bluuebunny/crossref_metadata_2025_split huggingface dataset:
A Combined Evaluation of Severity, Presentation and Duration of Chronic GvHD Predicts Transplant Outcome Among Patients with a Hematological Malignancy Undergoing Allogeneic Stem Cell Transplantation
Authors: Crocchiolo Roberto , Saillard Colombe , Furst Sabine , El-Cheikh Jean , Castagna Luca , Faucher Catherine , Oudin Claire , Harbi Samia , Granata Angela , Chabannon Christian , Blaise Didier 
Published: October 2019

Abstract
Abstract 4549

Introduction:
chronic graft-versus-host disease (cGvHD) after allogeneic hematopoietic stem cell transplantation (HSCT) for hematological malignancies is known to be associated with lower relapse risk, due to graft-versus-tumor effect. On the other hand, severe, extensive form of cGvHD is associated with high transplant-related mortality (TRM), mainly due to infectious complications as a consequence of the immunosuppressive treatment (IS) of cGvHD. Not only severity of cGvHD but also its presentation and duration of IS is responsible for deleterious effects on transplanted patients.
Here we built and evaluated a score taking into account both severity and duration of cGvHD (and related IS), looking for any association with TRM, OS and relapse/progression after HSCT.

Patients and Methods:
study population is represented by adult patients who received HSCT after fludarabine-busulfan-ATG conditioning between May 1999 and June 2010 at out Institution for a hematological malignancy, and who developed cGvHD. cGvHD was retrospectively classified according to NIH criteria into mild, moderate and severe form. Informations on cGvHD presentation (progressive, quiescent or de novo) were also collected. Duration of IS was taken into account as well as any change in cGvHD severity overtime. A combined mathematical function D was built, taking into account severity, type of presentation and duration of IS (appendix 1), and was treated as a covariate in univariate and multivariate Cox models for TRM, OS and relapse/progression.

Results:
on 313 transplanted patients transplanted in the above mentioned period, 130 developed cGvHD and represented our study population. The function D showed to predict TRM and OS both in univariate and multivariate analysis (table 1); a borderline association was found with relapse/progression, in univariate analysis.

Conclusion:
a function taking into account not only severity but also presentation of cGvHD and duration of IS seems to be predictive of TRM and OS after HSCT. Although further validation is needed, the analytical approach we used here allows to better describe the complexity of post-transplant reality, where severity and duration of IS can change overtime and are associated with patients' outcome.

Disclosures:
No relevant conflicts of interest to declare.
Pattern of Mir-155 and PU.1 Expression in CLL/SLL and Aggressive Lymphomas
Authors: Stopka Tomas , Vargova Karin , Huskova Hana , Burda Pavel , Curik Nikola , Vlckova Petra , Kulvait Vojtech , Pospisil Vit , Savvulidi Filipp , Necas Emanuel , Mraz Marek , Pospisilova Sarka , Mayer Jiri , Karban Josef , Obrtlikova Petra , Trneny Marek 
Published: October 2019

Abstract
Abstract 4600
MicroRNA miR-155 represents a candidate pathogenic factor in chronic lymphocytic leukemia (CLL) and lymphomas (Eis PS, et al. 2005, Fulci V, et al. 2007, Calin GA, et al. 2005, Kluiver J, et al. 2005, Metzler M, et al. 2004, van den Berg A, et al. 2003., Vargova K. et al 2011). In diffuse large B-cell lymphoma (DLBCL) expression of miR-155 was associated with NFkB pathway (Rai et al., 2008). In addition, increased miR-155 levels were associated with more aggressive (Activated B-cell like) DLBCL (Eis et al. 2005). The targets of up-regulated miR-155 apparently include a key hematopoietic transcription factor PU.1. Down-regulation of PU.1 represents an important and critical step for both myeloid as well as lymphoid tumorigenesis. We herein found the increased levels of miR-155 coupled with downregulation of its target PU.1 are frequent events in B cells of a vast majority of CLL patients (N=169). To advance understanding of miR-155 and PU.1 in lymphoid malignancies we studied lymph node biopsies derived from patients with lymphomas. We show that increased miR-155 levels are also found in DLBCL (N=31, P < 0.0001), Hodgkin lymphoma (HL, N=22, P < 0.0001), follicular lymphoma (FL, N=23, P < 0.001), small lymphocytic lymphoma (SLL, N=12, P < 0.01), and marginal zone lymphoma (MZL, N=11, P < 0.01). The miR-155-overexpressing tumors display downregulation of PU.1. In contrast, peripheral T cell lymphoma (T-NHL, N=6) and mantle cell lymphoma (MCL, N=7) expressed miR-155 and PU.1 comparably as control lymph nodes. Our data indicate that expression pattern of miR-155 and its target PU.1 represents a hallmark of some but not all lymphoid malignancies. Interestingly, the miR-155/PU1 levels vary substantially within each diagnosis, therefore, our aim is to further analyze the occurrence of this relationship in different prognostic groups and analyze the clinical outcome as well. (Grants: NT10310-3/2009 & NT11218-6/2010, MPO FR-TI2/509, GAUK251135 82210 & 251070 45410, SVV-2011-262507)

Disclosures:
No relevant conflicts of interest to declare.
The Expression of BAG3 Gene in Chronic Lymphocytic Leukemia and Its Clinical Significance
Authors: Zhu Huayuan , Liu Peng , Li Jianyong 
Published: October 2019

Abstract
Abstract 4605

Objective:
BAG3 (BCL2-associated athanogene 3) is a member of BAG family. Previously, many reports indicated that BAG3 was over-expressed in different human cancer tissues, such as thyroid carcinoma, pancreatic cancer, prostate cancer, and leukemic cells. However, there is no or low expression of BAG3 in normal tissues. These imply that BAG3 maybe have some important roles in the human cancers. This study was aimed to investigate the expression level of BAG3 mRNA in chronic lymphocytic leukemia (CLL) and its significance in evaluation of CLL prognosis.

Methods:
100 CLL human patients and 20 healthy controls were enrolled in this study. The BAG3 mRNA expressions were measured by real-time PCR with fluorescent dye SYBR Green I, the β-actin was used as internal control. The relative quantitative value of BAG3 expression was calculated by means of 2 (–ΔCt). We analyzed the results with established CLL prognostic factors, overall survival (OS) and treatment-free survival (TFS).

Results:
The expression of BAG3 mRNA in 100 CLL patients was significantly higher than 20 normal control (p = 0.038). Furthermore, The BAG3 expression was obviously increased in CD38 positive patients CD38 (n = 31) than CD38 negetive patients it (n = 61, p = 0.0238). We also found that the expression of BAG3 in the patients with unmutated immunoglobulin heavy-chain (n = 29) was markedly higher than those with mutated immunoglobulin heavy-chain (IgHV) (n = 62, p = 0.0326). It is also found that the patients with enlarged lymph nodes (n = 24) have higher BAG3 expression, compared with those without enlarged lymph nodes (n = 76, p = 0.0004). The increased BAG3 level can also be found in the patients with splenomegaly (n = 32),compared with those without splenomegaly (n=68, p = 0.0393). No association was found between BAG3 expression and patient clinical baseline information (gender and age), as well as other established prognostic factors (lymphocyte count, cytogenetics analysis and p53 mutation status). 25 of these patients were treated with fludarabine-based therapy, interestingly, BAG3 expression level was significantly increased in patients who were relapse and/or refractory to fludarabine-based treatment (n = 10, p = 0.019). However, there are no differences in OS and TFS between patients with low BAG3 expressions and high expression, which indicated BAG3 didn’t affect the patients’ OS and TFS.

Conclusion:
These results showed that the BAG3 expression in CLL patients is markedly higher than normal controls. CD38, mutation of IgHV, lymph nodes and splenomegaly are related to the expression of BAG3 in CLL. Furthermore, the patients, who were relapse and/or refractory to fludarabine, have a higher expression of BAG3. These imply that BAG3 may be involved in the pathogenesis of CLL and the roles of BAG3 in CLL need further investigation.

Disclosures:
No relevant conflicts of interest to declare.
Usability Testing of An Online Transition Program for Adolescents with Hemophilia
Authors: Breakey Vicky R. , Warias Ashley , Ignas Danial M , Blanchette Victor S. , Stinson Jennifer 
Published: October 2019

Abstract
Abstract 4777

Purpose:
This study explored the usability of a new Internet-based educational program of disease-specific information, self-management strategies and social support for adolescents with hemophilia.

Methods:
A comprehensive eight-module educational website was developed, based on results of an in-depth needs assessment. The website was tested for usability, using qualitative methods that included semi-structured, audio taped interviews and observation by a trained observer. To determine the usability and intuitiveness of the user interface of the “Teens Taking Charge: Managing Hemophilia Online” intervention, testing occurred in three cycles (4 participants per cycle). Participants were asked to work through standardized parts/features of the program, with changes to the prototype made after each cycle. Thematic analysis using a collaborative and iterative process was used to organize data into categories that reflected the emerging themes.

Results:
A purposive sample of twelve adolescents (range 12–18 years, mean 15.4 years) was recruited from a Canadian tertiary care center. All of the participants had access to a computer at home and felt comfortable using the Internet. Teens responded positively to the content, appearance and theme of the website (promoting self-management in youth with hemophilia). Subjects thought that it was easy to navigate, use and understand. Overall, they felt the content was appropriate and geared to meet the unique needs of adolescents with hemophilia. The multi-media components (videos, animations and quizzes) were thought to enrich the experience and make the program appealing. Adolescents provided ideas on how the website user-interface could be improved in terms of its usability (navigation, format and layout). Minor changes to the website user-interface were made and tested after the first and second cycles of testing. No further problems were identified in final cycle of testing. At the teens' suggestion, additional social media elements were added (discussion board, “ask the expert” section) to build in elements of support and increase interactivity. Most participants felt this program would be helpful prior to transition of care and beyond.

Conclusions:
Usability testing was the crucial first step in ensuring the acceptability and ease of use of “Teens Taking Charge: Managing Hemophilia Online”. Findings from this study were used to refine the website prototype. A pilot study is underway to determine the feasibility of using a randomized controlled trial to study the Internet-based self-management program.

Disclosures:
Breakey: Baxter Bioscience: Research Funding, Salary Support. Ignas:Baxter Bioscience: Salary Support.
γ-IFN May Act As a Positive Regulator to Hematogenesis Through PI3K/Akt and JAK2/STAT5 Pathways
Authors: Liu Hong , Zhou Lu , Wang Han 
Published: October 2019

Abstract
Abstract 4785
It is known to us all that γ-IFN is a growth inhibitory to hematogenesis and plays a critical role in the pathophysiology of aplastic anemia(AA). However, γ-IFN actually is a bilateral regulatory factor. In certain experimental situations, it can facilitate the proliferation and growth of hematopoitic stem cells. Interferon regulatory factor-1 (IRF-1) and IRF-2 are the key regulators of IFN system and IRF-1 has been shown to participate in development of CD8+ T cells, Th1 differentiation, and NK cell development. Thus, IRF-1 may has crucial functions against hematopoietic stem and/or progenitors cells. To investigate the function of IRF-1and the molecular mechanism of proliferative effect of γ-IFN, IRF-1-siRNA was designed to knocked out IRF-1 gene in 32D cells, mouse multipotent progenitor cells, and the apoptosis was assayed by flow cytometry 48 hr after incubation with γ-IFN in three different concentrations. It showed that after IRF-1 gene was knocked out, γ-IFN reversed its action on hematogenesis and apoptosis,it potentiated the proliferation and growth of 32D cells. Interestingly, at this situations, the inhibitory effect of γ-IFN on Akt activation was abolished completely and reversed to enhanced Akt phosphorylation significantly. The expression of pStat5 also changed when γ-IFN only exert proliferative effect.Nevertheless,The negative effect of γ-IFN on Erk1/2 activation was not blocked after IRF-1 gene was knocked out. Taken together, γ-IFN can enhance or antagonize cell proliferation depending on the function of IRF-1 and may act as a positive regulator to hematogenesis through PI3K/Akt and JAK2/STAT5 signaling pathways. Our findings may contribute to understanding the decreased number of stem cells characteristic of aplastic anemia and will provide novel therapeutic strategies for this disease.

Disclosures:
No relevant conflicts of interest to declare.
Expression of PTH in Bone Marrow Stem Cells in Adult SD Rats
Authors: Zou Weiying , Yang Bei , Wu Lei , Chen Ting , Zhang Dalei , Xiong Mingdi , Chen Jiaxiang , Zou Ting 
Published: October 2019

Abstract
Abstract 4817

Introduction:
Parathyroid hormone (PTH) is a major regulator of calcium and phosphate metabolism and is secreted by the cells of the parathyroid gland. Recent research showed that PTH treatment may regulate the hematopoietic stem cell niche, leading to beneficial effects on the HSC pool, and increasing stem cell number and the number of stem cells mobilized into the bloodstream. Many studies demonstrated that the serum PTH was decreased but maintained at the lower level suggesting persisting PTH secretion after total parathyroidectomy without autotransplantation. Some research showed that the thymus is another source of PTH. Can the bone marrow cells express PTH mRNA and release PTH and be a third source of PTH production? The aim of this study was to explore whether PTH could be expressed in bone marrow stem cells(BMSCs).

Method:
BMSCs were separated from adult male SD rat bone marrow and were cultured in DMEM supplemented with 10% fetal bovine serum. The total RNA was extracted using Trizol one-step method. RT-PCR was performed to examine the expression of PTH and GAPDH in BMSCs. The optical densities (ODs) value of RT-PCR product was measured using the Gel Imageware System. The signal intensity of PTH bands was normalized by corresponding GAPDH bands. The ODs in each sample were expressed as a ratio PTH over GAPDH densitometric intensities. The resultant PCR product of PTH was sequenced in both directions. Immunocytochemistry staining was utilized to examine the expression of PTH in the BMSCs from adherent cell fraction of the 2nd passage.

Results:
The expressions of PTH mRNA in BMSCs were detected in all 11 rats by RT-PCR. The PCR product was 193bp for PTH (Fig 1). The ratio of PTH over GAPDH ODs value ranged from 0.4407 to 3.1506, and the average ODs value (m ± SD) was 1.2617 ± 0.8953. The sequence of the PTH PCR products matched 100% with the PTH sequence in Genebank (Gene ID: 24694 Pth; gb K01268.1 RATPTH3, Rat parathyroid hormone gene, exons II and III). Some of the BMSCs were PTH positive by immunocytochemistry. The PTH positive cells were middle in size and spindle. PTH staining was localized in the cytoplasma and cell membrane of BMSCs cultured (Fig 2).

Conclusion:
The PTH mRNA and protein were detected in BMSCs. The results suggest that the BMSCs may be a third cell source responsible for the synthesis and secretion of PTH. It would be interesting to know what type of cells in BMSCs is secreating the PTH and to explore the role of PTH in bone marrow stem cell niche.

Disclosures:
No relevant conflicts of interest to declare.
Chromosomal Instability in Cultured Mouse Hematopoietic Cells Associated with Oxidative Stress
Authors: Liu Alice M. , Qu William W. , Liu Xia 
Published: October 2019

Abstract
Abstract 4819
Hematopoietic stem cells (HSCs) that give rise to all blood cell types are important vehicles for cell-based and gene therapies. After isolation from the bone marrow, HSCs are often cultured in laboratory settings for purposes of ex vivo expansion, gene transduction, and bone marrow transplantation for the treatment of various disorders of the blood and immune systems. While undergoing proliferation and differentiation in vitro, test tube and dish culturing can potentially induce genomic instability in HSCs due to prolonged culturing periods or the exposure to increased levels of oxygen. Here we demonstrate that in vitro culturing outside their bone marrow niches, HSCs may change even under very short durations of time. Lineage− Scal-1+ c-Kit+ (LSK) cells that are enriched with HSCs revealed significant levels of genomic instability in culture, as evidenced by the emergence of aneuploidy cells. To further determine the effects of in vitro culturing conditions, whole bone marrow cells were cultured in a hypoxic environment of 2–3% oxygen, mimicking conditions inside the body's bone marrow. In this case, cells proved to undergo less genetic alterations. Proper dosages of the antioxidant N-Acetyl-Cysteine (NAC) similarly decreased occurrences of chromosomal changes. Furthermore, in vitro normoxic culture-induced chromosomal instability was enhanced in aged hematopoietic cells compared to that in young hematopoietic cells due to noted increased oxidative stress in aged cells. These results reveal that in vitro cell culturing does indeed cause genomic instability in hematopoietic cells. Reduced oxygen levels and additions of antioxidants can be employed as a possible agent to lower oxidative stress and decrease chances of transformation. Additionally, since hematopoietic cells are commonly developed in laboratory settings before transplantation for patient treatment, our findings raise a concern for using cultured hematopoietic cells for therapeutic purposes.
Note: Alice Liu and William Qu contributed equally to this work.

Disclosures:
No relevant conflicts of interest to declare.
TIF1γ Knockdown Enhances Hematopoietic Stem Cell Self Renewal with Preferential Myeloid Differentiation and Delayed Erythropoiesis
Authors: Dorn David C , He Wei , Massague Joan , Moore Malcolm A.S. 
Published: October 2019

Abstract
Abstract 4829
The role of TIF1γ in hematopoiesis is still incompletely understood. We previously identified TIF1γ as a novel binding factor for Smad2/3 in the Transforming Growth Factor-β (TFGβ)-inducible signaling pathway implicated in the enhancement of erythropoiesis. To investigate the function of TIF1γ in regulation of hematopoietic stem cells we abrogated TIF1γ signaling by shRNA gamma-retroviral gene transfer in human umbilical cord blood-derived CD34+ hematopoietic stem/ progenitor cells (HCS/ HPCs). Upon blocking TIF1γ the self-renewal capacity of HSCs was enhanced two-fold in vitro as measured by week 5 CAFC assay and three-fold in vivo as measured by competitive engraftment in NOD/ SCID mice over controls. This was associated with a delay in erythroid differentiation and enhanced myelopoiesis. These changes were predominantly observed after TIF1γ knockdown and only mildly after Smad2 depletion but not after Smad3 or 4 reduction. Our data
reveal a role for TIF1γ-mediated signaling in the regulation of HSC self-renewal and differentiation.

Disclosures:
No relevant conflicts of interest to declare.
Optimizing Hydroxyurea Use in Children with Sickle Cell Disease: Least Tolerated Dose Is Effective
Authors: Sharef Sharef Waadallah , Al-Hajri Maya , Beshlawi Ismail , Al-Shahrabally Amer Qais , Wali Yasser 
Published: October 2019

Abstract
Abstract 4851

Background:
Hydroxyurea (HU) is an antimetabolite that effectively ameliorates the course of sickle cell disease (SCD). Starting dose is usually 20 mg/kg/day and the dose is escalated up to 35 mg/kg/day in most of the centers. In Oman, due to high level of consanguinity, ethnic neutropenia affects around 10% of population rendering higher doses difficult to maintain in most of the patients.

Materials and Methods:
161 patients (age 2–16 years, 61% male and 39% female) with SCD at Sultan Qaboos University Hospital, Muscat, Oman, were started on HU for clinically severe course (defined as: annual vaso occlusive crisis admissions more than 3, and/or occurrence of acute chest syndrome) with starting dose of 15 mg/kg and a mean dose of 18 mg/kg/day. All patients were included in the study except for 19 patients who were excluded for various reasons (8 lost follow up, 6 stopped HU by parents, and 5 stopped due to side effects: dizziness, abdominal pain, and myelosuppression). Remaining 142 patients were followed up for median treatment duration of 4 years (range, 1.5–10 years). Patients were assessed for clinical and laboratory response to HU.

Results:
There was significant reduction in the annual number of admissions due to vaso occlusive crisis (P <0.001, Wilcoxon Signed Ranks Test) with a mean of 4.7 and 1.5 before and after HU use respectively.
There was also observed clinical improvement regarding the incidence of acute chest syndrome. Only 12 out of 39 patients initiated on HU for acute chest syndrome had a recurrent attack during a follow up period of 3–7 years.
The laboratory parameters were consistent with previous reports: significant increase in hemoglobin level, fetal hemoglobin (HbF) level, Mean Corpuscular Volume, whereas significant decrease in platelet, absolute neutrophil, and absolute reticulocytic counts.
All 142 patients tolerated the treatment well. Observed side effects included abdominal pain, dizziness, rash, tremor in one patient each. Of note, 21 patients (11.1%) developed neutropenia (ANC<1000 × 10̂9/l) while 7 patients (3.7%) had thrombocytopenia (<100 × 10̂9/l) which interrupted the treatment for period ranging from 3 weeks to 3 months only.

Conclusion:
In SCD patients with background of ethnic neutropenia, lower starting dose and limited range of dose escalation of HU ensured safety and yet did not affect efficacy.

Disclosures:
No relevant conflicts of interest to declare.