Sung-Eun Yang

Mesenchymal stem cells feeder layer from human umbilical cord blood for ex vivo expanded growth and proliferation of hematopoietic progenitor cells.

Ex vivo expansion of hematopoietic stem cells was suggested as the best way of overcoming problems caused by limited hematopoietic cell number for cord blood transplantation. In this study, we quantified and characterized an ex vivo expansion capacity of umbilical cord blood (UCB)-derived mesenchymal stem cells (MSCs) as a cell feeder layer for support of UCB-derived committed hematopoietic progenitor cells (HPCs) in the absence or presence of recombinant cytokines. The UCB-derived MSCs used in the study differentiated into osteoblast, chondrocytes, and adipocytes under proper conditions. Frequencies in colony forming unit-granulocyte, macrophage, colony forming unit-granulocyte, erythrocyte, macrophage, megakaryocyte, burst forming unit-erythrocyte, and colony forming unit-erythrocyte increased to 3.46-, 9.85-, 3.64-, and 2.03-folds, respectively, only in culture supplemented by UCB-derived MSCs as a cell feeder layer without recombinant cytokines (culture condition C). Identified expansion kinetics in all kinds of committed HPCs showed plateaus at 7 culture days, suggesting some consumable components were required for the expansion. Physiological importance and different roles for different committed HPCs of UCB-derived MSCs as a cell feeder layer were revealed by a distinguished expansion capacity for colony forming unit-megakaryocyte. The preferred maintenance of CD33−CD34+ in culture condition C was also identified. The presence of cobblestone-like areas as hematopoietic microenvironment and various cell feeder layer-originated hematopoietic cytokines including interleukin-1β and granulocyte, macrophage-colony stimulating factor were suggested as underlying mechanisms for the identified expansion capacity. The present numeric and biological information about intrinsic expansion capacity for UCB-derived committed HPCs will increase further biological and clinical applications of UCB-derived MSCs.

Impact of Myocardial Infarct Proteins and Oscillating Pressure on the Differentiation of Mesenchymal Stem Cells : Effect of Acute Myocardial Infarction on Stem Cell Differentiation

Stem cell transplantation in acute myocardial infarction (AMI) has emerged as a promising therapeutic option. We evaluated the impact of AMI on mesenchymal stem cell (MSC) differentiation into cardiomyocyte lineage. Cord blood‐derived human MSCs were exposed to in vitro conditions simulating in vivo environments of the beating heart with acute ischemia, as follows: (a) myocardial proteins or serum obtained from sham‐operated rats, and (b) myocardial proteins or serum from AMI rats, with or without application of oscillating pressure. Expression of cardiac‐specific markers on MSCs was greatly induced by the infarcted myocardial proteins, compared with the normal proteins. It was also induced by application of oscillating pressure to MSCs. Treatment of MSCs with infarcted myocardial proteins and oscillating pressure greatly augmented expression of cardiac‐specific genes. Such expression was blocked by inhibitor of transforming growth factor β1 (TGF‐β1) or bone morphogenetic protein‐2 (BMP‐2). In vitro cellular and electrophysiologic experiments showed that these differentiated MSCs expressing cardiomyocyte‐specific markers were able to make a coupling with cardiomyocytes but not to selfbeat. The pathophysiologic significance of in vitro results was confirmed using the rat AMI model. The protein amount of TGF‐β1 and BMP‐2 in myocardium of AMI was significantly higher than that in normal myocardium. When MSCs were transplanted to the heart and analyzed 8 weeks later, they expressed cardiomyocyte‐specific markers, leading to improved cardiac function. These in vitro and in vivo results suggest that infarct‐related biological and physical factors in AMI induce commitment of MSCs to cardiomyocyte‐like cells through TGF‐β/BMP‐2 pathways.

Cotransplanted bone marrow derived mesenchymal stem cells (MSC) enhanced engraftment of hematopoietic stem cells in a MSC-dose dependent manner in NOD/SCID mice.

Transplantation of marrow-derived mesenchymal stem cells (MSCs), expanded by culture in addition to whole bone marrow, has been shown to enhance engraftment of human hematopoietic stem cells (HSCs). Our hypothesis was that there might be an optimum ratio range that could enhance engraftment. We examined the percent donor chimerism according to the ratio of HSCs to MSCs in non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice. We tested a series of ratios of co-transplanted CD34+-selected bone marrow cells, and marrow-derived MSCs into sublethally irradiated NOD/SCID mice. In all experiments, 1×105 bone marrow derived human CD34+ cells were administered to each mouse and human MSCs from different donors were infused concomitantly. We repeated the procedure three times and evaluated engraftment with flow cytometry four weeks after each transplantation. Serial ratios of HSCs to MSCs were 1:0, 1:1, 1:2 and 1:4, in the first experiment, 1:0, 1:1, 1:2, 1:4 and 1:8 in the second and 1:0, 1:1, 1:4, 1:8 and 1:16 in the third. Cotransplantation of HSCs and MSCs enhanced engraftment as the dose of MSCs increased. Our results suggest that the optimal ratio of HSCs and MSCs for cotransplantation might be in the range of 1:8-1:16; whereas, an excessive dose of MSCs might decrease engraftment efficiency.

Efficient intracytoplasmic labeling of human umbilical cord blood mesenchymal stromal cells with ferumoxides.

Mesenchymal stromal cells (MSCs) are multipotent cells found in several adult tissues; they have the capacity to differentiate into mesodermal, ectodermal, and endodermal tissues in vitro. There have been several reports that MSCs have therapeutic effects in a variety of diseases. Therefore, using a cell labeling technique, monitoring their temporal and spatial migration in vivo, would be useful in the clinical setting. Magnetic resonance imaging (MRI)—tracking of superparamagnetic iron oxide (SPIO)-labeled cells—is a noninvasive technique for determining the location and migration of transplanted cells. In the present study, we evaluated the influence and toxicity of SPIO (ferumoxides) labeling on multiple differentiated MSCs. To evaluate the influence and toxicity of ferumoxides labeling on differentiation of MSCs, a variety of concentrations of ferumoxides were used for labeling MSCs. We found that the cytoplasm of adherent cells was effectively labeled at low concentrations of ferumoxides. Compared with unlabeled controls, the ferumoxides-labeled MSCs exhibited a similar proliferation rate and apoptotic progression. The labeled MSCs differentiated into osteoblasts and adipocytes in an identical fashion as the unlabeled cells. However, chondrogenesis and neurogenesis were inhibited at high concentrations of ferumoxides. Our results suggest the effective concentration for ferumoxides use in tracking MSCs.

Distribution of haptoglobin phenotypes in a Korean population, using the semi-automated PhastSystem

We have established a new phenotyping method for haptoglobin, based on sodium dodecyl sulphate-polyacrylamide gel electrophoresis using the PhastSystem (Pharmacia Biotech, Uppsala, Sweden), followed by immunoblotting for detection. We measured haptoglobin concentrations and determined the haptoglobin phenotypes of 316 healthy Koreans using this method: 31 (9·8%) were of Hp 1-1 type, 140 (44·3%) of Hp 2-1 type and 145 (45·9%) of Hp 2-2 type. The haptoglobin allele frequencies were calculated to be 0·32 for Hp 1 and 0·68 for Hp 2. We were able to visualize up to 12 bands from the human Hp 2-2 polymeric series, with molecular weights in the range 171·9 x 103 to 802·2 x 103. The reference range of serum haptoglobin concentrations obtained by the IFCC (International Federation of Clinical Chemistry) standard method was 0·27 - 2·14 g/L. The serum haptoglobin concentration in Koreans was similar to that of Caucasians, but the Hp1 allele frequency was lower in Koreans. Our method could be used in clinical laboratories as a simple and practical method of haptoglobin phenotyping. In addition, the Hp 2-2 polymeric series could be used as high molecular weight standards.

Apolipoprotein E polymorphism and serum lipoprotein(a) concentrations in a Korean male population.

The purpose of this study was to investigate the effect of apolipoprotein E polymorphism on lipoprotein(a) metabolism by comparing serum lipoprotein(a) concentration with apolipoprotein E genotype in a Korean male population whose high molecular weight (HMW) lipoprotein(a) frequency was 95-98%. Serum lipoprotein(a), total cholesterol, triglyceride and high-density lipoprotein-cholesterol concentrations were measured and the apolipoprotein E genotype determined in 1189 healthy Korean males. The medians of serum lipoprotein(a) concentration in the apoE 2/3 group (0.105g/L) and the apoE 3/4 group (0·116g/L) were significantly lower than that in the apoE 3/3 group (0·155g/L; P<0·001). The medians of serum triglyceride were 1·497mmol/L in the apoE 2/3 group, 1·356mmol/L in the apoE 3/4 group, and 1·452mmol/L in the apoE 3/3 group (P<0·05). With the significant difference in the serum lipoprotein(a) concentration in Korean males according to apolipoprotein E genotype, and with the negative correlation between serum triglyceride concentration and serum lipoprotein(a) concentration, it is suggested that apolipoprotein E polymorphism and serum triglyceride participate in the metabolism of lipoprotein(a) with HMW.

A Qualitative Study on the Turnover Experiences of Teachers and Directors of Child Care Centers

This study is a qualitative research on job transfer experiences of child care teachers and directors of child care centers. The research was conducted on 21 participants: 10 child teachers and 11 directors of child care centers in Seoul and Gyeong-gi province. Focus group interviews were held at the subjects` child care centers in July 2011. Each focus group participated in one interview and the average length of these interviews was 2 hours. Four focus group interviews were recorded and transcribed. The gathered data was analyzed within and across groups according to focus group interview analysis methods. The analysis showed five categories in both the teacher group and the director group. Reorganizing this analysis revealed three phases: before, in the course of, and after the transfer. Further description about the job transfer was given for each phase of the two groups. This research is significantly meaningful in that it brings light to the experiences around job transfer for both groups at the same time.