63 research outputs found
Discussions about Limiting Treatment in a Geriatric Clinic
Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/111239/1/j.1532-5415.1993.tb06705.x.pd
Bacterial Pneumonias during an Influenza Pandemic: How Will We Allocate Antibiotics?
We are currently in the midst of the 2009 H1N1 pandemic, and a second wave of flu in the fall and winter could lead to more hospitalizations for pneumonia. Recent pathologic and historic data from the 1918 influenza pandemic confirms that many, if not most, of the deaths in that pandemic were a result of secondary bacterial pneumonias. This means that a second wave of 2009 H1N1 pandemic influenza could result in a widespread shortage of antibiotics, making these medications a scarce resource. Recently, our University of Michigan Health System (UMHS) Scarce Resource Allocation Committee (SRAC) added antibiotics to a list of resources (including ventilators, antivirals, vaccines) that might become scarce during an influenza pandemic. In this article, we summarize the data on bacterial pneumonias during the 1918 influenza pandemic, discuss the possible impact of a pandemic on the University of Michigan Health System, and summarize our committee's guiding principles for allocating antibiotics during a pandemic.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/78141/1/bsp.2009.0019.pd
FAN1 controls mismatch repair complex assembly via MLH1 retention to stabilize CAG repeat expansion in Huntington's disease.
CAG repeat expansion in the HTT gene drives Huntington's disease (HD) pathogenesis and is modulated by DNA damage repair pathways. In this context, the interaction between FAN1, a DNA-structure-specific nuclease, and MLH1, member of the DNA mismatch repair pathway (MMR), is not defined. Here, we identify a highly conserved SPYF motif at the N terminus of FAN1 that binds to MLH1. Our data support a model where FAN1 has two distinct functions to stabilize CAG repeats. On one hand, it binds MLH1 to restrict its recruitment by MSH3, thus inhibiting the assembly of a functional MMR complex that would otherwise promote CAG repeat expansion. On the other hand, it promotes accurate repair via its nuclease activity. These data highlight a potential avenue for HD therapeutics in attenuating somatic expansion
FAN1 modifies Huntington's disease progression by stabilising the expanded HTT CAG repeat
Huntingtonâs disease (HD) is an inherited neurodegenerative disease caused by an expanded CAG repeat in the HTT gene. CAG repeat length explains around half of the variation in age-at-onset, but genetic variation elsewhere in the genome accounts for a significant proportion of the remainder. Genome-wide association studies have identified a bidirectional signal on chromosome 15, likely underlain by FAN1 (FANCD2 and FANCI Associated Nuclease 1), a nuclease involved in DNA interstrand cross link repair. Here we show that increased FAN1 expression is significantly associated with delayed age-at-onset and slower progression of HD suggesting FAN1 is protective in the context of an expanded HTT CAG repeat. FAN1 overexpression in human cells reduces CAG repeat expansion in exogenously expressed mutant HTT exon 1, and in patient-derived stem cells and differentiated medium spiny neurons, FAN1 knockdown increases CAG repeat expansion. The stabilising effect is FAN1 concentration and CAG repeat length dependent. We show that FAN1 binds to the expanded HTT CAG repeat DNA and its nuclease activity is not required for protection against CAG repeat expansion. These data shed new mechanistic insights into how the genetic modifiers of HD act to alter disease progression, and show that FAN1 affects somatic expansion of the CAG repeat through a nuclease-independent mechanism. This provides new avenues for therapeutic interventions in HD and potentially other triplet repeat disorders
De la Casa de Orates al juzgado: pericia alienista y evaluaciĂłn judicial de la locura en Santiago de Chile hacia 1860
Evidence that 2 Nonoverlapping High-Affinity Calmodulin-Binding Sites are Present in the Head of Synapsin-1
Calmodulin is an important element in the regulation of nerve terminal exocytosis by Ca2+. Calmodulin has been shown to interact with the synaptic vesicle phosphoproteins synapsins Ia and Ib [Okabe, T. and Sobue, K. (1987) FEBS Lett. 213, 184-188; Hayes, N. V. L., Bennett, A. E and Baines, A. J. (1991) Biochem. J. 275, 93-97]. These proteins are thought to provide regulated linkages between synaptic vesicles and cytoskeletal elements. It is well established that calmodulin modulates synapsin I activities via calmodulin-dependent protein-kinase-LT-catalysed phosphorylation. The direct binding of calmodulin to synapsin I suggests a second mode of regulation in addition to phosphorylation. In this study, we present evidence indicating that two sites for calmodulin binding exist in the N-terminal head region of synapsins Ia and Ib. In unphosphorylated synapsin I, these sites had a K-d value of = 36+/-14 nM for binding to calmodulin labelled with acetyl-N'-(5-sulpho-1-naphthyl)ethylene diamine. The K-d values for synapsin I phosphorylated at various sites were as follows: site I 18+/-11 nM; sites II and III 35+/-14 nM; sites I-III 16+/-9 nM. The fluorescence data indicated a stoichiometry of not less than 2 mol calmodulin bound to 1 mol synapsin I at saturation in each case. Consistent with this stoichiometry, two chemically cross-linked species (96 kDa and 116 kDa) containing calmodulin and synapsin I were generated in vitro, corresponding to one and two calmodulin molecules bound/synapsin I. Defined fragments of synapsin I were generated with the reagent 2-nitro-5-thiocyanobenzoic acid, which cleaves at cysteine residues. Cysteine-specific cleavage of whole synapsin I after cross-linking to biotinylated calmodulin generated a pair of polypeptide complexes (approximately 46 kDa and 38 kDa), the masses of which indicated cross-linking of calmodulin to the N-terminal and middle regions of synapsin I. Purified N-terminal and middle fragments each showed a Ca2+-dependent interaction with calmodulin affinity columns. Two calmodulin-binding fragments (7.4 kDa and 6.5 kDa) were generated using Staphylococcus aureus V8 protease digestion of synapsin I. These fragments were isolated by calmodulin affinity chromatography and reverse-phase HPLC. N-terminal sequence analysis indicated that the fragments originated from two non-overlapping areas of the synapsin I head region, and that each was contained within one of the 2-nitro-5-thiocyanobenzoic-acid-derived calmodulin-binding fragments. The origins of these fragments are close to the putative sites of actin and tubulin interaction. Addition of calmodulin in the presence of Ca2+ to mixtures of F-actin and synapsin I reduced both the binding of synapsin I to F-actin and bundling of F-actin by synapsin I. Direct binding of calmodulin to synapsin I may represent a non-covalent. mode of regulation of synapsin I activity in addition to covalent regulation by phosphorylation
COORDINATED REGULATION OF SYNAPSIN-I INTERACTION WITH F-ACTIN BY CA2+ CALMODULIN AND PHOSPHORYLATION - INHIBITION OF ACTIN-BINDING AND BUNDLING
The synapsins are a family of synaptic vesicle phosphoproteins whose role seems to be to limit the availability of small synaptic vesicles for exocytosis by Linking them to the cytoskeleton. One member of the family, synapsin I, has been shown to bind calmodulin in a Ca2+-dependent manner. In this study, we have examined whether or not calmodulin can regulate one of the activities of synapsin I, namely, its interaction with F-actin. Synapsin I is an actin bundling protein: this activity is controlled by phosphorylation. Here we show that calmodulin in the presence of Ca2+ is a competitive inhibitor of both actin binding and bundling by synapsin I. Under the conditions of our assay (0.45 mu M synapsin I, 4 mu M F-actin), half-maximal inhibiton of actin binding and bundling by unphosphorylated synapsin I was found with 4.3 and 3.7 mu M calmodulin, respectively. The actin binding activity of synapsin I phosphorylated by cAMP-dependent protein kinase or by calmodulin-dependent protein kinase II showed similar sensitivity to calmodulin inhibition to unphosphorylated synapsin I. However, inhibition of bundling was potentiated. Half-maximal inhibition of bundling by synapsin I phosphorylated by cAMP-dependent kinase was achieved at approximately 0.5 mu M calmodulin. Half-maximal inhibition of bundling by synapsin I phosphorylated by calmodulin-dependent protein kinase II was achieved at less than 0.2 mu M calmodulin, although the maximum binding under the conditions of the assay was lower. Synapsin I phosphorylated by both cAMP-dependent kinase and calmodulin-dependent protein kinase II was half-maximally inhibited from actin binding by less than 100 nM calmodulin, although the binding and bundling characteristics of this form of synapsin I were weak even in the absence of calmodulin. We conclude that covalent (phosphorylation) and noncovalent (calmodulin) forms of regulation act together in regulating one of the activities of synapsin I
The neurofilament antibody RT97 recognises a developmentally regulated phosphorylation epitope on microtubule-associated protein 1B
Microtubules are important for the growth and maintenance of stable neuronal processes and their organisation is controlled partly by microtubule-associated proteins (MAPs). MAP 1B is the first MAP to be expressed in neurons and plays an important role in neurite outgrowth. MAP 1B is phosphorylated at multiple sites and it is believed that the function of the protein is regulated by its phosphorylation state. We have shown that the monoclonal antibody (mAb) RT97, which recognises phosphorylated epitopes on neurofilament proteins, fetal tau, and on Alzheimer's paired helical filament-tau, also recognises a developmentally regulated phosphorylation epitope on MAP 1B. In the rat cerebellum, Western blot analysis shows that mAb RT97 recognises the upper band of the MAP 1B doublet and that the amount of this epitope peaks very early postnatally and decreases with increasing age so that it is absent in the adult, despite the continued expression of MAP 1B in the adult. We confirmed that mAb RT97 binds to MAP 1B by showing that it recognises MAP 1B immunoprecipitated from postnatal rat cerebellum using polyclonal antibodies to recombinant MAP 1B proteins. We established that the RT97 epitope on MAP 1B is phosphorylated by showing that antibody binding was abolished by alkaline phosphatase treatment of immunoblots. Epitope mapping experiments suggest that the mAb RT97 site on MAP 1B is near the N-terminus of the molecule. Despite our immunoblotting data, immunostaining of sections of postnatal rat cerebellum with mAb RT97 shows a staining pattern typical of neurofilaments with no apparent staining of MAP 1B. For instance, basket cell axons and axons in the granule cell layer and white matter stained, whereas parallel fibres did not. These results suggest that the MAP 1B epitope is masked or lost under the immunocytochemical conditions in which the cerebellar sections are prepared. The upper band of the MAP 1B doublet is believed to be predominantly phosphorylated by proline-directed protein kinases(PDPKs). PDPKs are also good candidates for phosphorylating neurofilament proteins and tau and therefore we postulate that the sites recognised by RT97 on these neuronal cytoskeletal proteins may be phosphorylated by similar kinases. Important goals are to determine the precise location of the RT97 epitope on MAP 1B and the kinase responsible
A Multimedia Intervention on Cardiopulmonary Resuscitation and Advance Directives
Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/74528/1/j.1525-1497.1999.11208.x.pd
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