SYNOPSIS. Naturally occurring populations of the monarch butterfly, Danaus plexippus (L.) and the Florida queen butterfly D. gilippus berenice Cramer were found infected with Ophryocystis elektroscirrha sp. n. This neogregarine pathogen infects the hypodermal tissue, remains in micronclear schizogony until after pupation of the host, and then rapidly completes morphogenesis in the tissue that becomes the scales of the adult butterfly. The adult thus carries the spores externally; no internal infection was detected. The pathogen is unique in the genus Ophryocystis because: other species have all been reported from the Malphigian tubules of Coleoptera; no pseudopodial attachments of schizonts to host tissue were seen; the merozoites were motile; the cystic membrane surrounding the zygote was tenuous, and the sporont developed with no apparent membrane other than the spore wall: and the spore wall appeared amber in transmitted light when morphogenesis was completed.
Recently, we have developed an optimal decellularization protocol to generate 3D porcine myocardial scaffolds, which preserved natural extracellular matrix structure, mechanical anisotropy, and vasculature templates, and also showed good cell recellularization and differentiation potential. In this study, a multi-stimulation bioreactor was built to provide coordinated mechanical and electrical stimulations for facilitating stem cell differentiation and cardiac construct development. The acellular myocardial scaffolds were seeded with mesenchymal stem cells (106 cells/ml) by needle injection and subjected to 5-azacytidine treatment (3 μmol/L, 24 h) and various bioreactor conditioning protocols. We found that, after 2-day culture with mechanical (20% strain) and electrical stimulation (5 V, 1 Hz), high cell density and good cell viability were observed in the reseeded scaffold. Immunofluorescence staining demonstrated that the differentiated cells showed cardiomyocyte-like phenotype, by expressing sarcomeric α-actinin, myosin heavy chain, cardiac troponin T, connexin-43, and N-cadherin. Biaxial mechanical testing demonstrated that positive tissue remodeling took place after 2-day bioreactor conditioning (20% strain + 5 V, 1 Hz); passive mechanical properties of the 2-day and 4-day tissue constructs were comparable to the tissue constructs produced by stirring reseeding followed by 2-week static culture, implying the effectiveness and efficiency of the coordinated simulations in promoting tissue remodeling. In short, the synergistic stimulations might be beneficial not only for the quality of cardiac construct development, but also for patients by reducing the waiting time in future clinical scenarios.
These data support the hypothesis that chitosan-coatings are able to develop a close bony apposition or the osseointegration of dental/craniofacial and orthopedic implants.
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