Dong Il Won

Transplantation with higher dose of natural killer cells associated with better outcomes in terms of non-relapse mortality and infectious events after allogeneic peripheral blood stem cell transplantation from HLA-matched sibling donors.

Abstract:  Background: Little is known about the role of the CD56+ natural killer (NK) cell dose on the outcome of allogeneic peripheral blood stem cell transplantation (PBSCT). Recently, higher dose of NK cells has been associated with a lower incidence of severe graft‐versus‐host disease (GVHD). The current study attempted to evaluate the effect of the NK cell dose on transplant outcomes in allogeneic PBSCT setting. Methods and materials: Sixty‐one cytokine mobilized PBSC recipients were analyzed according to the infused dose of CD34+ cells and NK cells in relation to overall survival (OS), non‐relapse mortality (NRM), GVHD, and infectious events. Results: The group received a higher dose of NK cells (≥5 × 107/kg) showed a lower incidence of NRM (P = 0.0186) and infectious events (P = 0.0107). In a multivariate analysis, a higher dose of NK cells was correlated to better transplant outcomes for NRM (P = 0.042) with CD34+ cell dose (P = 0.018), and for infectious events (P = 0.013) with CD34+ cell dose (P = 0.016). Higher NK cell infusion group also showed a faster immune recovery in serial measurements at days +90, +180, and +365. Conclusions: High dose of NK cells may play an important role in improving transplant outcomes, in terms of reducing NRM and infectious events together with CD34+ cells.

Flow cytometric detection of erythrocyte osmotic fragility

Hereditary spherocytosis (HS) is the most common cause of inherited chronic hemolysis. The various tests developed for HS screening have many shortcomings. The purpose of this study was to develop a new, simple, reliable test using flow cytometry.

Trafficking Macrophage Migration Using Reporter Gene Imaging with Human Sodium Iodide Symporter in Animal Models of Inflammation

The aim of this study was to investigate the feasibility of nuclear molecular imaging using the human sodium iodide symporter (hNIS) as a reporter gene to monitor macrophage migration toward the inflammatory foci. Methods: A stable macrophage cell line coexpressing hNIS and green fluorescent protein (GFP) genes (RAW264.7/hNIS-GFP and RNIS cell) was established from an immortalized macrophage cell line (RAW264.7 cells). 125I uptake was determined (for hNIS protein functional activity), and flow cytometry analysis (to examine GFP gene expression), a cell proliferation assay, a cytokine assay, and a phagocytic activity assay were performed. 99mTc-pertechnetate images were acquired at 1 d after subcutaneous inoculation of RNIS cells in nude mice. Chemical inflammation was induced for in vivo imaging in the thigh of nude mice by turpentine oil injection. Small-animal PET with 18F-FDG and 124I was performed with an intravenous administration of RAW264.7 or RNIS cells in inflammation-induced animals. Results: The expression of hNIS and GFP genes was confirmed in RNIS cells by flow cytometry and immunofluorescent staining. 125I uptake was about 67 times higher in RNIS cells than in RAW264.7 cells. No significant difference was observed in cell proliferation, cytokine production, and phagocytic activity between RAW264.7 and RNIS cells. 99mTc-pertechnetate imaging revealed increased tracer uptake at the inoculation site. PET with 124I demonstrated a donut-shaped uptake, correlating with uptake shown by the 18F-FDG PET images, at the inflammation site of mice administered RNIS cells. 124I uptake (percentage injected dose per gram) was about 2.12 times higher at the inflammation site in the RNIS mice than in RAW264.7 mice. By immunohistochemistry, the migration of macrophages was further confirmed by positive staining for GFP and hNIS at the inflammation site of RNIS mice. Conclusion: These data support the feasibility of hNIS reporter gene imaging to monitor the macrophage migration toward an inflammatory lesion. Macrophages expressing hNIS may provide a new strategy to investigate the cellular behavior seen with inflammatory response in a preclinical model.

Living Donor Liver Transplantation for Acute Hepatic Failure Caused by Reactivation of Hepatitis B Virus Infection After Chemotherapy for Hematologic Malignancy: Case Reports

Cancer chemotherapy in chronic hepatitis B virus (HBV) carriers occasionally leads to acute hepatic failure (AHF) from viral reactivation resulting in an high mortality rate. In this situation, living donor liver transplantation (LDLT) can be life saving. Herein we have reported 2 cases of successful LDLT performed for AHF caused by reactivation of HBV infection during chemotherapy for hematologic malignancies. In case 1, a 38-year-old male HBV carrier with a neck mass was hisopathologically diagnosed as Hodgkins lymphoma. During 4 cycles of chemotherapy he developed right upper quadrant pain and jaundice. Laboratory data (alanine amino transferase, 701 U/L, total bilirubin: 7.92 mg/dL, positive hepatitis B e antigen showed that he had experienced an acute exacerbation of chronic hepatitis. Soon, he developed grade IV hepatic encephalopathy with a total bilirubin level of 50.56 mg/dL and a model for End-Stage Liver Disease score of 40. After LDLT, he has been free of relapse for 52 months so far. In case 2, a 49-year-old male HBV carrier was diagnosed in the chronic phase of chronic myeloid leukemia. The patient had been under Imatinib treatment for 1 year until he was admitted for AHF. He developed grade II encephalopathy with a total bilirubin of 50.8 mg/dL. We performed LDLT; the patient has been free of relapse for 17 months. LDLT was a life-saving procedure for AHF caused by reactivation of HBV during chemotherapy for hematologic malignancy. It can provide long-term survival if the coexistent hematologic malignancy has been controlled.

Flow cytometric measurements of TB-specific T cells comparing with QuantiFERON-TB gold†

Interferon‐γ (IFN‐γ) release assays and the detection of IFN‐γ synthesis in the cytoplasm of activated CD4+ T cells by flow cytometry have recently been used for tuberculosis (TB) diagnosis. The aim of this study was to compare the performance of IFN‐γ assay between ELISA (QuantiFERON‐TB Gold, QFT) and intracellular cytokine flow cytometry (ICCFC), and to investigate the significance of an optimal gating strategy in ICCFC.

Association of G-137C IL-18 promoter polymorphism with acute allograft rejection in renal transplant recipients.

Background. Interleukin 18 (IL-18) is a potent proinflammatory cytokine, which is postulated to play a role in mechanism of renal allograft rejection and strongly induces interferon (IFN)-&ggr; production. The aim of this study was to investigate the association of the G–137C IL-18 promoter polymorphism with acute allograft rejection in renal transplant recipients (RTRs). Methods. A total of 226 RTRs and 148 controls were recruited for association analysis. Genotyping was performed by using a real-time polymerase chain reaction. Patients were separated into two groups, the acute rejection (AR) (n=37) or the no AR group (n=189), depending on their history of AR episodes. IL-18 and IFN-&ggr; serum levels of 73 randomly selected RTRs were measured by ELISA. Results. No significant differences in genotype and allele frequencies were observed between the RTRs and the controls. Significant differences in genotype frequency and allele frequency between the AR and no AR group were observed. The frequency of the –137GG genotype was significantly increased in patients with AR (P=0.015, odds ratio=3.653). Serum levels of IL-18 and IFN-&ggr; were significantly elevated in the AR group with compared with the no AR group. In the AR group, patients with the –137GG genotype had significantly higher IL-18 serum levels compared to other genotypes. Conclusion. These data demonstrate that the –137GG genotype of the IL-18 gene, encoding higher IL-18 production, seems to be associated with AR and may be a useful marker of AR risk in RTRs.

Susceptibility of oxidative stress on red blood cells exposed to gamma rays: Hemorheological evaluation

Irradiation has been shown to induce biochemical changes in stored red blood cells (RBCs) and to generate reactive oxygen species (ROS). This study evaluated the hemorheological properties, the degree of lipid peroxidation and the oxidative susceptibility of irradiated RBCs. Furthermore, we investigated the radioprotective role of N-t-butyl hydroxylamine (NtBHA) against gamma-ray exposure of RBCs. RBC concentrates were irradiated with a minimum dose of 25 Gy, and were exposed to FeSO4 to examine the oxidative susceptibility. RBC deformability was evaluated by the use of a microfluidic ektacytometer, in relation to the hematological and biochemical properties. The deformability of the irradiated RBCs was significantly lower than that of control. Exposure to gamma rays significantly increased the mean corpuscular volume (MCV) and lipid peroxidation. Changes in RBC deformability were more prominent in irradiated RBCs than in non-irradiated RBCs also under conditions of oxidative stress. The deformability of NtBHA treated RBCs prior to irradiation was not altered as compared with irradiated RBCs not treated with NtBHA. In conclusion, irradiation reduces RBC deformability during storage and the irradiated RBCs seem susceptible to oxidative stress. NtBHA may not have a protective role against the effects of gamma-ray exposure in RBCs but further evaluation of NtBHA or another radioprotective compound is required.

Simultaneous detection of antibody binding and cytotoxicity in flow cytometry crossmatch for renal transplantation

The anti‐HLA antibody can be detected using either a complement‐dependent lymphocytotoxicity (CDC) assay or a flow cytometry crossmatch (FCXM) in renal transplantation. Discordant results are often obtained because the two methods detect different reaction phases between the donor lymphocytes and the recipient sera. This study was intended to confirm that antibody binding and cytotoxicity to the lymphocytes can be detected simultaneously in a single FCXM assay, cytotoxic FCXM.

Highly sensitive and novel point-of-care system, aQcare Chlamydia TRF kit for detecting Chlamydia trachomatis by using europium (Eu) (III) chelated nanoparticles.

Background The bacterium Chlamydia trachomatis is one of the leading causes of sexually transmitted diseases worldwide. Since no simple and effective tool exists to diagnose C. trachomatis infections, we evaluated a novel point-of-care (POC) test, aQcare Chlamydia TRF kit, which uses europium-chelated nanoparticles and a time-resolved fluorescence reader. Methods The test performance was evaluated by comparing the results obtained using the novel POC testing kit with those obtained using a nucleic acid amplification test (NAAT), using 114 NAAT-positive and 327 NAAT-negative samples. Results The cut-off value of the novel test was 20.8 with a detection limit of 0.27 ng/mL. No interference or cross-reactivity was observed. Diagnostic accuracy showed an overall sensitivity of 93.0% (106/114), specificity of 96.3% (315/327), positive predictive value (PPV) of 89.8% (106/118), and negative predictive value (NPV) of 97.5% (315/323). The sensitivity of the novel test was much higher than that of currently available POC tests. Furthermore, the relative ease and short turnaround time (30 min) of this assay enables C. trachomatis-infected individuals to be treated without a diagnostic delay. Conclusions This simple and novel test is a potential tool to screen a larger population, especially those in areas with limited resources.

CYP3A and ABCB1 genetic polymorphisms on the pharmacokinetics and pharmacodynamics of tacrolimus and its metabolites (M-I and M-III).

Background We prospectively studied renal transplant recipients receiving tacrolimus to determine the relationship between the CYP3A4, CYP3A5, and ABCB1 genetic polymorphisms and the pharmacokinetics (PK) and pharmacodynamics (PD) of tacrolimus and its metabolites. Methods Renal transplant recipients receiving tacrolimus were genotyped for CYP3A4*4, CYP3A4*5, CYP3A4*18, CYP3A5*3, ABCB1 c.1236C→T, ABCB1 c.2677G→A/T, and ABCB1 c.3435C→T. Dose-adjusted trough concentration (C0) of tacrolimus and its metabolites (M-I and M-III) and PK and PD (T-cell and monocyte subsets) were determined on transplantation days –2, 5, 30, and 90 and correlated with the corresponding genotypes. Results The dose-adjusted C0 of tacrolimus and its metabolites and AUC0–12 were significantly higher and the mean fluorescence intensity (MFI) of HLA/DR+ in monocytes was significantly lower in patients with CYP3A5*3/*3 than in patients with CYP3A5*1/*1 or CYP3A5*1/*3. However, there was no significant difference in the dose-adjusted C0 of tacrolimus and its metabolites, PK and PD among the ABCB1 genotypes. The MFI of HLA/DR+ in monocytes showed a significant negative correlation with dose-adjusted C0 of tacrolimus and its metabolites and AUC0–12. In a multiple regression analysis, the presence of the CYP3A5*3/*3 genotype was a significant independent variable determining the dose-adjusted C0 of tacrolimus and its metabolites, AUC0–12, and the MFI of HLA/DR+ in monocytes. Conclusions This study demonstrates that the CYP3A5 genetic polymorphisms are associated with the individual differences in PK and PD as well as in C0 of tacrolimus and its metabolites. The MFI of HLA/DR+ in monocytes might be considered to be a significant tool for monitoring tacrolimus efficacy.