• Dog
• FACS
• Liver
• Mesenchymal stromal cells
• qPCR
• RT-PCR
• Stem Cells

Within the field of Regenerative Medicine stem cells hold great potential for treating currently incurable diseases. One of these stem cells of interest is the Mesenchymal Stromal Cell (MSC). Major sources for MSCs are adipose tissue and bone-marrow although recently it has been proven that MSCs can also be isolated from different tissues such as liver (Pan Q, 2011), pancreas (Zanini C, 2011), muscles (Kisiel AH, 2011) etc. MSCs, with their well-known anti-inflammatory activity, are ideal for treatment of acute or fulminant liver diseases for which the only treatment option is liver transplantation. Moreover this multitude of source opportunities makes it possible to perform autologous treatment of patients with minimally invasive approaches. Due to the low number of available liver transplants, alternative (curative) treatments are necessary. Although the use of MSCs in mouse models seems promising, the lack of a large animal model limits the application of MSCs in a human clinical setting. For years the liver research group at the CSCA (Clinical Sciences for Companion Animals, Utrecht, The Netherland) has been investigating the dog as a large model animal. This research showed that canine liver diseases and regenerative mechanisms are highly comparable to their human counterparts. Up to now possibilities of treatments for liver diseases in the dog are limited, for this reason MSCs hold a great potential as a possible population for cell therapies. Prerequisite for the use of MSCs is their characterisation before clinical application. For this we started with the optimization of a protocol for the isolation of LMSCs, starting either from a wedge liver biopsy or from an intra-lobular perfused fraction. The focus of the second part was the characterisation of canine MSCs derived from adipose tissue (ASCs), bone-marrow (BMSCs), and liver (LMSCs). All cell types are in culture at the department of CSCA and multiple samples have been stored (liquid nitrogen). Characterization of the MSCs has been performed according to Dominici et al. (Horwitz EM & Therapy., 2005). The characterization started with gene-expression profiling of the different fractions (ASCs, BMSCs, and LMSCs). Canine specific primers have been designed and optimized and specificity tested by sequencing of the PCR product. Results indicated that all MSC fractions are positive for standard MSC markers (e.g. CD90, CD105) and negative for markers of haematopoietic cells, macrophages, endothelial cells (e.g. CD45). After this a selection of canine specific antibodies directed against Cluster of Differentiation (CD) markers (CD45-, CD90+, CD105+, CD29+, CD166+) has been optimised for Fluorescent Activated Cell Sorting (FACS) analysis. Finally the resulting LMSCs profile, for both qPCR and FACS analysis, has been compared to the human MSC profile (Fouraschen SM, Secreted factors of human liver-derived mesenchymal stem cells promote liver regeneration early after partial hepatectomy., 2012). This research will be the basis on which clinically applied mesenchymal stromal cells can become a reality within the veterinary field. This will, in turn, provide the necessary information to apply these cells in human clinics.