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Transcriptome profiling of NIH3T3 cell lines expressing opsin and the P23H opsin mutant identifies candidate drugs for the treatment of retinitis pigmentosa.

ABSTRACT: Mammalian cells are commonly employed in screening assays to identify active compounds that could potentially affect the progression of different human diseases including retinitis pigmentosa (RP), a class of inherited diseases causing retinal degeneration with compromised vision. Using transcriptome analysis, we compared NIH3T3 cells expressing wildtype (WT) rod opsin with a retinal disease-causing single P23H mutation. Surprisingly, heterologous expression of WT opsin in NIH3T3 cells caused more than a 2-fold change in 783 out of 16,888 protein coding transcripts. The perturbed genes encoded extracellular matrix proteins, growth factors, cytoskeleton proteins, glycoproteins and metalloproteases involved in cell adhesion, morphology and migration. A different set of 347 transcripts was either up- or down-regulated when the P23H mutant opsin was expressed suggesting an altered molecular perturbation compared to WT opsin. Transcriptome perturbations elicited by drug candidates aimed at mitigating the effects of the mutant protein revealed that different drugs targeted distinct molecular pathways that resulted in a similar phenotype selected by a cell-based high-throughput screen. Thus, transcriptome profiling can provide essential information about the therapeutic potential of a candidate drug to restore normal gene expression in pathological conditions.

SUBMITTER: Chen Y

PROVIDER: S-EPMC5205556 | biostudies-literature | 2017 Jan

REPOSITORIES: biostudies-literature

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Transcriptome profiling of NIH3T3 cell lines expressing opsin and the P23H opsin mutant identifies candidate drugs for the treatment of retinitis pigmentosa.

Chen Yuanyuan Y Brooks Matthew J MJ Gieser Linn L Swaroop Anand A Palczewski Krzysztof K

Pharmacological research 20161109

Mammalian cells are commonly employed in screening assays to identify active compounds that could potentially affect the progression of different human diseases including retinitis pigmentosa (RP), a class of inherited diseases causing retinal degeneration with compromised vision. Using transcriptome analysis, we compared NIH3T3 cells expressing wildtype (WT) rod opsin with a retinal disease-causing single P23H mutation. Surprisingly, heterologous expression of WT opsin in NIH3T3 cells caused mo ...[more]

PMID: 27838510

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Project description:Antipsychotic drugs haloperidol and clozapine have been reported to increase the sensitivity of retinal ganglion cells (RGCs) to flashes of light in the P23H rat model of retinitis pigmentosa. In order to better understand the effects of these antipsychotic drugs on the visual responses of P23H rat RGCs, I examined the responses of RGCs to a drifting sinusoidal grating of various contrasts. In-vitro multielectrode array recordings were made from P23H rat RGCs and healthy Sprague-Dawley (SD) rat RGCs. Retinas were stimulated with a drifting sinusoidal grating with eight values of contrast (0, 4, 6, 8.5, 13, 26, 51, and 83%). Contrast response functions based on response amplitudes were fitted with a hyperbolic ratio function and contrast thresholds were determined from the fitted curves. SD rat RGCs were divided into two categories, saturating and non-saturating cells, based on whether they showed saturation of responses at high contrast levels. Most SD rat RGCs (58%) were saturating cells. Haloperidol and clozapine decreased the responses of saturating SD rat RGCs to all grating contrasts, except for the highest contrast tested. Clozapine also decreased the responses of non-saturating SD rat RGCs to all grating contrasts, except for the highest contrast tested. Haloperidol did not however significantly affect the responses of non-saturating SD rat RGCs. Haloperidol and clozapine increased the contrast thresholds of both saturating and non-saturating cells in SD rat retinas. Most (73%) P23H rat RGCs could be categorized as either saturating or non-saturating cells. The remaining 'uncategorized' cells were poorly responsive to the drifting grating and were analyzed separately. Haloperidol and clozapine increased the responses of non-saturating and uncategorized P23H rat RGCs to most grating contrasts, including the highest contrast tested. Haloperidol and clozapine also increased the responses of saturating P23H rat RGCs to most grating contrasts but these increases were not statistically significant. Haloperidol and clozapine decreased the contrast thresholds of saturating cells, non-saturating cells and uncategorized cells in P23H rat retinas, although the decrease in contrast thresholds of saturating cells was not found to be statistically significant. Overall, the findings show that haloperidol and clozapine have differential effects on the contrast response functions of SD and P23H rat RGCs. In contrast to the effects observed on SD rat RGCs, both haloperidol and clozapine increased the responsiveness of P23H rat RGCs to both low and high contrast visual stimuli and decreased contrast thresholds.

Dataset Information

Transcriptome profiling of NIH3T3 cell lines expressing opsin and the P23H opsin mutant identifies candidate drugs for the treatment of retinitis pigmentosa.

Publications

Transcriptome profiling of NIH3T3 cell lines expressing opsin and the P23H opsin mutant identifies candidate drugs for the treatment of retinitis pigmentosa.

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