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Total RNA and MicroRNA Transcriptomic Responses of Human Retinal Müller Glial Cells to Infection with Toxoplasma gondii Tachyzoites

Year: 2025

Type: article

Abstract: Purpose: . The goal of this research was to delineate potential involvements of Müller glial cells in ocular toxoplasmosis at a molecular level. Design: Laboratory-based study. Samples: plus noninfected cells. Methods: tachyzoites (multiplicity of infection of 5), or incubated in parallel without infection. Total RNA and small RNA were extracted from cell monolayers, sequenced on the Illumina NovaSeq 6000 and NextSeq 550 platforms, respectively, and aligned to GRCh38. Transcriptomic responses to infection with each strain were compared for differential expression (false discovery rate <5% and twofold change). These data were interrogated for enrichment of Reactome and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, and Gene Ontology in InnateDB; putative transcription factor binding sites in HOMER; and potential microRNA-mRNA interactions in multiMiR. Main Outcome Measures: Total and small RNA transcriptomes. Results: 6.3% of total RNA and 2.4% of small RNA changed in GT1-infected cells (582 upregulated and 210 downregulated transcripts and 20 upregulated microRNAs) versus 4.3% of total RNA and 1.5% of small RNA in GPHT-infected cells (400 upregulated and 137 downregulated transcripts and 12 upregulated microRNAs). Seventy-six transcripts and 4 microRNAs were different between strains; most were increased by both, but GT1 induced higher levels than GPHT. Enriched pathways and ontologies were dominated by DNA replication and intracellular metabolic activities, and the immune response for GT1 and GPHT. Seven of 8 transcription factor binding sites were shared for GT1 and GPHT infections, all overexpressed, including sites for p65/RELA and E2F family members. Across the strains, miR-18a-5p was the most connected microRNA in predicted mRNA target networks. Conclusions: tachyzoites, consistent with a central role in the characteristic pathology of ocular toxoplasmosis. Financial Disclosures: Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Source: Ophthalmology Science

DOI: 10.1016/j.xops.2025.100971