We report a customized gene panel assay based on multiplex long-PCR followed by third generation sequencing on nanopore technology (MinION), designed to analyze five frequently mutated genes in ...chronic lymphocytic leukemia (CLL): TP53, NOTCH1, BIRC3, SF3B1 and MYD88. For this purpose, 12 patients were selected according to specific cytogenetic and molecular features significantly associated with their mutational status. In addition, simultaneous analysis of the targets genes was performed by molecular assays or Sanger Sequencing. Data analysis included mapping to the GRCh37 human reference genome, variant calling and annotation, and average sequencing depth/error rate analysis. The sequencing depth resulted on average higher for smaller amplicons, and the final breadth of coverage of the panel was 94.1%. The error rate was about 6% and 2% for insertions/deletions and single nucleotide variants, respectively. Our gene panel allows analysis of the prognostically relevant genes in CLL, with two PCRs per patient. This strategy offers an easy and affordable workflow, although further advances are required to improve the accuracy of the technology and its use in the clinical field. Nevertheless, the rapid and constant development of nanopore technology, in terms of chemistry advances, more accurate basecallers and analysis software, offers promise for a wide use of MinION in the future.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
The breakpoint cluster region–abelson 1 p190 fusion transcript is the most frequent variant observed in Philadelphia-positive (Ph+) acute lymphoblastic leukemia (ALL). Qualitative-PCR and real-time ...quantitative PCR are the currently used methods to monitor minimal residual disease (MRD) in Ph+ ALL patients; for the latter, full standardization and an international quality validation are lacking. Here, we developed a droplet digital PCR (ddPCR) assay for MRD monitoring in p190+ ALL cases. The analytical performance was assessed by the limit-of-detection determination, showing a reliability, sensitivity, and precision of the assay of up to 0.001%. Comparison of results obtained with qualitative PCR and ddPCR in 117 follow-up samples from 16 of 26 Ph+ ALL patients showed discordant results in 27% of cases (32 of 117). Real-time quantitative PCR analysis of 19 ddPCR-positive samples with a low tumor burden failed to provide quantitative results in 63% of cases (12 of 19). These results highlight that in p190+ ALL the ddPCR method has a sufficient analytical performance for very low MRD monitoring and for predicting molecular relapse several months before hematologic relapse. In conclusion, MRD monitoring by ddPCR may better stratify Ph+ ALL patients at risk of disease progression.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Sulfuric acid is a key contributor to new particle formation, though measurements of its gaseous concentrations are difficult to make. Several parameterizations to estimate sulfuric acid exist, all ...of which were constructed using measurements from the Northern Hemisphere. In this work, we report the first measurements of sulfuric acid from the Amazon basin. These measurements are consistent with concentrations measured in Hyytiälä, Finland, though, unlike Hyytiälä, there is no clear correlation of sulfuric acid with global radiation. There was a minimal difference in sulfuric acid observed between the wet and dry seasons in the Amazon basin. We also test the efficacy of existing proxies to estimate sulfuric acid in this region. Our results suggest that nighttime sulfuric acid production is due to both a stabilized Criegee intermediate pathway and oxidation of SO2 by OH, the latter of which is not currently accounted for in existing proxies. These results also illustrate the drawbacks of the common substitution of radiation for OH concentrations. None of the tested proxies effectively estimate sulfuric acid measurements at night. For estimates at all times of day, a recently published proxy based on data from the boreal forest should be used. If only daytime estimates are needed, several recent proxies that do not include the Criegee pathway are sufficient. More investigation of nighttime sulfuric acid production pathways is necessary to close the gap between measurements and estimates with existing proxies.
ATPase (SERCA2a) pumps. Furthermore, we show that NO influences cardiac response to preload through S-nitrosylation of phospholamban
and consequent activation of SERCA2a. This suggests that in the ...fish heart NO modulates the Frank–Starling response through
a beat-to-beat regulation of calcium reuptake and thus of myocardial relaxation., we previously showed that an endogenous NO release affects the Frank–Starling response making the heart more sensitive to
preload. Using the same bioassay, we now demonstrate that this effect is confirmed in the presence of the exogenous NO donor, We propose that this mechanism represents an important evolutionary step for the stretch-induced intrinsic regulation of the
vertebrate heart, providing, at the same time, a stimulus for mammalian-oriented studies., Anguilla anguilla, -acetyl penicillamine, is independent from endocardial endothelium and guanylate cyclase/cGMP/protein kinase G and cAMP/protein
kinase A pathways, involves a PI(3)kinase-mediated activation of endothelial NO synthase and a modulation of the SR-CA, In the, in vitro, The Frank–Starling mechanism is a fundamental property of the vertebrate heart, which allows the myocardium to respond to
increased filling pressure with a more vigorous contraction of its lengthened fibres. In mammals, myocardial stretch increases
cardiac nitric oxide (NO) release from both vascular endothelium and cardiomyocytes. This facilitates myocardial relaxation
and ventricular diastolic distensibility, thus influencing the Frank–Starling mechanism., working heart of the eel
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
•The goldfish is a well known champion of hypoxia and anoxia tolerance.•Hypoxic goldfish hearts show an increased basal performance.•Acute hypoxia enhances the sensitivity of the goldfish heart to ...the Starling response.•Intracardiac NO/NOS remarkably enhances the goldfish heart performance.
Goldfish tolerate prolonged and severe hypoxia, thus representing a well-suited model to study the maintenance of cardiac function when O2 availability represents a limiting factor.
Using a working heart preparation, we explored the role of the intracardiac nitric oxide synthase (NOS)-derived nitric oxide (NO) under normoxic and hypoxic conditions. Cardiac performance was examined both under basal (constant preload and afterload) and loading conditions, i.e. preload-induced increases in stroke volume (SV) and hence cardiac output at constant afterload (the Frank–Starling response).
Hypoxic hearts showed an increased basal mechanical performance compared to the normoxic counterpart. Under basal conditions, in both normoxic and hypoxic hearts, NOS and soluble guanylyl cyclase (sGC) inhibition increased SV, while exogenous NO supply decreased it.
The normoxic heart was very sensitive to filling pressure increases; the maximum SV = 1.08 ± 0.09 mL/kg body mass was obtained at 0.4 kPa. Acute hypoxia increased this sensitivity, SV reaching the maximum value (1.45 ± 0.12 mL/kg body mass) at 0.25 kPa. NOS inhibition by L-NMMA reduced the Frank–Starling response under normoxia, but was ineffective under acute hypoxia, where NO may come from nitrite reduction. In both conditions, sGC inhibition induced a reduction of the cardiac response to preload. Moreover, under acute hypoxia, NO scavenging significantly reduced the Frank–Starling response. The hypoxia-induced hemodynamic patterns were complemented by Western blotting analysis which revealed increased expressions of NOS and hypoxia inducible factor α(HIF-1α). In conclusion, we demonstrated that intracardiac NO/NOS enhances goldfish heart performance, remarkably expanding its hypoxic tolerance.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
Diastrophic dysplasia (DTD) is a recessive chondrodysplasia caused by pathogenic variants in the SLC26A2 gene encoding for a cell membrane sulfate/chloride antiporter crucial for sulfate uptake and ...glycosaminoglycan (GAG) sulfation. Research on a DTD animal model has suggested possible pharmacological treatment approaches. In view of future clinical trials, the identification of non-invasive biomarkers is crucial to assess the efficacy of treatments. Urinary GAG composition has been analyzed in several metabolic disorders including mucopolysaccharidoses. Moreover, the N-terminal fragment of collagen X, known as collagen X marker (CXM), is considered a real-time marker of endochondral ossification and growth velocity and was studied in individuals with achondroplasia and osteogenesis imperfecta. In this work, urinary GAG sulfation and blood CXM levels were investigated as potential biomarkers for individuals affected by DTD. Chondroitin sulfate disaccharide analysis was performed on GAGs isolated from urine by HPLC after GAG digestion with chondroitinase ABC and ACII, while CXM was assessed in dried blood spots. Results from DTD patients were compared with an age-matched control population. Undersulfation of urinary GAGs was observed in DTD patients with some relationship to the clinical severity and underlying SLC26A2 variants. Lower than normal CXM levels were observed in most patients, even if the marker did not show a clear pattern in our small patient cohort because CXM values are highly dependent on age, gender and growth velocity. In summary, both non-invasive biomarkers are promising assays targeting various aspects of the disorder including overall metabolism of sulfated GAGs and endochondral ossification.
•Urinary GAG sulfation and blood collagen X marker (CXM) have been tested as non-invasive biomarkers in DTD patients.•Urinary GAG sulfation is reduced in patients with DTD and is a promising biomarker for therapies that increase GAG sulfation.•Blood CXM is below age-matched control values in DTD patients being a suitable biomarker for bone growth velocity.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
PURPOSE: Sub-Tenon anesthesia is a form of local anesthetic techniques used in ophthalmic procedures, especially in cataract surgery. Few studies in our environment have reported the effects of ...sub-Tenon anesthesia on akinesia and analgesia as well as optimum volumes of anesthetic agents that are required for effective analgesia and akinesia. The objective of this study is to determine and compare the level of analgesia and akinesia and the effect of ocular compressions after sub-Tenon injection of 2 mL versus 3 mL of xylocaine among patients booked for cataract surgery in the University of Ilorin Teaching Hospital (UITH). METHODS: A cross-sectional comparative study among cataract patients booked for surgery in the Department of Ophthalmology at UITH was carried out from March 2017 to August 2017. A total of 200 cataract patients that met the inclusion criteria were recruited into the study over 6 months. Lists of cataract patients booked for surgery formed the sampling frame. Questionnaires were administered to obtain information on sociodemographic, ocular symptoms, and other histories. Visual acuity was checked, and ocular examination was done with a pen-torch, ophthalmoscope, and slit-lamp examination with 78D. Tonometry and optic nerve assessments were done to exclude any patient with preexisting glaucoma or suspicious disc. About 200 patients were randomly allocated into one of two volume groups (Group 1 had 2 mL sub-Tenon xylocaine injection whereas Group 2 had 3 mL); level of analgesia and akinesia was checked 12 min and 15 min, respectively. RESULTS: The age range was 20–107 years; mean age for Group 1 was 63.8 ± 12.64, whereas the mean age of Group 2 was 64.14 ± 14.14. There was a slight female preponderance given a M:f of 1:1.4 and more than half of the patients presented with right eye (RE) cataract. Levels of analgesia 12 min after sub-Tenon injections in both groups were similar. However, two patients were found to have severe pain in Group 1, and no patients had severe pain in Group 2. The level of akinesia which was assessed 15 min after injection in both groups was found to be significantly better in Group 2. CONCLUSION: There was a slight female preponderance with about two-third of the total patients being female and the percentage of females who had no pain was more than the male, more than 50% of the patients had RE cataracts. Sub-Tenon anesthesia whether with 2 mL or 3 mL led to a statistically significant rise in intraorbital pressure (IOP); however, with ocular compression, the IOP was found to decrease below the preinjection pressure in both groups. Levels of analgesia were similar in the two groups, whereas akinesia was statistically better in the group that had 3 mL injection.
Using morphological and physiological approaches we provided, for the first time, a structural and functional characterization of Carassius auratus L. heart. Besides to the classical four chambers, ...i.e. sinus venosus, atrium, ventricle, bulbus, we described two distinct structures corresponding to the atrio-ventricular (AV) region and the conus arteriosus. The atrium is very large and highly trabeculated; the ventricle shows an outer compacta, vascularized by coronary vessels, and an inner spongiosa; the bulbus wall is characterized by a high elastin/collagen ratio, which makes it extremely compliant. Immunolocalization revealed a strong expression of activated “eNOS-like” isoforms both at coronary endothelium and, to a lesser extent, in the myocardiocytes and the endocardial endothelium (EE). The structural design of the heart appears to comply with its mechanical function. Using an in vitro working heart preparation, cardiac performance was evaluated at different filling and afterload pressures. The hearts were very sensitive to filling pressure increases. Maximum Stroke volume (SV=1.08±0.09mL/kg body mass) was obtained with an input pressure of 0.4kPa. The heart was not able to sustain afterload increases, values higher than 1.5kPa impairing its performance. These morpho-functional features are consistent with a volume pump mechanical performance.
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IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, SAZU, SBCE, UL, UM, UPCLJ, UPUK
As part of the Observations and Modeling of the Green Ocean Amazon (GoAmazon 2014/5) Experiment, detailed aerosol and trace gas measurements were conducted near Manaus, a metropolis located in the ...central Amazon Basin. Measurements of aerosol particles and trace gases were done downwind Manaus at the sites T2 (Tiwa Hotel) and T3 (Manacapuru), at a distance of 8 and 70 km from Manaus, respectively. Based on in-plume measurements closer to Manaus (site T2), the chemical signatures of city emissions were used to improve the interpretation of pollutant levels at the T3 site. We derived chemical and physical properties for the city's atmospheric emission ensemble, taking into account only air masses impacted by the Manaus plume at both sites, during the wet and dry season Intensive Operating Periods (IOPs). At T2, average concentrations of aerosol number (CN), CO and SO2 were 5500 cm−3 (between 10 and 490 nm), 145 ppb and 0.60 ppb, respectively, with a typical ratio ΔCN/ΔCO of 60–130 particles cm−3 ppb−1. The aerosol scattering (at RH < 60%) and absorption at 637 nm at T2 ranged from 10 to 50 M m−1 and 5–10 M m−1, respectively, leading to a mean single scattering albedo (SSA) of 0.70. In addition to identifying periods dominated by Manaus emissions at both T2 and T3, the plume transport between the two sampling sites was studied using back trajectory calculations. Results show that the presence of the Manaus plume at site T3 was important mainly during the daytime and at the end of the afternoons. During time periods directly impacted by Manaus emissions, an average aerosol number concentration of 3200 cm−3 was measured at T3. Analysis of plume evolution between T2 and T3 indicates a transport time of 4–5 h. Changes of submicron organic and sulfate aerosols ratios relative to CO (ΔOA/ΔCO and ΔSO4/ΔCO, respectively) indicate significant production of secondary organic aerosol (SOA), corresponding to a 40% mass increase in OA and a 30% in SO4 mass concentration. Similarly, during air mass arrival at T3 the SSA increased to 0.83 from 0.70 at T2, mainly associated with an increase in organic aerosol concentration. Aerosol particle size distributions show a strong decrease in the Aitken nuclei mode (10–100 nm) during the transport from T2 to T3, in particular above 30 nm, as a result of efficient coagulation processes into larger particles. A decrease of 30% in the particle number concentration and an increase of about 50 nm in geometric mean diameter were observed from T2 to T3 sites. The study of the evolution of aerosol properties downwind of the city of Manaus improves our understanding of how coupling of anthropogenic and biogenic sources may be impacting the sensitive Amazonian atmosphere.
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•Manaus emissions significantly impacts Amazonian aerosols properties.•Organic aerosols increase 40% in mass downwind of Manaus, attributed to secondary formation.•Sulfate aerosols increase 30% in mass downwind of Manaus, attributed to SO2 oxidation.•Aerosol mean diameter increases about 60 nm during plume evolution, in the dry season.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
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