Transcriptome analysis on in vivo derived laser microbeam microdissected cells. Analysis of smooth muscle transcriptomes

Abstract

Smooth muscle cells (SMCs) are found in the respiratory, urogenital, circulatory and digestive systems. They provide contractility and structural support for those organs and perform multiple physiological important functions, such as modulation of blood pressure, regulation of airway resistance, and control of gastrointestinal and genitourinary motility. Moreover, SMC are involved in several pathological conditions such as atherosclerosis, restenosis, and asthma.SMC have a typical morphology with spindle shape, elongated nucleus and eosinophilic staining properties, and are treated as a distinct cell type in the literature. However, they originate from distant progenitor cell populations and have unique features in different organs. This challenges the view of SMC as being a distinct cell type. The aim of the thesis has been (i) to develop, refine, and evaluate protocols for in situ transcriptomics (IST), and (ii) to apply the method to resolve how SMC transcriptomes in different subpopulations relate to each other, and to identify genes that discriminate between subtypes.We used the combination of laser microdissection and cDNA microarray technology to monitor transcriptome profiles of SMC subpopulations in their true in vivo environment. SMC cells were captured from the abdominal aorta, the small intestines, urether, stomach, and bladder in embryonic mice. Cardiac muscle, skeletal muscle, cartilage, and whole blood were included as out-groups. The method was extensively validated in terms of its fidelity and reproducibility. Hierarchical clustering showed that the SMC typical phenotype is mediated by a limited number of genes, and that the transcriptomes otherwise appear to be unrelated. Moreover, a large number of vascular SMC specific/selective genes, and several bladder and urether SMC specific/selective genes were identified. However, we did not identify genes that were specific for gut or stomach SMC. This represents the largest systematic molecular characterization of SMC subtypes to date.