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Verteporfin treatment controls morphology, phenotype, and global gene expression for cells of the human nucleus pulposus.

ABSTRACT: Cells of the nucleus pulposus (NP) are essential contributors to extracellular matrix synthesis and function of the intervertebral disc. With age and degeneration, the NP becomes stiffer and more dehydrated, which is associated with a loss of phenotype and biosynthetic function for its resident NP cells. Also, with aging, the NP cell undergoes substantial morphological changes from a rounded shape with pronounced vacuoles in the neonate and juvenile, to one that is more flattened and spread with a loss of vacuoles. Here, we make use of the clinically relevant pharmacological treatment verteporfin (VP), previously identified as a disruptor of yes-associated protein-TEA domain family member-binding domain (TEAD) signaling, to promote morphological changes in adult human NP cells in order to study variations in gene expression related to differences in cell shape. Treatment of adult, degenerative human NP cells with VP caused a shift in morphology from a spread, fibroblastic-like shape to a rounded, clustered morphology with decreased transcriptional activity of TEAD and serum-response factor. These changes were accompanied by an increased expression of vacuoles, NP-specific gene markers, and biosynthetic activity. The contemporaneous observation of VP-induced changes in cell shape and prominent, time-dependent changes within the transcriptome of NP cells occurred over all timepoints in culture. Enriched gene sets with the transition to VP-induced cell rounding suggest a major role for cell adhesion, cytoskeletal remodeling, vacuolar lumen, and MAPK activity in the NP phenotypic and functional response to changes in cell shape.

SUBMITTER: Fearing BV

PROVIDER: S-EPMC7770208 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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Verteporfin treatment controls morphology, phenotype, and global gene expression for cells of the human nucleus pulposus.

Fearing Bailey V BV Speer Julie E JE Jing Liufang L Kalathil Aravind A P Kelly Michael M M Buchowski Jacob J P Zebala Lukas L Luhmann Scott S C Gupta Munish M A Setton Lori L

JOR spine 20200730 4

Cells of the nucleus pulposus (NP) are essential contributors to extracellular matrix synthesis and function of the intervertebral disc. With age and degeneration, the NP becomes stiffer and more dehydrated, which is associated with a loss of phenotype and biosynthetic function for its resident NP cells. Also, with aging, the NP cell undergoes substantial morphological changes from a rounded shape with pronounced vacuoles in the neonate and juvenile, to one that is more flattened and spread with ...[more]

PMID: 33392449

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Project description:Purpose: It has been observed that the drug verteporfin (VP), a known disruptor of YAP-TEAD signaling, causes cell rounding and changes in phenotype and biosynthesis in human nucleus pulposus cells. In this study the time-dependent effects of VP treatment on the transcriptome of human NP cells was explored. Methods: mRNA profiles of NP cells treated for 24 hours, 48 hours, or 4 days with VP or vehicle (DMSO) control were generated for cells from 3 adult human tissue samples, using Illumina NovaSeq. Unaligned reads were trimmed based on quality score (minimum quality level (Phred) = 20, minimum read length = 25) and aligned to the whole human genome (STAR 2.6.1d; hg19) using Partek Flow software. This software suite was also used to calculate the counts/normalized counts of genes and to perform gene specific analysis (GSA), principle component analysis (PCA), hierarchical clustering, and gene set enrichment. Differentially regulated genes were considered to be those with a fold change value (VP/DMSO) for a given time point that was greater than or equal to 2 or less than or equal to negative 2 and a FDR-adjusted p-value of less than or equal to 0.05. Results: Using a data analysis workflow buit in Partek Flow, we identified 1810 differentially regulated genes at 24 hours, 4287 differentially regulated genes at 48 hours, and 9272 differentially regulated genes at 4 days. Hierarchical clustering and PCA analysis demonstrated clustering of the data by patient and time point, but separation by treatment group. We further observed that while generally the expression of fibroblastic markers decreased with VP treatment, a number of markers of NP phenotype were increased. We also identified enriched gene sets (ex. 2026 at the 4 day time point) which represented a variety of cellular components, processes, and functions potentially implicated by VP treatment. Conclusions: The data from this RNA-sequencing demonstrate that VP treatment which induces cell rounding in NP cells was also accompanied by pronounced and time-dependent changes in the transcriptome of adult NP cells in culture. The enriched gene sets and differentially regulated genes suggest a major role for cell adhesion, cytoskeletal remodeling, vacuolar lumen, and M APK activity in the NP phenotypic and functional response to changes in cell shape.

Dataset Information

Verteporfin treatment controls morphology, phenotype, and global gene expression for cells of the human nucleus pulposus.

Publications

Verteporfin treatment controls morphology, phenotype, and global gene expression for cells of the human nucleus pulposus.

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