Molecular signaling of the HMGB1/RAGE axis contributes to cholesteatoma pathogenesis.
Cholesteatoma represents progressive expansion of the keratinizing squamous epithelium in the middle ear with subsequent chronic inflammation in subepithelial connective tissues. The hypothesis was tested that receptor for advanced glycation endproduct (RAGE) and its ligand, high-mobility box 1 (HMGB1), are overexpressed in cholesteatoma, and the RAGE/HMGB1 axis might contribute to its pathogenesis. Cholesteatoma samples (n = 36) and 27 normal skin specimens were studied by immunohistochemistry (IHC) for HMGB1 and RAGE expression. Effects of HMGB1 signaling on proliferation, migration, cytokine production, and apoptosis of human immortalized keratinocytes (HaCaTs) and normal keratinocytes were studied by quantitative reverse transcription (qRT)-PCR, IHC, Western blots, and flow cytometry after cell co-incubation with HMGB1. While all studied tissues expressed HMGB1, its expression was higher in cholesteatoma than in normal skin (p < 0.0001). All cases of cholesteatoma also showed elevated RAGE expression levels, and only 7/27 (26 %) of normal skin specimens were weakly positive for RAGE. Proliferation and migration of HaCaT cells incubated with HMGB1 were up-regulated (p < 0.05). HMGB1 also prevented HaCaT cell apoptosis and induced activation of several molecular signaling pathways in keratinocytes. The data suggest that in cholesteatoma, HMGB1 released from stressed or necrotic epithelial cells and binding to RAGE overexpressed in keratinocytes initiates molecular signaling that culminates in pro-inflammatory cytokine release and chronic inflammation.


CONCLUSIONS
HMGB1 signaling engages multiple activation pathways in RAGE-positive keratinocytes. HMGB1 protects RAGE-positive keratinocytes from drug-induced apoptosis. Keratinocyte proliferation is controlled via RAGE and HMGB1 molecular signaling. Molecular signaling of the HMGB1/RAGE axis contributes to cholesteatoma pathogenesis.
Department of Otorhinolaryngology, Faculty of Medicine and Dentistry, Medical University of Warsaw, Warsaw, Poland.
Miroslaw J
MJ
Szczepanski
Michal
M
Luczak
Ewa
E
Olszewska
Marta
M
Molinska-Glura
Mariola
M
Zagor
Antoni
A
Krzeski
Henryk
H
Skarzynski
Jan
J
Misiak
Karolina
K
Dzaman
Mikolaj
M
Bilusiak
Tomasz
T
Kopec
Malgorzata
M
Leszczynska
Henryk
H
Witmanowski
Theresa L
TL
Whiteside
D000293:Adolescent; D000328:Adult; D000368:Aged; D002460:Cell Line; D018424:Cholesteatoma, Middle Ear / Q000378:metabolism* / Q000473:pathology; D005260:Female; D015870:Gene Expression; D024243:HMGB1 Protein / Q000378:metabolism*; D006801:Humans; D015603:Keratinocytes; D008297:Male; D008875:Middle Aged; D009061:Mouth Mucosa / Q000378:metabolism / Q000473:pathology; D000067759:Receptor for Advanced Glycation End Products / Q000378:metabolism*; D015398:Signal Transduction; D012867:Skin / Q000378:metabolism; D055815:Young Adult
C000593871:AGER protein, human; D024243:HMGB1 Protein; C585491:HMGB1 protein, human; D000067759:Receptor for Advanced Glycation End Products