicine (TCM) prescription to empirically treat gouty arthritis (GA) in TCM clinical practice. However, the potential mechanisms of SMW on GA are not fully evaluated. AIM OF STUDY: The aim of this study is to investigate the role of macrophage polarization in the anti-GA activity of SMW. MATERIALS AND METHODS: Rats were intragastricly treated with SMW for consecutive 7 days. On day 6, monosodium urate (MSU) crystal-induced arthritis (MIA) in the ankle joint was prepared. Paw volume, gait score and histological score were measured. Levels of interleukin (IL)-1ß and IL-10 in serum were detected by enzyme-linked immunosorbent assay. Expressions of inducible nitric oxide synthase (iNOS), arginase (Arg)-1, phosphorylated (p)-p65, inhibitor of nuclear factor (NF)-κB (IκB)α, p-signal transducer and transcription activator (STAT)3 and p-Janus kinase (JAK)2 in synovial tissues were determined by Western blot. RESULTS: The elevated paw volume, gait score and histological score in MIA rats were significantly decreased by SMW treatment. Meanwhile, SMW significantly decreased the IL-1ß level and increased the IL-10 level in serum of MIA rats. Furthermore, SMW reduced the expressions of iNOS, p-p65 and enhanced the expressions of Arg-1, IκBα, p-STAT3 and p-JAK2 in synovial tissues of MIA rats. CONCLUSIONS: The results suggest that SMW attenuates the inflammation in MIA rats through promoting macrophage M2 polarization.
Assuntos
Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Antirreumáticos/farmacologia , Antirreumáticos/uso terapêutico , Artrite Experimental/prevenção & controle , Medicamentos de Ervas Chinesas/farmacologia , Medicamentos de Ervas Chinesas/uso terapêutico , Macrófagos/efeitos dos fármacos , Animais , Anti-Inflamatórios/química , Antirreumáticos/química , Arginase/metabolismo , Artrite Experimental/patologia , Medicamentos de Ervas Chinesas/química , Edema/induzido quimicamente , Edema/tratamento farmacológico , Edema/patologia , Extremidades/patologia , Marcha/efeitos dos fármacos , Proteínas I-kappa B/metabolismo , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Inflamação/patologia , Interleucina-10/metabolismo , Interleucina-1beta/metabolismo , Janus Quinase 2/metabolismo , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Proteínas de Neoplasias/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Proteínas de Transporte Nucleocitoplasmático/metabolismo , Ratos Sprague-Dawley , Fator de Transcrição STAT3/metabolismo , Membrana Sinovial/efeitos dos fármacos , Membrana Sinovial/metabolismo , Ácido Úrico/toxicidadeRESUMO
BACKGROUND: Monosodium urate (MSU)-mediated inflammatory response is a crucial inducing factor in gouty arthritis. Here, we explored the underlying mechanism of total glucosides of paeony (TGP) in MSU-induced inflammation of THP-1 macrophages in gouty arthritis. METHODS: 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to detect cell viability. Enzyme-linked immunosorbent assay (ELISA) was utilized to measure the production of interleukin 1ß (IL-1ß) and tumor necrosis factor α (TNF-α). Real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot assay were conducted to determine RNA and protein expression. Dual-luciferase reporter assay, RNA immunoprecipitation (RIP) assay and RNA pull down assay were used to confirm the interaction between miR-876-5p and MALAT1 or NLR family pyrin domain containing 3 (NLRP3). RESULTS: MSU-induced damage and inflammatory response in THP-1 macrophages were alleviated by the treatment of TGP in a dose-dependent manner. Overexpression of NLRP3 or MALAT1 reversed the protective effects of TGP in MSU-induced THP-1 macrophages. The binding relation between miR-876-5p and MALAT1 or NLRP3 was identified in THP-1 macrophages. MALAT1 up-regulated the expression of NLRP3 by sponging miR-876-5p in THP-1 macrophages. TGP suppressed MSU-induced inflammation in THP-1 macrophages through regulating MALAT1/miR-876-5p/NLRP3 axis. TGP suppressed MSU-induced activation of TLR4/MyD88/NF-κB pathway through regulating MALAT1/miR-876-5p/NLRP3 axis. CONCLUSION: In conclusion, TGP suppressed MSU-induced inflammation in THP-1 macrophages through regulating MALAT1/miR-876-5p/NLRP3 axis and TLR4/MyD88/NF-κB pathway, suggesting that TGP was a promising active ingredient for gouty arthritis treatment.
Assuntos
Artrite Gotosa/metabolismo , Glucosídeos/uso terapêutico , MicroRNAs/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Paeonia , RNA Longo não Codificante/metabolismo , Ácido Úrico/toxicidade , Artrite Gotosa/induzido quimicamente , Artrite Gotosa/prevenção & controle , Glucosídeos/isolamento & purificação , Glucosídeos/farmacologia , Humanos , Mediadores da Inflamação/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêutico , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Células THP-1/efeitos dos fármacos , Células THP-1/metabolismoRESUMO
The aim of the present study was to observe the changes of TTX-R, Nav1.8, and Nav1.9 Na+ currents in MSU-induced gouty arthritis mice, and to explore the possibility of Nav1.8 and Nav1.9 channels as potential targets for gout pain treatment. Acute gouty arthritis was induced by monosodium urate (MSU) in mice. Swelling degree was evaluated by measuring the circumference of the ankle joint. Mechanical allodynia was assessed by applying the electronic von Frey. Na+ currents were recorded by patch-clamp techniques in acute isolated dorsal root ganglion (DRG) neurons. MSU treatment significantly increased the swelling degree of ankle joint and decreased the mechanical pain threshold. The amplitude of TTX-R Na+ current was significantly increased and reached its peak on the 4th day after injection of MSU. For TTX-R Na+ channel subunits, Nav1.8 current density was significantly increased, but Nav1.9 current density was markedly decreased after MSU treatment. MSU treatment shifted the steady-state activation curves of TTX-R Na+ channel, Nav1.8 and Nav1.9 channels, and the inactivation curves of TTX-R Na+ channel and Nav1.8 channels to the depolarizing direction, but did not affect the inactivation curve of Nav1.9 channel. Compared with the normal group, the recovery of Nav1.8 channel was faster, while that of Nav1.9 channel was slower. The recovery of TTX-R Na+ channel remained unchanged after MSU treatment. Additionally, MSU treatment increased DRG neurons excitability by reducing action potential threshold. Nav1.8 channel, not Nav1.9 channel, may be involved in MSU-induced gout pain by increasing nerve excitability.
Assuntos
Artrite Gotosa/induzido quimicamente , Artrite Gotosa/metabolismo , Canal de Sódio Disparado por Voltagem NAV1.8/metabolismo , Canal de Sódio Disparado por Voltagem NAV1.9/metabolismo , Ácido Úrico/toxicidade , Animais , Artrite Gotosa/patologia , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/metabolismo , Gânglios Espinais/patologia , Hiperalgesia/induzido quimicamente , Hiperalgesia/metabolismo , Hiperalgesia/patologia , Masculino , Camundongos , Camundongos Endogâmicos ICR , Bloqueadores dos Canais de Sódio/farmacologiaRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Modified Simiaowan (MSW) is a traditional Chinese medicine formula that is composed of six herbs. It has been widely used in the treatment of gouty arthritis. AIM OF THE STUDY: This study was designed to investigate the effect of MSW on gouty arthritis and explore the possible mechanisms. MATERIAL AND METHODS: The rat gouty arthritis model was established by intra-articular injection of Monosodium Urate (MSU) crystal, and then treated with MSW for 5 days. The perimeter of the knee joints was measured in a time-dependent manner and serum samples were collected for the detection of TNF-α, IL-1ß, and IL-6 protein levels by ELISA. The protein expressions of MMP-3, TIMP-3, STAT3, and p-STAT3 in cartilage tissues and C28/I2 cells were detected by Western blot, and the levels of proteoglycan in primary chondrocytes and cartilage tissues were determined by toluidine blue staining. In addition, AG490 and IL-6 were used in vitro to explore the function of IL-6/STAT3 pathway in the protective effect of MSU. RESULTS: MSW reduced the joint swelling rate in gouty arthritis model and inhibited MSU induced up-regulation of IL-1ß, TNF-α, and IL-6 protein levels in serum and synovial fluid. IL-1ß induced an increase in p-STAT3 and MMP-3 protein expression in C28/I2 cells, as well as a decrease in TIMP-3. MSW serum inhibited the protein expression changes induced by IL-1ß in vitro. Furthermore, inhibition of STAT3 signaling negated the effect of MSW serum on p-STAT3, MMP-3, and TIMP-3 protein levels in C28/I2 cells. MSW also increased the content of proteoglycan significantly both in vivo and in vitro. CONCLUSION: Our data indicated that MSW protected rats from MSU-induced experimental gouty arthritis and IL-1ß/IL-6/STAT3 pathway played an essential role in the protective effect of MSU against GA.
Assuntos
Artrite Gotosa/tratamento farmacológico , Medicamentos de Ervas Chinesas/farmacologia , Medicamentos de Ervas Chinesas/uso terapêutico , Substâncias Protetoras/farmacologia , Substâncias Protetoras/uso terapêutico , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Artrite Gotosa/induzido quimicamente , Linhagem Celular , Condrócitos/efeitos dos fármacos , Citocinas/genética , Citocinas/metabolismo , Modelos Animais de Doenças , Edema/induzido quimicamente , Edema/tratamento farmacológico , Humanos , Interleucina-1beta/toxicidade , Masculino , Proteoglicanas/efeitos dos fármacos , Coelhos , Ratos Sprague-Dawley , Ácido Úrico/toxicidadeRESUMO
BACKGROUND: Celastrol, a pentacyclic triterpenoid quinonemethide isolated from several spp. of Celastraceae family, exhibits anti-inflammatory activities in a variety of diseases including arthritis. PURPOSE: This study aims to investigate whether the inhibition of NLRP3 inflammasome is engaged in the anti-inflammatory activities of celastrol and delineate the underlying mechanism. METHODS: The influence of celastrol on NLRP3 inflammasome activation was firstly studied in lipopolysaccharide (LPS)-primed mouse bone marrow-derived macrophages (BMDMs) and phorbol 12-myristate 13-acetate (PMA)-primed THP-1 cells treated with nigericin. Reconstituted inflammasome was also established by co-transfecting NLRP3, ASC, pro-caspase-1 and pro-IL-1ß in HEK293T cells. The changes of inflammasome components including NLRP3, ASC, pro-caspase-1/caspase-1 and pro-IL-1ß/IL-1ß were examined by enzyme-linked immunosorbent assay (ELISA), western blotting and immunofluorescence. Furthermore, Propionibacterium acnes (P. acnes)/LPS-induced liver injury and monosodium urate (MSU)-induced gouty arthritis in mice were employed in vivo to validate the inhibitory effect of celastrol on NLRP3 inflammasome. RESULTS: Celastrol significantly suppressed the cleavage of pro-caspase-1 and pro-IL-1ß, while not affecting the protein expressions of NLRP3, ASC, pro-caspase-1 and pro-IL-1ß in THP-1 cells, BMDMs and HEK293T cells. Celastrol suppressed NLRP3 inflammasome activation and alleviated P. acnes/LPS-induced liver damage and MSU-induced gouty arthritis. Mechanism study revealed that celastrol could interdict K63 deubiquitination of NLRP3, which may concern interaction of celastrol and BRCA1/BRCA2-containing complex subunit 3 (BRCC3), and thereby prohibited the formation of NLRP3, ASC and pro-caspase-1 complex to block the generation of mature IL-1ß. CONCLUSION: Celastrol suppresses NLRP3 inflammasome activation in P. acnes/LPS-induced liver damage and MSU-induced gouty arthritis via inhibiting K63 deubiquitination of NLRP3, which presents a novel insight into inhibition of celastrol on NLRP3 inflammasome and provides more evidences for its application in the therapy of inflammation-related diseases.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Artrite Gotosa/tratamento farmacológico , Fígado/efeitos dos fármacos , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Triterpenos/farmacologia , Animais , Artrite Gotosa/induzido quimicamente , Artrite Gotosa/metabolismo , Células HEK293 , Humanos , Inflamassomos/efeitos dos fármacos , Inflamassomos/metabolismo , Lipopolissacarídeos/toxicidade , Fígado/microbiologia , Fígado/patologia , Lisina/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Triterpenos Pentacíclicos , Propionibacterium acnes/patogenicidade , Células THP-1 , Ubiquitinação/efeitos dos fármacos , Ácido Úrico/toxicidadeRESUMO
Icariin (ICA) has anti-inflammatory effects in some diseases, but its role in gouty arthritis (GA) is not clear. This study investigated the effects of ICA in monosodium urate (MSU)-induced GA rat models. GA rat models were induced by MSU, and co-treated with ICA of low-dose (20 mg/kg), medium-dose (40 mg/kg), and high-dose (80 mg/kg), respectively. The ankle swelling rates, haematoxylin-eosin (HE) staining changes, inflammatory factors (interleukin (IL)-1ß, IL-6, tumour necrosis factor-α (TNF-α)) and prostaglandin E2 (PGE2 ) levels in synovial tissues were detected. The antioxidants levels in rat serum, and NF-κB pathway-related proteins and NALP3 inflammasome expressions in synovial tissues were also analysed. In cell experiments, chondrocytes were co-treated with different concentrations of ICA (1, 5, 10 µmol/L) on the basis of MSU. The activities and inflammatory cytokines, hydroxyproline (Hyp) and glycosaminogly (GAG) expressions in chondrocytes were measured. In rat experiments, MUS increased the ankle swelling rates, promoted inflammatory cells infiltration, and increased IL-1ß, IL-6, TNF-α, PGE2 levels in synovial tissues, which were all alleviated by ICA. Moreover, ICA also suppressed nuclear translocation of NF-κB pathway-related proteins and reduced the expression of NALP3 inflammasome in rat models. As for cell experiments, ICA decreased the activity, inflammatory cytokines and GAG levels, and suppressed nuclear translocation of NF-κB pathway-related proteins of MSU-treated chondrocytes. In general, medium and high concentrations of ICA showed good effects. ICA has an inhibitory effect in MSU-induced rat GA models through NF-κB/NALP3 pathway, which may provide a direction for the treatment of GA. SIGNIFICANCE: Icariin (ICA) has anti-inflammatory effects in some diseases, but its role in gouty arthritis (GA) is not clear. This study excogitated that monosodium urate (MSU) increased the ankle swelling rates of rats, promoted inflammatory cells infiltration, and increased cytokines levels in synovial tissues, which were all alleviated by ICA. In related mechanism, we found that ICA might exert the catabatic functions through the NF-κB/NALP3 pathway. The findings of this study clarified that ICA may provide a direction for the treatment of patients with GA and illustrated the relevant underlying mechanism of its role.
Assuntos
Artrite Gotosa , Flavonoides/farmacologia , NF-kappa B/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Transdução de Sinais/efeitos dos fármacos , Ácido Úrico/toxicidade , Animais , Artrite Gotosa/induzido quimicamente , Artrite Gotosa/metabolismo , Artrite Gotosa/patologia , Artrite Gotosa/prevenção & controle , Citocinas/metabolismo , Feminino , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Hyperuricaemia is a disorder of purine metabolism. Elevated serum uric acid is strongly associated with many diseases, including gout, abdominal obesity, insulin resistance, and cardiovascular and kidney disease. Our previous studies showed that high uric acid (HUA) induced insulin resistance in several peripheral organs, including the liver, myocardium and adipose tissue. However, whether HUA directly induces insulin resistance of pancreatic ß cells, the only source of insulin in the body and also a sensitive insulin target, is unknown. In this study, pancreatic ß cells pretreated with HUA showed impaired insulin expression/secretion, glucose uptake and the glycolytic pathway. RNA-seq revealed that HUA affected the biological processes of INS-1 cells broadly, including oxidoreduction coenzyme metabolic process, pyruvate metabolic process, and glycolytic process. In addition, HUA reduced mitochondrial membrane potential and increased the production of reactive oxygen species(ROS) in INS-1 cells. INS-1 cells pretreated with probenecid, an organic anion transporter inhibitor, protected INS-1 cells against HUA-induced insulin secretion decrease, Pretreatment with N-acetyl-L-cysteine(NAC), a globally used antioxidant, recovered HUA-decreased insulin secretion and glucose uptake by pancreatic ß cells. Insulin-like growth factor 1 (IGF-1), the phosphatidylinositol 3-kinase (PI3K) activator, rescues HUA-decreased insulin secretion by re-activating AKT phosphorylation. Thus, HUA induce insulin resistance, impaired insulin secretion and glycolytic pathway of pancreatic êµ cell through IRS2/AKT pathway.