bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Incidence of Chronic Lymphocytic Leukemia in the General Population.
Authors: Frezzato Maurizio , Tosetto Alberto , Giaretta Ilaria , Rodeghiero Francesco 
Published: April 2020

Abstract
Background. Chronic Lymphocytic Leukemia is the most frequent leukemia in the Western countries (Rozman, 1995) but its incidence is not well established. Epidemiological data are based on reports from tumour registries, with an estimated incidence lower than 3-cases/100000 pts-year. Tumour registries can however underestimate the incidence of CLL as highlighted by the 37% higher incidence reported in a selected population of Veterans (Zent, 2000). We aimed to assessing the incidence of CLL in a population-based cohort from an epidemiological survey on familial thrombosis, (Tosetto, 2003) which was followed for at least 8 years.
Material and Methods. 15109 healthy people from 18 to 65 years of age in the Vicenza township were enrolled in the VITA Project (8017 females and 7092 males, with a median age of 43 years). A clinical history was collected and blood and DNA samples were withdrawn and stored at the time of enrolment. The study begun in 1993 and ended in 1996. Patients with a diagnosis of CLL (ICD-9 code 204) were traced using data from the Veneto Region Health Service database, which records all hospital admission for residents in the enrolment area. Moreover, the database encompasses data of all the subjects visited in the Vicenza Hematology Department, the unique hematological facility of the area, even if not subsequently admitted to the hospital. Diagnosis of CLL was validated if based on morphological and immunophenotypic data according to currently accepted criteria (Cheson, 1988; IWCC, 1989; Rozman, 1995).
Results. Six out 15109 people were diagnosed with a CLL (4 males). The median follow-up for the entire cohort was 8.2 years and the total time of exposure was 121000 pts-year. Incidence of CLL is estimated to be 4.9 cases/100000 pts-year (CI: 1.8–10.7). Diagnosis of CLL was done from 36 to 97 months after the enrolment. Mean age at diagnosis was 66 years (61 – 70). At the time of enrolment in the study, all of the patients had a normal WBC. Three persons presented an immunoglobulin VH gene rearrangement, 32, 54 and 57 months before diagnosis. All of the patients were asymptomatic at diagnosis. The median follow-up of all the subjects was 28 months (7 – 57 months). Only one of them became symptomatic and needed a treatment, 40 months after the diagnosis.
Conclusions. According to our data, the incidence of CLL appears to be higher than assessed so far. In our opinion it’s noteworthy that the estimation is based on data of subjects evaluated in a clinical setting and not from tumour or disease registries. In fact it could be still underestimated comparing the young median age of our cohort with that usually reported at diagnosis (Rozman, 1995). Moreover, the comparison of our data with those available could suggest that only a quote of the cases might be usually diagnosed. Finally, the presence of the Ig VH gene rearrangement at the time of the enrolment suggests that the disease may be present a very long time before any clinical evidence.
Final Results of the Thai-NHLBI Epidemiological Study of Aplastic Anemia.
Authors: Issaragrisil Surapol , Kaufman David W. , Anderson Theresa E. , Chansung Kanchana , Leaverton Paul , Shapiro Samuel , Young Neal S. 
Published: April 2020

Abstract
Aplastic anemia, rare in the West, is a relatively more frequent hematologic diagnosis in Asia. A systematic epidemiological study of aplastic anemia was conducted in Thailand from 1989–2002 in order to determine both the incidence and environmental risk factors. We employed the methodology established by the International Agranulocytosis and Aplastic Anemia Study, conducted in Europe and Israel in the 1980s. In the first phase of the current study, we documented an annual incidence of 3.9 per million for the Bangkok metropolitan area and 5 per million in the northeast region of Khonkaen. Aplastic anemia therefore occurs at a rate at least 2–3 fold more frequent in Thailand than in Europe and Israel. We now report final results of the case-control portion of the study, in which 541 patients and 2,261 hospital controls were enrolled--the largest epidemiologic study of aplastic anemia to date. Environmental exposures were determined with a detailed personal interview, with emphasis on both known risk factors for aplastic anemia and novel factors that might provide clues to disease causation. Among previously suspected factors, we observed significantly elevated relative risk estimates for benzene (3.5) and other solvents (mostly unspecified thinners; 2.0), and among medical drugs for sulfonamides (5.6), thiazides (3.8), and mebendazole (3.0). Chloramphenicol use was infrequent among cases and controls, and no significant association was observed. A number of agricultural pesticides were implicated in Khonkaen: there were significant associations with organophosphates (2.1), DDT (6.7), and carbamates (7.4), and an association of borderline significance with paraquat (2.3). Among novel factors, we found significant risks for farmers and their family members who had been exposed to ducks and geese (3.7) and a borderline association with the use of animal fertilizer (2.1). There was a significant association confined to Khonkaen with drinking other than bottled and distilled water (2.8). Nonmedical needle exposure through tattooing, body piercing, or acupuncture was infrequent but associated in Bangkok and Khonkaen combined (3.8). Only two cases consistent with a diagnosis of post-hepatitis aplastic anemia were observed. Our results confirm the well known link between aplastic anemia and benzene, other solvents, and some medical drugs, but only a small proportion of cases were exposed to these agents. A variety of pesticides were implicated and these could account for up to 15% of cases in Khonkaen. Most striking was the large etiologic fraction of aplastic anemia in that region accounted for by animal exposures and especially drinking of water from sources such as wells, rural taps, and rainwater. These findings are consistent with an infectious etiology for many cases of aplastic anemia in rural Thailand.
PIG-A Mutations in Normal Hematopoiesis.
Authors: Hu Rong , Mukhina Galina , Piantadosi Steven , Jones Richard J. , Brodsky Robert A. 
Published: April 2020

Abstract
Introduction: Paroxysmal nocturnal hemoglobinuria (PNH) is caused by somatic mutations of the X-linked gene, PIG-A, in hematopoietic stem cells (HSCs). The product of this gene is necessary for the assembly of glycosylphosphatidylinositol (GPI) anchors. Consequently, PNH cells lack the expression of GPI-anchored proteins on their cell surface. PIG-A mutations have been found in granulocytes and T lymphocytes from most normal individuals. Although the significance of these mutations is unclear, it suggests that they are important in the pathogenesis of PNH.
Methods: We isolated CD34+ progenitors from 4 PNH patients, 18 healthy donors, and 9 non-PNH patients undergoing peripheral blood stem cell tranplantation. The frequency of PIG-A mutant progenitors was determined by assaying for colony forming cells (CFC) in methylcellulose containing toxic doses of aerolysin. Aerolysin is a pore-forming toxin that uses the GPI anchor as it receptor; hence, PNH cells are unique in their resistance to aerolysin. DNA was extracted from individual day 14 aerolysin resistant CFC and the PIG-A gene was sequenced to determine clonality. We performed a Poisson distribution of the mutational frequency to determine the probability that the PIG-A mutation in controls arose from HSC.
Results: In PNH patients, 67% of the CFC were aerolysin resistant. The frequency of aerolysin resistant CFC was 14.7 ± 4.0 x 10−6 in the bone marrow of healthy donors and was 57.0 ± 6.7 x 10−6 from mobilized peripheral blood. Aerolysin resistant CFC from PNH patients exhibited clonalPIG-A mutations, and thus, arose from HSC. In contrast, PIG-A mutations in the CFC from controls were polyclonal (up to 15 different mutations from one individual). Recent evidence suggests that humans and other large mammals possess only 10,000 primitive HSC, and that only 1000 of these cells are thought to contribute to hematopoiesis at any one time. Poisson statistics show that only 5% of normals would be expected to harbor a PIG-A mutation in 1000 HSC, and &lt; 1 x 10−9 persons would harbor 10 or more different PIG-A mutations even if all 10,000 hematopoietic stem cells were contributing to hematopoiesis. Thus, the high frequency of PIG-A mutations in controls, coupled with their polyclonality, suggests that they do not arise at the level of HSC; rather, PIG-A mutations in normals appear to arise as a consequence of hematopoietic differentiation, between the level of an HSC and a CFC.
Conclusion: Our data confirm the findings that PIG-A mutations are relatively common in normal hematopoiesis. Although we cannot rule out that a rare PIG-A mutant blood cell in normals does in fact arise from a mutant HSC, our data suggest that most of the mutations occur with differentiation. Genetic fidelity can be lost with differentiation without consequence, as mutations in differentiated cells would not be propagated. These data also call into question the relevance of PIG-A mutations in normals to the pathogenesis of PNH.
Nuclear Localization of Fanconi Anemia Protein FANCA Is Regulated by Hsc70/Hsp90 Chaperone Machinery.
Authors: Oda Tsukasa , Miyaso Hidenobu , Yamashita Takayuki 
Published: April 2020

Abstract
Fanconi anemia (FA) is a genetic disorder characterized by bone marrow failure, cancer susceptibility and cellular hypersensitivity to DNA crosslinkers such as mitomycin C (MMC). Current evidence indicates that formation of a nuclear multiprotein complex (core complex) including six FA proteins FANCA/C/E/F/G/L is essential for FANCL/PHF9 ubiquitin ligase-mediated activation of FANCD2 into a monoubiquinated form, which participates in BRCA1 and FANCD1/BRCA2-mediated DNA repair (the FA/BRCA pathway). Subcellular distribution of FANCA plays a crucial role in the regulation of the FA/BRCA pathway. However, the underlying molecular mechanisms are not fully understood. To address this issue, we tried to identify FANCA-associated proteins. To this end, Flag-FANCA ectopically expressed in HeLa cells was immunopurified from the cytoplasmic fraction, using anti-Flag antibody-conjugated sepharose beads. Analysis of the immune complex on SDS polyacrylamide gel electrophoresis revealed that several proteins of Mr. 60–70 kD specifically associated with Flag-FANCA. These proteins were identified as FANCG and Hsc (heat shock cognate protein) 70 by LC-MS/MS. Immunoblot analysis showed that FANCA associated with Hsp90 as well as Hsc70. Hsc70 is an ATP-dependent molecular chaperone highly homologous to Hsp70 and often cooperates with Hsp90 to form a chaperone machinery involved in the regulation of diverse protein functions. Patient-derived FANCA mutants failed to bind FANCC but associated with larger amounts of Hsc70 than wt-FANCA, indicating that the interaction between FANCA and Hsc70 is not mediated by FANCC, as suggested by previous observations of the interaction of FANCC with Hsp70. To study the role of Hsc70 and Hsp90 in the regulation of FANCA, we examined effects of a dominant-negative (dn) form of Hsc70 with inactivated ATPase activity, and a specific inhibitor of Hsp90, 17-AAG (a geldanamycin analog). Overexpression of dn-Hsc70 inhibited nuclear localization of FANCA and inhibited its core complex formation, whereas wt-Hsc70 did not. 17-AAG induced cytoplasmic distribution and proteosomal degradation of FANCA and suppressed FANCD2 mono-ubiquitination. Taken together, these results suggest that Hsc70/Hsp90 chaperone machinery interacts with FANCA and regulates its subcellular distribution and stability, thereby controlling activation of the FA/BRCA pathway.
Ex Vivo Expansion of Frozen Cord Blood Products without Selection.
Authors: McNiece Ian K. , Harrington Jenny , Kellner Joshua , Turney Jennifer , Shpall Elizabeth J. 
Published: April 2020

Abstract
Ex vivo expansion of cord blood products (CB) has been proposed as an approach to increase the number of cells available from a single CB unit. We and others have reported the requirement of CD34 selection for optimal expansion of CB products, however, the selection of frozen CB products results in significant losses of CD34+ cells with a median recovery of 43% (range 6 to 203%, N=40) and low purities resulting in decreased expansion. Therefore we explored approaches to expand CB without prior selection and have described the use of co-culture of CB mononuclear cells (MNC) on mesenchymal stem cells (MSC). In the present study we have evaluated the expansion of clinical CB products (provided by Duke University CB Bank CB). MNC were obtained after ficol separation of RBCs and 10% of the CB product was cultured on preformed layers of MSC in T150 flasks containing 50ml of defined media (Sigma Aldrich) plus 100 ng/ml each of rhSCF, rhG-CSF and rhTpo. After 6 days of culture, the non adherent cells were transferred to a Teflon bag and a further 50 ml of media and GFs added to the flask. Again at day 10, non adherent cells were transferred to the Teflon bag and media and growth factors replaced. At day 12 to 13 of incubation the cells were harvested, washed and total nucleated cell (TNC) counts and progenitor assays performed. In three separate experiments we have achieved greater than 20 fold expansion of TNC with a median of 22, and a median expansion of GM-CFC of 37 fold. Morphologic analysis demonstrated the expanded cells contained high levels of mature neutrophils and neutrophil precursors. In vivo studies in NOD/SCID mice also demonstrated that the expanded cells maintained in vivo engraftment potential. Clinical studies are being designed to evaluate the in vivo potential of CB MNC products expanded on MSC.
Recombinant TAT-HOXB4 Protein Promotes Ex Vivo Expansion of Primitive Human Hematopoietic Cells.
Authors: Krosl Gorazd , Giard Marie-Pier , Krosl Jana , Humphries R. Keith , Sauvageau Guy , Roy Denis-Claude 
Published: April 2020

Abstract
The clinical application of therapeutic protocols depending on hematopoietic stem cell (HSC) transplantation for long term reconstitution with donor-derived HSCs, particularly in patients previously exposed to intensive radiation or chemo-therapy, or when grafts are purged of infiltrating malignant or alloreactive T cells, can be severely hampered by limited numbers of HSCs in the graft. In mouse bone marrow transplantation models, engineered overexpression of HOXB4 has been one of the most potent stimulator of HSC expansion identified to date. The simple addition of soluble recombinant TAT-HOXB4 protein was also recently reported to enable rapid in vitro expansion of mouse HSCs that retain their in vivo proliferation and differentiation capacity. To test the feasibility of using TAT-HOXB4 as a stimulator of human HSC expansion, we performed a series of experiments using CD34+ populations isolated from healthy volunteers. The CD34+ cell populations were cultured in X-Vivo medium supplemented with Stem Cell Factor (300 ng/mL) and G-CSF (50 ng/mL) in the presence or absence of TAT-HOXB4 protein (50 nmol/L) for 4 days. In response to TAT-HOXB4, total numbers of mononuclear cells demonstrated a modest but distinct 2-fold increase compared to controls. TAT-HOXB4 treatment had the largest proliferation enhancing effect on more primitive cell populations such as CFU-GEMM, BFU-E and BFU-Meg, whose numbers increased 26.5 ± 1.4 fold (mean±S.D.), 2.2 ± 0.7 fold and 2.1 ± 0.2 fold, respectively, over their input values, and 19.1 ± 1.3 fold, 2.7 ± 0.7 and 31 ± 3.4 fold, respectively, compared to growth factor only controls. In response to TAT-HOXB4, the total numbers of CD34+CD38-Lin- cells increased 2.1 ± 0.7 fold above their starting numbers compared to a 1.5 ± 0.5 fold loss of this population in control cultures. HSC numbers were enumerated at the beginning, and after a 4-day TAT-HOXB4 treatment period using a NOD/SCID repopulation assay. In response to 50 nM TAT-HOXB4, NOD/SCID repopulating cell (SRC) numbers increased ~2-fold over their input values, compared to a 9-fold loss in control cultures without TAT-HOXB4. These results show that recombinant TAT-HOXB4 protein has the capacity to rapidly induce ex vivo expansion of primitive human bone marrow populations, and suggest that optimization of treatment conditions will rapidly lead to clinically useful expansion of human HSCs.
Phase II Study of a Single Pegfilgrastim Injection as Adjunct to Chemotherapy to Mobilize Stem Cells into Peripheral Blood of Resistant/Relapsed Lymphoma Patients.
Authors: Lemoli Roberto M. , Isidori Alessandro , Tani Monica , Bonifazi Francesca , Zinzani Pier Luigi , Curti Antonio , Motta Maria Rosa , Rizzi Simonetta , Giudice Valeria , Farese Oriana , Rovito Manuela , Alinari Lapo , Conte Roberto , Baccarani Michele 
Published: April 2020

Abstract
The primary end point of the study was the successful mobilization of a target cell dose of 2 x 106 CD34+ cells/kg in lymphoma patients receiving ifosfamide, epirubicin and etoposide (IEV) chemotherapy and a fixed dose (6 mg) of pegfilgrastim given as single subcutaneous injection. An open-label phase II study including 25 relapsed or refractory patients (Hodgkin’s disease=4; aggressive non-Hodgkin’s lymphoma=21) was conducted to evaluate the efficacy of pegfilgrastim, in combination with salvage chemotherapy, mobilizing CD34+ stem cells into peripheral blood. Following chemotherapy, all patients had grade 4 neutropenia with a median duration of 1.5 days (1–3). Pegfilgrastim treatment was well tolerated and only 2/25 patients required pain-control medication. CD34+ cells were mobilized in all patients. The median (range) peak value of peripheral blood CD34+cells after IEV chemotherapy and pegfilgrastim was 141/microL (12.8–386) and occurred almost invariably on day +14 (13–16). Twenty three/25 patients underwent a single apheresis to collect a median of 8.7 CD34+cells/Kg (1.8–17.3). Twenty four/25 patients (96%) reached the target cell dose of 2 x 106 CD34+ cells/kg. High concentrations of circulating CD34+ cells (&gt; 50/microL) were observed for several days after the achievement of the peak value. All patients have been transplanted with pegfilgrastim-mobilized CD34+ cells and all of them showed a rapid and sustained engraftment after high-dose chemotherapy. Our results show that pegfilgrastim as adjunct to chemotherapy is a predictable and highly effective mobilization regimen in lymphoma patients
Isolation of CD56+ Natural Killer Cells Using the CliniMACS® MARRS Tubing Set, a Newly Developed Tubing Set Including a Cross-Flow Filtration Module for Removal of Unbound Reagent.
Authors: Huppert Volker , Wolf Inka , Duwendag Julia , Arendt Andreas , Schmitz Jurgen 
Published: April 2020

Abstract
Use of Natural Killer (NK) cells in cellular immunotherapy requires processing of large numbers of NK cells in a closed system consisting of clinical grade components. We have previously developed a two step NK cell isolation process in which the CliniMACS® plus Instrument and CliniMACS® CD3 and CD56 Reagents are used for CD3+ cell depletion and subsequent CD56+ cell enrichment, respectively. We have recently optimized the CD3 depletion step for very large scale cell processing by development of the CliniMACS® Depletion Tubing Set (“DTS”) and appropriate cell separation software, reducing the processing time of the CD3 depletion step by 50%. We now have developed the CliniMACS® MARRS Tubing Set (MARRS = MAgnetic Reagent Removal System), a tubing set which includes a cross-flow filtration module for removal of unbound magnetic reagent after magnetic labelling. The cross-flow filtration module allows to directly connect a magnetically labelled cell product to the tubing set without centrifugational washing steps, reducing the processing time by 45 minutes. We isolated CD56+ cells from buffy coats as well as apheresis harvests using the CliniMACS® MARRS Tubing Set as compared to the standard CliniMACS® Tubing Set. Both tubing sets (MARRS vs. standard) yielded CD56+ cells of comparable purity (94.1% vs. 92.3%), comparable depletion of CD56−CD3+ T cells (3.2 log vs. 3.0 log) and comparable recovery (72.9% vs. 76.7%) with a trend towards better performance of the CliniMACS® MARRS Tubing Set. To assess the functionality of CliniMACS® separated CD56+ cells we performed flow cytometric cytotoxicity tests against the MHC class I negative target cell line K562. Specific killing could be increased from &lt;2% (unseparated sample) to 47% (MARRS) and 53% (standard) at an effector to target ratio of 50:1. Thus, K562 killing cells are efficiently enriched and cytotoxicity is similar for both CD56+ cells isolated using the standard CliniMACS® Tubing Set and the CliniMACS® MARRS Tubing Set. We additionally assessed whether the killing efficiency of CD56+ cells correlates with NK receptor expression (NCR, KIR, NKG2D, costimulatory molecules) on different NK and NKT subsets. Results of these analyses will be presented. We conclude that the CliniMACS® MARRS Tubing Set can be used to efficiently isolate functional CD56+ cells from buffy coats or medium scale apheresis harvests (e.g. non-mobilized donors), clearly reducing total processing time. For larger scale NK cell selection using apheresis harvests (e.g. mobilized donors) the combination of Depletion Tubing Set for CD3+ cell depletion and the standard CliniMACS® Tubing Set for CD56+ cell enrichment is suitable.