We present a modified method of embedded bioprinting, which allows maintaining freestanding threedimensional (3D) printed structures in cell culture conditions for extended periods of time. This method, termed CLASS (constructs laid in agarose slurry suspension), was tested using cell-laden alginate and gelatin methacrylate (GelMa)-based bioinks. A direct comparison of 3D printed constructs, supported by gelatin and agarose hydrogel slurries, revealed several advantages, including slurry stability across different print temperatures and blending times, increased slurry homogeneity, and the ability of CLASS to support freestanding constructs for an extended time in cell culture. We conclude that CLASS is a straightforward and cost-efficient way to print and support freestanding cell-laden biomaterials.

Aim-To 3D print heart tissue, one must understand how the main two types of cardiac cells are affected by the printing process. Materials & methods-Effects of gelatin methacryloyl (GelMA) concentration, extruder pressure and duration of UV exposure on survival of cardiac myocytes and fibroblasts were examined using lactate dehydrogenase and LIVE/DEAD assays, bioluminescence imaging and morphological assessment. Results & conclusion-Cell survival within 3D printed cardiomyocyte-laden GelMA constructs was more sensitive to extruder pressure and GelMA concentrations than within 3D fibroblast-laden GelMA constructs. Cells within both types of constructs were adversely impacted by the UV curing step. Use of mixed cell populations and enrichment of bioink formulation with fibronectin led to an improvement of cardiomyocyte survival and spreading.

Radiofrequency ablation (RFA) is a widely used treatment for atrial fibrillation, the most common cardiac arrhythmia. Here, we explore autofluorescence hyperspectral imaging (aHSI) as a method to visualize RFA lesions and interlesional gaps in the highly collagenous left atrium. RFA lesions made on the endocardial surface of freshly excised porcine left atrial tissue were illuminated by UV light (365 nm), and hyperspectral datacubes were acquired over the visible range (420–720 nm). Linear unmixing was used to delineate RFA lesions from surrounding tissue, and lesion diameters derived from unmixed component images were quantitatively compared to gross pathology. RFA caused two consistent changes in the autofluorescence emission profile: a decrease at wavelengths below 490 nm (ascribed to a loss of endogenous NADH) and an increase at wavelengths above 490 nm (ascribed to increased scattering). These spectral changes enabled high resolution, in situ delineation of RFA lesion boundaries without the need for additional staining or exogenous markers. Our results confirm the feasibility of using aHSI to visualize RFA lesions at clinically relevant locations. If integrated into a percutaneous visualization catheter, aHSI would enable widefield optical surgical guidance during RFA procedures and could improve patient outcome by reducing atrial fibrillation recurrence.

HSI based on changes in tissue autofluorescence is a highly effective tool for revealing-in vivo and with high spatial resolution-surface boundaries of myocardial scar and discriminating it from areas of acute necrosis caused by RF ablation.

BackgroundCurrently, there are limited means for high-resolution monitoring of tissue injury during radiofrequency ablation procedures.ObjectiveTo develop the next generation of visualization catheters that can reveal irreversible atrial muscle damage caused by ablation and identify viability gaps between the lesions.MethodsRadiofrequency lesions were placed on the endocardial surfaces of excised human and bovine atria and left ventricles of blood perfused rat hearts. Tissue was illuminated with 365nm light and a series of images were acquired from individual spectral bands within 420-720nm range. By extracting spectral profiles of individual pixels and spectral unmixing, the relative contribution of ablated and unablated spectra to each pixel was then displayed. Results of spectral unmixing were compared to lesion pathology.ResultsRF ablation caused significant changes in the tissue autofluorescence profile. The magnitude of these spectral changes in human left atrium was relatively small (< 10% of peak fluorescence value), yet highly significant. Spectral unmixing of hyperspectral datasets enabled high spatial resolution, in-situ delineation of radiofrequency lesion boundaries without the need for exogenous markers. Lesion dimensions derived from hyperspectral imaging approach strongly correlated with histological outcomes. Presence of blood within the myocardium decreased the amplitude of the autofluorescence spectra while having minimal effect on their overall shapes. As a result, the ability of hyperspectral imaging to delineate ablation lesions in vivo was not affected.ConclusionsHyperspectral imaging greatly increases the contrast between ablated and unablated tissue enabling visualization of viability gaps at clinically relevant locations. Data supports the possibility for developing percutaneous hyperspectral catheters for high-resolution ablation guidance.