Energy transfer in PPV-based conjugated polymers: a defocused widefield fluorescence microscopy study.

Both pendant and main chain conjugated MEH-PPV based polymers have been studied at the level of single chains using confocal and widefield fluorescence microscopy techniques. In particular, defocused widefield fluorescence is applied to reveal the extent of energy transfer in these polymers by identifying whether they act as single emitters. For main chain conjugated MEH-PPV, molecular weight and the surrounding matrix play a primary role in determining energy transport processes and whether single emitter behaviour is observed. Surprisingly in polymers with a saturated backbone but containing the same pendant MEH-PPV oligomer on each repeating unit, intra-chain energy transfer to a single emitter is also apparent. The results imply there is chromophore heterogeneity that can facilitate energy funneling to the emitting site. Both main chain conjugated and pendant MEH-PPV polymers exhibit changes in orientation of the emission dipole during a fluorescence trajectory of many seconds, whereas a model MEH-PPV oligomer does not. The results suggest that, in the polymers, the nature of the emitting chromophores can change during the time trajectory.

"This is a kind of betrayal": a qualitative study of disability after breast cancer.

OBJECTIVE: We proposed to document the effect of arm morbidity and disability in 40 Canadian women who were 12-24 months post breast cancer surgery. METHODS: We completed 40 qualitative interviews as one component of a multidisciplinary national longitudinal study of arm morbidity after breast cancer (n = 745) involving four research sites (Fredericton/Saint John, Montreal, Winnipeg, Surrey). During semi-structured interviews, participants who had reported arm morbidity and disability in earlier surveys were asked to discuss the effects of these conditions on everyday life. RESULTS: The interviewees reported making major adjustments to paid and unpaid work, which often involved the assistance of family members, thus demonstrating the effect of disability. Interview data resulted in the creation of a model that addresses arm morbidity and disability, and that holds implications for health care professionals. CONCLUSIONS: Based on the interview findings, we conclude that a robust measure of disability after breast cancer should be developed. In the absence of a validated measure of the effect of disability, evaluating qualitative responses to questions about everyday activities could provide the impetus for provision of physical therapy and emotional support.

The fodrin-ankyrin cytoskeleton of choroid plexus preferentially colocalizes with apical Na+K(+)-ATPase rather than with basolateral anion exchanger AE2.

A unique feature of the choroid plexus as a single-layer epithelium is its localization of Na+K(+)-ATPase at its apical (lumenal) surface. In contrast, a band 3 (AE1)-related anion exchanger protein has been localized to the basolateral surface of the choroid plexus. Both Na+K(+)-ATPase and AE1 in other tissues have been shown to bind via ankyrin to the spectrin-actin-based membrane cytoskeleton. Since linkage of integral membrane proteins to the membrane cytoskeleton is important for their restriction to specialized domains of the cell surface, we investigated the polarity of the choroid plexus membrane cytoskeleton. We developed isoform-specific antibodies to confirm the identity of choroid plexus band 3-related polypeptide as AE2. We demonstrated that ankyrin, fodrin/spectrin, actin, myosin, and alpha-actinin are predominantly apical in choroid plexus and preferentially colocalize with apical Na+K(+)-ATPase rather than with basolateral anion exchanger AE2. Colchicine administration did not alter the polarity of apical cytoskeletal and transport proteins or basolateral AE2 in choroid plexus, suggesting that biosynthetic targeting of these proteins is not microtubule dependent. In choroid plexus papilloma, Na+K(+)-ATPase and AE2 were decreased in amount and failed to preserve their polarized distributions.

Central point sampling from cattle in livestock markets in areas of human sleeping sickness.

We present the results of a study to determine the value of central point sampling in cattle markets as a means of estimating the trypanosomiasis (T. brucei s.l.) prevalence in the surrounding landscape in Uganda. We find that in the epidemic area studied, central point sampling is a good predictor of prevalence in surrounding villages, but not in endemic areas. We also find that animals infected with trypanosomiasis are more likely to be brought for sale in livestock markets in endemic areas; we discuss these results in relation to the prevention of the spread of sleeping sickness.

Functional consequences of mutations in the transmembrane domain and the carboxy-terminus of the murine AE1 anion exchanger.

We have characterized mouse AE1-mediated 36Cl- influx and surface AE1 polypeptide expression in Xenopus oocytes injected with cRNA encoding two classes of loss-of-function mutants. The first arose spontaneously. Chimeric mutants constructed with a functional AE1 cDNA localized the site of spontaneous mutation to the transmembrane domain, and DNA sequencing revealed two missense mutations encoding the double-mutant polypeptide V728F/M7301. Each mutation individually produced only partial loss of AE1 transport activity, and coexpression of the individual mutants did not restore full activity. The functional changes produced by the mutations correlated with reduced fractional accumulation of polypeptides at the oocyte surface. The V728F/M7301 polypeptide expressed in mammalian cells displayed complete endoH resistance and rapid degradation. We also examined the effect on AE1 function of engineered removal of its hydrophilic carboxy-terminus. Both delta(c)890 and the internal deletion delta(c)890-917 were functionally inactive in Xenopus oocytes. Lack of transport activity correlated with lack of detectable polypeptide accumulation at the oocyte surface. Coexpression with wt AE1 of some, but not all, of these AE1 mutants partially suppressed wt AE1-mediated 36Cl- uptake. In contrast, coexpression with wt AE1 of soluble N-terminal AE1 fragments was not inhibitory.

VHL tumor suppressor regulates Cl-/HCO3- exchange and Na+/H+ exchange activities in renal carcinoma cells.

Mutations in the von Hippel-Lindau (VHL) tumor suppressor gene are thought to play a critical role in the pathogenesis of both sporadic and VHL disease-associated clear-cell renal carcinomas (RCC). Differential display-PCR identified the AE2 anion exchanger as a candidate VHL target gene. AE2 mRNA and polypeptide levels were approximately threefold higher in 786-O VHL cells than in 786-O Neo cells. In contrast, Cl(-)/HCO(3)(-) exchange activity in 786-O VHL cells was 50% lower than in 786-O Neo cells. Since resting intracellular pH (pH(i)) values were indistinguishable, we postulated that Na(+)/H(+) exchange activity (NHE) might be similarly reduced in 786-O VHL cells. NHE-mediated pH(i) recovery from acid load was less than 50% that in 786-O Neo cells, whereas hypertonicity-stimulated, amiloride-sensitive NHE was indistinguishable in the two cell lines. The NHE3 mRNA level was higher in 786-O VHL than 786-O Neo cells, but NHE1 mRNA levels did not differ. AE2 and NHE3 are the first transcripts reported to be upregulated by pVHL. Elucidation of mechanisms responsible for downregulation of both ion exchange activities will require further investigation.

Recovery sleep at different times of the night following loss of the last four hours of sleep.

The sleep of 8 women was restricted to the first half of the night for 1 night on two separate occasions. On each occasion, heavy loss of REM (64%) and stage 2 (60%) with only a relatively light loss (20%) of slow wave sleep resulted. The purpose of the present study was to investigate whether or not the circadian timing of recovery sleep, particularly the second 4 h, would affect the response of the sleep system to the differential loss of the sleep stages. Recovery sleep commenced at 2000 h (i.e., after a normal daily span of 16 h of wakefulness) and was either continuous or interrupted after 4 h with 4 h of enforced wakefulness. Thus, the second 4 h of recovery sleep occurred between either 0000-0400 h or 0400-0800 h, two periods of the night normally associated with low and high levels of REM, respectively. The composition of recovery sleep, particularly the level of REM sleep, was found to be relatively unaffected by circadian factors. Instead, the response of the sleep system was mainly determined by stage 4 debt. It was suggested that obtaining a daily stage 4 quota acts as the primary drive mechanism of the sleep system.

Homeostatic changes in slow wave sleep during recovery sleep following restricted nocturnal sleep and partial slow wave sleep recovery during an afternoon nap.

There has been speculation and some evidence to suggest that certain fractions of sleep, notably, slow wave sleep (SWS), are under homeostatic control. In order to test this hypothesis, the sleep of eight subjects was terminated on four different occasions after 50% of their normal baseline SWS levels had been obtained. An afternoon nap then followed during which 100%, 50%, or 25% of the SWS debt was reclaimed. A fourth condition contained no afternoon nap. The change in SWS from baseline during subsequent recovery sleep was directly related to the outstanding SWS debt, thus demonstrating that SWS is under homeostatic control.

Use of mobile genetic elements as tools for molecular epidemiology.

Trypanosomiasis is a complex zoonotic disease where human-infective and non-human-infective strains of Trypanosoma brucei interact in the same transmission cycles. Differentiating these strains is paramount to understanding disease epidemiology. Restriction fragment length polymorphism analysis of repetitive DNA has provided such a method for distinguishing human and non-human isolates. Unfortunately, this approach requires large amounts of material and a more rapid approach is required. We have developed a novel technique, mobile genetic element-PCR, for assaying for positional variation of the mobile genetic element, RIME. The trypanosome genome contains up to 400 copies of RIME. Using this approach we have observed considerable variation between strains of T. brucei. Such a technique may offer potential as a method for differentiating non-human- and human-infective trypanosomes and shows promise as a rapid sensitive tool for investigating the epidemiology of sleeping sickness.

Sleep learning during stage 2 and REM sleep.

Pairs of subjects were presented with a 20-item picture series at bedtime. In the latter part of the night, a tape-recorded series of 10 words, the verbal equivalent of half the original series of pictures, was repeated 10 times during either Stage 2 or REM sleep. Morning recall and recognition for repeated words was found to be facilitated following repetition during Stage 2 sleep, but relatively unaffected following repetition during REM sleep. However, adjusting for recall, the number of additional words elicited through recognition was found to be significantly greater for REM repeated words than for Stage 2 repeated words. It was suggested that retrieval limitations, perhaps as a result of REM state dependency, rather than storage inhibition may be the main locus of the initial recall failure. By comparison, Stage 2 sleep would seem to present both a lower barrier to memory storage and retrieval compatibility with wakefulness.

REM sleep and memory consolidation.

One proposal referring to a specific function of REM sleep has been that it is necessary for, or at least conductive to, the progress of memory consolidation. This hypothesis was tested by comparing the effects on story retention of REM deprivation against an S4 deprived control. It was found that recall accuracy following REM deprivation was significantly poorer than following S4 deprivation. Furthermore, the degree of deterioration in recall accuracy during REM recovery sleep was less than during S4 recovery sleep. These findings were interpreted as evidence for active REM facilitation of memory consolidation. However, alternative explanations based upon proactive influences need to be investigated.

Retrieval from semantic memory at different times of day.

This study is concerned with retrieval from semantic memory at different times of day. Three groups of eight subjects classified high-, medium-, and low-dominance category members as positive or negative instances of specified test categories in the morning (09.00), afternoon (14.00), or evening (20.00). Classification latencies became faster over the day. Positive test instances were classified faster than negative test instances. There was an interaction such that the difference in the time taken to classify high- relative to low-dominance category members was greater for positive instances and smaller for negative instances in the morning compared with the afternoon and evening. We propose that retrieval efficiency is lower and that a different retrieval strategy is adopted in the morning compared with later in the day. We suggest that the retrieval strategy adopted in the morning serves to minimize cognitive load and effort in a state of relatively low circadian arousal.

The effect of prior sleep on retrieval.

Sleeping immediately prior to learning impaires subsequent retention. The purpose of this study was to ascertain if relatively short-term (0 to 20 mins) retention deficits are the result of impaired retrieval at the time of testing. 24 young women students were randomly allocated to two groups: prior sleep and awake. The prior sleep group was required to generate (retrieve) instances of specified categories immediately upon awakening from the first hour or so of nocturnal sleep and repeated the task again after a delay of 20 minutes. The awake control group performed the task at exactly the same times of night but without having had any prior sleep. The prior sleep group retrieved significantly fewer category instances immediately upon awakening and at delayed testing. However, retrieval had significantly improved during the 20 mins delay. It was concluded that the detrimental effects of prior sleep on retention over relatively short intervals may in part be due to retrieval difficulties at the time of testing but that longer-term retention deficits (less than 20 mins) are more likely to be due to impaired memory consolidation.

Twisting the night away: the effects of all night disco dancing on reaction time.

8 subjects performed an unprepared simple reaction time task for 20 min. every 2 hr. while taking part in an all night disco dancing marathon (continuous exercise) and during a control (no exercise) night. Mean reaction times were significantly faster and less variable during the dance marathon. It was suggested that the effects of continuous exercise in the form of disco dancing may be to reduce drowsiness which in turn assists performance.

Roles of phosphatidylinositol 3-kinase and NF-kappaB in human cytomegalovirus-mediated monocyte diapedesis and adhesion: strategy for viral persistence.

Infected peripheral blood monocytes are proposed to play a key role in the hematogenous dissemination of human cytomegalovirus (HCMV) to tissues, a critical step in the establishment of HCMV persistence and the development of HCMV-associated diseases. We recently provided evidence for a unique strategy involved in viral dissemination: HCMV infection of primary human monocytes promotes their transendothelial migration and differentiation into proinflammatory macrophages permissive for the replication of the original input virus. To decipher the mechanism of hematogenous spread, we focused on the viral dysregulation of early cellular processes involved in transendothelial migration. Here, we present evidence that both phosphatidylinositol 3-kinase [PI(3)K] and NF-kappaB activities were crucial for the HCMV induction of monocyte motility and firm adhesion to endothelial cells. We found that the beta(1) integrins, the beta(2) integrins, intracellular adhesion molecule 1 (ICAM-1), and ICAM-3 were upregulated following HCMV infection and that they played a key role in the firm adhesion of infected monocytes to the endothelium. The viral regulation of adhesion molecule expression is complex, with PI(3)K and NF-kappaB affecting the expression of each adhesion molecule at different stages of the expression cascade. Our data demonstrate key roles for PI(3)K and NF-kappaB signaling in the HCMV-induced cellular changes in monocytes and identify the biological rationale for the activation of these pathways in infected monocytes, which together suggest a mechanism for how HCMV promotes viral spread to and persistence within host organs.

Prolonged activation of NF-kappaB by human cytomegalovirus promotes efficient viral replication and late gene expression.

Infection of fibroblasts by human cytomegalovirus (HCMV) rapidly activates the NF-kappaB signaling pathway, which we documented promotes efficient transactivation of the major immediate-early promoter (DeMeritt, I.B., Milford, L.E., Yurochko, A.D. (2004). Activation of the NF-kappaB pathway in human cytomegalovirus-infected cells is necessary for efficient transactivation of the major immediate-early promoter. J. Virol. 78, 4498-4507). Because a second, sustained increase in NF-kappaB activity following the initial phase of NF-kappaB activation was also observed, we investigated the role that this prolonged NF-kappaB activation played in viral replication and late gene expression. We first investigated HCMV replication in cells in which NF-kappaB activation was blocked by pretreatment with NF-kappaB inhibitors: HCMV replication was significantly decreased in these cultures. A decrease in replication was also observed when NF-kappaB was inhibited up to 48 h post-infection, suggesting a previously unidentified role for NF-kappaB in the regulation of the later class of viral genes.

Trypanosoma brucei s.l.: characterisation of stocks from Central Africa by PCR analysis of mobile genetic elements.

To better understand the epidemiology of sleeping sickness in the Central African sub-region, notably the heterogeneity of Human African Trypanosomiasis (HAT) foci, the mobile genetic element PCR (MGE-PCR) technique was used to genotype Trypanosoma brucei s.l. (T. brucei s.l.) isolates from this sub-region. Using a single primer REV B, which detects positional variation of the mobile genetic element RIME, via amplification of flanking regions, MGE-PCR revealed a micro genetic variability between Trypanosoma brucei gambiense (T. b. gambiense) isolates from Central Africa. The technique also revealed the presence of several T. b. gambiense genotypes and allowed the identification of minor and major ubiquitous genotypes in HAT foci. The presence of several T. b. gambiense genotypes in HAT foci may explain the persistence and the resurgence phenomena of the disease and also the epidemic and the endemic status of some Central African sleeping sickness foci. The MGE-PCR technique represents a simple, rapid, and specific method to differentiate Central African T. brucei s.l. isolates.

Predicting mood change on night shift.

This study examined the impact of night shift on six dimensions of mood. The efficacy of a range of personality, behavioural, and social/organizational predictors of these responses was also examined. Thirty-five female student nurses were studied during their first period of night work. ANOVAs indicated that only fatigue-inertia and vigour-activity were significantly affected by night work. Multivariate profile analyses revealed that significant changes were predominantly confined to the interval between the preceding rest day (B) and the first night (N1), although vigour-activity dropped significantly between N1 and N2 and rose significantly by N5 for one group of subjects. Multiple regression analyses revealed that morningness, neuroticism, work-non-work conflict, and sleep quality between shifts predicted fatigue-inertia. Extroversion and social support from both co-workers and family predicted vigour-activity. Extroversion and social support from co-workers appeared to predict the positive affect component of vigour-activity, rather than the vigour and energy component.

Staged treatment of lymphedema praecox.

Lymphedema of the lower extremities poses a challenging problem in management. Gross deformities may be encountered in patients with filarial infestation. This degree of involvement is rare in native North Americans suffering from primary lymphedema.A case is presented of a patient with changes similar to those seen in filariasis, due to several episodes of acute lymphangitis over a period of years. The involved tissue was excised and the defects skin-grafted, employing a modified Charles procedure. The magnitude of the excision was such that it was carried out in three widely spaced stages. The result was satisfactory from a functional viewpoint, and also represented a marked cosmetic improvement.