Long‐Term Imaging of Wound Angiogenesis with Large Scale Optoacoustic Microscopy

Abstract

Wound healing is a well-coordinated process, necessitating efficient formation of new blood vessels. Vascularization defects are therefore a major risk factor for chronic, non-healing wounds. The dynamics of mammalian tissue revascularization, vessel maturation, and remodeling remain poorly understood due to lack of suitable in vivo imaging tools. A label-free large-scale optoacoustic microscopy (LSOM) approach is developed for rapid, non-invasive, volumetric imaging of tissue regeneration over large areas spanning up to 50 mm with a depth penetration of 1.5 mm. Vascular networks in dorsal mouse skin and full-thickness excisional wounds are imaged with capillary resolution during the course of healing, revealing previously undocumented views of the angiogenesis process in an unperturbed wound environment. Development of an automatic analysis framework enables the identification of key features of wound angiogenesis, including vessel length, diameter, tortuosity, and angular alignment. The approach offers a versatile tool for preclinical research in tissue engineering and regenerative medicine, empowering label-free, longitudinal, high-throughput, and quantitative studies of the microcirculation in processes associated with normal and impaired vascular remodeling, and analysis of vascular responses to pharmacological interventions in vivo.

Tracking Strain-Specific Morphogenesis and Angiogenesis of Murine Calvaria with Large-Scale Optoacoustic and Ultrasound Microscopy

“…Large‐scale OA microscopy (LSOM) reveals optical absorption of hemoglobin via acoustic detection, while pulse‐echo US delineates the skull bone surface based on its distinctive acoustic impedance. ( 23 ) A total of 15 mice (five per strain) were imaged at four time points (4, 8, 12, and 16 weeks old, Fig. 1 B ), resulting in 60 volumetric datasets.…”

“…A previously reported automatic vessel segmentation and analysis algorithm was subsequently applied. ( 24,25 ) Briefly, MIPs were binarized and skeletonized, and vessel central lines were found by spline fitting. Vessel edges were then determined by computing the image gradient along the normal direction of the central lines.…”

The Potential of Photoacoustic Imaging in Radiation Oncology

“…Beyond vascular imaging, the emerging capabilities of PAI for fibrosis imaging (129)(130)(131) may aid characterization of this late-stage skin toxicity manifestation (12,120). In addition, PAI has shown promise for assessment of burns (132), wound healing (133,134), and skin disorders such as psoriasis (135), all of which present with features similar to those in radiation-induced injuries. These studies highlight the promising potential of PAI to evaluate and assess vascular changes caused by radiation.…”

Quantification of vascular networks in photoacoustic mesoscopy

“…Preclinically, mesoscopic PAI has been used to monitor the development of vasculature in several tumour xenograft models [17] , [18] , [19] and can differentiate aggressive from slow-growing vascular phenotypes [19] . Studies to-date, however, have been largely restricted to qualitative analyses due to the challenges of accurate 3D vessel segmentation, quantification and robust statistical analyses [17] , [20] , [15] , [18] , [19] , [21] . Instead, PAI quantification is typically manual and ad-hoc , with 2D measurements often extracted from 3D PAI data [17] , [20] , [22] , [19] , [23] , reducing repeatability and comparability across datasets.…”