Beneficial and harmful effects of anthracyclines in the treatment of childhood acute lymphoblastic leukaemia: a systematic review and meta-analysis.
Anthracyclines are used to treat childhood acute lymphoblastic leukaemia (ALL) but non-randomized studies suggest that cardiotoxicity may be a problem. Individual patient data from trials in childhood ALL that randomized anthracyclines or methods of reducing cardiotoxicity were analysed by standard meta-analysis methods. Results were grouped and combined according to: addition of an anthracycline to standard therapy, type of anthracycline, mode of administration, and the use of a cardioprotectant. Data from 958 patients in 4 trials, recruiting between 1972 and 1984, showed that addition of an anthracycline reduced bone marrow relapse and, non-significantly, non-bone marrow relapse, resulting in an increased relapse-free interval. However there was a non-significant increase in induction failures, and in deaths in first remission. Event-free survival at 5 years was 56.7% with anthracycline versus 52.8% without (Odds Ratio = 0.91; 95% Confidence Interval = 0.76-1.10; P = 0.3). There were no significant differences found in other treatment comparisons. The limited data from trials did not demonstrate differences in clinically evident cardiotoxicity. Anthracyclines are effective against bone marrow relapse but have not been shown to significantly increase event free survival in childhood ALL. The evidence on type of anthracycline, method of administration or use of cardioprotectant was insufficient to be able to rule out important differences.


Evaluation of stroke volume variations obtained with the pressure recording analytic method.
To investigate whether stroke volume variations obtained with the pressure recording analytic method can predict fluid responsiveness in mechanically ventilated patients with circulatory failure. Prospective study. Surgical intensive care unit of a university hospital. Thirty-five mechanically ventilated patients with circulatory failure for whom the decision to give fluid was taken by the physician were included. Exclusion criteria were: Arrhythmia, tidal volume <8 mL/kg, left ventricular ejection fraction<50%, right ventricular dysfunction, and heart rate/respiratory rate ratio <3.6. Fluid challenge with 500 mL of saline over 15 mins. Stroke volume variations and cardiac output obtained with a pressure recording analytic method, pulse pressure variations, and cardiac output estimated by echocardiography were recorded before and after volume expansion. Patients were defined as responders if stroke volume obtained using echocardiography increased by ≥15% after volume expansion. Nineteen patients responded to the fluid challenge. Median [interquartile range, 25% to 75%] stroke volume variation values at baseline were not different in responders and nonresponders (10% [8-16] vs. 14% [12-16]), whereas pulse pressure variations were significantly higher in responders (17% [13-19] vs. 7% [5-10]; p < .0001). A 12.6% stroke volume variations threshold discriminated between responders and nonresponders with a sensitivity of 63% (95% confidence interval 38% to 84%) and a specificity of 69% (95% confidence interval 41% to 89%). A 10% pulse pressure variation threshold discriminated between responders and nonresponders with a sensitivity of 89% (95% confidence interval 67% to 99%) and a specificity of 88% (95% confidence interval 62% to 98%). The area under the receiver operating characteristic curves was different between pulse pressure variations (0.95; 95% confidence interval 0.82-0.99) and stroke volume variations (0.60; 95% confidence interval 0.43-0.76); p < .0001). Volume expansion-induced changes in cardiac output measured using echocardiography or pressure recording analytic method were not correlated (r = 0.14; p > .05) and the concordance rate of the direction of change in cardiac output was 60%. Stroke volume variations obtained with a pressure recording analytic method cannot predict fluid responsiveness in intensive care unit patients under mechanical ventilation. Cardiac output measured by this device is not able to track changes in cardiac output induced by volume expansion.


Viscoelasticity of the sarcomere matrix of skeletal muscles. The titin-myosin composite filament is a dual-stage molecular spring.
The mechanical roles of sarcomere-associated cytoskeletal lattices were investigated by studying the resting tension-sarcomere length curves of mechanically skinned rabbit psoas muscle fibers over a wide range of sarcomere strain. Correlative immunoelectron microscopy of the elastic titin filaments of the endosarcomeric lattice revealed biphasic extensibility behaviors and provided a structural interpretation of the multiphasic tension-length curves. We propose that the reversible change of contour length of the extensible segment of titin between the Z line and the end of thick filaments underlies the exponential rise of resting tension. At and beyond an elastic limit near 3.8 microns, a portion of the anchored titin segment that adheres to thick filaments is released from the distal ends of thick filament. This increase in extensible length of titin results in a net length increase in the unstrained extensible segment, thereby lowering the stiffness of the fiber, lengthening the slack sarcomere length, and shifting the yield point in postyield sarcomeres. Thus, the titin-myosin composite filament behaves as a dual-stage molecular spring, consisting of an elastic connector segment for normal response and a longer latent segment that is recruited at and beyond the elastic limit of the sarcomere. Exosarcomeric intermediate filaments contribute to resting tension only above 4.5 microns. We conclude that the interlinked endo- and exosarcomeric lattices are both viscoelastic force-bearing elements. These distinct cytoskeletal lattices appear to operate over two ranges of sarcomere strains and collectively enable myofibrils to respond viscoelastically over a broad range of sarcomere and fiber lengths.


The addition of arsenic trioxide to low-dose Ara-C in older patients with AML does not improve outcome.
Most patients with acute myeloid leukaemia (AML) are older, with many unsuitable for conventional chemotherapy. Low-dose Ara-C (LDAC) is superior to best supportive care but is still inadequate. The combination of arsenic trioxide (ATO) and LDAC showed promise in an unrandomised study. We report a randomised trial of LDAC versus LDAC+ATO. Patients with AML according to WHO criteria or myelodysplastic syndrome with >10% blasts, considered as unfit for conventional chemotherapy, were randomised between subcutaneous Ara-C (20 mg b.d. for 10 days) and the same LDAC schedule with ATO (0.25 mg/kg) on days 1-5, 9 and 11, for at least four courses every 4 to 6 weeks. Overall 166 patients were entered; the trial was terminated on the advice of the DMC, as the projected benefit was not observed. Overall 14% of patients achieved complete remission (CR) and 7% CRi. Median survival was 5.5 months and 19 months for responders (CR: not reached; CRi: 14 months; non-responders: 4 months). There were no differences in response or survival between the arms. Grade 3/4 cardiac and liver toxicity, and supportive care requirements were greater in the ATO arm. This randomised comparison demonstrates that adding ATO to LDAC provides no benefit for older patients with AML.


Perthes' or Perthes' resembling disease in 13-month-old boy.
Onset of Perthes' disease is reported frequently from the age of 2 years. Latest publications showed cases with onset of this disease in infancy at ages of 17 and 18 months. We report the case of a 13-month-old boy, who presented with left-sided limping. Radiological examination showed reduced height and fragmentation of the femoral head. Magnetic resonance imaging showed the typical signs of an avascular necrosis. Follow-up was done after 3, 7 and 15 months. Plain radiography showed the femoral head in a state of reparation. This is the youngest documented case of Legg-Calvé-Perthes' disease and is discussed under consideration of the current literature.


Degradation of an old human protein: age-dependent cleavage of γS-crystallin generates a peptide that binds to cell membranes.
Long-lived proteins exist in a number of tissues in the human body; however, little is known about the reactions involved in their degradation over time. Lens proteins, which do not turn over, provide a useful system to examine such processes. Using a combination of Western blotting and proteomic methodology, age-related changes to a major protein, γS-crystallin, were studied. By teenage years, insoluble intact γS-crystallin was detected, indicative of protein denaturation. This was not the only change, however, because blots revealed evidence of significant cross-linking as well as cleavage of γS-crystallin in all adult lenses. Cleavage at a serine residue near the C terminus was a major reaction that caused the release of a 12-residue peptide, SPAVQSFRRIVE, which bound tightly to lens cell membranes. Several other crystallin-derived peptides with double basic residues also lodged in the cell membrane fraction. Model studies showed that once cleaved from γS-crystallin, SPAVQSFRRIVE adopts a markedly different shape from that in the intact protein. Further, the acquired helical conformation may explain why the peptide seems to affect water permeability. This observation may help explain the changes to cell membranes known to be associated with aging in human lenses. Age-related cleavage of long-lived proteins may therefore yield peptides with untoward biological activity.


Passive and active vaccination strategies to prevent ricin poisoning.
Ricin toxin (RT) is derived from castor beans, produced by the plant Ricinus communis. RT and its toxic A chain (RTA) have been used therapeutically to arm ligands that target disease-causing cells. In most cases these ligands are cell-binding monoclonal antibodies (MAbs). These ligand-toxin conjugates or immunotoxins (ITs) have shown success in clinical trials [1]. Ricin is also of concern in biodefense and has been classified by the CDC as a Class B biothreat. Virtually all reports of RT poisoning have been due to ingestion of castor beans, since they grow abundantly throughout the world and are readily available. RT is easily purified and stable, and is not difficult to weaponize. RT must be considered during any "white powder" incident and there have been documented cases of its use in espionage [2,3]. The clinical syndrome resulting from ricin intoxication is dependent upon the route of exposure. Countermeasures to prevent ricin poisoning are being developed and their use will depend upon whether military or civilian populations are at risk of exposure. In this review we will discuss ricin toxin, its cellular mode of action, the clinical syndromes that occur following exposure and the development of pre- and post-exposure approaches to prevent of intoxication.


Phenotypic mismatches reveal escape from arms-race coevolution.
Because coevolution takes place across a broad scale of time and space, it is virtually impossible to understand its dynamics and trajectories by studying a single pair of interacting populations at one time. Comparing populations across a range of an interaction, especially for long-lived species, can provide insight into these features of coevolution by sampling across a diverse set of conditions and histories. We used measures of prey traits (tetrodotoxin toxicity in newts) and predator traits (tetrodotoxin resistance of snakes) to assess the degree of phenotypic mismatch across the range of their coevolutionary interaction. Geographic patterns of phenotypic exaggeration were similar in prey and predators, with most phenotypically elevated localities occurring along the central Oregon coast and central California. Contrary to expectations, however, these areas of elevated traits did not coincide with the most intense coevolutionary selection. Measures of functional trait mismatch revealed that over one-third of sampled localities were so mismatched that reciprocal selection could not occur given current trait distributions. Estimates of current locality-specific interaction selection gradients confirmed this interpretation. In every case of mismatch, predators were "ahead" of prey in the arms race; the converse escape of prey was never observed. The emergent pattern suggests a dynamic in which interacting species experience reciprocal selection that drives arms-race escalation of both prey and predator phenotypes at a subset of localities across the interaction. This coadaptation proceeds until the evolution of extreme phenotypes by predators, through genes of large effect, allows snakes to, at least temporarily, escape the arms race.


PLANETARY SCIENCE: Asteroids Come of Age.
Since the discovery of the first asteroid almost 200 years ago, these celestial bodies have advanced from nuisances hampering stellar studies to the focus of geological studies aimed at understanding the early solar system. Binzel describes the history of their study, which has now culminated in NASA's NEAR mission to the asteroid Eros. Four papers report first results from this first rendezvous mission with an asteroid.


The structural basis of fungal glucuronoyl esterase activity on natural substrates.
Structural and functional studies were conducted of the glucuronoyl esterase (GE) from Cerrena unicolor (CuGE), an enzyme catalyzing cleavage of lignin-carbohydrate ester bonds. CuGE is an α/β-hydrolase belonging to carbohydrate esterase family 15 (CE15). The enzyme is modular, comprised of a catalytic and a carbohydrate-binding domain. SAXS data show CuGE as an elongated rigid molecule where the two domains are connected by a rigid linker. Detailed structural information of the catalytic domain in its apo- and inactivated form and complexes with aldouronic acids reveal well-defined binding of the 4-O-methyl-a-D-glucuronoyl moiety, not influenced by the nature of the attached xylo-oligosaccharide. Structural and sequence comparisons within CE15 enzymes reveal two distinct structural subgroups. CuGE belongs to the group of fungal CE15-B enzymes with an open and flat substrate-binding site. The interactions between CuGE and its natural substrates are explained and rationalized by the structural results, microscale thermophoresis and isothermal calorimetry.


Hip Pain: Dry Needling Versus Cortisone Injections.
Greater trochanteric pain syndrome (GTPS) is chronic, intermittent pain and tenderness on the outside of the hip. The medical community once thought that a swollen hip bursa was the source of such pain, which led to the use of corticosteroid injections to the bursa to help decrease swelling and pain. However, researchers now believe that injuries to the muscles and tendons around the hip are the actual cause of this pain, and that inflammation is often not involved. A study published in the April 2017 issue of JOSPT explores dry needling as an alternative to cortisone injections to reduce pain and improve function in patients with GTPS. J Orthop Sports Phys Ther 2017;47(4):240. doi:10.2519/jospt.2017.0504.


Differential uptake of DNA-poly(ethylenimine) polyplexes in cells cultured on collagen and fibronectin surfaces.
Genetically modified bone marrow-derived mesenchymal stem cells (MSCs) have proven to be efficient cell carriers for local or systemic delivery of therapeutics as well as growth factors to augment tissue formation. However, efficient non-viral gene transfer to these cells is limiting their applicability. Although most studies have focused on designing more efficient condensation agents for DNA, our focus in this manuscript is to study the role of two extracellular matrix (ECM) proteins, collagen I (Col I) and fibronectin (Fn), on the ability of MSCs to become transfected. Here we report that plating MSCs on Col I-coated surfaces inhibits transfection, while plating MSCs on Fn-coated surfaces enhances transfection. The mechanism by which these ECM proteins affect non-viral gene transfer involves the endocytosis pathway used for polyplex uptake and intracellular tension. We found that Fn promoted internalization through clathrin-mediated endocytosis and that this pathway resulted in more efficient transfection than caveolae-mediated endocytosis and macropinocytosis. Further, the disruption of actin-myosin interactions resulted in an enhancement of gene transfer for cells plated on Fn-coated surfaces, but not for cells plated on Col I. We believe that the cellular microenvironment can be engineered to enhance the ability of cells to become transfected and that through understanding the mechanisms by which the ECM affects non-viral gene transfer better materials and transfection protocols can be realized.


Symmetric States Requiring System Asymmetry.
Spontaneous synchronization has long served as a paradigm for behavioral uniformity that can emerge from interactions in complex systems. When the interacting entities are identical and their coupling patterns are also identical, the complete synchronization of the entire network is the state inheriting the system symmetry. As in other systems subject to symmetry breaking, such symmetric states are not always stable. Here, we report on the discovery of the converse of symmetry breaking-the scenario in which complete synchronization is not stable for identically coupled identical oscillators but becomes stable when, and only when, the oscillator parameters are judiciously tuned to nonidentical values, thereby breaking the system symmetry to preserve the state symmetry. Aside from demonstrating that diversity can facilitate and even be required for uniformity and consensus, this suggests a mechanism for convergent forms of pattern formation in which initially asymmetric patterns evolve into symmetric ones.


Multiple functions for NGF receptor in developing, aging and injured rat teeth are suggested by epithelial, mesenchymal and neural immunoreactivity.
We have used immunocytochemistry to analyse expression of nerve growth factor receptor (NGFR) in developing, aging and injured molar teeth of rats. The patterns of NGFR immunoreactivity (IR) in developing epithelia and mesenchyme matched the location of NGFR mRNA assayed by in situ hybridization with a complementary S35-labeled RNA probe. The following categories of NGFR expression were found. (1) There was NGFR-IR in the dental lamina epithelium and in adjacent mesenchyme during early stages of third molar formation. (2) NGFR-IR nerve fibers were posterior and close to the bud epithelium. (3) During crown morphogenesis NGFR expression was prominent in internal enamel epithelium and preodontoblasts; it faded as preameloblasts elongated and as odontoblasts began to make predentin matrix; and it was weak or absent from outer enamel epithelium, the cervical loop, and differentiated ameloblasts and odontoblasts. (4) When NGFR-IR nerve fibers entered the molars late in the bell stage, they innervated the most mature peripheral pulp and dentin in an asymmetric pattern which correlated more with asymmetric enamel synthesis than with mesenchymal NGFR-IR distribution. (5) The mesenchymal pulp cells continued to have intense NGFR expression in adult teeth, especially near coronal tubular dentin. (6) The pulpal NGFR-IR decreased in very old rats or subjacent to reparative dentin (naturally occurring or experimentally induced). (7) During root formation, the preodontoblasts had NGFR-IR but most root mesenchymal cells and Hertwig's epithelial root sheath did not. This work suggests that there are important epithelial and mesenchymal targets of NGF regulation during molar morphogenesis that differ for crown and root development and that do not correlate with neural development. The continuing expression of NGFR-IR by pulpal mesenchymal cells in adult rats was most intense near coronal odontoblasts making tubular dentin; and it was lost during aging, or subjacent to sites of dentin injury that caused a phenotypic change in the odontoblast layer.


miR-346 controls release of TNF-α protein and stability of its mRNA in rheumatoid arthritis via tristetraprolin stabilization.
TNF-α is a major cytokine implicated in rheumatoid arthritis. Its expression is regulated both at the transcriptional and posttranscriptional levels and recent data demonstrated that miRNAs are implicated in TNF-α response in macrophages. LPS-activated FLS isolated from RA patients express TNF-α mRNA but not the mature protein. This prompted us to look for miRNAs which could be implicated in this anti-inflammatory effect. Using a microarray, we found two miRNAs, miR-125b and miR-939 predicted to target the 3'-UTR of TNF-α mRNA, to be up-regulated in RA FLS in response to LPS, but their repression did not restore mature TNF-α expression in FLS. We showed previously that miR-346, which is upregulated in LPS-activated FLS, inhibited Btk expression that stabilized TNF-α mRNA. Blocking miR-346 reestablished TNF-α expression in activated FLS. Interestingly, transfection of miR-346 in LPS-activated THP-1 cells inhibited TNF-α secretion. We also demonstrated that TTP, a RNA binding protein which inhibited TNF-α synthesis, is overexpressed in activated FLS and that inhibition of miR-346 decreases its expression. Conversely, transfection of miR-346 in LPS-activated THP-1 cells increased TTP mRNA expression and inhibited TNF-α release. These results indicate that miR-346 controls TNF-α synthesis by regulating TTP expression.


Lineage-specific late pleistocene expansion of an endemic subtropical gossamer-wing damselfly, Euphaea formosa, in Taiwan.
Pleistocene glacial oscillations have significantly affected the historical population dynamics of temperate taxa. However, the general effects of recent climatic changes on the evolutionary history and genetic structure of extant subtropical species remain poorly understood. In the present study, phylogeographic and historical demographic analyses based on mitochondrial and nuclear DNA sequences were used. The aim was to investigate whether Pleistocene climatic cycles, paleo-drainages or mountain vicariance of Taiwan shaped the evolutionary diversification of a subtropical gossamer-wing damselfly, Euphaea formosa. E. formosa populations originated in the middle Pleistocene period (0.3 Mya) and consisted of two evolutionarily independent lineages. It is likely that they derived from the Pleistocene paleo-drainages of northern and southern Minjiang, or alternatively by divergence within Taiwan. The ancestral North-central lineage colonized northwestern Taiwan first and maintained a slowly growing population throughout much of the early to middle Pleistocene period. The ancestral widespread lineage reached central-southern Taiwan and experienced a spatial and demographic expansion into eastern Taiwan. This expansion began approximately 30,000 years ago in the Holocene interglacial period. The ancestral southern expansion into eastern Taiwan indicates that the central mountain range (CMR) formed a barrier to east-west expansion. However, E. formosa populations in the three major biogeographic regions (East, South, and North-Central) exhibit no significant genetic partitions, suggesting that river drainages and mountains did not form strong geographical barriers against gene flow among extant populations. The present study implies that the antiquity of E. formosa's colonization is associated with its high dispersal ability and larval tolerance to the late Pleistocene dry grasslands. The effect of late Pleistocene climatic changes on the subtropical damselfly's historical demography is lineage-specific, depending predominantly on its colonization history and geography. It is proposed that the Riss and Würm glaciations in the late Pleistocene period had a greater impact on the evolutionary diversification of subtropical insular species than the last glacial maximum (LGM).


Correlative fluorescence and electron microscopy on ultrathin cryosections: bridging the resolution gap.
Microscopy has become increasingly important for analysis of cells and cell function in recent years. This is due in large part to advances in light microscopy that facilitate quantitative studies and improve imaging of living cells. Analysis of fluorescence signals has often been a key feature in these advances. Such studies involve a number of techniques, including imaging of fluorescently labeled proteins in living cells, single-cell physiological experiments using fluorescent indicator probes, and immunofluorescence localization. The importance of fluorescence microscopy notwithstanding, there are instances in which electron microscopy provides unique information about cell structure and function. Correlative microscopy in which a fluorescence signal is reconciled with a signal from the electron microscope is an additional tool that can provide powerful information for cellular analysis. Here we review two different methodologies for correlative fluorescence and electron microscopy using ultrathin cryosections and the advantages attendant on this approach. (J Histochem Cytochem 49:803-808, 2001)


Differential impact of EGFR-targeted therapies on hypoxia responses: implications for treatment sensitivity in triple-negative metastatic breast cancer.
In solid tumors, such as breast cancer, cells are exposed to hypoxia. Cancer cells adapt their metabolism by activating hypoxia-inducible factors (HIFs) that promote the transcription of genes involved in processes such as cell survival, drug resistance and metastasis. HIF-1 is also induced in an oxygen-independent manner through the activation of epidermal growth factor receptor tyrosine kinase (EGFR-TK). Triple-negative breast cancer (TNBC) is a subtype of invasive breast cancer characterized by negative expression of hormonal and HER2 receptors, and this subtype generally overexpresses EGFR. Sensitivity to three EGFR inhibitors (cetuximab, gefitinib and lapatinib, an HER2/EGFR-TK inhibitor) was evaluated in a metastatic TNBC cell model (MDA-MB-231), and the impact of these drugs on the activity and stability of HIF was assessed. MDA-MB-231 cells were genetically modified to stably express an enhanced green fluorescent protein (EGFP) induced by hypoxia; the Ca9-GFP cell model reports HIF activity, whereas GFP-P564 reports HIF stability. The reporter signal was monitored by flow cytometry. HIF-1 DNA-binding activity, cell migration and viability were also evaluated in response to EGFR inhibitors. Cell fluorescence signals strongly increased under hypoxic conditions (> 30-fold). Cetuximab and lapatinib did not affect the signal induced by hypoxia, whereas gefitinib sharply reduced its intensity in both cell models and also diminished HIF-1 alpha levels and HIF-1 DNA-binding activity in MDA-MB-231 cells. This gefitinib feature was associated with its ability to inhibit MDA-MB-231 cell migration and to induce cell mortality in a dose-dependent manner. Cetuximab and lapatinib had no effect on cell migration or cell viability. Resistance to cetuximab and lapatinib and sensitivity to gefitinib were associated with their ability to modulate HIF activity and stability. In conclusion, downregulation of HIF-1 through EGFR signaling seems to be required for the induction of a positive response to EGFR-targeted therapies in TNBC.


Case management of malaria: diagnosis.
Laboratory diagnosis of malaria in South Africa has traditionally relied on microscopic examination of stained blood films. More recently, rapid diagnostic tests (RDTs) have been introduced into routine use, and molecular methods like polymerase chain reaction are useful in certain situations. As with all laboratory tests, each technique has its advantages and disadvantages. Microscopy and RDTs, if appropriately quality assured, are adequate for clinical case management. For elimination, active surveillance will need to be expanded substantially, with wider use of more sensitive diagnostic nucleic acid amplification techniques, and/or serology. To facilitate surveillance activities, techniques suitable for field or near-field use would be ideal. A long-running external quality assessment programme in South Africa has shown some deficiencies in the quality of malaria diagnosis in routine laboratories. Quality systems across the spectrum of diagnostic facilities in South Africa need strengthening, to ensure progress towards elimination.


Expression patterns of chondrocyte genes cloned by differential display in tibial dyschondroplasia.
Tibial dyschondroplasia (TD) appears to involve a failure of the growth plate chondrocytes within growing long bones to differentiate fully to the hypertrophic stage, resulting in a mass of prehypertrophic chondrocytes which form the avascular TD lesion. Many biochemical and molecular markers of chondrocyte hypertrophy are absent from the lesion, or show reduced expression, but the cause of the disorder remains to be identified. As differentiation to the hypertrophic state is impaired in TD, we hypothesised that chondrocyte genes that are differentially expressed in the growth plate should show altered expression in TD. Using differential display, four genes, B-cadherin, EF2, HT7 and Ex-FABP were cloned from chondrocytes stimulated to differentiate to the hypertrophic stage in vitro, and their differential expression confirmed in vivo. Using semi-quantitative RT-PCR, the expression patterns of these genes were compared in chondrocytes from normal and TD growth plates. Surprisingly, none of these genes showed the pattern of expression that might be expected in TD lesion chondrocytes, and two of them, B-cadherin and Ex-FABP, were upregulated in the lesion. This indicates that the TD phenotype does not merely reflect the absence of hypertrophic marker genes, but may be influenced by more complex developmental mechanisms/defects than previously thought.


Hierarchical Micro/Nano-Porous Acupuncture Needles Offering Enhanced Therapeutic Properties.
Acupuncture as a therapeutic intervention has been widely used for treatment of many pathophysiological disorders. For achieving improved therapeutic effects, relatively thick acupuncture needles have been frequently used in clinical practice with, in turn, enhanced stimulation intensity. However due to the discomforting nature of the larger-diameter acupuncture needles there is considerable interest in developing advanced acupuncture therapeutical techniques that provide more comfort with improved efficacy. So motivated, we have developed a new class of acupuncture needles, porous acupuncture needles (PANs) with hierarchical micro/nano-scale conical pores upon the surface, fabricated via a simple and well known electrochemical process, with surface area approximately 20 times greater than conventional acupuncture needles. The performance of these high-surface-area PANs is evaluated by monitoring the electrophysiological and behavioral responses from the in vivo stimulation of Shenmen (HT7) points in Wistar rats, showing PANs to be more effective in controlling electrophysiological and behavioral responses than conventional acupuncture needles. Comparative analysis of cocaine induced locomotor activity using PANs and thick acupuncture needles shows enhanced performance of PANs with significantly less pain sensation. Our work offers a unique pathway for achieving a comfortable and improved acupuncture therapeutic effect.


Incorporation of NKT cell-activating glycolipids enhances immunogenicity and vaccine efficacy of Mycobacterium bovis bacillus Calmette-Guerin.
The attenuated strain of Mycobacterium bovis known as bacille Calmette-Guérin (BCG) has been widely used as a vaccine for prevention of disease by Mycobacterium tuberculosis, but with relatively little evidence of success. Recent studies suggest that the failure of BCG may be due to its retention of immune evasion mechanisms that delay or prevent the priming of robust protective cell-mediated immunity. In this study, we describe an approach to enhance the immunogenicity of BCG by incorporating glycolipid activators of CD1d-restricted NKT cells, a conserved T cell subset with the potential to augment many types of immune responses. A method was developed for stably incorporating two forms of the NKT cell activator alpha-galactosylceramide into live BCG organisms, and the impact of this on stimulation of T cell responses and protective antimycobacterial immunity was evaluated. We found that live BCG containing relatively small amounts of incorporated alpha-galactosylceramide retained the ability to robustly activate NKT cells. Compared with immunization with unmodified BCG, the glycolipid-modified BCG stimulated increased maturation of dendritic cells and markedly augmented the priming of Ag-specific CD8(+) T cells responses. These effects were correlated with improved protective effects of vaccination in mice challenged with virulent M. tuberculosis. These results support the view that mycobacteria possess mechanisms to avoid stimulation of CD8(+) T cell responses and that such responses contribute significantly to protective immunity against these pathogens. Our findings raise the possibility of a simple modification of BCG that could yield a more effective vaccine for control of tuberculosis.


Pregnancy and malaria exposure are associated with changes in the B cell pool and in plasma eotaxin levels.
Pregnancy triggers immunological changes aimed to tolerate the fetus, but its impact on B lymphocytes is poorly understood. In addition, exposure to the Plasmodium parasite is associated with altered distribution of peripheral memory B cell (MBC) subsets. To study the combined impact of high malaria exposure and pregnancy in B cell subpopulations, we analyzed PBMCs from pregnant and nonpregnant individuals from a malaria-nonendemic country (Spain) and from a high malaria-endemic country (Papua New Guinea). In the malaria-naive cohorts, pregnancy was associated with a significant expansion of all switched (IgD(-)) MBC and a decrease of naive B cells. Malaria-exposed women had more atypical MBC and fewer marginal zone-like MBC, and their levels correlated with both Plasmodium vivax- and Plasmodium falciparum-specific plasma IgG levels. Classical but not atypical MBC were increased in P. falciparum infections. Moreover, active atypical MBC positively correlated with proinflammatory cytokine plasma concentrations and had lower surface IgG levels than the average. Decreased plasma eotaxin (CCL11) levels were associated with pregnancy and malaria exposure and also correlated with B cell subset frequencies. Additionally, active atypical and active classical MBC expressed higher levels of eotaxin receptor CCR3 than the other B cell subsets, suggesting a chemotactic effect of eotaxin on these B cell subsets. These findings are important to understand immunity to infections like malaria that result in negative outcomes for both the mother and the newborn and may have important implications on vaccine development.


A personal journey with matrix metalloproteinases.
I was given the honor of delivering the 2015 Lifetime Membership Award lecture at the International Proteolysis Society's annual meeting held in Penang, Malaysia in October 2015. It gave me an opportunity to look back on how I started my research on matrix metalloproteinases (MMPs) and how I continued to work on these proteinases for the next 42 years. This is a series of sketches from the personal journey that I took with MMPs, starting from the purification of metalloproteinases, cloning, structural studies, then to a more recent encounter, endocytic regulation of matrix-degrading metalloproteinases.


A phase I vaccine trial with peptides reflecting ras oncogene mutations of solid tumors.
Mutations in the ras genes occur in 20% of all human cancers. These genes, in turn, produce mutated proteins that are unique to cancer cells, rendering them distinguishable from normal cells by the immune system. Thus, mutated Ras proteins may form potential targets for immune therapy. We conducted a phase I/pilot clinical trial in patients with advanced cancers to test the toxicity and the ability to induce an immune response by vaccination with 13-mer mutated Ras peptides reflecting codon 12 mutations. These peptides corresponded to each of the patient's own tumor Ras mutation. Patients were vaccinated monthly x3 subcutaneously with the specific Ras peptide along with Detox adjuvant (RiBi ImmunoChem Research, Inc., Hamilton, MT, U.S.A.) at one of five different peptide dose levels (100, 500, 1,000, 1,500, and 5,000 micrograms). Three out of 10 evaluable patients generated a mutant Ras specific CD4+ and/or CD8+ T-cell immune response. The CD8+ cytotoxic cells specific for Gly to Val mutation at codon 12 were capable of lysing an HLA-A2-matched tumor cell line carrying the corresponding mutant but not the wild-type ras gene. The treatment has been well tolerated with no evidence of serious acute or delayed systemic side effects on any of the five dose levels. We demonstrated that we can generate in cancer patients specific T-lymphocyte responses that detect single amino acid differences in Ras oncoproteins. Neither the immune responses nor the minor side effects seen were found to be dose dependent. This approach may provide a unique opportunity for generating a tumor-directed therapy. Also, in vitro stimulation of these cells with the corresponding peptide generated specific T-cell lines that could be used for adoptive immune therapy.


Epilepsy and psychogenic non-epileptic seizures in forcibly displaced people: A scoping review.
With a growing number of forcibly displaced people (FDP) globally, the focus on their medical needs has necessarily increased. Studies about the prevalence and incidence of epilepsy and psychogenic non-epileptic seizures (PNES) in this population are, though, sparse. This Review highlights the importance of exploring and managing both conditions in these vulnerable people to promote global health. We performed an exhaustive review of 10 databases, as well as a manual search of relevant websites related to global health and refugee-related organizations. We analyzed data related to the prevalence and incidence of epilepsy and PNES; health visits; costs of medical care and challenges faced by healthcare workers in relation to FDP with these conditions. Fifty six papers met our inclusion criteria. Of these, 53 reported directly or indirectly on the prevalence of epilepsy and its costs in FDP. Two articles reported on the prevalence of PNES in forcibly displaced people. The reported prevalence of epilepsy in FDP varied from 0.2% to 39.13%, being highest in people with pre-existing neurological or psychiatric comorbidities. Only one study reported on the incidence of epilepsy in internally displaced children. Data from the United Nations Relief and Works Agency for Palestine Refugees in the Near East (UNRWA) have identified a tendency to higher incidence of epilepsy in children following wars. While within displaced women without a history of sexual violence the rate of PNES was 16.7%, in FDP women with such a history the rate of PNES was 43.7% (p=0.02). The healthcare costs for epilepsy can be high, with recurrent health visits related to seizures being the most common cause of health encounters in refugee camps. Increasing awareness and further studies of multicultural aspects to improve shared understanding of seizure phenomenon in vulnerable displaced populations would seem crucial.


The Effect of Breakfast Prior to Morning Exercise on Cognitive Performance, Mood and Appetite Later in the Day in Habitually Active Women.
Pre-exercise nutritional practices for active females exercising for mood, cognitive and appetite benefits are not well established. Results from an initial field pilot study showed that higher energy intake at breakfast was associated with lower fatigue and higher overall mood and alertness post-exercise (all p < 0.05). In a follow-up, randomised, controlled trial, 24 active women completed three trials in a balanced, cross-over design. At 0815 h participants completed baseline cognitive tasks, mood and appetite visual analogue scales (VAS) and were administered a cereal breakfast (providing 118 or 236 kcal) or no breakfast. After 45 min, they completed a 30 min run at 65% heart rate reserve (HRR). Parameters were re-assessed immediately after exercise, then hourly until lunch (~1240 h), immediately post-lunch and at 1500 and 1900 h via a mobile phone. Breakfast enhanced feelings of relaxation before lunch (p < 0.05, d > 0.40), though breakfast was detrimental for working memory mid-afternoon (p = 0.019, d = 0.37) and mental fatigue and tension later in the day (all p < 0.05, d > 0.038). Breakfast was also beneficial for appetite control before lunch irrespective of size (all p < 0.05, d > 0.43). These data provide information on pre-exercise nutritional practices for active females and suggest that a small breakfast eaten prior to exercise can benefit post-exercise mood and subjective appetite ratings.


Development and Validation of Prognostic Indices for Recovery of Physical Functioning Following Stroke: Part 1.
To develop a prognostic index using Functional Independence Measure grades and stages that would enable clinicians to determine the likelihood of achieving a level of minimum assistance with physical functioning after a stroke. Grades define varying levels of physical function, and stages define varying levels of cognitive functioning. Retrospective cohort study. Veterans Affairs Medical Centers throughout the United States. Veterans with a diagnosis of a new stroke discharged between October 1, 2006, and September 30, 2008, who were below physical grade IV (requiring minimal assistance) at initial rehabilitation assessment. Achievement of physical grade IV or above at final rehabilitation assessment. Physical grade IV was reached by 25.8% of participants who were initially below this grade. Seven variables remained independently predictive of physical grade IV after adjustment. These variables were assigned the following points: age, ≤69 years = 2, 70-79 years = 1, ≥80 years = 0; initial physical grade, I = 0, II = 3, III = 4; initial cognitive stage, I or II = 0, III = 2, IV or V = 3, VI or VII = 4; absence of renal failure = 1; no serious nutritional compromise = 3; the type of rehabilitation services received, consultative = 0, comprehensive = 4; and recovery time between admission and discharge physical grade assessment, 1-2 days = 0, 3-7 days = 4, and ≥8 days = 5. The area under the receiver operating characteristic curve was 0.84 and 0.83 for the point system in the derivation and validation cohorts, respectively. The Hosmer-Lemeshow statistic was not significant (P = .93) in the derivation cohort, indicating that the regression model demonstrated adequate fit. The proportions of patients recovered to physical grade IV in the first (score ≥9), second (score = 10-12), third (score = 13-15), and fourth (score >15) score quartiles were 2.72%, 11.38%, 28.96%, and 60.34%, respectively. By using a simple tool, clinicians can forecast the likelihood of recovery to or above the physical grade IV benchmark by the conclusion of rehabilitation services during the acute stroke hospitalization.


Melittin, a major peptide component of bee venom, and its conjugates in cancer therapy.
Melittin (MEL), a major peptide component of bee venom, is an attractive candidate for cancer therapy. This agent has shown a variety of anti-cancer effects in preclinical cell culture and animal model systems. Despite a convincing efficacy data against variety of cancers, its applicability to humans has met with challenges due to several issues including its non-specific cytotoxicity, degradation and hemolytic activity. Several optimization approaches including utilization of nanoparticle based delivery of MEL have been utilized to circumvent the issues. Here, we summarize the current understanding of the anticancer effects of bee venom and MEL on different kinds of cancers. Further, we also present the available information for the possible mechanism of action of bee venom and/or MEL.


Longitudinal development of attention and inhibitory control during the first year of life.
Executive functions (EFs) are key abilities that allow us to control our thoughts and actions. Research suggests that two EFs, inhibitory control (IC) and working memory (WM), emerge around 9 months. Little is known about IC earlier in infancy and whether basic attentional processes form the "building blocks" of emerging IC. These questions were investigated longitudinally in 104 infants tested behaviorally on two screen-based attention tasks at 4 months, and on IC tasks at 6 and 9 months. Results provided no evidence that basic attention formed precursors for IC. However, there was full support for coherence in IC at 9 months and partial support for stability in IC from 6 months. This suggests that IC emerges earlier than previously assumed. A video abstract of this article can be viewed at: https://www.youtube.com/watch?v=rVE17hooANY.


Activity and functions of the human gluteal muscles in walking, running, sprinting, and climbing.
It has been suggested that the uniquely large gluteus maximus (GMAX) muscles were an important adaptation during hominin evolution based on numerous anatomical differences between humans and extant apes. GMAX electromyographic (EMG) signals have been quantified for numerous individual movements, but not across the range of locomotor gaits and speeds for the same subjects. Thus, comparing relative EMG amplitudes between these activities has not been possible. We assessed the EMG activity of the gluteal muscles during walking, running, sprinting, and climbing. To gain further insight into the function of the gluteal muscles during locomotion, we measured muscle activity during walking and running with external devices that increased or decreased the need to control either forward or backward trunk pitch. We hypothesized that 1) GMAX EMG activity would be greatest during sprinting and climbing and 2) GMAX EMG activity would be modulated in response to altered forward trunk pitch demands during running. We found that GMAX activity in running was greater than walking and similar to climbing. However, the activity during sprinting was much greater than during running. Further, only the inferior portion of the GMAX had a significant change with altered trunk pitch demands, suggesting that the hip extensors have a limited contribution to the control of trunk pitch movements during running. Overall, our data suggest that the large size of the GMAX reflects its multifaceted role during rapid and powerful movements rather than as a specific adaptation for a single submaximal task such as endurance running.


Power MOSFET Linearizer of a High-Voltage Power Amplifier for High-Frequency Pulse-Echo Instrumentation.
A power MOSFET linearizer is proposed for a high-voltage power amplifier (HVPA) used in high-frequency pulse-echo instrumentation. The power MOSFET linearizer is composed of a DC bias-controlled series power MOSFET shunt with parallel inductors and capacitors. The proposed scheme is designed to improve the gain deviation characteristics of the HVPA at higher input powers. By controlling the MOSFET bias voltage in the linearizer, the gain reduction into the HVPA was compensated, thereby reducing the echo harmonic distortion components generated by the ultrasonic transducers. In order to verify the performance improvement of the HVPA implementing the power MOSFET linearizer, we measured and found that the gain deviation of the power MOSFET linearizer integrated with HVPA under 10 V DC bias voltage was reduced (-1.8 and -0.96 dB, respectively) compared to that of the HVPA without the power MOSFET linearizer (-2.95 and -3.0 dB, respectively) when 70 and 80 MHz, three-cycle, and 26 dBm input pulse waveforms are applied, respectively. The input 1-dB compression point (an index of linearity) of the HVPA with power MOSFET linearizer (24.17 and 26.19 dBm at 70 and 80 MHz, respectively) at 10 V DC bias voltage was increased compared to that of HVPA without the power MOSFET linearizer (22.03 and 22.13 dBm at 70 and 80 MHz, respectively). To further verify the reduction of the echo harmonic distortion components generated by the ultrasonic transducers, the pulse-echo responses in the pulse-echo instrumentation were compared when using HVPA with and without the power MOSFET linearizer. When three-cycle 26 dBm input power was applied, the second, third, fourth, and fifth harmonic distortion components of a 75 MHz transducer driven by the HVPA with power MOSFET linearizer (-48.34, -44.21, -48.34, and -46.56 dB, respectively) were lower than that of the HVPA without the power MOSFET linearizer (-45.61, -41.57, -45.01, and -45.51 dB, respectively). When five-cycle 20 dBm input power was applied, the second, third, fourth, and fifth harmonic distortions of the HVPA with the power MOSFET linearizer (-41.54, -41.80, -48.86, and -46.27 dB, respectively) were also lower than that of the HVPA without the power MOSFET linearizer (-25.85, -43.56, -49.04, and -49.24 dB, respectively). Therefore, we conclude that the power MOSFET linearizer could reduce gain deviation of the HVPA, thus reducing the echo signal harmonic distortions generated by the high-frequency ultrasonic transducers in pulse-echo instrumentation.


Differentiation of tumour-promoting stromal myofibroblasts by cancer exosomes.
Activation of myofibroblast rich stroma is a rate-limiting step essential for cancer progression. The responsible factors are not fully understood, but TGFβ1 is probably critical. A proportion of TGFβ1 is associated with extracellular nano-vesicles termed exosomes, secreted by carcinoma cells, and the relative importance of soluble and vesicular TGFβ in stromal activation is presented. Prostate cancer exosomes triggered TGFβ1-dependent fibroblast differentiation, to a distinctive myofibroblast phenotype resembling stromal cells isolated from cancerous prostate tissue; supporting angiogenesis in vitro and accelerating tumour growth in vivo. Myofibroblasts generated using soluble TGFβ1 were not pro-angiogenic or tumour-promoting. Cleaving heparan sulphate side chains from the exosome surface had no impact on TGFβ levels yet attenuated SMAD-dependent signalling and myofibroblastic differentiation. Eliminating exosomes from the cancer cell secretome, targeting Rab27a, abolished differentiation and lead to failure in stroma-assisted tumour growth in vivo. Exosomal TGFβ1 is therefore required for the formation of tumour-promoting stroma.


Correlation between vocal functions and glottal measurements in patients with unilateral vocal fold paralysis.
Observations and analysis of glottal characteristics are critical in choosing the best modality for surgery in patients with unilateral vocal fold paralysis (UVP). This study suggests that multiple glottal characteristics influence the vocal product in patients with UVP. In addition to the horizontal position of the paralyzed vocal fold (deviation from the midline), the glottal area, degree of bowing of the paralyzed and contralateral vocal folds, maximum separation between vocal folds, compensatory glottal maneuvers, and the vertical glottic closure plane significantly influenced the quality of the voice. Clinicians should be aware of these observations to facilitate treatment planning and assessment of the results of surgical procedures used to improve voice quality in cases of UVP.


Mll-COMPASS complexes mediate H3K4me3 enrichment and transcription of the osteoblast master gene Runx2/p57 in osteoblasts.
Expression of Runx2/p57 is a hallmark of the osteoblast-lineage identity. Although several regulators that control the expression of Runx2/p57 during osteoblast-lineage commitment have been identified, the epigenetic mechanisms that sustain this expression in differentiated osteoblasts remain to be completely determined. Here, we assess epigenetic mechanisms associated with Runx2/p57 gene transcription in differentiating MC3T3 mouse osteoblasts. Our results show that an enrichment of activating histone marks at the Runx2/p57 P1 promoter is accompanied by the simultaneous interaction of Wdr5 and Utx proteins, both are components of COMPASS complexes. Knockdown of Wdr5 and Utx expression confirms the activating role of both proteins at the Runx2-P1 promoter. Other chromatin modifiers that were previously described to regulate Runx2/p57 transcription in mesenchymal precursor cells (Ezh2, Prmt5, and Jarid1b proteins) were not found to contribute to Runx2/p57 transcription in full-committed osteoblasts. We also determined the presence of additional components of COMPASS complexes at the Runx2/p57 promoter, evidencing that the Mll2/COMPASS- and Mll3/COMPASS-like complexes bind to the P1 promoter in osteoblastic cells expressing Runx2/p57 to modulate the H3K4me1 to H3K4me3 transition.


Perfectionism as a transdiagnostic process: a clinical review.
Perfectionism is a risk and maintaining factor for eating disorders, anxiety disorders and depression. The objective of this paper is to review the four bodies of evidence supporting the notion that perfectionism is a transdiagnostic process. First, a review of the literature was conducted that demonstrates the elevation of perfectionism across numerous anxiety disorders, depression, and eating disorders compared to healthy controls. Data is presented that shows perfectionism increases vulnerability for eating disorders, and that it maintains obsessive-compulsive disorder, social anxiety and depression as it predicts treatment outcome in these disorders. Second, evidence is examined showing that elevated perfectionism is associated with co-occurrence of psychopathology. Third, the different conceptualisations of perfectionism are reviewed, including a cognitive-behavioural conceptualisation of clinical perfectionism that can be utilised to understand this transdiagnostic process. Fourth, evidence that treatment of perfectionism results in reductions in anxiety, depression and eating pathology is reviewed. Finally, the importance of clinicians considering the routine assessment and treatment of perfectionism is outlined.


Glycaemic index knowledge and use among African Americans with type 2 diabetes.
This exploratory pilot study investigated the extent to which African Americans with type 2 diabetes receive information on the glycaemic index and their knowledge of the index and use of this information. The growing impact of type 2 diabetes has sparked the need for evidence-based policies that result in increased diabetes self-care among affected individuals. Because of the disproportionate prevalence and severity of type 2 diabetes among African Americans, specific focus on this population is warranted. The glycaemic index is a useful dietary tool for controlling blood glucose levels. However, information on knowledge and use of the glycaemic index among African Americans with type 2 diabetes is lacking. The data source was a convenience sample of 60 African American adults with type 2 diabetes from three churches located in the Midwest region of the United States. This was an exploratory study using survey design. Data collection was performed during the period from November 2007 to March 2008. Although 73% of participants attended diabetes education classes, only 50% reported receiving education about the glycaemic index. Those attending diabetes education classes had a statistically significant higher knowledge of the glycaemic index and use of it. No statistically significant differences were found in age, gender, education, income or body mass index on the participants' knowledge of glycaemic index or use of the glycaemic index. Use of certified nurse diabetes educators may be a positive step in providing consistent and up-to-date diabetes care information to African Americans with type 2 diabetes.


Assessment of activity levels for CYP2D6*1, CYP2D6*2, and CYP2D6*41 genes by population pharmacokinetics of dextromethorphan.
The pharmacokinetics of dextromethorphan (DM) is markedly influenced by cytochrome P450 2D6 (CYP2D6) enzyme polymorphisms. The aim of this study was to quantify the effects of the CYP2D6*1, *2, and *41 variants on DM metabolism in vivo and to identify other sources of pharmacokinetic variability. Concentrations of DM and dextrorphan (DO) in plasma and urine were evaluated in 36 healthy Caucasian men. These volunteers participated in three clinical studies and received a single oral dose of 30 mg DM-HBr. Data were modeled simultaneously using the population pharmacokinetics NONMEM software. A five-compartment model adequately described the data. The activity levels of the alleles assessed differed significantly. The clearance attributable to an individual CYP2D6*1 copy was 2.5-fold higher as compared with CYP2D6*2 (5,010 vs. 2,020 l/h), whereas the metabolic activity of CYP2D6*41 was very low (85 l/h). Urinary pH was confirmed as a significant covariate for DM renal clearance. These results refine genotype-based predictions of pharmacokinetics for DM and presumably for other CYP2D6 substrates as well.


Remodeling of the axon initial segment after focal cortical and white matter stroke.
Recovery from stroke requires neuroplasticity within surviving adjacent cortex. The axon initial segment (AIS) is the site of action potential initiation and a focal point for tuning of neuronal excitability. Remodeling of the AIS may be important to neuroplasticity after stroke. Focal cortical stroke in forelimb motor cortex was induced by photothrombosis and compared with sham controls. White matter stroke was produced through stereotactic injection of a vasoconstrictor together with biotinylated dextran amine to retrogradely label injured cortical neurons. AIS length, morphology and number were measured using immunofluorescence and confocal microscopy 2 weeks after stroke. Within the peri-infarct cortex and after white matter stroke, AIS length decreases. This shortening is accompanied by altered AIS morphology. In peri-infarct cortex, the decrease in AIS length after stroke occurs from the distal end of the AIS, resulting in a Nav1.6. γ-aminobutyric acid type A receptor-α2 subunit staining at axoaxonic synapses along the AIS is significantly decreased. In addition, a significant increase in small, immature initial segments is present in layers 2/3 of peri-infarct cortex, reflecting maturation of axonal sprouting and new initial segments from surviving neurons. Stroke alters the compartmental morphology of surviving adjacent neurons in peri-infarct cortex and in neurons whose distal axons are injured by white matter stroke. With a key role in modulation of neuronal excitability, these changes at the AIS may contribute to altered neuronal excitability after injury and prove crucial to increasing neuroplasticity in surviving tissue affected by stroke.


Urinary erythropoietin concentrations after early short-term infusion of high-dose recombinant epo for neuroprotection in preterm neonates.
High-dose recombinant human erythropoietin (rEpo) has first been administered in clinical trials for neuroprotection in very preterm neonates at high risk of brain injury and in (near-) term neonates with hypoxic-ischemic encephalopathy. However, recent trials in adults raised concerns about the safety of high-dose rEpo for neuro- and cardioprotection. To evaluate the putative accumulation or renal leakage of Epo as a function of developmental stage after repetitive early short-term infusion of high-dose rEpo (3 × 3,000 U/kg within 42 h after birth; NCT00413946) for neuroprotection in very preterm infants. Epo concentrations were measured using the ELISA technique in the first two consecutive urine specimens after each rEpo infusion. Renal Epo excretion was significantly higher in preterm infants with gestational ages <29 weeks than in more mature infants and reached up to 23% of the administered rEpo within 8 h after each infusion. The urinary Epo concentration did not increase after three repetitive infusions of high-dose rEpo. The ratio of urinary Epo to total protein concentrations was the same in infants with gestational ages <29 weeks and in those with gestational ages ≥29 weeks. Our data suggest that the higher renal Epo excretion in more immature infants may be attributed to a higher glomerular filtration leakage due to the lower maturation of the kidneys and argue against saturation kinetics after multiple doses of 3,000 U/kg rEpo. This information should be considered in future trials on the use of rEpo for neuroprotection in neonates.


Increased serum levels of soluble receptors for tumor necrosis factor in cancer patients.
Soluble forms of the two molecular species of the cell surface receptors for tumor necrosis factor (TNF) have been detected in normal urine. Using enzyme-linked immunosorbent assays for these soluble receptors, we determined their levels in the sera of 40 healthy subjects and 59 patients with solid tumors. The mean +/- SD concentrations of both the soluble type I (p55) and type II (p75) receptors were significantly higher in the cancer patients than in the healthy controls: 1.96 +/- 1.19 versus 0.79 +/- 0.19 ng/ml (P less than 0.001) and 6.43 +/- 4.8 versus 3.2 +/- 0.6 ng/ml (P less than 0.001), respectively. The incidence and the extent of the increase correlated with the staging of disease. Sera of the cancer patients had a marked inhibitory effect on the in vitro cytocidal activity of TNF. This inhibition was proportional to the content of soluble TNF receptors and could be fully abolished by the addition to the sera of specific antibodies against the receptors. Among the cancer patients, the incidence of increase in the concentrations of soluble TNF receptors (about 70%) greatly exceeded that of the serum carcinoembryonic antigen (about 26%), a commonly used cancer marker. The origin of the serum soluble TNF receptors in cancer patients and the physiological implications of their effect on TNF function remain to be elucidated.


COVID-19, hypertension and cardiovascular diseases: Should we change the therapy?
The coronavirus disease (COVID-19) has spread all around the world in a very short period of time. Recent data are showing significant prevalence of arterial hypertension and cardiovascular diseases (CVD) among patients with COVID-19, which raised many questions about higher susceptibility of patients with these comorbidities to the novel coronavirus, as well as the role of hypertension and CVD in progression and the prognosis of COVID-19 patients. There is a very limited amount of data, usually obtained from a small population, regarding the effect of the underlying disease on the outcome in patients with COVID-19. The evaluation of the treatment of these comorbidities at baseline and during COVID-19 is scarce and the results are conflicting. Hypertension and CVD, after the adjustment for other clinical and demographic parameters, primarily age, did not remain independent predictors of the lethal outcome in COVID-19 patients. Some investigations speculated about the association between the renin-angiotensin-aldosterone system (RAAS) and susceptibility to COVID-19, as well as the relationship between RAAS inhibitors and the adverse outcome in these patients. Withdrawing or switching RAAS inhibitors would have uncertain benefits, but it would definitely have many disadvantages such as uncontrolled hypertension, cardiac function deterioration and renal function impairment, which could potentially induce more complications in patients with COVID-19 than the infection of coronavirus itself. The aim of this review article was to summarize the prevalence of hypertension and CVD in patients with COVID-19, their influence on the outcome and the effect of treatment of hypertension and CVD in COVID-19 patients.


Soft-tissue hemangioblastoma of the retroperitoneum: a case study and review of the literature.
A 71-year-old woman was found to have a solid retroperitoneal mass measuring 7 x 6 cm on a computed tomography scan. The tumor was not connected to the spinal cord or large nerves. It was resected and showed the typical histology of hemangioblastoma, that is, numerous capillaries and stromal cells with focal cytoplasmic vacuolization. Immunohistochemical study revealed that the tumor stromal cells were positive for S-100 protein, neuron-specific enolase, and inhibin-α, supporting the diagnosis. In addition, the tumor cells labeled for carbonic anhydrase IX and brachyury. The diffuse strong carbonic anhydrase IX expression suggested that soft-tissue hemangioblastoma may share with central nervous system hemangioblastoma the tumorigenic mechanism involving the von Hippel-Lindau gene product and hypoxia-inducible factor. The brachyury expression observed in this case, albeit focal and weak, may implicate embryonic hemangioblasts as a potential line of differentiation, as was proposed in central nervous system hemangioblastoma. The patient is well 4 years after the surgery without any recurrence. A review of the literature revealed that soft-tissue hemangioblastoma is typically a solid tumor found in older females and often involves the retroperitoneum/pelvis.


Association of fat mass profile with natriuretic peptide receptor alpha in subcutaneous adipose tissue of medication-free healthy men: A cross-sectional study.
Background: Atrial natriuretic peptide increases lipolysis in human adipocytes by binding to natriuretic peptide receptor-A (NPRA). The aim of the current study was to examine the associations of NPRA mRNA of subcutaneous adipose tissue with fat mass, fat-free mass, body mass index (BMI) and arterial blood pressure in medication-free healthy men. Method: Thirty-two volunteers [age (years): 36.06±7.36, BMI: 27.60±4.63 (kg/m 2)] underwent assessments of body height/weight, % fat mass, fat-free mass (kg), blood pressure, and a subcutaneous adipose tissue biopsy via a surgical technique. Results: We found that NPRA mRNA was negatively associated with % fat mass (r=-0.40, R 2=0.16, p=0.03) and BMI (r=-0.45, R 2=0.20, p=0.01). Cohen's f 2 effect size analyses showed a small effect size between NPRA mRNA and BMI ( f 2 =0.25). One-way analysis of variance with Bonferroni post-hoc tests showed a tendency for mean differences of NPRA mRNA across BMI categories (p=0.06). This was confirmed by Cohen's d effect size analyses revealing a large effect size of NPRA mRNA between obese individuals (BMI≥30 kg/m 2) and either normal weight (BMI=19-25 kg/m 2; d=0.94) or overweight (BMI=25-30 kg/m 2; d=1.12) individuals. Conclusions: NPRA mRNA is negatively associated with % fat mass and BMI in medication-free healthy men, suggesting a possible role of NPRA in the control of fat mass accumulation.


Life without oxygen: gene regulatory responses of the crucian carp (Carassius carassius) heart subjected to chronic anoxia.
Crucian carp are unusual among vertebrates in surviving extended periods in the complete absence of molecular oxygen. During this time cardiac output is maintained though these mechanisms are not well understood. Using a high-density cDNA microarray, we have defined the genome-wide gene expression responses of cardiac tissue after exposing the fish at two temperatures (8 and 13 °C) to one and seven days of anoxia, followed by seven days after restoration to normoxia. At 8 °C, using a false discovery rate of 5%, neither anoxia nor re-oxygenation elicited appreciable changes in gene expression. By contrast, at 13 °C, 777 unique genes responded strongly. Up-regulated genes included those involved in protein turnover, the pentose phosphate pathway and cell morphogenesis while down-regulated gene categories included RNA splicing and transcription. Most genes were affected between one and seven days of anoxia, indicating gene regulation over the medium term but with few early response genes. Re-oxygenation for 7 days was sufficient to completely reverse these responses. Glycolysis displayed more complex responses with anoxia up-regulated transcripts for the key regulatory enzymes, hexokinase and phosphofructokinase, but with down-regulation of most of the non-regulatory genes. This complex pattern of responses in genomic transcription patterns indicates divergent cardiac responses to anoxia, with the transcriptionally driven reprogramming of cardiac function seen at 13 °C being largely completed at 8 °C.


Quantitative assessment of prolonged metabolic abnormalities in reperfused canine myocardium.
Prolonged metabolic abnormalities have been demonstrated previously in postischemic myocardium, including relative increases in glucose uptake and abnormal fatty acid kinetics. However, quantitative metabolic information is limited, and the time course of changes in MVO2 in postischemic myocardium is unknown. To address these issues, chronically instrumented dogs were studied serially over 1 month after transient left anterior descending coronary artery (LAD) occlusion, using positron emission tomography. Dynamic imaging protocols were used in conjunction with tracer kinetic models to quantify blood flow and metabolic rates. Myocardial sectors were defined as normal, predominantly reversibly injured, and infarct-containing, based on occlusion blood flow images and postmortem histochemistry. Myocardial blood flow and metabolism were homogeneous at baseline. During LAD occlusion for 3 hours, myocardial blood flow in reversibly injured and infarct-containing sectors (determined with 13NH3) was decreased to 46% and 23%, respectively, of blood flow in normal tissue. MVO2, determined with [1-11C]acetate, was decreased less than myocardial blood flow, consistent with increased oxygen extraction in the ischemic tissue. After reperfusion, blood flow normalized rapidly in reversibly injured tissue but remained depressed in infarct-containing sectors. Regional myocardial function, assessed by two-dimensional echocardiography, was severely depressed during occlusion and did not improve significantly until 1 week after reperfusion. MVO2 remained depressed after reperfusion in both reversibly injured and infarct-containing sectors, did not improve from occlusion levels until 1 week after reperfusion, and remained significantly depressed 1 month after reperfusion even in reversibly injured sectors; [1-11C]palmitate kinetics were also abnormal in postischemic tissue. As reported previously, glucose metabolic rates were increased relative to baseline in normal but not in postischemic tissue 3 hours after reperfusion. Subsequently, glucose metabolism tended to be higher in postischemic relative to normal myocardium. The results demonstrate decreased oxidative metabolism in postischemic tissue, with concomitant abnormalities in palmitate kinetics and glucose metabolism. Oxidative metabolism and regional function demonstrated a parallel recovery with time.


Cellular approaches to the interaction between cannabinoid receptor ligands and nicotinic acetylcholine receptors.
Cannabinoids are among the earliest known drugs to humanity. Cannabis plant contains various phytochemicals that bind to cannabinoid receptors. In addition, synthetic and endogenously produced cannabinoids (endocannabinoids) constitute other classes of cannabinoid receptor ligands. Although many pharmacological effects of these cannabinoids are mediated by the activation of cannabinoid receptors, recent studies indicate that cannabinoids also modulate the functions of various integral membrane proteins including ion channels, receptors, neurotransmitter transporters, and enzymes by mechanism(s) not involving the activation of known cannabinoid receptors. Currently, the mechanisms of these effects were not fully understood. However, it is likely that direct actions of cannabinoids are closely linked to their lipophilic structures. This report will focus on the actions of cannabinoids on nicotinic acetylcholine receptors and will examine the results of recent studies in this field. In addition some mechanistic approaches will be provided. The results discussed in this review indicate that, besides cannabinoid receptors, further molecular targets for cannabinoids exist and that these targets may represent important novel sites to alter neuronal excitability.


Wrapping of nanoparticles by the cell membrane: the role of interactions between the nanoparticles.
A fundamental understanding of the interactions between nanoparticles (NPs) and the cell membrane is essential to improve the performance of the NP-based biomedical applications and assess the potential toxicity of NPs. Despite the great progress in understanding the interaction between individual NP and the membrane, little is known about the interaction between multiple NPs and the membrane. In this work, we investigate the wrapping of two parallel elongated NPs by the membrane, taking the NP-NP electrostatic interaction and van der Waals (vdW) interaction into consideration. Three types of NPs, namely the rigid NPs with circular and elliptic cross-sections and the deformable NPs, are systematically investigated. The results show that the electrostatic interaction would enhance the tendency of the independent wrapping and inhibit the rotation of the elongated and equally charged NPs with elliptic cross-sections. Under the vdW interaction, the competition of the NP-NP adhesion and the membrane elastic energies with the NP-membrane adhesion energy leads the NPs to be wrapped cooperatively or independently. For the system with elongated NPs with elliptic cross-sections, the NPs are more likely to be wrapped independently as the shapes become more anisotropic and the NPs would rotate to contact each other with the flat sides in the cooperative wrapping configuration. Moreover, the soft NPs are more likely to be wrapped cooperatively compared with the stiff NPs. These results may provide guidelines to control the internalization pathway of NPs and improve the efficiency of NP-based drug delivery systems.


BMP1-like proteinases are essential to the structure and wound healing of skin.
Closely related extracellular metalloproteinases bone morphogenetic protein 1 (BMP1) and mammalian Tolloid-like 1 (mTLL1) are co-expressed in various tissues and have been suggested to have overlapping roles in the biosynthetic processing of extracellular matrix components. Early lethality of mice null for the BMP1 gene Bmp1 or the mTLL1 gene Tll1 has impaired in vivo studies of these proteinases. To overcome issues of early lethality and functional redundancy we developed the novel BTKO mouse strain, with floxed Bmp1 and Tll1 alleles, for induction of postnatal, simultaneous ablation of the two genes. We previously showed these mice to have a skeletal phenotype that includes elements of osteogenesis imperfecta (OI), osteomalacia, and deficient osteocyte maturation, observations validated by the finding of BMP1 mutations in a subset of human patients with OI-like phenotypes. However, the roles of BMP1-like proteinase in non-skeletal tissues have yet to be explored, despite the supposed importance of putative substrates of these proteinases in such tissues. Here, we employ BTKO mice to investigate potential roles for these proteinases in skin. Loss of BMP1-like proteinase activity is shown to result in markedly thinned and fragile skin with unusually densely packed collagen fibrils and delayed wound healing. We demonstrate deficits in the processing of collagens I and III, decorin, biglycan, and laminin 332 in skin, which indicate mechanisms whereby BMP1-like proteinases affect the biology of this tissue. In contrast, lack of effects on collagen VII processing or deposition indicates this putative substrate to be biosynthetically processed by non-BMP1-like proteinases.


Interleukin-30 Promotes Breast Cancer Growth and Progression.
The inflammatory tissue microenvironment that promotes the development of breast cancer is not fully understood. Here we report a role for elevated IL30 in supporting the breast cancer cell viability and invasive migration. IL30 was absent in normal mammary ducts, ductules, and acini of histologically normal breast and scanty in the few stromal infiltrating leukocytes. In contrast, IL30 was expressed frequently in breast cancer specimens where it was associated with triple-negative and HER2+ molecular subtypes. In stromal leukocytes found in primary tumors or tumor-draining lymph nodes, which included mainly CD14+ monocytes, CD68+ macrophages, and CD33+/CD11b+ myeloid cells, IL30 levels increased with disease stage and correlated with recurrence. A negative correlation was determined between IL30 expression by nodal stromal leukocytes and overall survival. In vitro studies showed that human recombinant IL30 upregulated expression of a pro-oncogenic program, including especially IL6 in both triple-negative and HER2+ breast cancer cells. In triple-negative breast cancer cells, IL30 boosted a broader program of proliferation, invasive migration, and an inflammatory milieu associated with KISS1-dependent metastasis. Silencing of STAT1/STAT3 signaling hindered the regulation of the primary growth and progression factors in breast cancer cells. IL30 administration in vivo fostered the growth of triple-negative breast cancer by promoting proliferation and vascular dissemination of cancer cells and the accumulation of intratumoral CD11b+/Gr1+ myeloid cell infiltrates. Overall, our results show how IL30 regulates breast cancer cell viability, migration, and gene expression to promote breast cancer growth and progression and its impact on patient outcome. Cancer Res; 76(21); 6218-29. ©2016 AACR.


Pessary use in stress urinary incontinence: a review of advantages, complications, patient satisfaction, and quality of life.
Stress urinary incontinnce (SUI) is a common condition among women. The usual approach to treatment of SUI is a stepwise plan from conservative to surgical procedures. A vaginal pessary is one of the commonly used conservative treatments that offer symptomatic improvement for women with incontinence. This review provides a critical analysis of the benefits and shortcomings offered by vaginal pessaries to patients affected by SUI, with a particular focus on indications, advantages, quality of life, patient satisfaction, and potential complications. To obtain the required information, an extensive search of PubMed and Cochrane databases was performed, covering the time frame from January 2000 to December 2016. We also surveyed the published guidelines of American Urological Association, Canadian Urological Association, American Urogynecological Society, National Institutes of Health (USA), and National Institute for Health and Care Excellence (UK). A total of 192 original research papers, review articles, and clinical trials were identified. The analysis of retrieved data provides evidence that vaginal pessaries constitute an effective nonsurgical option for SUI. The satisfaction rate with pessary use is high and only minor complications, if any, occur, vaginal discharge being the most common. The reviewed studies document that vaginal pessaries provide an adequate control of SUI if they are fit properly and managed by frequent replacements and regular checkups. They should be considered among the first line of treatment for SUI associated with exercise and increased intra-abdominal pressure.


Re-evaluation of Sepharose-insulin as a tool for the study of insulin action.
The biological activity of Sepharose-insulin in different assays in vitro, e.g., stimulation of glucose oxidation, lipogenesis, and antilipolysis and activation of pyruvate dehydrogenase (EC 188.8.131.52) activity, has been investigated. According to amino acid analysis, between 270 and 330 mug (6.9-8.2 U) of insulin were coupled per ml of packed beads. Related to the total insulin content, 0.2-0.7% of the insulin was biologically active. Comparable biological activity was observed with isolated fat cells and fat pad pieces. After incubation with tissue or cells, Sepharose-insulin particles were separated by centrifugation from the medium. The clear supernatant was assayed for biologically and immunologically reactive insulin and contained soluble insulin activity. A quantitative evaluation of the soluble biological and immunological insulin activity in the supernatant accounted for the total insulin activity of Sepharose-insulin.


Transplanted and repopulated retinal pigment epithelial cells on damaged Bruch's membrane in rabbits.
The authors studied how artificially damaged Bruch's membrane influenced growth and differentiation of transplanted embryonic retinal pigment epithelial (RPE) cells and of host RPE cells in rabbits. Embryonic RPE cells obtained from pigmented rabbits were transplanted into the subretinal space of adult albino rabbits. The host RPE was removed with a silicone cannula, and Bruch's membrane was damaged by scratching with a microhooked 27 gauge needle under the detached retina in closed vitrectomy. The transplantation sites were examined 3, 7, and 14 days after surgery by light and electron microscopy. Varying degrees of damage in Bruch's membrane were observed. Pigmented and hypopigmented RPE cells showed a normal polarity and tight junctions were seen at the sites of mild to moderate damage 3-7 days after the surgery. In contrast, fibroblast-like cells with no such features of RPE cells formed multiple layers at the sites of severe damage involving the full thickness of Bruch's membrane and the choriocapillaris even 14 days after the surgery. Without transplantation, host RPE cells repopulated the damaged areas in the same way as transplanted RPE cells. Transplanted embryonic RPE cells as well as host RPE cells grew and differentiated on the moderately damaged Bruch's membrane, while the severely damaged Bruch's membrane did not allow differentiation of RPE cells although these cells could grow and cover the damaged areas.


Techniques for Nonterminal Blood Sampling in Black-Tailed Prairie Dogs (Cynomys ludovicianus).
Black-tailed prairie dogs (Cynomys ludovicianus) are used as an animal model for research on gallbladder stones and several infectious diseases. A comprehensive, instructive resource regarding the appropriate techniques for venipuncture and collection of nonterminal blood samples in this species has not yet been published. Blood samples (1 mL or larger) were readily obtained from the jugular vein, femoral vein, or cranial vena cava, whereas peripheral sites, such as the cephalic vein, saphenous vein, and tarsal vein, mainly were useful for obtaining smaller volumes. The detailed and illustrated information presented here can aid clinicians and researchers in performing venipuncture, anesthesia, and handling of this species.


[Cytokines in bone diseases. Genetic defects of PTH/PTHrP receptor in chondrodysplasia].
Parathyroid hormone-related protein (PTHrP) signaling plays important roles in regulating the differentiation of chondrocytes in endochondral bone development. PTHrP signaling functions as an inhibitory effect on chondrocyte hypertrophy which is a terminal stage of differentiation at a growth plate. Mutations of the PTH÷PTHrP receptor have been identified in Jansen metaphyseal chondrodysplasia, Blomstrand's lethal chondrodysplasia, and enchondromatosis. Furthermore, genetic manipulations of the PTHrP and its receptor genes in mice have demonstrated the critical roles of these proteins in regulating both the switch between proliferation and differentiation of chondrocytes.


Payments for care at private for-profit and private not-for-profit hospitals: a systematic review and meta-analysis.
It has been shown that patients cared for at private for-profit hospitals have higher risk-adjusted mortality rates than those cared for at private not-for-profit hospitals. Uncertainty remains, however, about the economic implications of these forms of health care delivery. Since some policy-makers might still consider for-profit health care if expenditure savings were sufficiently large, we undertook a systematic review and meta-analysis to compare payments for care at private for-profit and private not-for-profit hospitals. We used 6 search strategies to identify published and unpublished observational studies that directly compared the payments for care at private for-profit and private not-for-profit hospitals. We masked the study results before teams of 2 reviewers independently evaluated the eligibility of all studies. We confirmed data or obtained additional data from all but 1 author. For each study, we calculated the payments for care at private for-profit hospitals relative to private not-for-profit hospitals and pooled the results using a random effects model. Eight observational studies, involving more than 350 000 patients altogether and a median of 324 hospitals each, fulfilled our eligibility criteria. In 5 of 6 studies showing higher payments for care at private for-profit hospitals, the difference was statistically significant; in 1 of 2 studies showing higher payments for care at private not-for-profit hospitals, the difference was statistically significant. The pooled estimate demonstrated that private for-profit hospitals were associated with higher payments for care (relative payments for care 1.19, 95% confidence interval 1.07-1.33, p = 0.001). Private for-profit hospitals result in higher payments for care than private not-for-profit hospitals. Evidence strongly supports a policy of not-for-profit health care delivery at the hospital level.


Epidemiology of rotavirus and astrovirus infections in children in northwestern Nigeria.
Recent estimates attribute 527,000 deaths in children less than five years of age to rotavirus diarrhea annually, with 145,000 occurring in sub-Saharan Africa. Human astroviruses have been identified as one of the most frequent causes of infantile diarrhea, second in incidence only to rotavirus. This study was conducted to determine the prevalence of rotavirus and astrovirus and also to establish the circulating strains of rotavirus in a community in Nigeria where most diarrheic patients do not visit clinics or health care centers. A total of 154 stool samples (134 diarrheic and 20 non-diarrheic) were collected from infants and young children less than 5 years of age from January-March 2002. Samples were obtained by house-to-house visit in randomly selected districts in Zaria, Northwestern Nigeria. The samples were screened for rotavirus and astrovirus antigens using commercially available Enzyme Linked Immunosorbent Assay (ELISA) kits. All positive group A rotavirus samples were further subjected to VP6 sub-group ELISA, Polyacrylamide gel electrophoresis (PAGE) to determine their RNA electropherotypes and Reverse transcription polymerase chain reaction (RT-PCR) to determine their VP7 and VP4 genotypes. Rotavirus and astrovirus antigens were detected in 9% (12) and 5% (7) of the 134 diarrheic stool samples respectively. No viral antigen was detected in the non-diarrheic stools. Rotavirus infection was more common in younger children than astrovirus infection. VP6 sub-group II specificity (58.3%), long RNA electropherotypes (41.6%), VP7 genotype G1 (33.3%) and VP4 genotype P [6] (33.3%) were the most common strains in circulation at that time in the community. Of significance is the fact that a large proportion of the rotavirus strains in circulation could not be assigned either a VP6 subgroup or RNA electrophoretic pattern probably as a result of low viral load. In this community-based study, rotavirus and astrovirus were significantly associated with diarrhea. However, the prevalence of rotavirus infection among children appears to be low while that of astrovirus falls in the range seen in hospital-based studies around the continent.


AN UPDATED DEBARCODING TOOL FOR MASS CYTOMETRY WITH CELL TYPE-SPECIFIC AND CELL SAMPLE-SPECIFIC STRINGENCY ADJUSTMENT.
Pooled sample analysis by mass cytometry barcoding carries many advantages: reduced antibody consumption, increased sample throughput, removal of cell doublets, reduction of cross-contamination by sample carryover, and the elimination of tube-to-tube-variability in antibody staining. A single-cell debarcoding algorithm was previously developed to improve the accuracy and yield of sample deconvolution, but this method was limited to using fixed parameters for debarcoding stringency filtering, which could introduce cell-specific or sample-specific bias to cell yield in scenarios where barcode staining intensity and variance are not uniform across the pooled samples. To address this issue, we have updated the algorithm to output debarcoding parameters for every cell in the sample-assigned FCS files, which allows for visualization and analysis of these parameters via flow cytometry analysis software. This strategy can be used to detect cell type-specific and sample-specific effects on the underlying cell data that arise during the debarcoding process. An additional benefit to this strategy is the decoupling of barcode stringency filtering from the debarcoding and sample assignment process. This is accomplished by removing the stringency filters during sample assignment, and then filtering after the fact with 1- and 2-dimensional gating on the debarcoding parameters which are output with the FCS files. These data exploration strategies serve as an important quality check for barcoded mass cytometry datasets, and allow cell type and sample-specific stringency adjustment that can remove bias in cell yield introduced during the debarcoding process.


Vegetables, fruit and phytoestrogens as preventive agents.
The practice of medicine-both past and present-often involves the prescription of specific foods (almost always plants) or their potent derivatives, to treat a wide spectrum of illnesses. Foods that have been ascribed healing properties include the Cruciferae, the allium family, celery, cucumber, endive, parsley, radish and legumes. Review of the epidemiological data, including both cohort and case-control studies, of all cancer sites strongly suggests that plant foods also have preventive potential and that consumption of the following groups and types of vegetables and fruits is lower in those who subsequently develop cancer: raw and fresh vegetables, leafy green vegetables, Cruciferae, carrots, broccoli, cabbage, lettuce, and raw and fresh fruit (including tomatoes and citrus fruit). Other data suggest that foods high in phytoestrogens, particularly soy (which contains isoflavones), or high in precursor compounds that can be metabolized by gut bacteria into active agents, particularly some grains and vegetables with woody stems (which contain precursors to lignans), are plausibly associated with a lower risk of sex-hormone-related cancers. The human evidence for these latter associations is not strong. There are many biologically plausible reasons why consumption of plant foods might slow or prevent the appearance of cancer. These include the presence in plant foods of such potentially anticarcinogenic substances as carotenoids, vitamin C, vitamin E, selenium, dietary fibre (and its components), dithiolthiones, isothiocyanates, indoles, phenols, protease inhibitors, allium compounds, plant sterols, and limonene. Phytoestrogens are also derived from some vegetables and berries as well as grains and seeds. Most of the data for the observations on the anticarcinogenic potential of all of these compounds have come from animal and in vitro studies. At almost every one of the stages of the cancer process, identified phytochemicals are known to be able to alter the likelihood of carcinogenesis-occasionally in a way that enhances risk but usually in a favourable direction. For example, glucosinolates and indoles, thiocyanates and isothiocyanates, phenols, and coumarins can induce a multiplicity of phase II (solubilizing and usually inactivating) enzymes; ascorbate and phenols block the formation of carcinogens such as nitrosamines; flavonoids and carotenoids act as antioxidants, essentially disabling the carcinogenic potential of specific compounds; lipid-soluble compounds such as carotenoids and sterols may alter membrane structure or integrity; some sulphur-containing compounds suppress DNA and protein synthesis; carotenoids can suppress DNA synthesis and enhance differentiation; and phytoestrogens compete with estradiol for estrogen receptors in a way that is generally antiproliferative. Consumption of diets low in plant foods results in a reduced intake of a wide variety of those substances that can plausibly lower cancer risk. In the presence of a diet and lifestyle high in potential carcinogens (whether derived from fungal contamination, cooking or tobacco) or high in promoters (such as salt and alcohol), overall risk of cancer at many epithelial sites is elevated. Plant foods appear to exert a general risk-lowering effect; the patterns of exposure to cancer initiators and promoters and of genetic susceptibility may determine the variations in the site-specific risks of cancer seen across populations.


Matrilin-3 induction of IL-1 receptor antagonist is required for up-regulating collagen II and aggrecan and down-regulating ADAMTS-5 gene expression.
Deletion or mutation of the gene encoding the cartilage extracellular matrix (ECM) protein matrilin-3 (MATN3) results in the early onset of osteoarthritis (OA), suggesting chondroprotective properties of MATN3. To understand the mechanisms underlying these properties, we determined the effects of MATN3 protein on the expression of several key anabolic and catabolic genes involved in chondrocyte homeostasis, and the dependence of such regulation on the anti-inflammatory cytokine: IL-1 receptor antagonist (IL-1Ra). The effects of recombinant human (rh) MATN3 protein were examined in C28/I2 immortalized human chondrocytes, primary human chondrocytes (PHCs), and primary mouse chondrocytes (PMCs). Messenger RNA levels of IL-1Ra, COL2A1, ACAN, MMP-13, and ADAMTS-4 and -5 were determined using real-time RT-PCR. Knocking down IL-1Ra was achieved by siRNA gene silencing. IL-1Ra protein levels were quantified by ELISA and the Bio-Plex Suspension Array System. COL2A1 protein level was quantified using Western blot analysis. Statistic analysis was done using the two-tailed t-test or one-way ANOVA. rhMATN3 protein induced gene expression of IL-1Ra in C28/I2 cells, PHCs, and PMCs in a dose- and time-dependent manner. Treatment of C28/I2 cells and PHCs with MATN3 protein stimulated gene expression of COL2A1 and ACAN. Conversely, mRNA levels of COL2A1 and ACAN were decreased in MATN3 KO mice. MATN3 protein treatment inhibited IL-1β-induced MMP-13, ADAMTS-4 and ADAMTS-5 in C28/I2 cells and PHCs. Knocking down IL-1Ra abolished the MATN3-mediated stimulation of COL2A1 and ACAN and inhibition of ADAMTS-5, but had no effect on MATN3 inhibition of MMP-13 mRNA. Our findings point to a novel regulatory role of MATN3 in cartilage homeostasis due to its capacity to induce IL-1Ra, to upregulate gene expression of the major cartilage matrix components, and to downregulate the expression of OA-associated matrix-degrading proteinases in chondrocytes. The chondroprotective properties of endogenous MATN3 depend partly on its induction of IL-1Ra. Our findings raise a possibility to use rhMATN3 protein for anti-inflammatory and chondroprotective therapy.


Porcine heterotopic composite tissue allograft transplantation using a large animal model for preclinical studies.
Composite tissue allograft (CTA) transplantation is currently limited by the risks of side effects resulting from long-term high-dose immunosuppression. Therefore, preclinical animal models are essential to help CTA transplantation advance into clinical reality. Evidence has shown that small-animal model (rodents) immunotherapy protocols cannot be directly applied to humans. This study investigated whether a miniature porcine model is reproducible for preclinical studies. Based on the concept of vascularized skeletal tissue allograft transplantation, limb heterotopic allograft tissue from a mismatched donor miniature pig consisting of the distal femur, knee joint, tibia, fibula, and surrounding muscle with a vascularized skin paddle model supplied by the superficial femoral vessels was transplanted into recipient pigs. Swine viability and rejection signs of the allograft were monitored postoperatively. Histopathological changes in the allograft tissues were examined using hematoxylin and eosin staining if the allo-skin flap was rejected. The recipient pigs were ambulatory immediately following surgery. The flaps showed no visible signs of rejection over the first 4 days of observation. The skin flaps appeared bluish-purple and edematous on postoperative days 5 approximately 7, and progressed to tissue necrosis and rejection on postoperative days 8 approximately 13. Histological examination revealed marked mononuclear cell infiltration and necrotic changes in the all rejected tissues, especial in the allograft skin tissues (skin > muscle > bone > cartilage). The results showed this the porcine CTA model is reproducible and suitable for preclinical training for human CTA transplantation. Monitoring of the allo-skin flap is a useful strategy to evaluate composite tissue allograft rejection.


Predicting CD4 lymphocyte count <200 cells/mm(3) in an HIV type 1-infected African population.
Clinical criteria are recommended to select HIV-infected patients for initiation of antiretroviral therapy when CD4 lymphocyte testing is unavailable. We evaluated the performance characteristics of WHO staging criteria, anthropometrics, and simple laboratory measurements for predicting CD4 lymphocyte count (CD4 count) <200 cells/mm(3) among HIV-infected patients in Tanzania. A total of 202 adults, diagnosed with HIV infection through community-based testing, underwent a detailed evaluation including staging history and examination, anthropometry, complete blood count, erythrocyte sedimentation rate (ESR), and CD4 count. Univariable analysis and recursive partitioning were used to identify characteristics associated with CD4 count 200 cells/mm(3). Of 202 participants 109 (54%) had a CD4 count <200 cells/mm(3). Characteristics most strongly associated with CD4 count <200 cells/mm(3) (p-value <0.0001) were the presence of mucocutaneous manifestations (72% vs. 28%), lower total lymphocyte count (TLC) (median 1,450 vs. 2,200 cells/mm(3)), lower total white blood cell count (median 4,200 vs. 5,500 cells/mm(3)), and higher ESR (median 95 vs. 53 mm/h). In a partition tree model, TLC <1,200 cells/mm(3), ESR >or=120 mm/h, or the presence of mucocutaneous manifestations yielded a sensitivity of 0.85 and specificity of 0.63 for predicting CD4 count <200 cells/mm(3). The sensitivity of the 2006 WHO Staging system improved from 0.75 to 0.93 with inclusion of these parameters, at the expense of specificity (0.36 to 0.26). The presence of mucocutaneous manifestations, TLC <1,200 cells/mm(3), or ESR >or=120 mm/h was a strong predictor of CD4 count <200 cells/mm(3) and enhanced the sensitivity of the 2006 WHO staging criteria for identifying patients likely to benefit from antiretrovirals.