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.Pressure Myography to Study the Function and Structure of Isolated Small Arteries
Small arteries play an important role in regulation of peripheral resistance and organ perfusion. Here we describe a series of methods allowing measurements in pressurized segments of small arteries from the systemic and coronary circulation of mice as well as other species. The pressure myography techniques described include measurements of wall structure, wall stress, strain, and myogenic tone. The pressurized perfused small arteries also allow evaluation of responses to increases in pressure, flow, and drugs, where the main readout is changes in vascular diameter. (Source: Springer protocols feed by Molecular Medicine)
Source: Springer protocols feed by Molecular Medicine - July 31, 2015 Category: Molecular Biology Source Type: news
Wire Myography to Study Vascular Tone and Vascular Structure of Isolated Mouse Arteries
Atherosclerosis is characterized by endothelial dysfunction and alterations in vascular reactivity, which can be investigated by wire myography. The method allows ex vivo monitoring of the transversal isometric tension developed by a vessel segment in response to different pathophysiological stimuli. Here we describe in detail how to use the wire myograph to evaluate endothelial function and vasoconstrictor or vasodilator properties of the vessel, as well as to identify and characterize different factors and molecular pathways that control vascular tone. We also describe how to use the wire myograph to analyze biomechanica...
Source: Springer protocols feed by Molecular Medicine - July 31, 2015 Category: Molecular Biology Source Type: news
Glucose and Insulin Tolerance Tests in the Mouse
In vivo metabolic tests are highly valuable to determine whether atherosclerosis progression in mouse models is accompanied by carbohydrate metabolism alterations such as glucose intolerance and insulin resistance. In this chapter, we describe protocols to perform in the mouse glucose and insulin tolerance tests, two metabolic assays which evaluate the glucose tolerance and the insulin sensitivity, respectively. (Source: Springer protocols feed by Molecular Medicine)
Source: Springer protocols feed by Molecular Medicine - July 31, 2015 Category: Molecular Biology Source Type: news
In Vitro Macrophage Phagocytosis Assay
The key roles of macrophages in atherosclerosis include the phagocytosis of apoptotic and necrotic cells and cell debris, whose accumulation in atherosclerotic lesions exacerbates inflammation and promotes plaque vulnerability. Evidence is accumulating that macrophage phagocytic functions peak at the early stages of atherosclerosis and that the reduced phagocytosis at the late stages of disease leads to the generation of necrotic cores and a defective resolution of inflammation, which in turn promotes plaque rupture, thrombus formation, and life-threatening acute ischemic events (myocardial infarction and stroke). The impa...
Source: Springer protocols feed by Molecular Medicine - July 31, 2015 Category: Molecular Biology Source Type: news
Quantification of In Vitro Macrophage Cholesterol Efflux and In Vivo Macrophage-Specific Reverse Cholesterol Transport
Promotion of reverse cholesterol transport (RCT) is thought to be a major HDL-mediated mechanism for protecting against atherosclerosis. Preclinical studies support the concept that increasing cholesterol efflux from macrophages may confer atheroprotective benefits independently of the plasma HDL-cholesterol concentration. The application of the macrophage-to-feces RCT method in genetically engineered mice has provided evidence that this major HDL property correlates closely with changes in atherosclerosis susceptibility. This chapter provides details on the methodologies currently used to measure in vitro cholesterol effl...
Source: Springer protocols feed by Molecular Medicine - July 31, 2015 Category: Molecular Biology Source Type: news
Quantification of Cellular Proliferation in Mouse Atherosclerotic Lesions
Excessive cell proliferation within atherosclerotic plaques plays an important role in the progression of atherosclerosis. Macrophage proliferation in particular has become a major focus of attention in the cardiovascular field because it appears to mediate most of macrophage expansion in mouse atherosclerotic arteries. Therefore, quantification of cell proliferation is an essential part of the characterization of atherosclerotic plaques in experimental studies. This chapter describes two variants of a simple immunostaining protocol that allow for the quantification of cellular proliferation in mouse atherosclerotic lesion...
Source: Springer protocols feed by Molecular Medicine - July 31, 2015 Category: Molecular Biology Source Type: news
Quantification of Apoptosis in Mouse Atherosclerotic Lesions
We describe the morphological and immunohistochemical methods used in our laboratory to detect apoptotic cells in animal and human atherosclerotic plaques. (Source: Springer protocols feed by Molecular Medicine)
Source: Springer protocols feed by Molecular Medicine - July 31, 2015 Category: Molecular Biology Source Type: news
In Vitro Differentiation of Naïve CD4+ T Cells: A Tool for Understanding the Development of Atherosclerosis
A complete knowledge of atherosclerosis requires a better understanding of how innate and adaptive immunity operate systemically and locally within the arterial wall. T-helper 1 (Th1) lymphocyte responses have proatherogenic effects in mice, contrasting with the responses of T regulatory cells (Tregs), which can suppress growth of atherosclerotic lesions. An imbalance in the differentiation of T-helper cells may therefore impact the development, size, and stability of atherosclerosis plaques. This chapter describes a method to isolate naïve CD4+ T cells from atherosclerosis-prone mouse peripheral blood lymphocytes and...
Source: Springer protocols feed by Molecular Medicine - July 31, 2015 Category: Molecular Biology Source Type: news
Flow Cytometric Analysis of Immune Cells Within Murine Aorta
The immune system plays a critical role in the modulation of atherogenesis at all stages of the disease. However, there are many technical difficulties when studying the immune system within murine aortas. Common techniques such as PCR and immunohistochemistry have answered many questions about the presence of immune cells and mediators of inflammation within the aorta yet many questions remain unanswered due to the limitations of these techniques. On the other hand, cumulatively the flow cytometry approach has propelled the immunology field forward but it has been challenging to apply this technique to aortic tissues. Her...
Source: Springer protocols feed by Molecular Medicine - July 31, 2015 Category: Molecular Biology Source Type: news
Immunostaining of Lymphocytes in Mouse Atherosclerotic Plaque
Immunostaining can be defined as a method for identification of constituents in situ, e.g., proteins, using specific antigen–antibody interactions, visualized by color or fluorescence. In experimental research of atherosclerosis, immunostaining is a key method used for the characterization of the cellular composition in the atherosclerotic plaques. This includes the quantification of macrophages, smooth muscle cells, and lymphocytes—especially T lymphocytes. In this chapter we focus on basic protocols for characterization of lymphocytes and/or local T lymphocyte-related antigens in mouse atherosclerotic lesions...
Source: Springer protocols feed by Molecular Medicine - July 31, 2015 Category: Molecular Biology Source Type: news
Peripheral Blood Mononuclear Cells: Isolation, Freezing, Thawing, and Culture
The work with peripheral blood mononuclear cells (PBMCs), which comprise lymphocytes and monocytes, is indispensable in immunological diagnostics and research. The isolation of PBMCs takes advantage of differences in cell density of the different blood components. Density gradient centrifugation of diluted whole blood layered over a density gradient medium yields PBMCs; two subsequent washing steps remove remaining platelets. To store the cells for future assays, they can be frozen and thawed when required. Dimethyl sulfoxide (DMSO) serves as a cryoprotectant for freezing PBMCs, but must be removed by washing after thawing...
Source: Springer protocols feed by Molecular Medicine - July 5, 2015 Category: Molecular Biology Source Type: news
Experimental Autoimmune Encephalomyelitis in Mice
Experimental autoimmune encephalitis (EAE), the animal model of multiple sclerosis (MS), has provided significant insight into the mechanisms that initiate and drive autoimmunity. Several central nervous system proteins and peptides have been used to induce disease, in a number of different mouse strains, to model the diverse clinical presentations of MS. In this chapter, we detail the materials and methods used to induce active and adoptive EAE. We focus on disease induction in the SJL/J, C57BL/6, and BALB/c mouse strains, using peptides derived from proteolipid protein, myelin basic protein, and myelin oligodendrocyte gl...
Source: Springer protocols feed by Molecular Medicine - July 5, 2015 Category: Molecular Biology Source Type: news
Actively Induced Experimental Autoimmune Encephalomyelitis in Rats
The rat and especially a number of inbred rat strains are very well suited for modeling multiple sclerosis (MS). Experimental autoimmune encephalomyelitis (EAE), the model of MS, can be induced by active or passive immunization. Active immunization can be performed with different myelin proteins or peptides thereof. Passive immunization is performed by transfer of myelin-specific T cells. Most known is EAE induced with myelin basic protein (MBP) in LEW (RT1l) rats that results in monophasic disease and EAE induced with myelin oligodendrocyte glycoprotein (MOG) in DA (RT1av1) rats that leads to relapsing remitting or chroni...
Source: Springer protocols feed by Molecular Medicine - July 5, 2015 Category: Molecular Biology Source Type: news
Optical Coherence Tomography to Assess Neurodegeneration in Multiple Sclerosis
Retinal spectral domain optical coherence tomography (OCT) has emerged as a clinical and research tool in multiple sclerosis (MS) and optic neuritis (ON). This chapter summarizes a short OCT protocol as included in international consensus guidelines. The protocol was written for hands-on style such that both clinicians and OCT technicians can make use of it. The protocol is suitable for imaging of the optic nerve head and macular regions as a baseline for follow-up investigations, individual layer segmentation, and diagnostic assessment. (Source: Springer protocols feed by Molecular Medicine)
Source: Springer protocols feed by Molecular Medicine - July 5, 2015 Category: Molecular Biology Source Type: news
Neuropathological Techniques to Investigate Central Nervous System Sections in Multiple Sclerosis
Immunohistochemical techniques (IHC) and in situ hybridization (ISH) are widely used techniques to study the expression of proteins and messenger RNAs in tissues and are extremely important to confirm and interpret biochemical and molecular results from the same tissues. Investigation of human brain by IHC and ISH therefore still plays an important role in the elucidation of pathogenetic mechanisms in diseases such as multiple sclerosis. In this review we describe the processing of human brain tissues as well as basic and advanced immunohistochemical staining and ISH techniques used for neuropathological analysis of such p...
Source: Springer protocols feed by Molecular Medicine - July 5, 2015 Category: Molecular Biology Source Type: news
