Unknown,Transcriptomics,Genomics,Proteomics

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Erbb2 regulates inflammation and proliferation in the skin after ultraviolet irradiation.

ABSTRACT: Exposure to ultraviolet (UV) irradiation is the major cause of nonmelanoma skin cancer, the most common form of cancer in the United States. UV irradiation has a variety of effects on the skin associated with carcinogenesis, including DNA damage and effects on signal transduction. The alterations in signaling caused by UV regulate inflammation, cell proliferation, and apoptosis. UV also activates the orphan receptor tyrosine kinase and proto-oncogene Erbb2 (HER2/neu). In this study, we demonstrate that the UV-induced activation of Erbb2 regulates the response of the skin to UV. Inhibition or knockdown of Erbb2 before UV irradiation suppressed cell proliferation, cell survival, and inflammation after UV. In addition, Erbb2 was necessary for the UV-induced expression of numerous proinflammatory genes that are regulated by the transcription factors nuclear factor-kappaB and Comp1, including interleukin-1beta, prostaglandin-endoperoxidase synthase 2 (Cyclooxygenase-2), and multiple chemokines. These results reveal the influence of Erbb2 on the UV response and suggest a role for Erbb2 in UV-induced pathologies such as skin cancer. Keywords: time course, ultraviolet irradiation, UV, erbB2, mouse skin The dorsal skin of adult female CD-1 mice was clipped one day before treatment and shaved on the day of treatment. DMSO or 4 mg AG825 dissolved in DMSO was applied topically to the shaved back of the mice 2 h prior to exposure to 10 kJ/m^2 UV or sham irradiation. The UV dose was approximately 30% UVA, 70% UVB and <1% UVC, with a total output of 470 uW/cm^2. Flash frozen skin was removed and total RNA expracted with TRIzol reagent (Invitrogen) and further purified with an RNeasy kit (Qiagen). Amplification, reverse-transcription, biotinylation, and hybridization were all carried out under standard conditions and procedures recommended by the manufacturer.

ORGANISM(S): Mus musculus

SUBMITTER: Laura Hansen

PROVIDER: E-GEOD-4066 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Erbb2 regulates inflammation and proliferation in the skin after ultraviolet irradiation.

Madson Justin G JG Lynch David T DT Tinkum Kelsey L KL Putta Sumanth K SK Hansen Laura A LA

The American journal of pathology 20061001 4

Exposure to ultraviolet (UV) irradiation is the major cause of nonmelanoma skin cancer, the most common form of cancer in the United States. UV irradiation has a variety of effects on the skin associated with carcinogenesis, including DNA damage and effects on signal transduction. The alterations in signaling caused by UV regulate inflammation, cell proliferation, and apoptosis. UV also activates the orphan receptor tyrosine kinase and proto-oncogene Erbb2 (HER2/neu). In this study, we demonstra ...[more]

PMID: 17003495

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Project description:Following UV irradiation of skin, dendritic cells (DCs) differentiating from the bone marrow (BM) of mice have a reduced ability to prime new immune responses; their reduced immunogenicity is maintained for at least 16 weeks in UV-chimeric mice. We hypothesized that different metabolic states underpin changes in DC function. Compared with DCs from the BM of non-irradiated mice, DCs from the BM of UV-irradiated mice produced more lactate and utilized greater amounts of glucose, a profile that was supported by greater glycolytic flux when incubated in low-serum-containing medium. Responses to a mitochondrial stress test were similar suggesting that the DCs from the BM of UV-irradiated mice had not switched from a profile of oxidative phosphorylation, but were imprinted for greater glycolytic responses. After microarray profiling, RT-qPCR confirmation and Ingenuity pathway analysis, greater expression of the enzyme, 3-hydroxyanthranilate 3,4-dioxygenase, was identified as a potential contributor to increased glycolysis by BM-differentiated DCs. This enzyme provides the final step of the biosynthetic pathway from tryptophan to quinolinate, the universal de novo precursor to the pyridine ring of nicotinamide adenine dinucleotide (NAD), and may provide a mechanism to ensure sufficient NAD is available to support enhanced glycolysis. Increased lactate production was also measured for DCs from the BM of 16-week engrafted UV-chimeric mice and suggests long-lasting imprinting of progenitor cells for altered immunometabolism in their progeny cells. This study provides evidence of changes to metabolic states that associate with altered DC function. Female C57BL/6J (CD45.2 alloantigen) and B6.SJL-Ptprca (CD45.1 alloantigen) mice were obtained from the Animal Resources Centre (Murdoch, Western Australia). A bank of TL40W/12RS lamps (Philips, Amsterdam, The Netherlands) emitting broadband UVR with 65% UVB (280-320 nm) and peak emission at 313 nm was used. Twenty-four h prior to irradiation, a uniform area of dorsal skin of mice was shaved (8 cm2). To administer UVR, mice were held in perspex compartments which were covered with 0.2 mm polyvinyl chloride plastic to eliminate wavelengths <290 nm. The compartments were placed 20 cm beneath the UV lamps and up to 8 kJ/m2 UVR was delivered. Eight kJ/m2 UVR is equivalent to 3-4 minimal erythemal doses . The UVB output by the lamps was measured using a UVX radiometer (Ultraviolet Products Inc., Upland, CA). Mice were 6-10 wk old at the time of irradiation unless otherwise stated. Mice were subcutaneously implanted with two PGE2 pellets, each containing 0.1 mg with a constant release of 4.76 Âµg/day over 21 days (total 9.52 Âµg/day, Innovative Research of America, Sarasota, FL), three days prior to isolation of BM cells. The pellets were inserted into the loose skin at the top of the back. Freshly-isolated BM cells were cultured for 5 days in RPMI-1640 (HyClone, GE Health Care Life Sciences, Logan, UT) supplemented with 10% FCS (RPMI-10), 10 ng/ml GM-CSF and 10 ng/ml IL-4 (Peprotech Inc, Rocky Hill, NJ) at a density of 8 x 105 cells/ml in 24 well plates to promote CD11c+ cell differentiation (medium replaced on days 2 and 4). Non-adherent cells were enriched to >95% CD11c+ cells (confirmed by flow cytometry) using anti-CD11c magnetic microbeads (Miltenyi Biotec, Bergisch Gladbach, Germany) and autoMACS-Pro (Miltenyi Biotec) separation. Total RNA was extracted from triplicate preparations of BM-differentiated CD11c+ cells. Two sample groups; BM-differentiated DC sample from C57BL/6J mice injected with 2 x 10^6 BM cells from a naÃ¯ve congenic B6.SJL-Ptprca mouse BM-differentiated DC sample from C57BL/6J mice injected with 2 x 10^6 BM cells from a UV irradiated congenic B6.SJL-Ptprca mouse

Project description:Dr. Liu's research group is interested in studying the expression and functions of galectin-3, -7 and -12, in particular the roles of these proteins in inflammation and neoplasm. Members of the galectin family are known to participate in cellular homeostasis by modulating cell growth, controlling cell cycle progression, and inducing or inhibiting apoptosis. It is known that some galectins have similar functions. However, it is not fully understood whether they work cooperatively or not. As the outermost barrier of the body, skin is directly and frequently exposed to a prooxidative environment, including solar ultraviolet A (UVA), ultraviolet B (UVB) radiation, and air pollution. Several reports have shown that exposure of cells to UV increase or decrease the levels of galectins. For example, the amounts of galectin-7 mRNA and protein are increased rapidly after UVB irradiation of keratinocytes (Proc. Natl. Acad. Sci. USA 1999; 96:11329-34). Heat shock and subculturing decrease, while alkylating agents and UV-light increase galectin-3 (Cell Physiol Biochem 2000; 10:149-58). To analyze the change of all galectin gene expression profiles after UVB irradiation and to determine the presence or absence of coordinate regulation, we analyzed the gene expression profiles of keratinocytes exposed to UVB. Normal human epidermal keratinocytes (NHEK) were irradiated with 200 J/m2 of UVB. Total RNA will be extracted at 0, 6, 12 and 24 h after irradiation (duplicate) for analysis on the Glyco gene chip. Several reports have shown that exposure to UV light can regulate levels of galectin in skin. This study seeks to analyze the changes in all galectin gene expression profiles post-UVB irradiation to determine the presence or absence of coordinate regulation. In this study, normal human keratinocytes were irradiated with 200J/m2 of UVB. Total RNA was extracted at 0, 6, 12, and 24-hour post irradiation time points, in duplicate. Samples were hybridized and analyzed using the GLYCOv2 array.

Dataset Information

Erbb2 regulates inflammation and proliferation in the skin after ultraviolet irradiation.

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Erbb2 regulates inflammation and proliferation in the skin after ultraviolet irradiation.

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