scholarly journals Punicalagin Prevents Inflammation in LPS-Induced RAW264.7 Macrophages by Inhibiting FoxO3a/Autophagy Signaling Pathway

Nutrients ◽  
2019 ◽  
Vol 11 (11) ◽  
pp. 2794 ◽  
Author(s):  
Cao ◽  
Chen ◽  
Ren ◽  
Zhang ◽  
Tan ◽  
...  
Keyword(s):  
Western Blot ◽  
Signaling Pathway ◽  
Inflammatory Cytokines ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Inflammatory Responses ◽  
Raw264.7 Macrophages ◽  
Tnf Α ◽  
Autophagy Inhibition ◽  
Pro Inflammatory Cytokines

Punicalagin, a hydrolysable tannin of pomegranate juice, exhibits multiple biological effects, including inhibiting production of pro-inflammatory cytokines in macrophages. Autophagy, an intracellular self-digestion process, has been recently shown to regulate inflammatory responses. In this study, we investigated the anti-inflammatory potential of punicalagin in lipopolysaccharide (LPS) induced RAW264.7 macrophages and uncovered the underlying mechanisms. Punicalagin significantly attenuated, in a concentration-dependent manner, LPS-induced release of NO and decreased pro-inflammatory cytokines TNF-α and IL-6 release at the highest concentration. We found that punicalagin inhibited NF-κB and MAPK activation in LPS-induced RAW264.7 macrophages. Western blot analysis revealed that punicalagin pre-treatment enhanced LC3II, p62 expression, and decreased Beclin1 expression in LPS-induced macrophages. MDC assays were used to determine the autophagic process and the results worked in concert with Western blot analysis. In addition, our observations indicated that LPS-induced releases of NO, TNF-α, and IL-6 were attenuated by treatment with autophagy inhibitor chloroquine, suggesting that autophagy inhibition participated in anti-inflammatory effect. We also found that punicalagin downregulated FoxO3a expression, resulting in autophagy inhibition. Overall these results suggested that punicalagin played an important role in the attenuation of LPS-induced inflammatory responses in RAW264.7 macrophages and that the mechanisms involved downregulation of the FoxO3a/autophagy signaling pathway.

FGF-7 facilitates the process of psoriasis by inducing TNF-α expression in HaCaT cells

2019 ◽  
Vol 51 (10) ◽  
pp. 1056-1063 ◽  
Author(s):  
Jiaojiao Pu ◽  
Rui Wang ◽  
Guanglin Zhang ◽  
Ju Wang
Keyword(s):  
Western Blot ◽  
Signaling Pathway ◽  
Specific Antibody ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Quantitative Polymerase Chain Reaction ◽  
Hacat Cells ◽  
Tnf Α

Abstract The purpose of this study was to uncover the mechanism of tumor necrosis factor (TNF)-α induction by fibroblast growth factor-7 (FGF-7) in human HaCaT cells and the potential role of FGF-7-specific antibody F-9 in psoriatic therapy. TNF-α expression in HaCaT cells induced by FGF-7 was analyzed by quantitative polymerase chain reaction, western blot analysis, and enzyme-linked immunosorbent assays. In vivo, the BALB/c mouse psoriasis model established by topical application of imiquimod (IMQ) was used to determine the role of FGF-7-specific antibody (F-9) in skin inflammation. We found that induction of TNF-α expression by FGF-7 in HaCaT cells was suppressed by FGF-7-specific antibody F-9. Western blot analysis results showed that FGF-7 induced TNF-α expression in HaCaT cells via the FGF receptor 2 (FGFR2)/AKT/NF-κB signaling pathway. In vivo, F-9 could significantly ameliorate the inflammations in a mouse psoriatic model evaluated by Psoriasis Area and Severity Index scores and ear thickness, which was consistent with the results of hematoxylin–eosin staining, immunohistochemistry assay, and western blot analysis. These results indicate that FGF-7 induces TNF-α expression in HaCaT cells and FGF-7 antibody F-9 alleviates IMQ-induced psoriasiform in mice. Therefore, FGF-7/FGFR2 signaling pathway is a potential target for psoriasis treatment.

scholarly journals Separation of Flavonoids and the Evaluation of the Anti-inflammatory Potential of Flavonoid-enriched Extracts from the Thlaspi arvense Linn by Inhibiting the Activation of TLR-4/NF-κ B

2021 ◽  
Author(s):  
Xiang Wang ◽  
Panke Zeng ◽  
Xuejiao Li ◽  
Liyuan Cheng ◽  
Haroon Rashidb ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Inflammatory Responses ◽  
Activity Assay ◽  
Nf Kappa B ◽  
Thlaspi Arvense ◽  
Raw264.7 Macrophages ◽  
Tnf Α ◽  
Anti Inflammatory ◽  
First Time ◽  
Pro Inflammatory Cytokines

Abstract Background: Thlaspi arvense Linn, belonging to the dicotyledonous cruciferous family, is distributed Europe and Asia. In this study, we evaluated for the first time the anti-inflammatory effects of Thlaspi arvense Linn on LPS-stimulated RAW264.7 macrophages, and explored the related mechanism.Methods: The extract was identified and quantified using the HPLC, NMR. The anti-inflammatory activities of crude extracts C11, C12, C13 were screened by xylene-induced ear swelling and carrageenan-induced foot swelling in mice. The inflammatory mediators, pro-inflammatory cytokines and TLR-4-mediated signals in LPS-stimulated RAW264.7 macrophages were determined using NO activity assay, MTS, ELISA and Western blot.Results:The extract of Thlaspi arvense Linn was found to enrich flavonoid（mainly Orientin,Isoorientin,Vitexin;,Isovitexin,Luteolin-7-O-β-D-glucoside,Apigenin-7-O-β-D-glucoside,Luteolin,Apigeni）.5,7-dihydroxy-flavone-4′-(6′′-β-O-glucopyranoside)-β-O-D-glucopyranoside was novel, whereas isosapogenin and 8-methoxyvitexin were isolated for the first time from Thlaspi arvense Linn. The extract (flavonoid-enriched) inhibited xylene-induced ear swelling and carrageenan-induced foot swelling in mice. And suppressed LPS-induced overrelease of iNOS,TNF-α,COX-2 and IL-6 in RAW264.7 macrophages. The extract inhibited the inflammatory response through the signaling pathway mediated by TLR-4/NF- kappa B pathway and its downstream signals, IκB-α, NF-κB-P65 and IL-1β in LPS-stimulated macrophages.Conclusions: The present results demonstrate that the extract of Thlaspi arvense Linn inhibit inflammatory responses via the TLR-4/NF-KB-mediated signaling pathway.

scholarly journals Puerarin Prevents Acute Liver Injury via Inhibiting Inflammatory Responses and ZEB2 Expression

2021 ◽  
Vol 12 ◽  
Author(s):  
Junfa Yang ◽  
Maomao Wu ◽  
Hui Fang ◽  
Yue Su ◽  
Lingling Zhang ◽  
...  
Keyword(s):  
Liver Injury ◽  
Signaling Pathway ◽  
Inflammatory Cytokines ◽  
Inflammatory Responses ◽  
Acute Liver Injury ◽  
Expression Level ◽  
Inflammatory Factors ◽  
Tnf Α ◽  
Zeb2 Expression ◽  
Pro Inflammatory Cytokines

Puerarin, an isoflavone component extracted from herb radix puerariae, is widely used in China in the treatment of immune diseases and inflammation. Previous studies have demonstrated that puerarin prevented acute lung injury by regulating inflammatory responses. However, the effect of puerarin on acute liver injury (ALI) was unclear. The purpose of this study was to explore the beneficial effects of puerarin when applied to ALI. We found that puerarin inhibited liver injury and inflammatory cell infiltration in lipopolysaccharide (LPS)/D-galactose (D-Gal)-induced acute liver failure and the liver pro-inflammatory cytokines interleukin (IL)-1β, IL-6, and tumor necrosis factor-alpha (TNF-α) in liver tissues with ALI and LPS-induced L-02 cells but upregulated the expression level of zinc finger E-box-binding homeobox 2 (ZEB2). Significantly, the results of this study showed that the inhibition of liver pro-inflammatory cytokine (IL-1β, IL-6, and TNF-α) production in LPS-induced L-02 cells was caused by ZEB2 overexpression. However, knocking down ZEB2 promoted LPS-mediated secretion of liver pro-inflammatory cytokines in L-02 cells. Additional experiments showed that puerarin inhibited the activation of the NF-κB signaling pathway by elevating ZEB2 expression in L-02 cells. In summary, puerarin most likely prevented activation of the pro-inflammatory factors and reduced LPS/D-Gal-induced liver injury by enhancing the ZEB2 expression level and, consequently, blocking activation of the NF-κB signaling pathway in the liver.

scholarly journals Long non-coding RNA MALAT1 enhances the apoptosis of cardiomyocytes through autophagy inhibition by regulating TSC2-mTOR signaling

2019 ◽  
Vol 52 (1) ◽  
Author(s):  
Hao Hu ◽  
Jiawei Wu ◽  
Xiaofan Yu ◽  
Junling Zhou ◽  
Hua Yu ◽  
...  
Keyword(s):  
Western Blot ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Noncoding Rna ◽  
Mtor Signaling ◽  
Cardiomyocyte Apoptosis ◽  
Tunel Staining ◽  
Promoter Regions ◽  
Related Proteins ◽  
Autophagy Inhibition

Abstract Background Our previous study showed that knockdown of long noncoding RNA (lncRNA) metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) attenuated myocardial apoptosis in mouse acute myocardial infarction (AMI). This study aims to explore whether MALAT1 enhanced cardiomyocyte apoptosis via autophagy regulation and the underlying mechanisms of MALAT1 regulating autophagy. Methods Cardiomyocytes were isolated from neonatal mice and then stimulated with hypoxia/reoxygenation (H/R) injury to mimic AMI. The autophagy level was assessed using GFP-LC3 immunofluorescence and western blot analysis of autophagy-related proteins. RNA pull-down and RNA immunoprecipitation (RIP) was performed to analyze the binding of MALAT1 and EZH2. Chromatin immunoprecipitation (ChIP) assay was performed to analyze the binding of TSC2 promoter and EZH2. The cell apoptosis was evaluated using TUNEL staining and western blot analysis of apoptosis-related proteins. Results H/R injury increased MALAT1 expression in cardiomyocytes. Furthermore, MALAT1 overexpression inhibited, whereas MALAT1 knockdown enhanced the autophagy of cardiomyocytes. Moreover, MALAT1 overexpression recruited EZH2 to TSC2 promoter regions to elevate H3K27me3 and epigenetically inhibited TSC2 transcription. Importantly, TSC2 overexpression suppressed mTOR signaling and then activated the autophagy. Further results showed that MALAT1 inhibited proliferation and enhanced apoptosis of cardiomyocytes through inhibiting TSC2 and autophagy. Conclusion These findings demonstrate that the increased MALAT1 expression induced by H/R injury enhances cardiomyocyte apoptosis through autophagy inhibition by regulating TSC2-mTOR signaling.

scholarly journals The Anti-Inflammatory Effects of Angiogenin in an Endotoxin Induced Uveitis in Rats

2020 ◽  
Vol 21 (2) ◽  
pp. 413
Author(s):  
Jihae Park ◽  
Jee Taek Kim ◽  
Soo Jin Lee ◽  
Jae Chan Kim
Keyword(s):  
Western Blot ◽  
Inflammatory Cytokines ◽  
Aqueous Humor ◽  
Rat Model ◽  
Inflammatory Responses ◽  
Inflammatory Cells ◽  
Ocular Tissue ◽  
Tnf Α ◽  
Anti Inflammatory

Angiogenin (ANG) is involved in the innate immune system and inflammatory disease. The aim of this study is to evaluate the anti-inflammatory effects of ANG in an endotoxin induced uveitis (EIU) rat model and the pathways involved. EIU rats were treated with balanced salt solution (BSS), a non-functional mutant ANG (mANG), or wild-type ANG (ANG). The integrity of the blood-aqueous barrier was evaluated by the infiltrating cell and protein concentrations in aqueous humor. Histopathology, Western blot, and real-time qRT-PCR of aqueous humor and ocular tissue were performed to analyze inflammatory cytokines and transcription factors. EIU treated with ANG had decreased inflammatory cells and protein concentrations in the anterior chamber. Compared to BSS and mANG, ANG treatment showed reduced expression of IL-1β, IL-8, TNF-α, and Myd88, while the expression of IL-4 and IL-10 was increased. Western blot of ANG treatment showed decreased expression of IL-6, inducible nitric oxide synthase (iNOS), IL-1β, TNF-α, and phosphorylated NF-κB and increased expression of IL-10. In conclusion, ANG seems to reduce effectively immune mediated inflammation in the EIU rat model by reducing the expression of proinflammatory cytokines, while increasing the expression of anti-inflammatory cytokines through pathways related to NF-κB. Therefore, ANG shows potential for effectively suppressing immune-inflammatory responses in vivo.

Peripheral interleukin-6 promotes resilience versus susceptibility to inescapable electric stress

2015 ◽  
Vol 27 (5) ◽  
pp. 312-316 ◽  
Author(s):  
Chun Yang ◽  
Yukihiko Shirayama ◽  
Ji-Chun Zhang ◽  
Qian Ren ◽  
Kenji Hashimoto
Keyword(s):  
Prefrontal Cortex ◽  
Western Blot ◽  
Inflammatory Cytokines ◽  
Learned Helplessness ◽  
Blot Analysis ◽  
Interleukin 6 ◽  
Serum Levels ◽  
Enzyme Linked Immunosorbent Assay ◽  
Electric Stress ◽  
Pro Inflammatory Cytokines

ObjectiveAccumulating evidences suggest that pro-inflammatory cytokines such as interleukin-6 (IL-6) play a role in the pathophysiology of depression. In the learned helplessness (LH) paradigm, ~35% rats are resilient to inescapable stress.MethodsLevels of IL-6 in the serum and medial prefrontal cortex (mPFC) of LH rats (susceptible) and non-LH rats (resilience) were measured using enzyme-linked immunosorbent assay and western blot analysis, respectively.ResultsSerum levels of IL-6 in the LH rats were significantly higher than those of control and non-LH rats. In contrast, tissue levels of IL-6 in the mPFC were not different among three groups.ConclusionThe results suggest that peripheral IL-6 may contribute to resilience versus susceptibility to inescapable stress.

scholarly journals Aging as a Risk Factor on the Immunoexpression of Pro-Inflammatory IL-1β, IL-6 and TNF-α Cytokines in Chronic Apical Periodontitis Lesions

Biology ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 14
Author(s):  
Quésia Euclides Teixeira ◽  
Dennis de Carvalho Ferreira ◽  
Alexandre Marques Paes da Silva ◽  
Lucio Souza Gonçalves ◽  
Fabio Ramoa Pires ◽  
...  
Keyword(s):  
Elderly Patients ◽  
Inflammatory Cytokines ◽  
Inflammatory Responses ◽  
The Elderly ◽  
Apical Periodontitis ◽  
Middle Aged ◽  
Tnf Α ◽  
Significant Difference ◽  
Cytokine Levels ◽  
Pro Inflammatory Cytokines

Persistent inflammatory responses in the elderly may act as modifiers on the progression and repair of chronic apical periodontitis lesions (CAPLs). While the involvement of IL-1β, IL-6 and TNF-α in inflammatory responses and, particularly, in CAPL has been documented, their expression in elderly patients needs to be further characterized. Therefore, the purpose of this study was to evaluate and compare the expressions of pro-inflammatory cytokines in CAPL from elderly individuals with young/middle-aged individuals. Thirty CAPL (15 cysts and 15 granulomas) from elderly patients (>60 years) and 30 CAPL (15 cysts and 15 granuloma) from young/middle-aged individuals (20–56 years) were selected. Immunohistochemical reactions were performed against IL-1β, IL-6 and TNF-α. The slides were subdivided into five high-magnification fields and analyzed. The number of positive stains was evaluated for each antibody. There was no significant difference between the cytokines when the cysts and granuloma were compared in the two groups. In the young/middle-aged, only IL-1β showed a difference and was significantly higher in granulomas (p = 0.019). CAPL pro-inflammatory cytokine levels in the elderly were significantly higher than in young/middle-aged individuals (p < 0.05). The pro-inflammatory cytokines IL-1β, IL-6 and TNF-α were significantly higher in CAPL in the elderly compared with the young/middle-aged group. Further elaborate research studies/analyses to elucidate the reasons for and consequences of inflammation in the elderly are recommended.

Adenosine monophosphate-activated protein kinase activator inhibits activation of fibroblast-like synoviocytes but promotes hyaluronan and proteoglycan link protein 1 secretion

2020 ◽  
Author(s):  
Yong Chen ◽  
Qiu Fujuan ◽  
Yu Beijia ◽  
Zuo Fangfang ◽  
Bi Yanan ◽  
...  
Keyword(s):  
Western Blot ◽  
Rna Sequencing ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Enrichment Analysis ◽  
Protective Mechanism ◽  
Sequencing Data ◽  
Positive Correlation ◽  
Tnf Α ◽  
Fibroblast Like Synoviocytes

Abstract Objectives: To determine whether any correlation exists between disease activity and AMPK levels in rheumatoid arthritis (RA) patients and investigate the effects of AMPK activator treatment on RA fibroblast-like synoviocytes (RA-FLS).Methods: Serum AMPK-α1, p-AMPK-α1, TNF-α and IL-17 levels between osteoarthritis (OA) and RA patients having different disease activities were compared by ELISA. Differentially expressed genes (DEGs) between RA and OA synovium from NCBI GEO Profiles (accession numbers: GSE1202112, GSE55235, GSE5545713) were identified and the genes intersecting in all the three datasets were selected for enrichment analysis. Immunohistochemical staining was done with synovium obtained from OA and RA patients for p-AMPK-α1. AMPK gene expression in synovium was semi-quantified by RT-qPCR. RNA sequencing of FLS was performed and DEGs were selected for KEGG enrichment analysis. AMPK activator, metformin, treated RA-FLS were tested for proliferation and migration by MTT and scratch test, respectively. Expression of IL-6, AMPK-α1, PKA-α, RAPTOR, mTOR, HAPLN1, RUNX1 and RUNX2 genes were determined by qPCR. Phosphorylated AMPK-α1 and HAPLN1 levels were determined by an automated electrophoresis-western blot analysis method.Results: In RA sera, a positive correlation between p-AMPK-α1 levels and DAS28 (r = 0.270, 95%CI: 0.142-0.492, p < 0.0001) as well as CRP levels (r = 0.259, 95%CI: 0.009-0.478, p < 0.05) was found. Similarly, a positive correlation was observed between AMPK-α1 and TNF-α levels (r = 0.460, 95% CI: 0.241-0.640, p = 0.0002). DEGs between OA and RA synovium from NCBI GEO profiles and our RNA sequencing data suggested activation of metabolic pathways specific to RA-FLS. AMPK-α1 was highly expressed in the synovium of RA but not OA patients. Metformin at higher concentrations inhibited RA-FLS proliferation in a dose dependant manner, however, at lower concentrations it has an opposite effect. On the other hand, AMPK inhibitor, dorsmophin, promoted the proliferation of RA-FLS significantly. Interestingly, both metformin and dorsmophin substantially inhibited the migration of RA-FLS. In FLS, relative expression level of IL-6 mRNA was significantly decreased after metformin treatment, while the expression of AMPK-α1, PKA-α and HAPLN1 genes were significantly increased. Western blot analysis confirmed increased expression of p-AMPK-α1 and HAPLN1 genes in the metformin treated FLS.Conclusions: Inflammatory stress in RA synovium leads to an increase in AMPK levels, possibly as a protective mechanism. AMPK activator but not metformin per se could be a potential therapeutic for RA by promoting HAPLN1 secretion to protect the joints.

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