Ex Vivo and in Vivo Administration of Fluorescent CNA35 Specifically Marks Cardiac Fibrosis

Abstract

Cardiac fibrosis is a major hallmark of cardiac diseases. For evaluation of cardiac fibrosis, the development of highly specific and preferably noninvasive methods is desired. Our aim was to evaluate CNA35, a protein known to specifically bind to collagen, as a specific marker of cardiac fibrosis. Fluorescently labeled CNA35 was applied ex vivo on tissue sections of fibrotic rat, mouse, and canine myocardium. After quantification of CNA35, sections were examined with picrosirius red (PSR) and compared to CNA35. Furthermore, fluorescently labeled CNA35 was administered in vivo in mice. Hearts were isolated, and CNA35 labeling was examined in tissue sections. Serial sections were histologically examined with PSR. Ex vivo application of CNA35 showed specific binding to collagen and a high correlation with PSR (Pearson r = .86 for mice/rats and r = .98 for canine; both p < .001). After in vivo administration, CNA35 labeling was observed around individual cardiomyocytes, indicating its ability to penetrate cardiac endothelium. High correlation was observed between CNA35 and PSR (r = .91, p < .001). CNA35 specifically binds to cardiac collagen and can cross the endothelial barrier. Therefore, labeled CNA35 is useful to specifically detect collagen both ex vivo and in vivo and potentially can be converted to a noninvasive method to detect cardiac fibrosis.

Construction of CNA35 Collagen-Targeted Phase-Changeable Nanoagents for Low-Intensity Focused Ultrasound-Triggered Ultrasound Molecular Imaging of Myocardial Fibrosis in Rabbits

“…As a molecular probe, the ability of CNA35 to target type I collagen in the myocardium has been demonstrated in basic research and animal studies. 36,41,42 The liquid fluorocarbon/phospholipid film-forming material is doped with a phospholipid carrying an amino group at the terminals. In the experimental procedure, we dissolved 1-ethyl-3- Recently, with the extensive application of the CEUS technique in clinical practice, ultrasound molecular imaging and therapeutic research have received increasing attention.…”

Local microRNA‐133a downregulation is associated with hypertrophy in the dyssynchronous heart

“…The volume percentage of collagen in cardiac tissue was histologically determined in tissue sections stained with 0.1% Sirius Red. 22…”

Multi-photon microscopy in cardiovascular research

“…Myocardial inflammation and ischemia can be considered precursors to clinical symptoms and complications which are intensely investigated in vivo at macroscopic resolution levels using non-invasive techniques [88]. Mechanistic studies on arrhythmia and inflammation are often conducted in vitro on cell cultures or ex vivo on tissue slices [89,90].…”

“…Mechanistic studies on arrhythmia and inflammation are often conducted in vitro on cell cultures or ex vivo on tissue slices [89,90]. Previously, TPLSM has been involved in some of these studies on ex vivo slabs of cardiac tissue [88,91,92] or ex vivo Langendorff-perfused hearts [93][94][95][96]. Even more so, intravital heart imaging with subcellular resolution has been very limited, but has been gaining interest due to technological advances in video rate microscopes, triggering systems, and innovative tissue preparation methods.…”

“…These studies exemplify that in vivo heart studies are indeed feasible and open new avenues for various research areas, such as cardiac fibrosis, angiogenesis, valve malfunction, and arrhythmia, also areas where fibrosis plays an important role and the possibility of collagen imaging comes into play [88,[101][102][103][104].…”