Autologous fat grafting procedures in plastic surgery have been extensively used to reinforce soft tissue in congenital or acquired tissue impairments. With this background, the aim of this study is firstly to examine the impact of a selective centrifugation on existing adipose stem cells (ASCs) in terms of stemness profile maintenance and, secondly, to investigate the effect of restoring volume in reconstruction on patients affected by soft tissue damage. After centrifugation, the fat graft products were separated into two layers and subsequently examined in vitro for the expression of CD34, CD90, CD117, CD105, CD29, CD31, CD44, CD73, CD133, CD14 and CD45 markers by flow cytometry and gene expression analyses were performed for Sox2, WNT3A, END, CD44, FUT4, COLL1, CTNNB1, hbEGF, KRTLG, MMP2 and VIM genes. The results showed that in the middle-high density (MHD) layer there was a peak concentration of ASCs, compared to another layer obtained after centrifugation. Research carried out on patients under treatment for soft tissue regeneration using cells obtained from MHD layer selection will be fundamental in comparative analysis. These studies will lead to an adequate standardization of outcomes, provided that treatment is performed through cell selection. Therefore, a unique procedure in tissue reconstruction and regeneration through fat grafting is presented here. Full article

The chemical composition and texture of titanium coatings can influence the growth characteristics of the adhered cells. An enhanced proliferation of the human mesenchymal stem cells (hMSCs) would be beneficial. The present study was aimed to investigate whether titanium deposited at different atmospheres would affect the cell growth properties, cellular morphology, and expression of surface markers of hMSCs. Titanium-based coatings were deposited on silicon wafers under oxygen, nitrogen, or argon atmospheres by ultra-short pulsed laser deposition using two different gas pressures followed by heating at 400 °C for 2 h. The characteristics of the coated surfaces were determined via contact angle, zeta potential, and scanning electron microscopy (SEM) techniques. Human MSCs were cultivated on differently coated silicon wafers for 48 h. Subsequently, the cell proliferation rates were analyzed with an MTT assay. The phenotype of hMSCs was checked via immunocytochemical stainings of MSC-associated markers CD73, CD90, and CD105, and the adhesion, spreading, and morphology of hMSCs on coated materials via SEM. The cell proliferation rates of the hMSCs were similar on all coated silicon wafers. The hMSCs retained the MSC phenotype by expressing MSC-associated markers and fibroblast-like morphology with cellular projections. Furthermore, no significant differences could be found in the size of the cells when cultured on all various coated surfaces. In conclusion, despite certain differences in the contact angles and the zeta potentials of various titanium-based coatings, no single coating markedly improved the growth characteristics of hMSCs. Full article

Autoimmune regulator (Aire) can promote the ectopic expression of peripheral tissue-restricted antigens (TRAs) in thymic medullary epithelial cells (mTECs), which leads to the deletion of autoreactive T cells and consequently prevents autoimmune diseases. However, the functions of Aire in the periphery, such as in dendritic cells (DCs), remain unclear. This study’s aim was to investigate the effect of Aire-overexpressing DCs (Aire cells) on the functions of CD4⁺ T cells and the treatment of type 1 diabetes (T1D). We demonstrated that Aire cells upregulated the mRNA levels of the tolerance-related molecules CD73, Lag3, and FR4 and the apoptosis of CD4⁺ T cells in STZ-T1D mouse-derived splenocytes. Furthermore, following insulin stimulation, Aire cells decreased the number of CD4⁺ IFN-γ⁺ T cells in both STZ-T1D and WT mouse-derived splenocytes and reduced the expression levels of TCR signaling molecules (Ca²⁺ and p-ERK) in CD4⁺ T cells. We observed that Aire cells-induced CD4⁺ T cells could delay the development of T1D. In summary, Aire-expressing DCs inhibited TCR signaling pathways and decreased the quantity of CD4⁺IFN-γ⁺ autoreactive T cells. These data suggest a mechanism for Aire in the maintenance of peripheral immune tolerance and provide a potential method to control autoimmunity by targeting Aire. Full article

Migration and chondrogenesis of human subchondral cortico-spongious progenitor cells (SPCs) are the key steps in the repair of microfracture-induced articular cartilage defects. The aim of this study was to evaluate the effect of human plasma-derived fibronectin (Fn) on the chondrogenic differentiation of SPCs, which was isolated from subchondrol cortico-spongious bone of late-stage osteoarthritis (OA) patients. SPCs were isolated and cultured for three passages. Stem cell surface antigens of SPCs were analyzed by flow cytometry. The osteogenic, chondrogenic and adipogenic differentiation potential were detected by histological staining. The chondrogenesis potential of SPCs with or without stimulation of either Fn or BMP-2 were studied by immunochemical staining and gene expression analysis. Cells isolated from subchondral bone presented to be positive for CD44, CD73, CD90, and CD166, and showed high capacity of osteogenic, adipogenic and chondrogenic differentiation, which suggested this cell population to be MSC-like cells. Stimulating with Fn increased the expression of SOX-9, aggrecan, collagen II while decreased the formation of collagen I by immunochemical staining. Gene expression analysis showed similar results. These results suggest that plasma-derived Fn can increase the chondrogenic differentiation of SPCs isolated from late-stage OA and improve cartilage repair after microfracture. Full article

Adenosine, an immunomodulatory biomolecule, is produced by the ecto-enzymes CD39 (nucleoside triphosphate dephosphorylase) and CD73 (ecto-5'-nucleotidase) by dephosphorylation of extracellular ATP. CD73 is expressed by many cell types during injury, infection and during steady-state conditions. Besides host cells, many bacteria also have CD39-CD73-like machinery, which helps the pathogen subvert the host inflammatory response. The major function for adenosine is anti-inflammatory, and most recent research has focused on adenosine’s control of inflammatory mechanisms underlying various autoimmune diseases (e.g., colitis, arthritis). Although adenosine generated through CD73 provides a feedback to control tissue damage mediated by a host immune response, it can also contribute to immunosuppression. Thus, inflammation can be a double-edged sword: it may harm the host but eventually helps by killing the invading pathogen. The role of adenosine in dampening inflammation has been an area of active research, but the relevance of the CD39/CD73-axis and adenosine receptor signaling in host defense against infection has received less attention. Here, we review our recent knowledge regarding CD73 expression during murine Salmonellosis and Helicobacter-induced gastric infection and its role in disease pathogenesis and bacterial persistence. We also explored a possible role for the CD73/adenosine pathway in regulating innate host defense function during infection. Full article

The use of amnion and amniotic fluid (AF) are abundant sources of mesenchymal stem cells (MSCs) that can be harvested at low cost and do not pose ethical conflicts. In human and veterinary research, stem cells derived from these tissues are promising candidates for disease treatment, specifically for their plasticity, their reduced immunogenicity, and high anti-inflammatory potential. This work aimed to obtain and characterize bovine amniotic fluid mesenchymal stem cells (AFMSC). The bovine AF from the amniotic cavity of pregnant gilts in the early stages of gestation (3- and 4-m-old bovine embryos) was collected. AFMSCs exhibit a fibroblastic-like morphology only starting from the fourth passage, being heterogeneous during the primary culture. Immunofluorescence results showed that AFMSCs were positive for β-integrin, CD44, CD73 and CD166, but negative for CD34, CD45. Meanwhile, AFMSCs expressed ES cell markers, such as Oct4, and when appropriately induced, are capable of differentiating into ectodermal and mesodermal lineages. This study reinforces the emerging importance of these cells as ideal tools in veterinary medicine; future studies aimed at a deeper evaluation of their immunological properties will allow a better understanding of their role in cellular therapy. Full article

Apoptosis and necrosis of intestinal epithelial cells (IECs), induced by ischemia-reperfusion (I/R) injury, can lead to dysfunction of the intestinal barrier, which could cause multiple organ dysfunction syndromes. Mesenchymal stem cells (MSCs) have the potential of providing protective effects on damaged IECs via paracrine action. This study investigated whether hypoxia can enhance the protective effect of placental-derived MSCs (pMSCs) on H₂O₂-treated-caco2 cells, and explored the possible mechanism. The pMSCs isolated by tissue culture were fibroblast-like, positive for CD73, CD90 and CD105 and can differentiate into chondrocytes and endothelial cells. Five days after treatment with H₂O₂, the numbers of living caco2 cells significantly decreased. More live H₂O₂-treated-caco2 cells were observed in pMSCs hypoxia culture medium (pMSCs-HCM) than pMSCs normoxia culture medium (pMSCs-NCM), and the application of a specific antibody that blocked insulin-like growth factor-1 (IGF-1) leads to a significant decrease of the protective effect of pMSCs-HCM. Hypoxia can promote IGF-1 expression of pMSCs at mRNA and protein levels, and caco2 stably expressed IGF-1 receptor. Knocking down IGF-1 expression in pMSCs by siRNA resulted in a significant attenuation of the increase in apoptosis of H₂O₂-treated-caco2 cultured in pMSCs-HCM. In conclusion, hypoxia can increase the protective effect of pMSCs on H₂O₂-treated-caco2 cells via a promotion of their paracrine actions, and the key cytokine involved is IGF-1. Full article