Dysregulation of CLOCK gene expression in hyperoxia-induced lung injury.
Hyperoxic acute lung injury (HALI) is characterized by inflammation and epithelial cell death. CLOCK genes are master regulators of circadian rhythm also implicated in inflammation and lung diseases. However, the relationship of CLOCK genes in hyperoxia-induced lung injury has not been studied. This study will determine if HALI alters CLOCK gene expression. To test this, wild-type and NALP3(-/-) mice were exposed to room air or hyperoxia for 24, 48, or 72 h. In addition, mice were exposed to different concentrations of hyperoxia (50, 75, or 100% O2) or room air for 72 h. The mRNA and protein levels of lung CLOCK genes, based on quantitative PCR and Western blot analysis, respectively, and their target genes are significantly elevated in mice exposed to hyperoxia compared with controls. Alterations in CLOCK genes are associated with increased inflammatory markers in bronchoalveolar lavage fluid of hyperoxic mice compared with controls. Histological examination of mice lungs exposed to hyperoxia show increased inflammation and alveolar congestion compared with controls. Our results indicate sequential increase in CLOCK gene expression in lungs of mice exposed to hyperoxia compared with controls. Additionally, data suggest a dose-dependent increase in CLOCK gene expression with increased oxygen concentrations. To validate if the expression changes related to CLOCK genes are indeed associated with inflammation, NALP3(-/-) was introduced to analyze loss of function in inflammation. Western blot analysis showed significant CLOCK gene downregulation in NALP3(-/-) mice compared with wild-type controls. Together, our results demonstrate that hyperoxia-mediated lung inflammation is associated with alterations in CLOCK gene expression.
Division of Allergy and Immunology, Internal Medicine, Morsani College of Medicine, University of South Florida, Tampa, Florida.
Venu
V
Lagishetty
Division of Allergy and Immunology, Internal Medicine, Morsani College of Medicine, University of South Florida, Tampa, Florida.
Prasanna Tamarapu
PT
Parthasarathy
Division of Allergy and Immunology, Internal Medicine, Morsani College of Medicine, University of South Florida, Tampa, Florida.
Oluwakemi
O
Phillips
Division of Allergy and Immunology, Internal Medicine, Morsani College of Medicine, University of South Florida, Tampa, Florida.
Jutaro
J
Fukumoto
Division of Allergy and Immunology, Internal Medicine, Morsani College of Medicine, University of South Florida, Tampa, Florida.
Young
Y
Cho
Division of Allergy and Immunology, Internal Medicine, Morsani College of Medicine, University of South Florida, Tampa, Florida.
Itsuko
I
Fukumoto
Division of Allergy and Immunology, Internal Medicine, Morsani College of Medicine, University of South Florida, Tampa, Florida.
Huynh
H
Bao
Division of Allergy and Immunology, Internal Medicine, Morsani College of Medicine, University of South Florida, Tampa, Florida.
Ruan
R
Cox
Division of Allergy and Immunology, Internal Medicine, Morsani College of Medicine, University of South Florida, Tampa, Florida.
Lakshmi
L
Galam
Division of Allergy and Immunology, Internal Medicine, Morsani College of Medicine, University of South Florida, Tampa, Florida.
Richard F
RF
Lockey
Division of Allergy and Immunology, Internal Medicine, Morsani College of Medicine, University of South Florida, Tampa, Florida nkollipu@health.usf.edu.
Narasaiah
N
Kolliputi
D000818:Animals; D056926:CLOCK Proteins / Q000096:biosynthesis* / Q000235:genetics; D005786:Gene Expression Regulation; D018496:Hyperoxia / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D055370:Lung Injury / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics
D012333:RNA, Messenger; D056926:CLOCK Proteins; C538944:Clock protein, mouse
acute lung injury; circadian rhythms; hyperoxia; inflammation