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RNA sequencing analysis of ARV825 target genes in senescent cells

ABSTRACT: RNA sequencing analysis of ARV825 target genes in senescent cells

PROVIDER: PRJNA591563 | ENA |

REPOSITORIES: ENA

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Project description:The presence of senescent cells in the aging/degenerating human disc is now well-recognized. Senescent cells are viable, cannot divide, remain metabolically active and accumulate within the disc over time. Molecular analysis of senescent cells in tissue, however, offers a special challenge since there are no cell surface markers for senescence which would let one use fluorescence-activated cell sorting as a method for separating out senescent cells. Here we use a novel experimental design using laser capture microdissection to selectively separately harvest senescent and non-senescent annulus cells in paraffin-embedded tissue, and then compare their gene expression with microarray analysis. An initial in vitro study using cultured human annulus cells was first performed to test whether there was any difference in identification of senescent cells using the accepted histochemical methodology vs. the immunofluoresent identification of cells positive for senescence-associated-ß-galactosidase in control cells and cells induced into stress-induced premature senescence via hydrogen peroxide exposure. No statistically significant difference was found between the 2 methods. Laser capture microdissection was used to separately harvest senescent and non-senescent cells from 11 human annulus specimens, and microarray analysis was used to determine gene expression levels. Genes with established relationships to senescence were found to be significantly upregulated in senescent cells vs. non-senescent cells. Additional genes related to cytokines, cell proliferation, and other cell processes were also identified. Disc Tissue samples were obtained from surgical disc procedures performed on patients with herniated discs and degenerative disc disease. Tissue was fixed and paraffin embedded. Standard laser capture microdissection (LCM) techniques were used to collect senescent cells. Remaining non-senescent cells were scraped from the histology slide. Total RNA was isolated and analyzed via mircoarray. Gene expression from senescent cells was compared to non-senescent cells. Eight histological samples were used to obtain both senescent and non-senescent cells. From an additional 3 samples, only senescent cells were harvested.

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RNA sequencing analysis of ARV825 target genes in senescent cells

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