bluuebunny/crossref_metadata_2025_split huggingface dataset:
Polyphosphate Enhances Fibrin Clot Structure.
Authors: Smith Stephanie A. , Morrissey James H. 
Published: October 2019

Abstract
Introduction: Inorganic polyphosphate (polyP) is a negatively charged polymer of phosphate units linked by high energy phosphoanhydride bonds. Dense granules of human platelets contain polyP which is released in response to thrombin stimulation. We recently reported that polyphosphate is a potent hemostatic regulator, accelerating blood clotting by activating the contact pathway and promoting the activation of factor V. Our previous studies found that polyP did not affect the time to clot formation when plasma was clotted with thrombin, however, suggesting that polyP exerts its procoagulant actions upstream of thrombin. We now report that polyP enhances fibrin clot structure.
Methods: Purified fibrinogen and polyP were preincubated for 15 min in multiwell plates in buffer containing CaCl2, after which clotting was initiated by adding 0.1 to 8 nM thrombin and fibrin clot formation was evaluated by quantifying the change in turbidity (A405). Mass-length ratios were calculated from scans of A400 to A800. The effect of polyP on fibrinolysis was examined by adding 8 nM plasmin to the reaction mixtures immediately prior to thrombin. Scanning electron microscopy (SEM) was employed to visualize clot structure, and time courses of covalent fibrin cross-linking were assessed by SDS-PAGE.
Results: PolyP had no effect on time to clot formation, but clots formed in the presence of polyP had markedly (up to threefold) higher turbidity than clots formed in the absence of polyP (see figure), irrespective of thrombin concentration. The increased turbidity in the presence of polyP was calcium-dependent and was enhanced when fibrinogen, CaCl2, and polyP were preincubated for up to 15 min prior to initiation of clotting with thrombin. PolyP increased the mass-length ratio of fibrin, and SEM confirmed that fibers formed with polyP were thicker than those formed without polyP. The ability of polyP to enhance fibrin clot turbidity was independent of factor XIIIa activity, and polyP did not alter the rate or extent of covalent fibrin cross-linking by factor XIIIa. When plasmin was included in clotting reactions containing polyP, mean times to 50% clot lysis were 28.5 ± 0.8 min for clots without polyP but 120.4 ± 5.6 min for clots with polyP.
Conclusions: PolyP alters polymerization of fibrin, resulting in fibers of higher mass-length ratio that are lysed more slowly. This effect is calcium-dependent and is enhanced by preincubation of fibrinogen with calcium and polyP. Release of polyP from activated platelets or infectious microorganisms may therefore enhance fibrin clot structure.
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Evidence for a Metabolic Stem Cell Niche.
Authors: Cross Michael , Alt Rudiger , Schnapke-Hille Lydia , Riemer Thomas , Niederwieser Dietger 
Published: October 2019

Abstract
The hematopoietic stem cell niche presents a localised environment supporting the balanced maintenance, self-renewal and occasional expansion of the stem cell pool. These options are widely assumed to be regulated exclusively by signalling from specific combinations of stroma-bound or soluble ligands. However, a consideration of the rare conditions under which absolute numbers of stem cells increase in vivo as well as the selective pressures acting on regenerative systems during evolution has led us to propose a metabolic component to the stem cell niche which serves to limit cumulative damage, to avoid the selection of potentially oncogenic mutations and to tie symmetric division to slow proliferation. This would mean that traditional cell culture media based on “systemic” substrates such as glucose and glutamine may actively prevent the symmetric amplification of high quality stem cells, offering a possible explanation for the limited success in this area to date. To investigate this possibility, we have examined the effects of range of carbon and energy sources on the proliferation and maintenance of stem and progenitor cells. Our strategy is to screen a wide variety of culture conditions using murine FDCPmix cells, which are non-tumorigenic but have an innate tendency to amplify symmetrically in the presence of IL-3, and then to test key observations in human UCB CD133+ cells provided with SCF, TPO and FLT-3L. In both cell systems, we do indeed find an unusually low requirement for the systemic substrates glucose and glutamine normally included as major energy and carbon sources in cell culture media. Reducing glucose reduces the yield of committed cells from CD133+ cultures without affecting the accumulation of CD133+CD34+cKit+ progenitors. When provided with alternative substrates more likely to reflect a “niche” type environment, FDCPmix cells can be maintained for long periods in media containing only the trace levels of glucose or glutamine derived from dialysed serum, and show improved self-renewal under these conditions. We have also found that raising osmolarity reduces glucose dependence and simultaneously favours the maintenance both of self-renewing CFU (FDCPmix culture) and of CAFCweek13 (CD133+ culture). In parallel, the use of NMR and mass spectrometry techniques to profile intracellular metabolites in self-renewing and differentiating FDCPmix cells reveals a shift in the metabolite balance indicating reduced glycolysis in the early cells. Taken together, these results suggest that hematopoietic stem cells do indeed have remarkable metabolic characteristics consistent with adaptation to a metabolically limiting niche environment. It may therefore be necessary to identify niche substrates and to combine these with the relevant signalling environment in vitro in order to effectively increase stem cell numbers for research, stem cell transplantation and tissue engineering applications.
Supportive Role of TNF-Family Death Receptors in Hematopoietic Cell Engraftment.
Authors: Pearl-Yafe Michal , Askenasy Nadir , Stein Jerry , Yaniv Isaac 
Published: October 2019

Abstract
The activity of death receptors of the tumor necrosis factor (TNF) superfamily as negative regulators of distal differentiation in the immuno-hematopoietic system is well characterized. The role of these receptors, classically associated with transduction of apoptotic signals, is unknown. A series of studies have attributed these receptors also negative regulatory functions in ex vivo expanded hematopoietic progenitors, showing variable supporting and suppressive interactions with growth factors. We observed robust upregulation of Fas, TNF-R1, TNF-R2 and TRAIL-R2 in bone marrow homed donor cells, reaching levels of 60–75% expression after 6 days. All putative SCA-1+c-kit+lin− progenitors ubiquitously expressed all four receptors. These receptor-positive cells were largely resistant to apoptotic signals delivered by the corresponding soluble and membranous ligands. Furthermore, there was no evidence of receptor cross-talk in sensitization of the cells to apoptotic death. The engraftment potential of cells was markedly diminished by ex vivo incubation, without any relationship to the viability of the putative progenitors. Taken together these data suggest a supportive role of the “death receptors” in hematopoietic cell engraftment and dissociate the deficient engraftment of cultured cells from apoptosis of progenitors.
Derivation, Characterization, and In Vitro Differentiation of Canine Embryonic Stem Cells.
Authors: Ramakrishnan Aravind , Hayes Brian , Fagerlie Sara R. , Baran Szczepan , Harkey Michael , Graf Lynn , Bar Merav , Bendoraite Ausra , Tewari Muneesh , Torok-Storb Beverly 
Published: October 2019

Abstract
Embryonic stem (ES) cells have created considerable excitement in the last few years due to their unlimited potential to produce cells for tissue repair and replacement. However, a large animal pre-clinical model is necessary to establish the safety and efficacy of ES cell-derived tissue replacement therapy. The canine model has long been used in medical research, has been well established to study adult stem cell transplantation and has been highly predictive of clinical outcomes in humans, more so than rodent models. Given the documented record for extrapolating from dog to man, we hypothesize that the dog would serve as an ideal pre-clinical in vivo model for studying the clinical applications of ESC derived tissue. Eleven putative ES cell lines were initiated from canine blastocysts harvested from natural matings. One line described here, FHDO-7, has been maintained through 34 passages and has many characteristics of ES cells from other species. FHDO-7 cells are alkaline phosphatase positive and express both message and protein for the Oct4 transcription factor. They also express message for Nanog and do not express message for Cdx2 which is associated with trophectoderm. Furthermore, they express a cluster of pluripotency-associated microRNAs (miR-302b, miR-302c and miR-367) that have been found to be characteristic of human and mouse ES cells. The FHDO-7 cells grow on feeder layers of modified mouse embryonic fibroblasts (MEF) as flat colonies that resemble ES cells from mink, a close phylogenetic relative of dog. When cultured in nonadherent plates without feeders the cells form embryoid bodies (EB). Under various culture conditions the EBs give rise to ectoderm-derived neuronal cells expressing β3-tubulin, mesoderm-derived osteocytes producing bone, and endoderm-derived cells expressing alpha feto protein or Clara cell specific protein. These results indicate that FHDO-7 is a pluripotent embryonic stem cell line.
Autologous Bone Marrow Transplant for Human Diabetes Mellitus.
Authors: Novoa Jose E. , Medina Maria A. , Gordillo Felipe , Ravera Jorge , Olivet Carlos 
Published: October 2019

Abstract
Background: stem cells have tha capacity of both self-renewal and multilineage differentiation. They have the ability to proliferate and differentiate into any type of cell. Bernat Soria and coworkers showed how to guide stem cells in animals to differentiate in specific cell lineages as neurons,cardiomyocytes an insulin-secreting cells (Diabetologia2001;44:407–415).Therapeutic angiogenesis has recently been developed as a new method of treatment for several ischaemic diseases performing stem cell transplantation. Ernesto Novoa et al. reported in march 2007 the Conzi’s Effect inhuman diabetes mellitus (Archivos de Medicina Interna2007;XXIX suppl1:S24.25). After therapeutic angiogenesis the patients discontinued antidiabetic treatment as a consequence of normalization in glucose metabolism.
Aims: to study the bahaviour in glucose metabolism in a group of patients receiving autologous bone marrow transplantation for arterial ischaemic limb disease and prior mobilization with G-CSF. To evaluate the safety of the procedure.
Methods: from july 2004 to july 2007, 71 patients developing severe arterial limb ischaemia (candidates to amputation) were included in this study. 40 men and 31 women. Median age 66 years old (44–86). Mobilized with filgrastim (Neupogen) 5 ug/kg weight daily (5 days). Bone marrow harvest at 6th day. Local anaesthesia was performed in all the patients. Unmanipulated cells, concentrated by gradient of density, were injected in the affected limb in 2 ml aliquots into the gastrocnemius muscle. Mean volume 40 ml. Follop up of each patient was done regularly evaluating: amputation, rest pain, frequency of analgesics intake, healing of the ulcers, peak walking time, insulin or oral antidiabetic drugs required. Doppler and angiographic test were also performed. The mean number of injected mononuclear cells was 1,9 x109/kg. All the patients received low molecular weight heparin (nadroparin, Fraxiparine)3800–5600 IU anti-Xa subcutaneously, aspirin 81 mg and pentoxifiline 400 mg daily, as medical treatment after the procedure for at least 60 to 90 days. A control population of 67 patients affected by severe arterial limb ischaemia was considered. They didn’t receive angiogenic treatment.
Results: there was no treatment related mortality. In the control group 65 patients were amputated. In the “ABMT Group” 3 patients received amputation of the lower limb. The statistical difference between the two groups was highly significant in favor of the angiogenic group. They were evaluated by the chi square test and log rank test with a p value < 0,05. In the ABMT Group, 33% of type 1 diabetes and 74% of type 2 diabetes, discontinued their antidiabetic treatment for more than 3 months after ABMT.
Conclusions: autologous bone marrow transplantatioan can be performed safely and appears to be a benefical therapy for selected patients with severe peripheral arterial occlusive disease and diabetes mellitus. novoa.je@gmail.com
Characterization of BioSafe SEPAX Manufactured Stem Cell Intended for Cardiac Cell Therapy.
Authors: O’Connor Sean L. , Sepulveda Carmen A. , Kaur Indreshpal , Sumari Robert D. , McMannis John D. 
Published: October 2019

Abstract
Numerous cardiology groups are actively pursuing the use of hematopoietic stem cells for myocardial delivery. Most published studies have used a manual ficoll density separation procedure to enrich the Mononuclear Cell (MNC) population thus enriching the stem cell content in the final therapeutic cell product. Manual ficoll procedures have certain intrinsic limitations including technologist variability, training time, and risk of contamination due to its open nature. Most open and manual cell processing procedures have been supplanted by recent technology developments in automated and closed devices. The BioSafe SEPAX is a closed system-automated ficoll density separation device that currently has FDA 510k clearance for the processing of umbilical cord blood products. In support of multi-center clinical studies being conducted by the NHLBI Cardiovascular Cell Therapy Research Network (CCTRN), the MD Anderson Cell Therapy Laboratory has evaluated bone marrow derived, MNC enriched, stem cells for their cardiovascular therapeutic potential. Traditional transplantation immunophenotyping was performed on these cells to evaluate hematopoietic stem cells (CD34+ and CD133+), T-Cell subsets, B-Cells, and NK-Cells. Colony Forming Units (CFU) as wells as surrogates used in cardiac regeneration studies including endothelial invasion assays and CXCR4 immunophenotyping (a chemokine receptor involved in cell migration) were also performed. Our data suggests the cells produced using the automated ficoll BioSafe SEPAX device are equivalent to cells produced using a manual ficoll procedure with respect to TNC, MNC, viability, and CD34+ recoveries. Cell subsets and function including invasion and CXCR4 analysis were also similar between the two procedures. The automated method produced a product with statistically higher CFU potential than the manual method suggesting the SEPAX isolates a more potent subset of the mononuclear fraction. These studies support the use of the SEPAX device as for enriching BM-MNCs for the CCTRN. Additional benefits might include the use of a closed system, ease of training, speed of cell manufacturing, and product consistency.
GlycoPEGylated Filgrastim XM22 Fixed Dose Demonstrates Similar Activity in Comparison to Pegfilgrastim in Healthy Volunteers after Single Administration.
Authors: Lubenau Heinz , Pokorny Rolf , Lopez-Lazaro Luis , Bias Peter , Wallin Bruce , Hinderer Walter , Maly Anne-Katrin , Seiberling Michael 
Published: October 2019

Abstract
XM22 is a glycoPEGylated recombinant human G-CSF that interacts with the G-CSF receptor by binding and activation. In pilot studies in non-neutropenic animals, XM22 demonstrated a similar PK/PD profile compared to pegfilgrastim. By its longer duration of action compared to filgrastim a once per cycle dosing as fixed dose in chemotherapy treated cancer patients for the reduction of duration of severe neutropenia and incidence of febrile neutropenia is intended.
METHODS: A single center, randomized, single-blind study of XM22, administered as single fixed dose given by the s.c. route, was conducted to assess safety, pharmacokinetic profile and pharmacodynamic properties based on absolute neutrophil count (ANC) and CD34+ cell count. Male or female healthy volunteers were assigned to either XM22 6 mg or pegfilgrastim 6 mg fixed dose treatment.
RESULTS: 36 healthy volunteers were enrolled in two treatment groups and completed 21 day follow-up. Injections were generally well-tolerated with no discontinuations due to adverse events or serious adverse events. Pharmacodynamic data were compared between the fixed doses of 6 mg XM22 and pegfilgrastim and in body weight strata within the treatment groups. Pharmacokinetic and antibody data will be presented.
CONCLUSIONS: Single doses of 6 mg fixed dose of XM22 administered to healthy volunteers were generally well-tolerated with regard to the expected side effect profile and demonstrated increases in absolute neutrophil count and CD34+ cell count. These results supplement data from another study in healthy volunteers in which XM22 was administered in ascending single doses adjusted according to body-weight.
NF-kappa B Plays a Key Role in the Hematopoietic Niche Function of Osteoblast after Ionizing Radiation.
Authors: Xiao Mang , Inal Cynthia E. , Whitnall Mark H. 
Published: October 2019

Abstract
Recent studies demonstrated that osteoblasts in the endosteal region of bone are a key cellular component of the hematopoietic niche that directly regulates hematopoietic stem cell (HSC) survival, proliferation and differentiation. In experiments with the human fetal osteoblast cell line hFOB1.19, we found survival of gamma-irradiated primary human hematopoietic CD34+ cells was significantly enhanced by co-culture with hFOB cells or by conditioned medium (CM) from hFOB cells, as shown by increased colony efficiency and colony size. In clonogenic assays of CD34+ cells starting with 5 × 103 cells/dish, co-culture with hFOB cells increased colony number from 428±53 (control) to 988±65 (co-culture) after 2 Gy (p<0.01), 301±44 to 739±56 after 4 Gy (p<0.01), and 45±15 to 72±10 after 6 Gy (p<0.05). Surprisingly, although radiation induces osteoblast cell damage, CM from 6 Gy-irradiated hFOB cells had a greater effect than CM from unirradiated hFOB cells in supporting CD34+ clonogenicity and resulted in colony number increase from 430±40 (CM from unirradiated hFOB cells) to 773±48 (CM from 6 Gy-irradiated hFOB cells) after 2 Gy IR (p<0.01). Using a cytokine array for 120 cytokines and chemokines, we detected 11 factors released from hFOB cells, and 4 of them (IL-6, G-CSF, GRO, and IL-8) were enhanced by ionizing radiation (IR). IR (2–8 Gy) caused hFOB cell cycle arrest in G2 phase after 24 h, and irreversible cell cycle block and apoptotic cell death were observed 72 h after IR in a radiation dose-dependent manner. IR induced NF-kappaB (NFkB) p65 phosphorylation and NFkB activity in hFOB cells. Inhibition of NFkB expression with siRNA blocked the ability of hFOB CM to support clonogenic survival of CD34+ cells, and abrogated the effect of IR on release of IL-6, G-CSF, GRO, and IL-8. Radiation upregulated p53 (ser15) phosphorylation within 4 h, and p21 expression after 24 h in hFOB cells. However, NFkB gene silencing accelerated p21 expression and induced a massive apoptosis of hFOB cells. We propose that IR induces NFkB and p53 activation simultaneously in hFOB cells. The fate of osteoblasts after irradiation depends on the amount and severity of injury, and on repair processes regulated by NFkB and p53. Silencing the NFkB gene disrupted this balance and resulted in hFOB cell apoptosis.
Ciprofloxacin-Induced G2 Arrest and Apoptosis in TK6 Lymphoblastoid Cells Is Not Dependent on Double-Strand Break Formation.
Authors: Halicka Dorota , Smart Daniel J. , Seiter Karen , Traganos Frank , Darzynkiewicz Zbigniew , Williams Gary M. 
Published: October 2019

Abstract
Drugs indicated for conditions other than tumors have shown promise in the potential treatment of leukemia and lymphoma. The fluoroquinolone antibiotic ciprofloxacin (CPFX) is known to modulate cycle cell progression and apoptosis in cancer cells and thought to induce DNA double-strand breaks (DSBs) via topoisomerase II (topo II) inhibition. DSBs trigger the phosphorylation of histone H2AX on serine 139 (γH2AX) by PI-3-like kinases including ATM; γH2AX can serve as a marker of DNA damage when measured in situ using immunocytochemistry and flow cytometry. The aim of the present study was to investigate the relationship between CPFX-mediated DNA damage and the induction of apoptosis in human lymphoblastoid cells and phytohaemagglutinin (PHA)-stimulated normal lymphocytes using γH2AX and multiparameter flow cytometry. Treatment of TK6 cells (wild-type p53) with 100 mg/ml CPFX for 2–10 hours produced no increase in γH2AX compared to controls; levels were in fact reduced in S phase cells at 6 and 10 hours. Phosphorylated ATM and G2 arrest was also observed in TK6 cells exposed to CPFX for 4 hours. γH2AX was dramatically increased after 24 hours treatment, indicating the onset of apoptosis (confirmed by presence of activated caspase-3). CPFX had a similar lack of effect on γH2AX at early time points in WTK-1 and NH32 lymphoblastoid cells (devoid of functional p53) and proliferating lymphocytes, however, induction of apoptosis was not as pronounced in these cells compared to TK6. The data suggest that, at early time points, CPFX may act a radical scavenger to reduce the level of endogenous oxidant-induced DSBs. However, phosphorylation of ATM (but lack of γH2AX induction) does indicate the presence of a non-DSB DNA modification. At later time points, induction of apoptosis appears to be facilitated by a functional p53 signaling pathway. CPFX may therefore have a potential use as a novel chemotherapeutic agent in the treatment of lymphoblast-derived cancer.