Up to 2.8 × 10(7) fibroblast-like cells displaying an abundant presence of mesenchymal stem cell (MSC) markers CD73, CD90, CD105 and a low level of HLA-I expression can be isolated from one whole human umbilical cord (UC) using a simple and highly reproducible explant culture approach. Cells derived from whole UC, similar to cells collected from separate compartments of UC, display a distinct chondrogenic and adipogenic potential. Therefore they are potential candidates for cartilage and adipose tissue engineering. Cell differentiation along the osteogenic pathway is, however, less efficient, even after the addition of 1.25-dihydroxyvitamin D3, a potent osteoinductive substance. Isolated cells are highly proliferative, tolerate cryopreservation with an average survival rate of about 75% and after thawing can be propagated further, at least over 20 population doublings before their proliferative activity begins to decline. More importantly, they synthesize numerous trophic factors including neurotrophins and factors which facilitate angiogenesis and hematopoiesis. In conclusion, cells isolated from whole UC satisfies all requirements essential for the generation of stem cell banks containing permanently available cell material for applications in the field of regenerative medicine. Nevertheless, further studies are needed to improve and adjust the methods which are already employed for adult MSC expansion and differentiation to specific properties and requirements of the primitive stem cells collected from UC. So, our data verify that the choice of individual parameters for cell propagation, such as duration of cell expansion and cell seeding density, has a substantial impact on the quality of UC-derived cell populations.

Background: A variety of cell types can be identified in the adherent fraction of bone marrow mononuclear cells including more primitive and embryonic-like stem cells, mesenchymal stem cells (MSC), lineage-committed progenitors as well as mature cells such as osteoblasts and fibroblasts. Different methods are described for the isolation of single bone marrow stem cell subpopulations -beginning from ordinary size sieving, long term cultivation under specific conditions to FACSbased approaches. Besides bone marrow-derived subpopulations, also other tissues including human umbilical cord (UC) have been recently suggested to provide a potential source for MSC. Although of clinical importance, these UC-derived MSC populations remain to be characterized. It was thus the aim of the present study to identify possible subpopulations in cultures of MSC-like cells obtained from UC. We used counterflow centrifugal elutriation (CCE) as a novel strategy to successfully address this question.

First isolated from bone marrow, mesenchymal stem or stromal cells (MSC) were shown to be present in several postnatal and extraembryonic tissues as well as in a large variety of fetal tissues (e.g., fatty tissue, dental pulp, placenta, umbilical cord blood, and tissue). In this study, an optimized protocol for the expansion of MSC-like cells from whole umbilical cord tissue under xeno-free culture conditions is proposed. Different fetal calf sera and human serum (HS) were compared with regard to cell proliferation and MSC marker stability in long-term expansion experiments, and HS was shown to support optimal growth conditions. Additionally, the optimal concentration of HS during the cultivation was determined. With regard to cell proliferative potential, apoptosis, colony-forming unit fibroblast frequency, and cell senescence, our findings suggest that an efficient expansion of the cells is carried out best in media supplemented with 10% HS. Under our given xeno-free culture conditions, MSC-like cells were found to display in vitro immunoprivileged and immunomodulatory properties, which were assessed by co-culture and transwell culture experiments with carboxyfluorescein diacetate succinimidyl ester-labeled peripheral blood mononuclear cells. These findings may be of great value for the establishment of biotechnological protocols for the delivery of sufficient cell numbers of high quality for regenerative medicine purposes.

Mesenchymal stem or stromal cells (MSCs) have a high potential for cell-based therapies as well as for tissue engineering applications. Since Friedenstein first isolated stem or precursor cells from the human bone marrow (BM) stroma that were capable of osteogenesis, BM is currently the most common source for MSCs. However, BM presents several disadvantages, namely low frequency of MSCs, high donor-dependent variations in quality, and painful invasive intervention. Thus, tremendous research efforts have been observed during recent years to find alternative sources for MSCs.In this context, the human umbilical cord (UC) has gained more and more attention. Since the UC is discarded after birth, the cells are easily accessible without ethical concerns. This postnatal organ was found to be rich in primitive stromal cells showing typical characteristics of bone-marrow MSCs (BMSCs), e.g., they grow as plastic-adherent cells with a fibroblastic morphology, express a set of typical surface markers, and can be directly differentiated at least along mesodermal lineages. Compared to BM, the UC tissue bears a higher frequency of stromal cells with a higher in vitro expansion potential. Furthermore, immune-privileged and immune-modulatory properties are reported for UC-derived cells, which open highly interesting perspectives for clinical applications.

BackgroundCD19 is a B cell lineage specific surface receptor whose broad expression, from pro-B cells to early plasma cells, makes it an attractive target for the immunotherapy of B cell malignancies. In this study we present the generation of a novel humanized anti-CD19 monoclonal antibody (mAb), GBR 401, and investigate its therapeutic potential on human B cell malignancies.MethodsGBR 401 was partially defucosylated in order to enhance its cytotoxic function. We analyzed the in vitro depleting effects of GBR 401 against B cell lines and primary malignant B cells from patients in the presence or in absence of purified NK cells isolated from healthy donors. In vivo, the antibody dependent cellular cytotoxicity (ADCC) efficacy of GBR 401 was assessed in a B cell depletion model consisting of SCID mice injected with healthy human donor PBMC, and a malignant B cell depletion model where SCID mice are xenografted with both primary human B-CLL tumors and heterologous human NK cells. Furthermore, the anti-tumor activity of GBR 401 was also evaluated in a xenochimeric mouse model of human Burkitt lymphoma using mice xenografted intravenously with Raji cells. Pharmacological inhibition tests were used to characterize the mechanism of the cell death induced by GBR 401.ResultsGBR 401 exerts a potent in vitro and in vivo cytotoxic activity against primary samples from patients representing various B-cell malignancies. GBR 401 elicits a markedly higher level of ADCC on primary malignant B cells when compared to fucosylated similar mAb and to Rituximab, the current anti-CD20 mAb standard immunotherapeutic treatment for B cell malignancies, showing killing at 500 times lower concentrations. Of interest, GBR 401 also exhibits a potent direct killing effect in different malignant B cell lines that involves homotypic aggregation mediated by actin relocalization.ConclusionThese results contribute to consolidate clinical interest in developing GBR 401 for treatment of hematopoietic B cell malignancies, particularly for patients refractory to anti-CD20 mAb therapies.