Thoracic endovascular aortic repair for aortic complications after esophagectomy for cancer: report of three cases.

Aortic complications after esophageal cancer surgery are rare and usually fatal. Here, we report three patients who underwent thoracic endovascular aortic repair (TEVAR) for aortic complications after esophagectomy for cancer. In the first case, aortic rupture was caused by pyothorax due to residual tumor after esophagectomy. In the second case, aortic rupture was caused by pyothorax due to anastomotic leakage. In the third case, a pseudoaneurysm was caused by surgical injury during esophagectomy. TEVAR was safe and effective for severe aortic complications when graft infection was avoided. The first case died of sepsis on the 84th postoperative day, and the other two cases have survived 4 years and 2 years to date.

Interaction of Rhei Rhizoma extract with cytochrome P450 3A and efflux transporters in rats.

Traditional Chinese herbal medicines are frequently prescribed in pharmacotherapy in Japan. In the present study, we evaluated the possible interaction of several herbal extracts including Rhei Rhizoma extract with cytochrome P450 (CYP) 3A and efflux transporters such as P-glycoprotein and multidrug resistance-associated protein (MRP) 2. Rhei Rhizoma extract (100 microg/ml) significantly suppressed the CYP3A-mediated 6beta-hydroxylation of testosterone in hepatic microsomes, and increased the extent of bioavailability of midazolam, a typical CYP3A substrate, in rats. Also, Rhei Rhizoma extract (300 microg/ml) significantly suppressed P-glycoprotein-mediated efflux transport of rhodamine 123 (Rho123) in rat everted intestine. In an in-vivo study, Rhei Rhizoma extract added to intestinal perfusate at a concentration of 300 microg/ml significantly suppressed the intestinal exsorption of Rho123, though it exerted no effect on the biliary excretion of Rho123. Furthermore, the in-vitro and in-vivo MRP2-mediated intestinal efflux of 2,4-dinitrophenyl-S-glutathione was significantly suppressed by Rhei Rhizoma extract (1000 microg/ml). In conclusion, Rhei Rhizoma extract, which is taken orally at doses of 0.5-1 g each or 1-3 g daily in clinical practice, may cause pharmacokinetic herb-drug interactions in the process of the intestinal and/or hepatic CYP3A-mediated drug metabolism and P-glycoprotein- and/or MRP2-mediated efflux transport in the intestine.

Absence of polymorphism at the ZFY locus on the human Y chromosome.

DNA polymorphism in the Y chromosome, examined at a 729-base pair intron located immediately upstream of the ZFY zinc-finger exon, revealed no sequence variation in a worldwide sample of 38 human males. This finding cannot be explained by global constraint on the intron sequence, because interspecific comparisons with other nonhuman primates revealed phylogenetically informative sequence changes. The invariance likely results from either a recent selective sweep, a recent origin for modern Homo sapiens, recurrent male population bottlenecks, or historically small effective male population sizes. A coalescence model predicts an expected time to a most recent common ancestral male lineage of 270,000 years (95 percent confidence limits: 0 to 800,000 years).

[Long-term results of aortic root replacement with a composite graft].

We analyzed the long-term results of aortic root replacement with a composite graft. Since 1992, 127 patients had undergone aortic root replacement with a composite graft. There were 92 men and 35 women with a mean age of 56.5 years. There were 69 patients with annuloaortic ectasia, 17 aortic dissections, and 41 ascending aortic dilatation due to aortic valve disease. Marfan syndrome was diagnosed in 19 patients. As surgical procedure, button technique was used in 90 patients, Cabrol technique in 11, and Piehler technique in 26. Open distal anastomosis was performed in 82 patients to avoid clamp injury of rest aorta. Early mortality was 3.1% and no major morbid events had occurred. Follow-up was completed in 95.9% of the patients and the mean follow-up period was 6.1 years. Actuarial survival at 5, 10, and 15 years was 86.2%, 83.4%, and 83.4%, respectively. Actuarial freedom from aortic valve reoperation at 10 and 15 years was 99.2% and 95.7%, respectively. The results of aortic root replacement with a composite graft were excellent. This procedure should be the 1st choice for surgical treatment of the aortic root disease.

Gene expression and molecular evolution.

The combination of complete genome sequence information and estimates of mRNA abundances have begun to reveal causes of both silent and protein sequence evolution. Translational selection appears to explain patterns of synonymous codon usage in many prokaryotes as well as a number of eukaryotic model organisms (with the notable exception of vertebrates). Relationships between gene length and codon usage bias, however, remain unexplained. Intriguing correlations between expression patterns and protein divergence suggest some general mechanisms underlying protein evolution.

Translational selection and molecular evolution.

An interplay among experimental studies of protein synthesis, evolutionary theory, and comparisons of DNA sequence data has shed light on the roles of natural selection and genetic drift in 'silent' DNA evolution.

Genetic recombination at different points in the Npro-coding region of bovine viral diarrhea viruses and the potentials to change their antigenicities and pathogenicities.

Cytopathogenic (cp) bovine viral diarrhea virus (BVDV) strain KS86-1 cp was isolated from a cow persistently infected with non-cytopathogenic (ncp) BVDV strain KS86-ncp after development of mucosal disease by superinfection with cp BVDV strain Nose. cp BVDV strains 799cp and 839cp were also isolated from independent cattle that developed mucosal disease by superinfection with cp BVDV KS86-1cp. In the present study, genetic analysis revealed that the genes of cp BVDV strains 799cp and 839cp were chimeras between the genes of the persisting ncp BVDVs and that of superinfecting KS86-1cp. The genetic recombination that generates 799cp occurred between the identical points in the N(pro) gene region, whereas genetic recombination that generates 839cp occurred between different points in the N(pro) gene region. Both 799cp and 839cp were inherited Jiv gene of KS86-1cp strain and envelope protein genes of the persisting viruses. In addition, neutralization test disclosed that antigenicities of 799cp, 839cp, and KS86-1cp were also similar to each persisting virus. These findings indicate that exogenous cp BVDV containing insertion of Jiv gene in the 5 terminal region can induce genetic recombination with the original ncp BVDV at different points in the N(pro) gene region, and those viruses have high potential to change those antigenicities and pathogenicities by RNA recombination.

Screening and determination of gene function using randomized ribozyme and siRNA libraries.

Rapid progress in the sequencing of the genomes of model organisms, such as the mouse, rat, nematode, fly, and Arabidopsis, as well as the human genome, has provided abundant sequence information, but functions of long stretches of these genomes remain to be determined. RNA-based technologies hold promise as tools that allow us to identify the specific functions of portions of these genomes. In particular, catalytic RNAs, known also as ribozymes, can be engineered for optimization of their activities in the intracellular environment. The introduction of a library of active ribozymes into cells, with subsequent screening for phenotypic changes, can be used for the rapid identification ofa gene function. Ribozyme technology complements another RNA-based tool for the determination of gene function, which is based on libraries of small interfering RNAs (siRNAs).

Enhancement of apoptosis via an extrinsic factor, TNF-alpha, in cells infected with cytopathic bovine viral diarrhea virus.

Isolates of bovine viral diarrhea virus (BVDV) are divided into cytopathic (cp) and noncytopathic (ncp) biotypes according to their effect on cultured cells. Calves persistently infected with ncp BVDV are known to develop lethal mucosal disease (MD) after superinfection by cp BVDV. Although the UV-irradiated supernatant of cp BVDV-infected cells has been reported to have no capacity to induce cell death, we found that it could enhance cell death through apoptosis. Up-regulation of tumor necrosis factor alpha (TNF-alpha) and inducible nitric oxide synthase (iNOS) mRNAs was detected specifically in cp BVDV-infected primary cell cultures. Suppression of TNF-alpha via antisense oligonucleotide transfection or incubation with a polyclonal antibody against TNF-alpha resulted in attenuation of apoptosis induced by cp BVDV, suggesting that TNF-alpha participates in apoptosis execution. Although TNF-alpha is one of the iNOS-inducible factors, the iNOS up-regulation was not regulated by TNF-alpha. And iNOS was revealed to serve as anti-apoptotic factor, contrary to our expectation. In addition, the expression level of both TNF-alpha and iNOS mRNAs in the ncp BVDV-infected cells was kept lower than that in the mock-infected cells, suggesting that ncp BVDV reduced or interfered with the factor triggering the expression of both mRNAs. These characteristic mRNA transcriptions would help to explain why BVDV acts differently in cells as well as in vivo, depending on its biotype. To elucidate viral factors inducing TNF-alpha and iNOS may be critical to understand the mechanism of MD development, which closely correlates with cp BVDV-induced apoptosis.

Synonymous codon usage in Drosophila melanogaster: natural selection and translational accuracy.

I present evidence that natural selection biases synonymous codon usage to enhance the accuracy of protein synthesis in Drosophila melanogaster. Since the fitness cost of a translational misincorporation will depend on how the amino acid substitution affects protein function, selection for translational accuracy predicts an association between codon usage in DNA and functional constraint at the protein level. The frequency of preferred codons is significantly higher at codons conserved for amino acids than at nonconserved codons in 38 genes compared between D. melanogaster and Drosophila virilis or Drosophila pseudoobscura (Z = 5.93, P < 10(-6)). Preferred codon usage is also significantly higher in putative zinc-finger and homeodomain regions than in the rest of 28 D. melanogaster transcription factor encoding genes (Z = 8.38, P < 10(-6)). Mutational alternatives (within-gene differences in mutation rates, amino acid changes altering codon preference states, and doublet mutations at adjacent bases) do not appear to explain this association between synonymous codon usage and amino acid constraint.

Inferring weak selection from patterns of polymorphism and divergence at "silent" sites in Drosophila DNA.

Patterns of codon usage and "silent" DNA divergence suggest that natural selection discriminates among synonymous codons in Drosophila. "Preferred" codons are consistently found in higher frequencies within their synonymous families in Drosophila melanogaster genes. This suggests a simple model of silent DNA evolution where natural selection favors mutations from unpreferred to preferred codons (preferred changes). Changes in the opposite direction, from preferred to unpreferred synonymous codons (unpreferred changes), are selected against. Here, selection on synonymous DNA mutations is investigated by comparing the evolutionary dynamics of these two categories of silent DNA changes. Sequences from outgroups are used to determine the direction of synonymous DNA changes within and between D. melanogaster and Drosophila simulans for five genes. Population genetics theory shows that differences in the fitness effect of mutations can be inferred from the comparison of ratios of polymorphism to divergence. Unpreferred changes show a significantly higher ratio of polymorphism to divergence than preferred changes in the D. simulans lineage, confirming the action of selection at silent sites. An excess of unpreferred fixations in 28 genes suggests a relaxation of selection on synonymous mutations in D. melanogaster. Estimates of selection coefficients for synonymous mutations (3.6 < magnitude of Nes < 1.3) in D. simulans are consistent with the reduced efficacy of natural selection (magnitude of Nes < 1) in the three- to sixfold smaller effective population size of D. melanogaster. Synonymous DNA changes appear to be a prevalent class of weakly selected mutations in Drosophila.

Inferring the fitness effects of DNA mutations from polymorphism and divergence data: statistical power to detect directional selection under stationarity and free recombination.

The fitness effects of classes of DNA mutations can be inferred from patterns of nucleotide variation. A number of studies have attributed differences in levels of polymorphism and divergence between silent and replacement mutations to the action of natural selection. Here, I investigate the statistical power to detect directional selection through contrasts of DNA variation among functional categories of mutations. A variety of statistical approaches are applied to DNA data simulated under Sawyer and Hartl's Poisson random field model. Under assumptions of free recombination and stationarity, comparisons that include both the frequency distributions of mutations segregating within populations and the numbers of mutations fixed between populations have substantial power to detect even very weak selection. Frequency distribution and divergence tests are applied to silent and replacement mutations among five alleles of each of eight Drosophila simulans genes. Putatively "preferred" silent mutations segregate at higher frequencies and are more often fixed between species than "unpreferred" silent changes, suggesting fitness differences among synonymous codons. Amino acid changes tend to be either rare polymorphisms or fixed differences, consistent with a combination of deleterious and adaptive protein evolution. In these data, a substantial fraction of both silent and replacement DNA mutations appear to affect fitness.

Molecular evolution between Drosophila melanogaster and D. simulans: reduced codon bias, faster rates of amino acid substitution, and larger proteins in D. melanogaster.

Both natural selection and mutational biases contribute to variation in codon usage bias within Drosophila species. This study addresses the cause of codon bias differences between the sibling species, Drosophila melanogaster and D. simulans. Under a model of mutation-selection-drift, variation in mutational processes between species predicts greater base composition differences in neutrally evolving regions than in highly biased genes. Variation in selection intensity, however, predicts larger base composition differences in highly biased loci. Greater differences in the G+C content of 34 coding regions than 46 intron sequences between D. melanogaster and D. simulans suggest that D. melanogaster has undergone a reduction in selection intensity for codon bias. Computer simulations suggest at least a fivefold reduction in Nes at silent sites in this lineage. Other classes of molecular change show lineage effects between these species. Rates of amino acid substitution are higher in the D. melanogaster lineage than in D. simulans in 14 genes for which outgroup sequences are available. Surprisingly, protein sizes are larger in D. melanogaster than in D. simulans in the 34 genes compared between the two species. A substantial fraction of silent, replacement, and insertion/deletion mutations in coding regions may be weakly selected in Drosophila.

Natural selection and the frequency distributions of "silent" DNA polymorphism in Drosophila.

In Escherichia coli, Saccharomyces cerevisiae, and Drosophila melanogaster, codon bias may be maintained by a balance among mutation pressure, genetic drift, and natural selection favoring translationally superior codons. Under such an evolutionary model, silent mutations fall into two fitness categories: preferred mutations that increase codon bias and unpreferred changes in the opposite direction. This prediction can be tested by comparing the frequency spectra of synonymous changes segregating within populations; natural selection will elevate the frequencies of advantageous mutations relative to that of deleterious changes. The frequency distributions of preferred and unpreferred mutations differ in the predicted direction among 99 alleles of two D. pseudoobscura genes and five alleles of eight D. simulans genes. This result confirms the existence of fitness classes of silent mutations. Maximum likelihood estimates suggest that selection intensity at silent sites is, on average, very weak in both D. pseudoobscura and D. simulans (magnitude of NS approximately 1). Inference of evolutionary processes from within-species sequence variation is often hindered by the assumption of a stationary frequency distribution. This assumption can be avoided when identifying the action of selection and tested when estimating selection intensity.

Histopathological Studies on the Neuropathogenicity of the Iriki and OBE-1 Strains of Akabane Virus in BALB/cAJcl Mice.

The OBE-1 strain of Akabane virus infects the fetus via the dam, resulting in abortion or congenital abnormalities in ruminants. In contrast, the Iriki strain of Akabane virus is highly virulent and causes encephalomyelitis by post-natal infection. To clarify the difference in pathogenicity between the two strains, BALB/cAJcl mice were inoculated either intraperitoneally or intracerebrally (IC) with either strain from 3 days to 8 weeks of age. Pathological examination revealed non-suppurative encephalitis in mice inoculated by either route with the Iriki strain. Virus antigens were distributed widely throughout the brain when the virus was inoculated into newborn mice, but distribution was limited to the brainstem in mice inoculated when 8 weeks old. However, brain lesions were observed only in mice inoculated with OBE-1 by the IC route when the mice were 3 days old, but these lesions were mild. To examine the manner of viral spreading, the Iriki strain was inoculated IC or intrastriatally into 8-week-old mice. Viral antigens were distributed prominently throughout the spinal cord as well as the brainstem and various cerebral nuclei, and were present with less prominence in the connective fibres. Virus antigens were also distributed in the subventricular zone, where neuronal stem cells exist. These results show that the neuroinvasiveness of the Iriki strain diminishes with age, while neurovirulence is maintained; however, for the OBE-1 strain both neuroinvasiveness and neurovirulence diminish with age. Furthermore, Akabane virus infects neuronal cells in the brainstem and spreads to the spinal cord via an unidentified transneuronal pathway.

Within- and between-species DNA sequence variation and the 'footprint' of natural selection.

Extensive DNA data emerging from genome-sequencing projects have revitalized interest in the mechanisms of molecular evolution. Although the contribution of natural selection at the molecular level has been debated for over 30 years, the relevant data and appropriate statistical methods to address this issue have only begun to emerge. This paper will first present the predominant models of neutral, nearly neutral, and adaptive molecular evolution. Then, a method to identify the role of natural selection in molecular evolution by comparing within- and between-species DNA sequence variation will be presented. Computer simulations show that such methods are powerful for detecting even very weak selection. Examination of DNA variation data within and between Drosophila species suggests that 'silent' sites evolve under a balance between weak selection and genetic drift. Simulated data also show that sequence comparisons are a powerful method to detect adaptive protein evolution, even when selection is weak or affects a small fraction of nucleotide sites. In the Drosophila data examined, positive selection appears to be a predominant force in protein evolution.

Codon bias evolution in Drosophila. Population genetics of mutation-selection drift.

Although non-random patterns of synonymous codon usage are a prominent feature in the genomes of many organisms, the relatives roles of mutational biases and natural selection in maintaining codon bias remain a contentious issue. In some species, patterns of codon bias and empirical findings on the biology of translation suggest 'major codon preference', a balance among mutation pressure, genetic drift, and weak selection in favor of translationally superior codons. Population genetics theory makes testable predictions to distinguish such a model from a strictly mutational model of codon bias. Major codon preference predicts two fitness classes of synonymous DNA changes: 'preferred' mutations from non-major to major codons and 'unpreferred' changes in the opposite direction. An extension of current statistical methods is employed to reveal differences in the within and between species dynamics of preferred and unpreferred silent mutations in Drosophila simulans. In this lineage, codon bias appears to be maintained under roughly equal magnitudes of natural selection and genetic drift. In the sibling species, D. melanogaster, however, a reduction in N(e)s, the product of effective population size and selection coefficient, appears to have allowed a genome-wide reduction in codon bias.

A test of translational selection at 'silent' sites in the human genome: base composition comparisons in alternatively spliced genes.

Natural selection appears to discriminate among synonymous codons to enhance translational efficiency in a wide range of prokaryotes and eukaryotes. Codon bias is strongly related to gene expression levels in these species. In addition, between-gene variation in silent DNA divergence is inversely correlated with codon bias. However, in mammals, between-gene comparisons are complicated by distinctive nucleotide-content bias (isochores) throughout the genome. In this study, we attempted to identify translational selection by analyzing the DNA sequences of alternatively spliced genes in humans and in Drosophila melanogaster. Among codons in an alternatively spliced gene, those in constitutively expressed exons are translated more often than those in alternatively spliced exons. Thus, translational selection should act more strongly to bias codon usage and reduce silent divergence in constitutive than in alternative exons. By controlling for regional forces affecting base-composition evolution, this within-gene comparison makes it possible to detect codon selection at synonymous sites in mammals. We found that GC-ending codons are more abundant in constitutive than alternatively spliced exons in both Drosophila and humans. Contrary to our expectation, however, silent DNA divergence between mammalian species is higher in constitutive than in alternative exons.

Antigenic diversity of Akabane virus detected by monoclonal antibodies.

The antigenic properties of 21 Japanese field isolates and two Australian strains of Akabane (AKA) virus (Simbu serogroup, bunyavirus) isolated from 1959 to 1990 were compared by enzyme-linked immunosorbent assay (ELISA), plaque-reduction neutralization (PRNT) and hemagglutination inhibition (HI) tests using monoclonal antibodies (Mabs) to the OBE-1 strain of AKA virus. Sixteen Mabs were established by fusing P3X63Ag8U1 mouse myeloma cells and spleen cells from BALB/c mice immunized with the OBE-1 strain. Of the 16 clones, 13 produced immunoglobulin (Ig) which precipitated glycoprotein G1 and three produced Ig which precipitated nucleoprotein (N). Twelve out of 13 Mabs had both NT and HI activities to not only the homologous OBE-1 strain but also the other isolates. By the competitive binding assay, at least five antigenic regions for G1, and two for N were defined. Some of the anti-G1 Mabs which reacted to the same antigenic region had unique reactivity while anti-N Mabs recognizing the same epitope reacted with almost the same degree to all of the isolates. Finally, nine epitopes of the G1 protein in five different antigenic regions have been identified. There was no striking correlation between isolation date and place of the isolates and their reactivity to Mabs. A most interesting result is that three isolates collected in the same place over a three week period had different reactivity patterns detected by ELISA, showing great antigenic variation of the virus. AKA virus may be a single gene pool consisting of different genotypes in the field.

Antigenic and genetic comparisons of Japanese and Australian Simbu serogroup viruses: evidence for the recovery of natural virus reassortants.

The antigenicity and RNA genome structures of five Simbu serogroup bunyaviruses isolated in Japan and Australia were analyzed using monoclonal antibodies (Mabs) raised to Akabane (AKA) virus and oligonucleotide fingerprinting. The virion surface glycoprotein (G1) and the nucleocapsid (N) protein of heterologous viruses showed no reactivity to the Mabs, while the AKA-derived anti-G1 Mab (2F1) reacted with Peaton virus and all three AKA anti-N Mabs reacted with Tinaroo (TIN) virus at almost the same antibody titers as the homologous virus. Oligonucleotide fingerprinting analyses indicated that the three RNA species of all the viruses were unique and distinguishable. However, AKA and TIN viruses exhibited very similar S RNA oligonucleotide fingerprints, while the L and M RNA fingerprints were quite different. The S RNA sequence of TIN virus has been determined and compared with that of AKA and Aino viruses. The results revealed 95.1% S sequence homology between the AKA and TIN viruses. The antigenic and genetic comparisons of AKA and TIN viruses suggest that the two viruses may represent naturally occurring reassortant viruses.