bluuebunny/crossref_metadata_2025 huggingface dataset:
The Association between Acute Lead Exposure from Indoor Firing Ranges and Iron Metabolism.
Authors: Merkel Drorit , Hirshhoren Nir , Shochat Tzippora , Vivante Asaf
Published: October 2019

Objective: To assess the impact of short term indoor firing ranges lead exposure and its relationship to iron, ferritin, lead, zinc protoporphyrin and hemoglobin concentration among young adults.

Methods: We report of a clinical observation that was carried out in 30 young and healthy soldiers serving in the Israel Defense Forces (IDF) Counter Terrorist Unit (CTU). Blood samples were drawn for Lead (Pb), Zinc Protoporphyrin (ZPP),Iron, Hemoglobin (Hb) and ferritin prior to and after a 6 weeks period of intensive shooting practice in indoor firing ranges.

Results: A mean Blood lead level increase (p< 0.0001) with a mean Iron (p<0.0005) and mean ferritin (p<0.0625) decrease simultaneously after 6 weeks period of Lead dusts exposure were demonstrated. We found a trend for inverse correlation between pre-exposure low ferritin levels and post exposure high blood lead levels.

Conclusion: We found decreased iron and ferritin levels after short term lead exposure among young adults. This can be explained by competition of iron and lead absorption viatransporters like DMT1 suggesting that lead poisoning can cause iron depletion and that iron depletion can aggravate lead poisoning. This synergistic effect should come to every physicians mind especially when treating patients with a potential risk for each problem separately.

Lead (Pb), Zinc Protoporphryn (ZPP), Hemoglobin (Hb), Iron and Ferritin among the CTU soldiers before and after the indoor firing ranges lead exposer variable N Pre-exposure N Post-exposure Difference *P value Pb (mcg/dl) 29 10.3±2.3 30 18.9±3.6 8.8±2.6 0.0001 ZPP(mcg/dl) 29 42±7.5 30 42.9±8.1 0.03±6.5 NS Hb (g/dl) 30 15±0.7 30 14.8±0.9 0.2±0.2 NS MCV (fl) 30 88.7±2.5 30 89±2.7 0.3±0.2 NS Iron (mcg/dl) 29 108.5±43.6 30 77.4±24.4 −30.4±41.2 0.0005 Ferritin (mcg/l) 27 58.1±27.7 16 51±19.9 −6.1±10.7 0.0625 Correlation between Pre-exposure Ferritin levels and Post exposure Lead level.(Person correlation coefficient = −0.28 r=0.14 number of observations = 27) Correlation between Pre-exposure Ferritin levels and Post exposure Lead level.(Person correlation coefficient = −0.28 r=0.14 number of observations = 27)
Epoetin alfa 80,000 U Every Three Weeks (Q3W) in Anemic Cancer Patients (pts) Not Receiving Chemotherapy (CT) or Radiation Therapy (RT).
Authors: Shasha Daniel , Dawkins Fitzroy , Wilhelm Francois E.
Published: October 2019

Anemia is reported at rates approaching 40% in cancer pts not receiving CT or RT. In two previous studies in anemic cancer pts not receiving CT or RT, epoetin alfa administered at 40,000 U sc once weekly1 or 60,000 U sc every two weeks2 was shown to be both safe and effective (hematopoietic response [HR] rates of 76.9% and 76.8%, respectively) in treating anemia. The primary objective of the current study was to evaluate the HR to epoetin alfa 80,000 U SC Q3W in this population; transfusions and quality of life were evaluated as well. Eligible pts had a non-myeloid malignancy, baseline hemoglobin (Hb) ≤ 11 g/dL, no CT administration within 8 weeks or RT within 4 weeks of study entry, and no plans to receive these therapies during the study period. Study drug was to be administered at Weeks 1, 4, 7, and 10, with follow-up continuing to Week 13. Epoetin alfa dose was reduced for Hb > 12 g/dL or Hb increase > 1.5 g/dL in any 3-week period; dose was withheld for Hb > 13 g/dL, then reduced when Hb ≤ 12 g/dL. All pts were to receive supplemental ferrous sulfate 325 mg po qd as tolerated. The primary efficacy endpoint was the proportion of pts achieving HR, defined as a ≥ 2 g/dL Hb increase and/or Hb ≥ 12 g/dL during the study, independent of transfusion within 28 days. 51 pts were enrolled (safety population) and 49 met the criteria for efficacy analyses (received ≥ 1 dose of epoetin alfa and had ≥ 1 post-baseline Hb value). Mean age was 71.0 ± 10.5 years, 53% of pts were female, 94% of pts had baseline ECOG status of 0 or 1, and mean baseline Hb was 10.3 ± 0.6 g/dL. The most common tumor types were breast (n=12, 24.5%) and prostate (n=12, 24.5%). HR was achieved in 37/49 (75.5%) pts. Mean time to first HR was 5.76 weeks. Hb change from baseline ≥ 1 g/dL was achieved in 39/49 (79.6%) pts, in a mean 3.77 weeks. 24 of 51 pts (47.1%) had at least one dose reduction or withhold. From Day 29 to end of study, 2/49 (4.1%) pts received PRBC transfusion. Statistically significant quality of life improvements from baseline in all 3 LASA scales (Energy Level, Daily Activities, and overall Quality of Life) were noted at Weeks 7 and 13/WD. Improvements were also seen with the FACT-An subscales, although not all changes were statistically significant at Weeks 7 and 13/WD. Adverse events occurring in ≥ 5% of the safety population were arthralgia (5 of 51 pts, 9.8%), back pain (3 of 51 pts, 5.9%), and vomiting (3 of 51 pts, 5.9%). One non-clinically relevant thrombotic vascular event of chest pain was reported. One pt died during the study, and 2 pts died during study follow-up; all deaths were considered unrelated to epoetin alfa therapy. In this population of cancer pts with anemia not receiving CT or RT, epoetin alfa 80,000 U sc administered Q3W safely increases Hb and improves pt quality of life. This extended dosing regimen further expands the flexible dosing options for epoetin alfa in this pt population.
AMD3100 and G-CSF Synergize To Promote Repopulating Stem Cell Mobilization in Fanca −/− and Fancc −/− Mice.
Authors: Pulliam Anna C. , Hobson M. Joe , Yang Fengchun , Broxmeyer Hal E. , Clapp D. Wade
Published: October 2019

Fanconi Anemia (FA) is an autosomal recessive DNA repair disorder clinically characterized by congenital abnormalities, progressive bone marrow (BM) failure and a propensity for malignancies. It is advantageous to bank FA patients’ cells for autologous treatment of pancytopenia, transplantation following chemotherapy or future gene therapy trials. Because the amount of cells able to be collected by BM harvesting is limited and the process is often problematic in children, mobilization of stem cells into the circulation would be an ideal means of collecting repopulating hematopoietic stem and progenitor cells (HSPC). Traditional mobilization protocols utilizing granulocyte colony-stimulating factor (G-CSF) as a mobilizing agent result in poor yields of phenotypic HSPC in FA patients. We have shown that analogous to the human patients with FA, administration of recombinant G-CSF is not sufficient to effectively mobilize myeloid progenitors from the BM of Fancc −/− mice (Broxmeyer et al, JEM, 2005). Antagonists to chemokine receptors, together with G-CSF, can enhance mobilization of murine wild type (WT) HSPC. AMD3100 is a selective antagonist of CXCR4, which is thought to function in the retention of HSPC in the BM. We hypothesized that the combination of AMD3100 and G-CSF would enhance mobilization of HSPC in Fanca −/− and Fancc −/− mice. Peripheral blood was collected to assess the number of myeloid progenitors and HSPC repopulating ability. Myeloid progenitor numbers were assessed using progenitor assays. Either G-CSF or AMD3100 alone resulted in poor mobilization of hematopoietic progenitors. In contrast, administration of AMD3100 and G-CSF together increased the number of colonies of myeloid progenitors up to 18 fold above that of either G-CSF or AMD3100 alone in both Fanca −/− and Fancc −/− mice. The stem cell activity of mobilized peripheral blood cells was evaluated using competitive repopulation assays, in which a common pool of isogenic ‘competitor cells’ and ‘test cells’ from mice of different genotypes or treatment conditions are co-transplanted. Fanca −/− and Fancc −/− cells mobilized with both G-CSF and AMD3100 yielded a 3–4 fold increase in peripheral blood chimerism as compared to the cells mobilized with G-CSF or AMD3100 alone. The latter mice also had a decreased survival. Phenotypic analysis of the peripheral blood of primary recipients confirmed the engraftment of multi-lineage test cells. Transplantation of chimeric bone marrow into secondary recipients maintained the test cell chimerism, confirming that long-term repopulating cells were mobilized. Collectively, these data from two FA murine models demonstrate that the AMD3100 and G-CSF protocol is an effective strategy to mobilize FA deficient repopulating HSPC. This has potential implications for more efficient banking of FA patients’ HSPC for future use or for use in gene transfer protocols.
Aplastic Anemia Due to Exposure to Benzene in Diamond Workers in Surat (India).
Authors: Shah Kiran C. , Naik Prashant K.
Published: October 2019

Benzene, used as a solvent in many industries is a known carcinogen which can lead to variety of hematological manifestations, like macrocytic anemia, aplastic anemia, myelodysplasticsyndrome and leukemia. However benzene induced aplastic anemia in diamond industry has not been reported so far in literature. We report series of cases of aplastic anemia from a single diamond factory, in Surat, India. All cases were diagnosed in June and July 2005. Benzene was used to remove whitener applied on diamonds before cutting them by Laser technology (Sarin technology from Israel). Diamond workers used benzene without any protective measures. Each worker used to clean about hundred diamonds daily by taking benzene in their hands. Most of the workers were using benzene for more than one year. The working environment used to have strong smell of benzene, however exact level of benzene in ppm was not determined. Out of 37 workers working in eight hour shift in a closed airconditioned room, eleven of them developed blood dyscrasia. Initially four patients presented with fever and pancytopenia. After confirmation of aplastic anemia in all of them remaining workers from same factory and other diamond factories using benzene were screened. From the same factory seven more cases of mild aplastic anemia were detected. Few cases of mild blood dyscrasia were detected from other diamond factories too. All cases were thoroughly examined clinically & investigated to rule out other causes of pancytopenia. Biochemical tests, liver functions, serum Iron, B12 study, tests for PNH, bone marrow aspiration & trephine biopsy and cytogenetics in severe cases were done. Three cases were treated with ATG and cyclosporine transfered to state capital, Ahmedabad. One patient died during treatment and two recovered. Two other cases were treated with cyclosporine alone and both of them recovered. No cases of MDS or leukemia detected from same factory so far. Since July 2005 benzene is no more used in diamond industry in India. After removing the workers from the exposure all other cases recovered. A close follow up for one year showed no recurrence or progression.
Hematological Characteristics of Infants and Young Children Presenting with Different Levels of Malarial Anemia at a Rural District Hospital in Western Kenya.
Authors: Novelli Enrico M. , Ong’echa John Michael , Davenport Greg , Keller Christopher C. , Were Tom , Ouma Collins , Otieno Richard , Rosano Caterina , Kaplan Sandra , Perkins Douglas J.
Published: October 2019

In areas of intense P. falciparum exposure, such as western Kenya, the most lethal complication of malaria is severe malarial anemia (SMA). The pathogenesis of SMA is multi-factorial and not fully understood. Our ongoing studies are, therefore, investigating the hematological manifestations of MA in a cohort of children presenting at a rural district hospital in western Kenya. A total of 619 children aged 3–33 mos. were categorized into 5 groups: SMA (Hb<6.0g/dL); moderate MA (MdMA; 6.0≤Hb<8.0g/dL); mild MA (MlMA; 8.0≤Hb<11.0g/dL); uncomplicated malaria (UM, Hb≥11.0g/dL), and healthy aparasitemic controls (HC). Parasite density, CBC, and reticulocyte count was obtained on all subjects (see table).

HC (n=62) UM (n=36) MlMA (n=139) MdMA (n=161) SMA (n=221) P valuea Data are presented as mean±SEM. aDifferences between the groups were compared using the Kruskal-Wallis test. *Denotes p<0.05 (Mann-Whitney U test) relative to the HC group. Parasitemia (/μL) 0 82471±15925 51711±5300 51513±5798) 62934±6416 0.107 Hb (g/dL) 1.81±0.11 11.66±0.14 9.27±0.07* 6.86±0.05* 4.78±0.06* <0.001 MCV (fL) 71.87±0.71 73.42±1.12 68.57±0.63* 69.25±0.76* 71.34±0.67 0.001 Reticulocyte count (%) 1.57±0.20 1.64±0.24 2.55±0.16* 4.21±0.29* 5.50±0.31* <0.001 RPI 1.83±0.33 1.90±0.49 1.44±0.09 1.50±0.10 1.15±0.07* 0.001

Peripheral blood smears (n=150) were randomly selected from all groups for examination of RBC and WBC morphology, and identification of any clinically significant abnormalities, such as presence of hemozoin (Hz). The majority of the children with malaria had inadequate bone marrow responses (77.6%), as determined by the reticulocyte production index (RPI). Relative to the UM group, the SMA group had increased microcytosis (p<0.005). Moreover, numbers of nucleated RBC (nRBC) and Hz-laden monocytes correlated with increasing degrees of anemia (p=0.08 and p<0.005, respectively). In contrast, levels of parasitemia, prevalence of high-density parasitemia (≥10,000 parasites/ml) and reactive lymphocytes did not differ among the four malaria groups. No significant increases were observed in the number of schistocytes, acanthocytes or spherocytes in any of the groups. Macrocytosis was rarely observed and sickle cells were not detected. These findings illustrate that parasitemia is not associated with the severity of anemia in this population. Conversely, the number of Hz-containing monocytes was a strong predictor of anemia severity, suggesting that SMA results from more chronic malaria infections. Finally, although nRBC were most commonly associated with SMA, the RPI was not appropriately increased for the degree of anemia, illustrating that these children have suppressed and/or ineffective erythropoiesis.
Successful Treatment of Pure Red Cell Aplasia (PRCA) Associated to Phenitoin without Blood Transfusions in a Jehovah’s Witness Patient.
Authors: Argote Victoria E. Ferrer , Ayala-Alcantara Noe , Ramos-Peñafiel CHristian O.
Published: October 2019

A 17 year-old Jehovah’s Witness girl was admitted to the neurosurgery ward with a subarachnoid hemorrhage Fisher IV, Hunt and Hess II, due to an arteriography-confirmed aneurism of the left posterior comunicating artery (LPCA). CT scan showed hemorrhages of the left lateral, 3rd and 4th ventriculi. Her blood counts were normal. She underwent surgery and during the procedure she developed neurologic deterioration due to brain edema. The procedure was stopped and she was started on prophylactic Phenitoin. Her hemoglobin levels began to drop progressively until they reached 3.6 gr/dL with 0.8% reticulocytes while her platelet and WBC counts remained within normal ranges (278 and 5.9 × 109/L respectively) with a normal differential count. No bleeding site was found, and hemorrhage was discarded as cause of the anemia. The serum iron was 218, the IBC 213 and the saturation index 102.5 (37.3%), all within normal values. The bone marrow aspirate showed a slight increase of the cellularity with normal megakariocytes and granulocytes with slight eosinophilia and a total absence of the erythroid series. The lymphocytes were also normal. This image was consistent with a PRCA. The existence of a thymoma as well as several viral infections including Parvovirus, EBV, HIV, HB and HC were discarded as cause of the PRCA. Phenitoin was stopped and treatment with steroids, cyclosporine, mesterolone and erythropoietin 8,000 U/d SC was started. Despite the evidence of very deep anemia with dispnea and prostration, which required red cell transfusions, the patient and her parents absolutely refused them for religious reasons and the supportive treatment was enhanced. Her clinical status improved after 48 hours and the Hb began to increase progressively. She was discharged one month later with a Hb of 13.1 gr/dL, Ht 39.6%, with a corrected reticulocyte count of 1.8%, 6.2 × 109/L WBC with a normal differential count and 284 × 109/L BP. Even though no tests could be done to confirm phenitoin as the triggering factor of the PRCA, the temporal association between its use and the development of the anemia very strongly suggests a causal association even though it can not be assured. The results we had with erythropoietin, high-dose iron, oxygen and parenteral feeding were very rewarding. Jehovah’s Witnesses patients with severe anemia like this one have a poor prognosis and pose a strong therapeutic challenge that may discourage the medical staff to continue a struggle that seems hopeless. We feel that by accepting it and sharing experiences like this one we can help these patients who will rather die than accept blood derivatives.
Early and Standard Intervention with Epoetin Alfa (PROCRIT®) 120,000 Units (U) Every Three Weeks (Q3W) in Patients with Cancer Receiving Chemotherapy - Results of a Planned Interim Analysis.
Authors: Glaspy John , Moyo Victor , Kamin Marc , Wilhelm Francois E.
Published: October 2019

Epoetin alfa has been used for chemotherapy-induced anemia (CIA) for over a decade, has an established efficacy and safety profile, and offers the flexibility of three times a week (TIW) or weekly (QW) dosing to match patient needs. Newer erythropoiesis stimulating therapies (ESTs) are being used for the treatment of CIA at dosing intervals of up to once every three weeks to coincide with commonly used chemotherapy regimens. A recent study suggested that initiating epoetin alfa every other week produced a similar hemoglobin (Hb) response as QW initiation1. We are conducting the first study to evaluate the efficacy and safety of initiating epoetin alfa Q3W for the treatment of CIA.

This 16-week study was designed to enroll 180 patients with non-myeloid malignancy, planned myelosuppressive chemotherapy for at least 9 weeks during the study, and with baseline Hb ≥ 11.0 and ≤ 12.0 g/dL. Patients were randomized to begin epoetin alfa 120,000 U SC immediately (early intervention) or when their Hb decreased to < 11 g/dL (standard intervention). In both groups, dosing was reduced to 80,000 U Q3W if Hb was > 12.0 g/dL at the time of a dosing visit or increased by more than 1.5 g/dL in a 3-week period. If a patient’s Hb was < 10.0 g/dL at any dosing visit after the first dose, the regimen was changed to 40,000 U QW. The objective of this planned interim analysis was to explore the safety and efficacy of the 120,000 U Q3W regimen.

We report results for the first 46 patients who received study drug and had at least one post-baseline Hb (26 early intervention and 20 standard intervention patients). Mean Hb values over time for patients receiving epoetin alfa Q3W are presented in the table below. Standard intervention patients had a mean Hb increase of 1.7 g/dL after two doses (week 7). Early intervention patients had weekly mean Hb values in the 11.0 to 12.0 g/dL range while receiving Q3W dosing. By week 5, weekly mean Hb levels were similar between the two groups. Eighteen patients (11 early and 7 standard) had at least one dose reduction to 80,000 U Q3W. Eleven patients (6 early and 5 standard) converted to 40,000 U QW. Epoetin alfa was well tolerated in both groups. A total of 4 (2 early and 2 standard) out of 47 treated patients (8.5%) experienced clinically relevant thrombotic vascular events.

The results of this interim analysis suggest that epoetin alfa can be initiated at 120,000 U Q3W in cancer patients with CIA, potentially offering the same Q3W initiation dosing option as the newer ESTs, yet with the added advantage of established TIW and QW dosing.

Mean Hb (g/dL) Over Time (N)* Week on Study Early Intervention (N=26) Standard Intervention (N=20) *Hb values censored after patients switched to 40,000 U QW Baseline (1) 11.5 (26) 10.4 (20) 4 11.7 (26) 10.9 (19) 7 11.6 (24) 12.1 (14) 10 11.6 (21) 11.8 (11) 13 11.9 (15) 12.3 (8) 16 12.3 (8) 11.8 (2)
Flippase Activity Decreases throughout Erythrocyte Life-Span in Normal and Sickle Mice.
Authors: DeJong Kitty , Bhagat Amrita , Emerson Renee K. , Kuypers Frans A.
Published: October 2019

Phosphatidylserine (PS) exposure contributes to recognition and removal of aged erythrocytes. PS exposure can only arise in cells in which the aminophospholipid translocase, or flippase, is defunct, or otherwise PS would be transported back into the inner cell monolayer. To provide evidence that flippase activity is reduced in old erythrocytes, we studied mouse erythrocytes that were biotinylated in vivo. This way we were able to distinguish the aging, biotinylated, RBC population from the newly formed, unbiotinylated RBC. Detection of the biotinylation with fluorescent streptavidin was combined with a flow cytometric assay for flippase activity using NBD-PS. We found that flippase activity decreased over time during the life-span of the RBC, as shown by a decrease in flippase activity of the biotinylated RBC compared to the newly formed RBC. A particularly high flippase activity was found in CD71-positive reticulocytes. Murine sickle cells show a much higher erythrocyte turnover than normal erythrocytes, with a 25–50% newly formed population each day and complete elimination of all biotinylated cells in 10 days. As in control mice, flippase activity decreased during the (short) life-span of sickle RBC and at all times, a subpopulation of sickle cells had a markedly reduced flippase activity. High flippase activity was only found in newly released erythrocytes. Our data indicate that the flippase activity decreases throughout the life-span of erythrocytes and can be markedly different in subpopulations of RBC in vivo. This decrease reduces the cell’s potential to maintain its phospholipid asymmetry, which results in PS exposure as a removal signal for the damaged cell. We conclude that the inability of sickle RBC to maintain flippase activity is related to the decreased survival time of those cells.
Long-Term Vitamin D Supplementation Improves Bone Mineral Density (BMD) in Patients with Sickle Cell Disease.
Authors: Adewoye Adeboye H. , Holick Michael F. , McMahon Lillian C. , Ma Qanli , Steinberg Martin H.
Published: October 2019

Among the many complications of sickle cell disease (SCD) are osteopenia and osteoporosis. We and others have previously documented a high prevalence of low serum 25-OH vitamin D level with accompanying osteopenia and osteoporosis in patients with SCD; however the long-term effects of treatment on BMD have not been reported. We hypothesized that supplementation with oral vitamin D and calcium will result in an improvement in BMD and accordingly, treated 6 adult subjects with SCD for 12 months with oral supplementation with vitamin D and calcium and studied the effects of treatment on BMD. Subjects with serum 25-OH D levels <20 ng/ml were placed on 50,000 IU of oral vitamin D weekly for 8 weeks, followed by 50,000 IU every other week for 44 weeks; subjects also received 1000 mg of elemental calcium daily. We evaluated BMD at weeks 1 and 52. Subjects had a mean pre-treatment plasma 25-OH vitamin D of 9.1 ng/ml. BMD was abnormal in all subjects; 4 subjects had osteoporosis with a Z-score <-2.5, and 2 subjects had osteopenia with a Z-score −1.9 to −2.5 (in the lumbo-sacral vertebrae). All subjects had mildly elevated intact PTH levels which were within the reference range but declined further post-treatment. Following one year of treatment with vitamin D and calcium, we observed a mean post-treatment increase in BMD of 5.4% (p value <0.02) and a 176% increase in plasma 25-OH D level (from 9.1ng/ml to 28ng/ml; p value <0.009). All patients with SCD we have studied have hypovitaminosis D that in adults is associated with low BMD. In patients followed for one year who received supplementation with calcium and vitamin D, 25-OH D level increased in all and BMD improved in four of six subjects. These results suggest that this treatment regimen can be effective is restoring BMD. The reasons for apparent unresponsiveness in some patients must be further evaluated. Clinical trials to document the effectiveness of treatment for this ubiquitous problem should be instituted.

Mean change in 25-OH vitamin D level Mean change in 25-OH vitamin D level Mean Change in BMD: Pre and Post Treatment Mean Change in BMD: Pre and Post Treatment
Genomic Instability in Sickle Cell Disease Patients Using Hydroxyurea Assessed by Micronucleus Assay.
Authors: Friedrisch Joao R. , Maluf Sharbel W. , Bittar Christina M. , Silva Maria A. , Silla Lucia M.
Published: October 2019

Hydroxyurea (HU) is an antineoplastic drug, which also plays an important role in treatment of sickle cell disease patients (SCD). Short-term HU toxicities primarily include transient myelosuppression, but long-term HU risks have not been defined. The mutagenic and carcinogenic potential of HU is not established, although HU has been associated with an increased risk of leukemia in some patients with myeloproliferative disorders.

In the present study we analyzed the presence of DNA damage in patients with SCD treated with HU, in peripheral blood lymphocytes, using micronucleus assay. We analyzed 36 patients with SCD (16 males and 20 females), aged 2–59 years (mean 25,75 ± 14,5), received oral HU median dose of 26,5 mg/Kg/day, for a period of 0.6– 11,8 years (mean 5,01). The control group was composed of 34 healthy individuals (16 males and 18 females), aged 4–52 years (mean 26,61 ± 7,20). The results revealed that frequency of micronucleus was significantly higher in SCD patients using HU than in controls(p=0,0182). This study indicates a possible genotoxicity of the HU, although further works are necessary to evaluate its mutagenicity.
Exercise Capacity and Pulmonary Function in Sickle Cell Patients with Pulmonary Arterial Hypertension.
Authors: van Beers Eduard J. , van der Plas Mart N. , Bogaard Harm J. , Biemond Bart J.
Published: October 2019

Pulmonary arterial hypertension (PAH) is increasingly recognised as a complication of sickle cell disease (SCD), with a prevalence of approximately 30%. In SCD, a tricuspid regurgitant jet velocity (TRV; determined by cardiac ultrasound) of 2.5 m/s or higher is used as a non-invasive marker of PAH and is associated with a poor prognosis. Since PAH in SCD is usually mild, it remains unclear whether PAH is directly responsible for the early death of these patients. Exercise capacity is closely related to survival in heart failure and other chronic cardiopulmonary diseases. We hypothesized that if PAH in SCD determines survival, it should be related to exercise capacity. The aim of this study was to determine whether exercise capacity is reduced in SCD patients with an increased TRV. In addition, it was determined whether exercise capacity is related to pulmonary function in these patients. Cardiac ultrasound, pulmonary function testing and a symptom-limited maximal cardiopulmonary exercise tests(CPET) using cycle ergometry were performed in 42 consecutive patients with SCD. PAH was present in 9/42 (21%) of the patients.

Exercise capacity was decreased in all patients, with a trend to a lower peak oxygen uptake (VO2peak) and oxygen uptake at the anaerobic threshold (VO2peak/AT) in patients with PAH compared to patients without PAH (74±18% and 57±18% versus 63±14% and 46±10%, respectively, p=0.16 and p=0.095). After matching for blood hemoglobin content, differences in exercise capacity disappeared. Pulmonary function was also impaired in both groups, with much larger differences between PAH and non-PAH patients: vital capacity (%predicted) 72±13% vs 85±15% and total lung capacity 70±11% vs. 81±11% (p=0.025 and p=0.020). No differences in FEV1 or membrane diffusion capacity were demonstrated. Multiple regression analysis, with the use of VO2peak as dependent variable and age, SCD phenotype, hemoglobin, spirometric parametres and TRV identified only vital capacity and hemoglobin as a significant independent correlates of VO2peak and not TRV. In conclusion, PAH in SCD is not related to exercise capacity, which could mean that it has no direct effect on mortality. SCD is associated with a restrictive pulmonary function impairment, especially in the group of patients with PAH. Interestingly, low VO2peak in SCD appears to be determined by the severity of the anemia instead of the prevalence of PAH.
Monocyte Chemoattractant Protein-1 and Interleukin-8 Levels in Acute Inflammation Induced by Prolonged Brisk Exercise.
Authors: Papassotiriou Ioannis , Tsironi Maria , Skenderi Katerina , Chrousos George
Published: October 2019

Monocyte chemoattractant protein-1 (MCP-1), also known as CCL2, a chemokine that regulates migration and infiltration by monocytes/macrophages, belongs to the CC chemokine subfamily. Interleukin-8 (IL-8), also known as CXCL8, a proinflammatory chemokine with angiogenesis-promoting properties belongs to the CXC chemokine superfamily. Both MCP-1 and IL-8 have important roles in the pathogenesis of many chronic inflammatory disorders, including atherosclerosis and obesity. Their proinflammatory effects are mediated mainly by the CC chemokine receptor 2 (CCR2) and CXCR1/R2, respectively. MCP-1 and IL-8 cause chronic vascular inflammation and induce thrombosis, proliferation and migration of vascular smooth muscle cells, angiogenesis, and oxidative stress. Previous studies indicate that:

MCP-1 production from endothelial cells, smooth muscle cells, and regional leukocytes increases in the presence of endothelial dysfunction and atherosclerosis risk factors; MCP-1 and IL-8 expression is increased in atherosclerotic lesions and injured arteries; and eliminating MCP-1 function decreases neointimal hyperplasia after injury and atheroma formation in mice.

We studied the association between MCP-1 and IL-8 levels and the degree of inflammation in 15 athletes that participated in the ultra-distance foot race of the 246 Km “Sparthathlon”. This race consists of continuous, prolonged, brisk exercise. We reported earlier that Interleukin-6 (IL-6), C-reactive protein (CRP), Serum amyloid A protein (SAA) and free plasma DNA levels markedly increased (by 8000–, 152– 108– and 10-fold, respectively) over the baseline at the end of the race1. However, IL-6 levels returned to normal by 48h, while CRP, SAA and free plasma DNA remained elevated. Circulated levels of MCP-1 and IL-8 were measured by means of a multi-analyte Biochip Array Technology, using the Evidence analyzer (Randox Laboratories, UK). The measurements were performed before (phase I), at the end (phase II) and 48h post-race (phase III). MCP-1 levels at phase I (216.9±48.5 ng/L) (mean±SE), increased significantly at phase II (592.9±115.7 ng/L) and subsequently decreased at phase III (278.1±62.7 ng/L). At the same time period, IL-8 followed a similar pattern (phase I: 9.4±4.5 ng/L, phase II: 28.5±8.8 and phase III: 8.9±4.3 ng/L). A significant positive correlation between MCP-1 and IL-8 was found at phase III (r=0.845, p<0.01), while this correlation was absent in the other two phases, indicating an independent response of each chemokine to inflammatory stimuli in each athlete.

In conclusion, prolonged exercise induces an inflammatory response that is expressed by an increase in circulating MCP-1 and IL-8 levels. Whether these changes have long-term negative effects on the vasculature remains unknown.
HNA−1a, −1b, −2a, −3a and −4a Frequencies in Brazilian Persons.
Authors: Norcia Angela M. , Kimura Elisa Y. , Chiba Akemi K. , Moritz Elyse , Yamamoto Mihoko , Bordin Jose Orlando
Published: October 2019

Granulocyte reactive antibodies have been found to cause clinical disorders such as transfusion related acute lung injury (TRALI), febrile transfusion reactions, alloimmune neonatal neutropenia, immune neutropenia after bone marrow transplantation, refractoriness to granulocyte transfusion, drug-induced neutropenia and autoimmune neutropenia. Using the granulocyte immunofluorescence test (GIFT) by microscopic and flow cytometric analysis, the frequencies of the neutrophil antigens HNA−1a, −1b, −2a, −3a and −4a were determined among 100 random Brazilian blood donors from the Blood Center of Universidade Federal de Sao Paulo, SP, Brazil. Granulocytes were separated from mononuclear cells and red cells by sedimentation with 5% dextran, followed by centrifugation on Ficoll-paque (d = 1.077), and then incubated with anti-sera (anti-HNA−1a, −1b, −2a, −3a, and −4a obtained from American Red Cross, North Central Blood Services, St. Paul, MN) conjugated with fluorescein isothiocyanate (FITC) labeled F(ab’)2 fragments of anti-human IgG. Only cell suspensions containing ≥95% neutrophils with viability ≥90% according to the trypan-blue staining were analyzed. The frequencies of HNA−1a, −1b and −2a were 65%, 83% and 94%, respectively, and for such alloantigens exact same results were observed using either the GIFT performed by microscopy or by flow cytometry. The frequency of HNA-3a was 86% by the microscopic GIFT, and 95% by the flow cytometry analysis; while the frequency of HNA−4a was 93% by the microscopic GIFT, and 94% by the flow cytometric technique. These results indicate that:

GIFT by flow cytometry is more sensitive than the GIFT by microscopy to detect HNA−3a; the phenotypic frequencies found for neutrophil antigens HNA−1a and −1b among Brazilian blood donors are quite similar to those reported among African Americans, but different from those reported for Japanese and Chinese individuals; the phenotype frequencies of the neutrophil antigens HNA−2a, −3a, and −4a in Brazilians are quite similar to those found among Caucasians (Table). (These studies were funded by FAPESP, SP, Brazil - 05/55237–9).

Asians Brazilians Antigen African Americans Chineses Hindus Japaneses Caucasians M FC M, microscopy; FC, flow cytometry; nd, not done HNA-1a 46 – 68 90 44 88 52 – 54 65 65 HNA-1b 78 – 84 52 83 51 – 64 87 – 89 83 83 HNA-2a nd 99 nd 89 87 – 97 94 94 HNA-3a nd nd nd nd 99 86 95 HNA-4a nd nd nd nd 96 93 94
Upregulation of Myeloperoxidase in End Stage Renal Disease and Its Modulation by Hemodialysis. A Paradoxical Paradigm.
Authors: Cunanan Josephine , Hoppensteadt Debra , Bansal Vinod , Fareed Jawed
Published: October 2019

Myeloperoxidase (MPO), a heme protein released from activated leucocytes mediates the generation of reactive intermediates promoting lipid peroxidation. MPO is secreted by activated neutrophils, monocytes and macrophages. MPO has been reported to be a marker of hemodialysis procedure biocompatibility and oxidative stress with discordant results. Different dialyzer membranes and the type of anticoagulation used contribute to the regulation of MPO in end stage renal disease (ESRD) patients. To validate the hypothesis that MPO is upregulated in ESRD and hemodialysis differentially modulates the MPO in patients with this syndrome, blood plasma samples from 70 patients on maintenance hemodialysis prior to and after hemodialysis session. The pre-dialysis levels of MPO antigen as measured by an ELISA method (Assay Design, ) ranged from 0.5–125 ng/ml (31.1±28.6 ng/ml), in contrast to the aged match normal controls where the range was 0.21–7.5 ng/ml, (2.8±1.3 ng/ml). After dialysis, 10/70 patients did not show any changes in the MPO level, whereas, 4 patients showed a decrease, the remaining 56 patients exhibited marked increase in the circulating MPO level ranging from 9.5–201 ng/ml (74.4±48.2 ng/ml). Progressive atherosclerotic changes are associated with the severity of ESRD where oxidative stress is known to play a major role. While MPO may mediated oxidative stress the beneficial consequences include its depletion from the vascular site. Hemodialysis in ESRD generally results in a downregulation of inflammatory mediators. The increased MPO level in ESRD represents a paradoxical and complex process which may contribute to the pathogenesis of ESRD. The role of different anticoagulants and dialyzer membrane and other factors in regulating circulating MPO level may be important in elucidating the pathogenesis and outcome of this syndrome.
Characterization of Dendritic Cell-Specific Genes p275 and p306.
Authors: Hilgendorf Ingo , Kurz Daniel , Bringmann Anita , Kanz Lothar , Grünebach Frank , Brossart Peter
Published: October 2019

Dendritic cells play an inimitable role in the functioning immune system as they are the most potent antigen presenting cells being able to prime naive T-cells. Their characteristic properties that enable them to take up antigens and present them to leukocytes are due to an expression of specific genes and thus specific proteins that are unique to this subset of antigen-presenting cells. Using a substractive cDNA library based on suppression hybridization between DC cDNA and the reference monocyte cDNA, we identified in DC two differentially expressed genes p275 and p306. p275 codes for a membrane protein and represents a splice isoform of the transport protein NAT-1. The predicted structure of protein p306 is globular, suggesting that the protein is either intracellular or secreted. The expression of both genes was confirmed by RT-PCR using cDNA isolated from peripheral blood monocytes and DC, generated in vitro from monocytes or CD34+ progenitor cells. To further analyze the protein expression polyclonal antibodies were generated by immunization with synthetic peptides deduced from the identified sequences. Interestingly, inhibition of DC differentiation using IL-10 or STI571 (Imatinib) resulted in an impaired expression of both proteins. Utilizing specific primers for two recently described splice variants of p306 we identified a new splice form expressed in DC. While the gene of p306 contains eight exons, splice variant 1 consists of the exons 1,2,4,5,6, and 7 and splice variant 2 contains the exons 1,2,3,4,5,6, a shortened exon 7, and exon 8. The new identified splice form includes the exons 1–7. However, as the open reading frame starts in exon 4, the expressed protein is identical with the one corresponding to splice variant 1. Analyzing different DC populations in peripheral blood we show that p306 is expressed in plasmacytoid, but not myeloid DC. Interestingly, the activation of DC with Toll-like receptor ligands (TLRL) Pam3Cys (TLR2L), Poly I:C (TLR3L), LPS (TLR4L) and R848 (TLR7L) has no influence on the expression of p306. Although the functions of p275 and p306 in DC have yet to be determined, both genes play a role in DC differentiation and are found in different hematopoietic cell populations. Especially p306 might be an interesting marker of plasmacytoid DC as the predicted protein does not resemble any known protein structure.
Enhanced Bacteriocidal Function by WKYMVm in Patients with Acute Leukemia.
Authors: Kim Hawk , Shin Soojin , Park Jae-Hoo , Noh Eui-Kyu , Min Young Joo
Published: October 2019

Acute leukemia (AL) is a disease of challenge in that many patients die not only from eventual relapse but also from infection during treatment. In this regard efforts for decreasing infectious mortality will help increasing survival. Our previous study revealed that leukocyte bacteriocidal functions in chemotherapy-treated cancer patients were decreased and were stimulated by a novel hexapeptide, WKYMVm. We evaluated the leukocyte bacteriocidal function in patients with AL and searched whether WKYMVm can enhance the bacteriocidal function in patients with AL. On induction chemotherapy, blood sampling was performed at diagnosis and repeated weekly from start day of chemotherapy until patient died or complete remission was achieved. Tests were done weekly until white blood cell (WBC) count reached up to 1000/mm3 and platelet counts were stable without transfusion after consolidation chemotherapy. Nineteen AL patients and 10 healthy controls were enrolled. Diseases were acute myeloid leukemia (17 patients) and acute lymphoblastic leukemia (2 patients). Median WBC, absolute blast count (ABC) and absolute neutrophil count (ANC) at diagnosis were 5280/mm3 (range 840–315100), 162.9/mm3 (range 0–286741) and 796.4/mm3 (range 126–7336), respectively. Mean values of bacreriocidal activity at diagnosis were increased by concentrations of WKYMVm (13.4 at 0 nM; 24.9 at 1 nM; 33.3 at 10 nM; 38.5 at 100 nM; p<0.001), which were also increased in normal samples (20.6 at 0 nM; 41.6 at 1 nM; 51.6 at 10 nM; 66.4 at 100 nM; p<0.001). At each concentrations of WKYMVm, the bacteriocidal activities were inferior to those of normal control (p=0.029, at 0 nM; p=0.015, at 1 nM; p=0.015, at 10 nM; p=0.015, at 100 nM). The bacteriocidal activities were increased (p=0.008, at 0 nM; p=0.015, at 1 nM; p=0.011, at 10 nM; p=0.021, at 100 nM) compared with the corresponding values when patients achieved complete remission (CR). However, bactericidal activities by stimulation of WKYMVm were inferior to normal control even in CR (p=0.036, at 1 nM; p=0.036, at 10 nM; p=0.036, at 100 nM) although base line value were similar (p=1.0, at 0 nM). After hematological recovery of consolidation chemotherapy, the bacteriocidal activities of patients were similar to those of normal control (p=0.533, at 0 nM; p=0.133, at 1 nM; p=0.133, at 10 nM; p=0.133, at 100 nM). In conclusion, the bacteriocidal activities in AL patients were severely decreased at diagnosis and could be enhanced by WKYMVm. Near normal bacteriocidal activities can be achieved when 1 nM or more concentration of WKYMVm is applied in patients with AL. At the end of consolidation, bacteriocidal activities and stimulation by WKYMVm in patients were almost recovered.
Adult Hemophagocytic Lymphohistiocytosis: Clinical Presentation and Follow-Up in a Consecutive Series of Ten Patients.
Authors: Shirai Keisuke , Montero Alberto , Powell Jesse , Christiansen Lydia , Lazarchick John
Published: October 2019

Hemophagocytic Lymphohistiocytosis (HLH) is very rare in adults but can be fatal without treatment. Reports in adults are limited to case reports and very small case series. Clinically it is characterized by fever, hepatosplenomegaly, lymphadenopathy, severe cytopenias, hepatic dysfunction, coagulopathy, as well as neurological involvement. This syndrome is associated with diverse processes including: infection, rheumatologic, and hematologic malignancies. Presently, the underlying cause of HLH is unknown. We present a cosecutive series of 10 adults with HLH diagnosed at our institution between 2004–2006. All diagnoses were confirmed by pathology. The median age was 59 years (range: 18–73 years), and a male: female ratio of 4:1. All patients uniformly presented with fever. Half of the patients presented with evidence of hepatomegaly or splenomegaly. The most predominant laboratory abnormalities included: leukopenia or thrombocytopenia (100%), and elevation of liver enzymes (50%). EBV IgG was positive in 8 of 10 patients. The underline illnesses associated with HLH were diverse. The underlying causes were as follows; acute leukemia (n=2), infection (n=2), rheumatologic (n=2), post transplant (n=2) sickle cell disease (n=2), unknown (n=2). Mortality rate was 60% with a median survival time since diagnosis of 58 days. One patient is still on maintenance cyclosporin after etoposide treatment. One patient is on steroid and cytoxan. One patient recovered just with supportive care. In conclusion, due to the high morality rate associated with HLH, early treatment with immunosuppressant is warranted and attempts to identify underlying cause.
Immune Suppression by Placenta Derived Adherent Cells (PDAC) Correlate with Monocyte Chemoattractant Protein-1 and IL-2 Secretion.
Authors: Paludan Casper , Harbacheuski Ryhor , Murray Rose Ann , Mendillo Megan , Soler Jorge , Wang Jia-Lun , Ye Qian , Edinger James , Hofgartner Wolfgang , Kaplan Gilla , Hariri Robert
Published: October 2019

The placenta is a readily available and ethically non-controversial source of large amounts of therapeutic stem cells. Placenta Derived Adherent Cells (PDACs) are isolated from the placenta by one of several methods including physical disruption of tissue from several different anatomical sites within the placenta that include the amniotic membrane, chorion, placental cotyledons, or any combination thereof. Flow cytometry analysis showed that PDACs isolated from certain sites exhibit defined phenotypes, including for example CD200+ CD105+ CD73+ CD34− CD45− at percentages ≥70% and constitutively secrete IL-6, IL-8, and Monocyte Chemoattractant Protein-1 (MCP-1). PDACs demonstrate in vitro pluripotency in the adipogenic, osteogenic, and chondrogenic lineages. Furthermore, PDACs suppress T cell proliferation in certain Mixed Leukocyte Reaction (MLR) and the autologous EBV regression assays. Because secreted factors can powerfully modify immune responses and influence therapeutic use of cells, we report on the cytokine secretion in certain PDAC MLR and regression assays.

Cytokines were measured on a Luminex system in supernatants from 6-day PDAC cultures, PDAC MLRs or PDAC regression assays. MLRs include PDACs, Dendritic Cells (DC)s, and T cells at DC/PDAC/T ratios 1/2/10. EBV regression assays included PDACs, EBV antigen-presenting cells (APC), and T cells at APC/PDAC/T ratios 1/2/10. Levels of IL-6 (11 ng/ml) and IL-8 (16 ng/ml) stayed constant in PDAC solo cultures, PDAC MLRs, and PDAC regression assays. MCP-1 concentration was 2 ng/ml in PDAC solo cultures, and non-suppressive control adherent cell MLRs and regression assays, but increased to 10 ng/ml in suppressed PDAC MLRs and PDAC regression assays. These values are consistent with reported MCP-1 serum levels.

Interleukin-2 (IL-2) is both a T cell survival factor and an obligate factor for CD4+CD25+ T regulatory cells. T regulatory cells are not required for PDAC T cell suppression, but IL-2 levels consistently increase when MLR suppression by PDACs occurs. The CD4 MLR supernatants contained 65 pg/ml IL-2, and the CD8 MLR contained 35 pg/ml IL-2. In the 85% and 75% suppressed CD4 and CD8 PDAC MLRs, the IL-2 levels rose 5-fold to 331 pg/ml (CD4) and 2-fold to 67 pg/ml(CD8).

These results indicate that IL-2 and MCP-1, traditionally known as stimulators of the immune response, may play a role in PDAC immune suppression. PDACs, which cause the secretion, may thus be useful therapeutic tools in the clinic.
The Study on Difference in Megakaryocyte Expansion between Cord Blood and Peripheral Blood.
Authors: Fang Jian-pei , Xia Ting , Chen Guo-hua , Wu Yan-fen , Xu Hong-gui , Wei Qing , Huang Shao-liang
Published: October 2019

Objective Currently, thrombocytopenia is typically observed in patients undergoing hematopioetic stem cell transplantation (HSCT), high-dose chemotherapy or irradiation. Severe thrombocytopenia can cause bowel hemorrhage and intracranial hemorrhage. To transfuse ex vivo-expanded Mk cells into patients can reinforce the ability of platelet formation and shorten the time of platelet recovery. So it is one of the effective approaches to protect the danger. To explore the differences in megakaryocyte (MK) expansion between CD34+ stem cells derived from cord blood (CB) and Peripheral Blood (PB) and to establish the most optimal culture system.

Methods Mononuclear cells were isolated by density gradient centrifugation over Ficoll. CD34+ cells were isolated by positive selection using an immunomagnetic separation system. selected CD34+ cells were seeded in Iscove’s modified Kulbecco’s medium (IMDM) supplemented with fetal calf serum (FCS) and all kinds of cytokines. After 15–17 days of culture, the cells were measured and determined the content of CD41+ cells by flow cytometry, and simultaneously measured the number of megakaryocyte colony-forming unit (CFU-MK).

Results After the definite days of culture, the cytokine combination of TPO+FL+IL-6+IL-3 is most suitable both for PB and CB to obtain high numbers of MK, and better than any other three groups(P<0.05). The fold-increase of CD41+ cells from CB on day 14 was 193.23±25.24 fold, and that from PB on day 10 was 131.23±18.26 fold respectively. With the time gone, the number of CD41+ cells both decreased.

Conclusion For PB and CB, the cytokine combination of TPO+FL+IL-6+IL-3 is most suitable to obtain high numbers of MK and is the best combination for MK ex vivo expansion. It is suggested that MKs from CB have higher proliferation potential than that from PB, and the optimal culture duration of MK from PB is shorter than that of MK from CB. The study explore the differences in MK expansion between CD34+ stem cells derived from CB and PB and establish experimental foundation for further application to pre-clinic and clinic.