Teacher to Teacher: Mentoring Preservice Teachers into the Profession

https://scholarworks.moreheadstate.edu/student_scholarship_posters/1003/thumbnail.jp

Direct monitoring of single-cell response to biomaterials by Raman spectroscopy

There is continued focus on the development of new biomaterials and associated biological testing methods needed to reduce the time taken for their entry to clinical use. The application of Raman spectroscopy to the study of individual cells that have been in contact with biomaterials offers enhanced in vitro information in a potentially non-destructive testing regime. The work presented here reports the Raman spectral analysis of discreet U-2 OS bone cells after exposure to hydroxyapatite (HA) coated titanium (Ti) substrates in both the as-deposited and thermally annealed states. These data show that cells that were in contact with the bioactive HA surface for 7 days had spectral markers similar to those cultured on the Ti substrate control for the same period. However, the spectral features for those cells that were in contact with the annealed HA surface had indicators of significant differentiation at day 21 while cells on the as-deposited surface did not show these Raman changes until day 28. The cells adhered to pristine Ti control surface showed no spectral changes at any of the timepoints studied. The validity of these spectroscopic results has been confirmed using data from standard in vitro cell viability, adhesion, and proliferation assays over the same 28-day culture period. In this case, cell maturation was evidenced by the formation of natural bone apatite, which precipitated intracellularly for cells exposed to both types of HA-coated Ti at 21 and 28 days, respectively. The properties of the intracellular apatite were markedly different from that of the synthetic HA used to coat the Ti substrate with an average particle size of 230 nm, a crystalline-like shape and Ca/P ratio of 1.63 ± 0.5 as determined by SEM-EDX analysis. By comparison, the synthetic HA particles used as a control had an average size of 372 nm and were more-rounded in shape with a Ca/P ratio of 0.8 by XPS analysis and 1.28 by SEM-EDX analysis. This study shows that Raman spectroscopy can be employed to monitor single U-2 OS cell response to biomaterials that promote cell maturation towards de novo bone thereby offering a label-free in vitro testing method that allows for non-destructive analyses. [Image: see text

Near-unity nuclear polarization with an open-source 129Xe hyperpolarizer for NMR and MRI

The exquisite NMR spectral sensitivity and negligible reactivity of hyperpolarized xenon-129 (HP129Xe) make it attractive for a number of magnetic resonance applications; moreover, HP129Xe embodies an alternative to rare and nonrenewable 3He. However, the ability to reliably and inexpensively produce large quantities of HP129Xe with sufficiently high 129Xe nuclear spin polarization (PXe) remains a significant challenge—particularly at high Xe densities. We present results from our “open-source” large-scale (∼1 L/h) 129Xe polarizer for clinical, preclinical, and materials NMR and MRI research. Automated and composed mostly of off-the-shelf components, this “hyperpolarizer” is designed to be readily implementable in other laboratories. The device runs with high resonant photon flux (up to 200 W at the Rb D1 line) in the xenon-rich regime (up to 1,800 torr Xe in 500 cc) in either single-batch or stopped-flow mode, negating in part the usual requirement of Xe cryocollection. Excellent agreement is observed among four independent methods used to measure spin polarization. In-cell PXe values of ∼90%, ∼57%, ∼50%, and ∼30% have been measured for Xe loadings of ∼300, ∼500, ∼760, and ∼1,570 torr, respectively. PXe values of ∼41% and ∼28% (with ∼760 and ∼1,545 torr Xe loadings) have been measured after transfer to Tedlar bags and transport to a clinical 3 T scanner for MR imaging, including demonstration of lung MRI with a healthy human subject. Long “in-bag” 129Xe polarization decay times have been measured (T1 ∼38 min and ∼5.9 h at ∼1.5 mT and 3 T, respectively)—more than sufficient for a variety of applications

Recurrent Tissue-Specific Mtdna Mutations are Common in Humans

Mitochondrial DNA (mtDNA) variation can affect phenotypic variation; therefore, knowing its distribution within and among individuals is of importance to understanding many human diseases. Intra-individual mtDNA variation (heteroplasmy) has been generally assumed to be random. We used massively parallel sequencing to assess heteroplasmy across ten tissues and demonstrate that in unrelated individuals there are tissue-specific, recurrent mutations. Certain tissues, notably kidney, liver and skeletal muscle, displayed the identical recurrent mutations that were undetectable in other tissues in the same individuals. Using RFLP analyses we validated one of the tissue-specific mutations in the two sequenced individuals and replicated the patterns in two additional individuals. These recurrent mutations all occur within or in very close proximity to sites that regulate mtDNA replication, strongly implying that these variations alter the replication dynamics of the mutated mtDNA genome. These recurrent variants are all independent of each other and do not occur in the mtDNA coding regions. The most parsimonious explanation of the data is that these frequently repeated mutations experience tissue-specific positive selection, probably through replication advantage

Comparative study of in situ N2 rotational Raman spectroscopy methods for probing energy thermalisation processes during spin-exchange optical pumping

Spin-exchange optical pumping (SEOP) has been widely used to produce enhancements in nuclear spin polarisation for hyperpolarised noble gases. However, some key fundamental physical processes underlying SEOP remain poorly understood, particularly in regards to how pump laser energy absorbed during SEOP is thermalised, distributed and dissipated. This study uses in situ ultra-low frequency Raman spectroscopy to probe rotational temperatures of nitrogen buffer gas during optical pumping under conditions of high resonant laser flux and binary Xe/N2 gas mixtures. We compare two methods of collecting the Raman scattering signal from the SEOP cell: a conventional orthogonal arrangement combining intrinsic spatial filtering with the utilisation of the internal baffles of the Raman spectrometer, eliminating probe laser light and Rayleigh scattering, versus a new in-line modular design that uses ultra-narrowband notch filters to remove such unwanted contributions. We report a ~23-fold improvement in detection sensitivity using the in-line module, which leads to faster data acquisition and more accurate real-time monitoring of energy transport processes during optical pumping. The utility of this approach is demonstrated via measurements of the local internal gas temperature (which can greatly exceed the externally measured temperature) as a function of incident laser power and position within the cell

XeNA: an automated ‘open-source’ 129Xe hyperpolarizer for clinical use

Here we provide a full report on the construction, components, and capabilities of our consortium’s “open-source” large-scale (~ 1 L/h) 129Xe hyperpolarizer for clinical, pre-clinical, and materials NMR/MRI (Nikolaou et al., Proc. Natl. Acad. Sci. USA, 110, 14150 (2013)). The ‘hyperpolarizer’ is automated and built mostly of off-the-shelf components; moreover, it is designed to be cost-effective and installed in both research laboratories and clinical settings with materials costing less than $125,000. The device runs in the xenon-rich regime (up to 1800 Torr Xe in 0.5 L) in either stopped-flow or single-batch mode—making cryo-collection of the hyperpolarized gas unnecessary for many applications. In-cell 129Xe nuclear spin polarization values of ~ 30%–90% have been measured for Xe loadings of ~ 300–1600 Torr. Typical 129Xe polarization build-up and T1 relaxation time constants were ~ 8.5 min and ~ 1.9 h respectively under our spin-exchange optical pumping conditions; such ratios, combined with near-unity Rb electron spin polarizations enabled by the high resonant laser power (up to ~ 200 W), permit such high PXe values to be achieved despite the high in-cell Xe densities. Importantly, most of the polarization is maintained during efficient HP gas transfer to other containers, and ultra-long 129Xe relaxation times (up to nearly 6 h) were observed in Tedlar bags following transport to a clinical 3 T scanner for MR spectroscopy and imaging as a prelude to in vivo experiments. The device has received FDA IND approval for a clinical study of chronic obstructive pulmonary disease subjects. The primary focus of this paper is on the technical/engineering development of the polarizer, with the explicit goals of facilitating the adaptation of design features and operative modes into other laboratories, and of spurring the further advancement of HP-gas MR applications in biomedicine

Analysis of the genetic phylogeny of multifocal prostate cancer identifies multiple independent clonal expansions in neoplastic and morphologically normal prostate tissue.

Genome-wide DNA sequencing was used to decrypt the phylogeny of multiple samples from distinct areas of cancer and morphologically normal tissue taken from the prostates of three men. Mutations were present at high levels in morphologically normal tissue distant from the cancer, reflecting clonal expansions, and the underlying mutational processes at work in morphologically normal tissue were also at work in cancer. Our observations demonstrate the existence of ongoing abnormal mutational processes, consistent with field effects, underlying carcinogenesis. This mechanism gives rise to extensive branching evolution and cancer clone mixing, as exemplified by the coexistence of multiple cancer lineages harboring distinct ERG fusions within a single cancer nodule. Subsets of mutations were shared either by morphologically normal and malignant tissues or between different ERG lineages, indicating earlier or separate clonal cell expansions. Our observations inform on the origin of multifocal disease and have implications for prostate cancer therapy in individual cases

Frailty-adjusted therapy in Transplant Non-Eligible patients with newly diagnosed Multiple Myeloma (FiTNEss (UK-MRA Myeloma XIV Trial)): a study protocol for a randomised phase III trial

INTRODUCTION: Multiple myeloma is a bone marrow cancer, which predominantly affects older people. The incidence is increasing in an ageing population.Over the last 10 years, patient outcomes have improved. However, this is less apparent in older, less fit patients, who are ineligible for stem cell transplant. Research is required in this patient group, taking into account frailty and aiming to improve: treatment tolerability, clinical outcomes and quality of life. METHODS AND ANALYSIS: Frailty-adjusted therapy in Transplant Non-Eligible patients with newly diagnosed Multiple Myeloma is a national, phase III, multicentre, randomised controlled trial comparing standard (reactive) and frailty-adjusted (adaptive) induction therapy delivery with ixazomib, lenalidomide and dexamethasone (IRD), and to compare maintenance lenalidomide to lenalidomide+ixazomib, in patients with newly diagnosed multiple myeloma not suitable for stem cell transplant. Overall, 740 participants will be registered into the trial to allow 720 and 478 to be randomised at induction and maintenance, respectively.All participants will receive IRD induction with the dosing strategy randomised (1:1) at trial entry. Patients randomised to the standard, reactive arm will commence at the full dose followed by toxicity dependent reactive modifications. Patients randomised to the adaptive arm will commence at a dose level determined by their International Myeloma Working Group frailty score. Following 12 cycles of induction treatment, participants alive and progression free will undergo a second (double-blind) randomisation on a 1:1 basis to maintenance treatment with lenalidomide+placebo versus lenalidomide+ixazomib until disease progression or intolerance. ETHICS AND DISSEMINATION: Ethical approval has been obtained from the North East-Tyne & Wear South Research Ethics Committee (19/NE/0125) and capacity and capability confirmed by local research and development departments for each participating centre prior to opening to recruitment. Participants are required to provide written informed consent prior to trial registration. Trial results will be disseminated by conference presentations and peer-reviewed publications