Posttranslational modifications of Rab proteins cause effective displacement of GDP dissociation inhibitor.
Intracellular vesicular trafficking is regulated by approximately 60 members of the Rab subfamily of small Ras-like GDP/GTP binding proteins. Rab proteins cycle between inactive and active states as well as between cytosolic and membrane bound forms. Membrane extraction/delivery and cytosolic distribution of Rabs is mediated by interaction with the protein GDP dissociation inhibitor (GDI) that binds to prenylated inactive (GDP-bound) Rab proteins. Because the Rab:GDP:GDI complex is of high affinity, the question arises of how GDI can be displaced efficiently from Rab protein in order to allow the necessary recruitment of the Rab to its specific target membrane. While there is strong evidence that DrrA, as a bacterially encoded GDP/GTP exchange factor, contributes to this event, we show here that posttranslational modifications of Rabs can also modulate the affinity for GDI and thus cause effective displacement of GDI from Rab:GDI complexes. These activities have been found associated with the phosphocholination and adenylylation activities of the enzymes AnkX and DrrA/SidM, respectively, from the pathogenic bacterium Legionella pneumophila. Both modifications occur after spontaneous dissociation of Rab:GDI complexes within their natural equilibrium. Therefore, the effective GDI displacement that is observed is caused by inhibition of reformation of Rab:GDI complexes. Interestingly, in contrast to adenylylation by DrrA, AnkX can covalently modify inactive Rabs with high catalytic efficiency even when GDP is bound to the GTPase and hence can inhibit binding of GDI to Rab:GDP complexes. We therefore speculate that human cells could employ similar mechanisms in the absence of infection to effectively displace Rabs from GDI.


[The natural history and laboratory parameters of HIV infection].
Since 1983 the morbidity and mortality rates as well as results of haematological, immunological and (later) HIV serological tests were recorded prospectively for 497 HIV-positive patients during 1837 clinic visits at least twice within at most six months for a median period of observation of 18 months (range 6-64 months). The rate of progression to a higher stage was calculated according to the method of Kaplan-Meier. The rate for asymptomatic patients was 16% after one and 33% after two years; for patients with persisting generalized lymphadenopathy it was 13% and 21%, respectively, for those with AIDS-related complex 28% and 47%, respectively, and for those with AIDS 33% and 82%, respectively. As for results of laboratory tests, patients with progressive disease had significantly lower titres of anti-HIV nuclear antibodies, as well as a higher incidence of HIV-p24 antigen. Haemoglobin levels, platelet and lymphocyte counts and number of CD-4-positive lymphocytes were significantly lower, Neopterin and beta 2-microglobulins higher (P less than 0.01).


Cyclin D3-associated kinase activity is regulated by p27kip1 in BALB/c 3T3 cells.
We report that cyclin D3/cdk4 kinase activity is regulated by p27(kip1) in BALB/c 3T3 cells. The association of p27(kip1) was found to result in inhibition of cyclin D3 activity as measured by immune complex kinase assays utilizing cyclin D3-specific antibodies. The ternary p27(kip1)/cyclin D3/cdk4 complexes do exhibit kinase activity when measured in immune complex kinase assays utilizing p27(kip1)-specific antibodies. The association of p27(kip1) with cyclin D3 was highest in quiescent cells and declined upon mitogenic stimulation, concomitantly with declines in the total level of p27(kip1) protein. The decline in this association could be elicited by PDGF treatment alone; this was not sufficient, however, for activation of cyclin D3 activity, which also required the presence of factors in platelet-poor plasma in the culturing medium. Unlike cyclin D3 activity, which was detected only in growing cells, p27(kip1) kinase activity was present throughout the cell cycle. Since we found that the p27(kip1) activity was dependent on cyclin D3 and cdk4, we compared the substrate specificity of the active ternary complex containing p27(kip1) and the active cyclin D3 lacking p27(kip1) by tryptic phosphopeptide mapping of GST-Rb phosphorylated in vitro and also by comparing the relative phosphorylation activity toward a panel of peptide substrates. We found that ternary p27(kip1)/cyclin D3/cdk4 complexes exhibited a different specificity than the active binary cyclin D3/cdk4 complexes, suggesting that p27(kip1) has the capacity to both inhibit cyclin D/cdk4 activity as well as to modulate cyclin D3/cdk4 activity by altering its substrate preference.


Developing machine-learned potentials to simultaneously capture the dynamics of excess protons and hydroxide ions in classical and path integral simulations.
The transport of excess protons and hydroxide ions in water underlies numerous important chemical and biological processes. Accurately simulating the associated transport mechanisms ideally requires utilizing ab initio molecular dynamics simulations to model the bond breaking and formation involved in proton transfer and path-integral simulations to model the nuclear quantum effects relevant to light hydrogen atoms. These requirements result in a prohibitive computational cost, especially at the time and length scales needed to converge proton transport properties. Here, we present machine-learned potentials (MLPs) that can model both excess protons and hydroxide ions at the generalized gradient approximation and hybrid density functional theory levels of accuracy and use them to perform multiple nanoseconds of both classical and path-integral proton defect simulations at a fraction of the cost of the corresponding ab initio simulations. We show that the MLPs are able to reproduce ab initio trends and converge properties such as the diffusion coefficients of both excess protons and hydroxide ions. We use our multi-nanosecond simulations, which allow us to monitor large numbers of proton transfer events, to analyze the role of hypercoordination in the transport mechanism of the hydroxide ion and provide further evidence for the asymmetry in diffusion between excess protons and hydroxide ions.


Performance of in vitro γH2AX assay in HepG2 cells to predict in vivo genotoxicity.
The γH2AX assay has recently been suggested as a new in vitro assay for detecting genotoxic (GTX) properties of chemicals. This assay is based on the phosphorylation of H2AX histone in response to DNA damage [i.e. induction of double-strand breaks (DSBs)]. Quantification of γH2AX foci using flow cytometry can rapidly detect DNA damage induced by chemicals that cause DNA DSBs. Up to now, only few compounds have been tested with this assay. The main goal of this study was to compare the performance of this automated γH2AX assay with that of standard in vitro genotoxicity assays in predicting in vivo genotoxicity. HepG2 cells were exposed to 64 selected compounds with known GTX properties and subsequently analysed for induction of γH2AX foci. The results of this assay were compared with public data from standard in vitro genotoxicity tests. Accuracy, sensitivity and specificity in predicting in vivo genotoxicity, using the γH2AX assay alone or in combinations with conventional assays, were calculated. Both the γH2AX assay and the bacterial mutagenicity test (Ames) were highly specific for in vivo GTX, whereas chromosomal aberration/micronucleus test (CA/MN) resulted in highest sensitivity. The currently widely used in vitro genotoxicity test battery-Ames test, mouse lymphoma assay (MLA) and CA/MN test-resulted in low accuracy (55-65%) to predict in vivo genotoxicity. Interestingly, the inclusion of γH2AX assay in the standard battery, instead of MLA assay, resulted in higher accuracy (62-70%) compared with other combinations. Advantage of the γH2AX assay in HepG2 cells is its high sensitivity to detect DNA-reactive GTX compounds, although the reduced sensitivity for compounds that require metabolic activation needs to be improved. In conclusion, the automated γH2AX assay can be a useful, fast and cost-effective human cell-based tool for early screening of compounds for in vivo genotoxicity.


rTSbeta as a novel 5-fluorouracil resistance marker of colorectal cancer: a preliminary study.
Colorectal cancer is the most common form of malignancy in Taiwan and the third leading cause of death from cancer, preceded only by lung and hepatic cancers. Colorectal cancer is typically treated by surgical intervention and/or chemotherapy and radiotherapy, if necessary. To date, 5-fluorouracil (5-FU) is the most commonly used anti-cancer chemotherapy drug. However, patients commonly experience resistance to the drug therefore limiting its efficiency. In this study, we measured the expression of rTSbeta in human colon cancer as a novel 5-FU resistance marker. We collected 172 colon cancer samples from 4 different hospitals (including 21 pairs of colon cancer biopsies and 151 pathologic slides of colon cancer). In vitro, we measured the cytotoxicity of 5-FU and 5-FU plus leucovorin in H630 and H630-1 colon cancer cell lines. The results revealed that rTSbeta was expressed in 115 (66.9 %) pathology samples and that tumour expression was higher than in corresponding normal tissue. Survival rates of up to 5 years following treatment was significantly higher for patients without rTSbeta expression than for those with rTSbeta expression (P = 0.0023). In vitro, H630-1 (with rTSbeta overexpression) had significantly higher IC50 of 5-FU than did H630. IC50 of 5-FU decreased when leucovorin was added. Results indicate a close relationship between rTSbeta expression and resistance to the drug 5-FU in human colorectal cancer. These results provide further evidence for rTSbeta expression as a novel 5-FU resistance marker of colorectal cancer.


Investigation of the electronic and physical properties of defect structures responsible for laser-induced damage in DKDP crystals.
Laser-induced damage at near operational laser excitation conditions can limit the performance of potassium dihydrogen phosphate (KH(2)PO(4), or KDP) and its deuterated analog (DKDP) which are currently the only nonlinear optical materials suitable for use in large-aperture laser systems. This process has been attributed to pre-existing damage precursors that were incorporated or formed during growth that have not yet been identified. In this work, we present a novel experimental approach to probe the electronic structure of the damage precursors. The results are modeled assuming a multi-level electronic structure that includes a bottleneck for 532 nm excitation. This model reproduces our experimental observations as well as other well-documented behaviors of laser damage in KDP crystals. Comparison of the electronic structure of known defects in KDP with this model allows for identification of a specific class that we postulate may be the constituent defects in the damage precursors. The experimental results also provide evidence regarding the physical parameters affecting the ability of individual damage precursors to initiate damage, such as their size and defect density; these parameters were found to vary significantly between KDP materials that exhibit different damage performance characteristics.


Comparative doses and costs of antipsychotic medication.
It is clinically useful to have a table listing the equivalent doses of the various neuroleptics to a standard, such as chlorpromazine. This article derives such data by reviewing double-blind controlled studies that used a flexible dosage schedule of neuroleptics in treating schizophrenic patients. Each neuroleptic is then converted to 100-mg chlorpromazine equivalents. This empirically derived dosage comparability table is compared with a similar table derived from the opinions of experts. Since these comparable doses produce equivalent amounts of antipsychotic activity, the cost to provide such medication was then calculated, and a table comparing the costs of the different neuroleptics was constructed. In absolute amounts, the cost differences between drugs are small. However, for any drug, large savings accrue when the largest possible capsule or tablet to achieve the desired dose is prescribed.


Exhaled acetone as a new biomaker of heart failure severity.
Heart failure (HF) is associated with poor prognosis, and the identification of biomarkers of its severity could help in its treatment. In a pilot study, we observed high levels of acetone in the exhaled breath of patients with HF. The present study was designed to evaluate exhaled acetone as a biomarker of HF diagnosis and HF severity. Of 235 patients with systolic dysfunction evaluated between May 2009 and September 2010, 89 patients (HF group) fulfilled inclusion criteria and were compared with sex- and age-matched healthy subjects (control group, n = 20). Patients with HF were grouped according to clinical stability (acute decompensated HF [ADHF], n = 59; chronic HF, n = 30) and submitted to exhaled breath collection. Identification of chemical species was done by gas chromatography-mass spectrometry and quantification by spectrophotometry. Patients with diabetes were excluded. The concentration of exhaled breath acetone (EBA) was higher in the HF group (median, 3.7 μg/L; interquartile range [IQR], 1.69-10.45 μg/L) than in the control group (median, 0.39 μg/L; IQR, 0.30-0.79 μg/L; P < .001) and higher in the ADHF group (median, 7.8 μg/L; IQR, 3.6-15.2 μg/L) than in the chronic HF group (median, 1.22 μg/L; IQR, 0.68-2.19 μg/L; P < .001). The accuracy and sensitivity of this method in the diagnosis of HF and ADHF were about 85%, a value similar to that obtained with B-type natriuretic peptide (BNP). EBA levels differed significantly as a function of severity of HF (New York Heart Association classification, P < .001). There was a positive correlation between EBA and BNP (r = 0.772, P < .001). EBA not only is a promising noninvasive diagnostic method of HF with an accuracy equivalent to BNP but also a new biomarker of HF severity.


Preclinical efficacy evaluation of potential chemopreventive agents in animal carcinogenesis models: methods and results from the NCI Chemoprevention Drug Development Program.
In the NCI, Chemoprevention Branch drug development program, potential chemopreventive agents are evaluated for efficacy against chemical carcinogen-induced tumors in animal models. This paper summarizes the results of 144 agents in 352 tests using various animal efficacy models. Of these results, 146 were positive, representing 85 different agents. The target organs selected for the animals model are representative of high-incidence human cancers. The assays include inhibition of tumors induced by MNU in hamster trachea, DEN in hamster lung, AOM in rat colon (including inhibition of AOM-induced aberrant crypts), MAM in mouse colon, DMBA and MNU in rat mammary glands, DMBA promoted by TPA in mouse skin, and OH-BBN in mouse bladder. The agents tested may be classified into various pharmacological and chemical structural categories that are relevant to their chemopreventive potential. These categories include antiestrogens, antiinflammatories (e.g., NSAIDs), antioxidants, arachidonic acid metabolism inhibitors, GST and GSH enhancers, ODC inhibitors, protein kinase C inhibitors, retinoids and carotenoids, organosulfur compounds, calcium compounds, vitamin D3 and analogs, and phenolic compounds (e.g., flavonoids). The various categories of compounds have different spectra of efficacy in animal models. In hamster lung, GSH-enhancing agents and antioxidants appear to have high potential for inhibiting carcinogenesis. In the colon, NSAIDs and other antiinflammatory agents appear particularly promising. Likewise, NSAIDs are very active in mouse bladder. In rat mammary glands, retinoids and antiestrogens (as would be expected) are efficacious. Several of the chemicals evaluated also appear to be promising chemopreventive agents based on their activity in several of the animal models. Particularly, the ODC inhibitor DFMO was active in the colon, mammary glands, and bladder models, while the dithiolthione, oltipraz, was efficacious in all the models listed above (i.e., lung, colon, mammary glands, skin, and bladder).


Characterization of ectonucleotidase expression in the rat carotid body: regulation by chronic hypoxia.
The carotid body (CB) chemoreflex maintains blood Po2 and Pco2/H+ homeostasis and displays sensory plasticity during exposure to chronic hypoxia. Purinergic signaling via P1 and P2 receptors plays a pivotal role in shaping the afferent discharge at the sensory synapse containing catecholaminergic chemoreceptor (type I) cells, glial-like type II cells, and sensory (petrosal) nerve endings. However, little is known about the family of ectonucleotidases that control synaptic nucleotide levels. Using quantitative PCR (qPCR), we first compared expression levels of ectonucleoside triphosphate diphosphohydrolases (NTPDases1,2,3,5,6) and ecto-5'-nucleotidase (E5'Nt/CD73) mRNAs in juvenile rat CB vs. brain, petrosal ganglia, sympathetic (superior cervical) ganglia, and a sympathoadrenal chromaffin (MAH) cell line. In whole CB extracts, qPCR revealed a high relative expression of surface-located members NTPDase1,2 and E5'Nt/CD73, compared with low NTPDase3 expression. Immunofluorescence staining of CB sections or dissociated CB cultures localized NTPDase2,3 and E5'Nt/CD73 protein to the periphery of type I clusters, and in association with sensory nerve fibers and/or isolated type II cells. Interestingly, in CBs obtained from rats reared under chronic hypobaric hypoxia (~60 kPa, equivalent to 4,300 m) for 5-7 days, in addition to the expected upregulation of tyrosine hydroxylase and VEGF mRNAs, there was a significant upregulation of NTPDase3 and E5'Nt/CD73 mRNA, but a downregulation of NTPDase1 and NTPDase2 relative to normoxic controls. We conclude that NTPDase1,2,3 and E5'Nt/CD73 are the predominant surface-located ectonucleotidases in the rat CB and suggest that their differential regulation during chronic hypoxia may contribute to CB plasticity via control of synaptic ATP, ADP, and adenosine pools.


Testing procedural invariance in the context of health.
Violations of procedural invariance, epitomised by 'classic' preference reversals, have been observed for more than 40 years, and yet the study of this phenomenon in health remains nascent. This is an oversight because such violations pose a challenge to health economics, where choice and valuation methodologies often are used interchangeably. This article reports two experiments that aim to test for preference reversals over outcomes defined by health status, using both 'open' and 'assisted' valuation procedures. Although systematic preference reversals in the direction generally reported in the literature were not observed, the rates of non-systematic reversal were substantial, measuring 35-40%. By analysing the respondents' explanations for their answers, it is clear that many of them used heuristics to answer the questions, and they may have good reasons to do so, which undermines the notion that preferences are always fixed and stable. These results, and others like it, pose a challenge to those who unquestioningly assume procedural invariance and maintain that population preferences should be used to inform health policy.


Role of coupled dynamics in the catalytic activity of prokaryotic-like prolyl-tRNA synthetases.
Prolyl-tRNA synthetases (ProRSs) have been shown to activate both cognate and some noncognate amino acids and attach them to specific tRNA(Pro) substrates. For example, alanine, which is smaller than cognate proline, is misactivated by Escherichia coli ProRS. Mischarged Ala-tRNA(Pro) is hydrolyzed by an editing domain (INS) that is distinct from the activation domain. It was previously shown that deletion of the INS greatly reduced cognate proline activation efficiency. In this study, experimental and computational approaches were used to test the hypothesis that deletion of the INS alters the internal protein dynamics leading to reduced catalytic function. Kinetic studies with two ProRS variants, G217A and E218A, revealed decreased amino acid activation efficiency. Molecular dynamics studies showed motional coupling between the INS and protein segments containing the catalytically important proline-binding loop (PBL, residues 199-206). In particular, the complete deletion of INS, as well as mutation of G217 or E218 to alanine, exhibited significant effects on the motion of the PBL. The presence of coupled dynamics between neighboring protein segments was also observed through in silico mutations and essential dynamics analysis. Altogether, this study demonstrates that structural elements at the editing domain-activation domain interface participate in coupled motions that facilitate amino acid binding and catalysis by bacterial ProRSs, which may explain why truncated or defunct editing domains have been maintained in some systems, despite the lack of catalytic activity.


Insights into the specificity of thioredoxin reductase-thioredoxin interactions. A structural and functional investigation of the yeast thioredoxin system.
The enzymatic activity of thioredoxin reductase enzymes is endowed by at least two redox centers: a flavin and a dithiol/disulfide CXXC motif. The interaction between thioredoxin reductase and thioredoxin is generally species-specific, but the molecular aspects related to this phenomenon remain elusive. Here, we investigated the yeast cytosolic thioredoxin system, which is composed of NADPH, thioredoxin reductase (ScTrxR1), and thioredoxin 1 (ScTrx1) or thioredoxin 2 (ScTrx2). We showed that ScTrxR1 was able to efficiently reduce yeast thioredoxins (mitochondrial and cytosolic) but failed to reduce the human and Escherichia coli thioredoxin counterparts. To gain insights into this specificity, the crystallographic structure of oxidized ScTrxR1 was solved at 2.4 A resolution. The protein topology of the redox centers indicated the necessity of a large structural rearrangement for FAD and thioredoxin reduction using NADPH. Therefore, we modeled a large structural rotation between the two ScTrxR1 domains (based on the previously described crystal structure, PDB code 1F6M ). Employing diverse approaches including enzymatic assays, site-directed mutagenesis, amino acid sequence alignment, and structure comparisons, insights were obtained about the features involved in the species-specificity phenomenon, such as complementary electronic parameters between the surfaces of ScTrxR1 and yeast thioredoxin enzymes and loops and residues (such as Ser(72) in ScTrx2). Finally, structural comparisons and amino acid alignments led us to propose a new classification that includes a larger number of enzymes with thioredoxin reductase activity, neglected in the low/high molecular weight classification.


Allergen immunotherapy in MASK-air users in real-life: Results of a Bayesian mixed-effects model.
Evidence regarding the effectiveness of allergen immunotherapy (AIT) on allergic rhinitis has been provided mostly by randomised controlled trials, with little data from real-life studies. To compare the reported control of allergic rhinitis symptoms in three groups of users of the MASK-air® app: those receiving sublingual AIT (SLIT), those receiving subcutaneous AIT (SCIT), and those receiving no AIT. We assessed the MASK-air® data of European users with self-reported grass pollen allergy, comparing the data reported by patients receiving SLIT, SCIT and no AIT. Outcome variables included the daily impact of allergy symptoms globally and on work (measured by visual analogue scales-VASs), and a combined symptom-medication score (CSMS). We applied Bayesian mixed-effects models, with clustering by patient, country and pollen season. We analysed a total of 42,756 days from 1,093 grass allergy patients, including 18,479 days of users under AIT. Compared to no AIT, SCIT was associated with similar VAS levels and CSMS. Compared to no AIT, SLIT-tablet was associated with lower values of VAS global allergy symptoms (average difference = 7.5 units out of 100; 95% credible interval [95%CrI] = -12.1;-2.8), lower VAS Work (average difference = 5.0; 95%CrI = -8.5;-1.5), and a lower CSMS (average difference = 3.7; 95%CrI = -9.3;2.2). When compared to SCIT, SLIT-tablet was associated with lower VAS global allergy symptoms (average difference = 10.2; 95%CrI = -17.2;-2.8), lower VAS Work (average difference = 7.8; 95%CrI = -15.1;0.2), and a lower CSMS (average difference = 9.3; 95%CrI = -18.5;0.2). In patients with grass pollen allergy, SLIT-tablet, when compared to no AIT and to SCIT, is associated with lower reported symptom severity. Future longitudinal studies following internationally-harmonised standards for performing and reporting real-world data in AIT are needed to better understand its 'real-world' effectiveness.


Development of a patient decision aid for the treatment of localised prostate cancer: a participatory design approach.
To develop a patient decision aid and to prepare an overview of requirements for implementation. We developed a decision aid that fits the preferences of patients and health care professionals to ensure adequate uptake in clinical practice. A participatory design approach was used to acquire insight into preferences regarding the content and design of a decision aid and into barriers and aspects of the decision aid that facilitate implementation in clinical practice. Three focus group interviews with patients, their partners and health care professionals were conducted. A prototype of the decision aid was developed and presented to patients (n = 14) and health care professionals (n = 13) in semi-structured interviews. Patients (n = 5) participated in a usability study. Data were analysed by two independent coders. Health care professionals considered medical information on treatments and side effects as the most important aspect to be included in the decision aid. Patients also focused on nonmedical considerations, such as location. Both expected the decision aid to support patients in making a treatment choice. According to health care professionals, the oncology nurse was the most suitable to discuss the decision aid with patients, while some patients preferred to discuss the patient decision aid with the urologist. The main barrier to implementation of the decision aid was said to be the expectation that it is time and money consuming, while the incorporation of the decision aid into clinical guidelines and basing the content on these guidelines, would promote implementation. By using a participatory design approach a patient decision aid was designed to meet patients' and health care professionals' needs. Insight was also gained on requirements for implementation. Wide-scale implementation of decision aids is desirable. An overview is provided of requirements for implementation to successfully incorporate a decision aid into clinical practice.


Intraoral Cutaneous Hamartomas-Clinicopathologic and Immunohistochemical Characteristics of 3 Cases.
Intraoral cutaneous hamartomas (ICHs) are uncommon mucosal lesions characterized microscopically by a combination of cutaneous structures, including various stages of follicular and sebaceous elements. Due to their rarity, the clinicopathologic and immunohistochemical attributes of ICHs have not been thoroughly delineated. Three cases of ICH were identified from our records, and formalin-fixed paraffin-embedded sections were immunohistochemically stained with antibodies against androgen receptor, estrogen receptor, and progesterone receptor, p63, factor XIIIα, and CD34. All 3 ICHs involved the buccal mucosa with an M:F ratio = 2:1 and mean age = 42.3 years (age range: 27-61 years). ICHs presented as thickened, painless, white and yellow plaques or nodules of long duration, measuring 0.6-1.5 cm. No history of skin graft in the area of the lesions was reported. Histopathologically, the lesions showed aggregates of rudimentary folliculosebaceous structures. Although well-defined piloerector muscles were present in all cases of ICH, bona fide hair follicles and isolated hair shafts were identified only in 1 case. The overlying oral epithelium exhibited epidermis-like morphological features, while inflammation was generally absent. Immunohistochemically, strong and diffuse nuclear staining for androgen receptor and factor XIIIα was observed in the sebaceous glands, and estrogen receptor and p63 reactivity were confined exclusively to the peripheral basal cells, while progesterone receptor staining was negative in ICHs. CD34 diffusely decorated the lesional stroma. In conclusion, ICH is a rare lesion composed of cutaneous elements in an abnormal location. A predilection for the buccal mucosa is reported in the current study.


Structure-activity relationship investigation for benzonaphthyridinone derivatives as novel potent Bruton's tyrosine kinase (BTK) irreversible inhibitors.
Through a structure-based drug design approach, a tricyclic benzonaphthyridinone pharmacophore was used as a starting point for carrying out detailed medicinal structure-activity relationhip (SAR) studies geared toward characterization of a panel of proposed BTK inhibitors, including 6 (QL-X-138), 7 (BMX-IN-1) and 8 (QL47). These studies led to the discovery of the novel potent irreversible BTK inhibitor, compound 18 (CHMFL-BTK-11). Kinetic analysis of compound 18 revealed an irreversible binding efficacy (kinact/Ki) of 0.01 μM-1s-1. Compound 18 potently inhibited BTK kinase Y223 auto-phosphorylation (EC50 < 100 nM), arrested cell cycle in G0/G1 phase, and induced apoptosis in Ramos, MOLM13 and Pfeiffer cells. We believe these features would make 18 a good pharmacological tool for studying BTK-related pathologies.


Noninvasive and quantitative evaluation of movement disorder disability using an infrared depth sensor.
Cerebellar ataxia and Parkinson's disease are neurodegenerative disorders clinically characterized by motor disabilities including gait disturbance. This study aimed to investigate the usefulness of an infrared depth sensor device to quantitatively evaluate gait disturbances in patients with movement disorders. 25 ataxic, 25 Parkinson's disease, and 25 control subjects were enrolled and evaluated their walk. Stride length, feet interval, gait rhythm, and a ratio of the actual walking route length to the linear distance between the start and goal points (A/L ratio) were assessed and compared. Outcome correlations with clinical scales were also analyzed. The average stride length was shorter in ataxic subjects or Parkinson's disease subjects than in control subjects. The average feet interval was larger in ataxic subjects than in control subjects. The stride length coefficient of variation (CV), gait rhythm CV, and average and standard deviations of the A/L ratio were larger in ataxic or Parkinson's disease subjects than in control subjects. Ataxic subjects exhibited significant positive correlations between the CV of stride length or average feet interval and scale for the assessment and rating of ataxia scores or international cooperative ataxia rating scale scores. Parkinson's disease subjects exhibited a significant correlation between the average stride length, CV of stride length, or standard deviation of A/L ratio and unified Parkinson's disease rating scale score. The device used in this study differentiated the characteristics of gait disturbance in each movement disorder and quantitatively evaluated ataxia or Parkinson's disease severity, indicating its potential clinical utility across applications.


The Nocturia Impact Diary: a self-reported impact measure to complement the voiding diary.
Nocturia is a chronic, fluctuating disease that disrupts sleep and has a wide-ranging impact on quality of life. Valid tools to measure the patient-reported impact of nocturia are essential for evaluating the value of treatment, but the available tools are suboptimal. This study reports the development and validation of the Nocturia Impact Diary-an augmented form of the Nocturia Quality of Life questionnaire designed to be completed in conjunction with the widely used 3-day voiding diary. The process comprised three steps: Step 1: Development of a concept pool using the Nocturia Quality of Life questionnaire and data from relevant studies; Step 2: Content validity study; Step 3: Psychometric testing of construct validity, reliability, and sensitivity of the diary in a randomized, placebo-controlled study in patients with nocturia. Step 1: Fourteen items and 4 domains were included in the first draft of the diary. Step 2: Twenty-three patients with nocturia participated in the cognitive debriefing study. Items were adjusted accordingly, and the content validity was high. Step 3: Fifty-six patients were randomized to desmopressin orally disintegrating tablet or placebo. The diary demonstrated high construct validity, with good sensitivity and a good fit to Rasch model, as well as high internal consistency, discriminatory ability, and acceptable sensitivity to change. Results indicated that the diary was unidimensional. The Nocturia Impact Diary is a convenient, validated patient-reported outcome measure. It should be used in conjunction with a voiding diary to capture the real-life consequences of nocturia and its treatment.


Temporal metatranscriptomic patterning in phototrophic Chloroflexi inhabiting a microbial mat in a geothermal spring.
Filamentous anoxygenic phototrophs (FAPs) are abundant members of microbial mat communities inhabiting neutral and alkaline geothermal springs. Natural populations of FAPs related to Chloroflexus spp. and Roseiflexus spp. have been well characterized in Mushroom Spring, where they occur with unicellular cyanobacteria related to Synechococcus spp. strains A and B'. Metatranscriptomic sequencing was applied to the microbial community to determine how FAPs regulate their gene expression in response to fluctuating environmental conditions and resource availability over a diel period. Transcripts for genes involved in the biosynthesis of bacteriochlorophylls (BChls) and photosynthetic reaction centers were much more abundant at night. Both Roseiflexus spp. and Chloroflexus spp. expressed key genes involved in the 3-hydroxypropionate (3-OHP) carbon dioxide fixation bi-cycle during the day, when these FAPs have been thought to perform primarily photoheterotrophic and/or aerobic chemoorganotrophic metabolism. The expression of genes for the synthesis and degradation of storage polymers, including glycogen, polyhydroxyalkanoates and wax esters, suggests that FAPs produce and utilize these compounds at different times during the diel cycle. We summarize these results in a proposed conceptual model for temporal changes in central carbon metabolism and energy production of FAPs living in a natural environment. The model proposes that, at night, Chloroflexus spp. and Roseiflexus spp. synthesize BChl, components of the photosynthetic apparatus, polyhydroxyalkanoates and wax esters in concert with fermentation of glycogen. It further proposes that, in daytime, polyhydroxyalkanoates and wax esters are degraded and used as carbon and electron reserves to support photomixotrophy via the 3-OHP bi-cycle.


Exploring the Contribution to ADHD of Genes Involved in Mendelian Disorders Presenting with Hyperactivity and/or Inattention.
Attention-deficit hyperactivity disorder (ADHD) is a complex neurodevelopmental disorder characterized by hyperactivity, impulsivity, and/or inattention, which are symptoms also observed in many rare genetic disorders. We searched for genes involved in Mendelian disorders presenting with ADHD symptoms in the Online Mendelian Inheritance in Man (OMIM) database, to curate a list of new candidate risk genes for ADHD. We explored the enrichment of functions and pathways in this gene list, and tested whether rare or common variants in these genes are associated with ADHD or with its comorbidities. We identified 139 genes, causal for 137 rare disorders, mainly related to neurodevelopmental and brain function. Most of these Mendelian disorders also present with other psychiatric traits that are often comorbid with ADHD. Using whole exome sequencing (WES) data from 668 ADHD cases, we found rare variants associated with the dimension of the severity of inattention symptoms in three genes: KIF11, WAC, and CRBN. Then, we focused on common variants and identified six genes associated with ADHD (in 19,099 cases and 34,194 controls): MANBA, UQCC2, HIVEP2, FOPX1, KANSL1, and AUH. Furthermore, HIVEP2, FOXP1, and KANSL1 were nominally associated with autism spectrum disorder (ASD) (18,382 cases and 27,969 controls), as well as HIVEP2 with anxiety (7016 cases and 14,475 controls), and FOXP1 with aggression (18,988 individuals), which is in line with the symptomatology of the rare disorders they are responsible for. In conclusion, inspecting Mendelian disorders and the genes responsible for them constitutes a valuable approach for identifying new risk genes and the mechanisms of complex disorders.


Establishing a new falls clinic--conflicting attitudes and inter-sectoral competition affecting the outcome.
Falls clinics are a newer model for falls management among the elderly. Few studies have addressed the impact of the strategy on falls prevention in the healthcare system. The aim of the present study was to describe the social processes that affect the implementation of new strategies in falls management. A newly established falls clinic was chosen as an instrumental case to describe the systematic and comprehensive approach to falls prevention among the elderly. The investigation had a qualitative case study research design with triangulation of sources and methods, including interviews, participant observation and analysis of documents. The study was conducted from January 2006 to June 2007 at a newly established falls clinic at a Danish hospital. Data were analysed qualitatively according to four main themes: The concept of falls, success/failure, competition and expertise. The study showed that the falls clinic was embedded in a context where interests varied at different levels of the organizational hierarchy. In contrast to the political agenda for falls prevention, patients and professionals did not associate falls with chronological age. The biomedical structure of the falls clinic and the hegemonic mode of handling falls prevention may have facilitated falls prevention services and patient trajectories across sectors, but if falls are associated with chronic disease secondary to an unhealthy lifestyle, the individual patient becomes responsible for the falls problem. This may subsequently prevent the patients from seeking timely help from the healthcare system, or patients may drop out of the existing programmes. Future research needs to look at sustainability and dropping out in relation to falls prevention programmes.


Management by objectives for the academic medical center.
Management by Objectives (MBO) is results-oriented, participatory management, which is appropriate for the current era of keen competition for resources. MBO was installed in the author's institution in clinical laboratory divisions meeting prerequisites and fit the system to the local management environment. The results were decidedly positive. Benefits included improved communication, organizational clarity, planning for deadlines, motivation of participants, work load distribution, and productivity. Projects were easier to visualize, track, and coordinate. A natural review cycle of three to four months evolved, with general review twice a year. MBO worked best with one- to two-year-long projects involving problem solving and innovation. The author expects maturity of his MBO application within three years. He recommends MBO for other academic medical center laboratories, provided executive management commits the necessary time and resources to install MBO carefully as a relatively loose, locally administered form of management philosophy, rather than management technology.


CT in the qualitative assessment of emphysema.
To assess the ability of CT to demonstrate the different types of emphysema, the CT appearance was compared with the corresponding barium-impregnated pathologic specimens cut in the transverse plane, in four patients with emphysema. The CT appearance was also compared with the chest radiographs of two patients with irregular emphysema associated with progressive massive fibrosis. The parenchymal destruction visualized on CT in the patients with mild and moderate centriacinar emphysema was distinctly different from that seen in the patient with panacinar emphysema in appearance and distribution. Paraseptal and irregular emphysema were also well demonstrated using CT. Window widths between 800 and 1,500 were found to demonstrate the parenchymal changes most accurately. We conclude that CT can help to identify the presence of emphysema and to distinguish radiologic characteristics of the different types of emphysema.


Characterization of the SRP68/72 interface of human signal recognition particle by systematic site-directed mutagenesis.
The signal recognition particle (SRP) is a ribonucleoprotein complex which is crucial for the delivery of proteins to cellular membranes. Among the six proteins of the eukaryotic SRP, the two largest, SRP68 and SRP72, form a stable SRP68/72 heterodimer of unknown structure which is required for SRP function. Fragments 68e' (residues 530 to 620) and 72b' (residues 1 to 166) participate in the SRP68/72 interface. Both polypeptides were expressed in Escherichia coli and assembled into a complex which was stable at high ionic strength. Disruption of 68e'/72b' and SRP68/72 was achieved by denaturation using moderate concentrations of urea. The four predicted tetratricopeptide repeats (TPR1 to TPR4) of 72b' were required for stable binding of 68e'. Site-directed mutagenesis suggested that they provide the structural framework for the binding of SRP68. Deleting the region between TPR3 and TPR4 (h120) also prevented the formation of a heterodimer, but this predicted alpha-helical region appeared to engage several of its amino acid residues directly at the interface with 68e'. A 39-residue polypeptide (68h, residues 570-605), rich in prolines and containing an invariant aspartic residue at position 585, was found to be active. Mutagenesis scanning of the central region of 68h demonstrated that D585 was solely responsible for the formation of the heterodimer. Coexpression experiments suggested that 72b' protects 68h from proteolytic digestion consistent with the assertion that 68h is accommodated inside a groove formed by the superhelically arranged four TPRs of the N-terminal region of SRP72.


Listener- Versus Speaker-Oriented Disfluencies in Autistic Adults: Insights From Wearable Eye-Tracking and Skin Conductance Within a Live Face-to-Face Paradigm.
Our study addresses three main questions: (a) Do autistics and neurotypicals produce different patterns of disfluencies, depending on the experimenter's direct versus averted gaze? (b) Are these patterns correlated to gender, skin conductance responses, fixations on the experimenter's face, alexithymia, or social anxiety scores? Lastly, (c) can eye-tracking and electrodermal activity data be used in distinguishing listener- versus speaker-oriented disfluencies? Within a live face-to-face paradigm combining a wearable eye-tracker with electrodermal activity sensors, 80 adults (40 autistics, 40 neurotypicals) defined words in front of an experimenter who was either staring at their eyes (direct gaze condition) or looking elsewhere (averted gaze condition). Autistics produce less listener-oriented (uh, um) and more speaker-oriented (prolongations, breath) disfluencies than neurotypicals. In both groups, men produce less um than women. Both autistics' and neurotypicals' speech are influenced by whether their interlocutor systematically looks at them in the eyes or not, but their reactions go in opposite directions. Disfluencies seem to primarily be linguistic phenomena as experienced stress, social attention, alexithymia, and social anxiety scores do not influence any of the reported results. Finally, eye-tracking and electrodermal activity data suggest that laughter could be a listener-oriented disfluency. This article studies disfluencies in a fine-grained way in autistic and neurotypical adults while controlling for social attention, experienced stress, and experimental condition (direct vs. averted gaze). It adds to current literature by (a) enlightening our knowledge of speech in autism, (b) opening new perspectives on disfluency patterns as important signals in social interaction, (c) addressing theoretical issues on the dichotomy between listener- and speaker-oriented disfluencies, and (d) considering understudied phenomena as potential disfluencies (e.g., laughter, breath). https://doi.org/10.23641/asha.23549550.


Advocating for Adolescent and Young Adult Male Sexual and Reproductive Health: A Position Statement From the Society for Adolescent Health and Medicine.
There is a critical need to improve the sexual and reproductive health (SRH) education and care of adolescent and young adult (AYA) males around the globe, as SRH is a basic human right for all AYAs. This special attention toward the SRH of AYA males is warranted given the fact that they often have difficulty accessing SRH services and education relative to their female counterparts and have higher rates of sexual risk behaviors than females. To promote AYA males' SRH and the health of their sexual partners and children, the Society for Adolescent Health and Medicine (SAHM) recommends that leaders in research, policy, public health, and clinical practice develop and implement evidence-based, comprehensive SRH education that supports AYA males at school, within communities and families, and through healthcare services that are developmentally appropriate, gender affirming, inclusive of, and informed by AYA males. Additionally, SAHM recommends that healthcare systems and healthcare professionals (HCPs) across disciplines establish and implement competencies for SRH education and skills preparation to meet the unique needs of AYA males across diverse healthcare and community settings. This statement examines multilevel barriers that AYA males face in accessing comprehensive SRH education and services and makes recommendations aligned with the World Health Organization's 2030 Sustainable Development Goals to address such barriers, improve AYA male SRH, and promote overall gender equity in SRH services.


Predictive value of single-fiber electromyography in the extensor digitorum communis muscle of patients with ocular myasthenia gravis: a retrospective study.
To determine if the jitter in the extensor digitorum communis (EDC) muscle in patients with purely ocular myasthenia gravis (OMG) predicts the subsequent development of generalized myasthenic weakness-Methods: Included in this retrospective study were patients who initially had purely ocular manifestations and had been followed for at least 2 years after onset of myasthenic symptoms. The group consisted of 50 patients with purely ocular weakness, 26 of whom later generalized. All underwent single-fiber electromyography (SFEMG) studies in the EDC muscle at the initial examination : Statistical analysis did not demonstrate that the amount of jitter in the EDC muscle predicted the development of generalized myasthenia gravis (GMG). Also, there was no threshold jitter value that predicted generalized weakness. : SFEMG measurement of jitter in the EDC muscle demonstrates abnormal neuromuscular transmission in many patients with OMG but is not useful in predicting generalization in these patients.


The treatment of hereditary multiple exostosis of the upper extremity.
Thirty of 50 patients with hereditary multiple exostosis developed significant deformities of the arm in one extremity. The degree of deformity is dependent on the location of the osteochondroma. If the osteochondroma is on the radius, deformity will usually be only minimal. If the osteochondroma is at the distal end of the ulna, the epiphysis usually stops growing. We believe that the ulnar collateral ligament then acts as a tether, very similar to that seen in the ulnar clubhand. The radius then has to either bow or dislocate at the elbow, and the wrist displaces ulnarly. We performed operations on 10 patients that consisted of cutting the ulnar collateral ligament, lengthening the ulna, osteotomizing the radius, and removing any osteochondromas. In the young, growing child, staples are placed across the lateral side of the distal radial epiphysis. The cosmetic results of the surgery were very gratifying. Nine patients also had osteochondromas removed from the hands and the forearms.


Selective induction of tyrosine hydroxylase by cell-cell contact in bovine adrenal chromaffin cells is mimicked by plasma membranes.
As a first step towards the identification and purification of the molecule(s) that are involved in cell contact-mediated tyrosine hydroxylase (TH) induction in cultures of bovine adrenal chromaffin cells, we have prepared plasma membranes (PM) from bovine adrenal medulla and tested their ability to mimick cell contact-mediated TH induction in low density chromaffin cultures. PM indeed induced TH in a manner similar to that observed in high density cultures. The maximal TH induction reached by PM corresponded to 69% of that of high density cultures, and half-maximal TH induction was obtained with 12 micrograms of PM per ml of medium. The induction of TH by PM was blocked by alpha-amanitin as observed in high density cultures. Since acetylcholinesterase was neither induced in high density nor in PM-treated low density cultures, an induction of TH as a result of a general increase in protein synthesis was excluded. The cell contact molecule(s) appear to be intrinsic membrane proteins. They were not removed by high or low salt extraction, but solubilized by 50 mM octylglucoside. They were resistant to 0.1% trypsin and heat denaturation but inactivated by 0.01% chymotrypsin. PM isolated from the adrenal cortex, kidney, and liver also induced TH in low density chromaffin cell cultures, although to a smaller extent than PM of the adrenal medulla. In contrast, muscle and erythrocyte PM were inactive. This shows that the cell contact molecule(s) are not restricted to the adrenal medulla, but are also present in some other but not all tissues.


Massively parallel single-molecule manipulation using centrifugal force.
Precise manipulation of single molecules has already led to remarkable insights in physics, chemistry, biology, and medicine. However, widespread adoption of single-molecule techniques has been impeded by equipment cost and the laborious nature of making measurements one molecule at a time. We have solved these issues by developing an approach that enables massively parallel single-molecule force measurements using centrifugal force. This approach is realized in an instrument that we call the centrifuge force microscope in which objects in an orbiting sample are subjected to a calibration-free, macroscopically uniform force-field while their micro-to-nanoscopic motions are observed. We demonstrate high-throughput single-molecule force spectroscopy with this technique by performing thousands of rupture experiments in parallel, characterizing force-dependent unbinding kinetics of an antibody-antigen pair in minutes rather than days. Additionally, we verify the force accuracy of the instrument by measuring the well-established DNA overstretching transition at 66 +/- 3 pN. With significant benefits in efficiency, cost, simplicity, and versatility, single-molecule centrifugation has the potential to expand single-molecule experimentation to a wider range of researchers and experimental systems.


Trial of Oral Diuretics Prior to Discharge Is Not Associated With Improved Outcomes in Decompensated Heart Failure.
Current heart failure guidelines recommend transition of intravenous (IV) diuretics to oral > 24 h prior to hospital discharge. The aim of this study was to determine whether transition to oral diuretics prior to discharge in patients hospitalized with decompensated systolic heart failure (SHF) was associated with improved 30-day events. This was a retrospective cohort study, in which adults admitted to the Loma Linda Medical Center for 3 - 14 days with a primary discharge diagnosis of acute on chronic SHF were included. Mortality data were obtained from the National Death Index, while readmission only to our facility was included. The t-test and Chi-square test were used for analyses. A total of 314 patients were studied. Patients who were managed with guideline-recommended trial of oral diuretics, and patients who continued to receive IV diuretics on the last full hospital day were overall similar in baseline characteristics. Patients who received oral diuretics on the day prior to discharge had longer length of stay, less weight loss, were discharged on lower diuretic doses (all P < 0.05), and had similar outcomes of 30-day readmission and 30-day hospitalization-free survival. The transition to oral diuretics prior to discharge in patients with decompensated SHF was not associated with improved 30-day outcomes. These results require validation in prospective trials but suggest that guideline recommendations regarding transitioning to oral diuretics prior to discharge may deserve re-evaluation.


Peptide-Based Subunit Vaccine Design of T- and B-Cells Multi-Epitopes against Zika Virus Using Immunoinformatics Approaches.
The Zika virus disease, also known as Zika fever is an arboviral disease that became epidemic in the Pacific Islands and had spread to 18 territories of the Americas in 2016. Zika virus disease has been linked to several health problems such as microcephaly and the Guillain-Barré syndrome, but to date, there has been no vaccine available for Zika. Problems related to the development of a vaccine include the vaccination target, which covers pregnant women and children, and the antibody dependent enhancement (ADE), which can be caused by non-neutralizing antibodies. The peptide vaccine was chosen as a focus of this study as a safer platform to develop the Zika vaccine. In this study, a collection of Zika proteomes was used to find the best candidates for T- and B-cell epitopes using the immunoinformatics approach. The most promising T-cell epitopes were mapped using the selected human leukocyte antigen (HLA) alleles, and further molecular docking and dynamics studies showed a good peptide-HLA interaction for the best major histocompatibility complex-II (MHC-II) epitope. The most promising B-cell epitopes include four linear peptides predicted to be cross-reactive with T-cells, and conformational epitopes from two proteins accessible by antibodies in their native biological assembly. It is believed that the use of immunoinformatics methods is a promising strategy against the Zika viral infection in designing an efficacious multiepitope vaccine.


Wheat stem rust recorded for the first time in decades in Ireland.
Wheat stem rust, caused by the fungus Puccinia graminis f. sp. tritici (Pgt), occurs in most wheat-growing areas worldwide, and, in western Europe since 2013, has started to re-emerge after many decades of absence. Following this trend across western Europe, in 2020, we also detected and recorded wheat stem rust for the first time in five decades in experimental plots across five locations in Ireland. To examine the potential origin of the Irish Pgt infection in 2020, we carried out transcriptome sequencing on 12 Pgt-infected wheat samples collected across Ireland and compared these to 76 global P. graminis isolates. This analysis identified a close genetic relationship between the Irish Pgt isolates and those from Ethiopia collected in 2015 after a severe stem rust epidemic caused by the TKTTF Pgt race, and with the UK-01 Pgt isolate that was previously assigned to the TKTTF race. Subsequent pathology-based race profiling designated two Irish isolates and recent UK and French Pgt isolates to the TKTTF Pgt race group. This suggests that the Irish Pgt occurrence most probably originated from recent long-distance windborne dispersal of Pgt urediniospores from neighbouring countries in Europe where we confirmed the Pgt TKTTF race continues to be prevalent. The identification of wheat stem rust in Ireland at multiple locations in 2020 illustrates that the disease can occur in Ireland and emphasizes the need to re-initiate local monitoring for this re-emergent threat to wheat production across western Europe.


Muscle or liver-specific Sirt3 deficiency induces hyperacetylation of mitochondrial proteins without affecting global metabolic homeostasis.
Sirt3 is a mitochondrial sirtuin, predominantly expressed in highly metabolic tissues. Germline ablation of Sirt3 has major metabolic consequences, including increased susceptibility to metabolic damage and oxidative stress after high fat feeding. In order to determine the contribution of liver and skeletal muscle to these phenotypes, we generated muscle-specific Sirt3 (Sirt3(skm-/-)) and liver-specific Sirt3 (Sirt3(hep-/-)) knock-out mice. Despite a marked global hyperacetylation of mitochondrial proteins, Sirt3(skm-/-) and Sirt3(hep-/-) mice did not manifest any overt metabolic phenotype under either chow or high fat diet conditions. Similarly, there was no evidence for increased oxidative stress in muscle or liver when Sirt3 was ablated in a tissue-specific manner. These observations suggest that the mitochondrial hyperacetylation induced by Sirt3-deletion in a tissue specific manner is not necessarily linked to mitochondrial dysfunction and does not recapitulate the metabolic abnormalities observed in the germline Sirt3 knock-out mice.


Differential effects of fibroblast growth factor and tumor promoters on the initiation and maintenance of adipocyte differentiation.
Fibroblast growth factor (FGF) has been shown to inhibit the differentiation of myogenic and adipogenic cell lines without inducing a proliferative response. We have previously shown that agents capable of activating protein kinase C (PKC), such as FGF and the phorbol ester tetradecanoyl phorbol-13-acetate (TPA), inhibit the differentiation of the adipogenic cell line TA1, as measured by the rapid loss of adipocyte-specific RNAs. We report here that the treatment of fully differentiated TA1 adipocytes with FGF at 10 ng/ml induces the reversal of adipocyte differentiation, even in cells where PKC activity has been down-regulated by TPA pretreatment. In contrast, TPA or lower concentrations of FGF (1 ng/ml), both effective inducers of c-fos RNA in adipocytes, fail to reverse adipocyte differentiation. The adipocytes, however, will extinguish differentiation-specific functions in response to TPA by the addition of a calcium ionophore. Therefore, we propose that there are two FGF-sensitive pathways in TA1 cells: one responsible for the initiation of differentiation (TPA sensitive) and another required for maintenance of the adipose phenotype (TPA insensitive). These results suggest that activation of two distinct signaling pathways--one PKC and calcium dependent, the other FGF activated but PKC independent--are capable of inhibiting the biochemical events responsible for the maintenance of adipocyte differentiation.


A multicentre comparison of the efficacy of terbutaline Turbuhaler and salbutamol pressurized metered dose inhaler in hot, humid regions.
Twenty seven adults and 20 children with previously diagnosed stable asthma, using a salbutamol pressurized metered dose inhaler (P-MDI) and living in Cairns, Townsville and Southport, Queensland, Australia participated in a randomized, open-label cross-over comparison of terbutaline administered via Turbuhaler and salbutamol administered via P-MDI. The aim of the study was to compare the clinical effectiveness and patient acceptance of the two treatments in hot, humid regions. Terbutaline was administered via Turbuhaler and salbutamol via P-MDI on at least two occasions per day during each four week treatment period. Spirometry was performed at the start of the study, after the two week run-in and at the end of each treatment period. Patients used diary cards to record morning and evening peak expiratory flows, daily symptom scores and daily intake of beta 2 agonist medication. At the end of the study, patients answered a treatment preference question. Forty six patients completed the study. No statistically significant differences were observed between the two treatments in peak expiratory flow, change in morning peak expiratory flow pre- and post-beta-agonist, daily symptom scores, diurnal variability and spirometry. Forty four percent of patients preferred the terbutaline Turbuhaler and 39% preferred salbutamol P-MDI. Both agents were similarly tolerated. Terbutaline delivered by Turbuhaler is as clinically effective as salbutamol delivered by P-MDI in patients with asthma living in hot, humid regions.


Targeted degrader technologies as prospective SARS-CoV-2 therapies.
COVID-19 remains a severe public health threat despite the WHO declaring an end to the public health emergency in May 2023. Continual development of SARS-CoV-2 variants with resistance to vaccine-induced or natural immunity necessitates constant vigilance as well as new vaccines and therapeutics. Targeted protein degradation (TPD) remains relatively untapped in antiviral drug discovery and holds the promise of attenuating viral resistance development. From a unique structural design perspective, this review covers antiviral degrader merits and challenges by highlighting key coronavirus protein targets and their co-crystal structures, specifically illustrating how TPD strategies can refine existing SARS-CoV-2 3CL protease inhibitors to potentially produce superior protease-degrading agents.


Efficacy of nonfetal human RPE for photoreceptor rescue: a study in dystrophic RCS rats.
This study determines the efficacy of nonfetal human retinal pigment epithelium (RPE) for photoreceptor rescue utilizing the dystrophic RCS rat as an animal model. Eyes from 10- and 49-year-old donors were obtained through the Rochester Eye and Human Parts Bank. The RPE was isolated by enzymatic treatment of the choroid-RPE with 2% dispase for 30 min at 37 degrees C. Mechanically dissociated RPE cells were injected at the superior hemisphere into the subretinal space of dystrophic RCS rats during the fourth postnatal week. Rats receiving vehicle injection served as sham controls. The animals were immunosuppressed with daily cyclosporine injections (10 mg/kg) and sacrificed 30 days posttransplantation for histologic evaluation of the RPE graft and its effect on photoreceptor survival. Transplantation of adult human RPE promoted the survival of photoreceptors in the dystrophic RCS rat. Morphometric analysis of the grafted superior hemisphere demonstrated a threefold increase in photoreceptor cell density (149.2 +/- 50 SD) compared to sham controls (39.7 +/- 31 SD) and the untouched inferior hemisphere (52.8 +/- 28 SD). RPE from the 49-year-old donor was as effective as RPE from the 10-year-old donor in promoting photoreceptor survival. The results of this study in RCS rats suggests that RPE from adult human donors of varied ages is suitable for transplantation and retains the capability to promote survival of photoreceptor cells. This finding opens the possibility of using nonfetal RPE cells in human retinal transplantation.


Immunodiagnosis of swine trichinellosis.
A rapid, sensitive and specific serologic test has been developed for the diagnosis of swine trichinellosis. The ELISA based test utilizes L1 stichosome antigens recovered as excretory-secretory (ES) products from in vitro cultivated muscle larvae. Field studies conducted with 20,000 commercial swine using crude ES antigen demonstrated that the test could detect 98% of the medically significant infections. The test had a false-positive rate of less than 3%. Because of difficulties in regulating the quality and quantity of ES antigen and the need to continually maintain infected laboratory animals for producing the diagnostic reagent, efforts have been made to clone and express the gene(s) encoding the immunodominant ES antigens. To date a cDNA sequence, designated TsA-12, which codes in part for a 53-kDa ES antigen, has been identified and expressed in bacteria. Results demonstrate that TsA-12 is recognized by immune sera and further suggest that the immunodominant 45-, 48- and 53-kDa ES proteins which share antigenic epitopes are distinct glycoproteins.


Double Pylorus: Report of a Case With Endoscopic Follow-Up and Review of the Literature.
Double pylorus is a rare endoscopic finding that has been reported in 0.001% to 0.4% of upper gastrointestinal endoscopies and can be either congenital or acquired. Acquired double pylorus is usually an uncommon complication of peptic ulcer that erodes and creates a fistula between the duodenal bulb and the prepyloric antrum. We describe a case of a 67-year-old man who experienced mild epigastric pain and dyspepsia over the last 6 months. The patient periodically took nonsteroidal anti-inflammatory drugs (NSAIDs) due to joint pain. Esophagogastroduodenoscopy revealed gastritis and a double pylorus. An accessory channel connected the lesser curvature of the prepyloric antrum to the duodenal bulb and the endoscope was able to be passed through both of the ducts. The Helicobacter pylori quick test proved positive. Two years later, a follow-up endoscopy showed that fistula fused with normal pylorus and there was a single large opening.


A double-blind, placebo-controlled trial of Fennel (Foeniculum vulgare) on menopausal symptoms: A high placebo response
The present study aimed to evaluate the effects of oral fennel on menopausal symptoms. This double-blind, randomized, placebo-controlled trial was conducted on 50 postmenopausal women in Mashhad (Iran). Patients were randomly divided into two groups of fennel (n=25) and placebo (n=25). Measurements were performed at baseline and after three months using the Menopause-Specific Quality of Life questionnaire. Both placebo and treatment groups revealed significant improvements in the hot flush score (p<0.001 for fennel and p<0.01 for placebo), night sweats (p=0.007 for fennel and p<0.01 for placebo), sweating (p=0.002 for fennel and p<0.01 for placebo), symptoms of anxiety (p=0.05 for fennel and p=0.001 for placebo), feeling depressed (p<0.01 for fennel and p=0.006 for placebo), and impatience with other people (p<0.01 for fennel and p=0.003 for placebo). There were no significant differences in any menopausal symptoms between the fennel and placebo groups, except for coughing and sneezing when urinating (p=0.03). The failure to indicate a significant effect may have been caused by a high placebo response. It is suggested that future trials should include a placebo run-in phase or design a sequential, parallel study with larger sample sizes to mitigate the placebo effect.


Fidelity and Timing of Spotted Lanternfly (Hemiptera: Fulgoridae) Attack Patterns on Ornamental Trees in the Suburban Landscape.
Invasive herbivores can have dramatic impacts in new environments by altering landscape composition, displacing natives, and causing plant decline and mortality. One of the most recent invasive insects in the United States, the spotted lanternfly (Lycorma delicatula), has the potential to cause substantial economic and environmental impacts in agriculture and forestry. Spotted lanternfly exhibits a broad host range, yet reports of late-season movement from the surrounding landscapes onto select tree species in suburban environments have been reported. In this study, we aimed to evaluate the fidelity of spotted lanternfly attack on specific, individual trees within the same species during this movement period. In 2018 and 2019, we observed that individual red (Acer rubrum L. [Sapindales: Sapindaceae]) and silver maple (Acer saccharinum L. [Sapindales: Sapindaceae]) trees were preferentially attacked over other nearby trees of the same species. Foliar elemental composition was a good predictor of spotted lanternfly attack numbers, indicating that individual variation in nutrients may influence spotted lanternfly attraction to and/or retention on maple trees. Our data also confirm reports of late-season movement from surrounding landscapes throughout autumn. Collectively, our results show that spotted lanternfly exhibits some fidelity to particular trees in the landscape during this movement period. While other potential mechanisms also contribute to host plant selection by spotted lanternfly, our data show that host nutritional profiles influence spotted lanternfly infestation of suburban trees at the landscape scale. Our data establish that late-season infestations of suburban trees by spotted lanternfly occurred and that variation in host quality should be further considered in the management of this invasive insect pest.


Morphological differentiation of tau-green fluorescent protein embryonic stem cells into neurons after co-culture with auditory brain stem slices.
Most types of congenital and acquired hearing loss are caused by loss of sensory hair cells in the inner ear and their respective afferent neurons. Replacement of spiral ganglion neurons (SGN) would therefore be one prioritized step in an attempt to restore sensory neuronal hearing loss. To initiate an SGN repair paradigm we previously transplanted embryonic neuronal tissue and stem cells (SC) into the inner ear in vivo. The results illustrated good survival of the implant. One such repair, however, would not have any clinical significance unless central connections from the implanted SGN could be established. For the purpose of evaluating the effects of cell transplantation on cochlear nucleus (CN) neurons we have established organotypic brain stem (BS) cultures containing the CN. At present we have used in vitro techniques to study the survival and differentiation of tau-green fluorescent protein (GFP) mouse embryonic stem cells (MESC) as a mono- or co-culture with BS slices. For the co-culture, 300 mum thick auditory BS slices encompassing the CN were prepared from postnatal Sprague-Dawley rats. The slices were propagated using the membrane interface method and the CN neurons labeled with DiI. After 5+/-2 days in culture a tau-GFP MESC suspension was deposited next to CN in the BS slice. Following deposition the MESC migrated towards the CN. One and two weeks after transplantation the co-cultures were fixed and immunostained with antibodies raised against neuroprogenitor, neuronal, glial and synaptic vesicle protein markers. Our experiments with the tau-GFP MESC and auditory BS co-cultures show a significant MESC survival but also differentiation into neuronal cells. The findings illustrate the significance of SC and auditory BS co-cultures regarding survival, migration, neuronal differentiation and connections.


miR-143-3p impacts on pulmonary inflammatory factors and cell apoptosis in mice with mycoplasmal pneumonia by regulating TLR4/MyD88/NF-κB pathway.
miR-143-3p is correlated with inflammatory pain responses, such as hsa-miR-143-3p expression reduction in fibromyalgia. The present study aimed to explore the effects of miR-143-3p and Toll-like receptor (TLR) 4/myeloid differentiation factor 88 (MyD88)/NF-κB signaling pathway on pulmonary inflammatory factors levels and alveolar epithelial cell apoptosis in mycoplasmal pneumonia mice. Twenty mice were selected as normal group. The 120 successfully modeled Mycoplasma pneumoniae (MP) infection mice were randomly divided into model group (without any treatment), negative control (NC) group (injected with NC mimic), miR-143-3p mimic group (injected with miR-143-3p mimic), miR-143-3p inhibitor group (injected with miR-143-3p inhibitor), TAK-242 group (treatment with TAK-242), and miR-143-3p inhibitor + TAK-242 group (treatment with miR-143-3p inhibitor + TAK-242). Compared with model group, model mice had up-regulated miR-143-3p expression and decreased MyD88 and p-NF-κB p50 protein expressions (all P<0.05); Model mice treated with miR-143-3p mimic and TAK-242 had reduced interleukin (IL)-2 and tumor necrosis factor (TNF)-α contents and protein expressions of MyD88, p-NF-κB p50, increased IL-10 content, fewer alveolar epithelial cell apoptosis, lower Bax expression and higher Bcl-2 expression (all P<0.05); however, mice with miR-143-3p inhibitor treatment showed opposite trends in terms of above indicators. The exacerbation of mycoplasmal pneumonia caused by miR-143-3p inhibitor was partly improved by miR-143-3p inhibitor + TAK-242 combination treatment (all P<0.05). Therefore, up-regulation of miR-143-3p expression may ameliorate pulmonary inflammatory factors levels and reduce alveolar epithelial cell apoptosis in mycoplasmal pneumonia mice by inhibiting TLR4/MyD88/NF-κB signaling pathway.


Malondialdehyde-acetaldehyde-haptenated protein induces cell death by induction of necrosis and apoptosis in immune cells.
Recent studies have demonstrated that circulating antibodies against malondialdehyde-acetaldehyde (MAA)-haptenated proteins are significantly increased in patients with alcohol-induced cirrhosis and hepatitis and correlate with the severity of liver damage. Additionally, when proteins are haptenated with MAA, they become highly immunogenic in vivo in the absence of adjuvants. However, the mechanism(s) of this immunogenicity are currently unknown. Initial in vitro studies on the effects of MAA-modified proteins on cells demonstrated an increase in cell death at concentrations that were cell type specific and time-dependent. Since immunogenicity due to cell death has been described, we investigated the mechanism(s) by which cell death was occurring. Assessment of cell death in splenocytes after 1 h found significant levels of apoptosis as compared to controls. After 5 h, a significant and dose-dependent necrosis occurred in which cells were exposed to >62.5 microg/ml (43.1 mM) MAA-haptenated protein. After 24 h, exposure to >31.3 microg/ml (21.6 mM) MAA-haptenated protein resulted in significant levels of necrosis, although DNA laddering studies found apoptosis was occurring as well. Morphological changes in the cells were observed by light microscopy that correlated with a "low" forward scatter population by flow cytometry. Since necrosis has been implicated in enhancing both primary and secondary immune responses, and necrosis was predominantly occurring in response to MAA-haptenated proteins, a possible mechanism by which the immunogenicity of MAA modification of proteins in vivo may occur is suggested. Specifically, MAA modification of self proteins may result in the death of various cell types, most likely those in the liver. These necrotic materials may induce anti-MAA antibodies and other auto antibodies, whose levels may then correlate with the severity of ALD.


Effects of melatonin in reducing the toxic effects of doxorubicin.
Anthracycline antibiotics, such as doxorubicin and daunorubicin, constitute a group of wide spectrum therapeutic agents. Application of these drugs in chemotherapy is limited because of their toxic effects. Melatonin, the main secretory product of pineal gland, was recently found as a free radical scavenger and antioxidant. We decided to evaluate the tissue protective effect of melatonin against toxic effects of doxorubicin in six groups of rats. Rats were given doxorubicin (Dx) (45 mg/kg dose), melatonin (MEL) (10 mg/kg), first doxorubicin and then melatonin (DM), first melatonin and then doxorubicin (MD). The degree of kidney, lung, liver and brain cells' alterations were examined biochemically. In doxorubicin-treated group, malondialdehyde (MDA) levels of kidney, lung, liver and brain tissues were significantly increased but glutathione (GSH) levels were decreased compared to control rats. In the group in which first doxorubicin and then melatonin were given, MDA levels were significantly decreased compared to the doxorubicin-treated group. In doxorubicin-treated group, serum levels of creatinine, uric acid, blood urea nitrogen (BUN), Gamma-glutamyl transpeptidase (GGT) and Lactic acid dehydrogenase (LDH) were significantly increased while serum albumin and total protein levels were significantly decreased compared to control rats. Melatonin decreased the intensity of the changes produced by the administration of doxorubicin alone. Melatonin was quite efficient in reducing the formation of lipid peroxidation, restoring the tissue GSH contents and alterations of serum levels.


[Trends in caesarean sections associated with non-clinical factors in a Birthing Educational Center in Mexico City].
To describe the trends in caesarian sections (CS) associated with non-clinical factors in women who attended a birthing educational facility, also known as 'Centro de Educación,' (CEPAPAR) in Mexico City. Data from 992 births of 847 women that occurred from 1988-2000 were analyzed using linear regression to identify the association between non-clinical factors and time-trends in CS. The overall incidence of CS was 32.8%, with an 8% average annual increase. Until 1994 the percentage of CS remained relatively steady at 30%, increasing to 40% in subsequent years. In analyzing trends in CS and their relationship with non-clinical factors from 1988-2000, the increased incidence of CS during this period was higher among births where no birth attendant was present, for first-time mothers, and in larger hospitals (>50 beds). Results show that the increased incidence of CS is associated with non-clinical factors such as size of the hospital and the presence of a birth attendant. A model of support for women which includes information, counseling, and the presence of a birth attendant during labor can contribute to reducing the risk of unnecessary CS in populations such as that studied.


Production and characterization of monoclonal antibodies specific for Shewanella colwelliana exopolysaccharide.
Six monoclonal antibodies were produced to whole cells of Shewanella colwelliana (Aco1 to Aco6) and two (Aco22 to Aco23) to purified exopolysaccharide (EPS). Aco1, -4 to -6, -22, and -23 bound to both the cell surface and the purified EPS, while Aco2 and -3 bound to cells only. The EPS of S. colwelliana was antigenically unique from those of nine other species of marine bacteria that were tested. Mapping studies revealed that all of the EPS-specific monoclonal antibodies bound to the same epitope. This EPS epitope was sensitive to cleavage of ester bonds, but neither pyruvate, acetate, nor terminal nonreducing sugars were required for antigenicity. When S. colwelliana was grown on rich media, most of its EPS was loosely associated with the cell surface.


Spirituality of hopeful risk-taking (Bahala na) among Filipino healthcare workers.
Mental health problems are prevalent among medical frontliners, as demonstrated in the study by Ghaleb et al. in the case of the Eastern Mediterranean Region. As mental health can be strengthened by spirituality, this study probed into the spirituality of bahala na of overseas Filipino health care workers. A phenomenological research was conducted that includes an interview of 17 Filipino Catholic migrant nurses in five countries, on the situations when they find themselves exclaiming "bahala na" in their work as frontliners, and what this means for them. Affirming the notion of bahala na as an expression of hopeful risk-taking, the study showed that the nurses usually exclaim bahala na when they have done everything in their power in a given situation and they are now leaving the rest to God. The spirituality of bahala na helped the nurses face the stresses brought about by the pandemic, and when tapped, can further aid them in fighting against institutional racism to promote health equity in the medical community.


Unique Features of Germline Variation in Five Egyptian Familial Breast Cancer Families Revealed by Exome Sequencing.
Genetic predisposition increases the risk of familial breast cancer. Recent studies indicate that genetic predisposition for familial breast cancer can be ethnic-specific. However, current knowledge of genetic predisposition for the disease is predominantly derived from Western populations. Using this existing information as the sole reference to judge the predisposition in non-Western populations is not adequate and can potentially lead to misdiagnosis. Efforts are required to collect genetic predisposition from non-Western populations. The Egyptian population has high genetic variations in reflecting its divergent ethnic origins, and incident rate of familial breast cancer in Egypt is also higher than the rate in many other populations. Using whole exome sequencing, we investigated genetic predisposition in five Egyptian familial breast cancer families. No pathogenic variants in BRCA1, BRCA2 and other classical breast cancer-predisposition genes were present in these five families. Comparison of the genetic variants with those in Caucasian familial breast cancer showed that variants in the Egyptian families were more variable and heterogeneous than the variants in Caucasian families. Multiple damaging variants in genes of different functional categories were identified either in a single family or shared between families. Our study demonstrates that genetic predisposition in Egyptian breast cancer families may differ from those in other disease populations, and supports a comprehensive screening of local disease families to determine the genetic predisposition in Egyptian familial breast cancer.


How to minimize the attack rate during multiple influenza outbreaks in a heterogeneous population.
If repeated interventions against multiple outbreaks are not feasible, there is an optimal level of control during the first outbreak. Any control measures above that optimal level will lead to an outcome that may be as sub-optimal as that achieved by an intervention that is too weak. We studied this scenario in more detail. An age-stratified ordinary-differential-equation model was constructed to study infectious disease outbreaks and control in a population made up of two groups, adults and children. The model was parameterized using influenza as an example. This model was used to simulate two consecutive outbreaks of the same infectious disease, with an intervention applied only during the first outbreak, and to study how cumulative attack rates were influenced by population composition, strength of inter-group transmission, and different ways of triggering and implementing the interventions. We assumed that recovered individuals are fully immune and the intervention does not confer immunity. The optimal intervention depended on coupling between the two population sub-groups, the length, strength and timing of the intervention, and the population composition. Population heterogeneity affected intervention strategies only for very low cross-transmission between groups. At more realistic values, coupling between the groups led to synchronization of outbreaks and therefore intervention strategies that were optimal in reducing the attack rates for each subgroup and the population overall coincided. For a sustained intervention of low efficacy, early intervention was found to be best, while at high efficacies, a delayed start was better. For short interventions, a delayed start was always advantageous, independent of the intervention efficacy. For most scenarios, starting the intervention after a certain cumulative proportion of children were infected seemed more robust in achieving close to optimal outcomes compared to a strategy that used a specified duration after an outbreak's beginning as the trigger.


Coccidioidomycosis in the U.S. Military: a review.
Coccidioidomycosis has had an impact on the military since the discovery of the causative agent, Coccidioides immitis, over a century ago. The first reports of Coccidioides outbreaks affecting U.S. military personnel were reported by Smith and others during World War II. Since that time, numerous outbreaks and sporadic cases have occurred, affecting both the health and readiness of our armed forces. This article summarizes the impact of Coccidioides sp. on our military troops with a review of the literature, a description of the experience at a tertiary referral hospital treating the disease, and a synopsis of incidence studies conducted at bases in the southwestern United States. The substantial effect that coccidioidomycosis has had on the military supports the development of both newer and more effective treatments as well as a Valley Fever vaccine for prevention of this disease.


The role of macrophages in the in vitro generation of extracellular DNA from apoptotic and necrotic cells.
Cell death is a ubiquitous process that occurs by apoptosis or necrosis depending on the triggering event. While apoptotic and necrotic cells differ biochemically, both are cleared by macrophages for elimination. The process is very efficient, although DNA can appear in the blood in various clinical conditions associated with cell death. To define the role of macrophages in the generation of extracellular DNA, in vitro experiments were performed, assessing the release of DNA into the media of apoptotic or necrotic Jurkat cells cultured with RAW264.7 or J774 macrophage cell lines. DNA was measured by a fluorimetric assay using the dye PicoGreen. In these experiments, while necrotic cells alone did not release DNA, in the presence of macrophages, significant amounts of DNA appeared in the medium. This DNA contained sequences from the Jurkat cells and had reduced molecular weight in comparison to cellular DNA. Furthermore, coculture of macrophages with enucleated necrotic Jurkat cells did not release DNA, suggesting that the DNA came from the dead cell. In contrast, Jurkat cells made apoptotic by treatment with either staurosporine or etoposide spontaneously released DNA, while coculture with macrophages caused a decrease in the DNA released. With apoptotic cells, the DNA in the medium showed low molecular weight and laddering whether or not macrophages were present. Together, these results indicate that macrophages play an important role in the generation of extracellular DNA from dead and dying cells, with the effect dependent on how the cell died.


Exercise Hemodynamics as a Predictor of Myocardial Recovery in LVAD Patients.
Mechanical circulatory support using left ventricular assist devices (LVADs) has been demonstrated to improve survival in patients with advanced heart failure. Left ventricular assist device therapy also promotes reverse ventricular remodeling, which in some cases has led to sufficient myocardial recovery to allow LVAD removal. Identification of suitable patients for LVAD removal however remains challenging. We investigated the hypothesis that invasive assessment of exercise hemodynamics may provide additional information in relation to the assessment of contractile reserve in potential candidates for LVAD explant.


Interdisciplinary treatment of encopresis in individuals with developmental disorders: need and efficacy.
The specific target behavior of encopresis in individuals with mental retardation and developmental disorders (MR/DD) is not well-defined in the literature on encopresis or in the literature on specific interventions with MR/DD populations. The criteria for establishing a diagnosis of encopresis in individuals with MR/DD are not clear. The literature on encopresis was reviewed for two factors: (1) definitions according to three major definitional criteria of age, defecation patterns and etiology; and (2) the relationship between etiology and intervention in specific, MR/DD, and nonspecific populations. This review yielded no clinical description of encopresis in MR/DD children, no guidelines for differentiating encopresis from generalized developmental delay and no substantive treatment guidelines. Although no definitions or treatment formulations specific to MR/DD were identified, information and definitional guidelines derived from the general literature on encopresis are relevant to identification, definition and treatment issues for MR/DD populations. Interdisciplinary or team approaches are appropriate for individuals with multiple problems or developmental disorders and intractable encopresis. An interdisciplinary approach, neurodevelopmental-behavioral intervention, which was successfully employed with a 5-year-old encopretic child with multiple developmental problems, is described.


Spectral clustering of risk score trajectories stratifies sepsis patients by clinical outcome and interventions received.
Sepsis is not a monolithic disease, but a loose collection of symptoms with diverse outcomes. Thus, stratification and subtyping of sepsis patients is of great importance. We examine the temporal evolution of patient state using our previously-published method for computing risk of transition from sepsis into septic shock. Risk trajectories diverge into four clusters following early prediction of septic shock, stratifying by outcome: the highest-risk and lowest-risk groups have a 76.5% and 10.4% prevalence of septic shock, and 43% and 18% mortality, respectively. These clusters differ also in treatments received and median time to shock onset. Analyses reveal the existence of a rapid (30-60 min) transition in risk at the time of threshold crossing. We hypothesize that this transition occurs as a result of the failure of compensatory biological systems to cope with infection, resulting in a bifurcation of low to high risk. Such a collapse, we believe, represents the true onset of septic shock. Thus, this rapid elevation in risk represents a potential new data-driven definition of septic shock.


Human Psychophysical Functions, an Update: Methods for Identifying their form; Estimating their Parameters; and Evaluating the Effects of Important Predictors.
Stevens' power law for the judgments of sensation has a long history in psychology and is used in many psychophysical investigations of the effects of predictors such as group or condition. Stevens' formulation [Formula: see text], where [Formula: see text] is the psychological judgment, P is the physical intensity, and [Formula: see text] is the power law exponent, is usually tested by plotting log [Formula: see text] against log (P). In some, but by no means all, studies, effects on the scale parameter, [Formula: see text], are also investigated. This two-parameter model is simple but known to be flawed, for at least some modalities. Specifically, three-parameter functions that include a threshold parameter produce a better fit for many data sets. In addition, direct non-linear computation of power laws often fit better than regressions of log-transformed variables. However, such potentially flawed methods continue to be used because of assumptions that the approximations are "close enough" as to not to make any difference to the conclusions drawn (or possibly through ignorance the errors in these assumptions). We investigate two modalities in detail: duration and roughness. We show that a three-parameter power law is the best fitting of several plausible models. Comparison between this model and the prevalent two parameter version of Stevens' power law shows significant differences for the parameter estimates with at least medium effect sizes for duration.


Estrogen replacement therapy continuously combined with four different dosages of dydrogesterone: effect on calcium and lipid metabolism.
Lipid and bone metabolism was studied in 165 healthy postmenopausal women treated with 2 mg 17 beta-estradiol continuously combined with one of four doses (2.5, 5, 10, or 15 mg) of dydrogesterone in a double blind randomized study design. Fasting blood and urine samples were drawn at baseline and after 3 and 6 months of treatment. Bone remodeling was significantly reduced in all four treatment groups, as indicated by the decrease in serum corrected calcium, phosphate, and alkaline phosphatase and the urinary calcium/creatinine ratio. A dose response of dydrogesterone on these indices was not found. With all four dosages of dydrogesterone, lipid profile (total cholesterol, high density lipoprotein cholesterol, low density lipoprotein cholesterol, and apolipoproteins) improved significantly; however, this was less pronounced with the highest dydrogesterone dose. Our data suggest that continuously applied dydrogesterone in combined hormone replacement therapy does not annihilate the beneficial effects on bone remodeling and lipid metabolism induced by estrogens.


Anti-Microbial Activity of Phytocannabinoids and Endocannabinoids in the Light of Their Physiological and Pathophysiological Roles.
Antibiotic resistance has become an increasing challenge in the treatment of various infectious diseases, especially those associated with biofilm formation on biotic and abiotic materials. There is an urgent need for new treatment protocols that can also target biofilm-embedded bacteria. Many secondary metabolites of plants possess anti-bacterial activities, and especially the phytocannabinoids of the Cannabis sativa L. varieties have reached a renaissance and attracted much attention for their anti-microbial and anti-biofilm activities at concentrations below the cytotoxic threshold on normal mammalian cells. Accordingly, many synthetic cannabinoids have been designed with the intention to increase the specificity and selectivity of the compounds. The structurally unrelated endocannabinoids have also been found to have anti-microbial and anti-biofilm activities. Recent data suggest for a mutual communication between the endocannabinoid system and the gut microbiota. The present review focuses on the anti-microbial activities of phytocannabinoids and endocannabinoids integrated with some selected issues of their many physiological and pharmacological activities.


Phosphorous(v) Lewis acids: water/base tolerant P3-trimethylated trications.
The water/base intolerance of the previously reported electrophilic phosphonium cations has been overcome by replacing the labile electron-withdrawing groups generally attached to phosphorus (e.g. -F, -OAr, -CF3) with methyl groups. Tri-phosphorus(v) tricationic species, accessible in one-pot from commercially available materials, are air and water/base tolerant, yet are sufficiently Lewis acidic for catalysis.


Enhanced levels of chemiluminescence and platelet activating factor in urease-positive gastric ulcers.
Helicobacter pylori are believed to play an important role in the formation of gastric ulcer in a syndrome characterized by a high urease activity. On the other hand, the production of oxygen radicals and platelet activating factor (PAF) is enhanced in gastric ulcers. The present study is designed to investigate the relationship between the different aspects of gastric mucosal injury, urease activity, oxygen radical production, and PAF content in gastric specimens. Biopsy specimens taken from 35 gastric ulcer patients were studied. Urease activity was detected by a rapid urease test (CLO). Oxygen radical production was measured as a value of luminol-dependent chemiluminescence (ChL) and PAF content was determined by radioimmunoassay in the biopsy samples. The CLO-positive rate was significantly higher in the gastric ulcer group in comparison with that in controls. ChL values and PAF content were significantly increased in gastric ulcers, especially in CLO-positive specimens. The CLO-positive rate, ChL values, and PAF content were also found to be increased at a distant site beyond the ulcer lesions. During the course of macroscopic ulcer healing of CLO-positive cases, the CLO positive level and the ChL values were not significantly decreased, although PAF content was significantly lower. Enhanced oxygen radical and PAF production were observed not only in the ulcer region but also at a distant site from the ulcer in the urease-positive gastric mucosa. The persistent enhancement of ChL values during the healing stage of urease-positive gastric ulcers suggests its involvement in the recurrence of gastric ulcers.


Application of Directigen FLU-A for the detection of influenza A virus in human and nonhuman specimens.
Directigen FLU-A, a new enzyme immunoassay membrane test, rapidly detects influenza A virus antigen in specimens from patients. Nasopharyngeal washes and pharyngeal gargles were used to determine the effectiveness of the assay as applied to different types of routinely collected clinical samples. All specimens had been previously shown to contain influenza A virus by virus isolation in tissue culture. Directigen FLU-A was 90% sensitive (95% confidence interval, 56 to 99.7%) with nasopharyngeal washes but only 39% sensitive (95% confidence interval, 17 to 64%) with pharyngeal gargles (P = 0.018) when used with samples containing similar amounts of infectious virus (50% tissue culture infective dose, 1.0 to 4.5). The intensity of the positive reaction with Directigen FLU-A did not correlate with the amount of virus in the specimens. Directigen FLU-A was found to detect cell-associated antigen more readily than free virus; only 20 infected cells were required to identify cell-associated influenza A virus antigen, whereas the limit of detection for free virus was 1.63 x 10(3) infectious virus particles. These findings suggest that Directigen FLU-A detects the cell-associated antigen present in clinical specimens rather than free virus. In addition, Directigen FLU-A detected avian and swine influenza A viruses in both cloacal swabs (75% sensitivity) and swine lung homogenates (86% sensitivity), indicating its potential usefulness in the surveillance of nonhuman influenza A viruses.