In vitro Cell-Based Assays for Potency Testing of Anti-TNF-α Biological Drugs

Psoriasis is a chronic inflammatory skin disease characterized by hyperproliferation of keratinocytes and expression of pro-inflammatory cytokines in the epidermis. New biological drugs were developed for the systemic treatment of moderate to severe psoriasis. However, products for the topical treatment of mild psoriasis are still required. Here, we examined the effect of natural compounds on psoriasis-like keratinocytes in vitro and ex vivo. Psoriasis-like keratinocytes were generated by treating human primary keratinocytes with the psoriasis-associated cytokines IL-17A, TNF-α and IL-22. Initially, 10 botanical extracts from Ayurvedic Medicine, Traditional Chinese Medicine, Northern American traditional medicine and Occidental Monastic Medicine were investigated using BrdU assays and IL-6 and IL-8 ELISAs. Curcuma amada, Humulus lupulus and Hypericum perforatum turned out to be the most effective plant extracts. In vitro, the plant extracts inhibited the expression of anti-microbial peptides (β-defensin 2), the hyperproliferation marker keratin 17, the glucose transporter 1 and downregulated the nuclear translocation of NF-κB and pSTAT3. In an ex vivo psoriasis model, Humulus lupulus displayed the most prominent anti-proliferative and anti-inflammatory effect. In conclusion, among the plant extracts investigated, Humulus lupulus showed the most promising anti-psoriatic effect. It is an interesting candidate for topical psoriasis treatment that should be further studied in clinical trials.

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Die RhoA-Effektorkinase PKN1 vermindert die durch TNF-α induzierte Permeabilitätssteigerung in einem in vitro Modell der intestinalen Barriere

Zeitschrift für Gastroenterologie ◽

10.1055/s-0032-1324101 ◽

2012 ◽

Vol 50 (08) ◽

Author(s):

M Gluth ◽

C Weber ◽

H Mukai ◽

D Baumgart ◽

J Turner ◽

...

Keyword(s):

Tnf Α

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Traditional Herbal Medicines, Newer Herbs and Other Novel Approaches Integrated in Herbal Medicine for Atopic Dermatitis-A Narrative Review

Current Drug Therapy ◽

10.2174/1574885514666191018165209 ◽

2020 ◽

Vol 15 (3) ◽

pp. 194-208

Author(s):

Pravin Kumar ◽

Dinesh Kumar Sharma ◽

Mahendra Singh Ashawat

Keyword(s):

Atopic Dermatitis ◽

Herbal Medicines ◽

Developed Countries ◽

Skin Lesions ◽

Scientific Data ◽

Herbal Drugs ◽

Biological Drugs ◽

Alternative Treatment Option

Atopic Dermatitis (AD) is a prolonged reverting skin ailment with characteristically distributed skin lesions. In the previous decades, researchers had shown a marked interest in AD due to its increased prevalence in developed countries. Although different strategies including biological and immune modulators are available for the treatment of AD, each has certain limitations. The researchers had shown considerable interest in the management of AD with herbal medicines. The establishment of herbal drugs for AD might eliminate local as well as systemic adverse effects associated with long term use of corticosteroids and also higher cost of therapy with biological drugs. The present review discusses the traditional East Asian herbal medicines and scientific data related to newer herbal extracts or compositions for the treatment of AD. In vivo animal models and in vitro cell cultures, investigated with herbal medicines to establish a possible role in AD treatment, have also been discussed in the paper. The paper also highlights the role of certain new approaches, i.e. pharmacopuncture, a combination of allopathic and herbal medicines; and novel carriers (liposomes, cubosomes) for herbal drugs on atopic skin. In conclusion, herbal medicines can be a better and safe, complementary and alternative treatment option for AD.

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AAnti-Inflammatory Effects of Plasma Circulating Exosomes Obtained from Normal-Weight and Obese Subjects on Hepatocytes

Endocrine Metabolic & Immune Disorders - Drug Targets ◽

10.2174/1871530320666200505121426 ◽

2020 ◽

Vol 20 ◽

Author(s):

Reza Afrisham ◽

Sahar Sadegh-Nejadi ◽

Reza Meshkani ◽

Solaleh Emamgholipour ◽

Molood Bagherieh ◽

...

Keyword(s):

Inflammatory Cytokines ◽

Hepg2 Cells ◽

Human Liver ◽

Liver Cells ◽

Normal Weight ◽

Protein Levels ◽

Human Liver Cells ◽

Tnf Α ◽

Anti Inflammatory

Introduction: Obesity is a disorder with low-grade chronic inflammation that plays a key role in the hepatic inflammation and steatosis. Moreover, there are studies to support the role of exosomes in the cellular communications, the regulation of metabolic homeostasis and immunomodulatory activity. Accordingly, we aimed to evaluate the influence of plasma circulating exosomes derived from females with normal-weight and obesity on the secretion of inflammatory cytokines in human liver cells. Methods: Plasma circulating exosomes were isolated from four normal (N-Exo) and four obese (O-Exo) women. The exosomes were characterized and approved for CD63 expression (common exosomal protein marker) and morphology/size using the western blot and TEM methods, respectively. The exosomes were used for stimulation of HepG2 cells in vitro. After 24 h incubation, the protein levels of TNF-α,IL-6, and IL-1β were measured in the culture supernatant of HepG2 cells using the ELISA kit. Results: The protein levels of IL-6 and TNF-α in the cells treated with O-Exo and N-Exo reduced significantly in comparison with control group (P=0.039 and P<0.001 respectively), while significance differences were not found between normal and obese groups (P=0.808, and P=0.978 respectively). However, no significant differences were found between three groups in term of IL-1β levels (P=0.069). Based on the correlation analysis, the protein levels of IL-6 were positively correlated with TNF-α (r 0.978, P<0.001). Conclusion: These findings suggest that plasma circulating exosomes have probably anti-inflammatory properties independently from body mass index and may decrease the secretion of inflammatory cytokines in liver. However, further investigations in vitro and in vivo are needed to address the anti-inflammatory function of N-Exo and O-Exo in human liver cells and/or other cells.

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TNF-α Suppresses Autophagic Flux in Acinar Cells in IgG4-Related Sialadenitis

Journal of Dental Research ◽

10.1177/0022034519871890 ◽

2019 ◽

Vol 98 (12) ◽

pp. 1386-1396 ◽

Cited By ~ 3

Author(s):

X. Hong ◽

S.N. Min ◽

Y.Y. Zhang ◽

Y.T. Lin ◽

F. Wang ◽

...

Keyword(s):

Acinar Cell ◽

Cell Injury ◽

Ex Vivo ◽

Secretory Granules ◽

Acinar Cells ◽

Autophagic Flux ◽

C6 Cells ◽

Tnf Α ◽

Α Level

IgG4-related sialadenitis (IgG4-RS) is a newly recognized immune-mediated systemic fibroinflammatory disease that affects salivary glands and leads to hyposalivation. Tumor necrosis factor–α (TNF-α) is a critical proinflammatory cytokine involved in several salivary gland disorders, but its role and mechanism regarding acinar cell injury in IgG4-RS are unknown. Here, we found that TNF-α level was significantly increased in serum and submandibular gland (SMG) of patients and that serum TNF-α level was negatively correlated with saliva flow rate. Ultrastructural observations of IgG4-RS SMGs revealed accumulation of large autophagic vacuoles, as well as dense fibrous bundles, decreased secretory granules, widened intercellular spaces, swollen mitochondria, and expanded endoplasmic reticulum. Expression levels of LC3 and p62 were both increased in patients’ SMGs. TNF-α treatment led to elevated levels of LC3II and p62 in both SMG-C6 cells and cultured human SMG tissues but did not further increase their levels when combined with bafilomycin A1 treatment. Moreover, transfection of Ad-mCherry-GFP-LC3B in SMG-C6 cells confirmed the suppression of autophagic flux after TNF-α treatment. Immunofluorescence imaging revealed that costaining of LC3 and the lysosomal marker LAMP2 was significantly decreased in patients, TNF-α–treated SMG-C6 cells, and cultured human SMGs, indicating a reduction in autophagosome-lysosome fusion. Furthermore, the ratio of pro/mature cathepsin D was elevated in vivo, ex vivo, and in vitro. TNF-α also appeared to induce abnormal acidification of lysosomes in acinar cells, as assessed by lysosomal pH and LysoTracker DND-26 fluorescence intensity. In addition, TNF-α treatment induced transcription factor EB (TFEB) redistribution in SMG-C6 cells, which was consistent with the changes observed in IgG4-RS patients. TNF-α increased the phosphorylation of extracellular signal–regulated kinase (ERK) 1/2, and inhibition of ERK1/2 by U0126 reversed TNF-α–induced TFEB redistribution, lysosomal dysfunction, and autophagic flux suppression. These findings suggest that TNF-α is a key cytokine related to acinar cell injury in IgG4-RS through ERK1/2-mediated autophagic flux suppression.

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Decitabine Promotes Modulation in Phenotype and Function of Monocytes and Macrophages That Drive Immune Response Regulation

Cells ◽

10.3390/cells10040868 ◽

2021 ◽

Vol 10 (4) ◽

pp. 868

Author(s):

Fabiana Albani Zambuzi ◽

Priscilla Mariane Cardoso-Silva ◽

Ricardo Cardoso Castro ◽

Caroline Fontanari ◽

Flavio da Silva Emery ◽

...

Keyword(s):

Immune Response ◽

Hematological Malignancies ◽

M2 Macrophage ◽

M2 Polarization ◽

Tnf Α ◽

Monocytes And Macrophages ◽

Ifn Γ ◽

And Function ◽

The Impact

Decitabine is an approved hypomethylating agent used for treating hematological malignancies. Although decitabine targets altered cells, epidrugs can trigger immunomodulatory effects, reinforcing the hypothesis of immunoregulation in treated patients. We therefore aimed to evaluate the impact of decitabine treatment on the phenotype and functions of monocytes and macrophages, which are pivotal cells of the innate immunity system. In vitro decitabine administration increased bacterial phagocytosis and IL-8 release, but impaired microbicidal activity of monocytes. In addition, during monocyte-to-macrophage differentiation, treatment promoted the M2-like profile, with increased expression of CD206 and ALOX15. Macrophages also demonstrated reduced infection control when exposed to Mycobacterium tuberculosis in vitro. However, cytokine production remained unchanged, indicating an atypical M2 macrophage. Furthermore, when macrophages were cocultured with lymphocytes, decitabine induced a reduction in the release of inflammatory cytokines such as IL-1β, TNF-α, and IFN-γ, maintaining IL-10 production, suggesting that decitabine could potentialize M2 polarization and might be considered as a therapeutic against the exacerbated immune response.

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Dipterocarpus tuberculatus Roxb. Ethanol Extract Has Anti-Inflammatory and Hepatoprotective Effects In Vitro and In Vivo by Targeting the IRAK1/AP-1 Pathway

Molecules ◽

10.3390/molecules26092529 ◽

2021 ◽

Vol 26 (9) ◽

pp. 2529

Author(s):

Haeyeop Kim ◽

Woo Seok Yang ◽

Khin Myo Htwe ◽

Mi-Nam Lee ◽

Young-Dong Kim ◽

...

Keyword(s):

Ethanol Extract ◽

Serum Levels ◽

Hepatoprotective Activity ◽

Tnf Α ◽

Anti Inflammatory ◽

Related Proteins ◽

Pro Inflammatory Cytokines ◽

Inflammatory Effect

Dipterocarpus tuberculatus Roxb. has been used traditionally as a remedy for many diseases, especially inflammation. Therefore, we analyzed and explored the mechanism of the anti-inflammatory effect of a Dipterocarpus tuberculatus Roxb. ethanol extract (Dt-EE). Dt-EE clearly and dose-dependently inhibited the expression of pro-inflammatory cytokines such as IL-6, TNF-α, and IL-1β in lipopolysaccharide (LPS)-treated RAW264.7 cells. Also, Dt-EE suppressed the activation of the MyD88/TRIF-mediated AP-1 pathway and the AP-1 pathway related proteins JNK2, MKK4/7, and TAK1, which occurred as a result of inhibiting the kinase activity of IRAK1 and IRAK4, the most upstream factors of the AP-1 pathway. Finally, Dt-EE displayed hepatoprotective activity in a mouse model of hepatitis induced with LPS/D-galactosamine (D-GalN) through decreasing the serum levels of alanine aminotransferase and suppressing the activation of JNK and IRAK1. Therefore, our results strongly suggest that Dt-EE could be a candidate anti-inflammatory herbal medicine with IRAK1/AP-1 inhibitory and hepatoprotective properties.

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Elevated plasma levels of soluble TNF receptors are associated with morbidity and mortality in patients with acute lung injury

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00302.2004 ◽

2005 ◽

Vol 288 (3) ◽

pp. L426-L431 ◽

Cited By ~ 128

Author(s):

Polly E. Parsons ◽

Michael A. Matthay ◽

Lorraine B. Ware ◽

Mark D. Eisner

Keyword(s):

Lung Injury ◽

Plasma Levels ◽

A549 Cells ◽

Multicenter Trial ◽

Morbidity And Mortality ◽

Lung Protective Ventilation ◽

Single Center ◽

Lung Protection ◽

Tnf Α

Ventilator-induced lung injury (VILI) is an inflammatory process that can be attenuated by lung protective ventilation strategies. Our objectives to further investigate the pathogenesis of ALI and VILI and the mechanism of lung protection in these syndromes were: 1) to determine if plasma measurements of soluble TNF receptor I (sTNFRI) and II (sTNFRII) would predict the development of ALI and mortality in a small single center trial; 2) to test the predictive value of these markers and of TNF-α in a larger, broader group of patients with ALI; 3) to test the hypothesis that low tidal volume ventilation (LTVV) would be associated with a decrease in plasma levels of TNF-α, sTNFRI, and sTNFRII. In the single center study, sTNFRI and II levels were higher in patients at risk for and with ALI, but they did not predict the development of the syndrome. In the multicenter trial sTNFRI and II were strongly associated with mortality (OR 5.76/1 log10 increment in receptor level; 95% CI 2.63–12.6 and OR 2.58; 95% CI 1.05–6.31, respectively) and morbidity measured as fewer nonpulmonary organ failure-free and ventilator-free days. The LTVV strategy was associated with an attenuation of plasma sTNFRI levels. In vitro, stimulated A549 cells release sTNFRI but not sTNRFII. In conclusion, plasma levels of sTNFRI and II can serve as biomarkers for morbidity and mortality in patients with ALI. Furthermore, LTVV is associated with a specific decrease in sTNFRI levels. This suggests that one beneficial effect of LTVV may be to attenuate alveolar epithelial injury.

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TNF-α Triggers RIP1/FADD/Caspase-8-Mediated Apoptosis of Astrocytes and RIP3/MLKL-Mediated Necroptosis of Neurons Induced by Angiostrongylus cantonensis Infection

Cellular and Molecular Neurobiology ◽

10.1007/s10571-021-01063-w ◽

2021 ◽

Author(s):

Hongli Zhou ◽

Minyu Zhou ◽

Yue Hu ◽

Yanin Limpanon ◽

Yubin Ma ◽

...

Keyword(s):

Cell Death ◽

Enrichment Analysis ◽

Angiostrongylus Cantonensis ◽

Gene Set Enrichment Analysis ◽

Caspase 8 ◽

Cns Injury ◽

Vitro Assay ◽

Tnf Α

AbstractAngiostrongylus cantonensis (AC) can cause severe eosinophilic meningitis or encephalitis in non-permissive hosts accompanied by apoptosis and necroptosis of brain cells. However, the explicit underlying molecular basis of apoptosis and necroptosis upon AC infection has not yet been elucidated. To determine the specific pathways of apoptosis and necroptosis upon AC infection, gene set enrichment analysis (GSEA) and protein–protein interaction (PPI) analysis for gene expression microarray (accession number: GSE159486) of mouse brain infected by AC revealed that TNF-α likely played a central role in the apoptosis and necroptosis in the context of AC infection, which was further confirmed via an in vivo rescue assay after treating with TNF-α inhibitor. The signalling axes involved in apoptosis and necroptosis were investigated via immunoprecipitation and immunoblotting. Immunofluorescence was used to identify the specific cells that underwent apoptosis or necroptosis. The results showed that TNF-α induced apoptosis of astrocytes through the RIP1/FADD/Caspase-8 axis and induced necroptosis of neurons by the RIP3/MLKL signalling pathway. In addition, in vitro assay revealed that TNF-α secretion by microglia increased upon LSA stimulation and caused necroptosis of neurons. The present study provided the first evidence that TNF-α was secreted by microglia stimulated by AC infection, which caused cell death via parallel pathways of astrocyte apoptosis (mediated by the RIP1/FADD/caspase-8 axis) and neuron necroptosis (driven by the RIP3/MLKL complex). Our research comprehensively elucidated the mechanism of cell death after AC infection and provided new insight into targeting TNF-α signalling as a therapeutic strategy for CNS injury.

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PSIII-38 Butyric acid and derivatives: In vitro anti-inflammatory effects tested in porcine alveolar macrophages

Journal of Animal Science ◽

10.1093/jas/skaa278.650 ◽

2020 ◽

Vol 98 (Supplement_4) ◽

pp. 370-370

Author(s):

Lauren L Kovanda ◽

Monika Hejna ◽

Yanhong Liu

Keyword(s):

Organic Acid ◽

Butyric Acid ◽

Sodium Butyrate ◽

Alveolar Macrophages ◽

Block Design ◽

Lps Challenge ◽

Tnf Α ◽

Anti Inflammatory ◽

Challenge Group

Abstract The aim of this experiment was to examine the anti-inflammatory effects of butyric acid, sodium butyrate, monobutyrin and tributyrin using porcine alveolar macrophages (PAMs). PAMs were isolated from the bronchial lavage of 6 piglets at 6 weeks of age, and then seeded at 106 cells/mL in 24-well plates. After 24 h incubation, cells were treated with different treatments in a randomized complete block design with 10 replicates. The treatments were in a factorial arrangement with 2 doses of lipopolysaccharide (LPS, 0 or 1 μg/mL) and 5 levels of organic acid (0, 0.5, 1, 2, 4 mM for butyric acid and tributyrin and 0, 1, 2, 4, 8 mM for sodium butyrate and monobutyrin). Supernatants were collected after another 24 h incubation and analyzed for tumor necrosis factor alpha (TNF-α). Cell viability was also tested by the MTT assay. Data were analyzed using the MIXED procedure of SAS. No cytotoxic effect was observed in LPS challenge and each organic acid with the percentage of live cells was more than 76% in comparison to the sham control. Sodium butyrate at 2 and 4 mM dose exhibited (P < 0.01) a stimulatory effect on cell proliferation. LPS challenge remarkably stimulated (P < 0.0001) TNF-α secretion from PAMs. In the non-challenge group, butyric acid, monobutyrin, and tributyrin linearly reduced TNF-α production from PAMs, whereas 2 mM sodium butyrate tended to increase (P = 0.056) TNF-α secretion from PAMs. In the LPS challenge group, all tested organic acid dose-dependently reduced (P < 0.001) TNF-α production from LPS-challenged PAMs, with the strongest inhibiting effect observed at the highest dose. Results indicated that butyric acid and its derivatives that were tested in the current experiment all had strong anti-inflammatory activities in vitro.

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