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Project ID: QLK6-CT-2002-02283
Finanziato nell'ambito di: FP5-LIFE QUALITY
Paese: Italy

Gene expression profiling in T cells with ageing

Genomic studies were carried out on T cell samples in collaboration with Unilever group in Colworth. The experimental design consisted in collecting total RNA from donors of different classes of age and compared them using microarray technology in order to evaluate differences in gene expression related to age. To avoid that gene expression data will be complicated by intrinsic intra-individual variation in samples we formerly conducted a pilot study of intra and inter-individual gene variability in young healthy donor-derived CD3+ T-cells. In fact the concept of individual variability was identified as a relevant problem when performing microarrays analysis and is crucial for the best evaluation of mRNA transcript levels when these determinations are performed employing fresh blood cells. PCA analysis, performed on one subject in a six-month period, revealed that a very small portion of gene expression profiles varies over-time. In fact the vast majority (>94%) of gene transcripts are stable over time of blood draw. Moreover, an in-depth investigation of gene expression profiles allowed us to quantify the individual variability using variance ratio or F statistic by dividing the variance of the expression levels among individuals by the variance within individuals and further confirmed using ANOVA. Finally we extracted a list of the most interesting genes, which identified an "individual molecular signature".

Additionally, to take into account the different percentage of T cells present in the blood of each donor at each time and for better understanding the contribution of the various T cell subsets to RNA expression profile, phenotypic analyses was assessed on part of the T cells used for microarray analysis. In particular, we assessed T cell subsets relevant to immunosenescence as the percentage of CD4+ or CD8+ and percentage of peripheral and central memory on CD4+ or CD8+ T cell subpopulations. Statistical results showed significant differences in lymphocytes composition between the various donors (P<0,01), and no significant differences for the same donor samples at different times (P>>0,01). In conclusion, each subject shows its own pattern of T cell subpopulations that it seem not to change over-time.
These results allowed us to progress with our experimental design aimed at characterizing T cells gene expression related to age.

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University of Bologna
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