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Keratinocyte Cell Line (keratinocyte + cell_line)
Kinds of Keratinocyte Cell Line Selected AbstractsThe Influence of Tetracycline Loading on the Surface Morphology and Biocompatibility of Films Made from P(3HB) Microspheres,ADVANCED ENGINEERING MATERIALS, Issue 7 2010Lydia Francis Tetracycline, an antibiotic used against a broad range of Gram positive and Gram negative bacteria was encapsulated in microspheres made of poly(3-hydroxybutyric acid) P(3HB), a microbial biodegradable polymer isolated from Bacillus cereus SPV. The drug loaded microspheres were prepared using an oil emulsion technique and compressed uniaxially to produce films. Although the same fabrication conditions were used for preparing the drug loaded and unloaded microspheres, the presence of the drug changed the surface morphology and roughness of the films. The surface morphology of the drug loaded films appeared uneven and coarser and the roughness, with an average root mean square value of 5.89,µm, was significantly higher than that of the unloaded film. The in vitro biocompatibility of the films was investigated using a human keratinocyte cell line (HaCaT) by comparing cell viability on the films to that on conventional tissue culture plastics. Both films appear to support cell growth but cell attachment and percentage cell viability were greater on the drug loaded films (32% of control) compared to the unloaded film (10% of control), possibly as a result of the non-uniform surface morphology and increased roughness of the drug loaded film. Thus, the above results illustrate that the drug loaded films, in addition to being a suitable matrix for drug delivery, represent an improved substrate for keratinocyte cell attachment. [source] CCL28 production in HaCaT cells was mediated by different signal pathways from CCL27EXPERIMENTAL DERMATOLOGY, Issue 2 2006Shinji Kagami Abstract:, Both CCL27 and CCL28 are ligands for CCR10 and attract CCR10+ lymphocytes. We previously demonstrated that CCL27 and CCL28 were strongly expressed in sera and lesional keratinocytes of patients with atopic dermatitis and psoriasis vulgaris. However, the regulation of CCL27 and CCL28 production in keratinocytes has not been well documented. In this study, we showed that CCL27 and CCL28 expression and production by a human keratinocyte cell line, HaCaT cells, were strongly induced by inflammatory cytokines tumor necrosis factor-, and interleukin-1,. CCL27 production was downregulated by inhibitors of p38 mitogen-activated protein kinase and nuclear factor-kappa B (NF-,B). By contrast, CCL28 production was downregulated by inhibitors of extracellular signal-regulated kinase and NF-,B. Our study results suggest that CCL28 produced by keratinocytes is mediated by different signal pathways from CCL27 and that both CCL27 and CCL28 are involved in the pathogenesis of inflammatory skin diseases. [source] Role of the IGF-II receptor in mediating acute, non-genomic effects of retinoids and IGF-II on keratinocyte cell deathEXPERIMENTAL DERMATOLOGY, Issue 4 2003F. Louafi Abstract:, In this study, we have examined the effects of retinoic acid (RA) on the human immortalized keratinocyte cell line (HaCaT). A significant twofold (P < 0.01) increase in apoptotic cell death compared with the control was found within 24 h of treatment with 10,5 M of RA. Apoptosis was confirmed by flow cytometry. Cycloheximide did not inhibit this acute RA-induced apoptosis. Interestingly, insulin-like growth factor-II (IGF-II, 50 ng/ml) was able to significantly (67.3%; P < 0.05) reduce RA effects, whereas IGF-I (50 ng/ml) and insulin (75 ng/ml) were without effect. Furthermore, analogues of IGF-II [leu27 IGF-II and Des(1-6) IGF-II], with altered affinities for the IGF-I receptor and IGF-binding proteins (IGFBPs), but retained affinities for the IGF-II receptor, also completely inhibited (100%; P < 0.01) RA-induced apoptosis, while an IGF-I receptor antagonist did not reduce the survival effects of IGF-II. Insulin pretreatment negates the survival effect of IGF-II. In contrast, mannose 6 phosphate (M6P) did not alter RA or IGF-II actions. These results indicate that rapid induction of cell death by RA is independent of production or secretion of new proteins. The inhibition of RA action by IGF-II was independent of its ability to signal through the IGF-I receptor or to interact with IGFBPs. [source] Genkwanin up-regulates the transcriptional activation of human type vii collagen gene promoterINTERNATIONAL JOURNAL OF COSMETIC SCIENCE, Issue 4 2007N. Takebayashi In a recent study, stimulating the formation of anchoring fibrils at the basement membrane zone in skin contributed to preventing skin ageing, such as wrinkle formation. Expression of the type VII collagen gene induces the formation of anchoring fibrils composed mainly of collagen type VII. We therefore transiently transfected a keratinocyte cell line with the plasmids containing type VII collagen gene promoter located upstream of the luciferase gene. We investigated the promoter activity under the presence of flavonoids and we found that Genkwanin up-regulates the transcriptional activation of human type VII collagen gene promoter. [source] Oncoprotein BMI-1 induces the malignant transformation of HaCaT cellsJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 1 2009Qian Wang Abstract BMI-1 (B-cell-specific Moloney murine leukemia virus integration site 1), a novel oncogene, has attracted much attention in recent years for its involvement in the initiation of a variety of tumors. Recent evidence showed that BMI-1 was highly expressed in neoplastic skin lesions. However, whether dysregulated BMI-1 expression is causal for the transformation of skin cells remains unknown. In this study, we stably expressed BMI-1 in a human keratinocyte cell line, HaCaT. The expression of wild-type BMI-1 induced the malignant transformation of HaCaT cells in vitro. More importantly, we found that expression of BMI-1 promoted formation of squamous cell carcinomas in vivo. Furthermore, we showed that BMI-1 expression led to the downregulation of tumore suppressors, such as p16INK4a and p14ARF, cell adhesion molecules, such as E-Cadherin, and differentiation related factor, such as KRT6. Therefore, our findings demonstrated that dysregulated BMI-1 could indeed lead to keratinocytes transformation and tumorigenesis, potentially through promoting cell cycle progression and increasing cell mobility. J. Cell. Biochem. 106: 16,24, 2009. © 2008 Wiley-Liss, Inc. [source] Cloning and identification of EDD gene from ultraviolet-irradiated HaCaT cellsPHOTODERMATOLOGY, PHOTOIMMUNOLOGY & PHOTOMEDICINE, Issue 6 2006Nishma Gupta Ultraviolet (UV) radiation is one of the most important external stimuli that affects skin by inducing cancer, inflammation and cell death. To identify the regulation of genes regulated by UV during transformation, normal human keratinocyte cell line, HaCaT, was exposed to multiple doses of UVA+B (UVA , 150,200 mJ/cm2 and UVB , 15,20 mJ/cm2× 6). Malignant transformation was confirmed by formation of colonies on soft agar and DNA methylation assay. To identify the genes involved in this process, random amplification of polymorphic DNA using RNA from unexposed and multiple exposed cells was performed after each exposure. A few up-regulated genes were identified, cloned and sequenced. One of the genes had homology to EDD (E3 identified by differential display) that was up-regulated at second exposure but was down-regulated in colony-forming cells (cells that received six or more exposures) as determined by RT-PCR. This is a progesterone-induced gene and progesterone treatment reduced the extent of colony formation on soft agar plate. It is possible that hormone therapy may have some effects on skin cancer in vivo. [source] TNF-, Drives Matrix Metalloproteinase-9 in Squamous Oral Carcinogenesis,THE LARYNGOSCOPE, Issue 8 2008Laurie Hohberger MD Abstract Objectives/Hypothesis: It is well known that invasion is a seminal event in the progression of oral and other head and neck carcinoma sites. We have previously demonstrated tumor necrosis factor (TNF)-, and its dependent cytokines are upregulated in saliva during oral carcinogenesis. TNF-dependent events stimulate nuclear factor (NF)-,B and many NF-,B-dependent genes are associated with cancer progression. Materials and Methods: In the present study, we examined NF-,B stimulation of matrix metalloproteinase (MMP)-9 in a precancerous keratinocyte cell line that models leukoplakia (Rhek cells). We stimulated Rhek cells with both TNF-, and phorbol myristate acetate, known stimulants of NF-,B. We then assayed MMP-9 transcription and secretion by luciferase reporter genes, quantitative real-time polymerase chain reaction, and fluorometric enzyme-linked immunosorbent serologic assay. Results: We discovered that the MMP-9 promoter was significantly stimulated by phorbol myristate acetate and TNF-, on luciferase reporter gene assays. Further, we uncovered that functional MMP-9 promoter activation was accompanied by significant increases in MMP-9 gene expression, as judged by quantitative real-time polymerase chain reaction. Functional activation of the MMP-9 protein was stimulated by TNF-, and PMA on a fluorescent enzyme-linked immunosorbent serologic assay. Finally, we searched our salivary proteomic database for increases in MMP-9 and discovered it was the third most significant protein in salivas of oral cavity cancer patients over normal controls. Conclusions: We conclude the milieu cytokine, TNF-,, has the capacity to provide stimulation of events related to early invasion of oral cavity cancer, as judged by its ability to stimulate MMP-9. [source] Opsonization of late apoptotic cells by systemic lupus erythematosus autoantibodies inhibits their uptake via an Fc, receptor,dependent mechanismARTHRITIS & RHEUMATISM, Issue 10 2007Esther Reefman Objective Decreased clearance of apoptotic cells is suggested to be a major pathogenic factor in systemic lupus erythematosus (SLE). The aim of this study was to investigate whether the binding of SLE autoantibodies to apoptotic cells influences the phagocytosis of these cells by macrophages. Methods Apoptosis was induced in a human T cell line (Jurkat) and a keratinocyte cell line (HaCaT) by ultraviolet B irradiation. Binding of purified IgG from 26 SLE patients and 15 healthy controls to apoptotic cells was assessed by flow cytometry and Western blotting. Phagocytosis of IgG-opsonized apoptotic cells by monocyte-derived macrophages was assessed by light microscopy. Similar experiments were performed with a monoclonal antibody against SSA/Ro and IgG fractions from 5 patients with Sjögren's syndrome (SS) and 5 patients with rheumatoid arthritis (RA). Results IgG fractions from all 26 SLE patients bound to late apoptotic, but not early apoptotic, cells. IgG fractions isolated from SLE patients with different autoantibody profiles showed comparable levels of binding. IgG fractions from healthy controls did not bind. Opsonization of apoptotic cells with IgG fractions from SLE patients resulted in a significant inhibition of phagocytosis as compared with healthy control IgG fractions. A monoclonal antibody directed against SSA/Ro and IgG isolated from 5 antinuclear antibody (ANA),positive patients with SS were also able to elicit these effects, whereas IgG from 5 ANA-negative patients with RA did not. The inhibitory effect of patient IgG was abolished by blocking either the Fc, receptors (Fc,R) or the constant region of IgG, using a specific Fc-blocking peptide. Conclusion Autoantibodies from SLE patients are able to opsonize apoptotic cells and inhibit their uptake by macrophages via an Fc,R-dependent mechanism. [source] Interleukin-4 and interleukin-13 enhance CCL26 production in a human keratinocyte cell line, HaCaT cellsCLINICAL & EXPERIMENTAL IMMUNOLOGY, Issue 3 2005S. Kagami Summary Eotaxin-2/CCL24 and eotaxin-3/CCL26 are CC chemokines and their receptor, CC chemokine receptor 3 is preferentially expressed on eosinophils. It was reported that vascular endothelial cells and dermal fibroblasts produced CCL26. However, the regulation of CCL24 and CCL26 production in keratinocytes has not been well documented. We investigated the expression and production of CCL24 and CCL26 in the human keratinocyte cell line, HaCaT cells. Reverse transcription and polymerase chain reaction was performed using these cells and Enzyme-linked immunosorbent assay was carried out using supernatant of these cells. The production of CCL24 in HaCaT cells was slightly enhanced by IL-4 and that of CCL26 was strongly enhanced by IL-4 and IL-13. Furthermore, TNF-, generated a synergistic effect on IL-4 enhanced CCL26 production. Dexamethasone, IFN-, and the p38 mitogen-activated protein kinase inhibitor SB202190 inhibited IL-4 enhanced CCL26 production. IL-4 enhanced production of CCL26 was inhibited by leflunomide and JAK inhibitor 1, but not by JAK3 inhibitor, which indicates that it is mediated by JAK1-STAT6-dependent pathway. This result also strongly suggests the involvement of the type 2 IL-4 receptor in IL-4 enhanced production of CCL26. These results suggest that keratinocytes are involved in the migration of CC chemokine receptor 3 positive cells such as eosinophils in a Th2-dominant situation like atopic dermatitis. [source] Cosmeceutical properties of levan produced by Zymomonas mobilisINTERNATIONAL JOURNAL OF COSMETIC SCIENCE, Issue 3 2006K. H. Kim Levan, a polysaccharide that can be produced by both plants and micro-organisms, is a sugar polymer composed of fructose, with-2,6 linkages. Here, we have attempted to assess the possible use of levan produced by Zymomonas mobilis as a cosmeceutical ingredient. In service of this goal, we assessed a host of levan's properties, including its moisturizing effects, cell cytotoxicity, cell proliferation effects and anti-inflammation effects. Levan exhibited a moisturizing effect that was almost exactly the same as that evidenced by hyaluronic acid, as well as a similar cell proliferation effect in human fibroblast and keratinocyte cell lines. Moreover, in our cell proliferation test, which was conducted using bio-artificial skin constructed via 3-dimensional (3-D) culture after the induction of primary skin inflammation with 0.05% sodium lauryl sulphate (SLS), cell viability in the presence of levan (0.01 and 0.05 mg mL,1) was determined to be higher than cell viability in the absence of levan. In our anti-inflammation test, which was also conducted using 3-D artificial skin, and which involved the measurement of a quantity of secreted interleukin-1 (IL-1), a pre-inflammatory mediator induced by SLS, we determined that the quantity of IL-1 in the 3-D artificial skin treated with 0.01 and 0.05 mg mL,1 of levan was less than that registered in a skin sample that had been treated only with SLS. In this study, we determined that levan exerted an anti-inflammatory effect against inflammatory reactions to skin irritants, and also that levan exerted a cell-proliferative effect in bio-artificial skin, thereby indicating its potential applicability as a cosmeceutical agent. [source] Establishment and characterization of immortalized human gingival keratinocyte cell linesJOURNAL OF PERIODONTAL RESEARCH, Issue 6 2008S. Gröger Background and Objective:, Primary human keratinocytes are used to analyze the properties of the oral epithelium and the early stages of oral bacterial infections. In vitro, these cells are characterized by their short life span and restricted availability. Approaches for culturing these cells will end after approximately 6,10 passages as a result of entry into apoptosis. For this reason, it is important to generate cell lines suitable for obtaining an unlimited source of cells. Therefore, the aim of the present study was to generate gingival keratinocyte cell lines and to compare their in vitro behaviour with those of primary human gingival keratinocytes. Material and Methods:, Primary human gingival keratinocytes were immortalized with a combination of the human papilloma virus onkoproteins E6 and E7. The pattern of the cytokeratins, involucrin and filaggrin was investigated by intracellular staining using flow cytometry. This method allows quantitative analysis of the expression of a variety of intracellular or extracellular markers. Results:, The immortalized cell lines showed many morphological similarities, expressing a cytokeratin pattern that is comparable with that of primary gingival keratinocytes. Furthermore, they developed transepithelial electrical resistance, which is a marker for the generation of tight junctions. These results indicate that the cells might be able to act as an epithelial barrier, reflecting the reaction of primary human cells. Conclusion:, The establishment of a continuous line of human gingival epithelial cells with functional characteristics of the epithelial barrier provides a valuable in vitro model for using to study the early steps of gingival/periodontal infections. [source] Analysis of the signal transduction pathway of nickel-induced matrix metalloproteinase-2 expression in the human keratinocytes in vitro: preliminary findingsJOURNAL OF CUTANEOUS PATHOLOGY, Issue 6 2007Brunella Perfetto Background:, Nickel can induce cellular and nuclear damages responsible for chronic diseases, like allergic contact dermatitis (ACD). We previously showed that matrix metalloproteinase-2 (MMP-2) gene expression was induced by nickel in nontumorigenic human keratinocytes cell line (HaCat). Objective:, To investigate the signal transduction pathways involved in gelatinolytic activity induced in HaCat under nickel stimulation. Methods:, We analyzed the involvement of protein kinase A (PKA), protein kinase C (PKC), tyrosine kinase (PTK), nuclear factor-kB (NF-kB) and activator protein-1 (AP-1) using specific inhibitors (H89, calphostin C, genistein, carpain and curcumin) by electrophoretic mobility shift assay, reverse transcription-polymerase chain reaction and gelatin zymography. Results:, Our results indicate that nickel-induced MMP-2 production was inhibited with PTK, PKC and AP-1 specific inhibitors. Moreover, both PKA and NF-kB were not involved in nickel pathway. Conclusions:, Using HaCat, we showed that curcumin and genistein can revert nickel-induced MMP-2 upregulation. Whether the use of PTK and AP-1 inhibitors has therapeutic ramifications in the management of ACD remains to be investigated. [source] | ||||||||||