[A Case of Idiopathic Thrombocytopenic Purpura during Adjuvant Chemotherapy for Colon Cancer].

A 76-year-old man underwent surgery for sigmoid colon cancer. The pathological finding was stage Ⅱ with a high-risk of recurrence (SI [bladder], l y0, v2, pN0, H0, P0, M0). He was treated with TS-1 as adjuvant chemotherapy. After the 1 course of chemotherapy, his platelet count was 4,000/mL. The high index of platelet associated IgG (PA-IgG) and bone marrow examination suggested that thrombocytopenia was caused by idiopathic thrombocytopenic purpura. The platelet count improved by prednisolone administration and Helicobacter pylori eradication treatment. After 6 months with no administration of adjuvant chemotherapy, the colon cancer recurred locally, and we performed a Hartmann's operation.

[Two cases of adult T-Cell leukemia/lymphoma with main lesion in stomach treated by VCAP-AMP-VECP regimen].

We report two cases of adult T-cell leukemia/lymphoma(ATLL)having their main lesions in the stomach. Case 1 was a 74-year-old man, complaining of left upper abdominal mass and pain. Upper gastrointestinal endoscopy revealed an ulcerous lesion in the stomach. Histological analysis and southern blotting for HTLV-1 pro-viral DNA led us to our diagnosis of ATLL. There were no apparent lesions in the bone marrow and other organs. He died of tumor lysis and multi-organ failure shortly after treatment with the VCAP-AMP-VECP regimen. Case 2 was a 68-year-old man complaining of abdominal bloating and pain. Upper gastrointestinal endoscopy disclosed an irregularity of the gastric mucosa. A biopsy sample was diagnosed pathohistologically as non-Hodgkin's lymphoma. We conducted total gastrectomy. Based on the results from the histological study and southern blotting for HTLV-1 p ro-viral DNA in the resected specimen, a diagnosis of ATLL was made. We treated him with a VCAP-AMP-VECP regimen, but multiple bone metastases and pathologic fracture occurred, proving that the disease was progressive. ATLL having a main lesion in the stomach is rare, and requires an accumulation of cases analyzed with careful diagnostic approach to establish a standard therapy for it.

Restrictive usage of monoclonal immunoglobulin lambda light chain germline in POEMS syndrome.

POEMS syndrome is a rare plasma cell disorder characterized by peripheral neuropathy, monoclonal gammopathy, and high levels of serum vascular endothelial growth factor, the pathogenesis of which remains unclear. A unique feature of this syndrome is that the proliferating monoclonal plasma cells are essentially lambda-restricted. Here we determined complete nucleotide sequences of monoclonal immunoglobulin lambda light chain (IGL) variable regions in 11 patients with POEMS syndrome. The V-region of the Ig lambda gene of all 11 patients was restricted to the V lambda 1 subfamily. Searching for homologies with IGL germlines revealed that 2 germlines, IGLV1-44*01 (9/11) and IGLV1-40*01 (2/10), were identified, with an average homology of 91.1%. The IGLJ3*02 gene was used in 11 of 11 re-arrangements with an average homology of 92.2%. These data suggest that the highly restricted use of IGL V lambda 1 germlines plays an important role in the pathogenesis of POEMS syndrome.

Automated precolumn derivatization system for analyzing physiological amino acids by liquid chromatography/mass spectrometry.

An automated method for high-throughput amino acid analysis, using precolumn derivatization high-performance liquid chromatography/electrospray mass spectrometry (HPLC/ESI-MS), was developed and evaluated. The precolumn derivatization step was performed in the reaction port of a home-built auto-sampler system. Amino acids were derivatized with 3-aminopyridyl-N-hydroxysuccinimidyl carbamate, and a 3 microm Wakosil-II 3C8-100HG column (100 x 2.1 mm i.d.) was used for separation. To achieve a 13 min cycle for each sample, the derivatization and separation steps were performed in parallel. The results of the method evaluation, including the linearity, and the intra- and inter-precision, were sufficient to measure physiological amino acids in human plasma samples. The relative standard deviations of typical amino acids in actual human plasma samples were below 10%.

Long-term prognosis of patients with endoscopically treated postoperative bile duct stricture and bile duct stricture due to chronic pancreatitis.

AIM: To compare the outcome of endoscopic therapy for postoperative benign bile duct stricture and benign bile duct stricture due to chronic pancreatitis, including long-term prognosis. METHODS: The subjects were 20 patients with postoperative benign bile duct stricture and 13 patients with bile duct stricture due to chronic pancreatitis who were 2 years or more after initial therapy. The patients underwent transpapillary drainage with tube exchange every 3 to 6 months until being free from the tube. Successful therapy was defined as a stent-free condition without hepatic disorder. RESULTS: Endoscopic therapy was successful in 90% (18/20) of the patients with postoperative bile duct stricture. The stent was removed (stent free) in 100% (20/20) of the patients, but jaundice resolved in only 10% (2/20) of patients while biliary enzymes kept increasing. Restructure occurred in 5% (1/20) of the patients, but after repeat treatment the stent could be removed. In patients with bile duct stricture due to chronic pancreatitis the therapy was successful in only 7.7% (1/13) of the patients; the stent was retained in 92.3% (12/13) of the patients during a long period. Severe acute pancreatitis occurred in 3.0% (1/33) of the patients as an accidental symptom attributable to endoscopic retrograde cholangiopancreatography (ERCP); however, it remitted after conservative treatment. CONCLUSION: Our results further confirm the usefulness of endoscopic therapy for postoperative benign bile duct strictures and good long-term prognosis of the patients.

Myeloid/Lymphoid Neoplasm with PDGFRB Rearrangement with t (5;10) (q33;q22) Harboring a Novel Breakpoint of the CCDC6-PDGFRB Fusion Gene.

Myeloid/lymphoid neoplasms with PDGFRB rearrangement are a distinct type of myeloid neoplasms that occur in association with rearrangement of PDGFRB at 5q32. The hematological features most often show prominent eosinophilia. We herein report a patient with myeloid/lymphoid neoplasms with PDGFRB rearrangement with t (5;10) (q33;q22) who showed atypical chronic myeloid leukemia-like clinical features without eosinophilia and achieved an optimal response to imatinib. A sequence analysis showed a CCDC6-PDGFRB fusion gene with a new break point in the PDGFRB gene. This is the sixth case of myeloid/lymphoid neoplasm with PDGFRB rearrangement harboring a CCDC6-PDGFRB fusion gene, and it has a new breakpoint in the PDGFRB fusion gene.

CD44 and hyaluronan engagement promotes dexamethasone resistance in human myeloma cells.

Dexamethasone (Dex) is an effective therapeutic agent against multiple myeloma (MM); however, resistance to it often becomes a clinical issue. CD44 is an adhesion molecule that serves as a cell surface receptor for extracellular matrix components, including hyaluronan (HA). HA is an extracellular matrix component that is involved in survival and progression in MM. In the present report, we describe isolation of a CD44-expressing population from a Dex-sensitive MM cell line, RPMI8226, in which the CD44-high population had a significantly higher potential to resist Dex than did the CD44-low population. Furthermore, we demonstrate that CD44 engagement by an anti-CD44 monoclonal antibody (mAb) or HA protects MM cells from Dex-induced growth inhibition. The activity of HA was partially inhibited by blocking its binding to CD44, indicating that CD44 mediates HA activity promoting MM cell survival. CD44 engagement by an anti-CD44 mAb led to phosphorylation and degradation of IkappaB-alpha, thus preventing its Dex-induced up-regulation. Our data suggest that CD44 is not only an important mediator for the survival activity of HA, but it may also contribute to MM cell resistance to Dex.

The effects of pre-analysis sample handling on human plasma amino acid concentrations.

BACKGROUND: The accurate and reliable quantification of amino acid concentrations in human plasma is important for the investigation of a number of diseases. However, few systematic studies investigating the changes in amino acid concentrations related to blood collection and storage conditions have been completed. METHODS: Blood samples were collected with EDTA-Na2 from 3 healthy volunteers and subjected to a number of different treatments; hemolysis, temperature after blood collection, time from blood collection to cooling, the influence of platelets, long term storage conditions, and repeated freeze-thaw cycles. Changes in the concentrations of 22 amino acids were determined using an Amino Acid Analyzer. RESULTS: Of the conditions influencing sample stability between blood collection and amino acid analysis, hemolysis, temperature after blood collection, and long-term storage at -20°C affected the concentrations of 11 amino acids. Time from blood collection to cooling, platelet contamination and repeated freeze-thaw cycles altered the levels of 4 amino acids. CONCLUSIONS: We observed changes in amino acid concentrations relating to blood collection and storage conditions. If attention is paid to 4 key factors (hemolysis, temperature immediately following blood collection, time from collection to cooling, and long-term storage temperature) 19 amino acids can be reliably quantified.

Validation of an analytical method for human plasma free amino acids by high-performance liquid chromatography ionization mass spectrometry using automated precolumn derivatization.

The analysis of human plasma free amino acids is important for diagnosing the health of individuals, because their concentrations are known to vary with various diseases. The development of valid, reliable, and high-throughput analytical methods for amino acids analysis is an essential requirement in clinical applications. In the present study, we have developed an automated precolumn derivatization amino acid analytical method based on high-performance liquid chromatography/electrospray ionization mass spectrometry (so-called UF-Amino Station). This method enabled the separation of at least 38 types of physiological amino acids within 8min, and the interval time between injections was 12min. We also validated this method for 21 major types of free amino acids in human plasma samples. The results of the specificity, linearity, accuracy, repeatability, intermediate precision, reproducibility, limits of detections, lower limits of quantification, carry over, and sample solution stability were sufficient to allow for the measurement of amino acids in human plasma samples. Our developed method should be suitable for use in clinical fields.

Detection and quantification of protein residues in food grade amino acids and nucleic acids using a dot-blot fluorescent staining method.

Food allergies represent an important health problem in industrialized countries, such that detection and quantitative analysis of the protein considered to be the main allergen is crucial. A dot-blot fluorescent staining method for the detection and quantitative analysis of protein residues in food grade amino acids and nucleic acids is presented. This method combines fluorescence staining with dot-blotting onto PVDF membrane. Several standard proteins, such as bovine serum albumin (66 kDa), lysozyme (14 kDa), ubiquitin (8.6 kDa), bovine insulin (5.7 kDa), and oxidized insulin B chain (3.5 kDa), were detectable at 0.1 ppm using SYPRO Ruby blot stain. Twenty-five different amino acids and two different nucleic acids of food grade were analyzed using this method combined with an internal standard addition method using BSA as an internal standard. All amino acids and nucleic acids were dissolved in 3.6% aqueous HCl and dot-blotted onto PVDF membrane before a large amount of amino acids and nucleic acid were removed. Protein residues and the internal standard protein immobilized on the membrane were stained using SYPRO ruby blot stain. The internal standard in all samples was detectable at 0.1 ppm. Samples were dissolved at 120 or 70 mg/mL, according to their solubility under acidic conditions. The detection limit of protein residues per weight was 0.8-1.4 ppm in amino acids and nucleic acids; residual protein was not detected in any sample.

Incidence, risk factors and outcomes of bronchiolitis obliterans after allogeneic stem cell transplantation.

Bronchiolitis obliterans (BO) after allogeneic stem cell transplantation (allo-SCT) is a late-onset, life-threatening respiratory complication that significantly reduces a patient's quality of life. We retrospectively analysed the incidence of and risk factors for BO in allo-SCT recipients. In 2087 patients who underwent allo-SCT between January 1994 and June 2005 and survived >90 days after transplantation, 57 patients developed BO with a 5-year cumulative incidence of 2.8%. The median time interval from transplantation to BO diagnosis was 335 days (range 83-907 days). The 5-year cumulative incidence of BO was 1.62% in bone marrow transplantation (BMT) from related donors, 3.83% in peripheral blood stem cell transplantation (PBSCT) from related donors (R-PBSCT), 2.91% in BMT from unrelated donors and 2.65% in unrelated cord blood transplantation. The incidence of BO after R-PBSCT was significantly higher than that after any other type of allo-SCT (p = 0.02). R-PBSCT (p = 0.019) and preceding chronic graft-versus-host disease (GVHD) (p < 0.001) were BO-associated risk factors. Overall 5-year survival of patients with BO from the time of diagnosis was 45.4%, significantly less than those without (77.5% from day 335, p < 0.001). R-PBSCT recipients with existent chronic GVHD have a high risk of developing BO, and need extensive care and repeated pulmonary function tests.

Advantage of LC-MS metabolomics methodology targeting hydrophilic compounds in the studies of fermented food samples.

The utility of a liquid chromatography mass spectrometry (LC-MS) method, using a pentafluorophenylpropyl (PFPP) bonded silica, was demonstrated in a metabolomics study of fermented food samples. Our LC-MS method was applied to Japanese fermented food (miso) of different stages of ripeness. The data acquired were evaluated by principal component analysis (PCA). The score plots indicated that the miso samples could be approximately classified into three groups, based on the stage of miso ripeness. The loading plots indicated that the ions responsible for group separation included not only amino acids and citric acid but also Amadori compounds. On the other hand, the miso samples were also analyzed by a conventional LC-MS method using an octadecyl (C(18)) column for comparison. The group separation of score plots from the conventional method was less clear than that from our method. The advantage of our LC-MS method is due to the different retention properties of the PFPP column and the C(18) column with hydrophilic compounds. Our LC-MS method will be useful for the metabolic profiling of fermented food samples.

Zoledronate has an antitumor effect and induces actin rearrangement in dexamethasone-resistant myeloma cells.

New strategies are needed to overcome the resistance of multiple myeloma (MM) to dexamethasone (Dex). Several recent in vitro studies demonstrated the antitumor effect of nitrogen-containing amino-bisphosphonates (N-BPs) in various tumor cell lines. Inhibition of the prenylation of small G proteins is assumed to be one of the principal mechanisms by which N-BPs exert their effects. There have been few reports on N-BP treatment of MM cells that are resistant to Dex. Additionally, it is not known how small G proteins are altered in N-BP-treated MM cells. In this study, we evaluated the effect of the most potent N-BP, zoledronate (ZOL), on a Dex-resistant human MM cell subline (Dex-R) that we established from the well-documented RPMI8226 cell line. ZOL reduced the viability and induced apoptosis of Dex-R cells. Some of the ZOL-treated RPMI8226 cells and ZOL-treated Dex-R cells were elongated; however, elongated cells were not seen among the Dex-treated RPMI8226 cells. Furthermore, we found that portions of the small G proteins, Rho and Rap1A, were unprenylated in the ZOL-treated MM cells. Geranylgeraniol reduced the above-mentioned ZOL-induced effects. These findings suggest that ZOL may be beneficial for the treatment of Dex-resistant MM by suppressing the processing of RhoA and Rap1A.

Chronic graft-versus-host disease after allogeneic bone marrow transplantation from an unrelated donor: incidence, risk factors and association with relapse. A report from the Japan Marrow Donor Program.

Chronic graft-versus-host disease (GVHD) remains the major cause of late morbidity and mortality after allogeneic stem cell transplantation. We retrospectively analysed 2937 patients who underwent bone marrow transplantation from an unrelated donor (UR-BMT) facilitated by the Japan Marrow Donor Program (JMDP) and survived beyond day 100 after transplantation. The cumulative incidence of chronic GVHD (limited + extensive) or extensive chronic GVHD at 5 years post-transplant was 45.8% and 28.2%, respectively. On multivariate analysis, seven variables predicting chronic GVHD were identified: recipient age over 20 years, donor age over 30 years, primary diagnosis of chronic myeloid leukaemia, human leucocyte antigen (HLA)-A or -B mismatch, total body irradiation-containing regimen, platelet count not having reached 50 x 10(9)/l by day 100, and prior acute GVHD. Among 2609 patients with haematological malignancy, overall survival was significantly higher in patients with limited chronic GVHD but lower in patients with extensive chronic GVHD compared with those without chronic GVHD. The cumulative incidence of relapse among patients with limited or extensive chronic GVHD was significantly lower than that among patients without chronic GVHD. Our results suggest that limited chronic GVHD provides a survival benefit to patients with haematological malignancies by reducing the risk of relapse without increasing the risk of death from chronic GVHD.