Effect of 1-MHz ultrasound on the proinflammatory interleukin-6 secretion in human keratinocytes.
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ABSTRACT:
SUBMITTER: Giantulli S
PROVIDER: S-EPMC8463532 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Skin is exposed to various physico-chemical cues. Keratinocytes, a major component of the skin epidermis, directly interact with the surrounding extracellular matrix, and thus, biochemical and biophysical stimulations from the matrix regulate the function of keratinocytes. Although it was reported that inflammatory responses of skin were altered by an applied mechanical force, understanding how the keratinocytes sense the mechanical stimuli and regulate a cytokine secretion remains unclear. Here, we designed a device that is able to apply chemo-mechanical cues to keratinocytes and assess their proinflammatory cytokine IL-6 production. We showed that when chemical stimuli were applied with mechanical stimuli simultaneously, the IL-6 production markedly increased compared to that observed with a single stimulus. Quantitative structural analysis of cellular components revealed that the applied mechanical stretch transformed the cell morphology into an elongated shape, increased the cell size, and dictated the distribution of focal adhesion complex. Our results suggest that the mechanical cue-mediated modulation of focal adhesion proteins and actin cytoskeleton translates into intracellular signaling associated with the IL-6 production particularly in skin sensitization. Our study can be applied to understand proinflammatory responses of skin under altered biophysical environments of the skin.
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2010-10-31 | E-GEOD-20706 | biostudies-arrayexpress
Project description:Analysis of cultured epidermal keratinocytes treated with interleukin-4 (IL-4) and interleukin-13 (IL-13). IL-4 and IL-13 are up-regulated in atopic dermatitis. Results provide insight into the role of IL-4 and IL-13 cytokines in the pathogenesis of atopic dermatitis. Analysis of epidermal keratinocytes transfected with dual oxidase 1 (DUOX1) siRNA knockdown before treatment with IL-4 and IL-13. DUOX1 is one of the NOX family members of NADPH oxidases whose primary function is ROS generation. Results provide insight into the role of the incraesed expression of DUOX1 in IL-4/IL-13-treated NHEK for IL4/IL13 signaling.
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Project description:Intrinsic β-cell circadian clocks are important regulators of insulin secretion and overall glucose homeostasis. Whether the circadian clock in β-cells is perturbed following exposure to prodiabetogenic stressors such as proinflammatory cytokines, and whether these perturbations are featured during the development of diabetes, remains unknown. To address this, we examined the effects of cytokine-mediated inflammation common to the pathophysiology of diabetes, on the physiological and molecular regulation of the β-cell circadian clock. Specifically, we provide evidence that the key diabetogenic cytokine IL-1β disrupts functionality of the β-cell circadian clock and impairs circadian regulation of glucose-stimulated insulin secretion. The deleterious effects of IL-1β on the circadian clock were attributed to impaired expression of key circadian transcription factor Bmal1, and its regulator, the NAD-dependent deacetylase, Sirtuin 1 (SIRT1). Moreover, we also identified that Type 2 diabetes in humans is associated with reduced immunoreactivity of β-cell BMAL1 and SIRT1, suggestive of a potential causative link between islet inflammation, circadian clock disruption, and β-cell failure. These data suggest that the circadian clock in β-cells is perturbed following exposure to proinflammatory stressors and highlights the potential for therapeutic targeting of the circadian system for treatment for β-cell failure in diabetes.
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Project description:Lipin-1 is an Mg2+-dependent phosphatidate phosphatase (PAP1) that catalyzes a critical step in the synthesis of glycerophospholipids and is also a cotranscriptional regulator. The role of lipin-1 in the regulation of inflammatory responses has been extensively studied in various cell types but not in skin cells. In the present study, the function of lipin-1 in UVB-induced proinflammatory responses was assessed in normal human epidermal keratinocytes (NHEKs). UVB radiation downregulated lipin-1 expression. Lipin-1 inhibition was mediated by UVB-dependent sterol-response element binding protein-1 (SREBP-1) inhibition. The UVB-dependent inhibition of lipin-1 and SREBP-1 was mediated by AMPK activation. UVB-induced activation of JNK was dependent on AMPK activation and mediated lipin-1 inhibition. Prevention of UVB-mediated lipin-1 repression by introducing a lipin-1 expression vector stimulated IL-6 and IL-8 production, suggesting that lipin-1 inhibition attenuates UVB-induced IL-6 and IL-8 production. The downregulation of lipin-1 ameliorated UVB-induced NF-ĸB phosphorylation, which might be attributed to the suppression of UVB-induced accumulation of free fatty acids (FFAs). Pharmacological inhibition of PAP1 with propranolol suppressed UVB-induced production of IL-6 and IL-8 in NHEKs and reconstituted human skin models. Taken together, lipin-1 is downregulated by exposure to UVB radiation, which confers protection against UVB-induced proinflammatory responses; therefore, the inhibition of lipin-1 is a potential strategy for photoaging.
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Project description:To investigate whether palmitic acid can be responsible for the induction of inflammatory processes, HaCaT keratinocytes were treated with palmitic acid at pathophysiologically relevant concentrations. Secretion levels of interleukin-6 (IL-6), tumor necrosis factor- ? (TNF- ?), interleukin-1 ? (IL-1 ?), NF- ? B nuclear translocation, NF- ? B activation, Stat3 phosphorylation, and peroxisome proliferator-activated receptor alpha (PPAR ?) mRNA and protein levels, as well as the cell proliferation ability were measured at the end of the treatment and after 24 hours of recovery. Pyrrolidine dithiocarbamate (PDTC, a selective chemical inhibitor of NF- ? B) and goat anti-human IL-6 polyclonal neutralizing antibody were used to inhibit NF- ? B activation and IL-6 production, respectively. Our results showed that palmitic acid induced an upregulation of IL-6, TNF- ? , IL-1 ? secretions, accompanied by NF- ? B nuclear translocation and activation. Moreover, the effect of palmitic acid was accompanied by PPAR ? activation and Stat3 phosphorylation. Palmitic acid-induced IL-6, TNF- ? , IL-1 ? productions were attenuated by NF- ? B inhibitor PDTC. Palmitic acid was administered in amounts able to elicit significant hyperproliferation and can be attenuated by IL-6 blockage. These data demonstrate for the first time that palmitic acid can stimulate IL-6, TNF- ? , IL-1 ? productions in HaCaT keratinocytes and cell proliferation, thereby potentially contributing to acne inflammation and pilosebaceous duct hyperkeratinization.
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Project description:Interleukin (IL)-38 is a member of the IL-1 family of cytokines, which was proposed to exert anti-inflammatory effects. IL-38 is constitutively expressed in the skin, where keratinocytes are the main producing cells. Little information is currently available concerning IL-38 biology. Here, we investigated the subcellular localization and interaction partners of the IL-38 protein in human keratinocytes. IL-38 expression was reduced in primary keratinocytes grown in monolayer (2D) cultures. We thus used IL-38 overexpressing immortalized normal human keratinocytes (NHK/38) to study this cytokine in cell monolayers. In parallel, differentiation of primary human keratinocytes in an in vitro reconstructed human epidermis (RHE) 3D model allowed us to restore endogenous IL-38 expression. In NHK/38 cells and in RHE, IL-38 was mainly cell-associated, rather than released into culture supernatants. Intracellular IL-38 was preferentially, although not exclusively, cytoplasmic. Similarly, in normal human skin sections, IL-38 was predominantly cytoplasmic in the epidermis and essentially excluded from keratinocyte nuclei. A yeast two-hybrid screen identified destrin/actin-depolymerizing factor (DSTN) as a potential IL-38-interacting molecule. Co-immunoprecipitation and proximity ligation assay confirmed this interaction. We further observed partial co-localization of IL-38 and DSTN in NHK/38 cells. Endogenous IL-38 and DSTN were also co-expressed in all epidermal layers in RHE and in normal human skin. Finally, IL-38 partially co-localized with F-actin in NHK/38 cells, in particular along the cortical actin network and in filopodia. In conclusion, IL-38 is found predominantly in the cytoplasm of human keratinocytes, where it interacts with DSTN. The functional relevance of this interaction remains to be investigated.
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