Biology subjects

Lee, P.-Y.

Publications and source records attributed to Lee, P.-Y..

7 recordsLinked to original sources

bioRxiv2024

Impact of anatomic variability and other vascular factors on lamina cribrosa hypoxia

Insufficient oxygenation in the lamina cribrosa (LC) may contribute to axonal damage and glaucomatous vision loss. To understand the range of susceptibilities to glaucoma, we aimed to identify key factors influencing LC oxygenation and examine if these factors vary with anatomical differences between eyes. We reconstructed 3D, eye-specific LC vessel networks from histological sections of four healthy monkey eyes. For each network, we generated 125 models varying vessel radius, oxygen consumption rate, and arteriole perfusion pressure. Using hemodynamic and oxygen supply modeling, we predicted blood flow distribution and tissue oxygenation in the LC. ANOVA assessed the significance of each parameter. Our results showed that vessel radius had the greatest influence on LC oxygenation, followed by anatomical variations. Arteriole perfusion pressure and oxygen consumption rate were the third and fourth most influential factors, respectively. The LC regions are well perfused under baseline conditions. These findings highlight the importance of vessel radius and anatomical variation in LC oxygenation, providing insights into LC physiology and pathology. Pathologies affecting vessel radius may increase the risk of LC hypoxia, and anatomical variations could influence susceptibility. Conversely, increased oxygen consumption rates had minimal effects, suggesting that higher metabolic demands, such as those needed to maintain intracellular transport despite elevated intraocular pressure, have limited impact on LC oxygenation.

bioengineering↗

bioRxiv2024

Impact of elevated IOP on lamina cribrosa oxygenation; A combined experimental-computational study on monkeys

PurposeOur goal is to evaluate how lamina cribrosa (LC) oxygenation is affected by the tissue distortions resulting from elevated IOP. DesignExperimental study on monkeys SubjectsFour healthy monkey eyes with OCT scans with IOP of 10 to 50 mmHg, and then with histological sections of LC. MethodsSince in-vivo LC oxygenation measurement is not yet possible, we used 3D eye-specific numerical models of the LC vasculature which we subjected to experimentally-derived tissue deformations. We reconstructed 3D models of the LC vessel networks of 4 healthy monkey eyes from histological sections. We also obtained in-vivo IOP-induced tissue deformations from a healthy monkey using OCT images and digital volume correlation analysis techniques. The extent that LC vessels distort under a given OCT-derived tissue strain remains unknown. We therefore evaluated two biomechanics-based mapping techniques: cross-sectional and isotropic. The hemodynamics and oxygenations of the four vessel networks were simulated for deformations at several IOPs up to 60mmHg. The results were used to determine the effects of IOP on LC oxygen supply, assorting the extent of tissue mild and severe hypoxia. Main Outcome MeasuresIOP-induced deformation, vasculature structure, blood supply, and oxygen supply for LC region ResultIOP-induced deformations reduced LC oxygenation significantly. More than 20% of LC tissue suffered from mild hypoxia when IOP reached 30 mmHg. Extreme IOP(>50mmHg) led to large severe hypoxia regions (>30%) in the isotropic mapping cases. ConclusionOur models predicted that moderately elevated IOP can lead to mild hypoxia in a substantial part of the LC, which, if sustained chronically, may contribute to neural tissue damage. For extreme IOP elevations, severe hypoxia was predicted, which would potentially cause more immediate damage. Our findings suggest that despite the remarkable LC vascular robustness, IOP-induced distortions can potentially contribute to glaucomatous neuropathy.

bioengineering↗

bioRxiv2023

Direct measurements of collagen fiber recruitment in the posterior pole of the eye

Collagen is the main load-bearing component of the peripapillary sclera (PPS) and lamina cribrosa (LC) in the eye. Whilst it has been shown that uncrimping and recruitment of the PPS and LC collagen fibers underlies the macro-scale nonlinear stiffening of both tissues with increased intraocular pressure (IOP), the uncrimping and recruitment as a function of local stretch have not been directly measured. This knowledge is crucial for the development of constitutive models associating micro and macro scales. In this project we measured local stretch-induced collagen fiber bundle uncrimping and recruitment curves of the PPS and LC. Thin coronal samples of PPS and LC of sheep eyes were mounted and stretched biaxially quasi-statically using a custom system. At each step, we imaged the PPS and LC with instant polarized light microscopy and quantified pixel-level (1.5 m/pixel) collagen fiber orientations. We used digital image correlation to measure the local stretch and quantified collagen crimp by the circular standard deviation of fiber orientations, or waviness. Local stretch-recruitment curves of PPS and LC approximated sigmoid functions. PPS recruited more fibers than the LC at the low levels of stretch. At 10% stretch the curves crossed with 75% bundles recruited. The PPS had higher uncrimping rate and waviness remaining after recruitment than the LC: 0.9{degrees} vs. 0.6{degrees} and 3.1{degrees} vs. 2.7{degrees}. Altogether our findings support describing fiber recruitment of both PPS and LC with sigmoid curves, with the PPS recruiting faster and at lower stretch than the LC, consistent with a stiffer tissue. Statement of SignificancePeripapillary sclera (PPS) and lamina cribrosa (LC) collagen recruitment behaviors are central to the nonlinear mechanical behavior of the posterior pole of the eye. How PPS and LC collagen fibers recruit under stretch is crucial to develop constitutive models of the tissues but remains unclear. We used image-based stretch testing to characterize PPS and LC collagen fiber bundle recruitment under local stretch. We found that fiber-level stretch-recruitment curves of PPS and LC approximated sigmoid functions. PPS recruited more fibers at a low stretch, but at 10% bundle stretch the two curves crossed with 75% bundles recruited. We also found that PPS and LC fibers had different uncrimping rates and non-zero wavinesss when recruited.

bioengineering↗

bioRxiv2023

Instant polarized light microscopy pi (IPOLπ) for quantitative imaging of collagen architecture and dynamics in ocular tissues

Collagen architecture determines the biomechanical environment in the eye, and thus characterizing collagen fiber organization and biomechanics is essential to fully understand eye physiology and pathology. We recently introduced instant polarized light microscopy (IPOL) that encodes optically information about fiber orientation and retardance through a color snapshot. Although IPOL allows imaging collagen at the full acquisition speed of the camera, with excellent spatial and angular resolutions, a limitation is that the orientation-encoding color is cyclic every 90 degrees ({pi}/2 radians). In consequence, two orthogonal fibers have the same color and therefore the same orientation when quantified by color-angle mapping. In this study, we demonstrate IPOL{pi}, a new variation of IPOL, in which the orientation-encoding color is cyclic every 180 degrees ({pi} radians). Herein we present the fundamentals of IPOL{pi}, including a framework based on a Mueller-matrix formalism to characterize how fiber orientation and retardance determine the color. The improved quantitative capability of IPOL{pi} enables further study of essential biomechanical properties of collagen in ocular tissues, such as fiber anisotropy and crimp. We present a series of experimental calibrations and quantitative procedures to visualize and quantify ocular collagen orientation and microstructure in the optic nerve head, a region in the back of the eye. There are four important strengths of IPOL{pi} compared to IPOL. First, IPOL{pi} can distinguish the orientations of orthogonal collagen fibers via colors, whereas IPOL cannot. Second, IPOL{pi} requires a lower exposure time than IPOL, thus allowing faster imaging speed. Third, IPOL{pi} allows visualizing non-birefringent tissues and backgrounds from tissue absorption, whereas both appear dark in IPOL images. Fourth, IPOL{pi} is cheaper and less sensitive to imperfectly collimated light than IPOL. Altogether, the high spatial, angular, and temporal resolutions of IPOL{pi} enable a deeper insight into ocular biomechanics and eye physiology and pathology. HighlightsO_LIWe introduce IPOL{pi}, addressing IPOL limitations for characterizing eye collagen. C_LIO_LIIPOL{pi} orientation-encoded color cycle is 180{degrees} ({pi} radians) instead of 90{degrees} in IPOL. C_LIO_LIIPOL{pi} requires a lower exposure time than IPOL, allowing faster imaging speed. C_LIO_LIIPOL{pi} visualizes non-birefringent tissues and backgrounds from brightness. C_LIO_LIIPOL{pi} is cheaper and less sensitive to imperfectly collimated light than IPOL. C_LI

bioengineering↗

bioRxiv2022

Stretch-Induced Uncrimping of Equatorial Sclera Collagen Bundles

Stretch-induced collagen uncrimping underlies the nonlinear mechanical behavior of the sclera according to what is often called the process of recruitment. We recently reported experimental measurements of sclera collagen crimp and pressure-induced uncrimping. Our studies, however, were cross-sectional, providing statistical descriptions of crimp with no information on the effects of stretch on specific collagen bundles. Data on bundle-specific uncrimping is necessary to better understand the effects of macroscale input on the collagen microscale and tissue failure. Our goal in this project was to measure bundle-specific stretch-induced collagen uncrimping of sclera. Three goat eyes were cryosectioned sagittally (30m). Samples of equatorial sclera were isolated, mounted to a custom uniaxial stretcher and imaged with polarized light microscopy at various levels of clamp-to-clamp stretch until failure. At each stretch level, local strain was measured using image tracking techniques. The level of collagen crimping was determined from the bundle waviness, defined as the circular standard deviation of fiber orientation along a bundle. Eye-specific recruitment curves were then computed using eye-specific waviness at maximum stretch before sample failure to define fibers as recruited. Nonlinear mixed effect models were used to determine the associations of waviness to local strain and recruitment to clamp-to-clamp stretch. Waviness decreased exponentially with local strain (p<0.001), whereas bundle recruitment followed a sigmoidal curve with clamp-to-clamp stretch (p<0.001). Individual bundle responses to stretch varied substantially, but recruitment curves were similar across sections and eyes. In conclusion, uniaxial stretch caused measurable bundle-specific uncrimping, with the sigmoidal recruitment pattern characteristic of fiber-reinforced soft tissues.

bioengineering↗

bioRxiv2021

Real-time imaging of optic nerve head collagen microstructure and biomechanics using instant polarized light microscopy

Current tools lack the temporal or spatial resolution necessary to image many important aspects of the architecture and dynamics of the optic nerve head (ONH). We evaluated the potential of instant polarized light microscopy (IPOL) to overcome these limitations by leveraging the ability to capture collagen fiber orientation and density in a single image. Coronal sections through the ONH of fresh normal sheep eyes were imaged using IPOL while they were stretched using custom uniaxial or biaxial micro-stretch devices. IPOL allows identifying ONH collagen architectural details, such as fiber interweaving and crimp, and has high temporal resolution, limited only by the frame rate of the camera. Local collagen fiber orientations and deformations were quantified using color analysis and image tracking techniques. We quantified stretch-induced collagen uncrimping of lamina cribrosa (LC) and peripapillary sclera (PPS), and changes in LC pore size (area) and shape (convexity and aspect ratio). The simultaneous high spatial and temporal resolutions of IPOL revealed complex ONH biomechanics: i) stretch-induced local deformation of the PPS was nonlinear and nonaffine. ii) under load the crimped collagen fibers in the PPS and LC straightened, without torsion and with only small rotations. iii) stretch-induced LC pore deformation was anisotropic and heterogeneous among pores. Overall, with stretch the pores were became larger, more convex, and more circular. We have demonstrated that IPOL reveals details of collagen morphology and mechanics under dynamic loading previously out of reach. IPOL can detect stretch-induced collagen uncrimping and other elements of the tissue nonlinear mechanical behavior. IPOL showed changes in pore morphology and collagen architecture that will help improve understanding of how LC tissue responds to load. HighlightsO_LIWe demonstrate that instant polarized light microscopy allows visualization and quantification of changes in optic nerve head collagen morphology and architecture under dynamic loading C_LIO_LIWe show crimped collagen fibers in the peripapillary sclera and lamina cribrosa straightening under load, without torsion and with only small rotations. C_LIO_LIWe show that stretch-induced local deformation of the peripapillary sclera was nonlinear and nonaffine. C_LIO_LIWe show that stretch-induced lamina cribrosa pore deformation was anisotropic and heterogeneous among pores. C_LIO_LIOur results show this novel imaging technique could help understand the role of collagen microstructure in eye physiology, aging, and in biomechanics-related diseases, such as glaucoma and myopia. C_LI

bioengineering↗

bioRxiv2018

Programmed Variations of Cytokinesis Contribute to Morphogenesis in the C. elegans embryo

While cytokinesis has been intensely studied, how it is executed during development is not well understood, despite a long-standing appreciation that various aspects of cytokinesis vary across cell and tissue types. To address this, we investigated cytokinesis during the invariant C. elegans embryo lineage and found several reproducibly altered parameters at different stages. During early divisions, furrow ingression asymmetry and midbody inheritance is consistent, suggesting specific regulation of these events. During morphogenesis, we find several unexpected alterations including migration of midbodies to the apical surface during epithelial polarization in different tissues. Aurora B kinase, which is essential for several aspects of cytokinesis, remains localized to the apical membrane after internalization of other midbody components. Inactivation of Aurora B causes cytokinesis failure, which disrupts polarization and tissue formation. Therefore, cytokinesis shows surprising diversity during development and is required during epithelial polarization to establish cellular architecture during morphogenesis.

cell biology↗