<p>Characterization and Immunomodulation of Canine Amniotic Membrane Stem Cells</p>

Abstract

Purpose: Amniotic membrane stem cells have a high capacity of proliferation, cell expansion, and plasticity, as well as immunomodulatory properties that contribute to maternal-fetal tolerance. Owing to the lack of research on human amniotic membrane at different gestational stages, the canine model is considered ideal because of its genetic and physiological similarities. We aimed to characterize the canine amniotic membrane (CAM) cell lineage in different gestational stages and evaluate the expression of immunomodulatory genes. Materials and Methods: Twenty CAMs from early (20-30 days) (n=7), mid-(31-45 days) (n=7), and late gestation (46-63 days) (n=6) stages were studied. The cell features were assessed by cell viability tests, growth curve, colony-forming units, in vitro differentiation, cell labeling for different immunophenotypes, and pluripotent potential markers. The cells were subjected to RT-PCR and qPCR analysis to determine the expression of IDO, HGF, EGF, PGE2, and IL-10 genes. Results: CAM cells exhibited a fibroblastoid morphology and adherence to plastic with an average cell viability of 78.5%. The growth curve indicated a growth peak in the second passage and we obtained an average of 138.2 colonies. Osteogenic, chondrogenic, and adipogenic lineages were confirmed by in vitro differentiation assays. Cellular immunophenotyping experiments confirmed the presence of positive mesenchymal markers (CD90 and CD105) and the low or negative expression of hematopoietic markers (CD45 and CD34). Qualitative analysis of the immunomodulatory functions indicated the expression of the IDO, HGF, EGF5, and PGE2 genes. When stimulated by interferon-gamma, CAM cells exhibited higher IDO levels throughout gestation. Conclusion: The CAMs from different gestational stages presented features consistent with mesenchymal stem cell lineage; better results were observed during the late gestation stage. Therefore, the gestational stage is a key factor that may influence the functionality of therapies when using fetal membrane tissues from different periods of pregnancy.

Biological characteristics and metabolic profile of canine mesenchymal stem cells isolated from adipose tissue and umbilical cord matrix

“…Colony formation was similar in both canine cell cultures. However, the mean number of colonies obtained in this study for AT and UC-MSCs is lower when compared with what found for canine amniotic membrane derived MSCs using the same protocol [ 23 ]. It is likely that different cells and culture medium influenced the results.…”

“…found for canine amniotic membrane derived MSCs using the same protocol [23]. It is likely that different cells and culture medium influenced the results.…”

“…To assess colony formation ability, 1x10 4 cells at P1 were seeded in a 90 mm petri dish and cultured for 13 days [ 23 ]. For each sample three replicates were carried out.…”

Conditioned Medium from Canine Amniotic Membrane-Derived Mesenchymal Stem Cells Improved Dog Sperm Post-Thaw Quality-Related Parameters

“…They are non-teratogenic [ 55 ], pluripotent, and also have regenerative properties and low immunogenicity [ 31 , 34 , 55 ]. Canine MSCs derived from the amniotic membrane have recently been isolated and proved to have the same characteristics, biosafety, and properties as other MSCs, which make them a good candidate for clinical trials [ 32 , 56 ]. The composition and potential use of cAMSC-CM have not been studied yet, but since CM contains cell paracrine factors [ 37 ], the application of cAMSC-CM in clinical studies seems to be promising.…”

Current Status on Canine Foetal Fluid and Adnexa Derived Mesenchymal Stem Cells

“…Since, interest in this tissue as a source of MSCs has also developed in dogs. As shown in Table 1 , canine AMMSCs expressed embryonic and MSCs markers, such as OCT4, CD44, CD184, and CD29 and could differentiate into neurocytes, osteocytes, adipocytes and chondrocytes based on cell morphology, specific stains, and molecular analysis [ 40 , 48 , 49 , 50 , 51 , 52 ]. Regarding the immunomodulatory properties of AMMSCs, Borghesi et al [ 51 ] found low MHC-I expression and no MHC-II expression, giving the MSCs the potential to escape recognition by CD4+T cells [ 55 ].…”