Assistant Professor University of Nebraska Omaha Omaha, Nebraska, United States
Introduction:: Elastin is the core protein of elastic fibers in the extracellular matrix in tissues such as the skin, lungs, and arteries, providing essential structural and mechanical support to these tissues. Elastic fiber production occurs during a short period in late fetal and early neonatal development. Elastic fibers have a remarkably stable half-life of around 70 years, and with aging, they experience degradation and fragmentation. In the human femoropopliteal artery (FPA), the main artery in the legs, elastic fibers are densely oriented longitudinally in the external elastic lamellae (EEL). Our team has investigated more than 1000 human FPAs and has observed long breaks in these elastic fibers that appear different than the common elastic fiber degradation and fragmentation that occurs with aging. In some samples, we have observed thin elastic fibers connecting the breaks. In this preliminary study, we aimed to examine these breaks and fibers, filling them in greater detail, and determine if they have characteristics of old or newly deposited elastic fibers.
Materials and Methods:: We obtained 10 human FPAs from organ donors (33±21 years; 50% male) within 24 hours postmortem after obtaining consent from their next of kin through an organ procurement organization, Live On Nebraska. The tissues were preserved in 0.9% phosphate-buffered saline at 4°C and transported to our laboratory. After dissection, 1-2 cm long axial strips were cut and fixed in methacarn for 48 hours, dehydrated in 70% ethanol, and embedded in paraffin. The specimens were then sectioned with a microtome and mounted on slides to prepare for histological staining. The samples were stained with Verhoeff-Van Gieson (VVG) and Movat’s Pentachrome to examine the extracellular matrix. Additionally, we stained the specimens with Hematoxylin and Eosin (H&E) to analyze the cell structures surrounding the intended region, as well as Periodic acid–Schiff (PAS) to examine the microfibrillar structures.
Results, Conclusions, and Discussions:: Most samples showed at least one break greater than 500 µm in length on the external elastic lamellae regardless of age or vascular health. Smooth muscles adjacent to the breaks often showed orientation, structure, and phenotype changes. Additionally, histology images showed that continuous elastic fibers around the breaks react negatively to Periodic acid–Schiff (PAS). However, when the breaks were filled with thin fibers, these fibers were frequently found to be PAS-positive.
Elastogenesis, i.e., the production and deposition of elastic fibers, is a hierarchical process that involves the formation and arrangement of the elastin core and the microfibrillar structures. Initially, the elastin core represents a small proportion of the fibers, but as the fibers mature, this ratio increases. Our results from PAS-stained histology sections show that the thin fibers filling the EEL breaks react positively to PAS and have a higher concentration of microfibrillar structures with heavy carbohydrates. This suggests that the thin fibers have characteristics of newly formed elastic fibers. In future studies, we will perform similar experiments on a larger sample size with a more comprehensive histology and immunohistochemistry analysis to better understand the breaks and the thin fibers that fill them.
