bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Distinct Stem Cell Population Derived From Chorionic Plate Of Human Placenta Which Has Differentiation Potential Into Erythroid Lineage Cells
Authors: Park Yong , Kim Ji Hye , Jung Ji Hye , Kang Ga-won , Kim Dae Sik , Kim Seok Jin , Kim Byung Soo 
Published: October 2019

Abstract

Background/Aims
Placenta is a specialized organ which has a property of fetal and adult tissues simultaneously. Mesenchymal stem cell and hematopoietic stem cells have been founded in the parenchyma of placenta tissue contributed by fetal side. Recently, it was reported that placenta is a major hematopoietic organ contributing to both generation and expansion of multipotent hematopoietic stem and progenitor cells. We hypothesized that hematopoietic stem cells in placenta might be originated from another placental stem cells which has been so-called “placenta mesenchymal stem cells”. To prove this hypothesis, this study is designed to induce in vitro differentiation of placental stem cells into the hematopoietic stem cells without genetic transduction.

Methods
Placental stem cells was obtained from surgically isolated placental chorionic plates from healthy women who had undergone abortion at 6-8 weeks of gestation. To prevent the contamination of hematopoietic stem cells, CD34-CD44+ cells were purified by flowcytometric isolation. After mesenchymal differentiation potential was identified in these cells by induction of adipogenic and osteogenic differentiation, they were cultured in vitro with serum-free media including cytokine cocktail which was known to induce differentiation of embryonic stem cells into hematopoietic lineage (erythropoietin, Flt-3/Flk-2 ligand, G-CSF, GM-CSF, IL-6/7/15, stem cell factor, and thrombopoietin). In vitro differentiation was performed in conventional cell culture dish (group 1) and Ultra-low attachment dish (group2) for 3 weeks, respectively (Figure 1). Expression of hematopoietic stem and progenitor cell markers were analyzed at every 1 week.

Results
In group 1 (conventional cell culture dish), no hematopoietic differentiation was identified regardless of the concentrations of cytokine cocktail. In group 2 (Ultra-low attachment dish), expression of pan-hematopoietic markers, CD34 and CD45 was also not observed. However, after 2 weeks of differentiation induction, red cells aggregates and expression of erythroid-lineage markers (CD235a and CD38) were detected in cells cultured using high concentration cytokines (Media 3 in Figure 1). The proportion of cells expressing erythroid-lineage markers became increased during in vitro differentiation (Figure 2).

Conclusion
Although this study failed to show the differentiation of placenta-derived stem cells into hematopoietic stem cells, the results demonstrated that placenta-derived stem cells could be differentiated into the erythroid lineage under certain condition (i.e. liquid culture + high concentration cytokines). Therefore, placenta-derived stem cells which have been so-called “placenta mesenchymal stem cells” is considered to be a distinct stem cell population including property of erythroid progenitor cells.

Disclosures:
No relevant conflicts of interest to declare.
The Hematopoietic Stem Cells Could Be Switched From Leukemic Cell Lines
Authors: Hu Jianda , Yang Ting , Zhu Liangfang , Luo Xiaofeng , Yuan Xiaohong , Chen Zhizhe 
Published: October 2019

Abstract

Background
The hematopoietic potential from induced pluripotent stem cells (iPS) has been explored for some years. However, the direct reprogramming from the blood cells to hematopoietic progenitors bypass the pluripotency is even more attractive for the clinical practice.

Methods
The lentiviral vector encoding human reprogramming factors (Oct-4, Sox2, c-Myc, Klf4FOSCK) were transfected along with packaging plasmid pCMV-dR8.91 and envelope plasmid pCMV-VSV-G in 293T package cells to produce lentiviral particles. The construction was then confirmed by RT-PCR and restriction enzyme digestion anaysis, and the lentivirus titer was determined using GFP/DAPI based on cell count by Image J software. Primary mouse embryo fibroblast (PMEF) feeder cells were isolated and prepared from CF-1 inbred mouse strain to help maintain pluripotency and to provide a cellular matrix for stem cells growing. Lastly, the lentivirus carrying OSCK factors was used totransduce HL-60 cells in three consecutive rounds of spin-infection on the prepared feeder layers with an interval of 12 hours, while the lentivirus carrying GFP served as a negative control. The cell clusters were picked based on morphology and alkaline phosphatase(AP) staining. The stem cell properties were tested by the expression of cell surface antigens using Flow cytometry. and the mRNA expression of OCT4, SOX2, C-MYC,KLF4,etc. by QT-PCR.

Results
The lentiviral particles were successful packaged and used to infect HL-60 cells. We occasionally observed some Human embryonic stem cells (hES) cell-like colonies in between the cells around 14 days after infection. These cell colonies also showed similarity to hES cells in feeder dependency. Detection of cell surface CD34 by FCM showed that HL-60 cells were switched to be a hematopoietic fate, expression of CD34 from 1.77% to 98.42%. Simultaneously, expressions of the myeloid antigen CD13, CD117 decreased. The gene expressions of OCT4, SOX2 indicated that the exogenous gene were down-regulation or silence while the endogenous gene were up-regulation. However, the cell colonies can survive only for a short time which might due to the absence of the survival factors they require or the first hematopoietic microenvironment, like the yolk sac.

Conclusion
After being reprogrammed, the HL-60 cell derived colonies showed the similarity to hES cells in morphology, feeder dependency and the expression of stem cell antigens. However, they may need an appropriate microenvironment at different time points to be programmed into primitive hematopoietic stem cells.

Disclosures:
No relevant conflicts of interest to declare.
Value Of Magnetic Resonance Imaging In The Study and Follow-Up Of Paroxysmal Nocturnal Hemoglobinuria
Authors: Villegas Ana , Gonzalez Ataulfo , Matute Fatima , Nieto Jorge Martinez , de la Fuente Felix 
Published: October 2019

Abstract

Introduction
Renal damage is relatively frequent in Paroxysmal Nocturnal Hemoglobinuria (PNH). Iron accumulation from hemolysis in the renal tubules has been found to be one of the triggering factors. Hemosiderin accumulates in the epithelial cells of the proximal renal tubules localized in the renal cortex, and they can be measured (and monitoring) using magnetic resonance imaging (MRI).

Methods
We studied 7 PNH patients with MR using T2-weighted gradient echo and multiecho sequences. We quantified renal, hepatic and myocardial iron with T2* relaxometry model. Four of these patients were studied at diagnosis and after 1 year of treatment with eculizumab. In 2 of them the MRI was done after 2 and 3 years respectively of treatment with eculizumab. One patient was studied after 8 months on eculizumab, and MRI was repeated at 12 and 24 months. Liver and myocardial iron deposition was quantified in all cases.

Results
At diagnosis the median hemoglobin of the 7 patients was 8.9 g/dl (8.2-12-4), the PNH median PNH clone size was 77% (60-90) and the median LDH level was 2340 U/L (1100-3600). Three patients presented an important accumulation of iron in the renal cortex, with much lower signal intensity than the medulla in the T2-weighted MR images. One patient with mild hemolysis (Hb 12.4 g/dl, 20% reticulocytes and 1,100 LDH) did not present iron accumulation in the renal cortex, and neither did the 2 patients studied after 2 a 3 years of eculizumab treatment. All the patients improved, presenting median of Hb of 11,2 g/dl ( 9,5-12,9) and of LDH of 520 U/L ( 430- 721). The patient studied after 8 months showed an increase of iron in the renal cortex that persisted, despite the improvement with the treatment with eculizumab, with increase of the Hb and decrease of the LDH levels; this patients had a C hepatitis and positivity on the hemochromatosis genes C282Y/H63D, with important iron hepatic overload (11-17 mgFe/g). In the rest of the patients the hepatic iron was normal, with values from 0.5 to 2 mgFe/g, except in one case where the levels were higher (10 mgFe/g) due to previous transfusions used to treat an aplastic anemia pre-PNH. Myocardial iron was normal in the 7 patients, with values ranging from 33 to 60 ms in T2. Only one of the patients presented several episodes of acute renal insufficiency, related with hemolytic crisis, with residual proteinuria, that disappeared when treated with eculizumab.

Conclusions
A diffuse signal loss in the renal cortex without liver and spleen alteration is indicative of intravascular hemolysis, suggesting PNH. Serial MRI studies of iron overload in the renal cortex can be used for the diagnosis, and for monitoring the effects of treatment.

Disclosures:
No relevant conflicts of interest to declare.
Comparison Of Flow Cytometric and Clinical Findings In Patients With Paroxysmal Nocturnal Hemoglobinuria
Authors: Demirer Taner , Baslar Suphi , Uysal Vasif akin , Beksac Meral , Ozen Mehmet , Konuk Nahide , Ilhan Osman , Arslan Onder , Ozcan Muhit , Topcuoglu Pervin , Gurman Gunhan 
Published: October 2019

Abstract
Paroxysmal Nocturnal Hemoglobinuria (PNH) is a rare disorder of blood cells associated with mutations of X-linked gene called phosphatidylinositol glycan class A, and presenting as hemoglobinuria, signs of cytopenias (fatigue, easy bleeding) or thrombosis.
In this study, 39 patients with PNH were retrospectively evaluated with regard to disease findings, laboratory investigations, complications and relationship between those and clone size. Median follow-up time was 26 months. Patients were divided into two groups. There were 24 classical PNH and 15 Aplastic Anemia (AA) or Myelodisplastic Syndrome (MDS) associated PNH patients. Evident signs of hemolysis at the time of diagnosis and thrombosis were seen in classical PNH patients. Sign of hemolysis developed in two AA patients. Hemoglobinuria, hemolysis test (increased reticulocyte and lactate dehydrogenase) were determined to be correlated with clone size. There was association between clone size and thrombosis. No patients with clone size smaller than 50% developed thrombosis and all patients with thrombosis were in classical PNH subtype which has greater clone size. PNH is a rare disease; therefore the effect of small number of patients on the statistical parameters must be taken into consideration. Over the entire course of follow-up time 8 patients died, 6 due to complications of allogeneic stem cell transplantation and 2 due to conditioning regimen before transplantation. Numbers of deaths are equal in each subtype.
In summary, in this study there was a linear correlation between hemoglobinuria and LDH levels with clone size, which was statistically significant (p=0,031 and p=0,001 respectively). Other clinical signs did not correlate with clone size. Thrombotic complications were seen only in classical type PNH patients. No patients with clone size smaller than 50% developed thrombosis, MPV levels were significantly higher than others (p=0,04). There was also statistically significant correlation between reticulocyte and LDH levels with thrombotic events (p=0,009 and 0,003 respectively). In addition, after allogeneic stem cell transplantation 9 patients were evaluated for PNH clone and in 7 the clone was disappeared.

Disclosures:
No relevant conflicts of interest to declare.
Memory B Cells Display Changes In The Expression Pattern Of Genes Related To Cell Survival In Elderly Individuals
Authors: Baez Alicia , Alvarez-Laderas Isabel , Piruat Jose Ignacio , Caballero-Velazquez Teresa , Millan-Ucles Africa , Barbado María Victoria , Medrano Mayte , Garcia-Guerrero Estefania , Sanchez-Abarca Luis Ignacio , Perez-Simón Jose Antonio , Espigado Ildefonso 
Published: October 2019

Abstract

Introduction
Memory B cells (MBCs) have a very long half-life, remaining viable in a quiescent state for years. However, elderly individuals have less amount of MBCs and they produce lower amounts of antibodies. Thus, in aged individuals immunization become less efficient in terms of quantity and quality. Furthermore, several hematologic malignancies involving B-cell lineage, such as non-Hodgkin lymphomas, chronic lymphocytic leukemia or multiple myeloma, are increasingly common in aging population. With this background, we analyzed the gene expression patterns of naïve B cells (NBCs) versus MBCs in both younger and older subjects, in order to identify genes related to longevity that could be altered in the elderly population and related to an increased risk of developing certain lymphoid malignancies.

Methods
NBCs and MBCs were obtained by immunomagnetic separation from buffy coats of 10 healthy donors: 5 young (20-25 years) and 5 elderly (65-70 years). By microarray techniques we analyzed the expression of 45000 genes in all samples. Unsupervised hierarchical clusters of gene expression data were performed using the average linkage and the Euclidean distance. To identify differentially expressed genes between experimental groups we applied non-parametric Mann-Whitney test. The differences in expression with a p value < 0.05 were considered significant. All analyses were performed using the Multi-experiment Viewer 4.7.1 software.

Results
Firstly, we confirmed that the expression pattern of NBCs versus MBCs is significanly different in 3548 genes in young and 2145 genes in elderly individuals. Next, in order to evaluate the effect of age in NBCs and MBCs, we compared the gene expression pattern of young versus aged subjects in both cell populations. Interestingly, we did not find significant differences in the naïve population between young and aged individuals, whereas we found 1593 genes differentially expressed in young versus elderly subjects within MBCs; therefore, age affects the gene expression pattern of MBCs but not NBCs. Furthermore, we identified 467 genes which displayed a higher or lower expression in MBCs from aged as compared to younger individuals and NBCs, most of them involved in proliferation and cell cycle, apoptosis, cell survival and hematological diseases (Table 1).

Conclusions
Gene expression profiles of NBCs and MBCs are different. While there are no significant differences in NBCs of young versus elderly individuals, we detected significant differences in expression patterns of MBCs between both age groups, which indicates that MBCs could be more susceptible to age related alterations.

Disclosures:
No relevant conflicts of interest to declare.
JAK2V617F Mutation In Hematologic Malignancies In Childhood and Myeloproliferative Disorders In Adulthood
Authors: Yigin Akin , Antmen Bulent Ali , Guvenmez Hatice Korkmaz , Yilmaz Mustafa , Sasmaz Ilgen , Kilinc Yurdanur 
Published: October 2019

Abstract

Introduction
Mutations in JAK2 have been implicated in polycythemia vera (PV), essential thrombocythemia (ES), primary myelofibrosis (PMF) as wll as other myeloproliferative disorders.

Aim
In this study we aimed to investigate the frequency of JAK2V617F mutation on 216 patients with hematologic malignancies in childhood and 176 patients with myeloproliferative disorders in adulthood.

Materials and method
Patient group consist of 164 ALL and 52 AML patients in childhood and 79 PV, 51 ES, 22 chronic myeloid leukemia patients (CML) and 24 PMF patients in adulthood. These patients followed by Cukurova University, Departments of Pediatric and Adult Hematology, are included in this study. Blood samples were collected in these patients group and DNA was isolated using high pure template preparation kit (Roche) and stored -80oC. Gene mutations were studied using TMB (TıbMolBiol) LightMix Kit JAK2V617F genomic and analyzed by Light Cycler 2.0 Roche Diagnostic, GmBh, Germany in both groups.

Findings
JAK2V617F mutation was found 1 of 164 ALL patients (0,6%), 0 of 52 AML patients (0%) in childhood. Nevertheless, JAK2V617F mutation was also found 71 of 79 PV patients (89,8%), 22 of 51 ET patients (43,1%), 1 of 22 CML patients (4,5%) and 15 of 24 PMF patients (62,5%) in adulthood.

Result
As a result we found high frequency of JAK2V617F mutation in PV patients than the other myeloproliferative disorders. JAK2V617F mutation was significantly high in myeloproliferative disorders in adulthood comparing with childhood acute leukemias (p<0,01). Also, this mutation was significantly high in PV patients comparing with the other myeloproliferative disorders (P<0,05). We found statistically significance between genotype and allels distribution of JAK2V617F mutation (p<0.001) and determined that T-allels may be risk of disease in terms of allel distrubition (p<0.001). Thsi study is one of the widest series to investigation of JAK2V617F mutation frequency especially in childhood hematologic malignancies and we showed that this mutation is almost all 0% in this age group. On the other hand, we showed that JAK2V617F mutation with using realtime PCR test is very useful test for diagnosing PV and ET desase.

Disclosures:
No relevant conflicts of interest to declare.
Role Of Stromal Cell-Mediated Notch Signaling In Hematological Malignancies
Authors: Bassi Giulio , Kamga Paul Takam , Kamdje Armel Nwabo , Stradoni Roberta , Malpeli Giorgio , Amati Eliana , Nichele Ilaria , Carusone Roberta , Zanoncello Jasmina , Pizzolo Giovanni , Krampera Mauro 
Published: October 2019

Abstract
Stromal cells are essential components of the bone marrow (BM) microenvironment regulating and supporting the survival of different tumors, including B-cell acute and chronic lymphocytic leukemia (B-ALL and CLL), and acute myeloid leukemia (AML). In this study, we investigated the role of Notch signalling in human BM-mesenchymal stromal cell (hBM-MSC)-promoted ALL, CLL and AML survival and chemoresistance. The block of Notch signalling through γ-secretase inhibitor (GSI) XII reverted the protective effect mediated by co-culture with BM-MSC. The treatment with combinations of anti-Notch neutralizing Abs resulted in the decrease of B-ALL cell survival, either cultured alone or cocultured in presence of BM-MSC from normal donors and B-ALL patients. The inhibition of Notch-3 and -4 or Jagged-1/-2 and DLL-1 resulted in a dramatic increase of apoptotic B-ALL cells by 3 days, similar to what is obtained by blocking all Notch signaling with the GSI XII. The same Notch receptors are involved in CLL survival except for Notch-1 that, in CLL, mediates a synergistic effect with other Notch receptors in inducing the anti-apoptotic phenotype. Some preliminary data showed that Notch system is involved in survival and chemoresistance of acute myeloid leukemia blasts. Overall, our ﬁndings show that stromal cell-mediated Notch signaling has a role in promoting ALL, CLL and AML survival and resistance to chemotherapy. Therefore, the target of Notch pathway activation may represent a useful strategy to overcome drug resistance and improve the efficacy of conventional treatments.

Disclosures:
No relevant conflicts of interest to declare.
Outcome Of Patients With Acute Myeloid Leukemia/High Risk Myelodysplastic Syndrome Based On Bone Mineral Density: Data From a Single center 2000-2012
Authors: Anand Ankit , Al-sharif Hussam , Abdelfatah Mohamed , Haller Nairmeen , Al-Ameri Ali 
Published: October 2019

Abstract

Background
Acute myeloid leukemia results from clonal proliferation of myeloid precursors, resulting in accumulation of leukemic blasts in the bone marrow. The effect of leukemic blasts in the bone have not been well studied. It is hypothesized that leukemic blasts may affect the osteoblast and osteoclast balance.

Aim
The aim of this study is to describe the effect of osteoporosis/osteopenia on the survival of patients with AML/ high risk MDS

Methods
A retrospective study of all AML and high-risk MDS patients treated at Akron General Medical Center, Ohio during 2001-2012. Following IRB approval, patients' charts were reviewed for information on demographics, cytogenetics, and treatment. Patients were classified into low risk, intermediate risk and high risk MDS based on WHO criteria. Charts were reviewed for a diagnosis of osteoporosis/osteopenia. Overall survival rates were determined by Kaplan-Meier survival analysis. Patients were classified into osteopenia/osteoporosis based on bone density scans. An additional three patients were identified with osteopenia on the basis of lumbar X rays and one patient was diagnosed with osteoporosis on the basis of typical compression fracture for osteoporosis.

Results
During the study period, 187 patients were identified with AML/high risk MDS. A total of 31 patients were found to have osteopenia/osteoporosis based on radiological findings. Eight patients had osteoporosis and 24 patients had osteopenia. A survival analysis was performed on the study population based on bone mineral density. Group one (osteoporosis) included eight patients; group two (osteopenia) included 23 patients and group three (no radiological evidence of osteoporosis/osteopenia) included 156 patients. Results are depicted in the survival curve below.

Conclusions
Bone mineral density remains an unidentified factor in patient's survival, suffering from AML/ High risk MDS. It appears that patients with normal bone density may live longer than patients with osteopenia/osteoporosis, but this is not confirmed here statistically. Future studies with a larger patient sample may be helpful in further exploring this aspect and the role of bisphosphonates in patients with AML/high risk MDS.

Disclosures:
No relevant conflicts of interest to declare.