The host defense peptide LL-37 is internalized by human periodontal ligament cells and prevents LPS-induced MCP-1 production
Aidoukovitch, Alexandra LU ; Anders, Emma LU ; Dahl, Sara LU
; Nebel, Daniel
LU
; Svensson, Daniel
LU
and Nilsson, Bengt Olof
LU
(2019)
In Journal of Periodontal Research
54(6).
p.662-670
- Abstract
Objective: The human host defense peptide LL-37 both shows antimicrobial effects and modulates host cell properties. Here, we assess the effects of synthesized LL-37 on lipopolysaccharide (LPS)-induced inflammation in human periodontal ligament (PDL) cells and investigates underlying mechanisms. Background: LL-37 has been detected in the periodontal tissues, but its functional importance for PDL cell innate immune responses is not known. Methods: Human PDL cells were obtained from premolars extracted on orthodontic indications. Cellular pro-inflammatory monocyte chemoattractant protein-1 (MCP-1) mRNA expression was determined using quantitative real-time RT-PCR. MCP-1 protein production was assessed by western blot and ELISA.... (More)
Objective: The human host defense peptide LL-37 both shows antimicrobial effects and modulates host cell properties. Here, we assess the effects of synthesized LL-37 on lipopolysaccharide (LPS)-induced inflammation in human periodontal ligament (PDL) cells and investigates underlying mechanisms. Background: LL-37 has been detected in the periodontal tissues, but its functional importance for PDL cell innate immune responses is not known. Methods: Human PDL cells were obtained from premolars extracted on orthodontic indications. Cellular pro-inflammatory monocyte chemoattractant protein-1 (MCP-1) mRNA expression was determined using quantitative real-time RT-PCR. MCP-1 protein production was assessed by western blot and ELISA. Internalization of LL-37 by PDL cells was visualized by immunocytochemistry. Nuclear factor kappa-light-chain-enhancer of activated B-cell (NF-κB) activity was assessed by western blot of phosphorylated p65, phosphorylated p105, and IκBα proteins. Binding of LL-37 to PDL cell DNA was determined by isolation and purification of DNA and dot blot for LL-37 immunoreactivity. Results: Treatment with LL-37 (1 µmol/L) for 24 hours prevented LPS-induced stimulation of MCP-1 expression analyzed both on transcript and on protein levels. Stimulation with LL-37 (1 µmol/L) for 24 hours had no effect on toll-like receptor (TLR)2 and TLR4 transcript expression, suggesting that LL-37 acts downstream of the TLRs. Preincubation with LL-37 for 60 minutes followed by stimulation with LPS for 24 hours in the absence of LL-37 completely prevented LPS-evoked MCP-1 transcript expression, implying that LL-37 acts intracellularly and not via binding and neutralization of LPS. In PDL cells stimulated with LL-37 for 60 minutes, the peptide was internalized as demonstrated by immunocytochemistry, suggesting an intracellular mechanism of action. LL-37 immunoreactivity was observed both in the cytosol and in the nucleus. Downregulation of LPS-induced MCP-1 by LL-37 was not mediated by reduction in NF-κB activity as shown by unaltered expression of phosphorylated p65, phosphorylated p105, and IκBα NF-κB proteins in the presence of LL-37. Immunoreactivity for LL-37 was observed in PDL cell DNA treated with but not without 0.1 and 1 µmol/L LL-37 for 60 minutes in vitro. Conclusion: LL-37 abolishes LPS-induced MCP-1 production in human PDL cells through an intracellular, NF-κB-independent mechanism which probably involves direct interaction between LL-37 and DNA.
(Less)
- author
- Aidoukovitch, Alexandra
LU
; Anders, Emma
LU
; Dahl, Sara
LU
; Nebel, Daniel
LU
; Svensson, Daniel
LU
and Nilsson, Bengt Olof
LU
- organization
- publishing date
- 2019-12
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- antimicrobial peptide, inflammation, innate immunity, NF-κB
- in
- Journal of Periodontal Research
- volume
- 54
- issue
- 6
- pages
- 9 pages
- publisher
- Wiley-Blackwell
- external identifiers
-
- pmid:31095741
- scopus:85066017963
- ISSN
- 0022-3484
- DOI
- 10.1111/jre.12667
- project
- Production and transport of the antimicrobial peptide LL-37 and other peptides in saliva: physiological and pathophysiological importance
- language
- English
- LU publication?
- yes
- id
- abfcf6b2-54a5-425a-9f90-13cc51307ae0
- date added to LUP
- 2019-06-14 12:29:25
- date last changed
- 2024-04-16 10:38:01
@article{abfcf6b2-54a5-425a-9f90-13cc51307ae0,
abstract = {{<p>Objective: The human host defense peptide LL-37 both shows antimicrobial effects and modulates host cell properties. Here, we assess the effects of synthesized LL-37 on lipopolysaccharide (LPS)-induced inflammation in human periodontal ligament (PDL) cells and investigates underlying mechanisms. Background: LL-37 has been detected in the periodontal tissues, but its functional importance for PDL cell innate immune responses is not known. Methods: Human PDL cells were obtained from premolars extracted on orthodontic indications. Cellular pro-inflammatory monocyte chemoattractant protein-1 (MCP-1) mRNA expression was determined using quantitative real-time RT-PCR. MCP-1 protein production was assessed by western blot and ELISA. Internalization of LL-37 by PDL cells was visualized by immunocytochemistry. Nuclear factor kappa-light-chain-enhancer of activated B-cell (NF-κB) activity was assessed by western blot of phosphorylated p65, phosphorylated p105, and IκBα proteins. Binding of LL-37 to PDL cell DNA was determined by isolation and purification of DNA and dot blot for LL-37 immunoreactivity. Results: Treatment with LL-37 (1 µmol/L) for 24 hours prevented LPS-induced stimulation of MCP-1 expression analyzed both on transcript and on protein levels. Stimulation with LL-37 (1 µmol/L) for 24 hours had no effect on toll-like receptor (TLR)2 and TLR4 transcript expression, suggesting that LL-37 acts downstream of the TLRs. Preincubation with LL-37 for 60 minutes followed by stimulation with LPS for 24 hours in the absence of LL-37 completely prevented LPS-evoked MCP-1 transcript expression, implying that LL-37 acts intracellularly and not via binding and neutralization of LPS. In PDL cells stimulated with LL-37 for 60 minutes, the peptide was internalized as demonstrated by immunocytochemistry, suggesting an intracellular mechanism of action. LL-37 immunoreactivity was observed both in the cytosol and in the nucleus. Downregulation of LPS-induced MCP-1 by LL-37 was not mediated by reduction in NF-κB activity as shown by unaltered expression of phosphorylated p65, phosphorylated p105, and IκBα NF-κB proteins in the presence of LL-37. Immunoreactivity for LL-37 was observed in PDL cell DNA treated with but not without 0.1 and 1 µmol/L LL-37 for 60 minutes in vitro. Conclusion: LL-37 abolishes LPS-induced MCP-1 production in human PDL cells through an intracellular, NF-κB-independent mechanism which probably involves direct interaction between LL-37 and DNA.</p>}},
author = {{Aidoukovitch, Alexandra and Anders, Emma and Dahl, Sara and Nebel, Daniel and Svensson, Daniel and Nilsson, Bengt Olof}},
issn = {{0022-3484}},
keywords = {{antimicrobial peptide; inflammation; innate immunity; NF-κB}},
language = {{eng}},
number = {{6}},
pages = {{662--670}},
publisher = {{Wiley-Blackwell}},
series = {{Journal of Periodontal Research}},
title = {{The host defense peptide LL-37 is internalized by human periodontal ligament cells and prevents LPS-induced MCP-1 production}},
url = {{http://dx.doi.org/10.1111/jre.12667}},
doi = {{10.1111/jre.12667}},
volume = {{54}},
year = {{2019}},
}