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Conformational effects of a cancer-linked mutation in pri-miR-30c RNA.
No full textMicroRNAs (miRNAs) are small, noncoding RNAs that mediate post-transcriptional downregulation of specific target genes. These transcripts are the products of a two-step processing pathway; primary miRNAs (pri-miRNAs) are processed by Drosha into individual precursor miRNA (pre-miRNA) hairpins, which are subsequently processed by Dicer into mature miRNAs. Single nucleotide polymorphisms (SNPs) that occur in pri-miRNAs, pre-miRNAs and mature miRNAs have been shown to affect the processing of specific target genes by modulating Drosha and Dicer processing or interactions with RNA binding proteins (RBPs). Using NMR and single-molecule optical tweezer experiments, we have investigated the conformational effects of a cancer-linked G/A mutation in the terminal loop of pri-miR-30c RNA, and how this influences binding by the SRSF3 and hnRNP A1 RBPs, which are implicated in its processing. Our results reveal that the wildtype and G/A variant pri-miR-30c RNAs adopt very similar elongated stem-loop structures, both of which are bound by SRSF3. However, while both wildtype and G/A pri-miR-30c RNAs can form dimeric kissing hairpin structures, the G to A mutation results in partial destabilization of the dimer in the variant transcript. This promotes recognition and binding by hnRNP A1, an RBP that enhances pri-miR-30c processing. Our data provide structural insight into the conformational effects of a G/A mutation in pri-miR-30c RNA and how this could affect processing and promote cancer
Identification of ocular regulatory functions of core histone variant H3.2.
No full textThe posttranscriptional modifications (PTM) of the Histone H3 family play an important role in ocular system differentiation. However, there has been no study on the nature of specific Histone H3 subtype carrying these modifications. Fortuitously, we had previously identified a dominant small-eye mutant Aey69 mouse with a mutation in the H3.2 encoding Hist2h3c1 gene (Vetrivel et al., 2019). In continuation, in the present study, the role of Histone H3.2 with relation to the microphtalmic Aey69 has been elaborated. Foremost, a transgenic mouse line expressing the fusion protein H3.2-GFP was generated using Crispr/Cas9. The approach was intended to confer a unique tag to the Hist2h3c1 gene which is similar in sequence and encoded protein structure to other histones. The GFP tag was then used for ChIP Seq analysis of the genes regulated by H3.2. The approach revealed ocular specific H3.2 targets including Ephrin family genes. Altered enrichment of H3.2 was found in the mutant Aey69 mouse, specifically around the ligand Efna5 and the receptor Ephb2. The effect of this altered enrichment on Ephrin signaling was further analyzed by QPCR and immunohistochemistry. This study identifies Hist2h3c1 encoded H3.2 as an important epigenetic player in ocular development. By binding to specific regions of ocular developmental factors Histone H3.2 facilitates the function of these genes for successful early ocular development
Providing metabolomics education and training: Pedagogy and considerations.
No full textBACKGROUND: Metabolomics is a highly multidisciplinary and non-standardised research field. Metabolomics researchers must possess and apply extensive cross-disciplinary content knowledge, subjective experience-based judgement, and the associated diverse skill sets. Accordingly, appropriate educational and training initiatives are important in developing this knowledge and skills base in the metabolomics community. For these initiatives to be successful, they must consider both pedagogical best practice and metabolomics-specific contextual challenges. AIM OF REVIEW: The aim of this review is to provide consolidated pedagogical guidance for educators and trainers in metabolomics educational and training programmes. KEY SCIENTIFIC CONCEPTS OF REVIEW: In this review, we discuss the principles of pedagogical best practice as they relate to metabolomics. We then discuss the challenges and considerations in developing and delivering education and training in metabolomics. Finally, we present examples from our own teaching practice to illustrate how pedagogical best practice can be integrated into metabolomics education and training programmes
Reflections on the future developments of research in retrospective physical dosimetry.
No full textElectron paramagnetic resonance, thermoluminescence, and optically stimulated luminescence, with biological tissues and inert materials are well established physical methods for retrospective dosimetry in acute accidental exposures. The objective of this article is to provide a view of the questions still open, the current challenges and the needed solutions. As research on emergency response methods is encountering increasing difficulties in terms of financial and human resources in many countries, it is essential to identify the research priorities and pay attention to cost-effective research paths. The intention of the paper is to stimulate discussion in the scientific community and to encourage collaboration among laboratories toward goals that address the real needs in retrospective dosimetry for acute exposures
The anorexigenic peptide neurotensin relates to insulin sensitivity in obese patients after BPD or RYGB metabolic surgery.
No full textNeurotensin is a peptide with effects on appetite and intestinal lipid absorption. Experimental data suggest a role in glucose homeostasis, while human data is missing. Here, 20 morbidly obese subjects either underwent biliopancreatic diversion with duodenal switch (BPD), or Roux-en-Y gastric bypass (RYGB) in a randomized fashion. Before and 1 year after surgery, anthropometric data, body composition, clinical biochemistry, insulin sensitivity by means of euglycemic hyperinsulinemic clamps (HEC) and fasting plasma proneurotensin 1-117 were analyzed. Plasma proneurotensin increased significantly more 1 year after BDP than RYGB (P = 0.028), while weight loss was comparable. After metabolic surgery, proneurotensin correlated positively with insulin sensitivity (M-value) (r = 0.55, P < 0.001), while an inverse relationship with fasting glucose, HOMA-IR and HbA1c was observed (P < 0.05 for all components). After adjustment for age and gender, proneurotensin and BMI remained independently related with delta of M-value (beta = 0.46 and beta = 0.51, P < 0.05, resp.). From these data we conclude that proneurotensin positively correlates with insulin sensitivity uniquely after weight loss induced by metabolic surgery in humans. BDP leads to a stronger increase in the anorexigenic peptide compared to RYGB
Simultaneous on-line vacuum single- and multi-photon ionization on an orthogonal acceleration time-of-flight mass spectrometer platform.
No full textThe development of orthogonal acceleration time-of-flight (oa-tof) technology is driven forward due to higher mass accuracy and resolving power than conventional linear/reflectron tof instruments. This is achieved with a more accurate definition of starting energies and coordinates of ions by spatial separation of ion generation and orthogonal ion extraction. Consequently, the ability to cover the whole mass spectral range without scanning is not given anymore. Therefore, continuous ion sources are favored for ensuring high duty cycles and thus high temporal resolution. For pulsed ion sources, high repetition rates are mandatory for covering large m/z ranges without losing their high temporal resolution. We have combined an oa-tof with deuterium lamp single-photon ionization (SPI) as a continuous ion source together with a pulsed 2000 Hz excimer (KrF) laser for resonance enhanced multi-photon ionization (REMPI). These two ionization techniques can be used simultaneously. To the best of our knowledge, this system is the first of its kind in combining a vacuum pulsed ionization source with an oa-tof instrument without any other ion storage hardware. The combination of a soft broadband ionization for organics (SPI) in combination with a very sensitive and selective soft ionization (REMPI) can be used for covering the whole mass range or in targeted on-line monitoring cases one or several smaller mass ranges. To demonstrate the simultaneous SPI/REMPI-oa-tof technique, two applicative areas are explored: on-line monitoring of coffee roast gas emissions and e-cigarette vapor. The complementary information from SPI and REMPI signals are combined in a way to exploit the advantages of both ionization types. In a further development step, a second data acquisition card is built into the system. This modification allows the independent storage of data from both ionization methods without mixing. For demonstration, a third example with a GC measurement is provided. The last example shows the possibility of modified sensitivities for different mass regions in REMPI data acquisition without affecting the SPI channel. The newly developed system shows high robustness in terms of measurements in real industrial environments. The simultaneous measurement technique provides a higher density of information in a single measurement, saving time and resources
The changes in <em>Lemna minor</em> metabolomic profile: A response to diclofenac incubation.
No full textMetabolomics is an emerging approach that investigates the changes in the metabolome profile. In the present study, Lemna minor —considered as an experimental aquatic plant model— was incubated with 10 and 100 μM diclofenac (DCF) for 96 h, respectively. Knowing that DCF is internationally often problematic in wastewater effluents and that it might affect particularly the metabolic profiles in aquatic plants, mainly the oxidoreductase, dehydrogenase, peroxidase, and glutathione reductase activities, here it was hypothesized (H) that in the common duckweed, DCF might increase the phenolic and flavonoids pathways, as an antioxidant response to this stress (H1). Also, it was expected DCF to alternate the physiological characteristics, especially the molecular interaction and biochemical properties, of Lemna (H2). Metabolic changes were investigated with target and untargeted screening analysis using RPLC-HILIC-ESI-TOF-MS. Twelve amino acids were identified in all treatments, together with three organic acids (p-coumaric, cinnamic, and sinapic acids). In untargeted screening, the important metabolites to discriminate between different treatments were assigned to Lemna such as organic acids, lignin, sugars, amino acids, dipeptides, flavonoids, biflavonoids, fatty acids, among others. In resume, Lemna responded to both DCF concentrations, showing different stress patterns. A similar metabolic response had already been identified in other studies in exposing Lemna to other anthropogenic stressors (like pesticides)
The influence of nuclear models and Monte Carlo radiation transport codes on stray neutron dose estimations in proton therapy.
No full textPurpose: This study investigates the influence of several Monte Carlo radiation transport codes and nuclear models on the simulation of secondary neutron spectra and its impact on calculating and measuring the neutron doses in proton therapy. Materials and methods: Three different multi-purpose Monte Carlo radiation transport codes (FLUKA, MCNPX, Geant4) were used together with different available nuclear models, to calculate secondary neutron energy spectra at various points inside a water tank phantom with PMMA walls using a 10 × 10 cm2 rectangular, mono-energetic proton beam (110 MeV, 150 MeV, 180 MeV, 210 MeV). Using Kerma approximation secondary neutron doses were calculated applying fluence-to-dose equivalent conversion coefficients in water. Moreover, the impact of varying spectra for electrochemically etched CR39 detector calibration was analyzed for different codes and models. Results: In distal positions beyond the Bragg peak, results show largest variations between the codes, which was up to 53% for the high energy neutron fluence at 16 cm from the Bragg peak of the 110 MeV proton beam. In lateral positions, the variation between the codes is smaller and for the total neutron fluence within 20%. Variation in the nuclear models in MCNPX was only visible for the proton beam energies of 180 and 210 MeV and modeling the high energy neutron fluence which reached up to 23% for 210 MeV at 11 cm lateral from the beam axis. Impact on neutron dose equivalent was limited for the different models used (<8%) while it was pronounced for the different codes (45% at 16 cm from the Bragg peak of the 110 MeV proton beam). CR39 calibration factors in lateral positions were on average varying 10% between codes and 5% between nuclear models. Conclusions: This study demonstrated a large impact on the neutron fluence spectra calculated by different codes while the impact of different models in MCNPX proved to be less prominent for the neutron modeling in proton therapy
Low-energy X-ray intraoperative radiation therapy (Lex-IORT) for resected brain metastases: A single-institution experience.
No full textBACKGROUND: Resection followed by local radiation therapy (RT) is the standard of care for symptomatic brain metastases. However, the optimal technique, fractionation scheme and dose are still being debated. Lately, low-energy X-ray intraoperative RT (lex-IORT) has been of increasing interest. METHOD: Eighteen consecutive patients undergoing BM resection followed by immediate lex-IORT with 16-30 Gy applied to the spherical applicator were retrospectively analyzed. Demographic, RT-specific, radiographic and clinical data were reviewed to evaluate the effectiveness and safety of IORT for BM. Descriptive statistics and Kaplan-Meyer analysis were applied. RESULTS: The mean follow-up time was 10.8 months (range, 0-39 months). The estimated local control (LC), distant brain control (DBC) and overall survival (OS) at 12 months post IORT were 92.9% (95%-CI 79.3-100%), 71.4% (95%-CI 50.2-92.6%) and 58.0% (95%-CI 34.1-81.9%), respectively. Two patients developed radiation necrosis (11.1%) and wound infection (CTCAE grade III); both had additional adjuvant treatment after IORT. For five patients (27.8%), the time to the start or continuation of systemic treatment was ≤15 days and hence shorter than wound healing and adjuvant RT would have required. CONCLUSION: In accordance with previous series, this study demonstrates the effectiveness and safety of IORT in the management of brain metastases despite the small cohort and the retrospective characteristic of this analysis
