Jung-Sook Ha

Calreticulin Exon 9 Mutations in Myeloproliferative Neoplasms

Background Calreticulin (CALR) mutations were recently discovered in patients with myeloproliferative neoplasms (MPNs). We studied the frequency and type of CALR mutations and their hematological characteristics. Methods A total of 168 MPN patients (36 polycythemia vera [PV], 114 essential thrombocythemia [ET], and 18 primary myelofibrosis [PMF] cases) were included in the study. CALR mutation was analyzed by the direct sequencing method. Results CALR mutations were detected in 21.9% of ET and 16.7% of PMF patients, which accounted for 58.5% and 33.3% of ET and PMF patients without Janus kinase 2 (JAK2) or myeloproliferative leukemia virus oncogenes (MPL) mutations, respectively. A total of five types of mutation were detected, among which, L367fs*46 (53.6%) and K385fs*47 (35.7%) were found to be the most common. ET patients with CALR mutation had lower leukocyte counts and ages compared with JAK2-mutated ET patients. Conclusion Genotyping for CALR could be a useful diagnostic tool for JAK2-or MPL-negative ET or PMF patients. CALR mutation may be a distinct disease group, with different hematological characteristics than that of JAK2-positive patients.

Subpopulations of regulatory T cells in rheumatoid arthritis, systemic lupus erythematosus, and Behcet's disease.

Recently, subpopulations of regulatory T (Treg) cells, resting Treg (rTreg) and activated Treg (aTreg), have been discovered. The authors investigated the relationship between the change of Treg, aTreg and rTreg and autoimmune diseases. Treg cells and those subpopulations were analyzed by using the human regulatory T cell staining kit and CD45RA surface marker for 42 rheumatoid arthritis (RA), 13 systemic lupus sclerosis (SLE), 7 Behcets disease (BD), and 22 healthy controls. The proportion of Treg cells was significantly lower in RA (3.8% ± 1.0%) (P < 0.001) and BD (3.3% ± 0.5%) (P < 0.01) compared to healthy controls (5.0% ± 1.3%). The proportion of aTreg cells was also significantly lower in RA (0.4% ± 0.2%) (P = 0.008) and BD (0.3% ± 0.1%) (P = 0.013) compared to healthy controls (0.6% ± 0.3%). The rTreg cells showed no significant differences. The ratio of aTreg to rTreg was lower in RA patients (0.4% ± 0.2%) than that in healthy controls (0.7% ± 0.4%) (P = 0.002). This study suggests that the decrement of aTreg not rTreg cells contributes the decrement of total Treg cells in peripheral blood of RA and BD autoimmune diseases. Detailed analysis of Treg subpopulations would be more informative than total Treg cells in investigating mechanism of autoimmune disease.

Correlations between Janus Kinase 2 V617F Allele Burdens and Clinicohematologic Parameters in Myeloproliferative Neoplasms

Background This study evaluated potential correlations between the allele burden of the Janus kinase 2 (JAK2) V617F mutation and clinicohematologic characteristics in patients with myeloproliferative neoplasms (MPN). Methods Clinical and hematologic features were reviewed for 103 MPN patients, including patients with polycythemia vera (PV, 22 patients), essential thrombocythemia (ET, 64 patients), and primary myelofibrosis (PMF, 17 patients). JAK2 V617F allele status and allele burdens were measured by allele-specific PCR and pyrosequencing, respectively. Results The JAK2 V617F mutation was detected in 95.5%, 68.8%, and 52.9% of PV, ET, and PMF patients, respectively. JAK2 V617F-positive ET patients were significantly older and exhibited higher neutrophil fractions, a higher frequency of thrombotic events, and a higher myelofibrosis rate than JAK2 V617F-negative patients (P <0.05). PV patients carried the highest mean T allele burden (66.0%±24.9%) compared with ET (40.5%±25.2%) and PMF patients (31.5%±37.0%) (P =0.00). No significant correlations were detected between V617F allele burden and patient age, white blood cell count, Hb, Hct, or the platelet count for PV, ET, or PMF patients. ET patients with organomegaly had a higher JAK2 V617F allele burden (53.4%±23.7%) than patients without organomegaly (35.6%±24.3%) (P =0.03). Conclusions The JAK2 V617F mutational status and its allele burden correlate with the clinicohematologic phenotypes of ET patients, including older age, higher neutrophil count, and greater rates of organomegaly, thrombotic events, and myelofibrosis. For PV and PMF patients, larger-scale studies involving more MPN patients are needed.

Anti-cancer properties of glucosamine-hydrochloride in YD-8 human oral cancer cells: Induction of the caspase-dependent apoptosis and down-regulation of HIF-1α.

Evidence suggests anti-tumor activities of glucosamine-hydrochloride (GS-HCl). In the present study, we investigated anti-proliferative, growth suppressive and/or pro-apoptotic effects of GS-HCl on YD-8 human oral squamous cell carcinoma (OSCC) cells. Fundamentally, treatment with GS-HCl strongly inhibited proliferation and induced apoptosis in YD-8 cells, as determined by MTS and DNA fragmentation analyses. Of further note, as measured by Western analyses, GS-HCl treatment led to activation of caspase-3, cytosolic accumulation of cytochrome c, down-regulation of Mcl-1 and HIF-1α, up-regulation of GRP78, an indicator of ER stress, and generation of ROS in YD-8 cells. Importantly, results of pharmacological inhibition studies showed that treatment with z-VAD-fmk, a pan-caspase inhibitor, but not with vitamin E, an anti-oxidant strongly blocked the GS-HCl-induced apoptosis in YD-8 cells. Analyses of additional cell culture works further revealed that GS-HCl had a strong growth suppressive effect on not only YD-8 but also YD-10B and YD-38, two other human OSCC cell lines. These findings collectively demonstrate that GS-HCl has anti-proliferative, anti-survival, and pro-apoptotic effects on YD-8 cells and the effects appear to be mediated via mechanisms associated with the mitochondrial-dependent activation of caspases, down-regulation of Mcl-1, and induction of ER stress. Considering HIF-1α as a tumor angiogenic transcription factor, the ability of GS-HCl to down-regulate HIF-1α in YD-8 cells may further support its anti-cancer property. It is thus suggested that GS-HCl may be used as a potential anti-cancer drug against human OSCC.

Prevalence of Metallo-β-lactamases in Imipenem-non-susceptible Pseudomonas aeruginosa and Acinetobacter baumannii

Background: Metallo-β-lactamases (MBLs) have been reported in gram negative bacilli and are becoming increasingly important clinically because the enzymes hydrolyse almost all β-lactams, including carbapenems. Thus, the present study was conducted to determine the prevalence of MBL types in imipenem-nonsusceptible Pseudomonas aeruginosa and Acinetobacter baumannii isolated from a tertiary teaching hospital. Methods: Imipenem-nonsusceptible strains, 128 P. aeruginosa and 93 A. baumannii, were collected from clinical specimens. Identification and susceptibility tests were determined by Vitek GNI and GNS cards. MBL production was determined by modified Hodge test and imipenem-EDTA synergy test. Multiplex PCR amplification of MBL genes including blaIMP-1, blaVIM-1 and blaVIM-2 were performed. Results: Thirty-one P. aeruginosa (24.2%) isolates and 3 A. baumannii (3.2%) were found to be MBL producers. In P. aeruginosa, 20 (15.6%) and 11 (8.6%) isolates were positive for blaIMP-1 and blaVIM-2, respectively whereas 1 (1.0%) and 2 (2.2%) isolates in A. baumannii, respectively. Conclusion: IMP-1 is more prevalent MBL type than VIM-2 among imipenem-nonsusceptible P. aeruginosa unlike in other studies. Larger numbers of isolates and sequential studies are strongly recommended for the useful evaluation and monitoring of MBL production in the hospital setting to infection-control. (Korean J Clin Microbiol 2010;13:169-172)

A Case of Pneumonia Caused by Ewingella americana in a Patient with Chronic Renal Failure

Though the pathogenic significance and the reservoir of Ewingella americana have not been clarified, this organism has caused several pathogenic infections, especially in immunocompromised patients. We report a pneumonia in a patient with chronic renal failure, who had chronic rejection of transplanted kidney. E. americana was identified to be the pathogen of pneumonia with clinical symptoms and signs and radiological examination. As soon as he was treated with ceftriaxone and isepamicin, clinical improvement was followed with no further growth of E. americana or other pathogenic isolates from sputum culture. This suggests to be the case of pneumonia caused by E. americana for the first time in the Korean literature.

[A case of partial trisomy 15q25.3-qter].

A 15q25-qter partial trisomy characterized by pre or postnatal overgrowth, tall stature, macrocephaly and craniosynostosis has rarely been reported. The cause of overgrowth has been thought to be the triplication of the insulin-like growth factor 1 receptor (IGF1R) gene located on the 15q26.3. We report a patient with partial trisomy 15q25.3-qter showing mental retardation, developmental delay, macrocephaly, long narrow face, ptosis, high palate arch, scoliosis, clinodactyly and overgrowth. Additional material located on terminal 2q was found in karyotyping analysis. In bacterial artificial chromosome (BAC) clone-based-array comparative genomic hybridization (aCGH) analysis, a gain of 31 clones on 15q25.3-qter and a loss of 2 clones on 2q37.3 were observed. An extra copy of IGF1R gene was observed on derivative chromosome 2 in FISH analysis. In conclusion, the patient was diagnosed to have de novo 46,XX,der(2)t(2;15)(q37.3;q25.3) chromosome complement. Adequate genetic counseling and regular follow-ups would be needed for the patient.

Comparison of the Automated cobas u 701 Urine Microscopy and UF‐1000i Flow Cytometry Systems and Manual Microscopy in the Examination of Urine Sediments

The cobas u 701, a new automated image‐based urine sediment analyzer, was introduced recently. In this study, we compared its performance with that of UF‐1000i flow cytometry and manual microscopy in the examination of urine sediments.

Evaluation of the Usefulness of Selective Chromogenic Agar Medium (ChromID VRE) and Multiplex PCR Method for the Detection of Vancomycin-resistant Enterococci

BACKGROUND Accurate and early detection of vancomycin-resistant enterococci (VRE) is critical for controlling nosocomial infection. In this study, we evaluated the usefulness of a selective chromogenic agar medium and of multiplex PCR for detection of VRE, and both these techniques were compared with the conventional culture method for VRE detection. METHODS We performed the following 3 methods for detecting VRE infection in stool specimens: the routine culture method, culturing in selective chromogenic agar medium (chromID VRE, bioMérieux, France), and multiplex PCR using the Seeplex® VRE ACE Detection kit (Seegene Inc., Korea) with additional PCR for vanC genes. RESULTS We isolated 109 VRE strains from 100 stool specimens by the routine culture method. In chromID VRE, all the isolates showed purple colonies, including Enterococcus gallinarum and E. raffinosus, which were later identified using the Vitek card. All VRE isolates were identified by the multiplex PCR method; 100 were vanA-positive E. faecium, 8 were vanA- and vanC-1-positive E. gallinarum, and 1 was vanA-positive E. raffinosus. CONCLUSIONS For VRE surveillance, culturing the isolates in chromID VRE after broth enrichment appears to be an accurate, rapid, and easy method for routine screening test. Multiplex PCR is relatively expensive and needs skilled techniques for detecting VRE, but it can be an auxiliary tool for rapid detection of genotype during a VRE outbreak.

mRNA Expression and RNA Editing (2451 C-to-U) of IL-12 Receptor β2 in Adult Atopic Patients

Interleukin (IL)-12 activates T helper (Th) 1 cells to produce interferon (IFN)-γ which inhibits atopic inflammation. IL-12 acts through interaction with its receptor, especially β2 subunit. In several studies, the low production of IFN-γ in peripheral mononuclear cells of atopic patients on response to IL-12 stimulation has been reported. Therefore we investigated the IL-12 receptor β2 (IL-12Rβ2) mRNA expression and RNA editing, nucleotide 2451 C-to-U conversion, to find the cause of low responsiveness to IL-12 in atopy. Quantitative real time PCR for mRNA expression and sequence analysis for RNA editing were performed in 80 atopic patients and 54 healthy controls. The expression of IL-12Rβ2 mRNA was significantly lower in atopic patients than healthy controls (p<0.05). In sequence analysis, RNA editing on nucleotide 2451 was not found from either atopic patients or healthy controls. In additional evaluation, there was no relationship between expression of IL-12Rβ2 mRNA and serum total IgE or blood eosinophil count. Reduced IL-12Rβ2 mRNA expression in atopic patients indicate the reduced capacity to respond to IL-12 which induce IFN-γ production and this may contribute to Th2-skewed immune response in atopy.