Covalent and non-covalent interaction of myofibrillar protein and cyanidin-3-O-glucoside: focus on structure, binding sites and in vitro digestion properties.
The aim of this study was to investigate the effects of covalent and non-covalent interactions between myofibrillar protein (MP) and cyanidin-3-O-glucoside (C3G) on protein structure, binding sites, and digestion properties. Four methods of inducing covalent cross-linking were used in the preparation of MP-C3G conjugates, including tyrosinase-catalyzed oxidation, alkaline pH shift treatment, free radical grafting, and ultrasonic treatment. A comparison was made between MP-C3G conjugates and complexes, and the analysis included sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), C3G binding ratio, liquid chromatography-tandem mass spectrometry (LC-MS/MS), protein side-chain amino acids, circular dichroism spectroscopy, three-dimensional fluorescence, particle size, and in vitro simulated digestion. Covalent bonding between C3G and amino acid side chains in MP was confirmed by LC-MS/MS. In covalent bonding, tryptophan residues, free amino groups and sulfhydryl groups were all implicated. Among the 22 peptides covalently modified by C3G, 30 modification sites were identified, located in lysine, histidine, tryptophan, arginine and cysteine. In vitro simulated digestion experiments showed that the addition of C3G significantly reduced the digestibility of MP, with the covalent conjugate showing lower digestibility than the non-covalent conjugate. Moreover, the digestibility of protein decreased more during intestinal digestion, possibly because covalent cross-linking of C3G and MP further inhibited trypsin targeting sites (lysine and arginine). Covalent cross-linking of C3G with myofibrillar proteins significantly affected protein structure and reduced protein digestibility by occupying more trypsin binding sites. © 2023 Society of Chemical Industry.


pH-dependent interaction of rhodopsin with cyanidin-3-glucoside. 1. Structural aspects.
Anthocyanins are a class of natural compounds common in flowers and vegetables. Because of the increasing preference of consumers for food containing natural colorants and the demonstrated beneficial effects of anthocyanins on human health, it is important to decipher the molecular mechanisms of their action. Previous studies indicated that the anthocyanin cyanidin-3-glucoside (C3G) modulates the function of the photoreceptor rhodopsin. In this paper, we show using selective excitation (1)H NMR spectroscopy that C3G binds to rhodopsin. Ligand resonances broaden upon rhodopsin addition and rhodopsin resonances exhibit chemical shift changes as well as broadening effects in specific resonances, in an activation state-dependent manner. Furthermore, dark-adapted and light-activated states of rhodopsin show preferences for different C3G species. Molecular docking studies of the flavylium cation, quinoidal base, carbinol pseudobase and chalcone forms of C3G to models of the dark, light-activated and opsin structures of rhodopsin also support this conclusion. The results provide new insights into anthocyanin-protein interactions and may have relevance for the enhancement of night vision by this class of compounds. This work is also the first report of the study of ligand binding to a full-length membrane receptor in detergent micelles by (1)H NMR spectroscopy. Such studies were previously hampered by the presence of detergent micelle resonances, a problem overcome by the selective excitation approach.


Glycoproteomic Analysis of MGL-Binding Proteins on Acute T-Cell Leukemia Cells.
C-type lectins are a diverse group of proteins involved in many human physiological and pathological processes. Most C-type lectins are glycan-binding proteins, some of which are pivotal for innate immune responses against pathogens. Other C-type lectins, such as the macrophage galactose-type lectin (MGL), have been shown to induce immunosuppressive responses upon the recognition of aberrant glycosylation on cancer cells. MGL is known to recognize terminal N-acetylgalactosamine (GalNAc), such as the Tn antigen, which is commonly found on malignant cells. Even though this glycan specificity of MGL is well described, there is a lack of understanding of the actual glycoproteins that bind MGL. We present a glycoproteomic workflow for the identification of MGL-binding proteins, which we applied to study MGL ligands on the human Jurkat leukemia cell line. In addition to the known MGL ligands and Tn antigen-carrying proteins CD43 and CD45 on these cells, we have identified a set of novel cell-surface ligands for MGL. Importantly, for several of these, O-glycosylation has hitherto not been described. Altogether, our data provide new insight into the identification and structure of novel MGL ligands that presumably act as modulatory molecules in cancer immune responses.


Lipid rafts and redox signaling.
In addition to the amplifying action of enzymes in the cell-signaling cascade, another important mechanism has been shown to amplify the signals massively when ligands bind to their receptors, which is characterized by clustering of membrane lipid microdomains or lipid rafts and formation of various signaling platforms. In this process, many receptor molecules aggregate on stimulation, thereby resulting in a very high density of the receptors and other signaling molecules to form signaling platforms and transmit and amplify the signals from receptor activation. Recent studies have indicated lipid rafts or lipid microdomain platforms may be importantly implicated in redox signaling of a variety of cells in response to agonists or stimuli. In this forum, we collected four original research communications, five review articles, and one news or views report to summarize recent progress in this research area. Information is offered for further understanding of the formation and function of lipid rafts and ceramide-enriched platforms and their roles in redox signaling. We hope that this forum could lead to more studies in this area and enhance our understanding of lipid rafts and redox regulation under physiologic and pathologic conditions.


Fas-induced B cell apoptosis requires an increase in free cytosolic magnesium as an early event.
Ligation of the Fas molecule expressed on the surface of a cell initiates multiple signaling pathways that result in the apoptotic death of that cell. We have examined Mg2+ mobilization as well as Ca2+ mobilization in B cells undergoing Fas-initiated apoptosis. Our results indicate that cytosolic levels of free (non-complexed) Mg2+ ([Mg2+]i) and Ca2+ ([Ca2+]i) increase in cells undergoing apoptosis. Furthermore, the percentages of cells mobilizing Mg2+, fragmenting DNA, or externalizing phosphatidylserine (PS) increase in parallel as the concentration of anti-Fas monoclonal antibody is raised. Kinetic analysis suggests that Mg2+ mobilization is an early event in apoptosis, clearly preceding DNA fragmentation and probably occurring prior to externalization of PS as well. The source of Mg2+ that produces the increases in [Mg2+]i is intracellular and most likely is the mitochondria. Extended pretreatment of B cells with carbonyl cyanide m-chlorophenylhydrazone, an inhibitor of mitochondrial oxidative phosphorylation, produces proportional decreases in the percentage of cells mobilizing Mg2+, fragmenting DNA, and externalizing PS in response to anti-Fas monoclonal antibody treatment. These observations are consistent with the hypothesis that elevated [Mg2+]i is required for apoptosis. Furthermore, we propose that the increases in [Mg2+]i function not only as cofactors for Mg2+-dependent endonucleases, but also to facilitate the release of cytochrome c from the mitochondria, which drives many of the post-mitochondrial, caspase-mediated events in apoptotic cells.


Motor Control Stabilisation Exercise for Patients with Non-Specific Low Back Pain: A Prospective Meta-Analysis with Multilevel Meta-Regressions on Intervention Effects.
Low-to-moderate quality meta-analytic evidence shows that motor control stabilisation exercise (MCE) is an effective treatment of non-specific low back pain. A possible approach to overcome the weaknesses of traditional meta-analyses would be that of a prospective meta-analyses. The aim of the present analysis was to generate high-quality evidence to support the view that motor control stabilisation exercises (MCE) lead to a reduction in pain intensity and disability in non-specific low back pain patients when compared to a control group. In this prospective meta-analysis and sensitivity multilevel meta-regression within the MiSpEx-Network, 18 randomized controlled study arms were included. Participants with non-specific low back pain were allocated to an intervention (individualized MCE, 12 weeks) or a control group (no additive exercise intervention). From each study site/arm, outcomes at baseline, 3 weeks, 12 weeks, and 6 months were pooled. The outcomes were current pain (NRS or VAS, 11 points scale), characteristic pain intensity, and subjective disability. A random effects meta-analysis model for continuous outcomes to display standardized mean differences between intervention and control was performed, followed by sensitivity multilevel meta-regressions. Overall, 2391 patients were randomized; 1976 (3 weeks, short-term), 1740 (12 weeks, intermediate), and 1560 (6 months, sustainability) participants were included in the meta-analyses. In the short-term, intermediate and sustainability, moderate-to-high quality evidence indicated that MCE has a larger effect on current pain (SMD = -0.15, -0.15, -0.19), pain intensity (SMD = -0.19, -0.26, -0.26) and disability (SMD = -0.15, -0.27, -0.25) compared with no exercise intervention. Low-quality evidence suggested that those patients with comparably intermediate current pain and older patients may profit the most from MCE. Motor control stabilisation exercise is an effective treatment for non-specific low back pain. Sub-clinical intermediate pain and middle-aged patients may profit the most from this intervention.


Mitochondrial instability during regional ischemia-reperfusion underlies arrhythmias in monolayers of cardiomyocytes.
Regional depolarization of the mitochondrial network can alter cellular electrical excitability and increase the propensity for reentry, in part, through the opening of sarcolemmal KATP channels. Mitochondrial inner membrane potential (ΔΨm) instability or oscillation can be induced in myocytes by exposure to reactive oxygen species (ROS), laser excitation, or glutathione depletion, and is thought to be a major factor in arrhythmogenesis during ischemia-reperfusion. Nevertheless, the correlation between ΔΨm recovery kinetics and reperfusion-induced arrhythmias has been difficult to demonstrate experimentally. Here, we investigate the relationship between subcellular changes in ΔΨm, cellular glutathione redox potential, electrical excitability, and wave propagation during coverslip-induced ischemia-reperfusion (IR) in neonatal rat ventricular myocyte (NRVM) monolayers. Ischemia led to decreased action potential amplitude and duration followed by electrical inexcitability after ~15min of ischemia. ΔΨm depolarization occurred in two phases during ischemia: in phase 1 (<30min ischemia), mitochondrial clusters within individual NRVMs depolarized, while phase 2 ΔΨm depolarization (30-60min) was characterized by global functional collapse of the mitochondrial network across the whole ischemic region of the monolayer, typically involving a propagating metabolic wave. Oxidation of the glutathione (GSSG:GSH) redox potential occurred during ischemia, followed by recovery upon reperfusion (i.e., lifting the coverslip). ΔΨm recovered in the mitochondria of individual myocytes quite rapidly upon reperfusion (<5min), but was highly unstable, characterized by subcellular oscillations or flickering of clusters of mitochondria in NRVMs across the reperfused region. Electrical excitability also recovered in a heterogeneous manner, providing an arrhythmogenic substrate which led to formation of sustained reentry. Treatment with 4'-chlorodiazepam, a peripheral benzodiazepine receptor ligand, prevented ΔΨm oscillation, improved GSH recovery rate, and prevented reentry during reperfusion, indicating that stabilization of mitochondrial network dynamics is important for preventing post-ischemic arrhythmias. This article is part of a Special Issue entitled "Mitochondria: From Basic Mitochondrial Biology to Cardiovascular Disease".


Trajectories of perinatal depressive symptoms in the context of the COVID-19 pandemic.
This study sought to advance understanding of the potential long-term consequences of the COVID-19 pandemic for child development by characterizing trajectories of maternal perinatal depression, a common and significant risk factor for adverse child outcomes. Data came from 393 women (86% White, 8% Latina; mean age = 33.51 years) recruited during pregnancy (n = 247; mean gestational age = 22.94 weeks) or during the first year postpartum (n = 146; mean child age = 4.50 months; 55% female). Rates of depression appear elevated, relative to published reports and to a pre-pandemic comparison group (N = 155). This study also provides evidence for subgroups of individuals who differ in their depressive symptom trajectories over the perinatal period. Subgroup membership was related to differences in maternal social support, but not to child birth outcomes.


The mitigating effects of maternal social support and paternal involvement on the intergenerational transmission of violence.
Childhood maltreatment is a strong risk factor for subsequent violence, including violent behaviors in young adulthood and offspring maltreatment after becoming a parent. Little is known about the specific circumstances under which supportive relationships may help disrupt this cycle of violence throughout the life course. We conducted two complementary analyses to assess whether maternal social support in early childhood, and also paternal involvement in middle childhood, could prevent the intergenerational transmission of violence, using data from the Avon Longitudinal Study of Parents and Children (n=11,384). We found that higher levels of maternal social support in the postpartum period reduced the odds of offspring maltreatment at ages 0-8 years (OR=0.95, 95% CI 0.93-0.96). When classifying mothers according to their abuse history, this protective association of social support was observed among mothers with no history of childhood maltreatment and among those with only childhood maltreatment (and not postpartum intimate partner violence [IPV]), but not among mothers who reported IPV since the child's birth. We then extended our analysis of these offspring forward in time and found that paternal involvement at ages 9-10 years was associated with a reduced risk of offspring self-reported violent perpetration at ages 18-20 years (OR=0.85, 95% CI=0.77-0.94). This protective association was generally apparent among all subgroups of children, including those with a history of childhood maltreatment. Together these results highlight the protective influence of supportive relationships against the intergenerational transmission of violence, depending on abuse history, context, and timing, with important implications for the prevention of childhood maltreatment and mitigation of its negative effects.


Evolving information priorities of hematologic cancer survivors, caregivers, and other relatives.
Little is known about information priorities of people touched by hematologic cancers. We interviewed and surveyed 29 survivors/patients, 13 caregivers, and 19 non-caregiver relatives. Qualitative interviews indicated limited information describing topics other than specific cancer subtypes and treatment options. The survey exercise revealed the following priorities: at diagnosis, cancer types and treatment options; during initial treatment, treatment options and coping with side effects; after treatment, follow-up tests and long-term side effects; at remission/during maintenance treatment at relapse, treatment options and follow-up tests; for patients, cancer types and treatment options; for caregivers, future outlook and support; for non-caregivers, finances. Information priorities vary by role and over time.


Living on the Edge: Diagnosing Sarcomatoid Melanoma Using Histopathologic Cues at the Edge of a Dedifferentiated Tumor: A Report of 2 Cases and Review of the Literature.
Sarcomatoid melanoma is a rare type of melanoma lacking typical histologic features of melanoma and often lacks expression of S100 protein and melanocyte-specific markers. Given the rarity of this entity, its clinicopathologic findings are not well defined. We report 2 cases of sarcomatoid melanoma received in consultation: a 65-year-old woman with a right breast mass and a 62-year-old man with a left plantar heel mass. Both lesions were ulcerated, pedunculated, highly cellular proliferations of atypical spindle cells arranged as fascicles and/or sheets. The tumor cells of the breast mass expressed CD10 and vimentin diffusely but S100 protein only focally. The tumor cells of the heel mass lacked expression of melanocytic markers altogether, except for weak, very focal S100 protein expression. At the junctional edge of the breast mass and in the ulcer base of the heel mass, focal precursor melanoma was present and exhibited melanocytic differentiation. We report these cases to emphasize the importance of meticulous histologic inspection at the lesion's edge and/or ulcer base to correctly identify the conventional precursor melanoma in these rare lesions to ensure appropriate diagnosis and subsequent clinical management as treatment options may be significantly different from those offered for sarcomas.


Development of an easy and rapid analytical method for the extraction and preconcentration of chloroquine phosphate from human biofluids prior to GC-MS analysis.
A vortex assisted spraying based fine droplet formation liquid phase microextraction (VA-SFDF-LPME) method was developed to determine chloroquine phosphate at trace levels in human serum, urine and saliva samples by gas chromatography-mass spectrometry (GC-MS) with single quadrupole mass analyzer. In the first part, several liquid phase microextraction (LPME) and magnetic solid phase extraction (MSPE) methods were compared to each other in order to observe their extraction ability for the analyte. VA-SFDF-LPME method was selected as an efficient and easy extraction method due to its higher extraction efficiency. Optimization studies were carried out for the parameters such as extraction solvent type, sodium hydroxide volume/concentration, sample volume, spraying number and mixing type/period. Tukey's method based on post hoc test was applied to all experimental data for the selection of optimum values. Optimum extraction parameters were found to be 12 mL initial sample volume, two sprays of dichloromethane, 0.75 mL of 60 g/kg sodium hydroxide and 15 s vortex. Under the optimum conditions, limit of detection and quantification (LOD and LOQ) were calculated as 2.8 and 9.2 μg/kg, respectively. Detection power of the GC-MS system was increased by approximately 317 folds with the developed extraction/preconcentration method. The applicability and accuracy of the proposed method was evaluated by spiking experiments and percent recovery results for human urine, serum and saliva samples were found in the range of 90.9% and 114.0% with low standard deviation values (1.9-9.4).


Aspirin Inhibits Brain Metastasis of Lung Cancer via Upregulation of Tight Junction Protein Expression in Microvascular Endothelial Cells.
The brain is one of the most vulnerable metastasis sites in lung cancer; approximately 40-50% of lung cancer patients develop brain metastasis during the disease course, contributing to the poor prognosis and high mortality of lung cancer patients. Therefore, it is important to clarify the molecular mechanism underlying brain metastasis of lung cancer for improving the overall survival of lung cancer patients. The present study aimed to investigate the potential role of blood-brain barrier (BBB) permeability in the development of brain metastasis of lung cancer and explore the effect of aspirin in an in-vitro BBB model. An in-vitro BBB model was established. The expression of heat shock protein 70 (HSP 70), zonula occludens-1 (ZO-1), and occludin in rat brain microvascular endothelial cells was detected using Western blot at different time points following the administration of aspirin. HSP70, ZO-1, and occludin expressions did not show significant changes before aspirin administration, but increased noticeably after aspirin administration. Tumor necrosis factor-α (TNF-α) could significantly attenuate the increased expression of these proteins induced by aspirin. Additionally, TNF-α also significantly reversed the aspirin-induced decrease of BBB permeability. Aspirin may inhibit brain metastasis of lung cancer in a time-dependent manner via upregulating tight junction proteins to reduce BBB permeability, and this effect can be reversed by TNF-α.


A tomato B-box protein SlBBX20 modulates carotenoid biosynthesis by directly activating PHYTOENE SYNTHASE 1, and is targeted for 26S proteasome-mediated degradation.
Carotenoids play important roles in many biological processes, such as light harvesting, photoprotection and visual attraction in plants. However, the regulation of carotenoid biosynthesis is still not fully understood. Here, we demonstrate that SlBBX20, a B-box (BBX) zinc-finger transcription factor, is a positive regulator of carotenoid accumulation in tomato (Solanum lycopersicum). Overexpression of SlBBX20 leads to dark green fruits and leaves and higher levels of carotenoids relative to the wild-type. Interactions between SlBBX20 and DE-ETIOLATED 1 (SlDET1) lead to the ubiquitination and 26S proteasome-mediated degradation of SlBBX20. Moreover, deficiencies in the components of the CUL4-DDB1-DET1 complex enhanced the stability of the SlBBX20 protein. Thus, we conclude that SlBBX20 is a substrate of the CUL4-DDB1-DET1 E3 ligase. SlBBX20 can activate the expression of PHYTOENE SYNTHASE 1, encoding a key enzyme in carotenoid biosynthesis, by directly binding to a G-box motif in its promoter, which results in the elevated levels of carotenoids in SlBBX20 overexpression lines. We identified a key regulator of carotenoid biosynthesis and demonstrated that the stability of SlBBX20 is regulated by ubiquitination. These findings provide us a new target for the genetic improvement of the nutritional quality of tomato fruit.


Effect of fiber source on cecal fermentation and nitrogen recycled through cecotrophy in rabbits.
The influence of fiber source on fiber digestion in rabbits was investigated. Six fibrous feedstuffs with wide differences in chemical composition and particle size were selected: paprika meal, olive leaves, alfalfa hay, soybean hulls, sodium hydroxide-treated barley straw, and sunflower hulls. Six diets were formulated to contain one of these ingredients as the sole source of fiber. To avoid nutrient imbalances, fiber sources were supplemented with different proportions of a fiber-free concentrate, based on soy protein isolate, wheat flour, lard, and a vitamin and mineral mix, to obtain diets containing at least 3% nitrogen and 5% starch. Daily soft feces excretion, and its NDF, and total and microbial nitrogen content were determined in 60 fattening rabbits (10 per diet). Seven days after the last cecotrophy control, the same animals were used to determine weight of stomach, cecum and their contents, and cecal fermentation traits (pH, VFA and ammonia concentrations, and buffer properties of cecal contents). Stepwise regression analysis showed a positive effect (P < .001) on soft feces excretion, total and microbial nitrogen concentrations in soft feces, cecal acidity, and total VFA in the cecum of dietary pectic constituents (2.9, 3.5, 2.5, .9, and 6.6%) and proportion of fine particles (< .315 mm) (1.8, .9, 1.3, .15, and .9%) per each increment of one percentage unit of the independent variables. Proportion of fine particles also increased weight of cecal contents (P < .001). Soft feces excretion and weight of stomach and of its contents increased (P < .001) by 5.2, 2.8, and 10.2% per each percentage unit increment of proportion of large particles (> 1.25 mm). Degree of lignification of NDF decreased total nitrogen concentration in soft feces and cecal VFA concentration (P < .001). Source of fiber affected cecal pH not only by its influence on the cecal concentrations of the final products of fermentation, but also through its effect on the pH of dry cecal contents (P < .001). The latter was negatively correlated with dietary proportion of fine particles, degree of lignification of NDF, and base-buffering capacity of dry cecal contents (r = -.52, -.37, and -.49, respectively). From these results, we conclude that pectic constituent concentration, degree of lignification of NDF, and particle size are the variables that best characterize the influence of the source of fiber on soft feces excretion and cecal fermentation traits in rabbits.


Renal tubular dysfunction in beta-thalassemia minor.
Persons with beta-thalassemia minor usually are symptomless. However, we previously reported renal tubular dysfunction in a patient with beta-thalassemia minor. The aim of this study is to investigate renal function in patients with beta-thalassemia minor. Forty-one subjects with beta-thalassemia minor and 20 sex- and age-matched healthy subjects were enrolled in the study. For analysis, patients were divided into 2 groups: group A, all patients with anemia (n = 19), and group B, patients without anemia (n = 22). Blood and 24-hour urine samples were obtained for hematologic and biochemical analysis. Anemic patients had increased urinary zinc excretion (U(zinc)) and fractional excretion of sodium (FE(Na)) and uric acid (FE(UA)) compared with both controls and patients without anemia. Hemoglobin levels correlated significantly in a negative manner with U(zinc), FE(Na), and FE(UA) in patients with beta-thalassemia minor. However, serum lactate dehydrogenase levels correlated significantly in a positive manner with the same parameters. In addition, 6 of 41 patients (14.6%) with beta-thalassemia minor showed significant signs of renal tubulopathy, such as hypercalciuria, decreased tubular reabsorption of phosphorus with hypophosphatemia, hypomagnesemia with renal magnesium wasting, hypouricemia with renal uric acid wasting, and tubular proteinuria. Proximal renal tubular dysfunction is not rare in patients with beta-thalassemia minor.


The modified-relative-dose-response values in serum and milk are positively correlated over time in lactating sows with adequate vitamin A status.
The modified-relative-dose-response (MRDR) test, which requires a blood sample after dosing with 3,4-didehydroretinyl acetate (DRA), has been used to determine vitamin A (VA) status of individuals and groups worldwide. Less invasive methods using milk are in development in a swine model. Swine are a good choice for studying VA metabolism because their gastrointestinal anatomy, morphology, physiology, and VA requirements are similar to those of humans. In this study, DRA was used as a VA tracer in lactating sows to follow the metabolism of newly ingested VA. Lactating sows (n = 6) were administered 35 micromol DRA after overnight food deprivation. Blood and milk were collected at 0, 1.5, 3, 5, 7, 9, 24, and 48 h; livers were obtained at the time of killing. Samples were analyzed for didehydroretinol (DR), retinol (R), and didehydroretinyl esters (DRE). Serum DR:R was compared with that in milk and other VA indicators. DRE rapidly increased in serum, corresponding to chylomicra, whereas DR increased at a slower rate corresponding to the holo-DR:retinol-binding protein complex released from the liver. An estimated 10-20% of the dose was irreversibly lost in milk over 48 h. The mean MRDR value was 0.018 +/- 0.013 at 5 h and the mean liver VA was 0.73 +/- 0.21 micromol/g, both signifying sufficient stores. Milk and serum DR:R values were directly correlated (r = 0.64, P < 0.0001). Thus, DR:R values in milk may be a potential alternative to serum in determining VA status in lactating women. Future work is required in VA-deficient sows and women of varying VA status to determine DR trafficking and to compare DR:R values in milk with those in serum.


Numerical simulations of the 3D virtual model of the human hip joint, using finite element method.
In this paper, we present a three-dimensional mathematical model for a normal hip joint. The three-dimensional finite element model has been constructed based on Computer Tomograph scans of the bones. The obtained 3D model is studied using the finite element method, taking into consideration the real structure of the bone and the mechanical characteristics of cortical and spongy. The FE model of hip joint, the material properties used to simulate the behavior of the cortical and trabecular bone (of femur and coxal bone) and the cartilage, as well as the boundary conditions are presented. The distribution map of the axial and global movements on the global model and the distribution map of the axial and von Misses strain in the cartilaginous surface of the femur are presented.


Comparative analysis of the global transcriptome of Anopheles funestus from Mali, West Africa.
Anopheles funestus is a principal vector of malaria across much of tropical Africa and is considered one of the most efficient of its kind, yet studies of this species have lagged behind those of its broadly sympatric congener, An. gambiae. In aid of future genomic sequencing of An. funestus, we explored the whole body transcriptome, derived from mixed stage progeny of wild-caught females from Mali, West Africa. Here we report the functional annotation and comparative genomics of 2,005 expressed sequence tags (ESTs) from An. funestus, which were assembled with a previous EST set from adult female salivary glands from the same mosquito. The assembled ESTs provided for a nonredundant catalog of 1,035 transcripts excluding mitochondrial sequences. Comparison of the An. funestus and An. gambiae transcriptomes using computational and macroarray approaches revealed a high degree of sequence identity despite an estimated 20-80 MY divergence time between lineages. A phylogenetically broader comparative genomic analysis indicated that the most rapidly evolving proteins--those involved in immunity, hematophagy, formation of extracellular structures, and hypothetical conserved proteins--are those that probably play important roles in how mosquitoes adapt to their nutritional and external environments, and therefore could be of greatest interest in disease control.


Resistance is futile: lack of predator switching and a preference for native prey predict the success of an invasive prey species.
Invasive species continue to severely impact biodiversity, yet predicting the success or failure of introduced species has remained elusive. In particular, the relationship between community invasibility and native species diversity remains obscure. Here, we apply two traditional ecological concepts that inform prey population stability and hence invasibility. We first show that the native predatory crustacean Gammarus duebeni celticus exhibited similar type II (destabilizing) functional responses (FRs) towards native mayfly prey and invasive amphipod prey, when these prey species were presented separately. However, when the two prey species were presented simultaneously, the predator did not exhibit prey switching, instead consuming disproportionately more native prey than expected from the relative abundance of native and invasive species. These consumptive propensities foster reductions of native prey, while simultaneously limiting biotic resistance against the invasive species by the native predator. Since our theoretical considerations and laboratory results match known field invasion patterns, we advocate the increased consideration of FR and prey switching studies to understand and predict the success of invasive species.


Serine Metabolism Tunes Immune Responses To Promote Oreochromis niloticus Survival upon Edwardsiella tarda Infection.
Overactive immune response is a critical factor triggering host death upon bacterial infection. However, the mechanism behind the regulation of excessive immune responses is still largely unknown, and the corresponding control and preventive measures are still to be explored. In this study, we find that Nile tilapia, Oreochromis niloticus, that died from Edwardsiella tarda infection had higher levels of immune responses than those that survived. Such immune responses are strongly associated with metabolism that was altered at 6 h postinfection. By gas chromatography-mass spectrometry-based metabolome profiling, we identify glycine, serine, and threonine metabolism as the top three of the most impacted pathways, which were not properly activated in the fish that died. Serine is one of the crucial biomarkers. Exogenous serine can promote O. niloticus survival both as a prophylactic and therapeutic upon E. tarda infection. Our further analysis revealed exogenous serine flux into the glycine, serine, and threonine metabolism and, more importantly, the glutathione metabolism via glycine. The increased glutathione synthesis could downregulate reactive oxygen species. Therefore, these data together suggest that metabolic modulation of immune responses is a potential preventive strategy to control overactive immune responses. IMPORTANCE Bacterial virulence factors are not the only factors responsible for host death. Overactive immune responses, such as cytokine storm, contribute to tissue injury that results in organ failure and ultimately the death of the host. Despite the recent development of anti-inflammation strategies, the way to tune immune responses to an appropriate level is still lacking. We propose that metabolic modulation is a promising approach in tuning immune responses. We find that the metabolomic shift at as early as 6 h postinfection can be predictive of the consequences of infection. Serine is a crucial biomarker whose administration can promote host survival upon bacterial infection either in a prophylactic or therapeutic way. Further analysis demonstrated that exogenous serine promotes the synthesis of glutathione, which downregulates reactive oxygen species to dampen immune responses. Our study exemplifies that the metabolite(s) is a potential therapeutic reagent for overactive immune response during bacterial infection.


Cardiovascular risk of adipokines: a review.
Over the last two decades, the understanding of adipose tissue has undergone radical change. The perception has evolved from an inert energy storage tissue to that of an active endocrine organ. Adipose tissue releases a cluster of active molecules named adipokines. The severity of obesity-related diseases does not necessarily correlate with the extent of body fat accumulation but is closely related to body fat distribution, particularly to visceral localization. There is a distinction between the metabolic function of central obesity (visceral abdominal) and peripheral obesity (subcutaneous) in the production of adipokines. Visceral fat accumulation, linked with levels of some adipokines, induces chronic inflammation and metabolic disorders, including glucose intolerance, hyperlipidaemia, and arterial hypertension. Together, these conditions contribute to a diagnosis of metabolic syndrome, directly associated with the onset of cardiovascular disease. If it is well known that adipokines contribute to the inflammatory profile and appetite regulation, this review is novel in synthesising the current state of knowledge of the role of visceral adipose tissue and its secretion of adipokines in cardiovascular risk.


A study of oxidative stress, antioxidant status and lipid profile in diabetic patient in the western region of Nepal.
Diabetes mellitus (DM) is often termed as a disease of premature aging. Several studies have indicated lopsided redox balance due to pro oxidant environment as one of the important etiological factors. Some recent researches also indicate a causal relationship with oxidative stress (OS). So far, no study has been undertaken on this aspect in Nepali populations. We, therefore, aimed this maiden study in Nepali population to examine redox balance by measuring OS and antioxidant status along with lipid profile in 37 patients of DM type- 2 and 30 matched normal subjects. Thirty seven patients of DM type-2 without any complications (mean age= 57.6+/- 10.6 years) and 30 normal subjects (mean age= 55.8 +/- 14.8 years) were included in this study. Body Mass Index (BMI) and Waist/Hip (W/H) ratio were measured. Fasting blood sample was collected for the analysis of total antioxidant activity (TAA), plasma and urinary thiobarbituric acid reactive substances (TBARS) and lipid profile by standard procedures in both the groups. The statistical analysis was done with SPSS 10 version. Total cholesterol, triglyceride, VLDL-cholesterol, LDL-cholesterol, plasma and urinary TBARS were significantly raised whereas, plasma TAA was significantly reduced in DM type-2 patients as compared to controls. The comparison of old and fresh cases revealed that though TAA was lower and PTBARS and UTBARS were higher in patients but did not attain the level of significance. W/H ratio is significantly higher in patients compared to normal subjects. But, no significant correlation of BMI and W/H with lipid profile is observed in both control and patients. Oxidative stress is raised in type 2 DM patients. This along with deranged lipid profile and decreased antioxidant status could be the risk factors in the development of complications associated with DM.


Immunotoxin treatment targeted to the high-molecular-weight melanoma-associated antigen prolonging the survival of immunodeficient rats with invasive intracranial human glioblastoma multiforme.
The aim of this study was to target immunotoxin treatment to the high-molecular-weight melanoma-associated antigen (HMW-MAA) and thereby examine any changes in the survival of immunodeficient rats with human glioblastoma multiforme (GBM). To target treatment specifically to human glioma cells, Pseudomonas exotoxin A (PE) was conjugated to the 9.2.27 antibody, which recognizes the HMW-MAA. Treatment of the antigen-positive glioma cell line U87MG with the resulting 9.2.27-PE caused cytotoxicity with a median inhibitory concentration of 1 ng/ml. Intratumoral 9.2.27-PE treatment of intracranial U87MG tumors in nude rats prolonged the survival of these animals by 43% compared with controls. In additional studies on the use of this targeted treatment, the authors precultured freshly dissected glioblastoma multiforme (GBM) biopsy tissue for 1 to 2 weeks. Inoculation of this tissue into the rat brain resulted in diffuse infiltrative gliomas. The markers glial fibrillary acidic protein and S100 protein were found to be expressed in the original biopsy specimens, as well as in the glioma xenografts in nude rat brains. Intratumoral immunotoxin treatment of such established tumors with 9.2.27-PE was effective and prolonged survival time from 30% to as high as 90% in animals with tumors originating from four different GBM specimens. Targeted treatment of highly invasive GBMs proved effective, and these results emphasize the clinical relevance of this antigen as a target molecule for immunotoxin treatment of human GBMs.


Effect of sequential surface irrigations on field-scale emissions of 1,3-dichloropropene.
A field experiment was conducted to measure subsurface movement and volatilization of 1,3-dichloropropene (1,3-D) after shank injection to an agricultural soil. The goal of this study was to evaluate the effect of sprinkler irrigation on the emissions of 1,3-D to the atmosphere and is based on recent research that has shown that saturating the soil pore space reduces gas-phase diffusion and leads to reduced volatilization rates. Aerodynamic, integrated horizontal flux, and theoretical profile shape methods were used to estimate fumigant volatilization rates and total emission losses. These methods provide estimates of the volatilization rate based on measurements of wind speed, temperature, and 1,3-D concentration in the atmosphere. The volatilization rate was measured continuously for 16 days, and the daily peak volatilization rates for the three methods ranged from 18 to 60 microg m(-2) s(-1). The total 13-D mass entering the atmosphere was approximately 44-68 kg ha(-1), or 10-15% of the applied active ingredient This represents approximately 30-50% reduction in the total emission losses compared to conventional fumigant applications in field and field-plot studies. Significant reduction in volatilization of 1,3-D was observed when five surface irrigations were applied to the field, one immediately after fumigation followed by daily irrigations.


Eating disorders patients' views on their disorders and on an outpatient service: a qualitative study.
The objective of the study was to determine sufferers' views of outpatient treatment for eating disorders and provide practical recommendations for treatment practice. Twenty NHS outpatients participated in semi-structured interviews, which were subjected to thematic analysis. Respondents expressed ambivalence about whether their eating disorder is a way of exerting control or a disorder that controls them and this leads to them seeking treatment. Sufferers preferred a practical and sensitive approach and began to rely on treatment for recovery. Treatment needs to facilitate sufferers' need for control by striking a balance between practical and empathetic approaches that both involve patients in treatment decisions and give authoritative guidance.


CO2 laser irradiation combined with fluoridated dentifrice improved its protective effect on caries lesion progression regardless of the acidulated phosphate fluoride gel application: An in situ study.
This in situ study aimed to investigate the efficacy of CO2 laser at a 10.6-µm wavelength combined with 1.23% acidulated phosphate fluoride (APF) and fluoridated dentifrice with 1100 µg F/g (FD) to control enamel caries progression. Sixteen volunteers wore palatal appliances containing eight demineralized enamel specimens for four 14-day phases under sucrose exposure. These specimens were submitted to CO2 laser irradiation and APF alone or combined with FD. Treatment groups were non-fluoridated dentifrice-NFD, NFD + CO2 laser, NFD + APF, NFD + CO2 laser + APF, FD, FD + CO2 laser, FD + APF, and FD + CO2 laser + APF. Mineral loss, calcium fluoride (CaF2), fluorapatite (FAp), and fluoride in the biofilm were analyzed by analysis of variance followed by the Student-Newman-Keuls test, p < 0.05. The highest mineral loss inhibition was noted when FD and CO2 laser irradiation were combined, which did not significantly differ from the FD + CO2 laser + APF group. The CaF2, FAp, and F in the biofilm were more pronounced when the FD and APF were combined. The CO2 laser irradiation promoted a slightly higher concentration of CaF2 in the enamel and F in the biofilm. Although APF promotes the high formation of CaF2 and FAp, the combined use of FD with CO2 laser overcomes the APF effect in inhibiting the progression of artificial caries-like lesions in situ. Under the in situ design of this study, remineralization of white spot lesions was achieved through CO2 laser irradiation and daily use of fluoridated dentifrice. Future clinical trials are encouraged to substantiate this finding.


The effects of soybean selenium proteinate on tissue selenium and meat quality traits in finishing pigs.
The purpose of the present study was to evaluate the effects of soybean selenium proteinate on Se tissue retention and meat quality in pigs. In group A (n = 11) the mixtures were supplemented with soybean selenium proteinate, in group B (n = 11) with sodium selenite and in group C (n = 11) with Se-enriched yeast (0.3 mg Se per kg in all groups). The use of soybean selenium proteinate resulted in lower retention of Se in tissues (liver, heart, muscle) compared to Se-enriched yeast. Selenium concentrations in tissues achieved by soybean selenium proteinate and sodium selenite were comparable. No differences in serum Se, serum GSH-Px and meat quality traits were found among the groups.


PTSD after severe vehicular crashes.
To describe predictors of PTSD after motor vehicle crashes (MVC). MVC patients were interviewed during their hospitalization and at 6 and 12 months post-injury. Interviews included information about behavioral factors, circumstances around the crash, recovery and PTSD screening. PTSD was defined as the development of 3 or more of 7 PTSD symptoms. Association of risk factors with PTSD development at 6 and 12 months was analyzed using contingency tables. Multiple regression models were built for the prediction of PTSD. 367 and 317 patients completed the 6 and 12 month interviews respectively. PTSD developed in 27.5 % (n=101) and 24.3 % (n=77) of the population at 6 and 12 months respectively. PTSD occurred more frequently among females, those with a previous history of depression, violent injury, or other traumatic events, and those whose crashes involved a fatality. Those who were culpable for the crash, age<30, and sustained brain injuries were less likely to develop PTSD at 6 months. Occupant position, education, marital status, alcohol problems, injury severity, heart rate, and blood alcohol + status did not show any significant association with PTSD. In the multiple logistic regression, female gender, history of depression, culpability, prior violent injury, and a fatality in the crash were associated with PTSD at 6 months. Only prior violent injury, and a death in same crash were predictors at one year. PTSD occurs frequently after MVCs. Female gender, prior violent injury, death of another occupant and history of depression are associated with PTSD development.


Assessment of salinity-induced photorespiratory glycolate metabolism in Anabaena sp. PCC 7120.
This paper reports an investigation of salinity-induced glycolate metabolism in the cyanobacterium Anabaena sp. PCC 7120 (hereafter Anabaena PCC 7120). Quantitative analysis of transcripts for the photosynthesis-associated genes encoding ribulose-1,5-bisphosphate carboxylase oxygenase (Rubisco), phosphoribulokinase and transketolase, as well as those involved in glycolate metabolism (phosphoglycolate phosphatase, glycolate oxidase, alanine-glyoxylate aminotransferase and serine hydroxymethyltransferase) was performed. The expression of all investigated photosynthesis-associated genes except Rubisco was downregulated after 24 h NaCl treatment. However, under the same conditions, the transcripts encoding enzymes involved in glycolate metabolism were overexpressed. This was further confirmed by the quantitative analysis of the intermediates involved in glycolate metabolism. The intracellular levels of organic acids (glyceric, glycolic and glyoxylic acids) and amino acids (glycine and serine) were elevated in salt-treated cells as compared to those in the control cells. Transcriptional inhibition of photosynthesis-associated genes, and upregulation of genes and enhanced synthesis of intermediates associated with glycolate metabolism, indicate the occurrence of this photorespiratory metabolic pathway metabolism in Anabaena PCC 7120 under salt stress.


Short horizons and obesity futures: disjunctures between public health interventions and everyday temporalities.
This paper examines the spatio-temporal disjuncture between 'the future' in public health obesity initiatives and the embodied reality of eating. Drawing upon ethnographic fieldwork in a disadvantaged community in South Australia (August 2012-July 2014), we argue that the future oriented discourses of managing risk employed in obesity prevention programs have limited relevance to the immediacy of poverty, contingencies and survival that mark people's day to day lives. Extending Bourdieu's position that temporality is a central feature of practice, we develop the concept of short horizons to offer a theoretical framework to articulate the tensions between public health imperatives of healthy eating, and local 'tastes of necessity'. Research undertaken at the time of Australia's largest obesity prevention program (OPAL) demonstrates that pre-emptive and risk-based approaches to health can fail to resonate when the future is not within easy reach. Considering the lack of evidence for success of obesity prevention programs, over-reliance on appeals to 'the future' may be a major challenge to the design, operationalisation and success of interventions. Attention to local rather than future horizons reveals a range of innovative strategies around everyday food and eating practices, and these capabilities need to be understood and supported in the delivery of obesity interventions. We argue, therefore, that public health initiatives should be located in the dynamics of a living present, tailored to the particular, localised spatio-temporal perspectives and material circumstances in which people live.


Reconciling the different roles of Gsk3β in "naïve" and "primed" pluripotent stem cells.
Signaling pathways orchestrated by PI3K/Akt, Raf/Mek/Erk and Wnt/β-catenin are known to play key roles in the self-renewal and differentiation of pluripotent stem cells. The serine/threonine protein kinase Gsk3β has roles in all three pathways, making its exact function difficult to decipher. Consequently, conflicting reports have implicated Gsk3β in promoting self-renewal, while others suggest that it performs roles in the activation of differentiation pathways. Different thresholds of Gsk3β activity also have different biological effects on pluripotent cells, making this situation even more complex. Here, we describe a further level of complexity that is most apparent when comparing "naïve" murine and "primed" human pluripotent stem cells. In naïve cells, Gsk3β activity is restrained by PI3K/Akt, but when released from inhibitory signals it antagonizes self-renewal pathways by targeting pluripotency factors such as Myc and Nanog. This situation also applies in primed cells, but, in addition, a separate pool of Gsk3β is required to suppress canonical Wnt signaling. These observations suggest that different Gsk3β-protein complexes shift the balance between naïve and primed pluripotent cells and identify fundamental differences in their cell signaling. Altogether, these findings have important implications for the mechanisms underpinning the establishment of different pluripotent cell states and for the control of self-renewal and differentiation.


The bidirectional relationship between sleep and physical activity following traumatic brain injury.
Sleep and physical activity are both modifiable behavioural factors that are associated with better health and are potentially related. Following traumatic brain injury, damage to the brain caused by an external force, sleep disturbances are common. Exploring bidirectional relationships between sleep and physical activity might provide insight into whether increasing physical activity could decrease these sleep disturbances. The current study, therefore, examined inter- and intra-individual temporal associations between sleep and daytime physical activity in 64 people with traumatic brain injury reporting sleep problems or fatigue (47 males; mean age, 40 years). Sleep and physical activity were measured using actigraphy with corroborating sleep diaries over 14 consecutive days. Multilevel models were used to examine inter- and intra-individual associations between physical activity and sleep. Inter-individual variations showed that earlier bedtimes, earlier wake-up times and lower sleep efficiency were associated with more physical activity. Intra-individual temporal variations showed no significant association of daytime physical activity with sleep duration or continuity. However, shorter sleep time and less wake after sleep onset than usual were associated with more time spent in light-intensity activity the next day. Therefore, sleep may have more of an influence on physical activity than physical activity has on sleep in people with traumatic brain injury. In conclusion, the results do not confirm a potential beneficial effect of physical activity on sleep but suggest that improving sleep quality might be relevant to support of a physically active lifestyle in people with traumatic brain injury. Further research is necessary to confirm these results.


[Personality factors that influence the working conditions in a hospital].
The author of this paper studied the structure of the personality of the doctors, nurses, social workers and auxiliar personnel of a hospital in Mexico City, through the test of incomplete sentences of Sacks-Cantú which investigate the attitude to the superior, values, repelled and accepted aspects of identity, personnel interrelations, success and downfall, character, liability, confiability and competence. The doctors has the following attributes to the performance of their work: initiative, altruism, personnel interrelations, liability and confiability. The nurses are abnegate and altruists and the social workers has initiative and acceptance of their identity. This investigation shows that the most structured personality, the best intellectual development and the best laboral effectiveness.


GDF11 restrains tumor growth by promoting apoptosis in pancreatic cancer.
Growth differentiation factor (GDF) acted as a factor that regulated proliferation, apoptosis and differentiation in several tumors. However, the effects of growth differentiation factor (GDF11) in pancreatic cancer remain unclear. To investigate the expression and significance of GDF11 in pancreatic cancer. Pancreatic cancer and corresponding paracancerous tissues (n=28) were collected from the Department of Hepatobiliary and Pancreatic Surgical Oncology of Chinese PLA General Hospital. Tissue microarray was obtained from Outdo Biotech Co., Ltd. (Shanghai, People's Republic of China). GDF11 mRNA and protein expressions in pancreatic cancer samples and cell lines were detected using qRT-PCR, Western-Blot and immunohistochemistry. Overexpression and knockdown of GDF11 were performed with lentiviral transduction system and siRNA technique in PANC-1 cell line and CFPAC-1 cell line. Proliferation, migration and invasion of pancreatic cancer cell lines were examinated by MTS and transwell assay, respectively. Flow cytometry was used for cell apoptosis analysis. The results of this study indicated that GDF11 was significantly down-regulated in pancreatic cancer tissues compared with adjacent tissues of pancreatic cancer. GDF11 was also associated with low expression in pancreatic cancer cell lines when compared with normal pancreatic cell line. In a cohort of 63 pancreatic cancer patients, high GDF11 expression levels was associated with favorable perineural invasion, T classification, N classification and overall survival (OS). Cox proportional hazards model revealed that high GDF11 expression was an independent predictor of favorable prognosis (HR: 0.496; 95% CI: 0.255-0.967; P=0.040). Overexpression of GDF11 in PANC-1 cells repressed the proliferation, migration and invasion abilities in vitro. Inhibition of GDF11 in CFPAC-1 showed inverse results. Furthermore, enhanced GDF11 expression promoted apoptosis and down-regulated GDF11 expression inhibited apoptosis in pancreatic cancer cell lines. These findings suggested that GDF11 acted as a tumor suppressor gene for pancreatic cancer.


Na+(Li+)-H+ exchange in rat renal cortical vesicles with endosomal characteristics.
Preparations of rat renal cortical endocytic vesicles, loaded with fluorescein isothiocyanate-labeled dextran (FITC-dextran) in vivo had an ATP-dependent N-ethylmaleimide (NEM)-sensitive H+ pump and, in addition, showed a limited activity of an electroneutral Na(+)-H+ and Li(+)-H+ exchanger. However, the majority of FITC-dextran-containing vesicles possessed only the H+ pump. The H+ pump and the electroneutral Na+(Li+)-H+ antiporter colocalized in only a small proportion of FITC-dextran-containing vesicles. These vesicles also exhibited marked electrically coupled Na+, Li+, and H+ movement due to conductances in the vesicle membrane for these cations. As measured by the acridine orange fluorescence quench method, the electroneutral Na(+)-H+ exchange in endosomal preparations obeyed hyperbolic kinetics with an apparent Michaelis constant (Km) for Na+ of 5.9 mM and resembled that of brush-border membrane vesicles (BBMV) isolated from the same tissue (Km for Na+, 3.7 mM). The endosomal Na+(Li+)-H+ exchange was not affected by the presence or absence of ATP or NEM. However, Li+, added before or simultaneously with Na+, inhibited the endosomal Na(+)-H+ antiporter. The Na+(Li+)-H+ antiporter was insensitive to extravesicular amiloride but was partially inhibited by intravesicular amiloride. Acidic buffers stimulated the activity of the electroneutral exchanger. We found that vesicles with similar characteristics contaminate preparations of right-side-out BBMV. We conclude that the vesicles with endosomal characteristics, which contain both a vacuolar-type H+ pump and the Na+(Li+)-H+ antiporter, are either a special pool of endocytic vesicles responsible for the trafficking of the exchanger between intracellular and luminal membranes or are inside-out BBMV that form during the homogenization of the tissue.


MEN1 in pancreatic endocrine tumors: analysis of gene and protein status in 169 sporadic neoplasms reveals alterations in the vast majority of cases.
Pancreatic endocrine tumors (PETs) may be part of hereditary multiple endocrine neoplasia type 1 (MEN1) syndrome. While MEN1 gene mutation is the only ascertained genetic anomaly described in PETs, no data exist on the cellular localization of MEN1-encoded protein, menin, in normal pancreas and PETs. A total of 169 PETs were used to assess the i) MEN1 gene mutational status in 100 clinically sporadic PETs by direct DNA sequencing, ii) immunohistochemical expression of menin in normal pancreas and 140 PETs, including 71 cases screened for gene mutations, and iii) correlation of these findings with clinical-pathological parameters. Twenty-seven PETs showed mutations that were somatic in 25 patients and revealed to be germline in 2 patients. Menin immunostaining showed strong nuclear and very faint cytoplasmic signal in normal islet cells, whereas it displayed abnormal location and expression levels in 80% of tumors. PETs harboring MEN1 truncating mutations lacked nuclear protein, and most PETs with MEN1 missense mutations showed a strong cytoplasmic positivity for menin. Menin was also misplaced in a significant number of cases lacking MEN1 mutations. In conclusion, the vast majority of PETs showed qualitative and/or quantitative alterations in menin localization. In 30% of cases, this was associated with MEN1 mutations affecting sequences involved in nuclear localization or protein-protein interaction. In cases lacking MEN1 mutations, the alteration of one of the menin interactors may have prevented its proper localization, as suggested by recent data showing that menin protein shuttles between the nucleus and cytoplasm and also affects the subcellular localization of its interactors.


Measuring performance of professional role substitution models of care against traditional medical care in healthcare-A systematic review.
To identify outcome measures used to evaluate performance of healthcare professional role substitution against usual medical doctor or specialist medical doctor care to facilitate our understanding of the adequacy of these measures in assessing quality of healthcare delivery. Using a systematic approach, we searched Medline, Cochrane Central Register of Controlled Trials, Embase, CINAHL, and Web of Science from database inception until May 2020. Studies that presented original comparative data on at least one outcome measure were included following screening by two authors. Findings were synthesized, and outcome measures classified into six domains which included: effectiveness, safety, appropriateness, access, continuity of care, efficiency, and sustainability which were informed by the Institute of Medicine dimensions of healthcare quality, the Australian health performance framework, and Levesque and Sutherland's integrated performance measurement framework. One thirty five articles met the inclusion criteria, describing 58 separate outcome measures. Safety of role substitution models of care was assessed in 80 studies, effectiveness (n = 60), appropriateness (n = 40), access (n = 36), continuity of care (n = 6), efficiency and productivity (n = 45). Two-thirds of the studies that assessed productivity and efficiency performed an economic analysis (n = 27). The quality and rigour of evaluations varied substantially across studies, with two-thirds of all studies measuring and reporting outcomes from only one or two of these domains. There are a growing number of studies measuring the performance of non-medical healthcare professional substitution roles. Few have been subject to robust evaluations, and there is limited evidence on the scientific rigour and adequacy of outcomes measured. A systematic and coordinated approach is required to support healthcare settings in assessing the value of non-medical role substitution healthcare delivery models.


Alteration of the serine protease PRSS56 causes angle-closure glaucoma in mice and posterior microphthalmia in humans and mice.
Angle-closure glaucoma (ACG) is a subset of glaucoma affecting 16 million people. Although 4 million people are bilaterally blind from ACG, the causative molecular mechanisms of ACG remain to be defined. High intraocular pressure induces glaucoma in ACG. High intraocular pressure traditionally was suggested to result from the iris blocking or closing the angle of the eye, thereby limiting aqueous humor drainage. Eyes from individuals with ACG often have a modestly decreased axial length, shallow anterior chamber and relatively large lens, features that predispose to angle closure. Here we show that genetic alteration of a previously unidentified serine protease (PRSS56) alters axial length and causes a mouse phenotype resembling ACG. Mutations affecting this protease also cause a severe decrease of axial length in individuals with posterior microphthalmia. Together, these data suggest that alterations of this serine protease may contribute to a spectrum of human ocular conditions including reduced ocular size and ACG.


Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana.
The Agrobacterium vacuum infiltration method has made it possible to transform Arabidopsis thaliana without plant tissue culture or regeneration. In the present study, this method was evaluated and a substantially modified transformation method was developed. The labor-intensive vacuum infiltration process was eliminated in favor of simple dipping of developing floral tissues into a solution containing Agrobacterium tumefaciens, 5% sucrose and 500 microliters per litre of surfactant Silwet L-77. Sucrose and surfactant were critical to the success of the floral dip method. Plants inoculated when numerous immature floral buds and few siliques were present produced transformed progeny at the highest rate. Plant tissue culture media, the hormone benzylamino purine and pH adjustment were unnecessary, and Agrobacterium could be applied to plants at a range of cell densities. Repeated application of Agrobacterium improved transformation rates and overall yield of transformants approximately twofold. Covering plants for 1 day to retain humidity after inoculation also raised transformation rates twofold. Multiple ecotypes were transformable by this method. The modified method should facilitate high-throughput transformation of Arabidopsis for efforts such as T-DNA gene tagging, positional cloning, or attempts at targeted gene replacement.


Suppression of luteinizing hormone and testosterone secretion in bulls following adrenocorticotropin hormone treatment.
The present investigation was conducted to evaluate the inhibitory effects of adrenal corticosteroids on testosterone production by the bull testis. Administration of a single i.v. dose of adrenocorticotropic hormone (ACTH; 80 IU) resulted in a corticosteroid peak which lasted approximately 6 h. During this 6 h period, no episodic increases in secretion of LH or testosterone were initiated and basal concentrations of testosterone were suppressed (P less than 0.05) below control values. Episodic secretion of LH and testosterone resumed 6--7 h after ACTH when concentrations of serum corticosteroids had returned to basal levels. These results suggest that ACTH-induced increases in serum corticosteroids suppress the episodic secretion of LH, resulting in a suppression of testosterone secretion by the bull testis.


Development of in vitro maturation techniques for clinical applications.
In vitro maturation (IVM) refers to maturation in culture of immature oocytes at different stages that may or may not have been exposed to short courses of gonadotropins. The source of immature oocytes is an important feature for subsequent embryonic development, pregnancy, and healthy live births. IVM is an effective treatment that has already achieved significant outcomes of acceptable pregnancy and implantation rates and has led to the births of several thousand healthy babies. As the development of IVM treatment continues, an attractive possibility for improving the already successful outcome is to combine a natural-cycle in vitro fertilization (IVF) treatment with immature-oocyte retrieval followed by IVM of those immature oocytes. If the treatment processes can be simplified for immature-oocyte retrieval, different types of infertile women may be able to take advantage of these treatments. Mild-stimulation IVF combined with IVM treatment may represent a viable alternative to the standard treatment. Although IVM treatment is still considered to be experimental, it is now time to reconsider the IVM technology and its development. Mild-stimulation IVF combined with IVM may prove to be not just alternatives to standard treatments, but potentially first-line treatment choices.


Etomidate infusion and adrenocortical function. A study in elective surgery.
The adrenocortical response to a short tetracosactrin (Synacthen) test was studied in 11 patients receiving either etomidate infusion or thiopentone infusion used to maintain anaesthesia for abdominal hysterectomy. Pethidine was used as the narcotic component. The results showed that etomidate infusion (median 28.5 ug/kg/min) completely blocked the adrenocortical response to corticotropin stimulation for at least 24 h after surgery. No suppression was found in patients receiving thiopentone infusion. It is concluded that etomidate cannot be recommended for routine induction and maintenance of anaesthesia.


Incidence and Estimated Prevalence of Endometriosis and Adenomyosis in Northeast Italy: A Data Linkage Study.
Despite being quite frequent and having serious implications in terms of symptomatology and fertility, data on incidence and prevalence of endometriosis and adenomyosis following gold standard definitions are dramatically lacking. The average time from onset of symptoms to diagnosis in industrialized countries still ranges from five to ten years. Using the regional centralized data linkage system, we calculated incidence and prevalence of endometriosis and adenomyosis in the female population of Friuli Venezia Giulia region, Italy, for the years 2011-2013. Cases were defined as new diagnoses from hospital discharge records, following procedures allowing direct visualization for endometriosis and hysterectomy for adenomyosis, with or without histological confirmation. Diagnoses were considered "new" after verifying women had not been diagnosed in the previous ten years. Incidence of endometriosis and adenomyosis in women aged 15-50 years is 0.14%. Prevalence, estimated from incidence, is 2.00%. Adenomyosis, representing 28% of all diagnoses, becomes increasingly prevalent after the age of 50 years. Our results shows how the study of both endometriosis and adenomyosis should not be limited to women of premenopausal age. Further efforts are needed to sensitize women and health professional, and to find new data linkage possibilities to identify undiagnosed cases.


Age and High-Fat Diet Effects on Glutamine Synthetase Immunoreactivity in Liver and Hippocampus and Recognition Memory in Mice.
High-Fat Diet (HFD)-induced obesity may promote agerelated memory impairment via disturbances of ammonia-glutamine metabolism. We studied the effects of age and long-term HFD exposure on Glutamine Synthetase (GS) expression in the liver and hippocampus and recognition memory in mice. Adult (5-month-old) and aged (15-month-old) male C57BL/6 mice were exposed to control diet (CD, 14% calories from fat) or HFD (60% fat). Novel place recognition testing was conducted and tissue was collected after 4 and 5 months on HFD, respectively. Tissue GS expression levels were assessed using immunohistochemistry and image analysis. The obese mice developed moderate/severe hepatic steatosis. GS immunoreactivity was observed in perivenous hepatocytes and in hippocampal astrocytes and neuropil. Hepatic GS immunoreactivity density was higher in aged mice on HFD (n = 8) than CD (n = 13, P = 0.004). In aged mice, hippocampal GS immunoreactivity density was higher with HFD than CD (P = 0.037). In the novel place recognition test, aged mice were classified into impaired (n = 7) and unimpaired (n = 12), relative to adult mice (n = 22). Hippocampal GS immunoreactivity density was higher in impaired than unimpaired aged mice (P < 0.05). Long-term exposure of aged mice to HFD was associated with increased GS expression in the liver and hippocampus. Novel place recognition impairment in aged mice was associated with increased hippocampal GS expression. These findings suggest that excess ammonia is involved in the age-related effects of HFD exposure and in neurotoxicity.


Trends in nosocomial bloodstream infections following health care restructuring in Alberta between 1999 and 2005.
A previous study at the University of Alberta Hospital/Stollery Children's Hospital in Edmonton, Alberta, revealed an increase in hospital-acquired bloodstream infection (BSI) rates associated with an increase in patient acuity during a period of public health care delivery restructuring between 1993 and 1996. The present study assessed trends in BSIs since the end of the restructuring. Prospective surveillance for BSIs was performed using Centers for Disease Control and Prevention (USA) criteria for infection. BSI cases between January 1, 1999, and December 31, 2005, were reviewed. Available measures of patient volumes, acuity and BSI risk factors between 1999 and 2005 were also reviewed from hospital records. The University of Alberta Hospital/Stollery Children's Hospital (617 adult and 139 pediatric beds, respectively). All pediatric and adult patients admitted during the above-specified period with one or more episodes of BSIs. There was a significant overall decline in the BSI number and rate over the study period between 1999 and 2005. The downward trend was widespread, involving both adult and pediatric populations, as well as primary and secondary BSIs. During this period, the number of hospital-wide and intensive care unit admissions, intensive care unit central venous catheter-days, total parenteral nutrition days and number of solid-organ transplants were either unchanged or increased. Gram-positive bacterial causes of BSIs showed significant downward trends, but Gram-negative bacterial and fungal etiologies were unchanged. These data imply that, over time, hospitals can gradually adjust to changing patient care circumstances and, in this example, control infectious complications of health care delivery.


Using dynamic programming to create isotopic distribution maps from mass spectra.
This article presents a method to identify the isotopic distributions within a mass spectrum using a probabilistic classifier supplemented with dynamic programming. Such a system is needed for a variety of purposes, including generating robust and meaningful features from mass spectra to be used in classification. The primary result of this article is that the dynamic programming approach significantly improves sensitivity, without harming specificity, of a probabilistic classifier for identifying the isotopic distributions. When annotating isotopic distributions where an expert has performed the initial 'peak-picking' (removal of noise peaks), the dynamic programming approach gives a true positive rate of 96% and a false positive rate of 0.0%, whereas the classifier alone has a true positive rate of only 47% when the false positive rate is 0.0%. When annotating isotopic distributions in machine peak-picked spectra, which may contain many noise peaks, the dynamic programming approach gives a true positive rate of only 22.0%, but it still keeps a low false positive rate of 1.0% and still outperforms the classifier alone. It is important to note that all these rates are when we require exact matches with the distributions in annotated spectra; in our evaluation a distribution is considered 'entirely incorrect' if it is missing even one peak or contains even one extraneous peak. We compared to the THRASH and AID-MS systems using a looser requirement: correctly identifying the distribution that contains the mono-isotopic mass. Under this measure, our dynamic programming approach achieves a true positive rate of 82% and a false positive rate of 1%, which again outperforms the classifier alone. The dynamic programming approach ends up being more conservative than THRASH and AID-MS, yielding both fewer true and false peaks, but the F-score of the dynamic programming approach is significantly better than those of THRASH and AID-MS. All results were obtained with 10-fold cross-validation of 99 sections of mass spectra with a total of 214 hand-annotated isotopic distributions. Programs are available via http://www.cs.wisc.edu/~mcilwain/IDM.


Inhibition of tumor necrosis factor-alpha-induced RANTES secretion by alkaline protease in A549 cells.
Pseudomonas aeruginosa is a gram-negative bacterium that is an opportunistic pathogen in patients with cystic fibrosis and in immunocompromised hosts. This bacterium produces a variety of proteolytic enzymes, including alkaline protease (AP), which has multiple biological effects. This study investigated the effects of AP on the A549 pulmonary epithelial cell line. Results demonstrate that AP inhibited tumor necrosis factor (TNF)-alpha-induced RANTES gene expression and secretion in a concentration-dependent manner. The TNF-alpha-induced RANTES gene expression and secretion was attenuated with a neutralizing monoclonal antibody directed against the TNF receptor type 1 (TNFR1). Conversely, a neutralizing monoclonal antibody directed against TNF receptor type II had no effect, suggesting that these events were regulated through the TNFR1 receptor. In addition, we observed that soluble TNF receptor type 1 (sTNFR1) levels were significantly increased in culture supernatants of AP-treated cells in a concentration-dependent manner. Finally, membrane-associated TNFR1 was decreased after AP exposures. In these studies, the enzymatically inactive form of AP had no effect on TNF-alpha-induced RANTES secretion, shedding of sTNFR1, or membrane-associated TNFR1. These results demonstrate that AP stimulates shedding of cell-surface TNFR1, resulting in an increase in sTNFR1. Consequently, these events decrease the cells' ability to stimulate RANTES gene expression and secretion through TNFR1.


Therapeutic leukocytapheresis for inflammatory bowel disease.
The inference that granulocytes and monocytes/macrophages (GM) are part of the immunopathogenesis of inflammatory bowel disease (IBD) and hence should be targets of therapy stems from observations of elevated, and activated GM in patients with IBD. The Adacolumn can selectively deplete GM by adsorption (GMA) and in patients with IBD, GMA has been associated with significant clinical efficacy together with sustained suppression of inflammatory cytokine profiles. Additionally, GMA depleted proinflammatory CD14(+)CD16(+) monocytes and was followed by an increase in CD4(+) T lymphocytes including the regulatory CD4(+)CD25(high+)Foxp3 phenotype. Hence, GMA could be a non-pharmacologic therapy for IBD with potential to spare steroids and other unsafe pharmacologic preparations.


Relationship between neuronal migration and cell-substratum adhesion: laminin and merosin promote olfactory neuronal migration but are anti-adhesive.
Regulation by the extracellular matrix (ECM) of migration, motility, and adhesion of olfactory neurons and their precursors was studied in vitro. Neuronal cells of the embryonic olfactory epithelium (OE), which undergo extensive migration in the central nervous system during normal development, were shown to be highly migratory in culture as well. Migration of OE neuronal cells was strongly dependent on substratum-bound ECM molecules, being specifically stimulated and guided by laminin (or the laminin-related molecule merosin) in preference to fibronectin, type I collagen, or type IV collagen. Motility of OE neuronal cells, examined by time-lapse video microscopy, was high on laminin-containing substrata, but negligible on fibronectin substrata. Quantitative assays of adhesion of OE neuronal cells to substrata treated with different ECM molecules demonstrated no correlation, either positive or negative, between the migratory preferences of cells and the strength of cell-substratum adhesion. Moreover, measurements of cell adhesion to substrata containing combinations of ECM proteins revealed that laminin and merosin are anti-adhesive for OE neuronal cells, i.e., cause these cells to adhere poorly to substrata that would otherwise be strongly adhesive. The evidence suggests that the anti-adhesive effect of laminin is not the result of interactions between laminin and other ECM molecules, but rather an effect of laminin on cells, which alters the way in which cells adhere. Consistent with this view, laminin was found to interfere strongly with the formation of focal contacts by OE neuronal cells.


Complex oiling-out behavior of procaine with stable and metastable liquid phases.
During the crystallization of a solute from solvent(s), spontaneous liquid-liquid phase separation (LLPS) might occur, under certain conditions. This phenomenon, colloquially referred to as "oiling-out" in the pharmaceutical industry, often leads to undesired outcomes, including undesired particle properties, encrustation, ineffective impurity rejection, and excessively long process time. Therefore, it is critical to understand the thermodynamic driving force and phase boundaries of this phenomenon, such that rational strategies can be developed to avoid oiling-out or minimize its negative impact. In this study, we systematically evaluated the oiling-out behavior of procaine, a low melting point drug, in the solvent systems heptane, and ethanol-heptane as a function of temperature and solvent composition. In the procaine-heptane binary system, we observed a region where the LLPS is metastable with respect to crystallization, which is most commonly observed in the crystallization of modern active pharmaceutical ingredients (APIs); however, we also identified a region of the phase diagram where the LLPS is stable with respect to crystallization, and therefore will persist indefinitely. In the procaine-ethanol-heptane ternary system we identified five different regions, including a homogeneous liquid (L) region, two solid-liquid (SLI and SLII) regions, a liquid-liquid (LILII) region, and a solid-liquid-liquid (SLILII) region. The binary and ternary phase diagrams were also predicted using a state-of-the-art thermodynamic model: the SAFT-γ-Mie equation of state, and the results were compared with experimental data. Our findings highlight the complexity of oiling-out behavior. This work also represents a combined modeling and experimental platform to identify phase boundaries that will enable rational selection of strategies to crystallize active pharmaceutical ingredients with oiling-out risks.


Dynamics of carbon monoxide recombination to fully reduced cytochrome c oxidase in plant mitochondria after low-temperature flash photolysis.
Rebinding of CO to reduced cytochrome c oxidase in plant mitochondria has been monitored optically at 590-630 nm after flash photolysis at low temperature from 160 to 200 K. (1) Under 100%-CO saturation, CO rebinding exhibits a four-step mechanism. The thermodynamic parameters of the first phase have been determined; its activation energy, Ea1, is 38.9 kJ.mol-1 and its enthalpy, delta H+/-1, and entropy, delta S+/-1, of activation are respectively 37.5 kJ.mol-1 and -75.8J.mol-1.K-1. (2) When the CO concentration is decreased to 0.2%, rebinding still occurs according to a four-step mechanism. The rate constant of the first phase is CO-concentration-independent. Under non-saturating conditions there is only one CO molecule per occupied site. The rebinding mechanism does not require additional CO molecules to be present in the haem pocket. (3) Dual-wavelength scanning experiments failed to detect optical forms correlated with the resolved phases. (4) Results are discussed with respect to previous work related to CO rebinding to mammalian cytochrome c oxidase and myoglobin.


Distributed Differentially-Private Algorithms for Matrix and Tensor Factorization.
In many signal processing and machine learning applications, datasets containing private information are held at different locations, requiring the development of distributed privacy-preserving algorithms. Tensor and matrix factorizations are key components of many processing pipelines. In the distributed setting, differentially private algorithms suffer because they introduce noise to guarantee privacy. This paper designs new and improved distributed and differentially private algorithms for two popular matrix and tensor factorization methods: principal component analysis (PCA) and orthogonal tensor decomposition (OTD). The new algorithms employ a correlated noise design scheme to alleviate the effects of noise and can achieve the same noise level as the centralized scenario. Experiments on synthetic and real data illustrate the regimes in which the correlated noise allows performance matching with the centralized setting, outperforming previous methods and demonstrating that meaningful utility is possible while guaranteeing differential privacy.


A randomized trial of intravenous n-acetylcysteine to prevent contrast induced nephropathy in acute coronary syndromes.
Pharmacokinetic data suggests that the intravenous form of n-acetylcysteine (NAC) may be more effective than the oral formulation in preventing contrast induced nephropathy (CIN). NAC owing to its anti-oxidant properties might be beneficial for patients with acute coronary syndromes (ACS) who are at increased risk for CIN. The aim of this prospective randomized, single-center, double-blind, placebo controlled trial (NCT00939913) was to assess the effect of high-dose intravenous NAC on CIN in ACS patients undergoing coronary angiography and/or percutaneous coronary intervention (PCI). We randomized 398 ACS patients scheduled for diagnostic angiography ± PCI to an intravenous regimen of high-dose NAC (1,200 mg bolus followed by 200 mg/hr for 24 hr; n = 206) or placebo (n = 192). The primary end-point was incidence of CIN defined as an increase in serum creatinine concentration ≥ 25% above the baseline level within 72 hr of the administration of intravenous contrast. There was no difference found for the primary end point with CIN in 16% of the NAC group and in 13% of the placebo group (p = 0.40). Change in serum cystatin-C, a sensitive marker for renal function, was 0.046 ± 0.204 in the NAC group and 0.002 ± 0.260 in the control group (p = 0.07). In ACS patients undergoing angiography ± PCI, high-dose intravenous NAC failed to reduce the incidence of CIN.


Finite element predictions of cartilage contact mechanics in hips with retroverted acetabula.
A contributory factor to hip osteoarthritis (OA) is abnormal cartilage mechanics. Acetabular retroversion, a version deformity of the acetabulum, has been postulated to cause OA via decreased posterior contact area and increased posterior contact stress. Although cartilage mechanics cannot be measured directly in vivo to evaluate the causes of OA, they can be predicted using finite element (FE) modeling. The objective of this study was to compare cartilage contact mechanics between hips with normal and retroverted acetabula using subject-specific FE modeling. Twenty subjects were recruited and imaged: 10 with normal acetabula and 10 with retroverted acetabula. FE models were constructed using a validated protocol. Walking, stair ascent, stair descent and rising from a chair were simulated. Acetabular cartilage contact stress and contact area were compared between groups. Retroverted acetabula had superomedial cartilage contact patterns, while normal acetabula had widely distributed cartilage contact patterns. In the posterolateral acetabulum, average contact stress and contact area during walking and stair descent were 2.6-7.6 times larger in normal than retroverted acetabula (P ≤ 0.017). Conversely, in the superomedial acetabulum, peak contact stress during walking was 1.2-1.6 times larger in retroverted than normal acetabula (P ≤ 0.044). Further differences varied by region and activity. This study demonstrated superomedial contact patterns in retroverted acetabula vs widely distributed contact patterns in normal acetabula. Smaller posterolateral contact stress in retroverted acetabula than in normal acetabula suggests that increased posterior contact stress alone may not be the link between retroversion and OA.


In vitro and in vivo radiosensitization of colorectal cancer HT-29 cells by the smac mimetic JP-1201.
The response to neoadjuvant chemoradiation in rectal cancer is variable and unpredictable. Resistance to chemoradiation has been directly correlated with the levels of the inhibitors of apoptosis (IAPs) in several malignancies. Because smac-DIABLO is a pro-apoptotic gene product that directly inhibits the activity of the IAPs, molecules with similar activity might radiosensitize rectal tumors with phenotypes that express high levels of IAPs. This study was undertaken to assess the radiosensitizing properties of the smac mimetic JP-1201 in radioresistant HT-29 colorectal cancer cells in vitro and established xenografts in SCID mice. Survival was determined by clonogenic assays. PARP-1, caspase-8 cleavage, and IAP levels were assessed by Western blot analysis. SCID mice bearing HT-29 xenografts were treated with ionizing radiation: 2.0 Gy x 5; (n = 6), JP-1201 (5.0 mg/Kg i.p., n = 5) or combination treatment (n = 7) and compared to control (n = 8). DNA repair mechanisms were interrogated by gammaH2AX positive foci. Pretreatment of HT-29 cells with JP-1201 (5.0 microM) prior to ionizing radiation (IR) significantly decreased the survival of these cells. SCID mice bearing HT-29 xenografts demonstrated no difference in tumor load in the group receiving exclusively JP-1201 versus control. At the end of the treatment (day 40), a 46% reduction of tumor load was observed in the IR+JP-1201-treated group compared to the IR-only treated group. Radiosensitization was achieved with a substantial elevation of cleaved PARP-1 in JP-1201- treated HT-29 cells versus control cells with a concomitant decrease of XIAP, but not of survivin or cIAP1/2. JP-1201-treated HT-29 cells had a reduced ability to repair double-stranded DNA breaks (DSBs). The smac mimetic JP-1201 decreased the survival of HT-29 cells and tumor growth by an additive effect in apoptosis and a reduction in the level of XIAP and an impairment of DNA repair mechanisms. The pathways leading to this response need to be further investigated.


Evaluation of a Microphysiological Human Placental Barrier Model for Studying Placental Drug Transfer.
Understanding drug transport across the placental barrier is important for assessing the potential fetal drug toxicity and birth defect risks. Current in vivo and in vitro models have structural and functional limitations in evaluating placental drug transfer and toxicity. Microphysiological systems (MPSs) offer more accurate and relevant physiological models of human tissues and organs on a miniature scale for drug development and toxicology testing. MPSs for the placental barrier have been recently explored to study placental drug transfer. We utilized a multilayered hydrogel membrane-based microphysiological model composed of human placental epithelial and endothelial cells to replicate the key structure and function of the human placental barrier. A macroscale human placental barrier model was created using a transwell to compare the results with the microphysiological model. Placental barrier models were characterized by assessing monolayer formation, intercellular junctions, barrier permeability, and their structural integrity. Three small molecule drugs (glyburide, rifaximin, and caffeine) that are prescribed or taken during pregnancy were studied for their placental transfer. The results showed that all three drugs crossed the placental barrier, with transfer rates in the order: glyburide (molecular weight, MW = 494Da) < rifaximin (MW = 785.9Da) < caffeine (MW = 194.19Da). Using non-compartmental analysis, we estimated human pharmacokinetic characteristics based on in vitro data from both MPS and transwell models. While further research is needed, our findings suggest that MPS holds potential as an in vitro tool for studying placental drug transfer and predicting fetal exposure, offering insights into pharmacokinetics.


Radiation induced cell loss in rat submandibular gland and its relation to gland function.
To understand early and late radiation-induced loss of function of the submandibular gland, changes in cell number were documented and correlated with data on gland function. Modulation of the radiation effect by sialogogues was used to investigate possible mechanisms of action. Rats were irradiated with a single dose of 15 Gy of X-rays after pre-treatment with either saline, the muscarinic receptor agonists methacholine or pilocarpine, the adrenergic receptor agonist phenylephrine or methacholine plus phenylephrine. Before and 1-240 days after irradiation, submandibular saliva flow rate was measured. At the same time points and from comparable animals submandibular glands were carefully extirpated, weighed and prepared for light microscopic examination. Soon after irradiation (<30 days) no significant loss of cells was observed, whereas the gland function was severely compromised. Sialogogue pre-treatment attenuated the radiation-induced loss of gland function. At later intervals a considerable loss of acinar cells and to a lesser extent loss of granular convoluted tubule cells were observed. Gland function subsequently declined slowly. Pre-treatment with sialogogues gave transient protection against cell loss and loss of gland function. The lack of cell loss observed soon after irradiation indicates that the observed reduction in gland function was caused by a compromised functioning of the acini. The later loss of cells is probably due to death of cells that normally proliferate, leading to a further reduced secretory capacity. Protection of gland morphology and function by sialogogues at later times must therefore involve resistance of progenitor cells to radiation-induced cell death.


Clinical pathogenesis and diagnosis of osteoarthritis.
New insights into the metabolism of cartilage have increased respect for efforts designed to prevent osteoarthritis. The aging of the population adds to the desire to diagnose and manage the disease before patients become seriously disabled.


The effect of pioglitazone on peroxisome proliferator-activated receptor-gamma target genes related to lipid storage in vivo.
Pioglitazone is a member of the thiazolidinediones (TZDs), insulin-sensitizing agents used to treat type 2 diabetes. The aim of this study was to define the effect of pioglitazone on the expression of genes related to carbohydrate and lipid metabolism in subcutaneous fat obtained from type 2 diabetic patients. Forty-eight volunteers with type 2 diabetes were divided into two groups treated for 12 weeks with placebo or pioglitazone (30 mg/day). The expression of several genes was quantified by real-time RT-PCR. Pioglitazone treatment increased the expression of genes involved in glycerol-3-phosphate synthesis. The mRNA expression of PEPCK-C and glycerol-3-phosphate dehydrogenase (GPDH) increased (P < 0.01) in patients treated with pioglitazone. There was no difference in glycerol kinase (GyK) mRNA levels. The expression of genes that regulate fatty acid availability in adipocytes, including lipoprotein lipase (LPL) and acetyl-CoA synthetase (ACS), was higher (P < 0.01) in pioglitazone-treated patients. Pioglitazone stimulated (P < 0.0001) expression of c-Cbl-associated protein (CAP), whereas tumor necrosis factor-alpha, leptin, resistin, angiopoietin like-4, and 11-beta-hydroxysteroid dehydrogenase type 1 (11beta HSD 1) were not affected by pioglitazone. The baseline peroxisome proliferator-activated receptor (PPAR)-gamma1 mRNA was significantly correlated with mRNA for LPL, CAP, ACS, 11beta HSD 1, GyK, fatty acid synthase, leptin, and GPDH, whereas PPAR-gamma2 mRNA was correlated with CAP, PEPCK-C, leptin, and GPDH. Treatment with pioglitazone increased body weight, and this is associated with upregulation of some, but not all, genes previously demonstrated as "TZD responsive" in subcutaneous fat. The results suggest that TZDs might increase body weight through the upregulation of genes facilitating adipocyte lipid storage in vivo.


Using notochordal cells of developmental origin to stimulate nucleus pulposus cells and bone marrow stromal cells for intervertebral disc regeneration.
Bone marrow stromal cells (BMSCs) have been proposed to complement the declining population of nucleus pulposus cells (NPCs) found in a degenerative intervertebral disc. Although able to stop degeneration, they could not produce enough matrix to restore a healthy state. Looking at development, when a large amount of matrix is produced, the disc also contains notochordal cells (NCs), potential progenitors or regulators of NPCs. The aim of the study was, therefore, to combine NCs to a BMSC/NPC mix and evaluate their effects on cell phenotype and matrix production, in long-term culture. In a 3D hydrogel, NCs were co-cultured in different ratios with BMSCs and/or NPCs. Matrix production, cell morphology, and gene expression of disc markers were assessed after 4 weeks of culture. At day 28, BMSCs/NPCs highly expressed disc matrix markers (type II collagen and aggrecan) and produced disc matrix up to 30 % of values obtained for the positive control (BMSCs under TGFβ stimulation). The addition of NCs only slightly up-regulated marker expression (6-12× increase); an up-regulation not reflected at the matrix level. During the 4 weeks of culture, however, the NC phenotype changed drastically (morphology, disc marker expression). In contrast to previously reported short-term studies, long-term co-cultures with NCs had no substantial effects on BMSCs and NPCs, most likely due to the loss of the NC native phenotype during culture. It, therefore, appears critical to maintain this specific phenotype for a long-term effect of the NCs.


Assessment of a Faculty and Preceptor Development Intervention to Foster Self-Awareness and Self-Confidence.
Objective. To conduct and evaluate the outcomes of a pharmacy faculty and preceptor development program to foster self-awareness and self-confidence. Methods. A faculty and preceptor development intervention was implemented in a multi-campus college of pharmacy to promote and assess for improvements in self-awareness and self-confidence. Faculty members and preceptors were surveyed regarding their self-perceptions and confidence at baseline and following an intervention in which they completed the Birkman Method self-assessment and participated in a training program with an active-learning component. A longitudinal follow-up survey was conducted to assess the long-term impacts of the intervention. Results. Faculty members and preceptors experienced significant improvements in self-awareness from baseline following the development intervention. They also experienced increases in self-confidence related to coaching. A survey evaluating the longitudinal impact of the intervention indicated a positive association between receiving a sufficient level of Birkman Method training and improved ability of both faculty members and preceptors to manage professional relationships. Similarly, a positive association was identified between the sufficiency of training and preceptors' confidence in their ability to manage personal relationships and stress following the intervention. Conclusion. Faculty members and preceptors teach students to be more self-aware and confident, yet both groups often need to grow in these areas themselves. A faculty and preceptor development intervention using the Birkman Method self-assessment is one approach to facilitating growth in these educators' self-awareness and self-confidence.