Layer-by-layer assembled fluorescent probes in the second near-infrared window for systemic delivery and detection of ovarian cancer.
Fluorescence imaging in the second near-infrared window (NIR-II, 1,000-1,700 nm) features deep tissue penetration, reduced tissue scattering, and diminishing tissue autofluorescence. Here, NIR-II fluorescent probes, including down-conversion nanoparticles, quantum dots, single-walled carbon nanotubes, and organic dyes, are constructed into biocompatible nanoparticles using the layer-by-layer (LbL) platform due to its modular and versatile nature. The LbL platform has previously been demonstrated to enable incorporation of diagnostic agents, drugs, and nucleic acids such as siRNA while providing enhanced blood plasma half-life and tumor targeting. This work carries out head-to-head comparisons of currently available NIR-II probes with identical LbL coatings with regard to their biodistribution, pharmacokinetics, and toxicities. Overall, rare-earth-based down-conversion nanoparticles demonstrate optimal biological and optical performance and are evaluated as a diagnostic probe for high-grade serous ovarian cancer, typically diagnosed at late stage. Successful detection of orthotopic ovarian tumors is achieved by in vivo NIR-II imaging and confirmed by ex vivo microscopic imaging. Collectively, these results indicate that LbL-based NIR-II probes can serve as a promising theranostic platform to effectively and noninvasively monitor the progression and treatment of serous ovarian cancer.


Exosomes Derived from Human Primary and Metastatic Colorectal Cancer Cells Contribute to Functional Heterogeneity of Activated Fibroblasts by Reprogramming Their Proteome.
Cancer-associated fibroblasts (CAFs) are a heterogeneous population of activated fibroblasts that constitute a dominant cellular component of the tumor microenvironment (TME) performing distinct functions. Here, the role of tumor-derived exosomes (Exos) in activating quiescent fibroblasts into distinct functional subtypes is investigated. Proteomic profiling and functional dissection reveal that early- (SW480) and late-stage (SW620) colorectal cancer (CRC) cell-derived Exos both activated normal quiescent fibroblasts (α-SMA- , CAV+ , FAP+ , VIM+ ) into CAF-like fibroblasts (α-SMA+ , CAV- , FAP+ , VIM+ ). Fibroblasts activated by early-stage cancer-exosomes (SW480-Exos) are highly pro-proliferative and pro-angiogenic and display elevated expression of pro-angiogenic (IL8, RAB10, NDRG1) and pro-proliferative (SA1008, FFPS) proteins. In contrast, fibroblasts activated by late-stage cancer-exosomes (SW620-Exos) display a striking ability to invade through extracellular matrix through upregulation of pro-invasive regulators of membrane protrusion (PDLIM1, MYO1B) and matrix-remodeling proteins (MMP11, EMMPRIN, ADAM10). Conserved features of Exos-mediated fibroblast activation include enhanced ECM secretion (COL1A1, Tenascin-C/X), oncogenic transformation, and metabolic reprogramming (downregulation of CAV-1, upregulation of glycogen metabolism (GAA), amino acid biosynthesis (SHMT2, IDH2) and membrane transporters of glucose (GLUT1), lactate (MCT4), and amino acids (SLC1A5/3A5)). This study highlights the role of primary and metastatic CRC tumor-derived Exos in generating phenotypically and functionally distinct subsets of CAFs that may facilitate tumor progression.


Ochratoxin A exerts neurotoxicity in human astrocytes through mitochondria-dependent apoptosis and intracellular calcium overload.
Astrocytes are the major glial cell type in the central nervous system (CNS), and the distal part of the astrocyte forms the blood-brain barrier with nearby blood vessels. They maintain the overall metabolism, growth, homeostasis of neurons, and signaling in the CNS. Ochratoxin A is considered a carcinogen and immunotoxic, nephrotoxic, and neurotoxic mycotoxin. Specifically, it exhibits neurotoxicity with high affinity for the brain. Despite some previous studies about the effects of ochratoxin A in glial cells, the intracellular working mechanism in astrocytes is not fully understood. In this study, we studied the specific working mechanism of ochratoxin A in the human astrocyte cell line, NHA-SV40LT. Ochratoxin A reduced cell proliferation with sub G0/G1 cell cycle arrest by inhibiting CCND1, CCNE1, CDK4, and MYC expression. It induced apoptosis of NHA-SV40LT cells through mitochondrial membrane potential (MMP) loss and up-regulation of BAX and TP53. In addition, ochratoxin A increased cytosolic and mitochondrial calcium levels, resulting in an increase in MMP2 and PLAUR mRNA expression in NHA-SV40LT cells. Furthermore, ochratoxin A regulated the phosphorylation of AKT, ERK1/2, and JNK signal molecules of human astrocytes. Collectively, ochratoxin A exerts neurotoxicity through anti-proliferation and mitochondria-dependent apoptosis in human astrocytes.


Growth rates and the prevalence and progression of scoliosis in short-statured children on Australian growth hormone treatment programmes.
STUDY DESIGN AND AIM: This was a longitudinal chart review of a diverse group (cohort) of patients undergoing HGH (Human Growth Hormone) treatment. Clinical and radiological examinations were performed with the aim to identify the presence and progression of scoliosis. 185 patients were recruited and a database incorporating the age at commencement, dose and frequency of growth hormone treatment and growth charts was compiled from their Medical Records. The presence of any known syndrome and the clinical presence of scoliosis were included for analysis. Subsequently, skeletally immature patients identified with scoliosis were followed up over a period of a minimum four years and the radiologic type, progression and severity (Cobb angle) of scoliosis were recorded. Four (3.6%) of the 109 with idiopathic short stature or hormone deficiency had idiopathic scoliosis (within normal limits for a control population) and scoliosis progression was not prospectively observed. 13 (28.8%) of 45 with Turner syndrome had scoliosis radiologically similar to idiopathic scoliosis. 11 (48%) of 23 with varying syndromes, had scoliosis. In the entire cohort, the growth rates of those with and without scoliosis were not statistically different and HGH treatment was not ceased because of progression of scoliosis. In this study, there was no evidence of HGH treatment being responsible for progression of scoliosis in a small number of non-syndromic patients (four). An incidental finding was that scoliosis, similar to the idiopathic type, appears to be more prevalent in Turner syndrome than previously believed.


Ethical issues in the economic assessment of health care technologies.
This paper challenges traditional views which oppose health economics and medical ethics by arguing that economic assessment is a necessary complement to medical ethics and can help to improve public participation and democratic processes in choices about resource allocation for health care technologies. In support of this argument, four points are emphasized: (1) Most current biomedical ethical debates implicitly deal with economic issues of resource allocation. (2) Clinical decisions, which usually respect the Hippocratic code of ethics, are nevertheless influenced by economic incentives and constraints. (3) Economic assessment is concerned with both efficiency and equity and potential trade-offs between the two, which means that ethical judgements are always embedded in welfare economics. (4) The real debate is not between economics on the one side and medical ethics on the other. Rather it is between different ethical conceptions of social justice and the contrasting approaches they entail to reconciling individual interests and preferences with collective goods and welfare.


The spatial relation of diabetic retinal neurodegeneration with diabetic retinopathy.
Diabetic retinal neurodegeneration (DRN) has been demonstrated in eyes of patients with diabetes mellitus (DM), even in the absence of diabetic retinopathy (DR). However, no studies have looked at the rate of change in retinal layers and presence/development of DR over time per quadrant of the macula. In this longitudinal study, we aimed to clarify whether the rate of DRN is associated with the development/presence of DR within 4 different quadrants of the retina. 80 eyes of 40 patients with type 1 DM and no/minimal DR were included. At 4 visits over 6 years, SD-OCT and fundus images were acquired. Thickness of the Retinal Nerve Fiber Layer (RNFL), Ganglion Cell Layer (GCL) and Inner Plexiform Layer (IPL) was measured in a 1-6mm circle around the fovea overall and for each quadrant (superior, nasal, inferior, temporal). Fundus images were scored for the presence/absence of DR in these areas. Multilevel analyses were performed to determine the rate of change for each layer overall and per quadrant for eyes/quadrants without and with DR during the follow-up period. RNFL and GCL showed significant thinning over time, IPL significant thickening. These changes were more pronounced for GCL and IPL in eyes/quadrants with DR during the follow-up period. RNFL and GCL both showed thinning over time, which was more pronounced in eyes with DR for GCL. This holds true even in regional parts of the retina, as quadrant analyses showed similar results, showing that structural DRN is associated with DR per quadrant independently.


High intensity focused ultrasound ablation of the vas deferens in a canine model.
High intensity focused ultrasound is an ablative technology capable of producing thermal coagulative necrosis of sub-surface structures without injuring intervening tissues. We assessed the feasibility of using high intensity focused ultrasound to produce occlusion of the canine vas deferens. A high intensity focused ultrasound transducer was incorporated into a hand held clip specially designed to grasp the vas deferens transcutaneously. Slots within the jaws of the clip ensured that the vas deferens and high intensity focused ultrasound target zone were properly co-located. We ablated 10 vasa using a range of power and time parameters. At 2 weeks after ablation each vas, epididymis and testis was surgically harvested en bloc. Retrograde vasography was performed to assess vasal occlusion, followed by pathological analysis. High intensity focused ultrasound occlusion of the vas deferens was confirmed in 4 specimens ablated with parameters at the upper end of the parameter range, 2 of the 2 ablated with 7 W. for 60 seconds and 2 of the 4 ablated with 7 W. for 30 seconds. Histological injury was noted in 8 of the 10 ablated specimens. Skin burns that developed over 4 of the targeted vasa were conservatively managed. Bilateral sham procedures in a control dog resulted in patent vasa and no associated skin burns. We demonstrated the feasibility of noninvasive, transcutaneous high intensity focused ultrasound occlusion of the vas deferens with ablation powers at the upper end of the tested range, that is 7 W. Modifications of the hand held clip and optimization of ablation parameters would likely improve the success rate of this procedure. Refinement of this technology may provide a rapid noninvasive alternative to conventional vasectomy.


Effect of propranolol on beta-adrenoceptors in rat hearts.
The question of whether the chronic administration of propranolol modifies the numbers or properties of cardiac beta-adrenoceptors was examined because of many reports suggestive of cardiac hypersensitivity following the withdrawal of beta-antagonists. In three studies, rats were given (+/-)-propranolol, 30 mg/day, orally or intraperitoneally, for 1 to 7 weeks. The numbers and affinities of specific binding sites for radioactive dihydroalprenolol in whole hearts from 60 control and 75 test animals were found to be the same. Thus catecholamine deprivation does not exert an important regulatory effect on most beta-receptors in the heart, as it does in the brain.


A model for the analysis of neighbor finding in pointer-based quadtrees.
A natural byproduct of the tree-like nature of the quadtree is that many basic image processing operations can be implemented as tree traversals which differ in the nature of the computation that is performed at each node. Some of these computations involve the inspection of a node's adjacent neighbors (termed neighbor finding). A new model is developed for images represented by quadtrees, and it is used to analyze various neighbor-finding techniques. The model's predicted costs for neighbor finding correlate very closely with empirical results and it is superior to the model that was used previously.


Learning the Structure of Mixed Graphical Models.
We consider the problem of learning the structure of a pairwise graphical model over continuous and discrete variables. We present a new pairwise model for graphical models with both continuous and discrete variables that is amenable to structure learning. In previous work, authors have considered structure learning of Gaussian graphical models and structure learning of discrete models. Our approach is a natural generalization of these two lines of work to the mixed case. The penalization scheme involves a novel symmetric use of the group-lasso norm and follows naturally from a particular parametrization of the model. Supplementary materials for this paper are available online.


Radiographic and functional results of osteosynthesis using the proximal femoral nail antirotation (PFNA) in the treatment of unstable intertrochanteric femoral fractures.
We evaluated the radiographic and functional results of the proximal femoral nail antirotation (PFNA) system in patients with unstable intertrochanteric femoral fractures. The study included 45 patients (25 women, 20 men; mean age 72 years; range 27 to 97 years) who underwent osteosynthesis using the PFNA for unstable intertrochanteric femoral fractures. The fractures were in the right hip in 25 patients, and in the left hip in 20 patients. The fractures were classified according to the AO system. One patient had an open fracture due to firearm injury (Gustilo-Anderson 3A). The patients underwent surgery within a mean of eight days (range 2 to 21 days) from injury. The mean hospital stay was 13.5 days (range 4 to 25 days). Closed reduction was achieved in all the patients. The results were assessed clinically and radiographically. The neck-shaft angle of the femur (collodiaphysial angle) and the tip-apex distance were measured. The position of the helical screw within the femoral head was determined using the method of Cleveland and Bosworth. Clinical evaluation was made using the Harris hip score. Perioperative and postoperative complications were recorded. The mean follow-up period was 17.3 months (range 6 to 23 months). The mean operation time was 37.8 min (range 22 to 118 min) and the mean blood loss was 225 ml (range 150 to 450 ml). During surgery, femoral shaft fracture occurred in three patients, and greater trochanter fracture occurred in nine patients. Union was obtained in all the patients. Reduction was poor in four patients (8.9%), acceptable in seven patients (15.6%), and good in 34 patients (75.6%). The mean collodiaphysial angle was 136.7 degrees (range 125 degrees to 148 degrees). The tip-apex distance was <25 mm in 36 patients (80%), and =or>25 mm in nine patients (20%). The position of the helical screw in the femoral head was appropriate in 38 patients (84.4%). Postoperative complications included secondary varus (n=2, 4.4%), calcification at the tip of the greater trochanter (n=7, 15.5%), sensitivity over the fascia lata (n=7), medial thigh pain (n=11, 24.4%), and screw cut-out (n=1, 2.2%). Nine patients developed femoral shortness (mean 9.4 mm; range 8 to 13 mm). Screws showed lateral displacement in five patients (11.1%), which was less than 5 mm in four patients. Secondary surgery was required in four patients (8.9%). The mean Harris hip score was 77.8. Harris hip scores were very good in 11 patients (24.4%), good in 19 patients (42.2%), moderate in nine patients (20%), and poor in six patients (13.3%). Due to advantages of high union rate, early postoperative mobilization, and short operation time, PFNA osteosynthesis is the method of choice for surgical treatment of unstable intertrochanteric femoral fractures..


[Unusual evolution of reactive arthritis after endovesical immunotherapy with BCG].
Intravesical instillation of BCG (Bacille Calmette-Guerin) is a therapy prepared with weakened strains of Mycobacterium bovis and is an effective complementary treatment for high-risk (non-musculoinfiltrating or non-invasive) bladder cancer. Although its safety for clinical use is high, endovesical immunotherapy is not without complications. Arthralgia and arthritis are infrequent, but potentially severe, complications, the early diagnosis of which can allow adequate medical treatment and avoid chronification of the pathology. This case shows the therapeutic management of hip osteoarthritis in a 59-year-old woman, a rare and serious complication, secondary to a rare pathology such as reactive arthritis due to BCG instillation.


Milestones: Critical Elements in Clinical Informatics Fellowship Programs.
Milestones refer to points along a continuum of a competency from novice to expert. Resident and fellow assessment and program evaluation processes adopted by the ACGME include the mandate that programs report the educational progress of residents and fellows twice annually utilizing Milestones developed by a specialty specific ACGME working group of experts. Milestones in clinical training programs are largely unmapped to specific assessment tools. Residents and fellows are mainly assessed using locally derived assessment instruments. These assessments are then reviewed by the Clinical Competency Committee which assigns and reports trainee ratings using the specialty specific reporting Milestones. The challenge and opportunity facing the nascent specialty of Clinical Informatics is how to optimally utilize this framework across a growing number of accredited fellowships. The authors review how a mapped milestone framework, in which each required sub-competency is mapped to a single milestone assessment grid, can enable the use of milestones for multiple uses including individualized learning plans, fellow assessments, and program evaluation. Furthermore, such a mapped strategy will foster the ability to compare fellow progress within and between Clinical Informatics Fellowships in a structured and reliable fashion. Clinical Informatics currently has far less variability across programs and thus could easily utilize a more tightly defined set of milestones with a clear mapping to sub-competencies. This approach would enable greater standardization of assessment instruments and processes across programs while allowing for variability in how those sub-competencies are taught. A mapped strategy for Milestones offers significant advantages for Clinical Informatics programs.


On the genesis of narcissistic and phobic character formation in an adult patient: a developmental view.
A case has been presented in which early maternal failures reconstructed from separation-individuation subphase developments revealed a narcissistic alliance between the child and her parents. This alliance stood in the way of separation and individuation processes and blocked the expression of hostility. Identifications with the phobic mother prevented the ego from developing a signal function for anxiety. A maternal father, who in many ways counteracted the deficiencies of the mother, could only partially become the principal love object during the phallic and oedipal phases. A strong bisexual orientation prevented the full emergence of a feminine gender identity. Only after the narcissistic alliance emerged in transference and was given no gratification did phobic symptoms appear. The transference recapitulated a segment of the infantile neurosis in which phobia-proneness was held in check by the 'narcissistic alliance' between mother and daughter.


Sequence analysis of active mariner elements in natural populations of Drosophila simulans.
Active and inactive mariner elements from natural and laboratory populations of Drosophila simulans were isolated and sequenced in order to assess their nucleotide variability and to compare them with previously isolated mariner elements from the sibling species Drosophila mauritiana and Drosophila sechellia. The active elements of D. simulans are very similar among themselves (average 99.7% nucleotide identity), suggesting that the level of mariner expression in different natural populations is largely determined by position effects, dosage effects and perhaps other factors. Furthermore, the D. simulans elements exhibit nucleotide identities of 98% or greater when compared with mariner elements from the sibling species. Parsimony analysis of mariner elements places active elements from the three species into separate groups and suggests that D. simulans is the species from which mariner elements in D. mauritiana and D. sechellia are most likely derived. This result strongly suggests that the ancestral form of mariner among these species was an active element. The two inactive mariner elements sequenced from D. simulans are very similar to the inactive peach element from D. mauritiana. The similarity may result from introgression between D. simulans and D. mauritiana or from selective constraints imposed by regulatory effects of inactive elements.


Isolation and characterization of turkey bone marrow-derived mesenchymal stem cells.
Flexor tendon injury is often associated with suboptimal outcomes and results in substantial digit dysfunction. Stem cells have been isolated from several experimental animals for the growing interest and needs of utilizing cell-based therapies. Recently, turkey has been developed as a new large animal model for flexor tendon research. In the present study, we reported the isolation and characterization of bone marrow-derived mesenchymal stem cells (BMSCs) from 8- to 12-month-old heritage-breed turkeys. The isolated cells demonstrated fibroblast-like morphology, clonogenic capacity, and high proliferation rate. These cells were positive for surface antigens CD90, CD105, and CD44, but were negative for CD45. The multipotency of turkey BMSCs was determined by differentiating cells into osteogenic, adipogenic, chondrogenic, and tenogenic lineages. There was upregulated gene expression of tenogenic markers, including mohawk, tenomodulin, and EGR1 as well as increased collagen synthesis in BMP12 induced cells. The successful isolation and verification of bone marrow-derived MSCs from turkey would provide opportunities of studying cell-based therapies and developing new treatments for tendon injuries using this novel preclinical large animal model. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:1419-1428, 2019.


The new RENT family of repetitive elements in Nicotiana species harbors gene regulatory elements related to the tCUP cryptic promoter.
The tCUP cryptic constitutive promoter was discovered in the tobacco genome by T-DNA (transfer DNA) tagging with a promoterless GUS-nos gene. Here, we show that the portion of the tCUP sequence containing a variety of cryptic gene regulatory elements is related to a new family of moderately repetitive sequences (10(2) copies), the RENT (repetitive element from Nicotiana tabacum) family. The RENT family is found only in certain Nicotiana species. Five RENT elements were cloned and sequenced. The RENT elements are a minimum of 5 kb in length and share 80-90% sequence similarity throughout their length. The 5' termini are the same in the isolated RENT family members and are characterized by a conserved border sequence (TGTTGA(T or C)ACCCAATTTT(T or C)). The 3' ends of RENT sequence similarity vary in location and sequence. The tCUP cryptic promoter originated from a unique truncated RENT element that interrupts a phytochelatin synthase-like gene that may have undergone rearrangements prior to or resulting from T-DNA insertion. No evidence was found for expressed coding regions within the RENT elements; however, like the cryptic gene regulatory elements within the tCUP sequence, the isolated RENT elements possess promoter activity and translational enhancer activity.


Non-animal sensitization testing: state-of-the-art.
Predictive tests to identify the sensitizing properties of chemicals are carried out using animals. In the European Union timelines for phasing out many standard animal tests were established for cosmetics. Following this policy, the new European Chemicals Legislation (REACH) favors alternative methods, if validated and appropriate. In this review the authors aim to provide a state-of-the art overview of alternative methods (in silico, in chemico, and in vitro) to identify contact and respiratory sensitizing capacity and in some occasions give a measure of potency. The past few years have seen major advances in QSAR (quantitative structure-activity relationship) models where especially mechanism-based models have great potential, peptide reactivity assays where multiple parameters can be measured simultaneously, providing a more complete reactivity profile, and cell-based assays. Several cell-based assays are in development, not only using different cell types, but also several specifically developed assays such as three-dimenionally (3D)-reconstituted skin models, an antioxidant response reporter assay, determination of signaling pathways, and gene profiling. Some of these assays show relatively high sensitivity and specificity for a large number of sensitizers and should enter validation (or are indeed entering this process). Integrating multiple assays in a decision tree or integrated testing system is a next step, but has yet to be developed. Adequate risk assessment, however, is likely to require significantly more time and efforts.


The mode of action of nerve growth factor in PC12 cells.
This review deals with the mechanism of nerve growth factor action. In view of the many and diversified effects of this growth factor, and since it could utilize different mechanism(s) in distinct types of cells, we have confined our analysis to the best characterized and more extensively studied target, the clonal cell line PC12. When exposed to NGF in vitro, these neoplastic cells recapitulate the last major steps of neuronal differentiation, i.e., the commitment to become a neuron and the acquisition of the neuronal phenotype. This is characterized by electrically excitable neurites, a display of a highly organized cytoskeleton, and the specific chemical and molecular neuronal properties. These effects are elicited upon the interaction of NGF with a receptor whose gene has been cloned and whose kinetic properties are now relatively well characterized. It is not yet clear, on the contrary, if and which of the several potential second messengers (cAMP, Ca, or phosphoinositides) that undergo marked fluctuations following NGF binding, transduce and amplify the NGF message. Among both the early and late effects of NGF is the modulation of expression of several genes. Some of the products of these genes are mainly restricted to nerve cells and others appear to play a crucial role in regulating the proper assembly of cytoskeletal elements. It is hypothesized that this complex array of chemical, molecular, and ultrastructural changes is triggered by NGF, not through activation of a single pathway, but more likely via combinatorial processes whereby several intracellular signals interplay before the irreversible commitment of becoming a neuron is undertaken.


Decreased expression of BRCA1 in SK-BR-3 cells is the result of aberrant activation of the GABP Beta promoter by an NRF-1-containing complex.
BRCA1 has recently been identified as a potential regulator of mammary stem/progenitor cell differentiation, and this function may explain the high prevalence of breast cancer in BRCA1 mutation carriers, as well as the downregulation of BRCA1 in a large proportion of sporadic breast cancers. That is, loss of BRCA1 function results in blocked differentiation with expansion of the mammary stem/progenitor cells. Because BRCA1 also maintains genomic integrity, its loss could produce a pool of genetically unstable stem/progenitor cells that are prime targets for further transforming events. Thus, elucidating the regulatory mechanisms of BRCA1 expression is important to our understanding of normal and malignant breast differentiation. Loss of BRCA1 expression in the ErbB2-amplified SK-BR-3 cell line was found to be the result of loss of activity of the ets transcription factor GABP, a previously characterized regulator of BRCA1 transcription. The expression of the non-DNA binding GABPβ subunit was shown to be deficient, while the DNA binding subunit, GABPα was rendered unstable by the absence of GABPβ. Deletion analysis of the GABPβ proximal promoter identified a potential NRF-1 binding site as being critical for expression. Supershift analysis, the binding of recombinant protein and chromatin immunoprecipitation confirmed the role of NRF-1 in regulating the expression of GABPβ. The siRNA knockdown of NRF-1 resulted in decreased GABPβ and BRCA1 expression in MCF-7 cells indicating that they form a transcriptional network. NRF-1 levels and activity did not differ between SK-BR-3 and MCF-7 cells, however the NRF-1 containing complex on the GABPβ promoter differed between the two lines and appears to be the result of altered coactivator binding. Both NRF-1 and GABP have been linked to the regulation of nuclear-encoded mitochondrial proteins, and the results of this study suggest their expression is coordinated by NRF-1's activation of the GABPβ promoter. Their linkage to BRCA1, a potential breast stem cell regulator, implies a connection between the induction of mitochondrial metabolism and breast differentiation.


Treatment of tibial defect and bone nonunion with limb shortening with external fixator and reconstituted bone xenograft.
To explore the effect of external fixator and reconstituted bone xenograft (RBX) in the treatment of tibial bone defect, tibial bone nonunion and congenital pseudarthrosis of the tibia with limb shortening. Twenty patients (13 males and 7 females) with tibial bone defect, tibial bone nonunion or congenital pseudarthrosis of the tibia with limb shortening were treated with external fixation. Two kinds of external fixators were used: a half ring sulcated external fixator used in 13 patients and a combined external fixator in 7 patients. Foot-drop was corrected at the same time with external fixation in 4 patients. The shortened length of the tibia was in the range of 2-9 cm, with an average of 4.8 cm. For bone grafting, RBX was used in 12 patients, autogenous ilium was used in 3 patients and autogenous fibula was implanted as a bone plug into the medullary canal in 1 case, and no bone graft was used in 4 patients. All the 20 patients were followed-up for 8 months to 7 years, averaging 51 months. Satisfactory function of the affected extremities was obtained. All the shortened extremities were lengthened to the expected length. For all the lengthening area and the fracture sites, bone union was obtained at the last. The average healing time of 12 patients treated with RBX was 4.8 months. Both the half ring sulcated external fixator and the combined external fixator have the advantages of small trauma, simple operation, elastic fixation without stress shielding and non-limitation from local soft tissue conditions, and there is satisfactory functional recovery of affected extremities in the treatment of tibial bone defects, tibial bone nonunion and congenital pseudarthrosis of the tibia combined with limb shortening. RBX has good biocompatibility and does not cause immunological rejections. It can also be safely used in treatment of bone nonunion and has reliable effect to promote bone healing.


Discovery of JN122, a Spiroindoline-Containing Molecule that Inhibits MDM2/p53 Protein-Protein Interaction and Exerts Robust In Vivo Antitumor Efficacy.
MDM2 and MDM4 cooperatively and negatively regulate p53, while this pathway is often hijacked by cancer cells in favor of their survival. Blocking MDM2/p53 interaction with small-molecule inhibitors liberates p53 from MDM2 mediated degradation, which is an attractive strategy for drug discovery. We reported herein structure-based discovery of highly potent spiroindoline-containing MDM2 inhibitor (-)60 (JN122), which also exhibited moderate activities against MDM4/p53 interactions. In a panel of cancer cell lines harboring wild type p53, (-)60 efficiently promoted activation of p53 and its target genes, inhibited cell cycle progression, and induced cell apoptosis. Interestingly, (-)60 also promoted degradation of MDM4. More importantly, (-)60 exhibited good PK properties and exerted robust antitumor efficacies in a systemic mouse xenograft model of MOLM-13. Taken together, our study showcases a class of potent MDM2 inhibitors featuring a novel spiro-indoline scaffold, which is promising for future development targeting cancer cells with wild-type p53.


Head and pelvic movement asymmetries at trot in riding horses in training and perceived as free from lameness by the owner.
Recent studies evaluating horses in training and considered free from lameness by their owners have identified a large proportion of horses with motion asymmetries. However the prevalence, type and magnitude of asymmetries when trotting in a straight line or on the lunge have not been investigated. The aim of this study was to objectively investigate the presence of motion asymmetries in riding horses in training by identifying the side and quantifying the degree and type (impact, pushoff) of forelimb and hind limb asymmetries found during straight line trot and on the lunge. In a cross-sectional study, vertical head and pelvic movement symmetry was measured in 222 Warmblood type riding horses, all without perceived performance issues and considered free from lameness by their owners. Body-mounted uni-axial accelerometers were used and differences between maximum and minimum head (HDmax, HDmin) and pelvic (PDmax, PDmin) vertical displacement between left and right forelimb and hind limb stances were calculated during straight line trot and on the lunge. Previously reported symmetry thresholds were used. The thresholds for symmetry were exceeded in 161 horses for at least one variable while trotting in a straight line, HDmin (n = 58, mean 14.3 mm, SD 7.1), HDmax (n = 41, mean 12.7 mm, SD 5.5), PDmax (n = 87, mean 6.5 mm, SD 3.10), PDmin (n = 79, mean 5.7 mm, SD 2.1). Contralateral and ipsilateral concurrent forelimb and hind limb asymmetries were detected in 41 and 49 horses, respectively. There was a linear association between the straight line PDmin values and the values on the lunge with the lame limb to the inside of the circle. A large proportion (72.5%) of horses in training which were perceived as free from lameness by their owner showed movement asymmetries above previously reported asymmetry thresholds during straight line trot. It is not known to what extent these asymmetries are related to pain or to mechanical abnormalities. Therefore, one of the most important questions that must be addressed is how objective asymmetry scores can be translated into pain, orthopedic abnormality, or any type of unsoundness.


Effect of gold compounds on phorbol myristic acetate (PMA) activated superoxide (O2-) production by mouse peritoneal macrophages.
A number of gold compounds, nonsteroidal antiinflammatory drugs (NSAID) and protease inhibitors have been examined for their ability to inhibit the respiratory burst of peritoneal macrophages. A potent (IC50 = 5 microM or less) dose dependent inhibitory activity was seen with a number of triethylphosphine gold I ( TEPG ) compounds. Nonphosphine gold compounds were less active. The ligands of TEPG were inactive. Several NSAID stimulated the respiratory burst, whereas 2 protease inhibitors tested were inhibitory. The structure activity relationship of TEPG is also presented.


Systematic screening of the LDL-PLA2 gene for polymorphic variants and case-control analysis in schizophrenia.
Systematic scans of the genome using microsatellite markers have identified chromosome 6p21.1 as a putative locus for schizophrenia in multiply affected families. There is also evidence from a series of studies for a role of abnormal phospholipid metabolism in schizophrenia. In light of these findings, and the role of platelet activating factor in neurotransmission and neurodevelopment, we have examined the LDL-PLA2 (plasma PAF acetylhydrolase, PAF-AH) gene, a serine dependent phospholipase that has been mapped by hybrid mapping to chromosome 6p21.1, as a positional candidate gene for schizophrenia. The gene was systematically screened using SSCP/HD analysis for polymorphisms associated with the disease. Four polymorphic variants were found within the gene and studied in a group of 200 schizophrenic patients and 100 controls. The variant in exon 7 (Iso195Thr) was found to be weakly associated with schizophrenia (p = 0.04) and the variant in exon 11 (Val379Ala) almost reached significance (p = 0.057). After correcting for multiple testing no significant associations were detected. Haplotype analysis combining pairs of polymorphisms also provided no evidence for association of this gene with schizophrenia in our sample of patients.


[Dengue and chikungunya acquired during travel in the tropics].
The global incidence of dengue and chikungunya has greatly increased over recent decades, partly due to the increase of geographic distribution of both vectors. These infections are endemic to the tropics and subtropics, however autochthonous transmission and outbreaks have been described in non-endemic areas. Currently, there is a large chikungunya outbreak in the western hemisphere which started in the Caribbean. Chikungunya had not previously been endemic to this region. Both arboviral infections are important causes of fever in Dutch travellers returning from tropical destinations. The clinical presentations of dengue and chikungunya overlap; both are characterised by high fever and arthralgia. Bleeding and plasma leakage are potentially life-threatening complications of dengue, while persistent arthralgia typifies chikungunya. The prevention of mosquito bites, by using protective clothing and insect repellents, is the only way to prevent infection. No vaccine is yet available.


Leukotriene B4 causes proliferation of interleukin 2-dependent T cells in the presence of suboptimal levels of interleukin 2.
We have recently found that endogenous leukotriene B4 (LTB4) production is a necessary component of mitogen-stimulated T-cell proliferation. In this report, we address the relationship between LTB4 and interleukin 2 (IL-2) in stimulating proliferation of IL-2 responsive T cells. We employed an IL-2 responsive T-cell line, HT-2 and also human peripheral blood T cells rendered IL-2 responsive by culture with phytohemagglutinin (PHA). In the presence of concentrations of IL-2 that resulted in suboptimal stimulation of HT-2 cells (5-20% of maximum proliferation), LTB4 at 10(-8) or 10(-10) M produced a several-fold increase in [3H]thymidine incorporation, resulting in levels of [3H]thymidine incorporation that were comparable to those produced by HT-2 cells stimulated with optimal concentrations of IL-2. Similar results were obtained with the peripheral blood T cells. Leukotriene C4, prostaglandin E2 (PGE2), and arachidonic acid did not share with LTB4 the ability to stimulate proliferation of HT-2 cells. Hydrocortisone and nordihydroguairetic acid (NDGA), drugs which block endogenous LTB4 production in cell cultures inhibited [3H]thymidine incorporation of HT-2 cells stimulated by suboptimal but not optimal levels of IL-2. This inhibition could be overcome by the readdition of LTB4 to the cultures. Thus, LTB4 in physiologic concentrations (10(-10) M) can substitute for IL-2 in stimulating proliferation of IL-2 responsive T cells, provided that some IL-2 is present in the culture. Endogenous LTB4 may act synergistically with endogenous IL-2 in promoting T-cell proliferation.


Mealtime Disruption and Caregiver Stress in Referrals to an Outpatient Feeding Clinic.
This investigation explores the feeding behaviors, comorbidities, and parenting stress in young children referred to an outpatient feeding clinic. Eligible participants (n =72) were primary caretakers of children ages 2-6 years referred to an interdisciplinary outpatient feeding disorder clinic, subcategorized according to the absence (n = 18) or presence (n = 54) of caretaker-reported medical or developmental comorbidities. This group was compared with an equivalent control sample of caretakers of age-matched children (n = 72). Measures included the Children's Eating Behavior Inventory (CEBI) and the Parenting Stress Index-Short Form (PSI-SF). The CEBI and PSI scores were higher in the feeding disorder group than in the control group. PSI total was incrementally increased for control vs feeding disorder without comorbidity vs feeding disorder with comorbidity. Increased maladaptive behaviors and parenting stress were evident in children with feeding disorder regardless of the presence of comorbidity. Parenting stress was exacerbated by the presence of comorbidity.


Comparative genomic analyses reveal diverse virulence factors and antimicrobial resistance mechanisms in clinical Elizabethkingia meningoseptica strains.
Three human clinical isolates of bacteria (designated strains Em1, Em2 and Em3) had high average nucleotide identity (ANI) to Elizabethkingia meningoseptica. Their genome sizes (3.89, 4.04 and 4.04 Mb) were comparable to those of other Elizabethkingia species and strains, and exhibited open pan-genome characteristics, with two strains being nearly identical and the third divergent. These strains were susceptible only to trimethoprim/sulfamethoxazole and ciprofloxacin amongst 16 antibiotics in minimum inhibitory tests. The resistome exhibited a high diversity of resistance genes, including 5 different lactamase- and 18 efflux protein- encoding genes. Forty-four genes encoding virulence factors were conserved among the strains. Sialic acid transporters and curli synthesis genes were well conserved in E. meningoseptica but absent in E. anophelis and E. miricola. E. meningoseptica carried several genes contributing to biofilm formation. 58 glycoside hydrolases (GH) and 25 putative polysaccharide utilization loci (PULs) were found. The strains carried numerous genes encoding two-component system proteins (56), transcription factor proteins (187~191), and DNA-binding proteins (6~7). Several prophages and CRISPR/Cas elements were uniquely present in the genomes.


Diagnostic Accuracy of MR Spectroscopic Imaging and 18F-FET PET for Identifying Glioma: A Biopsy-Controlled Hybrid PET/MRI Study.
Contrast-enhanced MRI is the method of choice for brain tumor diagnostics, despite its low specificity for tumor tissue. This study compared the contribution of MR spectroscopic imaging (MRSI) and amino acid PET to improve the detection of tumor tissue. Methods: In 30 untreated patients with suspected glioma, O-(2-[18F]fluoroethyl)-l-tyrosine (18F-FET) PET; 3-T MRSI with a short echo time; and fluid-attenuated inversion recovery, T2-weighted, and contrast-enhanced T1-weighted MRI were performed for stereotactic biopsy planning. Serial samples were taken along the needle trajectory, and their masks were projected to the preoperative imaging data. Each sample was individually evaluated neuropathologically. 18F-FET uptake and the MRSI signals choline (Cho), N-acetyl-aspartate (NAA), creatine, myoinositol, and derived ratios were evaluated for each sample and classified using logistic regression. The diagnostic accuracy was evaluated by receiver operating characteristic analysis. Results: On the basis of the neuropathologic evaluation of tissue from 88 stereotactic biopsies, supplemented with 18F-FET PET and MRSI metrics from 20 areas on the healthy-appearing contralateral hemisphere to balance the glioma/nonglioma groups, 18F-FET PET identified glioma with the highest accuracy (area under the receiver operating characteristic curve, 0.89; 95% CI, 0.81-0.93; threshold, 1.4 × background uptake). Among the MR spectroscopic metabolites, Cho/NAA normalized to normal brain tissue showed the highest diagnostic accuracy (area under the receiver operating characteristic curve, 0.81; 95% CI, 0.71-0.88; threshold, 2.2). The combination of 18F-FET PET and normalized Cho/NAA did not improve the diagnostic performance. Conclusion: MRI-based delineation of gliomas should preferably be supplemented by 18F-FET PET.


Co-Exsolution of Ni-Based Alloy Catalysts for the Valorization of Carbon Dioxide and Methane.
ConspectusThe reversible coexsolution mechanism of perovskite oxides is emerging as an alternative method for synthesizing alloy catalyst nanoparticles. Co-exsolution is a partial decomposition process where multiple B cations diffuse from the bulk of a solid precursor and nucleate on the surface. The unique properties of exsolved alloy catalysts, including improved dispersion, thermal stability, and compositional malleability, make them particularly useful for converting CO2 into chemical commodities and fuels. However, the coexsolution of alloys is still in development, and fundamental insights into the alloying mechanism, formation of nanoparticles, and defect chemistry are needed.This Account examines the solid-state chemistry of perovskite oxide precursors and reaction parameters that can be altered to control the assembly or exsolution of Ni-based alloys. The characteristics of bulk perovskite oxide precursors heavily influence the exsolved alloy catalyst nanoparticle assembly, growth, and composition. Inherent defects, such as oxygen vacancies and grain boundaries, primarily facilitate the transport of catalytic B-cation dopants from the bulk to the surface. An example of how bulk defects can affect the properties of Ni-based alloy catalysts is demonstrated through the formation of NiFe from La(Fe, Ni)O3. The A/B cation ratio plays a significant role in determining the size and composition of NiFe nanoparticles, which directly impacts their catalytic performance. Using in situ X-ray absorption spectroscopy (in situ XAS), the dynamic behavior of exsolved NiFe nanoparticles can be observed in different reaction environments (oxidation, reduction, and dry reforming of methane) by tracking the oxidation state and local environment of the Ni K-edge and Fe K-edge using X-ray absorption near-edge structure (XANES) and extended X-ray absorption fine structure (EXAFS), respectively. Time-resolved experiments with in situ XAS showed that NiFe nanoparticle growth starts at ∼280 °C and transforms from predominantly Ni to NiFe at higher reduction times and temperatures.The challenges of exsolution of higher-order Ni-based alloys, such as 3(NiFeCo), 4(NiCoCuPd), and 5(NiFeCoCuPd) element nanoparticles, to improve the catalyst properties are discussed. The size, concentration, and reducibility of the dopant cation can alter the exsolution kinetics, alloy nanoparticle growth dynamics, and catalyst performance. The size and composition of exsolved Ni-based alloys affect the effectiveness of catalysts in the dry reforming of methane. Large NiFeCo nanoparticles separated from Pd and Cu can lead to catalyst deactivation, but using a complex alloy with smaller NiFeCoPdCu nanoparticles results in a stable performance. The use of in situ XANES reveals how the dry reforming of methane reaction conditions can induce changes in the NiFe with the rapid redissolution of Fe back into the lattice.The dynamicity of the exsolved Ni-based alloy nanoparticles and implications for their regeneration after aging or exposure to waste gas contaminants are discussed. Finally, we summarize the Account and provide promising future directions.


Determination of silver in blood, urine, and tissues of volunteers and burn patients.
Silver sulfadiazine cream (SSD) has been used successfully in the management of burn wound sepsis. Silver deposition has been found in the skin, gingiva, cornea, liver, and kidney of patients treated with this cream, causing argyria, ocular injury, leukopenia, and toxicity in kidney, liver, and neurologic tissues. Monitoring concentrations of silver in blood and urine of patients receiving this treatment has become necessary, but sensitive and suitable methods adaptable to a clinical laboratory are still needed. We have developed a flameless thermal atomic absorption spectrophotometric method to measure silver concentrations in blood, urine, and other tissues. The detection limit is 0.4 microgram/L; the within-run precisions (CV) are 5.16%, 3.83%, and 2.79% for concentrations of 5, 13.5, and 42 micrograms/L, respectively; and the between-run precisions are 4.3% and 3.2% for concentrations of 13.5 and 42 micrograms/L. The concentrations of silver in blood, urine, liver, and kidney of subjects without industrial or medicinal exposure are less than 2.3 micrograms/L, 2 micrograms/day, 0.05 microgram/g wet tissue, and 0.05 microgram/g wet tissue, respectively. In SSD cream-treated burn patients, plasma concentrations may be as great as 50 micrograms/L within 6 h of treatment and can reach a maximum of 310 micrograms/L. Silver in urine is detectable after one day of treatment and may reach a maximum of 400 micrograms/day. After absorption, silver was found to be deposited in various tissues. Tissue silver concentrations in one burn patient who died of renal failure after eight days of treatment were 970, 14, and 0.2 micrograms/g wet tissue in cornea, liver, and kidney, respectively.


Postural Control Differences between Patients with Posterior Tibial Tendon Dysfunction and Healthy People during Gait.
Patients with posterior tibial tendon dysfunction (PTTD) may exhibit postural instability during walking likely due to a loss of medial longitudinal arch, abnormal foot alignment, and pain. While many studies have investigated gait alterations in PTTD, there is no understanding of dynamic postural control mechanisms in this population during gait, which will help guide rehabilitation and gait training programs for patients with PTTD. The purpose of the study was to assess dynamic postural control mechanisms in patients with stage II PTTD as compared to age and gender matched healthy controls. Eleven patients with stage II PTTD (4 males and 7 females; age 59 ± 1 years; height 1.66 ± 0.12 m; mass 84.2 ± 16.0 kg) and ten gender and age matched controls were recruited in this study. Participants were asked to walk along a 10 m walkway. Ten Vicon cameras and four AMTI force platforms were used to collect kinematic and center of pressure (COP) data while participants performed gait. To test differences between PTTD vs. control groups, independent t-tests (set at α < 0.05) were performed. Patients with PTTD had significantly higher double stance ratio (+23%) and anterior-posterior (AP) time to contact (TTC) percentage (+16%) as compared to healthy control. However, PTTD had lower AP COP excursion (-19%), AP COP velocity (-30%), and medial-lateral (ML) COP velocity (-40%) as compared to healthy controls. Mean ML COP trace values for PTTD were significantly decreased (-23%) as compared to controls, indicating COP trace for PTTD tends to be closer to the medial boundary than controls during single-support phase of walking. PTTD patients showed more conservative and cautious postural strategies which may help maintain balance and reduce the need for postural adjustment during PTTD gait. They also showed more medially shifted COP patterns than healthy controls during single-support phase of walking. Dynamic postural control outcomes could be used to develop effective gait training programs aimed at alleviating a medial shift of COP (everted foot) for individuals with PTTD in order to improve their functionality and gait efficiency.


Epigenetic maintenance of telomere identity in Drosophila: buckle up for the sperm ride.
A critical function of telomeres is to prevent the ligation of chromosome ends by DNA repair enzymes. In most eukaryotes, telomeric DNA consists in large arrays of G-rich tandem repeats that are recognized by DNA binding capping proteins. Drosophila telomeres are unusual as they lack short tandem repeats. However, Drosophila capping proteins can bind chromosome extremities in a DNA sequence-independent manner. This epigenetic protection of fly telomeres has been essentially studied in somatic cells where capping proteins such as HOAP or HP1 are essential in preventing chromosome end-to-end fusions. HipHop and K81 are two recently identified paralogous capping proteins with complementary expression patterns. While HipHop is involved in telomere capping in somatic cells, K81 has specialized in the protection of telomeres in post-meiotic male germ cells. Remarkably, K81 is required for the stabilization of HOAP and HP1 at telomeres during the massive paternal chromatin remodeling that occurs during spermiogenesis and at fertilization. We thus propose that the maintenance of capping proteins at Drosophila sperm telomeres is crucial for the transmission of telomere identity to the diploid zygote. :


Recent Bendamustine Treatment Before Apheresis Has a Negative Impact on Outcomes in Patients With Large B-Cell Lymphoma Receiving Chimeric Antigen Receptor T-Cell Therapy.
Approximately 30%-40% of patients with relapsed/refractory (R/R) large B-cell lymphoma (LBCL) infused with CD19-targeted chimeric antigen receptor (CAR) T cells achieve durable responses. Consensus guidelines suggest avoiding bendamustine before apheresis, but specific data in this setting are lacking. We report distinct outcomes after CAR T-cell therapy according to previous bendamustine exposure. The study included CAR T-cell recipients from seven European sites. Safety, efficacy, and CAR T-cell expansion kinetics were analyzed according to preapheresis bendamustine exposure. Additional studies on the impact of the washout period and bendamustine dose were performed. Inverse probability treatment weighting (IPTW) and propensity score matching (PSM) analyses were carried out for all efficacy comparisons between bendamustine-exposed and bendamustine-naïve patients. The study included 439 patients with R/R LBCL infused with CD19-targeted commercial CAR T cells, of whom 80 had received bendamustine before apheresis. Exposed patients had significantly lower CD3+ cells and platelets at apheresis. These patients had a lower overall response rate (ORR, 53% v 72%; P < .01), a shorter progression-free survival (PFS, 3.1 v 6.2 months; P = .04), and overall survival (OS, 10.3 v 23.5 months; P = .01) in comparison with the bendamustine-naïve group. Following adjustment methods for baseline variables, these differences were mitigated. Focusing on the impact of bendamustine washout before apheresis, those with recent (<9 months) exposure (N = 42) displayed a lower ORR (40% v 72%; P < .01), shorter PFS (1.3 v 6.2 months; P < .01), and OS (4.6 v 23.5 months; P < .01) in comparison with bendamustine-naïve patients. These differences remained significant after IPTW and PSM analysis. Conversely, the cumulative dose of bendamustine before apheresis did not affect CAR-T efficacy outcomes. Recent bendamustine exposure before apheresis was associated with negative treatment outcomes after CD19-targeted CAR T-cell therapy and should be therefore avoided in CAR T-cell candidates.


Electrophysiologic patterns of oral-pharyngeal swallowing in parkinsonian syndromes.
To assess the presence, severity, and differences in dysphagia in Parkinson disease (PD), Parkinson variant of multiple system atrophy (MSA-P), and progressive supranuclear palsy (PSP), and to study the pathophysiology of swallowing abnormalities in these disorders. We applied an electrophysiologic method to evaluate oral-pharyngeal swallowing. We analyzed the following measures: duration of EMG activity of suprahyoid/submental muscles (SHEMG-D); duration of laryngeal-pharyngeal mechanogram (LPM-D); duration of the inhibition of the cricopharyngeal muscle activity (CPEMG-ID); interval between onset of EMG activity of suprahyoid/submental muscles and onset of laryngeal-pharyngeal mechanogram (I-SHEMG-LPM); and swallowing reaction time (SRT). The prolongation of I-SHEMG-LPM was more typical in PD, whereas the most distinctive finding both in patients with PSP and MSA-P was the reduction or the absence of CPEMG-ID early in the course of the disease. Involvement of the peduncolo-pontine tegmental nucleus, with subsequent dysfunction of basal ganglia and of the medullary central pattern generator of swallowing, may account for the abnormalities detected in these parkinsonian syndromes. The method described was able to identify swallowing abnormalities also in patients without symptoms of dysphagia and to evaluate dysphagia severity in all patients.


Sulpiride, a D2/D3 blocker, reduces cataplexy but not REM sleep in canine narcolepsy.
Cataplexy, an abnormal manifestation of REM sleep atonia, is currently treated with antidepressants. These medications also reduce physiological REM sleep and induce nocturnal sleep disturbances. Because a recent work on canine narcolepsy suggests that the mechanisms for triggering cataplexy are different from those for REM sleep, we hypothesized that compounds which act specifically on cataplexy, but not on REM sleep, could be developed. Canine studies also suggest that the dopamine D2/D3 receptor mechanism is specifically involved in the regulation of cataplexy, but little evidence suggests that this mechanism is important for REM sleep regulation. We therefore assessed the effects of sulpiride, a commonly used D2/D3 antagonist, on cataplexy and sleep in narcoleptic canines to explore the possible clinical application of D2/D3 antagonists for the treatment of human narcolepsy. Both acute and chronic oral administration of sulpiride (300 mg/dog, 600 mg/dog) significantly reduced cataplexy without noticeable side effects. Interestingly, the anticataplectic dose of sulpiride did not significantly reduce the amount of REM sleep. Sulpiride (and other D2/D3 antagonists) may therefore be an attractive new therapeutic indication in human narcolepsy.


Large Group Exposure Treatment: A Feasibility Study of Exposure Combined with Diaphragmatic Breathing in Highly Dental Fearful Individuals.
A large-group one session treatment (LG-OST) combining exposure and diaphragmatic breathing as a bodily coping element was carried out to investigate its feasibility and effectiveness in a sample of 43 highly dental fearful individuals treated simultaneously. We assessed subjective dental fear, dysfunctional dental-related beliefs, and perceived control pre- and post-intervention and at four-month follow-up. Participants additionally performed a behavioural approach test (BAT) pre- and post-intervention. During the applied exposure exercises, four participants (9.3%) discontinued the program all reporting too high levels of distress. Regarding subjective dental fear and dysfunctional dental related beliefs post treatment effects, LG-OST showed medium to large effect sizes, ranging from Cohen's d = 0.51 to d = 0.84 in the Intention-to-Treat analysis. Subjective dental fear improved clinically significantly in about one fourth (25.6%) of therapy completers. All post-treatment effects remained stable over time. Concerning the behavioral fear dimension, we observed a strong ceiling effect. Already at pre-assessment, participants accomplished more than six out of seven BAT-steps. Thus, behavioral approach did not increase significantly following treatment. Overall, the LG-OST protocol proved feasible and efficient. Compared to other one-session individual and multi-session group treatments the observed LG-OST effects were smaller. However, if LG-OST could match the efficacy of highly intensive short treatments delivered in an individual setting in the future, for example, by applying a wider array of exposure exercises, it could be a very useful treatment option as an intermediate step within a stepped care approach.


[Proceedings: Microbiology of endocarditis].
The classical paper published by Schottmüller in 1903 has shown beyond doubt that blood cultures are a crucial diagnostic procedure for the diagnosis of endocarditis. Positive blood cultures also make it possible to distinguish between bacteremia and endocarditis: in the latter condition the number of organisms per ml of blood is relatively constant and all blood cultures are usually positive. Gram positive cocci are still the commonest organisms found in classical endocarditis. Gram negative organisms, yeasts and fungi are cultured with increasing frequency in cases of endocarditis secondary to valve replacement. Sensitivity testing by the disc diffusion method determines the minimal inhibitory concentration of the antibiotic towards the isolated organism. However, minimal bactericidal concentrations are of much greater prognostic value in endocarditis and should be performed routinely on the isolated organism.


[Dramatic reduction of myocardial infarction mortality. A retrospect of the past century].
Ischemic heart disease is the most common cause of death in the western world. Acute myocardial infarction is one of the most life-threatening manifestations of ischemic heart disease. Treatment of acute myocardial infarction has dramatically improved over the past 100 years by the introduction of coronary care units, mobile coronary care units, improved treatment of cardiac arrest plus the introduction of beta-blockers, fibrinolytic agents, aspirin, ACE-inhibitors and lipid lowering drugs. This has resulted in a marked reduction of mortality.


Diffusing wave spectroscopy used to study the influence of shear on aggregation.
In this study, diffusing wave spectroscopy (DWS) is used to investigate the effect of shear on a food-related aggregating emulsion. The principle of the method is validated using a nonaggregating, nearly monodisperse latex suspension. In general, with increasing shear rate the diffusive motion of the scatterers becomes negligible compared to the convective motion. This causes a decrease in the decay time of the autocorrelation curves and a change in the form of the autocorrelation curves from nearly exponential to Gaussian. This is reflected in the exponent of the mean square displacement that changes from 1 to 2. The effect of shear on the acidification of a sodium caseinate-stabilized emulsion was studied by DWS and by rheometry. The emulsion droplets in the food-related emulsion were uniformly dispersed at neutral pH. Upon acidification down to a pH of 5.2 +/- 0.05, the emulsion showed Newtonian behavior with constant viscosity over the whole pH range. At pH 5.17 +/- 0.05, independent of the applied shear rate during acidification, the viscosity suddenly increased. From this point on, the emulsion showed shear-thinning behavior. The photon-transport mean free path ( l*) was not influenced by the applied shear rate and did not change down to pH 5.2 +/- 0.05. Close to this pH, l* increased, and the decay of the autocorrelation curves shifted to longer correlation times when shear rates smaller than 1 s (-1) were applied. At lower pH (5.05 +/- 0.05), l* started to fluctuate, and the autocorrelation curves no longer decayed to zero, indicating that at these shear rates the system behaved nonergodicly. Assuming that the convective motion and the Brownian motion are independent of each other, the mean square displacement as a result of Brownian motion was determined. From this, the sol-gel point and the radius of the aggregates at this point as a function of the shear rate was determined. The results indicated that the radius of the aggregates at the sol-gel transition decreased with increasing shear rate and suggested that shear will result in a more open structure of the network formed by the aggregates.


[Chronic duodenal ulcer. Treatment by anterior fundus seromyotomy with posterior truncal vagotomy].
Anterior seromyotomy of the gastric lesser curvature with additional posterior truncal vagotomy has recently been proposed by Taylor in the elective surgical treatment of chronic duodenal ulcer disease. We report here a technical variant of this procedure. Our results are similar to those following proximal gastric vagotomy, with no operative mortality, minor functional disorders (100% Visick grade I or II at six months), significant reduction of gastric acidity (88.1% reduction of the basal acid output at 6 months) and no ulcer recurrence in the short-term follow-up. This technique offers several advantages over the operations previously designed. It is easy to perform and not time consuming; moreover, the procedure does not take into account possible variations of the vagus nervi, does not require dissection of the abdominal oesophagus and does not entail the risk of lesser curvature necrosis.


Comparison of the therapeutic effect of the Persian Medicine Protocol with the common treatment of chronic rhinosinusitis: a randomized clinical trial.
Chronic rhinosinusitis is one of the most common diseases affecting the quality of life of patients. Patients suffer from high costs in the diagnosis and treatment of the disease. Frequent recurrence and failure of therapeutic protocols are among the most important issues in the management of this disease. In view of this, the use of traditional and complementary therapies to promote the treatment of this disease has been increasingly taken into account. Comparison of the effectiveness of the Persian Medicine Protocol with the conventional therapy in the treatment of chronic rhinosinusitis. A randomized clinical trial was conducted at Imam Reza Hospital in Mashhad, Iran from July 2016 to March 2017. For patients with chronic rhinosinusitis symptoms, endoscopy of the sinuses was performed by an ENT specialist and in the case of negative endoscopy, paranasal sinus CT scan was requested. A total of 42 patients with chronic rhinosinusitis were randomly assigned to two groups. The first group (classical) received systemic and intranasal cortisone, and the second group (traditional) received a therapeutic Persian medicine protocol including intranasal lavender oil, and Liquorice Marjoram Tea (L. M. tea) for six weeks. The symptoms of the patients were evaluated using the SNOT-22 questionnaire at the beginning of the study and at the sixth week. If no improvement occurs, treatment continued for 12 weeks. Data were analyzed by SPSS version 16, using ANOVA, independent-samples and paired-samples t-test, Wilcoxon signed-rank test, and simple linear regression. In 20 patients in the traditional group, the decrease in SNOT score was observed as 56% after 6 weeks treatment (p=0.001), which is similar to the effect of the first group (classical). Although there was no statically significant difference between the two groups, in clinical terms, the difference in mean systemic symptoms such as confusion with 1.05 (p=0.5) and fatigue with 1.63 (p=0.01) had more improvement in the traditional group, and the difference in mean local symptoms such as nasal congestion with 2.37 (p=0.78) and runny nose with 1.95 (p=0.14) had a more decrease in the classical group. The results of this trial indicate the effectiveness of the Persian Medicine Protocol (including Lavender oil and L.M tea) in the treatment of chronic rhinosinusitis, especially on improving systemic symptoms. Nevertheless more clinical studies are necessary to support the acquired results. This trial was registered at the Iranian Center for Clinical Trials (ID: IRCT2015112425217N1). This research is part of a PhD thesis and is funded by the Vice-Chancellor for Research at Mashhad University of Medical Sciences, Grant No. 931673.


The role of extracellular calcium in drug-induced contractile responses of the rabbit pulmonary artery.
Concentration-response relationships in rabbit pulmonary arteries to norepinephrine (NE), histamine (H) and potassium chloride (KC1) were determined in Ringer's solution containing 2.4, 1.2, 0.6, 0.1 or OmM CaCl2. The EC50 for NE and H did not vary with the extracellular calcium concentrations [Ca2+ ext], but the EC50 for KC1 was greatly increased in 0.1mM Ca2+. Maximum isometric contractile force in response to NE and H was significantly decreased in OmM Ca2+. Addition of 0.1 mM EGTA to the Ca2+-free solution further depressed the responses. KC1-induced maximum tension was decreased in 1.2mM, 0.6mM, 0.1mM and 0mM Ca2+. When a maximum concentration of NE (5 X 10(-5)M), H(5 X 10(-4)M) or KC1 (6 X 10(-2) was added to the bath the initial rapid phase of contraction induced by NE or H appeared to be dependent upon the release of internal Ca2+ from EGTA-sensitive and EGTA-resistant sites. The slow phase of contraction was dependent upon extracellular Ca2+. Both the fast and slow phase of contraction induced by KC1 (60 mM) was dependent upon extracellular Ca2+. In the presence of 0mM Ca2+ the tension response to NE (5 X 10(-5)M) was maintained, but the tension response to H (5 X 10(-4)M) was not maintained.


[Guillain-Barré syndrome].
An exhaustive study of the Guillain-Barré syndrome that was based on the review of 22 clinical records from the "Manuel Ascunce Domenech" Camagüey Provincial Teaching Hospital files is presented. The clinical presentation of the syndrome is pointed out, and results are compared with those from other authors.


Efficient nuclear delivery of antisense oligodeoxynucleotides and selective inhibition of CETP expression by apo E peptide in a human CETP-stably transfected CHO cell line.
N,N-Dipalmitylglycyl-apolipoprotein E (129-169) peptide (dpGapoE) is an efficient gene delivery system for both plasmids and antisense oligodeoxynucleotides (ODNs). To develop a new and efficient approach to the regulation of cholesteryl ester transfer protein (CETP) expression, we used dpGapoE to transfect phosphorothioate antisense ODNs against nucleotides 329 to 349 of human CETP cDNA into a human CETP-stably transfected Chinese hamster ovary (CHO) cell line (hCETP-CHO). After transfection, translocation to the nuclei and concentration in nuclear structures were observed in >95% of the cells at 6 and 12 hours by fluorescence microscopy. No membrane disruption was observed after transfection of ODNs by dpGapoE. Although the translocation stability of phosphorothioate ODNs in the nuclei continued for >48 hours, it had weakened after 24 hours. Cellular CETP mRNA levels gradually declined, and the maximum reduction in the mRNA level (>50%) was observed at 36 hours, after which the mRNA level started to recover. CETP activity in the culture medium declined over 72 hours. The maximum reduction in CETP activity was observed at 36 hours (53.8% of control). Neither CETP mRNA nor CETP activities changed throughout the experiment after the transfection of sense phosphorothioate ODNs delivered by dpGapoE complex or naked antisense ODNs. We conclude that (1) the novel synthetic dpGapoE was a highly effective and nontoxic vehicle for the nuclear delivery of antisense ODNs into hCETP-CHO cells and (2) antisense ODNs selectively inhibited both CETP expression and activity in an hCETP-CHO cell line. This approach may enable gene regulation in vivo and could possibly be used as an antiatherosclerotic agent to alter high density lipoprotein metabolism.


Oncology and functional prognosis are both vital in the surgical treatment of RGCTs around the knee joint.
To explore and provide reasonable surgical options for recurrent giant cell tumors of bone (RGCTs) around the knee joint and compare the pros and cons of extended curettage (EC) and segmental resection (SR). A retrospective analysis was performed of 22 patients (11 male, 11 female; mean age, 34.1 years) with RGCT around the knee joint treated in our hospital between June 2007 and June 2017. Average recurrence time was 14.2 ± 4.7 months. Basic clinical data, including Campanacci grade, lesion location, filler materials, pathological fracture, were recorded. Based on different reoperation methods, patients were divided into the EC and SR groups. Patients were regularly followed up; and recurrence, metastasis, local complications such as osteoarthritis, infection, prosthesis loosening, were recorded. Patient function and surgical efficacy were evaluated using the musculoskeletal tumor society (MSTS) score and Mankin score, respectively. Postoperative recurrence occurred in one patient in both groups, and no difference in the prognosis of oncology was observed between the groups. In the EC group, seven patients developed postoperative complications, but required no surgical treatment, whereas in the SR group, five patients developed postoperative complications and surgical treatment was performed on two patients. There were significant differences in the functional prognosis and surgical efficacy between the two groups; however, the EC group showed more satisfactory results. The oncological and functional prognosis of patients with RGCT around the knee joint is vital. EC should be considered as the first-line treatment, unless the tumors severely invade the surrounding soft tissues or are accompanied by complex fractures with significant displacement leading to no surgical curettage boundary.


[How to assess the economic cost of tropical diseases in a rural community].
Many tropical diseases cause disability and hinder the socio-economic development of the Third World countries where they rage. A method of assessment of the economic cost of these diseases during epidemiological studies has been suggested here. It calls on the following factors: the type of activity of the community, the gross income of the community, the cost of treatment and the types of the disability (degree of disability, length of disability in relation to the period of production, and the degree of men's activity).


Sin recombinase from Staphylococcus aureus: synaptic complex architecture and transposon targeting.
The Sin recombinase from Staphylococcus aureus builds a distinctive DNA-protein synaptic complex to regulate strand exchange. Sin binds at two sites within an 86 basepair (bp) recombination site, resH. We propose that inverted motifs at the crossover site, and tandem motifs at the regulatory site, are recognized by structurally disparate Sin dimers. An essential architectural protein, Hbsu, binds at a discrete central site in resH. Positions of Hbsu-induced DNA deformation coincide with natural targets for Tn552 integration. Remarkably, Sin has the same topological selectivity as Tn3 and gammadelta resolvases. Our model for the recombination synapse has at its core an assembly of four Sin dimers; Hbsu plays an architectural role that is taken by two resolvase dimers in models of the Tn3/gammadelta synapse.


Risk of incarceration between cohorts of veterans with and without mental illness discharged from inpatient units.
This study examined the risk of incarceration among cohorts of veterans treated in the Department of Veterans Affairs (VA) Connecticut Healthcare System. Incarceration rates of persons with and without mental illness were compared and adjusted for various clinical and service utilization variables. Data were compared before and after the closure of over 80% of the Connecticut VA psychiatric inpatient beds in 1996. Data from five annual cohorts of patients (1993-1997) treated in an inpatient unit in the VA Connecticut Healthcare System (N=36,385) were merged with state Department of Correction data. Logistic regression models were used to identify risk factors for incarceration. Bivariate analysis showed that incarceration rates were higher for VA patients with psychiatric disorders and with substance use disorders than for those without such diagnoses, but there were no significant increases in likelihood of incarceration over these years of extensive closures. In multiple logistic regression analysis only diagnoses of substance use disorders and major depression were independently associated with an increased likelihood of incarceration, whereas schizophrenia, personality disorders, and co-occurring psychiatric and substance use disorders were not independently associated with increased likelihood in multivariate analysis. Alcohol and drug problems appeared to account for much of the risk of incarceration among hospitalized veterans during the study period. Unlike in previous studies, schizophrenia and related psychotic disorders were not independently associated with an increased risk of incarceration.


The hepatitis C virus (HCV) NS4B RNA binding inhibitor clemizole is highly synergistic with HCV protease inhibitors.
We recently identified a compound, clemizole hydrochloride, that inhibits NS4B's RNA binding and hepatitis C virus (HCV) replication. Although significant, clemizole's antiviral effect is moderate (50% effective concentration of 8 microM against an HCV genotype 2a clone). We hypothesized that the combination of clemizole with other anti-HCV agents can increase the antiviral effect over that achieved with each drug alone and could also decrease the emergence of viral resistance. Luciferase reporter-linked HCV replication assays were used to study the antiviral effects of drug combinations that included clemizole. Data were analyzed using Loewe additivity and Bliss independence models for synergy, and resistance studies were performed using HCV colony formation assays. Clemizole's antiviral effect was highly synergistic with the HCV protease inhibitors SCH503034 and VX950, without toxicity. In contrast, combinations of clemizole with either interferon, ribavirin, or the nucleoside (NM283) and nonnucleoside (HCV796) HCV polymerase inhibitors were additive. Furthermore, combination of clemizole with SCH503034 decreased the frequency of drug-resistant mutants, compared with treatment with either drug alone. Finally, no cross-resistance to clemizole of SCH503034-resistant mutants (or vice versa) was observed. Clemizole can yield high-level synergy with the protease inhibitor class. Inclusion of clemizole in future anti-HCV cocktails can represent an attractive paradigm for increasing current virologic response rates.


Adherence of Acinetobacter calcoaceticus RAG-1 to human epithelial cells and to hexadecane.
The ability of Acinetobacter calcoaceticus RAG-1 to adhere to human epithelial cells was investigated and compared with its ability to adhere to a test hydrocarbon (hexadecane). RAG-1, a microorganism originally isolated for growth on hydrocarbon, adhered to epithelial cells when grown under conditions which promote its adherence to hexadecane; similarly, RAG-1 cells adhered poorly to epithelial cells when grown under conditions which cause the cells to possess low affinity towards hexadecane. A mutant derived from RAG-1, MR-481, deficient in its ability to adhere to hydrocarbon, was similarly unable to adhere to epithelial cells. RAG-1 adherence to epithelial cells was not blocked by a number of sugars tested. Streptococcus pyogenes, whose adherence to epithelial cells has been previously attributed to hydrophobic interactions, was also able to adhere to hexadecane. Results suggest that hydrophobic interactions mediate adherence of the strains studied to both epithelial cells and hydrocarbon.


Histopathology practice and training in Nigeria – a model.
Contemporary histopathology practice and training in Nigeria have been plagued by the fundamental issue of inadequate exposure to surgical pathology material by both trainees and trainers. This paper critically examines the factors that affect the discipline and profers practical solutions to aid its advancement. This review is based on the authors experience and observations of histopathology practice in Nigeria. The Nigerian health sector is plagued by many ills including poor funding, weak policies, dilapidated structures, disgruntled and frustrated practitioners, amongst others - and pathologists are not immune to all these. In recent times, there has been a proliferation of accredited training centres as well as medical graduates interested in the specialty of histopathology. The busiest histopathology laboratories in the country ascession between 2200 and 5500 surgical samples yearly. Thus there is inadequate exposure by histopathologists and trainee pathologists to surgical materials with the attendant consequences. Many centres still rely principally on routine haematoxylineosin stains. There are no nationally agreed standard reporting formats for most diseases. The development of a deanery or regional system of accredited histopathology laboratories may form the fulcrum for improving the overall quality of histopathological services and training in Nigeria. This will help develop local expertise and ensure adequate exposure to teaching aids and surgical materials. We hope that the proffered solutions will help encourage local pathologists to continue and increase their efforts to raise the profession up to enviable heights.


Lactobacillus johnsonii ameliorates intestinal, extra-intestinal and systemic pro-inflammatory immune responses following murine Campylobacter jejuni infection.
Campylobacter jejuni infections are progressively increasing worldwide. Probiotic treatment might open novel therapeutic or even prophylactic approaches to combat campylobacteriosis. In the present study secondary abiotic mice were generated by broad-spectrum antibiotic treatment and perorally reassociated with a commensal murine Lactobacillus johnsonii strain either 14 days before (i.e. prophylactic regimen) or 7 days after (i.e. therapeutic regimen) peroral C. jejuni strain 81-176 infection. Following peroral reassociation both C. jejuni and L. johnsonii were able to stably colonize the murine intestinal tract. Neither therapeutic nor prophylactic L. johnsonii application, however, could decrease intestinal C. jejuni burdens. Notably, C. jejuni induced colonic apoptosis could be ameliorated by prophylactic L. johnsonii treatment, whereas co-administration of L. johnsonii impacted adaptive (i.e. T and B lymphocytes, regulatory T cells), but not innate (i.e. macrophages and monocytes) immune cell responses in the intestinal tract. Strikingly, C. jejuni induced intestinal, extra-intestinal and systemic secretion of pro-inflammatory mediators (such as IL-6, MCP-1, TNF and nitric oxide) could be alleviated by peroral L. johnsonii challenge. In conclusion, immunomodulatory probiotic species might offer valuable strategies for prophylaxis and/or treatment of C. jejuni induced intestinal, extra-intestinal as well as systemic pro-inflammatory immune responses in vivo.


Biomedical Ph.D. students enrolled in two elite universities in the United kingdom and the United States report adopting multiple learning relationships.
The ability to form multiple learning relationships is a key element of the doctoral learning environment in the biomedical sciences. Of these relationships, that between student and supervisor has long been viewed as key. There are, however, limited data to describe the student perspective on what makes this relationship valuable. In the present study, we discuss the findings of semi-structured interviews with biomedical Ph.D. students from the United Kingdom and the United States to: i) determine if the learning relationships identified in an Australian biomedical Ph.D. cohort are also important in a larger international student cohort; and ii) improve our understanding of student perceptions of value in their supervisory relationships. 32 students from two research intensive universities, one in the United Kingdom (n = 17), and one in the United States (n = 15) were recruited to participate in a semi-structured interview. Verbatim transcripts were transcribed, validated and analysed using a Miles and Huberman method for thematic analysis. Students reported that relationships with other Ph.D. students, post-doctoral scientists and supervisors were all essential to their learning. Effective supervisory relationships were perceived as the primary source of high-level project guidance, intellectual support and confidence. Relationships with fellow students were viewed as essential for the provision of empathetic emotional support. Technical learning was facilitated, almost exclusively, by relationships with postdoctoral staff. These data make two important contributions to the scholarship of doctoral education in the biomedical sciences. Firstly, they provide further evidence for the importance of multiple learning relationships in the biomedical doctorate. Secondly, they clarify the form of a 'valued' supervisory relationship from a student perspective. We conclude that biomedical doctoral programs should be designed to contain a minimum level of formalised structure to promote the development of multiple learning relationships that are perceived as key to student learning.


Nuclear polyadenylate-binding protein.
Polyadenylate-binding activity can be detected in eluates from sodium dodecyl sulfate gels by a nitrocellulose filter-binding assay. Nuclear extracts from rat liver show a single peak of binding activity at 50 to 55 kilodaltons; cytoplasmic extracts show a single peak at 70 to 80 kilodaltons, corresponding to a 75-kilodalton protein previously described. Similar results are obtained with yeast and mouse fibroblasts, indicating a high degree of conservation of both nuclear and cytoplasmic polyadenylate-binding proteins. The activity from rat liver nuclei has been purified 125-fold on the basis of specific binding to polyadenylate and shows two main bands in sodium dodecyl sulfate gels at 53 and 55 kilodaltons.


Structural Consequences of Variation in SARS-CoV-2 B.1.1.7.
New globally circulating SARS-CoV-2 strains are causing concern about evolution of virus transmissibility, fitness and immune evasion mechanisms. A variant emerging from the United Kingdom called SARS-CoV-2 VUI 202012/01, or B.1.1.7, is thought to exhibit increased transmissibility that results from replication 4-10 times faster than the original Wuhan virus (Wuhan-Hu-1). Although this property is suspected to result from a specific mutation in the spike glycoprotein, D614G, there are 9 mutations that distinguish the UK variant B.1.1.7 from Wuhan-Hu-1 yet to be evaluated for functional effects. We asked if mutated positions fixed in UK variant B.1.1.7 may be involved in the virus life cycle, or evasion of the immune response, by modeling the UK variant spike protein and conducting structural analysis of mutations on the spike glycoprotein trimer (protomer) complexed to ACE2. Importantly, 4 out of 9 differences between the UK variant B.1.1.7 and Wuhan-Hu-1 spike protein alter direct intermolecular interactions. N501Y increased affinity between the spike protein and ACE2. The mutations at A570D, D614G and S982A reduced contact between individual chains of the trimeric spike protomer, potentially enhancing cleavage into S1 and S2 subunits, dynamic structural rearrangement and host cell fusion mechanisms. These data suggest that combined characteristics of mutations unique to UK variant B.1.1.7 enable high affinity binding to ACE2 and enhanced replication properties. The D614G mutation, associated with enhanced virus transmissibility, opens a potentially druggable structural pocket at the interface between spike glycoprotein subunits S1 and S2.


The disulfide bonds in glycoprotein E2 of hepatitis C virus reveal the tertiary organization of the molecule.
Hepatitis C virus (HCV), a major cause of chronic liver disease in humans, is the focus of intense research efforts worldwide. Yet structural data on the viral envelope glycoproteins E1 and E2 are scarce, in spite of their essential role in the viral life cycle. To obtain more information, we developed an efficient production system of recombinant E2 ectodomain (E2e), truncated immediately upstream its trans-membrane (TM) region, using Drosophila melanogaster cells. This system yields a majority of monomeric protein, which can be readily separated chromatographically from contaminating disulfide-linked aggregates. The isolated monomeric E2e reacts with a number of conformation-sensitive monoclonal antibodies, binds the soluble CD81 large external loop and efficiently inhibits infection of Huh7.5 cells by infectious HCV particles (HCVcc) in a dose-dependent manner, suggesting that it adopts a native conformation. These properties of E2e led us to experimentally determine the connectivity of its 9 disulfide bonds, which are strictly conserved across HCV genotypes. Furthermore, circular dichroism combined with infrared spectroscopy analyses revealed the secondary structure contents of E2e, indicating in particular about 28% beta-sheet, in agreement with the consensus secondary structure predictions. The disulfide connectivity pattern, together with data on the CD81 binding site and reported E2 deletion mutants, enabled the threading of the E2e polypeptide chain onto the structural template of class II fusion proteins of related flavi- and alphaviruses. The resulting model of the tertiary organization of E2 gives key information on the antigenicity determinants of the virus, maps the receptor binding site to the interface of domains I and III, and provides insight into the nature of a putative fusogenic conformational change.


Chemical ecology of endophytic fungi: origins of secondary metabolites.
Endophytes constitute a remarkably multifarious group of microorganisms ubiquitous in plants and maintain an imperceptible association with their hosts for at least a part of their life cycle. Their enormous biological diversity coupled with their capability to biosynthesize bioactive secondary metabolites has provided the impetus for a number of investigations on endophytes. Here, we highlight the possible current and future strategies of understanding the chemical communication of endophytic fungi with other endophytes (fungi and bacteria) and with their host plants, which might not only allow the discovery and sustainable production of desirable natural products but also other mostly overlooked bioactive secondary metabolites.


Comparison of Costs of Radical Cystectomy vs Trimodal Therapy for Patients With Localized Muscle-Invasive Bladder Cancer.
Earlier studies on the cost of muscle-invasive bladder cancer treatments lack granularity and are limited to 180 days. To compare the 1-year costs associated with trimodal therapy vs radical cystectomy, accounting for survival and intensity effects on total costs. This population-based cohort study used the US Surveillance, Epidemiology, and End Results-Medicare database and included 2963 patients aged 66 to 85 years who had received a diagnosis of clinical stage T2 to T4a muscle-invasive bladder cancer from January 1, 2002, through December 31, 2011. The data analysis was performed from March 5, 2018, through December 4, 2018. Total Medicare costs within 1 year of diagnosis following radical cystectomy vs trimodal therapy were compared using inverse probability of treatment-weighted propensity score models that included a 2-part estimator to account for intrinsic selection bias. Of 2963 participants, 1030 (34.8%) were women, 2591 (87.4%) were white, 129 (4.4%) were African American, and 98 (3.3%) were Hispanic. Median costs were significantly higher for trimodal therapy than radical cystectomy in 90 days ($83 754 vs $68 692; median difference, $11 805; 95% CI, $7745-$15 864), 180 days ($187 162 vs $109 078; median difference, $62 370; 95% CI, $55 581-$69 160), and 365 days ($289 142 vs $148 757; median difference, $109 027; 95% CI, $98 692-$119 363), respectively. Outpatient care, radiology, medication expenses, and pathology/laboratory costs contributed largely to the higher costs associated with trimodal therapy. On inverse probability of treatment-weighted adjusted analyses, patients undergoing trimodal therapy had $136 935 (95% CI, $122 131-$152 115) higher mean costs compared with radical cystectomy 1 year after diagnosis. Compared with radical cystectomy, trimodal therapy was associated with higher costs among patients with muscle-invasive bladder cancer. The differences in costs were largely attributed to medication and radiology expenses associated with trimodal therapy. Extrapolating cost figures resulted in a nationwide excess spending of $468 million for trimodal therapy compared with radical cystectomy for patients who received a diagnosis of bladder cancer in 2017.


Interaction of murine BiP/GRP78 with the DnaJ homologue MTJ1.
The activity of Hsp70 proteins is regulated by accessory proteins, among which the most studied are the members of the DnaJ-like protein family. BiP/GRP78 chaperones the translocation and maturation of secreted and membrane proteins in the endoplasmic reticulum. No DnaJ-like partner has been described so far to regulate the function of mammalian BiP/GRP78. We show here that murine BiP/GRP78 interacts with the lumenal J domain of the murine transmembrane protein MTJ1 (J-MTJ1). J-MTJ1 stimulates the ATPase activity of BiP/GRP78 at stoichiometric concentrations. The C-terminal tail of BiP/GRP78 is not required for the interaction with J-MTJ1, leaving the function of this portion of the molecule still unclear. Physical interactions between J-MTJ1 and BiP/GRP78 are stable and can be abolished by a single histidine --> glutamine substitution in the highly conserved HPD motif shared by all DnaJ-like proteins. The J-MTJ1 fragment, but not the mutant J-MTJ1:H89Q fragment, stimulates the ATPase activity of Escherichia coli DnaK, although at a higher concentration than its genuine partner DnaJ. Full-length DnaJ does not stimulate BiP over the range of concentrations investigated. These results indicate that the J domain of MTJ1 is sufficient for its interaction with BiP/GRP78 and cannot be substituted by E. coli DnaJ.


Searching for optimal mitigation geometries for laser-resistant multilayer high-reflector coatings.
Growing laser damage sites on multilayer high-reflector coatings can limit mirror performance. One of the strategies to improve laser damage resistance is to replace the growing damage sites with predesigned benign mitigation structures. By mitigating the weakest site on the optic, the large-aperture mirror will have a laser resistance comparable to the intrinsic value of the multilayer coating. To determine the optimal mitigation geometry, the finite-difference time-domain method was used to quantify the electric-field intensification within the multilayer, at the presence of different conical pits. We find that the field intensification induced by the mitigation pit is strongly dependent on the polarization and the angle of incidence (AOI) of the incoming wave. Therefore, the optimal mitigation conical pit geometry is application specific. Furthermore, our simulation also illustrates an alternative means to achieve an optimal mitigation structure by matching the cone angle of the structure with the AOI of the incoming wave, except for the p-polarized wave at a range of incident angles between 30° and 45°.


Population pharmacokinetic-pharmacodynamic (PopPK/PD) modeling of risperidone and its active metabolite in Chinese schizophrenia patients.
Risperidone is a second-generation antipsychotic agent commonly used in the treatment of ~ 31.1% of schizophrenia patients in China, it is the most commonly-prescribed antipsychotic agent. Despite the abundant use of risperidone, population pharmacokinetic-pharmacodynamic models of risperidone have not been performed in Chinese schizophrenia patients. The objective of this study was to develop a population pharmacokinetic-pharmacodynamic (PopPK/PD) model to describe the PK behavior and efficacy of risperidone and 9-hydroxy-risperidone (active metabolite) in Chinese patients. Plasma concentration data (702 measurements from 131 patients) and positive and negative syndrome scale (PANSS) scores (258 observations from 56 patients) were analyzed using a nonlinear mixed-effects modeling (NONMEM) approach with first-order conditional estimation with interaction (FOCEI). The influence of potential covariates was evaluated. Model robustness was assessed using external validation, normalized prediction distribution error, nonparametric bootstrap, and visual predictive check approaches. Risperidone concentration data were well described by a one-compartmental model incorporating an additional compartment that refers to the concentration profiles of 9-hydroxy-risperidone. A complex absorption procedure was incorporated into the model to describe the metabolism of risperidone to 9-hydroxy-risperidone in the gastrointestinal (GI) tract. A binomial distribution in the estimated clearance (CL) of risperidone has been identified in our model. Decrease in PANSS score along with total AUC (AUCtotal) of risperidone and 9-hydroxy-risperidone was best characterized by an Emax model with 3 transit compartments describing the delay of drug effect. Considerable differences in PK behavior and drug effect of risperidone have been identified among Chinese extensive metabolizing (EM) and poor metabolizing (PM) patients. This PopPK/PD model may fulfill individualized treatment in clinical practice and may potentially be transferred to other antipsychotic therapies.


Highly Doped Upconversion Nanoparticles for In Vivo Applications Under Mild Excitation Power.
One of the major challenges in using upconversion nanoparticles (UCNPs) is to improve their brightness. This is particularly true for in vivo studies, as the low power excitation is required to prevent the potential photo toxicity to live cells and tissues. Here, we report that the typical NaYF4:Yb0.2,Er0.02 nanoparticles can be highly doped, and the formula of NaYF4:Yb0.8,Er0.06 can gain orders of magnitude more brightness, which is applicable to a range of mild 980 nm excitation power densities, from 0.005 W/cm2 to 0.5 W/cm2. Our results reveal that the concentration of Yb3+ sensitizer ions plays an essential role, while increasing the doping concentration of Er3+ activator ions to 6 mol % only has incremental effect. We further demonstrated a type of bright UCNPs 12 nm in total diameter for in vivo tumor imaging at a power density as low as 0.0027 W/cm2, bringing down the excitation power requirement by 42 times. This work redefines the doping concentrations to fight for the issue of concentration quenching, so that ultrasmall and bright nanoparticles can be used to further improve the performance of upconversion nanotechnology in photodynamic therapy, light-triggered drug release, optogenetics, and night vision enhancement.