bluuebunny/crossref_metadata_2025_split huggingface dataset:
Surviving Host CD4+CD25+Foxp3+ Cells Following Ablative Conditioning Expand and Comprise the Major Component of the Treg Compartment during the Lymphoid Reconstitution Period Following HCT.
Authors: Bayer Allison L. , Jones Monica , Urbieta Maite , Chirinos Jackeline , Schreiber Taylor , De Armas Leslie , Malek Thomas R. , Levy Robert B. 
Published: November 2019

Abstract
The capacity of CD4+CD25+Foxp3+ (Treg) cells to regulate adaptive and innate immune responses has led to studies investigating their application to regulate allogeneic T cell responses arising during hematopoietic stem cell transplants (HCT). With respect to HCT, a fundamental clinical concern is the reconstitution of the lymphoid compartment in recipients, particularly T cells since this process can be exceedingly delayed. We have previously found that host Treg cells can regulate resistance to engraftment following HCT, demonstrating that such cells survive and function at least transiently in recipients. The present studies investigated the residual host Treg compartment following HCT. Utilizing varying levels of total body irradiation (5.0 – 14Gy), we observed:
recipient CD4+CD25+Foxp3+ cells can survive ablative as well as reduced intensity conditioning, the surviving, i.e. residual Treg cells undergo expansion as assessed by BrdU uptake and cell numbers, and these cells comprise the majority of the Treg compartment in recipients for several months post-HCT during which time donor derived Treg cells gradually arise and cede this compartment.
Residual Tregs also dominated the compartment following allogeneic HCT of MHC-matched bone marrow depleted of T cells. To assess the functional capacity of the residual Treg cell compartment, the development of autoimmune disease following transplant of IL-2Rβ −/− (CD122−/−) bone marrow into syngeneic recipients with and without residual Tregs was examined. Autoimmune disease symptoms and T cell alterations were prevented in B6-wt but not T cell deficient recipients. Interestingly, the failure to transfer autoimmune disease following IL-2Rβ −/− HCT into lethally conditioned B6-CD4−/− recipients was associated with the presence of a peripheral CD8+FoxP3+ population not detected in B6-wt mice or the B6-wt mice transplanted with IL-2Rβ −/− BM. This finding indicates that in the genetic absence of CD4+ T cells, a CD8 regulatory population appears to emerge. In total, our observations support the notion that functioning host Tregs initially occupy a niche in the transplant recipient permitting lymphopenic expansion and an extended period of contribution to this compartment. Notably, this contribution reflected much greater levels than conventional T cell populations - even in aggressively conditioned recipients. Finally, these findings imply that the presence of host regulatory cells may be important to consider with respect to eliciting anti-tumor responses and vaccination in recipients during the early period post -hematopoietic cell transplantation.
MEK Inhibitor AZD6244 Induces Cell Growth Arrest and Synergizes Nutlin-3a-Mediated Cell Death by Upregulating p53 and PUMA Levels in Acute Myelogenous Leukemia.
Authors: Zhang Weiguo , Konopleva Marina , Schober Wendy D. , McQueen Teresa , Andreeff Michael 
Published: November 2019

Abstract
Raf/MEK/Erk signaling is constitutively activated in the majority of acute myeloid leukemias (AML), providing rationale for targeting this pathway with therapeutic intent. We investigated growth-inhibitory and pro-apoptotic effects of AZD6244, a small molecule MEK1/2 inhibitor, as well as potential synergistic effects with the MDM2 antagonist Nutlin-3a in AML. AZD6244 inhibited proliferation of AML cells by arresting cells in G1 phase, which resulted from inhibition of cell cycle checkpoint proteins cyclin E and cdk2, and was associated with the modulation of cyclin D1, cyclin A, cyclin B1, p27 and Rb phosphorylation levels. AZD6244 also induced apoptotic cell death in AML cell lines with constitutive activation of MEK/ERK signaling (OCI/AML3, HL60 and MOLM13) via decrease of anti-apoptotic Mcl-1 protein expression and upregulation of pro-apoptotic Bax, Bak, Bad and Bim proteins. Next we tested the hypothesis that activation of p53 signaling with a small molecule MDM2 inhibitor Nutlin-3a would enhance the pro-apoptotic effects of MEK inhibition in AML. Combinations of AZD6244 and Nutlin-3a triggered profound apoptotic responses in AML cells with wild-type p53 (OCI/AML3, MOLM13 and Baf3-FLT3-ITD), with combination indices (averaged ED50=0.37, ED75=0.38, ED90=0.43) suggesting strong synergism. In contrast, no synergism was seen in p53 null cell HL60 (ED50=1.27, ED75=1.38, ED90=1.66) or in OCI-AML3 cells stably transduced with p53 shRNA. Mechanistically, combined blockade of MEK pathway and activation of p53 signaling resulted in marked upregulation of p53 expression and of the p53 target protein PUMA, while only partially suppressing Nutlin-induced p21 upregulation. This resulted in loss of the mitochondrial membrane potential, cytochrome c release and cleavage of caspase-9 and -3. Importantly, combined AZD6244 and Nutlin-3a treatment potently induced apoptosis in blasts from primary AML patients. In summary, AZD6244 induced growth-inhibitory and pro-apoptotic effects in AML cells with constitutive activation of MEK/ERK signaling. These effects were synergistically enhanced by simultaneous activation of p53 signaling by Nutlin-3a, resulting in the activation of the intrinsic apoptotic pathway. Altogether, these results strongly suggest potential activity of AZD6244 alone or in combination with MDM2 antagonists as novel mechanism-based therapeutic agents in AML therapy.
The Decrease of Regulatory T Cells through the Low Levels of FOXP3 Expression Mediated by NFAT1 in Infancy with Autoimmune Neutropenia.
Authors: Nakamura Kazuhiro , Miki Mizuka , Karakawa Syuhei , Onodera Rie , Kurita Emi , Hiraoka Asako , Sato Takashi , Kobayashi Masao 
Published: November 2019

Abstract
CD4+ /CD25+ regulatory T cells have been recognized to regulate the maintenance of self-tolerance, and several human autoimmune diseases in adulthood. To date, little is known about the standard value of regulatory T cells in infancy and childhood and about its involvement in pediatric autoimmune disease. Autoimmune neutropenia (AIN) in infancy is a common disorder characterized by chronic neutropenia, the detection of antineutrophil antibodies in sera, and spontaneous resolution of neutropenia within several months to a few years. In this study we examined the frequencies of regulatory T cells in various normal age groups and AIN patients to elucidate the involvement of regulatory T cells in AIN known as a representative autoimmune disease in infancy and early childhood. Blood samples from healthy neonates (n = 22), infants (1 month through 3 years in age, n = 21), adults (n = 12) and AIN patients (1 month through 3 years in age, n = 22) were obtained after informed consent. The frequencies of regulatory T cells were assessed by the expression of CD4, CD25, and FOXP3 using flow cytometry. The mRNA expressions of FOXP3 and its transcription factor, NFAT in CD4+ /CD25+ regulatory T cells were determined by quantitative real-time PCR. The frequency of CD4+ /CD25high regulatory T cells known as pure regulatory T cells fraction was highest in neonates and decreased by aging. Notably, the number of CD4+ /CD25high regulatory T cells in AIN patients significantly decreased, compared with that of age-matched healthy infants (P < 0.01) and the ratio of intracytosolic FOXP3 expression of each regulatory T cell fraction in AIN patients was lower than that of age-matched healthy infants (P < 0.05). Next, we purified CD4+ /CD25+ T cells by using FACS-Aria (more than 95% of purity) and the mRNA expressions in CD4+ /CD25+ T cells were quantified. The expressions of FOXP3 and NFAT1 mRNA in AIN patients were significantly lower than those of healthy infants (P < 0.05). The low level of FOXP3 mRNA in patients well correlated with the expression level of NFAT1 mRNA, implying that the expression of FOXP3 was mediated by NFAT1. Furthermore, the decreased frequency of CD4+ /CD25high regulatory T cells and FOXP3-positive cells in AIN patients were restored to normal levels when the neutrophil count was spontaneously recovered. These results strongly suggest that regulatory T cells play an important role in the immunopathophysiology of AIN in childhood.
Lenalidomide Is Active in Patients with Relapsed/Refractory Chronic Lymphocytic Leukemia (CLL) Carrying Unfavorable Chromosomal Abnormalities.
Authors: Ferrajoli Alessandra , Keating Michael J. , Wierda William G. , O’Brien Susan M. , Padmanabhan Swaminathan , Czuczman Myron S. , Chanan-Khan Asher A. 
Published: November 2019

Genomic abnormalities, according to the Dohner hierarchical classification, are an important prognostic factor in chronic lymphocytic lymphoma (CLL). Specific chromosomal aberrations, such as a deletion 17p (P53 abnormalities) and deletion 11q (ATM defects), are associated with aggressive disease and inferior response to purine analogue-based chemotherapy. Lenalidomide is an immunomodulatory agent that has demonstrated clinical efficacy in patients with relapsed or refractory CLL in 2 separate phase II clinical trials (Chanan-Khan et al. JCO 2006; Ferrajoli et al. Blood 2006 abst). To investigate the clinical activity of lenalidomide in patients with high-risk cytogenetics (17p or 11q abnormalities), we reviewed data from 2 clinical studies. On these studies, lenalidomide was given orally either at 10 mg daily for 28 days followed by 5 mg increments every 28 days to a maximum dose of 25 mg (study A) or given at 25 mg on day 1–21 of each 28 day cycle (study B). The presence of deletion 17p or deletion 11q was demonstrated by interphase fluorescence in situ hybridization (FISH) analysis. Clinical responses were assessed using the NCI-WG 1996 criteria. Among the 80 patients treated, 40 patients with deletion 17p or deletion 11q were identified. Their characteristics are described in Table 1.
The overall response rate in patients carrying deletion 17p or 11q was 35% (14/40) and responses are depicted in Table 2. Median response duration was 12 months in both studies.
Based on our experience, single agent lenalidomide induces complete and partial responses in patients with unfavorable genomic aberrations, a group characterized by a poor outcome in several studies. The activity of this agent warrants further evaluation in this patient population.
Table 1. Characteristics of 40 patients with unfavorable cytogenetics Study A Study B N=24 N=16 Median age, years (range) 63 (49–86) 62 (56–75) Rai stage III/IV 13 (54%) 4 (25%) Median no. of prior therapies (range) 3 (1–15) 3 (1–10) Median beta 2 microglobulin (range) 4.5 (1.9–10.1) 4 (2–10) Table 2. Responses according to NCI-WG criteria in 40 patients with unfavorable cytogenetics. Response Study A Study B 17p- (N=8) 11q- (N=16) 17p-/11q- (n=24) 17p- (N=6) 11q- (n=10) 17p-/11q- (N=16) CR, n 0 2 2 0 1 1 PR, n 1 4 5 3 3 6 OR, n (%) 1 (13) 6 (38) 7 (29) 3 (50) 4 (40) 7 (44)
Alemtuzumab Increases Serious Infections in Patients with Previously Untreated Chronic Lymphocytic Leukemia (CLL) Receiving Fludarabine-Based Therapy: A Comparative Analysis of 3 Cancer and Leukemia Group B Studies (CALGB 9011, 9712, 19901).
Authors: Morrison Vicki A. , Peterson Bercedis L. , Rai Kanti R. , Byrd John C. , Larson Richard A. 
Published: November 2019

Abstract
Infections remain a critical issue in the care of CLL patients (pts). The specific agents used and their resultant immunosuppression impact the spectrum of infection. We compared the infectious complications reported in previously untreated CLL pts enrolled on 3 sequential CALGB treatment trials. On CALGB 9011, 188 pts received single agent fludarabine (F) therapy. On CALGB 9712, 104 pts received concurrent or sequential fludarabine plus rituximab (FR). On CALGB 19901, pts were to receive 4 cycles of F, followed by 6 weeks of alemtuzumab consolidation (FA); of the 85 enrolled patients, 59 proceeded to alemtuzumab consolidation. Infections requiring hospitalization and/or parenteral antibiotics were tabulated. The 85 pts on study 19901 (FA) had significantly more of these infections during protocol therapy (32/85, 38%) compared to the 188 pts receiving F on study 9011 (43/188, 23%, p=0.01). Similarly, the 19901 study pts also had more infections than the 104 FR-treated pts on study 9712 (21/104, 20%, p=0.0007). Among the 85 pts on study 19901, 14 (16%) had infections only during the F phase of therapy, 14 (16%) had infections only during alemtuzumab consolidation, and 4 (5%) had infections during both phases of therapy. The occurrence of cytomegalovirus (CMV) and Pneumocystis (PCP) infections was also examined on these trials. No CMV and only 3 PCP infections were diagnosed among the 188 pts receiving F on study 9011. For study 9712, 3/104 pts had PCP (one on concurrent FR, two on sequential FR), and 1 pt had CMV pneumonia on sequential FR. Among pts on study 19901, one case of PCP and no cases of CMV occurred during the F phase of therapy. However, 12/59 pts developed CMV infection during alemtuzumab consolidation. Four had resolution of infection and completed alemtuzumab therapy, while 8 were hospitalized and did not complete this therapy. We conclude that the addition of rituximab to F therapy does not increase the risk or severity of infections in previously untreated CLL pts, but the use of alemtuzumab consolidation following initial F therapy does increase the overall risk of serious infectious complications, and especially CMV infections. These issues are important not only for prophylactic antimicrobial therapy and monitoring during and after therapy, but also for the potential use of alemtuzumab as a consolidation therapy to eradicate residual CLL after F treatment.
Factor VIII, a Cofactor for Accelerating Proteolytic Cleavage of von Willebrand Factor by ADAMTS13 Metalloprotease.
Authors: Cao Wenjing , Krishnaswamy Sriram , Lenting Peter , Zheng X. Long 
Published: November 2019

Abstract
von Willebrand factor (VWF)-cleaving protease (ADAMTS13), a 195-kDa plasma glycoprotein, preferentially cleaves “unusually large” VWF multimers and generates a range of VWF multimers with sizes from approximately 500,000 Daltons to 20 million Daltons. However, the molecular basis how ADAMTS13 prefers the larger VWF multimers to the smaller ones under physiological condition remains unknown. We hypothesize that coagulation factor VIII, which binds VWF in high affinity, may alter the susceptibility of VWF to ADAMTS13 protease. In the present study, a plasma-derived or recombinant VWF at final concentration of 150 nM was incubated for 3 min with 50 nM of recombinant ADAMTS13 in the absence and the presence of various concentrations of recombinant factor VIII (0–40 nM) in 50 mM HEPES, pH 7.5 containing 150 mM NaCl and 0.5 mg/ml bovine serum albumin under constant vortex-induced mechanic shear stress at rotation rate of 2,500 rpm. The specific cleavage products (the dimer of 176 kDa) were determined by Western blotting with rabbit anti-VWF IgG, followed by a fluorescence (IRDye800)-labeled anti-rabbit IgG and an infrared fluorescent image system. We showed that the rate of proteolytic cleavage of VWF by ADAMTS13 was markedly accelerated by an addition of recombinant factor VIII into the reaction. This augmenting effect was factor VIII concentration-dependent. The maximal augmentation in proteolytic cleavage of VWF by ADAMTS13 in the presence of 20 nM of recombinant factor VIII was approximately 10 fold over the baseline in the absence of factor VIII. The concentration of factor VIII that achieved half of the maximal effect on proteolytic cleavage of both plasma and recombinant VWF was nearly identical (approximately 3.0 nM). The B domain-deleted factor VIII appeared as efficacious as wild type factor VIII in accelerating the proteolytic cleavage of plasma-derived VWF by ADAMTS13, suggesting that the large, central B-domain of factor VIII is not required for its cofactor activity. Moreover, after being activated by thrombin, the activated factor VIII retained its cofactor activity initially, but rapidly lost its ability to enhance the proteolysis of VWF by ADAMTS13 within minutes, which was correlated with the dissociation (or inactivation) kinetics of the activated factor VIII heterotrimer. These data demonstrate that both factor VIII and activated factor VIII may be the cofactors that regulate ADAMTS13 protease function under physiological condition. The findings may provide a novel insight into how the larger VWF multimers are more susceptible to ADAMTS13 protease, likely because the larger VWF multimers are able to bind more factor VIII. The findings may also help design an optimal therapeutic regimen for treatment of thrombotic thrombocytopenic purpura.
Expression of Coagulation FVIII and FIX in Human Adipose Stromal Cells (hASC) and hASC-Derived Cells - Potential Autologous Cell Therapy for Hemophilia.
Authors: Rosen Elliot D. , Liang Zhong , Song Hui , Bischoff Daniella , Kamocka Malgorzata , Zollman Amy 
Published: November 2019

Abstract
Pleuripotent Adipose Stromal Cells (ASCs) have the capacity to differentiate into mesodermal lineage cells such as adipocytes, myocytes, osteocytes and chondrocytes. In addition, the ASCs can trans-differentiate in vitro into hepatogenic, neurogenic, endothelial and endocrine cells. Since hASCs are relatively easy to isolate in large quantity and have high proliferative potential, they provide a potential source of stem/progenitor cells for autologous cell therapy for the regeneration of injured tissue. The goal of this study is to isolate and engraft adipose stromal cells (ASCs) to generate new tissue capable of secreting therapeutic plasma proteins. In particular, can one engraft genetically modified ASCs in a recipient that secrete coagulation factors for the treatment of hemophilia? Hemophilia A and B result from too low levels of plasma Factor VIII or FIX activity, respectively. The aim of the project is to generate hASC derived tissue capable of secreting FVIII or FIX to support hemostasis in hemophilic mice. Unlike murine ASCs, undifferentiated human ASCs express FVIII at low but significant levels (0.03 ug/ml per 105 cells in 48 hours; ∼30% of normal human plasma FVIII level). ASCs were transfected with a plasmid expressing a modified B-domainless FVIII cDNA. Following differentiation into preadipocytes, the level of FVIII secretion increased 3-fold. In preliminary studies, expression of FIX is detectable in hASCs and the level increases following differentiation into preadipocytes. These results indicate that ASCs and preadipocytes are capable of gamma carboxylation. FIX expression was further increased following infection with a lentivirus carrying a FIX expression cassette. Since HGF has been shown to elicit a potent proangiogenic response (Cai et al., in press), hASCs predifferentated into preadipocytes were mixed in hydrogel containing HGF and injected subcutaneously into SCID mice. Oil red O staining cells were recovered in a cellular nodule at the injection site 3 weeks post-transplantation. Preliminary studies indicate that co-injection of endothelial progenitor cells (EPCs) with ASCs promotes the development of more highly vascularized nodules. In summary, hASCs and hASC-derived preadipocytes cultured ex vivo express low levels of coagulation Factors VIII and IX. These levels can be increased following transfection with FVIII or FIX expression vectors. Furthermore, transplantation of hASCs cultured in adipogenic media results in the formation of preadipocyte/adipocyte containing nodules. These results suggest it may be possible to generate tissue derived from genetically modified hASCs capable of secreting coagulation factors for the treatment of hemophilia.
The Studies in Various Murine Strains with Defects in Activation of Complement Cascade (CC) Reveal Both Pivotal and Pleiotropic Role of CC in Mobilization of Hematopoietic Stem/Progenitor Cells.
Authors: Wysoczynski Marcin , Reca Ryan , Wan Wu , Kucia Magda , Botto Marina , Ratajczak Janina , Ratajczak Mariusz Z. 
Published: November 2019

Abstract
We reported that complement cascade (CC) becomes activated in bone marrow (BM) during mobilization of hematopoietic stem/progenitor cells (HSPC) by i) immunoglobulin (Ig)-dependent pathway and/or by ii) alternative Ig-independent pathway and, as result of this, iii) several potent bioactive CC anaphylatoxins (C3a, desArgC3a, C5a and desArgC5a) are released (Blood2003;101,3784; Blood2004;103,2071; Blood2005;105,40). To learn more on the role of CC and innate immunity in this process, we compared mobilization in mice that possess defects in CC activation by i) classical pathway (C1q−/−, Ig-deficient), ii) both classical and alternative pathway (C2fB−/−) and in animals iii) that do not generate CC-derived anaphylatoxins (C3−/−, C5−/−). For mobilization, we employed G-CSF and zymosan that activate classical and alternative pathways of CC, respectively. First, we found by ELISA that CC activation in fact correlates with the level of HSPC mobilization. Next, studies in mice deficient in CC activation revealed that CC plays both pivotal and pleiotropic roles in this process. Accordingly, while C1q−/− and C3−/− mice turned out to be easy mobilizers, mobilization was very poor in Ig-deficient, C2fB−/− and C5−/− mice that demonstrate that C3 and C5 cleavage fragments differently control the mobilization of HSPC. To explain this at molecular level, we found that C3 cleavage fragments (C3a, desArgC3a) directly interact with HSPC and increase their responsiveness to SDF-1 gradient and thus prevent uncontrolled egress of HSPC from BM. It explains why C1q−/− and C3−/− mice that do not generate C3 cleavage fragments in BM release easily HSPC into circulation. In contrast, C5 cleavage fragments (C5a, desArgC5a) increase permeability of BM-endothelium and thus are crucial for the egress of HSPC from BM to occur. This explains why mice that do not activate efficient CC such as Ig-deficient, C2fB−/− and C5−/− animals are poor mobilizers. We conclude that the mobilization of HSPC is i) dependent on C activation by the classical or alternative pathway and balanced differently by C3 and C5 cleavage fragments that enhance retention or promote egress of HSPC respectively. Thus, modulation of C activation in BM may help to develop new more efficient strategies for both HSPC mobilization and their homing/engraftment.