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The role of C/EBPβ phosphorylation in modulating membrane phospholipids repairing in LPS-induced human lung/bronchial epithelial cells
- Source :
- Gene
- Publication Year :
- 2017
- Publisher :
- Elsevier BV, 2017.
-
Abstract
- Acute lung injury/acute respiratory distress syndrome (ALI/ARDS) is a common critical emergency with high mortality in clinical practice. The key mechanism of ALI/ARDS is that the excessive inflammatory response damages the integrity of alveolar and bronchial cell membrane and thus affects their basic function. Phospholipids are the main component of cell membranes. Phospholipase A2 (PLA2), which catalyzes the cleavage of membrane phospholipids, is the most important inflammatory mediator of ALI. However, clara cell secretory protein 1 (CCSP1), an endogenous PLA2 inhibitor can increase the self-defense of membrane phospholipids. Thus, CCSP1 up-regulation and PLA2 inhibition constitutes an effective method for ensuring the stability of membrane phospholipids and for the treatment of ALI/ARDS. In the present study, we developed an in vitro model of ALI via lipopolysaccharide (LPS) stimulation of a human bronchial epithelial cell line, BEAS-2B, and assessed the mRNA and protein levels of CCSP1 and PLA2 in the model cells. The results demonstrated LPS induction inhibited the transcription and protein expression of CCSP1, but only the protein level of membrane associated PLA2 was increased, suggesting that in the in vitro ALI model, abnormally regulated CCSP1 transcription plays a crucial role in the damage of cell membrane. To find out the reason that CCSP1 expression was decreased in the ALI model, we predicted, by means of bioinformatics, putative transcription factors which would bind to CCSP1 promoter, examined their background and expression, and found that a transcription factor, CCAAT/enhancer binding protein β (C/EBP β), was correlated with the transcription of CCSP1 in the in vitro ALI model, and its phosphorylation in the model was decreased. CHIP-PCR and luciferase reporter assay revealed that C/EBP β bound to CCSP1 promoter and facilitated its transcription. Therefore, we conclude that there is a C/EBP β/CCSP1/PLA2 pathway in the in vitro ALI model. The study of underlying mechanism show that the activity of C/EBP β depends on its phosphorylation:LPS stimulation reduced C/EBP β phosphorylation and suppressed the transcription of CCSP1 in BEAS-2B cells, which resulted in enhanced PLA2 and the consequent membrane damage. And further study shows that overexpression of CDK2(Cyclindependent kinase 2), promoted the phosphorylation of C/EBP β and inhibited PLA2 through the C/EBP β/CCSP1/PLA2 pathway, so as to attenuate membrane damage. The significance of this study lies in that artificial C/EBP β phosphorylation regulation may ease the membrane damage in ALI and improve membrane repair.<br />Highlights • CDK2 over-expression promotes C/EBPβ phosphorylation and improves membrane repair through C/EBPβ/CCSP/PLA2 pathway in ALI.
- Subjects :
- Lipopolysaccharides
0301 basic medicine
ALI, acute lung injury
C/EBP β
Cell
AA, arachidonic acid
ECL, enhanced chemiluminescence
Cell membrane
LSD, Least Significant Difference
PCR, polymerase chain reaction
0302 clinical medicine
Enhancer binding
Uteroglobin
Phosphorylation
Promoter Regions, Genetic
Lung
Phospholipids
MOI, multiplicity of infection
GFP, green fluorescent protein
TNF, tumor necrosis factor
biology
Kinase
General Medicine
respiratory system
Cell biology
medicine.anatomical_structure
Biochemistry
030220 oncology & carcinogenesis
CDS, coding DNA sequence
LPS, lipopolysaccharide
lipids (amino acids, peptides, and proteins)
CCSP, clara cell secretory protein
CDK2
Acute Lung Injury
PLA2, Phospholipase A2
Lung injury
Article
Cell Line
C/EBP β, CCAAT/enhancer binding protein β
03 medical and health sciences
FBS, fetal bovine serum
CCSP1
ELISA, enzyme linked immunosorbent assay
Genetics
medicine
Humans
SARS, severe acute respiratory syndrome
Transcription factor
ARDS, acute respiratory distress syndrome
PAF, platelet activating factor
PLA2
CCAAT-Enhancer-Binding Protein-beta
Cyclin-dependent kinase 2
Epithelial Cells
BCA, bicinchoninic acid
IL, interleukin
respiratory tract diseases
Phospholipases A2
CDK2, Cyclindependent kinase 2
ALI
030104 developmental biology
DMEM, Dulbecco's modified eagle medium
biology.protein
ARDS
Subjects
Details
- ISSN :
- 03781119
- Volume :
- 629
- Database :
- OpenAIRE
- Journal :
- Gene
- Accession number :
- edsair.doi.dedup.....50c2795631004e015dd2fdd48f2a4c5d