bluuebunny/crossref_metadata_2025_split huggingface dataset:
Disruption of Bis Leads to Deterioration of Mesenchymal Stromal Cells Predominantly Affecting Vascular Compartment of Hematopoietic Stem Cell Niche.
Authors: Oh Il-Hoan , Kwon Kyong-Rim , Ahn Ji-Yeon , Kim Myungshin , Lee Jeong-Hwa 
Published: October 2019

Abstract
Abstract 3627
Poster Board III-563
The stem cell niche plays an important role in the microenvironmental regulation of hematopoietic stem cells (HSCs), but the integration of niche activity remains poorly understood. In this study explored the hematopoietic defect of mice disrupted with Bis/BAG-3/CAIR-1, a protein related to apoptosis and response to cellular stress and show that functional loss of bis leads to series of hematopoietic derangements due to perturbation of vascular stem cell niche. First, mice with targeted disruption of bis (bis−/−) exhibited severe hypocellularity in the bone marrows and spleen starting from 16 days after birth. Affected mice exhibited loss of primitive neonatal HSCs (CD34+Lin-Sca-1+c-kit+) and defect in early stage B-lymphopoiesis including common lymphoid progenitors (IL-17R+LSK), pre-B and pro-B cell populations, but not for mature recycling B-lymphocytes (IgD+B220+). However, this hematological defect of bis−/− mice was not reproduced when bis−/− bone marrow cells were transplanted into wild-type (WT) recipients, pointing to the microenvironmental origin of the phenotypes. Subsequent analysis of bis−/− mice bone marrow revealed a characteristic defect in the mesenchymal stromal component that included a quantitative loss of stromal cells (CD45-CD31-TER119-CD105+) in the bone marrows and rapid sensescence of stromal cells comprising colony forming unit-fibroblast (CFU-F) when re-plated in the ex-vivo culture. Moreover, mesenchymal stromal cells expressing CXCL-12 or IL-7 was lost in the affected bone marrows with lowered density of vascular development, together indicating a perturbation of peri-vascular stem cell niche in the bone marrow. In contrast, no abnormalities were observed in the growth and hematopoietic supporting activities of osteoblasts obtained from bis−/− mice. Collectively, these results indicate that Bis functions to mediate cellular regulation of the stem cell niche activities selectively on the vascular compartment without affecting osteoblastic niche, and suggest that Bis may serve as a molecule that can bridge the microenvironment niche and cellular stress/apoptotic signals during the in-vivo orchestration of hematopoiesis.

Disclosures:
No relevant conflicts of interest to declare.
MT1-MMP Is Required for Hematopoietic Maturation in the BM Niche.
Authors: Nishida Chiemi , Beate Heissig , Seiki Motoharu , Nakauchi Hiromitsu , Hattori Koichi 
Published: October 2019

Abstract
Abstract 3634
Poster Board III-570
Specialized niches, in which hematopoietic stem cells (HSCs) reside control the balance between HSC quiescence and self-renewal, yet little is known about the extrinsic signals provided by the niche and how these niche signals regulate such a balance. Activation of the fibrinolytic pathway via matrix metalloproteinase-9 (MMP-9) resulted in the release of kit ligand (KitL) in the BM niche. Membrane type 1-MMP (MT1-MMP) can activate e.g. MMP-9. To investigate the role of MT1-MMP in hematopoiesis, we used MT1-MMP deficient mice. MT1-MMP−/− mice examined 12 days after birth showed pancytopenia and reduced numbers of bone marrow mononuclear cells (BMMCs) and splenocytes. BM cytospins from MT1-MMP−/− mice showed mild perturbations in erythropoiesis and a more severe impairment of myelopoiesis. Although all lineages were present, the ratio of erythroid to myeloid precursors increased from 0.36 in wildtype to 0.60 in MT1-MMP−/− mice. Myeloid and erythroid cell differentiation was impaired in MT1-MMP−/− BMMCs. The numbers of colony forming unit cells (CFU-C) was reduced in MT1-MMP−/− BMMCs. In contrary, the number of immature hematopoietic cells (CFU-S8, KSL cells) was augmented in MT1-MMP−/− BMMCs. FACS analysis of BM cells showed a decrease in the percentage of mature B cells with an increase number of Pro-B and immature B cells in MT1-MMP−/− BMMCs relative to controls. MT1-MMP−/− BM cells showed lower expression of CXCL12 and KitL, typical niche growth factors important for myelopoiesis and lymphopoisis. Thus, MT1-MMP is required for normal hematopoietic differentiation of lymphoid and myeloid lineage cells, most likely due to growth factor defective niche cells.

Disclosures:
No relevant conflicts of interest to declare.
Modified International Prognostic Score (IPS) for Predicting the Need for Bone Marrow Biopsy (BMB) in Hodgkin's Lymphoma (HL).
Authors: Muslimani Alaa A , Spiro Timothy , Chaudhry Asif A. , Taylor Harris C , Jaiyesimi Ishmael , Vashist Praveen , Daw Hamed A 
Published: October 2019

Abstract
Abstract 3661
Poster Board III-597

Introduction
BMB is frequently performed during the staging process of patients (pts) with HL. Previous data suggested routine use of this procedure may be unnecessary. The Ann Arbor classification is currently used to detect pts requiring BMB, despite sensitivity (sen) of 100%, specificity (spec) is only 40%. We sought to determine whether a modified IPS [Hasenclever and Diehl (1998)] would yield greater spec while maintaining good sen.

Methods
We retrospectively reviewed charts of 1215 histologically proven HL pts from Jan 2000-Dec 2008 at Cleveland Clinic Taussig and Moll Cancer Centers. Of 1215 pts, 1089 had BMB (90%). 876 pts were included in our study. 213 were excluded due tor un-interpretable BMB or missing data. Each component of IPS was equally weighted such that one point is assigned to each positive variable [male sex, age ≥45, albumin (alb) <4 g/dL, hemoglobin (hem) <10.5 g/dL, stage IV, white blood cell (WBC) ≥15,000/mm3, lymphocyte (lymph) <600/mm3 and/or <8% of total WBC], Multivariable logistic regression was initially used to assess the association between bone marrow involvement (BMI) and each factor in the IPS. All 7 factors were significant (p<.001 for sex, age, albu, hem, stage and lymph; .07 for WBC); and therefore recursive partioning algorithm was used to identify a cutoff for determining BMI.

Result
88 pts (10%) had BMI, we used a modified IPS formally by calculating a score that uses “weights” based on the results of a multivariable model including all 7 factors and the relative magnitude of the associated regression coefficients (RC), [modified IPS= WBC+2x(sex+age+hem)+3x(stage+alb)+4xlymph].
This model suggests lymphopenia has a stronger impact on the presence of BMI, while leukocytosis has the least impact. The percentage of BMI was 0.1,17,48, and 87 % for pts with a score of 0-8, 9-10, 11-12, and >12 respectively. Using a score of >8 as a rule for BMI, sensitivity was 99% and specificity was 87%. The prevalence of BMI by histology was 4% lymphocyte-rich, 5% nodular sclerosis, 20% mixed-cellularity and 21% lymphocyte-depleted, further analysis showed using modified IPS, sensitivity and specificity were similar across all types of histologies.

Conclusions
Using a modified IPS of > 8 for predicting BMI in HL doubled the spec associated with Ann Arbor classification. The implementation of modified IPS is a practical and reliable tool that will allow physicians to predict BMI in HL pts. Therefore eliminate unnecessary painful procedure in many pts.

Disclosures:
No relevant conflicts of interest to declare.
Serum TARC Levels Correspond with Tumor Load and Response to Chemotherapy in Classical Hodgkin Lymphoma Patients.
Authors: Diepstra Arjan , van der Graaf Anne Marijn , Vos Hans JTWM , Visser Lydia , Van Den Berg Anke , van Imhoff Gustaaf 
Published: October 2019

Abstract
Abstract 3663
Poster Board III-599

Introduction
Thymus and Activation Related Chemokine (TARC) is produced in high amounts by Hodgkin and Reed-Sternberg (HRS) cells in classical Hodgkin lymphoma (cHL). It attracts CCR4 positive Th2 and Treg cells, which are typically present in large numbers in the reactive infiltrate in cHL. TARC can be measured in the serum of cHL patients and is usually elevated in the presence of active disease, as compared to healthy controls. Higher levels are observed in advanced stage disease, compared to stage I disease. The goal of the current study was to correlate TARC serum levels to metabolic response to therapy as measured by FDG-PET scanning.

Patients and methods
Newly diagnosed cHL patients treated in our hospital were asked to participate. Serum samples were taken before the start of treatment, at the time of the mid treatment PET scan (after 2 cycles of ABVD for stage I and II, or mid BEACOPP or ABVD treatment for stage III and IV disease) and at the time of the PET scan after completion of chemotherapy. TARC serum levels were measured by ELISA and TARC immunohistochemistry was done on primary involved lymph node tissue.

Results
A total of 59 cHL patients, median age 40 (range 16-80) and 49% male participated. Eight patients had stage I, 26 stage II, 14 stage III and 10 stage IV disease. The known association of high pretreatment TARC levels with advanced stage was also significantly present in our data, supporting the hypothesis that serum TARC levels reflect tumor burden. Moreover, in stage I and II patients, bulky disease (27%) was associated with a significant higher TARC level, lending further support to this hypothesis. Compared to pretreatment values, TARC levels had decreased significantly at mid treatment and treatment completion to the range of healthy controls. Only in stage I there was no significant difference between pretreatment and mid or after treatment samples (see figure), probably due to the relatively large number of patients without elevated TARC before the start of therapy. In patients with a positive PET scan at mid treatment (26%), there was a trend for a higher TARC level (P=0.08) compared to PET negative patients. After completion of chemotherapy PET positivity (19%) was significantly associated with a higher TARC level. Three patients had a relapse and at that time TARC levels had clearly increased in comparison to the levels after treatment.

Conclusions
serum TARC levels are associated with tumor burden and metabolic response to therapy as measured by FDG-PET scan. Measurement of serum TARC levels is a potential tool to evaluate the response to therapy and to be used in follow up after completion of chemotherapy. Inclusion of more patients is necessary to further assess the clinical value of TARC serum levels and is ongoing.

Disclosures:
No relevant conflicts of interest to declare.
CD4 and CD8 TCRαβ Cells Are selected On MHC Expressed On Thymocyte Precursors in OP9-DL1 Cultures.
Authors: Van Coppernolle Stefanie , Vandekerckhove Bart , Verstichel Greet , Velghe Imke , Taghon Tom , Leclercq Georges , Plum Jean , Kerre Tessa C. C. 
Published: October 2019

Abstract
Abstract 3670
Poster Board III-606
Human CD34+ haematopoietic precursor cells cultured on delta-like-ligand 1 expressing OP9 (OP9-DL1) stromal cells differentiate to T lineage cells. The nature of the T cells generated in these cultures has not been studied in detail. Since these cultures do not contain thymic epithelial cells (TEC) which are the main cell type mediating positive selection in vivo, generation of conventional helper CD4+ and cytotoxic CD8+ TCRαβ cells is not expected.
Phenotypically mature CD27+ CD1− TCRγδ as well as TCRαβ cells were generated in OP9-DL1 cultures. CD8 and few mature CD4 single positive (SP) TCRαβ cells were observed. Mature CD8 SP cells consisted of two subpopulations: one expressing mainly CD8αβ and one expressing CD8αα dimers. TCRαβ CD8αα and TCRγδ cells both expressed the IL2Rβ receptor constitutively and proliferated on IL-15, a characteristic of unconventional T cells. CD8αβ+ and CD4+ TCRαβ+ cells were unresponsive to IL-15, but could be expanded upon TCR stimulation as mature CD8αβ+ and CD4+ T cells. These T cells had the characteristics of conventional T cells: CD4+ cells expressed ThPOK, CD40L and high levels of IL-2 and IL-4; CD8+ cells expressed EOMES, RUNX3 and high levels of granzyme, perforin and IFN-γ.
Induction of murine or human MHC class I expression on OP9-DL1 cells had no influence on the differentiation of mature CD8+ cells, whereas blocking MHC class I antibodies and CD8 antibodies inhibited generation of mature CD27+ CD8+ SP cells. The presence of dendritic cells was not required for the generation of mature CD4+ or CD8+ T cells.
In conclusion, we have demonstrated positive selection of TCRαβ+ cells in the absence of TEC. The data suggest that positive selection is induced by interaction between T precursor cells without requirement for other haematologic or stromal elements.

Disclosures:
No relevant conflicts of interest to declare.
Incidence of Progressive Multifocal Leukoencephalopathy in Non-Hodgkin Lymphoma Patients Treated with Rituximab.
Authors: Focosi Daniele , Tuccori Marco , Blandizzi Corrado , Del Tacca Mario , Petrini Mario 
Published: October 2019

Abstract
Abstract 3675
Poster Board III-611
Progressive multifocal leukoencephalopathy (PML) is a rare complication of immunesuppression due to productive reactivation of JC polyomavirus (JCV) in glial cells. In the last decades many case reports of rituximab-associated PML have raised concerns, but no systematic incidence rate analysis has been ever performed. We retrospectively reviewed our clinical records with the aim of calculating the incidence rate of PML in patients with NHL who had been treated with regimens including rituximab. Data on HIV-negative patients who received the first dose of rituximab from January 1, 2000 to June 30, 2008, were analyzed. The follow-up period was from the first rituximab dose to the last recorded visit, up to September 30, 2008. PML cases were included if symptoms occurred at least 1 month after the first dose of rituximab and the diagnosis was supported by magnetic resonance imaging and detection of JCV DNA in stereotactic brain biopsies and/or cerebrospinal fluid. We collected data from 821 consecutive patients throughout the follow-up period. All patients received chemotherapy other than rituximab, and all completed their entire treatment course at the Hematology Unit in Pisa. No radiation therapy was administered. The median time of follow-up was 20 months (range = 1-106 month), resulting in 1725 patient-years at risk. Five cases of PML (two receiving maintenance rituximab) were identified, with an incidence rate of 2.89 cases per 1000 patient-years. We found that the incidence rate of PML in our population exceeded that observed in patients who are traditionally regarded as being at high risk of PML, namely patients with B-cell chronic lymphocytic leukaemia and AIDS. Rituximab might be a potential contributing factor to the development of PML in these patients. Nevertheless, rituximab currently represents an essential therapeutic tool that can positively affect the natural history of NHL. However, because of the expanding therapeutic indications, it seems reasonable to investigate the potential contribution of rituximab to PML occurrence.

Disclosures:
No relevant conflicts of interest to declare.
Latent Murine Herpesvirus-4 Infection Arms NK Cells.
Authors: White Douglas W , Keppel Catherine R , Schneider Stephanie E , Coder James , Reese Tiffany A , Payton Jacqueline E. , Ley Timothy J. , Virgin Herbert W , Fehniger Todd A 
Published: October 2019

Abstract
Abstract 3678
Poster Board III-614
Natural killer (NK) cells are lymphocytes originally identified by their ability to kill target cells without prior sensitization. However, murine NK cells require an antecedent ‘arming’ event to translate cytotoxic effector proteins (perforin and granzymes) resulting in potent cytotoxic capacity. In contrast to mice, most NK cells (CD56dim) from the peripheral blood of healthy humans are armed with pre-formed perforin and granzyme B proteins and mediate killing of NK sensitive targets directly ex vivo. This suggests that NK cells from specific pathogen free (SPF) laboratory mice are lacking a critical arming event present in healthy humans, and provides an experimental system to elucidate the events that result in NK cell arming in vivo. Since latent herpesviruses are highly prevalent and alter multiple aspects of host immunity, we hypothesized that the immune environment created by a latent herpesvirus infection arms NK cells. NK cells from mice latently infected with Murid herpesvirus 4 (MuHV-4), a virus closely related to the human viruses Kaposi's sarcoma associated herpesvirus (KSHV) and Epstein-Barr virus (EBV), were armed as evidenced by increased granzyme B protein expression, cytotoxicity, and interferon-gamma production. Adoptive transfer experiments with naïve NK cells indicated that NK arming occurred rapidly (≤ 72 hours) in the latently infected host and did not require acute infection. In addition, experiments utilizing a genetic variant of MuHV-4 that acutely infects mice similar to wild-type MuHV-4, but does not establish latency, confirmed that acute infection was not responsible for this NK cell modulation. Finally, NK cells armed by latent infection protected the host against a lethal RMA-S lymphoma challenge. Thus, the immune environment created by a latent herpesvirus infection arms NK cell function in vivo. These results indicate that some of the differences between human and SPF laboratory-mouse NK cell function may reflect a disparity in immune activation induced by chronic virus infection, as opposed to an intrinsic difference in the NK cells themselves. Moreover, these findings indicate that the activation state of the innate immune system needs to be taken into account when using mice to model immune responses against pathogens and tumors.

Disclosures:
No relevant conflicts of interest to declare.
Strong Inhibition of Alloreaction by Newly Generated Tolerogenic Dendritic Cells by Psoralen Plus UVA Treatment.
Authors: Maeba Hideaki , Nishimura Ryosei , Kuroda Rie , Araki Raita , Mase Shintaro , Ikawa Yasuhiro , Yachie Akihiro , Koizumi Shoichi 
Published: October 2019

Abstract
Abstract 3686
Poster Board III-622
Dendritic cells (DCs) are a heterogenous population of antigen-presenting cells (APCs) that contribute to innate immunity and initiate the adaptive immune response. In addition, recent studies demonstrated the existence of tolerogenic DCs (TDCs) that suppress the immunoreaction. Although the tolerogenic mechanisms are not fully understood, there are some methods reported to generate TDCs from conventional DCs by using cytokines such as IL-10/TGF-beta, vasoactive intestinal peptide (VIP), and IL-21. With the purpose of potential application of TDCs in a clinical bone marrow transplantation for preventing graft-versus-host disease (GVHD), which is caused by strong immunoreaction between host-typed conventional DCs and donor-typed naïve T cells, the establishment of much safer and easier, and more efficient TDCs culture system would be needed. To this end, we investigated whether treatment of conventional DCs with psoralen plus UVA (PUVA), which is widely and safely available in the treatment of some human immune disease and organ transplantation for preventing graft rejection, induced a subset of highly potent TDCs. Bone marrow cells obtained from Balb/c (H-2d) or C57BL/6 (H-2b) were incubated in complete RPMI containing GM-CSF for 10 days to generate bone marrow derived DCs (BM-DCs). BM-DCs were cultured with Psoralen (200 ng/mL) for 30 miniutes and then exposed to UVA light (2J/cm2). After 24 hours UVA irradiation, PUVA-treated DCs were collected and used in all experiments. First, PUVA-treated or untreated DCs with irradiatin were used as stimulator for allogenic splenocytes in mixed leukocyte reactions (MLR). The immunostimulatory capacity of PUVA-treated DCs was significantly diminished compared to those of untreated DCs (p<0.01). The expression levels of CD80 and CD86 by FACS, both of which are costimulatory molecules for T cell activation, was significantly reduced after PUVA treatment (p<0.05). This might explain for the induction of hyporesponsiveness in part. Next we further evaluated whether PUVA-treated DCs directly suppress T cell alloreaciton by cell-to-cell contact. Proliferation was inhibited when PUVA-treated DCs from the stimulator strain were added to the coculture, with a maximum reduction in proliferation of 95% at a 1:1 or higher ratio of untreated DCs to PUVA-treated DCs (Attached file). In conclusion, PUVA-treated DCs directly inhibit T cell alloreaction. Infusion of host-typed PUVA-treated DCs would be potent strategy for preventing lethal acute GVHD.

Disclosures:
No relevant conflicts of interest to declare.
The mTOR Pathway Inhibits Expression of the Tumor Suppressors ELF4 and KLF4 Downstream of TCR Signaling to Induce T Cell Proliferation.
Authors: Yamada Takeshi , Gierach Kirsten , Lacorazza H. Daniel 
Published: October 2019

Abstract
Abstract 3688
Poster Board III-624
Quiescence of circulating naïve T cells is maintained by the transcription factors ELF4 and KLF4 downstream of T-cell receptor (TCR) signaling. Hence, loss of ELF4 leads to increased proliferation of CD8+ T cells in response to homeostatic and antigen driven stimuli (Yamada et al, Nature Immunology, 2009). The identification of signals that suppress this restraint of proliferation will aid to enhance immunological memory during vaccination and to better understand development of T-cell acute lymphoblastic leukemias. Consistent with lower threshold of activation by ELF4 deletion in unstimulated naïve T cells, we identified a significant downregulation of the dual-specificity phosphatases DUSP1 and DUSP5 in a global gene expression study, which was confirmed at a protein level. Consequently, Elf4−/− CD8 T cells showed sustained phosphorylation of Erk1/2 upon TCR activation. In addition, we found that the PD98059 and LY294002 inhibitors, but not Cyclosporin A, blocked inhibition of ELF4 transcription upon TCR activation independently of CD28 co-stimulation and signals emanating from IL-2R. Furthermore, rapamycin also prevented downregulation of ELF4 transcripts following T cell activation, suggesting that mTORC1 inhibits ELF4 transcription downstream of MAPK and PI3K/Akt pathways. We conclude that the transcription factor ELF4 sets a proliferation threshold in naïve T cells by activating DUSPs and that ELF4 suppression upon TCR activation is mediated by mTORC1 downstream of MAPK and PI3K/Akt pathways. Our findings provide important targets of this novel control of T cell proliferation to enhance immune response to vaccination and to prevent expansion of pre-leukemic clones in pediatric patients that fail to respond to current therapies.

Disclosures:
No relevant conflicts of interest to declare.