Biology subjects

Overby, D. R.

Publications and source records attributed to Overby, D. R..

4 recordsLinked to original sources

bioRxiv2026

VEGFA-Positive Macrophages Regulate Aqueous Humor Outflow in Aged Mice and Humans

Elevated intraocular pressure (IOP) and aging are major risk factors for primary open-angle glaucoma (POAG), but how aging affects IOP regulation remains poorly understood. IOP remains within a narrow range despite age-associated changes predicted to increase aqueous humor outflow (AHO) resistance at the interface between the trabecular meshwork and Schlemm's canal (SC), suggesting compensatory mechanisms preserve AHO homeostasis during aging. Single-cell RNA sequencing of mouse ocular angle tissues revealed immunomodulatory transcriptional reprogramming of SC endothelial cells in older mice, while mouse and human imaging showed reduced SC size and increased peri-SC macrophage accumulation with aging. Ligand-receptor analysis predicted enhanced macrophage-to-SC VEGFA-VEGFR signaling in aged and Tie2-haploinsufficient mice, an independent model of vascular stress and glaucoma risk. Deletion of Vegfa in CX3CR1+ macrophages increased IOP and reduced AHO facility in 9-month-old mice, demonstrating that macrophage-derived VEGFA supports AHO homeostasis. Tie2 haploinsufficiency recapitulated key age-associated SC niche changes, including peri-SC macrophage accumulation, whereas gene therapy boosting TIE2 activity protected wild-type mice against age-related changes. Together, these findings identify peri-SC macrophage-derived VEGFA as a compensatory mechanism maintaining AHO homeostasis during aging and vascular stress and support TIE2 activation as a therapeutic strategy to preserve SC function and IOP regulation.

physiology↗

bioRxiv2025

Interdependent regulation of trabecular meshwork cell physiology and intraocular pressure by KALRN and TMCO1

Glaucoma is a leading cause of irreversible blindness. Primary open-angle glaucoma (POAG) is its most common form. Higher intraocular pressure (IOP), resulting from impaired aqueous humour outflow, is the cardinal mediating factor, and all proven treatments aim to lower IOP. POAG is a complex genetic disease with numerous loci linked to POAG and higher IOP. The mechanisms by which risk alleles cause disease remain unclear. Here, using primary trabecular meshwork (TM) cells, a cell type controlling outflow, and mice, we found that the POAG-associated gene KALRN encodes an endoplasmic reticulum (ER)-associated Rac regulator essential for TM homeostasis and normal IOP. KALRN loss caused widespread disruption from ER to calcium homeostasis and energy metabolism, leading to induction of cell senescence. KALRN-depletion also led to suppression of the ER translocase and calcium regulator TMCO1, encoded by a gene at one of the most significantly associated genomic loci for POAG. TMCO1-depletion in vitro and in vivo phenocopied KALRN-induced phenotypes, and reduced KALRN expression. These findings establish KALRN and TMCO1 as interdependent regulators of TM homeostasis and IOP, that link regulation of ER and intracellular calcium homeostasis to cell and tissue physiology, and illustrate how different genes linked to glaucoma can form regulatory pathways.

cell biology↗

bioRxiv2025

TRPV4 modulates substrate stiffness mechanosensing and transcellular pore formation in human Schlemm's canal cells

Pathological changes in the biomechanical environment of Schlemms canal (SC) inner wall cells, such as substrate stiffening and increased cellular stretch, are associated with ocular hypertension, a key risk factor for the development of glaucoma. Cell membrane stretch can trigger the activation of transient receptor potential vanilloid 4 (TRPV4) mechanosensitive ion channels, allowing calcium influx and initiating downstream signaling. However, the precise role of TRPV4 in SC cell mechanobiology remains unclear. Here, we demonstrate that sustained inhibition of TRPV4 activity modulates substrate stiffness mechanosensing to thereby affect the remodeling of the actin cytoskeleton and extracellular matrix of SC cells. This is accompanied by a reduction in cell stiffness and an increase in transcellular pore forming ability, potentially lowing outflow resistance and risk of ocular hypertension. Conversely, acute activation of TRPV4 channels induces Ca2+ influx, increasing transcellular pore formation in SC cells. Notaly, reduced TRPV4 mechanosensing was observed in glaucomatous SC cells, resulting in reduced transcellular pore forming ability. These findings suggest novel potential strategies based on targeting TRPV4 in SC cells for the treatment of ocular hypertension in glaucoma.

cell biology↗

bioRxiv2024

Endothelial cell stiffness and type drive the formation of biomechanically-induced transcellular pores

Formation of transcellular pores facilitates the transport of materials across endothelial barriers. In Schlemms canal (SC) endothelium, impaired pore formation is associated with glaucoma. However, our understanding of the cellular processes responsible for pore formation is limited by lack of in vitro assays. Here, we present a novel platform for studying transcellular pore formation in human endothelial cells. We induced pores in SC cells by seeding them atop micron-sized magnetic beads followed by application of a magnetic field to subject cells to a basal to apical force, mimicking in vivo biomechanical forces. The pore formation process was dynamic, with pores opening and closing. Glaucomatous cells exhibited impaired pore formation that correlated with their increased stiffness. We further discovered that application of forces from the apical to basal direction did not induce pores in SC cells but resulted in formation of pores in other types of endothelial cells. Our studies reveal the central role of cell mechanics in formation of transcellular pores in endothelial cells, and provide a new approach for investigating their associated underlying mechanism/s.

bioengineering↗