The FLK2/FLT3 ligand synergizes with interleukin-7 in promoting stromal- cell-independent expansion and differentiation of human fetal pro-B cells in vitro

The effects of a novel cytokine FLK2/FLT3 ligand (FL) on human fetal bone marrow-derived CD34+CD19+ pro-B cells were analyzed in a stromal- cell-independent, serum-deprived culture system. FL, like interleukin-3 (IL-3), synergized with IL-7 in promoting pro-B cell growth, and differentiation of these cells into CD34-CD19+clgM+slgM- pre-B cells, whereas a small proportion of these cells even differentiate into more mature slgM+ B cells. In contrast, KIT ligand (KL) and granulocyte- macrophage colony-stimulating factor (GM-CSF) were ineffective in promoting IL-7-dependent pro-B cell growth and differentiation. Maximal levels of pro-B cell expansion, generally resulting in 15- to 30-fold increases in cellularity, were obtained in cultures supplemented with optimal doses of FL + IL-7 + IL-3. The addition of mouse bone marrow stromal cells further enhanced the proliferation and differentiation of pro-B cells obtained in the presence of these three cytokines. Under these conditions, cultures could be maintained for more than 4 weeks, and in general 40- to 50-fold increases in cell numbers were observed by 3 weeks of culture. The percentages of clgM+ and slgM+ B cells increased 1.5- to 3-fold and 2-fold, respectively, suggesting that stromal cells may provide additional costimulatory signals for human B- cell growth and differentiation that are different from IL-7, IL-3, and FL. Collectively, our results indicate that FL, in contrast to KL, strongly promotes long-term expansion and differentiation of human pro- B cells in the presence of IL-7 or in combination of IL-7 and IL-3, which is a novel property of this hematopoietic growth factor.     

Unrelated donor marrow transplantation in children

Eighty-eight children 0.5 to 17 years of age (median, 9 years of age) received an unrelated donor marrow transplant for treatment of chronic myeloid leukemia (CML; n = 16), acute lymphoblastic leukemia (ALL) in first or second remission (n = 15) or more advanced stage (n = 28), acute myeloid leukemia (AML; n = 13), or other hematologic diseases (n = 16) between June 1985 and April 1993. All patients were conditioned with cyclophosphamide and total body irradiation and received a combination of methotrexate and cyclosporine as graft-versus-host disease (GVHD) prophylaxis. Fourty-six patients received transplants from HLA-identical donors and 42 patients received transplants from donors who were minor-mismatched at one HLA-A or B or D/DRB1 locus. The Kaplan-Meier estimates of disease-free survival and relapse were 75% and 0% for patients with CML, 47% and 20% for ALL in first or second remission, 10% and 60% for ALL in relapse or third remission, 46% and 46% for AML in first remission (n = 1) or more advanced disease (n = 12), and 29% and 69% for other diseases. HLA disparity was not significantly associated with lower disease-free survival, but the results suggest more relapses in HLA-matched recipients and there was significantly more transplant-related mortality in mismatched recipients (51% v 24%, P = .04). Most deaths were due to infections associated with acuteor chronic GVHD and occurred within the first 2 years after transplantation. Granulocyte engraftment occurred in all evaluable patients. Sixty-three percent of HLA-matched and 57% of HLA- mismatched recipients were discharged home disease-free at a median of 98 and 103 days, respectively, after transplantation (P = not significant [NS]). The incidence of grades II-IV acute GVHD was 83% in HLA-matched and 98% in HLA-mismatched recipients (P = .009). The incidence of chronic GVHD was 60% in HLA-matched and 69% in HLA- mismatched recipients (P = NS). One or multiple late adverse events such as cataracts, osteonecrosis of the hip or knee, restrictive or obstructive pulmonary disease, and hypothyroidism have occurred in 11 of 33 (33%) surviving patients. Immunosuppression was discontinued in 58% of surviving patients, including all 12 patients surviving more than 3.2 years, all of whom have a Lansky or Karnofsky score of 100%.(ABSTRACT TRUNCATED AT 400 WORDS)     

A competitive enzyme-linked immunosorbent assay specific for murine hepcidin-1: correlation with hepatic mRNA expression in established and novel models of dysregulated iron homeostasis

Mice have been essential for distinguishing the role of hepcidin in iron homeostasis. Currently, investigators monitor levels of murine hepatic hepcidin-1 mRNA as a surrogate marker for the bioactive hepcidin protein itself. Here, we describe and validate a competitive, enzyme-linked immunosorbent assay that quantifies hepcidin-1 in mouse serum and urine. The assay exhibits a biologically relevant lower limit of detection, high precision, and excellent linearity and recovery. We also demonstrate correlation between serum and urine hepcidin-1 values and validate the competitive enzyme-linked immunosorbent assay by analyzing plasma hepcidin response of mice to physiological challenges, including iron deficiency, iron overload, acute blood loss, and inflammation. Furthermore, we analyze multiple murine genetic models of iron dysregulation, including β-thalassemia intermedia (Hbb^th3/+), hereditary hemochromatosis (Hfe^−/−, Hjv^−/−, and Tfr2^Y245X/Y245X), hypotransferrinemia (Trf^hpx/hpx), heterozygous transferrin receptor 1 deficiency (Tfrc^+/−) and iron refractory iron deficiency anemia (Tmprss6^−/− and Tmprss6^hem8/hem8). Novel compound iron metabolism mutants were also phenotypically characterized here for the first time. We demonstrate that serum hepcidin concentrations correlate with liver hepcidin mRNA expression, transferrin saturation and non-heme liver iron. In some circumstances, serum hepcidin-1 more accurately predicts iron parameters than hepcidin mRNA, and distinguishes smaller, statistically significant differences between experimental groups.     

Multidrug-Resistant Organism Infections in Patients with Left Ventricular Assist Devices

Left ventricular assist devices improve survival prospects in patients with end-stage heart failure; however, infection complicates up to 59% of implantation cases. How many of these infections are caused by multidrug-resistant organisms is unknown. We sought to identify the incidence, risk factors, and outcomes of multidrug-resistant organism infection in patients who have left ventricular assist devices.We retrospectively evaluated the incidence of multidrug-resistant organisms and the independent risk factors associated with them in 57 patients who had permanent left ventricular assist devices implanted at our institution from May 2007 through October 2011. Outcomes included death, transplantation, device explantation, number of subsequent hospital admissions, and number of subsequent admissions related to infection. Infections were categorized in accordance with criteria from the Infectious Diseases Council of the International Society for Heart and Lung Transplantation.Multidrug-resistant organism infections developed in 18 of 57 patients (31.6%)—a high incidence. We found 3 independent risk factors: therapeutic goal (destination therapy vs bridging), P=0.01; body mass index, P=0.04; and exposed velour at driveline exit sites, P=0.004. We found no significant differences in mortality, transplantation, or device explantation rates; however, there was a statistically significant increase in postimplantation hospital admissions in patients with multidrug-resistant organism infection. To our knowledge, this is the first report in the medical literature concerning multidrug-resistant organism infection in patients who have permanent left ventricular assist devices.     

In‐hospital worsening heart failure

Acute worsening heart failure (WHF) is seen in a sizable portion of patients hospitalized for heart failure, and is increasingly being recognized as an entity that is associated with an adverse in‐hospital course. WHF is generally defined as worsening heart failure symptoms and signs requiring an intensification of therapy, and is reported to be seen in anywhere from 5% to 42% of heart failure admissions. It is difficult to ascertain the exact epidemiology of WHF due to varying definitions used in the literature. Studies indicate that WHF cannot be precisely predicted on the basis of baseline variables assessed at the time of admission. Recent data suggest that some experimental therapies may reduce the risk of development of WHF among hospitalized heart failure patients, and this is associated with a reduction in risk of subsequent post‐discharge cardiovascular mortality. In this respect, WHF holds promise as a endpoint for acute heart failure clinical trials to better elucidate the benefit of targeted novel therapies. Better understanding of the pathophysiology and a consensus on the definition of WHF will further improve our epidemiological and clinical understanding of this entity.