Dolutegravir induces FOLR1 expression during brain organoid development.

During the first month of pregnancy, the brain and spinal cord are formed through a process called neurulation. However, this process can be altered by low serum levels of folic acid, environmental factors, or genetic predispositions. In 2018, a surveillance study in Botswana, a country with a high incidence of human immunodeficiency virus (HIV) and lacking mandatory food folate fortification programs, found that newborns whose mothers were taking dolutegravir (DTG) during the first trimester of pregnancy had an increased risk of neural tube defects (NTDs). As a result, the World Health Organization and the U.S. Food and Drug Administration have issued guidelines emphasizing the potential risks associated with the use of DTG-based antiretroviral therapies during pregnancy. To elucidate the potential mechanisms underlying the DTG-induced NTDs, we sought to assess the potential neurotoxicity of DTG in stem cell-derived brain organoids. The gene expression of brain organoids developed in the presence of DTG was analyzed by RNA sequencing, Optical Coherence Tomography (OCT), Optical Coherence Elastography (OCE), and Brillouin microscopy. The sequencing data shows that DTG induces the expression of the folate receptor (FOLR1) and modifies the expression of genes required for neurogenesis. The Brillouin frequency shift observed at the surface of DTG-exposed brain organoids indicates an increase in superficial tissue stiffness. In contrast, reverberant OCE measurements indicate decreased organoid volumes and internal stiffness.

Acute alcohol consumption modulates corneal biomechanical properties as revealed by optical coherence elastography.

Acute alcohol ingestion has been found to impact visual functions, including eye movement, but its effects on corneal biomechanical properties remain unclear. This study aimed to investigate the influence of acute alcohol consumption on corneal biomechanical properties using optical coherence elastography (OCE). An air-coupled ultrasound transducer induced elastic waves in mice corneas in vivo, and a high-resolution phase-sensitive optical coherence tomography (OCT) system tracked the mechanical waves to quantify the elastic wave speed. In vivo measurements were performed on three groups of age- and gender-matched mice: control, placebo (administered saline), and alcohol (administered ethanol) groups. Longitudinal measurements were conducted over a one-hour period to assess acute temporal changes in wave speeds, which are associated with inherent biomechanical properties of the cornea. The results showed a significant decrease in wave speed for the alcohol group after 10 min of ingestion in comparison to pre-ingestion values (p = 0.0096), whereas the temporal wave speed changes for the placebo group were statistically insignificant (p = 0.076). In contrast, the control group showed no significant changes in elastic wave speed and corneal thickness. Furthermore, a significant difference was observed between the wave speeds of the placebo and alcohol groups at each measurement time point between 10 and 50 min (p < 0.05), though both groups exhibited a similar trend in corneal thickness change. The findings of this study have important implications for clinical assessments and research in corneal disorders, highlighting the potential of OCE as a valuable tool for evaluating such changes.

Optical coherence tomography-guided Brillouin microscopy highlights regional tissue stiffness differences during anterior neural tube closure in the Mthfd1l murine mutant.

Neurulation is a highly synchronized biomechanical process leading to the formation of the brain and spinal cord, and its failure leads to neural tube defects (NTDs). Although we are rapidly learning the genetic mechanisms underlying NTDs, the biomechanical aspects are largely unknown. To understand the correlation between NTDs and tissue stiffness during neural tube closure (NTC), we imaged an NTD murine model using optical coherence tomography (OCT), Brillouin microscopy and confocal fluorescence microscopy. Here, we associate structural information from OCT with local stiffness from the Brillouin signal of embryos undergoing neurulation. The stiffness of neuroepithelial tissues in Mthfd1l null embryos was significantly lower than that of wild-type embryos. Additionally, exogenous formate supplementation improved tissue stiffness and gross embryonic morphology in nullizygous and heterozygous embryos. Our results demonstrate the significance of proper tissue stiffness in normal NTC and pave the way for future studies on the mechanobiology of normal and abnormal embryonic development.

Disruption of Fuz in mouse embryos generates hypoplastic hindbrain development and reduced cranial nerve ganglia.

BACKGROUND: The brain and spinal cord formation is initiated in the earliest stages of mammalian pregnancy in a highly organized process known as neurulation. Environmental or genetic interferences can impair neurulation, resulting in clinically significant birth defects known collectively as neural tube defects. The Fuz gene encodes a subunit of the CPLANE complex, a macromolecular planar polarity effector required for ciliogenesis. Ablation of Fuz in mouse embryos results in exencephaly and spina bifida, including dysmorphic craniofacial structures due to defective cilia formation and impaired Sonic Hedgehog signaling. RESULTS: We demonstrate that knocking Fuz out during embryonic mouse development results in a hypoplastic hindbrain phenotype, displaying abnormal rhombomeres with reduced length and width. This phenotype is associated with persistent reduction of ventral neuroepithelial stiffness in a notochord adjacent area at the level of the rhombomere 5. The formation of cranial and paravertebral ganglia is also impaired in these embryos. CONCLUSIONS: This study reveals that hypoplastic hindbrain development, identified by abnormal rhombomere morphology and persistent loss of ventral neuroepithelial stiffness, precedes exencephaly in Fuz ablated murine mutants, indicating that the gene Fuz has a critical function sustaining normal neural tube development and neuronal differentiation.

Optical coherence elastography measures the biomechanical properties of the ex vivo porcine cornea after LASIK.

Significance: The biomechanical impact of refractive surgery has long been an area of investigation. Changes to the cornea structure cause alterations to its mechanical integrity, but few studies have examined its specific mechanical impact. Aim: To quantify how the biomechanical properties of the cornea are altered by laser assisted in situ keratomileusis (LASIK) using optical coherence elastography (OCE) in ex vivo porcine corneas. Approach: Three OCE techniques, wave-based air-coupled ultrasound (ACUS) OCE, heartbeat (Hb) OCE, and compression OCE were used to measure the mechanical properties of paired porcine corneas, where one eye of the pair was left untreated, and the fellow eye underwent LASIK. Changes in stiffness as a function of intraocular pressure (IOP) before and after LASIK were measured using each technique. Results: ACUS-OCE showed that corneal stiffness changed as a function of IOP for both the untreated and the treated groups. The elastic wave speed after LASIK was lower than before LASIK. Hb-OCE and compression OCE showed regional changes in corneal strain after LASIK, where the absolute strain difference between the cornea anterior and posterior increased after LASIK. Conclusions: The results of this study suggest that LASIK may soften the cornea and that these changes are largely localized to the region where the surgery was performed.

Nanobomb optical coherence elastography in multilayered phantoms.

Many tissues are composed of layered structures, and a better understanding of the changes in the layered tissue biomechanics can enable advanced guidance and monitoring of therapy. The advent of elastography using longitudinally propagating shear waves (LSWs) has created the prospect of a high-resolution assessment of depth-dependent tissue elasticity. Laser activation of liquid-to-gas phase transition of dye-loaded perfluorocarbon (PFC) nanodroplets (a.k.a., nanobombs) can produce highly localized LSWs. This study aims to leverage the potential of photoactivation of nanobombs to incudce LSWs with very high-frequency content in wave-based optical coherence elastography (OCE) to estimate the elasticity gradient with high resolution. In this work, we used multilayered tissue-mimicking phantoms to demonstrate that highly localized nanobomb (NB)-induced LSWs can discriminate depth-wise tissue elasticity gradients. The results show that the NB-induced LSWs rapidly change speed when transitioning between layers with different mechanical properties, resulting in an elasticity resolution of ∼65 µm. These results show promise for characterizing the elasticity of multilayer tissue with a fine resolution.

Multifocal acoustic radiation force-based reverberant optical coherence elastography for evaluation of ocular globe biomechanical properties.

Significance: Quantifying the biomechanical properties of the whole eye globe can provide a comprehensive understanding of the interactions among interconnected ocular components during dynamic physiological processes. By doing so, clinicians and researchers can gain valuable insights into the mechanisms underlying ocular diseases, such as glaucoma, and design interventions tailored to each patient's unique needs. Aim: The aim of this study was to evaluate the feasibility and effectiveness of a multifocal acoustic radiation force (ARF) based reverberant optical coherence elastography (RevOCE) technique for quantifying shear wave speeds in different ocular components simultaneously. Approach: We implemented a multifocal ARF technique to generate reverberant shear wave fields, which were then detected using phase-sensitive optical coherence tomography. A 3D-printed acoustic lens array was employed to manipulate a collimated ARF beam generated by an ultrasound transducer, producing multiple focused ARF beams on mouse eye globes ex vivo. RevOCE measurements were conducted using an excitation pulse train consisting of 10 cycles at 3 kHz, followed by data processing to produce a volumetric map of the shear wave speed. Results: The results show that the system can successfully generate reverberant shear wave fields in the eye globe, allowing for simultaneous estimation of shear wave speeds in various ocular components, including cornea, iris, lens, sclera, and retina. A comparative analysis revealed notable differences in wave speeds between different parts of the eye, for example, between the apical region of the cornea and the pupillary zone of the iris (p=0.003). Moreover, the study also revealed regional variations in the biomechanical properties of ocular components as evidenced by greater wave speeds near the apex of the cornea compared to its periphery. Conclusions: The study demonstrated the effectiveness of RevOCE based on a non-invasive multifocal ARF for assessing the biomechanical properties of the whole eyeball. The findings indicate the potential to provide a comprehensive understanding of the mechanical behavior of the whole eye, which could lead to improved diagnosis and treatment of ocular diseases.

Disruption of Fuz in mouse embryos generates hypoplastic hindbrain development and reduced cranial nerve ganglia.

The formation of the brain and spinal cord is initiated in the earliest stages of mammalian pregnancy in a highly organized process known as neurulation. Convergent and extension movements transforms a flat sheet of ectodermal cells into a narrow and elongated line of neuroepithelia, while a major source of Sonic Hedgehog signaling from the notochord induces the overlying neuroepithelial cells to form two apposed neural folds. Afterward, neural tube closure occurs by synchronized coordination of the surface ectoderm and adjacent neuroepithelial walls at specific axial regions known as neuropores. Environmental or genetic interferences can impair neurulation resulting in neural tube defects. The Fuz gene encodes a subunit of the CPLANE complex, which is a macromolecular planar polarity effector required for ciliogenesis. Ablation of Fuz in mouse embryos results in exencephaly and spina bifida, including dysmorphic craniofacial structures due to defective cilia formation and impaired Sonic Hedgehog signaling. In this work, we demonstrate that knocking Fuz out during embryonic mouse development results in a hypoplastic hindbrain phenotype, displaying abnormal rhombomeres with reduced length and width. This phenotype is associated with persistent loss of ventral neuroepithelial stiffness, in a notochord adjacent area at the level of the rhombomere 5, preceding the development of exencephaly in Fuz ablated mutants. The formation of cranial and paravertebral ganglia is also impaired in these embryos, indicating that Fuz has a critical function sustaining normal neural tube development and neuronal differentiation. SIGNIFICANCE STATEMENT: Neural tube defects (NTDs) are a common cause of disability in children, representing the second most common congenital structural malformation in humans following only congenital cardiovascular malformations. NTDs affect approximately 1 to 2 pregnancies per 1000 births every year worldwide, when the mechanical forces folding the neural plate fails to close at specific neuropores located anteriorly (cranial) or posteriorly (caudal) along the neural tube, in a process known as neurulation, which happens throughout the third and fourth weeks of human pregnancy.

Reverberant optical coherence elastography using multifocal acoustic radiation force.

In this study, we introduce a multifocal acoustic radiation force source that combines an ultrasound transducer and a 3D-printed acoustic lens for application in reverberant optical coherence elastography (Rev-OCE). An array of plano-concave acoustic lenses, each with an 11.8 mm aperture diameter, were used to spatially distribute the acoustic energy generated by a 1 MHz planar ultrasound transducer, producing multiple focal spots on a target plane. These focal spots generate reverberant shear wave fields detected by the optical coherence tomography (OCT) system. The effectiveness of the multifocal Rev-OCE system in probing mechanical properties with high resolution is demonstrated in layered gelatin phantoms.

The lens capsule significantly affects the viscoelastic properties of the lens as quantified by optical coherence elastography.

The crystalline lens is a transparent, biconvex structure that has its curvature and refractive power modulated to focus light onto the retina. This intrinsic morphological adjustment of the lens to fulfill changing visual demands is achieved by the coordinated interaction between the lens and its suspension system, which includes the lens capsule. Thus, characterizing the influence of the lens capsule on the whole lens's biomechanical properties is important for understanding the physiological process of accommodation and early diagnosis and treatment of lenticular diseases. In this study, we assessed the viscoelastic properties of the lens using phase-sensitive optical coherence elastography (PhS-OCE) coupled with acoustic radiation force (ARF) excitation. The elastic wave propagation induced by ARF excitation, which was focused on the surface of the lens, was tracked with phase-sensitive optical coherence tomography. Experiments were conducted on eight freshly excised porcine lenses before and after the capsular bag was dissected away. Results showed that the group velocity of the surface elastic wave, V , in the lens with the capsule intact ( V = 2.55 ± 0.23 m / s ) was significantly higher (p < 0.001) than after the capsule was removed ( V = 1.19 ± 0.25 m / s ). Similarly, the viscoelastic assessment using a model that utilizes the dispersion of a surface wave showed that both Young's modulus, E, and shear viscosity coefficient, η, of the encapsulated lens ( E = 8.14 ± 1.10 k P a , η = 0.89 ± 0.093 P a ∙ s ) were significantly higher than that of the decapsulated lens ( E = 3.10 ± 0.43 k P a , η = 0.28 ± 0.021 P a ∙ s ). These findings, together with the geometrical change upon removal of the capsule, indicate that the capsule plays a critical role in determining the viscoelastic properties of the crystalline lens.

Hyaluronan Modulates the Biomechanical Properties of the Cornea.

Purpose: Hyaluronan (HA) is a major constituent of the extracellular matrix (ECM) that has high viscosity and is essential for maintaining tissue hydration. In the cornea, HA is enriched in the limbal region and is a key component of the limbal epithelial stem cell niche. HA is upregulated after injury participating in the formation of the provisional matrix, and has a key role in regulating the wound healing process. This study investigated whether changes in the distribution of HA before and after injury affects the biomechanical properties of the cornea in vivo. Methods: Corneas of wild-type (wt) mice and mice lacking enzymes involved in the biosynthesis of HA were analyzed before, immediately after, and 7 and 14 days after a corneal alkali burn (AB). The corneas were evaluated using both a ring light and fluorescein stain by in vivo confocal microscopy, optical coherence elastography (OCE), and immunostaining of corneal whole mounts. Results: Our results show that wt mice and mice lacking HA synthase (Has)1 and 3 present an increase in corneal stiffness 7 and 14 days after AB without a significant increase in HA expression and absence of scarring at 14 days after AB. In contrast, mice lacking Has2 present a significant decrease in corneal stiffness, with a significant increase in HA expression and scarring at 14 days after AB. Conclusions: Our findings show that the mechanical properties of the cornea are significantly modulated by changes in HA distribution following alkali burn.

Multiple Optical Elastography Techniques Reveal the Regulation of Corneal Stiffness by Collagen XII.

Purpose: Collagen XII plays a role in regulating the structure and mechanical properties of the cornea. In this work, several optical elastography techniques were used to investigate the effect of collagen XII deficiency on the stiffness of the murine cornea. Methods: A three-prong optical elastography approach was used to investigate the mechanical properties of the cornea. Brillouin microscopy, air-coupled ultrasonic optical coherence elastography (OCE) and heartbeat OCE were used to assess the mechanical properties of wild type (WT) and collagen XII-deficient (Col12a1-/-) murine corneas. The Brillouin frequency shift, elastic wave speed, and compressive strain were all measured as a function of intraocular pressure (IOP). Results: All three optical elastography modalities measured a significantly decreased stiffness in the Col12a1-/- compared to the WT (P < 0.01 for all three modalities). The optical coherence elastography techniques showed that mean stiffness increased as a function of IOP; however, Brillouin microscopy showed no discernable trend in Brillouin frequency shift as a function of IOP. Conclusions: Our approach suggests that the absence of collagen XII significantly softens the cornea. Although both optical coherence elastography techniques showed an expected increase in corneal stiffness as a function of IOP, Brillouin microscopy did not show such a relationship, suggesting that the Brillouin longitudinal modulus may not be affected by changes in IOP. Future work will focus on multimodal biomechanical models, evaluating the effects of other collagen types on corneal stiffness, and in vivo measurements.

Micron-scale hysteresis measurement using dynamic optical coherence elastography.

We present a novel optical coherence elastography (OCE) method to characterize mechanical hysteresis of soft tissues based on transient (milliseconds), low-pressure (<20 Pa) non-contact microliter air-pulse stimulation and micrometer-scale sample displacements. The energy dissipation rate (sample hysteresis) was quantified for soft-tissue phantoms (0.8% to 2.0% agar) and beef shank samples under different loading forces and displacement amplitudes. Sample hysteresis was defined as the loss ratio (hysteresis loop area divided by the total loading energy). The loss ratio was primarily driven by the sample unloading response which decreased as loading energy increased. Samples were distinguishable based on their loss ratio responses as a function loading energy or displacement amplitude. Finite element analysis and mechanical testing methods were used to validate these observations. We further performed the OCE measurements on a beef shank tissue sample to distinguish the muscle and connective tissue components based on the displacement and hysteresis features. This novel, noninvasive OCE approach has the potential to differentiate soft tissues by quantifying their viscoelasticity using micron-scale transient tissue displacement dynamics. Focal tissue hysteresis measurements could provide additional clinically useful metrics for guiding disease diagnosis and tissue treatment responses.

In vivo assessment of corneal biomechanics under a localized cross-linking treatment using confocal air-coupled optical coherence elastography.

The localized application of the riboflavin/UV-A collagen cross-linking (UV-CXL) corneal treatment has been proposed to concentrate the stiffening process only in the compromised regions of the cornea by limiting the epithelium removal and irradiation area. However, current clinical screening devices dedicated to measuring corneal biomechanics cannot provide maps nor spatial-dependent changes of elasticity in corneas when treated locally with UV-CXL. In this study, we leverage our previously reported confocal air-coupled ultrasonic optical coherence elastography (ACUS-OCE) probe to study local changes of corneal elasticity in three cases: untreated, half-CXL-treated, and full-CXL-treated in vivo rabbit corneas (n = 8). We found a significant increase of the shear modulus in the half-treated (>450%) and full-treated (>650%) corneal regions when compared to the non-treated cases. Therefore, the ACUS-OCE technology possesses a great potential in detecting spatially-dependent mechanical properties of the cornea at multiple meridians and generating elastography maps that are clinically relevant for patient-specific treatment planning and monitoring of UV-CXL procedures.

Multimodal Heartbeat and Compression Optical Coherence Elastography for Mapping Corneal Biomechanics.

The biomechanical properties of the cornea have a profound influence on the health, structural integrity, and function of the eye. Understanding these properties may be critical for diagnosis and identifying disease pathogenesis. This work demonstrates how two different elastography techniques can be combined for a multimodal approach to measuring corneal biomechanical properties. Heartbeat optical coherence elastography (Hb-OCE) and compression OCE were performed simultaneously to measure the stiffness of the cornea in an in vivo rabbit model. Measurements were further performed after collagen crosslinking to demonstrate how the combined technique can be used to measure changes in corneal stiffness and map mechanical contrast. The results of this work further suggest that measurements from Hb-OCE and compression OCE are comparable, meaning that Hb-OCE and compression OCE may be used interchangeably despite distinct differences in both techniques.

Longitudinal assessment of the effect of alkali burns on corneal biomechanical properties using optical coherence elastography.

Eye injury due to alkali burn is a severe ocular trauma that can profoundly affect corneal structure and function, including its biomechanical properties. Here, we assess the changes in the mechanical behavior of mouse corneas in response to alkali-induced injury by conducting longitudinal measurements using optical coherence elastography (OCE). A non-contact air-coupled ultrasound transducer was used to induce elastic waves in control and alkali-injured mouse corneas in vivo, which were imaged with phase-sensitive optical coherence tomography. Corneal mechanical properties were estimated using a modified Rayleigh-Lamb wave model, and results show that Young's modulus of alkali-burned corneas were significantly greater than that of their healthy counterparts on days 7 (p = 0.029) and 14 (p = 0.026) after injury. These findings, together with the changes in the shear viscosity coefficient postburn, indicate that the mechanical properties of the alkali-burned cornea are significantly modulated during the wound healing process.

Multimodal imaging system combining optical coherence tomography and Brillouin microscopy for neural tube imaging.

To understand the dynamics of tissue stiffness during neural tube formation and closure in a murine model, we have developed a multimodal, coaligned imaging system combining optical coherence tomography (OCT) and Brillouin microscopy. Brillouin microscopy can map the longitudinal modulus of tissue but cannot provide structural images. Thus, it is limited for imaging dynamic processes such as neural tube formation and closure. To overcome this limitation, we have combined Brillouin microscopy and OCT in one coaligned instrument. OCT provided depth-resolved structural imaging with a micrometer-scale spatial resolution to guide stiffness mapping by Brillouin modality. 2D structural and Brillouin frequency shift maps were acquired of mouse embryos at gestational day (GD) 8.5, 9.5, and 10.5 with the multimodal system. The results demonstrate the capability of the system to obtain structural and stiffness information simultaneously.

Ultra-fast dynamic line-field optical coherence elastography.

In this work, we present an ultra-fast line-field optical coherence elastography system (LF-OCE) with an 11.5 MHz equivalent A-line rate. The system was composed of a line-field spectral domain optical coherence tomography system based on a supercontinuum light source, Michelson-type interferometer, and a high-speed 2D spectrometer. The system performed ultra-fast imaging of elastic waves in tissue-mimicking phantoms of various elasticities. The results corroborated well with mechanical testing. Following validation, LF-OCE measurements were made in in situ and in in vivo rabbit corneas under various conditions. The results show the capability of the system to rapidly image elastic waves in tissues.

Ultrasound Shear Wave Elastography and Transient Optical Coherence Elastography: Side-by-Side Comparison of Repeatability and Accuracy.

OBJECTIVE: We compare the repeatability and accuracy of ultrasound shear wave elastography (USE) and transient optical coherence elastography (OCE). METHODS: Elastic wave speed in gelatin phantoms and chicken breast was measured with USE and OCE and compared with uniaxial mechanical compression testing. Intra- and Inter-repeatability were analyzed using Bland-Altman plots and intraclass correlation coefficients (ICC). RESULTS: OCE and USE differed from uniaxial testing by a mean absolute percent error of 8.92% and 16.9%, respectively, across eight phantoms of varying stiffness. Upper and lower limits of agreement for intrasample repeatability for USE and OCE were ±0.075 m/s and -0.14 m/s and 0.13 m/s, respectively. OCE and USE both had ICCs of 0.9991. In chicken breast, ICC for USE was 0.9385 and for OCE was 0.9924. CONCLUSION: OCE and USE can detect small speed changes and give comparable measurements. These measurements correspond well with uniaxial testing.

Micro Air-Pulse Spatial Deformation Spreading Characterizes Degree of Anisotropy in Tissues.

In optical coherence elastography (OCE), air-pulse stimulation has been widely used to produce propagation of mechanical waves for elastic characterization of tissues. In this paper, we propose the use of spatial deformation spreading (SDS) on the surface of samples produced by air-pulse stimulation for the OCE of transverse isotropic tissues. Experiments in isotropic tissue-mimicking phantoms and anisotropic chicken tibialis muscle were conducted using a spectral-domain optical coherence tomography system synchronized with a confocal air-pulse stimulation. SDS measurements were compared with wave speeds values calculated at different propagation angles. We found an approximately linear relationship between shear wave speed and SDS in isotropic phantoms, which was confirmed with predictions made by the numerical integration of a wave propagation model. Experimental measurements in chicken muscle show a good agreement between SDS and surface wave speed taken along and across the axis of symmetry of the tissues, also called degree of anisotropy. In summary, these results demonstrated the capabilities of SDS produced by the air-pulse technique in measuring the shear elastic anisotropy of transverse isotropic tissues.