Characterization and Immunomodulatory Effects of Canine Adipose Tissue- and Bone Marrow-Derived Mesenchymal Stromal Cells

Abstract

BackgroundMesenchymal stromal cells (MSC) hold promise for both cell replacement and immune modulation strategies owing to their progenitor and non-progenitor functions, respectively. Characterization of MSC from different sources is an important and necessary step before clinical use of these cells is widely adopted. Little is known about the biology and function of canine MSC compared to their mouse or human counterparts. This knowledge-gap impedes development of canine evidence-based MSC technologies.Hypothesis and ObjectivesWe hypothesized that canine adipose tissue (AT) and bone marrow (BM) MSC (derived from the same dogs) will have similar differentiation and immune modulatory profiles. Our objectives were to evaluate progenitor and non-progenitor functions as well as other characteristics of AT- and BM-MSC including 1) proliferation rate, 2) cell surface marker expression, 3) DNA methylation levels, 4) potential for trilineage differentiation towards osteogenic, adipogenic, and chondrogenic cell fates, and 5) immunomodulatory potency in vitro.Results1) AT-MSC proliferated at more than double the rate of BM-MSC (population doubling times in days) for passage (P) 2, AT: 1.69, BM: 3.81; P3, AT: 1.80, BM: 4.06; P4, AT: 2.37, BM: 5.34; P5, AT: 3.20, BM: 7.21). 2) Canine MSC, regardless of source, strongly expressed cell surface markers MHC I, CD29, CD44, and CD90, and were negative for MHC II and CD45. They also showed moderate expression of CD8 and CD73 and mild expression of CD14. Minor differences were found in expression of CD4 and CD34. 3) Global DNA methylation levels were significantly lower in BM-MSC compared to AT-MSC. 4) Little difference was found between AT- and BM-MSC in their potential for adipogenesis and osteogenesis. Chondrogenesis was poor to absent for both sources in spite of adding varying levels of bone-morphogenic protein to our standard transforming growth factor (TGF-β3)-based induction medium. 5) Immunomodulatory capacity was equal regardless of cell source when tested in mitogen-stimulated lymphocyte reactions. Priming of MSC with pro-inflammatory factors interferon-gamma and/or tumour necrosis factor did not increase the lymphocyte suppressive properties of the MSC compared to untreated MSC.Conclusions/SignificanceNo significant differences were found between AT- and BM-MSC with regard to their immunophenotype, progenitor, and non-progenitor functions. Both MSC populations showed strong adipogenic and osteogenic potential and poor chondrogenic potential. Both significantly suppressed stimulated peripheral blood mononuclear cells. The most significant differences found were the higher isolation success and proliferation rate of AT-MSC, which could be realized as notable benefits of their use over BM-MSC.

Characterisation and In Vivo Safety of Canine Adipose-Derived Stem Cells

“…After five cell passages, they represented a phenotypically homogenous ASC population that was positive for typical MSC markers such as CD90, CD44 and CD73 but lacked the expression of CD34, CD45, CD14, and HLA class II. Comparable surface marker expression patterns of cASCs have been described previously (Vieira et al, 2010;Kisiel et al, 2012;Martinello et al, 2011;Reich et al, 2012;Russell et al, 2016;Takemitsu et al, 2012). Most of the previous studies have detected consistent expression of CD90 and CD44 in addition to other positive surface markers, but the expression of CD73 varies across the studies.…”

Immunomodulatory Effect of Adipose-Derived Stem Cells: The Cutting Edge of Clinical Application

“…Regarding the immunomodulatory capacity, it has been demonstrated that ASCs, as well as BMSCs, have a hypoimmunogenic phenotype, since they lack the major MHC class II molecules and express only low levels of MHC class I, this allowing them to evade immune recognition (Puissant et al, 2005;McIntosh et al, 2006). Furthermore, ASCs can act as modulators of the host response, showing a greater in vitro immunomodulatory ability than BMSCs derived from age-matched donors (Melief et al, 2013), since they are able to partially suppress lymphocytes proliferation, as well as to inhibit differentiation of monocyte-derived immature dendritic cells and NK cell cytotoxic activity (Russell et al, 2016;Valencia et al, 2016). Such effects are likely to depend on both cell contact-dependent mechanisms and paracrine effects through the production of cytokines and various soluble factors that regulate immune cells functions (Sotiropoulou et al, 2006), improve the microenvironment for tissue healing (Burlacu et al, 2013) and exert strong immunosuppressive effects by decreasing inflammatory cytokine production (Zhao et al, 2010).…”

The immunopathogenic and immunomodulatory effects of interleukin‐12 in periodontal disease

“…Russell et al . demonstrated the ability of MSCs obtained from adipose tissue and bone marrow to inhibit T‐cell proliferation in the absence of IFN‐ γ . Moreover, they showed that priming these MSCs with IFN‐ γ had no effect on MSC‐mediated T‐cell suppression .…”