PURPOSE: Mizoribine (MZR), an inhibitor of Inosine monophosphate (IMP) dehydrogenase which depletes cellular GTP, is clinically used as an immunosuppressive drug. This study was designed to evaluate the mechanism by which MZR exerts the cytotoxic effect on Jurkat T cells. METHODS: Jurkat T cell is a human T lymphocytic cell line. It was obtained from the Korean Type Culture Collection. Cell viability was measured by the MTT assay and flow cytometry. Caspase activity assay, Western blotting, 2-D PAGE, and mitochondrial membrane potential were detected using biochemical analysis. Morphologic finding was observed by Hoechst staining. RESULTS: The data demonstrated that the treatment of MZR decreased cell viability in a dose- and time-dependent manner. MZR-induced cell death was confirmed as apoptosis, which was characterized by chromatin condensation and H2AX phosphorylation. MZR increased the catalytic activity of caspase-3 protease, -8 protease and -9 proteases. The activation of caspase-3 protease was further confirmed by the degradation of polymerase (PARP), a substrate of caspase-3 protease by MZR in Jurkat T cells. Furthermore, MZR induced the changes of the mitochondrial transmembrane potential (MTP) and the cytosolic release of cytochrome c from the mitochondria. In addition, MZR induced the decrease of Bcl-X(L) expression whereas the increase of Bcl-X(S), Bak and Bim expression. Guanosine markedly inhibited cell viability and apoptosis through consistent suppression of the activity of caspase-8 protease, an upstream caspase among the caspase family, H2AX phosphorylation and PARP cleavage in MZR-treated cells. Also, I have screened the expression profile of proteins in the Jurkat T cells by using two-dimensional (2-D) gel electrophoresis. Among 300 spots resolved in the 2-D gels, the comparison of the control versus apoptotic cells revealed that the signal intensity of 10 spots was decreased and 5 spots was increased. CONCLUSION: The results suggest that MZR functions as an inhibitor of IMP dehydrogenase in apoptosis of Jurkat T cells via activation of intrinsic caspase cascades as well as mitochondrial dysfunction.
Apoptosis* ; Blotting, Western ; Caspase 3 ; Caspase 8 ; Cell Death ; Cell Line ; Cell Survival ; Chromatin ; Cytochromes c ; Cytosol ; Electrophoresis ; Flow Cytometry ; Gels ; Guanosine ; Guanosine Triphosphate ; Humans ; IMP Dehydrogenase ; Inosine Monophosphate ; Membrane Potential, Mitochondrial ; Membrane Potentials ; Mitochondria ; Oxidoreductases ; Peptide Hydrolases ; Phosphorylation ; T-Lymphocytes*

PURPOSE: Mizoribine (MZR), an inhibitor of Inosine monophosphate (IMP) dehydrogenase which depletes cellular GTP, is clinically used as an immunosuppressive drug. This study was designed to evaluate the mechanism by which MZR exerts the cytotoxic effect on Jurkat T cells. METHODS: Jurkat T cell is a human T lymphocytic cell line. It was obtained from the Korean Type Culture Collection. Cell viability was measured by the MTT assay and flow cytometry. Caspase activity assay, Western blotting, 2-D PAGE, and mitochondrial membrane potential were detected using biochemical analysis. Morphologic finding was observed by Hoechst staining. RESULTS: The data demonstrated that the treatment of MZR decreased cell viability in a dose- and time-dependent manner. MZR-induced cell death was confirmed as apoptosis, which was characterized by chromatin condensation and H2AX phosphorylation. MZR increased the catalytic activity of caspase-3 protease, -8 protease and -9 proteases. The activation of caspase-3 protease was further confirmed by the degradation of polymerase (PARP), a substrate of caspase-3 protease by MZR in Jurkat T cells. Furthermore, MZR induced the changes of the mitochondrial transmembrane potential (MTP) and the cytosolic release of cytochrome c from the mitochondria. In addition, MZR induced the decrease of Bcl-X(L) expression whereas the increase of Bcl-X(S), Bak and Bim expression. Guanosine markedly inhibited cell viability and apoptosis through consistent suppression of the activity of caspase-8 protease, an upstream caspase among the caspase family, H2AX phosphorylation and PARP cleavage in MZR-treated cells. Also, I have screened the expression profile of proteins in the Jurkat T cells by using two-dimensional (2-D) gel electrophoresis. Among 300 spots resolved in the 2-D gels, the comparison of the control versus apoptotic cells revealed that the signal intensity of 10 spots was decreased and 5 spots was increased. CONCLUSION: The results suggest that MZR functions as an inhibitor of IMP dehydrogenase in apoptosis of Jurkat T cells via activation of intrinsic caspase cascades as well as mitochondrial dysfunction.
Apoptosis* ; Blotting, Western ; Caspase 3 ; Caspase 8 ; Cell Death ; Cell Line ; Cell Survival ; Chromatin ; Cytochromes c ; Cytosol ; Electrophoresis ; Flow Cytometry ; Gels ; Guanosine ; Guanosine Triphosphate ; Humans ; IMP Dehydrogenase ; Inosine Monophosphate ; Membrane Potential, Mitochondrial ; Membrane Potentials ; Mitochondria ; Oxidoreductases ; Peptide Hydrolases ; Phosphorylation ; T-Lymphocytes*

PURPOSE: The formation of a liposarcoma is Known to be associated with a mutation of the p53 and MDM2 genes, and the histopathological subtypes of a liposarcoma are related to the prognosis of the patient. This study was performd to examine the relationship between the histopathological subtypes, the type of p53 mutation, and the proliferative rate. METHODS: Immunohistochemistry was used to measure the p53 protein and Ki-67 (Mib-1 labeling index) expression levels in 24 liposarcomas cases in which the liposarcoma developed primarily in the abdominal cavity. RESULTS: p53 expression was observed in 11.1% of the well- differentiated liposarcoma cases, 27.3% of the myxoid and round cell liposarcoma cases, and 50% of the pleomorphic liposarcoma cases. There were significant differences between the Ki-67 expression level according to the histopathological subtypes. There were significant differences between p53 positive or negative group and the Ki-67 expression level, and there was a quantitative correlation between them. CONCLUSION: The p53 protein was expressed in 25% of all liposarcomas, particularly in pleomorphic liposarcomas because it was expressed more frequently than in the other liposarcoma subtypes (in 2 cases out of 4 cases). The survival rate was much higher in the mucinous round cell liposarcomas which had high p53 and Ki-67 expression levels. The p53 expression level might be a prognostic predictor of a liposarcoma.
Abdominal Cavity ; Humans ; Immunohistochemistry ; Liposarcoma* ; Mucins ; Prognosis ; Survival Rate

BACKGROUND: Most imipenem-resistant Acinetobacter baumannii (IRAB) isolates are multiresistant, leaving few options for an effective antimicrobial therapy. We purposed to select possible candidates for the combinations of antimicrobials that are synergistic in vitro for inhibitory or bactericidal activities against IRAB and evaluate the usefulness of double disk synergy test (DDS) in predicting synergistic bactericidal activity. METHODS: Fifty-five IRAB isolates recovered from patients during the period from August 1999 to November 2000 were tested for susceptibilities to amikacin, gentamicin, tobramycin, piperacillin, piperacillin/tazobactam, cefotaxime, cefepime, cefoperazone/sulbactam (C/S), imipenem, meropenem, ciprofloxacin, levofloxacin, trimethoprim/sulfamethoxazole, chloramphenicol, minocycline, and colistin by the Clinical and Laboratory Standard Institute agar dilution method. Three isolates showing different susceptibility profiles were tested for antimicrobial synergy by DDS and then by timekill study (TKS) using DDS-positive combinations. RESULTS: Colistin, C/S, and minocycline were active in 50 (90.9%), 50, and 44 (80.0%) isolates, respectively, and all the other drugs were active in less than 20% of isolates. Minocycline-imipenem, minocycline-C/S, minocycline-amikacin, imipenem-tobramycin, C/S-amikacin, and C/S-tobramycin combinations showed synergistic inhibitory or bactericidal activity by TKS when the same combinations were synergistic in DDS; however, C/S-imipenem was found synergistic on DDS, but not by TKS. CONCLUSIONS: Colistin, C/S, and minocycline were relatively active against IRAB. DDS might help predict the synergistic antimicrobial effect of TKS if one of the combinations was susceptible.
Acinetobacter baumannii/*drug effects/isolation & purification ; Anti-Bacterial Agents/*pharmacology ; Drug Resistance, Bacterial ; Drug Synergism ; Humans ; Imipenem/*pharmacology ; Microbial Sensitivity Tests ; Time Factors

BACKGROUND AND PURPOSE: We investigated predictors of institutionalization in patients with Alzheimer's disease (AD) in South Korea. METHODS: In total, 2,470 patients with AD aged 74.5±7.8 years (mean±standard deviation, 68.1% females) were enrolled from November 2005 to December 2013. The dates of institutionalization were identified from the public Long-Term-Care Insurance program in January 2014. We used a Cox proportional-hazards model to identify predictors for future institutionalization among characteristics at the time of diagnosis in 2,470 AD patients. A similar Cox proportional-hazards model was also used to investigate predictors among variables that reflected longitudinal changes in clinical variables before institutionalization in 816 patients who underwent follow-up testing. RESULTS: A lower Mini Mental State Examination score [hazard ratio (HR)=0.95, 95% confidence interval (CI)=0.92–0.97] and higher scores for the Clinical Dementia Rating and Neuro-Psychiatric Inventory (HR=1.01, 95% CI=1.00–1.01) at baseline were independent predictors of institutionalization. The relationship of patients with their main caregivers, presence of the apolipoprotein E e4 allele, and medication at baseline were not significantly associated with the rate of institutionalization. In models with variables that exhibited longitudinal changes, larger annual change in Clinical Dementia Rating Sum of Boxes score (HR=1.15, 95% CI=1.06–1.23) and higher medication possession ratio of antipsychotics (HR=1.89, 95% CI=1.20–2.97) predicted earlier institutionalization. CONCLUSIONS: This study shows that among Korean patients with AD, lower cognitive ability, higher dementia severity, more-severe behavioral symptoms at baseline, more-rapid decline in dementia severity, and more-frequent use of antipsychotics are independent predictors of earlier institutionalization.
Alleles ; Alzheimer Disease* ; Antipsychotic Agents ; Apolipoproteins ; Behavioral Symptoms ; Caregivers ; Dementia ; Diagnosis ; Follow-Up Studies ; Humans ; Institutionalization* ; Insurance ; Korea*