A VIN1 GUS::GFP fusion reveals activated sucrose metabolism programming occurring in interspersed cells during tomato fruit ripening.
The tomato is a model for fleshy fruit development and ripening. Here we report on the identification of a novel unique cell autonomous/cellular pattern of expression that was detected in fruits of transgenic tomato lines carrying a GFP GUS driven by the fruit specific vacuolar invertase promoter VIN1. The VIN1 promoter sequence faithfully reproduced the global endogenous VIN expression by conferring a biphasic pattern of expression with a second phase clearly associated to fruit ripening. A closer view revealed a salt and pepper pattern of expression characterized by individual cells exhibiting a range of expression levels (from high to low) surrounded by cells with no expression. This type of pattern was detected across different fruit tissues and cell types with some preferences for vascular, sub-epidermal layer and the inner part of the fruit. Cell ability to show promoter activity was neither directly associated with overall ripening - as we find VIN+ and - VIN- cells at all stages of ripening, nor with cell size. Nevertheless the number of cells with active VIN-driven expression increased with ripening and the activity of the VIN promoter seems to be inversely correlated with cell size in VIN+ cells. Gene expression analysis of FACS-sorted VIN+ cells revealed a transcriptionally distinct subpopulation of cells defined by increased expression of genes related to sucrose metabolism, and decreased activity in protein synthesis and chromatin remodeling. This finding suggests that local micro heterogeneity may underlie some aspects (i.e. the futile cycles involving sucrose metabolism) of an otherwise more uniform looking ripening program.


AI-Assisted Screening of Oral Potentially Malignant Disorders Using Smartphone-Based Photographic Images.
The prevalence of oral potentially malignant disorders (OPMDs) and oral cancer is surging in low- and middle-income countries. A lack of resources for population screening in remote locations delays the detection of these lesions in the early stages and contributes to higher mortality and a poor quality of life. Digital imaging and artificial intelligence (AI) are promising tools for cancer screening. This study aimed to evaluate the utility of AI-based techniques for detecting OPMDs in the Indian population using photographic images of oral cavities captured using a smartphone. A dataset comprising 1120 suspicious and 1058 non-suspicious oral cavity photographic images taken by trained front-line healthcare workers (FHWs) was used for evaluating the performance of different deep learning models based on convolution (DenseNets) and Transformer (Swin) architectures. The best-performing model was also tested on an additional independent test set comprising 440 photographic images taken by untrained FHWs (set I). DenseNet201 and Swin Transformer (base) models show high classification performance with an F1-score of 0.84 (CI 0.79-0.89) and 0.83 (CI 0.78-0.88) on the internal test set, respectively. However, the performance of models decreases on test set I, which has considerable variation in the image quality, with the best F1-score of 0.73 (CI 0.67-0.78) obtained using DenseNet201. The proposed AI model has the potential to identify suspicious and non-suspicious oral lesions using photographic images. This simplified image-based AI solution can assist in screening, early detection, and prompt referral for OPMDs.


Dendritic Cells Overcome Cre/Lox Induced Gene Deficiency by Siphoning Material From Neighboring Cells Using Intracellular Monitoring-a Novel Mechanism of Antigen Acquisition.
Macrophages and dendritic cells (DCs) in peripheral tissue interact closely with their local microenvironment by scavenging protein and nucleic acids released by neighboring cells. Material transfer between cell types is necessary for pathogen detection and antigen presentation, but thought to be relatively limited in scale. Recent reports, however, demonstrate that the quantity of transferred material can be quite large when DCs are in direct contact with live cells. This observation may be problematic for conditional gene deletion models that assume gene products will remain in the cell they are produced in. Here, we investigate whether conditional gene deletions induced by the widely used Cre/Lox system can be overcome at the protein level in DCs. Of concern, using the human Langerin Cre mouse model, we find that epidermal Langerhans cells and CD11b+CD103+ mesenteric DCs can overcome gene deletion if the deleted gene is expressed by neighboring cells. Surprisingly, we also find that the mechanism of material transfer does not resemble known mechanisms of antigen uptake, is dependent on extra- and intracellular calcium, PI3K, and scavenger receptors, and mediates a majority of material transfer to DCs. We term this novel process intracellular monitoring, and find that it is specific to DCs, but occurs in all murine DC subsets tested, as well as in human DCs. Transferred material is successfully presented and cross presented on MHC-II and MHC-I, and occurs between allogeneic donor and acceptors cells-implicating this widespread and unique process in immunosurveillance and organ transplantation.


Fundamental Differences in Visual Perceptual Learning between Children and Adults.
It has remained uncertain whether the mechanisms of visual perceptual learning (VPL)1-4 remain stable across the lifespan or undergo developmental changes. This uncertainty largely originates from missing results about the mechanisms of VPL in healthy children. We here investigated the mechanisms of task-irrelevant VPL in healthy elementary school age children (7-10 years old) and compared their results to healthy young adults (18-31 years old). Subjects performed a rapid-serial-visual-presentation (RSVP) task at central fixation over the course of several daily sessions while coherent motion was merely exposed as a task-irrelevant feature in the visual periphery either at threshold or suprathreshold levels for coherent motion detection. As a result of this repeated exposure, children and adults both showed enhanced discrimination performance for the threshold task-irrelevant feature as in previous studies with adults.5-8 However, adults demonstrated a decreased performance for the suprathreshold task-irrelevant feature whereas children increased performance. One possible explanation for this difference is that children cannot effectively suppress salient task-irrelevant features because of weaker selective attention ability compared to that of adults.9-11 However, our results revealed to the contrary that children with stronger selective attention ability, as measured by the useful field of view (UFOV) test, showed greater increases in performance for the suprathreshold task-irrelevant feature. Together, these results suggest that the mechanisms of VPL change dramatically from childhood to adulthood due to a change in the way learners handle salient task-irrelevant features.


Functional outcomes after pyeloplasty in solitary kidneys: structured analysis with the implication of Acute Kidney Injury Network (AKIN) staging criteria to predict long-term renal function recoverability.
To assess the long-term functional outcomes after pyeloplasty in solitary kidney (SK) models and to define factors affecting postoperative renal function recoverability in adults. We retrospectively evaluated all adult patients who underwent pyeloplasty for pelvi-ureteric junction obstruction (PUJO) in SK. Long-term renal function was assessed by diuretic scintigraphy and morphological changes were also determined, e.g. renal parenchymal volume (RPV) and parenchymal thickness (PT). We considered a >20% increase in glomerular filtration rate (GFR) as improvement, >20% decrease as deterioration, and changes within 20% as stationary renal function. Univariate and multivariate analysis for factors affecting renal function recoverability, including the Acute Kidney Injury Network (AKIN) Staging System postoperatively, were performed. The study included 62 patients with a mean (SD) age of 29 (10.5) years. At a median of 48 months, the mean (SD) GFR increased from 41.3 (17) to 48.5 (20.1) mL/min (P < 0.001), with a significant increase in RPV and PT (P = 0.02 and P = 0.001, respectively). Follow-up renal function was static, improved and decreased in 39 (63.4%), 17 (27%) and six (9.6%) patients, respectively. Functional success was achieved in 90.4%. Patient's age ≥39 years, PT ≤0.75 cm and higher early postoperative AKIN staging were predictors associated with a 4.8-, 3.2- and 2.7-fold deterioration in renal function. Pyeloplasty in SK preserved renal function in 90.4% of the present patients. Pyeloplasty in SK when associated with older age, decreased PT preoperatively, and early higher AKIN staging postoperatively was associated with poor renal function recoverability.


Seizure susceptibility in the APP/PS1 mouse model of Alzheimer's disease and relationship with amyloid β plaques.
Alzheimer's disease is a common age associated neurodegenerative disorder associated with an elevated risk of seizures that may be fundamentally connected to cognitive dysfunction. We used 4-9month-old mice of the APP/PS1 mouse model of Alzheimer's disease to study the presence of epileptiform-like discharges and to establish if the amyloid-β plaques affect their generation. The EEG of the APP/PS1 transgenic mice revealed a higher incidence of epileptiform-like discharges i.e. seizure events (interictal spikes, sharp waves, or polyspikes) than in the controls. Also, APP/PS1 mice showed a lower latency to evoke seizure events than in the control animals when pentylenetetrazole (60mg/kg; i.p.) was injected. Moreover, physostigmine injection (1mg/kg; i.p.) also increased the frequency of spontaneous epileptiform-like discharges in the APP/PS1 mice. We also found a correlation between the frequency of epileptiform-like discharges and the number of amyloid-β plaques. Application of N-(2-chloroethyl)-N-ethyl-bromobenzylamine (50mg/kg) generated amyloid-β plaques in the cortex and seizure activity appeared. Taken together, these data indicate that deposits of amyloid-β plaques may be responsible for the epileptiform-like discharges recorded in the APP/PS1 mice and could be responsible for the elevated risk for seizures of Alzheimer's patients.


Multiparametric evaluation of dysphonic severity.
In recent years, the multiparametric approach for evaluating perceptual rating of voice quality has been advocated. This study evaluates the accuracy of predicting perceived overall severity of voice quality with a minimal set of aerodynamic, voice range profile (phonetogram), and acoustic perturbation measures. One hundred and twelve dysphonic persons (93 women and 19 men) with laryngeal pathologies and 41 normal controls (35 women and six men) with normal voices participated in this study. Perceptual severity judgement was carried out by four listeners rating the G (overall grade) parameter of the GRBAS scale. The minimal set of instrumental measures was selected based on the ability of the measure to discriminate between dysphonic and normal voices, and to attain at least a moderate correlation with perceived overall severity. Results indicated that perceived overall severity was best described by maximum phonation time of sustained /a/, peak intraoral pressure of the consonant-vowel /pi/ strings production, voice range profile area, and acoustic jitter. Direct-entry discriminant function analysis revealed that these four voice measures in combination correctly predicted 67.3% of perceived overall severity levels.


Site-specific conjugation of the quencher on peptide's N-terminal for the synthesis of a targeted non-spreading activatable optical probe.
Optical imaging offers high sensitivity and portability at low cost. The design of 'smart' or 'activatable' probes can decrease the background noise and increase the specificity of the signal. By conjugating a fluorescent dye and a compatible quencher on each side of an enzyme's substrate, the signal remains in its 'off ' state until it reaches the area where a specific enzyme is expressed. However, the signal can leak from that area unless the dye is attached to a molecule able to bind to a specific target also presented in that area. The aim of this study was to (i) specifically conjugate the quencher on the α-amino group of the peptide's N-terminus, (ii) conjugate the dye on the ε-amino group of a lysine in C-terminus, and (iii) conjugate the carboxyl group of the peptide's C-terminus to an amino group present on an antibody, using carbodiimide chemistry. The use of protecting groups, such as Boc or Fmoc, to allow site-specific conjugation, presents several drawbacks including 'on beads labeling', additional steps required for deprotection and removal from the resin, decreased yield, and dye degradation. A method of preferential labeling of α-amino N-terminal group in slightly acidic solution, proposed by Selo et al. (1996) has partially solved the problem. The present study reports improvements of the method allowing to (i) avoid the homo-bilabeling, (ii) increase the yield of the N-terminal labeling by two folds, and (iii) decrease the cost by 44-fold. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.


Role of cryptic rearrangements of human chromosomes in the aetiology of schizophrenia.
Schizophrenia (SZ) is a highly inherited disease that affects ~0.5% of the population. The genetic and environmental factors are involved in its aetiology and they interact with each other. Combination of symptoms is unique to each patient, the disease seriously interferes with the ability to function in society and affects the mental state of the patient. In most patients, the first manifestations of SZ appear during the adolescence or early adulthood. The hypothesis that SZ origin in impaired development of the nervous system is currently widely accepted. Some studies have identified several genetic and environmental factors that increase the risk of the disease manifestation, but none of them can be considered as the only cause of SZ. The genetics of the disease is complex and in last two decades it is assumed that the cryptic rearrangements could be one of its causes. Cryptic rearrangements (microdeletions and microduplications) are the chromosomal rearrangements smaller than 3-5 Mb. Their discovery was conditioned by the development of molecular genetic and molecular cytogenetic techniques. The aberrations affect one or more genes and change the gene dose. In this article, we present the rearrangements of the regions of human chromosomes more closely associated with the onset and development of SZ. Next, the candidate genes will be presented together with their inclusion in the context of theories trying to explain the origin of SZ through some important factors (e.g. action of dopamine or glutamate or GABA, formation of dendrites and neuronal synapses, etc.).


Azospirillum brasilense activates peroxidase-mediated cell wall modification to inhibit root cell elongation.
The molecular mechanism of beneficial bacterium Azospirillum brasilense-mediated root developmental remain elusive. A. brasilense elicited extensively transcriptional changes but inhibited primary root elongation in Arabidopsis. By analyzing root cell type-specific developmental markers, we demonstrated that A. brasilense affected neither overall organization nor cell division of primary root meristem. The cessation of primary root resulted from reduction of cell elongation, which is probably because of bacterially activated peroxidase that will lead to cell wall cross-linking at consuming of H2O2. The activated peroxidase combined with downregulated cell wall loosening enzymes consequently led to cell wall thickness, whereas inhibiting peroxidase restored root growth under A. brasilense inoculation. We further showed that peroxidase activity was probably promoted by cadaverine secreted by A. brasilense. These results suggest that A. brasilense inhibits root elongation by activating peroxidase and inducing cell wall modification in Arabidopsis, in which cadaverine released by A. brasilense is a potential signal compound.


Asymmetry of turning behaviour of rats induced by amphetamine and apomorphine.
Turning behaviour of rats following injection of amphetamine is in the same direction as following apomorphine. Unilateral common carotid artery ligation does not affect this asymmetry. Thus the reason for the asymmetry in turning behaviour following injection of amphetamine is probably due to postsynaptic dopaminergic asymmetry within the corpus striatum.


A case of ectopic cilia.
Ectopic cilia are the most rare of cilial anomalies. We report a 27-year-old man who presents with tarsal conjunctival symptomatic ectopic cilia. On slit-lamp examination eversion of the lower eyelid demonstrated that several black-white cilia were protruding from the conjunctiva.


Simultaneous live video presentation during knee arthroscopy.
To improve patient understanding, we evaluated prospectively the effect of patient-observed simultaneous surgeon-explained live video during knee arthroscopy. Patient satisfaction, understanding, and tolerance of the tenderness and pain were measured by a questionnaire completed immediately after the procedure. There were 72 patients (41 men, 31 women) with a mean age of 47 years; 76 joints were treated among them. All but one were comfortably able to watch the live video of the procedure, and 89% of the patients reported a good understanding of and satisfaction with the procedure. Moreover, if the patients ever had to have another arthroscopy, 66% answered that they "certainly" wanted a live video presentation, and 25% "somewhat" desired it. Live video-presented outpatient arthroscopic knee surgery could be considered a method for improving patient understanding of knee pathology.


Tocilizumab for treating severe COVID-19 pneumonia refractory to combined hydroxychloroquine, lopinavir plus ritonavir, and favipiravir: A case series.
Three patients diagnosed with severe COVID-19 pneumonia received treatment with hydroxychloroquine combined with lopinavir, ritonavir, and favipiravir. Two patients diagnosed early, received tocilizumab when the pneumonia became severe and survived. The thrid patient was diagnosed late and received tocilizumab when the disease progressed to acute respiratory distress syndrome, and died.


Contrasting patterns of within-species morphological variation in two cactophilic Drosophila species (Diptera: Drosophilidae).
In this work, we investigated the morphological variation of the intromittent male copulatory organ (aedeagus) of specimens from natural populations of two cactophilic Drosophila species distributed in the southeast region of Brazil, Drosophila gouveai Tidon-Sklorz & Sene and Drosophila antonietae Tidon-Sklorz & Sene. It was explored how the within-species variability is arranged for both species, considering their historical and ecological features. Our results showed two distinct aedeagal morphologies for these species, and differences within species were observed only in D. gouveai as specimens could be distinguished by their population origin. In contrast, after size discrepancies correction, this feature was not detected in D. antonietae. The contrasting patterns of intraspecific variation, together with the other features exhibited by these two species, are most likely to be explained by differences in the historical host plant association and distribution and in demographic events, which determined the evolutionary history of these two South American cactophilic Drosophila species.


Telomerase inhibition induces acute ATM-dependent growth arrest in human astrocytomas.
The purpose of the study was to examine the degree of hTERT, the catalytic subunit of telomerase, expression in paediatric high-grade astrocytoma and to explore the potential of telomerase inhibition as a therapy for these tumours. hTERT was expressed at high levels in 36 of 44 paediatric astrocytomas. Telomerase inhibition induced acute DNA damage and ATM-pathway-dependent G2/M cell cycle arrest in astrocytomas in vitro, both occurring prior to telomere shortening itself. Our data suggest that telomerase inhibition could be a useful adjuvant therapy for high-grade astrocytomas, potentially inducing tumour growth arrest following short-term treatment.


Pan LF-ELISA using BmR1 and BmSXP recombinant antigens for detection of lymphatic filariasis.
Anti-filarial IgG4 antibody has been shown to be a good marker for detection of lymphatic filaria infection. Previous studies demonstrated that anti-filarial IgG4 assay using BmR1 recombinant antigen was highly specific and sensitive for detection of brugian filariasis. For bancroftian filariasis, an equivalent assay employing recombinant antigen expressed from the ORF of SXP1 gene has been reported. In order to detect infections by all species of lymphatic filarial, BmR1 and BmSXP recombinant antigens were employed in the development of a pan LF-ELISA. BmR1 was previously produced while BmSXP recombinant antigen was produced by cloning the ORF of SXP1 gene from a Brugia malayi cDNA library, followed by expression in a bacterial expression system. Subsequently, each of the purified recombinant antigens (BmR1 and BmSXP) and mixture of different ratios of the two antigens (1:1, 2:1 and 1:2) were tested using IgG4-ELISA with various categories of infection and normal human serum samples. The results showed that both recombinant antigens were highly specific (99%-100%). For detection of brugian filariasis, BmR1 antigen alone and the mixture of BmR1 with BmSXP (1:1) gave 98% sensitivity; while BmSXP antigen alone showed 84% sensitivity. For detection of bancroftian filariasis, BmSXP antigen was more sensitive (95%) than assays using either BmR1 or mixtures of the two recombinant antigens. A sensitive and specific pan LF-ELISA for detection of lymphatic filariasis was successfully developed using two adjacent wells, each separately coated with BmR1 and BmSXP.


Glioma-Derived Platelet-Derived Growth Factor-BB Recruits Oligodendrocyte Progenitor Cells via Platelet-Derived Growth Factor Receptor-α and Remodels Cancer Stroma.
Glioma is an aggressive and incurable disease, and is frequently accompanied by augmented platelet-derived growth factor (PDGF) signaling. Overexpression of PDGF-B ligand characterizes a specific subclass of glioblastoma multiforme, but the significance of the ligand remains to be elucidated. For this end, we implanted a glioma-cell line transfected with PDGF-BB-overexpressing vector (GL261-PDGF-BB) or control vector (GL261-vector) into wild-type mouse brain, and examined the effect of glioma-derived PDGF on the tumor microenvironment. The volume of GL261-PDGF-BB rapidly increased compared with GL261-vector. Recruitment of many PDGF receptor (PDGFR)-α and Olig2-positive oligodendrocyte precursor cells and frequent hemorrhages were observed in GL261-PDGF-BB but not in GL261-vector. We then implanted GL261-PDGF-BB into the mouse brain with and without Pdgfra gene inactivation, corresponding to PDGFRα-knockout (KO) and Flox mice, respectively. The recruitment of oligodendrocyte precursor cells was largely suppressed in PDGFRα-KO than in Flox, whereas the volume of GL261-PDGF-BB was comparable between the two genotypes. Frequent hemorrhage and increased IgG-leakage were associated with aberrant vascular structures within the area where many recruited oligodendrocyte precursor cells accumulated in Flox. In contrast, these vascular phenotypes were largely normalized in PDGFRα-KO. Increased matrix metalloproteinase-9 in recruited oligodendrocyte precursor cells and decreased claudin-5 in vasculature may underlie the vascular abnormality. Glioma-derived PDGF-B signal induces cancer stroma characteristically seen in high-grade glioma, and should be therapeutically targeted to improve cancer microenvironment.


A Blinded, Cluster-Randomized, Placebo-Controlled School FeedingTrial in Burundi Using Rice Fortified With Iron, Zinc, Thiamine, and Folic Acid.
Iron-deficiency anemia is a major public health problem among school-aged children in sub-Saharan Africa. To evaluate the effectiveness of micronutrient-fortified rice to increase hemoglobin (Hb) concentration and reduce the prevalence of anemia among schoolchildren. Nine hundred four schoolchildren participated in this cluster-randomized trial during a 7-month intervention period. The study was conducted in 12 primary schools in rural Burundi. Hemoglobin, socioeconomic status, febrile illness, and dietary diversity were measured at baseline and follow-up. The changes in Hb concentration and anemia status were analyzed using linear and logistic mixed models, respectively. The micronutrient formulation contained an iron-to-zinc molar ratio of approximately 2.2. There was no significant difference in Hb concentration between the intervention and control groups (β = .09 g/dL; 95% confidence interval: -0.21 to 0.38) following the 7-month intervention. Nearly half the children reported having a fever within 2 weeks prior to baseline or follow-up. Children with febrile illness preceding follow-up were less than half as likely to show improvement in anemia status (odds ratio = 0.47, P < .001), with an average 0.56 g/dL smaller improvement in Hb at follow-up (P < .001). The high prevalence of fever and low iron-to-zinc molar ratio of the Ultra Rice formulation may have contributed to the lack of improvement in Hb. Alternatively, the detected anemia may not have been due to nutrient deficiencies. Anemia interventions in Burundi should implement multiple strategies to eliminate both iron deficiency and infectious causes of anemia.


Development and decision-making in the mammalian cerebral cortex.
One of the fundamental tasks of neurobiology is to understand how the precision and specificity of the adult nervous system is achieved during development. This paper reviews the progress that has been made toward this end in studies of the developing mammalian cerebral cortex. Particular attention is focused on the problem of how cortical neurons make decisions during development: the correlation between a neuron's 'birthday' and its final laminar destination and projection patterns has raised the possibility that young neurons may be committed to their adult fates very early on in development, perhaps prior to migration. Indeed, several lines of evidence reviewed here suggest that at least some of the decisions made by cortical neurons are intrinsic properties of the cell itself. These studies include experiments on the reeler mouse mutant, and more recent attempts to manipulate developmental fates by pharmacological interventions and transplantation techniques. It is concluded that early commitment events in the cerebral cortex may specify a neuron's laminar position and restrict the range of potential axonal projections that the cell may form, but that local positional cues direct neurons to select (or maintain) only certain of the possible projections.


The use of mucoadhesive polymers in vaginal delivery.
The vagina is an important area of the reproductive tract and serves as a potential route of drug administration. Beside locally acting drugs it is also of importance for systemic drug delivery, uterine targeting or even vaccination. Currently available dosage forms have several limitations, therefore novel concepts and dosage forms are needed. In this field mucoadhesive polymers will play a major role. After discussion of the anatomy and physiology of the vagina this review highlights the most important studies based on mucoadhesive polymer-systems like poly(acrylates), chitosan, cellulose derivatives, hyaluronic acid derivatives, pectin and traganth, starch, poly(ethylene glycol), sulfated polysaccharides, carrageenan, Na-alginate and gelatine.


Colony-stimulating factors in the management of neutropenia and its complications.
Granulocyte colony-stimulating factor (CSF) and granulocyte-macrophage CSF are potent drugs used to increase neutrophil counts after myelosuppressive chemotherapy. However, in various indications, the use of CSFs has no clinical benefit with regard to morbidity or mortality from infectious complications, frequency of antibiotic use, or rate of hospitalization. Thus, the application of CSFs should be limited to indications with proven clinical benefits or evidence of cost-effectiveness. This review will provide an overview of the state-of-the-art use of CSFs in chemotherapy-associated neutropenia, transplantation, and bone marrow failure syndromes. In addition, recently developed drugs for accelerated hematopoietic recovery will be presented.


Identifying the associated risks of pneumonia in COPD patients: ARCTIC an observational study.
Inhaled corticosteroids (ICS) are associated with an increased risk of pneumonia in patients with chronic obstructive pulmonary disease (COPD). Other factors such as severity of airflow limitation and concurrent asthma may further raise the possibility of developing pneumonia. This study assessed the risk of pneumonia associated with ICS in patients with COPD. Electronic Medical Record data linked to National Health Registries were collected from COPD patients and matched reference controls in 52 Swedish primary care centers (2000-2014). Levels of ICS treatment (high, low, no ICS) and associated comorbidities were assessed. Patients were categorized by airflow limitation severity. A total of 6623 patients with COPD and 48,566 controls were analyzed. Patients with COPD had a more than 4-fold increase in pneumonia versus reference controls (hazard ratio [HR] 4.76, 95% confidence interval [CI]: 4.48-5.06). ICS use increased the risk of pneumonia by 20-30% in patients with COPD with forced expiratory volume in 1 s ≥ 50% versus patients not using ICS. Asthma was an independent risk factor for pneumonia in the COPD population. Multivariate analysis identified independent predictors of pneumonia in the overall population. The highest risk of pneumonia was associated with high dose ICS (HR 1.41, 95% CI: 1.23-1.62). Patients with COPD have a greater risk of pneumonia versus reference controls; ICS use and concurrent asthma increased the risk of pneumonia further.


Matched analysis on orphan drug designations and approvals: cross regional analysis in the United States, the European Union, and Japan.
Orphan drugs have become a key area of focus in drug development for resolving unmet medical needs. The Orphan Drug Act in the USA and similar legislation in Japan, the European Union (EU), and several other countries has been enacted since 1983. This study provides a quantitative review of all orphan drug designations and approvals since the implementation of orphan drug legislation in key three regions. This study also identified and reviewed 'commonly designated' drugs across regions. Out of approximately 5000 designations, approximately 800 designations were common among the USA, EU, and/or Japan. Regional similarities, differences, and trends were identified. It is important to understand these aspects and the crucial role of orphan drug designation in global drug development.


The mechanical stop test and isovolumetric detrusor contractile reserve are associated with immediate spontaneous voiding after transurethral resection of prostate.
To identify urodynamic factors associated with the mechanical stop test and immediate spontaneous voiding following transurethral resection of prostate (TURP). We identified 90 men who underwent TURP over a 12-month period. Forty-three (mean age 68 years) underwent urodynamic evaluation prior to TURP. Isovolumetric detrusor contractile pressure (Piso) was obtained using the mechanical stop test during the voiding phase, and used to calculate detrusor contractile reserve (Pres = Piso - Pdet@Qmax). Primary outcome was spontaneous voiding after TURP. Preoperative catheter-free spontaneous voiding was present in 63% of men (27/43) with a urodynamic (mean ± SD): Qmax 6.2 ± 2.7 mL/s, Pdet@Qmax 102 ± 47 cmH2O, Piso 124 ± 49 cmH2O, Pres 22 ± 16 cmH2O, bladder outlet obstruction index (BOOI) 90 ± 49, and bladder contractility index (BCI) 132 ± 44. The remaining 16 catheter-dependent men demonstrated a urodynamic (mean ± SD): Qmax 3.6 ± 3.3 mL/s, Pdet@Qmax 87 ± 38 cmH2O, Piso 99 ± 51 cmH2O, Pres 10 ± 18 cmH2O, BOOI 82 ± 36, and BCI 106 ± 48. Following TURP, 67% of men voided spontaneously with their first void trial, and in receiver operator analysis of urodynamic measures (Pdet@Qmax, Piso, Pres, BOOI and BCI), only Pres was significantly associated with immediate spontaneous voiding after TURP (threshold Pres ≥ 9 cmH2O, AUC = 0.681, p = 0.035). In men who underwent TURP, a Pres ≥ 9 cmH2O was associated with immediate spontaneous voiding and may be easily incorporated into the postoperative pathway.


Does hearing loss after spinal anesthesia differ between young and elderly patients?
Fifty male patients scheduled for inguinal hernia repair with spinal anesthesia were included in this study. The patients were divided into two groups: 25 patients aged 30 yr or younger (Group Y) and 25 patients aged 60 yr or older (Group E). We performed subarachnoidal injection at the L3-4 interspace by using a 25-gauge Quincke needle with the patient in the sitting position, and 3 mL of 0.5% isobaric bupivacaine was administered. Patients were evaluated by pure tone audiometry (LdB [low frequencies], 125-500 Hz; SdB [speech frequencies], 500-2000 Hz; HdB [high frequencies], 2000-6000 Hz) on the day before and 2 days after spinal anesthesia. Low-frequency hearing loss observed in Group Y was significantly more common than in Group E (P < 0.01). There was no difference between the groups in speech and high frequencies. Mild hearing loss, defined as a hearing loss of 10-20 dB at two or more frequencies, was observed three times more frequently in Group Y than Group E (52% vs 16%; P = 0.014). We conclude that transient hearing loss was more common in young patients after spinal anesthesia, perhaps because the cerebrospinal fluid leakage after dural puncture is less in the elderly than in the young, a finding also associated with the infrequent incidence of postdural puncture headache in the elderly. Spinal anesthesia is one of the most frequently used regional anesthesia techniques in surgical interventions; however, rarely it may cause some transient or permanent neurological problems. One of these problems is headache, which is more frequent and severe in the young, and hearing loss, especially at low frequencies. Both the pain and the hearing loss are caused by leak of cerebrospinal fluid caused by the puncture in the membrane of the spinal cord during the procedure. We hypothesized that hearing loss might also be more frequent and severe in the young, and to test this hypothesis, we compared the hearing loss developing after spinal anesthesia between the young and the elderly. The implications of this study are as follows: First, spinal anesthesia must be performed carefully, especially in the young. Second, measures must be taken to avoid the leak of cerebrospinal fluid. This study reveals possible problems caused by spinal anesthesia in the young which can be easily overlooked.


Primitive reflexes in amyotrophic lateral sclerosis: prevalence and correlates.
Identifying frontal impairment in ALS is an important goal albeit disease-dedicated tools are still scarce. For this reason, we decided to consider primitive reflexes (PRs), variably regarded as correlates of frontal release and/or of upper motor neuron (UMN) impairment, often in the setting of dementias. Specifically, the aims of this work consisted in assessing the exact prevalence of the combination of seven PRs in ALS, trying to clarify their role as putative proxies of cognitive impairment or of UMN dysfunction. In this cross-sectional study, 50 consecutive ALS outpatients were evaluated for the presence of: palmomental (PM), corneomandibular (CM), glabella tap (MY), rooting, sucking, snout, and grasping reflexes. Cognitive screening was performed by the Frontal Assessment Battery (FAB) and the Weigl's Sorting test (WST); UMN dysfunction was concomitantly evaluated. PM, CM and MY were more frequently detected (62, 52, and 44 % of the ALS sample, respectively), while the other reflexes were under-represented. Patients displaying three or more PRs had significantly lower FAB and WST scores. On the other hand, UMN dysfunction was only moderately associated to PRs. In conclusion, PRs' assessment is a promising complementary tool for screening cognitive impairment in ALS; however, further work will be necessary to establish its added value with respect to already existing ALS-dedicated screening tools for cognition.


Aorto-left atrial tunnel: a rare entity.
Aorto-left atrial tunnel (ALAT) is a vascular channel that originates from 1 of the sinuses of Valsalva and terminates in the left atrium. The aorto-left atrial tunnel is an extremely rare anomaly. We describe here a case of congenital aorto-left atrial tunnel in a 4-year-old child who underwent successful surgical ligation with good immediate and early results.


Increased incidence of pemphigus vulgaris in southern Europeans living in Germany compared with native Germans.
The two major subtypes of pemphigus include: pemphigus vulgaris (PV) and pemphigus foliaceus. Only limited data are available on the epidemiology of these diseases. The aim of the present study was to estimate the gender- and age-specific incidences of PV in two well-defined regions of Germany and to compare the incidences among native Germans with those in citizens from other countries living in Germany. We performed a retrospective review of records from all patients that were diagnosed with PV at the Departments of Dermatology in Würzburg and Mannheim between 1989 and 1997. During the observation period, 14 patients were diagnosed with PV coming from an area with 1.46 million residents. With regard to the patients' age, the highest incidence for women was found in the 51-65-year-old-age group with 2.34 [95% confidence interval (CI): 0.36; 7.76] new cases per 1 million inhabitants per year. The highest incidence for men was observed in the over 65-year-old age group with 3.72 (95% CI: 0.95; 9.8) cases/million peryear. In the 51-65-year-old age group,we found a 25-foldhigher incidence of PV in foreigners living in Germany compared with native Germans. The age-adjusted incidence of PV was ninefold higher in foreigners compared with native Germans. Interestingly, all non-German patients came from two southern European countries (Turkey and Italy). The age-adjusted incidence of PV differs between native Germans and foreigners living in Germany. Further studies are necessary to address the risk factors (genetic and/or environmental) that contribute to this difference.


Nonviral gene delivery: Towards artificial viruses.
Several limitations to nonviral gene delivery have been overcome. Small nanometric particles have been obtained by condensation ofDNA with a polymerizable cation followed by DNA template-directed homopolymerization of the vector. Targeting of specific cell types has been achieved by using polyethylenimine (PEI), a cationic polymer, coupled to cell ligands such as galactose or small RGD peptide that allows cell entry by a receptor-mediated endocytosis pathway. Escape of the DNA complexes from the endosomes is favored by the 'proton sponge' effect as a consequence of the high buffering capacity of PEI. The last barrier to gene delivery, i.e. the nuclear membrane, can be crossed at the level of the nuclear pore complexes, by using nuclear localization signal DNA molecule conjugates.


Increases in body mass index, even within non-obese levels, raise the risk for Type 2 diabetes mellitus: a follow-up study in a Japanese population.
Obesity, the strongest risk factor for Type 2 diabetes mellitus, is less prevalent in Japanese than in other populations. We investigated the effects of body mass index (BMI, kg/m(2)) on the incidence of diabetes mellitus in a Japanese population. A follow-up study in 16 829 men and 8370 women who were apparently healthy at baseline (age 30-59 years, BMI 14.9-43.2 kg/m(2)). Incident diabetes mellitus was identified by 'fasting serum glucose > or = 7.00 mmol/l (126 mg/dl)' and/or 'under medical treatment for diabetes mellitus'. Hazard ratio, as an index for risk ratio, for incident diabetes mellitus according to BMI was estimated using Cox's proportional hazard models. Baseline age, smoking, drinking, exercise and education were computed as confounders. During mean follow-up periods of 7.4 years for men and 7.1 years for women, 869 men and 224 women had incident diabetes mellitus. Although the subjects were averagely non-obese [mean (sd) BMI 23.1 (2.6) kg/m(2) for men, 22.3 (2.7) kg/m(2) for women], hazard ratio for incident diabetes mellitus increased in parallel with increases in BMI. Multivariate-adjusted hazard ratios (95% confidence intervals) for increases in BMI of 1 kg/m(2) were 1.26 (1.24, 1.29) for men and 1.24 (1.20, 1.29) for women. BMI, even within the non-obese level, is a dose-dependent risk factor for diabetes mellitus in middle-aged Japanese. Increases in BMI of 1 kg/m(2) (= body-weight gain of 2.4-2.9 kg) may raise the risk by about 25%.


Thermosensitive midbrain neurons in the cat.
Responses of single neurons in the midbrain of cats anesthetized with chloral hydrate were studied during manipulations of midbrain temperature produced with a bilateral water-perfused thermode. Temperatures of the thermodes and the anterior hypothalamus were monitored while single neuron activity was recorded between the thermodes and correlated with the midbrain temperature. Q10's and thermal coefficients were calculated from the estimated temperature at the neuron itself. A surprisingly high percentage (72%) of the 72 neurons recorded in the caudal paramedian midbrain of 11 cats were thermoresponsive. Most of these were heat sensitive and exhibited a variety of frequency/temperature curves. Explorations of more rostral regions of the midbrain in 9 cats yielded only 18% thermoresponsive units out of the 99 neurons sampled. We suggest that the concentrated pool of warm-sensitive neurons in the caudal midbrain is part of an extensive system of brain stem thermosensors which are involved in establishing and controlling normal brain temperature.


Intracranial aneurysms and arterial hypertension: a review and hypothesis.
Intracranial aneurysms and systemic arterial hypertension coexist in a high percentage of patients. The relationship between intracranial aneurysms and hypertension is poorly defined. Therefore, we reviewed the role of hypertension in the pathogenesis of saccular aneurysms as previously reported in clinical, experimental, and autopsy studies. Among 24 relevant clinical and/or autopsy studies, the mean incidence of pre-existing hypertension was 43.5% in aneurysm patients compared to 24.4% in the normal population. Although definitive evidence is lacking, data from multiple types of investigations indicate that systemic arterial hypertension creates a greater risk for the development of intracranial aneurysms than previously believed. The underlying pathophysiological mechanism(s) are also poorly defined. We propose a unifying hypothesis: Endothelial injury, occlusion of the vasa vasorum, and disruption of the synthesis of collagen and elastin are likely the most important factors in initiating the development of aneurysms. Chronic hypertension potentially affects all of these factors. Consequently, chronic hypertension may cause intimal thickening, necrosis of the tunica media, changes in the compositional matrix, and degeneration of the internal elastic lamina to develop in the arterial wall. These structural changes could cause a focal weakening in the arterial wall with resultant bulging. This theory accounts for the high incidence of intracranial aneurysms in the absence of any known associated hereditary or connective-tissue disease. Nor does it exclude the possibility of other etiological factors. From the perspective of prevention, however, it offers clear opportunities for prophylaxis.


Distribution patterns and individual variations of callosal connections in the albino rat.
After complete callosotomy the distribution of degeneration products was re-investigated in adult albino rats. Three to seven days post operation, coronal, horizontal and "flattened" sections were impregnated according to the new methods of Gallyas et al. (1980) which stain degenerating axons and terminals, respectively. The regional distribution patterns of callosal terminals were directly visualized with dark field illumination at low magnification. With this technique the distribution pattern of axons and terminals could be compared between different cortical regions and individuals. Callosal terminals tend to accumulate in patches or bands along the borders of cortical regions and areas. The concentration of callosal terminals was especially high at the common corners of more than two cortical areas. The callosal system shows a rather constant distribution pattern which is composed of column shaped subunits. Considerable individual variations were recognized concerning the number, position, shape, density and contiguity of the columnar units either occupied by callosal connections or empty. Although the laminar distribution of callosal terminals shows some similarities in different areas of the cortex, there is no common laminar pattern characteristic either for the whole neocortex or for any cortical region. The comparison between consecutive sections stained either for degenerating fibers or degenerating axon terminals revealed that the callosal axons do not determine directly the arrangement and packing density of callosal synapses. Whatever determines the position and amount of callosal synapses this influence seems to be exerted via translation into the columnar organization.


Late evaluation of patients undergoing valve replacement with the Björk-Shiley prosthesis.
This study analyzes 484 patients who survived mitral, aortic, or mitral and aortic valve replacement using the Björk-Shiley prosthesis from January, 1970, through December 31, 1974. Long-term follow-up of 1 1/2 to 6 1/2 years (mean, 3.67 yr) was done on 435 patients (98.2%). Eighty to 85% of the patients have improved noticeably. Thromboembolic problems occurred in 6.9%, representing 1.5 emboli per 1,000 patient-months. Anticoagulant bleeding problems occurred in 6.4% of the patients; late mortality was 15%. Actuarial survival curves showed patients at risk to 6 years having a 79% chance of survival. The same analysis according to preoperative New York Heart Association Functional Classification showed a striking reduction in survival in class IV patients. The Björk-Shiley prosthesis is a good choice for valve replacement today. Earlier diagnosis and treatment are needed to obtain better long-term survival.


Negative effect of serotonin-norepinephrine reuptake inhibitor therapy on rat bone tissue after orchidectomy.
Our goal was to determine if venlafaxine has a negative effect on bone metabolism. Rats were divided into three groups. The sham-operated control group (SHAM), the control group after orchidectomy (ORX), and the experimental group after orchidectomy received venlafaxine (VEN ORX) in standard laboratory diet (SLD) for 12 weeks. Bone mineral content (BMC) was measured by dual energy X-ray absorptiometry (DXA). Bone marker concentrations of carboxy-terminal cross-linking telopeptide of type I collagen (CTX-I), osteoprotegerin (OPG), amino-terminal propeptide of procollagen type I (P1NP), bone alkaline phosphatase (BALP), sclerostin and bone morphogenetic protein 2 (BMP-2) were examined in bone homogenate. The femurs were used for biomechanical testing. Compared to the ORX group we found lower BMD in the diaphysis area of the femur in the VEN ORX group, suggesting a preferential effect on cortical bone. Of the bone metabolism markers, there was significant decrease (ORX control group versus VEN ORX experimental group) in BALP levels and increase in sclerostin and CTX-I levels, suggesting a decrease in osteoid synthesis and increased bone resorption. The results suggest that the prolonged use of venlafaxine may have a negative effect on bone metabolism. Further studies are warranted to establish whether venlafaxine may have a clinically significant adverse effect on bone.


Identification of deubiquitinase inhibitors via high-throughput screening using a fluorogenic ubiquitin-rhodamine assay.
Identification of selective deubiquitinase (DUB) inhibitors is critical for probe development to further understand and explore DUB biological function. Here, we detail the optimization and deployment of an in vitro fluorogenic ubiquitin-rhodamine assay to conduct high-throughput screening of a small molecule library against a panel of DUBs. In screening the compound library against multiple DUBs in parallel, we describe an approach for identifying selective DUB inhibitors and provide a roadmap for enabling selective DUB inhibitor discovery. For complete details on the use and execution of this protocol, please refer to Varca et al. (2021).


A multicenter Italian study of amineptine (Survector 100).
This study reports the antidepressant efficacy and the safety of amineptine (Survector 100) in a national multicenter open clinical trial (32 hospital centers). Three hundred twenty-four patients with depressive disorders, selected according to DSM-III Diagnostic Criteria and INSERM classification, were treated with amineptine (200 mg/day) for 40 days (mean). Amineptine proved to be significantly effective with a rapid onset of action (beginning on the 7th day) on depressive symptoms, as shown by the decrease in the Hamilton Depression Rating Scale total score. The assessment of the results in the different diagnostic subgroups confirmed the wide-spectrum antidepressant activity of amineptine. Particularly in reactive and neurotic depression (nonpsychotic depression according to INSERM) and dysthymic disorders according to DSM-III, amineptine induced an improvement in 90% of patients. Nevertheless, amineptine was also effective in psychotic depression (INSERM) and in major depressive episodes (DSM-III), in which improvement occurred in 76% of patients. The low frequency of side effects and the remarkable acceptability (clinical, biological, and cardiovascular) of amineptine were also confirmed.


Evaluation of facial morphology and sagittal relationship between dental arches in primary and mixed dentition.
To assess facial morphology (Pattern) and sagittal relationship between dental arches (Class), and establish a potential association between them and the variables sex, age and ethnicity, among schoolchildren aged between 4 and 9 years old (mean age of 6.7 years) in primary and mixed dentitions. The sample comprised 875 children (457 males and 418 females) attending schools in Descalvado, São Paulo, Brazil. An attempt was made with a view to establish a potential association between children's morphological features with sex, age and ethnicity. Descriptive analysis revealed a predominance of facial Pattern I (69.9 %) and Class I (67.4 %). Statistical tests (p < 0.001) showed that Class I was more frequent among Pattern I children, whereas Class II prevailed among Pattern II, and Class III was frequent among Pattern I and III children. Ethnicity was the only variable associated with facial pattern. Results suggest that facial pattern and sagittal relationship between dental arches tend to be correlated. Ethnicity was associated with facial pattern, with Pattern I being the most recurrent among Caucasians and facial Pattern II being recurrent among Afro-descendant subjects.


A writing group for female assistant professors.
The number of female medical school faculty being promoted and the speed at which they are promoted have not kept pace with their male counterparts at many institutions. One of the reasons is that these women are not publishing peer reviewed manuscripts at an equivalent rate. This study evaluates the impact of a women's writing group on faculty publication rates. The writing group was conducted by 2 senior faculty members at the University of Pennsylvania School of Medicine and targeted female junior faculty. The writing group consisted of a didactic skills curriculum, question sessions, and both faculty and peer support to improve publishing rates. Curriculum vitae were collected, and PubMed and Ovid searches were used to establish the publishing productivity of the writing group participants both before and after participation in the writing group. On average, women who completed the writing group showed a nearly 3-fold increase in average publishing rate from 1.5 papers per year preceding the course to 4.5 per year following completion of the writing group (p<.001). The results from our program suggest that a women's writing group is an effective intervention for increasing publishing rates of female junior faculty. In addition to the documented improvement in publication rates, we watched participants develop clearer writing styles, lose many of their inhibitions about writing, respond to group affiliation and collaboration, and gain tremendous self-confidence.


An Interview With Janeane N. Anderson Regarding Patient-Provider Communication and Health Disparities.
This month's column is an interview with Dr Janeane N. Anderson, assistant professor, College of Nursing, at the University of Tennessee Health Science Center (UTHSC) in Memphis who is a health communication scholar with a focus on patient-provider communication and healthcare disparities.


Photocatalytic Properties of g-C₃N₄-TiO₂ Heterojunctions under UV and Visible Light Conditions.
Graphitic carbon nitride (g-C₃N₄) and titanium dioxide (TiO₂) were chosen as a model system to investigate photocatalytic abilities of heterojunction system under UV and visible light conditions. The use of g-C₃N₄ has been shown to be effective in the reduction in recombination through the interaction between the two interfaces of TiO₂ and g-C₃N₄. A simple method of preparing g-C₃N₄ through the pyrolysis of melamine was employed, which was then added to undoped TiO₂ material to form the g-C₃N₄-TiO₂ system. These materials were then fully characterized by X-ray diffraction (XRD), Brunauer Emmett Teller (BET), and various spectroscopic techniques including Raman, X-ray photoelectron spectroscopy (XPS), Fourier transform infrared spectroscopy (FT-IR), diffuse absorbance, and photoluminescence analysis. Photocatalysis studies were conducted using the model dye, rhodamine 6G utilizing visible and UV light irradiation. Raman spectroscopy confirmed that a composite of the materials was formed as opposed to a mixture of the two. Using XPS analysis, a shift in the nitrogen peak to that indicative of substitutional nitrogen was detected for all doped samples. This is then mirrored in the diffuse absorbance results, which show a clear decrease in band gap values for these samples, showing the effective band gap alteration achieved through this preparation process. When g-C₃N₄-TiO₂ samples were analyzed under visible light irradiation, no significant improvement was observed compared that of pure TiO₂. However, under UV light irradiation conditions, the photocatalytic ability of the doped samples exhibited an increased reactivity when compared to the undoped TiO₂ (0.130 min-1), with 4% g-C₃N₄-TiO₂ (0.187 min-1), showing a 43.9% increase in reactivity. Further doping to 8% g-C₃N₄-TiO₂ lead to a decrease in reactivity against rhodamine 6G. BET analysis determined that the surface area of the 4% and 8% g-C₃N₄-TiO₂ samples were very similar, with values of 29.4 and 28.5 m²/g, respectively, suggesting that the actual surface area is not a contributing factor. This could be due to an overloading of the system with covering of the active sites resulting in a lower reaction rate. XPS analysis showed that surface hydroxyl radicals and oxygen vacancies are not being formed throughout this preparation. Therefore, it can be suggested that the increased photocatalytic reaction rates are due to successful interfacial interactions with the g-C₃N₄-doped TiO₂ systems.


A microcomputer based catheter laboratory system for analysis of coronary blood flow in man.
The most frequently and conveniently used technique for measurement of coronary blood flow in patients is that of continuous thermodilution applied to coronary sinus flow. A system is described in which a microcomputer is used to sample, analyse, display and store data obtained using this technique.


Evaluation of nucleotide derivatives and sugar nucleotide analogs as inhibitors of glycosyltransferases.
The inhibition of glycosyltransferases was studied using uridine monophosphate derivatives and uridine diphosphate sugar analogs. Modification in the nucleoside portion caused selective inhibition of glycosyltransferases.


Guinea pig maximization test for skin sensitisation: the use of fewer test animals.
Current European Community (Annex V) guidelines recommend the use of 20 test animals in the guinea pig maximisation test for skin sensitization. The suitability, for classification and labelling purposes, of reducing the number of test animals has been examined by analysing the results of 40 studies submitted to the Health and Safety Executive, and by the use of a mathematical model. Our results suggest that in most cases an experiment with ten test animals can be used to determine satisfactorily whether a substance should be labelled with the risk phrase "may cause sensitisation by skin contact". However, serious consideration should be given to the need for additional investigation if two or three of the ten test animals show a sensitisation response. The highest non-irritant concentration of a substance should be used at challenge. Clearer guidance in Annex V on evaluating challenge responses would be beneficial.


Angelica oil restores the intestinal barrier function by suppressing S100A8/A9 signalling in mice with ulcerative colitis.
Ulcerative colitis (UC) progression is driven by the activation of immune cells that release pro-inflammatory mediators to disrupt intestinal epithelial barrier integrity. This study aimed to investigate the potential protective effects of Angelica oil (AO) on the intestinal epithelial barrier in mice with UC and the underlying mechanisms. Improvement of the disease state and protective effect of AO on the intestinal epithelial barrier were observed in mice with dextran sulphate sodium salt (DSS)-induced UC. Protein microarrays were used to screen AO-affected cytokine pools and their recruited immune cells for accumulation in the tissues. Furthermore, quantitative proteomics was applied to search for AO-acting molecules and to verify in vitro the functions of key molecules between inflammation and the intestinal mucosal barrier. AO significantly alleviated intestinal inflammation, reduced intestinal permeability, and retained barrier function in mice with UC. Furthermore, cytokines inhibited by AO mainly promoted monocyte and neutrophil activation or chemotaxis. Moreover, proteomic screening revealed that S100A8/A9 was a key molecule significantly regulated by AO, and its mediated TLR4/NF-κB pathway was also inhibited. Finally, we verified that AO inhibited the activation of the S100A8/A9/TLR4 signalling pathway and enhanced the expression of tight junctions (TJs) proteins using a cellular model of intestinal barrier damage induced by S100A8/A9 or macrophage-derived medium. And the enhancement of TJs in intestinal epithelial cells and the inhibition of inflammatory signalling by AO were significantly attenuated due to the application of S100A8/A9 monoclonal antibody. These results demonstrated that AO improves intestinal mucosal barrier damage in the inflammatory environment of mice with UC by inhibiting the expression of S100A8/A9 and the activation of its downstream TLR4/NF-κB signalling pathway.


Characterization of ciprofibrate and clofibric acid as peroxisomal proliferators in primary cultures of rat hepatocytes.
We have determined the comparative activities of peroxisomal proliferators, ciprofibrate and clofibric acid on various hepatic parameters associated with endoplasmic reticulum, mitochondria and peroxisomes in primary cultures of rat hepatocytes. We have measured the activities of carnitine acetyltransferase and fatty acylCoA oxidase, and the amount of 60 and 80 kD polypeptides as biochemical markers of the peroxisomal function; laurate hydroxylase and cytochrome P-450 as markers of the endoplasmic reticulum; and carnitine palmitoyltransferase as a marker of mitochondria in primary cultures of hepatocytes. Ciprofibrate (0.01 to 0.3 mM) and clofibric acid (0.1 to 3 mM) produced similar changes in several components of cultured hepatocytes within 72 hr. Increases of protein (18 and 11%), carnitine palmitoyltransferase (23 and 97%), cytochrome P-450 (37 and 49%), carnitine acetyltransferase (484 and 614%), fatty acylCoA oxidase (529 and 931%) and laurate hydroxylase (624 and 671%) were obtained in hepatocytes after a 72-hr exposure to 0.1 mM ciprofibrate and 1.0 mM clofibric acid, respectively. In cultured hepatocytes, ciprofibrate was about 30-fold more active than clofibric acid for the stimulation of carnitine acetyltransferase, laurate hydroxylase and fatty acylCoA oxidase activities. Ciprofibrate was also more potent than clofibric acid as an inducer of the 60 and 80 kD proteins in hepatocytes. The maximal drug-induced increases in carnitine acetyltransferase activity were not additive, and the induction of carnitine acetyltransferase by ciprofibrate was blocked by addition (1 micrograms per ml) of cycloheximide or actinomycin D. Changes in protein and RNA synthesis preceded the drug-induced increases of carnitine acetyltransferase activity.(ABSTRACT TRUNCATED AT 250 WORDS)


Association of hemoglobin chains with the cell membrane as a cause of red cell distortion in thalassemia.
Hemoglobin chains were separated and their interaction with membrane ghosts was studied using their ability to quench the fluorescence intensity of a membrane embedded probe. It was observed that alpha chains bind faster and with higher affinity to the membrane sites than do beta chains. The fast reversible interaction of both chains with the membrane was followed by a time-dependent partial loss of reversibility. Band 3 cytoplasmic fragments (B3F) were isolated and their reaction with separated Hb chains was studied using fluorescence quenching techniques as well. The data demonstrate that the relative affinity of the chains for B3F and loss of reversibility of the reaction followed patterns similar to the corresponding interaction of the chains with whole membranes. Band 3 cytoplasmic poles are therefore suggested as the high-affinity sites on the membrane for hemoglobin chains. When globin was reacted with B3F, it was observed that this protein binds strongly to the same membrane sites, but practically irreversibly. Exchange of the HbA content of normal cells by separated alpha or beta chains resulted in membrane distortions in both cases, but alpha chains caused greater morphological changes than did beta chains. The results of this study may provide one explanation for the differences in the thalassemia syndromes when excess of either alpha or beta chains is involved.


Monoclonal antibodies against calcitonin. Characterization and application in light and electron microscopy immunocytochemistry.
Twenty-seven monoclonal antibodies (MAbs) to synthetic human calcitonin (CT) were characterized for their reactivities with human CT peptide fragments by dot-blot analysis on nitrocellulose paper. Most of the antibodies bound to the C-terminus and fewer to the mid-region of CT. We have studied thyroid tissue specimens from several animal species after fixation in paraformaldehyde-, glutaraldehyde- or picric acid-containing mixtures and cryostat sectioning or embedment in paraffin or plastic (Epon 812 or Lowicryl 4KM) using this panel of MAbs. The site of antigen-antibody reaction was revealed either by immunoperoxidase, immunoalkaline phosphatase or by silver-enhanced immunogold staining methods. All MAbs were able to localize CT in human, rat and mouse thyroid C cells. Nineteen MAbs recognizing synthetic salmon CT and synthetic [Asu1,7]-eel CT by dot-blot, reacted with chicken ultimobranchial body C cells. One MAb recognizing native porcine CT by dot-blot, stained C cells in hog thyroid. Immunopositivity was confined to the cytoplasm and ultrastructural immunogold labelling demonstrated that cytoplasmic secretory granules were stained. Surgical specimens from human medullary thyroid carcinoma were also analysed for the presence of CT and a variable number of positive cells was found. Furthermore, Congo red-positive areas were shown to react with the MAbs. All conventional staining and immunoabsorption controls were negative. Hence, these MAbs may be suitable for use in routine immunopathological diagnosis of CT-producing tumors and for immunocytochemical localization of the three major CT variants in different animal species.


Adipose tissue biglycan as a potential anti-inflammatory target of sodium salicylate in mice fed a high fat diet.
Inflammation in adipose tissue (AT) during obesity causes impaired AT function. Although multiple extracellular matrix (ECM) proteins are expressed in AT their potential role in adipose tissue inflammation is unclear. Biglycan, a pro-inflammatory ECM gene, is highly enriched in adipose tissue. However, whether it is correlated with adipose tissue inflammation is unknown. We provide evidence in support of a strong association between biglycan expression and inflammatory status of adipose tissue. C57BL6 mice were fed either a control (10% fat calories) or a high fat diet (HFD) (60% fat calories) for 8 weeks. Adipose tissue was analyzed for the expression of biglycan, IL-6 and TNFα. Biglycan knockout or wild type were also fed a high fat diet for 8 weeks and the expression of inflammatory genes in the mesenteric adipose tissue was examined. To test anti-inflammatory treatment on biglycan expression, a group of mice were fed either the low fat or high fat diet for eight weeks supplemented with either saline or sodium salicylate @ 25mg/100ml in their drinking water. Mice on HFD had an increase in ECM genes (BGN and COL1A1), inflammatory genes (IL-6 and TNFα) in both the subcutaneous and epididymal depots. However, correlation analysis only shows a positive correlation between biglycan, IL-6 and TNFα expression. In addition, lower expression of IL-6 and CD68 was found in the mesenteric adipose tissue of biglycan knockout mice compared to the wild type. Sodium salicylate treatment reduced subcutaneous adipose tissue expression of BGN, COL1A1, and COL6A1 and a concurrent downregulation of TNFα and IL-6 and TLR4 expression. Salicylate also lowered the serum TGFβ1 levels. Biglycan expression correlates with adipose tissue inflammation, especially in the subcutaneous depot compared to the epididymal depot. This is supported by the greater effect of sodium salicylate in attenuating both inflammatory and ECM gene expression the subcutaneous adipose depot compared to the epididymal depot. These results show that inflammatory state may explain the induction of biglycan, and perhaps, other ECM genes in adipose tissue.


New polymorphic microsatellite loci revealed for the dusky shark Carcharhinus obscurus through Ion Proton double-digest RAD sequencing.
The non-model shark species, dusky shark Carcharhinus obscurus, is a bio-economically and recreationally important shark in many areas of its range. Despite of the fishery importance of C. obscurus few genetic resources are currently available for the species. Here, we report on the isolation of eight novel microsatellite loci from C. obscurus using a double-digest restriction site associated DNA (RAD) sequencing approach on the Ion Proton semiconductor platform (ddRADseq-ion). We characterised the loci in 26 individuals and all loci were polymorphic, exhibiting 5-10 alleles (average 6.6), and observed and expected heterozygosities of 0.385-0.962 and 0.479-0.847, respectively. We found that all pairs of loci were in linkage equilibrium and conformed to Hardy-Weinberg expectations. The loci reported in this study are only the second set of microsatellite loci ever characterized for C. obscurus and will be valuable for molecular ecology studies for this vulnerable species.


DNA polymorphism in the conserved 190 kDa antigen gene repeat region among spotted fever group Rickettsiae.
The 190 kDa protein gene of R. rickettsii (R strain) contains a large region of tandemly arranged repeats. Homologous sequences were discovered within the genomes of all but one species of spotted fever group Rickettsiae. Southern blots showed restriction fragment length polymorphisms between species of Rickettsiae indicating structural differences among repeat regions. Further analysis using polymerase chain reaction techniques indicated the repeat regions vary in size among Rickettsiae. In addition, strains of R. rickettsii that vary in virulence were found to contain polymorphisms in this region.


Effect of vitamin B-6 nutrition on the levels of dopamine, dopamine metabolites, dopa decarboxylase activity, tyrosine, and GABA in the developing rat corpus striatum.
Evidence presented in this manuscript indicates that prenatal and postnatal vitamin B-6 undernutrition produces the loss of dopamine (DA) in the corpus striatum of the developing rat brain. The concentration of striatal DA in rats fed vitamin B-6 inadequate diets was significantly lower than rats fed optimal diets at 28 and 56 days of age. The differences in striatal DA concentrations among vitamin B-6 dietary groups was more evident with age. Measurements of the major metabolites of DA, homovanillic acid (HVA), and 3,4-dihydroxyphenylacetic acid (DOPAC) in rat corpus striatum showed a significant decrease in HVA level in B-6 restricted rats compared to B-6 sufficient groups. Striatal DOPAC levels were not significantly different among any of the groups at any age. The activity of dopa decarboxylase holoenzyme was found to be significantly lower in the corpus striatum of rats fed suboptimal B-6 diets. Whether this finding has a significant effect on DA levels has yet to be determined since dopa decarboxylase is not the rate-limiting enzyme in the synthesis of DA. In an effort to determine the underlying mechanism for the loss of striatal DA, the concentration of its precursor tyrosine (TYR) was measured. Vitamin B-6 undernutrition had no significant effect on the levels of TYR in rat corpus striatum. The concentration of the inhibitory neurotransmitter GABA was also measured. The results indicate that as expected the levels of striatal GABA were significantly lower than controls at 14 and 28 days of age. On the other hand, striatal GABA levels in B-6 restricted rats did not differ from controls at 56 days of age.(ABSTRACT TRUNCATED AT 250 WORDS)


A circadian clock in murine bone marrow-derived mast cells modulates IgE-dependent activation in vitro.
Circadian rhythm is expressed in most organisms, and many functions and parameters in the immune system are associated with time-of-day. However, it is largely unknown if local circadian clocks in immune cells directly control physiological outcomes. We hypothesized that a circadian clock in murine bone marrow derived mast cells (BMMCs) modulates IgE-dependent activation in vitro. Mature BMMCs, grown from bone marrow of C57BL/6 mice, were synchronized with serum rich media (50% horse serum). Total RNA was harvested from BMMCs at 4 h intervals for up to 72 h following synchronization and expression of circadian genes (mPer1, mPer2, Bmal1, Rev-erbα, and Dbp) was measured by quantitative PCR. Serum shock synchronized expression of circadian genes (mPer2, Bmal1, Rev-erbα, and Dbp) in BMMCs. Synchronized BMMCs stimulated via the high affinity IgE receptor (FcεRI) at different time intervals display circadian rhythms in IL-13 and IL-6 mRNA expression. The expression of fcer1a gene and FcεRIα protein displayed a circadian pattern following serum shock, with mean periods of 18.9 and 28.6 h, respectively. These results demonstrate that synchronized BMMCs provide an in vitro model to study circadian mechanism(s) associated with allergic disease and that circadian oscillation of cytokine production following IgE-dependent activation is at least in part due to circadian oscillation of FcεRIα.


Microparticles as biomarkers of lung disease: enumeration in biological fluids using lipid bilayer microspheres.
Extracellular vesicles, specifically microparticles (MPs), are rapidly gaining attention for their capacity to act as biomarkers for diagnosis, prognosis, or responsiveness to therapy in lung disease, in keeping with the concept of precision medicine. However, MP analysis by high-sensitivity flow cytometry (FCM) is complicated by a lack of accurate means for MP enumeration. To address this gap, we report here an enhanced FCM MP gating and enumeration technique based on the use of novel engineered lipid bilayer microspheres (LBMs). By comparison of LBM-based MP enumeration with conventional bead- or fluorescent-based FCM enumeration techniques and a gravimetric consumption gold standard, we found LBMs to be superior to commercial bead preparations, showing the smallest fixed bias and limits of agreement in Bland Altman analyses. LBMs had simultaneous capacity to aid FCM enumeration of MPs in plasma, BAL, and cell culture supernatants. LBM enumeration detected differences in MP counts in mice exposed to intraperitoneal lipopolysaccharide or saline. LBMs provided for 1) higher sensitivity for gating MPs populations, 2) reduced background within MP gates, 3) more appropriate size, and 4) an inexpensive alternative amenable to different fluorescent tags. LBM-based MP enumeration was useful for a series of different FCM systems assessed, whereas LBM gating benefited high- but not low-sensitivity FCM systems compared with fluorescence gating. By offering exclusive advantages over current means of gating and enumerating MPs, LBMs are uniquely suited to realizing the potential of MPs as biomarkers in biological lung fluids and facilitating precision medicine in lung disease.


[T2HD Study. Oral anticoagulants and antiplatelet agents: Practices, benefits, and risks in the chronic hemodialysis population. Observational data].
Chronic hemodialysis patients have disorders of hemostasis associated with End Stage Renal Disease, cardiovascular disease, and treatments they receive. They have more thrombotic and bleeding events than the general population. The balance of risks and benefits of drugs is not precisely defined. The aim of this study is to describe the practices patterns concerning oral anticoagulants (AVK) and antiplatelet agents (APA), and to investigate their impact on the survival and the occurrence of thrombotic and bleeding events. Based on REIN registry, we performed a multicenter study of 502 incident patients on hemodialysis in Lorraine from 2009 to 2010. Until June 2013, we collected retrospectively the prescription of oral anticoagulants and APA, the occurrence of thrombotic or hemorrhagic events. During follow-up, there were 550 events, including 454 thrombotic events and 96 bleeding events. The most common event was the dialysis vascular access thrombosis; 60.8% of patients were treated with APA, and 29.5% with AVK. Bleeding and thrombosis occurred the most in the APA plus AVK patient group. Events had often recurrences. Patients survived longer in no anticoagulant or antiplatelet group, but with no statistic difference. Although literature is centered on bleeding events, we found a more significant occurrence of thrombosis than bleeding. Patients without APA or AVK have less comorbidities, less bleeding and thrombotic events, and better survival. Analytic description of our data is expected to get a risk/benefit ratio of APA and AVK treatment in their different uses.


Molecular skins: a new concept for quantitative shape matching of a protein with its small molecule mimics.
A novel analytical method for comparing molecular shapes by optimizing the intersection of molecular "SKINS" has been developed. This method provides a quantitative measure of the shape similarity by maximizing the intersection volume of molecular surfaces with a finite thickness; a molecular skin. We report shape matching of a small tripeptide inhibitor (DFKi) of elastase class proteins with the 56 residue turkey ovomucoid inhibitor (TOMI). To match a large elastase inhibitor such as TOMI with a small inhibitor or drug, we found that it is necessary to use a skin match rather than molecular volume. Skin based comparisons of TOMI protein with DFKi successfully found the alignment expected from comparison of their respective crystallographic complexes with elastase (i.e. HLE/TOMI complex and PPE/tripeptide complex). In the skin comparison of the tripeptide with the TOMI protein, blind searching for skin matches involved optimization of the skin intersection from 172 starting positions randomly selected from a set of 500 points on the TOMI van der Waals surface [within 9.5 A of the Leu-18 on the TOMI binding loop (1 point/A2)]. The tripeptide center of mass was placed at these points and its orientation was randomized before optimization was initiated. The best skin intersection, 86.4 A3, was found three times and corresponds to the experimental alignment. The next best skin intersection was 78.1 A3 giving a discrimination factor in this case of 10%. Searches over the entire surface of the TOMI protein did not identify any new matches with skin intersection greater than 78.1 A3. Matching the DFKi with a TOMI structure relaxed from its crystal conformation by molecular dynamics gives similar results.


Prevalence of Intestinal Parasitic Infections in the Southwest of Iran: A Four-year Retrospective Study.
The current study was aimed to assess the prevalence of intestinal parasitic infections (IPIs) in persons who were referred to selected hospitals of Abadan and Khorramshahr cities, in the southwest of Iran. In this descriptive cross-sectional study, 43200 and 28821 stool specimens were gathered from Abadan and Khorramshahr cities, respectively, during four consecutive years (2014-2017). A decreasing trend was seen in the prevalence rate of IPIs in Abadan and Khorramshahr cities. The overall prevalence of IPIs was estimated to be 5.82% and 6.11% in Abadan and Khorramshahr cities, respectively. In Abadan city, the proportion of IPIs in male persons with 56% was higher than females 44%, while in Khorramshahr city, more prevalence was observed in females (54% vs. 46%). Moreover, according to the season, the highest prevalence was recorded in Spring (30.4%) and Summer (31.1%) in Abadan and Khorramshahr cities, respectively. We found Giardia lamblia, as the most frequent parasite in studied individuals of both cities (Abadan: 2.92%; 1261/43200 and Khorramshahr: 3.15%; 908/28821). Other parasites were as follows: Entamoeba histolytica/dispar, E. coli, E. hartmanni, Blastocystis hominis, Hymenolepis nana, Enterobius vermicularis, Strongyloides stercoralis, and Taenia spp. This was the first study on the prevalence of IPIs in Abadan and Khorramshahr cities, southwest of Iran. Intestinal parasites are still the main public health concern in these regions. Health education, improving sanitary conditions, screening and treatment of infected people, particularly in rural and semi-urban areas can be helpful.


Bovine trypanosomiasis in southern Tanzania: parasitological and serological survey of prevalence.
In a survey for bovine trypanosomiasis blood smears from 1,617 cattle at 72 sites were examined. Trypanosomes were found in 93 cattle, representing 16% of the cattle in herds in which trypanosomiasis was confirmed. Of the positive cattle 56% had infections with T. congolense, 17% T. vivax and 2.2% T. brucei. Five cattle had mixed infections and in 18 cattle the species was not identified. Sera from 1,352 cattle were tested using microelisa. Ten out of 16 sites, at which no trypanosomes were found in blood smears and at which trypanocides were in use, had over 15% seropositive cattle compared with five of 19 sites at which trypanocides were not in use. It was concluded that the microelisa was a useful aid to the diagnosis of bovine trypanosomiasis and that there is a need for accurate records of drug use and livestock movements to be kept. The serious risk of drug resistant strains of trypanosomes emerging due to the uncontrolled use of trypanocides is emphasised.


On the importance of topological descriptors in understanding structure-property relationships.
It has been generally observed in our work that molecular descriptors derived from a molecular graph theory or topological representation of structure play an important and often key role in many QSAR and QSPR models we have developed. These descriptors do not only provide the means to generate a good fit to the observed data used to train the models, but they also provide information that is needed to generate a clear physical interpretation of the underlying structure-activity or property relationships. In addition, these descriptors provide a conformation-independent method of measuring the key features of molecular structure that affect the observed properties of the molecules. These characteristics are exemplified in a model developed to predict critical micelle concentration (CMC). A model is described that exhibits excellent predictive strength, is independent of conformation of the structures used, and that yields a great deal of detail regarding the underlying structure-property relationship driving the observed CMC.


Calculation of the molecular masses of two newly synthesized thermostable enzymes isolated from thermophilic microorganisms.
Two thermostable enzymes synthesized by thermophilic microorganisms were isolated and purified. A thermostable beta-galactosidase was produced in a continuous fermentation process by Bacillus stearothermophilus TP 32 as an intracellular enzyme. After applying different concentration procedures the raw extract enzyme was prepurified on a Sephadex G-200 size exclusion column. The isolated beta-galactosidase fraction was then separated with HPLC on a TSK G 3000 SW size exclusion column to determine the molecular mass based on calibration curves of standard proteins. The other enzyme, a thermostable protease, was synthesized by Bacillus stearothermophilus TP 26 as an extracellular enzyme. After its concentration, the enzyme was purified on a classical size exclusion column (Sephacryl S-200) and on a HPLC size exclusion column (BIO-SIL TSK-250). The micropreparatively isolated fraction was separated again on this HPLC column to determine its molecular mass. The optimum temperature of both enzymes was approximately 75 degrees C.


Deletion of a cyclic AMP receptor protein homologue diminishes germination and affects morphological development of Streptomyces coelicolor.
Open reading frame SCO3571 of Streptomyces coelicolor encodes a protein of the cyclic AMP (cAMP) receptor protein (CRP) superfamily of regulatory proteins. A mutant revealed a dramatic defect in germination, followed by growth delay and earlier sporulation. This phenotype correlates with those of an adenylate cyclase (cya) mutant that cannot synthesize cAMP. This finding suggests that S. coelicolor may use a Cya-cAMP-CRP system to trigger complex physiological processes such as morphogenesis.


Mental illness, problem behaviour, needs and service use in adults with intellectual disability.
Mental ill health in adults with intellectual disability (ID) is a neglected field in psychiatry and thus still widely understudied. This paper provides data on the prevalence of mental illness and problem behaviour and analyses support needs, mental health service use and psychotropic medication in a representative sample of adults with mild to moderate ID. A set of well-established instruments was used to assess the main parameters in n = 371 participants recruited within a cross-sectional epidemiological multicentre study using a stratified randomised cluster sampling. Point prevalence of mental disorders was 10.8 %, that of problem behaviour 45.3 %. Most study participants needed help in specific lower order need areas (e.g., money budgeting, food, accommodation), and these need areas were mostly rated as met. The highest ratios of unmet to met need were found with respect to sexuality issues and with respect to mental health problems. The focus of psychiatric treatment was psychotropic medication. Referring to ICD-10 based diagnostic criteria and consequently avoiding confusing problem behaviour with mental disorders, point prevalence of mental disorders was lower than in the general population. A systematic deficit in meeting mental health problems in adults with ID indicates the need for implementing strategies to maximise the quality of identification and management of mental disorders.


Bisphosphonates for the treatment of postmenopausal osteoporosis: clinical studies of etidronate and alendronate.
Several large, prospective, controlled trials have demonstrated that bisphosphonates such as etidronate and alendronate are effective and generally safe for the treatment of postmenopausal osteoporosis. The results of these trials show a remarkable degree of consistency and indicate that long-term bisphosphonate therapy significantly increases BMD of the spine and of other skeletal sites, reduces the risk of vertebral and nonvertebral fractures, and generally is safe and well tolerated by patients. Data supporting the use of risedronate for the treatment of postmenopausal osteoporosis are presented in an accompanying article.


Clinical performance of the carbohydrate-deficient transferrin (CDT) assay by the Sebia Capillarys2 system in case of cirrhosis. Interest of the Bio-Rad %CDT by HPLC test and Siemens N-Latex CDT kit as putative confirmatory methods.
The CDT assay used to detect chronic alcohol abuse is difficult with cirrhotic patients. This article describes the performances of several CDT assays in case of cirrhosis. The CDT-Capillarys assay by capillary zone electrophoresis was used for initial testing. Two additional methods were tested as putative confirmatory methods. 110 patients with known hepatic status had their CDT measured by the Capillarys2 or alternative methods. Self-reported alcohol intake was used to assess the performances of CDT assays. Capillarys2 performance was lower in case of cirrhosis, many electropherograms displaying various abnormalities. We used the proper separation of the di- and tri-sialotransferrin peaks to select reliable profiles. This selection led to the classification of cirrhotic and non-cirrhotic patients in abusers and abstainers with similar performances. However, no interpretation was available for 54% of the cirrhotic patients and neither the BioRad %CDT by HPLC test, nor the Siemens N-Latex CDT kit was suitable as confirmatory methods for these samples. An attentive profile examination is required for the validation of Capillarys CDT results of cirrhotic patients. Reliability is significantly improved when samples with an improper separation are excluded. To date, no commercial test can confirm the excluded samples.


A novel approach with holmium laser ablation for endoscopic management of intrahepatic biliary stricture.
Hepatolithiasis, featuring high incidence, severe symptoms, and common recurrence, poses a heavy disease burden. Endoscopic management provides an opportunity to cure hepatolithiasis, but fails to properly resolve biliary stricture without additional interventional techniques. An innovative approach towards endoscopic management of biliary stricture is required. Holmium laser ablation was applied to biliary strictures via endoscopic access. Patients' demographic, operative, and follow-up data after receiving holmium laser ablation were retrospectively collected for analysis. A total of 15 patients (4 males and 11 females) underwent stricture ablation by holmium laser via cholangioscopy. All the patients successfully received holmium laser ablation, indicating a technical success rate of 100%. No postoperative mortality or no major perioperative complication was observed. During the follow-up period, the recurrence-free rate was 73% at 2 years and 67% at 5 years. We successfully developed a novel technique of biliary stricture removal by cholangioscopic holmium laser ablation with satisfying clinical outcomes.


The intergenerational transmission of smoking in adulthood: a 25-year study of maternal and offspring maladaptive attributes.
While smoking is a major cause of mortality and morbidity, and maternal smoking is a risk factor for smoking among their offspring, the mechanisms involved in the intergenerational transmission of smoking are not well understood. This study examines the pathways from maternal and adolescent child factors, and the parent-child relationship, to smoking among the adult offspring, approximately 25 years later. Data for the present analysis were based on time waves 2 (T2; 1983) and 7 (T7; 2007-2009) of an on-going study of a community sample of mothers and their children. Offspring and mother X¯ ages were 14.1 and 40.0 years, respectively, at T2, and 36.6 and 65.0 years, respectively, at T7. At T2, trained interviewers administered individual structured interviews. Psychosocial questionnaires were self-administered at T7. Structural equation modeling (SEM) was used to analyze the interrelationships among maternal and offspring attributes (T2 and T7). SEM results indicated a satisfactory model fit (RMSEA=0.052; CFI=0.91; SRMR=0.057), and confirmed hypothesized pathways. One pathway linked maternal maladaptive attributes (T2) to the mother-adolescent child attachment relationship (T2), which was associated with the offspring's maladaptive attributes over time (T2 to T7), which then predicted the adult offspring's smoking (T7). Other pathways highlighted the stability of maternal smoking, the continuity of maladaptive attributes, and less offspring educational attainment as predictors of offspring smoking at T7. Findings suggest the importance of early interventions to treat maternal smoking, maternal and offspring maladaptive attributes, and the mother-child relationship in order to reduce risk factors for the intergenerational transmission of smoking behavior. Interventions which enhance educational success should also prove effective in reducing smoking.


Detection of human histocompatibility (HLA) antigens with an indirect rosette microassay.
An indirect rosette assay which utilizes erythrocytes coupled with purified antiimmunoglobulin (Ig) antibodies was modified into a microassay for detecting HLA allo-and xenoantibodies. The test, which is performed in microtiter plates, is specific, reproducible and can handle large numbers of samples. As an inhibition assay the test can detect HLA antibodies even if mixed with antibodies to other cell surface structures. The rosette microassay is at least 10 times more sensitive than the complement-dependent microcytotoxic test and can use target cells which exhibit low viability or abnormal susceptibility to lysis.


Does the timing of shoulder manipulation under ultrasound-guided cervical nerve root block for frozen shoulder affect the clinical outcome?
Although the clinical outcomes of manipulation under ultrasound-guided fifth and sixth cervical nerve root block for frozen shoulder have been reported, few studies have focused on the timing of manipulation. This study aimed to determine whether the timing of manipulation impacts the clinical outcomes. We retrospectively reviewed the outcomes of 103 frozen shoulder patients (mean age 51.5 years) who underwent manipulation in one shoulder (n = 103 shoulders) between January 2012 and April 2019. Stiff shoulder was defined as limited range of motion in at least three directions, i.e., passive forward flexion of ≤100°, passive external rotation at the side of ≤10°, and internal rotation of ≤L5. The patients were categorized into two groups: those mobilized within 6 months after symptom onset (early group, 44 shoulders) and those mobilized >6 months after symptom onset (late group, 59 shoulders). The range of motion (forward flexion, external rotation, and internal rotation), Japanese Orthopaedic Association shoulder scores, Constant Shoulder Score, and University of California, Los Angeles scores before and 3, 6, and 12 months after manipulation were compared between groups. The late group exhibited significant improvement in forward flexion, external rotation, internal rotation, Japanese Orthopaedic Association scores, Constant Shoulder Score, and University of California, Los Angeles scores at 3 months; forward flexion at 6 months; and forward flexion and University of California, Los Angeles scores at 12 months after manipulation compared to the early group. Our results indicate that timing has a significant influence on the outcome of manipulation for frozen shoulders. The optimal time for manipulation may be >6 months after symptom onset. These findings can be applied in counselling for frozen shoulder patients and for improved outcomes after manipulation.


The establishment of appropriate methods for egg-activation by human PLCZ1 RNA injection into human oocyte.
Phospholipase C-zeta (PLCZ1), a strong candidate of egg-activating sperm factor, can induce Ca2+ oscillations and cause egg activation. For the application of PLCZ1 to clinical use, we examined the pattern of Ca2+ responses and developmental rate by comparing PLCZ1 RNA injection methods with the other current methods, such as cytosolic aspiration, electrical stimulation and ionomycin treatment in human oocytes. We found that the pattern of Ca2+ oscillations after PLCZ1 RNA injection exhibited similar characteristics to that after ICSI treatment. We also determined the optimal concentration of human PLCZ1 RNA to activate the human oocytes. Our findings suggest that human PLCZ1 RNA is a better therapeutic agent to rescue human oocytes from failed activation, leading to normal and efficient development.


Myomaker, and Myomixer-Myomerger-Minion modulate the efficiency of skeletal muscle development with melatonin supplementation through Wnt/β-catenin pathway.
Melatonin, a pleiotropic hormone secreted from the pineal gland, has been shown to exert beneficial effects in muscle regeneration and repair due to its functional diversity, including anti-inflammation, anti-apoptosis, and anti-oxidative activity. However, little is known about the negative role of melatonin in myogenesis. Here, using skeletal muscle cells, we found that melatonin promoted C2C12 cells proliferation and inhibits differentiation both in C2C12 cells and primary myoblasts in mice. Melatonin administration significantly down-regulated differentiation and fusion related genes and inhibited myotube formation both in C2C12 cells and primary myoblasts in mice. RNA-seq showed that melatonin down-regulated essential fusion pore components Myomaker and Myomixer-Myomerger-Minion. Moreover, melatonin suppressed Wnt/β-catenin signaling. Inhibition of GSK3β by LiCl rescued the influence of melatonin on differentiation efficiency, Myomaker, but not Myomxier in C2C12 cells. In conclusion, melatonin inhibits myogenic differentiation, Myomaker, and Myomixer through reducing Wnt/β-catenin signaling. These data establish a link between melatonin and fusogenic membrane proteins Myomaker and Myomixer, and suggest the new perspective of melatonin in treatment or preventment of muscular diseases.


Insulin secretion in vitro and insulin binding to isolated hepatocytes in congenic mice with different H-2 complexes.
Congenic male mice with differences in the H-2 complex have been used to investigate insulin secretion in vitro, insulin binding to isolated hepatocytes, plasma glucose, and serum insulin. Plasma glucose and serum insulin did not show consistent differences in the B10.BR, B10.D2, B10.A, B10.G, B10.M, B10.S, C57/10SCSN, and C3H.OH strains. Isolated islets of Langerhans responded to stimulation with 400 mg/dl glucose with a 3-5-fold increase in insulin secretion rates (2P less than 0.01): B10.BR greater than B10.M greater than C57BL/10SCSN greater than B10.G greater than C3H.OH, B10.D2, B10.A, B10.S. The biphasic pattern of insulin secretion was less distinct in B10.M, B10.G, and C3H.OH mice. The high-affinity constants of insulin binding to isolated hepatocytes at 37 degrees C varied between 4.5 X 10(7) L X mol-1 and 4.5 X 10(8) L X mol-1 (2P less than 0.01): B10.A greater than B10.BR greater than C57BL/10SCSN, B10.S, B10.D2 greater than B10.M, B10.G. The glucose-stimulated insulin secretion from isolated islets of Langerhans and the binding of insulin to isolated hepatocytes correlate to the H-2 complex independently.


[Sarcoma botryoides (embryonal rhabdomyosarcoma) as an unusual diagnosis of cervix polyps].
Embryonal rhabdomyosarcomas of the female genital tract usually occur during infancy in the vagina. Only in rare cases can they be found in the cervix, where they are most commonly seen in adolescence. Their prognosis seems to be good since the introduction of a combination of surgery and chemotherapy. We report a case of cervical embryonal rhabdomysarcoma in IRS stage Ia in a 32-year-old female. After surgical resection and chemotherapy there has been no evidence of disease for 7 months. The case is compared with the published literature and prognostically relevant features, and possible problems in the differential diagnosis in biopsies are discussed.