Role of the local structure in superconductivity of LaO0.5F0.5BiS2-x Se x system.

We have studied the local structure of LaO0.5F0.5BiS2-x Se x by Bi L1-edge extended x-ray absorption fine structure (EXAFS). We find a significant effect of Se substitution on the local atomic correlations with a gradual elongation of average in-plane Bi-S bondlength. The associated mean square relative displacement, measuring average local distortions in the BiS2 plane, hardly shows any change for small Se substitution, but decreases significantly for [Formula: see text]. The Se substitution appears to suppress the local distortions within the BiS2 plane that may optimize in-plane orbital hybridization and hence the superconductivity. The results suggest that the local structure of the BiS2-layer is one of the key ingredients to control the physical properties of the BiS2-based dichalcogenides.

The effect of RE substitution in layered REO(0.5)F(0.5)BiS2: chemical pressure, local disorder and superconductivity.

We have studied the effect of RE substitution on the structure and the local atomic disorder in REO0.5F0.5BiS2 (RE = rare-earth) to understand their correlation with the bulk superconductivity in these materials. The mean RE size, affecting the chemical pressure, has been varied in two series namely Ce1-xNdxO0.5F0.5BiS2 and Nd1-ySmyO0.5F0.5BiS2. The lattice parameters evolve anomalously, showing an anisotropic shrinkage (elongation) of the c-axis (a-axis) to an isotropic expansion of the lattice with increasing mean RE size. The Bi L3-edge extended X-ray absorption fine structure (EXAFS) measurements are performed to investigate local displacements in the BiS2 lattice, revealing a large disorder and a sharp boundary between the Ce-containing and Sm-containing series with a distinct local structure. The results suggest that the bulk superconductivity in REO0.5F0.5BiS2 is correlated with anomalous atomic displacements in the Bi-S1 network, likely to be a combined effect of active Bi 6s electronic states and a possible Jahn-Teller-like instability of the Bi 6px/6py electrons.

Cord blood transplantation is associated with rapid B-cell neogenesis compared with BM transplantation.

Hematopoietic cell transplantation (HCT) is used for treatment of hematopoietic diseases. Assessment of T- and B-cell reconstitution after HCT is crucial because poor immune recovery has a major effect on the clinical course. In this study, we retrospectively analyzed T-cell receptor excision circles (TRECs) as well as signal and coding joint kappa-deleting recombination excision circles (sjKRECs and cjKRECs, respectively) as markers of newly produced lymphocytes in 133 patients (56 primary immunodeficient and 77 malignant cases, median (range): 12 (0-62) years old). We analyzed the kinetics of TREC and KREC recovery and determined the factors that contributed to better immune recovery. KRECs became positive earlier than TRECs and increased thereafter. Younger recipient age had a favorable effect on recovery of sjKRECs and cjKRECs. Compared with BM and peripheral blood, our data suggested that cord blood (CB) provided rapid B-cell recovery. CB also provided better B-cell neogenesis in adult HCT recipients. Chronic GVHD was associated with low TRECs, but not increased sjKRECs/cjKRECs. Finally, positive sjKRECs 1 month after HCT were associated with fewer infectious episodes. Monitoring of TRECs and KRECs may serve as a useful tool for assessment of immune reconstitution post HCT.

Functional analysis of cytomegalovirus-specific T lymphocytes compared to tetramer assay in patients undergoing hematopoietic stem cell transplantation.

In order to evaluate whether we could predict reactivation of CMV by monitoring the number of CMV-specific cytotoxic T-lymphocytes (CTL), tetramer analysis was performed in 37 patients who underwent hematopoietic stem cell transplantation (HSCT). The results disclosed that the mean number of CMV-specific CTL at day 30 did not differ among patients who developed CMV antigenemia (22/microl) and those who did not (12/microl). Serial tetramer analysis showed that 21% of the patients had >10/microl CMV-specific CTL at the first detection of CMV antigenemia and 67% of the patients had more than 10/microl CMV-specific CTL at the onset of CMV disease. Intracellular staining upon stimulation by CMV lysates and peptide in patients with CMV colitis revealed that both IFN-gamma producing CD4+ and CD8+ lymphocytes were suppressed at the onset of CMV colitis (1.6 and 8/microl), which increased with recovery of the disease (19 and 47/microl). These data suggest that it is difficult to predict CMV reactivation solely by the number of CMV-specific CTL. We suggest that additional functional analysis by intracellular cytokine assay may be useful for immunomonitoring against CMV.

Glycogen synthase kinase-3beta and p38 phosphorylate cyclin D2 on Thr280 to trigger its ubiquitin/proteasome-dependent degradation in hematopoietic cells.

Cyclin D2 plays an important role in regulation of hematopoietic cell proliferation by cytokines and is implicated in oncogenesis of various hematopoietic malignancies. However, mechanisms regulating cyclin D2 stability and its expression level have remained to be known. Here, we demonstrate that interleukin-3 signaling stabilizes cyclin D2 by inhibition of glycogen synthase kinase-3beta (GSK3beta) through Janus kinase2-dependent activation of phosphatidylinositol 3'-kinase (PI3K)/Akt signaling pathway in hematopoietic 32Dcl3 cells. On the other hand, osmotic stress was shown to induce a rapid proteasomal degradation of cyclin D2, which was mediated by activation of p38. GSK3beta and p38 was demonstrated to phosphorylate cyclin D2 on Thr280 in vitro, while a cyclin D2 mutant with this residue substituted with Ala was found to be resistant to ubiquitination and proteasome-dependent degradation in 32Dcl3 cells. Inhibition of the PI3K pathway or induction of osmotic stress also caused a rapid proteasomal degradation of cyclin D2 in primary leukemic or myeloma cells. These results indicate that cyclin D2 expression in normal and malignant hematopoietic cells is regulated by ubiquitin/proteasome-dependent degradation that is triggered by Thr280 phosphorylation by GSK3beta or p38, which is induced by inhibition of the PI3K pathway or by osmotic stress, respectively.

Rottlerin synergistically enhances imatinib-induced apoptosis of BCR/ABL-expressing cells through its mitochondrial uncoupling effect independent of protein kinase C-delta.

Although the BCR/ABL tyrosine kinase inhibitor imatinib is highly effective for treatment of chronic myeloid leukemia (CML) and Philadelphia-chromosome positive acute lymphoblastic leukemia (ALL), relapse with emerging imatinib-resistance mutations in the BCR/ABL kinase domain poses a significant problem. Here, we demonstrate that rottlerin, a putative protein kinase C-delta (PKCdelta)-specific inhibitor, acts synergistically with imatinib to induce apoptosis of BCR/ABL-expressing K562 and Ton.B210 cells. However, rottlerin inhibited neither PKCdelta nor BCR/ABL in these cells. On the other hand, rottlerin, previously characterized also as a mitochondrial uncoupler, transiently but significantly reduced mitochondrial membrane potential and gradually induced mitochondrial membrane permeabilization. Moreover, two other mitochondrial uncouplers, FCCP and DNP, very similarly induced apoptosis of BCR/ABL-expressing cells in a synergistic manner with imatinib. Imatinib synergistically enhanced mitochondrial membrane permeabilization induced by mitochondrial uncouplers, which led to release of cytochrome c into the cytoplasm and activation of caspases-3 and -9. Rottlerin also enhanced the cytotoxic effect of imatinib in leukemic cells from patients with CML blast crisis and Ph-positive ALL or a cell line expressing the imatinib-resistant E255K BCR/ABL mutant. The present study indicates that rottlerin synergistically enhances imatinib-induced apoptosis through its mitochondrial uncoupling effect independent of PKCdelta and may contribute to the development of new treatment strategy to overcome the imatinib resistance and to cure the BCR/ABL expressing leukemias.

BCR/ABL and IL-3 activate Rap1 to stimulate the B-Raf/MEK/Erk and Akt signaling pathways and to regulate proliferation, apoptosis, and adhesion.

The Ras family small GTPase Rap1 is activated by hematopoietic cytokines, such as interleukin (IL)-3, to induce beta1 integrin-mediated cell adhesion or by the BCR/ABL fusion tyrosine kinase to stimulate the MEK/Erk signaling pathway. Here, we demonstrate that the abrogation of Rap1 activation by SPA-1, a Rap1-specific GAP, inhibits activation of B-Raf, MEK, Erk, and Akt in a murine hematopoietic cell line, Ton.B210, stimulated with IL-3 or inducibly expressing BCR/ABL. Furthermore, Rap1 inactivation had an inhibitory effects on proliferation and survival of Ton.B210 cells, which were more remarkable when cells were stimulated by BCR/ABL than by IL-3. Induction of BCR/ABL expression increased adhesion of Ton.B210 cells to fibronectin in a manner at least partly dependent on its kinase activity, and Rap1 inhibition by SPA-1 partially inhibited BCR/ABL-induced adhesion of cells. Thus, IL-3- or BCR/ABL-induced activation of Rap1 may play important roles in regulation of cell proliferation and survival through activation of the B-Raf/MEK/Erk and Akt signaling pathways and in induction of integrin-mediated cell adhesion. Furthermore, as compared with IL-3, BCR/ABL is more dependent on Rap1-mediated signaling to induce cell proliferation and survival and, thus, Rap1 may represent an attractive target for novel therapies for leukemias caused by BCR/ABL.

p38 MAP kinase plays a role in G2 checkpoint activation and inhibits apoptosis of human B cell lymphoma cells treated with etoposide.

p38 MAPK is mainly activated by stress stimuli and mediates signals that regulate various cellular responses, including cell-cycle progression and apoptosis, depending on cell types and stimuli. Here we examine the role of p38 in regulation of apoptosis and cell cycle checkpoint in Daudi B-cell lymphoma cells treated with the topoisomerase II inhibitor etoposide. Etoposide activated p38, inhibited the G2/M transition with the persistent inhibitory phosphorylation of Cdc2 on Tyr15, and caused apoptosis of Daudi cells. Inducible expression of a dominant negative p38alpha mutant in Daudi cells reduced the inhibition of Cdc2 as well as G2/M arrest and augmented apoptosis induced by etoposide. SB203580, a specific inhibitor of p38alpha and p38beta, similarly reduced the inhibitory phosphorylation of Cdc2 as well as G2/M arrest and augmented apoptosis of Daudi cells treated with etoposide. These results suggest that p38 plays a role in G2/M checkpoint activation through induction of the persistent inhibitory phosphorylation of Cdc2 and, thereby, inhibits apoptosis of Daudi cells treated with etoposide. The present study, thus, raises the possibility that p38 may represent a new target for sensitization of lymphoma cells to DNA-damaging chemotherapeutic agents.

Evaluation of cytomegalovirus-specific cytotoxic T-lymphocytes in patients with the HLA-A*02 or HLA-A*24 phenotype undergoing hematopoietic stem cell transplantation.

Cytomegalovirus-specific cytotoxic T-lymphocytes (CMV-CTL) are essential for the control of CMV reactivation. To monitor the quantity and function of CMV-CTL after hematopoietic stem cell transplantation (HSCT), two CMV epitopes that bind to HLA-A*0201 NLVPMVATV (A*02NLV) and HLA-A*2402 QYDPVAALF (A*24QYD) were evaluated for their immunological potential. Samples from patients with the HLA-A*02 or HLA-A*24 serotype were analyzed by tetramer, intracellular cytokine staining and enzyme-linked immunospot (ELISPOT) assay. There were significantly more A*02NLV-specific CMV-CTL than A*24QYD (23 x 10(6) vs 0.4 x 10(6)/l). The frequency of IFN-gamma-producing cells was also higher upon stimulation with A*02NLV than with A*24QYD (2.5 vs 0.1%/CD8). Furthermore, the magnitude of CMV-CTL expansion was two- to 50-fold when cells were cultured with A*02NLV, while only an insignificant increase was observed in culture with A*24QYD. Although the number of A*24QYD-specific CMV-CTL was very low in most of the HLA-A*24 patients, the incidence of CMV reactivation did not differ between those with HLA-A*02 and HLA-A*24 serotype alone. These results suggest that an epitope other than A*24QYD plays a major role in patients with HLA-A*24. Our study also showed that A*02NLV may be a useful epitope for monitoring CMV-CTL not only in patients with HLA-A*0201 but also in those with the A*0206 genotype.

Recovery of Valpha24+ NKT cells after hematopoietic stem cell transplantation.

Human Valpha24+ natural killer T (NKT) cells have an invariant T-cell receptor-alpha chain and are activated in a CD1d-restricted manner. Valpha24+ NKT cells are thought to regulate immune responses and to play important roles in the induction of allograft tolerance. In this report, we analyzed the recovery of Valpha24+ NKT cells after hematopoietic stem cell transplantation and its correlation with graft-versus-host disease (GVHD). Patients who received a dose-reduced conditioning regimen, antithymocyte globulin- or CAMPATH-1H-containing conditioning regimen were excluded. NKT cells were reconstituted within 1 month after transplantation in peripheral blood stem cell transplantation recipients, while their numbers remained low for more than 1 year in bone marrow transplantation (BMT) recipients. The number of Valpha24+ NKT cells in BMT recipients with acute GVHD was lower than that in patients without acute GVHD, and both the CD4+ and CD4- Valpha24+ NKT subsets were significantly reduced. With regard to chronic GVHD, BMT recipients with extensive GVHD had significantly fewer Valpha24+ NKT cells than other patients. Furthermore, the number of CD4+ Valpha24+ NKT cells was also significantly reduced in patients with chronic extensive GVHD. Our results raise the possibility that the number of Valpha24+ NKT cells could be related to the development of GVHD.

High prevalence of Epstein-Barr virus in gastric remnant carcinoma after Billroth-II reconstruction.

BACKGROUND: Epstein-Barr virus (EBV) has been detected in about 10% of gastric carcinoma cases worldwide, and a high prevalence of EBV involvement in gastric remnant carcinoma has been reported recently. Details of the background remnant stomach of EBV-positive lesions, however, have not been well clarified. METHODS: We screened 17 consecutive gastric remnant carcinoma lesions resected surgically. To detect EBV, we used in situ hybridization (ISH) for EBV-encoded small RNA1 (EBER-1) and we compared the clinicopathologic feature between EBV-positive and -negative gastric remnant carcinoma cases. RESULTS: EBV was detected in 41.8% (7 of 17) of the lesions by EBER-1 ISH. All 7 EBV-positive lesions developed in the anastomotic site had undergone Billroth-II reconstruction excess 20 years previously (mean 26.4 years). Histologically, all EBV-positive lesions were poorly differentiated adenocarcinomas with intense lymphocyte infiltration. In the adjacent mucosa of carcinomas, moderate or marked intestinal metaplasia was found in 85.7% (6 of 7) of EBV-positive lesions and in 40% (4 of 10) of EBV-negative lesions. CONCLUSIONS: EBV infection is strongly associated with gastric remnant carcinoma. Atrophic change of remnant gastritis in Billroth-II anastomoses is considered to be the carcinogenic background for EBV-positive gastric remnant carcinoma.

Rac is activated by tumor necrosis factor alpha and is involved in activation of Erk.

Tumor necrosis factor alpha (TNFalpha) activates various signal transduction pathways including those involving phosphatidylinositol 3-kinase (PI3K), extracellular signal-regulated kinases (Erk), c-Jun N-terminal protein kinases (JNK), and p38 kinases. Using the Rac binding domain of PAK (PAK-RBD) as an activation-specific probe, here we demonstrate that TNFalpha very rapidly and transiently activates the Rho family GTPase Rac in L929 cells. The PI3K inhibitor LY294002 significantly inhibited TNFalpha activation of Rac as well as Erk and abolished that of the PI3K target Akt, without showing any inhibitory effects on JNK and p38 activation. Furthermore, TNFalpha activation of Erk was abolished by a dominant negative Rac mutant, Rac17N, or by an activated Rac mutant, Rac12V. These findings suggest that Rac is activated by a mechanism that is at least partly dependent on PI3K in TNFalpha stimulated cells and plays a critical role in activation of the Erk signaling pathway.

CrkL is recruited through its SH2 domain to the erythropoietin receptor and plays a role in Lyn-mediated receptor signaling.

The erythropoietin (Epo) receptor transduces its signals by activating physically associated tyrosine kinases, mainly Jak2 and Lyn, and thereby inducing tyrosine phosphorylation of various substrates including the Epo receptor (EpoR) itself. We previously demonstrated that, in Epo-stimulated cells, an adapter protein, CrkL, becomes tyrosine-phosphorylated, physically associates with Shc, SHP-2, and Cbl, and plays a role in activation of the Ras/Erk signaling pathway. Here, we demonstrate that Epo induces binding of CrkL to the tyrosine-phosphorylated EpoR and SHIP1 in 32D/EpoR-Wt cells overexpressing CrkL. In vitro binding studies showed that the CrkL SH2 domain directly mediates the EpoR binding, which was specifically inhibited by a synthetic phosphopeptide corresponding to the amino acid sequences at Tyr(460) in the cytoplasmic domain of EpoR. The CrkL SH2 domain was also required for tyrosine phosphorylation of CrkL in Epo-stimulated cells. Overexpression of Lyn induced constitutive phosphorylation of CrkL and activation of Erk, whereas that of a Lyn mutant lacking the tyrosine kinase domain attenuated the Epo-induced phosphorylation of CrkL and activation of Erk. Furthermore, Lyn, but not Jak2, phosphorylated CrkL on tyrosine in in vitro kinase assays. Together, the present study suggests that, upon Epo stimulation, CrkL is recruited to the EpoR through interaction between the CrkL SH2 domain and phosphorylated Tyr(460) in the EpoR cytoplasmic domain and undergoes tyrosine phosphorylation by receptor-associated Lyn to activate the downstream signaling pathway leading to the activation of Erk and Elk-1.

Rap1 is activated by erythropoietin or interleukin-3 and is involved in regulation of beta1 integrin-mediated hematopoietic cell adhesion.

The CrkL adaptor protein is involved in signaling from the receptor for erythropoietin (Epo) as well as interleukin (IL)-3 and activates beta(1) integrin-mediated hematopoietic cell adhesion through its interaction with C3G, a guanine nucleotide exchange factor for Rap1. We demonstrate here that Epo as well as IL-3 activates Rap1 in an IL-3-dependent hematopoietic cell line, 32D, expressing the Epo receptor. The cytokine-induced activation of Rap1 was augmented in cells that inducibly overexpress CrkL or C3G. The CrkL-mediated enhancement of cell adhesion was inhibited by expression of a dominant negative mutant of Rap1, Rap1A-17N, whereas an activated mutant of Rap1, Rap1A-63E, activated beta(1) integrin-dependent adhesion of hematopoietic cells. In 32D cells, Rap1 was also activated by phorbol 12-myristate 13-acetate and ionomycin, which also enhanced cell adhesion to fibronectin, whereas, an inhibitor of phospholipase C, inhibited both cytokine-induced activation of Rap1 and cell adhesion. It was also demonstrated that Rap1 as well as CrkL is involved in signaling from the EpoR endogenously expressed in a human leukemic cell line, UT-7. These results suggest that Epo and IL-3 activate Rap1 at least partly through the CrkL-C3G complex as well as through additional pathways most likely involving phospholipase Cgamma and strongly implicate Rap1 in regulation of beta(1) integrin-mediated hematopoietic cell adhesion.

Mobilizaçäo precoce da mäo pós-reimplantes ou revascularizaçöes / Early mobilization of the post-replantation hand or revascularizations

A alta frequência de complicaçöes pós-operatórias nos traumatismos das mäos e dedos por aderências cicatriciais, sugerem a aplicaçäo de técnicas de reabilitaçäo precoce, uma vez que a mobilizaçäo da mäo neste período fornece melhor aporte vascular à área lesada, promove melhor qualidade nutricional, mantém as superfícies de deslizamento e evita a täo rotineira aderência tendinosa, que impossibilita a integridade funcional da mäo e dedos comprometidos, além de evitar edemas e outros problemas pós-cirúrgicos. No entanto, há divergências em relaçäo ao início devido, talvez, à insegurança dos cirurgiöes na aplicaçäo da técnica de cinesioterapia pós-operatória imediata após uma cirurgia que normalmente teve muitas horas de duraçäo. Sugere-se neste estudo que a mobilizaçäo pós-reimplante ou revascularizaçäo dos dedos e/ou da mäo seja iniciada após 48 horas, conforme o protocolo utilizado nos 3 casos apresentados neste trabalho, justificando, dessa forma, a integridade funcional e a reabilitaçäo funcional da mäo e dedos traumatizados. Preconizou-se o ínico das atividades diárias a partir do 21§ dia após a cirurgia, sem qualquer incidente traumático em funçäo da precocidade e com integridade funcional da mäo.