PubMed · 14323488
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G IOANNIDES. 1965. APPENDIX EPICARDICA.. https://doi.org/10.1093/ajcp%2F44.2.160
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Decellularized tissues provide biological cues owing to the wealth of structural and regulatory factors that promote angiogenesis, adipogenesis, and myogenesis and facilitate neurite outgrowth. Here, we demonstrated the advantages of decellularized adipose tissue (adipoECM) over defined collagen-based biomaterials for host tissue integration. Three batches of human adipose tissue were decellularized using a rapid decellularization protocol and analyzed using mass spectrometry. To assess the biological activity of the decellularized materials, adipoECM and a reference standard of care biomaterial (Integra®DRT, also containing collagen I and glycosaminoglycans) were implanted subcutaneously, but far from the wound bed (in anatomical potential spaces) of immunocompetent BALB/c mice. The mice were euthanized in the acute (1 day) and chronic (day 60) inflammatory reaction phases, followed by biomaterial excision and Masson’s trichrome immunohistofluorescence imaging of the paraffin-embedded specimens. Each batch of processed tissue passed a quality control check, showing a low level of donor genomic DNA, lack of nuclei, lipids, endotoxins, and bacterial contamination. Mass spectrometry revealed that all batches of decellularized tissue mainly contained collagen I and, to a lesser degree, collagen III, collagen IV, collagen V, laminin, fibrillin, fibronectin, tenascin, and elastin. No acute inflammatory reaction was observed in either material one day post-transplantation. At 60 days post-implantation, different cell types were detected in adipoECM specimens, whereas Integra®DRT remained acellular. Additional immunohistochemical staining of adipoECM revealed CD31-positive cells in the blood vessels. Mesenchymal (CD90 positive) and myeloid (CD14 positive) cells were also detected. Primary cell types involved in soft tissue healing and remodeling were found in the adipoECM-treated group. The ingrowth of blood vessels and mesenchymal cells confirmed the effective integration of adipoECM with host tissues. Our results demonstrate that decellularized adipose tissue implanted away from the wound bed possesses contextual biological activities that promote efficient integration with host tissues.
We have cloned and functionally characterized an Na+-coupled citrate transporter from Caenorhabditis elegans (ceNAC-2). This transporter shows significant sequence homology to Drosophila Indy and the mammalian Na+-coupled citrate transporter NaCT (now known as NaC2). When heterologously expressed in a mammalian cell line or in Xenopus oocytes, the cloned ceNAC-2 mediates the Na+-coupled transport of various intermediates of the citric acid cycle. However, it transports the tricarboxylate citrate more efficiently than dicarboxylates such as succinate, a feature different from that of ceNAC-1 (formerly known as ceNaDC1) and ceNAC-3 (formerly known as ceNaDC2). The transport process is electrogenic, as evidenced from the substrate-induced inward currents in oocytes expressing the transporter under voltage-clamp conditions. Expression studies using a reporter-gene fusion method in transgenic C. elegans show that the gene is expressed in the intestinal tract, the organ responsible for not only the digestion and absorption of nutrients but also for the storage of energy in this organism. Functional knockdown of the transporter by RNAi (RNA interference) not only leads to a significant increase in life span, but also causes a significant decrease in body size and fat content. The substrates of ceNAC-2 play a critical role in metabolic energy production and in the biosynthesis of cholesterol and fatty acids. The present studies suggest that the knockdown of these metabolic functions by RNAi is linked to an extension of life span and a decrease in fat content and body size.
Better understanding of the mechanisms involved in adipose tissue growth and metabolism is critical for the development of more effective treatments for obesity. However, because of its high lipid and low protein content, adipose tissue can present unique problems in some experimental procedures. We describe three protocols that provide new or improved methods for analysis of DNA, RNA, and protein from different adipose tissues. The first protocol provides a simple and rapid method for separation of fragmented DNA and visualization of apoptotic DNA laddering without the need for radioisotopes. This technique allows for an estimate of the amount of DNA fragmentation, and hence, apoptosis. The second protocol details subcellular fractionation of adipose tissue for the extraction of protein in the mitochondrial and cytosol fractions and the measurement of apoptotic protein (Bcl-2 and Bax) levels in each fraction. The last protocol involves extraction of total RNA from adipose tissue and the measurement of uncoupling protein mRNA using real-time RT-PCR, a method that has not previously been used to measure expression of uncoupling proteins in adipose tissue.