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Cardiovascular diseases, including myocardial infarction (MI), persist as the leading cause of mortality and morbidity worldwide. The limited regenerative capacity of the myocardium presents significant challenges specifically for the treatment of MI and, subsequently, heart failure (HF). Traditional therapeutic approaches mainly rely on limiting the induced damage or the stress on the remaining viable myocardium through pharmacological regulation of remodeling mechanisms, rather than replacement or regeneration of the injured tissue. The emerging alternative regenerative medicine-based approaches have focused on restoring the damaged myocardial tissue with newly engineered functional and bioinspired tissue units. Cardiac regenerative medicine approaches can be broadly categorized into three groups: cell-based therapies, scaffold-based cardiac tissue engineering, and scaffold-free cardiac tissue engineering. Despite significant advancements, however, the clinical translation of these approaches has been critically hindered by two key obstacles for successful structural and functional replacement of the damaged myocardium, namely: poor engraftment of engineered tissue into the damaged cardiac muscle and weak electromechanical coupling of transplanted cells with the native tissue. To that end, the integration of micro- and nanoscale technologies along with recent advancements in stem cell technologies have opened new avenues for engineering of structurally mature and highly functional scaffold-based (SB-CMTs) and scaffold-free cardiac microtissues (SF-CMTs) with enhanced cellular organization and electromechanical coupling for the treatment of MI and HF. In this review article, we will present the state-of-the-art approaches and recent advancements in the engineering of SF-CMTs for myocardial repair.
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Management of autofluorescence in formaldehyde-fixed myocardium: choosing the right treatment
Autofluorescence (AF) poses challenges for detecting proteins of interest in situ when employing immunofluorescence (IF) microscopy. This interference is particularly pronounced in strongly autofluorescent tissues such as myocardium, where tissue AF can be comparable to IF. Although various histochemical methods have been developed to achieve effective AF suppression in different types of tissue, their applications on myocardial samples have not been well validated. Due to inconsistency across different autofluorescent structures in sometypes of tissue, it is unclear if these methods can effectively suppress AF across all autofluorescent structures within the myocardium. Here, we quantitatively evaluated the performance of several commonly used quenching treatments on formaldehyde-fixed myocardial samples, including 0.3 M glycine, 0.3% Sudan Black B (SBB), 0.1% and 1% sodium borohydride (NaBH4), TrueVIEW® and TrueBlack®. We further assessed their quenching performance by employing the pre-treatment and post-treatment protocols, designed to cover two common IF staining scenarios where buffers contained detergents or not. The results suggest that SBB and TrueBlack® outperform other reagents in AF suppression on formaldehyde-fixed myocardial samples in both protocols. Furthermore, we inspected the quenching performance of SBB and TrueBlack® on major autofluorescent myocardial structures and evaluated their influence on IF imaging. The results suggest that SBB outperforms TrueBlack® in quenching major autofluorescent structures, while TrueBlack® excels in preserving IF labeling signal. Surprisingly, we found the treatment of NaBH4 increased AF signal and enhanced the AF contrast of major autofluorescent structures. This finding suggests that NaBH4 has the potential to act as an AF enhancer and may facilitate the interpretation of myocardial structures without the need for counterstaining.
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- Award ID(s):
- 1655740
- PAR ID:
- 10479395
- Publisher / Repository:
- PAGEPress
- Date Published:
- Journal Name:
- European Journal of Histochemistry
- Volume:
- 67
- Issue:
- 4
- ISSN:
- 1121-760X
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.4081/ejh.2023.3812
