Small RNA next generation sequencing of mice skin biopsies from acetone-treated healthy skins and DMBA/PMA-treated hyperplastic skins, papillomas and cutaneous Squamous Cell Carcinomas
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ABSTRACT: FVB/N mice were subjected to 7,12-Dimethylbenz[a]anthracene (DMBA) two-hit multistage skin carcinogenesis protocol. Mice were topically treated with 200 nmol of DMBA in 0.2 mL acetone then twice weekly for six weeks with 5 nmol PMA (Phorbol 12-myristate 13-acetate). A second hit of DMBA was performed on the eighth week followed by the resumption of PMA treatment for 14 more weeks. Control mice were only topically treated with 0.2 mL acetone vehicle. Healthy skins (acetone-treated), hyperplastic skins, papillomas and tumors were harvested throughout the protocol and biopsies were frozen for RNA extraction.
Project description:FVB/N mice were subjected to 7,12-Dimethylbenz[a]anthracene (DMBA) two-hit multistage skin carcinogenesis protocol. Mice were topically treated with 200 nmol of DMBA in 0.2 mL acetone then twice weekly for six weeks with 5 nmol PMA (Phorbol 12-myristate 13-acetate). A second hit of DMBA was performed on the eighth week followed by the resumption of PMA treatment for 14 more weeks. Control mice were only topically treated with 0.2 mL acetone vehicle. Healthy skins (acetone-treated), hyperplastic skins, papillomas and tumors were harvested throughout the protocol and biopsies were frozen for RNA extraction. Small RNA libraries were generated from total RNAs of control skins, hyperplastic skins, papillomas and cSCCs biopsies (n=5 in each group) corresponding to a total of 20 samples, and sequenced on the Applied Biosystems SOLiD System.
Project description:Small RNA next generation sequencing of mice skin biopsies from acetone-treated healthy skins and DMBA/PMA-treated hyperplastic skins, papillomas and cutaneous Squamous Cell Carcinomas
Project description:We profiled the transcriptomes of sorted neutrophils from precancerous and tumors in a DMBA/PMA cutenous squamous cell carcinoma, as well as the healthy adjacent tissue.
Project description:Human α1-antitrypsin (AAT) is an abundant acute phase glycoprotein performing anti-protease, anti-apoptotic and immunomodulatory functions. Purified human plasma AAT used as a specific therapy for patients with lung emphysema and panniculitis due to inherited AAT deficiency, and it is beneficial in treating of other disorders. This therapy administered intravenously although other routes of administration are tested. Herein, we examined the transdermal application of native or recombinant AAT by using epiCS®, the 3D human epidermis equivalent reconstructed from human primary epidermal keratinocytes. Topically applied AAT protein (50µl, prepared in Hank`s balance solution, HBSS) freely diffused across epidermis layers in a concentration and time-dependent manner. We concluded that within 18 hours topically added 0.2 mg AAT well penetrates the stratum corneum and can be detected inside the cytosol of keratinocytes. Importantly, treatments with AAT did not induce obvious morphological changes in keratinocyte layers. Next, we supplemented culture medium with 100µg/ml of combined bacterial lipopolysaccharide (LPS) and peptidoglycan (PNG) mixture and incubated epiCS for 18h with or without the topical application of 0.2 mg AAT. Under these conditions, LPS/PNG triggered a strongly activation of epidermis model. Even though AAT exhibited a limited capacity to neutralize the effect of LPS/PNG, it lowered expression of IL-18 and IL-8, and preserved levels of filaggrin, an important protein for maintaining the epidermal barrier integrity. Our results suggest that the transdermal route for delivering AAT is worth exploration. If successful, this approach may offer easy-to-use therapy with AAT.
Project description:Chronic stimulation of innate immune pathways by microbial agents or damaged tissue is known to promote inflammation-driven tumorigenesis by unclarified mechanisms1-3. Here we demonstrate that mutagenic 7,12-dimethylbenz(a)anthracene (DMBA), etoposide or cisplatin induces nuclear DNA leakage into the cytosol to intrinsically activate STING (Stimulator of Interferon Genes) dependent cytokine production. Inflammatory cytokine levels were subsequently augmented in a STING-dependent extrinsic manner by infiltrating phagocytes purging dying cells. Consequently, STING-/- mice, or wild type mice adoptively transferred with STING-/- bone marrow, were almost completely resistant to DMBA-induced skin carcinogenesis compared to their wild type counterparts. Our data emphasizes, for the first time, a role for STING in the induction of cancer, sheds significant insight into the causes of inflammation-driven carcinogenesis, and may provide therapeutic strategies to help prevent malignant disease Total RNA obtained from DMBA or acetone treated wild type (WT) or STING deficient (SKO) mouse skin or skin tumor was examined for gene expression.
Project description:This study aimed to understand the in vitro behaviour of epidermal cells of African and Caucasian skin types in the context of 3D reconstructed skin. Reconstructed skins epidermis made with cells isolated from skin of African or Caucasian skin type exhibited high differences in stratification and differentiation. The objective of this study is a first global approach to identify at the protein level the differences between reconstructed skins.
Project description:DNA microarrays were used to investigate global gene expression patterns in cultured human umbilical artery endothelial cells (HUAECs) exposed to 1 nmol/L estradiol and/or 100 µg/ml oxidized low density lipoprotein (oxLDL) for 24 hours compared to control cells.
Project description:Skin is the largest organ of body, and one function of skin is protecting underly organs away from ultraviolet (UV) radiation damage. Loss of melanocyte will reduce ability of skin to against UV radiation damage. We found Bama Pig can be an ideal model studying loss of melanocyte. In this study, we performed transcriptome profiling of mRNA and long noncoding RNA in Bama pig white skin (absence of melanocyte) and black skin (existence of melanocyte) to provide dataset for clinical researches. Total of 14,900 mRNAs and 7,566 lncRNAs were expressed in the study. Results of hierarchical cluster analysis and principal component analysis (PCA) demonstrated expression of mRNAs and lncRNAs varied greatly between two color skins. 2,342 mRNA were identified as being differentially expressed, including 1,309 genes that were down-regulated in white skin and 1,033 gene that were up-regulated in white skin (P <0.05; |log2(fold change)| > 1). The genes down-regulated in white skin were associated with melanocyte biology, melanocyte function and keratin, while genes up-regulated in white skin were main associated with metabolism pathway, oxidative phosphorylation and citrate cycle (TCA cycle). In addition, we identified 120 differentially expressed lncRNAs. In those lncRNAs, 4 lncRNAs may function in skin biology (TCONS_00019024 and TCONS_00077733) or metabolic (TCONS_00042201 and TCONS_00060772). Our research provides Bama pig skins datasets and could promote clinical researches utilizing Bama pig as model.