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.Intravital Multiphoton Microscopy for Dissecting Cellular Dynamics in Arthritic Inflammation and Bone Destruction
Osteoclasts are giant bone-resorbing polykaryons that differentiate from mononuclear macrophage/monocyte-lineage hematopoietic precursors. They play critical roles not only in normal bone homeostasis (remodeling) but also in the pathogenesis of bone-destructive disorders such as osteoporosis and rheumatoid arthritis. However, how the activity of mature osteoclasts is regulated in vivo remains unclear. To answer this question, we recently developed an advanced imaging system to visualize living bone tissues with intravital multiphoton microscopy. Using this system, we succeeded in visualization of mature osteoclasts in livi...
Source: Springer protocols feed by Molecular Medicine - April 7, 2014 Category: Molecular Biology Source Type: news
Label-Free Imaging of Adipogenesis by Coherent Anti-Stokes Raman Scattering Microscopy
Label-free imaging technologies to monitor the events associated with early, intermediate and late adipogenic differentiation in multipotent mesenchymal stromal cells (MSCs) offer an attractive and convenient alternative to conventional fixative based lipid dyes such as Oil Red O and Sudan Red, fluorescent labels such as LipidTOX, and more indirect methods such as qRT-PCR analyses of specific adipocyte differentiation markers such as peroxisome PPARγ and LPL. Coherent anti-Stokes Raman scattering (CARS) microscopy of live cells is a sensitive and fast imaging method enabling evaluation of the adipogenic differentiati...
Source: Springer protocols feed by Molecular Medicine - April 7, 2014 Category: Molecular Biology Source Type: news
Sample Processing, Protocol, and Statistical Analysis of the Time-of-Flight Secondary Ion Mass Spectrometry (ToF-SIMS) of Protein, Cell, and Tissue Samples
Time-of-flight secondary ion mass spectrometry (ToF-SIMS) is increasingly being used in analysis of biological samples. For example, it has been applied to distinguish healthy and osteoarthritic human cartilage. This chapter discusses ToF-SIMS principle and instrumentation including the three modes of analysis in ToF-SIMS. ToF-SIMS sets certain requirements for the samples to be analyzed; for example, the samples have to be vacuum compatible. Accordingly, sample processing steps for different biological samples, i.e., proteins, cells, frozen and paraffin-embedded tissues and extracellular matrix for the ToF-SIMS are presen...
Source: Springer protocols feed by Molecular Medicine - April 7, 2014 Category: Molecular Biology Source Type: news
Bayesian Systems-Based Genetic Association Analysis with Effect Strength Estimation and Omic Wide Interpretation: A Case Study in Rheumatoid Arthritis
Rich dependency structures are often formed in genetic association studies between the phenotypic, clinical, and environmental descriptors. These descriptors may not be standardized, and may encompass various disease definitions and clinical endpoints which are only weakly influenced by various (e.g., genetic) factors. Such loosely defined complex intermediate clinical phenotypes are typically used in follow-up candidate gene association studies, e.g., after genome-wide analysis, to deepen the understanding of the associations and to estimate effect strength. (Source: Springer protocols feed by Molecular Medicine)
Source: Springer protocols feed by Molecular Medicine - April 7, 2014 Category: Molecular Biology Source Type: news
Genome-Wide Genetic Study in Autoimmune Disease-Prone Mice
Mouse models of autoimmune diseases provide invaluable insights into the cellular and molecular bases of autoimmunity. Genetic linkage studies focusing on their abnormal quantitative phenotypes in relation to the loss of self-tolerance will lead to the identification of polymorphic genes that play pivotal roles in the genetic predisposition to autoimmunity. In this chapter, we first overview the basic concepts in the statistical genetics and then provide guides to genotyping microsatellite DNA markers and to quantitative trait loci mapping using a MAPMAKER program. (Source: Springer protocols feed by Molecular Medicine)
Source: Springer protocols feed by Molecular Medicine - April 7, 2014 Category: Molecular Biology Source Type: news
Screening for Novel Serum Biomarker for Monitoring Disease Activity in Rheumatoid Arthritis Using iTRAQ Technology-Based Quantitative Proteomic Approach
Useful biomarkers, which enable the prediction of drug susceptibility, identification of side effects, and/or evaluation of disease activity during drug treatment, are urgently needed to select adequate drugs for patients. Gene mutation status, protein expression levels in a biopsy, and serum proteins are often used as biomarkers. One of the methods to screen for protein biomarkers involves quantitative proteomic approaches using mass spectrometry. Owing to the development of quantitative proteomic approaches, the efficiency of identifying novel biomarkers from clinical samples has improved. In particular, isobaric tag for...
Source: Springer protocols feed by Molecular Medicine - April 7, 2014 Category: Molecular Biology Source Type: news
Mouse Model of Experimental Dermal Fibrosis: The Bleomycin-Induced Dermal Fibrosis
Relevant animal models are essential tools to investigate in depth the pathogenesis of autoimmune disease. Systemic sclerosis (SSc) is an autoimmune connective tissue disorder that affects particularly the skin. SSc is characterized by vasculopathy, immune disturbances, and fibrosis. Expression of each of the three pathologic features varies among SSc patients leading to disease heterogeneity and variable organ manifestations. Several animal models of SSc are available; however, some models display inflammation followed by fibrosis, whether some others primarily mimic autonomous fibroblast activation. Here, we describe the...
Source: Springer protocols feed by Molecular Medicine - April 7, 2014 Category: Molecular Biology Source Type: news
Induction of De Novo Autoimmune Disease in Normal Mice upon Repeated Immunization with Antigen
We describe here a novel and completely reproducible experimental technique that can induce systemic autoimmunity or systemic lupus erythematosus (SLE) in mice otherwise not prone to spontaneous autoimmune disease. This protocol involves the repeated immunization of mice with the same antigen. This rather simple technique enables us to perform exact and quantitative in vivo animal experiments with great accuracy. (Source: Springer protocols feed by Molecular Medicine)
Source: Springer protocols feed by Molecular Medicine - April 7, 2014 Category: Molecular Biology Source Type: news
ERRATUM TO: Whole-Mount In Situ Hybridization on Murine Skeletogenic Tissues
(Source: Springer protocols feed by Molecular Medicine)
Source: Springer protocols feed by Molecular Medicine - April 3, 2014 Category: Molecular Biology Source Type: news
Hematopoietic Stem Cell Cultures and Assays
The adult hematopoietic system is repopulated in its entirety from a rare cell type known as hematopoietic stem cells (HSCs) that reside in the marrow space throughout the skeletal system. Here we describe the isolation and identification of HSCs both phenotypically and functionally. (Source: Springer protocols feed by Molecular Medicine)
Source: Springer protocols feed by Molecular Medicine - February 3, 2014 Category: Molecular Biology Source Type: news
RANKL-Based Osteoclastogenic Assays from Murine Bone Marrow Cells
Osteoclasts are the only cell type in the body to resorb bone. Osteoclasts play a critical role in physiologic and pathologic bone remodeling. Many genetic mouse models affect the skeleton by regulating osteoclast function directly or indirectly. This protocol describes a procedure for generating osteoclasts from mouse bone marrow cells using macrophage colony stimulating factor and receptor activator of NF-κB ligand. (Source: Springer protocols feed by Molecular Medicine)
Source: Springer protocols feed by Molecular Medicine - February 3, 2014 Category: Molecular Biology Source Type: news
Isolation and Culture of Neonatal Mouse Calvarial Osteoblasts
This chapter describes the isolation and culture of neonatal mouse calvarial osteoblasts. This primary cell population is obtained by sequential enzymatic digestion of the calvarial bone matrix and is capable of differentiating in vitro into mature osteoblasts that deposit a collagen extracellular matrix and form mineralized bone nodules. Maturation of the cultures can be monitored by gene expression analyses and staining for the presence of alkaline phosphatase or matrix mineralization. This culture system, therefore, provides a powerful model to test how various experimental conditions, such as the manipulation of gene e...
Source: Springer protocols feed by Molecular Medicine - February 3, 2014 Category: Molecular Biology Source Type: news
Bone Marrow Stromal Cell Assays: In Vitro and In Vivo
Populations of bone marrow stromal cells (BMSCs, also known as bone marrow-derived “mesenchymal stem cells”) contain a subset of cells that are able to recapitulate the formation of a bone/marrow organ (skeletal stem cells, SSCs). The biological properties of BMSC cultures are assessed by a variety of assays, both in vitro and in vivo. Application of these assays in an appropriate fashion provides a great deal of information on the role of BMSCs, and the subset of SSCs, in health and in disease. (Source: Springer protocols feed by Molecular Medicine)
Source: Springer protocols feed by Molecular Medicine - February 3, 2014 Category: Molecular Biology Source Type: news
Isolation and Culture of Murine Primary Chondrocytes
To identify factors that are necessary and sufficient for chondrocyte hypertrophic differentiation and cartilage matrix mineralization, primary chondrocyte culture models have been developed. Here we describe the isolation, short-term and long-term culture, and analysis of primary costal chondrocytes from the mouse. Briefly, sternae and rib cages from neonatal pups are dissected, and chondrocytes are isolated via enzymatic digestions. Chondrocytes are then plated at high density and cultured in the presence of ascorbic acid and beta-glycerophosphate as well as various recombinant proteins to promote or inhibit hypertrophic...
Source: Springer protocols feed by Molecular Medicine - February 3, 2014 Category: Molecular Biology Source Type: news
Analysis of Chondrogenesis Using Micromass Cultures of Limb Mesenchyme
High-density micromass cultures of embryonic mesenchymal cells have proved to be an invaluable model for studying the entire chondrogenic program, from precartilaginous condensations through to chondrocyte hypertrophy. This culture model also provides a powerful system in which to explore the function of various factors in the commitment and differentiation of mesenchymal cells to the chondrogenic lineage. In this regard, micromass cultures provide a consistent and robust model for investigating the effects of genetic manipulations on skeletal phenotypes and for delineating their molecular basis. In this methods chapter, t...
Source: Springer protocols feed by Molecular Medicine - February 3, 2014 Category: Molecular Biology Source Type: news
