U9 - Brillouin microscopy and optical coherence elastography reveal the regulation of corneal stiffness by collagen XIV

Friday, October 10, 2025

10:00 AM - 11:00 AM PDT

Location: Exhibit Hall F, G & H

Presenting Author(s)

SS

Sajedeh Saeidi Fard

Research Assistant
University of Houston
Houston, Texas, United States

Co-Author(s)

AS

Amandeep Singh

Postdoctoral fellow
University of Houston
Houston, Texas, United States
MS

Manmohan Singh

Research assistant professor
University of Houston
Houston, Texas, United States
AW

Alexander W. Schill

Senior Research Scientist
University of Houston
Houston, Texas, United States
SR

Salavat R. Aglyamov

Research Associate Professor
University of Houston
Houston, Texas, United States
MS

Mei Sun

Research associate
University of South Florida, United States
EM

Edgar M. Espana

Professor
University of South Florida, United States
GS

Giuliano Scarcelli

Professor
University of Maryland, United States
KL

Kirill Larin

Professor
University of Houston
Houston, Texas, United States

Last Author(s)

KL

Kirill Larin

Professor
University of Houston
Houston, Texas, United States

Primary Investigator(s)

KL

Kirill Larin

Professor
University of Houston
Houston, Texas, United States

Introduction: : Evaluating the mechanical characteristics of the cornea is important for understanding its structural role and contribution to overall eye health. Collagen XIV is a key component of the cornea, but its function remains poorly understood. According to the literature, this protein plays a vital role in regulating corneal structure and function, and it is believed to be essential for sustaining mechanical strength. Different methods have been proposed to evaluate the mechanical characteristics of the cornea. Among these techniques, Brillouin microscopy and optical coherence elastography (OCE) have shown particular promise for measuring corneal stiffness. This research employed a dual-prong optical elastography approach that integrated Brillouin microscopy and OCE to examine the mechanical impact of collagen XIV deficiency in the mouse cornea. These two methods showed the ability to distinguish the corneal stiffness of wild type (WT) and collagen XIV-deficient (Col14a1–/–) mouse corneas and offer additional insights into the mechanical characteristics of the tissue.

Materials and

Methods: : Brillouin microscopy was conducted on WT (n=4) and Col14a1–/– (n=4) corneas utilizing a custom-built system where a 660 nm single-mode laser (Torus; Laser Quantum Inc., Fremont, CA, USA) illuminated the specimen. A microscope featuring a 0.42 numerical aperture concentrated the light onto the specimen. The backscattered light was sent to the dual-stage virtually imaged phase array spectrometer featuring a 30 GHz free spectral range and was recorded with an electron-multiplying charge-coupled device camera (iXon Andor, Belfast, UK). Before each measurement, spectral calibration was performed using standard substances: water, acetone, and ethanol. In each measurement, the sample was scanned along the axial and lateral directions employing a motorized stage. After data acquisition, it was processed in MATLAB R2024a (MathWorks, Natick, MA, USA) to measure the Brillouin frequency shift for each measurement location.
OCE assessments were conducted in WT (n=8) and Col14a1–/– (n=7) corneal samples. The OCE system contains an optical coherence tomography setup utilizing a superluminescent diode (S840-B-I-20; Superlum Diodes Ltd., Carrigtwohill, Ireland) which functions at a central wavelength of approximately 840 nm and a bandwidth of around 49 nm. The A-line rate was fixed at 200 kHz, and a quasi-harmonic square wave signal of 20 kHz was generated by a function generator (DG4162, RIGOL Tech, Beijing, China) for excitation. The produced signal was amplified by a radio-frequency power amplifier (1040L, Electronics & Innovation, Ltd., Rochester, NY, United States) prior to powering a 1 MHz air-coupled ultrasound transducer (ACUS). Figure 1 illustrates the schematics for the OCE and Brillouin systems.

Results, Conclusions, and Discussions::

Results:
Figure 2 shows 2D Brillouin microscopy image of a representative WT, Col14a1–/– cornea, and Col14a1–/– cornea B-mode OCT intensity map and wave propagation snapshots at t= 2.40 s and t = 2.44 s. Figure 3 plots the quantification of Brillouin frequency shift and the velocity of the elastic waves obtained through OCE for each sample. Results from both OCE and Brillouin indicate that a lack of collagen XIV decreases the cornea's overall stiffness. The OCE data indicates that the mean elastic wave velocity in the WT cornea and Col14a1–/– knockout corneas was 5.06±0.51 m/s and 3.51±0.26 m/s, respectively. There were notable disparities (p < 0.001 via Mann-Whitney U-test) in the wave speeds of the WT and Col14a1–/– corneas. Additionally, the Brillouin frequency shift was notably lower in the Col14a1–/– than in the WT (p < 0.001 via Mann-Whitney U-test). To conclude, our two-prong approach suggests that collagen XIV significantly provides normal stiffness to the cornea. The complementary mechanical characterization via OCE is directly associated with the shear elastic modulus (or Young’s modulus) of the cornea, while the Brillouin frequency shift is closely connected to the longitudinal modulus (i.e., compressibility). The essential connection between the Brillouin-measured longitudinal modulus and Young’s modulus remains an unresolved issue, though an empirical relationship may exist between them in specific materials. Upcoming research will concentrate on multimodal biomechanical models and assess the impact of various collagen types on corneal stiffness.

Conclusion:
Collagen XIV is essential for maintaining corneal structure and function and is thought to contribute to its mechanical strength. Nonetheless, the present understanding of its biomechanical function is constrained. Brillouin microscopy and optical coherence elastography have demonstrated considerable potential among the methods for evaluating corneal stiffness. In this research, we applied both techniques to assess the effect of collagen XIV lack on the stiffness of murine corneas. Our results show that the lack of collagen XIV results in decreased overall corneal stiffness. These findings emphasize the ability of these methods to identify mechanical irregularities, potentially aiding in the clinical identification of corneal disorders.

Acknowledgements and/or References (Optional): :