Induction of apoptosis in keratinocytes by UV light is a critical event in photocarcinogenesis. Although p53 is of importance in this process, evidence exists that other pathways play a role as well. Therefore, we studied whether the apoptosis-related surface molecule CD95 (Fas/APO-1) is involved. The human keratinocyte cell line HaCaT expresses CD95 and undergoes apoptosis after treatment with UV light or with the ligand of CD95 (CD95L). Incubation with a neutralizing CD95 antibody completely prevented CD95L-induced apoptosis but not UV-induced apoptosis, initially suggesting that the CD95 pathway may not be involved. However, the protease CPP32, a downstream molecule of the CD95 pathway, was activated in UV-exposed HaCaT cells, and UV-induced apoptosis was blocked by the ICE protease inhibitor zVAD, implying that at least similar downstream events are involved in CD95- and UV-induced apoptosis. Activation of CD95 results in recruitment of the Fas-associated protein with death domain (FADD) that activates ICE proteases. Immunoprecipitation of UV-exposed HaCaT cells revealed that UV light also induces recruitment of FADD to CD95. Since neutralizing anti-CD95 antibodies failed to prevent UV-induced apoptosis, this suggested that UV light directly activates CD95 independently of the ligand CD95L. Confocal laser scanning microscopy showed that UV light induced clustering of CD95 in the same fashion as CD95L. Prevention of UV-induced CD95 clustering by irradiating cells at 10°C was associated with a significantly reduced death rate. Together, these data indicate that UV light directly stimulates CD95 and thereby activates the CD95 pathway to induce apoptosis independently of the natural ligand CD95L. These findings further support the concept that UV light can affect targets at the plasma membrane, thereby even inducing apoptosis.
Regulatory CD4(+)CD25(+) T cells are important in suppressing immune responses. The requirements for the maintenance of peripheral CD4(+)CD25(+) T cells remain incompletely understood. Receptor activator of NF-kappaB (RANK) and its ligand (RANKL; also known as CD254, OPGL and TRANCE) are key regulators of bone remodeling, mammary gland formation, lymph node development and T-cell/dendritic cell communication. Here we report that RANKL is expressed in keratinocytes of the inflamed skin. RANKL overexpression in keratinocytes resulted in functional alterations of epidermal dendritic cells and systemic increases of regulatory CD4(+)CD25(+) T cells. Thus, epidermal RANKL expression can change dendritic cell functions to maintain the number of peripheral CD4(+)CD25(+) regulatory T cells. Epidermal RANKL mediated ultraviolet-induced immunosuppression and overexpression of epidermal RANKL suppressed allergic contact hypersensitivity responses and the development of systemic autoimmunity. Therefore, environmental stimuli at the skin can rewire the local and systemic immune system by means of RANKL.
SummaryTumor necrosis factor a (TNF-a), in addition to being cytotoxic for certain tumor cells, has turned out as a multifunctional cytokine that is involved in the regulation of immunity and inflammation. Since human keratinocytes have been demonstrated to be a potent source of various cytokines, it was investigated whether epidermal cells synthesize and release TNF-m Supernatants derived from normal human keratinocytes (HNK) and human epidermoid carcinoma cell lines (KB, A431) were tested both in a TNF-a-specific ELISA and a bioassay. In supernatants of untreated epidermal cells, no or minimal TNF-a activity was found, while after stimulation with lipopolysaccharide (LPS) or ultraviolet (UV) light, significant amounts were detected. Western blot analysis using an antibody directed against human TNF-a revealed a molecular mass of 17 kD for keratinocyte-derived TNF-a. These biological and biochemical data were also confirmed by Northern blot analysis revealing mRNA specific for TNF-a in LPS-or ultraviolet B (UVB)-treated HNK and KB cells. In addition, increased TNF-a levels were detected in the serum obtained from human volunteers 12 and 24 h after a single total body UVB exposure, which caused a severe sunburn reaction . These findings indicate that keratinocytes upon stimulation are able to synthesize and release TNF-a, which may gain access to the circulation. Thus, TNF-a in concert with other epidermal cell-derived cytokines may mediate local and systemic inflammatory reactions during host defense against injurious events caused by microbial agents or UV irradiation.
IKK gamma/NEMO is the essential regulatory subunit of the I kappa B kinase (IKK), encoded by an X-linked gene in mice and humans. It is required for NF-kappa B activation and resistance to TNF-induced apoptosis. Female mice heterozygous for Ikk gamma/Nemo deficiency develop a unique dermatopathy characterized by keratinocyte hyperproliferation, skin inflammation, hyperkeratosis, and increased apoptosis. Although Ikk gamma+/- females eventually recover, Ikk gamma- males die in utero. These symptoms and inheritance pattern are very similar to those of incontinentia pigmenti (IP), a human genodermatosis, synthenic with the IKK gamma/NEMO locus. Indeed, biopsies and cells from IP patients exhibit defective IKK gamma/NEMO expression but normal expression of IKK catalytic subunits. This unique self-limiting disease, the first to be genetically linked to the IKK signaling pathway, is dependent on X-chromosome inactivation. We propose that the IKK gamma/NEMO-deficient cells trigger an inflammatory reaction that eventually leads to their death.
ABSTRACT:Rivaroxaban is a novel, oral, direct factor Xa inhibitor for the prevention and treatment of thromboembolic disorders. The objective of this study was to investigate the in vivo metabolism and excretion of rivaroxaban in rats, dogs, and humans. Single doses of [ 14 C]rivaroxaban (3 and 1 mg/kg) were administered to rats (orally/ intravenously) and dogs (orally), respectively. A single oral dose of [ 14 C]rivaroxaban (10 mg) was administered to healthy human males (n ؍ 4). Plasma and excreta were collected and profiled for radioactivity. Recovery of total radioactivity was high and >92% in all species. Unchanged rivaroxaban was the major compound in plasma at all time points investigated, across all species. No major or pharmacologically active circulating metabolites were detected. Rivaroxaban and its metabolites were rapidly excreted; urinary excretion of radioactivity was 25 and 52%, and fecal excretion was 67 and 43% of the dose in rats and dogs, respectively. In humans, 66% of the dose was excreted renally (36% unchanged drug) and 28% in the feces. Radioactivity profiles in excreta were similar across species. Three metabolic pathways were identified: oxidative degradation of the morpholinone moiety (major pathway) and hydrolysis of the central amide bond and of the lactam amide bond in the morpholinone ring (minor pathways). M-1, the main metabolite in excreta of all species, was eliminated via both renal and fecal/biliary routes. In total, 82 to 89% of the dose administered was assigned to unchanged rivaroxaban and its metabolites in the excreta of rats, dogs, and humans.Rivaroxaban is a novel, oral, direct factor Xa (FXa) inhibitor in advanced clinical development for the prevention and treatment of thromboembolic disorders. More recently, it has received approval in Canada and the European Union for use in the prevention of venous thromboembolism in patients undergoing elective total hip or knee replacement surgery. Rivaroxaban is a selective inhibitor not only of free FXa (K i 0.4 nM), but also of prothrombinase activity and fibrinassociated FXa activity Perzborn et al., 2005). In vitro, rivaroxaban was shown to inhibit thrombin generation and prolong clotting times Perzborn et al., 2005) and, in vivo, it had potent antithrombotic effects in a variety of animal venous and arterial thrombosis models Biemond et al., 2007).Pharmacokinetic (PK) studies of rivaroxaban in rats and dogs have been reported previously (Weinz et al., 2005). These studies demonstrated that rivaroxaban was absorbed rapidly after oral dosing (absolute bioavailability 57-66 and 60 -86% in rats and dogs, respectively), had a favorable PK profile with dose proportional increases in area under the concentration-time curve (AUC) and was rapidly excreted via renal and fecal/biliary routes. Rivaroxaban has also been shown to demonstrate dose-proportional pharmacokinetics and predictable pharmacodynamics in single (up to 80 mg)-and multipledose studies in healthy subjects and patients with no evidence of accumulation (Kubitza et ...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
customersupport@researchsolutions.com
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
This site is protected by reCAPTCHA and the Google Privacy Policy and Terms of Service apply.
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.