Supplements to support clinical-grade cultures of mesenchymal stem cells (MSC) are required to promote growth and expansion of these cells. Platelet lysate (PL) is a human blood component which may replace animal serum in MSC cultures being rich in various growth factors. Here, we describe a plasma poor pathogen-free platelet lysate obtained by pooling 12 platelet (PLT) units, to produce a standardized and safe supplement for clinical-grade expansion of MSC.

C-type lectin receptors (CLRs) on antigen-presenting cells (APCs) facilitate uptake of carbohydrate antigens for antigen presentation, modulating the immune response in infection, homeostasis, autoimmunity, allergy, and cancer. In this review, we focus on the role of the macrophage galactose type C-type lectin (MGL) in the immune response against self-antigens, pathogens, and tumor associated antigens (TAA). MGL is a CLR exclusively expressed by dendritic cells (DCs) and activated macrophages (MØs), able to recognize terminal GalNAc residues, including the sialylated and nonsialylated Tn antigens. We discuss the effects on DC function induced throughout the engagement of MGL, highlighting the importance of the antigen structure in the modulation of immune response. Indeed modifying Tn-density, the length, and steric structure of the Tn-antigens can result in generating immunogens that can efficiently bind to MGL, strongly activate DCs, mimic the effects of a danger signal, and achieve an efficient presentation in HLA classes I and II compartments.

Background: Mesenchymal stromal cells (MSCs) have been largely investigated, in the past decade, as potential therapeutic strategies for various acute and chronic pathological conditions. MSCs isolated from different sources, such as bone marrow (BM), umbilical cord tissue (UCT) and adipose tissue (AT), share many biological features, although they may show some differences on cumulative yield, proliferative ability and differentiation potential. The standardization of MSCs growth and their functional amplification is a mandatory objective of cell therapies. The aim of this study was to evaluate the cumulative yield and the ex vivo amplification potential of MSCs obtained from various sources and different subjects, using defined culture conditions with a standardized platelet lysate (PL) as growth stimulus. Methods: MSCs isolated from BM, UCT and AT and expanded in human PL were compared in terms of cumulative yield and growth potential per gram of starting tissue. MSCs morphology, phenotype, differentiation potential, and immunomodulatory properties were also investigated to evaluate their biological characteristics. Results: The use of standardized PL-based culture conditions resulted in a very low variability of MSC growth. Our data showed that AT has the greater capacity to generate MSC per gram of initial tissue, compared to BM and UCT. However, UCT-MSCs replicated faster than AT-MSCs and BM-MSCs, revealing a greater proliferation capacity of this source irrespective of its lower MSC yield. All MSCs exhibited the typical MSC phenotype and the ability to differentiate into all mesodermal lineages, while BM-MSCs showed the most prominent immunosuppressive effect in vitro. Conclusions: The adoption of standardized culture conditions may help researchers and clinicians to reveal particular characteristics and inter-individual variability of MSCs sourced from different tissues. These data will be beneficial to set the standards for tissue collection and MSCs clinical-scale expansion both for cell banking and for cell-based therapy settings.

Circulating CD34⁺ cell populations characterized by a low rate (up to five) or high rate (more than five) of cell divisions were isolated from 8 d cultures in the presence of stem cell factor (SCF), interleukin‐3 (IL‐3), granulocyte‐macrophage colony stimulating factor (GM‐CSF), granulocyte colony stimulating factor (G‐CSF), erythropoietin (EPO), Flt3 ligand and Peg‐rHu megakaryocyte growth and development factor (Peg‐rHuMGDF) using the fluorescent dye 5,6‐carboxyfluorescein diacetate succinimidyl ester (CFDA‐SE) and flow cytometric cell sorting. Phenotypic characterization of cells which had experienced up to five divisions (CFDA‐SE^bright) showed a similar surface antigen expression to starting, freshly isolated CD34⁺ cells. Conversely, cells which experienced more than five divisions (CFDA‐SE^dim) showed a differentiating behaviour, down‐regulating CD34 antigen and acquiring differentiation markers. CFDA‐SE^bright cells were significantly enriched in CD105 (endoglin) positive precursors as compared to both freshly isolated CD34⁺ and CFDA‐SE^dim cells. Functional analysis indicated that CFDA‐SE^bright had a 3‐fold and 10‐fold greater cumulative cloning efficiency as compared to freshly isolated CD34⁺ cells and CFDA‐SE^dim cells, respectively. CFDA‐SE^bright cells retained the vast majority of LTC‐IC and showed a LTC‐IC frequency 2.8‐fold higher than that found in freshly isolated CD34⁺ cells. RT‐PCR and Western blot analyses showed significantly higher bcl‐2 RNA and protein levels in CFDA‐SE^bright cells as compared to freshly isolated CD34⁺ and CFDA‐SE^dim cells. This study indicates that cytokine low‐responding circulating CD34⁺ cells (CFDA‐SE^bright cells) represent a functionally, phenotypically and molecularly distinct multipotent progenitor population with biological properties associated with primitive precursors.