| 1. |
- Aidoukovitch, Alexandra, et al.
(författare)
-
Desquamated Epithelial Cells of Unstimulated Human Whole Saliva Express Both EGF Transcript and Protein
- 2022
-
Ingår i: International Journal of Dentistry. - : Hindawi Limited. - 1687-8728 .- 1687-8736. ; 2022, s. 1-9
-
Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVE: The aim of this study was to investigate if desquamated oral epithelial cells (DOECs) express the epidermal growth factor (EGF) and if these cells thereby may contribute to salivary EGF contents.BACKGROUND: DOECs have recently been shown to harbor the antimicrobial peptide LL-37, proposing that they may also store other biologically important salivary peptides/proteins. The EGF peptide is a growth factor which plays a critical role to maintain epithelial integrity and promote epithelial healing. The EGF is produced by salivary glands, but it is not known whether DOECs contain the EGF and thereby contribute to salivary EGF levels.MATERIALS AND METHODS: DOECs were isolated from unstimulated whole saliva collected from four healthy volunteers. EGF protein expression was determined in cell lysates by dot blot and ELISA. Cellular distribution of cytokeratin, the proliferation marker Ki67, and EGF immunoreactivity were assessed by immunocytochemistry. EGF gene expression was investigated by qPCR. Expression of EGF transcript and protein in DOECs was compared to that in the human cultured keratinocyte cell line (HaCaT) cells.RESULTS: EGF protein expression was detected in DOEC cell lysates by both dot blot and ELISA. Strong cytoplasmic EGF immunoreactivity was observed in DOECs, although some cells showed only a weak immunoreactive signal for EGF. Moreover, DOECs, besides containing EGF protein, also expressed transcript for EGF. Interestingly, ELISA analysis revealed that EGF protein contents were higher in DOECs than in HaCaT cells. ELISA analysis also disclosed that EGF concentration was about 10 times higher in whole saliva compared to DOECs. EGF transcript expression was about 50% lower in HaCaT cells stimulated with high (10%) compared to low (0.1%) concentration of fetal bovine serum, representing growth-stimulated and growth-restricted conditions, respectively, implying that growth-stimulus exerts negative feedback on EGF gene activity in HaCaT cells.CONCLUSION: Here, we show for the first time that DOECs express the EGF, arguing that these cells contribute to salivary EGF contents and hence may play a role in gingival epithelial repair and wound healing.
|
|
| 2. |
- Bodahl, Sara
(författare)
-
Effects of the host defense peptide LL-37 on human cells : Immunomodulation and cytotoxicity
- 2022
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The human host defense peptide LL-37 has an essential role in the first line of defense against invading pathogens. This cathelicidin is mainly produced by immune cells and epithelial cells aligning the mucosal areas and is normally upregulated upon infection and inflammation. LL-37 displays direct antimicrobial activity against a variety of pathogens, but also modulates the immune responses by acting as a chemoattractant, influencing the production of inflammatory mediators and by affecting immune receptor signaling. Abnormally high local levels of LL-37 have been linked to inflammatory diseases, such as psoriasis, atherosclerosis, periodontitis and asthma. However, little is known about the role of LL-37 in the development and progression of inflammatory and autoimmune diseases and more research is needed to clarify and establish how LL-37 mediates host cell functions in these conditions. In this thesis we have studied LL-37-induced cell toxicity, cell membrane permeability and immune receptor signaling in several types of human cells. We show that LL-37 reduces cell viability in a dose-dependent manner and permeabilizes the cell membranes in all human cell types studied. Interestingly, we demonstrate that LL-37 reduces the cell viability and generates cell membrane permeabilization in osteoblast-like cells even though LL-37 import via clathrin-mediated endocytosis is prevented. In mast cells, LL-37-induced cell membrane permeabilization results in release of both cytosolic and nucleic components, indicating that LL-37 permeabilizes cellular compartments such as the nucleus. Furthermore, we show that LL-37 influences TLR3 signaling in both coronary artery smooth muscle cells and bronchial epithelial cells. More specifically, LL-37 enhances viral dsRNA signaling on TLR3, resulting in upregulation of TLR3 expression and downstream pro-inflammatory signaling that involves the transcription factor NF-κB. Finally, we show that LL-37 induces an increased import of dsRNA in bronchial epithelial cells, suggesting that this mechanism of action is associated with upregulated expression of TLR3. Overall, the work in this thesis provides new insights on underlying mechanisms behind LL-37-induced host cytotoxicity and cell membrane permeability and proposes a novel LL-37-driven pro-inflammatory mechanism of action involving potentiation of dsRNA-stimulated TLR3 expression and signaling
|
|
| 3. |
- Bodahl, Sara, et al.
(författare)
-
LL-37 and Double-Stranded RNA Synergistically Upregulate Bronchial Epithelial TLR3 Involving Enhanced Import of Double-Stranded RNA and Downstream TLR3 Signaling
- 2022
-
Ingår i: Biomedicines. - : MDPI AG. - 2227-9059. ; 10:2
-
Tidskriftsartikel (refereegranskat)abstract
- The human host defense peptide LL-37 influences double-stranded RNA signaling, but this process is not well understood. Here, we investigate synergistic actions of LL-37 and synthetic double-stranded RNA (poly I:C) on toll-like receptor 3 (TLR3) expression and signaling, and examine underlying mechanisms. In bronchial epithelial BEAS-2B cells, LL-37 potentiated poly I:C-induced TLR3 mRNA and protein expression demonstrated by qPCR and Western blot, respectively. Interestingly, these effects were associated with increased uptake of rhodamine-tagged poly I:C visualized by immunocytochemistry. The LL-37/poly I:C-induced upregulation of TLR3 mRNA expression was prevented by the endosomal acidification inhibitor chloroquine, indicating involvement of downstream TLR3 signaling. The glucocorticoid dexamethasone reduced LL-37/poly I:C-induced TLR3 expression on both mRNA and protein levels, and this effect was associated with increased IκBα protein expression, suggesting that dexamethasone acts via attenuation of NF-κB activity. We conclude that LL-37 potentiates poly I:C-induced upregulation of TLR3 through a mechanism that may involve enhanced import of poly I:C and that LL-37/poly I:C-induced TLR3 expression is associated with downstream TLR3 signaling and sensitive to inhibition of NF-κB activity.
|
|
