Phase-domain Photoacoustic Mechanical Imaging for Quantitative Elastography and Viscography.

The role and importance of mechanical properties of cells and tissues in pathophysiological processes have widely been acknowledged. However, current elastography techniques most based on transverse elastic waves, diminish the translation of wave speed into elastic modulus due to its limited wave propagation direction. Here, we propose phase-domain photoacoustic mechanical imaging (PD-PAMI), leveraging the initial time and phase response characteristics of an omnidirectional photoacoustic elastic wave to quantitatively extract elastic and viscous moduli. Theoretical simulations and experiment on tissue-mimicking phantoms with different levels of viscoelastic properties were conducted to validate the approach with a precision in elasticity and viscosity estimation of 4.6% and 6.6%, respectively. The trans-scale viscoelasticity mappings over three length scales-covering cell, tissue section, and in vivo organ, were provided to demonstrate the scalability of the technique with different implementations of PD-PAMI. Experiments on animal models of breast tumour and atherosclerosis reveal that PD-PAMI technique enables effective monitoring of the viscoelastic parameters for examinations of the diseases involved with the variations in collagen or lipid composition and in inflammation level. PD-PAMI technique opens new perspectives of conventional PA imaging and provides new technical way for biomechanical imaging, prefiguring potential clinical applications in mechanopathology-involved disease diagnosis.

Photoacoustic micro-viscoelastography for mapping mechanocellular properties.

Cellular biomechanical properties provide essential insights into biological functions regarding health and disease. Current measurements of the biomechanical properties of cells require physical contact with cells or pre-loading on the cells. Here, we have developed photoacoustic micro-viscoelastography (PAMVE), which utilizes the phase characteristics of photoacoustic (PA) response, for mapping mechanocellular properties in a load-free manner. PAMVE realizes the local viscoelasticity measurement on the macrophages and red blood cells with micrometer scale. Furthermore, PAMVE can successfully identify the adipose cell and skeletal muscle cell due to the difference in their composition-related biomechanical properties. PAMVE represents an irreplaceable option for interrogating characteristic mechanocellular properties, opening the possibility of studying cellular mechanobiology and pathophysiology.

Design of Plasmonic Photonic Crystal Fiber for Highly Sensitive Magnetic Field and Temperature Simultaneous Measurement.

A high-sensitivity plasmonic photonic crystal fiber (PCF) sensor is designed and a metal thin film is embedded for achieving surface plasmon resonance (SPR), which can detect the magnetic field and temperature simultaneously. Within the plasmonic PCF sensor, the SPR sensing is accomplished by coating both the upper sensing channel (Ch1) and the lower sensing channel (Ch2) with gold film. In addition, the temperature-sensitive medium polydimethylsiloxane (PDMS) is chosen to fill in Ch1, allowing the sensor to respond to the temperature. The magnetic field-sensitive medium magnetic fluid (MF) is chosen to fill in Ch2, allowing this sensor to respond to the magnetic field. During these processes, this proposed SPR-PCF sensor can achieve dual-parameter sensing. The paper also investigates the electrical field characteristics, structural parameters and sensing performance using COMSOL. Finally, under the magnetic field range of 50-130 Oe, this sensor has magnetic field sensing sensitivities of 0 pm/Oe (Ch1) and 235 pm/Oe (Ch2). In addition, this paper also investigates the response of temperature. Under the temperature range of 20-40 °C, Ch1 and Ch2 have temperature sensitivities of -2000 pm/°C and 0 pm/°C, respectively. It is noteworthy that the two sensing channels respond to only a single physical parameter; this sensing performance is not common in dual-parameter sensing. Due to this sensing performance, it can be found that the magnetic field and temperature can be detected by this designed SPR-PCF sensor simultaneously without founding and calculating a sensing matrix. This sensing performance can solve the cross-sensitivity problem of magnetic field and temperature, thus reducing the measurement error. Since it can sense without a matrix, it further can solve the ill-conditioned matrix and nonlinear change in sensitivity problems in dual-parameter sensing. These excellent sensing capabilities are very important for carrying out multiparameter sensing in complicated environments.

Automatic and label-free detection of meningioma in dura mater using the combination of multiphoton microscopy and image analysis.

Significance: To prevent meningioma recurrence, it is necessary to detect and remove all corresponding tumors intraoperatively, including those in the adjacent dura mater. Aim: Currently, the removal of meningiomas from the dura mater depends solely on cautious visual identification of lesions by a neurosurgeon. Inspired by the requirements for resection, we propose multiphoton microscopy (MPM) based on two-photon-excited fluorescence and second-harmonic generation as a histopathological diagnostic paradigm to assist neurosurgeons in achieving precise and complete resection. Approach: Seven fresh normal human dura mater samples and 10 meningioma-infiltrated dura mater samples, collected from 10 patients with meningioma, were acquired for this study. First, multi-channel mode and lambda mode detection were utilized in the MPM to characterize the architectural and spectral features of normal and meningioma-infiltrated dura mater, respectively. Three imaging algorithms were then employed to quantify the architectural differences between the normal and meningioma-infiltrated dura mater through calculations of the collagen content, orientation, and alignment. Finally, MPM was combined with another custom-developed imaging algorithm to locate the meningioma within the dura mater and further delineate the tumor boundary. Results: MPM not only detected meningioma cells in the dura mater but also revealed the morphological and spectral differences between normal and meningioma-infiltrated dura mater, providing quantitative information. Furthermore, combined with a self-developed image-processing algorithm, the precise borders of meningiomas in the dura mater could be accurately delineated. Conclusions: MPM can automatically detect meningiomas in the dura mater label-free. With the development of advanced multiphoton endoscopy, MPM combined with image analysis can provide decision-making support for histopathological diagnosis, as well as offer neurosurgeons more precise intraoperative resection guidance for meningiomas.

Effects of Different Spectrum of LEDs on Retinal Degeneration Through Regulating Endoplasmic Reticulum Stress.

Purpose: To evaluate the impact of full-spectrum light-emitting diodes (LEDs) on albino guinea pigs' retina and investigate the roles of short-wavelength opsin (S-opsin) and endoplasmic reticulum (ER) stress in light-induced retinal degeneration (LIRD). Methods: Three-week-old albino guinea pigs (n = 30) were distributed into five groups under 12/12 light/dark cycles with indoor natural light (NC; 300-500 lux, n = 6), full-spectrum LEDs (FL; 300 lux, n = 6; 3000 lux, n = 6), and commercial cold-white LEDs (CL; 300 lux, n = 6; 3000 lux, n = 6) and raised for 28 days. Hematoxylin and eosin staining and transmission electron microscopy evaluated the morphological changes of retinas. The immunofluorescence and real-time quantitative polymerase chain reaction (RT-qPCR) measured the expression and content of S-opsin and ER stress-related genes and proteins. Results: We found that albino guinea pigs exposed to FL at either 300 lux or 3000 lux developed less severe retinal morphological damage than animals exposed to the CL light, which emerged as a significant characteristic of LIRD. Meanwhile, the damage on the ventral retina was more serious, mainly due to its ability to absorb the blue light in the LEDs more easily. Compared to the FL-exposed groups, the CL light increased the aggregation of S-opsin and the expression of ER stress-related factors. Conclusions: Commercial cold-white LEDs can induce ER stress and unfolded protein response in LIRD, and full-spectrum LED attenuates LIRD by regulating ER stress in albino guinea pig retinas in vivo. Translational Relevance: Full-spectrum LEDs offer specific eye protection and eye adaptability that can well replace commercial cold-white LEDs in both clinical practice and research. It should be further developed for lighting used in health care facilities.

All-optical noncontact phase-domain photoacoustic elastography.

Mechanical properties such as elasticity are important indicators of tissue functions that can be used for clinical diagnosis and disease monitoring. However, most current elastography techniques are limited in their ability to distinguish localized microstructural mechanical variations due to employing elastic wave velocity measurement. In addition, their contact-based measurement manner is not favored and may even be prohibited in many applications. In this Letter, we propose all-optical noncontact phase-domain photoacoustic elastography (NPD-PAE), leveraging the temporal response characteristics of laser-induced thermoelastic displacement using optical interferometric detection to calculate the elastic modulus. The all-optical pump-probe method allows the capture of the initial displacement profiles generated at the origin, thus enabling the extraction of in situ elasticity. The feasibility of the method was verified using a tissue-mimicking phantom. The capability to map the mechanical contrast was demonstrated on an ex vivo biological tissue. NPD-PAE opens a new avenue for development of a noncontact elastography technique, holding great potential in the biomedical field and materials science.

Optical coherence hyperspectral microscopy with a single supercontinuum light source.

In the paper, we have developed an optical coherence hyperspectral microscopy with a single supercontinuum light source. The microscopy consists of optical coherence tomography (OCT) and hyperspectral imaging (HSI), which can visualize the structural and functional characteristics of biological tissues. The 500 to 700 nm band is selected for HSI and OCT imaging, where HSI enables imaging of oxygen saturation and hemoglobin (Hb) content, while OCT acquires structural characteristics to assess the morphology of biological tissues. The system performance of the optical coherence hyperspectral microscopy is verified by normal mice ears, and the practical applications of the microscopy is further performed in 4T1 and inflammation Balb/c mice ears in vivo. The experimental results demonstrate that the microscopy has potential to provide complementary information for clinical applications.

Spectral interferometric depth-resolved photoacoustic viscoelasticity imaging.

Viscoelasticity is closely related to the physiological characteristics of biological tissues. In this Letter, we propose a novel spectral interferometric depth-resolved photoacoustic viscoelasticity imaging (SID-PAVEI) method, to the best of our knowledge for the first time, which breaks the plight of surface viscoelasticity imaging and achieves an internal visible microscale SID-PAVEI in a noncontact fashion. In this work, we employ a high-sensitive and depth-resolved spectral domain low coherence interferometry (SDLCI) to remotely track photoacoustic-induced strain response of absorbers in situ. By decoupling the phase and amplitude of the photoacoustic-encoded spectral interference signal, the SID-PAVEI and scattering structure imaging (SSI) can be obtained simultaneously. Depth-resolved performance of the SID-PAVEI and the SSI in one scan were demonstrated by imaging biological tissues. The method opens new perspectives for three-dimensional microscale viscoelasticity imaging and provides a great potential in multi-parametric characterizing pathological information.

Towards quantitative assessment of burn based on photoacoustic and optical coherence tomography.

Accurate and timely assessment of the severity of burn is essential for the treatment of burns. Currently, although most first-degree and third-degree burns are easily diagnosed through visual inspection or auxiliary diagnostic methods, the second-degree burn is still difficult to distinguish due to the ambiguity boundaries of second-degree with first-degree and third-degree burns. In this study, we proposed a non-invasive technique by combing photoacoustic imaging (PAI) and optical coherence tomography (OCT) to multi-parameter quantitatively assess the burns. The feasibility and capacity of the dual-mode PAT/OCT for assessing the burns was first testified by tissue-mimicking phantom and burn wounds in mouse pinna in vivo. The further experiments conducted on the back of rats showed that the changes in skin scattering structure, vascular morphology and blood flow provided by the dual-mode PAI/OCT system can determine distinct boundaries and depth of the burns. The experimental results prove that combined PAI/OCT as a novel method can be used to assess the severity of burn, which has the potential to diagnose the burns in clinic.

Multi-parameter characterization of atherosclerotic plaques based on optical coherence tomography, photoacoustic and viscoelasticity imaging.

Detection of atherosclerotic plaque vulnerability is the critical step in prevention of acute coronary events. Fibrous cap thickness, lipid core size, and inflammation extent are three key parameters for assessing plaque vulnerability. Here, we report on multimodality imaging of mice aortic plaques using a system that integrates optical coherence tomography (OCT), photoacoustic imaging (PAI), and photoacoustic viscoelasticity imaging (PAVEI). The thickness of fibrous cap is accurately evaluated by OCT, and PAI helps to determine the distribution and size of lipid core. The mechanical properties of plaques are closely related to the plaque compositions and the content and distribution of macrophages, while PAVEI can characterize the plaque viscoelasticity through the phase delay of photoacoustic signal. Experimental results demonstrate that the OCT-PAI-PAVEI system can comprehensively characterize the three traits of atherosclerotic plaques, thereby identifying high-risk lesions.

Integrated photoacoustic and hyperspectral dual-modality microscopy for co-imaging of melanoma and cutaneous squamous cell carcinoma in vivo.

Skin carcinoma such as melanoma (MM) and cutaneous squamous cell carcinoma (cSCC) are considered as the highest mortality and the most aggressive skin cancers in dermatology. In view that early diagnosis and treatment can greatly improve the survival rate and life quality of the patients, developing noninvasive and effective evaluation methods is of great significance for the detection and identification of early stage cutaneous cancers. In this article, we propose a hybrid photoacoustic and hyperspectral dual-modality microscopy to evaluate and differentiate skin carcinoma by structural and multiphysiological parameters. The proposed system's imaging abilities are verified by mimic phantoms and normal mice experiments. Furthermore, in vivo characterization and evaluation results of MM and cSCC mice are obtained successfully, which prove this novel method could be used as a reliable and useful method for skin cancer detection in early stages.

Single-Cell Photoacoustic Microrheology.

Rheological properties, such as elasticity and viscosity, are fundamental biomechanical parameters that are related to the function and pathological status of cells and tissues. In this paper, an innovative photoacoustic microrheology (PAMR), which utilized the time and phase characteristics of photoacoustic (PA) response, was proposed to extract elastic modulus and viscosity. The feasibility and accuracy of the method were validated by tissue-mimicking agar-gelatin phantoms with various viscoelasticity values. PAMR realized single cell elasticity and viscosity mappings on the adipocyte and myocyte with micrometer scale. In clinical samples, normal blood cells and iron deficiency anemia cells were successfully distinguished due to their various rheological properties. This method expands the scope of conventional PA imaging and opens new possibilities for developing microrheological technology, prefiguring great clinical potential for interrogating mechanocellular properties.

Optical Biopsy of Melanoma and Basal Cell Carcinoma Progression by Noncontact Photoacoustic and Optical Coherence Tomography: In Vivo Multi-Parametric Characterizing Tumor Microenvironment.

Measuring the structural and functional status of tumor microenvironment for malignant melanoma (MM) and basal cell carcinoma (BCC) is of profound significance in understanding dermatological condition for biopsy. However, conventional optical imaging techniques are limited to visualize superficial skin features and parameter information is deficient to depict pathophysiology correlations of skin diseases. Here, we demonstrate a preclinical device, all-optically integrated photoacoustic and optical coherence tomography (AOPA/OCT), that, for the first time, can simultaneously provide label-free biomarkers of vascular patterns, temporal and spatial heterogeneity of blood flow, and tissue micro-structure changes during tumor growth with pathophysiological correlations in mice models. We found that tumor microenvironment of MM and BCC led to the alternation in spatial-temporal heterogeneity that affected morphological and functional parameters, performing the AOPA/OCT quantitative metrics. A robust correlation between imaging biomarkers derived from this in vivo technique and histopathology validation ex vivo in distinguishing benign from malignant is also presented. In receiver operating characteristics (ROC) analysis, multi-parametric AOPA/OCT yields improved diagnostic accuracy of 98.4% and 95.8% for MM and BCC respectively, which indicate that AOPA/OCT represents a high-performance and clinically translatable technique for accurate diagnosis and therapy monitoring in dermatology.

Co-impulse multispectral photoacoustic microscopy and optical coherence tomography system using a single supercontinuum laser.

A combination of multispectral photoacoustic microscopy (PAM) and optical coherence tomography (OCT) by a single light source was previously realized discretely; however, this is unfavorable for visualizing vital physiological and pathological activities in vivo. Here, a co-impulse dual-mode imaging system that simultaneously enables multispectral PAM and OCT using a megahertz supercontinuum pulse laser in vivo is presented. The 500-600 nm band is used for functional PAM imaging, which can flexibly switch between different wavelengths, while the 600-840 nm band is selected for OCT imaging. A mimicking phantom experiment and in vivo imaging of normal and melanoma mouse ears demonstrate that the co-impulse multispectral PAM-OCT system can simultaneously provide structural and functional information for bioimaging.

Extended depth-of-field all-optical photoacoustic microscopy with a dual non-diffracting Bessel beam.

All-optical photoacoustic microscopy (AOPAM) facilitates high-sensitivity, wide-bandwidth, volumetric imaging without coupling media. However, the rapid divergence of the Gaussian beam restricts the stability and depth-of-field in typical Gaussian AOPAM (G-AOPAM). Here we report an extended depth-of-field AOPAM using a dual non-diffracting Bessel beam (B-AOPAM). Benefiting from the designing, the B-AOPAM has the unique advantages of increasing depth resolving ability and improving photoacoustic detection sensitivity. The proposed scheme shows optimal lateral resolution of 2.4 µm and a long depth-of-focus of 635 µm, which is 10-fold larger than that of the G-AOPAM. The scattering phantoms and in vivo animal experiments demonstrated the imaging feasibility and capability of the B-AOPAM, which can provide noncontact, high spatial resolution imaging of non-flat tissue and contribute to future clinical applications.

Cartilage oligomeric matrix protein is a novel notch ligand driving embryonic stem cell differentiation towards the smooth muscle lineage.

Cartilage oligomeric matrix protein (COMP), a protective component of vascular extracellular matrix (ECM), maintains the homeostasis of mature vascular smooth muscle cells (VSMCs). However, whether COMP modulates the differentiation of stem cells towards the smooth muscle lineage is still elusive. Firstly, purified mouse COMP directly induced mouse embryonic stem cell (ESC) differentiation into VSMCs both in vitro and in vivo, while the silencing of endogenous COMP markedly inhibited ESC-VSMC differentiation. RNA-Sequencing revealed that Notch signaling was significantly activated by COMP during ESC-VSMC differentiation, whereas the inhibition of Notch signaling attenuated COMP-directed ESC-VSMC differentiation. Furthermore, COMP deficiency inhibited Notch activation and VSMC differentiation in mice. Through silencing distinct Notch receptors, we identified that Notch1 mainly mediated COMP-initiated ESC-VSMC differentiation. Mechanistically, COMP N-terminus directly interacted with the EGF11-12 domain of Notch1 and activated Notch1 signaling, as evidenced by co-immunoprecipitation and mammalian two-hybrid assay. In conclusion, COMP served as a potential ligand of Notch1, thereby driving ESC-VSMC differentiation.

Deficiency of FAM3D (Family With Sequence Similarity 3, Member D), A Novel Chemokine, Attenuates Neutrophil Recruitment and Ameliorates Abdominal Aortic Aneurysm Development.

OBJECTIVE: Chemokine-mediated neutrophil recruitment contributes to the pathogenesis of abdominal aortic aneurysm (AAA) and may serve as a promising therapeutic target. FAM3D (family with sequence similarity 3, member D) is a recently identified novel chemokine. Here, we aimed to explore the role of FAM3D in neutrophil recruitment and AAA development. APPROACH AND RESULTS: FAM3D was markedly upregulated in human AAA tissues, as well as both elastase- and CaPO4-induced mouse aneurysmal aortas. FAM3D deficiency significantly attenuated the development of AAA in both mouse models. Flow cytometry analysis indicated that FAM3D-/- mice exhibited decreased neutrophil infiltration in the aorta during the early stage of AAA formation compared with their wild-type littermates. Moreover, application of FAM3D-neutralizing antibody 6D7 through intraperitoneal injection markedly ameliorated elastase-induced AAA formation and neutrophil infiltration. Further, in vitro coculture experiments with FAM3D-neutralizing antibody 6D7 and in vivo intravital microscopic analysis indicated that endothelial cell-derived FAM3D induced neutrophil recruitment. Mechanistically, FAM3D upregulated and activated Mac-1 (macrophage-1 antigen) in neutrophils, whereas inhibition of FPR1 (formyl peptide receptor 1) or FPR2 significantly blocked FAM3D-induced Mac-1 activation, indicating that the effect of FAM3D was dependent on both FPRs. Moreover, specific inhibitors of FPR signaling related to Gi protein or ß-arrestin inhibited FAM3D-activated Mac-1 in vitro, whereas FAM3D deficiency decreased the activation of both FPR-Gi protein and ß-arrestin signaling in neutrophils in vivo. CONCLUSIONS: FAM3D, as a dual agonist of FPR1 and FPR2, induced Mac-1-mediated neutrophil recruitment and aggravated AAA development through FPR-related Gi protein and ß-arrestin signaling.

All-optical photoacoustic Doppler transverse blood flow imaging.

The method of measuring blood flow in photoacoustic microscopy usually relies on ultrasonic transducers in contact fashion, which is not favored in many applications, such as wound areas, burns, and anabrosis. Here we present a noncontact photoacoustic velocity measurement method to quantitatively map transverse blood flow based on the photoacoustic Doppler (PAD) bandwidth broadening method with an all-optical photoacoustic microscopy system. It is validated that the PAD bandwidth broadening is proportional to the transverse flow within a certain range. The transverse flow speed ranging from 0 to 5.5 mm/s, as well as sectional flow images, was obtained in the blood-mimicking flow phantoms. Furthermore, the blood flow image of the mouse ear demonstrates that the all-optical photoacoustic Doppler method can acquire the information of blood flow in vivo, which could significantly broaden the scope of applications for obtaining the blood flow velocity of the microvasculature in biomedicine.

Optical biopsy approach to basal cell carcinoma and melanoma based on all-optically integrated photoacoustic and optical coherence tomography.

Basal cell carcinoma (BCC) and melanoma (MM), with the highest morbidity and mortality, respectively, are considered as two skin cancers of concern in dermatology. Histological studies have demonstrated that vascular patterns and collagenous stroma serve as key parameters for BCC and MM classification. In this Letter, we sought to identify BCC and MM based on the dual parameters of vascular patterns and scattering structures provided by all-optically integrated photoacoustic and optical coherence tomography (AOPA/OCT). The imaging capability of the AOPA/OCT was verified by the mimic phantoms. Furthermore, in vivo characterization of vasculatures and tissue structures from BCC and MM mice were successfully achieved with high resolution. Results prove the feasibility of AOPA/OCT as a novel method to dedicate to the in vivo biopsy of skin cancers which shows new insights into the study of skin diseases in early stages.

Autophagy regulatory molecule, TMEM74, interacts with BIK and inhibits BIK-induced apoptosis.

TMEM74 (Transmembrane protein 74), a lysosome transmembrane protein, induces cell autophagy. Knockdown of TMEM74 abolished EBSS-induced autophagy. BIK, belonging to BOP (BH3-only protein) protein family, has been reported to induce cell apoptosis. Autophagy and apoptosis, as different pathways regulated by extra- or intra-cellular signals precisely, both play a crucial role in processes of intra-cellular substrates degradation, energy metabolism and cell survival. However, the relationship between autophagy and apoptosis still remains elusive. To elucidate the putative new relationship and further identify the function of TMEM74, we performed the study mainly using co-immunoprecipitation, immunoblotting, fluorescent location and basic cell biologic experimental techniques. In the present study, for the first time, it is demonstrated that autophagy-related protein TMEM74 co-localizes with apoptosis-related protein BIK in subcellular organelles. The data indicated that TMEM74 associates with BIK via TM domains of TMEM74 and BH3 domain of BIK. Further investigations revealed that TMEM74 inhibits BIK-induced apoptosis by interacting with BIK, as evidenced by the results that autophagosome formation inhibitor could not block the inhibition effect completely. On the contrary, knockdown of TMEM74 and the TM domain-deficient mutant led to deprivation of the function. Overall, the results revealed the autophagy modulator TMEM74 interrelates with apoptosis inducer BIK and inhibits its function, which provides a novel crosstalk point between autophagy and apoptosis to enlarge our understanding of the programmed cell death.