bluuebunny/crossref_metadata_2025_split huggingface dataset:
Intravenous Infusion of Liposomal Clodronate Blocks Clearance of Opsonized Erythrocytes in Dogs.
Authors: Mathes Mark A. , Dow Steven W. , Jordan Michael 
Published: October 2019

Abstract
Immune-Mediated Hemolytic Anemia (IMHA) is a spontaneously occurring disease of man and dogs in which immunoglobulin-coated erythrocytes are destroyed in the spleen. Immunosuppressive therapies and splenectomy have been previously used to combat premature destruction of erythrocytes. Liposomal clodronate has been previously shown to be a potent anti-macrophage agent when administered intravenously in a murine model. Splenic macrophages were selectively depleted through the induction of apoptosis following phagocytosis of liposomes containing clodronate. We hypothesized that such an approach might be useful in blocking clearance of erythrocytes in dogs with spontaneous IMHA.
Clodronate (dichloromethyl bisphosphonate) was formulated in liposomes (phosphatidyl choline, cholesterol and mannose), according to a previously published technique. The liposomal clodronate was evaluated in vitro for activity using canine and murine histiocytic cell lines, as assessed by cell killing and blocking of erythrophagia. Cell killing was monitored using microscopic evaluation of cell change, MTT and by propidium iodide staining. Blocking of erythophagia was monitored using flow cytometry. In vivo in normal dogs, increasing doses of liposomal clodronate were administered intravenously and evaluated to determine efficacy in blocking erythrocyte clearance and toxicity. Twenty-four hours post infusion, dye-labeled erythrocytes were opsonized using rabbit anti-canine erythrocyte antibody. Erythrocytes clearance was monitored using flow cytometry. The safe and effective dose of liposomal clodronate was determined to be 0.5 ml/kg administered IV via CRI over a 90 minute time period. After administration of intravenous liposomal clodronate in normal dogs, clearance of opsonized erythrocytes was nearly completely blocked at the 0.5 ml/kg dose. In vitro assays indicated that liposomal clodronate induced death of canine macrophage and dendritic cell lines. Five dogs with spontaneous IMHA were treated once with liposomal clodronate at the 0.5 ml/kg dose. Though erythrocyte clearance was not completely blocked, the drug was well-tolerated and all 5 dogs survived to leave the hospital. These studies suggest that liposomal clodronate may be an effective agent for temporary suppression of erythrocyte destruction in IMHA. Additional studies, including evaluation of higher doses of liposomal clodronate, are warranted in dogs with spontaneous IMHA.
Safety of Liver Biopsies in Children with Transfusional Iron Overload.
Authors: Mueller Brigitta Ursula , Williams Jennifer , Finegold Milton J. , Teruya Jun , Karpen Saul J. 
Published: October 2019

Abstract
Secondary iron overload, caused by chronic transfusions, is common among patients who have chronic hemolytic disorders, especially sickle cell disease and thalassemias. Accurate assessment of the hypertransfused patient’s body iron burden is important in deciding when and whether to start chelation therapy in order to prevent iron -related toxicities such as heart failure and endocrine abnormalities. The gold standard to measure iron content is through a liver biopsy. Though shown to be safe, many physicians are still reluctant to use this invasive approach in children to evaluate liver iron content.
Study Design: A retrospective chart review of chronically transfused patients at Texas Children’s Hospital (1998–2003) was performed to assess possible biopsy complications, including pain, hemorrhage, bilious peritonitis, puncturing of other organs, and bacterial infections, in patients with hemolytic disorders. The study was approved by the Institutional Review Board.
Results: There were 65 biopsies, 43 patients had one biopsy and 11 patients had two biopsies. 56 % were male, 44 % were female; 52% (28/54) were African American, 15 % (8/54) were Oriental/Asian, and 22 % (12/54) were Hispanic. 30 patients (56%) had sickle cell disease, 16 (30%) thalassemias, 5 (14%) other forms of anemia, 1 patient had leukemia, and 1 had lymphoma with the sickle cell trait. Eight of these patients (15%) are not on or have never been on chelation therapy, 11 (20%) used to be on chelation therapy, 35 (65%) are currently on chelation therapy. The average age when the first biopsy was performed was 11 years (range 2 to 18.9 years); and the average age at the time of the second biopsy was 12.7 years (range, 5.8 to 21.1 years). The reasons for their biopsies included possible iron overload due to hypertransfusions, pre-bone marrow transplant evaluation, and elevated liver enzymes. The average serum ferritin level before biopsy was 2681.8 ng/mL ranging from 348 to 35,332 ng/mL, and the average liver iron content (LIC) determined through biopsy was 12,278 ug/g dry wt. Ultrasound guidance was used for all 65 biopsies; 46 (71%) biopsies were done percutaneously, 8 (12%) laparoscopically, and 6 (10%) through an open procedure. There were no complications during the biopsy. There were no major and only 5 minor complications after the biopsy: 3 patients (5%) had transient abdominal pain at the biopsy site which subsided over a couple of days, and 2 (3%) had a fever with other (non-biopsy-related) complications requiring an increased hospital stay.
Conclusions: Liver biopsies in the pediatric population are safe and feasible and they should be considered for the accurate assessment of iron content in chronically transfused patients. Certain factors contributing to the success of the biopsies include having an experienced physician, using ultrasound guidance, and fully informing the patient or patient’s parents about the biopsy before the procedure.
Treatment of Severe Aplastic Anaemia (SAA) with Antilymphocyte Globulin (ALG), Cyclosporine (CyA) and Granulocyte Colony Stimulating Factor (G-CSF) 5 μG vs 10 μG/Kg : A Gitmo Prospective Randomized Study.
Authors: Locasciulli Anna , Bruno Barbara , Rambaldi Alessandro , Saracco Paola , Dufour Carlo , Finelli Carlo , Sica Simona , Varotto Stefania , Arcese William , Locatelli Franco , Soligo Davide , Bacigalupo Andrea 
Published: October 2019

Abstract
Background and objective. In a previous study we showed encouraging outcome in severe aplastic anemia (SAA) patients treated with anti-lymphocyte globulin (ALG), cyclosporin (CyA) and G-CSF 5mg/Kg/day (Blood2000; 95: 1931–4.). However failure to respond, delayed responses, partial responses, relapses and early deaths remain signifcant problems. The aim of the present study was to test whether an increased dose of G-CSF (10 mg/Kg/day) would reduce these complications.
Design and methods. This is a multicenter prospective trial in 77 SAA patients treated with horse ALG (15 mg/kg/day day1–5) and CyA (5 mg/kg/day day 1–180). Patients were randomized to receive G-CSF 5 mg/Kg/day (n=38, group A) or 10 mg/Kg/day (n=39, group B) from day +1 to day +30. All patients then received G-CSF 5 mg/Kg/day from day +31 to day +90. Primary end point was response at day +120. Secondary end points were early deaths , blood counts at day +120, and survival.
Results. At day +120 responses were classified as absent, partial, complete in 12, 22, 4 patients in group A and in 23, 7, 9 patients in group B (p=0.001). At last follow up these figures were respectively 9,12,17 vs 19,2,18 (p=0.004). Thirteen patients (5 in group A and 8 in group B) died before day 120 (p=0.3). Median peripheral blood counts an day 120 were comparable in the two groups: Hb 10.5 vs 9.5 gr% (p=0.6), Neutrophils 2.4 vs 1.9x10^9/l (p=0.4) and platelets 42 vs 36 x10^9/l (p=0.3). The actuarial survival at 4 years is 72% in group A vs 67% in group B (p=0.3).
An additional finding of this study is a strong age effect, with an actuarial 4 year survival of 81% in patients aged 0–20 , 80% in patients aged 21–40 and 34% in patients over 40 (p=0.0002). This correlated with the inability of older patients to increase their white blood cell (WBC) counts above 5x10^9/L, during G-CSF treatment. In a multivariate COX analysis patient age and highest WBC counts during G-CSF, were both significant predictors of survival.
Interpretation and conclusions. Increasing the dose of G-CSF does not appear to reduce early deaths, does not improve peripheral blood counts nor survival, and may reduce the response rate in patients with SAA receiving ALG and CyA. Older age and failure to improve WBC are negative predictors of survival.
Abnormal Erythropoiesis in Microgravity.
Authors: Xu Kun , Holubec Keith V. , Love John E. , Goodwin Thomas J. , Sytkowski Arthur J. 
Published: October 2019

Abstract
Humans and experimental animals subjected to microgravity, such as experienced during space flight, exhibit alterations in erythropoiesis, including changes in red blood cell morphology, survival and a reduction in red blood cell mass. Some of these alterations have been attributed to a disruption of normal in vivo erythropoietin physiology. However, human bone marrow cells grown on orbit showed a profound reduction in the number of erythroid cells, suggesting a cellular component. We now report the results of a study carried out on orbit on the International Space Station (ISS UF-1) in which an erythroid cell line was induced to differentiate. Rauscher murine erythroleukemia cells, a continuous cell line that can undergo erythropoietin (Epo)- or chemical-induced differentiation similar to normal erythropoiesis, were cultured for 6 days either in microgravity on board the ISS or on earth and then for 3 days in the absence or presence of 50 U Epo/ml or 0.7% dimethyl sulfoxide (DMSO). The cells were fixed, stored on orbit and returned to earth for study. Compared to ground-based controls, cells cultured in microgravity exhibited a greater degree of differentiation (hemoglobinization) (p<0.01). However, TER-119 antigen, a specific marker of the late stages of murine erythroid differentiation, was not detected on the surface of cells grown in microgravity. A significantly higher percentage (p<0.05) of cell clusters formed on orbit, whereas actin content appeared reduced. Furthermore, there was a more profound loss of actin stress fibers in microgravity following Epo or DMSO treatment. These results demonstrate abnormal erythropoiesis in vitro in microgravity and are consistent with the hypothesis that erythropoiesis is affected by gravitational forces at the cellular level.(Supported by NASA Grants NAG9-1368 and NAG2-1592 to AJS)
Pathogenesis and Therapy of a New Model of SOD2 Deficiency-Induced Sideroblastic Anemia.
Authors: Martin Florent M. , Welsh Megan L. , Friedman Jeff 
Published: October 2019

Abstract
Manganese superoxide dismutase (SOD2) is a major antioxidant protein which protects cells against the toxicity of superoxide anion, a by-product of mitochondrial respiration. While knockout of SOD2 results in late embryonic or early neonatal death, SOD2 deficient hematopoeitic stem cells (HSC) are capable of rescuing lethally irradiated normal hosts. The resulting hematopoeitic chimeric mice show a persistent hemolytic anemia similar to human sideroblastic anemia. Loss of SOD2 in erythroid progenitor cells results in mitochondrial proliferation and excess iron deposition within mitochondria of reticulocytes. Mature red blood cell (RBC) show abundant iron inclusions with evidence of a defect in iron incorporation into heme and altered survival. SOD2 deficient cells show both enhanced reactive oxygen species production and protein oxidative damage. We recently published a comparative RBC proteome analysis. Our results show that the expression of several proteins involved in folding/chaperone function, redox regulation, ATP synthesis and RBC metabolism is altered in SOD2 deficient cells. To define early events in the pathogenesis, we are currently performing a gene expression array strategy. The overall goal of this procedure is to characterize differential gene expression at the level of TER-119+ CD71+ erythroid precursor cells from the bone marrow of SOD2 deficient HSC mice versus wild-type and antioxidant-treated SOD2 deficient recipients. Preliminary analysis generated a list of 718 genes as significantly discriminating between wild-type and SOD2 deficient RBC progenitors. Taken together, our results and subsequent validation of candidate genes in in vivo experiments will identify genes involved in response of erythroid progenitors to oxidative stress as well as determinants of RBC lifespan in other hemolytic disorders.
Is EPO Receptor Expressed in Human Tumor Cells?.
Authors: Sinclair Angus M. , Busse Leigh , Rogers Norma , Sarosi Ildiko , Van Gwyneth , Elliott Steve 
Published: October 2019

Abstract
EPO stimulates formation of red blood cells by binding to and activating the EPO receptor (EPOR) expressed on the surface of erythroid progenitor cells. Recently EPOR was reportedly expressed in other, non-hematopoietic tissues including brain, kidney, heart and endothelium. EPOR expression has also been reported in tumor tissues and tumor cell lines and many investigators have suggested that EPO may promote tumor growth. This has raised concerns about the use of rHuEPO to treat anemia associated with chemotherapy in cancer patients. To evaluate this, we have examined EPOR expression and binding in tissues and cell lines by quantitative PCR, Western blot analysis and EPO binding assays. Quantitative RT-PCR analysis revealed that the EPOR gene is transcribed in most tissues and cell lines. However non-hematopoietic EPOR transcript levels were lower than that found in hematopoietic cells. No evidence of elevated EPOR gene transcription in tumor vs. non-tumor tissues or in tumor cell lines analyzed was observed. Using Western analysis, we found that commercial anti-EPOR antibodies commonly used to examine EPOR protein expression in published tumor studies, cross-reacted with multiple proteins and/or reacted with proteins of sizes different from that predicted for EPOR. This suggests that in many publications where these antibodies were used, claims of expression of EPOR protein in tumors are of questionable significance. A rabbit polyclonal antibody was developed that specifically reacted with a protein band on Western blots of the size predicted for EPOR. The antibody reacted specifically with a band in COS cells transfected with a human EPOR expressing construct but not in transfection controls. This antibody also reacted with the same sized protein in non-hematopoietic cells. We concluded that EPOR mRNA and EPOR protein were present in most normal cell types examined. Thus the detection of EPOR in tumors and tumor cell lines likely represents the continued expression reflecting the tissue of tumor origin. Since expression does not equate to the presence of surface receptor, binding assays with 125I EPO to non-hematopoietic tumor cell lines were performed. Tumor cell lines showed weak to no detectable EPO binding. These results suggest that although EPOR protein is synthesized in these cells, little or no receptor is expressed on their cell surface. Furthermore, if limited amounts of EPO bound EPOR in tumor lines any biological effect may be different or muted compared to classical responses in hematopoietic cells.
Placental Growth Factor Concentrations in Bone Marrow of Sickle Cell Patients.
Authors: Means Robert T. , Fleury Thomas W. , Dallalio Gail A. , Brunson Chris Y. 
Published: October 2019

Abstract
Although sickle cell anemia (SCA) is the result of an abnormality in the beta globin chain of hemoglobin, its clinical manifestations cannot be explained solely on that basis. It has become clear over the last several years that inflammation and inflammatory mechanisms contribute to the clinical syndromes associated with SCA, and that sickle cell disease, even in steady-state, is an inflammatory process. Placental growth factor (PlGF) is a member of the vascular endothelial growth factor family and is associated with inflammation and with pathologic angiogenesis. Perelman and colleagues have reported that PlGF is released from marrow erythroid cells and that its circulating concentration is 50% higher in patients with severe SCA than it is in healthy controls, and also that PlGF induces the expression of inflammatory cytokines like interleukin-1 (IL-1) and tumor necrosis factor (TNF) (Blood2003;102:1506–14; Blood2003;102:1514–24). As part of our studies of the regulation of erythropoiesis by inflammation in sickle disease, we collected bone marrow aspirates from homozygous SCA patients not currently in crisis (n=5) and from healthy volunteers (n=10), and evaluated PlGF concentrations in the marrow aspirate plasma. The SCA patients studied were all adults, and were not on chronic transfusion, erythropoietin, or hydroxyurea. PlGF concentrations were significantly higher in the marrow aspirate plasma from SCA patients compared to healthy controls (range 38.7 – 90.2 pg/mL vs. 8.7–21.9 pg/mL; p = 0.002). In contrast, plasma PlGF concentrations were overlapping (range 6.2– 11.3 pg/mL for SCA patients vs. 2.2–27.0 pg/mL; p = 0.59). When PlGF concentration was normalized to marrow CFU-E concentration, mean PlGF protein per CFU-E was substantially higher for SCA patients (0.86 pg/105 CFU-E vs. 0.13 pg/105 CFU-E ), although the difference did not achieve statistical significance (p = 0.07). Since cytokines induced by PlGF, such as TNF and IL-1, are known to inhibit CFU-E colony formation, the effects of PlGF on erythroid colony formation in vitro by marrow from SCA patients and healthy volunteers were studied. At concentrations 10 – 1000 pg/mL, no inhibition of CFU-E colony formation was observed, and there was no difference in the response of progenitors from SCA patients and from normal volunteers. In summary, PlGF concentrations in bone marrow aspirate plasma are significantly greater than those observed in healthy controls, and the degree of this difference is more substantial than has been reported for circulating PlGF concentrations. Erythroid progenitors from SCA patients tend to produce PlGF at a higher level then do progenitors from healthy controls. PlGF does not inhibit erythroid colony formation in vitro, and would not be expected to contribute directly to impairment of erythropoiesis in SCA patients.
L-Carnitine and Apoptosis in Thalassemia Major Patients.
Authors: Taher Ali , Seoud H. , Ibrahim A. , Gabre H. , Ibrahim I. Youssry , ElGawada G. Abd , Aoun E. , El-Beshlawy A. 
Published: October 2019

Abstract
Background and Objectives:
Thalassemia major patients suffer from severe anemia due to intramedullary hemolysis of erythroid progenitors. Depending on the severity of their disease, thalassemic patients exhibit different patterns of apoptosis. In cases of β-thalassemia major, apoptosis appears to be greatly enhanced in the early-stage erythroid precursors in the bone marrow and it is considered to be the machinery via which ineffective erythropoiesis develops. L-Carnitine is a butyrate derivative found to strongly reduce apoptosis in different diseases. We investigated the effect of oral L-carnitine therapy on apoptosis in thalassemia major patients.
Methods:
eighteen Thalassemia major patients were included. Detection of apoptosis was done by Photometric enzyme immunoassay (Elisa) and agarose gel electrophoresis before and after 6 months oral therapy with L-Carnitine (50mg/kg/day).
Results:
The mean age was 12.2±6.6 years. All patients tolerated L-carnitine with no side effects. A significant decrease of apoptosis rates in the erythroid precursors in the bone marrow of studied cases was noted after therapy by. The enrichment factor measured by the photometric enzyme immunoassay dropped from 3.65±1.338 before therapy to 1.60±0.65 after therapy (p=0.005). A positive ladder pattern reflecting apoptosis on agarose gel electrophoresis was detected in 88.9% of cases prior to treatment (figure 1) versus 11.1% after therapy (figure 2) (p=0.006). Patients also had significant decrease in the frequency of transfusions and increase in the pre-transfusion hemoglobin levels after therapy. These results were found to significantly correlate with the reduction in apoptosis rates (p=0.006, r = 0.543 and p=0.027 &r = −0.7762 respectively).
Conclusion:
Apoptosis plays a major role in the underlying pathophysiology of anemia in thalassemic patients. L-Carnitine seems to be a good modulator of apoptotic processes in such patients leading to decreased rates of programmed erythroblast death and general improvement of the disease condition. Further large-scale studies are needed to evaluate the overall role of L-Carnitine in the management of these patients.
Figure Figure
Hydroxyurea to Increase Hemoglobin Level in Thalassemia Intermedia.
Authors: Baronciani Donatella , Vaquer Stefania , Carta Paola , Lai Eliana 
Published: October 2019

Abstract
Hydroxyurea (HU) has been shown to increase fetal hemoglobin (HbF) levels in patients with Sickle Cell Disease and Thalassemia Intermedia. Because there are no defined protocols for the use of this drug, we studied the effect of HU in a group of thalassemia intermedia (genetically defined) patients with at least one of the following features:
Low hemoglobin level ( hemoglobin < 7 gr/dl) Bulky extramedullary erythroid masses
We started the enrollment in February 2003 and 8 patients with a minimum follow up of 8 months have been evaluated ( range 8–16 months ) ( mean 13,6 months; median 14 months ). Mean age of the patients was 39.8 years ( range 28–49 years); all were not transfused at the moment of the study entry. HU was administered orally at a mean daily dose of 8.8 mg/kg (range 4–12 mg/kg). The drug was well tolerated; only one patient discontinued the treament for 2 months for pleural effusion. Table summarizes the pre and post treatment characteristics of this group of patients. Data reported are median (range), the differences between pre and post treatment values were analyzed with the the Wilcoxon test for non parametric observations. In this group of patients a mean increase of hemoglobin level of 1.2 g/dl was registered. Extramedullary erythroid expansion, studied by computed tomography scan did not show any change in this follow-up period.
These results confirm that HU may improve Hb level in thalassemia intermedia patients even if other aspects must be studied and longer follow-up is necessary.
Treatment results
Pre treatment Post treatment Two tailed P value Values are expressed as median (range). P value = Wilcoxon test Hemoglobin level (g/dl) 6.6 (4.9–7.9) 7.8 (7.1–9.8) 0.05 Erythropoietin value (mU/ml) 679 (98–2855) 191 (71–1690) 0.2 Erythroblasts % leucocytes 191 (80–567) 153 (2–426) 0.2 Mean Corpuscolar volume (micron3) 77 (66–87) 82 (69–91) 0.08 Mean Hemoglobin Concentration (pg/rbc) 22 (21–25) 25 (22–27) 0.02 Hemoglobin F (%) 94.7 (52.9–97.7) 96.8 (58.7–98.4) 0.06