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A comprehensive understanding of how Staphylococcus aureusadapts to cause infections in humans can inform development of diagnostic, therapeutic, and preventive approaches. Expression analysis of clinical strain libraries depicts in vitroconditions that differ from those in human infection, but low bacterial burden and the requirement for reverse transcription or nucleic acid amplification complicate such analyses of bacteria causing human infection.

Reactivation of telomerase reverse transcriptase (TERT) expression is found in more than 85% of human cancers. The remaining cancers rely on the alternative lengthening of telomeres (ALT), a recombination-based mechanism for telomere-length maintenance. Prevalence of TERT reactivation over the ALT mechanism was linked to secondary TERT function unrelated to telomere length maintenance. To characterize this non-canonical function, we created a panel of ALT cells with recombinant expression of TERT and TERT variants: TERT-positive ALT cells showed higher tolerance to genotoxic insults compared with their TERT-negative counterparts. We identified telomere synthesis-defective TERT variants that bestowed similar genotoxic stress tolerance, indicating that telomere synthesis activity is dispensable for this survival phenotype. TERT expression improved the kinetics of double-strand chromosome break repair and reduced DNA damage-related nuclear division abnormalities, a phenotype associated with ALT tumors. Despite this reduction in cytological abnormalities, surviving TERT-positive ALT cells were found to have gross chromosomal instabilities. We sorted TERT-positive cells with cytogenetic changes and followed their growth. We found that the chromosome-number changes persisted, and TERT-positive ALT cells surviving genotoxic events propagated through subsequent generations with new chromosome numbers. Our data confirm that telomerase expression protects against double-strand DNA (dsDNA)-damaging events, and show that this protective function is uncoupled from its role in telomere synthesis. TERT expression promotes oncogene-transformed cell growth by reducing the inhibitory effects of cell-intrinsic (telomere attrition) and cell-extrinsic (chemical- or metabolism-induced genotoxic stress) challenges. These data provide the impetus to develop new therapeutic interventions for telomerase-positive cancers through simultaneous targeting of multiple telomerase activities.

A condition-based maintenance optimization approach is developed for the road-cracking problem in order to derive optimal maintenance policies that minimize a total discounted maintenance cost. The approach is based on a Markov decision process that takes into account multiple actions with varying effects on future road performance. Maintaining the road consists of adding a new asphalt layer; however, as resurfacing actions are constrained by a maximum total road thickness, the maintenance decision is not only how thick a layer to apply, but also how much old road to remove. Each combination of these actions leads to different maintenance costs and different future degradation behaviours. The road state is modelled by a dependent bivariate deterioration variable (the longitudinal cracking percentage and the deterioration growth rate), for taking these different changes in the cracking patterns into account. Moreover, the sensitivity to cracking for existing roads can be reduced with the addition of new layers, and thus actions that can lead to states better than good-as-new have to be considered. A numerical analysis is provided to illustrate the benefits of the introduction of the deterioration speed in the decision framework, as well as the belief that initially building a road to its maximum thickness is not optimal. The trade-offs in the design decisions and the exploitation/maintenance costs are also explored.

The formulation of a partially observed Markov decision process (POMDP) model to adaptively schedule testing, minimal repairs and overhauls for a two-state process with age-dependent degradation is presented. Structurally, the optimal maintenance policy is of control-limit type with respect to the repair and overhaul actions in both age and the probability of being out-of-control. A tailored solution algorithm is developed that iteratively determines an upper bound on the truncation age under the optimal policy. Numerical examples highlight the flexibility of the model, possible complexities of the optimal policy, and cost savings in comparison with less flexible models.

It has long been puzzling regarding the physical origin of the thermally induced red-shift of Raman optical phonons in group III nitride crystals despite some possible mechanisms such as phonon decay, thermal expansion, interfacial effects, or their combinations. Here we show that an extension of the recent approach Sun, C. Q.; Pan, L. K.; Li, C. M.; Li, S. Phys. Rev.B2005, 72, 134301 to the functional dependence of the frequency of Raman optical modes on the atomic bonding identities (bond length, bond strength, and atomic coordination numbers) to the temperature domain has enabled us to gain a simple solution with improved understanding of the thermally induced red-shift of Raman optical phonons. Reproduction of the measured temperature dependence of the Raman shift of AlN, GaN, and InN reveals that the thermally driven red-shift arises simply from bond vibration and bond expansion, together with derived information about the modecohesive energy.

Epidermal growth factor (EGF) stimulates the homodimerization of EGF receptor (EGFR) and the heterodimerization of EGFR and ErbB2. The EGFR homodimers are quickly endocytosed after EGF stimulation as a means of down-regulation. However, the results from experiments on the ability of ErbB2 to undergo ligand-induced endocytosis are very controversial. It is unclear how the EGFR-ErbB2 heterodimers might behave. In this research, we showed by subcellular fractionation, immunoprecipitation, Western blotting, indirect immunofluorescence, and microinjection that, in the four breast cancer cell lines MDA453, SKBR3, BT474, and BT20, the EGFR-ErbB2 heterodimerization levels were positively correlated with the ratio of ErbB2/EGFR expression levels. ErbB2 was not endocytosed in response to EGF stimulation. Moreover, in MDA453, SKBR3, and BT474 cells, which have very high levels of EGFR-ErbB2 heterodimerization, EGF-induced EGFR endocytosis was greatly inhibited compared with that in BT20 cells, which have a very low level of EGFR-ErbB2 heterodimerization. Microinjection of an ErbB2 expression plasmid into BT20 cells significantly inhibited EGF-stimulated EGFR endocytosis. Coexpression of ErbB2 with EGFR in 293T cells also significantly inhibited EGF-stimulated EGFR endocytosis. EGF did not stimulate the endocytosis of ectopically expressed ErbB2 in BT20 and 293T cells. These results indicate that ErbB2 and the EGFR-ErbB2 heterodimers are impaired in EGF-induced endocytosis. Moreover, when expressed in BT20 cells by microinjection, a chimeric receptor composed of the ErbB2 extracellular domain and the EGFR intracellular domain underwent normal endocytosis in response to EGF, and this chimera did not block EGF-induced EGFR endocytosis. Thus, the endocytosis deficiency of ErbB2 is due to the sequence of its intracellular domain.

SEC13 encodes a 33 kDa protein that participates in vesicle budding from the endoplasmic reticulum (ER). In order to purify a functional form of Sec13p, a SEC13‐dihydrofolate reductase (mouse) fusion gene (SEC13:DHFR) was constructed that complements both sec13 temperature sensitive and null mutations. Methotrexate‐agarose affinity chromatography facilitated the purification of two forms of the Sec13‐dhfrp fusion protein: a monomeric form and a high molecular weight complex. The complex form consists of two subunits: Sec13‐dhfrp and a 150 kDa protein (p150). Native immunoprecipitation experiments confirm that Sec13p exists in a complex with p150 in wild type cells. Functional analysis supports a role for both subunits in protein transport. Vesicle budding from the ER in a cell‐free reaction is inhibited by Fab antibody fragments directed against either Sec13p or p150. The purified Sec13‐dhfrp/p150 complex, but not the Sec13‐dhfrp monomer, in combination with two other pure protein fractions (Sar1p and a Sec23/Sec24 protein complex) satisfies the requirement for cytosol in a cell‐free vesicle budding reaction. The vesicles formed with the purified protein fractions are competent to fuse with the Golgi and are biochemically distinct from the ER membrane fraction from which they derive.

The Escherichia coli RecR protein participates in a recombinational DNA repair process. Its gene is located in a region of chromosome that extends from 502 to 509 kilobases on the physical map and that contains apt, dnaX, orf12-recR, htpG, and adk. Most, if not all, of these are involved in nucleic acid metabolism. The orf12-recR reading frames consist of 935 base pairs and overlap by one nucleotide, with the 3' A of the orf12 termination codon forming the 5' nucleotide of the recR initiation codon. The orf12-recR promoter was located upstream of orf12 by sequence analysis, promoter cloning, and S1 nuclease protection analysis. The start point of transcription was determined by primer extension. The transcript 5' end contained a long, apparently untranslated region of 199 nucleotides. Absence of a detectable promoter specific for recR and the overlap of the orf12 and recR reading frames suggest that translation of recR is coupled to that of orf12. By maxicell analysis, it was determined that both orf12 and recR are translated.

SEC13 encodes a 33 kDa protein that participates in vesicle budding from the endoplasmic reticulum (ER). In order to purify a functional form of Sec13p, a SEC13‐dihydrofolate reductase (mouse) fusion gene (SEC13:DHFR) was constructed that complements both sec13 temperature sensitive and null mutations. Methotrexate‐agarose affinity chromatography facilitated the purification of two forms of the Sec13‐dhfrp fusion protein: a monomeric form and a high molecular weight complex. The complex form consists of two subunits: Sec13‐dhfrp and a 150 kDa protein (p150). Native immunoprecipitation experiments confirm that Sec13p exists in a complex with p150 in wild type cells. Functional analysis supports a role for both subunits in protein transport. Vesicle budding from the ER in a cell‐free reaction is inhibited by Fab antibody fragments directed against either Sec13p or p150. The purified Sec13‐dhfrp/p150 complex, but not the Sec13‐dhfrp monomer, in combination with two other pure protein fractions (Sar1p and a Sec23/Sec24 protein complex) satisfies the requirement for cytosol in a cell‐free vesicle budding reaction. The vesicles formed with the purified protein fractions are competent to fuse with the Golgi and are biochemically distinct from the ER membrane fraction from which they derive.

Summary A prospective observational study of the pathophysiology of sodium and water disorders in patients with pituitary region tumours after surgical excision was carried out in 20 patients. Serial pre-operative and post-operative fluid and sodium balance, plasma and urine elctrolyte biochemistry and their derived parameters, and circulating hormones associated with fluid balance, atrial natriureic peptide (ANP) and antidiuretic hormone (ADH) were documented to correlate with the patients' clinical conditions. Ten out of these twenty cases developed diabetes insipidus (DI) requiring ADH replacement therapy, although in the majority (6 cases), this way only a transient event. Of the nine patients who developed hyponatraemia, six had symptoms such as impaired consciousness and convulsions. Four patients developed alternating hypoatraemia and hypernatraemia, which constituted a difficult group, where appropriate sodium and fluid management, and ADH replacement therapy were based upon twice daily plasma and urine biochemistry and their derived parameters. Whilst DI in this group of patients was the result of a low circulating ADH level, hyponatraemia was not associated with an exaggerated ADH activity (6.0 ± 2.3 vs 7.4 ± 2.3 pmol/ml, mean ± SEM). Rather, hyponatraemia was strongly associated with an elevated circulating ANP concentration (82.4±10.5 vs 30.0 ±3.1 pmol/ml, mean ± SEM, p < 0.001), resulting in salt wasting and hypovolaemia.

A histological assessment has been made of both time- and dose-related changes in the choroid plexus after the local irradiation of the rat brain with single doses of 17.5–25 Gy of X rays. These investigations involved the serial killing of animals 1–52 weeks after irradiation and the quantitative and semiquantitative evaluation of histological sections. Counts of the relative number of cells in the choroid plexus of the lateral ventricles showed an atrophy of the epithelial layer after 13 weeks. However, this was not as marked as the reduction in the number of endothelial cells in the wall of blood vessels. Moreover, the epithelium had recovered by 39 weeks after irradiation, while the dose-related depletion in endothelial cells tended to be progressive. A highly correlated group of changes in the vascular-connective tissue was used to produce a numerical “factor”. This represented a combined score of radiation damage which was both time- and dose-related. These data suggest that, as an expression of late radiation damage to the choroid plexus, the effect on the endothelium was more important than that to the epithelial cells.

Following the local irradiation of the rat brain with single doses of 17.5–25 Gy of X rays, necrosis of the white matter was seen after a latent interval of > 26 weeks. At 39 weeks and 52 weeks after irradiation the incidence of necrosis was doserelated. The doses associated with a 50% incidence of necrosis in the white matter (ED50) at these times were 23.45 ± 0.49 and 20.98 ± 0.91 Gy, respectively. At both these times the incidence of necrosis was higher in the fimbria than in the capsula interna and the corpus callosum. This reflects a variation in the latency time for the appearance of necrosis. Necrosis occurs earlier in the fimbria. In the corpus callosum and the capsula interna the latency times for the appearance of necrosis were also dosedependent. In the latent period prior to the onset of necrosis of the fimbria, a number of dose-related changes were seen in the vasculature and the associated astroglial cells. These changes, which included blood vessel dilation, blood vessel wall thickening, endothelial cell nuclear enlargement and the hypertrophy of perivascular astrocytes, were highly correlated and when combined appeared to represent a “unit of tissue injury”. The incidence and severity of this “unit of tissue injury” apparently increased with time after irradiation until necrosis ensued. These dose-related vascular/glial changes were preceded by a reduction in the endothelial cell and vascular density. No early changes were seen in the number of glial parenchymal cells.

The toxicity of single doses of the anthracycline epirubicin was compared in the rat after either the intravenous (i.v.: 2–6 mg/kg) or intraperitoneal (i.p.; 4–8 mg/kg) administration of the drug. These doses produced comparable acute toxicity that was characterised by a dose-dependent, transient reduction in body weight (<15%) in the first 2 weeks after drug administration. Sequential measurements of cardiac output in animals that received i.v. doses of epirubicin showed that the time-related changes in cardiac function were biphasic. There was an initial decline in cardiac output in the first 12 weeks, which was followed by a phase of persistent depression in cardiac output between 12 weeks and 20 weeks. The time-related changes in cardiac function after i.p. doses of the drug were more variable, although the trend of changes, as after i.v. administration, appeared to be dose-dependent. Recovery of cardiac function occurred at 20 weeks after an i.p. dose of 4 mg/kg; however, after 6 mg/kg, cardiac function was significantly depressed after ?16 weeks. For both routes of administration, the likelihood of late cardiac complications was dependent on the level of the reduction in cardiac output at 12 weeks. A study of the impairment of cardiac output and the incidence of cardiac-related mortality demonstrated that epirubicin was more cardiotoxic after i.v. administration. Equivalent cardiotoxic doses of epirubicin after i.v. and i.p. administration were highly linearly correlated (r=0.998), although there appeared to be a threshold dose of 3.33 mg/kg after i.p. administration of the drug. Thus, the relative cardiotoxicity between the two routes of administration was found to be dependent on the drug dose and, hence, the level of effect. The difference in the effect was less for high drug doses. The LD50 for deaths due to cardiotoxicity at 20 weeks was 4.42±0.42 mg/kg after i.v. administration, which was significantly lower than the value of 6.28±0.41 mg/kg obtained after i.p. administration of the drug (P<0.01). There was no qualitative difference in the histological lesions induced in the myocardium after the i.v. vs i.p. administration of epirubicin.

It has been proposed that one of metronidazole's partially reduced intermediates interacts either with DNA to exert a bactericidal effect or with water to form acetamide. To test this hypothesis we have examined the effect of metronidazole on several mutants of Escherichia coli that are defective in DNA repair. UV-susceptible RecA- and UvrB- point mutants have an increased susceptibility to metronidazole as manifested by both a decreased minimal inhibitory concentration and a greater bactericidal response to metronidazole in resting cultures. By these criteria, however, we find that UvrB- deletion mutants, which lack the ability to reduce nitrate and chlorate, are no more susceptible to metronidazole than is the wild type. We find, however, that these deletion mutants also lack the ability to reduce metronidazole and thus possibly to form its reactive species. When metronidazole's bactericidal effect is expressed in terms of the concurrent accumulation of acetamide derived from metronidazole, then all RecA- and UvrB- mutants are killed more efficiently than their wild types. The data are consistent, therefore, with metronidazole's lethal effect being mediated by a partially reduced intermediate on the metabolic pathway between metronidazole and acetamide. Defects in other aspects of the DNA repair system do not confer this increased susceptibility to the proposed intermediate. A Tag- mutant, for example, which is defective in 3-methyl-adenine-DNA glycosylase, does not have this increased susceptibility to the presumed precursor of acetamide. Thus, these results provide further support for the hypothesis that the bactericidal effect of metronidazole is mediated by a partially reduced intermediate in the metabolic conversion of metronidazole to acetamide and suggest that this intermediate interacts with DNA to produce a lesion similar to that caused by UV light.

The charge transfer process between negative atomic ions of iodine and iodine molecules has been studied in the afterglow of a pulsed discharge in iodine vapour, A wire probe technique was used. The cross section of the process was found to be 3·7±0·8 × 10-17 cm2 at room temperature.

The chemical reactions between caesium and halogen vapours at pressures of the order of 10-1 mm Hg or lower have been investigated. It was found that only the reaction in caesium and iodine mixture was sufficiently moderate for its recombination coefficient to be measured. Since caesium has the lowest ionization potential among the elements and iodine has a large value of electron affinity, it is to be expected that in the afterglow following a pulsed radio-frequency discharge in such a vapour mixture, the positive carriers will be caesium positive ions and the negative carriers will be iodine molecular ions, and the de-ionization will proceed mainly by their mutual recombination. On making these assumptions the recombination coefficient of was found to be 2·3±0·7 × 10-8 cm3 sec-1 for the process, Cs+ +I2-=Cs+I2, at room temperature.

High enantioselectivities have been achieved in the 1,4-addition of dialkylzincs to 2-cyclopentenone, 2-cyclohexenone, and 2-cycloheptenone with ee values up to 99% by using chiral aryl diphosphite ligands derived from H8-binaphthol.