Wilkinson, James Maxwell; (2003) Serial analysis of genes expressed in normal human glomerular mesangial cells. Doctoral thesis (Ph.D.), University College London (United Kingdom).

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Abstract

Advances in sequencing based genomics like the Human Genome Mapping Project (HGMP) have meant that the majority of the estimated human genes have been at least partially sequenced. The variation in expression of a set of essentially identical genes will provide information on the molecular basis of phenotype. Serial analysis of gene expression (SAGE) is based on the ability to assign an individual transcript to a ten base pair 'tag', and the technology facilitating rapid sampling of such tags. Glomerular mesangial cells (MC) are considered to play a major role in the development of renal disease and in vitro culturing of MC's has become a model system with which to study the molecular mechanisms of glomerular pathology. To this end, a SAGE project was undertaken to identify genes expressed in normal human mesangial cells (NHMC). Primary normal human mesangial cells were cultured for periods up to 96 hrs. A total of 46,219 tags were sampled (14,953 unique tags). Tags were mapped to 20,382 sequences. Of these 79% of tags mapped to characterised cDNAs, 16% tags mapped to ESTs. 5% of tags failed to match any database entry. The most abundant tags mapped to ribosomal genes or genes associated with the cytoskeleton. Represented in the top ten tags were the matricellular genes transgelin (1.2%), SPARC (1%) type IV collagen (0.5%) and fibronectin (0.53%), which support the notion that the MC is a producer and re-modeller of the glomerular extracellular matrix (ECM). The contractile nature of MC was apparent with the high abundance of contractile proteins like myosins and tropomyosins. Also apparent in the transcriptome were lineage specific isoforms of several genes, supporting the myoblastoid linage of MC. Comparing the transcriptomes of the MC to other libraries revealed a high correlation between cells in the same lineage as MC, such as astrocytes, smooth muscle cells and fibroblasts when compared to libraries sampled from heart, liver and various other unrelated cell lines. Understanding gene expression in the mesangial cell facilitates a greater understanding of its role in renal pathology.

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