In vitro generation of transplantable insulin-producing cells from canine adipose-derived mesenchymal stem cells

Abstract

Canine mesenchymal stem cells (cMSCs) have potential applications for regenerative therapy, including the generation of insulin-producing cells (IPCs) for studying and treating diabetes. In this study, we established a useful protocol for generating IPCs from canine adipose mesenchymal stem cells (cAD-MSCs). Subsequently, in vitro preservation of pluronic F127-coated alginate (ALGPA)-encapsulated cAD-MSC-derived IPCs was performed to verify ready-to-use IPCs. IPCs were induced from cAD-MSCs with the modulated three-stepwise protocol. The first step of definitive endoderm (DE) induction showed that the cooperation of Chir99021 and Activin A created the effective production of Sox17-expressed DE cells. The second step for pancreatic endocrine (PE) progenitor induction from DE indicated that the treatment with taurine, retinoic acid, FGF2, EGF, TGFβ inhibitor, dorsomorphin, nicotinamide, and DAPT showed the significant upregulation of the pancreatic endocrine precursor markers Pdx1 and Ngn3. The last step of IPC production, the combination of taurine, nicotinamide, Glp-1, forskolin, PI3K inhibitor, and TGFβ inhibitor, yielded efficiently functional IPCs from PE precursors. Afterward, the maintenance of ALGPA-encapsulated cAD-MSC-derived IPCs with VSCBIC-1, a specialized medium, enhanced IPC properties. Conclusion, the modulated three-stepwise protocol generates the functional IPCs. Together, the encapsulation of cAD-MSC-derived IPCs and the cultivation with VSCBIC-1 enrich the maturation of generated IPCs.

Optimization of differentiation protocols of dental tissues stem cells to pancreatic β-cells

“…However, the differentiation efficiency and the maturation capacity of those generated cells, are still faced with challenges that need to be addressed. With the purpose of improving the differentiation protocol for adipose derived mesenchymal stem cells, Dang le et al, proposed a three step differentiation protocol through manipulating the culturing microenvironment [ 35 ]. Also, in an attempt to mimic 3D cultures, Ojaghi et al, reported that the application of PLLA/PVA scaffolds significantly enhanced the differentiation potential of adipose derived mesenchymal stem cells into IPCs [ 36 ].…”

Dissecting specific Wnt components governing osteogenic differentiation potential by human periodontal ligament stem cells through interleukin-6

“…The differentiation potential of hPDLSCs toward the osteogenic, adipogenic, and chondrogenic lineages was investigated in accordance with published protocols with slight modifications 32 , 69 , 86 , 87 . For osteogenic differentiation, cells were seeded into the wells of 24-well culture plates at 3 × 10 4 cells/well.…”

Influence of the Anatomical Site on Adipose Tissue-Derived Stromal Cells’ Biological Profile and Osteogenic Potential in Companion Animals

“…Numerous published studies have consistently indicated that ADSCs isolated from the subcutaneous region exhibit robust expression of CD90, CD44, CD29, and CD13, along with minimal or absent expression of CD54, CD34, CD45, MHC class II, CD11b, CD117, and CD3 ( Table 2 ), which is in line with recognized standards for human ADSCs [ 46 ]. Nonetheless, CD105 and CD73, also acknowledged MSC markers highly expressed in human ADSCs, present a controversial expression in canine ADSCs [ 45 , 56 ]. Different authors have demonstrated both positive and negative expressions of CD73 and CD105 [ 45 , 49 , 51 , 54 , 55 , 56 , 57 , 60 , 90 , 100 ].…”

“…Nonetheless, CD105 and CD73, also acknowledged MSC markers highly expressed in human ADSCs, present a controversial expression in canine ADSCs [ 45 , 56 ]. Different authors have demonstrated both positive and negative expressions of CD73 and CD105 [ 45 , 49 , 51 , 54 , 55 , 56 , 57 , 60 , 90 , 100 ]. CD73, an ecto-5’-nucleotidase, is attached to the outer plasma membrane by a glycosylphosphatidylinositol (GPI) anchor and acts as a signal and adhesion molecule to support cells’ anchoring [ 55 , 101 ].…”