bluuebunny/crossref_metadata_2025_split huggingface dataset:
Expression of CXCR4 on Intrahepatic Stem Cells from Organ Donors and Patients with Chronic Liver Disease.
Authors: Porretti Laura , Lopa Raffaella , Gramignoli Roberto , Ambrosone Antonella , Mosca Annamaria , Cattaneo Alessandra , Scalamogna Mario , Gatti Stefano , Rossi Giorgio , Colli Agostino , Prati Daniele 
Published: October 2019

Abstract
Replication of mature hepatocytes can ensure hepatic tissue renewal in normal liver. However, in certain physiopathologic conditions, cells with stem-like properties can proliferate and/or be recruited from extra-hepatic compartments. For a better understanding of the mechanisms related to liver regeneration, more data on the morphological and differentiative profiles of liver stem cells are needed. This study was aimed at the immunophenotypic characterization of circulating and intrahepatic stem cells in organ donors and in patients with chronic liver disease.
We performed a four-colors flow cytometric evaluation of liver mononuclear cell suspensions (L-MNC) obtained after enzymatic digestion of liver biopsy specimens from 8 patients and 6 multi-organ donors. Paired peripheral blood mononuclear cells (PB-MNC) from the same subjects were also examined. L-MNC and PB-MNC were incubated with antibodies to the following markers: CD34, CD133, CD45, CD117 (c-kit) and CXCR4 (Stromal derived factor-1 receptor). Cell viability was performed using 7-aminoActinomycin D to exclude dead cells.
In patients, the stem cell marker CD34 was detected on the surface of 0.6±0.9% of the L-MNC, a proportion similar to that observed in donors (0.6±0.5%). In both patients and donors, the majority of CD34+ cells co-expressed CD45 (86±9% and 60±31% respectively). The subset of CD34+/CD45- cells expressing c-kit was increased in L-MNC from patients as compared to donors (13.8±12% vs.1.5±1% of the total CD34+ cells; p<0.05 by Student t test), while it was scarcely represented or not detectable in PB-MNC. The proportion of CD34+ cells with a definite hematopoietic profile (CD45+/c-kit+) did not differ between patients and donors, both in L-MNC (50±32% vs. 57±13%) and in PB-MNC (66±20% vs. 45±23%). However, in the latter compartment we identified a subset of cells expressing both CD133 and CXCR4. This subset of CD34+ cells was more frequently found in diseased (74±25%) and normal liver (50±18%) as compared to peripheral blood from both patients and donors (24±22% and 30±30% respectively), (p<0.01; paired-t test).
Flow cytometry analysis of intrahepatic CD34+ stem cells allows the identification of at least two phenotipically distinct populations. The first population co-expresses c-kit, is increased in the presence of chronic hepatocellular damage and is likely resident in the liver, being rarely detectable in peripheral blood. The second population resembles the classical hematopoietic stem cell and harbors CD133 and the chemokine receptor CXCR4 for trafficking to the liver in response to liver injury.
Erythropoietin Augments Antibody Response to DNP in Immunized Mice.
Authors: Yeyni Lidor , Katz Odelia , Prutchi-Sagiv Sari , Skaat Alon , Barzilay Eran , Mittelman Moshe , Neumann Drorit 
Published: October 2019

Abstract
Recombinant human erythropoietin (rHuEpo), the main growth regulator of red blood cells, has been used in clinical practice for the treatment of anemia of various etiologies. Over the last decade several reports have indicated that the action of Epo is not restricted to the erythroid lineage. Studies indicate that rHuEpo treatment may have an immunomodulatory effect on hemodialysis patients. Here we address the effects of rHuEpo on the humoral immune response of mice. Naïve BALB/c mice (two groups of 16 mice each) were immunized with 2mg antigen dinitrophenylated keyhole limpet hemocyanin (DNP-KLH). Concomitant to antigen injection, the mice were injected three times weekly with rHuEpo (180U per injection) or with diluent (control). Blood samples were taken 2 and 4 weeks after immunization and anti-DNP antibody levels were measured by ELISA. Production of anti DNP antibodies increased significantly (1.5–2 fold) in the initial phase of the response (2 weeks following DNP-KLH injection) in mice injected with rHuEpo compared to their counterpart control antigen-injected mice, that received diluent only.
These data indicate that erythropoietin may have a role in stimulating the humoral arm of the immune system, suggesting an immunomodulatory and/or immunostimulatory role for rHuEpo and may open new avenues in improving the immune response of immunocompromised patients.
Bcr-Abl Protection of Fas-Induced Apoptosis.
Authors: Brumatti Gabriela , Castro Fabiola A. , Bueno-da-Silva Ana Elisa B. , Vaccarezza Gabriela , Amarante-Mendes Gustavo P. 
Published: October 2019

Abstract
The Bcr-Abl tyrosine kinase found in Philadelphia chromosome (Ph1)-positive leukaemia is the result of a reciprocal translocation between chromosomes 9 and 22. Depending on the breakpoint of the bcr gene, different isoforms of Bcr-Abl appear and are associated with specific types of leukemias; in chronic myelogenous leukaemia (CML), a 210kDa protein is responsible for a deregulated proliferation and consequently accumulation of myeloid cells and their precursors, whereas in acute lymphocytic leukaemia (ALL) a 185kDa Bcr-Abl protein is the oncogenic counterpart. This chimeric protein not only induces transformation of cell lines, but also regulates cell proliferation and is one of the most potent anti-apoptotic molecules responsible for resistance to a variety of apoptosis-inducing agents. The signalling pathway of apoptosis which may be regulated by Bcr-Abl is still unknown, but our latest findings suggested that Bcr-Abl act at the mitochondrial level, preserving the mitochondria integrity during several apoptogenic stimuli thereby preventing release of cytochrome c and the consequent activation of effector caspases. To investigate the mechanisms whereby Bcr-Abl interferes with the cell apoptotic machinery, we asked where in Fas signalling cascade that Bcr-Abl blocks the apoptogenic signal. For this purpose, HL-60.neo, HL-60.Bcr-Abl, HL-60.Bcl-2 and HL-60.Bcl-xL cells were stimulated with anti-Fas antibodies (CH11), in the presence or not of cycloheximide, and the levels of apoptosis and activation of caspases where compared between wild-type and transfected cells with anti-apoptotic molecules. Our results demonstrate that Bcr-Abl blocks both mitochondria dependent and independent Fas pathway, suggesting that Bcr-Abl has a bigger spectrum of action than Bcl-2 and Bcl-xL.
CNPq, FAPESP and IEP-HIAE
Mutations Innate Immune System NOD/CARD15 Gene and Toll-Like-Receptor-4 Gene Are Associated with Increased Risk for Severe Acute Gvhd in Patients Who Underwent Allogeneic Transplantation.
Authors: Elmaagacli Ahmet H. , Koldehoff Michael , Steckel Nina K. , Hindahl Heidrun , Trenschel Rudolf , Ottinger Hellmut , Beelen Dietrich W. 
Published: October 2019

Abstract
The innate immune system detects invading pathogens through several pattern-recognition receptors and represents the first line of mucosal host defense. Mutations of the NOD2/CARD15 and Toll like receptor −2, −4, −9 genes have been associated with an increased incidence of inflammatory bowel disease due to a diminished response to bacterial cell wall products. Moreover, it was reported that mutations of NOD2 gene loci might be associated with an increased risk for TRM and severe GVHD in patients who underwent allogeneic blood stem cell transplantation. Here we analyzed 201 patients and their respective donors for NOD2, TLR-2, TLR-4 and TLR-9 mutations and correlated the results with the incidence of overall acute GVHD and intestinal GVHD. Further, we evaluated if the occurrence of NOD2/CARD15 and TLR allele mutations are accompanied with an increased risk for transplant-related mortality and overall survival.
Mutated alleles of the NOD2 gene were observed in 17% of the patients and 16% of the donors. NOD2 gene mutations in patients and donors together were associated with an increased incidence of severe acute GVHD grade III-IV (6/11 patients; 55% versus 35/145 patients; 24%). If a mutated allele of the NOD2 gene was found either in patients or in donors only no correlation with increased incidence of severe acute GVHD was seen in this study.
Patients with mutated alleles in TLR-4 genes (Asp299Gly and Thr399Ile) had a higher incidence of intestinal GVHD (p<0.02), whereas mutations of donor alleles only had no influence on the occurrence of intestinal GVHD in this analysis.
No correlation with intestinal GVHD or overall acute GVHD was seen in patients with mutated alleles for TLR-2 and TLR-9. Overall survival, TRM and relapse risks was not influenced in none of the patients with mutated alleles of the innate immune system genes. Multivariate analysis including all potential factors, confirmed that that mutations of NOD2/CARD15 in patients and donors together influenced the occurrence of severe acute GVHD, whereas the occurrence of intestinal GVHD was influenced in patients by mutated alleles of TLR-4 gene. But mutations in alleles of the NOD2 or TLR-2,−4,−9 had no influence on the outcome of allogeneic transplant (TRM and overall survival) in the multivariate analysis.
Since all patients in our institution had received an intestinal bacterial decontamination using metronidazole and ciprofloxacin after allogeneic stem cell transplantation, it might be speculative, if the reduction of concentrations of anaerobic bacteria in the intestinum might have protected patients from the occurrence of increased TRM. Decontamination of anaerobic bacteria is associated with a reduced risk for severe acute GVHD as reported earlier (Blood1999; 93:3267–75).
CD133 Positive Mesenchymal Stem Cells in Mobilized Peripheral Blood and Cord Blood: Proliferation, Oct4 Expression and Plasticity.
Authors: Tondreau Tatiana , Meuleman Nathalie , Dejeneffe Marielle , Delforge Alain , Bron Dominique , Lagneaux Laurence 
Published: October 2019

Abstract
Background: Mesenchymal Stem Cells (MSC) can be isolated from bone marrow, adipose and fetal tissues, but their presence in Mobilized Peripheral Blood (MPB) and in Umbilical Cord Blood (UCB) remains controversial.
Methods: In this study we evaluated whether MPB (n=6) and UCB (n=8) could be two other sources of MSC beside Bone Marrow (BM). CD133 positive cell fraction was isolated through immunomagnetic system and MNC were seeded in medium supplemented or not with 5% of conditioned medium durong the first 48 hours of adhesion. MSC derived from MPB or UCB were identified by their expression of mesenchymal (SH2, SH3) and hematopoietic markers (CD14, CD34, CD45 and CD62-E). MPB and CB-MSC were also tested for their capacity to generate CFU-F (Fibroblast colony-forming units) and to differentiate into adipocytes, osteocytes, chondrocytes and neuronal/glial cells after specific induction. Finally we tested through RT-PCR the gene expression of Oct4, a transcriptional binding factor present in undifferentiated cells with high proliferative capacity.
Results: Through CFU-F, we observed that the selection of CD133 positive cells allows to obtain a great amount of MSC in comparison with the MNC fraction seeded (MPB-MSC, 139 versus 72 and UCB-MSC 165 versus 8 after the primoculture). After four passages, more than 1.108 cells were obtained by the cell expansion assays from 1.106 cells seeded in primoculture. During the first culture, whatever the medium used or cell fraction seeded, the cell population was composed of osteoblasts, osteoclasts, marcophages and hematopoietic cells with less than 5%SH3 positive cells. After two passages, cells derived from MPB or UCB expressed mesenchymal markers (SH3 and SH2) and Oct4. Under appropriate conditions these cells were able to differentiate into adipocytes, osteocytes, chondrocytes and neuronal/glial cells.
Conclusion: These results indicate that MSC are present in MPB and UCB, with similar characteristics to bone marrow (self-renewal capacity and multi-diffentiation potential), and may have a major clinical importance due to their accessibility. They could be an attractive target for cellular and gene therapies
Mesenchymal marker expression (%)
T0 (%) second passage (%) MPB SH2 2,7±1,1 96,1±2,9 SH3 4,9±1,5 95,7±3,3 UCB SH2 1,4±0,6 98±0,7 SH3 5,2±1,8 96,5±2,5
Establishment and Characterization of Human Fetal Liver Derived Stem Cell Line.
Authors: Chung Hyung-Min , Rhee Yun-Hee , Cha Kwang-Yul , Baek Jin-Young 
Published: October 2019

Abstract
Mesenchymal stem cells (MSC) have been derived from different sources, including bone marrow and liver. To further support the hypothesis that MSC may also exist in most postnatal tissues, we isolated a clonogenic, multipotent, rapidly proliferating population of cells from a fetal liver and defined them as mesenchymal-like stem cells (FL-MLSC) derived from human fetal liver. In this study, FACS analysis showed that FL-MLSC are positive for CD105 (endoglin/SH-2), CD73 (5′ terminal nucleotidase/SH-3), CD166 (ALCAM-1), CD44 (the hyaluronate receptor), HLA class-I(HLA-ABC), CDw90 (thy-1), CD13, and CD106 but negative for CD34 and HLA class-II(HLA-DR). Cell-cycle analysis showed that more than 81.9% cells were arrested in G0 and G1 phases, whereas an only small subpopulation of cells was actively engaged in proliferation (S + G2 + M = 18%) and doubling in about 48 hours. FL-MLSC retained normal diploid karyotypes and growth characteristics over the successive culture. In addition, under differentiation culture conditions, these cells showed the capability of differentiation into various cell types including chondrogenic, adipogenic, osteogenic, neuronal and endocrine lineages. These results demonstrated that FL-MLSC could be isolated from human fetal livers by means of their adherent ability and suggested that they were capable of self-renewal and multipotent differentiation.
Analyzing the MLL Recombinome.
Authors: Meyer Claus , Schneider Bjoern , Reichel Martin , Angermuller Sieglinde , Schnittger Susanne , Schoch Claudia , Jansen Mieke W.J.C. , van Dongen Jacques J.. M. , Pieters Rob , Haas Oskar , Strehl Sabine , Dingermann Theo , Klingebiel Thomas , Marschalek Rolf 
Published: October 2019

Abstract
Acute leukemias are frequently associated with specific chromosomal translocations of the human MLL gene. In general, MLL translocations define a distinct disease entity that needs to be diagnosed with precision to facilitate rapid clinical decisions. Here we present data about a new PCR based method that uses patient genomic DNA to identify any MLL fusion. Fourty different MLL translocations were successfully analyzed. We will present three novel MLL translocation partner genes and a new MLL deletion. The benefits of this novel technique for diagnosis and MRD analyses will be discussed. Supported by grant 2001.061.1 from the Wilhelm Sander foundation.
Long Range PCR Analysis for the t (8;14)(q24;q32) in Burkitts Lymphomas.
Authors: Mahlangu Johnny N. , Stevens Wendy S. , Stevens Gwyneth 
Published: October 2019

Abstract
Background: The breakpoints in the cytogenetic lesion in Burkitts lymphoma are very far apart such that the t(8,14)9q24;q32) is not amenable to standard polymerase chain reaction analysis. Long range polymerase chain reaction (LD-PCR) with its enzyme mix, appears to be capable of amplifying the t (8;14)(q24;32) in the majority of published sporadic Burkitts lymphoma analyses. The utility of t(8;14)(q24;q32) LD-PCR for routine use in the diagnosis of African and AIDS-related Burkitts lymphoma has not been studied. This study aims to analyze bone marrow of known African and AIDS-related Burkitts Lymphomas using LD-PCR and to establish if this procedure is suitable for routine diagnostic use.
Materials and methods: High molecular weight DNA was extracted from stored unstained bone marrow slides of previously diagnosed Burkitts lymphoma. Three hundred nanograms of patient and control DNA were amplified as per published LD-PCR methods. Each DNA sample was amplified with up to three pairs of MYC/IgH primer sets. The resulting amplicons were separated on a 0.8% agarose gel and their size compared to that of positive t(8;14)(q24;q32) controls, tPa controls and molecular weight markers.
Results: DNA was extracted from 74 Burkitts lymphoma bone marrow slides. Only 41 of these were amplifiable with the tPa primers and therefore suitable for further analysis. A t(8;14)(q24;q32) specific product was demonstrable in only 6 of 41 patients.
Conclusion: In this t(8;14)(q24;32) LD-PCR analysis of known Burkitts lymphomas comprising largely of African and AIDS-related variants, the procedure appears to be labour intensive and costly with a low diagnostic yield. These results may reflect the fundamental molecular differences between the sporadic and African or AIDS-related Burkitts lymphomas.
Differential Effects of Three Small Molecules Blocking Phosphatidylinositol-3 Kinase or AKT in Hodgkin Disease Cell Lines: Induction of Apoptosis and Cell Cycle Arrest.
Authors: Georgakis Georgios V. , Li Yang , Rassidakis Georgios Z. , Medeiros Jeffrey L. , Mills Gordon B. , Younes Anas 
Published: October 2019

Abstract
Several types of tumor cells utilize the phosphatidylinositol 3 Kinase (PI3K)/AKT pathway to support their growth and survival, and blockade of this pathway has been shown to have an antiproliferative effect in these cells. The significance of this survival pathway in Hodgkin disease (HD), and in particular, the neoplastic Hodgkin and Reed-Sternberg (H/RS) of Hodgkin disease (HD) is unknown. We studied routinely processed tissue sections of 42 cases of primary HD (n=42, 35 classical HD and 7 nodular lymphocyte predominant) for the expression of the active phosphorylated form of AKT. We also studied AKT expression in a panel of 4 well-characterized HD-derived cell lines (HD-MyZ, HD-LM2, L-428, and KM-H2). 27 of 42 (64.3%) cases of primary HD sections expressed phospho AKT (23/35 classical and 4/7 nodular lymphocyte predominant). Cultured H/RS cells also showed constitutive phosphorylation of AKT on Ser473. CD30 ligand (CD30L), CD40L and RANKL, increased the phosphorylation of AKT and downstream molecules as early as 1 hour. The effect of 3 small molecule inhibitors of PI3K/AKT pathway (PI3K inhibitor LY294002; AKT inhibitors QLT0394 and QLT0395 from QLT Inc, Vancouver, Canada) on cell proliferation and survival of cultured H/RS cells was examined by the MTS assay. The PI3K inhibitor LY294002 showed antiproliferative activity in a time and dose dependent manner in all cell lines tested. This antiproliferative effect was primarily due to cell cycle arrest in G0/G1 phase, as determined by propidium iodide staining, and to a less extent due to induction of apoptosis, as determined by the Annexin-V binding assay. However, when the AKT inhibitors QLT0394 and QLT0395 were used, they primarily induced apoptosis even at nanomolar doses. Apoptosis was mediated by caspase 8 activation as determined by western blot and was partially reversed by the pancaspase inhibitor ZVAD-FMK. Both AKT inhibitors alteredf AKT phosphorylation within 24 hours, and reduced phosphorylation of downstream molecules, such as 4E-BP1. Moreover, the AKT inhibitor QLT0395 showed downregulation of MDM2 and cyclin D1 within 24 hours, and increased ERK1/2 phosphorylation with subsequent decrease of total ERK1/2 levels at the same time frame, but had no effect in BCL-2, cFLIP or Bax. Importantly, both AKT inhibitors (QLT0394 and QLT0395) maintained their killing activity even in the presene of CD30L, CD40L, and RANKL. These preclinical data suggest that small molecules hitting different targets within PI3K/AKT pathway may have different effects on proliferation and apoptosis and that blockade of this pathway may be of therapeutic value in the treatment of patients with HD.
Escherichia Coli Expressing Recombinant WT1 Protein and Listeriolysin O Stimulate Antigen-Specific T Cells Following Vaccination in C57BL/6 Mice.
Authors: Nitcheu Josianne , Ramirez-Jimenez Francisco S. , Stauss Hans J. , Vassaux Georges 
Published: October 2019

Abstract
The Wilm’s Tumor (WT1) gene is expressed at high levels in most types of leukemia, but also in various types of solid tumors. We tested the efficacy of recombinant Escherichia Coli (E. Coli) expressing Listeriolysin O (LLO) as an antigen delivery vector for tumor specific immunotherapy targeting WT1 gene products. C57BL/6 (B6) mice were immunised with E. Coli expressing a form of WT1 protein truncated of 70 amino acids at the N-Terminus portion, and then challenged subcutaneously with 5X106 WT1-expressing TRAMP-C tumor cells. A significant inhibitory effect on TRAMP-C tumor growth was observed in vivo in vaccinated mice, with no signs of toxicity in tissues such as liver and kidney on histopathlogic examination. Furthermore, splenocytes from immunised mice were stimulated in vitro either with peptides that have been shown to contain major histocompatibility (MHC) class I binding anchor motifs (P126: RMFPNAPYL and P330: CNKRYFKL), or with irradiated TRAMP-C tumor cells. Specific CD8-positive cytotoxic T lymphocytes (CTLs) responses against TRAMP-C tumor cells were seen in cultures stimulated with irradiated tumor cells. However, no CTL activity was found in cultures stimulated with the relevant WT1 peptides that specifically lyse TRAMP-C tumor cells or the leukemia cell line RMA-S cells pulsed with each of the peptides. These results suggest that P126 and P330 might not be naturally processed WT1 epitopes for B6 mice and supports the possibility of other B6 CTLs epitopes. We are currently using a WT1 peptide library to screen immunised animals to look for novel peptides. Immunisation with E. Coli expressing WT1 protein but not LLO did not show any effect on TRAMP-C tumor growth in vivo. In this latter group, western blot analysis revealed WT1 specific antibodies directed against the N-terminus portion of the WT1 protein in the sera of immunised mice. Altogether the results presented here showed that E. Coli expressing WT1 could stimulated antigen-specific T cells dependant on LLO co-expression and suggest possible vaccine strategies for generating CD8+ T cell responses against tumors.