| 1. |
- Dahl, Sara, et al.
(författare)
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The host defense peptide LL-37 triggers release of nucleic acids from human mast cells
- 2018
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Ingår i: Peptides. - : Elsevier BV. - 0196-9781 .- 1873-5169. ; 109, s. 39-45
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Tidskriftsartikel (refereegranskat)abstract
- The human host defense peptide LL-37 possesses antimicrobial activity but also affects host cell function and viability. Mast cells are involved in innate immunity but no data have been presented on effects of LL-37 on human mast cell viability and export of nucleic acids. Here, we demonstrated by immunofluorescence microscopy that synthesized LL-37 was internalized by human LAD2 mast cells and detected both in cytoplasm and nucleus. Treatment with high (4 and 10 μM) but not low (1 μM) concentrations of LL-37 for 4 h reduced cell viability assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Stimulation with 10 μM LL-37 for 4 h enhanced export of nucleic acids, total protein and lactate dehydrogenase (LDH), suggesting that both nuclear and plasma membranes are permeabilized by LL-37. Although LL-37 triggered release of nucleic acids, no extracellular trap-like structures were observed by laser scanning confocal microscopy of cells incubated with the plasma membrane impermeable nucleic acid fluorophore SYTOX-Green, indicating that LL-37 promotes export of nucleic acids but not formation of extracellular traps. On the other hand, phorbol-12-myristate-13-acetate (PMA), which is a well-known inducer of extracellular traps, stimulated export of nucleic acids and also formation of extracellular trap-like structures. However, PMA had no effect on export of either total protein or LDH. Hence, LL-37 and PMA seem to stimulate export of nucleic acids from LAD2 mast cells through different pathways. In conclusion, we demonstrate that LL-37 triggers release of nucleic acids from human mast cells but not the formation of extracellular trap-like structures.
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| 2. |
- Aidoukovitch, Alexandra, et al.
(författare)
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The host defense peptide LL-37 is internalized by human periodontal ligament cells and prevents LPS-induced MCP-1 production
- 2019
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Ingår i: Journal of Periodontal Research. - : Wiley. - 0022-3484 .- 1600-0765. ; 54:6, s. 662-670
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Tidskriftsartikel (refereegranskat)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. 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.
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| 3. |
- Anders, Emma, et al.
(författare)
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Globular C1q receptor (p33) binds and stabilizes pro-inflammatory MCP-1 : a novel mechanism for regulation of MCP-1 production and function
- 2018
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Ingår i: Biochemical Journal. - 0264-6021. ; 475:4, s. 775-786
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Tidskriftsartikel (refereegranskat)abstract
- The protein gC1qR (globular C1q receptor), also named p33, was originally identified as a binding partner of the globular heads of C1q in the complement system. gC1qR/p33 is abundantly expressed in many cell types, but the functional importance of this protein is not completely understood. Here, we investigate the impact of gC1qR/p33 on the production and function of the pathophysiologically important chemokine monocyte chemoattractant protein-1 (MCP-1) and the underlying molecular mechanisms. Knockdown of gC1qR/p33 negatively regulated the production of MCP-1, but had no effect on the expression of transcript for MCP-1 in human periodontal ligament cells, suggesting a translational/post-translational mechanism of action. Laser scanning confocal microscopy showed considerable cytosolic co-localization of gC1qR/p33 and MCP-1, and co-immunoprecipitation disclosed direct physical interaction between gC1qR/p33 and MCP-1. Surface plasmon resonance analysis revealed a high-affinity binding (KD = 10.9 nM) between gC1qR/p33 and MCP-1. Using a transwell migration assay, we found that recombinant gC1qR/p33 enhances MCP-1-induced migration of human THP-1 monocytes, pointing to a functional importance of the interaction between gC1qR/p33 and MCP-1. An in vitro assay revealed a rapid turnover of the MCP-1 protein and that gC1qR/ p33 stabilizes MCP-1, hence preventing its degradation. We propose that endogenous gC1qR/p33 physically interacts with MCP-1 causing stabilization of the MCP-1 protein and stimulation of its activity in human periodontal ligament cells, suggesting a novel gC1qR/p33-mediated pro-inflammatory mechanism of action.
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| 4. |
- Anders, Emma, et al.
(författare)
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LL-37-induced human osteoblast cytotoxicity and permeability occurs independently of cellular LL-37 uptake through clathrin-mediated endocytosis
- 2018
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Ingår i: Biochemical and Biophysical Research Communications. - : Elsevier BV. - 0006-291X. ; 501:1, s. 280-285
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Tidskriftsartikel (refereegranskat)abstract
- The host defense peptide LL-37 is cytotoxic for bacteria but it has also been reported to reduce host cell viability through an intracellular mechanism. LL-37-evoked cytotoxicity may be involved in the loss of bone tissue in periodontitis which is an inflammatory disease characterized by high concentrations of LL-37 observed locally in the periodontal tissue at the inflammation process. Here, we showed that LL-37 reduced human osteoblast-like MG63 cell viability assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and increased plasma membrane permeability determined by measuring intracellular Ca2+ levels and lactate dehydrogenase (LDH) release. Treatment with chlorpromazine, a well-recognized inhibitor of clathrin-mediated endocytosis, reduced cellular uptake of synthesized LL-37 b y about 30% assessed by Western blotting and ELISA, while filipin, an inhibitor of caveolin-mediated endocytosis, had no effect. The chlorpromazine-induced attenuation of LL-37 uptake was not associated with modulation of LL-37-induced cytotoxicity and LL-37-evoked plasma membrane permeability. Clathrin heavy chain 2 is a major protein of the polyhedral coat of coated pits and vesicles encoded by clathrin heavy chain like 1 gene. Down-regulation of clathrin heavy chain like 1 gene activity by siRNA reduced uptake of LL-37 but did not affect LL-37-induced cytotoxicity and permeability. Thus, we show, using both a pharmacological approach and knockdown of clathrin heavy chain like 1 expression, that LL-37-induced MG63 cell cytotoxicity and permeability occurs independently of LL-37 uptake via clathrin-mediated endocytosis.
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| 5. |
- Grossi, Mario, et al.
(författare)
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Pyk2 inhibition promotes contractile differentiation in arterial smooth muscle
- 2017
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Ingår i: Journal of Cellular Physiology. - : Wiley. - 0021-9541. ; 232:11, s. 3088-3102
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Tidskriftsartikel (refereegranskat)abstract
- Modulation from contractile to synthetic phenotype of vascular smooth muscle cells is a central process in disorders involving compromised integrity of the vascular wall. Phenotype modulation has been shown to include transition from voltage-dependent toward voltage-independent regulation of the intracellular calcium level, and inhibition of non-voltage dependent calcium influx contributes to maintenance of the contractile phenotype. One possible mediator of calcium-dependent signaling is the FAK-family non-receptor protein kinase Pyk2, which is activated by a number of stimuli in a calcium-dependent manner. We used the Pyk2 inhibitor PF-4594755 and Pyk2 siRNA to investigate the role of Pyk2 in phenotype modulation in rat carotid artery smooth muscle cells and in cultured intact arteries. Pyk2 inhibition promoted the expression of smooth muscle markers at the mRNA and protein levels under stimulation by FBS or PDGF-BB and counteracted phenotype shift in cultured intact carotid arteries and balloon injury ex vivo. During long-term (24–96 hr) treatment with PF-4594755, smooth muscle markers increased before cell proliferation was inhibited, correlating with decreased KLF4 expression and differing from effects of MEK inhibition. The Pyk2 inhibitor reduced Orai1 and preserved SERCA2a expression in carotid artery segments in organ culture, and eliminated the inhibitory effect of PDGF stimulation on L-type calcium channel and large-conductance calcium-activated potassium channel expression in carotid cells. Basal intracellular calcium level, calcium wave activity, and store-operated calcium influx were reduced after Pyk2 inhibition of growth-stimulated cells. Pyk2 inhibition may provide an interesting approach for preserving vascular smooth muscle differentiation under pathophysiological conditions.
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| 6. |
- Karbalaei, Mardjaneh, et al.
(författare)
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Detrusor Induction of miR-132/212 following Bladder Outlet Obstruction: Association with MeCP2 Repression and Cell Viability.
- 2015
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Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 10:1
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Tidskriftsartikel (refereegranskat)abstract
- The microRNAs (miRNAs) miR-132 and miR-212 have been found to regulate synaptic plasticity and cholinergic signaling and recent work has demonstrated roles outside of the CNS, including in smooth muscle. Here, we examined if miR-132 and miR-212 are induced in the urinary bladder following outlet obstruction and whether this correlates with effects on gene expression and cell growth. Three to seven-fold induction of miR-132/212 was found at 10 days of obstruction and this was selective for the detrusor layer. We cross-referenced putative binding sites in the miR-132/212 promoter with transcription factors that were predicted to be active in the obstruction model. This suggested involvement of Creb and Ahr in miR-132/212 induction. Creb phosphorylation (S-133) was not increased, but the number of Ahr positive nuclei increased. Moreover, we found that serum stimulation and protein kinase C activation induced miR-132/212 in human detrusor cells. To identify miR-132/212 targets, we correlated the mRNA levels of validated targets with the miRNA levels. Significant correlations between miR-132/212 and MeCP2, Ep300, Pnkd and Jarid1a were observed, and the protein levels of MeCP2, Pnkd and Ache were reduced after obstruction. Reduction of Ache however closely matched a 90% reduction of synapse density arguing that its repression was unrelated to miR-132/212 induction. Importantly, transfection of antimirs and mimics in cultured detrusor cells increased and decreased, respectively, the number of cells and led to changes in MeCP2 expression. In all, these findings show that obstruction of the urethra increases miR-132 and miR-212 in the detrusor and suggests that this influences gene expression and limits cell growth.
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| 7. |
- Müller, Dieter K., 1968-, et al.
(författare)
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Making "wilderness" in a northern natural resource periphery : on restructuring and production of a pleasure periphery in northern Sweden
- 2019
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Ingår i: The politics of Arctic resources. - London : Routledge. - 9781138040601 - 9781315174969 ; , s. 99-118
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Bokkapitel (refereegranskat)abstract
- Chapter 6 takes up the more recently developed sector of tourism, problematizing the assumptions inherent in conceptions such as "resource periphery" and "pleasure periphery". Instead, the chapter shows that resource use and tourism may well interact, and that tourism even largely relates to mining or mining infrastructure: the extensive existence of infrastructure related to resource uses and industrial as well as post-industrial development at large can even be seen as the basis for tourists being able to access the "wilderness".
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| 8. |
- Müller, Dieter, et al.
(författare)
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Making "Wilderness" in a northern natural resource periphery: on restructuring and the production of a pleasure periphery in northern Sweden
- 2019
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Ingår i: The Politics of Arctic Resources: Change and Continuity in the "Old North" of Northern Europe. - Abingdon, Oxon ; New York, NY : Routledge, 2019. | Series: Transforming environmental politics and policy : Routledge. - 9781315174969 ; , s. 99-118
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Bokkapitel (övrigt vetenskapligt/konstnärligt)abstract
- Historically, northern peripheries have been colonized in order to access northern resources. The use of these resources promised economic wealth and community development even in northern parts of the Scandinavian Peninsula (Sörlin, 1988). However, already in the late 19th century the Swedish Tourist Association started promoting the area as also being worth a visit (Aldskogius 1993). Still, tourism has developed slowly, and as an industry remained in the shadow of resource-extractive activities until the 1990s when automatization within resource industries, but also a general transition of the Swedish economy towards a domination of service industries, caused a search for alternative development options (Müller, 2011, 2013a). In this context, an abundant resource – a seemingly “pristine nature” or wilderness – is in focus and utilized as a resource for tourism development. Today, a renewed focus on the North, because of climate change and new industrial opportunities, is creating a discursive environment where ideas of the region as pristine and important for a global environment compete with the idea of a resource periphery. This development is cutting across scales, creating new stakeholder constellations, and globalizing the issue of northern development far beyond what has been witnessed to date. Against this background, the purpose of this chapter is to analyze tourism development in northern Sweden with regard to factors on different scales influencing the transition from a resource periphery to a pleasure periphery, and particularly the role of path dependence. This is achieved through a review of relevant literature and case studies conducted in the Swedish North.
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| 9. |
- Nebel, Daniel, et al.
(författare)
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1α,25-dihydroxyvitamin D3 promotes osteogenic activity and downregulates proinflammatory cytokine expression in human periodontal ligament cells
- 2015
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Ingår i: Journal of Periodontal Research. - : John Wiley & Sons. - 0022-3484 .- 1600-0765. ; 50:5, s. 666-673
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Tidskriftsartikel (refereegranskat)abstract
- Background and ObjectiveThe aim of this study was to assess the impact of 1,25-dihydroxyvitamin D3 (vitamin D3) on osteogenic and inflammatory properties of human periodontal ligament (PDL) cells and investigate underlying mechanisms. Material and MethodsHuman PDL cells, obtained from four subjects, were stimulated with vitamin D3 for 4-48h. The bone markers osteopontin and osteocalcin and proinflammatory cytokine/chemokine expression was determined by quantitative real-time polymerase chain reaction and enzyme-linked immunosorbent assay. Cytokine and chemokine expression was determined after stimulation with the inflammation promoter lipopolysaccharide (LPS) in the presence or absence of vitamin D3. Alkaline phosphatase activity was assessed using p-nitrophenylphosphate substrate. ResultsTreatment with 30ng/mL of vitamin D3, corresponding to an optimal plasma concentration of vitamin D, for 24h had no effect on PDL cell number and morphology but increased PDL cell osteopontin and osteocalcin mRNA expression by about 70 and 40%, respectively, and, moreover, treatment with vitamin D3 for 48h enhanced PDL cell alkaline phosphatase activity by about two times showing that vitamin D3 exerts pro-osteogenic effects in human PDL cells. Stimulation with LPS (1g/mL) for 4h increased PDL cell interleukin (IL)-6 cytokine and chemokine ligand 1 (CXCL1) chemokine mRNA expression several fold. The LPS-induced increase in IL-6 and CXCL1 transcripts was attenuated by vitamin D3 (30ng/mL). Treatment with vitamin D3 (3-300ng/mL) for 24h reduced the LPS-evoked increase in PDL cell IL-6 protein by about 50%. Vitamin D3 (30ng/mL) had no effect on LPS-induced IL-1 and MCP-1 mRNA expression. ConclusionsVitamin D3 promotes osteogenic differentiation but also downregulates inflammation promoter-induced IL-6 cytokine and CXCL1 chemokine expression in human PDL cells, suggesting that vitamin D3 both stimulates bone regeneration and antagonizes inflammation in human periodontal tissue.
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| 10. |
- Svensson, Daniel, et al.
(författare)
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Apolipoprotein A-I attenuates LL-37-induced endothelial cell cytotoxicity
- 2017
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Ingår i: Biochemical and Biophysical Research Communications. - : Elsevier BV. - 0006-291X. ; 493:1, s. 71-76
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Tidskriftsartikel (refereegranskat)abstract
- The human cathelicidin peptide LL-37 has antimicrobial and anti-biofilm functions, but LL-37 may also damage the host by triggering inflammation and exerting a cytotoxic effect, thereby reducing host cell viability. Human plasma mitigates LL-37-induced host cell cytotoxicity but the underlying mechanisms are not completely understood. Apolipoprotein A-I (ApoA-I) is a plasma protein endowed with atheroprotective effects. Here, we investigate the interaction between ApoA-I and LL-37 by biochemical techniques, and furthermore assess if ApoA-I protects against LL-37-evoked cytotoxicity in human umbilical vein endothelial cells (HUVEC). Our results demonstrated that ApoA-I effectively binds LL-37. The binding of ApoA-I to LL-37 resulted in a structural rearrangement of the protein, but this interaction did not cause lower ApoA-I stability. Recombinant ApoA-I protected against LL-37-induced cytotoxicity in HUVEC and endogenous ApoA-I knockdown in HepG2 cells made the cells more sensitive to LL-37-evoked cytotoxicity. We conclude that ApoA-I physically interacts with LL-37 and antagonizes LL-37-induced down-regulation of endothelial cell viability suggesting that this mechanism counteracts endothelial cell dysfunction.
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