The conversion of adenosine to inosine is catalyzed by adenosine deaminase (ADA) (EC 3.5.4.4), which has two isoforms in humans (ADA1 and ADA2) and belongs to the zinc-dependent hydrolase family. ADA ...modulates lymphocyte function and differentiation, and regulates inflammatory and immune responses. This study investigated ADA activity in lymphocyte-rich peripheral blood mononuclear cells (PBMCs) in the absence of disease. The viability of lymphocyte-rich PBMCs isolated from humans and kept in 0.9% saline solution at 4-8°C was analyzed over 20 h. The incubation time and biochemical properties of the enzyme, such as its Michaelis-Menten constant (Km) and maximum velocity (Vmax), were characterized through the liberation of ammonia from the adenosine substrate. Additionally, the presence of ADA protein on the lymphocyte surface was determined by flow cytometry using an anti-CD26 monoclonal human antibody, and the PBMCs showed long-term viability after 20 h. The ADA enzymatic activity was linear from 15 to 120 min of incubation, from 2.5 to 12.5 µg of protein, and pH 6.0 to 7.4. The Km and Vmax values were 0.103±0.051 mM and 0.025±0.001 nmol NH3·mg-1·s-1, respectively. Zinc and erythro-9-(2-hydroxy-3-nonyl) adenine (EHNA) inhibited enzymatic activity, and substrate preference was given to adenosine over 2'-deoxyadenosine and guanosine. The present study provides the biochemical characterization of ADA in human lymphocyte-rich PBMCs, and indicates the appropriate conditions for enzyme activity quantification.
The MLL-AF9 fusion gene is associated with aggressive leukemias of both the myeloid and lymphoid lineage in infants, whereas in adults, this translocation is mainly associated with acute myeloid ...leukemia. These observations suggest that differences exist between fetal and adult tissues in terms of the 'cell of origin' from which the leukemia develops. Here we show that depending on extrinsic cues, human neonatal CD34(+) cells are readily immortalized along either the myeloid or lymphoid lineage upon MLL-AF9 expression and give rise to mainly lymphoid leukemia in immunocompromised mice. In contrast, immortalization of adult bone marrow CD34(+) cells is more difficult to achieve and is myeloid-biased, even when MLL-AF9 is expressed in purified hematopoietic stem cells (HSCs). Transcriptome analysis identified enrichment of HSC but not progenitor gene signatures in MLL-AF9-expressing cells. Although not observed in adult cells, neonatal cells expressing MLL-AF9 were enriched for gene signatures associated with poor prognosis, resistance to chemotherapeutic agents and MYC signaling. These results indicate that neonatal cells are inherently more prone to MLL-AF9-mediated immortalization than adult cells and suggest that intrinsic properties of the cell of origin, in addition to extrinsic cues, dictate lineage of the immortalized cell.
Fluorochrome choices for multi-color flow cytometry Flores-Montero, Juan; Kalina, Tomas; Corral-Mateos, Alba ...
Journal of immunological methods,
December 2019, 2019-12-00, 20191201, Letnik:
475
Journal Article
Recenzirano
Fluorochrome selection is a key step in designing multi-color antibody panels. The list of available fluorochromes is continuously growing, fitting current needs in clinical flow cytometry to ...simultaneously use more markers to better define multiple leukocyte subpopulations in a single tube. Several criteria guide fluorochrome selection: i) the fluorescence profiles (excitation and emission), ii) relative brightness, iii) fluorescence overlap, iv) fluorochrome stability, and v) reproducible conjugation to antibodies. Here we used 75 samples (45 bone marrow and 30 blood) to illustrate EuroFlow strategies for evaluation of compatible fluorochromes, and how the results obtained guide fluorochrome selection as a critical step in the antibody-panel building process. Our results allowed identification of optimal fluorescence profiles (e.g. higher fluorescence intensity and/or resolution with limited fluorescence overlap into neighbor channels) for brilliant violet (BV)421 and BV510 in the violet laser and allophycocyanin (APC) hilite 7 (H7) or APC C750 in the red laser vs. other candidate fluorochromes generally applied for the same detectors and here evaluated. Moreover, evaluation of the same characteristics for another group of fluorochromes (e.g. BV605, BV650, PE CF594, AF700 or APC AF700) guided selection of the most appropriate fluorochrome conjugates to be combined in a multi-color antibody panel. Albeit this is a demanding approach, it could be successfully applied for selection of fluorochrome combinations for the EuroFlow antibody panels for diagnosis, classification and monitoring of hematological malignancies and primary immunodeficiencies. Consequently, sets of 8-, 10- and 12-color fluorochrome combinations are proposed as frame of reference for initial antibody panel design.
Background Radiofrequency ablation (RFA) is safe and effective for eradicating Barrett's esophagus (BE) and BE-associated early neoplasia. Most RFA studies have limited the baseline length of BE (<10 ...cm), and therefore little is known about RFA for longer BE. Objective To assess the safety and efficacy of RFA with or without prior endoscopic resection (ER) for BE ≥10 cm containing neoplasia. Design Prospective trial. Setting Two tertiary-care centers. Patients This study involved consecutive patients with BE ≥10 cm with early neoplasia. Intervention Focal ER for visible abnormalities, followed by a maximum of 2 circumferential and 3 focal RFA procedures every 2 to 3 months until complete remission. Main Outcome Measurements Complete remission, defined as endoscopic resolution of BE and no intestinal metaplasia (CR-IM) or neoplasia (CR-neoplasia) in biopsy specimens. Results Of the 26 patients included, 18 underwent ER for visible abnormalities before RFA. The ER specimens showed early cancer in 11, high-grade intraepithelial neoplasia (HGIN) in 6, and low-grade intraepithelial neoplasia (LGIN) in 1. The worst residual histology, before RFA and after any ER, was HGIN in 16 patients and LGIN in 10 patients. CR-neoplasia and CR-IM were achieved in 83% (95% confidence interval CI, 63%-95%) and 79% (95% CI, 58%-93%), respectively. None of the patients had fatal or severe complications and 15% (95% CI, 4%-35%) had moderate complications. During a mean (± standard deviation) follow-up of 29 (± 9.1) months, no neoplasia recurred. Limitations Tertiary-care center, short follow-up. Conclusion ER for visible abnormalities, followed by RFA of residual BE is a safe and effective treatment for BE ≥10 cm containing neoplasia, with a low chance of recurrence of neoplasia or BE during follow-up.
Three developmental stages of the Mediterranean fruit fly,
Ceratitis capitata
(Wiedemann) (Diptera: Tephritidae), can be found in the soil (late third instar larvae, pupae, and teneral adults). These ...stages are susceptible to predation by generalist ground-dwelling predators. Our objectives have been to investigate the association between ground cover management (bare soil, a seeded cover of
Festuca arundinacea
and a mulch of straw), the emergence success of
C. capitata
, and the activity density of the most important groups of ground-dwelling predators (spiders, beetles, ants and earwigs). As expected,
C. capitata
emergence was lower in a seeded cover of
Festuca arundinacea
(FA) and a mulch of straw (M) (10.2%) relative to bare soil (BS) (13.2%). This was related to higher diversity and activity density of ground-dwelling predators in FA and M compared to BS. The contribution of the ground-dwelling predators considered in this study to this reduction highlights the key role of beetles, earwigs and, to a lesser extent, ants, while the contribution of the most abundant group of ground-dwelling predators, spiders, remains unclear. Ground covers appear as a strong and sustainable conservation biological control method that should be taken into consideration for the management of
C. capitata
populations.
We introduce a general concept for the design of all-optical wavelength converters with pulse reformatting functionality. The novel wavelength converters are based on a single semiconductor optical ...amplifier followed by an optical filter. A microelectromechanical system-based realization is shown and simultaneous 40 Gb/s wavelength conversion, switching and signal format conversion is demonstrated. The new pulse reformatting optical filter device outperforms current schemes with respect to input-power requirements, input-power dynamic range and signal quality.
Although NOD-SCID IL2Rγ
(NSG) xenograft mice are currently the most frequently used model to study human leukemia in vivo, the absence of a human niche severely hampers faithful recapitulation of the ...disease. We used NSG mice in which ceramic scaffolds seeded with human mesenchymal stromal cells were implanted to generate a human bone marrow (huBM-sc)-like niche. We observed that, in contrast to the murine bone marrow (mBM) niche, the expression of BCR-ABL or MLL-AF9 was sufficient to induce both primary acute myeloid leukemia (AML) and acute lymphocytic leukemia (ALL). Stemness was preserved within the human niches as demonstrated by serial transplantation assays. Efficient engraftment of AML MLL-AF9 and blast-crisis chronic myeloid leukemia patient cells was also observed, whereby the immature blast-like phenotype was maintained in the huBM-sc niche but to a much lesser extent in mBM niches. We compared transcriptomes of leukemias derived from mBM niches versus leukemias from huBM-like scaffold-based niches, which revealed striking differences in the expression of genes associated with hypoxia, mitochondria and metabolism. Finally, we utilized the huBM-sc MLL-AF9 B-ALL model to evaluate the efficacy of the I-BET151 inhibitor in vivo. In conclusion, we have established human niche models in which the myeloid and lymphoid features of BCR-ABL
and MLL-AF9
leukemias can be studied in detail.
Pebble beds have been employed in thermal storage and energy systems, where they are typically used to promote heat exchange in high-temperature environments. Understanding the heat conduction of the ...entire pebble bed could aid in the material selection of the pebbles themselves and structural components, system design, and safety monitoring. However, the thermal conductivity of pebble beds can change significantly near geometric boundaries. Using a complex multilayer analytical model in conjunction with a line source probe, we found a substantial increase in the thermal conductivity of a sintered bauxite pebble bed in the near-wall region (7.6 W m–1 K–1) compared to the bulk (0.59 W m–1 K–1). We investigated this difference by comparing porosity results, acquired with micro-CT, of 33.18 and 33.31% at approximately one pebble width surrounding the probe (near-wall) and the bulk of the pebble bed, suggesting that the thermal conductivity is largely altered by thermal contact resistance in the near-wall regime.
Dispersal can be an essential factor affecting the biological control of pests. Tetranychus urticae Koch (Acari: Tetranychidae) is a cosmopolitan and polyphagous species that may reach the pest ...status in many cropping systems including clementine orchards, where it may be found both in the trees and the associated flora. In a previous study, we demonstrated that the use of a ground cover of Festuca arundinacea Schreber (Poaceae) offered a better regulation of T. urticae populations than traditional alternatives (bare soil, multifloral wild cover). Therefore, we decided to study the ambulatory dispersal of mites crawling up and down tree trunks in a clementine mandarin orchard grown in association with a F. arundinacea cover for one season. The highest ambulatory migration rate was upward from the cover to the canopy. Multivariate regressions showed that the dynamics of T. urticae populations in the trees was strongly related to that of Phytoseiidae mites, their main natural predators. Surprisingly, canopy populations were not related to those on the ground cover or to those dispersing from it. When T. urticae individuals collected from the ground cover, the tree trunk, and the canopy were subjected to molecular analyses, the optimal number of genetic clusters (demes) was two. One cluster grouped individuals dispersed from the ground cover (e.g. collected on tree trunks) and 27.5 % of individuals collected in the ground cover. The second cluster grouped all the individuals collected from trees and 72.5 % of those collected in the cover. Interestingly, none of the individuals collected from the tree canopies was grouped with the first deme. This result may be taken as indicative that grass-adapted T. urticae individuals are unable to satisfactorily colonize and establish on the trees and provides evidence that host adaptation can hamper dispersal and establishment of the ground cover deme on trees, contributing to a better natural regulation of this pest species in citrus.
All-optical XOR functionality has been demonstrated experimentally using an integrated SOA-based Mach-Zehnder interferometer (SOA-MZI) at 20 and 40 Gb/s. The performance of the XOR results has been ...analyzed by solving the rate equation of the SOA numerically. The high-speed operation is limited by the carrier lifetime in the SOA. In order to solve the limitations imposed by carrier lifetime, a differential scheme for XOR operation has been experimentally investigated. This scheme is potentially capable of XOR operation to >100 Gb/s.
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