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Stem Cell-Induced Inflammation in Cholesteatoma is Inhibited by the TLR4 Antagonist LPS-RS.

ABSTRACT: Cholesteatoma is a severe non-cancerous lesion of the middle ear characterized by massive inflammation, tissue destruction, and an abnormal growth of keratinized squamous epithelium. We recently demonstrated the presence of pathogenic stem cells within cholesteatoma tissue, unfortunately their potential roles in regulating disease-specific chronic inflammation remain poorly understood. In the presented study, we utilized our established human in vitro cholesteatoma stem cell model for treatments with lipopolysaccharides (LPS), tumor necrosis factor ? (TNF?), and the TLR4-antagonist LPS from R. sphaeroides (LPS-RS) followed by qPCR, western blot, and immunocytochemistry. Middle ear cholesteatoma stem cells (ME-CSCs) showed a significantly increased expression of TLR4 accompanied by a significantly enhanced LPS-dependent pro-inflammatory gene expression pattern of TNF?, IL-1?, IL-1ß, IL-6, and IL-8 compared to non-pathogenic control cells. LPS-dependent pro-inflammatory gene expression in ME-CSCs was driven by an enhanced activity of NF-B p65 leading to a TNF?-mediated feed-forward-loop of pro-inflammatory NF-B target gene expression. Functional inactivation of TLR4 via the TLR4-antagonist LPS-RS blocked chronic inflammation in ME-CSCs, resulting in a nearly complete loss of IL-1ß, IL-6, and TNF? expression. In summary, we determined that ME-CSCs mediate the inflammatory environment of cholesteatoma via TLR4-mediated NF-B-signaling, suggesting a distinct role of ME-CSCs as drivers of cholesteatoma progression and TLR4 on ME-CSCs as a therapeutic target.

SUBMITTER: Schurmann M

PROVIDER: S-EPMC7017370 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Stem Cell-Induced Inflammation in Cholesteatoma is Inhibited by the TLR4 Antagonist LPS-RS.

Schürmann Matthias M Greiner Johannes F W JFW Volland-Thurn Verena V Oppel Felix F Kaltschmidt Christian C Sudhoff Holger H Kaltschmidt Barbara B

Cells 20200114 1

Cholesteatoma is a severe non-cancerous lesion of the middle ear characterized by massive inflammation, tissue destruction, and an abnormal growth of keratinized squamous epithelium. We recently demonstrated the presence of pathogenic stem cells within cholesteatoma tissue, unfortunately their potential roles in regulating disease-specific chronic inflammation remain poorly understood. In the presented study, we utilized our established human in vitro cholesteatoma stem cell model for treatments ...[more]

PMID: 31947538

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Project description:Pneumonia is an inflammatory disease that occurs in the lungs associated with pathogens or other factors. It has been well established that long noncoding RNA X inactivate-specific transcript (XIST) is involved in several cancers. The present study focused on the effect and detailed mechanism of XIST in lipopolysaccharide (LPS)-induced injury in pneumonia. Here, XIST was silenced by transfection with XIST-targeted siRNA, and then, mRNA expression, cell viability, apoptosis, and protein expression were, respectively, assessed by qRT-PCR, CCK-8, flow cytometry, and Western blotting. Luciferase reporter, RIP, and RNA pull-down assays were used to detect the combination of miR-370-3p and XIST. Besides, the tested proinflammatory factors were analysed by qRT-PCR and Western blot, and their productions were quantified by ELISA. The results showed that XIST expression was robustly increased in serum of patients with acute-stage pneumonia and LPS-induced WI-38 human lung fibroblasts cells. Functional analyses demonstrated that knockdown of XIST remarkably alleviated LPS-induced cell injury through increasing cell viability and inhibiting apoptosis and inflammatory cytokine levels. Mechanistically, XIST functioned as a competitive endogenous RNA (ceRNA) by effectively binding to miR-370-3p and then restoring TLR4 expression. More importantly, miR-370-3p inhibitor abolished the function of XIST knockdown on cell injury and JAK/STAT and NF-?B pathways. Taken together, XIST may be involved in progression of cell inflammatory response, and XIST/miR-370-3p/TLR4 axis thus may shed light on the development of novel therapeutics to the treatment of acute stage of pneumonia. SIGNIFICANCE OF THE STUDY: Our study demonstrated that XIST was highly expressed in patients with acute stage of pneumonia. Knockdown of XIST remarkably alleviated LPS-induced cell injury through increasing cell viability and inhibiting apoptosis and inflammatory cytokine levels through regulating JAK/STAT and NF-?B pathways.

Dataset Information

Stem Cell-Induced Inflammation in Cholesteatoma is Inhibited by the TLR4 Antagonist LPS-RS.

Publications

Stem Cell-Induced Inflammation in Cholesteatoma is Inhibited by the TLR4 Antagonist LPS-RS.

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