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Next-generation genetic sequencing (NGS) technologies facilitate the screening of multiple genes linked to neurodegenerative dementia, but there are few reports about their use in clinical practice. Which patients would most profit from testing, and information on the likelihood of discovery of a causal variant in a clinical syndrome, are conspicuously absent from the literature, mostly for a lack of large-scale studies. We applied a validated NGS dementia panel to 3241 patients with dementia and healthy aged controls; 13,152 variants were classified by likelihood of pathogenicity. We identified 354 deleterious variants (DV, 12.6% of patients); 39 were novel DVs. Age at clinical onset, clinical syndrome and family history each strongly predict the likelihood of finding a DV, but healthcare setting and gender did not. DVs were frequently found in genes not usually associated with the clinical syndrome. Patients recruited from primary referral centres were compared with those seen at higher-level research centres and a national clinical neurogenetic laboratory; rates of discovery were comparable, making selection bias unlikely and the results generalisable to clinical practice. We estimated penetrance of DVs using large-scale online genomic population databases and found 71 with evidence of reduced penetrance. Two DVs in the same patient were found more frequently than expected. These data should provide a basis for more informed counselling and clinical decision making.

The primary progressive aphasias (PPA) are a group of clinically, genetically and pathologically heterogeneous neurodegenerative disorders caused by FTLD-tau, FTLD-TDP or Alzheimers disease pathology. Clinically, three subtypes are recognized, the semantic, logopenic and nonfluent variants but there remains ongoing discussions over how the clinical subtypes should be dissected. This review looks at the genetic and pathological basis of PPA and argues that with the advent of clinical trials in PPA, establishing the underlying pathology accurately during life will become increasingly important. Current and future biomarkers that may help make a pathological diagnosis in life, i.e. PPA-tau, PPA-TDP and PPA-AD, are reviewed including clinical and neuropsychological data, neuroimaging, blood and CSF markers.

There is currently much interest in biomarkers of disease activity in frontotemporal lobar degeneration (FTLD). We assessed MRI and behavioral measures of progression in a longitudinal FTLD cohort.

Frontotemporal lobar degeneration (FTLD) is a genetically and pathologically heterogeneous neurodegenerative disorder.

Frontotemporal lobar degeneration (FTLD) is a clinically, genetically, and pathologically heterogeneous neurodegenerative disorder. Two subtypes commonly present with a language disorder: semantic dementia (SemD) and progressive nonfluent aphasia (PNFA).

Semantic dementia is a sporadic neurodegenerative disorder characterized by the progressive erosion of semantic processing and is one of the canonical subtypes of frontotemporal lobar degeneration. This study aimed to characterize the pattern of global and regional longitudinal brain atrophy in semantic dementia and to identify imaging biomarkers that could underpin therapeutic trials.

The article discusses the life and works of American photographer Asahel Curtis. According to the author, Curtis is capable of capturing the natural splendor and the emerging 20th century civilization of the Pacific Northwest wherein his prolific commercial works allow him to go to different places. In addition, the author notes that many of the works of Curtis document the different aspects of Pacific Northwest life as well as yachting assignments.

Background & Aims: The molecular targets for the protective actions of conjugated linoleic acid (CLA) on experimental inflammatory bowel disease (IBD) are unknown. We used a loss-of-function approach to investigate whether CLA ameliorated colitis through a peroxisome proliferator-activated receptor @c (PPAR @c)-dependent mechanism. Methods: The expression of PPAR @c, @d, and their target genes in the colon of mice fed control or CLA-supplemented diets was assayed after a 7-day dextran sodium sulfate (DSS) challenge by quantitative real-time polymerase chain reaction (PCR). Additionally, nuclear factor-@k B (NF-@kB) p65 activation was quantified in the colon. To determine the involvement of PPAR @c in the mechanism of action of CLA directly, specific deletions of PPAR @c in the colon were performed in mice by using the Cre-lox recombination system. Colonic PPAR @c null mice and wild-type littermates were fed either a CLA-supplemented or a control diet for 42 days and challenged with 2.5% DSS. The therapeutic efficacy of CLA also was examined by using the CD4^+CD45RB^h^i transfer colitis model. Results: CLA induced PPAR @c and @d, transcriptionally modulated PPAR @c and @d-responsive gene clusters involved in lipid metabolism (uncoupling protein [UCP]1, UCP3, PPAR @c coactivator 1@a [PGC-1@a], and CD36) and epithelial cell maturation (Gob-4 and Keratin 20). Additionally, CLA repressed tumor necrosis factor @a (TNF-@a) expression and NF-@kB activation while inducing the immunoregulatory cytokine transforming growth factor @b 1 (TGF-@b"1). Clinically, CLA ameliorated DSS- and CD4^+-induced colitis. Loss of the PPAR @c gene in the colon abrogated the beneficial effects of CLA in DSS colitis. Conclusions: Our studies provide molecular evidence in vivo, suggesting that CLA ameliorates colitis through a PPAR @c-dependent mechanism.

Market research is an important element of the strategic marketing process. By understanding the healthcare needs of a market area, hospital and health system managers can set priorities for new services and allocate resources appropriately. The process of market research often begins with an evaluation of health status and socioeconomic indicators collected from secondary sources. Unfortunately, indicators that have been recommended in the literature may not be feasible for use in rural markets because of their lack of statistical precision or inability to differentiate healthcare service needs. This study evaluated the statistical precision and variability of 79 secondary health status and socioeconomic measures reported at the county level in Iowa, USA, a largely rural state. Our findings suggest that many readily available health status and socioeconomic indicators do not discriminate need among rural health care markets. Only six health status and two socioeconomic indicators met our statistical precision and variability criteria. These findings have important implications for managers planning health services in rural localities. Managers of rural health systems may need to employ alternative market research methods, such as analysis of claims-based utilization rates or community health surveys.

Rough terrain in mid‐ and high latitudes is often covered with highly reflective snow, giving rise to a very complex transfer of incident sunlight. In order to simplify the radiative transfer in weather and climate models, snow is generally treated as an isotropically reflecting material. We develop a new model of radiative transfer over mountainous terrain, which considers for the first time the forward scattering properties of snow. Combining ground‐measured meteorological data and high resolution digital elevation models, we show that the forward scattering peak of snow leads to a strong local redistribution of incident terrain reflected radiation. In particular, the effect of multiple terrain reflections is enhanced. While local effects are large, area‐wide albedo is only marginally decreased. In addition, we show that solar panels on snowy ground can clearly benefit from forward scattering, helping to maximize winter electricity production. Forward scattering of light by snow rewards slopes facing the sun at the expense of shady slopesForward scattering amplifies the multiple scattering and increases terrain‐wide irradianceForward scattering increases the potential for winter solar panel yield in mountainous terrain Forward scattering of light by snow rewards slopes facing the sun at the expense of shady slopes Forward scattering amplifies the multiple scattering and increases terrain‐wide irradiance Forward scattering increases the potential for winter solar panel yield in mountainous terrain

XCE, a new member of the endothelin-converting enzyme and neutral endopeptidase family, is preferentially expressed in specific areas of the central nervous system including spinal chord and medulla. To elucidate the importance and function of XCE, we disrupted its gene in mouse embryonic stem cells by homologous recombination and created mice deficient in XCE. The resulting phenotype is characterized by neonatal lethality. All XCE -/- homozygous mice died of respiratory failure shortly after birth, and in most cases their lungs were never ventilated. Apart from the atelectasis, anatomical and histological examinations of embryonic day 18.5 XCE -/- embryos and newborn homozygotes did not reveal any obvious abnormalities in organs and tissues. Malformations that are related to the knock-out were also not found in the skeletons of XCE -/- mice. In addition, XCE knock-out animals showed no deficiency of pulmonary surfactant proteins and had normal heart beat frequencies. Taken together, our results demonstrate that XCE is an essential gene. The phenotype of the XCE-deficient mice together with the central nervous system-specific expression further suggest that XCE may play a vital role in the control of respiration.

Monitoring the performance of the health care delivery system is a public health function that becomes more important as organized delivery systems begin to take control over large portions of the medical care market. The study reported here illustrates how standard medical care epidemiology can be applied to analysis of health care system performance to aid governmental efforts to monitor new developments in the medical care market. In order to evaluate the efficiency of hospital care delivered in Iowa, age- and sex-adjusted population admission rates for five common procedures were generated for all 99 counties. The five common procedures were defined as follows: hernia, tonsillectomy, cesarean section, hysterectomy, and cholecystectomy. In addition, variations in 11 ambulatory care-sensitive condition rates were analyzed. Residents of at least 15 counties were at significantly greater than average risk for receiving each of the common procedures (χ2test, P< 0.05). Counties that had a high rate for one procedure tended to have a high rate for at least one other procedure. Several counties had more than twice the mean rate. Even a 10% reduction, when added across all five common procedures, amounts to well over 2000 hospitalizations avoided. It is assumed that reductions would be concentrated in high-rate counties. If a 50% reduction could be achieved in only part of the ambulatory care-sensitive procedures, more than 10 000 hospital admissions could be avoided.

The outcomes of cardiovascular care for individual patients are known to be better when treatment is provided in hospitals where the volume of such care is greater. We examined the impact of establishing safe volume thresholds on hospitals now performing such procedures in the state of Iowa by analysing the relationship between various volume thresholds and the number of hospitals that would continue to provide the service if the thresholds were enforced. Four procedures performed in hospitals in Iowa in 1990 were studied: coronary artery bypass graft surgery, vascular surgery, cardiac valve surgery, and cardiac catheterization. The analysis was conducted assuming current per capita procedure rates and repeated assuming reduced procedure rates. The study finds that the 12 hospitals now performing coronary artery bypass graft surgery in Iowa could be reduced to two, providers of vascular surgery could be reduced to as low as four from the current 33, the 10 providers of cardiac valve surgery might be reduced to one, and catheterization could be provided at two rather than 22 hospitals.

Thrombocytopenia may be the presenting finding for both Wiskott-Aldrich syndrome and Fanconi anemia. We examined a sibship of four boys who had features of both of these hematologic disorders. Peripheral blood lymphocytes from three of the boys demonstrated DNA instability when cultured with diepoxybutane, confirming the diagnosis of Fanconi anemia in these patients. However, results of linkage analysis and X chromosome inactivation studies were consistent with the diagnosis of Wiskott-Aldrich syndrome in two of the boys, including one of the boys with Fanconi anemia. These findings could be attributed to the occurrence of two rare genetic disorders in a single family or to an unusual variant of Fanconi anemia. The recent identification of the Wiskott-Aldrich gene permitted us to address this question directly. Epstein-Barr virus-transformed cell lines from the two boys thought to have Wiskott-Aldrich syndrome on the basis of linkage analysis failed to express transcripts for the Wiskott-Aldrich gene. Genomic DNA from these two patients demonstrated a G insertion in the tenth exon of the Wiskott-Aldrich gene, resulting in a frameshift and a premature stop codon. Surprisingly, the patient with Fanconi anemia and a null mutation in the Wiskott-Aldrich gene had typical Fanconi anemia but mild Wiskott-Aldrich syndrome. (J PEDIATR 1996;129:50-5)

Serum riboflavin-binding protein, a phosphoglycoprotein from the blood of laying hens, contains two Asn-Xaa-(Thr)Ser sequons in very similar but well-separated regions of amino acid sequence. In order to evaluate the effect of local amino acid sequence on the structure of the attached oligosaccharides, serum riboflavin-binding protein was purified to homogeneity, reduced and alkylated, digested with trypsin, and the two glycopeptides were separated by reversed-phase chromatography. After digestion with peptide-N-glycosidase F the released oligosaccharides were separated by high-pH anion-exchange chromatography and the oligosaccharide profiles of the two glycopeptides were compared. Although the two asparagine residues that are glycosylated are contained in pentapeptide segments in which four out of five amino acids are identical, the array of oligosaccharides present at each site show differences in both type and distribution. This suggests that local secondary or tertiary structure, or the order of glycosylation, influences the oligosaccharide structure more than does the primary structure flanking the attachment site.

The Saccharomyces cerevisiae alpha-pheromone receptor, a polytopic, G protein-coupled, membrane protein, is internalized after binding of alpha-factor. Mutational analysis suggested that the first 39 residues of the receptor's cytoplasmic tail carries sufficient information for internalization. A point mutation in one of these 39 residues, K337 to R337, renders the receptor nonfunctional for endocytosis. Other residues, D335 and S338, contribute to the efficiency of internalization. When the sequence DAKSS is added onto a severely truncated receptor, endocytosis of the receptor is restored, showing that this sequence functions to mediate or to signal interaction with the endocytic machinery. Analysis of pheromone response and recovery in strains expressing mutant receptors suggests that receptor internalization is not important for response but contributes to recovery from pheromone.

p63 is a type II integral membrane protein that has previously been suggested to be a resident protein of a membrane network interposed between the ER and the Golgi apparatus. In the present study, we have produced a polyclonal antibody against the purified human p63 protein to reassess the subcellular distribution of p63 by confocal immunofluorescence, immunoelectron microscopy, and cell fractionation. Double immunofluorescence of COS cells showed significant colocalization of p63 and a KDEL-containing lumenal ER marker protein, except for differences in the staining of the outer nuclear membrane. Immunoelectron microscopy of native HepG2 cells and of COS cells transfected with p63 revealed that both endogenous and overexpressed p63 are predominantly localized in the rough ER. While p63 was colocalized with protein disulfide isomerase, an ER marker protein, very little overlap of p63 was found with ERGIC-53, an established marker for the ER-Golgi intermediate compartment. When rough and smooth membranes were prepared from rat liver, p63 was found to copurify with ribophorin II, a rough ER protein. Both p63 and ribophorin II were predominantly recovered in rough microsomes and were largely separated from the intermediate compartment marker protein p58. From these results it is concluded that p63 is localized in the rough ER.

The recently identified 63 kDa membrane protein, p63, is a resident protein of a membrane network interposed in between rough ER and Golgi apparatus. To characterize p63 at the molecular level a 2.91 kb cDNA encoding p63 has been isolated from a human placenta lambda gt10 cDNA library. Sequence analysis of tryptic peptides prepared from isolated p63 confirmed the identify of the cloned gene. The translated amino acid sequence consists of 601 amino acids (65.8 kDa) with a single putative membrane-spanning region and a N-terminal cytoplasmic domain of 106 amino acids. The human p63 cDNA exhibits a high level of sequence identify to the pig hepatic cDNA 3AL (accession number M27092) whose expression is enhanced after resuscitation from circulatory shock. An additional remarkable feature of p63 is that it becomes reversibly palmitoylated when intracellular protein transport is blocked by the drug brefeldin A. Overexpression of p63 in COS cells led to the development of a striking tubular membrane network in the cytoplasm. This suggests that the protein may be determinant for the structure of the p63 compartment.

Palmitoylation of p63, a type II membrane protein localized in the endoplasmic reticulum, is induced in a reversible manner by the drug brefeldin A. To study the requirements for palmitoylation, mutant forms of p63 were expressed in COS cells and analyzed by metabolic labeling with [3H]palmitate, immunoprecipitation, and SDS-polyacrylamide gel electrophoresis. By investigating deletion and point mutations, Cys100 in the 106-amino acid cytoplasmic tail of p63 has been identified as the site of acylation. Site-directed mutagenesis of residues 99-105 together with cytoplasmic tail truncation mutants showed that the amino acids surrounding Cys100 are not critical for palmitoylation of this residue. Analysis of a chimeric construct between p63 and the plasma membrane protein dipeptidylpeptidase IV further revealed that p63 palmitoylation is not dependent on its transmembrane domain. In contrast, the six-amino acid distance between the end of the predicted transmembrane domain and the palmitoylation site was found to be essential for proper acylation of p63.

The nucleotide sequence of a 5,317-bp fragment which includes the region required for mobilization of broad-host-range plasmid pTF-FC2 was determined. A region of approximately 3.5 kb was required for plasmid mobilization, and oriT was localized on a 138-bp fragment. Polypeptides which corresponded in size and location to several of the open reading frames were detected in an in vitro transcription-translation system. Three open reading frames essential for plasmid mobilization and two which affect the mobilization frequency were identified. There was a distinct similarity in the sizes, amino acid sequences, and locations of the proteins from the mobilization region of pTF-FC2 and the Tra1 region of IncP plasmid RP4. Similarity in the structures and sequences of the oriT regions was also apparent. A sequence with 37-of-38-bp homology to the inverted repeated sequences of Tn21 and an open reading frame with strong homology to the MerR regulatory protein was identified outside of the region required for mobilization.

alpha-factor, one of two peptide hormones responsible for synchronized mating between MATa and MAT alpha-cell types in Saccharomyces cerevisiae, binds to its cell surface receptor and is internalized in a time-, temperature-, and energy-dependent manner (Chvatchko, Y., I. Howald, and H. Riezman. 1986. Cell. 46:355-364). After internalization, alpha-factor is delivered to the vacuole via vesicular intermediates and degraded there consistent with an endocytic mechanism (Singer, B., and H. Riezman. 1990. J. Cell Biol. 110:1911-1922; Chvatchko, Y., I. Howald, and H. Riezman. 1986. Cell. 46:355-364). We have isolated two mutants that are defective in the internalization process. Both mutations confer a recessive, temperature-sensitive growth phenotype upon cells that cosegregates with their endocytosis defect. Lucifer yellow, a marker for fluid-phase endocytosis, shows accumulation characteristics in the mutants that are similar to the uptake characteristics of 35S-alpha-factor. The endocytic defect in end4 cells appears immediately upon shift to restrictive temperature and is reversible at permissive temperature if new protein synthesis is allowed. Furthermore, the end4 mutation only affects alpha-factor internalization and not the later delivery of alpha-factor to the vacuole. Other vesicle-mediated processes seem to be normal in end3 and end4 mutants. END3 and END4 are the first genes shown to be necessary for the internalization step of receptor-borne and fluid-phase markers in yeast.

Molecules travel through the yeast endocytic pathway from the cell surface to the lysosome-like vacuole by passing through two sequential intermediates. Immunofluorescent detection of an endocytosed pheromone receptor was used to morphologically identify these intermediates, the early and late endosomes. The early endosome is a peripheral organelle that is heterogeneous in appearance, whereas the late endosome is a large perivacuolar compartment that corresponds to the prevacuolar compartment previously shown to be an endocytic intermediate. We demonstrate that inhibiting transport through the early secretory pathway in sec mutants quickly impedes transport from the early endosome. Treatment of sensitive cells with brefeldin A also blocks transport from this compartment. We provide evidence that Sec18p/N-ethylmaleimide-sensitive fusion protein, a protein required for membrane fusion, is directly required in vivo for forward transport early in the endocytic pathway. Inhibiting protein synthesis does not affect transport from the early endosome but causes endocytosed proteins to accumulate in the late endosome. As newly synthesized proteins and the late steps of secretion are not required for early to late endosome transport, but endoplasmic reticulum through Golgi traffic is, we propose that efficient forward transport in the early endocytic pathway requires delivery of lipid from secretory organelles to endosomes.

Presence or absence of N-acetylneuraminic acid (Neu5Ac) can change a sialylated glycoprotein's serum half-life and possibly its function. We evaluated the linearity, sensitivity, reproducibility, and accuracy of a HPAEC/PAD method to determine its suitability for routine simultaneous analysis of Neu5Ac and N-glycolylneuraminic acid (Neu5Gc). An effective internal standard for this analysis is 3-deoxy-d-glycero-d-galacto-2-nonulosonic acid (KDN). We investigated the effect of the Au working electrode recession and determined that linear range and sensitivity were dependent on electrode recession. Using an electrode that was 350 microm recessed from the electrode block, the minimum detection limits of Neu5Ac, KDN, and Neu5Gc were 2, 5, and 2 pmol, respectively, and were reduced to 1, 2, and 0.5 pmol using a new electrode. The response of standards was linear from 10 to 500 pmol (r2>0.99) regardless of electrode recession. When Neu5Ac, KDN, and Neu5Gc (200 pmol each) were analyzed repetitively for 48 h, area RSDs were <3%. Reproducibility was unaffected when injections of glycoprotein neuraminidase and acid digestions were interspersed with standard injections. Area RSDs of Neu5Ac and Neu5Gc improved when the internal standard was used. We determined the precision and accuracy of this method for both a recessed and a new working electrode by analyzing Neu5Ac and Neu5Gc contents of bovine fetuin and bovine and human transferrins. Results were consistent with published values and independent of the working electrode. The sensitivity, reproducibility, and accuracy of this method make it suitable for direct routine analysis of glycoprotein Neu5Ac and Neu5Gc contents.

Stabilization of iron in a bioavailable form is the function of ferritin, a protein of 24 subunits forming a coat around a core of less than or equal to 4500 hydrated iron atoms. The core of ferritin isolated from tissues contains Fe3+, but Fe2+ is required for experimental core formation in protein coats; reduction of Fe3+ to Fe2+ facilitates iron removal from protein coats. Using the differences in x-ray absorption spectra (x-ray absorption near edge structure) between Fe2+ and Fe3+ to monitor reconstitution of ferritin from Fe2+ and protein coats, we observed stabilization of Fe2+, apparently inside the coat. Mixtures of Fe2+ and Fe3+ persisted for greater than or equal to 16 h in air indicating that, in vivo, some iron in ferritin could be stored as Fe2+ and with Fe3+ could yield magnetite.