Prevalence of CD44+/CD24-/low cells in breast cancer may not be associated with clinical outcome but may favor distant metastasis.
Breast cancer is composed of phenotypically diverse populations of cancer cells. The ability to form breast tumors has been shown by in vitro/in vivo studies to be restricted to epithelial tumor cells with CD44(+)/CD24(-/low) characteristics. Validation of these findings with respect to detection in clinical samples, prognosis, and clinical relevance is in demand. We investigated breast cancer tissues for the prevalence of CD44(+)/CD24(-/low) tumor cells and their prognostic value. The study included paraffin-embedded tissues of 136 patients with and without recurrences. In addition, a breast cancer progression array with normal, carcinoma in situ, and carcinoma tissues was analyzed. We applied double-staining immunohistochemistry for the detection of CD44(+)/CD24(-/low) cells. Evaluation was by microscopic pathologic inspection and automated image analysis. CD44(+)/CD24(-/low) cells ranged from 0% to 40% in normal breast and from 0% to 80% in breast tumor tissues. The prevalence of CD44(+)/CD24(-/low) tumor cells in 122 tumors was < or =10% in the majority (78%) of cases and >10% in the remainder. There was no significant correlation between CD44(+)/CD24(-/low) tumor cell prevalence and tumor progression. Although recurrences of tumors with high percentages of CD44(+)/CD24(-/low) tumor cells were mainly distant, preferably osseous metastasis, there was no correlation with the event-free and overall survival. There was no influence on the response to different treatment modalities. Our findings suggest that the prevalence of CD44(+)/CD24(-/low) tumor cells in breast cancer may not be associated with clinical outcome and survival but may favor distant metastasis.


MicroRNA regulation of progesterone receptor in breast cancer.
Hormone receptor status is of significant value when deciding on anti-estrogenic adjuvant therapy for breast cancer tumors. However, while estrogen receptor (ER) regulation was intensively studied, the regulation of progesterone receptor (PR) levels has not been extensively investigated. MicroRNAs (miRNAs, miRs) are post-transcriptional negative regulators of gene expression involved in diverse cellular processes. The aim of this study was to identify miRNAs that regulate PR in breast cancer.We mapped potential miRNA binding sites for miR-181a, miR-23a and miR-26b on PR mRNA and demonstrated a direct regulation of PR by these three miRNAs by in-vitro Luciferase binding assays. Over-expression of each miRNA in MCF-7 cells resulted in a reduction in the expression levels of PR mRNA. Then, expression levels of these miRNAs were measured in Formalin-Fixed, Paraffin-Embedded (FFPE) samples of 29 ER-positive breast cancer tumors and adjacent normal breast tissues. A significant reciprocal correlation between PR mRNA and the miRNA levels were identified suggesting a role for miR-181a, miR-23a and miR-26b in PR regulation in breast cancer. Moreover, the average expression fold-changes of the three miRNAs between cancerous and normal tissues displayed an opposite trend when analyzing according to Immuno-histochemistry(IHC) status. Furthermore, miR-181a and miR-26b were found to be over-expressed in most tumor tissues supporting their role in ER-positive breast cancer development. We conclude that miR-181a, miR-23a and miR-26b act as negative regulators of PR expression in ER-positive breast cancer. The diagnostic and prognostic potential of these miRNAs in breast cancer should be further evaluated.


Azulene as an ingredient for visible-light- and stimuli-responsive photoswitches.
The azulene molecule features a unique combination of optical, luminescence, and stimuli-responsive properties. This makes the azulene motif a promising functional group to be introduced in photoswitches. Recent challenges in the field of photochromic compounds require the development of new approaches to molecules that are switched by visible light (400-760 nm), are proton responsive and have advanced luminescent properties. Merging azulene with photoswitches opens prospects for fulfilling these requirements. Herein, we highlight recent results on the application of this hydrocarbon motif in various photochromic systems, such as stilbenes, diarylethenes, and azobenzenes.


Roscovitine: a novel regulator of P/Q-type calcium channels and transmitter release in central neurons.
Roscovitine is widely used for inhibition of cdk5, a cyclin-dependent kinase expressed predominantly in the brain. A novel function of roscovitine, i.e. an effect on Ca(2+) channels and transmitter release in central neurons, was studied by whole-cell voltage-clamp recordings and time-lapse fluorescence imaging techniques. Extracellular application of roscovitine markedly enhanced the tail calcium current following repolarization from depolarized voltages. This effect was rapid, reversible and dose dependent. Roscovitine dramatically slowed the deactivation kinetics of calcium channels. The deactivation time constant was increased 3- to 6-fold, suggesting that roscovitine could prolong the channel open state and increase the calcium influx. The potentiation of tail calcium currents caused by roscovitine and by the L-channel activator Bay K 8644 was not occluded but additive. Roscovitine-induced potentiation of tail calcium currents was significantly blocked by the P/Q-channel blocker CgTx-MVIIC, indicating that the major target of roscovitine is the P/Q-type calcium channel. In mutant mice with targeted deletion of p35, a neuronal specific activator of cdk5, roscovitine regulated calcium currents in a manner similar to that observed in wild-type mice. Moreover, intracellular perfusion of roscovitine failed to modulate calcium currents. These results suggest that roscovitine acts on extracellular site(s) of calcium channels via a cdk5-independent mechanism. Roscovitine potentiated glutamate release at presynaptic terminals of cultured hippocampal neurons detected with the vesicle trafficking dye FM1-43, consistent with the positive effect of roscovitine on the P/Q-type calcium channel, the major mediator of action potential-evoked transmitter release in the mammalian CNS.


Dynamic regulation of CTCF stability and sub-nuclear localization in response to stress.
The nuclear protein CCCTC-binding factor (CTCF) has diverse roles in chromatin architecture and gene regulation. Functionally, CTCF associates with thousands of genomic sites and interacts with proteins, such as cohesin, or non-coding RNAs to facilitate specific transcriptional programming. In this study, we examined CTCF during the cellular stress response in human primary cells using immune-blotting, quantitative real time-PCR, chromatin immunoprecipitation-sequence (ChIP-seq) analysis, mass spectrometry, RNA immunoprecipitation-sequence analysis (RIP-seq), and Airyscan confocal microscopy. Unexpectedly, we found that CTCF is exquisitely sensitive to diverse forms of stress in normal patient-derived human mammary epithelial cells (HMECs). In HMECs, a subset of CTCF protein forms complexes that localize to Serine/arginine-rich splicing factor (SC-35)-containing nuclear speckles. Upon stress, this species of CTCF protein is rapidly downregulated by changes in protein stability, resulting in loss of CTCF from SC-35 nuclear speckles and changes in CTCF-RNA interactions. Our ChIP-seq analysis indicated that CTCF binding to genomic DNA is largely unchanged. Restoration of the stress-sensitive pool of CTCF protein abundance and re-localization to nuclear speckles can be achieved by inhibition of proteasome-mediated degradation. Surprisingly, we observed the same characteristics of the stress response during neuronal differentiation of human pluripotent stem cells (hPSCs). CTCF forms stress-sensitive complexes that localize to SC-35 nuclear speckles during a specific stage of neuronal commitment/development but not in differentiated neurons. We speculate that these particular CTCF complexes serve a role in RNA processing that may be intimately linked with specific genes in the vicinity of nuclear speckles, potentially to maintain cells in a certain differentiation state, that is dynamically regulated by environmental signals. The stress-regulated activity of CTCF is uncoupled in persistently stressed, epigenetically re-programmed "variant" HMECs and certain cancer cell lines. These results reveal new insights into CTCF function in cell differentiation and the stress-response with implications for oxidative damage-induced cancer initiation and neuro-degenerative diseases.


Synthesis and optical properties of subphthalocyanine homo- and heterodimers axially connected via a hydroquinone linker.
Axially linked trifluoroethoxy (TFEO)-coated subphthalocyanine (SubPc) homo- and heterodimers were synthesized by two different axial ligand substitution methods. TFEO-SubPc homodimers were obtained directly from TFEO-SubPc boron chloride with o-, m-, and p-hydroquinones, while TFEO-SubPc heterodimers were synthesized via stepwise construction using a combination of the TFEO method and Torres' triflate method. The optical properties of the dimers obtained were investigated using UV-Vis and fluorescence spectrometry. Electron transfer was observed in the heterodimers, and TFEO-SubPc acted as an electron acceptor in the process. The electron transfer process differs depending on the o-, m-, and p-geometries of the hydroquinone linker, and is supported by computational calculations.


Effects of ovarian fluid and genetic differences on sperm performance and fertilization success of alternative reproductive tactics in Chinook salmon.
In many species, sperm velocity affects variation in the outcome of male competitive fertilization success. In fishes, ovarian fluid (OF) released with the eggs can increase male sperm velocity and potentially facilitate cryptic female choice for males of specific phenotypes and/or genotypes. Therefore, to investigate the effect of OF on fertilization success, we measured sperm velocity and conducted in vitro competitive fertilizations with paired Chinook salmon (Oncorhynchus tshawytscha) males representing two alternative reproductive tactics, jacks (small sneaker males) and hooknoses (large guarding males), in the presence of river water alone and OF mixed with river water. To determine the effect of genetic differences on fertilization success, we genotyped fish at neutral (microsatellites) and functional [major histocompatibility complex (MHC) II ß1] markers. We found that when sperm were competed in river water, jacks sired significantly more offspring than hooknoses; however, in OF, there was no difference in paternity between the tactics. Sperm velocity was significantly correlated with paternity success in river water, but not in ovarian fluid. Paternity success in OF, but not in river water alone, was correlated with genetic relatedness between male and female, where males that were less related to the female attained greater paternity. We found no relationship between MHC II ß1 divergence between mates and paternity success in water or OF. Our results indicate that OF can influence the outcome of sperm competition in Chinook salmon, where OF provides both male tactics with fertilization opportunities, which may in part explain what maintains both tactics in nature.


Maternal expression and early zygotic regulation of the Hox3/zen gene in the grasshopper Schistocerca gregaria.
In insects, a key step in the early patterning of the egg is to distinguish the primordium of the embryo proper from those regions that will form extra-embryonic membranes. In Drosophila, where these processes are well understood, the structure of the extra-embryonic membranes is highly derived. The distinct amnion and serosa typical of lower insects is replaced by a single, fused, and much reduced membrane, the amnioserosa, which never secretes an embryonic cuticle. We have used the Zen gene as a marker to study the formation of the extra-embryonic membranes, and other aspects of early embryonic patterning, in the grasshopper Schistocerca gregaria (African Plague Locust). Zen genes are derived from Hox genes, but in Drosophila they appear to have lost any role in patterning the A/P axis of the embryo; instead, they are involved in D/V patterning and the specification of the extra-embryonic membranes. We show that the Schistocerca zen gene is expressed during embryogenesis in three distinct phases. The first of these is during cleavage, when Sgzen is transiently expressed in all energids that reach the cell surface. The second phase of expression initiates in a ring of "necklace cells" that surround the forming embryo, and demarcate the boundary between the amnion and serosa. This leads to expression throughout the serosa. The final phase of expression is in the amnion, after this has separated from the serosa. This complex pattern implies that the role of Sgzen in Schistocerca is not limited solely to the specification of cell identity in the extra-embryonic membranes. We also report that the Schistocerca zen gene is expressed maternally, unlike its Drosophila and Tribolium counterparts. A distinct maternal transcript, and maternal Zen protein, accumulate in the developing oocyte from early post-meiotic stages. They remain uniformly distributed in the oocyte cytoplasm until late vitellogenic stages, when the protein and RNA become somewhat concentrated at the egg cortex and in the posterior polar cap of the oocyte, probably by passive exclusion from the yolk. The cytoplasmic localization of Sgzen protein in the oocyte, and at some stages during embryogenesis, implies that nuclear exclusion of this transcription factor is specifically controlled.


Symplekin, a constitutive protein of karyo- and cytoplasmic particles involved in mRNA biogenesis in Xenopus laevis oocytes.
Symplekin is a dual location protein that has been localized to the cytoplasmic plaques of tight junctions but also occurs in the form of interchromatin particles in the karyoplasm. Here we report the identification of two novel and major symplekin-containing protein complexes in both the karyo- and the cytoplasm of Xenopus laevis oocytes. Buffer-extractable fractions from the karyoplasm of stage IV-VI oocytes contain an 11S particle, prepared by immunoselection and sucrose gradient centrifugation, in which symplekin is associated with the subunits of the cleavage and polyadenylation specificity factor (CPSF). Moreover, in immunofluorescence microscopy nuclear symplekin colocalizes with protein CPSF-100 in the "Cajal bodies." However, symplekin is also found in cytoplasmic extracts of enucleated oocytes and egg extracts, where it occurs in 11S as well as in ca. 65S particles, again in association with CPSF-100. This suggests that, in X. laevis oocytes, symplekin is possibly involved in both processes, 3'-end processing of pre-mRNA in the nucleus and regulated polyadenylation in the cytoplasm. We discuss the possible occurrence of similar symplekin-containing particles involved in mRNA metabolism in the nucleus and cytoplasm of other kinds of cells, also in comparison with the nuclear forms of other dual location proteins in nuclei and cell junctions.


Herbivory induces systemic production of plant volatiles that attract predators of the herbivore: Extraction of endogenous elicitor.
It was previously shown that in response to infestation by spider mites (Tetranychus urticae), lima bean plants produce a volatile herbivoreinduced synomone that attracts phytoseiid mites (Phytoseiulus persimilis) that are predators of the spider mites. The production of predator-attracting infochemicals was established to occur systemically throughout the spider mitein-fested plant. Here we describe the extraction of a water-soluble endogenous elicitor from spider mite-infested lima bean leaves. This elicitor was shown to be transported out of infested leaves and was collected in water in which the petiole of the infested leaf was placed. When the petioles of uninfested lima bean leaves were placed in water in which infested leaves had been present for the previous seven days, these uninfested lima bean leaves became highly attractive to predatory mites in an olfactometer when an appropriate control of uninfested lima bean leaves was offered as alternative. The strength of this effect was dependent on the number of spider mites infesting the elicitor-producing leaves. Higher numbers of spider mites resulted in an elicitor solution with a stronger effect. In addition, spider mite density was important. The elicitor obtained from one leaf with 50 spider mites had a stronger effect on the attractiveness of uninfested leaves than the elicitor obtained from three leaves with 17 spider mites each. This suggests that the stress intensity imposed on a plant is an important determinant of the elicitor quantity. While the elicitor has a strong effect on the attractiveness of uninfested leaves, spider mite-infested leaves are still much more attractive to predatory mites than elicitor-exposed leaves. The data are discussed in the context of systemic effects in plant defense and the biosynthesis of herbivore-induced terpenoids in plants.


Packed red blood cells suppress T-cell proliferation through a process involving cell-cell contact.
Packed red blood cell (PRBC) transfusion suppresses immunity and increases morbidity and mortality. Leukocyte reduction has failed to abrogate these effects, thus implicating red blood cells themselves or their components. PRBC impair proliferation of immortal (Jurkat) T cells by depleting arginine from the extracellular environment. The effect of PRBC on isolated ex vivo T-cell proliferation has not been reported. We hypothesize that PRBCs depress mitogen-stimulated proliferation in isolated human and mouse T cells. Human peripheral T cells were isolated by Ficoll-Hypaque gradient, purified by magnetic separation, and stimulated with anti-CD3 or anti-CD28. DO11.10 transgenic mouse splenic T cells were stimulated with ovalbumin. Cells were cultured at 1 x 10(6)/mL in 96-well plates or in 24-transwell plates in the presence of PRBC (0.015-5% by volume, stored for 4-6 weeks). In culture media, arginine and citrulline were varied. Proliferation was measured at 72 hours by thymidine incorporation. T-cell viability, apoptosis, and receptor zeta chain were measured by flow cytometry. PRBC significantly depressed human peripheral and mouse splenic T-cell proliferation in a dose-dependent manner. PRBC arginase blockade by N-omega-hydroxy-nor-l-arginine only partly restored proliferation. Cell contact was required in both cell types for maximal effect. Depressed zeta chain in human peripheral T cells was partly restored by arginase blockade. Salvage by high-dose arginine and citrulline was unsuccessful. Decreased proliferation was not related to cell death. PRBC suppresses mitogen-stimulated human and antigen-stimulated mouse T-cell proliferation by mechanisms independent of arginine depletion. This is a novel mechanism for transfusion-associated immune suppression.


Widespread microbial invasion of the chorioamniotic membranes is a consequence and not a cause of intra-amniotic infection.
Acute chorioamnionitis is a response to amniotic fluid (AF) infection. However, it remains unclear whether substantial bacterial propagation in the chorioamniotic membranes (CAMs) precedes microbial invasion of the amniotic cavity (MIAC), which is inconsistent with characteristic 'amniotropic neutrophil migration' in acute chorioamnionitis. This study was performed to determine whether CAMs have widespread bacterial infection during MIAC and whether bacteria normally colonize CAMs. AF pellets and CAMs from the following groups were studied: group 1, patients with positive (n=18) or negative (n=22) AF cultures; group 2, patients with or without acute chorioamnionitis in which the amnion and chorion were studied separately (n=60); and group 3, patients at term who underwent a cesarean delivery (n=30). SYTO 9/propidium iodide fluorescent staining and fluorescent in situ hybridization for 16S rRNA were performed. Real-time quantitative PCR for 16S rDNA and PCR for genital mycoplasmas were also conducted. Bacteria were more frequently detected in AF than in CAMs of patients with positive AF culture (100 vs. 33%; P<0.0001). Bacteria were detected more frequently in CAMs as the severity of chorioamnionitis increased (P<0.01). The median 16S rRNA gene copy number in the amnion was significantly greater than in the chorion (group 2; P<0.0001). Bacteria were not detected in CAMs or AF in women at term before labor (group 3). A fraction of patients with chorioamnionitis or MIAC did not have bacteria in CAMs. Collectively, the findings herein indicate that MIAC does not follow widespread infection of CAMs, but precedes it. We propose a model of MIAC: the initial stage is intra-amniotic bacterial invasion through a discrete region of the CAMs, followed by intra-amniotic proliferation, and bacterial invasion of CAMs primarily extends from the amniotic fluid. This study emphasizes the importance of assessing the intra-amniotic compartment for diagnosis and treatment of preterm birth.


Cytotoxicity of snake venom enzymatic toxins: phospholipase A2 and l-amino acid oxidase.
The phospholipase A2 (PLA2) and l-amino acid oxidase (LAAO) are two major enzymes found in the venoms from most snake species. These enzymes have been structurally and functionally characterised for their pharmacological activities. Both PLA2 and LAAO from different venoms demonstrate considerable cytotoxic effects on cancer cells via induction of apoptosis, cell cycle arrest and suppression of proliferation. These enzymes produce more pronounced cytotoxic effects in cancer cells than normal cells, thus they can be potential sources as chemotherapeutic agents. It is proposed that PLA2 and LAAO contribute to an elevated oxidative stress due to their catalytic actions, for instance, the ability of PLA2 to produce reactive oxygen species during lipolysis and formation of H2O2 from LAAO catalytic activity which consequently lead to cell death. Nonetheless, the cell-death signalling pathways associated with exposure to these enzymatic toxins are not fully elucidated yet. Here in this review, we will discuss the cytotoxic effects of PLA2 and LAAO in relationship to their catalytic mechanisms and the underlying mechanisms of cytotoxic actions.


Pancreatic Fluid Interleukin-1β Complements Prostaglandin E2 and Serum Carbohydrate Antigen 19-9 in Prediction of Intraductal Papillary Mucinous Neoplasm Dysplasia.
We sought to determine if interleukin (IL)-1β and prostaglandin E2 (PGE2) (inflammatory mediators in pancreatic fluid) together with serum carbohydrate antigen (CA) 19-9 could better predict intraductal papillary mucinous neoplasm (IPMN) dysplasia than individual biomarkers alone. Pancreatic cyst fluid (n = 92) collected via endoscopy or surgery (2003-2016) was analyzed for PGE2 and IL-1β (enzyme-linked immunosorbent assay). Patients had surgical pathology-proven IPMN. Threshold values (PGE2 [>1100 pg/mL], IL-1β [>20 pg/mL], and serum CA 19-9 [>36 U/mL]) were determined. Levels of IL-1β were higher in high-grade dysplasia (HGD)/invasive-IPMN (n = 42) compared with low/moderate IPMN (n = 37) (median [range], 54.6 [0-2671] vs 5.9 [0-797] pg/mL; P < 0.001; area under curve [AUC], 0.766). Similarly, PGE2 was higher in HGD/invasive IPMN (n = 45) compared with low/moderate IPMN (n = 47) (median [range], 1790 [20-15,180] vs. 140 [10-14,630] pg/mL; P < 0.001; AUC, 0.748). Presence of elevated PGE2 and IL-1β (AUC, 0.789) provided 89% specificity and 82% positive predictive value (PPV) for HGD/invasive IPMN. Elevated levels of all 3 provided 100% specificity and PPV for HGD/invasive IPMN. Cyst fluid PGE2, IL-1β, and serum CA 19-9 in combination optimize specificity and PPV for HGD/invasive IPMN and may help build a panel of markers to predict IPMN dysplasia.


FGF-2 controls the differentiation of resident cardiac precursors into functional cardiomyocytes.
Recent evidence suggests that the heart possesses a greater regeneration capacity than previously thought. In the present study, we isolated undifferentiated precursors from the cardiac nonmyocyte cell population of neonatal hearts, expanded them in culture, and induced them to differentiate into functional cardiomyocytes. These cardiac precursors appear to express stem cell antigen-1 and demonstrate characteristics of multipotent precursors of mesodermal origin. Following infusion into normal recipients, these cells home to the heart and participate in physiological and pathophysiological cardiac remodeling. Cardiogenic differentiation in vitro and in vivo depends on FGF-2. Interestingly, this factor does not control the number of precursors but regulates the differentiation process. These findings suggest that, besides its angiogenic actions, FGF-2 could be used in vivo to facilitate the mobilization and differentiation of resident cardiac precursors in the treatment of cardiac diseases.


Multi-omics approaches reveal the molecular mechanisms underlying the interaction between Clonorchis sinensis and mouse liver.
Clonorchiasis remains a serious global public health problem, causing various hepatobiliary diseases. However, there is still a lack of overall understanding regarding the molecular events triggered by Clonorchis sinensis (C. sinensis) in the liver. BALB/c mouse models infected with C. sinensis for 5, 10, 15, and 20 weeks were constructed. Liver pathology staining and observation were conducted to evaluate histopathology. The levels of biochemical enzymes, blood routine indices, and cytokines in the blood were determined. Furthermore, alterations in the transcriptome, proteome, and metabolome of mouse livers infected for 5 weeks were analyzed using multi-omics techniques. The results of this study indicated that adult C. sinensis can cause hepatosplenomegaly and liver damage, with the most severe symptoms observed at 5 weeks post-infection. However, as the infection persisted, the Th2 immune response increased and symptoms were relieved. Multi-omics analysis of liver infected for 5 weeks identified 191, 402 and 232 differentially expressed genes (DEGs), proteins (DEPs) and metabolites (DEMs), respectively. Both DEGs and DEPs were significantly enriched in liver fibrosis-related pathways such as ECM-receptor interaction and cell adhesion molecules. Key molecules associated with liver fibrosis and inflammation (Cd34, Epcam, S100a6, Fhl2, Itgax, and Retnlg) were up-regulated at both the gene and protein levels. The top three metabolic pathways, namely purine metabolism, arachidonic acid metabolism, and ABC transporters, were associated with liver cirrhosis, fibrosis, and cholestasis, respectively. Furthermore, metabolites that can promote liver inflammation and fibrosis, such as LysoPC(P-16:0/0:0), 20-COOH-leukotriene E4, and 14,15-DiHETrE, were significantly up-regulated. Our study revealed that the most severe symptoms in mice infected with C. sinensis occurred at 5 weeks post-infection. Moreover, multi-omics analysis uncovered predominant molecular events related to fibrosis changes in the liver. This study not only enhances our understanding of clonorchiasis progression but also provides valuable insights into the molecular-level interaction mechanism between C. sinensis and its host liver.


Patient reported health status and all-cause mortality in patients with coronary heart disease.
Patients with coronary heart disease (CHD) experience reduced quality of life which may be associated with mortality in the longer term. This study explores whether patient-rated physical and mental health status was associated with mortality at 6-year follow-up among patients with CHD attending primary care in Ireland and Northern Ireland. This study is a secondary data analysis of patients with CHD recruited to a cluster randomized controlled trial from 2004 to 2010. Data collected included patient-rated physical component summary (PCS) and mental component summary (MCS) scores of health status (from the 12-Item Short-Form Health Survey (SF-12)), demographics and clinical parameters at baseline, and all-cause mortality at 6-year follow-up. Multivariate regression was conducted using generalized estimating equations (GEE) with a log-link function. Results are presented as odds ratios (ORs) and 95% confidence intervals (CIs). The study consisted of 762 individuals with mean age 67.6 years [standard deviation (SD): 9.8], and was 29% female. Mean baseline SF-12 mental (MCS) and physical (PCS) component scores were 50.0 (SD: 10.8) and 39.6 (SD: 11.2), respectively. At 6-year follow-up, the adjusted OR for the baseline MCS for mortality was 0.97 (95% CI: 0.95-0.99) and for the PCS 0.97 (95% CI: 0.95-0.99). For every five-point increase in MCS and PCS scores, there was a 14% reduction in the likelihood of all-cause mortality. Overall, the magnitude of effect for both mental health status and physical health status was similar; higher scores were significantly associated with a lower risk of mortality at 6-year follow-up.


Biallelic expression of the Gsalpha gene in human bone and adipose tissue.
Mutations of the Gsalpha gene inherited from the mother lead to pseudohypoparathyroidism (PHP) type Ia (PHP Ia), in which Albright's hereditary osteodistrophy is associated to resistance to the action of different hormones, whereas the same mutations inherited from the father lead to isolated Albright's hereditary osteodistrophy [pseudo-PHP (PPHP)]. Accordingly, it has been suggested that Gsalpha is under tissue-specific imprinting control, and recent studies provided evidence for a predominant maternal origin of Gsalpha transcripts in different endocrine organs involved in the PHP Ia phenotype. To establish whether Gsalpha is imprinted also in tissues that are site of alteration both in PHP Ia and PPHP, we selected 20 bone and 10 adipose tissue samples, which were heterozygous for a known polymorphism in exon 5. Expression from both parental alleles was evaluated by RT-PCR and enzymatic digestion of the resulting fragments. By this approach, the great majority of the samples analyzed showed an equal expression of the two alleles. Our results provide evidence for the absence of Gsalpha imprinting in human bone and fat and suggest that the clinical finding of osteodystrophy and obesity in PHP Ia and PPHP patients despite the presence of a normal Gsalpha allele is likely due to Gsalpha haploinsufficiency in these tissues.


Recognition of disgust is selectively preserved in Alzheimer's disease.
The neural substrates that subserve decoding of different emotional expressions are subject to different rates of degeneration and atrophy in Alzheimer's disease (AD), and there is therefore reason to anticipate that a differentiated profile of affect recognition impairment may emerge. However, it remains unclear whether AD differentially affects the recognition of specific emotions. Further, there is only limited research focused on whether affect recognition deficits in AD generalize to more ecologically valid stimuli. In the present study, relatively mild AD participants (n=24), older controls (n=30) and younger controls (n=30) were administered measures of affect recognition. Significant AD deficits were observed relative to both the younger and older control groups on a measure that involved labeling of static images of facial affect. AD deficits on this measure were observed in relation to all emotions assessed (anger, sadness, happiness, surprise and fear), with the exception of disgust, which was preserved even relative to the younger adult group. The relative preservation of disgust could not be attributed to biases in the choice of labels made, and it is suggested instead that this finding might reflect the relative sparing of the basal ganglia in AD. No significant AD effect was observed for the more ecologically valid measure that involved dynamic displays of facial expressions, in conjunction with paralinguistic and body movement cues, although a trend for greater AD difficulty was observed.


Prediction of intracranial hemorrhages after mechanical thrombectomy of basilar artery occlusion.
Mechanical thrombectomy (MT) achieves high recanalization rates in basilar artery occlusion (BAO). A severe complication of MT in BAO is intracranial hemorrhage (ICH). Yet, knowledge of risk factors for ICH after MT in BAO is limited. To evaluate clinical and procedural parameters of patients treated with MT owing to BAO to identify potential risk factors for ICH-in particular, symptomatic ICH (sICH), and assess their clinical relevance. We conducted a retrospective analysis of 101 consecutive patients presenting with BAO, who were treated with MT in our centre. Important clinical and procedural parameters were analysed as possible predictors for any ICH and sICH according to the Heidelberg Bleeding Classification using univariate tests and multivariate logistic regressions. ICH occurred in 25 (24.8%) patients, with a total of 7 (6.9%) developing sICH. Treatment with glycoprotein IIb/IIIa (GPIIb/IIIa) inhibitors was independently associated with any ICH (OR=24.67, 95% CI 4.90 to 124.03) and sICH (OR=7.08, 95% CI 1.36 to 36.78). Also, a longer onset-to-recanalization time increased the risk of both any ICH (OR=1.17, 95% CI 1.07 to 1.31) and sICH (OR=1.22, 95% CI 1.08 to 1.42). Higher serum glucose levels were associated with a higher incidence of any ICH (OR=1.39, 95% CI 1.06 to 1.85) and a higher risk of a fatal outcome (OR=1.03, 95% CI 1.01 to 1.05). Administration of GPIIb/IIIa inhibitor during the course of MT of BAO was identified as an important risk factor in the development of any ICH and sICH.


Joblessness, decreased income, and disability in intensive care unit survivors of nontraumatic intracranial hemorrhage in South Korea.
To examine the proportions of unemployment, decreased household income, and newly acquired disability, and their impact on long-term mortality after intensive care unit (ICU) admission due to nontraumatic intracranial hemorrhage (IH). This nationwide population-based retrospective cohort study enrolled adult patients admitted to the ICU because of nontraumatic IH between 2010 and 2018 in South Korea. Patients who were alive ≥365 days after ICU admission were defined as nontraumatic IH survivors. In total, 104,086 nontraumatic IH survivors were included in the final analysis. Among them, 7,225 (6.9 %) experienced job loss, 25,709 (24.7 %) experienced decreased household income, and 20,938 (20.1 %) had newly acquired disabilities, of whom 14,188 (13.6 %) had newly acquired brain disabilities. Male sex, increased duration of intensive care unit stay, comorbid status, hospital admission through the emergency room, nontraumatic intracerebral hemorrhage, receipt of surgery, mechanical ventilatory support, and increased total cost of hospitalization were associated with job loss, decreased household income, and newly acquired disabilities. However, these changes were not significantly associated with 2-year all-cause mortality (adjusted hazard ratio: 1.00, 95 % confidence interval: 0.95, 1.06; P = 0.997). Many nontraumatic IH survivors experienced unemployment, decreased household income, and newly acquired disability one year after ICU admission in South Korea. Some factors were potential risk factors for these changes, but the changes were not associated with 2-year all-cause mortality.


Immunohistochemical Analysis of the Drosophila Larval Neuromuscular Junction.
Synapses are specialized junctions between cells that mediate neurotransmission to modify brain activity and body function. Studies on synapse structure and function play an important role in understanding how neurons communicate and the consequences of their dysfunction in neurological disorders. The Drosophila larval neuromuscular junction is an excellent model for dissecting the cellular and molecular mechanisms of the synapse, with its large size, accessibility, and well-characterized genetics. This protocol describes the steps required for morphological and immunohistochemical analysis of the Drosophila larval neuromuscular junction including its dissection and multiplex labeling of synaptic proteins. This technique can be used to assess the impact of genetic manipulations on synaptic development, integrity, and plasticity, thus providing a valuable tool for probing complex neurological processes in a whole animal system.


The critical role of IFN-gamma in experimental coronavirus retinopathy.
Experimental coronavirus retinopathy (ECOR) is an animal model of progressive retinal disease that is first manifest as an acute retinal inflammation followed by chronic, immune-associated retinal degeneration in genetically susceptible, BALB/c mice. In retinal degeneration-resistant CD-1 mice, only the acute infection is seen. In the present study, interferon (IFN)-gamma production during ECOR was studied and its role evaluated in the clearance of infectious virus from the retina. BALB/c, CD-1, and IFN-gamma-deficient (IFN-gamma gko) mice were inoculated with the JHM strain of murine coronavirus by the intravitreal route. Mouse eyes were evaluated for infiltrating cells and major histocompatibility complex (MHC) expression by immunocytochemical staining. Isolated retinas were analyzed for IFN-gamma mRNA by RT-PCR, and sera were evaluated for IFN-gamma protein by ELISA assays. Virus infection in BALB/c mice was associated with an increase in the incidence and levels of systemic IFN-gamma. Moreover, IFN-gamma mRNA was detected within the retinas of infected animals during the acute phase of the disease but was not detected in normal or mock-injected animals. IFN-gamma mRNA was detected at the time of T-cell infiltration, and earlier studies have shown that this is temporally related to granzyme B gene expression and the clearance of infectious virus from the retina. Retinal IFN-gamma mRNA was also associated with the upregulation of MHC class I and II molecules within the retina. When this infection occurred in IFN-gamma gko mice, the virus was unchecked, and the infection led to death. These studies indicate that generation of IFN-gamma by cells infiltrating the retina is an essential part of an immune mechanism responsible for noncytolytic clearance of infectious virus from the retina.


Predictive Value of Ex Vivo Chemosensitivity Assays for Individualized Cancer Chemotherapy: A Meta-Analysis.
Current treatment strategies for chemotherapy of cancer patients were developed to benefit groups of patients with similar clinical characteristics. In practice, response is very heterogeneous between individual patients within these groups. Precision medicine can be viewed as the development toward a more fine-grained treatment stratification than what is currently in use. Cell-based drug sensitivity testing is one of several options for individualized cancer treatment available today, although it has not yet reached widespread clinical use. We present an up-to-date literature meta-analysis on the predictive value of ex vivo chemosensitivity assays for individualized cancer chemotherapy and discuss their current clinical value and possible future developments.


Human melatonin MT1 receptor induction by valproic acid and its effects in combination with melatonin on MCF-7 breast cancer cell proliferation.
We have reported that valproic acid upregulates melatonin MT1 receptor expression in rat C6 glioma cells. In addition to its anticonvulsant and mood stabilizing properties, valproic acid can also inhibit the growth of cancer cells. Since the melatonin MT1 receptor has been implicated in the oncostatic action of melatonin on human MCF-7 breast cancer cells, the effect of valproic acid on its expression was examined in this cell line. Treatment of MCF-7 cells with valproic acid (0.5 or 1 mM) for 24 or 72 h caused a significant increase in melatonin MT1 receptor mRNA or protein expression, as shown by reverse transcription-polymerase chain reaction (RT-PCR) analysis and western blotting, respectively. MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assays revealed a concentration-dependent inhibition of MCF-7 cell proliferation by valproic acid (0.5, 1.0 or 5 mM), but melatonin (1 or 10 nM) was ineffective alone or in combination with valproic acid, in the first (MCF-7A) subline examined. However, in subsequent experiments using a different (MCF-7B) subline, which expressed higher levels of MT1 receptor mRNA and showed modest sensitivity to melatonin, a combination of this hormone with valproic acid produced a significant synergistic inhibition of cell proliferation. These findings indicate that clinically relevant concentrations of valproic acid upregulate melatonin MT1 receptor expression in human breast cancer cells. Moreover, the enhanced antiproliferative effect observed with a combination of valproic acid and melatonin suggests that a similar therapeutic approach may be beneficial in breast cancer.


Overexpression of interleukin-15 in vivo enhances antitumor activity against MHC class I-negative and -positive malignant melanoma through augmented NK activity and cytotoxic T-cell response.
Interleukin (IL)-15, a pleiotropic cytokine, is involved in the development and maintenance of NK cells and memory CD8+ T cells. We examined the effects of in vivo overexpression of IL-15 on protection against 2 types of murine B16 melanoma lines, MHC class I-negative B16.44 and MHC class I-positive B16F10 cells, using IL-15 transgenic (Tg) mice that we have recently constructed. The tumor growth was severely retarded in IL-15 Tg mice after subcutaneous (s.c.) inoculation with B16.44 or B16F10 cells. IL-15 Tg mice showed an augmented NK cell activity against B16.44 cells, and in vivo depletion of NK cells by anti-asialoGM1 Ab treatment abrogated the antitumor activity in IL-15 Tg mice. On the other hand, IL-15 Tg mice inoculated with B16F10 cells developed a significant level of CTL response against B16F10 cells, and in vivo depletion of CD8+ T cells by anti-CD8 MAb treatment abrogated the antitumor activity. Thus, overexpression of IL-15 augmented antitumor activity against different tumors via augmentation of different antitumor mechanisms. These results suggest a possible therapeutic application of IL-15 for human neoplasms expressing a wide range of MHC class molecules.


[Minibrain/DYRK1A gene: candidate gene for mental retardation in Down's syndrome?].
DYRK1A is the first member of a novel subfamily of protein kinases with dual specificity. The human gene for DYRK1A is located in the "Down syndrome critical region" (21q22.2). Due to its relationship to the Drosophila gene minibrain (Mnb), whose mutation results in specific defects in neurogenesis, and based on functional experiments on transgenic mice, DYRK1A is discussed as a candidate gene for mental retardation in Down syndrome. The kinase is characterized by its ability to catalyze tyrosine-directed autophosphorylation as well as phosphorylation of serine/threonine residues in substrates. Its exact cellular function is yet unknown. DYRK1A is, however, known to be translocated into the nucleus and supposed to be involved in the control of cell growth and development. The pathogenetic impact of DYRK1A on Down syndrome needs further elucidation.


Liraglutide-Induced Weight Loss May be Affected by Autonomic Regulation in Type 1 Diabetes.
The role of the autonomic nervous system in the efficacy of glucagon-like peptide-1 receptor agonists (GLP-1 RA) in patients with type 1 diabetes is unknown. We assessed the association between autonomic function and weight loss induced by the GLP-1 RA liraglutide. Methods: Lira-1 was a randomized, double-blind, placebo-controlled trial assessing the efficacy and safety of 1.8 mg liraglutide once-daily for 24 weeks in overweight patients with type 1 diabetes. Autonomic function was assessed by heart rate response to deep breathing (E/I ratio), to standing (30/15 ratio), to the Valsalva maneuver and resting heart rate variability (HRV) indices. Associations between baseline the cardiovascular autonomic neuropathy (CAN) diagnosis (> 1 pathological non-resting test) and levels of test outcomes on liraglutide-induced weight loss was assessed by linear regression models. Results: Ninety-nine patients with mean age 48 (SD 12) years, HbA1c 70 (IQR 66;75) mmol/mol and BMI of 30 (SD 3) kg/m2 were assigned to liraglutide (N = 50) or placebo (N = 49). The CAN diagnosis was not associated with weight loss. A 50% higher baseline level of the 30/15 ratio was associated with a larger weight reduction by liraglutide of -2.65 kg during the trial (95% CI: -4.60; -0.69; P = 0.009). Similar significant associations were found for several HRV indices. Conclusions: The overall CAN diagnosis was not associated with liraglutide-induced weight loss in overweight patients with type 1 diabetes. Assessed separately, better outcomes for several CAN measures were associated with higher weight loss, indicating that autonomic involvement in liraglutide-induced weight loss may exist.


The relationship between EQ-5D, HAQ and pain in patients with rheumatoid arthritis.
This study aims to provide robust estimates of EQ-5D as a function of the HAQ and pain in patients with RA. Repeated observations were made of patients diagnosed with RA in a US observational cohort (n = 100 398 observations) who provided data on HAQ, pain on a visual analogue scale and the EQ-5D questionnaire. We used a bespoke statistical method based on mixture modelling to appropriately reflect the characteristics of the EQ-5D instrument and to compare this with results from standard multiple regression. EQ-5D can be predicted from summary HAQ and pain scores. We identify four different classes of respondents who differ in terms of disease severity. Unlike the multiple regression, the mixture model exhibits very good fit to the data and does not suffer from problems of bias or predict values outside the feasible range. It is appropriate to model the relationship between HAQ and EQ-5D but only if suitable statistical methods are applied. Linear models underestimate the quality-adjusted life year benefits, and therefore the cost-effectiveness, of therapies. The bespoke mixture model approach outlined here overcomes this problem. The addition of pain as an explanatory variable greatly improves the estimates. Reimbursement agencies rely on these types of analyses when formulating policy on the use of new drug therapies. Clinicians as well as economists should be concerned with these issues.


Circadian rhythms and their association with body temperature and time awake when performing a simple task with the dominant and non-dominant hand.
The two main aims of the study were to compare the dominant and non-dominant hand with regard to circadian rhythms of accuracy of performance at a task that required eye-hand coordination and sub-maximum muscle contraction, as well as to investigate if there were differences between the dominant and non-dominant hands in the associations between circadian rhythms of performance and core temperature and time awake. The task consisted of using a larger counter to flick a set of 20 smaller counters to land as near as possible to the center of a target. The nearer to the center of the target a counter landed, the higher the score awarded. Three measures of accuracy were calculated: the total score, the number of times the counter missed the target altogether, and the mean score for those counters that hit the target. Seventy-eight healthy participants performed the task at each of six test sessions distributed every 4 h throughout the day (at 08:00, 12:00... . 04:00 h), the participants then having been awake for about 1, 4... . 20 h, respectively. Before each test session, sub-lingual temperature (an estimate of core temperature) was measured, and estimates of the individual's alertness and fatigue were obtained. Temperature, alertness, and fatigue all showed circadian rhythms that were phased conventionally. Measures of accuracy of performance also showed significant circadian rhythms that were phased closer to the rhythms of alertness and fatigue than to that of oral temperature. In addition, and in support of our previous work, there were significant associations between performance and temperature (positive) and time awake (negative) for most measures of accuracy. Even though circadian rhythms of performance accuracy and effects of oral temperature and time awake were generally very similar between the dominant and non-dominant hand, there was a suggestion that time awake affected some aspects of performance of the non-dominant hand to a greater extent. There was little evidence to support the view that the 24-h rhythmicity was less marked in the non-dominant hand, which argues against internal desynchronization, at least for the task used in this study.


Early-branching or fast-evolving eukaryotes? An answer based on slowly evolving positions.
The current paradigm of eukaryotic evolution is based primarily on comparative analysis of ribosomal RNA sequences. It shows several early-emerging lineages, mostly amitochondriate, which might be living relics of a progressive assembly of the eukaryotic cell. However, the analysis of slow-evolving positions, carried out with the newly developed slow-fast method, reveals that these lineages are, in terms of nucleotide substitution, fast-evolving ones, misplaced at the base of the tree by a long branch attraction artefact. Since the fast-evolving groups are not always the same, depending on which macromolecule is used as a marker, this explains most of the observed incongruent phylogenies. The current paradigm of eukaryotic evolution thus has to be seriously re-examined as the eukaryotic phylogeny is presently best summarized by a multifurcation. This is consistent with the Big Bang hypothesis that all extant eukaryotic lineages are the result of multiple cladogeneses within a relatively brief period, although insufficiency of data is also a possible explanation for the lack of resolution. For further resolution, rare evolutionary events such as shared insertions and/or deletions or gene fusions might be helpful.


Distribution of glutamate receptor subunits in the primate temporal cortex and hippocampus.
The distribution of subunits for the N-methyl-D-aspartate (NR1, NR2A/B), alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (GluR1, GluR2/3, GluR4) and low affinity kainate (GluR5/6/7) ionotropic glutamate receptors was examined by immunocytochemistry in the temporal cortex and hippocampus of the rhesus macaque (Macaca mulatta). Neurons expressing NR1, NR2A/B, GluR2/3, and GluR4 subunits were widely distributed in all of the cortical layers but the overall density of the GluR4-immunopositive neurons was very low. Neurons expressing the GluR1 subunit were found predominantly in cortical layers V and VI while those expressing the GluR5/6/7 subunits were concentrated in layer V and were readily distinguishable by the thick elongate shape of their primary apical dendrites. Subcellular differences in the immunostaining pattern were also noted between the different glutamate receptor subunits. NR1 and NR2A/B immunoreactivity was most pronounced in somatic and primary dendritic compartments and to a lesser extent in cortical and hippocampal molecular layers. GluR1 immunoreactivity was more intense than GluR2/3 in the hippocampal molecular layers whereas GluR4 was undetectable. GluR5/6/7 immunoreactivity was very intense in the dentate molecular layer, and the CA1 pyramidal cells had a subcellular distribution of GluR5/6/7 that was similar to the cortical neurons. Overall, the distribution patterns of the different glutamate receptor subunits was identical in animals that had been ovariectomized and in ovariectomized animals that had subsequently undergone estradiol or estradiol/progesterone hormone replacement. Taken together, these findings demonstrate a differential spatial arrangement of glutamate receptor subunits in the primate temporal cortex and hippocampus, which may have functional significance for the integration of excitatory inputs to these areas. Furthermore, they show that in adult macaques, sex steroids do not play a major role in determining the distribution patterns of these receptor subunits.


The Effect of Speed and Rotation for Protaper File Systems on Postobturation Pain in a Single Visit and Multiple (Two) Visits in Root Canal Therapy: An In Vivo Study.
The basic idea of a root canal treatment is to alleviate the pain and heal the infection within the infected tooth, which can be resolved in a single visit or multiple visits in root canal therapy. Therefore, in this article, an in vivo comparison of single visits and multiple visits using different rotation and speed for two Protaper universal design file systems is done on the basis of time taken and incidence of pain. One hundred single-rooted premolars with irreversible symptomatic pulpitis were assigned to two groups of 50 patients each using the odd-even method, GA to be treated endodontically in a single visit and GB to be treated endodontically in multiple visits. Each group was further divided into two subgroups of 25 patients each on the basis of two different variations of speed and rotation for two Protaper file systems of the same design, GA1 (Hand Protapers) and GA2 (Rotary Protapers), GB1 (Hand Protapers) and GB2 (Rotary Protapers), respectively. After proper biomechanical shaping and cleaning, obturation was done with Gutta-percha cones and Ah plus sealer using Fast Pack obturation pen for warm vertical compaction. The pain was measured by a 100 mm modified visual analogue scale, and time was measured using a stopwatch. At 6-h intervals, post-obturation pain was more in single-visit root canal therapy than multiple-visit root canal therapy (P < 0.01). Single-visit rotary Protaper (GA2) had less incidence of post-obturation pain as compared with single-visit hand protapers (GA1) (P < 0.05). There was no significant difference in post-obturation pain in the multiple-visit hand protaper subgroup (GB1) and multiple-visit rotary protaper subgroup (GB2) (P > 0.05). Preoperative pain significantly influences the post-obturation pain. The statistical analysis was done using SPSS (Statistical Package for Social Sciences) version 17.0 statistical Analysis Software. The presence of preoperative pain can significantly influence the presence of postoperative pain. Most of the pain in both single-visit and multiple-visits root canal therapy occurred in the first 48 h after obturation, which decreases thereafter. Single-visit rotary protaper (GA2) had less incidence of post-obturation pain as compared with single-visit hand protapers (GA1). There was no significant difference in post-obturation pain in the multiple-visit hand protaper subgroup (GB1) and multiple-visit rotary protaper subgroup (GB2). Presence of sealer puff influences the duration of pain.


Ambulatory oximetry monitoring in patients with severe COPD: a preliminary study.
The benefits of long-term oxygen supplementation in COPD patients with hypoxemia are well established. The standard approach to prescribing oxygen uses a static assessment of oxygen requirements in a hospital or clinic setting. The assumption behind this approach is that patients will maintain a "therapeutic" hemoglobin oxygen saturation (SpO2) in the outpatient setting. We questioned the validity of this assumption, and hypothesized that many patients may demonstrate significant oxygen desaturation during normal activities of daily living. STUDY DESIGN, METHODS, AND MEASUREMENTS: We determined if oxygen supplementation maintained a therapeutic SpO2 level in patients with COPD (n = 27), using the technique of ambulatory oximetry monitoring (AOM). AOM consisted of using a portable oximeter to monitor SpO2, pulse rate, and patient activity while patients were engaged in normal activities of daily living over an extended time period (approximately 18 h). The portable oximeter collected and stored these data every 15 s over the monitored time period. Each AOM recording was manually scored for desaturation events and other key variables, including average SpO2 over the monitoring period, the average number of desaturation events per hour, and the percentage of monitored time deleted secondary to artifacts. University-affiliated Veterans Affairs Medical Center. All subjects were patients with stable COPD with no recent history of hospitalization or exacerbation of their lung disease. This cohort of patients demonstrated a surprising frequency of desaturation below the recommended target SpO2 value (90%), which averaged approximately 25% of AOM recording time. There was wide variability among patients in the percentage of time SpO2 was below the target value (range, 3 to 67% of AOM recording time). Motion artifact on the AOM recordings was not a major problem; an average of 8% of the recording time was deleted secondary to artifacts in this patient cohort. The results demonstrate that AOM is feasible and accurate with an acceptable level of motion artifact. These results also suggest that the standard approach for prescribing oxygen may lead to subtherapeutic SpO2 values in the outpatient setting. AOM holds promise as a tool to monitor the adequacy of oxygen prescriptions in the outpatient setting in patients with lung disease.


Validity and reproducibility of a tool for assessing clinical competencies in physical therapy students.
The evaluation of competencies in the clinical field is essential for health professionals, as it allows the acquisition of these competencies to be tracked. The objective of this study was to create and evaluate the validity and reliability of a tool for measuring clinical competencies in physical therapy (PT) students to assess the quality of their performance in a professional context. A descriptive study was designed. The Measurement Tool for Clinical Competencies in PT (MTCCP) was developed based on the evaluation of 39 experts: 15 clinicians and 24 instructors. The content validity was evaluated using the Content Validity Index (CVI). Three professors were invited to apply the tool to 10 students. Cronbach's alpha, exploratory factor analysis, and the intraclass correlation coefficient were used to determine the reliability and validity of the scale. The CVI was positive-higher than 0.8. Principal component analysis confirmed the construct validity of the tool for two main factors: clinical reasoning (first factor) and professional behavior (second factor). With regard to reliability, the MTCCP achieved an internal congruence of 0.982. The inter-evaluator reproducibility for clinical reasoning, professional behavior, and the total MTCCP score was almost perfect; the ICCs were 0.984, 0.930, and 0.983, respectively. The MTCCP is a valid and reliable instrument for assessing the performance of PT students in hospital settings and can be used to determine what skills students feel less confident using and what additional training/learning opportunities could be provided. Further research is needed to determine whether the MTCCP has similar validity and reproducibility in other Spanish-speaking national and international PT programs.


Plasminogen activator inhibitor-1 expression is regulated by the angiotensin type 1 receptor in vivo.
The fibrinolytic system plays an important role in degrading fibrin-rich thrombi and in vascular and tissue remodeling. Elevated levels of plasminogen activator inhibitor-1 (PAI-1) can reduce the efficiency of the endogenous fibrinolytic system. Angiotensin (Ang) has been shown to regulate PAI-1 expression via the Ang type 1 (AT1) receptor in some tissues and via the AT4 receptor in cultured endothelium. The purpose of this study was to examine the tissue-specific pattern of PAI-1 expression in response to infusion of Ang II in vivo. Adult male Sprague-Dawley rats (N = 5 in each group) were treated with four hours of intravenous infusions of Ang II or vehicle control while mean arterial pressure (MAP) was monitored: group 1, 600 ng/kg/min Ang II; group 2, Ang II + 10 mg/kg of the AT1 receptor antagonist (AT1RA) L158-809 q2 hour; group 3, Ang II + 0.01 to 0.1 mg/kg hydralazine as required to maintain normal blood pressure; and group 4, saline-infused controls. After infusion, tissue was harvested for Northern blotting, immunohistochemical analysis, and in situ hybridization. In group 1, Ang II infusion increased MAP from 105 +/- 8 to 160 +/- 9 mm Hg (mean +/- SE, P < 0. 01). Ang II induced increased expression of PAI-1 mRNA in all tissues examined from 5.1-fold in the heart, 9.7-fold in the kidney, 10.0-fold in the aorta, and up to 30.0-fold in the liver (all P < 0. 01 vs. control). While both AT1RA (group 3) and hydralazine (group 4) prevented Ang II-induced elevation in blood pressure, the Ang II-dependent expression of PAI-1 mRNA was reduced by only AT1 receptor blockade. We conclude that in the rat, PAI-1 is induced in a variety of tissues by Ang II directly through the AT1 receptor, independent of its effects on blood pressure.


Does the way concussion is portrayed affect public awareness of appropriate concussion management: the case of rugby league.
It is important to identify factors that might adversely affect appropriate medical consultation and management of concussion. One factor that might present a barrier to timely intervention is media portrayal of concussion in sporting events, such as professional rugby league. Accordingly, the current study employed a surveillance method of publicly available broadcast information to establish the incidence rate of probable concussion in the National Rugby League's Telstra Premiership 2010 season and examined how these injury events were shown to be managed with respect to return-to-play procedures. The incident rate for probable concussion was higher than previously reported: 11.10\1000 player hours (95% CI 5.78 to 16.40) and was likely underestimated. Importantly, most injured players were shown to continue playing or return to play despite being visibly concussed and described as such by the commentary team. Although 'return-to-play decisions' for professional players are guided by medical assessment, the authors discuss whether the way concussion is portrayed might affect public awareness of appropriate concussion management.


Surface-active phospholipid as the lubricating component of lubricin.
To resolve the apparent conflict between a lubricating glycoprotein, 'lubricin', as the active ingredient in synovial fluid (SF) and surface-active phospholipid (SAPL) present in SF (and adsorbed to articular cartilage) as the boundary lubricant reducing friction to such low physiological levels, lubricin was isolated from bovine SF following the original procedure of Swann et al. (Arthritis Rheum 1981;24:22-30). Analysis of the lipid extract by thin-layer chromatography and phosphorus determination demonstrated a phospholipid component of 11.1 +/- 1.7% (N = 5) which corresponds very closely to the 9.2-13.0% of lubricin which had hitherto remained unidentified and which has previously been shown to be transferable to the articular surface to impart lubrication. These results would appear to resolve any theoretical conflict in that lubricin is, indeed, an active ingredient within SF. Yet, as a large water-soluble molecule, it really functions as a carrier for the highly insoluble SAPL which it deposits on the articular surface as the oligolamellar layer visualized in previous studies. However, it is this deposited SAPL, rather than lubricin, which actually lubricates.


An agnostic reevaluation of the amyloid cascade hypothesis of Alzheimer's disease pathogenesis: The role of APP homeostasis.
To reassess the role of amyloid beta (Aβ) and the amyloid precursor protein (APP) system in the pathogenesis of Alzheimer's disease (AD). APP is a cell adhesion molecule that has been highly conserved over the course of phylogeny that has critical roles in brain development, synaptic plasticity, and the brain's intrinsic immune system. The amyloid cascade hypothesis describes a relatively linear, deterministic sequence of events triggered by a gain of Aβ peptide fragment toxicity that results in neurodegeneration and cognitive loss, yet well designed immunotherapy and beta secretase inhibitor trials that have successfully targeted Aβ have failed to have any consistent effects on the steady decline of cognition. Mutations of the APP and presenilin genes not only alter the ratio of longer to shorter Aβ fragments (resulting in a gain of Aβ toxicity), but also disrupt the normal homeostatic roles of their respective proteins. The evolutionary history, physiological importance, and complexity of the APP and presenilin systems, as well as other critical components including tau and apolipoprotein E (APOE) imply that altered function of such systems could have severe consequences that include but need not be limited to a gain of Aβ toxicity and would more generally result in altered homeostasis of APP-related functions. Challenges that a loss of APP homeostasis addresses better than the more limited gain of Aβ toxicity model include the topographic mismatches between Aβ and tau pathology, the profile and chronology of cognitive and biomarker changes that precede the clinical expression of mild cognitive impairment and dementia, and the disappointments of Aβ targeted therapeutics among others. The importance of APP, α- and β-secretases, the presenilins and γ-secretase, as well as tau was recognized by the authors of the amyloid cascade hypothesis, and has since led multiple investigators to propose alternative, more balanced hypotheses including reduced homeostasis and frank loss-of-function of key components that include but go beyond the currently envisioned linear model of Aβ toxicity.


Developmental trajectories of achievement emotions in mathematics during adolescence.
This study examined how adolescents' emotions in mathematics develop over time. Growth curve modeling was applied to longitudinal data collected annually from 2002 to 2006 (Grades 5-9; N = 3425 German adolescents; Mage = 11.7, 15.6 years at the first and last waves, respectively; 50.0% female). Results indicated that enjoyment and pride decreased over time (Glass's Δs = -.86, -.71). In contrast, negative emotions exhibited more complex patterns: Anger, boredom, and hopelessness increased (Δs = .52, .79, .26), shame decreased (Δ = -.12), and anxiety remained stable (Δ = .00). These change trajectories of emotions were associated with change trajectories of perceived control, intrinsic value, achievement value, and achievement in mathematics. Implications and future directions are discussed.


Virgin Olive Oil Study (VOLOS): vasoprotective potential of extra virgin olive oil in mildly dyslipidemic patients.
In vitro, olive phenols exert potent antioxidant and enzyme-modulating activities. We comparatively evaluate, in mildly dyslipidemic patients, the vasoprotective potential of extra virgin olive oil. 22 patients were administered 40 mL/day of either extra-virgin, i. e. phenol rich, or refined, i. e. phenol poor, olive oils (EVOO or ROO, respectively, with nearly identical fatty acid composition), with a crossover design. Each treatment was carried out for seven weeks, with four weeks of washout in between. Plasma antioxidant capacity, serum thromboxane B2 (TXB2) formation, and urinary isoprostane excretion were evaluated as surrogate markers of cardioprotective potential and vascular function. No effects on plasma lipid/lipoprotein profile were observed. Conversely, EVOO consumption was associated with favorable effects on circulating markers. Namely, decreased serum TXB2 production and increased plasma antioxidant capacity were observed when EVOO was administered in both treatment arms. Neither treatment had any significant effect on isoprostane excretion. EVOO consumption by mildly dyslipidemic patients is associated with favorable changes in circulating markers of cardiovascular condition. Based on current knowledge, these effects may be associated with cardioprotection.


[Use of the urea increment as an index of the effectiveness of parenteral feeding].
The influence exerted by the composition of intravenously introduced mixture of amino acids on so-called urea increment, i.e. the difference between the urea concentration in the blood on an empty stomach an on termination of its introduction, was studied in tests on adult mongrel male-dogs. On the other hand, upon introduction of an amino acids mixture imitating the Swedish preparation Vamin, which includes all essential and non-essential amino acids (except for glutamine and asparagine) in proportion as found in the egg white the urea increment values turned out to be inferior than on administration of a mixture containing from among essential amino acids only arginine and histidine, and from among non-essential ones-glycine and glutamic acid. The urea increment was also lower following combined introduction of an amino acids mixture together with an energy-producing (calorific) material, than when the latter was fed on prior to introduction of the mixture. The totality of findings suggests that as to its sensitivity to the composition of the parenterally introduced nutritional mixture and, possibly, dietary regimen-the urea increment is in no way inferior to the cumbersome, and difficulty realizable under clinical conditions method of nitrogen balance as a criterion for the efficacy of parenteral nutrition and deserves a detailed study as regards its clinical application.


Bisphenol S remodels red blood cell membrane lipids by altering plasma lipid levels, causing the risk of venous thrombosis in SD rats and zebrafish embryos.
Bisphenol S (BPS) is a raw material that is used extensively in various manufacturing processes but possesses a high detection rate in human red blood cells (RBCs). Accordingly, BPS is a potential toxicant in disturbing the function of RBCs and causing RBC-related diseases. To date, the effects and mechanisms of BPS-induced RBC-related diseases have not been elucidated. Here, using different models, including rats, zebrafish embryos and RBCs, the underlying mechanism of RBC-related diseases induced by BPS was explored. The accumulation of BPS in tissue was colon > kidney > liver > plasma > testicle > heart > brain in SD rats orally administered BPS (10 and 50 mg/kg bw/day) for 32 days, which was similar in both 10 mg/kg bw/day and 50 mg/kg bw/day group. Rats given BPS orally developed hyperlipidemia and increased RBC membrane cholesterol, as well as changes in RBC morphology and function. Moreover, BPS at the concentrations measured in rats plasma caused oxidative stress and phosphatidylserine exposure in vitro RBCs. These combined factors led to RBC aggregation in blood and an increasing in the number of RBCs in the blood vessels of the liver in rats. The dynamic visual observation of RBCs in vein vessels of zebrafish embryos exposed to BPS at 0, 1, 10 and 100 μg/L further found that the flow of RBCs in the tail vein is slow or even immobile, posing the risk of venous thrombosis. The present study provides new insight into the links between environmental pollutants and venous thrombosis.


Role of protein transamidation in serotonin-induced proliferation and migration of pulmonary artery smooth muscle cells.
Pulmonary hypertension is characterized by elevated pulmonary artery pressure and pulmonary artery smooth muscle cell (SMC) proliferation and migration. Clinical and experimental evidence suggests that serotonin (5-HT) is important in these responses. We previously demonstrated the participation of the 5-HT transporter and intracellular 5-HT (5-HTi) in the pulmonary vascular SMC-proliferative response to 5-HT. However, the mechanism underlying the intracellular actions of 5-HT is unknown. We speculated that 5-HTi activates SMC growth by post-translational transamidation of proteins via transglutaminase (TGase) activity, a process referred to as serotonylation. To test this hypothesis, serotonylation of pulmonary artery SMC proteins, and their role in 5-HT-induced proliferative and migratory responses, were assessed. 5-HT caused dose- and time-dependent increase in serotonylation of multiple proteins in both bovine and rat pulmonary artery SMCs. Inhibition of TGase with dansylcadaverin blocked this activity, as well as SMC-proliferative and migratory responses to 5-HT. Serotonylation of proteins also was blocked by 5-HT transporter inhibitors, and was enhanced by inhibition of monoamine oxidase, an enzyme known to degrade 5-HTi, indicating that 5-HTi levels regulate serotonylation. Immunoprecipitation assays and HPLC-mass spectral peptide sequencing revealed that a major protein serotonylated by TGase was fibronectin (FN). 5-HT-stimulated SMC serotonylation and proliferation were blocked by FN small interfering (si) RNA. These findings, together with previous observations that FN expression in the lung strongly correlates with the progression of pulmonary hypertension in both experimental animals and humans, suggest an important role of FN serotonylation in the pathogenesis of this disease.


Prediction of atherosclerotic cardiovascular risk in early childhood.
Atherosclerotic lesions are present even in very young individuals and therefore, risk stratification for cardiovascular (CV) disease should be implemented in childhood to promote early prevention strategies. In this review we critically appraise clinical, biochemical and genetic biomarkers available for pediatric clinical practice, which might be integrated to build effective algorithms to identify children at risk of future CV events. The first critical issue is to characterize in children aged 2-5 years, those CV risk factors/clinical conditions associated with dramatically accelerated atherosclerosis. Presence of clinical conditions such as obesity, diabetes mellitus, Kawasaki disease, etc., or positive family history for premature CV disease should be evaluated. Subsequently, a complete lipid profile and Lipoprotein(a) determination are recommended for children with increased baseline CV risk. Individuals with altered lipid profile could then undergo genetic testing for monogenic dyslipidemias to identify children with high CV genetic risk, who will be directed to appropriate therapeutic options. In perspective, calculation of a polygenic risk score, based on the analysis of several common single-nucleotide polymorphisms, could be integrated for better risk assessment. We here emphasize the importance of a "holistic" strategy integrating biochemical, anamnestic and genetic data to stratify CV risk in early childhood.


Aglossia: Case Report.
Introduction Aglossia is a rare condition caused by failure of the tongue embryogenesis process (in the fourth to eighth weeks of gestation). The tongue is an organ used in different activities such as sucking, swallowing, chewing, and talking. It is also responsible for shaping palate dental arches (in its absence, they become atrophic). There are few similar cases reported in the literature. Objective To describe a rare case of aglossia and the multidisciplinary professionals working together for 5 years to treat the patient. Resumed Report An 8-year-old girl with aglossia had an assessment comprising: (1) clinical assessment of the stomatognathic system related to resting posture, tonus, and mobility; (2) orthodontic assessment; (3) surface electromyography of the chewing muscles; (4) swallowing videofluoroscopy. Conclusion The authors confirmed the need of multidisciplinary cooperation to improve the patient's quality of life, because agenesia implicates many activities/functions that depend on the tongue to fully work. Multiprofessional cooperation helps the patient learn compensation mechanisms.


Oxygen sensing and conducted vasomotor responses in mouse cremaster arterioles in situ.
This study examines mechanisms by which changes in tissue oxygen tension elicit vasomotor responses and whether localized changes in oxygen tension initiates conducted vasomotor responses in mouse cremaster arterioles. Intravital microscopy was used to visualize the mouse cremaster microcirculation. The cremaster was superfused with Krebs' solution with different oxygen tensions, and a gas exchange chamber was used to induce localized changes in oxygen tension. In arterioles where red blood cells were removed by buffer perfusion, arterioles responded with same magnitudes of vasodilatation (DeltaD = 16.0 +/- 4.9 microm) when changing from high (PO(2) = 242.5 +/- 13.3 mm Hg) to low (PO(2) = 22.5 +/- 4.8 mm Hg) oxygen tension as seen in the intact cremaster circulation (DeltaD = 18.7 +/- 1.0 microm). Blockade of NO synthases by L: -NAME and adenosine receptors by DPCPX had no effects on vasomotor responses to low or high oxygen. Induction of localized low (PO(2) = 23.3 +/- 5.7 mmHg) or high (PO(2) = 300.0 +/- 25.7 mm Hg) oxygen tension caused vasodilatation or -constriction locally and at a site 1,000 microm upstream (distantly). Glibenclamide blocker of ATP-sensitive K(+) channels inhibited vasodilatation and -constriction to low (PO(2) = 16.0 +/- 6.4 mm Hg) and high (PO(2) = 337.4 +/- 12.8 mm Hg) oxygen tension. 1) ATP-sensitive K(+) channels seem to mediate, at least in part, vasodilatation and vasoconstriction to low and high oxygen tension; 2) Red blood cells are not necessary for inducing vasodilatation and vasoconstriction to low or high oxygen tension; 3) localized changes in the oxygen tension cause vasomotor responses, which are conducted upstream along arterioles in mouse cremaster microcirculation.


New sources of dietary fibre.
Two sources of dietary fibre were discussed in this presentation: soybean and fenugreek. Soybean dietary fibre (SDF) was found to be effective in reducing plasma glucose levels in diabetic and fa/fa rats, ob/ob mice and in non-insulin dependent diabetes mellitus (NIDDM) subjects. Supplementation of SDF in bread was more effective in glucose reduction than powder. SDF was also found to be more effective in subjects with fasting blood glucose levels above 7.2 mmol/l. SDF had no effect on insulin levels in rats or NIDDM subjects although the insulin levels in ob/ob mice were lower after 180 d feeding. SDF had no effect on body weight or lipid levels in rats and human subjects. However, in diabetic rats with high levels of blood cholesterol, SDF feeding decreased the cholesterol levels after 45 d SDF administration. Addition of powdered fenugreek to an oral glucose tolerance test significantly reduced the subsequent postprandial blood glucose level in diabetic rats. Inclusion of fenugreek to the meal tolerance test given to NIDDM also decreased the postprandial blood glucose levels. Fenugreek was found to reduce the rate of gastric emptying and to inhibit glucose transport, indicating the blood modulating effect of fenugreek to be due mainly to delayed gastric emptying with direct interference with intestinal glucose absorption. Soybean and fenugreek dietary fibres reveal a potential benefit for the control of glucose metabolism in diabetes with additional advantages resulting from their ease in usage either in a mixture of water or milk products or in cooking.


Analysis of a vector-bias model on malaria transmission.
We incorporate a vector-bias term into a malaria-transmission model to account for the greater attractiveness of infectious humans to mosquitoes in terms of differing probabilities that a mosquito arriving at a human at random picks that human depending on whether he is infectious or susceptible. We prove that transcritical bifurcation occurs at the basic reproductive ratio equalling 1 by projecting the flow onto the extended centre manifold. We next study the dynamics of the system when incubation time of malaria parasites in mosquitoes is included, and find that the longer incubation time reduces the prevalence of malaria. Also, we incorporate a random movement of mosquitoes as a diffusion term and a chemically directed movement of mosquitoes to humans expressed in terms of sweat and body odour as a chemotaxis term to study the propagation of infected population to uninfected population. We find that a travelling wave occurs; its speed is calculated numerically and estimated for the lower bound analytically.


Tree Modeling with Real Tree-Parts Examples.
We introduce a 3D tree modeling technique that utilizes examples of real trees to enhance tree creation with realistic structures and fine-level details. In contrast to previous works that use smooth generalized cylinders to represent tree branches, our method generates realistic looking tree models with complex branching geometry by employing an exemplar database consisting of real-life trees reconstructed from scanned data. These trees are sliced into representative parts (denoted as tree-cuts), representing trunk logs and branching structures. In the modeling process, tree-cuts are positioned in space in an intuitive manner, serving as efficient proxies that guide the creation of the complete tree. Allometry rules are taken into account to ensure reasonable relations between adjacent branches. Realism is further enhanced by automatically transferring geometric textures from our database onto tree branches as well as by guided growing of foliage. Our results demonstrate the complexity and variety of trees that can be generated with our method within few minutes. We carry a user study to test the effectiveness of our modeling technique.


Bariatric surgery and endometrial pathology in asymptomatic morbidly obese women: a prospective, pilot study.
To determine the prevalence of occult uterine pathology in asymptomatic, morbidly obese women before and after bariatric surgery-induced weight loss. Prospective, blinded, non-interventional cohort. Urban teaching hospital. Morbidly obese women. Endometrial biopsies were obtained at the time of Roux-en-Y gastric bypass and again 1 year later. Both the patient and the physician were blinded to the results of the initial biopsy until the conclusion of the study. Specimens were independently reviewed by two blinded pathologists. Effect of bariatric surgery-induced weight loss on the prevalence of endometrial pathology at 1 year. Fifty-nine women underwent an endometrial biopsy during bariatric surgery. The mean (range) age, weight, and body mass index (BMI) were 42 years (22-62 years), 127 kg (87-176 kg), and 46.8 kg/m(2) (36-64.3 kg/m(2) ), respectively. Four women had hyperplasia (three simple and one complex), for an overall prevalence of 6.8%. The prevalence among women not receiving some anti-estrogen therapy was 9.5%. Forty-six women (78%) underwent follow-up biopsy after a mean (range) weight loss of 42 kg (19-67 kg). Simple hyperplasia was identified in 3/46 women at the 1-year follow-up (6.5%). Two women had resolution of hyperplasia, two women had persistent, simple hyperplasia, and one had had a normal initial biopsy. No woman showed progressive pathology or cancer. At the end of the follow-up all but one patient had a documented resolution of endometrial pathology. Asymptomatic morbidly obese women are at relatively high risk of harbouring occult endometrial hyperplasia. Bariatric surgery-associated weight loss reduced but did not eliminate this risk for endometrial pathology.


Mechanisms of neurodegeneration in Huntington's disease.
Huntington's disease (HD) is caused by an expansion of cytosine-adenine-guanine (CAG) repeats in the huntingtin gene, which leads to neuronal loss in the striatum and cortex and to the appearance of neuronal intranuclear inclusions of mutant huntingtin. Huntingtin plays a role in protein trafficking, vesicle transport, postsynaptic signaling, transcriptional regulation, and apoptosis. Thus, a loss of function of the normal protein and a toxic gain of function of the mutant huntingtin contribute to the disruption of multiple intracellular pathways. Furthermore, excitotoxicity, dopamine toxicity, metabolic impairment, mitochondrial dysfunction, oxidative stress, apoptosis, and autophagy have been implicated in the progressive degeneration observed in HD. Nevertheless, despite the efforts of a multidisciplinary scientific community, there is no cure for this devastating neurodegenerative disorder. This review presents an overview of the mechanisms that may contribute for HD pathogenesis. Ultimately, a better understanding of these mechanisms will lead to the development of more effective therapeutic targets.


A new spectrin, beta IV, has a major truncated isoform that associates with promyelocytic leukemia protein nuclear bodies and the nuclear matrix.
We isolated cDNAs that encode a 77-kDa peptide similar to repeats 10-16 of beta-spectrins. Its gene localizes to human chromosome 19q13.13-q13.2 and mouse chromosome 7, at 7.5 centimorgans. A 289-kDa isoform, similar to full-length beta-spectrins, was partially assembled from sequences in the human genomic DNA data base and completely cloned and sequenced. RNA transcripts are seen predominantly in the brain, and Western analysis shows a major peptide that migrates as a 72-kDa band. This new gene, spectrin betaIV, thus encodes a full-length minor isoform (SpbetaIVSigma1) and a truncated major isoform (SpbetaIVSigma5). Immunostaining of cells shows a micropunctate pattern in the cytoplasm and nucleus. In mesenchymal stem cells, the staining concentrates at nuclear dots that stain positively for the promyelocytic leukemia protein (PML). Expression of SpbetaIVSigma5 fused to green fluorescence protein in cells produces nuclear dots that include all PML bodies, which double in number in transfected cells. Deletion analysis shows that partial repeats 10 and 16 of SpbetaIVSigma5 are necessary for nuclear dot formation. Immunostaining of whole-mount nuclear matrices reveals diffuse positivity with accentuation at PML bodies. Spectrin betaIV is the first beta-spectrin associated with a subnuclear structure and may be part of a nuclear scaffold to which gene regulatory machinery binds.


Total cavopulmonary and atriopulmonary connections are associated with reduced heart rate variability.
To determine whether cavopulmonary connections are associated with abnormalities of heart rate variability. Heart rate variability was studied by 24 hour Holter monitoring in 39 patients (mean (SD) age 12.2 (4.1) years) who underwent cavopulmonary connection operations (partial in 12, total in 13, and atriopulmonary in 14). Two control groups were used: 18 healthy children (11.1 (2.5) years) and 16 patients (11.7 (4.3) years) undergoing cardiovascular surgery for biventricular repair of congenital heart disease. All patients were in sinus rhythm and had normal left ventricular function. Four time domain indices were calculated: mean duration of RR intervals (RR), standard deviation of all RR intervals (SD), square root of the mean squared differences of successive RR intervals (r-MSSD), and percentage differences of successive RR intervals of > 50 ms duration (pNN50). Four frequency domain indices were calculated: total power (TP), low frequency (LF), high frequency (HF), and the LF:HF ratio. Heart rate variability indices were identical in the two control groups. Significantly reduced heart rate variability was found in patients with total cavopulmonary connections and atriopulmonary connections compared with the two control groups. In patients with partial cavopulmonary connections, heart rate variability was reduced compared with healthy controls. No differences in heart rate variability could be related to clinical status (New York Heart Association functional class), number of surgical interventions, or presence of right atrial enlargement. Patients with cavopulmonary connections have significantly reduced heart rate variability and a particularly low vagal drive.


Streptozocin-induced diabetes affects rat urinary bladder response to autonomic agents.
The response of the urinary bladder body and base to autonomic agents was studied in streptozocin (STZ)-diabetic rats. The bladder body region from 6-wk diabetic rats showed no changes in response to acetylcholine, phenylephrine, or isoproterenol. In contrast, the bladder base region showed a 39% increase in contractile response to acetylcholine and a 37% increased response to phenylephrine. In tissues from 47-wk diabetic animals, the bladder body showed a 51% increased contractile response to acetylcholine and a 37% increased relaxation response to isoproterenol. The bladder base showed a 66% increased contraction to acetylcholine. Thus, in the bladder base, enhanced responses to acetylcholine are detected soon after induction of diabetes and continue to increase as the diabetic state progresses. Moreover, in the same bladder region, an increase in responsiveness to alpha-adrenergic stimuli occurs. In the bladder body, enhanced responses to cholinergic and to beta-adrenergic stimuli occur, but are only observed in a more chronic diabetic state. The data suggest that an effect associated with autonomic diabetic neuropathy of the urinary bladder is an increased postsynaptic responsiveness to cholinergic stimuli in both regions.


Recent advances in the molecular genetics of frontotemporal lobar degeneration.
The term frontotemporal lobar degeneration (FTLD) describes a spectrum of neurodegenerative disorders associated with deposition of misfolded proteins in the frontal and temporal lobes. Up to 40% of FTLD patients reports a family history of neurodegeneration, and approximately 1/3 of familial cases shows an autosomal dominant pattern of inheritance of the phenotype. Over the past two decades, several causative and susceptibility genes for FTLD have been discovered, supporting the notion that genetic factors are important contributors to the disease processes. Genetic variants in three genes, MAPT, GRN and C9orf72, account for about half of familial FTLD cases. In addition, rare defects in the CHMP2B, VCP, TARDBP, SQSTM1, FUS, UBQLN, OPTN, TREM2, CHCHD10 and TBK1 genes have been described. Additional genes are expected to be found in near future. The purpose of this review is to describe recent advances in the molecular genetics of the FTLD spectrum and to discuss implications for genetic counseling.


Peptide conformation and supramolecular organization in amylin fibrils: constraints from solid-state NMR.
The 37-residue amylin peptide, also known as islet amyloid polypeptide, forms fibrils that are the main peptide or protein component of amyloid that develops in the pancreas of type 2 diabetes patients. Amylin also readily forms amyloid fibrils in vitro that are highly polymorphic under typical experimental conditions. We describe a protocol for the preparation of synthetic amylin fibrils that exhibit a single predominant morphology, which we call a striated ribbon, in electron microscopy and atomic force microscopy images. Solid-state nuclear magnetic resonance (NMR) measurements on a series of isotopically labeled samples indicate a single molecular structure within the striated ribbons. We use scanning transmission electron microscopy and several types of one- and two-dimensional solid-state NMR techniques to obtain constraints on the peptide conformation and supramolecular structure in these amylin fibrils and to derive molecular structural models that are consistent with the experimental data. The basic structural unit in amylin striated ribbons, which we call the protofilament, contains four layers of parallel beta-sheets, formed by two symmetric layers of amylin molecules. The molecular structure of amylin protofilaments in striated ribbons closely resembles the protofilament in amyloid fibrils with a similar morphology formed by the 40-residue beta-amyloid peptide that is associated with Alzheimer's disease.


The long-term cognitive consequences of adolescent exposure to recreational drugs of abuse.
During adolescence, the brain continues to undergo vital developmental processes. In turn, complex behavioral and cognitive skills emerge. Unfortunately, neurobiological development during adolescence can be influenced by environmental factors such as drug exposure. Engaging in drug use during adolescence has been a long-standing health concern, especially how it predicts or relates to drug using behavior later in life. However, recent findings suggest that other behavioral domains, such as learning and memory, are also vulnerable to adolescent drug use. Moreover, it is becoming increasingly apparent that deficits in learning and memory following adolescent drug use endure into adulthood, well after drug exposure has subsided. Although persistent effects suggest an interaction between drug exposure and ongoing development during adolescence, the exact acute and long-term consequences of adolescent drug exposure on substrates of learning and memory are not fully understood. Thus, this review will summarize human and animal findings on the enduring cognitive deficits due to adolescent drug exposure. Moreover, due to the fact that adolescents are more likely to consume drugs of abuse legally available to adults, this review will focus on alcohol, nicotine, and marijuana. Further, given the critical role of the frontal cortex and hippocampus in various learning and memory domains, the impact adolescent use of the previous listed drugs on the neurobiology within these regions will also be discussed.


[Organization and differentiation of the somite mesoderm cultured in the presence of adjacent tissues (notocord, spinal cord, ectoderm, endoderm) in anurans].
The explantation of pieces of somitic mesoderm alone or in association with neighbouring tissues (notochord, spinal cord, ectoderm, endoderm) shows slearly the power of self-differentiation of the somitic tissue. Only, the spinal cord does not stimulate the differentiation of the somitic mesoderm into muscular tissue. Only the notochord does not stimulate the formation of typical myotomes. It seems that ii does not possess biochemical activity, its effect if reduced to a mechanical role.


OTOF encodes multiple long and short isoforms: genetic evidence that the long ones underlie recessive deafness DFNB9.
We have recently reported that OTOF underlies an autosomal recessive form of prelingual sensorineural deafness, DFNB9. The isolated 5-kb cDNA predicted a 1,230 amino acid (aa) C-terminus membrane-anchored cytosolic protein with three C2 domains. This protein belongs to a family of mammalian proteins sharing homology with the Caenorhabditis elegans fer-1. The two other known members of this family, dysferlin and myoferlin, both have six predicted C2 domains. By northern blot analysis, a 7-kb otoferlin mRNA could be detected in the human brain. We isolated the corresponding cDNA, which is expected to encode a 1,977-aa-long form of otoferlin with six C2 domains. A 7-kb cDNA derived from the murine orthologous gene, Otof, was also identified in the inner ear and the brain. The determination of the exon-intron structure of the human and murine genes showed that they are composed of 48 coding exons and extend approximately 90 kb and approximately 80 kb, respectively. Alternatively spliced transcripts could be detected that predict several long isoforms (six C2 domains) in humans and mice and short isoforms (three C2 domains) only in humans. Primers were designed to explore the first 19 OTOF exons, henceforth permitting exploration of the complete coding sequence of the gene in DFNB9 patients. In a southwestern Indian family affected by DFNB9, a mutation in the acceptor splice site of intron 8 was detected, which demonstrates that the long otoferlin isoforms are required for inner ear function.


Late effects and quality of life after chemo-radiation for the treatment of anal cancer.
The aim of this study was to evaluate the late effects and quality of life of patients following chemo-radiation treatment for anal cancer. All surviving anal cancer patients treated within NHS Lothian between 1990 and 2007 were invited to participate. Data were collected using the EORTC QLQ-C30, the EORTC QLQ-CR38 and the Memorial Sloan-Kettering Cancer Centre Bowel Function Instrument (MSKCC). Overall response rate was 46% (n = 42); mean age 54.7 years and with a median time interval of 63.8 months between treatment and completion of the questionnaires. Thirty-five percent of the participants were 'rarely' or 'never' able to wait 15 min to get to the toilet; 50% were 'rarely' or 'never' able to control the passage of gas; 35% limited the types of solid foods they eat; 22% had leakage of stool during the day; 39% required to use a protective pad and 29% altered their daily activities because of bowel function 'always' or 'most of the time'. Seventeen percent of patients reported financial difficulties ('quite a bit' and 'very much'). Both men and women reported high symptomology for sexual problems with a median score of 83.3 (50.0. 100.0). This study has shown that in a sub-set of patients treated with chemo-radiation for anal cancer, persistent treatment related issues are reported at a medium time interval of 5.3 years. Further work is now required to understand the impact of symptoms on day-to-day life and the challenges that people face in managing these inter-related and complex problems.


Prediction of DNA rejoining kinetics and cell survival after proton irradiation for V79 cells using Geant4-DNA.
Track structure Monte Carlo (MC) codes have achieved successful outcomes in the quantitative investigation of radiation-induced initial DNA damage. The aim of the present study is to extend a Geant4-DNA radiobiological application by incorporating a feature allowing for the prediction of DNA rejoining kinetics and corresponding cell surviving fraction along time after irradiation, for a Chinese hamster V79 cell line, which is one of the most popular and widely investigated cell lines in radiobiology. We implemented the Two-Lesion Kinetics (TLK) model, originally proposed by Stewart, which allows for simulations to calculate residual DNA damage and surviving fraction along time via the number of initial DNA damage and its complexity as inputs. By optimizing the model parameters of the TLK model in accordance to the experimental data on V79, we were able to predict both DNA rejoining kinetics at low linear energy transfers (LET) and cell surviving fraction. This is the first study to demonstrate the implementation of both the cell surviving fraction and the DNA rejoining kinetics with the estimated initial DNA damage, in a realistic cell geometrical model simulated by full track structure MC simulations at DNA level and for various LET. These simulation and model make the link between mechanistic physical/chemical damage processes and these two specific biological endpoints.