Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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A Human Fetal Cochlear Cell Atlas Reveals a Regulatory Blueprint for Spatial Patterning - scRNA-seq

ABSTRACT: Human fetal cochleae were collected from 11-16 PCW donors (each n=2) and immediately preserved in ice-cold oxygenated artificial tissue preservation solution. Under microscopic guidance, the non-osseous cochlear tissues were meticulously dissected to expose internal structures. Tissue dissociation was achieved through 0.125% trypsin digestion at 37°C for 20 minutes, followed by mechanical trituration. The resulting cell suspension was filtered through a 40 μm strainer and subjected to red blood cell lysis. After viability assessment using LUNA-FL automated counting with LIVE/DEAD staining, single-cell suspensions exceeding 80% viability were processed for library preparation. Libraries were constructed using the 10X Genomics Chromium Single Cell 3' platform (v3.1) and sequenced on Illumina NovaSeq 6000 with 150 bp paired-end reads.

INSTRUMENT(S): Illumina NovaSeq 6000

ORGANISM(S): Homo sapiens

SUBMITTER: Jiacheng Fan

PROVIDER: E-MTAB-16336 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:We performed a microarray analysis of auditory midbrain (inferior colliculus, IC) mRNA from young adult CBA mice (controls) with good hearing, middle aged (MA) with good hearing, and old mild (MP) and severe (SP) presbycusic CBA mice. Fold Change data derived from RMA normalization revealed that the overall GABA receptor alpha 6 expression profiles for MA, MP and SP were down-regulated relative to young adult controls with good hearing. Relative real-time PCR for five GABA receptors confirmed this age-related down regulation quantitatively. Functional hearing data: Auditory Brainstem Responses (ABR) enriched the analysis to select the probe-sets that changed with age and hearing loss by the linear regression best-fit line model technique. GABA receptor genotype-phenotype correlations with auditory functional data indicated that GABA-receptor subtypes are under expressed in SP mice. Hierarchical clustering (HC) analyses yielded statistical significance of normalized GeneChip data Real-time PCR showed that Gabra6, GABA B receptor 1 (Gabbr1), and Gaba transporter protein Slc32a1 may be involved in physiological changes that occur in age-related hearing loss. Presbycusis – age-related hearing loss – is the number one communicative disorder of our aged population. In this study we analyzed gene expression for a set of GABA receptors in the inferior colliculus of aging CBA mice using the Affymetrix GeneChip MOE430A. Functional phenotypic hearing changes from RMA normalized microarray data (39 replicates) in four age-groups, Young Controls and Middle aged mice with good hearing, mild and sever e presbycusis from old mice. Fold change gene expression derived from RMA normalized data were first subjected to one-way ANOVA, and then linear regression was performed. The selected gene expression changes were confirmed by relative real-time relative to young adult controls with good hearing. Statistically significant and real time PCR confirmed GABA receptor genes; Gabra6, GABA B receptor 1 (Gabbr1), and Gaba transporter protein Slc32a1, may be involved in physiological changes that occur in age-related hearing loss. Lastly, gene expression measures of each age group were correlated with pathway/network relationships relevant to the inferior colliculus using Pathway Architect, to identify key pathways consistent with the gene expression changes observed In the study of Expression changes in IC GABA receptors in the Auditory Midbrain of young adult and aging presbycusis mice total of thirty nine chips were used. The normal aging mice were in Four groups Young adults Controls with good hearing (8 mice, 8 MOE430A GeneChips), Middle aged group with good hearing ( 17 mice, 17 MOE430A GeneChips), Mild Presbycusis with limited hearing loss (9 mice, 9 MOE430A GeneChips) and Severe Presbycusis (5 mice, 5 MOE430A GeneChips).

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A Human Fetal Cochlear Cell Atlas Reveals a Regulatory Blueprint for Spatial Patterning - scRNA-seq

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