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Background & Aim Background Stem cell therapy has been considered as a potential strategy in regenerative for the patient with neurological deficits. Mesenchymal Stem/Stromal Cells (MSC) have been extensively investigated as a treatment in stroke. However, the MSC lost its potency for neural repair in aged patients due to the limited numbers of Neural Stem Cell (NSC). NSC is essential for neurogenesis of the injured brain. In vitro studies have shown that MSCs which were cultured in the laboratory has the capability to transform into different cells. Adding stimulants such as growth factors into the culture media could induce the trans-differentiation of MSC into NSC. However, to produce NSC in the clinical setting on a large scale is still a challenge. Objective In this study, we like to evaluate the use of epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), heregulin B27, and N2 the trans-differentiation mechanism of umbilical cord MSC (UC-MSC) into NSC. Methods, Results & Conclusion Method MSC was transferred into a 24-well plate at 2 × 104 cells per well. Induction of NSC is done using DMEM containing N2/B27 supplement, 20 ng/ml epidermal growth factor (EGF), and 10 ng/ml basic fibroblast growth factor (bFGF). Identification of success trans-differentiation is analyzed using NeuroFlour CDr3 antibody. Result and Discussion MSC transdifferentiates into NSC as an important source of cell in therapy for neurological diseases. UC-MSC was induced using N2 to helps the cell to commit to differentiation and its survival. B27 is used for the long term survival of neurons cells and helps the NSC to grow and proliferates. bFGF beliefs as pre-inductive molecules that enhance neuronal differentiation and self-renewal. Upon bFGF stimulation, NGFβR and NRP1 will upregulate and promotes differentiation. On the contrary, EGF induces differentiation at lower concentrations. EGF is used for inducing the expression of the neuron and glial marker. To confirm the successful transdifferentiating process, the expression of CDr3 in the cell culture is analyzed. CDr3 positive result as the membrane-permeable fluorescent probe was shown in Figure 1. NeuroFluor CDr3 binds to human fatty acid-binding protein 7 (FABP7) which use to confirm the neural induction from human stem cells. Conclusion UC-MSC could be cultured and differentiate into NSC in vitro when supplemented with N2/B27, bFGF, and EGF, which may produce NSC in the clinical setting on a large scale.

Background & Aim Umbilical cord tissue is a potential source of Mesenchymal Stem Cells (MSCs). These cells have been widely used especially in pre-clinical and clinical applications resulting its various effectiveness. Banking process of umbilical cord involves transportation from the hospital where the cord is received to the banking facility to be processed further. Prior to processing, pre-sterility test must be conducted and while waiting for the result, the umbilical cord is quarantined. This quarantine process has to be carried out carefully to maintain cell's recovery and viability. Suitable range of time should be chosen to deliver high quality cells. In this study, we aim to determine the maximum time for quarantined umbilical cord to be processed. Methods, Results & Conclusion One umbilical cord (UC2018030001) was used in this study. Medium transportation is used to carry the umbilical cord to banking facility. Prior to processing, pre sterility was done using spread plate method by spreading the medium transport onto TSA agar. After the results were out and passed, the tissues were put into quarantine medium and were processed at day 0, 7, 14, and 21 for MSCs isolation. The tissue was processed using explant method and sub-cultured until 3rd passage. Graphic shown below are the cell doubling level from different passages at various days of quarantine. The doubling level remain constant with maximum deviation at 0.512. Results showed that tissues which was quarantined until day 21 still produce cells normally. The cell number are slowly increasing because the higher the passage, the more pure the cells are so they can propagate easily. This shows normal growth of normal cells. Their doubling level are also on track with no significant difference. In brief, umbilical cord tissue can be quarantined until 21 days and still produce cells normally.

Background & Aim Coronary artery disease (CAD) is one of the leading cause of death in the world. Currently, the common therapy for this disease is to use drugs and surgical procedures however the result has been less optimum. Therefore, it is necessary to develop an alternative treatment using Mesenchymal Stem Cell (MSC) and Conditioned Medium (CM). Methods, Results & Conclusion MSCs were obtained from the umbilical cord of eligible donor and process in a cGMP-standardized laboratory certified by the Indonesian Ministry of Health. A 52-year-old subject who experienced Coronary Artery Disease was treated with PCI four years before the stem cell therapy. The subject signed an informed consent for stem cell therapy before the injection. Allogeneic UC-MSC for CAD was packed in a syringe containing 20 ml NaCl 0.9% with cell concentration of 20 × 106. Umbilical Cord-Mesenchymal Stem Cells were administered intravenously and conducted three times over 4 months period. Meanwhile, Conditioned Medium (CM) was applied intranasally with a dose of 0.4 cc/day for a week after each UC-MSCs administrated. Assessment with Magnetic Resonance Imaging (MRI) shows improvement on the Left Ventricular Ejection Fraction (LVEF) from 55% at baseline and 75% at 3 weeks after first stem cell transplantation. Endothelial Cell Progenitor (EPC) count show decreasing the number of EPC cells from 11,893 cells/mL into 4,932 cells/mL. This shows the angiogenesis process takes place after the UC-MSC injection. On the twelve weeks after the first injection, the subject undergo spirometry test and shows improvement in the vital capacity (85% into 94%), forced capacity (85% into 94%), and forced expansion volume (93% into 103%). There are no adverse events occur and the patient experience improvement in physic, MRI, laboratory and spirometry assessment.

Background & Aim Background: Type 2 diabetes mellitus (T2DM) is a metabolic disease caused by insulin disruption, insulin secretion or both. Hyperglycemia occurs in diabetes, where long-term hyperglycemia causes an increase in oxidative stress, a decrease in antioxidants and an increase in pro-inflammatory factors that contribute to endothelial dysfunction. In these conditions where microenvironment is not right, the amount of available ATP is not adequate, causing a decrease in the number and potential of Endothelial Progenitor Cells (EPC) in playing its function and role in the decreased colonization process. In this preliminary study, the aim was to prove changes in microenvironment by adding growth media to improve the condition of the microenvironment (niche) in an in-vitro able to increase the number and potential of EPC Methods, Results & Conclusion Method: Circulating EPC counts were calculated using a combination of expressions from CD34+ and CD133+ antigen surfaces using BD FACS Canto II flow cytometry, and the potential of EPC was done by adding ALDH bright staining. Then the EPC was isolated and cultured on the growth medium, then cultured for seven days. After that, the number of EPC circulating and the EPC potential is recalculated Result The results of this study indicate that after the addition of growth medium and incubated for seven days, the number of EPCs increased by 413.20%, and 1,401.1% in the controlled and uncontrolled group. Also, the potential of EPC showed by express of ALDHbright was obtained at the end of incubation of 99.9%, in each group in diabetes. Discussion This study illustrates that by adding a growth medium, it can restore microenvironment conditions to be better characterised by the ability of EPC to express bright ALDH which increases very high. Bright ALDH activity shows the ability to re-populating and differentiation and illustrates the operation of retinoids that have a pro-angiogenic effect with modulation of endothelial cell cytokines, the impact of migration and vasculogenesis. Conclusion The addition of growth medium in EPC culture from peripheral blood improves microenviral conditions and can increase the amount of EPC and the potential of EPC associated with its role in the colonisation function.

Background & Aim Umbilical Cord derived Mesenchymal Stem Cells (UC-MSCs) and Stromal Vascular Fraction (SVF) has been widely used in preclinical and clinical applications, with varying degrees of effectiveness (Menasche, 2011). Due to freshness condition and different cell doses for therapy of each patient, the laboratory have to provide cell product while maintaining the optimal quality of the cells. The effects of storage conditions such as storage solutions, temperature, and duration of cell storage greatly influence the cell quality for transplant. The quality of cells directly can be evaluated from the cell viability rapid measurement. High cell viability shows that proportion of live cells is higher than dead cells, while low cell viability indicates that many cells have begun to be critical or even death. Therefore, our research aims to evaluate of several mediums and temperatures storage to maintain stability of UC-MSCs and SVF viability for transplantation. Methods, Results & Conclusion Both of UC-MSCs and SVF were divided into 4 groups with two variables temperature and medium storage. Cells have been evaluated in room temperature (RT) (18-22°C) and cold temperature (2-8°C) with 0.9% NaCl and conditioned medium (UC-MSCs) or autologous serum (SVF). The viability from each group was assessed day by day using Tryphan Blue. At room temperature, viability of cells in both storage mediums ware significantly decreased while stored at cold temperature the viability was more stable. The highest viability of UC-MSCs at RT was obtained from cells in 0.9% NaCl solution (77% viable). However, the highest UC-MSCs viability at cold temperature was obtained from cell storaged in 0.9% NaCl solution (93% viable). Data shown, for SVF viability from day 0 (when SVF was produced) to day 6, cells are most stable in 0.9% NaCl and autologous serum suspension. Both of them which stored at cold temperature stable on 90.4% viability. SVF in 0.9% NaCl at room temperature has a less stable viability where cell viability decrease from 96% at first day until 77% at sixth day. Cold temperature (2-8°C) can maintain stability of UC-MSCs and SVF cell viability. The most stable of UC-MSCs viability was obtained from cell storage in 0.9% NaCl solution and SVF in autologous serum.

Background & Aim Adipose tissue provides an abundant source of stromal vascular fraction (SVF), which resides a potential regenerative medicine named mesenchymal stem cells (MSC). Storage period and temperature before isolation may impact the quality of isolated cells. Here, we identify the effect of storage periods and temperatures to SVF viability and cell number along with the MSC recovery. Methods, Results & Conclusion A total of 70 ml donor-adipose tissue was divided into two different groups, room temperature (RT) and 4-8°C. SVF was isolated using manual method. Each group was process after stored for 4, 5, 6, 7, and 12 days. After it passed each incubation time, adipose samples were processed enzymatically until SVF was obtained. The viability of SVF stored at RT was decreasing until 40%, but relatively constant when stored at 4-8°C (>75%). Nevertheless, the sample stored at RT revealed to have better proliferation and colonization than sample stored at 4-8°C. The sample retained the proliferation capability when stored at RT up to 7 days, but only up to 5 days when stored at 4-8°C. The average of SVF number was higher in the sample stored at RT (8,552,315 cell/mL) compared to the sample at 4-8°C (4,581,111 cell/mL). The highest SVF number was observed in sample stored at 4 days before isolation. The average of MSC recovery was also higher at RT compared to sample stored at 4-8°C, which was 6% and 3%, respectively. In conclusion, adipose tissue is preferable to be stored at RT before isolation for higher SVF total number and MSC recovery. The adipose tissue should be processed immadiately or stored only up to 4 days after sample collection.