scholarly journals Stem Cell-Induced Inflammation in Cholesteatoma Is Inhibited by the TLR4 Antagonist LPS-RS

Cells ◽  
2020 ◽  
Vol 9 (1) ◽  
pp. 199 ◽  
Author(s):  
Matthias Schürmann ◽  
Johannes F. W. Greiner ◽  
Verena Volland-Thurn ◽  
Felix Oppel ◽  
Christian Kaltschmidt ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Stem Cell ◽  
Chronic Inflammation ◽  
Middle Ear ◽  
Gene Expression Pattern ◽  
Inflammatory Gene Expression ◽  
Inflammatory Gene ◽  
Middle Ear Cholesteatoma ◽  
Tlr4 Antagonist

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 α (TNFα), 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 TNFα, IL-1α, 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-κB p65 leading to a TNFα-mediated feed-forward-loop of pro-inflammatory NF-κB 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 TNFα expression. In summary, we determined that ME-CSCs mediate the inflammatory environment of cholesteatoma via TLR4-mediated NF-κB-signaling, suggesting a distinct role of ME-CSCs as drivers of cholesteatoma progression and TLR4 on ME-CSCs as a therapeutic target.

scholarly journals Chronic inflammation of middle ear cholesteatoma promotes its recurrence via a paracrine mechanism

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Matthias Schürmann ◽  
Felix Oppel ◽  
Senyao Shao ◽  
Verena Volland-Thurn ◽  
Christian Kaltschmidt ◽  
...  
Keyword(s):  
Stem Cells ◽  
Middle Ear ◽  
Culture Conditions ◽  
Epidermal Differentiation ◽  
Epidermal Stem Cells ◽  
Middle Ear Cholesteatoma ◽  
Surgical Extraction ◽  
Standard Culture ◽  
Lps Treatment ◽  
Tlr4 Antagonist

Abstract Background Cholesteatoma disease is an expanding lesion in the middle ear. Hearing loss and facial paralysis alongside with other intracranial complications are found. No pharmaceutical treatment is available today and recurrence after surgical extraction occurs. We investigated possible TLR4-based mechanisms promoting recurrence and explore possible treatments strategies. Methods We isolated fibroblasts and epidermal stem cells from cholesteatoma tissue and healthy auditory canal skin. Subsequently, their expression under standard culture conditions and after stimulation with LPS was investigated by RT-qPCR. Cell metabolism and proliferation were analysed upon LPS treatment, with and without TLR4 antagonist. An indirect co-culture of fibroblasts and epidermal stem cells isolated from cholesteatoma tissue was utilized to monitor epidermal differentiation upon LPS treatment by RT-qPCR and immunocytochemistry. Results Under standard culture conditions, we detected a tissue-independent higher expression of IL-1β and IL-8 in stem cells, an upregulation of KGF and IGF-2 in both cell types derived from cholesteatoma and higher expression of TLR4 in stem cells derived from cholesteatoma tissue. Upon LPS challenge, we could detect a significantly higher expression of IL-1α, IL-1β, IL-6 and IL-8 in stem cells and of TNF-a, GM-CSF and CXCL-5 in stem cells and fibroblasts derived from cholesteatoma. The expression of the growth factors KGF, EGF, EREG, IGF-2 and HGF was significantly higher in fibroblasts, particularly when derived from cholesteatoma. Upon treatment with LPS the metabolism was elevated in stem cells and fibroblasts, proliferation was only enhanced in fibroblasts derived from cholesteatoma. This could be reversed by the treatment with a TLR4 antagonist. The cholesteatoma fibroblasts could be triggered by LPS to promote the epidermal differentiation of the stem cells, while no LPS treatment or LPS treatment without the presence of fibroblasts did not result in such a differentiation. Conclusion We propose that cholesteatoma recurrence is based on TLR4 signalling imprinted in the cholesteatoma cells. It induces excessive inflammation of stem cells and fibroblasts, proliferation of perimatrix fibroblasts and the generation of epidermal cells from stem cells thru paracrine signalling by fibroblasts. Treatment of the operation site with a TLR4 antagonist might reduce the chance of cholesteatoma recurrence.

scholarly journals Chronic inflammation of middle ear cholesteatoma promotes its recurrence via a paracrine mechanism

2020 ◽  
Author(s):  
Matthias Schuermann ◽  
Felix Oppel ◽  
Senyao Shao ◽  
Verena Volland-Thurn ◽  
Christian Kaltschmidt ◽  
...  
Keyword(s):  
Stem Cells ◽  
Middle Ear ◽  
Culture Conditions ◽  
Epidermal Differentiation ◽  
Epidermal Stem Cells ◽  
Middle Ear Cholesteatoma ◽  
Surgical Extraction ◽  
Standard Culture ◽  
Lps Treatment ◽  
Tlr4 Antagonist

Abstract BackgroundCholesteatoma disease is an expanding lesion in the middle ear. Hearing loss and facial paralysis alongside with other intracranial complications are found. No pharmaceutical treatment is available today and recurrence after surgical extraction occurs. We investigated possible TLR4-based mechanisms promoting recurrence and explore possible treatments strategies.MethodsWe isolated fibroblasts and epidermal stem cells from cholesteatoma tissue and healthy auditory canal skin. Subsequently, their expression under standard culture conditions and after stimulation with LPS was investigated by RT-qPCR. Cell metabolism and proliferation were analysed upon LPS treatment, with and without TLR4 antagonist. An indirect co-culture of fibroblasts and epidermal stem cells isolated from cholesteatoma tissue was utilized to monitor epidermal differentiation upon LPS treatment by RT-qPCR and immunocytochemistry.ResultsUnder standard culture conditions, we detected a tissue-independent higher expression of IL-1β and IL-8 in stem cells, an upregulation of KGF and IGF-2 in both cell types derived from cholesteatoma and higher expression of TLR4 in stem cells derived from cholesteatoma tissue. Upon LPS challenge, we could detect a significantly higher expression of IL-1α, IL-1β, IL-6 and IL-8 in stem cells and of TNF-a, GM-CSF and CXCL-5 in stem cells and fibroblasts derived from cholesteatoma. The expression of the growth factors KGF, EGF, EREG, IGF-2 and HGF was significantly higher in fibroblasts, particularly when derived from cholesteatoma. Upon treatment with LPS the metabolism was elevated in stem cells and fibroblasts, proliferation was only enhanced in fibroblasts derived from cholesteatoma. This could be reversed by the treatment with a TLR4 antagonist. The cholesteatoma fibroblasts could be triggered by LPS to promote the epidermal differentiation of the stem cells, while no LPS treatment or LPS treatment without the presence of fibroblasts did not result in such a differentiation.ConclusionWe propose that cholesteatoma recurrence is based on TLR4 signalling imprinted in the cholesteatoma cells. It induces excessive inflammation of stem cells and fibroblasts, proliferation of perimatrix fibroblasts and the generation of epidermal cells from stem cells thru paracrine signalling by fibroblasts. Treatment of the operation site with a TLR4 antagonist might reduce the chance of cholesteatoma recurrence.

scholarly journals Pro-inflammatory gene expression in solid glioblastoma microenvironment and in hypoxic stem cells from human glioblastoma

2011 ◽  
Vol 8 (1) ◽  
pp. 32 ◽  
Author(s):  
Marco Tafani ◽  
Maura Di Vito ◽  
Alessandro Frati ◽  
Laura Pellegrini ◽  
Elena De Santis ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Inflammatory Gene Expression ◽  
Human Glioblastoma ◽  
Inflammatory Gene

The Severity of Spinal Cord Injury Determines the Inflammatory Gene Expression Pattern after Immunization with Neural-Derived Peptides

2018 ◽  
Vol 65 (2) ◽  
pp. 190-195 ◽  
Author(s):  
Elisa García ◽  
Raúl Silva-García ◽  
Adrian Flores-Romero ◽  
Liliana Blancas-Espinoza ◽  
Roxana Rodríguez-Barrera ◽  
...  
Keyword(s):  
Gene Expression ◽  
Spinal Cord ◽  
Spinal Cord Injury ◽  
Expression Pattern ◽  
Gene Expression Pattern ◽  
Inflammatory Gene Expression ◽  
Inflammatory Gene ◽  
Cord Injury

scholarly journals T9. EPIGENETIC PROFILING IN SCHIZOPHRENIA DERIVED HUMAN INDUCED PLURIPOTENT STEM CELLS (HIPSCS) AND NEURONS

2020 ◽  
Vol 46 (Supplement_1) ◽  
pp. S234-S234
Author(s):  
Lorna Farrelly ◽  
Shuangping Zhang ◽  
Erin Flaherty ◽  
Aaron Topol ◽  
Nadine Schrode ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mass Spectrometry ◽  
Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Neural Progenitor Cells ◽  
Gene Expression Pattern ◽  
Early Gene ◽  
Label Free ◽  
Induced Pluripotent

Abstract Background Schizophrenia (SCZ) is a severe psychiatric disorder affecting ~1% of the world’s population. It is largely heritable with genetic risk reflected by a combination of common variants of small effect and highly penetrant rare mutations. Chromatin modifications are known to play critical roles in the mediation of many neurodevelopmental processes, and, when disturbed, may also contribute to the precipitation of psychiatric disorders, such as SCZ. While a handful of candidate-based studies have measured changes in promoter-bound histone modifications, few mechanistic studies have been carried out to explore how these modifications may affect chromatin to precipitate behavioral phenotypes associated with the disease. Methods We applied an unbiased proteomics approach to evaluate the epigenetic landscape of SCZ in human induced pluripotent stem cells (hiPSC), neural progenitor cells (NPCs) and neurons from SCZ patients vs. matched controls. We utilized proteomics-based, label free liquid chromatography mass spectrometry (LC-MS/MS) on purified histones from these cells and confirmed our results by western blotting in postmortem SCZ cortical brain tissues. Furthermore we validated our findings with the application of histone interaction assays and structural and biophysical assessments to identify and confirm novel chromatin ‘readers’. To relate our findings to a SCZ phenotype we used a SCZ rodent model of prepulse inhibition (PPI) to perform pharmacological manipulations and behavioral assessments. Results Using label free mass spectrometry we performed PTM screening of hiPSCs, NPCs and matured neurons derived from SCZ patients and matched controls. We identified, amongst others, altered patterns of hyperacetylation in SCZ neurons. Additionally we identified enhanced binding of particular acetylation ‘reader’ proteins. Pharmacological inhibition of such proteins in an animal model of amphetamine sensitization ameliorated PPI deficits further validating this epigenetic signature in SCZ. Discussion Recent evidence indicates that relevance and patterns of acetylation in epigenetics advances beyond its role in transcription and small molecule inhibitors of these aberrant interactions hold promise as useful therapeutics. This study identifies a role for modulating gene expression changes associated with a SCZ epigenetic signature and warrants further investigation in terms of how this early gene expression pattern perhaps determines susceptibility or severity of the SCZ disease trajectory.

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