Surface glycan pattern of canine, equine, and ovine bone marrow-derived mesenchymal stem cells.

The use of bone marrow-derived mesenchymal stem cells (MSCs) for clinical and experimental studies is increasing, but full characterization of MSCs in veterinary species is hindered by the variability in species-specific cell surface marker expression and antibody cross reactivity. Recent studies demonstrated that the glycans in the glycocalyx of MSCs are promising candidates as cell biomarkers. In the present study, we analyzed the glycocalyx of canine MSCs (cMSCs), ovine MSCs (oMSCs), and equine MSCs (eMSCs) using a cell microarray procedure in which MSCs were spotted on microarray slides and incubated with a panel of 14 biotinylated lectins and Cy3-conjugated streptavidin. The signal intensity was then detected using a microarray scanner. The lectin-binding signals indicated that the MSC surface of the investigated species contained both N- and O-linked glycan types, with N-glycosylation predominating over O-glycosylation and fucosylation being more abundant than sialylation. Relative quantification revealed an interspecific difference between these glycans. In addition, cMSCs expressed more α2,3-linked sialic acid (MAL II), terminal lactosamine (RCA120 ), and α1,6 and α1,3 fucosylated oligosaccharides (PSA, LTA); oMSCs exhibited more T antigen (Jacalin), GalNAcα1,3(LFucα1,2)Galß1,3/4GlcNAcß1 (DBA), chitotriose (succinylated WGA), and α1,2-linked fucose (UEA I); and eMSCs showed a higher density of α2,6 sialic acids (SNA) and high mannose N-glycans (Con A). Using cell microarray methodology, we have for the first time demonstrated differences in the glycosylation profiles of cMSC, oMSC, and eMSC surfaces. These results could be valuable as resources and references for MSC differentiation and molecular remodeling in clinical cell-based therapy and tissue engineering studies. © 2017 International Society for Advancement of Cytometry.

Probiotic supplementation affects the glycan composition of mucins secreted by Brunner's glands of the pig duodenum.

The effect of a dietary probiotic blend on the carbohydrate composition of mucins secreted by the Brunner's glands in the duodenum of growing-finishing pigs was investigated by means of conventional (periodic acid-Schiff, Alcian Blue pH 2.5, high iron diamine staining) and lectin (15 lectins) histochemistry. Pigs were assigned to two dietary treatments: a control basal diet without the probiotic blend (No-Pro) and a test diet that included the probiotic blend (Pro). Duodenal tissue fragments were fixed in 4% phosphate-buffered-saline-buffered paraformaldehyde, dehydrated through a graded alcohol series, and embedded in paraffin wax. The secretory cells of the Brunner's glands from No-Pro pigs primarily produced neutral glycoproteins and a small amount of acidic non-sulphated mucins. This glycan pattern was opposite that of the Brunner's glands from Pro animals. A comparison of lectin-binding profiles of the secretory cells of Brunner's glands in these two groups showed that in Pro pigs, there was (i) a decrease in N-linked glycans containing α1,2-linked fucose (Con A, UEA I); (ii) a loss of complex types of N-glycans (PHA-L, PHA-E) terminating with lactosamine (RCA120), α1,6- and α1,3-linked fucose (LTA), and α-galactose (GSA I-B4), as well as of O-glycans with terminal Galß1,3GalNAc (PNA); and (iii) an increase in O-glycans containing GalNAc HPA. No-Pro and Pro samples showed no change in the expression of α2,6 sialoglycans and terminal GlcNAc residues and no affinity for MAL II, DBA, and SBA. These results indicate that probiotic supplementation affects the glycan composition of mucins produced in the Brunner's glands of growing-finishing pigs. These changes could effectively act on the gastrointestinal function and health status of these animals because the probiotic blend induced higher growth performance and meat quality in the test probiotic group than it did in the control basal diet group (Tufarelli et al., 2017).

Ultrastructural characteristics of ovine bone marrow-derived mesenchymal stromal cells cultured with a silicon stabilized tricalcium phosphate bioceramic.

Bioceramics are being used in experimental bone engineering application in association with bone marrow derived mesenchymal stem cells (BM-MSCs) as a new therapeutic tool, but their effects on the ultrastructure of BM-MSCs are yet unknown. In this study we report the morphological features of ovine (o)BM-MSCs cultured with Skelite, a resorbable bioceramic based on silicon stabilized tricalcium phosphate (SiTCP), able to promote the repair of induced bone defect in sheep model. oBM-MSCs were isolated from the iliac crest, cultured until they reached near-confluence and incubated with SiTCP. After 48 hr the monolayers were highly damaged and only few cells adhered to the plastic. Thus, SiTCP was removed, and after washing the cells were cultured until they became confluent. Then, they were trypsinizated and processed for transmission electron microscopy (TEM) and RT-PCR analysis. RT-PCR displayed that oBM-MSCs express typical surface marker for MSCs. TEM revealed the presence of electron-lucent cells and electron-dense cells, both expressing the CD90 surface antigen. The prominent feature of electron-lucent cells was the concentration of cytoplasmic organelles around the nucleus as well as large surface blebs containing glycogen or profiles of endoplasmic reticulum. The dark cells had a multilocular appearance by the presence of peripheral vacuoles. Some dark cells contained endocytic vesicles, lysosomes, and glycogen aggregates. oBM-MSCs showed different types of specialized interconnections. The comparison with ultrastructural features of untreated oBM-MSCs suggests the light and dark cells are two distinct cell types which were differently affected by SiTCP bioceramic. Skelite cultured ovine BM-MSCs display electron-dense and electron-lucent cells which are differently affected by this bioceramic. This suggests that they could play a different role in bioceramic based therapy.