In experimental models of pancreatic growth and recovery, changes in pancreatic size are assessed by euthanizing a large cohort of animals at varying time points and measuring organ mass. However, to ascertain this information in clinical practice, patients with pancreatic disorders routinely undergo non-invasive cross-sectional imaging of the pancreas using magnetic resonance imaging (MRI) or computed tomography (CT). The aim of the current study was to develop a thin-sliced, optimized sequence protocol using a high field MRI to accurately calculate pancreatic volumes in the most common experimental animal, the mouse. Using a 7 Telsa Bruker micro-MRI system, we performed abdominal imaging in whole-fixed mice in three standard planes: axial, sagittal, and coronal. The contour of the pancreas was traced using Vitrea software and then transformed into a 3-dimensional (3D) reconstruction, from which volumetric measurements were calculated. Images were optimized using heart perfusion-fixation, T1 sequence analysis, and 0.2 to 0.4 mm thick slices. As proof of principle, increases in pancreatic volume among mice of different ages correlated tightly with increasing body weight. In summary, this is the first study to measure pancreatic volumes in mice, using a high field 7 Tesla micro-MRI and a thin-sliced, optimized sequence protocol. We anticipate that micro-MRI will improve the ability to non-invasively quantify changes in pancreatic size and will dramatically reduce the number of animals required to serially assess pancreatic growth and recovery.
INTRODUCTION: In plastic and reconstructive surgery, surgeons are often challenged with a shortage of bone tissue at the site of bone defect. Human skeletal muscle-derived cells (skMDCs) undergo osteogenic differentiation in response to bone morphogenetic proteins (BMP) and have been explored for use in bone reconstructions.1,2 Before cell therapy can be considered for clinical use, delivery of cells and control of bone growth must be optimized. The aim of this study was to determine the effect of BMP2 on differentiation of skMDCs fabricated into 3D constructs, thereby validating this novel model of bone formation. 3D constructs provide a more accurate representation of in vivo setting than monolayer cultures and may be used in bone grafting without additional structural support. METHODS: skMDCs isolated from 20, 25, and 32 year-old male donors were purchased from Cook MyoSite and cultured in growth media. 1x10^5 skMDCs were fabricated into 3D constructs with extracellular matrix components using Flexcell Tissue Train System then cultured in osteogenic media with or without BMP2 (50ng/mL) for 3 days. RNA isolation and reverse transcription were then performed. Expression of marker genes of osteogenic and myogenic differentiation, Osx and MyoD, respectively, were evaluated via qPCR. They were normalized to GAPDH, and fold differences of BMP2 groups over corresponding controls were computed. The data was analyzed using ANOVA and Tukey’s test with P<0.05 considered statistically significant. RESULTS: All skMDCs groups treated with BMP2 showed statistically significant difference in both Osx and MyoD expression when compared to corresponding groups not treated with BMP2. Osx expression was increased with BMP2 treatment with mean fold difference over control ± standard deviation of 5.12±1.01 for 20 yo, 2.85±0.10 for 25 yo, and 3.67±1.09 for 32 yo (Figure 1). MyoD expression was decreased with BMP2 treatment with mean fold difference over control ± standard deviation of 0.61±0.06 for 20 yo, 0.51±0.10 for 25 yo, and 0.25±0.11 for 32 yo (Figure 2).Figure 1: Osx Expression.Figure 2: MyoD Expression.CONCLUSION: The results indicate that BMP2 promotes osteogenic differentiation of skMDCs in 3D constructs as evidenced by increased Osx expression and decreased MyoD expression in groups treated with BMP2. Human skMDCs fabricated into 3D constructs showed an osteogenic response to BMP2 comparable to other models and can therefore serve as an excellent in vitro model of bone formation to identify factors that promote osteogenesis.
We investigate cardiovascular (CV) developmental physiology and biomechanics in order to understand the dramatic acquisition of form and function during normal development and to identify the adaptive mechanisms that allow embryos to survive adverse genetic and epigenetic events. Cardiovascular patterning, morphogenesis, and growth occur via highly conserved genetic mechanisms. Structural and functional maturation of the embryonic heart is also conserved across a broad range of species with evidence for load dependence from onset of the heartbeat. The embryonic heart dynamically adapts to changes in biomechanical loading conditions and for reasons not yet clear, adapts better to increased than to decreased mechanical load. In mammals, maternal cardiovascular function dynamically impacts embryonic/fetal growth and hemodynamics and these interactions can now be studied longitudinally using high-resolution noninvasive techniques. Maternal exposure to hypoxia and to bioactive chemicals, such as caffeine, can rapidly impact embryonic/fetal cardiovascular function, growth, and outcome. Finally, tissue engineering approaches can be applied to investigate basic developmental aspects of the embryonic myocardium. We use isolated embryonic and fetal chick, mouse, or rat cardiac cells to generate 3D engineered early embryonic cardiac tissues (EEECT). EEECT retains the morphologic and proliferative features of embryonic myocardium, responds to increased mechanical load with myocyte hyperplasia, and may be an excellent future material for use in cardiac repair and regeneration. These insights into cardiovascular embryogenesis are relevant to identifying mechanisms for congenital cardiovascular malformations and for developing cell- and tissue-based strategies for myocardial repair.
