| 1. |
- Moberg, Christina, et al.
(författare)
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De unga gör helt rätt när de stämmer staten
- 2022
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Ingår i: Aftonbladet. - 1103-9000.
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Tidskriftsartikel (populärvet., debatt m.m.)abstract
- 1 620 forskare och lärare i forskarvärlden: Vi ställer oss bakom Auroras klimatkrav
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| 2. |
- Bankell, Elisabeth, et al.
(författare)
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LL-37-induced caspase-independent apoptosis is associated with plasma membrane permeabilization in human osteoblast-like cells
- 2021
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Ingår i: Peptides. - : Elsevier BV. - 0196-9781 .- 1873-5169. ; 135
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Tidskriftsartikel (refereegranskat)abstract
- The host defense peptide LL-37 is active against both gram-positive and gram-negative bacteria, but it has also been shown to reduce human host cell viability. However, the mechanisms behind LL-37-induced human host cell cytotoxicity are not yet fully understood. Here, we assess if LL-37-evoked attenuation of human osteoblast-like MG63 cell viability is associated with apoptosis, and if the underlying mechanism may involve LL-37-induced plasma membrane permeabilization. MG63 cell viability and plasma membrane permeabilization were investigated by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method and by measuring lactate dehydrogenase (LDH) release, respectively. Apoptosis was assessed by the terminal deoxynucleotidyl dUTP nick end labeling (TUNEL) assay and Annexin V flow cytometry, and caspase-3 and poly (ADP-ribose) polymerase (PARP) cleavage were determined by Western blot. LL-37 (4 and 10 μM) reduced both cell number and cell viability, and these effects were associated with a pro-apoptotic effect demonstrated by positive TUNEL staining and Annexin V flow cytometry. LL-37-induced apoptosis was not coupled to either caspase-3 or PARP cleavage, suggesting that LL-37 causes caspase-independent apoptosis in MG63 cells. Both LL-37 and the well-known plasma membrane permeabilizer Triton X-100 reduced cell viability and stimulated LDH release. Triton X-100-treated cells showed positive TUNEL staining, and the detergent accumulated cells in late apoptosis/necrosis. Similar to LL-37, Triton X-100 caused no PARP cleavage. We conclude that LL-37 promotes caspase-independent apoptosis, and that this effect seems coupled to plasma membrane permeabilization in human MG63 cells.
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| 3. |
- Aidoukovitch, Alexandra, et al.
(författare)
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Antimicrobial peptide LL-37 and its pro-form, hCAP18, in desquamated epithelial cells of human whole saliva
- 2020
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Ingår i: European Journal of Oral Sciences. - : Wiley. - 0909-8836 .- 1600-0722. ; 128:1, s. 1-6
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Tidskriftsartikel (refereegranskat)abstract
- The antimicrobial peptide LL-37 is active against oral bacteria and has been demonstrated to be present in human saliva, but its distribution in different fractions of saliva is not known. LL-37 is formed from its intracellular pro-form, hCAP18, in an extracellular enzymatic reaction catalyzed by proteinase 3 and kallikrein 5. Here, we prepared cell-containing and cell-free fractions of unstimulated human whole saliva by centrifugation after depolymerization of mucins with dithiothreitol, and measured the levels of hCAP18/LL-37 in these fractions using ELISA. Cellular expression of hCAP18/LL-37 was determined by western blotting and immunocytochemistry. The ELISA analyses demonstrated that both cells and cell-free saliva contained hCAP18/LL-37. Western blot analysis of cell-pellet homogenates showed a strong band corresponding to hCAP18 at the correct molecular weight and a weak band corresponding to LL-37. Phase-contrast and light microscopy revealed that the cells consisted of desquamated epithelial cells. These cells expressed cytoplasmic immunoreactivity for hCAP18/LL-37. The peripheral part of the cytoplasm, corresponding to the plasma membrane, was particularly rich in hCAP18/LL-37 immunoreactivity. No immunoreactivity was observed after omission of the primary antibody. We conclude that desquamated epithelial cells of human whole saliva contain antimicrobial hCAP18/LL-37, suggesting that these cells may take part in the innate immune system by harboring and releasing these peptides.
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| 4. |
- Aidoukovitch, Alexandra, et al.
(författare)
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Desquamated Epithelial Cells of Unstimulated Human Whole Saliva Express Both EGF Transcript and Protein
- 2022
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Ingår i: International Journal of Dentistry. - : Hindawi Limited. - 1687-8728 .- 1687-8736. ; 2022, s. 1-9
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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.
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| 5. |
- Aidoukovitch, Alexandra, et al.
(författare)
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Exogenous LL-37 but not homogenates of desquamated oral epithelial cells shows activity against Streptococcus mutans
- 2021
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Ingår i: Acta Odontologica Scandinavica. - : Informa UK Limited. - 0001-6357 .- 1502-3850. ; 79:6, s. 466-472
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Tidskriftsartikel (refereegranskat)abstract
- Objective: The antimicrobial peptide hCAP18/LL-37 is detected in desquamated epithelial cells of human whole saliva, but the functional importance of this pool of hCAP18/LL-37 is not understood. Here, we assess the impact of homogenates of desquamated oral epithelial cells and exogenous, synthetic LL-37 on two oral bacteria: S. mutans and S. gordonii. Material and methods: Desquamated epithelial cells of unstimulated whole saliva were isolated and cellular and extracellular levels of hCAP18/LL-37 analyzed by ELISA. Bacterial viability was determined by BacLight Live/Dead staining and confocal laser scanning microscopy. Results: Desquamated oral epithelial cells harboured hCAP18/LL-37, and they spontaneously released/leaked the peptide to their medium. Exogenous, synthetic LL-37 showed cytotoxic activity against S. mutans but not S gordonii, suggesting that LL-37 acts differentially on these two types of oral bacteria. Homogenates of desquamated oral epithelial cells had no effect on S. mutans viability. Treatment with exogenous, synthetic LL-37 (8 and 10 μM) reduced S. mutans viability, whereas lower concentrations (0.1 and 1 µM) of the peptide lacked effect. Conclusions: Desquamated oral epithelial cells contain hCAP18/LL-37, but their cellular levels of hCAP18/LL-37 are too low to affect S. mutans viability, whereas exogenous, synthetic LL-37 has a strong effect on these bacteria.
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| 6. |
- Aidoukovitch, Alexandra, et al.
(författare)
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Vitamin D triggers hCAP18/LL-37 production : Implications for LL-37-induced human osteoblast cytotoxicity
- 2024
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Ingår i: Biochemical and Biophysical Research Communications. - 0006-291X. ; 712-713
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Tidskriftsartikel (refereegranskat)abstract
- The human cathelicidin LL-37 shows activity against microorganisms, but it is also cytotoxic to host cells. The CAMP gene codes for the LL-37 precursor hCAP18 which is processed extracellularly to active LL-37. It has previously been shown that vitamin D stimulates CAMP gene activity, but less information is available demonstrating that vitamin D also can increase hCAP18/LL-37 protein production. Here, we show with RT-qPCR that a physiological concentration of vitamin D (50 nM) enhances CAMP mRNA levels by about 170 times in human THP-1 monocyte cells. Stimulation with 50 nM vitamin D increases hCAP18/LL-37 protein contents 3–4 times in THP-1 cell lysates demonstrated by both dot blot analysis and ELISA applying two different hCAP18/LL-37 antibodies. Treatment with the proteasome inhibitor MG132 enhances hCAP18/LL-37 levels, suggesting that turnover of hCAP18/LL-37 protein is regulated by the proteasome. The hCAP18/LL-37 concentration in vitamin D-stimulated THP-1 cells corresponds to 1.04 μM LL-37. Interestingly, synthetic LL-37, at this concentration, reduces viability of human osteoblast-like MG63 cells, whereas the THP-1 cells are less sensitive as demonstrated by the MTT assay. In summary, we show that vitamin D enhances hCAP18/LL-37 production, and that this effect can be of physiological/pathophysiological relevance for LL-37-induced human osteoblast toxicity.
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| 7. |
- Bankell, Elisabeth, et al.
(författare)
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Suppression of smooth muscle cell inflammation by myocardin-related transcription factors involves inactivation of TANK-binding kinase 1
- 2024
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Ingår i: Scientific Reports. - 2045-2322. ; 14:1
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Tidskriftsartikel (refereegranskat)abstract
- Myocardin-related transcription factors (MRTFs: myocardin/MYOCD, MRTF-A/MRTFA, and MRTF-B/MRTFB) suppress production of pro-inflammatory cytokines and chemokines in human smooth muscle cells (SMCs) through sequestration of RelA in the NF-κB complex, but additional mechanisms are likely involved. The cGAS-STING pathway is activated by double-stranded DNA in the cytosolic compartment and acts through TANK-binding kinase 1 (TBK1) to spark inflammation. The present study tested if MRTFs suppress inflammation also by targeting cGAS-STING signaling. Interrogation of a transcriptomic dataset where myocardin was overexpressed using a panel of 56 cGAS-STING cytokines showed the panel to be repressed. Moreover, MYOCD, MRTFA, and SRF associated negatively with the panel in human arteries. RT-qPCR in human bronchial SMCs showed that all MRTFs reduced pro-inflammatory cytokines on the panel. MRTFs diminished phosphorylation of TBK1, while STING phosphorylation was marginally affected. The TBK1 inhibitor amlexanox, but not the STING inhibitor H-151, reduced the anti-inflammatory effect of MRTF-A. Co-immunoprecipitation and proximity ligation assays supported binding between MRTF-A and TBK1 in SMCs. MRTFs thus appear to suppress cellular inflammation in part by acting on the kinase TBK1. This may defend SMCs against pro-inflammatory insults in disease.
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| 8. |
- Bankell, Elisabeth, et al.
(författare)
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The antimicrobial peptide LL-37 triggers release of apoptosis-inducing factor and shows direct effects on mitochondria
- 2022
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Ingår i: Biochemistry and Biophysics Reports. - : Elsevier BV. - 2405-5808. ; 29
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Tidskriftsartikel (refereegranskat)abstract
- The human antimicrobial peptide LL-37 permeabilizes the plasma membrane of host cells, but LL-37-induced direct effects on mitochondrial membrane permeability and function has not been reported. Here, we demonstrate that LL-37 is rapidly (within 20 min) internalized by human osteoblast-like MG63 cells, and that the peptide co-localizes with MitoTracker arguing for accumulation in mitochondria. Subcellular fractionation and Western blot disclose that stimulation with LL-37 (8 μM) for 2 h triggers release of the mitochondrial protein apoptosis-inducing factor (AIF) to the cytosol, whereas LL-37 causes no release of cytochrome C oxidase subunit IV of the inner mitochondrial membrane, suggesting that LL-37 affects mitochondrial membrane permeability in a specific manner. Next, we investigated release of AIF and cytochrome C from isolated mitochondria by measuring immunoreactivity by dot blot. The media of mitochondria treated with LL-37 (8 μM) for 2 h contained 50% more AIF and three times more cytochrome C than that of control mitochondria, showing that LL-37 promotes release of both AIF and cytochrome C. Moreover, in vesicles reflecting mitochondrial membrane lipid composition, LL-37 stimulates membrane permeabilization and release of tracer molecules. We conclude that LL-37 is rapidly internalized by MG63 cells and accumulates in mitochondria, and that the peptide triggers release of pro-apoptotic AIF and directly affects mitochondrial membrane structural properties.
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| 9. |
- Bodahl, Sara, et al.
(författare)
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LL-37 and Double-Stranded RNA Synergistically Upregulate Bronchial Epithelial TLR3 Involving Enhanced Import of Double-Stranded RNA and Downstream TLR3 Signaling
- 2022
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Ingår i: Biomedicines. - : MDPI AG. - 2227-9059. ; 10:2
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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.
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| 10. |
- Dahl, Sara, et al.
(författare)
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Human host defense peptide LL-37 facilitates double-stranded RNA pro-inflammatory signaling through up-regulation of TLR3 expression in vascular smooth muscle cells
- 2020
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Ingår i: Inflammation Research. - : Springer Science and Business Media LLC. - 1420-908X .- 1023-3830. ; 69:6, s. 579-588
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVE: The importance of human host defense peptide LL-37 in vascular innate immunity is not understood. Here, we assess the impact of LL-37 on double-stranded RNA (dsRNA) signaling in human vascular smooth muscle cells.MATERIALS AND METHODS: Cellular import of LL-37 and synthetic dsRNA (poly I:C) were investigated by immunocytochemistry and fluorescence imaging. Transcript and protein expression were determined by qPCR, ELISA and Western blot. Knockdown of TLR3 was performed by siRNA.RESULTS: LL-37 was rapidly internalized, suggesting that it has intracellular actions. Co-stimulation with poly I:C and LL-37 enhanced pro-inflammatory IL-6 and MCP-1 transcripts several fold compared to treatment with poly I:C or LL-37 alone. Poly I:C increased IL-6 and MCP-1 protein production, and this effect was potentiated by LL-37. LL-37-induced stimulation of poly I:C signaling was not associated with enhanced import of poly I:C. Treatment with poly I:C and LL-37 in combination increased expression of dsRNA receptor TLR3 compared to stimulation with poly I:C or LL-37 alone. In TLR3 knockdown cells, treatment with poly I:C and LL-37 in combination had no effect on IL-6 and MCP-1 expression, showing loss of function.CONCLUSIONS: LL-37 potentiates dsRNA-induced cytokine production through up-regulation of TLR3 expression representing a novel pro-inflammatory mechanism.
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