Objective To investigate the relationship between interleukin-1β (IL-1β) and miR-185-5p in the process of joint injury in acute gouty arthritis (AGA). Methods The serum miR-185-5p levels of 89 AGA patients and 91 healthy volunteers were detected by real-time quantitative PCR. The correlation between miR-185-5p expression level and VAS score or IL-1β expression level was evaluated by Pearson correlation coefficient method. Receiver operating characteristic (ROC) curve was used to evaluate the diagnostic value of miR-185-5p in AGA. THP-1 cells were induced by sodium urate (MSU) to construct an in vitro acute gouty inflammatory cell model. After the expression level of miR-185-5p in THP-1 cells was upregulated or downregulated by transfection of miR-185-5p mimics or inhibitors in vitro, inflammatory cytokines of THP-1 cells, such as IL-1β, IL-8 and tumor necrosis factor α (TNF-α), were detected by ELISA. The luciferase reporter gene assay was used to determine the interaction between miR-185-5p and the 3'-UTR of IL-1β. Results Compared with the healthy control group, the expression level of serum miR-185-5p in AGA patients was significantly reduced. The level of serum miR-185-5p was negatively correlated with VAS score and IL-1β expression level. The area under the curve (AUC) was 0.905, the sensitivity was 80.17% and the specificity was 83.52%. Down-regulation of miR-185-5p significantly promoted the expression of IL-1β, IL-8 and tumor necrosis factor (TNF-α), while overexpression of miR-185-5p showed the opposite results. Luciferase reporter gene assay showed that IL-1β was the target gene of miR-185-5p, and miR-185-5p negatively regulated the expression of IL-1β. Conclusion miR-185-5p alleviates the inflammatory response in AGA by inhibiting IL-1β.
Humans ; 3' Untranslated Regions ; Arthritis, Gouty/genetics* ; Interleukin-1beta/genetics* ; Interleukin-8 ; Luciferases ; MicroRNAs/genetics* ; Tumor Necrosis Factor-alpha

OBJECTIVE: To investigate the effects of Danmu Extract Syrup (DMS) on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in mice and explore the mechanism. METHODS: Seventy-two male Balb/C mice were randomly divided into 6 groups according to a random number table (n=12), including control (normal saline), LPS (5 mg/kg), LPS+DMS 2.5 mL/kg, LPS+DMS 5 mL/kg, LPS+DMS 10 mL/kg, and LPS+Dexamethasone (DXM, 5 mg/kg) groups. After pretreatment with DMS and DXM, the ALI mice model was induced by LPS, and the bronchoalveolar lavage fluid (BALF) were collected to determine protein concentration, cell counts and inflammatory cytokines. The lung tissues of mice were stained with hematoxylin-eosin, and the wet/dry weight ratio (W/D) of lung tissue was calculated. The levels of tumor necrosis factor-α (TNF-α), interleukin (IL)-6 and IL-1 β in BALF of mice were detected by enzyme linked immunosorbent assay. The expression levels of Claudin-5, vascular endothelial (VE)-cadherin, vascular endothelial growth factor (VEGF), phospho-protein kinase B (p-Akt) and Akt were detected by Western blot analysis. RESULTS: DMS pre-treatment significantly ameliorated lung histopathological changes. Compared with the LPS group, the W/D ratio and protein contents in BALF were obviously reduced after DMS pretreatment (P<0.05 or P<0.01). The number of cells in BALF and myeloperoxidase (MPO) activity decreased significantly after DMS pretreatment (P<0.05 or P<0.01). DMS pre-treatment decreased the levels of TNF-α, IL-6 and IL-1 β (P<0.01). Meanwhile, DMS activated the phosphoinositide 3-kinase/protein kinase B (PI3K/Akt) pathway and reversed the expressions of Claudin-5, VE-cadherin and VEGF (P<0.01). CONCLUSIONS DMS attenuated LPS-induced ALI in mice through repairing endothelial barrier. It might be a potential therapeutic drug for LPS-induced lung injury.
Mice ; Male ; Animals ; Proto-Oncogene Proteins c-akt/metabolism* ; Lipopolysaccharides ; Phosphatidylinositol 3-Kinases/metabolism* ; Interleukin-1beta/metabolism* ; Vascular Endothelial Growth Factor A/metabolism* ; Tumor Necrosis Factor-alpha/metabolism* ; Claudin-5/metabolism* ; Acute Lung Injury/chemically induced* ; Lung/pathology* ; Interleukin-6/metabolism* ; Drugs, Chinese Herbal

Neuropathic pain(NP) has similar phenotypes but different sequential neuroinflammatory mechanisms in the pathological process. It is of great significance to inhibit the initiation of neuroinflammation, which has become a new direction of NP treatment and drug development in recent years. Mongolian drug Naru-3 is clinically effective in the treatment of trigeminal neuralgia, sciatica, and other NPs in a short time, but its pharmacodynamic characteristics and mechanism of analgesia are still unclear. In this study, a spinal nerve ligation(SNL) model simulating clinical peripheral nerve injury was established and the efficacy and mechanism of Naru-3 in the treatment of NPs was discussed by means of behavioral detection, side effect evaluation, network analysis, and experimental verification. Pharmacodynamic results showed that Naru-3 increased the basic pain sensitivity threshold(mechanical hyperalgesia and thermal radiation hyperalgesia) in the initiation of SNL in animals and relieved spontaneous pain, however, there was no significant effect on the basic pain sensitivity threshold and motor coordination function of normal animals under physiological and pathological conditions. Meanwhile, the results of primary screening of target tissues showed that Naru-3 inhibited the second phase of injury-induced nociceptive response of formalin test in mice and reduced the expression of inflammatory factors in the spinal cord. Network analysis discovered that Naru-3 had synergy in the treatment of NP, and its mechanism was associated with core targets such as matrix metalloproteinase-9(MMP9) and interleukin-1β(IL-1β). The experiment further took the dorsal root ganglion(DRG) and the stage of patho-logical spinal cord as the research objects, focusing on the core targets of inducing microglial neuroinflammation. By means of Western blot, immunofluorescence, agonists, antagonists, behavior, etc., the mechanism of Naru-3 in exerting NP analgesia may be related to the negative regulation of the MMP9/IL-1β signaling pathway-mediated microglia p38/IL-1β inflammatory loop in the activation phase. The relevant research enriches the biological connotation of Naru-3 in the treatment of NP and provides references for clinical rational drug use.
Rats ; Mice ; Animals ; Matrix Metalloproteinase 9/metabolism* ; Rats, Sprague-Dawley ; Neuroinflammatory Diseases ; Interleukin-1beta/metabolism* ; Spinal Cord/metabolism* ; Signal Transduction ; Hyperalgesia/metabolism* ; Neuralgia/metabolism*

The aim of this study is to investigate the effects of Gynostemma pentaphyllum dried with two different methods(air drying and heating) on inflammation in acute lung injury(ALI) mice in vivo and in vitro. Lipopolysaccharide(LPS) was sprayed into the airway of wild type C57BL/6J male mice to establish the model, and the drug was injected into the tail vein 24 h after modeling. Lung function, lung tissue wet/dry weight(W/D) ratio, the total protein concentration, interleukin 6(IL-6), IL-1β, and tumor necrosis factor-α(TNF-α) in the bronchoalveolar lavage fluid(BALF), and pathological changes of the lung tissue were used to evaluate the effects of different gypenosides on ALI mice. The results showed that total gypenosides(YGGPs) and the gypenosides substituted with one or two glycosyl(GPs_(1-2)) in the air-dried sample improved the lung function, significantly lowered the levels of IL-1β and TNF-α in BALF, and alleviated the lung inflammation of ALI mice. Moreover, GPs_(1-2) had a more significant effect on inhibiting NO release in RAW264.7 cells. This study showed that different drying methods affected the anti-inflammatory activity of G. pentaphyllum, and the rare saponins in the air-dried sample without heating had better anti-inflammatory activity.
Male ; Mice ; Animals ; Tumor Necrosis Factor-alpha/metabolism* ; Gynostemma ; Mice, Inbred C57BL ; Lung ; Anti-Inflammatory Agents/metabolism* ; Interleukin-6/metabolism* ; Interleukin-1beta/metabolism* ; Lipopolysaccharides/pharmacology*

OBJECTIVE: To investigate whether Salvianolic acid A (SAA) can restore cartilage endplate cell degeneration of intervertebral discs and to identify the mechanism via regulation of micro-RNA. METHODS: Cartilage endplate cells were isolated from lumbar intervertebral disc surgical samples and were treated with serum containing a series of concentrations of SAA (2, 5, and 10 ?M) for 24, 48, and 72 h to identify a proper dose and treatment time of SAA. The effect SAA on interlenkin-1β (IL-1β)-induced extracellular matrix degradation of cartilage endplate cells were analyzed by Alcian blue staining and assessment of the expression levels of ADAMTS-5, MMP3 and Col2a1. Further, the potential target miRNAs were preliminarily screened by micro-RNA sequencing combining qRT-PCR and Western blot, and then, the miRNAs mimics and inhibitors were used to verify the regulatory effect of SAA on potential target miRNAs. RESULTS: The 10 μM SAA treatment for 48 h significantly enhanced the viability of cartilage endplate cells, and increased Col2a1 expression and glycosaminoglycan accumulation that were repressed by IL-1β, and reduced the effect of IL-1β on ADAMTS-5, and MMP3. Screening analysis based on micro-RNA sequencing and Venny analysis identified the downstream micro-RNAs, including miR-940 and miR-576-5p. Then, the miR-940-mimic or miR-576-5p-mimic were transfected into CEPCs. Compared with the SAA group, the expression of ADAMTS-5 and MMP3 increased significantly and the expression of COL2A1 obviously decreased after overexpression of miR-940 or miR-576-5p in CEPCs. CONCLUSION Salvianolic acid A attenuated the IL-1β-induced extracellular matrix degradation of cartilage endplate cells by targeting regulate the miR-940 and the miR-576-5p.
Humans ; Apoptosis ; Cartilage/metabolism* ; Chondrocytes/metabolism* ; Interleukin-1beta/metabolism* ; Matrix Metalloproteinase 3/metabolism* ; MicroRNAs/metabolism*

OBJECTIVE: To explore and verify that transient receptor potential vanilloid 4(TRPV4) affects chondrocyte degeneration. METHODS: Neonatal SD rats were selected, primary chondrocytes were extracted, and identified by toluidine blue staining and alcian blue staining;an in vitro chondrocyte inflammation model was constructed by IL-1β, and TRPV4 inhibitor was used to treat chondrocytes under inflammatory conditions, and the chondrocytes were treated by RT-PCR method was used to detect matrix metallopeptidase 13(MMP-13), a disintegrin and metalloproteinase with thrombospondin 5, (ADAMTS-5)、nitric oxide synthase 2(NOS2)、Collagen, type II alpha 1(Col2α1)and aggrecan (Acan) mRNA in chondrocytes; primary chondrocytes were treated with different concentrations of TRPV4 overexpression plasmid, and the optimal overexpression dose was screened. The mRNA expressions of TRPV4, MMP-13, ADAMTS-5, NOS2, Col2α1 and Acan in chondrocytes under the optimal TRPV4 overexpression dose were detected. RESULTS: Toluidine blue staining and Alcian blue staining identified the extracted cells as primary chondrocytes;RT-PCR showed that TRPV4, MMP-13, ADAMTS-5, NOS2 mRNA in chondrocytes treated with TRPV4 inhibitor under inflammatory conditions. The expression of Col2α1 mRNA was significantly decreased (P<0.05), and the expression of Col2α1 mRNA was increased (P<0.05). Although there was no significant difference in the expression of Acan mRNA, the overall trend was also increasing. The expression of Col2α1 and Acan mRNA in chondrocytes was significantly decreased (P<0.05), and the expression of NOS2 mRNA was increased(P<0.05), but there was no significant difference in MMP-13 and ADAMTS-5 (P>0.05). CONCLUSION Inhibiting the expression of TRPV4 can down-regulate the expression of genes related to chondrocyte degeneration.
Animals ; Rats ; Aggrecans/metabolism* ; Cartilage, Articular ; Cells, Cultured ; Chondrocytes ; Interleukin-1beta/metabolism* ; Matrix Metalloproteinase 13/metabolism* ; Rats, Sprague-Dawley ; RNA, Messenger/metabolism* ; TRPV Cation Channels/metabolism*

OBJECTIVE: To observe the effects of moxibustion at "Baihui" (GV 20) and "Dazhui" (GV 14) at different time points on the serum level of β-endorphin (β-EP), substance P (SP) and expression of interleukin-1β (IL-1β) and cyclooxygenase-2 (COX-2) protein in brainstem in rats with migraine, and to explore the effect and mechanism of moxibustion in preventing and treating migraine. METHODS: Forty male SD rats were randomly divided into a blank group, a model group, a prevention+treatment (PT) group and a treatment group, 10 rats in each group. Except the blank group, the rats in the remaining groups were injected with nitroglycerin subcutaneously to prepare migraine model. The rats in the PT group were treated with moxibustion 7 days before modeling (once a day) and 30 min after modeling, while the rats in the treatment group were treated with moxibustion 30 min after modeling. The "Baihui" (GV 20) and "Dazhui" (GV 14) were taken for 30 minutes each time. The behavioral scores in each group were observed before and after modeling. After intervention, ELISA method was used to detect the serum level of β-EP and SP; the immunohistochemistry method was used to detect the number of positive cells of IL-1β in brainstem; the Western blot method was used to detect the expression of COX-2 protein in brainstem. RESULTS: Compared with the blank group, the behavioral scores in the model group were increased 0-30 min, 60-90 min and 90-120 min after modeling (P<0.01); compared with the model group, in the treatment group and the PT group, the behavioral scores were decreased 60-90 min and 90-120 min after modeling (P<0.01). Compared with the blank group, in the model group, the serum level of β-EP was decreased (P<0.01), while the serum level of SP, the number of positive cells of IL-1β in brainstem and the expression of COX-2 protein were increased (P<0.01). Compared with the model group, in the PT group and and the treatment group, the serum level of β-EP was increased (P<0.01), while the serum level of SP, the number of positive cells of IL-1β and the expression of COX-2 protein in brainstem were decreased (P<0.01, P<0.05). Compared with the treatment group, in the PT group, the serum level of β-EP was increased and COX-2 protein expression was decreased (P<0.05). CONCLUSION Moxibustion could effectively relieve migraine. The mechanism may be related to reduce the serum level of SP, IL-1β and COX-2 protein expression in brainstem, and increase the serum level of β-EP, and the optimal effect is observed in the PT group.
Rats ; Male ; Animals ; Moxibustion ; Rats, Sprague-Dawley ; Cyclooxygenase 2 ; beta-Endorphin ; Substance P ; Interleukin-1beta ; Migraine Disorders ; Brain Stem

OBJECTIVE: To investigate the effect of electroacupuncture (EA) at Neiguan (PC 6) on myocardial fibrosis in spontaneously hypertensive rats (SHRs), and to explore the contribution of interleukin-1 β (IL-1 β), insulin-like growth factor 1 (IGF-1), and transforming growth factor β 1 (TGF- β 1) to the effects. METHODS: Nine 12-weeks-old Wistar Kyoto (WKY) male rats were employed as the normal group. Twenty-seven SHRs were equally randomized into SHR, SHR+EA, and SHR + sham groups. EA was applied at bilateral PC 6 once a day 30 min per day in 8 consecutive weeks. After 8-weeks EA treatment at PC 6, histopathologic changes of collagen type I (Col I), collagen type 1 (Col 1) and the levels of IGF-1, 1L-1 β, TGF- β 1, matrix metalloproteinase (MMP)-2 and MMP-9 were examined in myocardial tissure respectively. RESULTS: After 8-weeks EA treatment at PC 6, the enhanced myocardial fibrosis in SHRs were characterized by the increased mean fluorescence intensity of Col I and Col 1 in myocardium tissue (P<0.01). All these abnormal alterations above in SHR + EA group was significantly lower compared with the SHR group (P<0.01). Meanwhile, the increased levels of IL-1 β, IGF-1, TGF-β 1 in serum or myocardial tissue of SHRs, diminished MMP 9 mRNA expression in SHRs were also markedly inhibited after 8 weeks of EA treatment (P<0.05 or P<0.01). Furthermore, the contents of IL-1 β, IGF-1, TGF-β 1 in myocardial tissue were positively correlated with the systolic blood pressure and hydroxyproline respectively (P<0.01). CONCLUSION EA at bilateral PC 6 could ameliorate cardiac fibrosis in SHRs, which might be mediated by regulation of 1L-1 β/IGF-1-TGF- β 1-MMP9 pathway.
Rats ; Animals ; Male ; Rats, Inbred WKY ; Electroacupuncture ; Hypertension/therapy* ; Insulin-Like Growth Factor I ; Interleukin-1beta ; Rats, Inbred SHR ; Essential Hypertension ; Myocardium/pathology* ; Collagen Type I ; Fibrosis

10,11-Dehydrocurvularin (DCV) is a natural-product macrolide that has been shown to exert anti-inflammatory activity. However, the underlying mechanism of its anti-inflammatory activity remains poorly understood. Aberrant activation of the NLRP3 inflammasome is involved in diverse inflammation-related diseases, which should be controlled. The results showed that DCV specifically inhibited the activation of the NLRP3 inflammasome in association with reduced IL-1β secretion and caspase-1 activation, without effect on the NLRC4 and AIM2 inflammasomes. Furthermore, DCV disturbed the interaction between NEK7 and NLRP3, resulting in the inhibition of NLRP3 inflammasome activation. The C=C double bond of DCV was required for the NLRP3 inflammasome inhibition induced by DCV. Importantly, DCV ameliorated inflammation in vivo through inhibiting the NLRP3 inflammasome. Taken together, our study reveals a novel mechanism by which DCV suppresses inflammation, which indicates the potential role of DCV in NLRP3 inflammasome-driven inflammatory disorders.
Animals ; Mice ; Inflammasomes ; NLR Family, Pyrin Domain-Containing 3 Protein/genetics* ; Inflammation/drug therapy* ; Anti-Inflammatory Agents/pharmacology* ; Interleukin-1beta/genetics* ; Mice, Inbred C57BL

This study aims to explore the effect of "Trichosanthis Fructus-Allii Macrostemonis" combination(GX) on the activation of NOD-, LRR-, and pyrin domain-containing protein 3(NLRP3) inflammasome, the release of inflammatory cytokines, and the level of autophagy in RAW264.7 macrophage damaged by lipopolysaccharide(LPS), and the mechanism of GX against inflammatory response in macrophages. To be specific, LPS was used to induce the injury of RAW264.7 cells. Cell Counting Kit-8(CCK-8) assay was employed to measure the survival rate of cells, and Western blot to detect the protein expression of NLRP3, apoptosis-associated speck-like protein(ASC), cysteine-aspartic acid protease(caspase)-1, interleukin(IL)-18, IL-1β, microtubule-associated protein light chain 3(LC3)-Ⅱ, and selective autophagy junction protein p62/sequestosome 1 in RAW264.7 macrophages. ELISA was used to measure the levels of IL-18 and IL-1β in RAW264.7 cells. Transmission electron microscopy was applied to observe the number of autophagosomes in RAW264.7 cells. Immunofulourescence staining was used to detect the expression of LC3-Ⅱ and p62 in RAW264.7 cells. The result showed that GX significantly reduced the protein expression of NLRP3, ASC, and caspase-1 in RAW264.7 cells, significantly increased the protein expression of LC3Ⅱ, decreased the expression of p62, significantly inhibited the secretion of IL-18 and IL-1β, significantly increased the number of autophagosomes, significantly enhanced the immunofluorescence of LC3Ⅱ, and reduced the immunofluorescence of p62. Furthermore, 3-methyladenine(3-MA) could reverse the inhibitory effect of GX on NLRP3, ASC, and caspase-1 and reduce the release of IL-18 and IL-1β. In summary, GX can increase of the autophagy activity of RAW264.7 and inhibit the activation of NLRP3 inflammasome, thereby reducing the release of inflammatory cytokines and suppressing inflammatory response in macrophages.
Inflammasomes/metabolism* ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Interleukin-18/metabolism* ; Lipopolysaccharides/pharmacology* ; Macrophages ; Cytokines/metabolism* ; Caspase 1/metabolism* ; Autophagy ; Interleukin-1beta/metabolism*