Investigating the Effect of Surgical Menopause on Gene Expression Changes in Retinal Ganglion Cells and Astrocytes Öffentlichkeit
Jiang, Xinyi (Spring 2024)
Published
Abstract
Purpose: Menopause is one of the risk factors for glaucoma. Animal studies have shown significantly worse visual function in ovariectomized (OVX) rodents after optic nerve injury. There is evidence the estrogen may interaction with retinal ganglion cell (RGC) and retinal astrocytes. However, the gene expression changes in RGC and retinal astrocytes after loss of estrogen by menopause have not been explored.
Method: Young (2-4months) female Brown Norway rats (Charles River) equally divided into naïve (n=6) and OVX (n=6) groups. The retinas have been dissected and dissociated. RGC and astrocytes were sorted, and their mRNA were isolated for bulk RNA sequencing. Principle Component analysis (PCA) was used for verifying sample purity. DESeq2 was used for finding differentially expressed genes between OVX and control group in RGC and Astrocyte separately. Gene Ontology (GO) Enrichment Analysis and (Kyoto Encyclopedia of Genes and Genomes) KEGG pathway analysis were performed.
Results: Samples are clustered and separated by cell type along PC1, capturing 39.5% of the variation. RGC samples exhibited significantly higher counts for RGC specific markers and no photoreceptor marker expression. The RGC OVX samples showed 135 upregulated genes (0.91%) and 130 downregulated genes (0.87%) compared to the RGC control group. The Astrocyte OVX group had 143 upregulated genes (0.96%) and 54 downregulated genes (0.36%) compared to the astrocyte control group (adjusted p <0.01, log2 fold change > 0.5). Both groups showed a significant increase in immune receptor activity and purinergic nucleotide receptor activity (adjusted p <0.05, log2 fold change > 0.5). Analysis of the top 40 gene pathways revealed distinct clusters correlated within RGC (22 pathways) and astrocyte groups (13 and 7 pathways), (adjusted p <0.01, log2 fold change > 0.5). Overlapped pathways between RGC and astrocytes include the TNF pathway, NFκB pathway, IL-7 pathway, MAPK pathway, and pathways in cancer. In the TNF pathway, 17 genes overlapped between the two groups, playing roles in leukocyte recruitment and activation, inflammatory cytokine production, and cell survival. In the NFκB pathway, 11 overlapping genes are responsible for apoptosis, inflammation, and cell survival.
Conclusion: Our results suggest that OVX induces significant changes in gene expression and pathway activations related to immune and inflammatory responses in RGCs and astrocyte cells. These alterations after OVX suggest a systemic adjustment in the retinal environment, potentially affecting the response of these cell populations to injury or disease. Future research should be conducted in vitro to better understand the transcriptomic changes affected by estradiol (E2).
Table of Contents
Table of Contents
Introduction and Background
Glaucoma
Sex as an Important Aspect in Glaucoma
Retinal Ganglion Cell (RGC) and Retinal Astrocytes
Estrogen, Astrocytes and RGC
Motivation and goal
Methods and Material
Animals
Surgical Menopause
Retinal dissection and dissociation
RGC and Astrocyte isolation
RNA isolation
RNA-sequencing
Data Analysis
Results
RGC and retinal Astrocytes purity
Differently Expressed Genes between OVX and Control
Pathway Changes
Discussion
Works Cited
Acknowledgement
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