bluuebunny/crossref_metadata_2025 huggingface dataset:
Prominent Prevalence of Coagulase-Negative Staphylococcus Bloodstream Infections in Neutropenic Hematology/Oncology Children with Central Venous Catheter
Authors: Lee Young-Ho , Lim Yeon Jung , Song Soon-young , Kim Ji-hye , Oh Sung-hee , Koo Hong Hoe
Published: April 2020

A retrospective analysis was performed on febrile neutropenic episodes in children with hematologic and oncologic diseases from 2005 to 2008. We reviewed total 255 febrile episodes occurred during the 3-year period in a total of 114 patients. Twenty-eight episodes of bacteremia occurred during neutropenic period in a total of 16 patients. All these patients had central venous catheter. There were 18 febrile episodes of Grampositive bacteria (64.3%), 9 episodes of Gram-negative bacteria (32.1%), and 1 episode of Candida (3.6%). The isolated organisms were as follows: Coagulase-negative Staphylococcus (CNS, N=17, 60.7%), Staphylococcus aureus (N=1, 3.6%), Enterobacter spp. (N=2, 7.1%), Pseudomonas spp. (N=2, 7.1%), Klebsiella (N=2, 7.1%), E. coli (N=2, 7.1%), Acinetobacter (N=1, 3.6%), Candida. (N=1, 3.6%). The bloodstream infection with CNS were more prevalent in children with Chemo-port (12 episodes in 6 from 34 patients, 17.6%) than in children with Hickman catheter (5 episodes in 5 from 22 patents, 22.7%), even though statistically not significant. In antibiotic susceptibility tests, all gram positive bacteria were resistant to penicillin and oxacillin, and sensitive to vancomycin and teicoplanin. In the case of Gram-negative bacteria, resistant rates to amikacin, gentamicin, imipenem, and piperacillin/tazobactam were 4.8%, 38.1%, 14.3%, and 42.9%, respectively. Based on this retrospective analysis, which reflects predominance of CNS bacteremia in febrile neutropenic patients, more emphasis needs to be laid on the empirical antibiotic regimen including vancomycin or teicoplanin as well as the strict skin preparations during procedures in children with central venous catheter.
Manageable Side Effects of Intravenous Iron in a Community Practice
Authors: Brown Nicole , Henry David H. , Ford Patricia A.
Published: April 2020

Our community hematology oncology practice uses intravenous (IV) iron in a wide variety of patients. A review of the literature on the adverse events (AE) of IV iron found 0.0–32.0% of patients were experiencing an AE with 0.0–0.5 events per patient. In our experience with IV iron, we felt that the actual incidence of AE was much higher than reported. Between June 2006 and April 2007, 156 patient surveys were completed by a total of 120 patients. Surveys were given to patients who had received IV iron in the past week. Those who experience adverse events were asked to note the type, onset, duration and intervention used to alleviate symptoms. Six patients were excluded for reporting symptoms occurring prior to treatment or completing the survey immediately after IV iron treatment. Of the remaining patients, 100 were female and 14 were male with an average age of 48.9 years (17–95 years). Eighty-six patients received iron sucrose, 22 sodium ferric gluconate and 6 iron dextran (InFed). In our patients, 48.2% experienced at least one AE with an average of 1.3 symptoms per patient (see table). The majority of these events (94.5%) were delayed and mild or moderate with only 5.5% of patients requiring a visit to the hospital or physician’s office. Common side effects were muscle or bone aches, dizziness, numbness, tingling and swelling of the hands and feet, abdominal pain and nausea/vomitting. There was no statistically significant difference in the number or severity of AE related to the different preparations. In our experience, AE occur at an increased frequency compared to the published literature. We feel it is important for health care providers to be aware that AE are likely to occur, but that they are usually minor and quite manageable without the need for physician intervention. In our practice the nurses trained in chemotherapy are well equipped to handle most events that arise. We find IV iron to be an extremely useful treatment for patients suffering from both absolute and functional iron deficiency and offer this data as evidence that parenteral iron is safe in a wide variety of patients.

Comparison of AE Iron Sucrose n=86 Sodium Ferric Gluconate n=22 Iron Dextran n=6 Our Study n=114 Previously Reported Muscle or bone aches 17.4% 22.7% 50.0% 20.2% 0.0–3.8% Dizzy/Lightheadedness 16.3% 22.7% 33.3% 18.4% 0.0–0.9% Numbness/Tingling in hands/feet 14.0% 9.1% 16.7% 13.2% N/A Swelling in hands/feet 12.8% 13.6% 16.7% 13.2% 0.0–12.9% Stomach pain/cramping 10.5% 18.2% 16.7% 12.3% 0.0–4.5% Nausea/vomiting 10.5% 13.6% 16.7% 11.4% 0.0–25% Itching 10.5% 4.5% 16.7% 9.6% 0.0–1.5% Chest tightness/discomfort 4.7% 9.1% 0.0% 5.3% 0.0–1.0% Shortness of breath 3.5% 13.6% 0.0% 5.3% 0.0–1.5% Facial Flushing 3.5% 0.0% 0.0% 2.6% 0.0–.3% Hives 1.2% 4.5% 0.0% 1.8% 0.0–6.45% Other 12.8% 13.6% 50.0% 14.9% 0.0–30.0% Any symptom 44.2% 59.1% 66.7% 48.2% 0.0–32.26% Total symptoms 1.2 symptoms/patient 1.5 symptoms/patient 2.2 symptoms/patient 1.3 symptoms/patient 0.0–.45 symptoms/patient
Mega-Analysis of 50 Years of Diagnosis and Therapy of Childhood Acute Lymphoblastic Leukemia in Bydgoszcz, Poland: From Prednisolone Therapy to Microarray Studies
Authors: Styczynski Jan , Jatczak-Gaca Agnieszka , Debski Robert , Koltan Andrzej , Wysocki Mariusz
Published: April 2020

Background: The beginnings of pediatric oncology in Bydgoszcz, Poland dates back to the year 1958. The first patient was treated with prednisolone only.

Objective: Analysis of results of childhood acute lymphoblastic leukemia therapy in Bydgoszcz region in 1958–2008.

Patients and methods: A total number of 570 children were diagnosed and treated over the period of 50 years, including: 215 children treated with different drugs and combinations in 1958–1972, 49 children treated with Memphis V–VII protocols in 1976–1983, 114 children treated with ALL-BFM83 or ALL-BFM86 protocols in 1983– 1995 and 192 children treated with ALL-BFM90, New York I–II or ALL-IC-2002 between 1995–2008.

Results: A number of patients with „events” in respective time periods was: 205/215 (95.3%, in 1958–1972); 44/49 (89.8%, in 1976–1983), 54/114 (47.4%, in 1983–1995), 56/192 (29.2%, in 1995–2008). The values of 5-year pEFS in consecutive therapeutic periods were: p=0.012±0.008 (1958–1972), p=0.102±0.043 (in 1976–1983), p=0.526±0.047 (in 1983–1995), p=0.700±0.037 (in 1995–2008) (p<0.0001). A median survival time was, respectively: 0.8 year (for patients treated in 1958–1972); 1.8 year (in1976–1983); 5.7 years (in 1983–1995); 7.4 years (for patients treated in 1995–2008).

Conclusions: Complex therapy based on international strategy of multiagent total therapy and supportive care, has led to an improved long-term survival in childhood ALL in Bydgoszcz region, Poland, from 1% in sixties up to 70% nowadays

Figure 1. pEFS in ALL with respect to time period Figure 1. pEFS in ALL with respect to time period
Assessment of the Anemia Work up Prior to the Use of Erythropoiesis Stimulating Agents (ESA) in Cancer Patients Undergoing Chemotherapy in a Community Oncology Practice
Authors: Davila Carine , Brown Nicole , Rossi Suzanne , Henry David H.
Published: April 2020

Introduction: Patients undergoing chemotherapy frequently experience chemotherapy induced anemia, which is commonly treated with erythropoiesis stimulating agents (ESAs). However, patients undergoing chemotherapy may also have anemia due to other factors. This study describes how often a proper anemia workup was performed to ensure that the patient’s anemia is due to chemotherapy and not renal insufficiency or iron, B12, or folate deficiency, before administering ESAs in a community hematology-oncology practice of 8 physicians.

Patients and Methods: Medical charts of 278 cancer patients receiving chemotherapy between January and June 2007 who were also given injections of either darbepoetin alfa (Aranesp) or epoetin alfa (Procrit) during the same time period were reviewed. Abnormal levels for each measure were defined as follows: ferritin (under 100ng/mL), B12 (under 200pg/mL), folate (under 5ng/mL), creatinine (over 1.4mg/dL), reticulocyte count percentage (over 5%), C-reactive protein (over 5mg/dL), erythrocyte sedimentation rate (over 20mm), and erythropoietin (over 20mU/dL).

Results: N=278 Patients

Test Performed Not Performed Normal* Abnormal* *Percentages shown in these two columns indicate the percentage of patients in which the test result was normal or abnormal out of the patients who had the test performed. Ferritin 85.5% (136) 14.5% (23) 42.8% (119) B12 96.4% (132) 3.6% (5) 50.7% (141) Folate 94.7% (126) 5.4% (7) 52.2% (145) Reticulocyte count 91.9% (125) 8.1% (11) 51.1% (142) Creatinine 89.1% (246) 10.9% (30) 0.7% (2) C-reactive protein 20.8% (5) 79.2% (19) 91.4% (254) Erythrocyte sedimentation rate 75% (3) 25% (1) 98.6% (274) Erythropoietin 28.9% (13) 71.1% (32) 83.8% (233)

Conclusion: We conclude that a significant number of patients do not have a proper anemia workup prior to ESA use (or before continued ESA after the first dose). In those patients who did, some laboratory results indicate that there may be another cause for the anemia, in addition to chemotherapy such as iron, B12, or folate deficiency, or renal insufficiency. Given the recent controversy over ESA use, it is particularly important to consider other treatment options prior to administering ESAs, especially given the cost-effectiveness of some of these treatments (FDA Alert: Erythropoietin Stimulating Agents 2006). Oncologists may want to consider identifying and treating other causes of anemia in their cancer patients on chemotherapy prior to, or in conjunction with, ESA use.
Stem Cell Factor Signaling Collaborates in the Development of FIP1L1/PDGFRα Induced Chronic Eosinophilic Leukemia in a Murine Model
Authors: Yamada Yoshiyuki , Rothenberg Marc E , McBride Melissa , Sanchez-Aguilera Abel , Cancelas Jose
Published: April 2020

We have previously demonstrated the synergistic role of stem cell factor (SCF) in systemic mastocytosis associated with FIP1L1/PDGFRA (F/P)- chronic eosinophilic leukemia (CEL) (Yamada Y et al., Blood 2008). The aim of this study is to determine if SCF is also required for eosinophilia development in CEL. First, F/P- or vector control-transduced bone marrow hematopoietic stem cell/progenitors (BM-HSC/P) were cultured in SCF in the presence of hydrocortisone (10−5M, HC-SCF) for 2 or 3 weeks. Expression of F/P in HSC/P induced a 27,000-fold higher cell expansion in the presence of HC-SCF compared with HC-SCF -cultured control vector-transduced HSC/P. The cultured cells were then assayed in an IL-5-containing methylcellulose for analysis of CFU-Eosinophil formation, while the culture was continued with SCF in the absence or presence of HC. Only F/P+ HSC/P formed CFU-Eosinophil after ex-vivo culture in the presence of SCF (19 ± 2.2 CFU-eosinophils/105 cells) and HC-SCF-cultured BM-HSC/P quickly differentiated into mast cells after deprivation of HC (98% c-kit+/EGFP+ cells at two weeks of culture after HC withdrawal), suggesting that SCF in collaboration with F/P fusion, may expand progenitor cells of both eosinophil and mast cell lineages. In addition, we analyzed the role of SCF/ckit signaling in disease development of F/P+ CEL in vivo. CEL mice were developed using retroviral introduction of F/P fusion into HSC/P of IL-5 transgenic mice as previously reported (Yamada Y et al., Blood 2006). CEL mice were treated with a blocking anti-c-kit antibody, ACK-2, or an isotype-matched control antibody. ACK-2 treatment suppressed the circulating leukocyte count, eosinophil count and spleen weight significantly in CEL mice, whereas control (vector control-IL-5 transgenic BM-HSC/P transplanted wild type mice) did not show significant diference between ACK-2 treatment and isotype control one (Table). These results suggest that SCF signaling is required for eosinophilic leukemia development induced by FIP1L1/PDGFRa.

Table Circulating total leukocyte and eosinophilc counts and spleen weight of mice treated with antibodies

Control CEL isotype-control ACK-2 isotype-control ACK-2 Data are shown as mean ± SD, **p < 0.001 compared with isotype control, † p<0.001 compared with control WBC, white blood cells, CEL, chronic eosinophilic leukemia. WBC(mm3) 8805±4377 5480±1433 176278±1210† 9602±12177±68.3** Eosinophil(mm3) 2609±671 804±235 29940±18735† 3480±4292±68.3** Spleen weight (mg) 137±11.3 89.9±211.2 2±11.2** 222.8±68.3**
Proteome Analysis of Apoptotic MR2 Cells, Acute Promyelocytic Leukemia Cell Line with All-Trans Retinoic Acid Resistance, Induced by Arsenic Trioxide
Authors: He Pengcheng , Zhang Mei , Wang Xiaoning , Wang Huaiyu , Xi Jieying , Li Jing , Wu Di , Wei Kaihua , Wang Hongli
Published: April 2020

Although all-trans retinoic acid(ATRA) provides complete remission in 90% patients with untreated acute promyelocytic leukemia(APL), it becomes ineffective to quite a few APL patients who have received ATRA before when their disease relapsed and used ATRA again. Arsenic trioxide(ATO) can make APL patients with ATRA-resistance obtain complete remission again by inducing APL cells apoptosis. However, the molecular mechanisms of apoptosis in ATRA-resistance APL cells induced by ATO remain unclear. For this reason, we take the apoptotic MR2 cells (APL cell line with ATRA-resistance) induced by ATO as a model, to screen and identify the proteins related with ATO-induced apoptosis by comparative proteomics. After MR2 cells were dyed with annexin V and PI staining, the percentage of the apoptotic MR2 cells induced by 1.0μmol/L ATO for 0h, 6h, 12h, 24h and 48h respectively was detected by Flow cytometry. The results showed that the majority of the apoptotic cells were in the earlier and later stage of apoptosis respectively, when MR2 cells were treated with ATO for 24 and 48 hours in sequence. The total proteins of MR2 cells of the control group, the earlier stages apoptosis group and the later stages apoptosis group were separated by two-dimensional electrophoresis(2-DE) respectively. Then, the differences in proteome profile among three groups were analyzed by ImageMaster™ 2D Platinum software. 14 protein pots were selected to be identified by Matrix-assisted laser desorption/ionization time of flight-mass spectrometry(MALDI-TOF-MS), in which the quantity of the protein differentially expressed was more than two times(≥2 or ≤0.5) among MR2-0h, MR2-24h and MR2-48h cells’ 2-DE map. However, only 11 proteins were successfully identified and their definite information was obtained. Among them, there were 8 proteins that were all probably involved in the mechanisms of apoptosis in MR2 cells and they were Calreticulin(CRT), Heat shock 70 kDa protein(HSP70), High mobility group protein B1(HMGB1), Ran-specific GTPase-activating protein(RanGAP1), Elongation factor 1-beta(EF-1β), Beta-tubulin, Cofilin-1, and Prolyl 4-hydroxylase(P4H) respectively. CRT was probably related with the early stage of apoptosis in MR2 cells, while RanGAP1 and HSP70 might related with the late stage of apoptosis in MR2 cells. Moreover, so far there was no related report on the roles of CRT, HMGB1, RanGAP1, cofilin-1 and beta-tubulin in the mechanisms of APL cells apoptosis. These differential proteins identified provide the new clues for further researching the molecular mechanisms of apoptosis in the ATRA-resistance APL cells induced by ATO.
Coordination of Fenretinide-Induced Apoptosis by Stress-Responsive Regulators in Leukemia Cells
Authors: Wang Kankan , Fang Hai , Xiao Da-Kai , Zhu Xue-Hua , He Miao- Miao , Pan Xiao-Ling , Chen Sai-Juan , Zhang Ji
Published: April 2020

Pharmacological intervention affecting intracellular redox balance often results in oxidative stress-mediated apoptosis, which also represents a new direction for cancer therapeutic designs. However, mechanistic links between redox signals and their downstream effects on apoptosis have not yet been elucidated in cancer cells. Here, we report a detailed analysis of fenretinide-induced apoptosis in leukemia-derived cells through a systems approach, integrating experimental and computational methods together with advanced data mining tools. Robust transcriptome profiling reveals numerous stress-responsive events at the temporal and spatial levels, including oxidative stress, endoplasmic reticulum stress/unfolded protein response, translational repression, proteasome activation, and apoptosis induction. Moreover, stress-responsive transcription factors, as highlighted by NRF2 and HSF1, play prominent roles in the configuration of these relevant events. Several lines of evidence suggest that these stress-responsive regulators and thus their target genes are involved not only in converting oxidative signaling into downstream effects but also in coordinating the progression of cell apoptosis. This study provides a roadmap for understanding oxidative stress-mediated apoptosis in cancer cells, which may be further developed into more sophisticated therapeutic protocols, as implicated by synergistic induction of cell apoptosis using proteasome inhibitors with fenretinide.
Identification and Purification of Leukemic Stem-Like Cells in Acute Myeloid Leukemia Cell Line
Authors: She Miaorong , Niu Xingqing , Chen Xilin , Kunyuan Guo , Zhou Maohua , He Yanjie , Le Yi
Published: April 2020

Acute myeloid leukemia (AML) is initiated and maintained by a rare population of (leukemic stem cells) LSCs. LSCs play the central role in the relapse and refractory of AML and highlight the critical need for the new therapeutic strategies to directly target the LSC population for ultimately curing leukemia which is it is important to identify and study LSCs. However, relatively little is known about the unique molecular mechanisms of survival and self-renewal of LSCs because of very small number of LSCs in bone marrow. In this study, we investigated whether established leukemia cell lines contain LSCs. We showed that leukemia cell line contain leukemic stem-like cells which have been phenotypically restricted within the CD34+CD38− fraction. We demonstrated that CD34+CD38− cells could generate CD34+CD38+ cells in culture medium and had proliferation function. Moreover, CD34+CD38− cells had self-renewal potential both in vitro soft agar colonies formation assay and in vivo NOD/SCID mouse xenotransplant model serial transplantation. Furthermore, CD34+CD38− cells isolated from leukemia cell line were found resistant to conventional chemotherapy and NK cells-mediated cytotoxicity and these were related to up-regulation of ABCG2 and MRP-1 and antiapoptotic proteins of Bcl2. Down-regulation of NKG2D ligand also played a critical role in NK cytotoxicity resistance. Taken together, our studies provide a novel cell model for leukemic stem cells research. Our data also shed light on mechanism of double resistant to resistant to chemotherapy and NK cell immunotherapy, which was helpful for developing novel effective strategies for LSCs.
Immnuoregulatory Effect of Human Mesenchymal Stem Cell Is Mediated Via STAT3 Molecule
Authors: Yoon Jinsun , Kim Seoju , Kim Eun Shil , PARK Byoungbae , Choi Junghye , Kim Insoon , Kim Byoung Kook , Lee Young
Published: April 2020

The best curative treatment modality in hematologic malignancies is allogeneic hematopoietic stem cell transplantation (HSCT). Graft-versus-host disease (GVHD) is a major obstacle of allogenic HSCT. Bone marrow derived human mesenchymal stem cells (MSCs) is known to have immunoregulatory effect in vitro and in vivo via inhibiting alloreactive T lymphocytes, leading to their clinical use for the prevention of GVHD in HSCT. However, the molecular mechanism of immunoregulatory effect of human MSCs is not fully understood. In this study we investigated the signaling of immunoregulatory effect by co-culture of human MSCs with lymphocytes. The proliferation of allogeneic T cells was strongly inhibited. The fraction of CD4+CD25+Foxp3+ cells (Treg cells) was increased, while the fraction of CD4+, CD8+, CD25+ was decreased. In addition, induction of Th1 to Th2 shift was observed. Western blot study showed that phosphorylation of STAT1, STAT3, STAT6 was up-regulated, but STAT1, STAT4, ERK, AKT, NF-κB (p65, p50 subunits) was down-regulated. While expression of STAT3 was observed in culture of MSCs only, no expression of STAT3 was shown in co-cultured human MSCs with lymphocytes. In order to validate our results, expression of STAT3 in human MSCs co-cultured with lymphocytes was ablated using small interfering RNA. As a result, inhibition of CD4+CD25+FoxP3+ cells, proliferation of allogenic T lymphocytes, inhibited induction of Th1 to Th2 shift and proliferation of CD4+, CD8+, CD25+ cells. Furthermore, expressions of Th1 and Th17-related cytokines were increased, while expressions of Th2-related cytokines were decreased. In summary, these results suggest that STAT 3 may be an indispensable molecule in the immunoregulatory effects in human MSCs via modulation of regulatory T cells.
Mesenchymal Stem Cell Differentiation Markers Shared with Soft Tissue Sarcomas
Authors: Wirths Stefan , Buehring Hans-Joerg , Kanz Lothar , Hartmann Joerg T , Kopp Hans-Georg
Published: April 2020

Malignant tumors are hypothesized to harbor small populations of self-renewing cancer stem cells. Targeting these cells may be the decisive step to overcome treatment resistance and achieve tumor eradication in cancer patients. Advanced soft tissue sarcomas (STS) are rare tumors with a dismal prognosis and a small number of systemic treatment options. STS may originate from mesenchymal stem cells (MSC); the latter have mainly been isolated from adult bone marrow (BM) as non-hematopoietic, self-renewing cells whose in vitro progeny comprises osteoblasts, chondroblasts, myocytes, and adipocytes. While in vitro expression profiles of MSC have been investigated extensively, the in vivo counterparts of MSC are still hypothetical. To target rare human cell BM populations including MSC, an exclusive antibody panel was developed. The target antigens include platelet-derived growth factor receptor-β (CD140b), HER-2/erbB2 (CD340), the recently described W8B2 antigen as well as several surface antigens identified by novel antibodies. To define the expression pattern of MSC-markers in STS, three STS cell lines were tested for expression of these antigens. In addition, snap-frozen primary STS sections were analyzed by immunohistochemistry using the same antibody panel. All cell lines revealed expression of selected markers including CD340, W8B2, and CD140b. Several MSC markers were restricted to a subpopulation of cells. In addition, leiomyosarcoma cells displayed a different expression pattern as compared to liposarcoma and Ewing’s sarcoma cells. Results of immunohistochemistry analysis of primary leiomyosarcoma tumor samples correlated strongly with expression patterns established by FACS analysis. However, important cytoarchitectural features regarding selected markers were revealed by immunohistochemistry: while primary leiomyosarcomas displayed uniform expression of W7C6, HEK3D6, CD10, and CD318, other markers such as CD34, W5C5, and 57D2 were expressed by tumor endothelia only. Moreover, a population of perivascular tumor cells was found to express the MSC-markers W4A5, W8B2, CD140b, W3D5, and W5C4. Novel MSC-markers are expressed by subpopulations in STS cell lines as well as in primary sarcoma tissue. Further studies on the functional significance of these phenotypical studies are underway and may help to identify novel specific targets recognizing the self-renewing STS-compartment.
Gata1 Regulates Erythroid Transcription by Cooperating with Chromatin Remodeling Protein Snf2h
Authors: Podskocova Jarmila , Burda Pavel , Vargova Karin , Kokavec Juraj , Curik Nikola , Zavadil Jiri , Skoultchi Arthur , Stopka Tomas
Published: April 2020

Gata1 is transcription factor that regulates erythropoiesis and its direct interaction with chromatin remodeling protein Snf2h may affect chromatin structure (Rodriguez 2005). Snf2h belongs to SWI/SNF2 superfamily of ATPases regulating structure of nuclear chromatin by nucleosome movement and assembly. Snf2h knockout in mice is embryonic lethal and heterozygotes display mild growth retardation (Stopka 2003). We studied nuclear localization of Snf2h and detected its presence in euchromatin and to a lesser extent in heterochromatin. Decreased Snf2h levels in Snf2h heterozygotes and Snf2h-null embryos exhibit significantly decreased heterochromatin size. In addition, histone modifications associated with transcription activation (histone H3K79 dimethylation and H4K16 acetylation) are globally decreased in Snf2h mutants. To test the involvement of Snf2h in hematopoiesis, ectopically expressed Snf2h mutants were tested in Gata1-mediated transcription assay in HeLa cells and demonstrated that Snf2h efficiently repressed Gata1 transactivation. Testing whether the ATPase domain is required for the repression mechanism we found the Snf2h dominant negative mutant (DN) can also repress Gata1-dependent transcription in both HeLa and Snf2h +/− fibroblasts. We next studied the effect of Snf2h DN mutant on histone modifications downstream the Gata1 binding site and found that Snf2h DN further increases H3K79 dimethylation induced by Gata1. In contrast, an occupancy of histone H3 downstream the Gata1 binding site was significantly reduced by Snf2h DN mutant indicated it caused a defect in chromatin remodeling. Collectively, our data demonstrate a cooperative role of Gata1 and Snf2h in erythroid transcription regulation and propose that Snf2h in both ATP-dependent and ATPindependent manner represses transcription by disrupting the regular array of nucleosomes near Gata1 binding sites.
Identification of a Novel Acetyl-CoA Binding Protein (EST1) Which Epigenetically Regulates the Expression of Erythroid-Specific Genes
Authors: Fujiwara Tohru , Ikura Tsuyoshi , Ichikawa Satoshi , Takahashi Shinichiro , Furuyama Kazumichi , Igarashi Kazuhiko , Harigae Hideo
Published: April 2020

(Introduction) Embryonic stem (ES) cells that lack 5-aminolevulinate synthase-2 (ALAS2) gene provide a valuable model in dissecting molecular events in which heme is required during erythroid differentiation. Recently, we identified a novel acetyltransferase-like gene (EST1) through differential expression analysis between wild-type and heme-deficient erythroblasts as a potential downstream target gene of heme (BBRC2006;340:105–110, ASH 2007). EST1 belongs to the GNAT (GCN5-related N-acetyltransferase) superfamily since it contains the highly conserved amino acid residues (motif A), known as acetyl-CoA binding domain. Since the GNAT superfamily contains a wide variety of acetyltransferases with different substrates or with unknown functions, probing how EST1 might be required for biological events in erythroid cells, could be informative. Here, we investigated the role of EST1 during erythroid differentiation.

(Methods) EST1 was constitutively expressed using Flag/HA-tagged retroviral vector into mouse erythroleukemia (MEL) cell line. EST1 protein complex was purified by affinity chromatography from nuclear extract of EST1-expessed MEL cells. To obtain dominant-negative EST1-expressing cells, both Arg-62 and Gly-65 within EST1 were substituted to glutamic acid, and similarly transduced into MEL cells. These mutations have been widely applied for abolishment of acetyl-CoA binding activity. For depletion of endogenous EST1, siRNAs specific for EST1 were introduced into Hepa1c1c7 cells

(Results) Although recombinant EST1 protein did not have acetylase activity for free histones in vitro (ASH 2007), EST1 protein forms a multimeric protein complex. Western blot analysis using FLAG-eluted polypeptides revealed the presence of GCN5, TRRAP, SPT3 and GATA-1, implying that this protein complex might participate in the transcriptional regulation of erythroid-specific genes. Following EST1 depletion in Hepa1c1c cells, a significant decrease in the acetylation of H3 and a mild decrease in that of H4, were observed by Western blot analyses. Similarly, a significant decrease in the acetylation of H3 and a mild decrease in that of H4 were also observed in dominant-negative than in wild-type EST1-expressing MEL cells. Furthermore, the level of bmajor and ALAS2 mRNA were significantly lower in dominant-negative than in wild-type EST1-expressing MEL cells upon treatment with 1.5% DMSO for both 48h and 72h. We are currently exploring mechanisms of how EST1 participates in the regulation of histone modification in erythroid cells.

(Conclusion) EST1 may epigenetically regulate a subset of erythroid-specific genes under control of heme. Further elucidation of the function of the EST1 gene would enhance our understanding of the transcriptional network involving erythroid differentiation.
A Probabilistic Integrated Gene Network Model of Multiple Hematopoietic Lineage Specification
Authors: Fang Hai , Wang Kankan , Zhang Ji
Published: April 2020

Hematopoietic stem cells and their progenitor hierarchy are highly controlled by the underlying gene regulatory network. In past decades, great progress has been made in elucidating lineage-restricted transcription factors and lineage-specific gene expression patterns. With accumulation of genome-wide biological data, it is of great value to expand beyond mere transcriptional regulatory analysis to the systemic understanding. Here, we utilized a probabilistic integrated gene network for integrating heterogeneous functional genomic and proteomic data sources into predictive model of hematopoietic lineage diversification. We first constructed a naïve Bayesian network by incorporating disparate biological data, including time-series gene expression during re-dedifferentiation of leukemia along alternative paths into granulocyte or monocyte upon the treatment of differentiation-inducing agents, computationally generated transcription factor regulatory sites and microRNA targets, well-curated physical protein-protein interaction, and functional annotation data. The resultant network, coupled with binomial-based statistical analysis of the interplay between node properties and the network topology, predicted the specific hematopoietic lineage, generating testable hypotheses regarding their unified reprogramming principles. This study demonstrates the utility of the growing biological data in detailed elucidations of the orchestration of distinct hematopoietic cell fates.
Hematopoietic Stem/Progenitor Cell Mobilization in Donors and Recipients after Living Donor Liver Transplantation
Authors: Masuya Masahiro , Shiraki Katsuya , Ohishi Kohshi , Nakamura Shiho , Miyazaki Hiroshi , Katayama Naoyuki
Published: April 2020

Combination therapy with interferon-α and anti-viral drug is the current treatment of choice for chronic hepatitis viral infection. However, an important number of patients fail to respond to this therapeutic strategy and develop cirrhosis or liver cancer. Liver transplantation (mainly living donor liver transplantation (LDLT) in Japan) is known to be the ultimate therapy for them. To clarify the association of liver regeneration and hematopoietic stem/progenitor cells (HSCs), we quantified the number of circulating CD34+ cells (CD34+ cell number) and serum concentration of thrombopoietin (TPO) before and after surgery in 10 donors and 9 recipients undergoing LDLT. The evaluation of CD34+ cells has been carried out by flow cytometry, by applying conventional protocols. In donors, CD34+ cell number increased from 3.69 ± 2.87 cells/μL at pre-LDLT to 8.25 ± 4.94 cells/μL at post-operative day (POD) 7. Platelet count increased from 209 ± 50 × 109/L at pre-LDLT to 273 ± 23 × 109/L at POD 7–11. In recipients, although CD34+ cell number was 0.78 ± 0.73 cells/μL at pre-LDLT and significantly lower than that in healthy volunteers (3.78 ± 3.93 cells/μL), the number at POD 9–42 increased to 4.25 ± 3.55 cells/μL. Platelet count increased from 64 ± 35 × 109/L at pre-LDLT to 197 ± 91 × 109/L at POD 9–42. Serum TPO concentration at pre-LDLT was 0.98 ± 0.61 fmol/L in donors and 1.00 ± 0.35 fmol/L in recipients. TPO level peaked at POD 3–7 (3.14 ± 1.67 fmol/L) in donors and at POD 7–22 (6.74 ± 3.13 fmol/L) in recipients. It has been reported that mobilization of HSCs after partial hepatectomy in donors or LDLT in recipients was likely enhanced by increased serum levels of granulocyte colony-stimulating factor (peaked at POD 1), interleukin-6 and stem cell factor (POD 7). However, in our study, TPO (peaked at POD 7–22) was strongly associated with not only the increase of platelet count (peaked at POD 9–42) but also the mobilization of HSCs (POD 9–42) in recipients. Both mobilized HSCs and platelets may contribute to the liver regeneration after LDLT.
Array-CGH Identifies Regions, Including the FOXP1 Locus, Associated with Different Clinical Outcome in Diffuse Large B-Cell Lymphomas (DLBCL) Treated with R-CHOP
Authors: Scandurra Marta , Rancoita Paola M.V. , Greiner Timothy C. , Chan Wing C. , Vose Julie M. , Inghirami Giorgio , Chiappella Annalisa , Baldini Luca , Ponzoni Maurilio , Ferreri Andres JM , Montes-Moreno Santiago , Piris Miguel A. , Franceschetti Silvia , Gaidano Gianluca , Facchetti Fabio , Tucci Alessandra , Lazure Thierry , Lambotte Olivier , Uccella Silvia , Pinotti Graziella , Pruneri Giancarlo , Martinelli Giovanni , Nomdedeu Josep , Tibiletti Maria Grazia , Chigrinova Ekaterina , De Campos Cassio P. , Rinaldi Andrea , Zucca Emanuele , Kwee Ivo , Bertoni Francesco
Published: April 2020

Background. Diffuse large B-cell lymphoma (DLBCL) consists of a heterogeneous group of tumors. More than 30% of cases are not cured with R-CHOP chemotherapy.

Aim. To identify genomic aberrations that could affect the response to therapy, we performed an arrayCGH study on uniformly treated DLBCL patients.

Materials and Methods. Tumor samples were analyzed with Affymetrix Human Mapping 250K SNP arrays. Eligibility criteria were diagnosis of de novo DLBCL, first line treatment with R-CHOP or R-CHOP-like regimens, availability of frozen biopsy and of clinical baseline and follow-up data. Exclusion criteria were: primary mediastinal DLBCL, primary central nervous system DLBCL, HIV-positivity. Material has been collected according to the local IRB guidelines. Results. Genomic tumor profiles have been obtained in 163 samples from 10 Institutions; 23/163 cases were excluded from this analysis because the information on response was not available; 140/163 cases fulfilled the study requirements. The clinical parameters of the patients reflected the normal DLBLC population, as shown by the IPI score distribution (0–1 in 30%, 2 in 37%, 3 in 23% and 4–5 in 10%). The median follow-up was 23 months (range 1–1251). Complete remission was observed in 108/140 (77%) patients, partial response in 21 (15%), stable disease in 6 (4%) and progression in 5 (2%). Genomic differences were observed between complete responders (108 cases) and the remaining patients, grouped together as poor responders (32 cases). The latter group had more gains of 3p14.1 (FOXP1 locus), 3q29, 11q24.3, and losses of 2p11.2-p13.3, 8p23.1- pter, 10p12.31-p13, 15q11.2-q14, 15q21.1 and copy neutral LOH of chromosome 9p. On the converse, 1q gains were more common among patients achieving complete remission. No differences were observed for other common region of gains (7, 12, 18q/BCL2) or losses (1p, 6q, 17p/TP53).

Conclusions. Specific genomic aberrations are associated with the response to R-CHOP in patients with DLBCL. In particular, the gain of the 3p14.1 (FOXP1 locus), which is associated with a lack of response to R-CHOP, suggests that the role of FOXP1 should be further investigated in DLBCL.
Identification and Characterization of Hematopoietic Stem (HSC)/Progenitor Cells in Human Solid Tumors
Authors: Leung Roland C , Dorn David C , Downey Robert J , Levine Douglas A , Moore Malcolm A.S.
Published: April 2020

It has recently been shown that a solid tumor recruits cells to the tumor mass, which facilitate vascularization, suppress immune responses, aid its growth, and influence metastatic potential. These recruited cells include mesenchymal stem cells, endothelial progenitor cells and tumor infiltrating leukocytes. By performing multiparameter FACS analysis on primary lung carcinoma and pleural effusion material, we demonstrate for the first time the consistent presence of CD45+/CD34+ cells (mean 1.6% of total CD45+cells range 0.24–13.2% n=12). In primary tumor samples, CD45+ lymphohematopoietic cells were purified by the MACS system and examined for progenitor colony formation in methycellulose. Erythroid and myeloid colony formation was observed in the CD45+ selected subfraction. We assayed for HSC activity by cobblestone formation (CAFC) on MS-5 stroma. Week 2 and week 5 CAFC potential was present in CD45+ purified ovarian cancer ascites. In non-irradiated NOD/SCID IL2 gamma null mice injected subcutaneously with dissociated and unfractionated lung tumor and ovarian peritoneal ascites, small numbers of human CD45+ cells were recovered from the bone marrow after 2–3 months which gave rise to hematopoietic colonies. Human CD45+ cells also persisted within the retransplanted subcutaneous tumors. In conclusion, we report the recruitment of HSCs and progenitors to primary tumor sites. The differentiation and self-renewal potential of these HSCs indicate a potential alternative source of tumor-infiltrating leukocytes as opposed to or in addition to recruitment of mature cells from the circulation.

The role of solid tumors in mobilizing HSCs and progenitors from the bone marrow and their selective chemotaxis to the primary and metastatic sites require further investigation. Targeting this novel tumor-HSC interaction may offer new therapeutic opportunities for controlling tumor growth and metastasis.
Nitric Oxide and Vascular Endothelial Growth Factor during Pregnancy in Sickle Cell Disease
Authors: Brittain Julia E. , Ataga Kenneth I , Lee Sean , Thorp John M. , Orringer Eugene P.
Published: April 2020

BACKGROUND: Vascular Endothelial Growth Factor (VEGF) is a multifunctional cytokine that may be clinically relevant in sickle cell disease (SCD). In additional to its role as a potent activator of angiogenesis, VEGF is also a potent inducer of Nitric Oxide (NO) release. Patients with SCD have impaired NO metabolism and decreased bioavailability of NO due to chronic hemolysis which may contribute to the vasculopathy that is present in this disorder. High levels of plasma arginase and the resultant decreased arginine/ornithine ratios have also been proposed to contribute to low NO levels in SCD patients. We have previously reported that pregnant SCD patients have higher circulating levels of the naturally occurring VEGF antagonist, sFLT-1, when compared to pregnant African American controls. High levels of sFLT-1 are associated with poor pregnancy outcomes including preeclampsia and delivery of small for gestational age (SGA) infants.

OBJECTIVE: In this study we wanted to assess the levels of NO, free VEGF, sFLT-1 and arginase activity during pregnancy in SCD. We also sought to explore the relationship between NO and sFLT-1 with pregnancy outcome.

METHODS: We performed an analysis of previously collected plasma/serum samples obtained from the Pregnancy, Infection and Nutrition (PIN) study cohort at UNC- Chapel Hill. Subjects were matched on race, gestational age, BMI and smoking status. Levels of free VEGF were quantified by luminex analysis and confirmed by ELISA. NO was quantified via its metabolite NOx using a modified Griess method in de-proteinated serum/plasma. Arginase activity was quantified by ELISA. Pregnancy outcomes were previously recorded by PIN study personnel.

RESULTS: We found that levels of free VEGF and NO were significantly lower in pregnant women with SCD than in matched pregnant controls (VEGF: 5.8 pg/ml vs. 13.56 pg/ml, p = 0.03, NOx: 14.31 μM vs. 26.99 μM, p=0.0009). We also found a significant inverse correlation between NOx levels and sFLT-1 levels (r2=0.6421, p = 0.0094), but no relationship between NOx and arginase activity in plasma. There was also no difference in arginase activity between pregnant SCD subjects and controls. Finally, pregnant SCD subjects that delivered an SGA infant had significantly lower levels of NOx than those women that did not.

CONCLUSIONS: These results suggest that VEGF may play a role in NO release during pregnancy in SCD. In addition, low NO levels may contribute to poor pregnancy outcomes in pregnant SCD women.
Markers of Coagulation Activation and Inflammation in Sickle Cell Disease and Sickle Cell Trait
Authors: Adam Soheir S. , Zaher Galila , Ibrahim Mahmoud , Ataga Kenneth I , De Castro Laura M. , Key Nigel S.
Published: April 2020

It has been well-described that sickle cell disease (SCD) due to HbSS or HbSC is associated with systemic activation of coagulation. A single study has reported a similar, albeit lesser, degree of chronic hyperactivation of coagulation in subjects with sickle cell trait (SCT) (Am J Hem2002; 69(2):89–94), and we have shown that SCT is a risk factor for venous thromboembolism (Blood2007;110(3):908–12). The aim of this study was to simultaneously compare markers of activation of coagulation and inflammation in SCD and SCT. Ninety Saudi Arabian individuals (40 SCD (HbSS), 20 SCT (HbAS) and 30 controls (HbAA)) were recruited. SCD patients were not in vaso-occlusive crisis at the time of sample collection. None of the subjects was taking any relevant medication (anticoagulants, hormones or hydroxyurea), nor had they received blood transfusion for at least 6 weeks. On all subjects, complete blood count (CBC), prothrombin time (PT), activated partial thromboplastin time (APTT), D-dimer antigen, protein C activity, protein S activity, antithrombin (AT) activity, fibrinogen concentration, activated protein C resistance ratio (APCR), and C-reactive protein (CRP) levels were obtained. Student’s “t” test was used to compare two independent means. Multivariate analysis “ANOVA” test was done to test statistical difference between more than 2 means.

Results: D-dimer and CRP were significantly higher, whereas protein C, protein S, and fibrinogen were significantly lower in SS patients compared to controls (Table). In SCT, D-dimer levels were higher, and levels of proteins C and S were significantly lower than controls. APC resistance was more pronounced in both SCD and SCT compared to controls. In summary, this study provides further evidence of hypercoagulability in SCD, but more intriguingly, confirms that significant changes in coagulation parameters may be found in SCT. We speculate that loss of asymmetry of phospholipids in the red cell membrane may occur in certain hypoxic vascular beds (such as the renal vasculature) in SCT. We are currently exploring other possible underlying mechanisms that may contribute to activation of coagulation in SCT.

Table: Comparison of Markers of Inflammation and Coagulation Activation in all Groups

Group I (SS) (n=40) Group II (AS) (n=20) Group III (AA) (n=30) *P < 0.05 comparing SCD and control groups by paired t- test **P <0.05 comparing SCT and control groups by paired t- test CRP 22.5 ± 10.9* 0.3 ± 0.2 0.3 ± 0.1 PT 12.5 ± 1.6* 12.2 ± 1.0** 11.5 ± 0.6 APTT 33.3 ± 4.2 35.8 ± 4.6** 32.9 ± 3.2 D-dimer 287 ± 140* 278 ± 207** 141 ± 65 Protein C 77.9 ± 20.3* 97.7 ± 21.7** 109.4 ± 17.2 APCR 0.82 ± 0.12* 0.79 ± 0.12** 1.05 ± 0.19 Protein S 54.0 ± 18.4* 64.6 ± 15.4** 89.7 ± 14.9 AT 94.4 ± 16.6* 102.2 ± 7.4** 109.3 ± 10.7 Fibrinogen 305 ± 101 361 ± 146 358 ± 80
Pulmonary Hypertension as a Risk Factor for Death in Patients with Sickle Cell Anemia
Authors: Cancado Rodolfo D , Olivato Maria Cristina A , Filho Newton Nunes Lima , Campos Orlando , Chiattone Carlos
Published: April 2020

Pulmonary hypertension develops in most forms of hereditary and chronic hemolytic anemia, including sickle cell disease, thalassemia, hereditary spherocytosis, and paroxysmal nocturnal hemoglobinuria, suggesting that there is a clinical syndrome of hemolysis-associated pulmonary hypertension. Retrospective studies from tertiary care referral centers suggest a prevalence of pulmonary hypertension in adults with sickle cell disease ranging from 20 to 40%. Despite the fact the elevations in pulmonary artery pressures are slight, morbidity and mortality are high. In adult sickle cell anemia patients, pulmonary hypertension is emerging as a major risk factor for death. We performed Doppler echocardiographic assessments of pulmonary-artery systolic pressure in 80 consecutive patients (20 men and 60 women; mean [±SD] age, 30 ± 10.8 years) between 1/20/2006 and 1/20/2008. The genotype on the basis of hematologic and hemoglobin characteristics was hemoglobin SS in all patients. Pulmonary hypertension was prospectively defined as a tricuspid regurgitant Jet velocity (TFJV) of at least 2.5 m per second. Patients were followed for a mean of 18 months (6–24 months), and data were censored at the time of death or loss to follow-up. Doppler-defined pulmonary hypertension occurred in 37.5 percent of patients (30/80). Multiple logistic-regression analysis, with the use of the dichotomous variable of a tricuspid regurgitant jet velocity of less than 2.5 m per second or 2.5 m per second or more, identified age, female sex, deferasirox therapy, left ventricular mass index, pulmonary artery systolic pressure, reticulocytes, white-cell count, platelet count, lactate dehydrogenase (a marker of hemolysis), blood urea nitrogen, creatinine, uric acid and self-reported history of cardiovascular complication, billiary stones, retinopathy and acute chest syndrome, as significant independent correlates of pulmonary hypertension. The hemoglobin level, fetal hemoglobin level, hydroxyurea therapy and serum ferritin level were unrelated to pulmonary hypertension. Hazard rate for death according to the TFJV of at least 2.5 m per second, as compared with a velocity of less than 2.5 m per second, was associated with an increased risk of death (0.00 versus 2.54; P=0.998). Mortality rate in 24 months was 6.7% (2/30) for patients with TRJ velocity ≥ 2.5 m/sec versus 0.0% (0/50) for patients without pulmonary hypertension. Pulmonary hypertension, diagnosed by Doppler echocardiography, is common in adults with sickle cell disease. It appears to be a complication of chronic hemolysis, is resistant to hydroxyurea therapy, and confers a high risk of death. Large trials evaluating the effects of treatment for pulmonary hypertension in the sickle cell anemia population are indicated.