Normal Cord Blood Samples Research Articles

Environmental Toxins Found Historically in the Polycythemia Vera Cluster Area and their Potential for Inducing DNA Damage.

In 2006, the Agency for Toxic Substances and Disease Registry received a request to determine whether a cluster of polycythemia vera patients existed in a northeast Pennsylvania community. A significant cluster of PV cases was identified at the nexus of three counties near several hazardous waste sites. The current study evaluated the potential for a select number of environmental contaminants previously detected in the cluster area to induce DNA damage using in vitro assays with hematopoietic stem-cell derived progenitor cells.CD34+ cells were isolated from normal cord blood samples and were cultured for 48–72 hours to generate erythroid progenitor cells. Eighteen compounds were chosen for the assay; arsenic trioxide, benzo(a)pyrene, benzene, methylene chloride, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), trichloroethylene, potassium chloride, ethylbenzene, benzo[k]fluoranthene, styrene, cadmium chloride, hydroquinone, 1,1,1-trichloroethane, sodium cyanide, manganese chloride, chromium oxide, lead oxide, and sodium arsenite. Genotoxicity of the compounds was determined using the comet assay, and toxicity determined via the cell viability assay.Using the comet assay, 16 compounds at 10 nM concentration, induced a significant amount of DNA damage compared to the control. When evaluating whether a dose-dependent relationship was present, seventeen of the eighteen compounds led to greater DNA damage with increasing exposure concentrations. 2,3,7,8-TCDD was particularly potent, inducing DNA damage in virtually all cells at 1 μM. In conclusion, most of the toxins evaluated using the comet assay showed potential to induce DNA damage in hematopoietic cells, and the genotoxic effects were dose-dependent.

Dexpramipexole As a Steroid-Sparing Agent in Hypereosinophilic Syndromes (HES): An Open-Label Proof-of-Concept Study

Hypereosinophilic syndromes (HES) are a heterogeneous group of disorders characterized by peripheral eosinophilia and eosinophil-related end organ damage. Whereas most patients respond to steroid therapy, high doses are often necessary and serious side effects are common. Dexpramipexole (KNS-760704) is an orally bioavailable synthetic amino-benzothiazole that was in development for the treatment of amyotrophic lateral sclerosis (ALS). Despite failure to meet the primary efficacy endpoint in a phase 3 trial in ALS patients, dexpramipexole showed an excellent safety profile and was found to significantly decrease absolute eosinophil counts (AEC) in >70% of the study participants. Consequently, a proof-of-concept study was designed to evaluate the safety and efficacy of dexpramipexole as a steroid-sparing agent in 10 HES subjects.Subjects with HES on steroid monotherapy were eligible for the study if they required ≥10 mg prednisone or equivalent for control of symptoms and eosinophilia. Subjects with AEC <1000/uL and no active symptoms entered a lead-in period, consisting of a standardized steroid taper with weekly assessment of AEC and symptoms to establish a "minimally effective corticosteroid dose (MECD)". Those with AEC ≥1000/uL entered directly into the treatment phase. Once the MECD was established, treatment with dexpramipexole (150 mg twice daily) was initiated. After 12 weeks of treatment, a standardized corticosteroid taper was attempted to determine the "MECD on dexpramipexole". Clinical assessments, including bone marrow and tissue biopsies of affected organs (when possible), were performed prior to and after 12 weeks of dexpramipexole. The primary efficacy endpoint was defined as a ≥50 % change in prednisone dose to maintain AEC at or below baseline levels and control clinical symptoms.Study enrollment is complete. Median age at enrollment was 51 years (22-72) with 40% women. Baseline clinical manifestations included eosinophilic gastrointestinal disease (60%), pulmonary involvement (50%), skin, muscle, sinus, and cardiac disease. Median baseline AEC was 670/uL (280-2540) and median MECD at baseline was 18.75 mg of prednisone (10-25). To date, the MECD on dexpramipexole has been determined for 7 subjects, and 3 subjects continue on study and have not completed the steroid taper on dexpramipexole. Two of the 7 evaluable subjects met the primary end point. Both subjects reached a MECD of 0 mg of prednisone and had complete symptom resolution with AEC of 0/uL within 3 months of initiating treatment with dexpramipexole. They remain in clinical remission on dexpramipexole monotherapy for 7 and 12 months, respectively. One additional subject who did not meet the primary endpoint at 3 months, was able to maintain a stable dose of 12.5 mg of prednisone with clinical improvement. She has remained on dexpramipexole and has demonstrated a delayed response with AEC 100/uL on 8.75 mg prednisone (50% of her baseline MECD) at 6 months. The remaining 4 evaluable subjects failed to respond and were taken off study. No deaths or drug related adverse events were observed, although 2 subjects reported transient palpitations and insomnia that resolved without drug discontinuation.Bone marrow biopsies performed at 12 weeks revealed markedly decreased AEC and basophils in responders as compared to baseline samples, and residual eosinophils appeared left-shifted. Other cell lineages were unchanged. Gastrointestinal biopsies in 1 responder demonstrated complete resolution of tissue eosinophilia after 5 months on dexpramipexole. Flow cytometry demonstrated no difference in CD34+ or CD34+IL-5R+ cell numbers after 12 weeks of treatment, but a decrease in eosinophil expression of Siglec 8, an inhibitory receptor expressed only on mature eosinophils. These changes were not observed in the non-responders. CD34+ cell cultures using normal cord blood samples also suggest a delay in maturation of the eosinophil lineage in the presence of dexpramipexole.Given the frequency of steroid-induced morbidity in patients with HES, alternate therapeutic options are critical. In this pilot study, dexpramipexole showed remarkable efficacy as a steroid-sparing agent without apparent toxicity in a subset of subjects with steroid-responsive HES. Although the mechanism of action is unknown, preliminary data suggest that dexpramipexole may affect eosinophil maturation in the bone marrow. DisclosuresBozik:Knopp Biosciences: Employment. Brown:Leidos Biomedical Research Inc.: Employment. Demarco:Knopp Biosciences: Employment. Komarov:Knopp Biosciences: Employment. Magee:Leidos Biomedical Research Inc.: Employment. Prussin:Knopp Biosciences: Employment. Signore:Knopp Biosciences: Employment. Sullivan:Knopp Biosciences: Employment. Wetzler:Leidos Biomedical Research Inc.: Employment. Dunbar:Novartis: Research Funding. Dworetzky:Knopp Biosciences: Employment.

Reference values of fetal serum β2-microglobulin in the Chinese: evaluation of its clinical usefulness

The level of β2-microglobulin was generally used to evaluate the renal function in adults. Elevated levels of β2-microglobulin were also applied to assess the perinatal situations in neonates and fetuses. The aim of our study was to establish and determine the reference values of fetal serum β2-microglobulin in the Chinese and to assess its clinical benefits in abnormal fetuses. Data from 308 normal cord blood samples were obtained to calculate the normal reference values of fetal serum β2-microglobulin. According to the equations we obtained, we analyzed the level of β2-microglobulin in four case groups: renal malformation, hydrops, cytomegalovirus (CMV) infection and rubella virus (RV) infection. In the normal group, the concentration of β2-microglobulin decreased with the gestational age, with a mean value of 4.35±0.59 mg/L. The upper limit of β2-microglobulin was calculated as 7.55-0.074×gestational age in weeks. The levels of β2-microglobulin were significantly higher in the four case groups than the normal group. For the four groups, the sensitivity is 72.7%, 69.6%, 86.7% and 100%, respectively. Fetal serum β2-microglobulin may be used as a predictor to evaluate the situations of fetal diseases.

Identification of Nine Novel Alternative Splicing Isoforms of RHD mRNA

To investigate the gene structures of RHD mRNA alternative splicing isoforms, total RNA was extracted from normal cord blood samples and RHD mRNA were detected using reverse transcription and polymerase chain reaction (RT-PCR) method. The PCR products were cloned and directly sequenced. The splicing signals of various isoforms were also analyzed by sequencing of exon-intron boundaries using RHD gene specific primers, and the expressed sequence tags (ESTs) databank was screened to find out other RHD alternative splicing isoforms. Nine novel and three recurrent RHD alternative splicing isoforms were identified, among which six isoforms were RHD-RHCE hybrid genes. Three types of alternative splicing were found including exon skipping, alternative 5' splice sites and alternative 3' splice sites. Exons 2 to 9 were involved in RHD mRNA alternative splicing. Two other isoforms of intron retention type were also found in human ESTs databank. Thus, the mRNA splicing mechanism of RHD gene was very complex, and nine novel alternative splicing isoforms were identified including several isoforms based on homologous RHD-RHCE hybrid gene.

Heterogeneity of fetal hemoglobin in severe alpha-thalassemia.

The heterogeneity of residue 136 of the gamma-chain of human hemoglobin has been determined for a patient afflicted with severe alpha-thalassemia. Separation of the cord blood sample into the various constituent hemoglobins A, F, FI and Bart's were done on a column packed with DEAE Sephadex. The amount of glycine or alanine at position 136 was determined for hemoglobins F, FI and Bart's. The ratios determined for all three hemoglobins indicated that the G gamma/A gamma ratio is the same for all three fractions and is similar to that observed in normal cord blood samples.

Fetal immune response following prematurely ruptured membranes

Concentrations of immunoglobulins (Ig)A1, and IgA2, IgD, IgE, IgG, and IgM have been determined in cord blood, amniotic fluid, and maternal serum in a group of patients with a history of prematurely ruptured membranes (PRM) prior to the onset of labor and in a control group of patients undergoing normal delivery and without a history of infection during pregnancy. IgA and IgD were determined by sensitive hemagglutination-inhibition tests; IgG and IgM, by radial immunodiffusion; IgE, by a radiommunoassay. There was evidence for an immune response in 10 of 16 cases of PRM: five of 16 had increased IgA but normal IgM; three of 16 had increased IgA and IgM; two of 16 had high IgM and normal IgA in cord blood. In patients with significantly increased levels of either IgA or IgM or both, there was a decreased level of IgD. These changes are most likely the result of the immune response to ascending infection from the maternal genitals. The sensitive testing method employed could demonstrate the presence of IgD in 53 per cent of normal cord blood samples and 72 per cent of amniotic fluid samples obtained at term. IgE was found in all normal cord blood and amniotic fluid samples tested. By concentrating the amniotic fluid up to 180-fold, IgM was demonstrated in all normal samples tested. The potential importance of IgA determinations in cord blood in addition to IgM determination for detection of intrauterine infections is stressed.