Stemness Evaluation of Mesenchymal Stem Cells from Placentas According to Developmental Stage: Comparison to Those from Adult Bone Marrow

This study was done to evaluate the stemness of human mesenchymal stem cells (hMSCs) derived from placenta according to the development stage and to compare the results to those from adult bone marrow (BM). Based on the source of hMSCs, three groups were defined: group I included term placentas, group II included first-trimester placentas, and group III included adult BM samples. The stemness was evaluated by the proliferation capacity, immunophenotypic expression, mesoderm differentiation, expression of pluripotency markers including telomerase activity. The cumulative population doubling, indicating the proliferation capacity, was significantly higher in group II (P<0.001, 31.7+/-5.8 vs. 15.7+/-6.2 with group I, 9.2+/-4.9 with group III). The pattern of immunophenotypic expression and mesoderm differentiation into adipocytes and osteocytes were similar in all three groups. The expression of pluripotency markers including ALP, SSEA-4, TRA-1-60, TRA-1-81, Oct-4, and telomerase were strongly positive in group II, but very faint positive in the other groups. In conclusions, hMSCs from placentas have different characteristics according to their developmental stage and express mesenchymal stemness potentials similar to those from adult human BMs.

The effectiveness of ethylene glycol as cryoprotectant in mouse embryo freezing with slow freezing method / 대한산부인과학회지

OBJECTIVE: We intended to know how the cryoprotectant ethylene glycol (EG) would affect the outcome of the embryo development when used in slow freezing method. And to know if there is any difference in the outcome of frozen-thawed embryos according to freezing methods and the timing. METHODS: We used 5-6 weeks old ICR female mice and T6 containing 0.4% BSA for basic culture media. The embryos at the developmental stages of 1-cell, 8-cell and blastocyst were cryopreserved respectively by slow freezing method using EG, propylene glycol (PROH), and glycerol as a cryoprotectant. We also compared the results of slow freezing and vitrification methods with the same cryoprotectant, EG. And finally, we evaluated the quality of blastocysts by counting the cell numbers in each group. RESULTS: The post-thaw embryo development were better in EG group when they were frozen at 1-cell and blastocyst stage (P<0.05). Although there were no differences in the recovery rate, the survival rate in vitrification group was significantly higher (P<0.05). Post-thaw embryo development to morula and blastocyst were better in vitrification group when frozen at 1-cell embryo (P<0.05), not at 8-cell and blastocyst group. The cell counts of blastocyst derived from 1-cell stage frozen EG group were significantly increased than that of PROH-glycerol groups (P<0.05), however, there was no difference between the two freezing methods. CONCLUSION: These results suggest that EG may be advantageous comparing with the conventional cryoprotectants, PROH and glycerol in slow freezing method for mouse embryo cryopreservation. In terms of freezing method, vitrification is better than slow freezing.

Hematopoietic Differentiation of Embryoid Bodies Derived from the Human Embryonic Stem Cell Line SNUhES3 in Co-culture with Human Bone Marrow Stromal Cells

Human embryonic stem (ES) cells can be induced to differentiate into hematopoietic precursor cells via two methods: the formation of embryoid bodies (EBs) and co-culture with mouse bone marrow (BM) stromal cells. In this study, the above two methods have been combined by co-culture of human ES-cell-derived EBs with human BM stromal cells. The efficacy of this method was compared with that using EB formation alone. The undifferentiated human ES cell line SNUhES3 was allowed to form EBs for two days, then EBs were induced to differentiate in the presence of a different serum concentration (EB and EB/high FBS group), or co- cultured with human BM stromal cells (EB/BM co-culture group). Flow cytometry and hematopoietic colony-forming assays were used to assess hematopoietic differentiation in the three groups. While no significant increase of CD34+/CD45- or CD34+/CD38- cells was noted in the three groups on days 3 and 5, the percentage of CD34+/CD45- cells and CD34+/ CD38- cells was significantly higher in the EB/BM co-culture group than in the EB and EB/high FBS groups on day 10. The number of colony-forming cells (CFCs) was increased in the EB/BM co-culture group on days 7 and 10, implying a possible role for human BM stromal cells in supporting hematopoietic differentiation from human ES cell-derived EBs. These results demonstrate that co-culture of human ES-cell-derived EBs with human BM stromal cells might lead to more efficient hematopoietic differentiation from human ES cells cultured alone. Further study is warranted to evaluate the underlying mechanism.

Patient-Controlled Sedation versus Nurse-Administered Sedation with Propofol during Colonoscopy / 대한소화기내시경학회지

BACKGROUND/AIMS: Patient-controlled sedation (PCS) allows the patients to titrate the dosages of sedative drug according to their needs. The objective of this study was to compare the safety and the efficacy of nurse-administered propofol sedation (NAPS) with those of PCS. METHODS: Eighty one patients were randomly assigned to two groups. All patients received meperidine 25 mg and propofol 40 mg as an initial dose for sedation. Patients in PCS group were subsequently infused with propofol 15 mg over 80 seconds through infusion pump whenever they required. Patients in NAPS group were injected with 10~20 mg propofol by nurse with supervision by endoscopist. The dosage of propofol, cardiopulmonary parameters, procedure time, sedation score, pain score, the patients' and endoscopists' satisfaction scores were assessed. RESULTS: With regard to blood pressure, pulse rate and oxygen saturation, serious complications were not observed. Especially, there was no significant difference of mean total dose between two groups (NAPS group and PCS group received 76.7+/-24.7 mg and 82.5+/-26.6 mg respectively). Pain score was higher in woman than in man (p=0.03). CONCLUSIONS: 1.2~1.5 mg/kg of propofol with small dose of opioid during colonoscopy was effective and safe. NAPS was more practical and useful method of sedation than PCS during colonoscopy.

Comparative study on the development of frozen-thawed mouse embryo using vitrification at each developmental stage / 대한산부인과학회잡지

OBJECTIVE: To find out the optimal freezing time in terms of developmental stage of mouse embryo and the effectiveness of vitrification method for freezing them. METHODS: Superovulation induction was performed using PMSG and hCG. Total 1,437 mouse embryos (vitrification group: 743, slow-freezing group: 694) were obtained and cultured with the T6 containing 0.4% BSA medium. Each developmental stage of embryos (1-, 2-, 4-, 8-cell, morula and blastocyst) were cryopreserved by vitrification and also by slow freezing method for comparison of the results. After thawing, the recovery rate, the survival rate and the blastocyst developmental rate were analysed and compared in two different settings. Student's t-test was used for statistical analysis. RESULTS: The survival and developmental rates at all subgroups of vitrification method are significantly higher than those of slow-freezing groups, but not in the recovery rates. In vitrification group, the survival rate and the blastocyst developmental rate are highest when frozen at morula stage, 98.4% and 86.4%, respectively. In slow-freezing group, the survival rate is also highest when frozen at morula stage, 87.2%, and the blastocyst developmental rate is highest when frozen at 8-cell stage, 78.1%. CONCLUSION: The vitrification method is more efficient for mouse embryo freezing compared with slow freezing one. Among various developmental stages of mouse embryos, morula stage seems to be the optimal stage for cryopreservation, whatever the freezing method applied. Therefore, we recommend embryo freezing at morula stage by vitrification method.

Effect of Intrathecal Clonidine in Hyperbaric Bupivacaine Spinal Anesthesia / 대한마취과학회지

BACKGROUND: Vasoconstrictors have been used as an adjunct to local anesthetics to prolong the duration of spinal anesthesia. Recently, clonidine, an 2-receptor agonist has been shown to prolong the duration of spinal anesthesia following intrathecal administration. Bupivacaine has been used for spinal anesthesia and compared with tetracaine in recent studies. We have undertaken this study to further evaluate the effect of clonidine in hyperbaric 0.5% bupivacaine spinal anesthesia. METHODS: Thirty patients who were scheduled for lower limb or urologic operation were divided into 2 groups: Group A (hyperbaric bupivacaine 13 mg, 2.6 ml + N/S 1 ml), Group B (hyperbaric bupivacaine 13 mg, 2.6 ml + clonidine 150 g, 1 ml). We used standardized techniques and injected above drugs to group A and B intrathecally for spinal anesthesia. We investigated the onset and the duration of spinal anesthesia along with hemodynamic changes (blood pressure and heart rate) in patients. RESULTS: There were no significant differences in the onset of spinal anesthesia and hemodynamic changes between two groups. The time taken to recover from the nerve block was more prolonged in the group B (touch 225, pain 262, foot dorsiflexion 271, knee flexion 290 minutes) than group A (touch 154, pain 188, foot dorsiflexion 198, knee flexion 216 minutes). There were no significant differences in sedation, and in experiencing dry mouth and other side effects between two groups. CONCLUSION: Intrathecal clonidine 150 g has been proved to prolong the duration of hyperbaric 0.5% bupivacaine spinal anesthesia without neurotoxicity or dangerous hemodynamic depression. Therefore, clonidine can be used as an effective adjunct in hyperbaric bupivacaine spinal anesthesia.