Bone marrow mesenchymal stem cells (BM-MSCs) are multipotent stem cells that can differentiate into some kinds of mesenchymal cells such as osteoblasts, chondroblasts and adipocytes. These cells were discovered for a long time and considered as the first discovered source of MSCs. BM-MSC transplantation was used to treat some diseases such as bone disease, myocardial infarction, stroke, diabetes mellitus… This study aimed to provide a new method to in vitro primarily culture and secondary culture of BM-MSCs that were compliant with good manufacturing practice for clinical applications. Bone marrow was aspirated from the hip bone using special needles and syringes. Mononuclear cells (MNCs) were isolated from bone marrow by BM-MSC extraction kit. These MNCs were cultured in the commercial medium – MSC-Cult supplemented with 10% activated platelet rich plasma (aPRP) to obtain BM-MSCs. The results showed that 100% samples of bone marrow can be successfully cultured to produce BM-MSCs. Obtained cells exhibited the MSC phenotypes, and maintained the stable and normal karyotype up to 10th passages, and failed to cause tumor in the mouse models. This research provided an advanced protocol for clinical applications of BM-MSCs.

Bone marrow mesenchymal stem cells; Clinical application; GMP; Mesenchymal stem cells; Stem cell culture

Human umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) are a promising stem cell source with the potential to modulate the immune system as well as the capacity to differentiate into osteoblasts, chondrocytes, and adipocytes. In previous publications, UCB-MSCs have been successfully differentiated into cardiomyocytes. This study aimed to improve the efficacy of differentiation of UCB-MSCs into cardiomyocytes by combining 5-azacytidine (Aza) with mouse fetal heart extract (HE) in the induction medium. UCB-MSCs were isolated from umbilical cord blood according to a published protocol. Murine fetal hearts were used to produce fetal HE using a rapid freeze-thaw procedure. MSCs at the 3rd to 5th passage were differentiated into cardiomyocytes in two kinds of induction medium: complete culture medium plus Aza (Aza group) and complete culture medium plus Aza and fetal HE (Aza + HE group). The results showed that the cells in both kinds of induction medium exhibited the phenotype of cardiomyocytes. At the transcriptional level, the cells expressed a number of cardiac muscle-specific genes such as Nkx2.5, Gata 4, Mef2c, HCN2, hBNP, α-Ca, cTnT, Desmin, and β-MHC on day 27 in the Aza group and on day 18 in the Aza + HE group. At the translational level, sarcomic α-actin was expressed on day 27 in the Aza group and day 18 in the Aza + HE group. Although they expressed specific genes and proteins of cardiac muscle cells, the induced cells in both groups did not contract and beat spontaneously. These properties are similar to properties of heart muscle precursor cells in vivo. These results demonstrated that the fetal HE facilitates the differentiation process of human UCB-MSCs into heart muscle precursor cells.

Adipose-derived stem cells (ADSCs) are excellent for regenerative medicine. Like mesenchymal stem cells, ADSCs possess multi-potent differentiation capacity that enables them to differentiate into osteoblasts, chondrocytes and adipocytes, as well as trans-differentiation into other cells. ADSC transplantation has gained attention in recent years, especially in vitro expanded ADSC transplantation. This study aimed to provide a new method to in vitro primarily culture and secondary culture of ADSCs that were compliant with good manufacturing practice for clinical applications. Stromal vascular fraction (SVF) was extracted from adipose tissue by commercial kits. SVF was expanded in vitro in medium with non-allogeneic supplements. Cultured ADSCs maintained immune-phenotype, karyotype, and differentiation potential after ten passages. Moreover, ADSCs at 15th passage could not form tumors in NOD/SCID mice. This research produced a suitable protocol for clinical applications of expanded ADSCs.

Adipose derived stem cells; Stem cell transplantation; ADSC transplantation; ADSC application; Stem cell culture

Currently, immune cell-based therapies, particularly those that utilize dendritic cells (DCs), are a promising therapy approach for cancer treatment. Therefore, DC therapy is the focus of many studies in many laboratories worldwide that are developing novel cancer therapies. This study aimed to develop a reproducible procedure to produce functional DCs from mouse bone marrow for DC therapy research. Bone marrow was collected from mouse femur bones by flushing with buffered saline. These cells were used to isolate mononuclear cells (MNCs) by Ficoll gradient centrifugation. MNCs were cultured in RPMI 1640 medium supplemented with 20 ng/mL of IL-4 and 20 ng/mL of GMCSF to induce maturation of immature DCs. The results showed that this procedure induced cells exhibiting the DC phenotypes, such as the expression of CD40, CD80, and CD86, high phagocytic capacity, strong production of IL-12, and efficient stimulation of T-CD4 lymphocytes. These results suggest that this procedure can be used to produce functional DCs in future studies that use DCs for immune therapy.

Breast cancer stem cells, Dendritic cells, IL-12 mRNA, Immune therapy, Tumor lysates primed DCs.

Authors Pham PV, Nguyen NT, Nguyen HM, Khuat LT, Le PM, Pham VQ, Nguyen ST, Phan NK

Published Date August 2014 Volume 2014:7 Pages 1455—1464

DOI http://dx.doi.org/10.2147/OTT.S67057

Received 30 April 2014, Accepted 9 June 2014, Published 18 August 2014

Type 1 diabetes mellitus is characterized by the destruction of pancreatic islet beta cells, which leads to insulin insufficiency, hyperglycemia, and reduced metabolic glucose level. Insulin replacement is the current standard therapy for type 1 diabetes mellitus but has several limitations. Pancreatic islet transplantation can result in the production of exogenous insulin, but its use is limited by immune-rejection and donor availability. Recent studies have shown that mesenchymal stem cells (MSCs) can trans-differentiate into insulin-producing cells (IPCs), which could be utilized for diabetes mellitus treatment. Previously published reports have demonstrated that MSC or IPC transplantation could produce significant improvement in mouse models of diabetes mellitus. This study was aimed at determining the effects of two different methods of MSC transplantation on the efficacy of diabetes mellitus treatment in mouse models. The MSCs were isolated from umbilical cord blood and were proliferated following a previously published procedure. Diabetes mellitus was induced in mice by streptozotocin (STZ) injection. Thirty days after transplantation, the weight of the mice treated by intra-venous infusion and intra-pancreatic injection was found to be 22% and 14% higher than that of the un-treated mice. The blood glucose concentrations in both intra-venous infusion and intra-pancreatic injection groups decreased and remained more stable than those in the control group. Moreover, insulin was detected in the serum of the treated mice, and the pancreas also showed gradual recovery. Based on the results of this preliminary investigation, intra-venous infusion seems more suitable than intra-pancreatic injection for MSC transplantation for diabetes mellitus treatment.

 

Type 1 diabetes mellitus; umbilical cord blood; mesenchymal stem cells; intravenous transplantation; intra-pancreatic injection

The ability of dendritic cells to efficiently present tumor-derived antigens when primed with tumor cell lysates makes them attractive as an approach for cancer treatment. This study aimed to evaluate the effects of dendritic cell transfusion dose on breast cancer tumor growth in a murine model. Dendritic cells were produced from allogeneic bone marrow-derived mononuclear cells that were cultured in RPMI 1640 medium supplemented with 20 ng/mL GMCSF and 20 ng/mL IL-4 for 7 days. These cells were checked for maturation before being primed with a cancer cell-derived antigen. Cancer cell antigens were produced by a rapid freeze-thaw procedure using a 4T1 cell line. Immature dendritic cells were loaded with 4T1 cell-derived antigens. Dendritic cells were transfused into mice bearing tumors at three different doses, included 5.10⁴, 10⁵, and 10⁶ cells/mouse with a control consisting of RPMI 1640 media alone. The results showed that dendritic cell therapy inhibited breast cancer tumors in a murine model; however, this effect depended on dendritic cell dose. After 17 days, in the treated groups, tumor size decreased by 43%, 50%, and 87.5% for the doses of 5 × 10⁴, 10⁵, and 10⁶ dendritic cells, respectively, while tumor size in the control group decreased by 44%. This result demonstrated that dendritic cell therapy is a promising therapy for breast cancer treatment.

4T1 cells; Breast cancer; Dendritic cells; Immune cells based therapy; Immunotherapy; Targeting treatment