Putative role of intracellular Zn(2+) release during oxidative stress: a trigger to restore cellular thiol content that is decreased by oxidative stress.

Although the ability of zinc to retard the oxidative process has been recognized for many years, zinc itself has been reported to induce oxidative stress. In order to give some insights into elucidating the role of intracellular Zn(2+) in cells suffering from oxidative stress, the effects of N-ethylmaleimide (NEM) and ZnCl(2) on cellular thiol content and intracellular Zn(2+) concentration were studied by use of 5-chloromethylfluorescein diacetate (5-CMF-DA) and FluoZin-3 pentaacetoxymethyl ester (FluoZin-3-AM) in rat thymocytes. The treatment of cells with NEM attenuated 5-CMF fluorescence and augmented FluoZin-3 fluorescence in a dose-dependent manner. These NEM-induced phenomena were observed under external Zn(2+)-free conditions. Results suggest that NEM decreases cellular thiol content and induces intracellular Zn(2+) release. Micromolar ZnCl(2) dose-dependently augmented both FluoZin-3 and 5-CMF fluorescences, suggesting that the elevation of intracellular Zn(2+) concentration increases cellular thiol content. Taken together, it is hypothesized that intracellular Zn(2+) release during oxidative stress is a trigger to restore cellular thiol content that is decreased by oxidative stress.

[Remission induced by dose-reduction of immunosuppressants alone in a patient with post-transplant lymphoproliferative disorder of central nervous system origin].

A 43-year-old male renal transplant recipient, who received a living related renal transplant 7 years ago and had been maintained with tacrolimus, mycophenolate mofetil (MMF), and prednisolone, was admitted to our hospital complaining of headache and nausea. MRI showed a large mass in the right hemisphere with ring-enhancement indicating brain abscess, tumor or lymphoma. Open biopsy was performed and pathological examination demonstrated diffuse proliferation of polymorphic cells, positive for CD20, bcl-2, EBER, and LMP-1. Based on these findings, primary central nervous system post-transplant lymphoproliferative disorder (PCNS-PTLD) was diagnosed. MMF was discontinued and tacrolimus was tapered. After 2 weeks, MRI showed regression of the tumor size and after 9 months, the tumor had disappeared. Though many reports have shown the severity of PCNS-PTLD, and recommend aggressive treatments such as chemotherapy and/or radiotherapy, our case shows that reduction of immunosuppressant alone with close observation could be a choice of treatment.

Some characteristics of membrane Cd²+ transport in rat thymocytes: an analysis using Fluo-3.

Although cadmium-induced apoptosis of lymphocytes is one of common features in the immunotoxicity of cadmium, the membrane pathway for intracellular cadmium accumulation is not fully elucidated. To characterize membrane Cd(2+) transport of rat thymocytes, the change in intracellular Cd(2+) concentration under various conditions was examined by the use of Fluo-3, a fluorescent probe for monitoring the change in intracellular concentration of divalent metal cations. The membrane Cd(2+) transport was estimated by the augmentation of Fluo-3 fluorescence induced by bath application of CdCl(2). Lowering temperature strongly suppressed the augmentation of Fluo-3 fluorescence by CdCl(2), suggesting that the metabolic process can be involved in membrane Cd(2+) transport. External acidification (decreasing pH) and membrane depolarization by adding KCl attenuated the augmentation, indicating the requirement of electrochemical driving force for membrane Cd(2+) transport into the cells. Bath application of CaCl(2) and ZnCl(2) equally decreased the augmentation, suggesting their competition with Cd(2+) at the membrane transport. The augmentation by CdCl(2) was lesser in the cells treated with N-ethylmaleimide inducing chemical depletion of cellular thiols. The result suggests the contribution of sulfhydryl groups to membrane Cd(2+) transport. Taken together, it is suggested that the cells possess a temperature-sensitive membrane Cd(2+) pathway, driven by electrochemical gradient of Cd(2+) and transmembrane potential, with competitive binding site. Based on the characteristics described above, it is unlikely that the membrane Cd(2+) transport in rat thymocytes is attributed to a single transport system although it has characteristics that are similar to those of divalent cation transporter 1.

Cytometric analysis on cytotoxicity of curcumin on rat thymocytes: Proapoptotic and antiapoptotic actions of curcumin.

Curcumin exhibits various pharmacological actions including anti-inflammatory, anti-infectious, and anticancer actions. Furthermore, the supplements containing curcumin are supplied for persons consuming alcoholic beverage. A primary criterion for an ingredient ingested by general population is that it exerts no harmful effect. In this study, we examined the effect of curcumin on rat thymocytes to see if curcumin exerts cytotoxicity on normal cells. The incubation with 10 µM curcumin for 24h increased the population of dead cells while it was not the case for 5 µM or less. Curcumin at 5-10 µM increased the populations of shrunken cells and the cells positive to annexin V, phenomena for early stage of apoptosis. However, the incubation with 10 µM curcumin suppressed the increase in population of cells with hypodiploid DNA, a phenomenon for late stage of apoptosis. Thus, curcumin at 10 µM may show both proapoptotic and antiapoptotic actions. The simultaneous incubation with 5 µM, but not 3 µM, curcumin and 0.5% ethanol increased the population of shrunken cells. It is likely that curcumin at 5 µM or more exerts cytotoxic action on normal cells although many studies show some anticancer actions of curcumin at 10 µM or more on cancer cells.

Apoptosis-inducing action of two products from oxidation of sesamol, an antioxidative constituent of sesame oil: a possible cytotoxicity of oxidized antioxidant.

Many effects of sesamol, an antioxidative constituent of sesame oil, have been reported for human health benefits due to its antioxidative action. However, we recently isolated two cytotoxic products, trimer and tetramer of sesamol, from oxidation of sesamol by an assay-guided purification. In this study, we have revealed some cytotoxic characteristics of these products in rat thymocytes and human leukemia K562 cells. Incubation of cells with trimer or tetramer at 10-30 microM for 24h significantly increased cell lethality and population of rat thymocytes containing hypodiploid DNA, suggesting cell death with DNA fragmentation, while it was not the case for 30 microM sesamol. The cytotoxic action of tetramer was more potent than that of trimer in rat thymocytes when their concentrations were 10-30 microM. The incubation of cells with 10 microM tetramer for 24h increased the population of cells with exposed phosphatidylserine, the activity of caspases, and the nick of DNA. These results indicate tetramer-induced apoptosis. In K562 cells, the incubation with tetramer at 10 microM for 72 h significantly inhibited the growth without affecting the lethality. However, tetramer at 30 microM significantly increased cell lethality. It is likely that tetramer exerts more cytotoxic action on normal non-proliferative cells (rat thymocytes) rather than proliferative cancer cells (human leukemia K562 cells). It may be necessary to consider the condition for preservation of sesamol and the safety of products from in vivo oxidation of sesamol for human health.

Reduction in adverse reactions to platelets by the removal of plasma supernatant and resuspension in a new additive solution (M-sol).

BACKGROUND: Leukodepletion reduces but does not eliminate adverse reactions to platelet concentrate (PC). As an alternative strategy, plasma reduction or washing of platelets should be considered. However, the efficacy of this strategy is still unclear. STUDY DESIGN AND METHODS: A total of 12 patients who experienced adverse reactions at a 29 to 100 percent reaction rate for plasma-PC were enrolled. The reactions were allergic reactions and nonhemolytic transfusion reactions, such as chills. Plasma-removed PC (W/R-PC), which was suspended in a recently developed additive solution (M-sol) containing less than 20 mL plasma, was prepared. W/R-PCs in M-sol were then transfused into patients after an overnight storage period; the occurrence of adverse reactions was monitored and 1- and 24-hour corrected count increment (CCI) values were evaluated. RESULTS: Although plasma-PC caused reaction in 12 patients, W/R-PC prevented reactions in 11 of 12 patients, with 1 patient having one minor allergic reaction of 15 transfusions. There was a significant difference in the incidence of reaction (p < 0.0001, Fisher's exact test). On a per-transfusion basis, the reaction rate for W/R-PC (1/156, 0.64%; 95% confidence interval [CI], 0.02%-3.5%) was reduced significantly compared to that for plasma-PC (117/276, 42%; 95% CI, 36%-48%; p < 0.0001). W/R-PC gave findings of satisfactory CCI at 1 hour (22,400 +/- 8,000/microL) and 24 hours (15,400 +/- 8,000/microL). No clinically evident bleeding episodes were recorded. CONCLUSIONS: W/R-PC suspended in M-sol in the presence of less than 20 mL plasma can be transfused safely and eliminate a wide range of adverse reactions to plasma-PC.

Effects of the mean daily doses of imatinib during the first year on survival of patients with chronic myeloid leukemia in Japan: a study of the Hokkaido Hematology Study Group.

In this study, we retrospectively analyzed 213 Japanese patients with chronic myeloid leukemia (CML) treated with imatinib mesylate. In 150 evaluable patients, mean daily doses were 400 mg or more in 42 patients, 300-400 mg in 42 patients, 200-300 mg in 44 patients and <200 mg in 22 patients. Complete hematologic response was observed in all the 84 patients treated with mean daily doses of 300 mg or more and complete cytogenetic response was achieved in 94.8% of those patients. In comparison with the effects of 300 mg or more, mean daily doses of 200-300 mg led to less complete cytogenetic response (78.6% vs. 94.8%, P < 0.01), shorter complete cytogenetic remission duration (81.3% vs. 95.6% at 24 months, P = 0.01), and lower overall survival (90.0% vs. 98.8% at 36 months, P = 0.03). This study suggests that the mean daily doses of 300 mg (roughly equivalent to 100,000 mg/yr) or more may improve overall survival and that mean daily doses of imatinib during the first year may be one of the prognostic factors for CML in Japan.

Cutaneous-type adult T-cell leukemia/lymphoma presenting as a solitary large skin nodule: a review of the literature.

Adult T-cell leukemia/lymphoma (ATLL) often involves the skin. Cases with skin lesions without either leukemic nor lymph node involvement have been categorized into a cutaneous type. While the clinical manifestations of the cutaneous-type ATLL are variable, including multiple papules, nodules, plaques, or erythroderma, a solitary skin nodule alone is rare, and only 2 cases have been reported in the literature. We present a 58-year-old Japanese patient with cutaneous-type ATLL that presented as a large, solitary skin nodule as the sole clinical feature. The skin tumor was completely resolved after treatment with x-ray and electron beam irradiation.

[ALK-negative primary gastric anaplastic large cell lymphoma].

A 56-year-old Japanese woman was admitted to our hospital with upper abdominal pain. Gastroendoscopy revealed a Borrmann III type tumor which was diagnosed from the biopsied specimen as an anaplastic large cell lymphoma (ALCL). CT scan revealed para-aortic and left-supra clavicular lymph node swelling, so her clinical stage was defined as IIIE according to the Ann Arbor classification. The patient first of all received CHOP therapy, however her lymphoma lesions increased in size. Therefore she underwent salvage chemotherapy regimens and autologous peripheral blood stem cell transplantation (APBSCT). She has remained in complete clinical remission (CCR) for eleven months after the APBSCT.

Delayed redistribution of CD27, CD40 and CD80 positive B cells and the impaired in vitro immunoglobulin production in patients with non-Hodgkin lymphoma after rituximab treatment as an adjuvant to autologous stem cell transplantation.

Recent studies have indicated that patients who received rituximab as an adjuvant to stem cell transplantation (SCT) demonstrated an increased risk of developing severe hypogammaglobulinaemia, which was found to be a result of delayed recovery of CD27 positive memory B cells and impaired isotype expression. It appears that rituximab influences both the quantity and quality of B-cell redistribution. Precisely how the B-cell repertoire regenerates after anti-CD20-mediated transient B-cell depletion in patients with non-Hodgkin lymphoma (NHL) remains to be elucidated. This study performed a phenotypical analysis of B cells in 17 NHL patients who received rituximab as an adjuvant to autologous SCT. The median period after final administration of rituximab was 36 months (range, 12-43 months). Surface antigen expression of CD27, CD40 and CD80 in NHL patients was statistically significantly different from healthy controls (n = 14). Moreover, B cells from NHL patients showed significantly impaired IgG and IgA production upon engagement of surface immunoglobulin receptors in the presence of interleukin (IL)-2, IL-10 and CD40 ligand in comparison with samples from healthy controls. The delayed recovery of memory B cells with an abnormal cell marker expression and function demonstrates that naive B cells may fail to differentiate into plasma cells, resulting in hypogammaglobulinaemia after autologous SCT and rituximab therapy.

Hypogammaglobulinemia with a selective delayed recovery in memory B cells and an impaired isotype expression after rituximab administration as an adjuvant to autologous stem cell transplantation for non-Hodgkin lymphoma.

OBJECTIVES: Some studies have indicated patients who received rituximab as adjuvant to stem cell transplantation had an increased risk of developing severe hypogammaglobulinemia. The mechanism of this hypogammaglobulinemia is unknown, although investigators have hypothesized a further delay in the B-cell recovery as one potential etiology. The aim of this study is to clarify the mechanism(s) of this hypogammaglobulinemia. METHODS: A total of 14 patients with high-risk CD20+ lymphoma underwent an autologous peripheral blood stem cell transplantation (APBSCT). After a hematological recovery, rituximab was given weekly for up to four doses as an adjuvant therapy. RESULTS: After a median follow up of 33.5 months, we found six patients (group A) who had hypogammaglobulinemia, while the eight other patients (group B) had normal serum immunoglobulin levels. A phenotypical analysis revealed that group A patients had already achieved B-cell recovery. However, we found a severe delay in the recovery of CD27+ memory B cells, especially in the IgD-/CD27+ switched populations in group A, but CD27 negative naive B-cells reverted to a normal range in both groups. Consistent with this, reverse transcriptase-polymerase chain reaction studies with peripheral blood mononuclear cells revealed that most patients in group A lacked more than two classes of isotype transcripts. CONCLUSIONS: Abnormal repertoires and impaired isotype expression are seen in patients with common variable immunodeficiency, these data suggested that rituximab after APBSCT can affect not only the B-cell quantities, but also the recovery of the B-cell repertoires.

Effective high-dose chemotherapy combined with CD34+-selected peripheral blood stem cell transplantation in a patient with cutaneous involvement of nasal NK/T-cell lymphoma.

The prognosis of nasal natural killer (NK)/T-cell lymphoma with cutaneous involvement especially is morbid despite intensive chemotherapy and radiotherapy. We treated a 52-yr-old Japanese woman with cutaneous dissemination of nasal NK/T-cell lymphoma. Six cycles of chemotherapy, irradiation to skin lesion were administered and complete remission (CR) was attained. High-dose chemotherapy (HDC; etoposide 750 mg/m(2) x 2 d, cyclophosphamide 60 mg/kg x 2 d, total body irradiation 12 Gy two daily fractions x 3 d) followed by CD34(+)-selected autologous peripheral blood stem cell transplantation (CD34(+)-APBSCT) was then prescribed. Complete remission (CR) was obtained and she has been free of disease for 34 months since CD34(+)-APBSCT. We suggest that marrow-ablative chemotherapy facilitated by autologous stem cell transplantation should be considered part of the primary therapy for subjects with a poor prognosis for nasal NK/T-cell lymphoma with cutaneous involvement.

Tumor necrosis factor-alpha inhibits generation of glycophorin A+ cells by CD34+ cells.

OBJECTIVE: The inhibitory effects of tumor necrosis factor-alpha (TNF-alpha) on cytokine-induced proliferation and differentiation of normal human erythroid progenitors have been characterized extensively, yet little is known about the maturation level of erythroid progenitors that are sensitive to TNF-alpha or of the expression of TNF receptors (TNFRs) in erythroid lineage. The aim of this study was to determine the extent to which human erythroid progenitor cells are sensitive to TNF-alpha, and to relate this to the expression of TNFRs in the erythroid lineage. MATERIALS AND METHODS: Highly purified human CD34+ cells underwent erythroid differentiation, with or without TNF-alpha. We used colony assay as well as a method by which colony-forming unit-erythroid (CFU-E) and glycophorin A (GPA; a specific marker for erythroid lineage) positive cells can be generated in liquid phase from purified human CD34+ cells in the presence of multiple cytokines, including stem cell factor (SCF), interleukin-3 (IL-3), and erythropoietin (EPO). During erythroid differentiation of CD34+ cells, TNFRs expression were monitored. RESULTS: TNF-alpha inhibited the generation of GPA+ cells by CD34+ cells as well as the proliferative capacity of GPA+ cells supported by EPO, IL-3, and SCF. Erythroid progenitors became resistant to the inhibitory effect of TNF-alpha as they matured. The detectable expression of TNFR-I was transient in the early phase of erythroid differentiation, whereas TNFR-II was expressed through the entire course of erythroid differentiation of CD34+ cells. CONCLUSIONS: TNF-alpha suppresses erythropoiesis by inhibiting the generation of GPA+ cells derived from CD34+ cells as well as by inhibiting the proliferative capacity of GPA+ cells. Although the presence of TNFRs does not directly indicate that the receptor(s) mediates death signaling, altered expression of TNFRs depending on the level of maturation may imply altered sensitivities to TNF-alpha in various stage of erythroid progenitors.