Cytotoxic response of the relatively difluoromethylornithine-resistant human lung tumor cell line NCI H157 to the polyamine analogue N1,N8-bis(ethyl)spermidine.
Difluoromethylornithine (DFMO), an enzyme activated irreversible inhibitor of ornithine decarboxylase (ODC), depletes intracellular putrescine, and spermidine (Spd), but not spermine, and generally leads to an inhibition of cell proliferation, without cell death, in both normal and neoplastic cells. This is the case with a culture line of human large cell undifferentiated lung cancer, NCI H157, which will survive indefinitely in DFMO containing medium and ultimately actually grows through the DFMO block. We now provide evidence that a Spd analogue, N1,N8-bis(ethyl)spermidine (BES) also suppresses ODC activity in H157 cells but leads not only to complete depletion of putrescine and Spd but also reduces intracellular spermine to 20-30% of control levels. This depletion of polyamines is accompanied by a rapid decrease in cell proliferation and ultimately cell death. The cell death resulting from BES treatment is in direct contrast to results obtained with DFMO and occurs at concentrations of less than 10 microM, whereas 5 mM DFMO is required to maintain growth inhibition in NCI H157. The observed suppression of ODC activity by BES is consistent with mechanisms by which Spd itself regulates ODC activity. Our data suggest that although both agents, DFMO and BES, interfere with polyamine metabolism, the differential sensitivities to these agents indicate susceptibility to polyamine depletion may be agent and cell type specific. Such differences may be related to the different requirement of individual cell types for polyamines and different regulatory events in polyamine biosynthesis. These differences may be exploitable in the treatment of neoplastic disease with polyamine analogues or inhibitors of polyamine biosynthesis.


Caged Polyprenylated Xanthones in Garcinia hanburyi and the Biological Activities of Them.
The previous phytochemical analyses of Garcinia hanburyi revealed that the main structural characteristic associated with its biological activity is the caged polyprenylated xanthones with a unique 4-oxatricyclo [4.3.1.03,7] dec-2-one scaffold, which contains a highly substituted tetrahydrofuran ring with three quaternary carbons. Based on the progress in research of the chemical constituents, pharmacological effects and modification methods of the caged polyprenylated xanthones, this paper presents a preliminary predictive analysis of their drug-like properties based on the absorption, distribution, metabolism, excretion and toxicity (ADME/T) properties. It was found out that these compounds have very similar pharmacokinetic properties because they possess the same caged xanthone structure, the 9,10-double bond in a,b-unsaturated ketones are critical for the antitumor activity. The author believes that there is an urgent need to seek new breakthroughs in the study of these caged polyprenylated xanthones. Thus, the research on the route of administration, therapeutic effect, structural modification and development of such active ingredients is of great interest. It is hoped that this paper will provide ideas for researchers to develop and utilize the active ingredients derived from natural products.


Hsp72 and Nek6 Cooperate to Cluster Amplified Centrosomes in Cancer Cells.
Cancer cells frequently possess extra amplified centrosomes clustered into two poles whose pseudo-bipolar spindles exhibit reduced fidelity of chromosome segregation and promote genetic instability. Inhibition of centrosome clustering triggers multipolar spindle formation and mitotic catastrophe, offering an attractive therapeutic approach to selectively kill cells with amplified centrosomes. However, mechanisms of centrosome clustering remain poorly understood. Here, we identify a new pathway that acts through NIMA-related kinase 6 (Nek6) and Hsp72 to promote centrosome clustering. Nek6, as well as its upstream activators polo-like kinase 1 and Aurora-A, targeted Hsp72 to the poles of cells with amplified centrosomes. Unlike some centrosome declustering agents, blocking Hsp72 or Nek6 function did not induce formation of acentrosomal poles, meaning that multipolar spindles were observable only in cells with amplified centrosomes. Inhibition of Hsp72 in acute lymphoblastic leukemia cells resulted in increased multipolar spindle frequency that correlated with centrosome amplification, while loss of Hsp72 or Nek6 function in noncancer-derived cells disturbs neither spindle formation nor mitotic progression. Hence, the Nek6-Hsp72 module represents a novel actionable pathway for selective targeting of cancer cells with amplified centrosomes. Cancer Res; 77(18); 4785-96. ©2017 AACR.


"I like to be an informed person but..." negotiating responsibility for treatment decisions in cancer care.
Social expectations surrounding sickness have undergone a transformation in Western welfare states. Emerging discourses about patients' roles and responsibilities do not however always map neatly onto patients' actions, experiences or desires. This paper emerges from a study in Ontario, Canada. Drawing on in-depth interviews with 5 women diagnosed with breast cancer we explore the activity and effort prompted for patients by the routine professional practice of outlining treatment options and encouraging patients to choose between them. We highlight research participants' complex responses to their responsibility for treatment decisions: their accepting, deflecting and reframing and their active negotiation of responsibility with professionals. The literature on treatment decision making typically characterizes people who resist taking an active role as overwhelmed, misinformed about the nature of treatment decisions, or more generally lacking capacity to participate. In this paper we suggest that patients' expressions of ambivalence about making treatment choices can be understood otherwise: as efforts to recast the identities and positions they and their physicians are assigned in the organization of cancer care. We also begin to map key features of this organization, particularly discourses of patient empowerment, and evidence-based medicine.


Development of endometrial cancer following radiation therapy for cervical carcinoma.
The clinical and histologic findings in five cases of endometrial cancer, which developed following radiation therapy for squamous cell carcinoma of the cervix, are described. The mean age at endometrial cancer diagnosis was 69 years and average latency period from initial therapy to development of endometrial carcinoma was 13.4 years. For endometrial cancer, one patient had Stage Ib, one patient had Stage IIIa, two patients had Stage IIIc, and one patient did not undergo laparotomy. The histological types were carcinosarcoma in two patients, endometrioid adenocarcinoma, grade 3 in one patient, and clear cell carcinoma in one patient. All patients died of disease within 33 months of diagnosis. Endometrial cancers that develop after radiation treatment have a preponderance of high-risk histological subtypes, and consequently have a poor prognosis. Long-term follow-up should be mandatory for patients surviving radiation therapy for cervical cancer in order to detect and effectively treat second malignancies.


Curve-skeleton extraction using iterative least squares optimization.
A curve skeleton is a compact representation of 3D objects and has numerous applications. It can be used to describe an object's geometry and topology. In this paper, we introduce a novel approach for computing curve skeletons for volumetric representations of the input models. Our algorithm consists of three major steps: 1) using iterative least squares optimization to shrink models and, at the same time, preserving their geometries and topologies, 2) extracting curve skeletons through the thinning algorithm, and 3) pruning unnecessary branches based on shrinking ratios. The proposed method is less sensitive to noise on the surface of models and can generate smoother skeletons. In addition, our shrinking algorithm requires little computation, since the optimization system can be factorized and stored in the pre-computational step. We demonstrate several extracted skeletons that help evaluate our algorithm. We also experimentally compare the proposed method with other well-known methods. Experimental results show advantages when using our method over other techniques.


Bluetongue virus infection in naïve cattle: Identification of circulating serotypes and associated Culicoides biting midge species in Trinidad.
To better understand risks associated with trading cattle, it is important to know which serotypes of Bluetongue virus (BTV) are circulating within the exporting and importing country. Hence, this study was conducted to identify the circulating serotypes of BTV in Trinidad. Blood samples were collected monthly from sixty BTV- naïve imported cattle over a six month period after their arrival in the country. Virological (PCR and virus isolation) and serological (ELISA) analyses were carried out on the samples and CDC light traps were placed near the cattle enclosure to trap and identify the species of Culicoides biting midges that were present. All of the cattle seroconverted for BTV antibodies within three months of their arrival in the country and real-time reverse transcription PCR (rRT-PCR) detected BTV-RNA in the samples throughout the remainder of the study. The patterns of infection observed in the cattle indicated serial infections with multiple serotypes. A combination of BTV serotype-specific rRT-PCR on the original blood samples and virus isolation followed by serotype-specific rRT-PCR on selected samples, confirmed the presence of BTV serotypes 1, 2, 3, 5, 12 and 17. This is the first report of BTV-2 and BTV-5 in Trinidad. Light-suction traps placed in close proximity to the cattle predominantly trapped Culicoides insignis Lutz 1913 species (96%), with a further six Culicoides species making up the remaining 4% of trapped samples. The circulation of multiple BTV serotypes in Trinidad underlines the need for regular surveillance, which will contribute to the development of risk assessments for trade in livestock.


Shared Decision-Making Training in Internal Medicine: A Multisite Intervention Study.
Research shows that when patients and health care providers share responsibility for clinical decisions, both patient satisfaction and quality of care increase, and resource use decreases. Yet few studies have assessed how to train residents to use shared decision-making (SDM) in their practice. We developed and evaluated a SDM training program in internal medicine. Senior internal medicine residents from 3 hospitals in Switzerland were assessed shortly before and 2 months after completing a program that included a 2-hour workshop and pocket card use in clinical practice. Encounters with standardized patients (SPs) were recorded and SDM performance was assessed using a SDM completeness rating scale (scores ranging from 0 to 100), a self-reported questionnaire, and SPs rating the residents. Of 39 eligible residents, 27 (69%) participated. The mean (SD) score improved from 65 (SD 13) to 71 (SD 12; effect size [ES] 0.53; P = .011). After training, participants were more comfortable with their SDM-related knowledge (ES 1.42, P < .001) and skills (ES 0.91, P < .001), and with practicing SDM (ES 0.96, P < .001). Physicians applied SDM concepts more often in practice (ES 0.71, P = .001), and SPs felt more comfortable with how participants discussed their care (ES 0.44, P = .031). The SDM training program improved the competencies of internal medicine residents and promoted the use of SDM in clinical practice. The approach may be of interest for teaching SDM to residents in other disciplines and to medical students.


Iron, siderophores, and the pursuit of virulence: independence of the aerobactin and enterochelin iron uptake systems in Escherichia coli.
Many strains of Escherichia coli isolated from extraintestinal infections of humans and domestic animals are able to synthesize two siderophores, aerobactin and enterochelin. Although aerobactin has a dramatically lower affinity for iron than enterochelin, it has been shown to provide a significant selective advantage for bacterial growth in conditions of iron limitation, such as in the body fluids and tissues of an infected animal. We have used streptonigrin, which is bactericidal in the presence of iron, as a probe to determine levels of free intracellular iron during bacterial growth promoted by the two siderophores. A strain with only enterochelin remained sensitive to the bactericidal action of streptonigrin, suggesting that assimilated iron was contributed to an intracellular pool from which the rate of its withdrawal for growth is probably concentration dependent. On the other hand, a strain that synthesized aerobactin alone became resistant to streptonigrin, indicating that iron complexed with aerobactin was not made accessible to streptonigrin and suggesting that it may be channeled directly to where it is required for growth. Aerobactin, probably because it is repeatedly reusable, efficiently stimulated bacterial growth at external concentrations some 500-fold lower than those of enterochelin. Moreover, the effective concentration, and thus the siderophore activity, of enterochelin but not of aerobactin was significantly reduced by the presence of human serum in the medium. Differential regulation of the genetic determinants of the two siderophores resulted in preferential induction of the aerobactin system in the presence of unsaturated levels of transferrin and lactoferrin.


Regulation of the autophagy system during chronic contractile activity-induced muscle adaptations.
Skeletal muscle is adaptable to exercise stimuli via the upregulation of mitochondrial biogenesis, and recent studies have suggested that autophagy also plays a role in exercise-induced muscle adaptations. However, it is still obscure how muscle regulates autophagy over the time course of training adaptations. This study examined the expression of autophagic proteins in skeletal muscle of rats exposed to chronic contractile activity (CCA; 6 h/day, 9V, 10 Hz continuous, 0.1 msec pulse duration) for 1, 3, and 7 days (n = 8/group). CCA-induced mitochondrial adaptations were observed by day 7, as shown by the increase in mitochondrial proteins (PGC-1α, COX I, and COX IV), as well as COX activity. Notably, the ratio of LC3 II/LC3 I, an indicator of autophagy, decreased by day 7 largely due to a significant increase in LC3 I. The autophagic induction marker p62 was elevated on day 3 and returned to basal levels by day 7, suggesting a time-dependent increase in autophagic flux. The lysosomal system was upregulated early, prior to changes in mitochondrial proteins, as represented by increases in lysosomal system markers LAMP1, LAMP2A, and MCOLN1 as early as by day 1, as well as TFEB, a primary regulator of lysosomal biogenesis and autophagy flux. Our findings suggest that, in response to chronic exercise, autophagy is upregulated concomitant with mitochondrial adaptations. Notably, our data reveal the surprising adaptive plasticity of the lysosome in response to chronic contractile activity which enhances muscle health by providing cells with a greater capacity for macromolecular and organelle turnover.


TRPV1, CGRP and SP in scalp arteries of patients suffering from chronic migraine.
The transient receptor potential vanilloid type-1 receptor (TRPV1) and the neuropeptides calcitonin gene-related peptide (CGRP) and substance P (SP) appear to be differently involved in migraine pain. A role of neurovascular scalp structures is also suggested by several data. We performed a quantitative study of TRPV1-like immunoreactive (LI), CGRP-LI and SP-LI innervation of scalp arterial samples from patients affected with chronic migraine (CM). Short segments of scalp arteries were collected from 17 participants undergoing vascular surgery for treatment-resistant CM and from 6 controls who underwent neurosurgery for various indications. The immunoreactivity of the arterial innervation to TRPV1, CGRP, SP and to the pan-neuronal marker protein gene product 9.5 (PGP9.5) was examined. Immunoreactive nerve fibres in vessel cross-sections were quantified by computerised image analysis. A significant increase of TRPV1-LI nerve fibres was found in the arterial wall from CM compared with control patients (p<0.05), while no significant difference was found for CGRP and SP. This study yields the first evidence for the existence of a TRPV1-LI innervation in human scalp arteries and provides the first quantitative assessment of the TRPV1-LI, CGRP-LI and SP-LI innervation of those vessels. The increase of TRPV1-LI periarterial nociceptive fibres of scalp arteries may represent, at least in some participants, a structural condition favouring CM (and possibly migraine), for example, by causing a higher sensitivity to algogenic agents.


Effect of the HIV protease inhibitor amprenavir on the growth and differentiation of primary gingival epithelium.
HIV-positive patients taking antiretroviral drugs, including protease inhibitors, have shown a significant increase in the development of oral complications; these complications are a major health issue for affected patients. The effect of these drugs on oral epithelium growth and differentiation is presently unknown. In this study, we explore for the first time the effect of the HIV protease inhibitor amprenavir on gingival epithelium growth and differentiation. Organotypic (raft) cultures of gingival keratinocytes were established and the raft cultures treated with a range of amprenavir concentrations. Haematoxylin and eosin staining was performed to examine the effect of amprenavir on gingival epithelium growth and stratification. The raft cultures were also immunohistochemically analysed to determine the effect of amprenavir on the expression of key differentiation and proliferation markers, including cytokeratins, proliferating cell nuclear antigen (PCNA) and cyclin A. Amprenavir severely inhibited the growth of gingival epithelium when the drug was present throughout the growth period of the tissue. When the drug was added at day 8, amprenavir treatment altered the proliferation and differentiation of gingival keratinocytes. Expression of the cytokeratins 5, 14, 6 and 10, PCNA and cyclin A was increased; their expression pattern was also altered over time in treated rafts. Biochemically, the tissue exhibited characteristics of increased proliferation in the suprabasal layers of amprenavir-treated tissue. Our results suggest that amprenavir treatment deregulates the cell cycle/proliferation and differentiation pathways, resulting in abnormal epithelial repair and proliferation. Our system could be developed as a potential model for studying the effects of highly active antiretroviral therapy in vitro.


Glucose transporter expression differs between bovine monocyte and macrophage subsets and is influenced by milk production.
The peripartal period of dairy cows is characterized by negative energy balance and higher incidences of infectious diseases such as mastitis or metritis. With the onset of lactation, milk production is prioritized and large amounts of glucose are transported into the mammary gland. Decreased overall energy availability might impair the function of monocytes acting as key innate immune cells, which give rise to macrophages and dendritic cells and link innate and adaptive immunity. Information on glucose requirements of bovine immune cells is rare. Therefore, this study aims to evaluate glucose transporter expression of the 3 bovine monocyte subsets (classical, intermediate, and nonclassical monocytes) and monocyte-derived macrophages and to identify influences of the peripartal period. Blood samples were either collected from nonpregnant healthy cows or from 16 peripartal German Holstein cows at d -14, +7, and +21 relative to parturition. Quantitative real-time PCR was applied to determine mRNA expression of glucose transporters (GLUT) 1, GLUT3, and GLUT4 in monocyte subsets and monocyte-derived macrophages. The low GLUT1 and GLUT3 expression in nonclassical monocytes was unaltered during differentiation into macrophages, whereas in classical and intermediate monocytes GLUT expression was downregulated. Alternatively activated M2 macrophages consumed more glucose compared with classically activated M1 macrophages. The GLUT4 mRNA was only detectable in unstimulated macrophages. Neither monocytes nor macrophages were insulin responsive. In the peripartum period, monocyte GLUT1 and GLUT3 expression and the GLUT3/GLUT1 ratio were negatively correlated with lactose production. The high-affinity GLUT3 transporter appears to be the predominant glucose transporter on bovine monocytes and macrophages, especially in the peripartal period when blood glucose levels decline. Glucose transporter expression in monocytes is downregulated as a function of lactose production, which might impair monocyte to macrophage differentiation.


Short-term versus long-term triple antithrombotic therapy for patients with coronary stents and requiring oral anticoagulation: a meta-analysis of randomized clinical trials.
The cornerstone therapy for patients with coronary stents is dual antiplatelet therapy (DAPT). In 5-10% of these patients, oral anticoagulation (OAC) is clearly indicated in addition to DAPT. However, the optimal duration of this triple antithrombotic therapy (TAT) remains uncertain. Scientific databases and websites were searched for randomized clinical trials (RCTs). RCTs were included if patients undergoing coronary stent placements with additional indications of chronic OAC were randomly assigned to either short-term TAT or long-term TAT. Short-term TAT was defined as no more than 6 weeks of TAT, and long-term TAT was defined as 6-12 months of TAT RESULTS: Using data from three RCTs and 1883 patients, short-term TAT was associated with decreased rates of major adverse cardiovascular events, cardiac mortality, all-cause mortality, and any-bleeding events compared to long-term TAT, but similar rates of myocardial infarction, stroke, stent thrombosis, and thrombolysis in myocardial infarction major bleeding. Furthermore, in subgroup analysis, short-term TAT was associated with decreased rates of major adverse cardiovascular events, cardiac mortality, all-cause mortality, and any-bleeding compared to 12-month TAT, but similar rates compared to 6-month TAT. In patients who require chronic OAC therapy and undergo coronary stent placement, short-term TAT was associated with better efficacy and safety outcomes compared to long-term TAT.


Computer-based Learning of Neuroanatomy: A Longitudinal Study of Learning, Transfer, and Retention.
A longitudinal experiment was conducted to evaluate the effectiveness of new methods for learning neuroanatomy with computer-based instruction. Using a 3D graphical model of the human brain, and sections derived from the model, tools for exploring neuroanatomy were developed to encourage adaptive exploration. This is an instructional method which incorporates graphical exploration in the context of repeated testing and feedback. With this approach, 72 participants learned either sectional anatomy alone or whole anatomy followed by sectional anatomy. Sectional anatomy was explored either with perceptually continuous navigation through the sections or with discrete navigation (as in the use of an anatomical atlas). Learning was measured longitudinally to a high performance criterion. Subsequent tests examined transfer of learning to the interpretation of biomedical images and long-term retention. There were several clear results of this study. On initial exposure to neuroanatomy, whole anatomy was learned more efficiently than sectional anatomy. After whole anatomy was mastered, learners demonstrated high levels of transfer of learning to sectional anatomy and from sectional anatomy to the interpretation of complex biomedical images. Learning whole anatomy prior to learning sectional anatomy led to substantially fewer errors overall than learning sectional anatomy alone. Use of continuous or discrete navigation through sectional anatomy made little difference to measured outcomes. Efficient learning, good long-term retention, and successful transfer to the interpretation of biomedical images indicated that computer-based learning using adaptive exploration can be a valuable tool in instruction of neuroanatomy and similar disciplines.


Billing, coding, and practice management: a primer for today's reproductive medicine professional.
Despite years of recognition that many physicians are woefully unprepared to face challenges regarding the business of medicine, marginal progress has been made. In this piece, we aim to provide the contemporary reproductive medicine physician with an understanding of billing, coding, and, most importantly, cost containment for a typical fertility practice. It is critical for modern practices to not forego hard-earned revenue to insurance companies or not be aware of critical rules and regulations. While running a successful fertility practice requires good medical care, a profitable practice is necessary for overall long-term success. This article provides a brief history of medical insurance and billing, explains the process of updating billing codes, and reviews the revenue cycle, cost containment, and contract negotiations with insurance companies.


CCR10+ ILC2s with ILC1-like properties exhibit a protective function in severe allergic asthma.
We previously showed that patients with severe allergic asthma have high numbers of circulating ILC2s expressing CCR10. Herein, CCR10+ ILC2s were further analyzed in the blood of healthy individuals or patients with allergic and non-allergic asthma. Characteristics of human CCR10+ and CCR10- ILC2s were assessed by flow cytometry as well as single-cell multiplex RT-qPCR. The role of CCR10+ ILC2s in asthma pathophysiology was studied in allergen-treated mice. When compared to healthy controls, CCR10+ ILC2s are enriched in the blood of both allergic and non-allergic severe asthmatic patients, and these cells are recruited to the lungs. Plasma concentrations of the CCR10 ligand CCL27 are significantly increased in severe asthmatics when compared to non-asthmatic patients. CCR10+ ILC2s secrete little TH 2 cytokines, but exhibit ILC1-like properties, including a capacity to produce IFN-γ. Also, single-cell analysis reveals that the CCR10+ ILC2 subset is enriched in cells expressing amphiregulin. CCR10+ ILC2 depletion, as well as blocking of IFN-γ activity, exacerbates airway hyperreactivity in allergen-challenged mice, providing evidence for a protective role of these cells in allergic inflammation. Frequencies of circulating CCR10+ ILC2s and CCL27 plasma concentrations represent candidate markers of asthma severity. The characterization of CCR10+ ILC2s in human samples and in mouse asthma models suggests that these cells downregulate allergic inflammation through IFN-γ production.


Detecting panic disorder in emergency department chest pain patients: a validated model to improve recognition.
To develop and validate a detection model to improve the probability of recognizing panic disorder in patients consulting the emergency department for chest pain. Through logistic regression analysis, demographic, self-report psychological, and pain variables were explored as factors predictive of the presence of panic disorder in 180 consecutive patients consulting an emergency department with a chief complaint of chest pain. The detection model was then prospectively validated on a sample of 212 patients recruited following the same procedure. Panic-agoraphobia (Agoraphobia Cognitions Questionnaire, Mobility Inventory for Agoraphobia), chest pain quality (Short Form McGill Pain Questionnaire), pain loci, and gender variables were the best predictors of the presence of panic disorder. These variables correctly classified 84% of chest pain subjects in panic and non-panic disorder categories. Model properties: sensitivity 59%; specificity 93%; positive predictive power 75%; negative predictive power 87% at a panic disorder sample prevalence of 26%. The model correctly classified 73% of subjects in the validation phase. The scales in this model take approximately ten minutes to complete and score. It may improve upon current physician recognition of panic disorder in patients consulting for chest pain.


The cost effectiveness of an early transition from hospital to nursing home for stroke patients: design of a comparative study.
As the incidence of stroke has increased, its impact on society has increased accordingly, while it continues to have a major impact on the individual. New strategies to further improve the quality, efficiency and logistics of stroke services are necessary. Early discharge from hospital to a nursing home with an adequate rehabilitation programme could help to optimise integrated care for stroke patients.The objective is to describe the design of a non-randomised comparative study evaluating early admission to a nursing home, with multidisciplinary assessment, for stroke patients. The study is comprised of an effect evaluation, an economic evaluation and a process evaluation. The design involves a non-randomised comparative trial for two groups. Participants are followed for 6 months from the time of stroke. The intervention consists of a redesigned care pathway for stroke patients. In this care pathway, patients are discharged from hospital to a nursing home within 5 days, in comparison with 12 days in the usual situation. In the nursing home a structured assessment takes place, aimed at planning adequate rehabilitation. People in the control group receive the usual care. The main outcome measures of the effect evaluation are quality of life and daily functioning. In addition, an economic evaluation will be performed from a societal perspective. A process evaluation will be carried out to evaluate the feasibility of the intervention as well as the experiences and opinions of patients and professionals. The results of this study will provide information about the cost effectiveness of the intervention and its effects on clinical outcomes and quality of life. Relevant strengths and weaknesses of the study are addressed in this article. Current Controlled Trails ISRCTN58135104.


Beta-galactosidase from termination and deletion mutant strains.
beta-Galactosidase fragments were isolated from strains of Escherichia coli with mutations in the lacZ gene. The polypeptide obtained from a termination mutant (lacZNG125) appeared to be the intact gene product, containing the first half of the beta-galactosidase amino acid sequence. From an internal deletion mutant strain (lacZU163), an aggregate was obtained of several partially degraded polypeptides. Each of these was smaller than predicted from genetic data for the fragment. Introduction of the lacZU163 mutation into a protein degradation-deficient strain (Deg(-)) resulted in the protection of the amino-terminal region of the protein. Some of the BrCN peptides from the U163 polypeptides were separated and identified. From such experiments it was shown that in both Deg(-) and Deg(+) strains the COOH-terminal region is rapidly degraded. This indicates that the complete gene product of lacZU163 has not been detected. The use of genetically defined enzyme fragments in studying structure-function relationships and in determination of primary structure is discussed.


Threshold position control of arm movement with anticipatory increase in grip force.
The grip force holding an object between fingers usually increases before or simultaneously with arm movement thus preventing the object from sliding. We experimentally analyzed and simulated this anticipatory behavior based on the following notions. (1) To move the arm to a new position, the nervous system shifts the threshold position at which arm muscles begin to be recruited. Deviated from their activation thresholds, arm muscles generate activity and forces that tend to minimize this deviation by bringing the arm to a new position. (2) To produce a grip force, with or without arm motion, the nervous system changes the threshold configuration of the hand. This process defines a threshold (referent) aperture (R(a)) of appropriate fingers. The actual aperture (Q(a)) is constrained by the size of the object held between the fingers whereas, in referent position R(a), the fingers virtually penetrate the object. Deviated by the object from their thresholds of activation, hand muscles generate activity and grip forces in proportion to the gap between the Q(a) and R(a). Thus, grip force emerges since the object prevents the fingers from reaching the referent position. (3) From previous experiences, the system knows that objects tend to slide off the fingers when arm movements are made and, to prevent sliding, it starts narrowing the referent aperture simultaneously with or somewhat before the onset of changes in the referent arm position. (4) The interaction between the fingers and the object is accomplished via the elastic pads on the tips of fingers. The pads are compressed not only due to the grip force but also due to the tangential inertial force ("load") acting from the object on the pads along the arm trajectory. Compressed by the load force, the pads move back and forth in the gap between the finger bones and object, thus inevitably changing the normal component of the grip force, in synchrony with and in proportion to the load force. Based on these notions, we simulated experimental elbow movements and grip forces when subjects rapidly changed the elbow angle while holding an object between the index finger and the thumb. It is concluded that the anticipatory increase in the grip force with or without correlation with the tangential load during arm motion can be explained in neurophysiological and biomechanical terms without relying on programming of grip force based on an internal model.


Effect of pramipexole in treatment of resistant restless legs syndrome.
To report the results of an open-label trial with a dopaminergic agent, pramipexole, in patients with treatment-resistant restless legs syndrome (RLS). We studied the response to pramipexole in a consecutive series of 16 patients with symptomatic RLS who had previously experienced failure with other dopaminergic therapies. Patients assessed their posttreatment change in symptoms of RLS on a visual analog scale and indicated drug-related side effects with use of a checklist. With a mean dose of pramipexole of 0.3 mg, most patients reported clinically significant improvement. From 2 to 3 months after initiation of pramipexole therapy, nocturnal leg restlessness, involuntary leg movements, and insomnia had decreased in 12, 10, and 11 patients, respectively. The most frequent adverse effects were fatigue and stiffness, which occurred in a third of the patients. Overall, the drug was well tolerated. On the basis of these findings, we propose that pramipexole, a D2 subgroup receptor agonist, is an effective agent for treatment of RLS.


Robotic-assisted McKeown esophagectomy.
Over the last few decades, minimally invasive approaches have gained in popularity for the surgical treatment of esophageal cancer, with the hopes of reducing the morbidity and mortality of this formidable procedure. The traditional McKeown approach includes thoracic esophageal dissection, abdominal mobilization of the gastric conduit and a cervical anastomosis. It presents several advantages over the Ivor Lewis approach: it is suitable for proximal tumors, allows a 3-field lymph node dissection, and avoids the morbidity associated with an intra-thoracic anastomotic leak. Here, we discuss the technical aspects of robotic-assisted minimally invasive McKeown esophagectomy (McKeown RAMIE).


Long-chain ceramide is a potent inhibitor of the mitochondrial permeability transition pore.
The sphingolipid ceramide has been implicated in mediating cell death that is accompanied by mitochondrial functional alterations. Moreover, ceramide has been shown to accumulate in mitochondria upon induction of apoptotic processes. In this study, we sought to evaluate the effects of natural, highly hydrophobic long-chain ceramides on mitochondrial function in vitro. Ceramide in a dodecane/ethanol delivery system inhibited the opening of the mitochondrial permeability transition pore (PTP) induced by either oxidative stress, SH group cross-linking, or high Ca2+ load, suggesting that the inhibitory point is at a level at which major PTP regulatory pathways converge. Moreover, ceramide had no effect on well known mitochondrial components that modulate PTP activity, such as cyclophilin D, voltage-dependent anion channel, adenine nucleotide transporter, and ATP synthase. The inhibitory effect of ceramide on PTP was not stereospecific, nor was there a preference for ceramide over dihydroceramide. However, the effect of ceramide on PTP was significantly influenced by the fatty acid moiety chain length. These studies are the first to show that long-chain ceramide can influence PTP at physiologically relevant concentrations, suggesting that it is the only known potent natural inhibitor of PTP. These results suggest a novel mechanism of ceramide regulation of mitochondrial function.


Polar lipids of Burkholderia pseudomallei induce different host immune responses.
Melioidosis is a disease in tropical and subtropical regions of the world that is caused by Burkholderia pseudomallei. In endemic regions the disease occurs primarily in humans and goats. In the present study, we used the goat as a model to dissect the polar lipids of B. pseudomallei to identify lipid molecules that could be used for adjuvants/vaccines or as diagnostic tools. We showed that the lipidome of B. pseudomallei and its fractions contain several polar lipids with the capacity to elicit different immune responses in goats, namely rhamnolipids and ornithine lipids which induced IFN-γ, whereas phospholipids and an undefined polar lipid induced strong IL-10 secretion in CD4(+) T cells. Autologous T cells co-cultured with caprine dendritic cells (cDCs) and polar lipids of B. pseudomallei proliferated and up-regulated the expression of CD25 (IL-2 receptor) molecules. Furthermore, we demonstrated that polar lipids were able to up-regulate CD1w2 antigen expression in cDCs derived from peripheral blood monocytes. Interestingly, the same polar lipids had only little effect on the expression of MHC class II DR antigens in the same caprine dendritic cells. Finally, antibody blocking of the CD1w2 molecules on cDCs resulted in decreased expression for IFN-γ by CD4(+) T cells. Altogether, these results showed that polar lipids of B. pseudomallei are recognized by the caprine immune system and that their recognition is primarily mediated by the CD1 antigen cluster.


Examining the Dynamic Structure of Daily Internalizing and Externalizing Behavior at Multiple Levels of Analysis.
Psychiatric diagnostic covariation suggests that the underlying structure of psychopathology is not one of circumscribed disorders. Quantitative modeling of individual differences in diagnostic patterns has uncovered several broad domains of mental disorder liability, of which the Internalizing and Externalizing spectra have garnered the greatest support. These dimensions have generally been estimated from lifetime or past-year comorbidity patters, which are distal from the covariation of symptoms and maladaptive behavior that ebb and flow in daily life. In this study, structural models are applied to daily diary data (Median = 94 days) of maladaptive behaviors collected from a sample (N = 101) of individuals diagnosed with personality disorders (PDs). Using multilevel and unified structural equation modeling, between-person, within-person, and person-specific structures were estimated from 16 behaviors that are encompassed by the Internalizing and Externalizing spectra. At the between-person level (i.e., individual differences in average endorsement across days) we found support for a two-factor Internalizing-Externalizing model, which exhibits significant associations with corresponding diagnostic spectra. At the within-person level (i.e., dynamic covariation among daily behavior pooled across individuals) we found support for a more differentiated, four-factor, Negative Affect-Detachment-Hostility-Disinhibition structure. Finally, we demonstrate that the person-specific structures of associations between these four domains are highly idiosyncratic.


Novel Anti-TM4SF1 Antibody-Drug Conjugates with Activity against Tumor Cells and Tumor Vasculature.
Antibody-drug conjugates (ADC) represent a promising therapeutic modality for managing cancer. Here, we report a novel humanized ADC that targets the tetraspanin-like protein TM4SF1. TM4SF1 is highly expressed on the plasma membranes of many human cancer cells and also on the endothelial cells lining tumor blood vessels. TM4SF1 is internalized upon interaction with antibodies. We hypothesized that an ADC against TM4SF1 would inhibit cancer growth directly by killing cancer cells and indirectly by attacking the tumor vasculature. We generated a humanized anti-human TM4SF1 monoclonal antibody, v1.10, and armed it with an auristatin cytotoxic agent LP2 (chemical name mc-3377). v1.10-LP2 selectively killed cultured human tumor cell lines and human endothelial cells that express TM4SF1. Acting as a single agent, v1.10-LP2 induced complete regression of several TM4SF1-expressing tumor xenografts in nude mice, including non-small cell lung cancer and pancreas, prostate, and colon cancers. As v1.10 did not react with mouse TM4SF1, it could not target the mouse tumor vasculature. Therefore, we generated a surrogate anti-mouse TM4SF1 antibody, 2A7A, and conjugated it to LP2. At 3 mpk, 2A7A-LP2 regressed several tumor xenografts without noticeable toxicity. Combination therapy with v1.10-LP2 and 2A7A-LP2 together was more effective than either ADC alone. These data provide proof-of-concept that TM4SF1-targeting ADCs have potential as anticancer agents with dual action against tumor cells and the tumor vasculature. Such agents could offer exceptional therapeutic value and warrant further investigation. Mol Cancer Ther; 14(8); 1868-76. ©2015 AACR.


Liver Transplantation for Homozygous Familial Hypercholesterolemia.
Pharmacological treatments to decrease low-density lipoprotein (LDL) cholesterol (LDL-C) have limited effects on patients with homozygous familial hypercholesterolemia (HoFH). Since LDL receptors are located mainly in the liver, liver transplantation is considered to be the only way to correct the hepatic cholesterol metabolism abnormalities in HoFH. Liver transplantations, including those combined with heart transplantation, for HoFH have been increasing since 1984, making this a globally established therapeutic option for HoFH. Plasma LDL-C is reported to be dramatically lowered, by 80%, after transplantation, with the rapid regression of cutaneous and tendinous xanthomas. However, long-term cardiovascular benefits remain unclear. The major concerns about liver transplantation include surgical complications, the need for lifelong immunosuppressive therapy, and rejection. In addition, organ transplantations from deceased donors are extremely rare in Japan. We experienced two pediatric siblings with HoFH who received living-donor liver transplantations from their heterozygous parents. Their plasma LDL-C levels decreased immediately and stabilized at approximately 200 mg/dL. Both developed normally with the administration of lipid-lowering medications and have been free of severe problems for more than 10 years, to date, since transplantation. In Japan, where the shortage of deceased donors is critical, the combination of living-donor liver transplant from a heterozygous donor, that is, usually a parent, and medication is regarded as a valid therapeutic option for HoFH. Further studies and clinical experience are required to establish liver transplantation as a safe and effective treatment for HoFH.


Nanoparticulate formulations for paclitaxel delivery across MDCK cell monolayer.
Paclitaxel appears to be a potential substrate of the multidrug resistance protein p-glycoprotein, thus preventing itself from entry into the brain and penetrating blood-brain barrier poorly. In this study, the main objective was to design paclitaxel formulation using PLGA-based nanoparticles with different additives and surface coatings to facilitate the paclitaxel transport through MDCK cell monolayer. PLGA-based nanoparticles of around 200 nm without and with additives and surface coatings were developed by direct dialysis. The transendothelial electrical resistance (TEER) variation of MDCK cell monolayer on the cell inserts imposed by paclitaxel-loaded nanoparticles with and without additives was investigated. (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, a tetrazole) (MTT) assay was used to quantify the cell viability of C6 glioma cells after administration of formulations on the topical side. Investigations showed that particles with additives were able to enhance cellular uptake more than surface-coated particles. TEER values dropped upon the introduction of paclitaxel-loaded PLGA nanoparticles to the cell inserts. After incubation for 24 h, the cell viability of C6 glioma cells in the wells treated with nanoparticles was lower than that of the control wells without particles. Taken together, PLGA nanoparticles with vitamin E TPGS and polysorbate 80 as additives were successfully obtained as paclitaxel formulations, demonstrating great potential for the delivery of paclitaxel through MDCK cell monolayer in vitro.


Salivary Testosterone Levels and Health Status in Men and Women in the British General Population: Findings from the Third National Survey of Sexual Attitudes and Lifestyles (Natsal-3).
Salivary T (Sal-T) measurement by liquid chromatography-tandem mass spectroscopy resents the opportunity to examine health correlates of Sal-T in a large-scale population survey. This study sought to examine associations between Sal-T and health-related factors in men and women age 18-74 years. Morning saliva samples were obtained from participants in a cross-sectional probability-sample survey of the general British population (Natsal-3). Self-reported health and lifestyle questions were administered as part of a wider sexual health interview. Study participants included 1599 men and 2123 women. Sal-T was measured using liquid chromatography-tandem mass spectroscopy. Linear regression was used to examine associations between health factors and mean Sal-T. In men, mean Sal-T was associated with a range of health factors after age adjustment, and showed a strong independent negative association with body mass index (BMI) in multivariable analysis. Men reporting cardiovascular disease or currently taking medication for depression had lower age-adjusted Sal-T, although there was no association with cardiovascular disease after adjustment for BMI. The decline in Sal-T with increasing age remained after adjustment for health-related factors. In women, Sal-T declined with increasing age; however, there were no age-independent associations with health-related factors or specific heath conditions with the exception of higher Sal-T in smokers. Sal-T levels were associated, independently of age, with a range of self-reported health markers, particularly BMI, in men but not women. The findings support the view that there is an age-related decline in Sal-T in men and women, which cannot be explained by an increase in ill health. Our results demonstrate the potential of Sal-T as a convenient measure of tissue androgen exposure for population research.


Is dietary fat a major determinant of body fat?
The percentage of energy from dietary fat is widely believed to be an important determinant of body fat, and several mechanisms have been proposed to account for such a relation. Comparisons of both diets and the prevalence of obesity between affluent and poor countries have been used to support a causal association, but these contrasts are seriously confounded by differences in physical activity and food availability. Within areas of similar economic development, regional intake of fat and prevalence of obesity have not been positively correlated. Randomized trials are the preferable method to evaluate the effect of dietary fat on adiposity, and are feasible because the number of subjects needed is not large. In short-term trials, a modest reduction in body weight is typically seen in individuals randomly assigned to diets with a lower percentage of energy from fat. However, compensatory mechanisms appear to operate because in trials lasting > or = 1 y, fat consumption within the range of 18-40% of energy appears to have little if any effect on body fatness. Moreover, within the United States, a substantial decline in the percentage of energy from fat consumed during the past two decades has corresponded with a massive increase in obesity. Diets high in fat do not appear to be the primary cause of the high prevalence of excess body fat in our society, and reductions in fat will not be a solution.


The prevalence and risk factors of kidney disease in first degree relatives of patients with ESRD treatment - single center study.
To determine the prevalence and risk factors of kidney disease in first degree relatives of patients undergoing treatment for end-stage renal disease. The prospective, cross-sectional study was conducted at the Sindh Institute of Urology and Transplantation, Karachi, from May 1 to July 31, 2021, and comprised patients undergoing treatment for end-stage renal disease at the pre-transplant out-patients clinic, and their first degree relatives. Risk factors of chronic kidney disease, including age, gender, body mass index, hypertension, diabetes mellitus, and the causes of index cases were investigated alongside proteinuria, haematuria and estimated glomerular filtration rate. Diagnosis was made according to the criteria of the National Kidney Foundation-Kidney Disease Outcomes Quality Initiative. The participants were divided chronic kidney disease group 1, and healthy group 2. The risk factors were compared between the groups. Data was analysed using SPSS 22. Of the 1,406 subjects assessed, 266(19%) were ESRD patients; 175(65.8%) males and 91(34.2%) females with mean age 34.04±11.19 years. 1,140(81%) first degree relatives of these 266 ESRD patient were assessed; 595(52.2%) males and 545(47.8%) females with mean age 36.78±13.76 years. Among the relatives, 146 (12.8%) had chronic kidney disease out of which 54 (4.7%) were already aware of their underlying disease. Older age, hypertension and diabetes mellitus were among the risk factors for chronic kidney disease (p<0.05), while gender was not significantly different between groups 1 and 2 (p>0.05). The relatives of index cases with underlying stone disease were at higher risk of haematuria 39(22.4%), whereas the relatives of index cases with chronic glomerulonephritis were at higher risk of proteinuria 67(28.03%) compared to index cases of other kinds (p<0.05). Screening in the high-risk population might help to identify early chronic kidney disease patients for making suitable interventions to prevent disease progression.


Nano electro-mechanical optoelectronic tunable VCSEL.
We report a novel electrostatic actuated nano-electromechanical optoelectronic (NEMO) tunable vertical-cavity surface-emitting laser (VCSEL) centered at 850 nm. By integrating a movable, single-layer (230 nm), high-index-contrast subwavelength grating (HCG) as the VCSEL top mirror, single mode emission (SMSR >40 dB) and continuous wavelength tuning (~2.5 nm) was obtained at room temperature under CW operation. The small footprint of HCG enables the scaling down of each of the cantilever dimensions by a factor of 10, leading to 1000 times reduction in mass, which potentially increases the mechanical resonant frequency and tuning speed.


[Prenatal diagnosis of congenital diaphragmatic hernia: an update].
Congenital diaphragmatic hernia (CDH) has an incidence of approximately 1:4000 live births. Most frequently the diaphragmatic defect is a left and posterolateral (Bochdalek) one. Prenatal diagnosis is made at ultrasonography; the relevant sonographic features will be described in the paper. Cystic adenomatoid malformation of the lung (CAML), pulmonary sequestration, bronchogenic cysts, pulmonary hypoplasia/agenesia need to be considered in differential diagnosis. In some cases, diagnosis of CDH is not possible "in utero": in such cases, herniation of abdominal viscera into the thorax takes place presumably just at delivery through a small diaphragmatic defect. CDH may be associated with intrauterine growth retardation (IUGR), chromosomal abnormalities (3%) and/or other malformations (10-50%): such as Central Nervous System, digestive, cardiac and urogenital anomalies. Therefore, search of associated malformations and amniocentesis with analysis of fetal karyotype are mandatory, whenever a CDH is diagnosed. CDH is still at present characterised by a high mortality (reportedly, about 45%). Many prognostic factors have been correlated to postnatal outcome of CDH: some of them are valuable prenatally by ultrasonography. However, the role of sonography in the prediction of neonatal outcome is still controversial: in particular, although many ultrasonographic parameters have been proposed, prenatal evaluation of pulmonary hypoplasia (a crucial factor related to postnatal survival) has not proved to be very accurate so far. Nevertheless, it is undisputable that prenatal diagnosis itself represents a crucial prognostic factor for CDH, since it allows birth of the affected fetuses in 3d level Perinatologic Centres provided with a Neonatal Intensive Care Unit and Neonatal Surgery.


Screening for abdominal aortic aneurysm--areas where information is still inadequate.
Abdominal aortic aneurysm (AAA) fulfils the criteria for a disease suitable for screening. However, important aspects need to be further analysed; the optimal age of the male population considered for screening has not yet been established, and whether women or specific high risk groups would benefit from screening has not been sufficiently evaluated. The impact of the current shift toward a high proportion of AAA repair done with endovascular technique and the long-term effect on QoL are additional issues that have not been adequately studied. Furthermore, therapeutic options for small AAA as well as secondary prevention programmes have to be developed.


Elevated plasma homocysteine and low vitamin B-6 status in nonsupplementing older women with rheumatoid arthritis.
The purpose of this study was to determine if nonsupplementing older women (aged >or=55 years) with rheumatoid arthritis had higher plasma homocysteine and lower B-vitamin status compared to healthy controls. Elevated plasma homocysteine, a risk factor for cardiovascular disease, may help explain why individuals with rheumatoid arthritis have an increased risk of cardiovascular disease. Older, free-living women were classified as rheumatoid arthritis (n=18) or healthy control (n=33). Participants were not using B-vitamin supplements. Fasting blood samples were measured for pyridoxal 5'phosphate (PLP) (the metabolically active coenzyme form of vitamin B-6), folate, red blood cell folate, vitamin B-12, transcobalamin II, homocysteine, C-reactive protein, and lipid concentrations. Participants completed 7-day weighed food records, the Stanford Health Assessment Questionnaire (HAQ), and a visual analog pain scale. PLP concentrations were lower in the rheumatoid arthritis vs healthy control participants (4.93+/-3.85 vs 11.35+/-7.11 ng/mL [20+/-16 vs 46+/-29 nmol/L]; P<0.01) whereas plasma homocysteine was higher in the rheumatoid arthritis group (1.63+/-0.74 vs 1.15+/-0.38 mg/L [12.1+/-5.5 vs 8.5+/-2.8 micromol/L]; P=0.02). Red blood cell folate concentrations were lower in the rheumatoid arthritis vs healthy control participants [414+/-141 vs 525+/-172 ng/mL [938+/-320 vs 1,190+/-390 nmol/L]; P=0.02). No significant differences were found for plasma folate, vitamin B-12, and transcobalamin II. An inverse correlation was found between PLP concentrations and the HAQ disability index (r=-0.37; P<0.01). A positive correlation was found between homocysteine concentrations and the HAQ disability index (r=0.36; P=0.01). Total cholesterol and low-density lipoprotein cholesterol levels were lower in the rheumatoid arthritis group (cholesterol 191+/-43 vs 218+/-33 mg/dL [4.95+/-1.11 vs 5.65+/-0.85 mmol/L]; P=0.02; low-density lipoprotein cholesterol 110+/-36 vs 137+/-29 mg/dL [2.85+/-0.93 vs 3.55+/-0.75 mmol/L]; P<0.01). No significant differences were seen between groups for protein (g/day), fat (g/day), cholesterol (mg/day), folate (microg/day), vitamin B-12 (microg/day), and vitamin B-6 (mg/day) dietary intakes. Poor vitamin B-6 status and elevated plasma homocysteine concentrations were seen in older women with rheumatoid arthritis compared to healthy controls and may contribute to their increased risk of cardiovascular disease.


Two new species of nematode parasites, Cucullanus epinepheli sp. n. (Cucullanidae) and Procamallanus (Spirocamallanus) sinespinis sp. n. (Camallanidae), from marine serranid and haemulid fishes off New Caledonia.
Based on light and scanning electron microscopical studies, two new species of parasitic nematodes are described from marine perciform fishes off New Caledonia: Cucullanus epinepheli sp. n. (Cucullanidae) from the intestine of the brownspotted grouper Epinephelus chlorostigma (Valenciennes) (Serranidae) and Procamallanus (Spirocamallanus) sinespinis sp. n. from the intestine of the silver grunt Pomadasys argenteus (Forsskål) (Haemulidae). Cucullanus epinepheli sp. n. differs from its congeners mainly in possessing a unique structure of the anterior, elevated cloacal lip with a large posterior outgrowth covering the cloacal aperture and in the presence of cervical alae and two small preanal papillae on the median dome-shaped precloacal elevation. This is the second known nominal species of this genus parasitising fishes of the family Serranidae and the second representative of Cucullanus Müller, 1777 recorded from fishes in New Caledonian waters. Procamallanus (Spirocamallanus) sinespinis sp. n. is mainly characterised by 10-12 spiral ridges in the buccal capsule, the presence of wide caudal alae, three pairs of pedunculate preanal papillae, two unequally long spicules (465-525 µm and 218-231 µm) and by the tail tip with a knob-like structure in the male, and the broad, rounded tail with a terminal digit-like protrusion without cuticular spikes in the female. This is the fifth nominal species of the subgenus Spirocamallanus Olsen, 1952 reported from fishes in New Caledonian waters.


Carfilzomib in combination with daratumumab in the management of relapsed multiple myeloma.
The therapeutic landscape of relapsed multiple myeloma (MM) is constantly evolving. To date, a large proportion of patients present with lenalidomide refractory disease at the time of first or second relapse. In this context, few efficient options are currently available. Carfilzomib and daratumumab are approved in the relapse setting. Recently, Phase Ib and Phase III trials evaluated the triplet drug combination daratumumab-carfilzomib-dexamethasone in the relapse setting and demonstrated strong clinical efficacy, especially in lenalidomide refractory patients. Based on these results, this combination has been approved by the US FDA for relapsed MM patients. The present review discusses the safety and efficacy of daratumumab-carfilzomib-dexamethasone in MM.


Secondary Corneal Myxoma After DMEK (Descemet Membrane Endothelial Keratoplasty).
BACKGROUND This is a clinical case of secondary corneal myxoma, which developed 18 months after Descemet membrane endothelial keratoplasty (DMEK). The DMEK was performed to treat viral endotheliitis and, postoperatively, a diagnosis of Crohn's disease was made. CASE REPORT A 52-year-old male, with no prior clinical history, presented with an endotheliitis in the left eye. The hypothesis of an undetected herpes infection was favored, and an antiviral treatment was prescribed using valacyclovir (1 g orally, 3 times daily). After 3 months of antiviral treatment, the endotheliitis was successfully controlled and a combined intervention of DMEK endothelial graft and phacoemulsification was performed. A corneal tumor was found 18 months after a successful DMEK procedure and was surgically removed. A pathological examination revealed a secondary corneal myxoma. CONCLUSIONS Corneal myxomas are rare lesions, often secondary to trauma in the Bowman's membrane, which is why it is often called a myxomatous corneal degeneration. In this instance, there is no link with Carney complex. However, myxomas under the eyelid or within the orbit are often associated with cardiac myxomas. Treatment is strictly surgical, either by simple excision or by surgical excision followed by graft. To the best of our knowledge, this is the first time that such an association between DMEK and secondary corneal myxoma has been described in the literature.


Biogenesis of mitochondria. A requirement for mitochondrial protein synthesis for the formation of a normal adenine nucleotide transporter in yeast mitochondria.
1. Parameters of ATP uptake by fully functional Saccharomyces cerevisiae mitochondria, including kinetic constants, binding constants and sensitivity to atractylate, closely resemble those of mammalian mitochondria. Scatchard plots of atractylate-sensitive adenine nucleotide binding indicate two distinct sites of high affinity (binding constant, K(D)'=1mum), and low affinity (binding constant, K(D)''=20mum) in the ratio 1:3. Uptake has high Arrhenius activation energies (+35 and +57kJ/mol), above and below a transition temperature of 11 degrees C. Atractylate-insensitive ATP uptake is apparently not saturable and has a low Arrhenius activation energy (6kJ/mol), suggesting a non-specific binding process. 2. Kinetic and binding constants for ATP uptake are not significantly changed in catabolite-repressed or anaerobic mitochondrial structures. 3. Inhibition of the mitochondrial protein-synthesizing system by growth of cells in the presence of erythromycin, or loss of mitochondrial DNA by mutation profoundly alters the adenine nucleotide transporter. ATP uptake becomes completely insensitive to atractylate, and the high-affinity binding site is lost. However, the adenine nucleotide transporter does not appear to be totally eliminated, as a moderate amount of saturable low-affinity ATP binding remains. 4. It is concluded that products of the mitochondrial protein-synthesizing system, probably coded by mitochondrial DNA, are required for the normal function of the adenine nucleotide transporter.


[Structural and functional myocardial alterations in patients with arterial hypertension associated with thyroid pathology under conditions of the Far North].
The remodeling types of left ventricle (LV), types and periodicity of heart arrhythmias have been studied in patients with arterial hypertension (AH) associated with thyroid pathology (mainly autoimmune thyroiditis (AIT)), residing in the Far North. It is shown, that in patients with AH associated with thyroid pathology, residing in Far North, defects of LV geometry, adverse in prognostic plan, are revealed significantly more often. The functional conditions of thyroid gland influences on distribution of LV geometry types (concentric LV hypertrophy is registered significantly more often at hypothyroidism). Predominant type of heart arrhythmias in patient with AH and AIT is superventricular extrasistoly (SVE). At presence of thyroid dysfunctions SVE number grows in 1.2 times.


Mechanisms of biological S-nitrosation and its measurement.
Nitric oxide (NO) exhibits multiple biological actions through formation of various oxidized intermediates derived from NO. Among them, nitrosothiol adducts (RS-NOs) with the sulfhydryl moiety of proteins and amino acids appears to be an important species in view of its unique chemical reactivity. Understanding of the biologically relevant S-nitrosation mechanism is essential because RS-NOs seem to be critically involved in modulation of intracellular and intercellular signal transduction, including gene transcription, cell apoptosis, and oxidative stress. RS-NOs have been recently found to be formed efficiently via one-electron oxidation of NO catalyzed by ceruloplasmin, a major copper-containing protein in mammalian plasma. Ceruloplasmin is synthesized mainly by hepatocytes, but it is also expressed by other cells such as macrophages and astrocytes. Once RS-NOs are formed, they function as NO transporters in biological systems, the NO being transferred to different sulfhydryls of various biomolecules. This transfer may be mediated by transnitrosation reactions occurring chemically or enzymatically by a means of specific enzymes such as protein disulfide isomerase. The molecular mechanism of biological S-nitrosation is discussed as related to the important physiological and pathophysiological functions of RS-NOs. Also, RS-NO assays that are being successfully used for detection of biological S-nitrosation are briefly reviewed.


Relation of heart rate recovery after exercise to C-reactive protein and white blood cell count.
This study was conducted to test the hypothesis that slow and abnormal heart rate recovery (HRR), an indicator of decreased autonomic nervous system activity, after exercise is associated with inflammatory markers. Subjects who underwent exercise treadmill testing (n = 5,527, mean age 50.4 +/- 8.5 years) were studied in a cross-sectional design. HRR was calculated as the difference between maximum heart rate during the test and heart rate 1 minute after the cessation of exercise. Abnormal HRR was defined as < or =12 beats/min. Subjects with abnormal HRR had higher levels of log C-reactive protein (CRP; 1.38 +/- 0.6 vs 1.11 +/- 0.4 mg/dl, p <0.001) and higher white blood cell counts (6.9 +/- 2.1 vs 6.2 +/- 1.7 x 10(9) cells/L, p <0.001) than those with normal HRR. HRR was associated with CRP (r = -0.21, p <0.001) and white blood cell count (r = -0.19, p <0.001). HRR was independently associated with CRP (beta = -0.13, p = 0.001) in a stepwise multiple regression. In a logistic multivariate model, the group within the highest quartile of CRP (odds ratio 1.54, 95% confidence interval 1.05 to 2.27) was more likely to have abnormal HRR than those within the lowest quartile. In conclusion, slow and abnormal HRR after exercise testing is associated with inflammatory markers, which could contribute to the high incidence of cardiovascular disease in these subjects.


[Stenosis of the right renal artery caused by the crura of the diaphragm. Report of a case].
We present one case of arterial hypertension that had rapidly developed in a 23-year-old patient. Arteriography demonstrates a light stenosis of the root of the right renal artery and an obstruction of the superior mesenteric artery. After an attempt of intraluminal dilatation had failed, we decided to carry out double revascularization with a venous graft. Postoperative control arteriography demonstrated early thrombosis in both grafts. Our second operation directly approached the lesion on the root of the renal artery, which was stenosed by muscle fibers from the right column of the diaphragm muscle. The resection of these fibers released the renal artery, which was otherwise normal, as was confirmed by the postoperative arteriogram. In a second part, we discuss the etiology of this double stenosis and our surgical strategy.


Highly efficient alkylation to ketones and aldimines with Grignard reagents catalyzed by zinc(II) chloride.
A highly efficient alkylation to ketones and aldimines with Grignard reagents in the presence of catalytic trialkylzinc(II) ate complexes derived from ZnCl2 (10 mol %) in situ was developed. This simple Zn(II)-catalyzed alkylation could minimize the well-known but serious problems with the use of only Grignard reagents, which leads to reduction and aldol side products, and the yield of desired alkylation products could be improved.


Primary cardiac angiosarcoma: A case report.
Primary cardiac angiosarcoma is rare, but it is the most aggressive type of cardiac malignant tumor. When disease is confirmed it is often late in its stage resulting in a poor prognosis. We reported a 52-year-old woman who complained of dyspnea for 2 months and dry cough for 10 days. Computed tomography (CT), F-fluorodeoxyglucose (F-FDG), positron emission tomography-computed tomography (PET-CT), and transthoracic echocardiography showed a space-occupying lesion 7 cm × 5 cm × 5 cm in the right atrium. Malignant cells can be seen in pericardial fluid by pericardiocentesis in local hospital, but not be diagnosed definitely. We conducted CT-guided biopsy of the tumor and observed the necrotic tissues. An exploratory thoracotomy was performed and histological examination confirmed angiosarcoma. Unable to be surgical excision due to infiltrating widely, the patients received chemotherapy with docetaxel for 2 cycles and the disease was progressive. Antiangiogenesis-targeted therapy was combined with chemotherapy for 3 months. Unfortunately, the tumor was still in progress. She died of respiratory failure, survived for 10 months. It is difficult to diagnose the primary cardiac angiosarcoma in early stage. Chemotherapy and target therapy are very limited in this case. Surgery remains the preferred treatment for patients with primary cardiac angiosarcoma. More efforts should be carried out on the effective diagnosis and multimodality treatment for this malignant disease.


A huge ovarian cyst in a hysterectomized bitch.
A 11-year-old, spayed, female mixed breed-dog was presented with an abdominal mass that was detected 1 month ago. Upon abdominal palpation a large, firm, oval shaped, movable mass was found in the mid-abdominal region. Survey radiograph of the abdomen demonstrated an oval soft tissue dense mass located on the right side of the abdominal cavity. A large, heteregenous and cystic mass with solid components occupying the majority of the abdomen and a small, cystic mass with solid components caudal to the left kidney were identified by transabdominal ultrasonography. Computed tomography scans revealed bilateral ovarian masses, and a small volume of retroperitoneal fluid on the right side. A cystic, but otherwise solid mass located in the right ovary and small retained left ovary encapsulated in the ovarian bursa were excised surgically by midline laparotomy. Histopathological examination of the excised mass from the right side revealed a large cystic structure consistent with an ovarian cyst and multiple corpora lutea and follicles at different maturational stages were detected in the left ovary. The precise origin of the ovarian cyst could not be determined by morphological appearance. Immunohistochemical staining suggested a cyst of surface epithelial origin. At re-examination 6 months after the surgery, the bitch appeared healthy and the clinical findings were all normal. To our knowledge, the cyst described here is the largest reported in an incompletely ovariohysterectomized bitch.


Application of probe electrospray ionization-tandem mass spectrometry to ultra-rapid determination of glufosinate and glyphosate in human serum.
Glufosinate and glyphosate, which are non-selective herbicides that include an amino acid moiety in their structures, are frequently used worldwide to control unwanted vegetation. Unfortunately, these readily available herbicides are also used by people to commit suicide, and thus represent important chemicals of interest in the fields of clinical medicine and forensics. Because of the high water solubility of these herbicides, most analytical methods for their detection require a derivatization step, which results in longer analysis times. Therefore, derivatization-based methods do not currently contribute to judgements on treatment decisions in emergency medicine. In this study, we addressed this limiting factor by developing an ultra-rapid and simple analytical technique using a combination of probe electrospray ionization (PESI) and tandem mass spectrometry (MS/MS), which gives quantitative results within 0.3 min. Herbicide standards were added to human serum that was then subjected to analysis (N = 5 per concentration). The analysis was repeated daily over eight consecutive days. The limit of detection (LOD) was 0.59 μg/mL for glufosinate and 0.20 μg/mL for glyphosate. The limit of quantitation (LOQ), i.e., the lowest point on the calibration curves, was 1.56 μg/mL for both the herbicides. The matrix effects were observed at three different concentrations (between 95.7%-104% for glufosinate, and between 90.7%-95.7% for glyphosate). When applied to samples taken from actual poisoning cases (six samples for each herbicide), the present method gave almost the same quantitative values as those obtained by conventional high-performance liquid chromatography with fluorescence detection. Thus, we believe that PESI-MS/MS could emerge as a rapid diagnosis method in the clinical emergency field.


Screening the ToxCast Phase 1, Phase 2, and e1k Chemical Libraries for Inhibitors of Iodothyronine Deiodinases.
Deiodinase enzymes play an essential role in converting thyroid hormones between active and inactive forms by deiodinating the pro-hormone thyroxine (T4) to the active hormone triiodothyronine (T3) and modifying T4 and T3 to inactive forms. Chemical inhibition of deiodinase activity has been identified as an important endpoint to include in screening chemicals for thyroid hormone disruption. To address the lack of data regarding chemicals that inhibit the deiodinase enzymes, we developed robust in vitro assays that utilized human deiodinase types 1, 2, and 3 and screened over 1800 unique chemicals from the U.S. EPA's ToxCast phase 1_v2, phase 2, and e1k libraries. Initial testing at a single concentration identified 411 putative deiodinase inhibitors that produced inhibition of 20% or greater in at least 1 of the 3 deiodinase assays, including chemicals that have not previously been shown to inhibit deiodinases. Of these, 228 chemicals produced enzyme inhibition of 50% or greater; these chemicals were further tested in concentration-response to determine relative potency. Comparisons across these deiodinase assays identified 81 chemicals that produced selective inhibition, with 50% inhibition or greater of only 1 of the deiodinases. This set of 3 deiodinase inhibition assays provides a significant contribution toward expanding the limited number of in vitro assays used to identify chemicals with the potential to interfere with thyroid hormone homeostasis. In addition, these results set the groundwork for development and evaluation of structure-activity relationships for deiodinase inhibition, and inform targeted selection of chemicals for further testing to identify adverse outcomes of deiodinase inhibition.


Temporal dynamics in the formation and interpretation of Nodal and BMP morphogen gradients.
One of the most powerful ideas in developmental biology has been that of the morphogen gradient. In the classical view, a signaling molecule is produced at a local source from where it diffuses, resulting in graded levels across the tissue. This gradient provides positional information, with thresholds in the level of the morphogen determining the position of different cell fates. While experimental studies have uncovered numerous potential morphogens in biological systems, it is becoming increasingly apparent that one important feature, not captured in the simple model, is the role of time in both the formation and interpretation of morphogen gradients. We will focus on two members of the transforming growth factor-β family that are known to play a vital role as morphogens in early vertebrate development: the Nodals and the bone morphogenetic proteins (BMPs). Primarily drawing on the early zebrafish embryo, we will show how recent studies have demonstrated the importance of feedback and other interactions that evolve through time, in shaping morphogen gradients. We will further show how rather than simply reading out levels of a morphogen, the duration of ligand exposure can be a crucial determinant of how cells interpret morphogens, in particular through the unfolding of downstream transcriptional events and in their interactions with other pathways.


Potential detrimental effects of a phytoestrogen-rich diet on male fertility in mice.
Soy and soy-based products are widely consumed by infants and adult individuals. There has been speculation that the presence of isoflavone phytoestrogens in soybean cause adverse effects on the development and function of the male reproductive system. The purpose of this study was to examine the influence of dietary soy and phytoestrogens on testicular and reproductive functions. Male mice were fed from conception to adulthood with either a high soy-containing diet or a soy-free diet. Although adult mice fed a soy-rich diet exhibited normal male behaviour and were fertile, we observed a reduced proportion of haploid germ cells in testes correlating with a 25% decrease in epididymal sperm counts and a 21% reduction in litter size. LH and androgens levels were not affected but transcripts coding for androgen-response genes in Sertoli cells and Gapd-s, a germ cell-specific gene involved in sperm glycolysis and mobility were significantly reduced. In addition, we found that dietary soy decreased the size of the seminal vesicle but without affecting its proteolytic activity. Taken together, these studies show that long-term exposure to dietary soy and phytoestrogens may affect male reproductive function resulting in a small decrease in sperm count and fertility.


Obstacle classification and 3D measurement in unstructured environments based on ToF cameras.
Inspired by the human 3D visual perception system, we present an obstacle detection and classification method based on the use of Time-of-Flight (ToF) cameras for robotic navigation in unstructured environments. The ToF camera provides 3D sensing by capturing an image along with per-pixel 3D space information. Based on this valuable feature and human knowledge of navigation, the proposed method first removes irrelevant regions which do not affect robot's movement from the scene. In the second step, regions of interest are detected and clustered as possible obstacles using both 3D information and intensity image obtained by the ToF camera. Consequently, a multiple relevance vector machine (RVM) classifier is designed to classify obstacles into four possible classes based on the terrain traversability and geometrical features of the obstacles. Finally, experimental results in various unstructured environments are presented to verify the robustness and performance of the proposed approach. We have found that, compared with the existing obstacle recognition methods, the new approach is more accurate and efficient.


Penetrating the air-liquid interface is the key to colonization and wrinkly spreader fitness.
In radiating populations of Pseudomonas fluorescens SBW25, adaptive wrinkly spreader (WS) mutants are able to gain access to the air-liquid (A-L) interface of static liquid microcosms and achieve a significant competitive fitness advantage over other non-biofilm-forming competitors. Aerotaxis and flagella-based swimming allows SBW25 cells to move into the high-O2 region located at the top of the liquid column and maintain their position by countering the effects of random cell diffusion, convection and disturbance (i.e. physical displacement). However, wild-type cells showed significantly lower levels of enrichment in this region compared to the archetypal WS, indicating that WS cells employ an additional mechanism to transfer to the A-L interface where displacement is no longer an issue and a biofilm can develop at the top of the liquid column. Preliminary experiments suggest that this might be achieved through the expression of an as yet unidentified surface active agent that is weakly associated with WS cells and alters liquid surface tension, as determined by quantitative tensiometry. The effect of physical displacement on the colonization of the high-O2 region and A-L interface was reduced through the addition of agar or polyethylene glycol to increase liquid viscosity, and under these conditions the competitive fitness of the WS was significantly reduced. These observations suggest that the ability to transfer to the A-L interface from the high-O2 region and remain there without further expenditure of energy (through, for example, the deployment of flagella) is a key evolutionary innovation of the WS, as it allows subsequent biofilm development and significant population increase, thereby affording these adaptive mutants a competitive fitness advantage over non-biofilm-forming competitors located within the liquid column.


Synthesis of soluble poly-yne polymers containing zirconium and silicon and corresponding conversion to nanosized ZrC/SiC composite ceramics.
Soluble organometallic polymers containing zirconium and silicon were synthesized by a salt metathesis reaction. The molecular weight of the polymers was measured by GPC and the corresponding structures were identified by (13)C NMR and FT-IR. After heat treatment of the polymers under argon at 1400 °C for 2 h, ZrC/SiC composites with different molar ratios of crystalline phases were obtained and characterized by XRD, elemental analysis, SEM and Raman spectroscopy. The crystalline size of the composites was approximately 100 nm-200 nm and the elements were well distributed at the different sites in the ceramics. The Raman results indicated that the ceramic residue could be considered as ZrC/SiC/C ternary composites.


Experiences of Children with Fetal Alcohol Spectrum Disorder and Their Families: A Critical Review.
Evidence suggests that children with fetal alcohol spectrum disorder (FASD) experience challenges across many areas of their daily lives and often require interprofessional supports. Recent studies have emphasized the need for an integrated system of care for children with FASD, incorporating medical, allied health, and education services, to facilitate open communication and support for the complex needs that many children experience. To develop such a system of care, it is important to first understand the impact of FASD on children's functioning during daily activities in different environmental contexts. A critical review of existing research was conducted using a critical interpretive synthesis approach. Results revealed that while many studies discussed impacts at the body functions and structures level of children with FASD, they often did not consider the activity, participation, and environmental factors also contributing to the daily functioning of this population. Several studies discussed caregiver experiences and challenges raising a child with FASD; however, no studies investigated the lived experiences relating to impacts across activities and environments from children's perspectives. In addition, the focus on deficits overshadowed investigation into the strengths of children with FASD, leaving a gap in the picture of their daily lives. Further research is required to determine the strengths that children with FASD demonstrate and the challenges impacting their daily functioning within different environmental contexts. Insights gleaned from such research would support intervention practices to become more holistic and interprofessional.


Peritoneal equilibration test results are different in infants, children, and adults.
Peritoneal equilibration test (PET) curves have been standardized in adult peritoneal dialysis (PD) patients. However, it appears that norms for pediatric PD patients may be different. A series of PET in 29 stable, chronic PD patients < or = 14 yr old performed at dwell volumes of 33 +/- 6 mL/kg with 2.5% Dianeal is reported. PET results for glucose and creatinine transport were compared between patients age < or = 2 and those 3 to 14 and published adult values by analysis of variance. Children < or = 2 transport glucose and creatinine more rapidly than do children 3 to 14 and adults. Children 3 to 14 transport glucose more rapidly than do adults; creatinine transport is not significantly different. These data demonstrate that transport characteristics differ between very young children, older children, and adults. Because PET are usually performed to plan mode of therapy, to address inadequate ultrafiltration, or to increase clearance, awareness of these results should assist in the clinical care of children on PD.


[Actions geared to tobacco control: a review of their implementation in Primary Health Care].
This survey examined the status of the implementation of policies for tobacco control in a medium-sized municipality through an assessment of actions recommended by the National Tobacco Control Program (NTCP) promoted by Primary Health Care (PHC) units. The coordinator of the local NTCP and professionals from 44 PHC units of Juiz de Fora in the state of Minas Gerais were interviewed, between May and July 2011. It was observed that the actions of treatment and health promotion were being duly executed, the highlight being the training of professionals to implement treatment in PHC. In 40.9%, there is the provision of treatment for smokers, and in 88.6% the actions are concentrated in activities in waiting rooms, groups and individual consultations. Limitations identified are probably not restricted to the municipality, such as inadequate structuring of units, high turnover, the varying degree of involvement of professionals in implementing the service and also the lack of mechanisms to enable them to implement and give continuity to care among their other responsibilities. The major challenge for tobacco control is to conduct intersectoral actions and in primary health care. The results can serve for the formulation of strategic health actions in other parts of the country.


Anthropometric, clinical and molecular determinants of treatment outcomes in postmenopausal, hormone receptor positive metastatic breast cancer patients treated with fulvestrant: Results from a real word setting.
To characterize determinants of treatment outcome in a real world population of 161 post-menopausal hormone receptor-positive metastatic breast cancer patients treated with fulvestrant. Descriptive statistics for demographics, anthropometrics, clinical and molecular characteristic were compared across subgroups of sensitivity/resistance to prior endocrine therapy and tested in uni/multivariate models. Clinical benefit was more common in sensitive patients with higher estrogen receptor expression and when fulvestrant was given in first line (p=0.02 and 0.046). In resistant patients, PFS was longer with lower BMI (p=0.01). Among endocrine sensitive women, longer PFS was associated with fulvestrant in first-line, single metastasis and no visceral involvement (p=0.01, 0.003 and 0.01). OS was shorter in resistant patients with HER2-positive disease and if fulvestrant was given in second and subsequent line (p=0.03). In sensitive patients, we observed worse OS with multiple metastases (p=0.008). Multivariate analyses confirmed longer PFS in resistant patients with lower BMI and older age (p=0.002 and 0.007). OS in resistant patients was negatively influenced by HER2 positivity and fulvestrant in second and subsequent line (p=0.04). In sensitive women, multiple metastases were associated with poorer survival (p=0.002). This evidence encourages considering patient and disease characteristics in decision making and outcome interpretation for patients candidate to fulvestrant.


Current and potential pest threats for canola in the Canadian Prairies.
Canola/oilseed rape (Brassica napus L.) production in Canada has increased to become a foundational crop in the Canadian Prairies and an important economic driver of this region. The increase in seeded area, and by association its reduction in-crop rotation frequency, has made it easier for pests to overcome current recommended agronomic management practices. The Canola Council of Canada has been successful in involving the entire commodity value chain in promoting and strengthening the Canadian canola industry; however, because of this production increase it is critically important to understand, evaluate and mitigate the potential risks of canola yield losses to current and potential pests. This Perspective provides an overview of what are currently the most damaging insects, pathogens and weeds to canola in the Canadian Prairies, potential future threats and opportunities farmers, agronomists and researchers can take to minimize these risks. © 2023 Society of Chemical Industry.


The Protein Chaperone ClpX Targets Native and Non-native Aggregated Substrates for Remodeling, Disassembly, and Degradation with ClpP.
ClpX is a member of the Clp/Hsp100 family of ATP-dependent chaperones and partners with ClpP, a compartmentalized protease, to degrade protein substrates bearing specific recognition signals. ClpX targets specific proteins for degradation directly or with substrate-specific adaptor proteins. Native substrates of ClpXP include proteins that form large oligomeric assemblies, such as MuA, FtsZ, and Dps in Escherichia coli. To remodel large oligomeric substrates, ClpX utilizes multivalent targeting strategies and discriminates between assembled and unassembled substrate conformations. Although ClpX and ClpP are known to associate with protein aggregates in E. coli, a potential role for ClpXP in disaggregation remains poorly characterized. Here, we discuss strategies utilized by ClpX to recognize native and non-native protein aggregates and the mechanisms by which ClpX alone, and with ClpP, remodels the conformations of various aggregates. We show that ClpX promotes the disassembly and reactivation of aggregated Gfp-ssrA through specific substrate remodeling. In the presence of ClpP, ClpX promotes disassembly and degradation of aggregated substrates bearing specific ClpX recognition signals, including heat-aggregated Gfp-ssrA, as well as polymeric and heat-aggregated FtsZ, which is a native ClpXP substrate in E. coli. Finally, we show that ClpX is present in insoluble aggregates and prevents the accumulation of thermal FtsZ aggregates in vivo, suggesting that ClpXP participates in the management of aggregates bearing ClpX recognition signals.


Cas-CLOVER is a novel high-fidelity nuclease for safe and robust generation of TSCM-enriched allogeneic CAR-T cells.
The use of T cells from healthy donors for allogeneic chimeric antigen receptor T (CAR-T) cell cancer therapy is attractive because healthy donor T cells can produce versatile off-the-shelf CAR-T treatments. To maximize safety and durability of allogeneic products, the endogenous T cell receptor and major histocompatibility complex class I molecules are often removed via knockout of T cell receptor beta constant (TRBC) (or T cell receptor alpha constant [TRAC]) and B2M, respectively. However, gene editing tools (e.g., CRISPR-Cas9) can display poor fidelity, which may result in dangerous off-target mutations. Additionally, many gene editing technologies require T cell activation, resulting in a low percentage of desirable stem cell memory T cells (TSCM). We characterize an RNA-guided endonuclease, called Cas-CLOVER, consisting of the Clo051 nuclease domain fused with catalytically dead Cas9. In primary T cells from multiple donors, we find that Cas-CLOVER is a high-fidelity site-specific nuclease, with low off-target activity. Notably, Cas-CLOVER yields efficient multiplexed gene editing in resting T cells. In conjunction with the piggyBac transposon for delivery of a CAR transgene against the B cell maturation antigen (BCMA), we produce allogeneic CAR-T cells composed of high percentages of TSCM cells and possessing potent in vivo anti-tumor cytotoxicity.


Clinical impact of postoperative loss in psoas major muscle and nutrition index after radical cystectomy for patients with urothelial carcinoma of the bladder.
Although the significance of preoperative nutritional status has been investigated, there is no report regarding the relationship of their postoperative changes on outcomes in patients who underwent radical cystectomy for bladder cancer. Here, we report the clinical impact of the change, from baseline, in nutritional status and volume of abdominal skeletal muscle mass and adipose tissue after radical cystetomy. A retrospective analysis of 89 patients with bladder cancer, who underwent curative radical cystectomy, was conducted to assess the time course of change, from baseline, in body composition and nutritional status at 1, 3, 6, 12, and 24 months, after surgery. Skeletal muscle mass and abdominal adipose tissue mass were quantified by unenhanced computed tomography images. Two different nutritional indices, the Prognostic Nutritional Index and the Controlling Nutritional Status score were calculated from laboratory blood tests. We evaluated the prognostic value of the rate of change in the body composition and nutritional status after radical cystectomy. The cross-sectional area at the level of the third lumbar vertebra of the psoas major muscle and nutritional indices showed a transient deterioration at 1 and 3 months after radical cystectomy, with a return to baseline values from 6 to 24 months. A ≤ -10% loss in the area of the psoas muscle was associated with a shorter overall survival, compared to those with a > -10 change [hazard ratio (HR) 2.2, P = 0.02]. Multivariate analyzes identified sarcopenia status at baseline (HR 2.2, P = 0.03) and a ≤ -10% loss in the psoas muscle (HR 2.4, P = 0.02) were identified as independent prognostic factors for overall survival. A subanalysis of patients without sarcopenia identified a worse survival outcome for patients with a ≤ -10% loss in the psoas muscle (HR 2.6, P = 0.03) and ≤ - 5 change in the Prognostic Nutritional Index (HR 3.6, P = 0.01). Further research is required to establish appropriate rehabilitation protocols and nutritional interventions after radical cystectomy for maintaining skeletal muscle mass and nutrition status which could counteract physical deterioration and improve outcomes.


Lethality in mice infected with recombinant vaccinia virus expressing hepatitis C virus core protein.
To establish a mouse model of HCV core expression and investigate the toxicity of HCV core protein or the possible pathogenic effects. A series of vaccinia viral expression vectors were engineered to express 5' portion of HCV genes including 5' non-translated region (NTR), core protein, and portion of the E1 gene. These HCV sequences were fused to a luciferase reporter gene and inserted into a vaccinia virus expression vector (pSC11) adjacent to the vaccinia virus promoter, p7.5. The recombinant DNA constructs were packed into infectious recombinant chimeric viruses. The expression of HCV core protein was examined in cultured cells after infection with these viruses. Death of the infected mice was investigated by specific correlation to the expression of HCV core protein and its expression levels. The recombinant virus (VNCE-LUA) expressed HCV core protein and an envelopeluciferase fusion protein in cultured cells. When Balb/c mice were inoculated intraperitoneally with more than 10(7) pfu per mouse of VNCE-LUA, death occurred immediately. The mortality was dependent on the amount of VNCE-LUA virus inoculated. All mice inoculated with 3 x 10(8) pfu of VNCE-LUA died within 4 days of infection and 50% of mice inoculated with 3 x 10(7) pfu of VNCE-LUA died within 7 days of infection. No death occurred in mice inoculated with 3 x 10(8) pfu of a control recombinant vaccinia virus, which expressed luciferase but not the HCV core and envelope proteins. Deletion of core sequences from VNCE-LUA rapidly reduced the mortality of infected mice whereas deletion of envelope sequence did not. SCID mice infected with VNCE-LUA died 2-3 days after infection, suggesting that the HCV-core induced mortality is not dependent on host T- or B-cell responses to core protein. HCV core protein can be lethal to mice when expressed in vivo and this specific lethality is independent of T-cells or B-cells. The findings and model itself provide a useful tool for further investigation on potential pathological effects as well as the potential toxicity of the HCV core protein.


Medulloblastoma: clinical presentation and management. Experience at the hospital for sick children, toronto, 1950-1980.
The authors review the cases of 144 children with medulloblastoma treated between 1950 and 1980. Duration of time between onset of symptoms and initial treatment was less than 1 1/2 months in 51% of cases, and less than 3 months in 76%. The tumor was located in the cerebellar vermis in 93% of patients. Brainstem infiltration was noted in 32%. Classical medulloblastomas comprised 82% of the total number reviewed, and desmoplastic medulloblastomas 15%. The majority of desmoplastic medulloblastomas were found in the midline of the cerebellum and in patients under 10 years of age. The prognosis for patients with desmoplastic medulloblastomas was worse than that for children with classical medulloblastomas. Spontaneous hemorrhage associated with primary or recurrent medulloblastoma occurred in 5.6% of the patients. Supratentorial metastases were found in 14.6% of cases, spinal cord metastases in 12.5%, and systemic metastases in 9%. The overall 5-year survival rate was 47%, and the 10-year survival rate 42%. Extent of surgical excision proved to be a statistically significant prognostic factor. Two patients developed recurrence after the "period of risk" as defined by Collins' rule. Delayed complications of radiotherapy were found to be substantial. Intelligence quotient (IQ) testing on 16 survivors revealed verbal IQ, performance IQ, and full-scale IQ to be within the normal range in 11, seven, and nine cases, respectively. Two were retarded on all scores.