Biology subjects

Bertrand, J. A.

Publications and source records attributed to Bertrand, J. A..

6 recordsLinked to original sources

bioRxiv2025

Corticosteroids elevate intraocular pressure through suppression of TREK-1 signaling

Clinicians are often forced into the dilemma of whether to battle ocular inflammation or preserve vision imperiled by elevated intraocular pressure (IOP). Anti-inflammatory treatments utilizing glucocorticosteroid regimens may induce glaucoma by chronically elevating IOP via increased trabecular meshwork (TM) resistance to the flow of aqueous humor, but it is not known whether pressure transduction itself is impacted by steroids and how changes in TM mechanosignaling affect conventional outflow resistance and IOP. To address this, we investigated the role of TREK-1 (TWIK-related potassium channel-1), a mechanosensitive K+ channel, in regulation of outflow facility, transmembrane signaling and dexamethasone (DEX)-induced ocular hypertension (OHT). The expression of tandem-pore potassium channels in mouse TM cells was dominated by Trek-1 (Kcnk2) mRNA, with residual expression of Traak, Tresk2 and Twik3 and vanishingly low levels of Task1 and Trek2. DEX suppressed Trek1 transcription by [~]80% but did not affect expression of Trpv4 and Piezo1 genes. Chronic DEX administration depolarized the membrane potential of TM cells and elevated IOP in mice whereas the selective TREK-1 agonist ML-402 lowered IOP in rodent OHT models. ML-402 doubled the outflow facility in perfused mouse eyes at all applied pressures and hyperpolarized DEX-treated TM cells. These in vitro, ex vivo and in vivo results implicate TREK-1 channels in homeostatic regulation of TM mechanosignaling, conventional outflow regulation and IOP homeostasis. Suppression of TREK-1 signaling by corticosteroids underlies OHT and could contribute to steroid glaucoma but this can be obviated by pharmacological stimulation of the channel with cornea-permeant ML-402 eye drops.

physiology↗

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↗

bioRxiv2023

Gene flow throughout the evolutionary history of a polymorphic and generalist clownfish

Even seemingly homogeneous on the surface, the oceans display high environmental heterogeneity across space and time. Indeed, different soft barriers structure the marine environment, which offers an appealing opportunity to study various evolutionary processes such as population differentiation and speciation. Here, we focus on Amphiprion clarkii (Actinopterygii; Perciformes), the most widespread of clownfishes that exhibits the highest colour polymorphism. Clownfishes can only disperse during a short pelagic larval phase before their sessile adult lifestyle, which might limit connectivity among populations, thus facilitating speciation events. Consequently, the taxonomic status of A. clarkii has been under debate. We used whole-genome resequencing data of 67 A. clarkii specimens spread across the Indian and Pacific Oceans to characterise the species population structure, demographic history, and colour polymorphism. We found that A. clarkii spread from the Indo-Pacific Ocean to the Pacific and Indian Oceans following a stepping-stone dispersal and that gene flow was pervasive throughout its demographic history. Moreover, edge populations exhibited more similar colouration patterns compared to central populations. However, we demonstrate that colour polymorphism is not associated with population structure, thus, colour phenotype is unreliable in assessing the taxonomic status of A. clarkii. Our study further highlights the power of whole-genome comparative studies to determine the taxonomy of geographically wide-ranging and phenotypically diverse species, supporting the status of A. clarkii as a single species.

evolutionary biology↗

bioRxiv2021

TRPM4 is required for calcium oscillations downstream from the stretch-activated TRPV4 channel

Transduction of mechanical information is influenced by physical, chemical and thermal cues but the molecular mechanisms through which transducer activation shapes temporal signaling remain underexplored. In the present study, electrophysiology, histochemistry and functional imaging were combined with gene silencing and heterologous expression to gain insight into calcium signaling downstream from TRPV4 (Transient Receptor Potential Vanilloid 4), a stretch-activated nonselective cation channel. We show that trabecular meshwork (TM) cells, which employ mechanotransduction to actively regulate intraocular pressure, respond to the TRPV4 agonist GSK1016790A with fluctuations in intracellular Ca2+ concentration ([Ca2+]i) and an increase in [Na+]i. ([Ca2+]i oscillations coincided with a monovalent cation current that was suppressed by BAPTA, Ruthenium Red and 9-phenanthrol, an inhibitor of TRPM4 (Transient Receptor Potential Melastatin 4) channels. Accordingly, TM cells expressed TRPM4 mRNA, protein at the expected 130-150 kDa and showed punctate TRPM4 immunoreactivity at the membrane surface. Genetic silencing of TRPM4 antagonized TRPV4-evoked oscillatory signaling whereas TRPV4 and TRPM4 co-expression in HEK-293 cells reconstituted the oscillations. Membrane potential recordings indicated that TRPM4-dependent oscillations required release of Ca2+ from internal stores. 9-phenanthrol did not affect the outflow facility in mouse eyes. Collectively, our results show that TRPV4 activity initiates dynamic calcium signaling in TM cells by stimulating TRPM4 channels and intracellular Ca2+ release. These findings provide insight into the complexity of membrane-cytosolic interactions during TRPV4 signaling and may foster strategies to promote homeostatic regulation and counter pathological remodeling within the conventional outflow pathway of the mammalian eye.

cell biology↗

bioRxiv2020

Gene Deletion Studies Reveal A Critical Role For Prostamide/Prostaglandin F2α Synthase In Prostamide F2α Formation

Prostamide/prostaglandin F synthase (PM/PGFS) is an enzyme with very narrow substrate specificity and is dedicated to the biosynthesis of prostamide and prostaglandin F2. The importance of this enzyme, relative to the aldoketoreductase (AKR) series, in providing functional tissue prostamide F2 levels was determined by creating a line of PM/PGFS gene deleted mice. Deletion of the gene encoding PM/PGFS (fam213b) was accomplished by a two exon disruption. Prostamide F2 levels in WT and PM/PGFS KO mice were determined by LC/MS/MS. Intraocular pressure was measured by tonometry and outflow facility was measured by the iPerfusion method in enucleated mouse eyes. Deletion of fam213b had no observed effect on behavior, appetite, or fertility. Intraocular pressure was significantly elevated by approximately 4 mmHg in PM/PGFS KO mice compared to littermate WT mice. No effect on pressure dependent outflow facility occurred, which is consistent with the typically reported prostanoid effect of increasing outflow through the unconventional pathway. The elevation of intraocular pressure caused by deletion of the gene encoding the PM/PGFS enzyme likely results from a diversion of the endoperoxide precursor pathway to provide increased levels of those prostanoids that raise intraocular pressure, namely prostaglandin D2 (PGD2), thromboxane A2 (TxA2). It follows that PM/PGFS may serve an important regulatory role in the eye by providing PGF2 and prostamide F2 to constrain the influence of those prostanoids that raise intraocular pressure.

pharmacology and toxicology↗