Voltage-jump relaxation kinetics for wild-type and chimeric beta subunits of neuronal nicotinic receptors.
We have studied the voltage-jump relaxation currents for a series of neuronal nicotinic acetylcholine receptors resulting from the coexpression of wild-type and chimeric beta 4/beta 2 subunits with alpha 3 subunits in Xenopus oocytes. With acetylcholine as the agonist, the wild-type alpha 3 beta 4 receptors displayed five- to eightfold slower voltage-jump relaxations than did the wild-type alpha 3 beta 2 receptors. In both cases, the relaxations could best be described by two exponential components of approximately equal amplitudes over a wide range of [ACh]'s. Relaxation rate constants increased with [ACh] and saturated at 20- to 30-fold lower concentrations for the alpha 3 beta 2 receptor than for the alpha 3 beta 4 receptor, as observed previously for the peak steady state conductance. Furthermore, the chimeric beta 4/beta 2 subunits showed a transition in the concentration dependence of the rate constants in the region between residues 94 and 109, analogous to our previous observation with steady state conductances. However, our experiments with a series of beta-subunit chimeras did not localize residues that govern the absolute value of the kinetic parameters. Hill coefficients for the relaxations also differed from those previously measured for steady state responses. The data reinforce previous conclusions that the region between residues 94 and 109 on the beta subunit plays a role in binding agonist but also show that other regions of the receptor control gating kinetics subsequent to the binding step.


Diagnostic Accuracy of Videofluoroscopy for Symptomatic Cervical Spine Injury Following Whiplash Trauma.
Intervertebral instability is a relatively common finding among patients with chronic neck pain after whiplash trauma. Videofluoroscopy (VF) of the cervical spine is a potentially sensitive diagnostic tool for evaluating instability, as it offers the ability to examine relative intervertebral movement over time, and across the entire continuum of voluntary movement of the patient. At the present time, there are no studies of the diagnostic accuracy of VF for discriminating between injured and uninjured populations. Symptomatic (injured) study subjects were recruited from consecutive patients with chronic (>6 weeks) post-whiplash pain presenting to medical and chiropractic offices equipped with VF facilities. Asymptomatic (uninjured) volunteers were recruited from family and friends of patients. An ethical review and oversight were provided by the Spinal Injury Foundation, Broomfield, CO. Three statistical models were utilized to assess the sensitivity, specificity, positive and negative predictive values (PPV and NPV) of positive VF findings to correctly discriminate between injured and uninjured subjects. A total of 196 subjects (119 injured, 77 uninjured) were included in the study. All three statistical models demonstrated high levels of sensitivity and specificity (i.e., receiver operating characteristic (ROC) values of 0.71 to 0.95), however, the model with the greatest practical clinical utility was based on the number of abnormal VF findings. For 2+ abnormal VF findings, the ROC was 0.88 (93% sensitivity, 79% specificity) and the PPV and NPV were both 88%. The highest PPV (1.0) was observed with 4+ abnormal findings. Videofluoroscopic examination of the cervical spine provides a high degree of diagnostic accuracy for the identification of vertebral instability in patients with chronic pain stemming from whiplash trauma.


Perceived barriers to and ideas for weight control interventions in Malay homemakers: results from focus groups.
To develop a better understanding of perceived barriers to and ideas for weight loss and maintenance among Malay homemakers in Singapore. Interventions for weight management were reviewed, and 5 focus groups were conducted among overweight and obese Malay homemakers (n = 24) to explore barriers to and concepts and ideas pertaining to weight loss and maintenance. Barriers identified included the lack of time, finances, and motivation. Ideas that were most appealing included free health screening, workshops on how best to manage children and family, regular fun group activities, monetary incentives for participation in activities for healthy living, and flexible part-time jobs. The focus groups provided useful insights pointing toward possible interventions that could appeal to Malay homemakers in Singapore. The most appealing approaches were those that were time flexible and had the least financial burden or had a financial benefit.


Requirements for measles virus induction of RANTES chemokine in human astrocytoma-derived U373 cells.
Interferons and chemokines play a critical role in regulating the host response to viral infection. Measles virus, a member of the Paramyxoviridae family, induces RANTES expression by astrocytes. We have examined the mechanism of this induction in U373 cells derived from a human astrocytoma. RANTES was induced in a dose- and time-dependent manner by measles virus infection. Inhibition of receptor binding by the anti-CD46 antibody TRA-2.10 and of virus-membrane fusion by the tripeptide X-Phe-Phe-Gly reduced RANTES expression. Formalin-inactivated virus, which can bind but not fuse, and extensively UV-irradiated virus, which can bind and fuse, were both ineffective. Therefore, virus binding to the cellular receptor CD46 and subsequent membrane fusion were necessary, but not sufficient, to induce RANTES. UV irradiation of virus for less than 10 min proportionally inhibited viral transcription and RANTES expression. RANTES induction was decreased in infected cells treated with ribavirin, which inhibits measles virus transcription. However, RANTES mRNA was superinduced by measles virus in the presence of cycloheximide. These data suggest that partial transcription of the viral genome is sufficient and necessary for RANTES induction, whereas viral protein synthesis and replication are not required. This hypothesis was supported by the fact that RANTES was induced through transient expression of the measles virus nucleocapsid gene but not by measles genes encoding P or L proteins or by leader RNA in A549 cells. Thus, transcription of specific portions of measles virus RNA, such as the nucleocapsid gene, appears able to generate the specific signaling required to induce RANTES gene expression.


The adverse effects of bisphosphonates in breast cancer: A systematic review and network meta-analysis.
Bisphosphonate drugs can be used to improve the outcomes of women with breast cancer. Whilst many meta-analyses have quantified their potential benefits for patients, attempts at comprehensive quantification of potential adverse effects have been limited. We undertook a meta-analysis with novel methodology to identify and quantify these adverse effects. We systematically reviewed randomised controlled trials in breast cancer where at least one of the treatments was a bisphosphonate (zoledronic acid, ibandronate, pamidronate, alendronate or clodronate). Neoadjuvant, adjuvant and metastatic settings were examined. Primary outcomes were adverse events of any type or severity (excluding death). We carried out pairwise and network meta-analyses to estimate the size of any adverse effects potentially related to bisphosphonates. In order to ascertain whether adverse effects differed by individual factors such as age, or interacted with other common adjuvant breast cancer treatments, we examined individual-level patient data for one large trial, AZURE. We identified 56 trials that reported adverse data, which included a total of 29,248 patients (18,301 receiving bisphosphonate drugs versus 10,947 not). 24 out of the 103 different adverse outcomes analysed showed a statistically and practically significant increase in patients receiving a bisphosphonate drug compared with those not (2 additional outcomes that appeared statistically significant came only from small studies with low event counts and no clinical suspicion so are likely artifacts). Most of these 24 are already clinically recognised: 'flu-like symptoms, fever, headache and chills; increased bone pain, arthralgia, myalgia, back pain; cardiac events, thromboembolic events; hypocalcaemia and osteonecrosis of the jaw; as well as possibly stiffness and nausea. Oral clodronate appeared to increase the risk of vomiting and diarrhoea (which may also be increased by other bisphosphonates), and there may be some hepatotoxicity. Four additional potential adverse effects emerged for bisphosphonate drugs in this analysis which have not classically be recognised: fatigue, neurosensory problems, hypertonia/muscle spasms and possibly dysgeusia. Several symptoms previously reported as potential side effects in the literature were not significantly increased in this analysis: constipation, insomnia, respiratory problems, oedema or thirst/dry mouth. Individual patient-level data and subgroup analysis revealed little variation in side effects between women of different ages or menopausal status, those with metastatic versus non-metastatic cancer, or between women receiving different concurrent breast cancer therapies. This meta-analysis has produced estimates for the absolute frequencies of a range of side effects significantly associated with bisphosphonate drugs when used by breast cancer patients. These results show good agreement with previous literature on the subject but are the first systematic quantification of side effects and their severities. However, the analysis is limited by the availability and quality of data on adverse events, and the potential for bias introduced by a lack of standards for reporting of such events. We therefore present a table of adverse effects for bisphosphonates, identified and quantified to the best of our ability from a large number of trials, which we hope can be used to improve the communication of the potential harms of these drugs to patients and their healthcare providers.


Electrocardiographic diagnosis of the left ventricular hypertrophy in patients with left bundle branch block: is it necessary to verify old criteria?
The diagnosis of left ventricular hypertrophy (LVH) in the presence of the left bundle branch block (LBBB) is difficult. In our study we compared commonly used ECG criteria with left ventricular mass index (LVMI) calculated during cardiac magnetic resonance imaging (CMRI) to verify their clinical value or the need to recalibrate. CMRI and ECG data of 36 patients were included in this study. Based on the ECG measures we used in our study selected ECG criteria for LVH detection in cases with LBBB: QRS duration, amplitude of S waves in V1, V2, V3; R waves in aVL, V5, V6 and combinations of amplitudes. LVH was defined as LVMI (CMRI) exceeding reference values. LVH was diagnosed in 17 (47%) patients (6 women and 11 men). Following ECG parameters correlated the most prominently with LVMI - RV5: r = 0.5 (p = 0.002), RV6: r = 0.61 (p = 0.0001), SV1+RV5, 6: r = 0.64 (p = 0.001), RaVL+SV3: r = 0.5 (p = 0.002), SV2+RV5, 6: r = 0.71 (p = 0.0001), SV2, 3+RV5, 6: r = 0.75 (p = 0.0001). Based on the results of ROC analysis we proposed new cut points for LVH parameters. The highest diagnostic accuracy achieved S2+SV3 〉 6 mV, SV2,V3+RV5,V6 〉 4 mV, RaVL+SV3 〉 3.5 mV (86-89%). In patients with LBBB the ECG parameters based on the amplitude of S wave in V2 or V3 and R wave in the leads aVL, V5, V6 have the highest clinical value in the prediction of the LVH determined by CMRI. It was necessary to recalibrate these parameters and it is needed to verify them in larger number of LBBB patients.


Social inequalities associated with COVID-19 case fatality rate in Fortaleza, Ceará state, Brazil, 2020.
To analyze the association among social and health inequalities, socioeconomic status, spatial segregation and Case Fatality Rate (CFR) due to COVID-19 in Fortaleza, the state capital of Ceará, Brazil. This was an ecological study of confirmed cases and deaths due to COVID-19. The 119 neighborhoods of Fortaleza were used as units of analysis. Incidence, mortality and apparent CFR indicators due to COVID-19 were calculated between January 1 and June 8, 2020. Socioeconomic indicators were obtained from the 2010 Brazilian Demographic Census. Spatial analysis was performed and local and global Moran's indexes were calculated. There were 22,830 confirmed cases, 2,333 deaths and the apparent CFR was 12.7% (95% CI 11.6;13.9). Significant spatial autocorrelations between apparent CFR (I=0.35) and extreme poverty (I=0.51), overlapping in several neighborhoods of the city, were found. The apparent CFR due to COVID-19 is associated with the worst socioeconomic and health status, which shows the relationship between social inequalities and health outcomes in times of pandemic.


In vivo imaging of oskar mRNA transport reveals the mechanism of posterior localization.
oskar mRNA localization to the posterior of the Drosophila oocyte defines where the abdomen and germ cells form in the embryo. Although this localization requires microtubules and the plus end-directed motor, kinesin, its mechanism is controversial and has been proposed to involve active transport to the posterior, diffusion and trapping, or exclusion from the anterior and lateral cortex. By following oskar mRNA particles in living oocytes, we show that the mRNA is actively transported along microtubules in all directions, with a slight bias toward the posterior. This bias is sufficient to localize the mRNA and is reversed in mago, barentsz, and Tropomyosin II mutants, which mislocalize the mRNA anteriorly. Since almost all transport is mediated by kinesin, oskar mRNA localizes by a biased random walk along a weakly polarized cytoskeleton. We also show that each component of the oskar mRNA complex plays a distinct role in particle formation and transport.


Intracellular localization of the UL31 protein of herpes simplex virus type 2.
We studied intracellular localization of the UL31 protein of herpes simplex virus type 2 (HSV-2) in infected and transfected cells. The UL31 protein localized diffusely throughout the nucleus in infected Vero cells and the distribution patterns of the UL31 protein appeared to be different from those of either replication protein ICP8 or capsid protein ICP35. In transfected Vero cells it localized diffusely throughout the nucleus except the nucleolus at early times after transfection. At very low efficiency, it accumulated in the nucleolus. At intermediate times after transfection, the UL31 protein showed punctate staining in the nucleus. These punctate forms fused and became larger. At later times after transfection, granular forms further fused and a nuclear diffuse pattern virtually disappeared. We also constructed five N and C terminal deletion mutants of the UL31 protein for transfection assays and showed that the region containing amino acids 44 to 110 was important for nuclear and nucleolar localization. Moreover, green fluorescent protein (GFP)-targeting experiments showed that the UL31 protein was able to transport nonnuclear GFP to the nucleus and nucleolus as a fusion protein.


Metabolic flux elucidation for large-scale models using 13C labeled isotopes.
A key consideration in metabolic engineering is the determination of fluxes of the metabolites within the cell. This determination provides an unambiguous description of metabolism before and/or after engineering interventions. Here, we present a computational framework that combines a constraint-based modeling framework with isotopic label tracing on a large scale. When cells are fed a growth substrate with certain carbon positions labeled with (13)C, the distribution of this label in the intracellular metabolites can be calculated based on the known biochemistry of the participating pathways. Most labeling studies focus on skeletal representations of central metabolism and ignore many flux routes that could contribute to the observed isotopic labeling patterns. In contrast, our approach investigates the importance of carrying out isotopic labeling studies using a more comprehensive reaction network consisting of 350 fluxes and 184 metabolites in Escherichia coli including global metabolite balances on cofactors such as ATP, NADH, and NADPH. The proposed procedure is demonstrated on an E. coli strain engineered to produce amorphadiene, a precursor to the antimalarial drug artemisinin. The cells were grown in continuous culture on glucose containing 20% [U-(13)C]glucose; the measurements are made using GC-MS performed on 13 amino acids extracted from the cells. We identify flux distributions for which the calculated labeling patterns agree well with the measurements alluding to the accuracy of the network reconstruction. Furthermore, we explore the robustness of the flux calculations to variability in the experimental MS measurements, as well as highlight the key experimental measurements necessary for flux determination. Finally, we discuss the effect of reducing the model, as well as shed light onto the customization of the developed computational framework to other systems.


RNA binding protein IGF2BP2 expression is induced by stress in the heart and mediates dilated cardiomyopathy.
The IGF2BP family of RNA binding proteins consists of three paralogs that regulate intracellular RNA localization, RNA stability, and translational control. Although IGF2BP1 and 3 are oncofetal proteins, IGF2BP2 expression is maintained in many tissues, including the heart, into adulthood. IGF2BP2 is upregulated in cardiomyocytes during cardiac stress and remodeling and returns to normal levels in recovering hearts. We wondered whether IGF2BP2 might play an adaptive role during cardiac stress and recovery. Enhanced expression of an IGF2BP2 transgene in a conditional, inducible mouse line leads to dilated cardiomyopathy (DCM) and death within 3-4 weeks in newborn or adult hearts. Downregulation of the transgene after 2 weeks, however, rescues these mice, with complete recovery by 12 weeks. Hearts overexpressing IGF2BP2 downregulate sarcomeric and mitochondrial proteins and have fragmented mitochondria and elongated, thinner sarcomeres. IGF2BP2 is also upregulated in DCM or myocardial infarction patients. These results suggest that IGF2BP2 may be an attractive target for therapeutic intervention in cardiomyopathies.


First ray dorsal mobility in relation to hallux valgus deformity and first intermetatarsal angle.
The hypermobile first ray has been implicated as contributing to the cause and progression of hallux valgus deformity. Deformity of the hallux is often accompanied by an enlarged first intermetatarsal (IM 1-2) angle. It has been hypothesized that subjects having an abnormally large IM 1-2 angle have laxity of the first ray. Objectives of this study were to compare dorsal mobility of the first ray in subjects with hallux valgus to asymptomatic controls, and to investigate the relationship between dorsal mobility and the IM 1-2 angle. Fourteen subjects (age 23-81) with hallux valgus were matched by gender and age to control subjects. The IM 1-2 angle was measured from radiographs. A load-cell device measured dorsal mobility of the first ray under a standard load of 55 N. Pearson's correlation coefficient identified a marginal correlation (r = .51) between IM 1-2 angle and dorsal mobility. An independent t-test showed a statistically (P < 0.01) larger amount of dorsal mobility in the group of subjects having hallux valgus. Mobility of the first ray was increased in subjects with hallux valgus and a large IM 1-2 angle may be an indicator of increased dorsal mobility.


Hierarchical regulation of mRNA partitioning between the cytoplasm and the endoplasmic reticulum of mammalian cells.
The mRNA transcriptome is currently thought to be partitioned between the cytosol and endoplasmic reticulum (ER) compartments by binary selection; mRNAs encoding cytosolic/nucleoplasmic proteins are translated on free ribosomes, and mRNAs encoding topogenic signal-bearing proteins are translated on ER-bound ribosomes, with ER localization being conferred by the signal-recognition particle pathway. In subgenomic and genomic analyses of subcellular mRNA partitioning, we report an overlapping subcellular distribution of cytosolic/nucleoplasmic and topogenic signal-encoding mRNAs, with mRNAs of both cohorts displaying noncanonical subcellular partitioning patterns. Unexpectedly, the topogenic signal-encoding mRNA transcriptome was observed to partition in a hierarchical, cohort-specific manner. mRNAs encoding resident proteins of the endomembrane system were clustered at high ER-enrichment values, whereas mRNAs encoding secretory pathway cargo were broadly represented on free and ER-bound ribosomes. Two distinct modes of mRNA association with the ER were identified. mRNAs encoding endomembrane-resident proteins were bound via direct, ribosome-independent interactions, whereas mRNAs encoding secretory cargo displayed predominantly ribosome-dependent modes of ER association. These data indicate that mRNAs are partitioned between the cytosol and ER compartments via a hierarchical system of intrinsic and encoded topogenic signals and identify mRNA cohort-restricted modes of mRNA association with the ER.


A retrospective analysis of American football hyperthermia deaths in the United States.
Over the period 1980-2009, there were 58 documented hyperthermia deaths of American-style football players in the United States. This study examines the geography, timing, and meteorological conditions present during the onset of hyperthermia, using the most complete dataset available. Deaths are concentrated in the eastern quadrant of the United States and are most common during August. Over half the deaths occurred during morning practices when high humidity levels were common. The athletes were typically large (79% with a body mass index >30) and mostly (86%) played linemen positions. Meteorological conditions were atypically hot and humid by local standards on most days with fatalities. Further, all deaths occurred under conditions defined as high or extreme by the American College of Sports Medicine using the wet bulb globe temperature (WBGT), but under lower threat levels using the heat index (HI). Football-specific thresholds based on clothing (full football uniform, practice uniform, or shorts) were also examined. The thresholds matched well with data from athletes wearing practice uniforms but poorly for those in shorts only. Too few cases of athletes in full pads were available to draw any broad conclusions. We recommend that coaches carefully monitor players, particularly large linemen, early in the pre-season on days with wet bulb globe temperatures that are categorized as high or extreme. Also, as most of the deaths were among young athletes, longer acclimatization periods may be needed.


MicroRNAs in cancer.
MicroRNAs (miRNAs) are small noncoding RNAs that typically inhibit the translation and stability of messenger RNAs (mRNAs), controlling genes involved in cellular processes such as inflammation, cell-cycle regulation, stress response, differentiation, apoptosis, and migration. Thus, miRNAs have been implicated in the regulation of virtually all signaling circuits within a cell, and their dysregulation has been shown to play an essential role in the development and progression of cancer. Here, after a brief description of miRNA genomics, biogenesis, and function, we discuss the effects of miRNA dysregulation in the cellular pathways that lead to the progressive conversion of normal cells into cancer cells and the potential to develop new molecular miRNA-targeted therapies.


Nutritional efficiency of alpha-ketoisocaproate relative to leucine, assessed isotopically.
The efficiency of alpha-ketoisocaproate as a dietary substitute for leucine was assessed in rats by two techniques: first, the minimal dose of alpha-ketoisocaproate required, as a supplement to a leucine-free diet, to achieve a growth rate as great as animals receiving leucine was found to be between 2.2 and 4.4 times larger. Therefore the nutritional efficiency of alpha-ketoisocaproate lies between 0.23 and 0.46. Second, alpha-[1-14C]-ketoisocaproate and [3H]leucine were administered orally and the ratio of 14C/3H incorporated into the leucine of whole-body protein and fibrin was measured. This ratio, divided by the ratio 14C/3H injected, was the same in fibrin as in whole-body protein and averaged 0.39. Thus both techniques yield the same value, within the error of measurement, for the relative nutritional efficiency of alpha-ketoisocaproate. We also found that alpha-ketoisocaproate feeding at varying dosage did not alter this ratio in whole-body protein (measured in rats fasted overnight), suggesting that neither wide variations in growth rate nor exposure for 10 days to alpha-ketoisocaproate (in a diet of constant protein content) alters the relative rates of utilization (or oxidation) of alpha-ketoisocaproate vs. leucine.


[Extraintestinal manifestations in inflammatory bowel disease].
Chronic inflammatory bowel diseases (IBD) are inflammatory gastrointestinal disorders that are not limited to the gastrointestinal tract. Many different organ systems may be involved, which makes IBD a systemic disease. The most common extraintestinal manifestations (EIM) include musculoskeletal, ophthalmological, dermatological, and hepato-biliary disorders. EIM considerably contribute to the morbidity of patients with IBD, and they limit quality of life of affected patients. Due to the diversity of the organ systems involved, care should be provided by an interdisciplinary team. Early detection of EIM allows targeted therapy and reduces overall morbidity. Of importance is the fact that EIM can occur in up to 25% of all IBD patients before the onset of the first Crohn's episode or ulcerative colitis. Therefore, all doctors, especially dermatologists, ophthalmologists and rheumatologists should be aware of this possible association between EIM and the simultaneous occurrence of intestinal symptoms.


Endogenous brain-derived neurotrophic factor in the nucleus tractus solitarius tonically regulates synaptic and autonomic function.
Brain-derived neurotrophic factor (BDNF) and its receptor, TrkB, are highly expressed in the nucleus tractus solitarius (nTS), the principal target of cardiovascular primary afferent input to the brainstem. However, little is known about the role of BDNF signaling in nTS in cardiovascular homeostasis. We examined whether BDNF in nTS modulates cardiovascular function in vivo and regulates synaptic and/or neuronal activity in isolated brainstem slices. Microinjection of BDNF into the rat medial nTS (mnTS), a region critical for baroreflex control of sympathetic outflow, produced dose-dependent increases in mean arterial pressure (MAP), heart rate (HR), and lumbar sympathetic nerve activity (LSNA) that were blocked by the tyrosine kinase inhibitor K252a. In contrast, immunoneutralization of endogenous BDNF (anti-BDNF), or microinjection of K252a alone, decreased MAP, HR, and LSNA. The effects of anti-BDNF were abolished by blockade of ionotropic glutamate receptors, indicating a role for glutamate signaling in the response to BDNF. In vitro, BDNF reduced the amplitude of miniature EPSCs as well as solitary tract (TS) evoked EPSC amplitude and action potential discharge (APD) in second-order nTS neurons. BDNF effects on EPSCs were independent of GABAergic signaling and abolished by AMPA receptor blockade. In contrast, K252a increased spontaneous EPSC frequency and TS evoked EPSC amplitude. BDNF also attenuated APD evoked by injection of depolarizing current into second-order neurons, indicating reduced intrinsic neuronal excitability. Our data demonstrate that BDNF signaling in mnTS plays a tonic role in regulating cardiovascular function, likely via modulation of primary afferent glutamatergic excitatory transmission and neural activity.


An Assessment of the Rapid Decline of Trust in US Sources of Public Information about COVID-19.
We conducted a longitidinal assessment of 806 respondents in March, 2020 in the US to examine the trustworthiness of sources of information about COVID-19. Respondents were recontacted after four months. Information sources included mainstream media, state health departments, the CDC, the White House, and a well-known university. We also examined how demographics, political partisanship, and skepticism about COVID-19 were associated with the perceived trustworthiness of information sources and decreased trustworthiness over time. At baseline, the majority of respondants reported high trust in COVID-19 information from state health departments (75.6%), the CDC (80.9%), and a university (Johns Hopkins, 81.1%). Mainstream media was trusted by less than half the respondents (41.2%), and the White House was the least trusted source (30.9%). At the 4-month follow-up, a significant decrease in trustworthiness in all five sources of COVID-19 information was observed. The most pronounced reductions were from the CDC and the White House. In multivariate analyses, factors associated with rating the CDC, state health department, and a university as trustworthy sources of COVID-19 information were political party affiliation, level of education, and skepticism about COVID-19. The most consistent predictor of decreased trust was political party affiliation, with Democrats as compared to Republicans less likely to report decreased trust across all sources.


Thalamostriatal projections from the medial posterior and parafascicular nuclei have distinct topographic and physiologic properties.
The dorsolateral striatum (DLS) is critical for executing sensorimotor behaviors that depend on stimulus-response (S-R) associations. In rats, the DLS receives it densest inputs from primary somatosensory (SI) cortex, but it also receives substantial input from the thalamus. Much of rat DLS is devoted to processing whisker-related information, and thalamic projections to these whisker-responsive DLS regions originate from the parafascicular (Pf) and medial posterior (POm) nuclei. To determine which thalamic nucleus is better suited for mediating S-R associations in the DLS, we compared their input-output connections and neuronal responses to repetitive whisker stimulation. Tracing experiments demonstrate that POm projects specifically to the DLS, but the Pf innervates both dorsolateral and dorsomedial parts of the striatum. The Pf nucleus is innervated by whisker-sensitive sites in the superior colliculus, and these sites also send dense projections to the zona incerta, a thalamic region that sends inhibitory projections to the POm. These data suggest that projections from POm to the DLS are suppressed by incertal inputs when the superior colliculus is activated by unexpected sensory stimuli. Simultaneous recordings with two electrodes indicate that POm neurons are more responsive and habituate significantly less than Pf neurons during repetitive whisker stimulation. Response latencies are also shorter in POm than in Pf, which is consistent with the fact that Pf receives its whisker information via synaptic relays in the superior colliculus. These findings indicate that, compared with the Pf nucleus, POm transmits somatosensory information to the DLS with a higher degree of sensory fidelity.


Stevens-Johnson syndrome and toxic epidermal necrolysis associated with the use of macrolide antibiotics: a review of published cases.
Macrolides are one of the most commonly prescribed antibiotics. In several studies, their use was associated with the occurrence of Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN). This review aimed to explore and summarize available cases of SJS/TEN suspected to be associated with the use of macrolide antibiotics reported in the literature. Electronic searches were conducted in PubMed/MEDLINE, Web of Science, Scopus, and Serbian Citation Index (SCIndeks). Twenty-five publications describing a total of 27 patients were included. Cases of SJS/TEN which satisfied inclusion criteria were found for azithromycin (n = 11), clarithromycin (n = 7), erythromycin (n = 5), roxithromycin (n = 2), and telithromycin (n = 2). The age of the patients ranged from 2 to 77 years (median: 29 years). There were 14 female (51.9%) and 13 male (48.1%) patients. SJS was diagnosed in 16 patients (59.3%), TEN in 10 patients (37.0%), and SJS/TEN overlap in one patient (3.7%). Time to onset of the first symptoms ranged from 1 to 14 days (median: 3 days). All patients received some form of supportive and symptomatic care. Systemic corticosteroids were reported to be administered in 12 patients (44.4%) and intravenous immunoglobulin in five patients (18.5%). Three patients (11.1%) died. Considering that SJS/TEN is a severe and potentially life-threatening reaction, physicians should be aware that they could be adverse effects of macrolide antibiotics and keep in mind that prompt recognition of SJS/TEN and discontinuation of the culprit drug in combination with supportive care is essential.


Pediatric Miller Fisher Syndrome; Characteristic Presentation and Comparison with Adult Miller Fisher Syndrome.
We aimed to investigate the characteristic presentation of Miller Fisher syndrome (MFS) in pediatrics and compare it with that in adults. We performed a retrospective review of medical records, laboratory findings, and disease course of pediatric MFS. The data were compared with those of adult MFS, and literature review was done. Unpaired and paired comparisons between groups were made using Wilcoxon rank-sum and signed-rank tests, respectively. Median age for pediatric MFS was 9.8 ± 6.5 years. There were 5 (45.5%) male and 6 (54.5%) female patients. All patients had preceding infection. Two patients (22.2%) had tested positive for anti-GQ1b antibody. Ten patients (90.1%) were treated with intravenous immunoglobulin, and 2 (18.2%) also received intravenous methylprednisolone. Within one month, 8 (72.7%) patients showed recovery, and all 11 (100%) recovered fully within 3 months. Further, the pediatric group had higher frequency of unilateral involvement of ophthalmoplegia, ataxia, and autonomic symptoms but lower antiganglioside antibody positivity and manifestations of areflexia than the adult group. Neuro-ophthalmic manifestations and disease course of pediatric MFS were similar to those of adult MFS as stated in the literature. However, the presence of autonomic symptoms was higher and anti-GQ1b antibody positivity was lower in pediatric MFS than in adult MFS.


Interleukin-17 limits hypoxia-inducible factor 1α and development of hypoxic granulomas during tuberculosis.
Mycobacterium tuberculosis (Mtb) is a global health threat, compounded by the emergence of drug-resistant strains. A hallmark of pulmonary tuberculosis (TB) is the formation of hypoxic necrotic granulomas, which upon disintegration, release infectious Mtb. Furthermore, hypoxic necrotic granulomas are associated with increased disease severity and provide a niche for drug-resistant Mtb. However, the host immune responses that promote the development of hypoxic TB granulomas are not well described. Using a necrotic Mtb mouse model, we show that loss of Mtb virulence factors, such as phenolic glycolipids, decreases the production of the proinflammatory cytokine IL-17 (also referred to as IL-17A). IL-17 production negatively regulates the development of hypoxic TB granulomas by limiting the expression of the transcription factor hypoxia-inducible factor 1α (HIF1α). In human TB patients, HIF1α mRNA expression is increased. Through genotyping and association analyses in human samples, we identified a link between the single nucleotide polymorphism rs2275913 in the IL-17 promoter (-197G/G), which is associated with decreased IL-17 production upon stimulation with Mtb cell wall. Together, our data highlight a potentially novel role for IL-17 in limiting the development of hypoxic necrotic granulomas and reducing disease severity in TB.


An imperfect dopaminergic error signal can drive temporal-difference learning.
An open problem in the field of computational neuroscience is how to link synaptic plasticity to system-level learning. A promising framework in this context is temporal-difference (TD) learning. Experimental evidence that supports the hypothesis that the mammalian brain performs temporal-difference learning includes the resemblance of the phasic activity of the midbrain dopaminergic neurons to the TD error and the discovery that cortico-striatal synaptic plasticity is modulated by dopamine. However, as the phasic dopaminergic signal does not reproduce all the properties of the theoretical TD error, it is unclear whether it is capable of driving behavior adaptation in complex tasks. Here, we present a spiking temporal-difference learning model based on the actor-critic architecture. The model dynamically generates a dopaminergic signal with realistic firing rates and exploits this signal to modulate the plasticity of synapses as a third factor. The predictions of our proposed plasticity dynamics are in good agreement with experimental results with respect to dopamine, pre- and post-synaptic activity. An analytical mapping from the parameters of our proposed plasticity dynamics to those of the classical discrete-time TD algorithm reveals that the biological constraints of the dopaminergic signal entail a modified TD algorithm with self-adapting learning parameters and an adapting offset. We show that the neuronal network is able to learn a task with sparse positive rewards as fast as the corresponding classical discrete-time TD algorithm. However, the performance of the neuronal network is impaired with respect to the traditional algorithm on a task with both positive and negative rewards and breaks down entirely on a task with purely negative rewards. Our model demonstrates that the asymmetry of a realistic dopaminergic signal enables TD learning when learning is driven by positive rewards but not when driven by negative rewards.


Racial disparities in risk of second breast tumors after ductal carcinoma in situ.
The purpose of the study was to examine the impact of race/ethnicity on second breast tumors among women with ductal carcinoma in situ (DCIS). We identified 102,489 women diagnosed with primary DCIS between 1988 and 2009 from the 18 NCI-SEER Registries. Cox proportional hazard regression was used to estimate race/ethnicity-associated relative risks (RRs) and their 95 % confidence intervals (CI) of ipsilateral breast tumors (IBT; defined as DCIS or invasive carcinoma in the ipsilateral breast) and contralateral breast tumors (CBT; defined as DCIS or invasive carcinoma in the contralateral breast). Overall, 2,925 women had IBT and 3,723 had CBT. Compared with white women, black (RR 1.46; 95 % CI 1.29-1.65), and Hispanic (RR 1.18; 95 % CI 1.03-1.36) women had higher IBT risk, which was similar for invasive IBT and ipsilateral DCIS. A significant increase in IBT risk among black women persisted, regardless of age at diagnosis, treatment, tumor grade, tumor size, and histology. The CBT risk was significantly increased among black (RR 1.21; 95 % CI 1.08-1.36) and Asian/PI (RR 1.16; 95 % CI 1.02-1.31) women compared with white women. The association was stronger for invasive CBT among black women and for contralateral DCIS among Asian/PI women (P heterogeneity < 0.0001). The black race-associated CBT risk was more pronounced among women ≥50 years at diagnosis and those with comedo DCIS; in contrast, a significant increase in risk among Asian/PI women was restricted to those <50 years and those with noncomedo DCIS. Racial/ethnic differences in risks of second breast tumors after DCIS could not be explained by pathologic features and treatment.


Quantitative determination of acylphosphatase levels in horse tissues by enzyme-linked immunosorbent assay.
A non competitive enzyme-linked immunosorbent assay (ELISA) specific for horse muscle acylphosphatase (E.C. 220.127.116.11.) has been developed. The purified anti-acylphosphatase antibodies were immobilized by passive absorption to a solid-phase support and incubated with known and unknown amounts of antigen. The antibody-acylphosphatase complex was quantified using the same antibody conjugated to horseradish peroxidase. The assay yields positive reactions with as little as 0.05 ng of antigen, with intra- and interassay coefficients of variation of 5% and 7%, respectively. On the basis of this assay we developed a more sensitive test than the optical one, using benzoyl-phosphate as substrate, for acylphosphatase determination. By means of this test, the presence of the enzyme in horse tissue homogenates was evaluated under conditions in which the optical test failed to distinguish the acylphosphatase activity from that of other enzymes.


Glutathione preservation during storage of rat lenses in optisol-GS and castor oil.
Glutathione concentration in the lens decreases in aging and cataractous lenses, providing a marker for tissue condition. Experimental procedures requiring unfrozen lenses from donor banks rely on transportation in storage medium, affecting lens homeostasis and alterations in glutathione levels. The aim of the study was to examine the effects of Optisol-GS and castor oil on lens condition, determined from their ability to maintain glutathione concentrations. Rat lenses were stored in the two types of storage media at varying time intervals up to 3 days. Glutathione concentration was afterwards determined in an enzymatic detection assay, specific for both reduced and oxidized forms. Lenses removed immediately after death exhibited a glutathione concentration of 4.70±0.29 mM. In vitro stored lenses in Optisol-GS lost glutathione quickly, ending with a concentration of 0.60±0.34 mM after 3 days while castor oil stored lenses exhibited a slower decline and ended at 3 times the concentration. A group of lenses were additionally stored under post mortem conditions within the host for 6 hours before its removal. Total glutathione after 6 hours was similar to that of lenses removed immediately after death, but with altered GSH and GSSG concentrations. Subsequent storage of these lenses in media showed changes similar to those in the first series of experiments, albeit to a lesser degree. It was determined that storage in Optisol-GS resulted in a higher loss of glutathione than lenses stored in castor oil. Storage for more than 12 hours reduced glutathione to half its original concentration, and was considered unusable after 24 hours.


[The effect of physical training on the indices of the general hemodynamics and kinin-kallikrein system activity in hypertension patients].
Forty men aged 30-60 years suffering from stage-II essential hypertension were examined for the general hemodynamics, the level of prekallikrein (PK) and kallikrein-like activity (KLA) of blood plasma, and kallikrein excretion with urine. The examinations were made before and after 30 weeks of exercise. Before such a course of exercise the response of the cardiovascular system and of the kinin-kallikrein system (KKS) to exercise was characterized by a significant rise of systolic, diastolic medium-hemodynamic arterial pressure, heart rate and kallikrein excretion with urine together with the lack changes in PK and KLA of blood plasma. No correlations were established between the KKS, which is likely to be determined by the disease. After the course of exercise there was a significant reduction of all the initial characteristics of the general hemodynamics, KLA, a close to significant increase of the PK level and a well-defined tendency towards lowering of kallikrein excretion with urine, and less remarkable response of all the parameters of both systems to standard exercise. A reverse correlation (r = -0.57) was established between the PK level and KLA at rest which may attest to a positive influence of exercise on the recovery of the correlations between the parameters of the KKS.


Bacterial diversity associated with freshwater zooplankton.
Bacterial community compositions (BCC) associated with the cladoceran Bosmina coregoni and the cyclopoid copepod Thermocyclops oithonoides in oligotrophic Lake Stechlin versus eutrophic Lake Dagow (northeastern Germany) were compared using molecular techniques. We also transplanted the zooplankton from their native lake to the other lake, and studied changes in their associated BCC as a result of the modified ambient environment. Bacterial community composition associated with B. coregoni was quite conservative between the oligotrophic and eutrophic lakes, and most of the bacteria belonged to Alpha- and Betaproteobacteria. In contrast, BCC associated with T. oithonoides was highly variable and dependent on the environment, and Betaproteobacteria and Bacteroidetes were the most represented among the bacteria. Thermocyclops oithonoides from the oligotrophic lake was more able to retain its bacteria after being transplanted to the eutrophic lake than vice versa. This suggests that bacteria in oligotrophic water were more firmly attached to the copepod and better in resisting environmental fluctuations than those in eutrophic water.


Promoter analysis and differential expression of the Candida rugosa lipase gene family in response to culture conditions.
Five lipase genes have been identified and sequenced from Candida rugosa. However, as the sequences of LIP multigene family are extremely closely related, it is difficult to characterize the expression spectrum of LIP genes. In the present work we have cloned, sequenced, and analyzed the promoters of these five LIP isoform genes, and several putative transcriptional elements including oleate response element (ORE) and upstream activation sequence 1 (UAS1) were identified. A quantitative real-time RT-PCR method was developed for determining the differential expression of C. rugosa lipase family genes in response to various environmental and nutritional factors. While all five LIP genes display significant changes in mRNA expression under oleic acid and/or olive oil culture conditions, LIP2 showed the strongest induction (456-fold) in response to oleic acid. LIP transcription and promoter regulation were studied by assaying the beta-galactosidase activities of promoter-lacZ fusions in Saccharomyces cerevisiae. Three of the LIP genes, LIP3, LIP4, and LIP5, showed significant induction by oleic acid, and their ORE and UAS1 elements are essential for induction by oleic acid. Together, this suggests that the multiple lipase expression profiles may be due to differential transcriptional regulation of the LIP genes in response to environment or nutritional factors.


Hemithyroidectomy versus total thyroidectomy in the intermediate-risk differentiated thyroid cancer: the Italian Societies of Endocrine Surgeons and Surgical Oncology Multicentric Study.
The surgical treatment of the intermediate-risk DTC (1-4 cm) remains still controversial. We analyzed the current practice in Italy regarding the surgical management of intermediate-risk unilateral DTC to evaluate risk factors for recurrence and to identify a group of patients to whom propose a total thyroidectomy (TT) vs. hemithyroidectomy (HT). Among 1896 patients operated for thyroid cancer between January 2017 and December 2019, we evaluated 564 (29.7%) patients with unilateral intermediate-risk DTC (1-4 cm) without contralateral nodular lesions on the preoperative exams, chronic autoimmune thyroiditis, familiarity or radiance exposure. Data were collected retrospectively from the clinical register from 16 referral centers. The patients were followed for at least 14 months (median time 29.21 months). In our cohort 499 patients (88.4%) underwent total thyroidectomy whereas 65 patients (11.6%) underwent hemithyroidectomy. 151 (26.8%) patients had a multifocal DTC of whom 57 (10.1%) were bilateral. 21/66 (32.3%) patients were reoperated within 2 months from the first intervention (completion thyroidectomy). Three patients (3/564) developed regional lymph node recurrence 2 years after surgery and required a lymph nodal neck dissection. The single factor related to the risk of reoperation was the histological diameter (HR = 1.05 (1.00-1-09), p = 0.026). Risk stratification is the key to differentiating treatment options and achieving better outcomes. According to the present study, tumor diameter is a strong predictive risk factor to proper choose initial surgical management for intermediate-risk DTC.


Total extract and standardized fraction from the stem bark of Butea monosperma have osteoprotective action: evidence for the nonestrogenic osteogenic effect of the standardized fraction.
The aim of this study was to determine the skeletal effects of Butea total extract (BTE) and its acetone soluble fraction (ASF) from Butea monosperma, which is rich in methoxyisoflavones, in ovariectomized (OVx) rats, a model for postmenopausal bone loss. BTE (1.0 g kg d) and ASF (100 mg kg d) were given orally for 12 weeks to adult OVx rats. The sham-operated and ovariectomy + vehicle groups served as controls. Bone mineral density, osteoid formation (mineral apposition rate and bone formation rate), bone microarchitecture, and bone turnover/resorption markers were studied. Phytoestrogens in rats given BTE and ASF were analyzed by high-performance liquid chromatography. One-way analysis of variance was used to test significance of effects. OVx rats treated with either BTE or ASF exhibited increased bone mineral density in trabecular bones and improved trabecular microarchitecture compared with the ovariectomy + vehicle group. ASF treatment was more efficient than BTE treatment in maintaining trabecular microarchitecture. Serum osteocalcin and urinary type 1 collagen levels in OVx rats treated with either BTE or ASF were significantly lower than those of the ovariectomy + vehicle group. ASF treatment led to increased mineral apposition rate and bone formation rate compared with ovariectomy + vehicle, whereas BTE had no such effect. In the uterotropic assay, BTE was mildly estrogenic in adult OVx rats. In immature rats, BTE exhibited both estrogenicity and antiestrogenicity. ASF had neither uterine estrogenicity nor antiestrogenicity. Analysis of phytoestrogens revealed significant enrichment of cladrin, isoformononetin, and medicarpin in ASF over BTE. Derived from B monosperma, ASF at a 10-fold lower dose than that of BTE was effective in preventing OVx-induced bone loss and stimulated new-bone formation.


Assessing uncertainty in pollutant wash-off modelling via model validation.
Stormwater pollution is linked to stream ecosystem degradation. In predicting stormwater pollution, various types of modelling techniques are adopted. The accuracy of predictions provided by these models depends on the data quality, appropriate estimation of model parameters, and the validation undertaken. It is well understood that available water quality datasets in urban areas span only relatively short time scales unlike water quantity data, which limits the applicability of the developed models in engineering and ecological assessment of urban waterways. This paper presents the application of leave-one-out (LOO) and Monte Carlo cross validation (MCCV) procedures in a Monte Carlo framework for the validation and estimation of uncertainty associated with pollutant wash-off when models are developed using a limited dataset. It was found that the application of MCCV is likely to result in a more realistic measure of model coefficients than LOO. Most importantly, MCCV and LOO were found to be effective in model validation when dealing with a small sample size which hinders detailed model validation and can undermine the effectiveness of stormwater quality management strategies.


Role of the Filters in the Formation and Stabilization of Semiquinone Radicals Collected from Cigarette Smoke.
The fractional pyrolysis of Bright tobacco was performed in nitrogen atmosphere over the temperature range of 240 - 510 °C in a specially constructed, high temperature flow reactor system. Electron paramagnetic resonance (EPR) spectroscopy was used to analyze the free radicals in the initially produced total particular matter (TPM) and in TPM after exposure to ambient air (aging). Different filters have been used to collect TPM from tobacco smoke: cellulosic, cellulose nitrate, cellulose acetate, nylon, Teflon and Cambridge. The collection of the primary radicals (measured immediately after collection of TPM on filters), the formation and stabilization of the secondary radicals (defined as radicals formed during aging of TPM samples on the filters) depend significantly on the material of the filter. A mechanistic explanation about different binding capability of the filters decreasing in the order: cellulosic < cellulose nitrate < cellulose acetate < nylon ~ teflon is presented. Different properties were observed for the Cambridge filter. Specific care must be taken using the filters for identification of radicals from tobacco smoke to avoid artifacts in each case.


ICP, BMI, surgical repair, and CSF diversion in patients presenting with spontaneous CSF otorrhea.
To assess intracranial pressure (ICP), body mass index (BMI), surgical repair, and cerebrospinal fluid (CSF) diversion in patients presenting with spontaneous CSF otorrhea. Retrospective series review. Tertiary referral center. Thirty-two patients were treated surgically from 2004 to 2013 for spontaneous CSF otorrhea by the principal investigators. Patients with a history of chronic ear disease and cholesteatoma, previous mastoid surgery, head trauma, or iatrogenic injury were excluded. Average age was 56 years. Twenty-two patients (69%) were female. Middle fossa repair, transmastoid repair, lumbar puncture, V-P shunt, L-P shunt, and magnetic resonance imaging. Patients underwent middle fossa or transmastoid repair of tegmen defects. Intracranial pressures were determined with lumbar puncture at time of surgical repair or shortly after surgery. CSF diversion procedures were performed in patients who were found to have elevated ICP, which was not controlled medically, presented with recurrent leak or had ICP of 25 cm or greater of H2O. Preoperative BMI was calculated. Thirty-two patients underwent 37 operations. Average BMI was 35.0 kg/m2 (median, 34.7; range, 18.7-53.2 kg/m2). There were 21 repairs on the left and 16 on the right. The majority underwent a middle fossa craniotomy for repair (27/32). Two patients had bilateral repairs. Three patients (8%) underwent revision surgery, of which, 2 had untreated intracranial hypertension (ICP 24.5 and 24 cm H2O). ICP measurements were available for 29 patients. The mean ICP was 23.4 cm H2O (median, 24; range, 13-36 cm H20). Twenty-two patients (69%) had ICP of 20 cm or greater of H20; of those, 13 had an ICP of 25 cm or greater of H20. Seventeen patients (53%) underwent CSF diversion procedures. Our findings of elevated ICP and BMI in patients presenting with spontaneous CSF otorrhea are consistent with previous reports in the literature. The percentage of patients that underwent CSF diversion procedures was high at 53% and represents an aggressive stance in managing elevated ICP in a population that may be at risk for subsequent leaks.


Effects of parity and litter size on cortisol measures in commercially housed sows and their offspring.
Breeding sows are regularly exposed to on-farm stressors throughout the duration of their production period. The impact of such stressors may differ for primi- and multiparous sows, as sows could learn to cope with stressors as they gain experience with them. If parity affects stress in sows, it may also impact their prenatal offspring through differential maternal stress. In addition to parity, litter size is another potential factor involved in stress of sows and piglets. Larger litters may be a source of discomfort for gestating sows, while it can result in intra-uterine growth restriction of piglets. In the current study, we aimed to assess whether parity and litter size affect cortisol measures in breeding sows and their offspring. To do this, we measured salivary cortisol concentrations of 16 primiparous and 16 multiparous sows at three time points: 1) while sows were group housed, 2) after sows were separated from the group prior to moving to the farrowing unit and 3) after handling procedures. In addition, hair cortisol concentration was determined for the sows during late gestation and for their low birth weight (n = 63) and normal birth weight (n = 43) offspring on day 3 after birth, to reflect in-utero cortisol exposure. It was expected that if sows adapt to on-farm stressors, the more experienced, multiparous sows would show decreased stress responses in comparison to primiparous sows. However, we found a comparable acute stress response of primi- and multiparous sows to separation from the group. Handling procedures did not influence sows' salivary cortisol concentrations. Sows' hair cortisol concentration was positively correlated with litter size. Future research is needed to assess whether this finding reflects increased stress in sows carrying larger litters. Parity or litter size did not have a direct effect on their offspring's hair cortisol concentration. Larger litters did have a higher occurrence of low birth weight piglets. For these piglets, females had higher neonatal hair cortisol concentrations than males. Overall, our results indicate that breeding sows do not adapt to all on-farm stressors. In addition, litter size may influence HPA axis activity in both sows and piglets.


An efficient chemoenzymatic approach to (S)-gamma-fluoroleucine ethyl ester.
[reaction: see text] An asymmetric synthesis of (S)-gamma-fluoroleucine ethyl ester 1 is described. The key transformation involves a lipase-catalyzed dynamic ring-opening of 2-(3-butenyl)azlactone 7b with EtOH to give amide ester (S)-6b in 84% enantiomeric excess. Removal of the N-pentenoyl group with N,N'-dibromodimethylhydantoin in the presence of trifluoroacetic acid afforded the titled compound, which was isolated as its hydrogen sulfate salt in 75% yield and >97% ee.


Decline in US stroke mortality in the era before antihypertensive therapy.
This study was done to determine if reported declines in stroke mortality in the era before antihypertensive therapy are artifactual. This study involved analyses of national and state vital statistics data using adjusted and specific rates. Adoption of the third revision of the International List of Causes of Death in 1921 produced an abrupt 6.6% decrease in stroke mortality rates, but otherwise, changes in disease classification systems had little effect on stroke mortality rates. Adoption of the second revision of the joint-cause manual produced a 9.2% drop in stroke death rates, but other revisions of the joint-cause selection rules had little effect. While rates for the expanding group of states in the death registration area progressively declined, rates for fixed component areas remained constant until around 1925 and then declined. Reselection of the underlying cause from aggregate multiple cause data for 1917, 1925, and 1940 using uniform selection rules confirmed a decline after 1925. Correlation analyses of rates of change for stroke and heart disease rates did not support a shift in diagnosis to explain the divergent trends. The apparent decline in stroke mortality rates before 1925 is an artifact of changes in disease classification systems, joint-cause selection rules, and nonrandom incorporation of states with different mortality rates into the expanding registration area. The decline after 1925 could not be explained by changes in coding systems or joint-cause selection rules or by a shift in diagnosis from stroke to heart disease.


The Colors of Health: Chemistry, Bioactivity, and Market Demand for Colorful Foods and Natural Food Sources of Colorants.
There is an increasing consumer demand for natural colors in foods. However, there is a limited number of available natural food sources for use by the food industry because of technical and regulatory limitations. Natural colors are less stable and have less vibrant hues compared to their synthetic color counterparts. Natural pigments also have known health benefits that are seldom leveraged by the food industry. Betalains, carotenoids, phycocyanins, and anthocyanins are major food colorants used in the food industry that have documented biological effects, particularly in the prevention and management of chronic diseases such as diabetes, obesity, and cardiovascular disease. The color industry needs new sources of stable, functional, and safe natural food colorants. New opportunities include sourcing new colors from microbial sources and via the use of genetic biotechnology. In all cases, there is an imperative need for toxicological evaluation to pave the way for their regulatory approval.


Changes in physical activity and all-cause mortality in COPD.
Little is known about changes in physical activity in subjects with chronic obstructive pulmonary disease (COPD) and its impact on mortality. Therefore, we aimed to study changes in physical activity in subjects with and without COPD and the impact of physical activity on mortality risk. Subjects from the Copenhagen City Heart Study with at least two consecutive examinations were selected. Each examination included a self-administered questionnaire and clinical examination. 1270 COPD subjects and 8734 subjects without COPD (forced expiratory volume in 1 s 67±18 and 91±15% predicted, respectively) were included. COPD subjects with moderate or high baseline physical activity who reported low physical activity level at follow-up had the highest hazard ratios of mortality (1.73 and 2.35, respectively; both p<0.001). In COPD subjects with low baseline physical activity, no differences were found in survival between unchanged or increased physical activity at follow-up. In addition, subjects without COPD with low physical activity at follow-up had the highest hazard ratio of mortality, irrespective of baseline physical activity level (p≤0.05). A decline to low physical activity at follow-up was associated with an increased mortality risk in subjects with and without COPD. These observational data suggest that it is important to assess and encourage physical activity in the earliest stages of COPD in order to maintain a physical activity level that is as high as possible, as this is associated with better prognosis.


Heparin and antiaggregating therapy in unstable angina.
Heparin and antiaggregating therapy represent an effective treatment of unstable angina. Aspirin is able to prevent myocardial infarction and death in patients with unstable angina, but is not able to control myocardial ischemia. On the contrary, heparin administered by continuous intravenous infusion (about 1,000 U/h) controls myocardial ischemia by very early lowering anginal attacks and silent ischemic episodes. Heparin intermittently administered (6,000 U i.v. four times daily) did not significantly affect anginal attacks and myocardial ischemia. Thrombolytic therapy alone (Alteplase 1.75 mg/kg) reduced, but not significantly, the number of anginal attacks. The reduction of myocardial ischemia induced by continuous infusion of heparin is associated with a decrease of fibrinopeptide A plasma levels, whereas the thromboxane B2 production by platelets seems to be irrelevant in order to obtain a reduction of myocardial ischemia. Thus thrombin formation seems to be a primary factor in the occurrence of refractory unstable angina and in the activation of platelets.


The nutritive value of biscuits protein-enriched with soy-bean meal.
The researches were carried out on 4 groups of 15 weanling rats and 2 groups of 8 adult rats "pair fed" in individual cages, with a complete isocaloric and isoproteic (10%) diet, the nitrogenous material being supplied for one group from casein, for another from gluten and for the two others this was supplemented with soy-bean meal (5 or 10%). The results obtained in the growth test and in adult rats (nitrogen balance) show that the incorporation of soy-bean into the gluten biscuits increased the gain in weight and the PER lowering the consumption index (p less than 0.001), and doubled the nitrogen retention.


[On the therapeutic methods of facial bone injuries].
The authors present clinicomorphological characteristics of 2293 patients with benign neoplasms of the vocal cords. Nodules predominated in females, contact granulomas in males. Histological and histochemical evidence indicates that polyp-like new growths of the vocal cords known as fibromas, angiofibromas and nodules are not tumors. They are rather edematous, edematous-fibrous and angiomatous polyps arising as a reaction of vocal cords mucosa to phonatory trauma or chemical irritants.


Isolation of an IHF-deficient mutant of a Pseudomonas aeruginosa mucoid isolate and evaluation of the role of IHF in algD gene expression.
The role of integration host factor (IHF) in the regulation of alginate synthesis was investigated in a mucoid strain of Pseudomonas aeruginosa (strain CHA) isolated from a cystic fibrosis patient. Escherichia coli strain BL21(DE3) was made IHF-deficient by inactivation of its chromosomal IHF genes, himA and himD, then used as host strain to overproduce P. aeruginosa IHF. The purified recombinant IHF protein was used to determine the affinity of IHF for the two IHF binding sites in the algD promoter. The Kd values were determined to be 130 nM for algD IHF site 2 and about 2 microM for algD IHF site 1. Two IHF-deficient mutants of P. aeruginosa strain CHA were constructed by insertional inactivation of the himA gene, and the activity of the algD promoter was determined using transcriptional fusion with xylE as reporter gene. The expression of algD, the structural gene for GDP-mannose dehydrogenase, was decreased three- to fourfold in the himA mutants under conditions of high salinity and nitrogen limitation. Assays of alginate production by cultures grown on agar plates indicated that the IHF-deficient mutants synthesized 50% less polymer than the mucoid parental strain. These results demonstrate clearly that although IHF is dispensable for alginate production, himA expression is required for full activation of algD expression.


1,25-Dihydroxyvitamin D(3) prevents puromycin aminonucleoside-induced apoptosis of glomerular podocytes by activating the phosphatidylinositol 3-kinase/Akt-signaling pathway.
Accumulating evidence suggests that vitamin D and its analogs reduce proteinuria and slow the decline in kidney function in chronic kidney disease. Given a rich literature identifying podocyte apoptosis as an early step in the pathophysiological progression to proteinuria and glomerulosclerosis, we hypothesized that vitamin D protects podocytes from undergoing apoptosis. A rat model of podocyte apoptosis was created by a single intravenous injection of 100 mg x kg(-1) puromycin aminonucleoside (PAN) and received either solvent or 1,25(OH)(2)D(3) treatment. Proteinuria, podocyte apoptosis, the expression of nephrin protein and mRNA, TGF-beta/Smad and phosphatidylinositol 3-kinase (PI3K)/Akt-signaling pathway were evaluated, respectively. PAN induced massive proteinuria, serum creatinine elevation and podocyte apoptosis in PAN nephropathy rats, which was associated with the loss of nephrin, an adhesion molecule specific for the glomerular slit and the reduced of p-Akt/Akt ratio. Moreover, PAN induced foot process retraction, redistribution of nephrin and the activation of TGF-beta/Smad-signaling pathway. Compared with PAN nephropathy rats, 1,25(OH)(2)D(3) significantly prevented loss of nephrin, foot process retraction and podocyte apoptosis by stimulating Akt phosphorylation and suppressing TGF-beta/Smad-signaling pathway. 1,25(OH)(2)D(3) reduced the PAN-induced podocyte apoptosis and loss of nephrin in PAN nephropathy rat. The anti-apoptotic effects of 1,25(OH)(2)D(3 )on podocytes may be partly attributable to activation of a PI3K/Akt survival pathway.


Characterization of a new class of tetracycline-resistance gene tet(S) in Listeria monocytogenes BM4210.
The nucleotide sequence of the tetracycline (Tc)-minocycline (Mc)-resistance determinant of plasmid pIP811 from Listeria monocytogenes BM4210 has been determined. The gene, designated tet(S), was identified by analysis of the start and stop codons as a coding sequence of 1923 bp, corresponding to a protein with a calculated M(r) of 72,912. The apparent 68-kDa size estimated by sodium dodecyl sulfate-polyacrylamide-gel electrophoresis of the protein characterized in a cell-free coupled transcription-translation system was in good agreement with the calculated value. The tet(S) gene product exhibits 79 and 72% amino acid identity with Tet(M) from Streptococcus pneumoniae and Tet(O) from Campylobacter coli, respectively. The distribution of tet(S) in strains of Gram+ and Gram- genera resistant to Tc (TcR) and Mc (McR) was studied by hybridization under high stringency using a 590-bp intragenic probe. Homology with tet(S) was detected in two clinical isolates of L. monocytogenes isolated in different geographical areas.


The biomechanics of fast-starts during ontogeny in the common carp cyprinus carpio
Common carp Cyprinus carpio L. were reared a constant temperature of 20 degrees C from the larval (7 mm total length) to the juvenile (80 mm) stage. Body morphology and white muscle mass distribution were measured. Fast-start escape responses were recorded using high-speed cinematography from which the velocities, accelerations and hydrodynamic power requirements were estimated. All three measures of fast-start performance increased during development. White muscle contraction regimes were calculated from changes in body shape during the fast-starts and used to predict the muscle force and power production for all longitudinal positions along the body. Scaling arguments predicted that increases in body length would constrain the fish to bend less rapidly because the cross-sectional muscle area, and hence force production, does not increase at the same rate as the inertial mass that resists bending. As predicted, the increases in body length resulted in decreases in muscle shortening velocity, and this coincided with increases in both the force and power produced by the muscles. The hydrodynamic efficiency, which relates the mechanical power produced by the muscles to the inertial power requirements in the direction of travel, showed no significant change during ontogeny. The increasing hydrodynamic power requirements were thus met by increases in the power available from the muscles. The majority of the increases in fast-start swimming performance during ontogeny can be explained by size-dependent increases in muscle power output. For all sizes, there was a decrease in muscle-mass-specific power output and an increase in muscle stress in a posterior direction along the body due to systematic variations in fibre strain. These changing strain regimes result in the central muscle bulk producing the majority of the power requirements during the fast-start, and this power is transmitted to the tail region of the fish and ultimately to the water via muscle in the caudal myotomes.


Chemical composition and antimicrobial and spasmolytic properties of Poliomintha longiflora and Lippia graveolens essential oils.
In the present study, we reported a comparative analysis of the chemical composition and pharmacological properties of the essential oils obtained from 2 Mexican oreganos, Poliomintha longiflora and Lippia graveolens. The gas chromatography-mass spectrometry (GC-MS) profiles of the oils showed high amounts of oxygenated monoterpenes, mainly carvacrol (%[mg/100 g dry matter]) (18.36 [459.0] in P. longiflora and 13.48 [164.7] in L. graveolens). In addition, these oils contained marked quantities of p-cymene (14.09 [352.2] and 7.46 [37.3], respectively), β-caryophyllene oxide, β-caryophyllene, and carvacrol acetate. Headspace analyses of the leaves of both species using different coated fibers revealed that γ-terpinene, eucalyptol, and p-cymene were the principal light volatile components. Chromatographic fingerprints and a suitable analytical method for quantifying the main components of both essences were established using high-performance liquid chromatography (HPLC) as analytical tool. The essential oils of both species were not toxic in the acute toxicity studies in mice performed according to the Lorke procedure (DL(50) > 5000 mg/kg). The oils and the major constituents, carvacrol and p-cymene, displayed a moderate in vitro antibacterial activity, with minimum inhibitory concentration values ranging from 128 to 512 μg/mL. In addition, these samples demonstrated a marginal antispasmodic activity in vivo and provoked a concentration-dependent inhibition of the carbachol- and histamine-induced contractions using the isolated guinea-pig ileum preparation. In particular, p-cymene exerts good selective inhibitory activity on the carbachol-induced contractions (IC(50) = 9.85 μg/mL). The analytical methods using GC-MS and HPLC techniques will be useful for establishing quality control as well as preclinical pharmacological and toxicological parameters of the crude drug P. longiflora, which is widely used as substitute of L. graveolens for medicinal and flavorings purposes. This overall information will be also useful for elaborating scientific and pharmacopoeic monographs of this very Mexican medicinal plant.


Contributions to the biodiversity of Vietnam - Results of VIETBIO inventory work and field training in Cuc Phuong National Park.
VIETBIO [Innovative approaches to biodiversity discovery and characterisation in Vietnam] is a bilateral German-Vietnamese research and capacity building project focusing on the development and transfer of new methods and technology towards an integrated biodiversity discovery and monitoring system for Vietnam. Dedicated field training and testing of innovative methodologies were undertaken in Cuc Phuong National Park as part and with support of the project, which led to the new biodiversity data and records made available in this article collection. VIETBIO is a collaboration between the Museum für Naturkunde Berlin - Leibniz Institute for Evolution and Biodiversity Science (MfN), the Botanic Garden and Botanical Museum, Freie Universität Berlin (BGBM) and the Vietnam National Museum of Nature (VNMN), the Institute of Ecology and Biological Resources (IEBR), the Southern Institute of Ecology (SIE), as well as the Institute of Tropical Biology (ITB); all Vietnamese institutions belong to the Vietnam Academy of Science and Technology (VAST). The article collection "VIETBIO" (https://doi.org/10.3897/bdj.coll.63) reports original results of recent biodiversity recording and survey work undertaken in Cuc Phuong National Park, northern Vietnam, under the framework of the VIETBIO project. The collection consist of this "main" cover paper - characterising the study area, the general project approaches and activities, while also giving an extensive overview on previous studies from this area - followed by individual papers for higher taxa as studied during the project. The main purpose is to make primary biodiversity records openly available, including several new and interesting findings for this biodiversity-rich conservation area. All individual data papers with their respective primary records are expected to provide useful baselines for further taxonomic, phylogenetic, ecological and conservation-related studies on the respective taxa and, thus, will be maintained as separate datasets, including separate GUIDs also for further updating.


Responsiveness of the Duke Social Support sub-scales in older women.
an abbreviated form of the Duke Social Support Index (DSSI) as used in a large longitudinal study of older Australian women was examined with respect to factors that might be expected to affect social support for older women over time. in this large cohort study two sub-scales of the DSSI, one describing the size and structure of the social network (four items) and the other perceiving satisfaction with social support (six items), were analysed in relation to outcome and exploratory variables. over a 3-year period, the network score increased among women whose life circumstances meant that they were likely to receive more support (e.g. recent widowhood). Likewise, those women at risk of becoming more socially isolated (e.g. those with sensory loss) became less satisfied with their social support. Changes in both measures were tempered by women's mental health and optimism. although the sub-scales of the DSSI may not fully reflect the complexity of social support paradigms, they are responsive to changes in the lives of older women and can be useful in community-based epidemiological studies.


Effect of two different concentrations of sodium hypochlorite on postoperative pain following single-visit root canal treatment: a triple-blind randomized clinical trial.
To compare the effects of two different concentrations of NaOCl solution on postoperative pain following single-visit root canal treatment in mandibular molars with irreversible pulpitis. A total of 122 patients who had mandibular molars with irreversible pulpitis were treated. The patients were randomly divided into two groups according to the concentration of NaOCl used during root canal instrumentation - either 2.5% or 5.25%. RaCe rotary instruments were used for root canal preparation, and all root canals were filled in one visit. Postoperative pain was evaluated using the visual analogue scale. Data were analysed by independent t-test, chi-square and Mann-Whitney tests. Twelve patients were excluded for various reasons. Pain reported by 110 patients who were eligible to be included in the study was analysed. No significant differences were found in the age and gender of the patients between the two groups (P = 0.50, P = 0.51, respectively). The patients who had 5.25% NaOCl reported significantly lower postoperative pain compared to those who had 2.5% NaOCl during the first 72 h following treatment (P = 0.021); however, there was no significant difference in pain felt by the patients during the rest of the study period, that is 4-7 days following treatment (P = 0.185) when the four-level pain categorization method was used. When the two-level pain categorization method was used, the results revealed that patients who had 5.25% NaOCl reported significantly less pain for the first 3 days after treatment (P = 0.026). The number of analgesics taken by patients who had 2.5% NaOCl was significantly higher than that taken by patients who had 5.25% NaOCl (P = 0.001). 5.25% NaOCl was associated with significantly lower postoperative pain compared to 2.5% NaOCl during the first 72 h following one-visit root canal treatment of mandibular molars with irreversible pulpitis.


Brain region-specific expression of Fxyd1, an Mecp2 target gene, is regulated by epigenetic mechanisms.
Fxyd1 encodes a trans-membrane protein that modulates Na(+) ,K(+) -ATPase activity and is a substrate for multiple protein kinases. Fxyd1 expression is repressed by methyl CpG-binding protein 2 (Mecp2) in the frontal cortex (FC) but not in the cerebellum (CB) of the mouse brain. Consistently with these observations, FXYD1 mRNA abundance is increased in the FC of Rett syndrome (RTT) patients with MECP2 mutations. Because Fxyd1 is implicated in the regulation of neuronal excitability, understanding how Fxyd1 expression is controlled is important. Here we report that basal expression of Fxyd1a and Fxyd1b, the two main alternatively spliced forms of Fxyd1 mRNA, is lower in the FC than in the CB. This difference is accompanied by increased Mecp2 recruitment to the promoter region of these two Fxyd1 mRNA forms. DNA methylation of both promoters is more frequent in the FC than in the CB, and in both cases the most frequently methylated CpG dinucleotides are adjacent to [A/T](4) sequences required for high-affinity Mecp2 binding. Consistently with these features of epigenetic silencing, histone 3 acetylated at lysines 9 and 14 (H3K9/14ac) and histone 3 methylated at lysine 4 (H3K4me3), both activating histone marks, were associated with the Fxyd1 promoter to a lesser degree in the FC than in the CB. These results indicate that differential Fxyd1 expression in these two brain regions is, at least in part, regulated by an epigenetic mechanism involving increased DNA methylation of the two alternative Fxyd1 promoters, enhanced Mecp2 recruitment, and reduced association of activating histones.


Persistence of selectable herpesvirus saimiri in various human haematopoietic and epithelial cell lines.
Herpesvirus (h.) saimiri, an infectious agent of squirrel monkeys, is capable of persisting in T lymphocytes of various primate species. It has been used as a vector for the functional analysis of regulatory genes in primary human T lymphocytes. As it is not yet known whether other cell types are capable of supporting viral persistence, various human cell lines were investigated using selectable h. saimiri recombinants. The lines chosen represent cells from the epithelium and connective tissue as well as from all haematopoietic lineages, i.e. cells of B and T lymphoid origin as well as myeloid-, fibroblast- and carcinoma-derived cultures converted to Geneticin or hygromycin B resistance, and harbouring episomal DNA of the selectable recombinants. The Burkitt's lymphoma-derived cell line Raji also contained simultaneously persisting episomes of the Epstein-Barr virus. Most of the cell cultures except a pancreatic carcinoma line and foreskin fibroblasts did not produce infectious virus. These observations show that a herpesvirus genome can persist episomally in a broad range of cultured cell types. The variety of infectable cell types and species suggests the presence of a widely distributed and well conserved virus receptor for h. saimiri. Thus the h. saimiri genome could be applied more generally as a vector.


Common salt aggravated pathology of testosterone-induced benign prostatic hyperplasia in adult male Wistar rat.
Benign prostatic hyperplasia (BPH) is a major health concern associated with lower urinary tract symptoms and sexual dysfunction in men. Recurrent inflammation, decreased apoptotic rate and oxidative stress are some of the theories that explain the pathophysiology of BPH. Common salt, a food additive, is known to cause systemic inflammation and redox imbalance, and may serve as a potential risk factor for BPH development or progression. This study examined the effect of common salt intake on the pathology of testosterone-induced BPH. Forty male Wistar rats were randomly divided into four equal groups of 10: a control and three salt diet groups-low-salt diet (LSD), standard-salt diet (SSD) and high-salt diet (HSD). The rats were castrated, allowed to recuperate and placed on salt-free diet (control), 0.25% salt diet (LSD), 0.5% salt diet (SSD) and 1.25% salt diet (HSD) for 60 days ad libitum. On day 33, BPH was induced in all the rats with daily injections of testosterone propionate-Testost® (3 mg/kg body weight) for 28 days. The rats had overnight fast (12 h) on day 60 and were euthanized the following day in order to collect blood and prostate samples for biochemical, molecular and immunohistochemistry (IHC) analyses. Mean ± SD values were calculated for each group and compared for significant difference with ANOVA followed by post hoc test (Tukey HSD) at p < 0.05. This study recorded a substantially higher level of IL-6, IL-8 and COX-2 in salt diet groups and moderate IHC staining of COX-2 in HSD group. The prostatic level of IL-17, IL-1β, PGE2, relative prostate weight and serum PSA levels were not statistically different. The concentrations of IGF-1, TGF-β were similar in all the groups but there were multiple fold increase in Bcl-2 expression in salt diet groups-LSD (13.2), SSD (9.5) and HSD (7.9) and multiple fold decrease in VEGF expression in LSD (-6.3), SSD (-5.1) and HSD (-14.1) compared to control. Activity of superoxide dismutase (SOD) and concentration of nitric oxide rose in LSD and SSD groups, and SSD and HSD groups respectively. Activities of glutathione peroxidase and catalase, and concentration of NADPH and hydrogen peroxide were not significantly different. IHC showed positive immunostaining for iNOS expression in all the groups while histopathology revealed moderate to severe prostatic hyperplasia in salt diet groups. These findings suggest that low, standard and high salt diets aggravated the pathology of testosterone-induced BPH in Wistar rats by promoting inflammation, oxidative stress, while suppressing apoptosis and angiogenesis.


A comparison of thin-plate spline deformation and finite element modeling to compensate for brain shift during tumor resection.
Brain shift during tumor resection can progressively invalidate the accuracy of neuronavigation systems and affect neurosurgeons' ability to achieve optimal resections. This paper compares two methods that have been presented in the literature to compensate for brain shift: a thin-plate spline deformation model and a finite element method (FEM). For this comparison, both methods are driven by identical sparse data. Specifically, both methods are driven by displacements between automatically detected and matched feature points from intraoperative 3D ultrasound (iUS). Both methods have been shown to be fast enough for intraoperative brain shift correction (Machado et al. in Int J Comput Assist Radiol Surg 13(10):1525-1538, 2018; Luo et al. in J Med Imaging (Bellingham) 4(3):035003, 2017). However, the spline method requires no preprocessing and ignores physical properties of the brain while the FEM method requires significant preprocessing and incorporates patient-specific physical and geometric constraints. The goal of this work was to explore the relative merits of these methods on recent clinical data. Data acquired during 19 sequential tumor resections in Brigham and Women's Hospital's Advanced Multi-modal Image-Guided Operating Suite between December 2017 and October 2018 were considered for this retrospective study. Of these, 15 cases and a total of 24 iUS to iUS image pairs met inclusion requirements. Automatic feature detection (Machado et al. in Int J Comput Assist Radiol Surg 13(10):1525-1538, 2018) was used to detect and match features in each pair of iUS images. Displacements between matched features were then used to drive both the spline model and the FEM method to compensate for brain shift between image acquisitions. The accuracies of the resultant deformation models were measured by comparing the displacements of manually identified landmarks before and after deformation. The mean initial subcortical registration error between preoperative MRI and the first iUS image averaged 5.3 ± 0.75 mm. The mean subcortical brain shift, measured using displacements between manually identified landmarks in pairs of iUS images, was 2.5 ± 1.3 mm. Our results showed that FEM was able to reduce subcortical registration error by a small but statistically significant amount (from 2.46 to 2.02 mm). A large variability in the results of the spline method prevented us from demonstrating either a statistically significant reduction in subcortical registration error after applying the spline method or a statistically significant difference between the results of the two methods. In this study, we observed less subcortical brain shift than has previously been reported in the literature (Frisken et al., in: Miller (ed) Biomechanics of the brain, Springer, Cham, 2019). This may be due to the fact that we separated out the initial misregistration between preoperative MRI and the first iUS image from our brain shift measurements or it may be due to modern neurosurgical practices designed to reduce brain shift, including reduced craniotomy sizes and better control of intracranial pressure with the use of mannitol and other medications. It appears that the FEM method and its use of geometric and biomechanical constraints provided more consistent brain shift correction and better correction farther from the driving feature displacements than the simple spline model. The spline-based method was simpler and tended to give better results for small deformations. However, large variability in the spline results and relatively small brain shift prevented this study from demonstrating a statistically significant difference between the results of the two methods.


Critical role of TXNIP in oxidative stress, DNA damage and retinal pericyte apoptosis under high glucose: implications for diabetic retinopathy.
Diabetic retinopathy (DR) is characterized by early loss of retinal capillary pericytes and microvascular dysfunction. We recently showed that pro-oxidative stress and pro-apoptotic thioredoxin interacting protein (TXNIP) is significantly up-regulated in rat retinas in experimental diabetes and mediates inflammation and apoptosis. Therefore, we hypothesize here that TXNIP up-regulation in pericyte plays a causative role in oxidative stress and apoptosis under sustained high glucose exposure in culture. We maintained a rat retinal capillary pericyte cell line (TR-rPCT1) for 5 days under low glucose (LG, 5.5mM) or high glucose (HG, 25 mM) with or without anti-oxidant N-acetylcysteine (5mM, NAC), Azaseine (2 μM, AzaS), an inhibitor of TXNIP, and TXNIP siRNA (siTXNIP3, 20 nM). The results show that HG increases TXNIP expression in TR-rPCT1, which correlates positively with ROS generation, protein S-nitrosylation, and pro-apoptotic caspase-3 activation. Furthermore, pericyte apoptosis is demonstrated by DNA fragmentation (alkaline comet assay) and a reduction in MTT survival assay. Treatment of TR-rPCT1 with NAC or an inhibition of TXNIP by AzaS or siTXNIP3 each reduces HG-induced ROS, caspase-3 activation and DNA damage demonstrating that TXNIP up-regulation under chronic hyperglycemia is critically involved in cellular oxidative stress, DNA damage and retinal pericyte apoptosis. Thus, TXNIP represents a novel gene and drug target to prevent pericyte loss and progression of DR.


Physical activity promotion in primary care: bridging the gap between research and practice.
While a growing literature supports the effectiveness of physical activity interventions delivered in the primary care setting, few studies have evaluated efforts to increase physician counseling on physical activity during routine practice (i.e., outside the context of controlled research). This paper reports the results of a dissemination trial of a primary care-based physical activity counseling intervention conducted within the context of a larger, multi-strategy, Australian community-based, physical activity intervention, the 10,000 Steps Rockhampton Project. All 23 general practices and 66 general practitioners (GPs, the Australian equivalent of family physicians) were invited to participate. Practice visits were made to consenting practices during which instruction in brief physical activity counseling was offered, along with physical activity promotion resources (print materials and pedometers). The evaluation, guided by the RE-AIM framework, included collection of process data, as well as pre- and post-intervention data from a mailed GP survey, and data from the larger project's random-digit-dialed, community-based, cross-sectional telephone survey that was conducted in Rockhampton and a comparison community. Ninety-one percent of practices were visited by 10,000 Steps staff and agreed to participate, with 58% of GPs present during the visits. General practitioner survey response rates were 67% (n =44/66 at baseline) and 71% (n =37/52, at 14-month follow-up). At follow-up, 62% had displayed the poster, 81% were using the brochures, and 70% had loaned pedometers to patients, although the number loaned was relatively small. No change was seen in GP self-report of the percentage of patients counseled on physical activity. However, data from the telephone surveys showed a 31% increase in the likelihood of recalling GP advice on physical activity in Rockhampton (95% confidence interval [CI]=1.11-1.54) compared to a 16% decrease (95% CI=0.68-1.04) in the comparison community. This dissemination study achieved high rates of GP uptake, reasonable levels of implementation, and a significant increase in the number of community residents counseled on physical activity. These results suggest that evidence-based primary care physical activity counseling protocols can be translated into routine practice, although the initial and ongoing investment of time to develop partnerships with relevant healthcare organizations, and the interest generated by the overall 10,000 Steps program should not be underestimated.


Mechanism of inactivation of TEM-1 beta-lactamase by 6-acetylmethylenepenicillanic acid.
The interaction between 6-acetylmethylenepenicillanic acid (compound Ro 15-1903; AMPA) and TEM-1 beta-lactamase was investigated in order to elucidate the mechanism of action of AMPA. Formation of the enzyme-inhibitor complex (EA) was accompanied by a shift of the absorbance maximum from 292 nm to 303 nm and an increase in the absorption. Regeneration of activity was very slow and incomplete, reaching about one-third of the initial activity after 48 h at 37 degrees C. This behaviour indicated a branched pathway of the decay of the inactivated enzyme. Kinetic and isoelectric-focusing experiments proved this assumption. The first-order constant of regeneration of active enzyme was 6 X 10(-6)-10 X 10(-6) s-1, whereas the rate constant leading to inactive enzyme (EA') was 10 X 10(-6)-15 X 10(-6) s-1 at pH 7.0. Both constants became larger at higher pH. Inactive enzyme (EA') consisted of two major species, with pI 5.36 (EA'1) and 5.30 (EA'2). The former increased at the beginning of incubation but decreased after prolonged incubation. From consideration of these results and previous data [Arisawa & Then (1983) Biochem. J. 209, 609-615], a likely mechanism of inactivation of TEM-1 beta-lactamase by AMPA is discussed.


Endocytosis by the asialoglycoprotein receptor is independent of cytoplasmic serine residues.
The human asialoglycoprotein (ASGP) receptor, like most other plasma membrane receptors, has previously been shown to be phosphorylated at serine residues within the cytoplasmic domain. Phorbol esters, which activate protein kinase C, cause hyperphosphorylation and down-regulation of the ASGP receptor in HepG2 cells. To test the importance of serine residues for receptor traffic and function, we have mutated all the cytoplasmic serines of the two receptor subunits H1 (at positions 16 and 37) and H2 (at positions 12, 13, and 55) to alanines or glycines. Stable transfected fibroblast cell lines expressing either mutant H1 alone or both mutant subunits together were created and compared to cell lines expressing the respective wild-type proteins. Mutant and wild-type subunits were found to have very similar distributions between the cell surface and intracellular compartments. Constitutive internalization of H1 alone and ligand uptake and degradation by cells expressing both receptor subunits were not affected by the mutations. Cytoplasmic serines and serine phosphorylation are thus not essential for receptor function and intracellular traffic. Analysis of individual serine mutations identified serine-12 of subunit H2 as the major site of phosphorylation in the ASGP receptor.


Immunotherapy for ovarian cancer.
All work referenced herein relates to treatment of epithelial ovarian carcinomas, as their treatment differs from ovarian germ cell cancers and other rare ovarian cancers, the treatments of which are addressed elsewhere. Fallopian tube cancers and primary peritoneal adenocarcinomatosis are also generally treated as epithelial ovarian cancers. The standard of care initial treatment of advanced stage epithelial ovarian cancer is optimal debulking surgery as feasible plus chemotherapy with a platinum plus a taxane agent. If this front-line approach fails, as it too often the case, several FDA-approved agents are available for salvage therapy. However, because no second-line therapy for advanced-stage epithelial ovarian cancer is typically curative, we prefer referral to clinical trials as logistically feasible, even if it means referring patients outside our system. Immune therapy has a sound theoretical basis for treating carcinomas generally, and for treating ovarian cancer in particular. Advances in understanding the immunopathogenic basis of ovarian cancer, and the immunopathologic basis for prior failures of immunotherapy for it and other carcinomas promises to afford novel treatment approaches with potential for significant efficacy, and reduced toxicities compared with cytotoxic agents. Thus, referral to early phase immunotherapy trials for ovarian cancer patients that fail conventional treatment merits consideration.


Marginal Adaptation of Provisional CAD/CAM Restorations Fabricated Using Various Simulated Digital Cement Space Settings.
The ideal digital cement space value for the fabrication of provisional computer-aided design/computer-aided manufacturing (CAD/CAM) crowns with clinically acceptable marginal adaptation is not well known. The aim of this study was to evaluate the effect of different simulated cement space settings on the marginal fit of poly(methyl methacrylate) (PMMA) provisional CAD/CAM restorations. An extracted premolar tooth was prepared using ceramic crown preparation guidelines and represented both natural teeth and/or custom implant abutments. The prepared tooth abutment was scanned with a three-dimensional (3D) laboratory scanner (D900, 3Shape). CAD design software was used to subsequently design a premolar crown core with three different simulated cement space settings (20 to 40 μm, 20 to 50 μm, 20 to 60 μm). PMMA blocks were used to mill the specimens (n = 9, N = 27). Using a stereo zoom microscope, a total of 36 images for each of the 3 groups (9 crowns per group, 4 sites per crown) were captured to measure the mean vertical marginal discrepancy for every group. One-way analysis of variance (ANOVA) was used to analyze the data, and the post hoc Tukey multiple comparison test was performed. The marginal gap values of the PMMA cores fabricated using the three cement space settings were significantly different from each other (P < .001). The marginal gap was smaller with a 20- to 60-μm setting compared with 20 to 50 μm and 20 to 40 μm, and the 20- to 50-μm setting allowed for smaller marginal gaps compared with 20 to 40 μm (P < .001). Within the limitations of this study, the marginal gaps of CAD/CAM-fabricated PMMA cores were smaller when the cement space was larger. The smallest marginal gaps were achieved when a 20- to 60-μm cement space was used (P < .001).