Project description:Together with TEAD4 the Hippo pathway effector YAP stimulates chromosomal instability. Verteporfin is known to disrupt the physical YAP/TEAD interaction and tehrefore might prevent from chromosomal instability in liver cancer. Immortalized HCC cell line hepG2 is treated with Verteporfin for 24 hours.
Project description:Skin is usually exposed during human exposures to ionizing radiation, however there are few experiments that evaluate the radiation responsiveness of the cells of the epidermis (keratinocytes) and those of the dermis (fibroblasts) in the same studies. We evaluated the transcriptional responses of quiesent primary keratinocytes and fibroblasts from the same individual and compared them with quiescent keratinocytes and fibroblasts that were immortalized by human telomerase (hTert). The primary transcriptional responses to 10-500 cGy ionizing radiation were p53-mediated responses; however, we did identify distinct responses between the keratinocytes and the fibroblasts. Experiment Overall Design: Four cell types (primary keratinocytes, hTert immortalized keratinocytes, primary fibroblasts, hTert immortalized fibroblasts) grown to quiescence, treated with 0, 10, 100 or 500 cGy gamma irradiation, RNA collected at 4 hrs.
Project description:Immortalized keratinocytes HaCaT is popular model for skin research (toxicity, irritation, allergic reactions or interaction of cells). They maintain stable keratinocyte phenotype and respond to keratinocyte differentiation stimuli. However, the programs of stratification and expression of differentiation markers in HaCaT keratinocytes are aberrant. HaCaT cells bear two mutant p53 alleles (R282Q and H179Y) which contain Gain-of-Function (GOV) mutations acquired as a result of spontaneous immortalization (mutp53). At the same time, mutp53 acts as a transcription factor, and also affects interaction of p63 protein with its transcription targets. It is known that proteins of the P53 family play an important role in regulating processes of proliferation and differentiation of human keratinocytes.At the same time, the role of mutp53 in these processes is not fully clear. We present data sets obtained as a result of high-performance proteomic analysis of immortalized HaCaT keratinocytes with p53 knockout in two different states: sub-confluent and confluent, which are characterized by different intensity of cell differentiation processes. As a protocol for proteomic profiling of cells, we used the approach of obtaining LC-MS/MS measurements followed by their processing with Progenesis LC-MS software (Nonlinear Dynamics Ltd.)
Project description:Previous studies have shown that normal and HPV immortalized keratinocytes are sensitive to TNF anti-proliferative effect. Conversely, HPV18-immortalized keratinocytes are resistant to the cytostatic effect mediated by this cytokine. In this study we have compared gene expression patterns in primary cultures of human foreskin epidermal keratinocytes (PHK or NHFK) and HPV16 (HF698) and HPV18(HF18Nco)-immortalized cell lines, and profile the transcriptional changes 3 and 60 hours after TNF treatment. Keywords: global expression profile, microarray, HPV, TNF
Project description:The UVB component of the sunlight (290-320 nm) plays an important role in carcinogenesis through generation of high levels of bipyrimidine DNA photoproducts, while UVA (320-400 nm) has been associated with photoaging and tumor progression through generation of low, but continuous levels of DNA damage and oxidative stress. However, the contribution of UVA light to epidermal cell fate in the context of photoaging remains poorly understood. Here, by using proteomic analyses and biochemical assays for validation, we show that UVA induces proteome remodeling and senescence in primary keratinocytes, eliciting potent antioxidant and pro-inflammatory responses. As a model of early skin tumorigenesis during aging, immortalized non-malignant keratinocytes, bearing potentially oncogenic mutations and dysfunctional components of the senescent machinery , are resilient to UVA-induced stress, but are sensitive to paracrine oxidative stress and immune system activation induced by senescent neighboring keratinocytes. These observations reveal a new cellular mechanism by which UVA induces photoaging in the epidermis.
Project description:Photobiomodulation (PBM) with blue light induces a biphasic dose response curve in proliferation of immortalized human keratinocytes (HaCaT), with a maximum proliferative effect reached with 7.5min (10.35J/cm2). The aim of this study was to test the photobiomodulatory effect of 10.35J/cm2 blue light irradiation on cell proliferation, ROS production and gene expression at different time points after irradiation of HaCaT cells in vitro and to compare the results with the anti-proliferative phase of PBM using blue light with 41.4J/cm2. For 10.35J/cm2 cell proliferation was increased for all measured harvesting time points up to at least 24h after irradiation. ROS concentration was increased 30min after irradiation. However, already 1h after irradiation, cells were able to decrease ROS and stabilize the concentration to the basal level. The sudden increase in ROS was even higher when compared to 41.4J/cm2. However, gene expression analysis did not indicate any sign of apoptosis and even indicated an activation of cell survival mechanisms. Furthermore, comparable to 41.4J/cm2, aryl hydrocarbon receptor (AHR) “battery genes” showed a deregulation after blue light irradiation strengthening the hypothesis of AHR as possible target for blue light irradiation.
Project description:Photobiomodulation (PBM) with blue light induces a biphasic dose response curve in proliferation of immortalized human keratinocytes (HaCaT), with a maximum proliferative effect reached with 7.5min (10.35J/cm2). The aim of this study was to test the photobiomodulatory effect of 10.35J/cm2 blue light irradiation on cell proliferation, ROS production and gene expression at different time points after irradiation of HaCaT cells in vitro and to compare the results with the anti-proliferative phase of PBM using blue light with 41.4J/cm2. For 10.35J/cm2 cell proliferation was increased for all measured harvesting time points up to at least 24h after irradiation. ROS concentration was increased 30min after irradiation. However, already 1h after irradiation, cells were able to decrease ROS and stabilize the concentration to the basal level. The sudden increase in ROS was even higher when compared to 41.4J/cm2. However, gene expression analysis did not indicate any sign of apoptosis and even indicated an activation of cell survival mechanisms. Furthermore, comparable to 41.4J/cm2, aryl hydrocarbon receptor (AHR) “battery genes” showed a deregulation after blue light irradiation strengthening the hypothesis of AHR as possible target for blue light irradiation.