Biology subjects

Vahabikashi, A.

Publications and source records attributed to Vahabikashi, A..

5 recordsLinked to original sources

bioRxiv2021

Surface Engineering of FLT4-Targeted Nanocarriers Enhances Cell-Softening Glaucoma Therapy

Primary open-angle glaucoma is associated with elevated intraocular pressure (IOP) that damages the optic nerve and leads to gradual vision loss. Several agents that reduce the stiffness of pressure-regulating Schlemms canal endothelial cells, in the conventional outflow pathway of the eye, lower IOP in glaucoma patients and are approved for clinical use. However, poor drug penetration and uncontrolled biodistribution limit their efficacy and produce local adverse effects. Compared to other ocular endothelia, FLT4/VEGFR3 is expressed at elevated levels by Schlemms canal endothelial cells and can be exploited for targeted drug delivery. Here, we validate FLT4 receptors as a clinically relevant target on Schlemms canal cells from glaucomatous human donors and engineer polymeric self-assembled nanocarriers displaying lipid-anchored targeting ligands that optimally engage this receptor. Targeting constructs were synthesized as lipid-PEGX-peptide, differing in the number of PEG spacer units (x), and were embedded in micelles. We present a novel proteolysis assay for quantifying ligand accessibility that we employ to design and optimize our FLT4-targeting strategy for glaucoma nanotherapy. Peptide accessibility to proteases correlated with receptor-mediated targeting enhancements. Increasing the accessibility of FLT4-binding peptides enhanced nanocarrier uptake by Schlemms canal cells while simultaneously decreasing uptake by off-target vascular endothelial cells. Using a paired longitudinal IOP study in vivo, we show this enhanced targeting of Schlemms canal cells translates to IOP reductions that are sustained for a significantly longer time as compared to controls. Histological analysis of murine anterior segment tissue confirmed nanocarrier localization to Schlemms canal within one hour after intracameral administration. This work demonstrates that steric effects between surface-displayed ligands and PEG coronas significantly impact targeting performance of synthetic nanocarriers across multiple biological scales. Minimizing the obstruction of modular targeting ligands by PEG measurably improved the efficacy of glaucoma nanotherapy and is an important consideration for engineering PEGylated nanocarriers for targeted drug delivery.

bioengineering↗

bioRxiv2021

Nuclear lamin isoforms differentially contribute to LINC complex-dependent nucleocytoskeletal coupling and whole cell mechanics

The ability of a cell to regulate its mechanical properties is central to its function. Emerging evidence suggests that interactions between the cell nucleus and cytoskeleton influence cell mechanics through poorly understood mechanisms. Here we show that A- and B-type nuclear lamin isoforms distinctively modulate both nuclear and cellular volume and selectively stabilize the linker of nucleoskeleton and cytoskeleton (LINC) complexes that couple the nucleus to cytoskeletal actin and vimentin. We reveal, further, that loss of each of the four-known lamin isoforms in the mouse embryonic fibroblasts differentially affects cortical and cytoplasmic stiffness as well as cellular contractility, and then propose a LINC complex mediated model that explains these impaired mechanical phenotypes. Finally, we demonstrate that loss of each lamin isoform softens the nucleus in a manner that correlates with loss of heterochromatin. Together, these findings uncover distinctive roles for each lamin isoform in maintaining cellular and nuclear mechanics.

biophysics↗

bioRxiv2019

Adaptive-Resolution Multi-Orientation Analysis of Complex Filamentous Network Images

Microscopy images of cytoskeletal, nucleoskeletal, and other filamentous structures contain complex junctions where multiple filaments with distinct orientations overlap, yet state-of-the-art software generally uses single orientation analysis to segment these structures. We describe an image analysis approach to simultaneously segment both filamentous structures and their intersections in microscopy images, based on analytically resolving coincident multiple orientations upon image filtering in a manner that balances orientation resolution and spatial localization.

bioinformatics↗

bioRxiv2019

Vimentin protects the structural integrity of the nucleus and suppresses nuclear damage caused by large deformations

Mammalian cells frequently migrate through tight spaces during normal embryogenesis, wound healing, diapedesis or in pathological situations such as metastasis. The nucleus has recently emerged as an important factor in regulating 3D cell migration. At the onset of migratory behavior, cells often initiate the expression of vimentin, an intermediate filament protein which forms networks extending from a juxtanuclear cage to the cell periphery. However, the role of vimentin intermediate filaments (VIFs) in regulating nuclear shape and mechanics remains unknown. Here, we used wild type and vimentin-null mouse embryonic fibroblasts to show that VIFs regulate nuclear shape, motility, and the ability of cells to resist large deformations. The results show that loss of VIFs alters nuclear shape, reduces perinuclear stiffness, and enhances motility in 3D. These changes increase nuclear rupture and activation of DNA damage repair mechanisms, which are rescued by exogenous re-expression of vimentin. Our findings show that VIFs provide mechanical support to protect the nucleus and genome during migration.

cell biology↗

bioRxiv2018

Probe Sensitivity to Cortical versus Intracellular Cytoskeletal Network Stiffness

In development, wound healing, and pathology, cell biomechanical properties are increasingly recognized as being of central importance. To measure these properties, experimental probes of various types have been developed, but how each probe reflects the properties of heterogeneous cell regions has remained obscure. To better understand differences attributable to the probe technology, as well as to define the relative sensitivity of each probe to different cellular structures, here we took a comprehensive approach. We studied two cell types --Schlemms canal (SC) endothelial cells and mouse embryonic fibroblasts (MEFs) - using four different probe technologies: 1) atomic force microscopy (AFM) with sharp-tip; 2) AFM with round-tip; 3) optical magnetic twisting cytometry (OMTC); and 4) traction microscopy (TM). Perturbation of SC cells with dexamethasone treatment, a-actinin overexpression, or Rho-A overexpression caused increases in traction reported by TM and stiffness reported by sharp-tip AFM, as compared to corresponding controls. By contrast, under these same experimental conditions, stiffness reported by round-tip AFM and by OMTC indicated little change. Knock out (KO) of vimentin in MEFs caused a diminution of traction reported by TM, as well as stiffness reported by sharp-tip and round-tip AFM. However, stiffness reported by OMTC in vimentin KO MEFs was greater than in wild-type. Finite element analysis demonstrated that this paradoxical OMTC result in vimentin KO MEFs could be attributed to reduced cell thickness. Our results also suggest that vimentin contributes not only to intracellular network stiffness but also cortex stiffness. Taken together, this evidence suggests that AFM sharp-tip and TM emphasize properties of the actin-rich shell of the cell whereas round-tip AFM and OMTC emphasize those of the non-cortical intracellular network.

biophysics↗