MiJung Kim

Age-related alterations in mesenchymal stem cells related to shift in differentiation from osteogenic to adipogenic potential: Implication to age-associated bone diseases and defects

Mesenchymal stem cells (MSC) have attracted considerable attention in the fields of cell and gene therapy due to their intrinsic ability to differentiate into multiple lineages. The various therapeutic applications involving MSC require initial expansion and/or differentiation in vitro prior to clinical use. However, serial passages of MSC in culture lead to decreased differentiation potential and stem cell characteristics, eventually inducing cellular aging which will limit the success of cell-based therapeutic interventions. Here we review the age-related changes that occur in MSC with a special focus on the shift of differentiation potential from osteogenic to adipogenic lineage during the MSC aging processes and how aging causes this preferential shift by oxidative stress and/or energy metabolism defect. Oxidative stress-related signals and some microRNAs affect the differentiation potential shift of MSC by directly targeting key regulatory factors such as Runx-2 or PPAR-γ, and energy metabolism pathway is involved as well. All information described here including transcription factors, microRNAs and FoxOs could be used towards development of treatment regimens for age-related bone diseases and related defects based on mutually exclusive lineage fate determination of MSC.

Angiogenesis Facilitated by Autologous Whole Bone Marrow Stem Cell Transplantation for Buerger's Disease

We hypothesized that angiogenesis can be triggered by autologous whole bone marrow stem cell transplantation. Twenty‐seven patients (34 lower limbs) with Buergers disease, who were not candidates for surgical revascularization or radiologic intervention, were enrolled in this study. Six sites of the tibia bone were fenestrated using a 2.5‐mm‐diameter screw under fluoroscopic guidance. Clinical status and outcome were determined using the “Recommended Standards for Reports.” To mobilize endothelial progenitor cells (EPCs) from bone marrow, recombinant human granulocyte colony‐stimulating factor (r‐HuG‐CSF) was injected subcutaneously as a dose of 75 μg, preoperatively. During the follow‐up period (19.1 ± 3.5 months), one limb showed a markedly improved outcome (+3), and 26 limbs showed a moderately improved outcome (+2). Thirteen limbs among 17 limbs with nonhealing ulcers healed. Postoperative angiograms were obtained for 22 limbs. Two limbs showed marked (+3), five limbs moderate (+2), and nine limbs slight (+1) collateral development. However, six limbs showed no collateral development (0). Peripheral blood and bone marrow samples were analyzed for CD34 and CD133 molecules to enumerate potential EPCs, and EPC numbers were found to be increased in peripheral blood and in bone marrow after r‐HuG‐CSF injection. In conclusion, the transplantation of autologous whole BMCs by fenestration of the tibia bone represents a simple, safe, and effective means of inducing therapeutic angiogenesis in patients with Buergers disease.

The effect of VEGF on the myogenic differentiation of adipose tissue derived stem cells within thermosensitive hydrogel matrices.

We investigated the combination of human adipose tissue derived stem cells (ADSC) and in vivo gel-forming methoxy poly (ethyleneglycol)-poly (epsilon-caprolactone) (MPEG-PCL) as a muscle regeneration matrix, with and without inclusion of vascular endothelial cell growth factor (VEGF). VEGF(165)-treated stem cell grafts showed significant proliferation and differentiation into muscle tissue in vivo. Importantly, the inclusion of VEGF enhanced vascularization. This scaffold supported preconditioned ADSC, and allowed them to differentiate into mature muscle tissues in vivo, indicating that ADSC of human origin and MPEG-PCL scaffolds provided an appropriate environment for cellular growth and expansion. Our results thus provide a potential solution to the major obstacle encountered in the engineering of thick complex tissues, which require an adequate blood supply to maintain cell viability during tissue growth and to induce appropriate structural organization. Therefore, the combination of ADSC and in vivo gel-forming MPEG-PCL with VEGF(165) might serve as a suitable non-invasive biomaterial for clinical muscle regeneration applications.

Idiopathic Thrombocytopenic Purpura: Better Therapeutic Responses of Patients with B- or T-Cell Clonality than Patients without Clonality

Results of recent studies of the pathogenesis of idiopathic thrombocytopenic purpura (ITP) have suggested activated helper T-cells drive B-lymphocytes to produce antibodies. Twenty-eight children and 85 adults with ITP entered this study. We performed polymerase chain reaction (PCR) using framework III variable region (VH FRIII)- and joining region (JH)-specific primers to analyze immunoglobulin heavy-chain gene rearrangement (IgH GR) for B-cell clonality. We used multiplex PCR to analyze T-cell receptor (TCR) γ-chain gene rearrangement (TCRγ GR) for T-cell clonality. We diagnosed 10 cases as acute ITP and 97 cases as chronic ITP. The IgH GR result was positive in 77.8% of the acute-form cases and in 58.8% of the chronic-form cases. The TCRγ GR result was positive in 11.1% of the acute cases and in 10.6% of the chronic cases. There was no dif-ference in frequency of clonality between the acute and chronic forms. After treatment the platelet count normalized in 81.8% (36/44) of the chronic ITP cases with B-cell clonality and in 88.9% (8/9) of the chronic ITP cases with T-cell clonality, compared with a normalized platelet count in 46.2% (12/26) of the chronic ITP cases without clonality. The patients with T- or B-cell clonality appeared to have better therapeutic responses than patients without clonality. In conclusion,T- and B-cell clonality may play a positive role in determining therapeutic response.

Direct international normalized ratio determination using multicalibrators is more responsive than the conventional method for measuring prothrombin time.

Direct international normalized ratio (INR) determination using certified INR plasmas was shown to improve precision and accuracy. We evaluated the utility of a multicalibrator in determining INR. INR values were measured in 493 blood samples from patients subjected to anticoagulation therapy (320) and control subjects (173). Study was performed using CA‐7000 coagulation analyzer (Sysmex, Japan) with Thromborel S (Dade Behring, Germany). Direct INR values were obtained using PT‐Multi Calibrator (Dade Behring) composed of five lyophilized calibrant plasmas. Conventional INR values were calculated from mean normal prothrombin time and instrument/reagent‐specific international sensitivity index (ISI). We compared the difference between the INR results obtained with the two methods. The mean INR value of direct INR method was significantly higher than that of conventional method. The differences in values (direct INR – conventional INR) generated using the two methods increased in proportion to the INR values. Elevation of INR was observed in data obtained with the direct INR method, compared with conventional INR values. Accordingly, we conclude that direct INR method is more responsive than conventional method in determining INR.

Minimal Residual Disease Detection in Acute Leukemia Patients by Flow Cytometric Assay of Cross-lineage Antigen Expression

BACKGROUND It has been demonstrated that flow cytometric detection of minimal residual disease (MRD) has a prognostic significance in the treatment of patients with acute leukemia. We investigated the significance of flow cytometric MRD detection for the first time in Korea. METHODS We analyzed the results of MRD detection in morphologically complete remission bone marrow aspirates from 89 patients with newly-diagnosed or relapsed acute leukemia, in which leukemic cells had cross-lineage antigen expression. Patients were grouped based on MRD frequencies: ≥ 1.0%, high MRD; <1.0%, low MRD. RESULTS Forty-seven ALL patients consisted of 10 with high and 37 with low MRD levels. Patients with high MRD levels showed a tendency of more frequent relapse than those with low MRD levels (40.0% and 13.5%, respectively) (P=0.08). High MRD group showed a tendency of short relapse-free survival (RFS) and overall survival (OS), although the differences were not statistically significant. Forty-two AML patients consisted of 16 with high and 26 with low MRD levels. There were no correlations between the MRD levels and relapse rate, RFS or OS. AML patients with high MRD levels showed significantly higher rate of unfavorable cytogenetic risk categories and lower rate of favorable risk categories (P=0.03). CONCLUSIONS MRD detection by flow cytometric assay of cross-lineage antigen expression would be useful in predicting treatment outcome in patients with ALL rather than AML. We expect that the establishment of the standardization of methods, time to test or antibody combination would be achieved through further trials in this country.

Evaluation of the TEST 1 erythrocyte sedimentation rate system and intra- and inter-laboratory quality control using new latex control materials

Abstract Background: The erythrocyte sedimentation rate (ESR) test has been considered to be a simple procedure, not requiring quality control (QC). However, QC is essential for accuracy and precision. We evaluated the TEST 1 ESR system and performed QC procedures using newly developed latex control materials in three hospitals. Methods: Using tripotassium ethylenediaminetetraacetic acid blood samples (n=184), we compared TEST 1 ESR values with Westergren ESR data and evaluated intra-assay precision. Three levels of latex control materials were used to assess inter-assay precision. Reference range assessment was done using samples from 220 healthy individuals. Inter-laboratory QC with latex control materials in three hospitals was performed. Results: Correlation between TEST 1 ESR and Westergren ESR results was good (p<0.001). Intra-assay precision [coefficients of variation (CV) 6.6%–21.7%] with patient samples and inter-assay precision (CV 0.0%–6.8%) with latex control materials were satisfactory. The reference ranges of 2–10 mm/h for males and 2–19 mm/h for females were established. Inter-laboratory QC data with latex control materials in three hospitals demonstrated good accuracy and satisfactory precision (CV 0.0%–14.4%). Conclusions: Our results demonstrate that the TEST 1 QC is reliable and the latex control materials are valuable for inter-laboratory proficiency testing. Clin Chem Lab Med 2010;48:1043–8.