Biology subjects

Narendran, S.

Publications and source records attributed to Narendran, S..

3 recordsLinked to original sources

bioRxiv2021

The learning curve of murine subretinal injection among clinically trained ophthalmic surgeons

PO_SCPLOWURPOSEC_SCPLOWSubretinal injection (SRI) in mice is widely used in retinal research, yet the learning curve (LC) of this surgically challenging technique is unknown. MO_SCPLOWETHODSC_SCPLOWTo evaluate the LC for SRI in a murine model, we analyzed training data from 3 clinically trained ophthalmic surgeons from 2018 to 2020. Successful SRI was defined as either the absence of retinal pigment epithelium (RPE) degeneration after phosphate buffer saline injection and the presence of RPE degeneration after Alu RNA injection. Multivariable survival-time regression models were used to evaluate the association between surgeon experience and success rate, with adjustment for injection agents, and to calculate an approximate case number to achieve a 95% success rate. A Cumulative Sum (CUSUM) analysis was performed and plotted individually to monitor each surgeons simultaneous performance. RO_SCPLOWESULTSC_SCPLOWDespite prior microsurgery experience, the combined average success rate of the first 50 cases in mice was only 27%. The predicted SRI success rate did not reach a plateau above 95% until approximately 364 prior cases. Using the 364-training case as a "cutoff" point, the predicted probability of success before and after the 364th case was 65.38% and 99.32%, respectively (P < 0.0001). CUSUM analysis showed an initial upward slope and then remained within the decision intervals with an acceptable success rate set at 95% in the late stage. CO_SCPLOWONCLUSIONSC_SCPLOWThis study demonstrates the complexity and substantial LC for successful SRI in mice with high confidence. A systematic training system could improve the reliability and reproducibility of SRI-related experiments and improve the interpretation of experimental results using this technique. Translational RelevanceOur prediction model and monitor system allow objective quantification of technical proficiency in the field of subretinal drug delivery and gene therapy for the first time.

neuroscience↗

bioRxiv2021

Identification of Fluoxetine as a direct NLRP3 inhibitor to treat atrophic macular degeneration

The atrophic form of age-related macular degeneration (dry AMD) affects nearly 200 million people worldwide. There is no FDA-approved therapy for this disease, which is the leading cause of irreversible blindness among people over 50 years of age. Vision loss in dry AMD results from degeneration of the retinal pigmented epithelium (RPE). RPE cell death is driven in part by accumulation of Alu RNAs, which are noncoding transcripts of a human retrotransposon. Alu RNA induces RPE degeneration by activating the NLRP3-ASC inflammasome. We report that fluoxetine, an FDA-approved drug for treating clinical depression, binds NLRP3 in silico, in vitro, and in vivo, and that it inhibits activation of the NLRP3-ASC inflammasome in RPE cells and macrophages, two critical cell types in dry AMD. We also demonstrate that fluoxetine, unlike several other anti-depressant drugs, reduces Alu RNA-induced RPE degeneration in mice. Finally, by analyzing two health insurance databases comprising more than 100 million Americans, we report a reduced hazard of developing dry AMD among patients with depression who were treated with fluoxetine. Collectively, these studies triangulate to link fluoxetine as a potential drug repurposing candidate for a major unmet medical need that causes blindness in millions of people in the United States and across the world. Significance StatementDry age-related macular degeneration (AMD) affects the vision of millions of people worldwide. There is currently no FDA-approved treatment for dry AMD. The inflammasome components NLRP3 and ASC have been implicated in the pathogenesis of dry AMD. We report that fluoxetine, which is approved for the treatment of clinical depression, directly binds the NLRP3 protein and prevents the assembly and activation of the NLRP3-ASC inflammasome. As a result, it also blocks the degeneration of retinal pigmented epithelium (RPE) cells in an animal model of dry AMD. Furthermore, we demonstrate through an analysis of health insurance databases that use of this FDA-approved anti-depressant drug is associated with reduced incidence of dry AMD. These studies identify that fluoxetine is a potential repurposing candidate for AMD, a prevalent cause of blindness.

immunology↗

bioRxiv2020

Start codon disruption with CRISPR/Cas9 prevents murine Fuchs' endothelial corneal dystrophy

A missense mutation of collagen type VIII alpha 2 chain (COL8A2) gene leads to early onset Fuchs endothelial corneal dystrophy (FECD), which progressively impairs vision through loss of corneal endothelial cells. We demonstrate that CRISPR/Cas9-based postnatal gene editing achieves structural and functional rescue in a mouse model of FECD. A single intraocular injection of an adenovirus encoding both the Cas9 gene and guide RNA (Ad-Cas9-Col8a2gRNA), efficiently knocked down mutant COL8A2 expression in corneal endothelial cells, prevented endothelial cell loss, and rescued corneal endothelium pumping function in adult Col8a2 mutant mice. There were no adverse sequelae on histology or electroretinography. Col8a2 start codon disruption represents a non-surgical strategy to prevent vision loss in early-onset FECD. As this demonstrates the ability of Ad-Cas9-gRNA to restore phenotype in adult post-mitotic cells, this method may be widely applicable to adult-onset diseases, even in tissues affected with disorders of non-reproducing cells.

genomics↗