Investigation of the Effect of PKC Activation on In vitro Prostate Cell Metabolism using 13C NMR

PKC isozymes have been implicated in regulating everything from transformation and proliferation of prostate cancer cells to apoptosis. Many recent studies have implicated PKC-e, a novel PKC, in supporting cell survival and proliferation in addition to having an anti-apoptotic effect through interactions with BAX. PKC-d is another novel PKC that has been shown to promote apoptosis in LNCaP cells, and thus antagonizing the antiapoptotic effect of PKC-e. 13C-NMR and 13C(3)-aspartate supplemented media were utilized to examine the metabolism of LNCaP, DU-145 and BPH cell lines with and without activation of PKC by phorbol 12-myristate 13-acetate (PMA). Equivalent amounts of 13C-lactate were produced by the BPH cell line irrespective of addition of PMA (0.50±0.0.06 mM without PMA and 0.50±0.04 mM with PMA). The LNCaP cells produced significantly less 13C-lactate on PMA treatment from 0.40±0.04 mM to 0.25±0.10 mM, while the DU-145 cells nearly doubled the production of 13C-lactate on PMA treatment from 0.27±0.09 mM to 0.53±0.09 mM. Real-time PCR experiments showed the dramatic effect of PMA on cell metabolism could not be directly explained by the relative expression level of PKC-e and PKC-d mRNA as there was no statistically significant difference in levels of PKC-e and PKC-d mRNA. These results suggest an alternative explanation, such as 2nd messenger expression levels, need to be explored

Gaps and opportunities in refractory status epilepticus research in children: A multi-center approach by the Pediatric Status Epilepticus Research Group (pSERG)

PURPOSE: Status epilepticus (SE) is a life-threatening condition that can be refractory to initial treatment. Randomized controlled studies to guide treatment choices, especially beyond first-line drugs, are not available. This report summarizes the evidence that guides the management of refractory convulsive SE (RCSE) in children, defines gaps in our clinical knowledge and describes the development and works of the \u27pediatric Status Epilepticus Research Group\u27 (pSERG). METHODS: A literature review was performed to evaluate current gaps in the pediatric SE and RCSE literature. In person and online meetings helped to develop and expand the pSERG network. RESULTS: The care of pediatric RCSE is largely based on extrapolations of limited evidence derived from adult literature and supplemented with case reports and case series in children. No comparative effectiveness trials have been performed in the pediatric population. Gaps in knowledge include risk factors for SE, biomarkers of SE and RCSE, second- and third-line treatment options, and long-term outcome. CONCLUSION: The care of children with RCSE is based on limited evidence. In order to address these knowledge gaps, the multicenter pSERG was established to facilitate prospective collection, analysis, and sharing of de-identified data and biological specimens from children with RCSE. These data will allow identification of treatment strategies associated with better outcomes and delineate evidence-based interventions to improve the care of children with SE

Investigation of zinc in biological systems using X-ray absorption spectroscopy. I. EXAFS characterization of zinc-dependent alkyltransferase proteins. II. Exploring the role of zinc in the early development of <italic>Danio rerio</italic> embryos using X-ray fluorescence microprobe imaging and micro-xanes.

Zinc is essential for the growth, development and differentiation of all species. Due to the d10 nature of Zn2+, X-ray absorption spectroscopy (XAS) has proven to be critical for obtaining structural and mechanistic information, especially in the absence of diffraction quality crystals. One of the most exciting recent developments in zinc bioinorganic chemistry has been the discovery of an entirely new class of catalytic zinc sites. The first studies have centered on determining structural information about the zinc binding sites in the methionine synthase enzymes of Escherichia coli methylcobamide:CoM methyltransferase, epoxypropane:CoM transferase and the Ada protein both in their native form and with added substrate. Mechanistic implications of the structural changes observed on substrate binding are explored. The second part of these studies begins to examine the role of Zn in embryonic development. For these studies, zebrafish (Danio rerio) embryos have been used as a model system, since these embryos develop outside the mother, and thus must contain all of the nutrients needed for development at the time of fertilization. With the information potential of XAS and the ability to focus the beam to a 4 mum2 spot using the X-ray fluorescence microprobe facilities at the Advanced Light Source, spatial and temporal changes of all of the zinc in zebrafish embryos have been measured at different stages of development. XANES measurements on whole embryos have shown that the bulk zinc environment undergoes significant changes during the first four hours of development. Zinc fluorescence images of the 1-cell, 4-cell, 16-cell, 64-cell, 128-cell and MBT zebrafish embryos (20 minutes--3.3 hours after fertilization) demonstrate that the majority of the zinc is located within the yolk sac, but there is a distinct increase in the zinc fluorescence from the dividing blastomeres as development progresses. Micro-XANES measurements made with the beam positioned in the blastomeres versus the yolk region of the cell show that the zinc environments in the yolk and the blastomere of the same embryo are distinct from one another.Ph.D.BiochemistryInorganic chemistryPhysical chemistryPure SciencesUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/132864/2/9990963.pd

Case of Small Vessel Disease Associated with COL4A1 Mutations following Trauma

With this case report, we would like to heighten the awareness of clinicians about COL4A1 as a single-gene disorder causing cerebral small vessel disease and describe a previously unreported pathogenic missense substitution in COL4A1 (p.Gly990Val) and a new clinical presentation. We identified a heterozygous putatively pathogenic mutation of COL4A1 in a 50-year-old female with a history of congenital cataracts and glaucoma who presented with multiple diffusion-positive infarcts and areas of contrast enhancement following mild head trauma. We believe that this presentation of multiple areas of acute brain and vascular injury in the setting of mild head trauma is a new manifestation of this genetic disorder. Imaging findings of multiple acute infarcts and regions of contrast enhancement with associated asymptomatic old deep microhemorrhages and leukomalacia in adults after head trauma should raise a high suspicion for a COL4A1 genetic disorder. Radiographic patterns of significant leukoaraiosis and deep microhemorrhages can also be seen in patients with long-standing vasculopathy associated with hypertension, which our patient lacked. Our findings demonstrate the utility of genetic screening for COL4A1 mutations in young patients who have small vessel vasculopathy on brain imaging but who do not have significant cardiovascular risk factors