Springer protocols feed by Molecular Medicine 
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This is an RSS file. You can use it to subscribe to this data in your favourite RSS reader or to display this data on your own website or blog.Animal Model for Cutaneous Leishmaniasis
Using cutaneous leishmaniasis of mice, the existence of so-called T helper (Th) cells type 1 and type 2 had been identified more than 20 years ago. Nowadays, it is well accepted that additional T cell populations as well as B cell-mediated immunity is required for immunity against Leishmania major. Finally, using inbred mouse strains, the relevance of genetical factors that influence anti-pathogen immunity as well as elements of the skin-immune system have been identified. This protocol describes a model for murine experimental leishmaniasis that tries to mimic natural parasite transmission by several means: (1) utilizatio...
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
Induction of Experimental Epidermolysis Bullosa Acquisita by Immunization with Murine Collagen VII
Epidermolysis bullosa acquisita (EBA) is an autoimmune subepidermal blistering disease caused by an autoreactive response against collagen VII, the major constituent of the anchoring fibrils at the epidermal basement membrane. The pathogenic relevance of collagen VII-specific autoantibodies has been conclusively demonstrated ex vivo and in experimental animals using antibody passive transfer models. To study the mechanisms of autoantibody production and tissue damage an animal model reproducing both the autoimmune response and the active skin disease is needed. In the present protocol, we describe the induction of an autoi...
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
Mouse Models of Autoimmune Blistering Diseases Induced by the Passive Transfer of Antibodies
Passive transfer of IgG into neonatal mice is a potential method of reproducing antibody-mediated blistering skin diseases. The major autoantigen for bullous pemphigoid is collagen XVII (COL17)/BP180, which is an epidermal linker transmembrane protein. A single intraperitoneal injection of human or rabbit IgG against pathogenic epitopes for COL17 can induce skin fragility in neonatal mice that express human COL17. Since amino acid sequences of the pathogenic epitopes for COL17 significantly differ between humans and rodents, the required antibodies are those that correctly target the molecule to induce the blistering pheno...
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
RNAi-Mediated Gene Function Analysis in Skin
We have recently developed a method for RNAi-mediated gene function analysis in skin (Beronja et al., Nat Med 16:821–827, 2010). It employs ultrasound-guided in utero microinjections of lentivirus into the amniotic cavity of embryonic day 9 mice, which result in rapid, efficient, and stable transduction into mouse skin. Our technique greatly extends the available molecular and genetic toolbox for comprehensive functional examination of outstanding problems in epidermal biology. In its simplest form, as a single-gene function analysis via shRNA-mediated gene knockdown, our technique requires no animal mating and may n...
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
Generation of Functional Multipotent Keratinocytes from Mouse Induced Pluripotent Stem Cells
Recent advances in reprogramming somatic cells into induced pluripotent stem cells (iPSCs) offer the possibility of developing new therapeutic approaches for the treatment of a variety of diseases, including inherited skin disorders. While the ultimate goal is the use of iPSCs in the treatment of human diseases, extensive research is still required with preclinical mouse models before iPSC technology can be introduced into the clinic. Therefore, the methodology for the derivation of multipotent keratinocytes from mouse iPSCs is of particular importance since it may allow for the assessment of the feasibility of using iPSCs...
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
Induction of Contact Hypersensitivity in the Mouse Model
Contact hypersensitivity (CHS) in the mouse model is a standard method to assess delayed type hypersensitivity (DTH) responses in the skin induced by low molecular weight chemicals that in humans cause contact dermatitis. These responses are clinically important and present as eczematous skin reactions. Here, this chapter describes the standard protocol for T cell-mediated CHS and a variation thereof, which allows to address more specific questions regarding immunologic pathomechanisms. (Source: Springer protocols feed by Molecular Medicine)
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
Bioengineered Skin Humanized Model of Psoriasis
This protocol describes the generation of a skin humanized mouse model for psoriasis using bioengineering approaches. This method is relatively simple, highly reproducible and ensures the obtention of a large and homogenous number of engrafted animals bearing a portion of human skin with psoriatic phenotype. The technique can employ cells from skin biopsies and blood samples from non-related healthy human donors (allogeneic version), as well as skin and blood cells from psoriatic patients (autologous version). In both cases, the psoriatic phenotype was developed after intradermal administration of in vitro derived T1 lymph...
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
Molecular Dermatology Comes of Age
This article highlights several major developments in molecular experimental and clinical dermatology. (Source: Springer protocols feed by Molecular Medicine)
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
Reconstitution of Skin Fibrosis Development Using a Tissue Engineering Approach
Skin fibrosis is involved in several pathologies as hypertrophic scar or scleroderma. The determination of the mechanisms at the origin of these problems is however difficult due to the low number of in vivo models. To bypass this absence of animal models, studies typically use human pathological cells cultured in a monolayer way on plastic. However, cell behavior is different according to the fact that cells are on plastic or embedded in matrix. Using a tissue engineering method, we have developed new in vitro models to study these pathologies of the skin. Human pathological cells are used to reconstitute a three dimensio...
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
Analysis of Cell Movement Between Skin and Other Anatomical Sites In Vivo Using Photoconvertible Fluorescent Protein “Kaede”-Transgenic Mice
Clarification of the spatiotemporal regulation and function of immune cells within the skin is critical to the understanding of the role of immune cells and the skin in immune homeostasis. Here, we describe a novel assay system for monitoring cell movements in the entire body using the photoconvertible fluorescent protein “Kaede”-transgenic (Tg) mice. We can label immune cells by the change in color of Kaede from green to red in these cells following exposure to violet light and track these cells in the entire body. The Kaede-Tg system is an ideal tool for monitoring precise cellular movements between the skin ...
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
Detection, Enumeration, and Characterization of Immune Cells Infiltrating Melanoma Tumors
Tumor-infiltrating immune cells have long been thought to affect tumor growth. In recent years, large retrospective studies have shown that the nature and polarization of the immune cells found within the tumor microenvironment impact not only the growth of the primary tumor, but also disease progression and patient survival. This has triggered considerable interest for an in depth analysis of the tumoral immune microenvironment and has created a need for standardized methods to characterize tumor-infiltrating immune cells. Here, we describe three approaches that can be used in mouse and human melanoma tumors. (Source: Spr...
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
Isolation of Melanoma Tumor-Initiating Cells from Surgical Tissues
A new model of cancer progression has been put forward that predicts existence of tumor stem cells (TSCs) in the heterogeneous bulk tumor mass that self-renew, are resistant to chemo- and radiotherapies, and sustain tumor growth during the course of its progression or relapse (Ailles and Weissman, Curr Opin Biotechnol 18:460–466, 2007; Chan et al., Proc Natl Acad Sci U S A 106:14016–14021, 2009; D’Angelo and Wicha, Prog Mol Biol Transl Sci 95:113–158, 2010; O’Brien, Semin Radiat Oncol 19:71–77, 2009; Park et al., Mol Ther 17:219–230, 2009). Using most advanced methods of cell purif...
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
Analysis of Collective Invasion of Carcinoma Cells in a 3D Organotypic Model
Cancer cell invasion and dissemination from primary tumors are complex multistep mechanisms which remain poorly understood. It is now clear that cancer cells can adapt their mode of invasion to the signalling provided by the surrounding stroma. Single and collective cancer cell invasion are the two invasion features most currently observed and described by pathologists. Here we describe a three-dimensional organotypic assay that allows the study of squamous cell carcinoma cell collective invasion induced by the carcinoma associated fibroblasts. This model preserves the relationship between epithelial and mesenchymal cells,...
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
Induction of Granulocyte-Dependent Dermal-Epidermal Separation by Autoantibodies Ex Vivo
Disease models represent powerful tools used by biomedical researchers to address various questions related to the pathogenesis and the possible therapeutic targets in different diseases. To get a complete picture of the process one needs complementary animal and ex vivo disease models. While subsequent chapters dwell upon animal models, this chapter describes an ex vivo model for granulocyte-dependent dermal-epidermal separation induced by autoantibodies to the basal membrane. The strength of the described ex vivo model resides in the fact that it uses human material (the skin, autoantibodies, and cells), thus better mimi...
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
In Vitro Pathogenicity Assay for Anti-desmoglein Autoantibodies in Pemphigus
We describe here two assays that measure the pathogenic strength of autoantibodies in blister formation: an in vitro dissociation assay using primary human epidermal keratinocytes to assess pathogenicity of anti-Dsg3 autoantibodies, and an alternative method whereby anti-Dsg3 and Dsg1 autoantibodies are injected into organ-cultured human skin specimen. (Source: Springer protocols feed by Molecular Medicine)
Source: Springer protocols feed by Molecular Medicine - December 19, 2012 Category: Molecular Biology Source Type: news
