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PubMed1999

Structure and promoter activity of the 5' flanking region of ace-1, the gene encoding acetylcholinesterase of class A in Caenorhabditis elegans.

We report the structure and the functional activity of the promoter region of ace-1, the gene encoding acetylcholinesterase of class A in the nematode Caenorhabditis elegans. We found that ace-1 was trans -spliced to the SL1 spliced leader and that transcription was initiated at a cluster of multiple starts. There was neither a TATA nor a CAAT box at consensus distances from these starts. Interspecies sequence comparison of the 5' regions of ace-1 in C. elegans and in the related nematode Caenorhabditis briggsae identified four blocks of conserved sequences located within a sequence of 2.4 kilobases upstream from the initiator ATG. In vitro expression of CAT reporter genes in mammalian cells allowed the determination of a minimal promoter in the first 288 nucleotides. In phenotype rescue experiments in vivo, the ace-1 gene containing 2.4 kilobases of 5' flanking region of either C. elegans or C. briggsae was found to restore a coordinated mobility to the uncoordinated double mutants ace-1(-);ace-2(-)of C. elegans. This showed that the ace-1 promoter was contained in 2.4 kilobases of the 5' region, and indicated that cis -regulatory elements as well as coding sequences of ace-1 were functionally conserved between the two nematode species. The pattern of ace-1 expression was established through microinjection of Green Fluorescent Protein reporter gene constructs and showed a major mesodermal expression. Deletion analysis showed that two of the four blocks of conserved sequences act as tissue-specific activators. The distal block is a mesodermal enhancer responsible for the expression in body wall muscle cells, anal sphincter and vulval muscle cells. Another block of conserved sequence directs expression in pharyngeal muscle cells pm5 and three pairs of cephalic sensory neurons.

5' Untranslated Regions↗
PubMed1995

The differences in structural specificity for recognition and binding between asialoglycoprotein receptors of liver and macrophages.

The Gal/GalNAc-specific lectin on the surface of rat peritoneal macrophages (macrophage asialoglycoprotein binding protein, M-ASGP-BP), which consists of a single polypeptide chain of 42 kDa, can form a homo-oligomeric receptor exhibiting high affinity for asialoorosomucoid (ASOR) [Ozaki K., Ii M., Itoh N., Kawasaki T. (1992) J Biol Chem 267: 9229-35]. In this study, the binding affinity of M-ASGP-BP was studied by using a series of synthetic or natural glycosides as inhibitors of 125I-ASOR binding to recombinant M-ASGP-BP expressed on COS-1 cells (rM-ASGP-BP), and the results were compared with those of human hepatic lectin (HHL) on Hep G2 cells. Clustering of multiple Gal (or GalNAc) residues increased the binding affinity to M-ASGP-BP as well as to HHL. In contrast to HHL and other mammalian hepatic lectins, rM-ASGP-BP bound Gal residues tighter than GalNAc residues. A galactose-terminated triantennary N-glycoside, having one N-acetyl-lactosamine unit on the 6 branch and two N-acetyl-lactosamine units on the 3 branch of the trimannosyl core structure, showed affinity enhancement of approximately 10(5) over a monovalent ligand for HHL, while the same glycopeptide showed enhancement of about 2000-fold for rM-ASGP-BP. These results suggest that spatial arrangements of sugar combining sites and subunit organization of macrophage and hepatic lectins are different.

Animals↗
PubMed1994

Purification of the channel component of the mitochondrial calcium uniporter and its reconstitution into planar lipid bilayers.

The purification of the channel-forming component of the mitochondrial calcium uniporter and its channel properties are described. After ethanol and 50% ethanol-water extraction of mitochondria from beef heart or perfused rat liver, the extract was passed through thiopropyl-Sepharose 6B column, and absorbed components were eluted with 2-mercaptoethanol, followed by gel-filtration on Sephadex G-15. The last fraction eluted (M(r) about 2000) was then subjected to reverse-phase high-performance liquid chromatography. Of the more than 10 distinct peaks, only one showed specific Ca(2+)-channel activity in BLM with properties similar to earlier, less extensively purified preparations, i.e., conductance of 20 pS and multiples thereof, clustering of channels, participation of 2 or more subunits in channel formation, and sensitivity to 1 microM ruthenium red. Voltage sensitivity and cooperativity between channels are described. The Ca(2+)-binding glycoprotein with which the peptide was associated was found to have high homology with human acid alpha 1-glycoprotein (orosomucoid) and to show identity with beef plasma orosomucoid in the Ouchterlony immunodiffusion test.

Amino Acid Sequence↗
PubMed1996

PWP2, a member of the WD-repeat family of proteins, is an essential Saccharomyces cerevisiae gene involved in cell separation.

WD-repeat proteins contain four to eight copies of a conserved motif that usually ends with a tryptophan-aspartate (WD) dipeptide. The Saccharomyces cerevisiae PWP2 gene, identified by sequencing of chromosome III, is predicted to contain eight so-called WD-repeats, flanked by nonhomologous extensions. This gene is expressed as a 3.2-kb mRNA in all cell types and encodes a protein of 104 kDa. The PWP2 gene is essential for growth because spores carrying the pwp2 delta 1::HIS3 disruption germinate before arresting growth with one or two large buds. The growth defect of pwp2 delta 1::HIS3 cells was rescued by expression of PWP2 or epitope-tagged HA-PWP2 using the galactose-inducible GALI promoter. In the absence of galactose, depletion of Pwp2p resulted in multibudded cells with defects in bud site selection, cytokinesis, and hydrolysis of the septal junction between mother and daughter cells. In cell fractionation studies, HA-Pwp2p was localized in the particulate component of cell lysates, from which it would be solubulized by high salt and alkaline buffer but not by nonionic detergents or urea. Indirect immunofluorescence microscopy indicated that HA-Pwp2p was clustered at multiple points in the cytoplasm. These results suggest that Pwp2p exists in a proteinaceous complex, possibly associated with the cytoskeleton, where it functions in control of cell growth and separation.

Base Sequence↗
PubMed1993

DSM-III-R personality disorders in outpatients with non-bipolar depression: the frequency in a sample of Japanese and the relationship to the 4-month outcome under adequate antidepressant therapy.

We investigated the frequency of personality disorders (PDs) and the relationship between the presence of PD and the 4-month outcome of depression under adequate antidepressant therapy in a Japanese sample of 96 outpatients with non-bipolar major depression. The diagnosis of PD was made using a structured interview method (the Structured Clinical Interview for DSM-III-R Personality Disorders) and after severe depressive symptoms were reduced. Any one kind of PD was found in 54.2% of the sample. The most frequent was avoidant (35.4%), obsessive-compulsive (22.9%), narcisstic (18.8%), and dependent (16.7%) PDs. The frequencies of these PDs in our study, except narcisstic PD, were about the same as those reported in previous studies with a matched setting for the PD diagnosis. Compared with patients without PD, a worse outcome was found in patients with PD, especially patients with multiple PDs from multiple PD clusters. There was no evidence that a specific PD or PD cluster especially worsens the outcome of depression.

Adult↗
PubMed1999

Challenges and future directions for tailored communication research.

As informatics technology advances, a growing number of research trials on tailored communications provide an accumulation of promising evidence to support their efficacy. These trials also reveal gaps and opportunities for future research. The scope and boundaries of tailoring must be redefined in terms of both new technology and the trade-offs between complexity, demand burden on participants, and the minimal information required for effective and efficient tailoring. Basic and methods research is needed to broaden theory, develop a common language, standardize measures, and isolate the key mediating mechanisms that facilitate tailored communications. Applied research must consider more rigorous research designs for efficacy trials and conduct more effectiveness trials to investigate the mechanisms of technology transfer to enhance large-scale diffusion of tailored communications. The role of contextual variables needs to be examined, as well as their interaction with different population groups, and also the channels, modes, and methods of tailored message delivery. Research is also needed on the feasibility of tailoring across clusters of multiple risk factors to identify the commonalities, differences, and interrelations among diverse behaviors. The potential cost-effectiveness of tailored communications must also be examined. No matter how efficacious, tailored communications delivered to large populations (i.e. mass-customization) will not make a public health impact unless proven to be practical and cost-efficient.

Cost-Benefit Analysis↗
PubMedDate not supplied

Multiple chromosomal associations and paracentromeric region instability in a case of acute leukemia.

A case of acute myelomonocytic leukema is described, which was characterized cytogenetically by the presence of centromeric elongations, somatic crossovers, selective endoreduplication figures, and multiple chromosomal clusters. The demonstration of these phenomena by selective staining techniques for the chromosome bands (Q, C, G and S) and the nucleolar areas (acridine-orange, amido black B 10) raises some biological aspects involved in the proliferation of leukemic cells, such as nucleolar persistance during the metaphase and the non-separation of chromatids in the clusters during the anaphase. These structural abnormalities may represent the background for the explanation of the appearance of subclones in neoplastic disorders.

Adult↗
PubMed2002

Microsatellites in starch-synthesizing genes in relation to starch physicochemical properties in waxy rice ( Oryza sativa L.).

Rice starch is composed of amylose and amylopectin. Amylose content, an important determinant of rice starch quality, is primarily controlled by the waxy gene, encoding granule-bound starch synthase (GBSS). The starch branching enzyme (SBE) and soluble starch synthase (SSS) play major roles in the synthesis of amylopectin. Microsatellite polymorphisms in the three genes, the wx gene encoding granule-bound starch synthase I, the SBE gene encoding starch branching enzyme I and the SSS gene encoding soluble starch synthase I, were studied for 56 accessions of waxy rice ( Oryza sativa L.). Four (CT)(n) microsatellite alleles, (CT)(16), (CT)(17), (CT)(18) and (CT)(19), at the wx locus were detected in this set of waxy rice, of which (CT)(17) was the most frequent. Three (CT)(n) microsatellite allele classes were found at the SBE locus, (CT)(8) or (CT)(10) together with an insertion sequence of CTCTCGGGCGA, and (CT)(8) alone without the insertion. There were multiple microsatellites clustered at the SSS locus. However, these alleles can also be grouped into three classes, i.e. the allele class SSS-A = (AC)(2) em leader TCC(TC)(11) em leader (TC)(5)C(ACC)(11), the allele class SSS-B = (AC)(3) em leader TCT(TC)(6) em leader (TC)(4)C(ACC)(9), and the allele class SSS-C = (AC)(3) em leader TCT(TC)(6) em leader (TC)(4)C(ACC)(8). The analyses of starch physicochemical properties among different microsatellite genotypes indicated that the waxy rice group with the (CT)(19) allele, the SBE-A allele and the SSS-B allele was quite different from other groups. Nine out of 15 accessions with a high gelatinization temperature (GT) belonged to the wx (CT)(19) group, all of them belonged to the SBE-A group and 13 of them belonged to the SSS-B group. These microsatellites might be useful in marker-assisted breeding for the improvement of rice grain quality.

Journal Article↗
PubMed2006

Microsatellites, single nucleotide polymorphisms and a sequence tagged site in starch-synthesizing genes in relation to starch physicochemical properties in nonwaxy rice (Oryza sativa L.).

Starch characteristics determine the quality of various products of rice, e.g., eating, cooking and processing qualities. Our previous study indicated that molecular markers inside or close to starch synthesizing genes can differentiate the starch properties of 56 waxy rices. Here we report microsatellite (or simple sequence repeat, SSR) polymorphism in the Waxy (Wx) gene, soluble starch synthase I gene (SS1) and starch branching enzyme 1 gene (SBE1), single nucleotide polymorphism (SNP) in Wx and starch branching enzyme 3 gene (SBE3), and a sequence tagged site (STS) in starch branching enzyme 1 gene (SBE1) among 499 nonwaxy rice samples and their relationships with starch physicochemical properties. The nonwaxy rice samples consist of landraces (n = 172) obtained from germplasm centers and cultivars and breeding lines (n = 327) obtained from various breeding programs. Ten (CT)( n ) microsatellite alleles, (CT)(8), (CT)(10), (CT)(11), (CT)(12, )(CT)(17), (CT)(18), (CT)(19), (CT)(20), (CT)(21), and (CT)(22), were found at the Wx locus, of which (CT)(11) was the most frequent, and (CT)(12), (CT)(21) and (CT)(22) were identified for the first time. Four (CT)( n ) microsatellite alleles were found at the SBE1 locus, (CT)(8), (CT)(9), and (CT)(10 )together with an insertion sequence of CTCTCGGGCGA, and (CT)(8) alone without the insertion, of which (CT)(9) and the insertion was a new allele identified in only one rice, IR1552. Multiple microsatellites clustered at the SS1 locus, and in addition to the three alleles previously detected (SSS-A = (AC)(2)...TCC(TC)(11)...(TC)(5)C(ACC)(11), SSS-B = (AC)(3)...TCT(TC)(6)...(TC)(4)C(ACC)(9), and SSS-C = (AC)(3)...TCT(TC)(6)...(TC)(4)C(ACC)(8)), one new allele (SSS-D = (AC)(2)...TCC(TC)(10)...(TC)(4)C(ACC)(9)) was found. Analysis of the starch physicochemical properties of the samples with different microsatellites, SNPs and STS groups indicated that these molecular markers can differentiate almost all the physicochemical properties examined, e.g., apparent amylose content (AAC), pasting viscosity characteristics, and gel textural properties. Wx SSR and Wx SNP alone explained more variations for all physicochemical properties than the other molecular markers. The total six markers could explain 92.2, 81 and 86% of total variation of AAC, gel hardness (HD), and gel cohesiveness (COH), respectively, and they could explain more than 40% of the total variation of hot paste viscosity (HPV), cool paste viscosity (CPV), breakdown viscosity (BD), setback viscosity (SB) and gel adhesiveness (ADH). However, only 29% of the total variation of peak viscosity (PV) and 37% of pasting temperature (PT) could be explained by all the molecular markers. Some of these markers can differentiate the starch physicochemical properties among the rice samples with the same Wx allele, indicating that the variation within Wx allele classes can be explained by other starch synthesizing genes. These SSRs, SNPs and STS are useful in marker-assisted breeding for the improvement of starch quality of rice.

1,4-alpha-Glucan Branching Enzyme↗
PubMed2001

Saturated non-esterified fatty acids stimulate de novo diacylglycerol synthesis and protein kinase c activity in cultured aortic smooth muscle cells.

AIMS/HYPOTHESIS: Insulin resistance is linked with a cluster of multiple risk factors and excessive acceleration of atherosclerosis. The underlying mechanism is not, however, fully understood. METHODS: To determine the link between insulin resistance and altered vascular function, we focused on the effect of various non-esterified fatty acids on diacylglycerol-protein kinase C pathway and mitogen-activated protein kinase activity in cultured aortic smooth muscle cells. RESULTS: Incubation of the cells with saturated non-esterified fatty acids (200 micromol/l) for 24 h, such as palmitate or stearate, induced a significant increase in diacylglycerol concentrations by about fivefold or eightfold, respectively, whereas oleate induced a slight increase in diacylglycerol concentrations by 1.8-fold and arachidonate induced none. In addition, the increased diacylglycerol concentrations induced by palmitate were completely restored to control concentrations by triacsin C, acyl-CoA synthetase inhibitor. These results suggest that saturated non-esterified fatty acids may increase diacylglycerol concentrations through de novo pathway by stepwise acylation. In parallel with the increased diacylglycerol, incubation of the cells with saturated non-esterified fatty acids significantly induced the activation of protein kinase C and mitogen-activated protein kinase. The palmitate-induced increase in mitogen-activated protein kinase activity was restored to control concentrations by GF109203X (5 x 10(-7) mol/l), a specific protein kinase C inhibitor, suggesting a protein kinase C-dependent activation of mitogen-activated protein kinase. CONCLUSION/INTERPRETATION: Saturated non-esterified fatty acids induced an increase in de novo diacylglycerol synthesis and subsequent activation of protein kinase C and mitogen-activated protein kinase in cultured aortic smooth muscle cells. This could contribute to the altered vascular functions in the insulin resistant state.

Animals↗
PubMed2005

How are volume-outcome associations related to models of health care funding and delivery? A comparison of the United States and Canada.

How models of health care financing and delivery affect patterns of procedure volumes, outcomes, and volume-outcome associations is not known. We compared volume-outcome studies done in Canada, which provides residents with universal, single-payer health care, with those done in the United States, to determine whether there was a difference in the likelihood of finding statistically significant volume-outcome associations. We analyzed 142 articles, most (90.1%) of which were from the United States. The articles described a total of 291 separate analyses. After adjusting for the clustering of multiple analyses in the same study, the likelihood of finding a statistically significant volume-outcome association was substantially lower in Canadian studies as compared with those from the United States (odds ratio 0.24, 95% confidence interval 0.08 to 0.74, p = 0.01). This result persisted after adjustment for the procedure/condition studied, and the number of study subjects. Canadian volume-outcome analyses are less likely to identify statistically significant volume-outcome associations than US studies, possibly because of the smaller size of some Canadian studies. It is also possible that different models of health care financing and delivery affect patterns of procedure volumes and volume-outcome associations. By promoting competition between hospitals and providers, market-based models may exacerbate existing variations in the quality of hospital care.

Canada↗
PubMed2002

The phylogenetic position of the Theileria buffeli group in relation to other Theileria species.

Theileria parasites known as either T. buffeli, T. orientalis or T. sergenti share many characteristics and are referred to as the T. buffeli group. The 18S ribosomal RNA and merozoite-piroplasm surface protein-encoding genes display a wider genetic variation within the T. buffeli group than between well defined species like T. annulata and T. parva. Analysis of 18S rRNA gene sequences from the database showed that similar groups of related Theileria parasites occur in wild ruminants in Japan and the USA. Moreover, a recently discovered Theileria species pathogenic for small ruminants in China is phylogenetically related to these Japanese Theileria parasites. Phylogenetic analysis of all Theileria 18S rRNA genes reported thus far allowed a subdivision into eight clusters. Some of these clusters contain multiple different species, whereas others appear to contain parasites with similar biological properties whose true speciation remains at present unresolved. The consequences for 18S rRNA based diagnostic assays is discussed.

Animals↗
PubMed2005

Identification and characterization of novel gut-associated lymphoid tissues in rat small intestine.

BACKGROUND: The crypt lamina propria of the mouse small intestine has been shown to harbor multiple tiny clusters filled with c-kit- and interleukin 7 receptor (IL-7R)-positive lympho-hemopoietic cells (cryptopatches; CPs). However, it has remained an open question whether similar lymphoid tissue are present in the gastrointesitinal tract in other animals. In the present study, we investigated whether the small intestine of rats harbored lymphoid tissues similar to mouse CPs. METHODS: Immunohistochemical and flow cytometric analyses were carried out using various antibodies, including those to c-kit and IL-7R molecules. RESULTS: Lymphocyte-filled villi (LFVs), populated predominantly with c-kit- and IL-7 receptor (IL-7R)-positive cells and less with T cell receptor (TCR)-alphabeta T cells were found throughout the small intestine of young adult rats. Although LFVs were absent from fetal rat intestine, they were first detected at around 2 weeks after birth. Notably, in most LFVs that settled in the antimesenteric wall of the small intestine in young adult rats, immunoglobulin M-positive B cells were also detectable at the bottom of the LFVs. In aged rats, lymphocytes in some LFVs displayed a different phenotype, comprising a large B-cell area that included a germinal center. Thus, these clusters represent the first description of isolated lymphoid follicles (ILFs) in the rat small intestine. CONCLUSIONS: The present study provides the first evidence for c-kit- and IL-7R-positive lymphocyte clusters in the rat small intestine. Our data also indicating that LFVs and ILFs may constitute novel organized gut-associated lymphoid tissues in lamina propria of the rat small intestine.

Animals↗
PubMedDate not supplied

Histiocytoid cardiomyopathy: three new cases and a review of the literature.

Histiocytoid cardiomyopathy (HC), a rare arrhythmogenic disorder, presents as difficult-to-control arrhythmias or sudden death in infants and children, particularly girls. Three cases are described with autopsy findings. In two cases, yellow-tan nodules were grossly visible in the myocardium; in the third case, no gross lesions were identified. Microscopic examination in all three cases revealed multiple, scattered clusters of histiocytoid myocytes which on ultrastructural examination were filled with abnormal mitochondria, scattered lipid droplets, and scanty myofibrils. These pathologic findings are similar to those previously described. The pathogenesis of this entity remains controversial. It was recently proposed that this disorder is X-linked dominant with the associated gene located in the region of Xp22.

Arrhythmias, Cardiac↗
PubMed2000

Mechanism of differential effects of antihypertensive agents on serum lipids.

Essential hypertension is frequently associated with the metabolic abnormalities of insulin resistance and dyslipidemia. This prevalent clustering of multiple cardiovascular risk factors may help explain the less-than-expected improvement in coronary heart disease mortality provided by simple blood pressure reduction alone. Many antihypertensive medications effectively reduce blood pressure while providing no benefit or even causing a detrimental effect on the associated metabolic abnormalities. beta-Blockers and diuretics tend to negatively affect both glucose tolerance and plasma lipids. Calcium channel blockers, angiotensin converting enzyme inhibitors, and angiotensin II receptor blockers are most often found to be metabolically neutral. alpha-Blockers provide the most favorable metabolic effects of antihypertensive agents by improving both insulin sensitivity and dyslipidemia. The multiple physiologic mechanisms by which blood pressure medications alter plasma lipids are discussed in detail. The effects of antihypertensive medications on postprandial lipid metabolism and the associated postprandial lipemia-induced endothelial dysfunction deserve special attention.

Adrenergic beta-Antagonists↗
PubMed1989

Low-conductance states of K+ channels in adult mouse skeletal muscle.

Single-channel currents were recorded from Ca2+-activated or ATP-sensitive K+ channels in inside-out membrane patches excised from isolated mouse toe muscles. In addition to the closed and fully open configurations, both types of channels may exhibit several intermediate low-conductance states which are clustered near multiples of elementary conductance units. The units are 1/8 or 1/6 of the channel conductance for Ca2+-activated channels and 1/4 or 1/3 for ATP-sensitive channels. Normally, low-conductance states are rare, but they occur more frequently directly after patch excision. An increased probability of low-conductance states of ATP-sensitive K+ channels was also observed in the presence and during washout of the internal channel blocker adenine. The results suggest that Ca2+-activated and ATP-sensitive K+ channels are composed of several membrane pores with strong positive cooperativity among the elementary conductance units.

Adenosine Triphosphate↗
PubMed1984

Distribution and quantification of ACh receptors and innervation in diaphragm muscle of normal and mdg mouse embryos.

Muscular dysgenesis (mdg) in the mouse is an autosomal recessive mutation, expressed in the homozygous state (in vivo and in vitro) as an absence of skeletal muscle contraction. The distribution of acetylcholine receptors (ACh R) in the diaphragms of phenotypically normal and dysgenic (mdg/mdg) embryos was studied from the 14th to 19th day of gestation by binding of 125I-alpha-bungarotoxin to the muscle, followed by autoradiography of longitudinally sectioned hemidiaphragms and/or of isolated muscle fibers. Localization of ACh R at putative motor end-plate regions begins 14 to 15 days in utero in both normal and dysgenic diaphragms. The distribution of high ACh R density patches is aberrantly scattered beyond the normal innervation pattern in dysgenic diaphragms. Isolated mutant fibers possess (1) multiple ACh R clusters, up to five per single fiber, (2) larger clusters of more variable morphology and variable receptor density than normal clusters, and (3) higher levels of extrajunctional receptors than normal fibers. These autoradiographic results correlate well with higher total level of toxin binding sites per diaphragm and per milligram protein in dysgenic vs normal muscle, as quantified from gamma counting of sucrose density gradient isolation of 125I-toxin-ACh R complexes. The dispersed distribution of ACh R patches on dysgenic muscle may be correlated with extensive phrenic nerve branching as demonstrated by silver impregnation technique. We suggest that the aberrant ACh R cluster distribution is a result of multiple innervation of single fibers from the branched nerve terminals. Possible causes of the excessive nerve branching in the mutant are discussed in light of generalized nerve sprouting found in paralyzed muscle.

Animals↗
PubMed1984

XANES study of iron displacement in the haem of myoglobin.

The XANES (X-ray absorption near edge structure) spectra of deoxy human adult haemoglobin (HbA) and myoglobin (Mb) have been measured at the wiggler beam line of the Frascati synchrotron radiation facility. The XANES are interpreted by the multiple scattering cluster theory. The variations in the XANES between HbA and Mb are assigned to changes in the Fe-porphyrin geometry.

Animals↗