Spatio-temporal climate-based model of dengue infection in Southern, Thailand.
This study explored the spatio-temporal patterns of dengue infection in southern Thailand. Data on monthly-notified cases of dengue fever, over the period of January 1981- May 2014 were collected from the Bureau of Epidemiology, Department of Disease Control, Ministry of Public Health. Weather data over the period of January 2007-May 2014 were obtained from the Thai Meteorological Department. Box and whisker plots were used to study the spatial temporal patterns of dengue incidence. Spearman correlation analysis and time-series adjusted Poisson regression analysis were performed to quantify the relationship between weather and the number of dengue cases. The results show that the highest dengue cases occurred in July in the Gulf of Thailand. Conversely, for the Andaman Sea, the highest dengue cases occurred in June in Phang-Nga, Phuket and Ranong provinces. Only Krabi province had the most dengue cases occurring in July. When we compared dengue transmission duration between the Andaman Sea and the Gulf of Thailand, we found that for the Andaman Sea, Trang province had the longest dengue transmission duration (i.e. June-September) and for the Gulf of Thailand, Nakhon Si Thammarat had the longest dengue transmission duration (i.e. June-September). The number of rainy days and relative humidity were the main predictors of dengue incidence by the Gulf of Thailand, while the amount of rainfall and the temperature were the main predictors of dengue incidence by the Andaman Sea. The time series Poisson regression models provided such goodness-of-fit that the correlation between observed and predicted numbers of dengue incidence exceeded 80%. These models could be used to optimise dengue prevention by predicting trends in dengue incidence. Accurate predictions, for even a few months, provide an invaluable opportunity to mount a vector control intervention or to prepare for hospital demand in the community.


Evaluation of piperonyl butoxide as a deltamethrin synergist for pyrethroid-resistant bed bugs.
An understanding of the mechanisms of insecticide resistance in the bed bug, Cimex lectularius L., has the potential to lead to new approaches for the control of resistant populations. We used the cytochrome P450 monooxygenase (P450) inhibitor piperonyl butoxide (PBO) to assess the role of P450s in deltamethrin resistance in three field-collected bed bug strains, LA-1, CIN-1 and WOR-1. In addition, we exposed two highly resistant strains, CIN-1 and WOR-1 (resistance ratio [RR] >2,500-fold), to dry residues of piperonyl butoxide-synergized pyrethroid formulations to determine the utility of synergism by PBO. Piperonyl butoxide synergized deltamethrin in all three strains, but its impact was variable. The synergistic ratio varied from 40 in CIN-1 to 176 in WOR-1. Because the resistance ratio for each strain after piperonyl butoxide treatment was 174 and 39, respectively, our results suggest that P450s have some involvement in deltamethrin resistance, but other resistance mechanisms must be involved as well. No significant synergistic effect of formulated deltamethrin was observed with the addition of synergized pyrethrins or formulated piperonyl butoxide in the CIN-1 strain, but synergism occurred in the WOR-1 strain. Addition of PBO to pyrethroids is not a comprehensive solution to pyrethroid resistance because strains vary in both overall resistance level and the proportion of that resistance attributable to P450s.


Sleep-disordered breathing in a population-based cohort: behavioral outcomes at 4 and 7 years.
Examine statistical effects of sleep-disordered breathing (SDB) symptom trajectories from 6 months to 7 years on subsequent behavior. Parents in the Avon Longitudinal Study of Parents and Children reported on children's snoring, mouth breathing, and witnessed apnea at ≥2 surveys at 6, 18, 30, 42, 57, and 69 months, and completed the Strengths and Difficulties Questionnaire at 4 (n = 9140) and 7 (n = 8098) years. Cluster analysis produced 5 "Early" (6-42 months) and "Later" (6-69 months) symptom trajectories ("clusters"). Adverse behavioral outcomes were defined by top 10th percentiles on Strengths and Difficulties Questionnaire total and subscales, at 4 and 7 years, in multivariable logistic regression models. The SDB clusters predicted ≈20% to 100% increased odds of problematic behavior, controlling for 15 potential confounders. Early trajectories predicted problematic behavior at 7 years equally well as at 4 years. In Later trajectories, the "Worst Case" cluster, with peak symptoms at 30 months that abated thereafter, nonetheless at 7 years predicted hyperactivity (1.85 [1.30-2.63]), and conduct (1.60 [1.18-2.16]) and peer difficulties (1.37 [1.04-1.80]), whereas a "Later Symptom" cluster predicted emotional difficulties (1.65 [1.21-2.07]) and hyperactivity (1.88 [1.42-2.49]) . The 2 clusters with peak symptoms before 18 months that resolve thereafter still predicted 40% to 50% increased odds of behavior problems at 7 years. In this large, population-based, longitudinal study, early-life SDB symptoms had strong, persistent statistical effects on subsequent behavior in childhood. Findings suggest that SDB symptoms may require attention as early as the first year of life.


MALAT1 affects ovarian cancer cell behavior and patient survival.
Epithelial ovarian cancer (EOC) is one of the most lethal malignancies of the female reproductive organs. Increasing evidence has revealed that long non‑coding RNAs (lncRNAs) participate in tumorigenesis. Metastasis associated lung adenocarcinoma transcript 1 (MALAT1) is an lncRNA and plays a role in various types of tumors. However, the function of MALAT1 on cellular behavior in EOC remains unclear. The current study explored the expression of MALAT1 in ovarian cancer tissues and in EOC cell lines. Quantitative RT‑PCR analysis revealed that the expression of MALAT1 was higher in human ovarian malignant tumor tissues and EOC cells than in normal ovarian tissues and non‑tumorous human ovarian surface epithelial cells, respectively. By analyzing the online database Kaplan‑Meier Plotter, MALAT1 was identified to be correlated with the overall survival (OS) and progression‑free survival (PFS) of patients with ovarian cancer. Furthermore, knockdown of MALAT1 by small interfering RNA (siRNA) significantly decreased EOC cell viability, migration, and invasion. Finally, dual‑luciferase reporter assays demonstrated that MALAT1 interacted with miR‑143‑3p, a miRNA that plays a role in EOC as demonstrated in our previous study. Inhibition of MALAT1 resulted in an increase of miR‑143‑3p expression, leading to a decrease of CMPK protein expression. In conclusion, our results indicated that MALAT1 was overexpressed in EOC. Silencing of MALAT1 decreased EOC cell viability and inhibited EOC cell migration and invasion. These data revealed that MALAT1 may serve as a new therapeutic target of human EOC.


Genome-enabled insights into legume biology.
Legumes are the third-largest family of angiosperms, the second-most-important crop family, and a key source of biological nitrogen in agriculture. Recently, the genome sequences of Glycine max (soybean), Medicago truncatula, and Lotus japonicus were substantially completed. Comparisons among legume genomes reveal a key role for duplication, especially a whole-genome duplication event approximately 58 Mya that is shared by most agriculturally important legumes. A second and more recent genome duplication occurred only in the lineage leading to soybean. Outcomes of genome duplication, including gene fractionation and sub- and neofunctionalization, have played key roles in shaping legume genomes and in the evolution of legume-specific traits. Analysis of legume genome sequences also enables the discovery of legume-specific gene families and provides a framework for genome-wide association mapping that will target phenotypes of special importance in legumes. Translating genomic resources from sequenced species to less studied but still important "orphan" legumes will enhance prospects for world food production.


Developmental plasticity of the carotid chemoafferent pathway in rats that are hypoxic during the prenatal period.
The chemoreflex pathway undergoes postnatal maturation, and the perinatal environment plays a critical role in shaping respiratory control system. We investigated the role of prenatal hypoxia on the maturation of the chemoreflex neural circuits regulating ventilation in rat. Effects of hypoxia (10% O2) from the 5th to the 20th day of gestation were studied on male offspring at birth and on postnatal days 3, 7, 21 and 68. Maturation of the respiratory control system was assessed by in vivo tyrosine hydroxylase (TH) activity measurement in peripheral chemoreceptors (carotid bodies, petrosal ganglia), and in brainstem catecholaminergic cell groups (A2C2c and A1C1 areas in the medulla, A5 and A6 areas in the pons). Resting ventilation and ventilatory response to hypoxia were evaluated as functional sequelae. In peripheral structures, prenatal hypoxia reduced TH activity within the first postnatal week and enhanced it later. In contrast, in central areas, prenatal hypoxia upregulated TH activity within the first postnatal week and downregulated it later. The in vivo TH activity impairment is therefore tissue specific, with an opposite effect on the peripheral and central neural circuits. A shift of the effect of prenatal hypoxia occurred between 1 and 3 weeks, indicating a postnatal temporal effect of prenatal hypoxia. An important period in the development of the chemoafferent pathway occurred between the first and the third postnatal week. Functionally, prenatal hypoxia impaired resting ventilation and ventilatory response to hypoxia. The alterations of the catecholaminergic components of the chemoafferent pathway resulting from prenatal hypoxia might contribute to impair postnatal respiratory behaviour.


Early-life soy exposure and gender-role play behavior in children.
Soy-based infant formula contains high levels of isoflavones. These estrogen-like compounds have been shown to induce changes in sexually dimorphic behaviors in animals exposed in early development. We examined gender-role play behavior in relation to soy-based and non-soy-based infant feeding methods among children in the Avon Longitudinal Study of Parents and Children. We studied 3,664 boys and 3,412 girls. Four exposure categories were created using data from questionnaires administered at 6 and 15 months postpartum: primarily breast, early formula (referent), early soy, and late soy. Gender-role play behavior was assessed using the Pre-School Activities Inventory (PSAI). Associations between infant feeding and PSAI scores at 42 months of age were assessed using linear regression. Post hoc analyses of PSAI scores at 30 and 57 months were also conducted. Early-infancy soy use was reported for approximately 2% of participants. Mean [95% confidence interval (CI)] PSAI scores at 42 months were 62.3 (62.0, 62.6) and 36.9 (36.6, 37.2) for boys and girls, respectively. After adjustment, early soy (vs. early formula) feeding was associated with higher (less feminine) PSAI scores in girls (β = 2.66; 95% CI: 0.19, 5.12) but was not significantly associated with PSAI scores in boys. The association between soy exposure and PSAI scores in girls was substantially attenuated at 30 and 57 months. Although not consistent throughout childhood, early-life soy exposure was associated with less female-typical play behavior in girls at 42 months of age. Soy exposure was not significantly associated with play behavior in boys.


[Inhalation anesthesia and intravenous anesthesia from the medical and economic viewpoint].
The unique pharmacokinetic properties of propofol gave rise to a widespread use of the technique of total intravenous anaesthesia. These properties of propofol are reviewed and compared to those of barbiturates and benzodiazepines. Based on this comparison and with respect to their respective intra- and postoperative effects, a cost-benefit analysis of both inhalational- and intravenous anaesthesia is presented. The choice of an anaesthetic technique must not only be made with regard to medical implications; economical aspects have also to be taken into account without challenge to the quality of care. A consequent use of low-flow techniques and a market oriented purchase of drugs and disposables may allow cost savings in anesthesia.


Heterospecific expression of misrepair-enhancing activity of mucAB in Escherichia coli and Bacillus subtilis.
Enterobacterial plasmid genes mucAB, which possess error-prone repair activity, were cloned and sequenced independently of a sequence previously determined (K.L. Perry, S.J. Elledge, B.B. Mitchell, L. Marsh, and G.C. Walker, Proc. Natl. Acad. Sci. USA 82:4331-4335, 1985). The survival- and mutation-enhancing activities of mucAB ligated to the MLSr promoter of a Bacillus subtilis plasmid in the shuttle vector pTE22R were expressed in B. subtilis as well as in Escherichia coli after mutagenic treatment. mucAB fragments with 5' deletions of various lengths up to the base sequence encoding Ala-26-Gly-27, the putative RecA-mediated cleavage site of the MucA protein, showed mutation-enhancing activity for noninducible lexA3 E. coli when ligated to the MLSr promoter in frame. This activity was lost by extending the deletion downstream. The formations of MucA and MucB proteins in B. subtilis and E. coli were demonstrated by Western blot (immunoblot) analysis. MucA cleavage in Rec+ B. subtilis was observed only after treatment with an alkylating agent and was not observed in RecA- and RecE- strains, whereas in E. coli cleavage was observed in Rec+ cells after treatment with either mitomycin C or an alkylating agent but was not detected in RecA- cells. Common activity of B. subtilis Rec and E. coli RecA in the induction of mutants is suggested.


Array Microcell Method (AMCM) for Serial Electroanalysis.
We describe a method for electrochemical measurement and synthesis based on the combination of a mobile micropipette and a microelectrode array, which we term the array microcell method (AMCM). AMCM has the ability to address single electrodes within a microelectrode array (MEA) and provides a simple, low-cost format to enable versatile electrochemical measurements. In AMCM, a droplet at the tip of a movable micropipette (inner diameter of 50 μm) functions as an electrochemical cell, in which the electrode area is defined by a microelectrode of the array. We also report carbon MEAs that are well suited for AMCM and are fabricated from pyrolyzed photoresist films (PPFs). PPF-MEAs with nominal electrode diameters of 5.5 μm are characterized by AMCM, standard macroscale electrochemical methods, and finite element modeling. The versatility of AMCM is demonstrated by measurement of single Pt microparticles and by electrodeposition of shapecontrolled Pt nanoparticles.


Anomalous composition of musculature of the first dorsal fibro-osseous compartment of the wrist.
The anomalous orientation of musculature of the first dorsal fibro-osseous compartment of the wrist is clinically relevant to De Quervian's stenosing tenosynovitis and reconstructive surgeries. Split insertion of the abductor pollicis longus (APL) is commonly found in chimpanzees, gorillas and gibbons. A comparable identical pattern of anomalous slips in humans is of anthropological and phylogenetic importance and could be a result of atavism. This case report describes an unusual fused muscle belly of the APL and extensor pollicis brevis (EPB), which split into three slips--medial, intermediate and lateral. Further, the medial slip was seen to divide into two tendons, inserting on the base of the first metacarpal along with the intermediate slip. The lateral slip divided into three tendons, inserting into the base of the proximal phalanx, base of the first metacarpal and abductor pollicis brevis muscle. The fusion and unusual insertion pattern of the APL and EPB merits documentation for reconstructive procedures such as tendon transfer and interposition arthroplasty.


Ultrafast and Selective Nanofiltration Enabled by Graphene Oxide Membranes with Unzipped Carbon Nanotube Networks.
Carbon nanomaterials have proven their wide applicability in molecular separation and water purification techniques. Here, an unzipped carbon nanotubes (CNT) embedded graphene oxide (GO) membrane (uCNTm) is reported. The multiwalled CNTs were longitudinally cut into multilayer graphene oxide nanoribbons by a modified Hummer method. To investigate the varying effects of different bandwidths of unzipped CNTs on their properties, four uCNTms were prepared by a vacuum-assisted filtration process. Unzipped-CNTs with different bandwidths were made by unzipping multiwalled CNTs with outer diameters of 0-10, 10-20, 20-30, and 30-50 nm and named uCNTm-1, uCNTm-2, uCNTm-3, and uCNTm-4, respectively. The uCNTms exhibited good stability in different pH solutions, and the water permeability of the composite membranes showed an increasing trend with the increase of the inserted uCNTm's bandwidth up to 107 L·m-2·h-1·bar-1, which was more than 10 times greater than that of pure GO membranes. The composite membranes showed decent dye screening performance with the rejection rate of methylene blue and rhodamine B both greater than 99%.


[Endoscopy, shockwave lithotripsy and local lysis in complicated pigmentary calculi of extra- and intrahepatic bile ducts].
Endoscopy, extracorporeal shockwave lithotripsy (ESWL) and local lysis with alkaline solution of EDTA and bile salts in water were applied in combination in four patients with extra- and intrahepatic pigment stones as well as calcium bilirubinate covered concrements of the biliary tract. In the first patient (a man aged 80 years) a giant concrement of the bile duct was broken up after ESWL by three weeks of local chemical lysis and the fragments were removed by endoscopy. In the second case (man, aged 72), a nonextractable pigment stone was at first reduced in size by four-day local lysis and then removed endoscopically. Intrahepatic pigment stones were completely removed in the other two patients (boy of 12, man of 62) by local lysis only in 3 and 15 weeks, respectively. Even long-term use of the alkaline solution may not cause any serious side effects. Breaking up of stones after size reduction with ESWL of giant stones, size reduction of intact stones and contact lysis of intrahepatic stones are three important indications for chemical dissolution of biliary tract stones, respectively.


DYRK1B blocks canonical and promotes non-canonical Hedgehog signaling through activation of the mTOR/AKT pathway.
Hedgehog (Hh) signaling plays important roles in embryonic development and in tumor formation. Apart from the well-established stimulation of the GLI family of transcription factors, Hh ligands promote the phosphorylation and activation of mTOR and AKT kinases, yet the molecular mechanism underlying these processes are unknown. Here, we identify the DYRK1B kinase as a mediator between Hh signaling and mTOR/AKT activation. In fibroblasts, Hh signaling induces DYRK1B protein expression, resulting in activation of the mTOR/AKT kinase signaling arm. Furthermore, DYRK1B exerts positive and negative feedback regulation on the Hh pathway itself: It negatively interferes with SMO-elicited canonical Hh signaling, while at the same time it provides positive feed-forward functions by promoting AKT-mediated GLI stability. Due to the fact that the mTOR/AKT pathway is itself subject to strong negative feedback regulation, pharmacological inhibition of DYRK1B results in initial upregulation followed by downregulation of AKT phosphorylation and GLI stabilization. Addressing this issue therapeutically, we show that a pharmacological approach combining a DYRK1B antagonist with an mTOR/AKT inhibitor results in strong GLI1 targeting and in pronounced cytotoxicity in human pancreatic and ovarian cancer cells.


Diabetes among Inuit migrants in Denmark.
The study aimed to estimate the prevalence of diabetes and impaired glucose intolerance (IGT) among Inuit migrants living in Denmark, and to compare with findings from Greenland. Further, we analyzed determinants for diabetes and impaired glucose metabolism. Cross-sectional, population-based epidemiological study. This cross-sectional study included randomly selected Inuit migrants in Denmark aged 34 years and above. Diabetes and IGT were diagnosed using the oral glucose tolerance test. Body mass index (BMI) and waist circumference were measured, and blood samples were taken from each subject. Socio-demographic characteristics were investigated using a questionnaire. For comparison, data from the Greenland Population Study were used (n = 917). Of 506 eligible subjects, 256 (51%) participated. Twenty-six subjects had diabetes (10.2%) and twenty-eight had IGT (10.9%). Of those with diabetes, 64% had not been previously diagnosed. The prevalences of diabetes and IGT were not significantly different from those among Inuit in Greenland. Significant predictors of diabetes and impaired glucose metabolism (IGM) were found to be age, waist circumference and physical inactivity. The association between waist circumference and diabetes was significantly stronger among Inuit migrants in Denmark than among Inuit in Greenland. The prevalence of diabetes is high among the Inuit migrants in Denmark. However, unlike that reported in most studies, the prevalence was not significantly higher in the migrant population compared with the population of origin.


The adhesion G protein-coupled receptor G2 (ADGRG2/GPR64) constitutively activates SRE and NFκB and is involved in cell adhesion and migration.
Adhesion G protein-coupled receptors (ADGRs) are believed to be activated by auto-proteolytic cleavage of their very large extracellular N-terminal domains normally acting as a negative regulator of the intrinsically constitutively active seven transmembrane domain. ADGRG2 (or GPR64) which originally was described to be expressed in the epididymis and studied for its potential role in male fertility, is highly up-regulated in a number of carcinomas, including breast cancer. Here, we demonstrate that ADGRG2 is a functional receptor, which in transfected HEK293 cells signals with constitutive activity through the adhesion- and migration-related transcription factors serum response element (SRE) and nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB) presumably via coupling to Gα12/13 and Gαq. However, activation of these two pathways appears to occur through distinct molecular activation mechanisms as auto-proteolytic cleavage is essential for SRE activation but not required for NFκB signaling. The overall activation mechanism for ADGRG2 is clearly distinct from the established ADGR activation mechanism as it requires the large extracellular N-terminal domain for proper intracellular signal transduction. Knockdown of ADGRG2 by siRNA in the highly motile breast cancer cell lines Hs578T and MDA-MB-231 resulted in a strong reduction in cell adhesion and subsequent cell migration which was associated with a selective reduction in RelB, an NFκB family member. It is concluded that the adhesion GPCR ADGRG2 is critically involved in the adhesion and migration of certain breast cancer cells through mechanisms including a non-canonical NFkB pathway and that ADGRG2 could be a target for treatment of certain types of cancer.


cDNA microarray in isolation of novel differentially expressed genes related to human glioma and clone of a novel full-length gene.
This investigation was undertaken to obtain differentially expressed genes related to human glioma using cDNA microarray and the characterization of one novel full-length gene. Total RNA was extracted from human glioma tissues and normal brain tissues, and mRNA was used to make probes. After hybridization and washing, the results were scanned using a computer system. The gene named 681F05 clone was an expressed gene to human glioma through four-time hybridization and scanning. Subsequently northern blot analysis was performed by northern blot, 5'RACE and bioinformatics. Fifteen differentially expressed genes to human glioma were obtained through four-time hybridization and scanning. Northern blot analysis confirmed that 681F05 clone was low-expressed in human brain tissues and over-expressed in human glioma tissues. The analysis of BLASTn and BLASTx showed that 681F05 clone is two cDNA clones encoding two novel proteins that are highly identified to the cyclophilin isoform 10 of C. Elgans, respectively. Sequence analysis revealed the two cDNA clones are two different splicing variants of a novel cycophilin-like gene (PPIL3a and PPIL3b). cDNA microarray technology can be successfully used to identify differentially expressed genes. The novel full-length gene of human PPIL3 may be correlated with the formation of human glioma.


A new liquid homogeneous assay for HDL cholesterol determination evaluated in seven laboratories in Europe and the United States.
We evaluated a new liquid homogeneous assay for the direct measurement of high density lipoprotein cholesterol (HDL-C Plus) in seven laboratories. The assay includes two reagents which can be readily used in most available clinical chemistry analyzers. The total CVs of the new method were below 4.6% and the bias in relation to the designated comparison method was below 3.9%. The total error ranged between 4 to 7%. HDL-C values determined by this method were in good agreement with those obtained by the old homogeneous assay using lyophilized reagents, and other homogeneous and precipitation assays (0.944 < r < 0.996). The assay was linear up to at least 3.89 mmol/l HDL-C. Hemoglobin did not interfere, whereas in icteric samples slight deviations were observed. Lipemia up to 11.3 to 22.6 mmol/l triglycerides did not interfere with this homogeneous HDL-C assay. In samples of patients with paraproteinemia, discrepant results were seen. This liquid homogeneous HDL-C assay was easy to handle and produced similar results in all laboratories participating in this study. This method will enable clinical laboratories to reliably measure HDL-C for risk assessment of coronary heart disease.


Identification, evolution, and expression of GDSL-type Esterase/Lipase (GELP) gene family in three cotton species: a bioinformatic analysis.
GDSL esterase/lipases (GELPs) play important roles in plant growth, development, and response to biotic and abiotic stresses. Presently, an extensive and in-depth analysis of GELP family genes in cotton is still not clear enough, which greatly limits the further understanding of cotton GELP function and regulatory mechanism. A total of 389 GELP family genes were identified in three cotton species of Gossypium hirsutum (193), G. arboreum (97), and G. raimondii (99). These GELPs could be classified into three groups and eight subgroups, with the GELPs in same group to have similar gene structures and conserved motifs. Evolutionary event analysis showed that the GELP family genes tend to be diversified at the spatial dimension and certain conservative at the time dimension, with a trend of potential continuous expansion in the future. The orthologous or paralogous GELPs among different genomes/subgenomes indicated the inheritance from genome-wide duplication during polyploidization, and the paralogous GELPs were derived from chromosomal segment duplication or tandem replication. GELP genes in the A/D subgenome underwent at least three large-scale replication events in the evolutionary process during the period of 0.6-3.2 MYA, with two large-scale evolutionary events between 0.6-1.8 MYA that were associated with tetraploidization, and the large-scale duplication between 2.6-9.1 MYA that occurred during diploidization. The cotton GELPs indicated diverse expression patterns in tissue development, ovule and fiber growth, and in response to biotic and abiotic stresses, combining the existing cis-elements in the promoter regions, suggesting the GELPs involvements of functions to be diversification and of the mechanisms to be a hormone-mediated manner. Our results provide a systematic and comprehensive understanding the function and regulatory mechanism of cotton GELP family, and offer an effective reference for in-depth genetic improvement utilization of cotton GELPs.


Use of recombinant factor VIIa in patients with warfarin-associated intracranial hemorrhage.
Warfarin-associated intracranial hemorrhage (ICH) requires rapid normalization of clotting function. Current therapies are associated with significant complications and/or prolonged time to correction of coagulopathy. Recombinant factor VIIa (FVIIa) might allow faster and safer correction of coagulopathy. This article presents a retrospective chart review of all patients with warfarin-associated ICH treated in a neurology/neurosurgery intensive care unit over an 11-month period. All patients were treated to rapidly reverse the warfarin effect. Fifteen patients received vitamin K and fresh frozen plasma (FFP) alone (FFP group). Twelve patients also received FVIIa (FVIIa group). The median times from presentation to an international normalization ratio (INR) of less than 1.3 were 32.2 and 8.8 hours in the FFP the FVIIa groups, respectively (p = 0.016). INR normalized slowly (at 110 and 130 hours, respectively) in two patients with end-stage renal failure who were given FVIIa, one of whom developed disseminated intravascular coagulation after three doses of FVIIa. No other complications occurred from FVIIa administration. One patient in the FFP group developed severe pulmonary edema. FVIIa may be an effective adjunct to FFP in warfarin-related ICH, facilitating faster correction of INR and decreasing FFP requirements. A prospective, randomized trial is needed to confirm these preliminary findings and to determine whether there is a clinical benefit.


Structural and functional energetic linkages in allosteric regulation of muscle pyruvate kinase.
The understanding of the molecular mechanisms of allostery in rabbit muscle pyruvate kinase (RMPK) is still in its infancy. Although, there is a paucity of knowledge on the ground rules on how its functions are regulated, RMPK is an ideal system to address basic questions regarding the fundamental chemical principles governing the regulatory mechanisms about this enzyme which has a TIM (α/β)(8) barrel structural motif [Copley, R. R., and Bork, P. (2000). Homology among (βα)8 barrels: Implications for the evolution of metabolic pathways. J. Mol. Biol.303, 627-640; Farber, G. K., and Petsko, G. A. (1990). The evolution of α/ß barrel enzymes. Trends Biochem.15, 228-234; Gerlt, J. A., and Babbitt, P. C. (2001). Divergent evolution of enzymatic function: Mechanistically diverse superfamilies and functionally distinct superfamilies. Annu. Rev. Biochem.70, 209-246; Heggi, H., and Gerstein, M. (1999). The relationship between protein structure and function: A comprehensive survey with application to the yeast genome. J. Mol. Biol.288, 147-164; Wierenga, R. K. (2001). The TIM-barrel fold: A versatile framework for efficient enzymes. FEB Lett.492, 193-198]. RMPK is a homotetramer. Each subunit consists of 530 amino acids and multiple domains. The active site resides between the A and B domains. Besides the basic TIM-barrel motif, RMPK also exhibits looped-out regions in the α/β barrel of each monomer forming the B- and C-domains. The two isozymes of PK, namely, the kidney and muscle isozymes, exhibit very different allosteric behaviors under the same experimental condition. The only amino acid sequence differences between the mammalian kidney and muscle PK isozymes are located in the C-domain and are involved in intersubunit interactions. Thus, embedded in these two isozymes of PK are the rules involved in engineering the popular TIM (α/β)(8) motif to modulate its allosteric properties. The PK system exhibits a lot of the properties that will allow mining of the ground rules governing the correlative linkages between sequence-fold-function. In this chapter, we review the approaches to acquire the fundamental functional and structural energetics that establish the linkages among this intricate network of linked multiequilibria. Results from these diverse approaches are integrated to establish a working model to represent the complex network of multiple linked reactions which ultimately leads to the observation of allosteric regulation of PK.


The nucleolus as a multiphase liquid condensate.
The nucleolus is the most prominent nuclear body and serves a fundamentally important biological role as a site of ribonucleoprotein particle assembly, primarily dedicated to ribosome biogenesis. Despite being one of the first intracellular structures visualized historically, the biophysical rules governing its assembly and function are only starting to become clear. Recent studies have provided increasing support for the concept that the nucleolus represents a multilayered biomolecular condensate, whose formation by liquid-liquid phase separation (LLPS) facilitates the initial steps of ribosome biogenesis and other functions. Here, we review these biophysical insights in the context of the molecular and cell biology of the nucleolus. We discuss how nucleolar function is linked to its organization as a multiphase condensate and how dysregulation of this organization could provide insights into still poorly understood aspects of nucleolus-associated diseases, including cancer, ribosomopathies and neurodegeneration as well as ageing. We suggest that the LLPS model provides the starting point for a unifying quantitative framework for the assembly, structural maintenance and function of the nucleolus, with implications for gene regulation and ribonucleoprotein particle assembly throughout the nucleus. The LLPS concept is also likely useful in designing new therapeutic strategies to target nucleolar dysfunction.


Curcumin and o-Vanillin Exhibit Evidence of Senolytic Activity in Human IVD Cells In Vitro.
Curcumin and o-Vanillin cleared senescent intervertebral disc (IVD) cells and reduced the senescence-associated secretory phenotype (SASP) associated with inflammation and back pain. Cells from degenerate and non-mildly-degenerate human IVD were obtained from organ donors and from patients undergoing surgery for low back pain. Gene expression of senescence and SASP markers was evaluated by RT-qPCR in isolated cells, and protein expression of senescence, proliferation, and apoptotic markers was evaluated by immunocytochemistry (ICC). The expression levels of SASP factors were evaluated by enzyme-linked immunosorbent assay (ELISA). Matrix synthesis was verified with safranin-O staining and the Dimethyl-Methylene Blue Assay for proteoglycan content. Western blotting and ICC were used to determine the molecular pathways targeted by the drugs. We found a 40% higher level of senescent cells in degenerate compared to non-mildly-degenerate discs from unrelated individuals and a 10% higher level in degenerate compared to non-mildly-degenerate discs from the same individual. Higher levels of senescence were associated with increased SASP. Both drugs cleared senescent cells, and treatment increased the number of proliferating as well as apoptotic cells in cultures from degenerate IVDs. The expression of SASP factors was decreased, and matrix synthesis increased following treatment. These effects were mediated through the Nrf2 and NFkB pathways.


Atomic modeling of cryo-electron microscopy reconstructions--joint refinement of model and imaging parameters.
When refining the fit of component atomic structures into electron microscopic reconstructions, use of a resolution-dependent atomic density function makes it possible to jointly optimize the atomic model and imaging parameters of the microscope. Atomic density is calculated by one-dimensional Fourier transform of atomic form factors convoluted with a microscope envelope correction and a low-pass filter, allowing refinement of imaging parameters such as resolution, by optimizing the agreement of calculated and experimental maps. A similar approach allows refinement of atomic displacement parameters, providing indications of molecular flexibility even at low resolution. A modest improvement in atomic coordinates is possible following optimization of these additional parameters. Methods have been implemented in a Python program that can be used in stand-alone mode for rigid-group refinement, or embedded in other optimizers for flexible refinement with stereochemical restraints. The approach is demonstrated with refinements of virus and chaperonin structures at resolutions of 9 through 4.5 Å, representing regimes where rigid-group and fully flexible parameterizations are appropriate. Through comparisons to known crystal structures, flexible fitting by RSRef is shown to be an improvement relative to other methods and to generate models with all-atom rms accuracies of 1.5-2.5 Å at resolutions of 4.5-6 Å.


Double-hadron leptoproduction in the nuclear medium.
The first measurements of double-hadron production in deep-inelastic scattering within the nuclear medium were made with the HERMES spectrometer at DESY HERA using a 27.6 GeV positron beam. By comparing data for deuterium, nitrogen, krypton, and xenon nuclei, the influence of the nuclear medium on the ratio of double-hadron to single-hadron yields was investigated. Nuclear effects on the additional hadron are clearly observed, but with little or no difference among nitrogen, krypton, or xenon, and with smaller magnitude than effects seen on previously measured single-hadron multiplicities. The data are compared with models based on partonic energy loss or prehadronic scattering and with a model based on a purely absorptive treatment of the final-state interactions. Thus, the double-hadron ratio provides an additional tool for studying modifications of hadronization in nuclear matter.


The effect of Early Head Start on child welfare system involvement: A first look at longitudinal child maltreatment outcomes.
The high societal and personal costs of child maltreatment make identification of effective early prevention programs a high research priority. Early Head Start (EHS), a dual generational program serving low-income families with children prenatally through age three years, is one of the largest federally funded programs for infants and toddlers in the United States. A national randomized trial found EHS to be effective in improving parent and child outcomes, but its effectiveness in reducing child maltreatment was not assessed. The current study used administrative data from state child welfare agencies to examine the impact of EHS on documented abuse and neglect among children from seven of the original seventeen programs in the national EHS randomized controlled trial. Results indicated that children in EHS had significantly fewer child welfare encounters between the ages of five and nine years than did children in the control group, and that EHS slowed the rate of subsequent encounters. Additionally, compared to children in the control group, children in EHS were less likely to have a substantiated report of physical or sexual abuse, but more likely to have a substantiated report of neglect. These findings suggest that EHS may be effective in reducing child maltreatment among low-income children, in particular, physical and sexual abuse.


A role for cholinesterases in tumorigenesis?
Hydrolysis of the neurotransmitter acetylcholine by acetylcholinesterase (ACHE) and butyrylcholinesterase (BCHE) is the rate-limiting step in the termination of cholinergic signaling at neuromuscular junctions. A growing body of evidence suggests that these enzymes also play a role in tumorigenesis. The ACHE and BCHE genes are amplified, mutated, and/or aberrantly expressed in a variety of human tumor types. These changes could be the result of chromosome breakage, since there is an unusually high frequency of chromosomal abnormalities near the map positions of these genes (3q26-ter and 11p-ter, respectively) in such tumors, particularly hemopoietic malignancies. Both ACHE and BCHE contain the consensus peptide motif S/T-P-X-Z, which is found in many substrates of cdc2-related protein kinases. Here we consider the intriguing possibility that phosphorylation by cdc2-related kinases may be the molecular mechanism linking cholinesterases with tumor cell proliferation. We also discuss the notion that inhibition of these enzymes by commonly used organophosphorous poisons may be tumorigenic in humans.


Personalized automated treatment planning for breast plus locoregional lymph nodes using Hybrid RapidArc.
Breast cancer patients who require locoregional lymph node (LLN) irradiation can be treated using a hybrid RapidArc technique combining 2 tangential and 3 RapidArc fields. Because the creation of hybrid RapidArc plans is complex and labor-intensive, we developed an automated treatment planning workflow using the scripting application programming interface of the Eclipse treatment planning system. Fifteen patients (5 right- and 10 left-sided) previously treated with breast + LLN radiation therapy were replanned using the script. The automated workflow included1 optimal placement of the tangential fields based on the planning target volume and organ-at-risk contours, followed by optimization of field weights and beam energy2; positioning of the RapidArc fields; and3 subsequent RapidArc optimization using the RapidPlan knowledge-based planning solution. Average total planning times were 163 ± 97 and 33 ± 5 minutes for the manual and automated plans, respectively, with approximately 130 and 5 minutes of user interaction. Dosimetrically, both sets of plans were very similar, with comparable planning target volume dose homogeneity values and organ-at-risk mean dose differences of ≤1.9 Gy. In 14/15 patients, the physician judged that the automated plan was either preferred (n = 4) or equal (n = 10) to the manual plan. The complex hybrid RapidArc planning process for patients requiring breast + LLN irradiation was automated by optimizing the tangential field setup and integrating RapidPlan. The quality of the automated and manual plans was comparable, whereas automated planning times were substantially shorter. The principles described here could be used to automate other planning workflows.


Beet-vinasse (condensed molasses solubles) as an ingredient in diets for cattle and pigs - nutritive value and limitations.
Sugar beet vinasse, originating as the condensed molasses residue from yeast production, was fed to bulls and pigs to determine the feed value. It contained, per kg dry matter (DM content 66.8%), 293 g crude protein, 305 g N-free extract (therefrom 34.4 g total sugar), and 395 g crude ash. The experiment was carried out with three fattening bulls (German Holstein) and four adult pigs (Gottingen Minipig). In bulls, two balance trials lasting 20 days each (10 days adaptation; 10 days collection) were conducted in which subsequently a basal ration without and with vinasse (14% in DM) was fed. In pigs, the experiment consisted of three 12-day balance trials with the final 5 days as collection period. In the first trial, pigs received a basal diet without vinasse and in the following, a mixed diet with 16 and 43% vinasse (in DM) respectively. Digestibilities of components in vinasse in bulls were: organic matter 73.5%, crude protein 72.6%, and N-free extract 52.3%. In bulls, no influence on faeces quality could be detected while feeding 14% vinasse in DM. While feeding 16% vinasse in DM to pigs, the digestibility of organic matter amounted to 72.3% (crude protein 71.8%; NfE 74.6%). Feeding higher amounts of vinasse (43% in DM) in the diet of pigs, digestibilities were reduced (organic matter: 61.6%), which might be a result of a reduced retention time of the chyme as a result of an osmotically driven diarrhoea because of unexpectedly high sulphate (136 g/kg DM) content in the vinasse. It is concluded that because of organic matter digestibility >70% for cattle and pigs, the use of vinasse in these species in small proportions is reasonable.


Increasing aging content in social work curriculum: perceptions of key constituents.
This mixed methodology study examines the perceptions of key constituents regarding methods for effectively integrating aging content into the foundation curriculum of the BSW and MSW program at the University of Georgia School of Social Work. Students were asked to complete a survey to determine their perception of geriatric content that existed within the foundation coursework. Following an analysis of the survey results, eight semi-structured focus group discussions were conducted with a purposeful sample of students, faculty, field instructors, social work alumni, older adults from the community, and representatives from aging agencies. The intention of these focus groups was to find out what aging content should be infused within the curriculum. The focus group meetings were held in various locations throughout Northeast Georgia and in one remote location in South Georgia. Participants were interviewed about the necessary skills and knowledge for social workers practicing with an aging population in the areas of: essential intervention skills, program policies and regulations, critical information needed to develop client service plans, strategies for addressing service delivery fragmentation, and community collaboration to support intergenerational family needs. The results of this study will be discussed to provide suggestions on how existing foundation courses can integrate aging content.


Protracted viral shedding and viral load are associated with ICU mortality in Covid-19 patients with acute respiratory failure.
Protracted viral shedding is common in hospitalized patients with COVID-19 pneumonia, and up to 40% display signs of pulmonary fibrosis on computed tomography (CT) after hospital discharge. We hypothesized that COVID-19 patients with acute respiratory failure (ARF) who die in intensive care units (ICU) have a lower viral clearance in the respiratory tract than ICU patients discharged alive, and that protracted viral shedding in respiratory samples is associated with patterns of fibroproliferation on lung CT. We, therefore, conducted a retrospective observational study, in 2 ICU of Lyon university hospital. 129 patients were included in the study, of whom 44 (34%) died in ICU. 432 RT-PCR for SARS-CoV-2 were performed and 137 CT scans were analyzed. Viral load was significantly higher in patients deceased as compared to patients alive at ICU discharge (p < 0.001), after adjustment for the site of viral sampling and RT-PCR technique. The median time to SARS-CoV-2 negativation on RT-PCR was 19 days [CI95 %:15-21] in patients alive at ICU discharge and 26 days [CI95 %:17-infinity] in non-survivors at ICU discharge. Competitive risk regression identified patients who died in ICU and age as independent risk factors for longer time to SARS-CoV-2 negativation on RT-PCR, while antiviral treatment was independently associated with shorter time. None of the CT scores exploring fibroproliferation (i.e., bronchiectasis and reticulation scores) were significantly associated with time to SARS-CoV-2 negativation. Viral load in respiratory samples is significantly lower and viral shedding significantly shorter in ICU survivors of COVID-19 associated acute respiratory failure. Protracted viral shedding is unrelated to occurrence of fibrosis on lung CT.


Inversion concept of the origin of life.
The essence of the inversion concept of the origin of life can be narrowed down to the following theses: 1) thermodynamic inversion is the key transformation of prebiotic microsystems leading to their transition into primary forms of life; 2) this transformation might occur only in the microsystems oscillating around the bifurcation point under far-from-equilibrium conditions. The transformation consists in the inversion of the balance "free energy contribution / entropy contribution", from negative to positive values. At the inversion moment the microsystem radically reorganizes in accordance with the new negentropy (i.e. biological) way of organization. According to this approach, the origin-of-life process on the early Earth took place in the fluctuating hydrothermal medium. The process occurred in two successive stages: a) spontaneous self-assembly of initial three-dimensional prebiotic microsystems composed mainly of hydrocarbons, lipids and simple amino acids, or their precursors, within the temperature interval of 100-300°C (prebiotic stage); b) non-spontaneous synthesis of sugars, ATP and nucleic acids started at the inversion moment under the temperature 70-100°C (biotic stage). Macro- and microfluctuations of thermodynamic and physico-chemical parameters able to sustain this way of chemical conversion have been detected in several contemporary hydrothermal systems. A minimal self-sufficient unit of life on the early Earth was a community of simplest microorganisms (not a separate microorganism).


The MRI findings of iris metastasis in patients with breast cancer.
Breast cancer and lung cancer are the most common tumors that metastasize to iris. The metastasis of iris was generally diagnosed on ophthalmologic examination. In this case, we reported iris metastasis of patients with adenocarcinoma of breast cancer and MRI findings. We report a case of a 51-year-old. She was diagnosed breast cancer two years ago. After adjuvant chemotherapy, radiotherapy and trastuzumab, she was admitted to hospital with the complaints of headache on February 2012. The magnetic resonance imaging (MRI ) revealed multiple brain metastasis. Whole brain radiotherapy and palliative chemotherapy were applied to the patients. In follow-up, on ophthalmological examination, there was a solid lesion on iris. The orbital MRI was performed and it revealed the thickness on iris of left eye. After diagnostic procedure final pathological rewiev reported that invasive ductal carcinom metastasis. Iris metastasis may be considered by MRI findings following: The thickness on iris and contrast enhanced lesion. This reason may be resulted that the fine niddle aspiration biopsy for diagnosis of iris metastasis is not need.


[Histological study of splenic colonies at different times following the transplantation of hematopoietic cells from embryonic liver].
Histological analysis was carried out on the hemopoietic spleen colonies within 7 and 11 days after transplantation of the embryonic liver cells. Large superficial colonies were always present and were, predominantly, erythroid and mixed. Small superficial colonies consisted of undifferentiated cells and, unlike large colonies, appeared on the 11th day only. In the spleen thickness erythroid colonies predominated. The possibility of formation of small superficial 11 day colonies at the expense of pre-CFU-S is discussed.


Can anti-speeding messages based on protection motivation theory influence reported speeding intentions?
The study investigated the effects of anti-speeding messages based on protection motivation theory (PMT) components: severity, vulnerability, rewards, self-efficacy, response efficacy, and response cost, on reported speeding intentions. Eighty-three participants aged 18-25 years holding a current Australian driver's license completed a questionnaire measuring their reported typical and recent speeding behaviors. Comparisons were made between 18 anti-speeding messages used on Australian roads and 18 new anti-speeding messages developed from the PMT model. Participants reported their reactions to the 36 messages on the perceived effectiveness of the message for themselves and for the general population of drivers, and also the likelihood of themselves and other drivers driving within the speed limit after viewing each message. Overall the PMT model-derived anti-speeding messages were better than jurisdiction-use anti-speeding messages in influencing participants' reported intention to drive within the speed limit. Severity and vulnerability were the most effective PMT components for developing anti-speeding messages. Male participants reported significantly lower intention to drive within the speed limit than did female participants. However, males reported significantly higher intention to drive within the speed limit for PMT-derived messages compared with jurisdiction-based messages. Third-person effects were that males reported anti-speeding messages to be more effective for the general driving population than for themselves. Females reported the opposite effect - that all messages would be more effective for themselves than for the general driving population. Findings provided support for using a sound conceptual basis as an effective foundation for anti-speeding message development as well as for evaluating proposed anti-speeding messages on the target driver population.


Genetic variants in p53-related genes confer susceptibility to second primary malignancy in patients with index squamous cell carcinoma of head and neck.
Because of their important roles in mediating the stabilization and expression of p53, we hypothesized that high-risk genotypes of polymorphisms in p53-related genes, including p53, p73, p14(ARF), MDM2 and MDM4, may be associated with an increased risk of second primary malignancy (SPM) after index squamous cell carcinoma of the head and neck (SCCHN). We analyzed data from a cohort of 1283 patients with index SCCHN who were recruited between 1995 and 2007 at MD Anderson Cancer Center and followed for SPM development. Patients were genotyped for nine polymorphisms of p53-related genes. A log-rank test and Cox models were used to compare SPM-free survival and risk. Our results demonstrated that each p53-related polymorphism had a moderate effect on increased SPM risk, but when we combined risk genotypes of these nine polymorphisms together, we found that SPM-free survival was significantly shorter among risk groups with a greater number of combined risk genotypes. SPM risk increased with increasing number of risk genotypes (P < 0.0001 for trend). Compared with the low-risk group (0-3 combined risk genotypes), both the medium-risk (4-5 combined risk genotypes) and high-risk (6-9 combined risk genotypes) groups had significantly increased SPM risk [hazard ratio (HR): 1.6; 95% confidence interval (CI): 1.0-2.6 and HR: 3.0; 95% CI: 1.8-5.0, respectively]. Moreover, such significant associations were even higher in several subgroups. Our findings suggest that combined risk genotypes of p53-related genes may jointly modify SPM risk, especially in patients who are smokers and those with index non-oropharyngeal cancers. However, larger studies are needed to validate our findings.


Bicarbonate transport by amphibian nephron.
To determine the site of bicarbonate reabsorption, tubular fragments from five different segments of the salamander nephron (Ambystoma) were perfused in vitro. Bicarbonate contents (total CO2) of tubular fluid were determined by microcalorimetry. Bicarbonate was not transported by the diluting segment or the mid-distal tubule. Although proximal tubule fragments did appear to reabsorb bicarbonate (10.8 +/- 3.7 pmol X mm-1 X min-1, P less than 0.01, n = 14), the chemical gradients observed were small with respect to the sensitivity of our methods. In the late distal tubule (LDT), bicarbonate reabsorption averaged 28.9 +/- 8.2 pmol X mm-1 X min-1 (P less than 0.01, n = 16). Transport in this segment was inhibited by ethoxzolamide or by perfusing with a sodium-free solution. Ethoxzolamide had no effect on the trans-epithelial voltage. Thus bicarbonate is reabsorbed by the LDT of amphibia via a mechanism that is dependent on carbonic anhydrase and the presence of luminal sodium. The 13 initial collecting tubules (ICT) studied did not appear to reabsorb bicarbonate. On the other hand, some ICT developed substantial bicarbonate gradients. Therefore this nephron segment has the capacity to reabsorb bicarbonate.


Beneficial effects of GH replacement therapy in adults.
Growth hormone deficiency in adults is associated with psychosocial maladjustment, reduced muscle strength, and reduced exercise capacity. Body composition is significantly altered, with increased fat and decreased muscle volume as compared with healthy subjects. Kidney function is subnormal. Epidemiological data suggest premature mortality owing to cardiovascular disease in hypopituitary patients. Short-term GH treatment trials have shown improved psychosocial performance, normalization of body composition, increased muscle strength, improved exercise capacity, increased cardiac performance, and normalization of kidney function. Thus GH replacement therapy in GH-deficient adults exhibits potential long-term beneficial effects. A number of important questions have to be addressed before long-term GH replacement therapy in GH-deficient adults can be considered on a routine basis.


Increasing the meaningful involvement of women in HIV cure-related research: a qualitative interview study in the United States.
Cisgender women represent over half of people living with HIV globally. However, current research efforts toward a cure for HIV focus predominantly on cisgender men. The under-representation of women in HIV cure clinical studies is particularly problematic given data suggesting that sex-dependent phenotypes limit scientific discovery. We aimed to generate considerations to increase the meaningful involvement of women in HIV cure-related research. We conducted in-depth interviews with biomedical researchers and community members to better understand factors that could increase the meaningful involvement of women in HIV cure clinical trials. Participants were affiliated with academia, industry, community advisory boards, and community-based organizations, and were identified using listings from the AIDS Clinical Trials Group and the Martin Delaney Collaboratories. We used conventional content analysis to analyze the qualitative data. We recruited 27 participants, of whom 11 were biomedical researchers and 16 were community members. Participants included 25 cisgender women, 1 transgender woman, and 1 cisgender man. Key considerations emerged, including the need to ensure that HIV cure studies reflect HIV epidemiologic trends and having accurate representation by sex and gender in HIV cure research. To increase the meaningful involvement of women, recommendations included instituting intentional enrollment goals, frequent and mandatory reporting on enrollment, and incentives for sites to enroll women. Additional themes included the need for agency and self-determination, attention to lived experiences, trauma and healing, and adequate support for women (e.g. logistical, psychosocial, mental, emotional, and physical). Participants noted that women would be willing to participate in HIV cure trials, related procedures (e.g. biopsies), and analytical treatment interruptions. They also expressed a desired for women-centered and holistic clinical trial designs that account for intersectionality. Our empirical inquiry extends recent calls to action to increase diversity of people involved in HIV cure research. Redressing the under-inclusion of women in HIV cure research is an urgent imperative. The entire field must mobilize and reform to achieve this goal. Meaningfully involving women across the gender spectrum in HIV cure research is needed to ensure that interventions are safe, effective, scalable, and acceptable for all people with HIV.


Protection of inactivated vaccine against SARS-CoV-2 infections in patients with comorbidities: a prospective cohort study.
Protection against severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2) infection of inactivated vaccines is not well characterized in people with comorbidities, who are at high risk of severe infection. We compared the risk of SARS-CoV-2 infection after complete vaccination with Sinopharm/BBIBP in people with comorbidities (e.g., autoimmune diseases, cardiovascular disease, chronic lung disease, and diabetes) with healthy individuals using a Cox-proportional hazard model. In July-September 2021, a total of 10 548 people (comorbidities, 2143; healthy, 8405) receiving the complete primary series of vaccination with Sinopharm/BBIBP in Bangkok, Thailand were prospectively followed for SARS-CoV-2 infection through text messaging and telephone interviewing for 6 months. A total of 295 infections from 284 participants were found. HRs (95% CI) of individuals with any comorbidities did not increase (unadjusted, 1.02 (0.77-1.36), P = 0.89; adjusted, 1.04 (0.78-1.38), P = 0.81). HRs significantly increased in the subgroup of autoimmune diseases (unadjusted, 2.64 (1.09-6.38), P = 0.032; adjusted, 4.45 (1.83-10.83), P = 0.001) but not in cardiovascular disease, chronic lung disease, or diabetes. The protection against SARS-CoV-2 infection of the Sinopharm vaccine was similar in participants with any comorbidities vs. healthy individuals. However, the protection appeared lower in the subgroup of autoimmune diseases, which may reflect suboptimal immune responses among these people.


[Professional medical education in Russia].
There is a tendency to increase the role of education process in the life of the individual, caused by necessity of new knowledge, experience and skills, which is the effective measure to adapt human being to the current social and economic conditions. The idea of education as a relatively short period of life is gone. It becomes obvious, that use of forms and types of adult education becomes limited and inefficient. The development of the modern education system involves training with a high level of independence and leadership of the individual student; provision by vocational education institutions a wide range of educational services; adequate to the needs of the labor market; variability of methods and forms of education; active use of the modern educational technology as one of the most convenient ways of training.


Elaborated spaced-retrieval and prospective memory in mild Alzheimer's disease.
Prospective memory, or the timely remembering of a planned action, is conceptualised as a cognitive process demanding episodic memory and executive attention. Impairments in these skills are characteristic of the cognitive decline in early Alzheimer's disease, providing an expectation of prominent prospective memory difficulties in this population, and yet surprisingly, memory performance in early Alzheimer's disease has rarely been evaluated within a prospective memory framework. In a preliminary study we demonstrated that older adults with early Alzheimer's disease (n=14), as compared to healthy older adults (n=14), were significantly impaired in a simple experimental paradigm of prospective remembering (a text-reading task). In a subsequent intervention study, we investigated the efficacy of spaced-retrieval for improving the prospective remembering performance of older adults with early Alzheimer's disease (n=16) compared to healthy older adults (n=16) under two learning conditions: a spaced-retrieval technique alone or spaced-retrieval combined with elaborated encoding of task. The majority of the Alzheimer's disease group (63%) demonstrated benefit in prospective remembering in the combined condition as compared to spaced-retrieval alone. Participants with Alzheimer's disease who demonstrated better executive attention (Trail Making- set-shifting) and/or better retrospective memory (Hopkins Verbal Learning Test-Revised- recognition) were more successful in the combined learning condition.


Hypoxia-resistant profile implies vulnerability of cancer stem cells to physiological agents, which suggests new therapeutic targets.
We have previously shown that peculiar metabolic features of cell adaptation and survival in hypoxia imply growth restriction points that are typical of embryonic stem cells and disappear with differentiation. Here we provide evidence that such restrictions can be exploited as specific antiblastic targets by physiological factors such as pyruvate, tetrahydrofolate, and glutamine. These metabolites act as powerful cytotoxic agents on cancer stem cells (CSCs) when supplied at doses that perturb the biochemical network, sustaining the resumption of aerobic growth after the hypoxic dormant state. Experiments were performed in vivo and in vitro using CSCs obtained from various anaplastic tumors: human melanoma, leukemia, and rat hepatoma cells. Pretreatment of melanoma CSCs with pyruvate significantly reduces their self-renewal in vitro and tumorigenicity in vivo. The metabolic network underlying the cytotoxic effect of the physiological factors was thoroughly defined, principally using AH130 hepatoma, a tumor spontaneously reprogrammed to the embryonic stem stage. This network, based on a tight integration of aerobic glycolysis, cellular redox state, and folate metabolism, is centered on the cellular NADP/NADPH ratio that controls the redox pathway of folate utilization in purine synthesis. On the whole, this study indicates that pyruvate, FH 4, and glutamine display anticancer activity, because CSCs are committed to survive and maintain their stemness in hypoxia. When CSC need to differentiate and proliferate, they shift from anaerobic to aerobic status, and the few mitochondria available makes them susceptible to the injury of the above physiological factors. This vulnerability might be exploited for novel therapeutic treatments.


Transmission of Streptococcus dysgalactiae subsp. equisimilis between child and dog in an Aboriginal Australian community.
Streptococcus dysgalactiae subsp. equisimilis (SDSE), also known as group G and C streptococci, is becoming increasingly recognized as a pathogen in humans. We report here the finding of an identical strain of SDSE in the throat of a child and their dog in an Aboriginal Australian community. The strain was identified using the API 20Strep system, 16S rRNA gene sequencing, emm sequence typing (emmST) and multilocus sequence typing (MLST) as a group C SDSE, stC839.5 and ST-3. Carriage of this strain by a human and dog in the same household justifies detailed epidemiological studies using molecular typing to clarify the extent of cross-species transmission and sharing of SDSE and other group G and C streptococci, and its impact in these communities.


Head and neck IMPT probabilistic dose accumulation: Feasibility of a 2 mm setup uncertainty setting.
To establish optimal robust optimization uncertainty settings for clinical head and neck cancer (HNC) patients undergoing 3D image-guided pencil beam scanning (PBS) proton therapy. We analyzed ten consecutive HNC patients treated with 70 and 54.25 GyRBE to the primary and prophylactic clinical target volumes (CTV) respectively using intensity-modulated proton therapy (IMPT). Clinical plans were generated using robust optimization with 5 mm/3% setup/range uncertainties (RayStation v6.1). Additional plans were created for 4, 3, 2 and 1 mm setup and 3% range uncertainty and for 3 mm setup and 3%, 2% and 1% range uncertainty. Systematic and random error distributions were determined for setup and range uncertainties based on our quality assurance program. From these, 25 treatment scenarios were sampled for each plan, each consisting of a systematic setup and range error and daily random setup errors. Fraction doses were calculated on the weekly verification CT closest to the date of treatment as this was considered representative of the daily patient anatomy. Plans with a 2 mm/3% setup/range uncertainty setting adequately covered the primary and prophylactic CTV (V95 ≥ 99% in 98.8% and 90.8% of the treatment scenarios respectively). The average organ-at-risk dose decreased with 1.1 GyRBE/mm setup uncertainty reduction and 0.5 GyRBE/1% range uncertainty reduction. Normal tissue complication probabilities decreased by 2.0%/mm setup uncertainty reduction and by 0.9%/1% range uncertainty reduction. The results of this study indicate that margin reduction below 3 mm/3% is possible but requires a larger cohort to substantiate clinical introduction.


Exploring the Influence of Social Determinants, Social Capital, and Health Expertise on Health and the Rural Church.
In rural communities, religious places can significantly shape health for individuals, families, and communities. Rural churches are prominent community centers in rural communities and are deeply woven into rural culture. Thus, health influences arising from the rural church likely have health implications for the greater community. This article explores health influences emerging from rural churches using social determinants of health, social capital, and health expertise. Although nurses are important health resources for all populations, their value in rural areas may be exceedingly significant. The contribution of nurses to church-based health capital in rural communities may be quite significant and underestimated, although it remains poorly understood.


Measles-mumps-rubella vaccine at 6 months of age, immunology, and childhood morbidity in a high-income setting: study protocol for a randomized controlled trial.
Measles is a highly contagious and serious infection. Before the introduction of vaccination, measles caused yearly epidemics putting vulnerable children at risk of brain damage and death. Despite safe and cost-effective vaccines, measles remains a leading cause of death in children globally. Due to insufficient vaccine coverage and low levels of in utero transferred antibodies from vaccinated mothers, outbreaks of measles in Denmark and other high-income countries are observed at increasing frequency. The current vaccine was introduced in Denmark in 1987 as a one-shot measles-mumps-rubella vaccine at 15 months, a timing chosen to avoid inhibition of the infant's immune response by maternal antibodies. One generation later, the MMR vaccinated mothers have lower antibody levels compared to the naturally infected, and their infants are already susceptible at 6 months of age or earlier, thus increasing the risk of epidemics. The Danish MMR trial is a double-blind randomized clinical trial recruiting between March 2019 and December 2021 with last patient last visit in February 2022. Altogether N = 6500 infants aged 6 months will be randomly assigned to intramuscular vaccination with routine MMR (M-M-R VaxPro) or placebo (solvent only). According to the Danish Childhood vaccination program, all infants will receive a routine MMR vaccination at 15 months of age. At randomization, 1 month later, and 1 month after routine MMR vaccination at 15 months of age, a blood sample is drawn from app. 10% (N = 600) of the population. Additionally, hair, saliva, and urine are sampled at randomization. The co-primary study outcomes are immunogenicity 1 month after MMR vaccination at 6 months of age assessed as plaque-reduction neutralization test, and incidence of infectious disease hospitalizations from randomization to 12 months of age. Six weeks post randomization, all participants are interviewed regarding adverse events. The trial is registered in the EU Clinical Trials Registry. EudraCT registration number: 2016-001901-18 . Registered on 14 February 2017.


Creatinine clearance, walking speed, and muscle atrophy: a cohort study.
Chronic kidney disease is associated with malnutrition and inflammation. These processes may lead to loss of skeletal muscle and reduced physical performance. Associations of kidney function with muscle composition and longitudinal measures of physical performance are unknown. Prospective cohort study. We evaluated 826 community-dwelling older adults enrolled in the Invecchiare in Chianti (InCHIANTI) Study who were free of baseline stroke or activities of daily living disability. Baseline creatinine clearance (Clcr) based on 24-hour urine collection. Cross-sectional and longitudinal trajectories of physical performance measured by 7-m usual gait speed, 400-m fast gait speed, and knee extension strength using isometric dynamometry. Calf muscle composition assessed by quantitative computed tomography. Mean age of participants was 74 ± 7 (SD) years, with 183 having Clcr < 60 mL/min/1.73 m(2). After adjustment, each 10-mL/min/1.73 m(2) decrement in Clcr was associated with 0.01 (95% CI, 0.004-0.017) m/s slower 7-m usual walking speed and 0.008 (95% CI, 0.002-0.014) m/s slower 400-m walking speed. Each 10-mL/min/1.73 m(2) decrement in Clcr was associated with 28 (95% CI, 0.8-55) mm(2) lower muscle area and 0.15 (95% CI, 0.04-0.26) mg/cm(3) lower muscle density. After adjustment, lower Clcr was associated with slower mean 7-m (P=0.005) and 400-m (P=0.02) walk and knee extension strength (P=0.001) during the course of follow-up. During a mean follow-up of 7.1 ± 2.5 years, each 10-mL/min/1.73 m(2) lower baseline Clcr was associated with 0.024 (95% CI, 0.01-0.037) kg/y greater decline in knee strength. Single baseline measurement of Clcr and 3-year interval between follow-up visits may lead to nondifferential misclassification and attenuation of estimates. Among older adults, lower Clcr is associated with muscle atrophy, reduced walking speed, and more rapid declines in lower-extremity strength over time.


[Reproducibility of perimetric study results].
Theoretical and experimental aspects of the reproducibility of visual fields in static automated perimetry are described. Special emphasis was placed on factors which could potentially influence reproducibility, such as cooperation, the patient's age and the size of his/her pupils, learning and fatigue effects, the depth of scotomas and the influence of drugs.


Gene expression profile study in CFTR mutated bronchial cell lines.
Cystic fibrosis (CF) is caused by mutations in the gene encoding the cystic fibrosis conductance transmembrane regulator (CFTR). Symptoms are pancreatic insufficiency, chronic obstructive lung disease, liver disease, chronic sinusitis and infertility in male patients. The phenotypic variability may be explained only in part by the more than 1200 CFTR mutations, which are grouped into six different classes, according to their effect on the protein ranging from a severe (no synthesis or blocked processing) to mild mutation (altered conductance or reduced synthesis). However, it is now accepted that other genes (CF modifiers) influence the phenotypic spectrum of the disease. In order to identify CF modifier genes, we built a low-density home-made oligoarray containing 144 genes selected according to biochemical criteria and evaluated their expression in two CF bronchial epithelial cell lines (CuFi1 F508del/F508del; CuFi3 F508del/R553X). If we consider both cell lines, 38 genes (26.3%) show an altered expression pattern with a threshold > +/-1.5. Of these 38 genes, 12 are altered in CuFi1, and 26 in CuFi3. Some of these genes share the same expression pattern in both cell lines, while others have a different behaviour. These results were validated by a QRT-PCR assay (R2 CuFi1 = 0.81 and R2 CuFi3 = 0.91). These data could suggest that the presence of a class I allele (R553X) determines a more profound alteration of gene expression pattern than the presence of a class II allele (F508del). The identification of the genes altered by a specific CF mutation could lead to the development of a pharmacological approach specific for different CFTR genotypes.


The language of vaccination campaigns during COVID-19.
Understanding what makes communication effective when designing public health messages is of key importance. This applies in particular to vaccination campaigns, which aim to encourage vaccine uptake and respond to vaccine hesitancy and dispel any myth or misinformation. This paper explores the ways in which the governments of Great Britain (England, Scotland and Wales) promoted COVID-19 vaccination as a first-line strategy and studies health message effectiveness by examining the language of official vaccination campaigns, vaccine uptake across the different nations and the health message preferences of unvaccinated and vaccine sceptic individuals. The study considers communications beginning at the first lockdown until the point when daily COVID-19 updates ended for each nation. A corpus linguistic analysis of official government COVID-19 updates is combined with a qualitative examination of the expression of evaluation in governmental discourses, feedback from a Public Involvement Panel and insights from a nationally representative survey of adults in Great Britain to explore message production and reception. Fully vaccinated, unvaccinated and sceptic respondents showed similar health messaging preferences and perceptions of health communication efficacy, but unvaccinated and sceptic participants reported lower levels of compliance for all health messages considered. These results suggest that issues in health communication are not limited to vaccination hesitancy, and that in the future, successful vaccination campaigns need to address the determining factors of public attitudes and beliefs besides communication strategies.


Psychiatric disorder and cognitive impairment in SLE.
Poor performance on cognitive testing is common in SLE but it is not progressive in the majority of patients and may fluctuate or resolve without specific treatment. Cognitive impairment in patients without overt CNS-SLE may result from generalised disease activity or psychiatric disorder which reduce speed, concentration and motivation. This emphasises once again the importance of recognising and treating psychiatric disorder in these patients. Although mean cognitive scores are lower in SLE patients with overt CNS involvement than those without, an individual's cognitive score is a poor predictor of the presence of CNS involvement because of considerable overlap between groups. It has been suggested that the pattern of cognitive impairment, rather than simply whether it is present or absent, may be more helpful in identifying patients with CNS involvement but this requires further investigation in prospective studies. Cognitive impairment at one point in time is not predictive of future CNS events during 1 or 2 years of follow-up. Routine cognitive testing in SLE does not therefore appear to be helpful either for identifying patients with current CNS involvement or for identifying those at future risk of this complication. In the absence of double-blind randomised controlled trials, treatment of neuropsychiatric SLE is based on clinical experience and anecdotal case reports. Aggressive immunosuppression with high-dose corticosteroids in conjunction with either azathioprine or cyclophosphamide may be indicated in patients with life-threatening CNS-SLE but, on the basis of current evidence, is not justified in those with lone subtle cognitive abnormalities.


Preliminary characterization of an antibiotic produced by Xanthomonas albilineans which inhibits DNA synthesis in Escherichia coli.
Chlorosis-inducing isolates of Xanthomonas albilineans, the sugarcane leaf scald pathogen, produced a mixture of antibacterial compounds in culture. The antibiotic mixture, which eluted as a single strongly retarded peak from Sephadex LH-20 in methanol, was bactericidal to Escherichia coli. Inhibition of E. coli was not reversed by added nutrients, and affected cells were not lysed but many accumulated polyphosphate granules. The major antibacterial component, isolated in crystalline form after HPLC, is given the trivial name albicidin. Near the minimum inhibitory concentration, albicidin caused a complete block to DNA synthesis, followed by partial inhibition of RNA and protein synthesis, as assessed by incorporation of radioactive precursors. Spontaneous antibiotic-resistant mutants of E. coli showed no cross-resistance between albicidin and inhibitors of either subunit of DNA gyrase. Mixing albicidin with purified DNA from E. coli did not alter the thermal denaturation behaviour of the DNA, or the absorption spectrum of the antibiotic. PolA+ and PolA - strains of E. coli were equally sensitive to albicidin, indicating that the antibiotic does not bind to or modify DNA. Selective inhibition of DNA synthesis without evidence of DNA binding suggests a specific interaction of albicidin with an essential replication protein.


Neuropsychological impairments on the NEPSY-II among children with FASD.
We examined the pattern of neuropsychological impairments of children with FASD (compared to controls) on NEPSY-II measures of attention and executive functioning, language, memory, visuospatial processing, and social perception. Participants included 32 children with FASD and 30 typically developing control children, ranging in age from 6 to 16 years. Children were tested on the following subtests of the NEPSY-II: Attention and Executive Functioning (animal sorting, auditory attention/response set, and inhibition), Language (comprehension of instructions and speeded naming), Memory (memory for names/delayed memory for names), Visual-Spatial Processing (arrows), and Social Perception (theory of mind). Groups were compared using MANOVA. Children with FASD were impaired relative to controls on the following subtests: animal sorting, response set, inhibition (naming and switching conditions), comprehension of instructions, speeded naming, and memory for names total and delayed, but group differences were not significant on auditory attention, inhibition (inhibition condition), arrows, and theory of mind. Among the FASD group, IQ scores were not correlated with performance on the NEPSY-II subtests, and there were no significant differences between those with and without comorbid ADHD. The NEPSY-II is an effective and useful tool for measuring a variety of neuropsychological impairments among children with FASD. Children with FASD displayed a pattern of results with impairments (relative to controls) on measures of executive functioning (set shifting, concept formation, and inhibition), language, and memory, and relative strengths on measures of basic attention, visual spatial processing, and social perception.


Isolation and purification of chiamydospores of Candida albicans.
Two methods were investigated for their efficiency in isolating and purifying chlamydospores of Candida albicans. Chlamydospores were disconnected from pseudomycelial cells either enzymatically using beta-glucuronidase or mechanically by ultrasonic treatment. Free chlamydospores were separated from other cell material by sucrose gradient centrifugation. The resulting preparations were inspected by light-microscopy and electron-microscopy. Both methods yielded preparations with a level of over 90% chlamydospore cells. Ultrasonic treatment caused little change to the ultrastructure of the chlamydospores, whereas the enzyme treatment profoundly affected the cell wall. It is concluded that ultrasonic treatment is an efficient method for obtaining pure preparations of chlamydospores.


A validated PROM in genetic counselling: the psychometric properties of the Dutch version of the Genetic Counselling Outcome Scale.
Patient empowerment has been identified as a key outcome goal in genetic counselling, and a patient reported outcome measure (PROM) has been developed to measure empowerment in genetic services: the Genetic Counselling Outcome Scale (GCOS). Here we validate the GCOS for a large and diverse Dutch study sample of 2194 patients referred to two clinical genetic centres for counselling about a wide range of conditions (heart disease, neurological disorders, cancer, congenital syndromes, intellectual disability and prenatal pathology). Our results suggest that the GCOS consists of a hierarchical 6-factor structure, with a main scale for empowerment and six subscales: uncertainty about heredity, hope, negative emotions, knowledge about the condition, knowledge about genetic services and uncertainty about the treatment. Six of the original 24 GCOS items were removed due to low factor loadings and small inter-item correlations. Internal consistency and test-retest reliability of the main scale and most subscales were satisfactory. Convergent validity was confirmed by moderate positive and moderate/strong negative associations between the GCOS main scale and other validated outcome measures. Responsiveness was comparable to that of other validated outcome measures. We saw significant improvement in the GCOS main scale and all the subscales after the first genetic counselling session. This study contributes to the international validation process of the GCOS, with the ultimate goal of using this instrument as a PROM, with empowerment as an outcome measure, to evaluate and improve the quality of genetic counselling in various clinical genetics settings.


Fungal Cell Wall Proteins and Signaling Pathways Form a Cytoprotective Network to Combat Stresses.
Candida species are part of the normal flora of humans, but once the immune system of the host is impaired and they escape from commensal niches, they shift from commensal to pathogen causing candidiasis. Candida albicans remains the primary cause of candidiasis, accounting for about 60% of the global candidiasis burden. The cell wall of C. albicans and related fungal pathogens forms the interface with the host, gives fungal cells their shape, and also provides protection against stresses. The cell wall is a dynamic organelle with great adaptive flexibility that allows remodeling, morphogenesis, and changes in its components in response to the environment. It is mainly composed of the inner polysaccharide rich layer (chitin, and β-glucan) and the outer protein coat (mannoproteins). The highly glycosylated protein coat mediates interactions between C. albicans cells and their environment, including reprograming of wall architecture in response to several conditions, such as carbon source, pH, high temperature, and morphogenesis. The mannoproteins are also associated with C. albicans adherence, drug resistance, and virulence. Vitally, the mannoproteins contribute to cell wall construction and especially cell wall remodeling when cells encounter physical and chemical stresses. This review describes the interconnected cell wall integrity (CWI) and stress-activated pathways (e.g., Hog1, Cek1, and Mkc1 mediated pathways) that regulates cell wall remodeling and the expression of some of the mannoproteins in C. albicans and other species. The mannoproteins of the surface coat is of great importance to pathogen survival, growth, and virulence, thus understanding their structure and function as well as regulatory mechanisms can pave the way for better management of candidiasis.


Dynamic Light Scattering Study of a Laser-Induced Phase-Separated Droplet of Aqueous Glycine.
Tightly focusing a continuous-wave, near-infrared laser beam at the air/solution interface of a millimeter-thick layer of glycine in D2O forms a crystal through a polymorphically and spatially controlled nucleation process known as gradient-force laser-induced nucleation or optical-tweezer laser-induced nucleation. However, when this same beam is focused at the glass/solution interface of a film of aqueous glycine, a highly concentrated laser-induced phase-separated (LIPS) solution droplet is formed that does not nucleate while the focusing beam remains on. Two competing theories have emerged about the nature of the LIPS droplet: one proposes that it is a merger of prenucleation metastable nanodroplets and clusters into one large homogeneous "dense liquid droplet", and the other stipulates that it is the result of the partitioning of larger droplets into the new phase, but not a merging of droplets, around the focal point of the beam. In order to determine the nature of the LIPS droplet, dynamic light scattering was used to detect the presence of nanodroplets undergoing Brownian motion within the droplet and to measure their relative size following a range of laser exposure times. The observation of nanodroplets in motion in the center of the LIPS droplet revealed that the application of optical tweezers at the glass/solution interface forms a relatively monodisperse collection of large nanodroplets (>700 nm) concentrated around the focal point of the beam with smaller particles (<100 nm) depleted within the first 2 min of laser exposure. The LIPS droplet quickly reaches a steady state and is not affected by increasing focusing times. These findings allow for a better understanding of the interactions of optical tweezers with aqueous glycine nanodroplets. This understanding will help in studying the fundamental nature of metastable nanodroplets. More practically, laser-induced phase separation makes possible the nucleation-free separation of large nanodroplets from small clusters, facilitating materials technologies such as high purity, polymorphically selective nucleation of crystals and co-crystals used for pharmaceuticals, dyes, and photovoltaics.


A serine alkaline protease from the fungus Conidiobolus coronatus with a distinctly different structure than the serine protease subtilisin Carlsberg.
In view of the functional similarities between subtilisin Carlsberg and the alkaline protease from Conidiobolus coronatus, the biochemical and structural properties of the two enzymes were compared. In spite of their similar biochemical properties, e.g., pH optima, heat stability, molecular mass, pI, esterase activity, and inhibition by diisopropyl fluorophosphate and phenylmethlysulfonylfluoride, the proteases were structurally dissimilar as revealed by (1) their amino acid compositions, (2) their inhibition by subtilisin inhibitor, (3) their immunological response to specific anti-Conidiobolus protease antibody, and (4) their tryptic peptide maps. Our results demonstrate that although they are functionally analogous, the Conidiobolus protease is structurally distinct from subtilisin Carlsberg. The Conidiobolus protease was also different from other bacterial and animal proteases (e.g. pronase, protease K, trypsin, and chymotrypsin) as evidenced by their lack of response to anti-Conidiobolus protease antibody in double diffusion and in neutralization assays. The Conidiobolus serine protease fails to obey the general rule that proteins with similar functions have similar primary sequences and, thus, are evolutionarily related. Our results strengthen the concept of convergent evolution for serine proteases and provide basis for research in evolutionary relationships among fungal, bacterial, and animal proteases.


Expression of the metabotropic glutamate receptor 5 (mGluR5) induces melanoma in transgenic mice.
Glutamate is the major excitatory neurotransmitter in the mammalian CNS and mediates fast synaptic transmission upon activation of glutamate-gated ion channels. In addition, glutamate modulates a variety of other synaptic responses and intracellular signaling by activating metabotropic glutamate receptors (mGluRs), which are G protein-coupled receptors. The mGluRs are also expressed in nonneuronal tissues and are implicated in a variety of normal biological functions as well as diseases. To study mGluR-activated calcium signaling in neurons, we generated mGluR5 transgenic animals using a Thy1 promoter to drive expression in the forebrain, and one founder unexpectedly developed melanoma. To directly investigate the role of mGluR5 in melanoma formation, we generated mGluR5 transgenic lines under a melanocyte-specific promoter, tyrosinase-related protein 1. A majority of the founders showed a severe phenotype with early onset. Hyperpigmentation of the pinnae and tail could be detected as early as 3-5 d after birth for most of the mGluR5 transgene-positive mice. There was 100% penetrance in the progeny from the tyrosinase-related protein 1-mGluR5 lines generated from founders that developed melanoma. Expression of mGluR5 was detected in melanoma samples by RT-PCR, immunoblotting, and immunohistochemistry. We evaluated the expression of several cancer-related proteins in tumor samples and observed a dramatic increase in the phosphorylation of ERK, implicating ERK as a downstream effector of mGluR5 signaling in tumors. Our findings show that mGluR5-mediated glutamatergic signaling can trigger melanoma in vivo. The aggressive growth and severe phenotype make these mouse lines unique and a potentially powerful tool for therapeutic studies.


Effects of feeding high volumes of milk replacer on reproductive performance and on concentrations of metabolites and hormones in blood of Japanese black heifer calves.
We evaluated the effects of feeding high volumes of milk replacer on growth and reproductive performances in Japanese black heifers. Fifty-one heifers were fed milk replacer at 9 L/day for 60 days (9 L × 60 days; n = 18) or 41 days (9 L × 41 days; n = 15), or at 7 L/day for 40 days (7 L × 40 days; n = 18). Artificial insemination (AI) was performed on heifers with ≥270 kg body weight and ≥116 cm body height at 300 days of age. The age at the first AI was 0.35 month later for 7 L × 40 days than the other groups (p < .01). However, age at calving did not differ among treatments (22.1 months). The interval from the first AI to pregnancy tended to be ~2 months longer for the 9 L × 60 days than the other groups (p = .07). Our results showed that feeding high volumes of milk replacer may reduce the age at calving via an improved rate of growth. In addition, we propose that feeding a maximum of 7 L milk replacer for 40 days may be the most appropriate rearing regime because the success of pregnancy per AI may be reduced in calves fed a maximum of 9 L for 41 and 60 days.


The design of a stress-management programme for nursing personnel.
This study identifies the stressors and coping strategies of nursing staff (students, trained staff and those who had left the profession before qualification) in a variety of ward specialisms. The research instruments included an open-ended interview concerning pre-nursing experience, perceived stressors and satisfactions, and ways of coping, and psychometric tests of self-esteem, assertion, ways of coping and personality. The five most frequently cited stressors were understaffing, conflict with nurses, dealing with death and dying, overwork and conflict with doctors. Experience of stressors was related to role and seniority of respondents, with different aspects of the same stressor differentially affecting nurses at different levels of experience. Coping strategies also depended on experience. Trained staff showed more use of problem-focused ways of coping, whilst students and leavers relied more on emotion-focused strategies to deal with stressful situations. These differences were related to personality characteristics of respondents and to self-esteem as well as to situational characteristics of the stressful episode. Social support was important in times of work-related stress, with students in particular making good use of peer group support. Respondents were generally lacking in assertiveness and high in anxiety. Although self-esteem was generally high, leavers scored markedly less than other subject groups in the areas of personal and social self-esteem. Leavers had little prior knowledge or experience of nursing before entering training and knew few nurses or doctors: consequently, nursing failed to meet their expectations. Stress was identified as the major cause of attrition and the sources of stress are identified. This study informed a major programme of stress-management training for student nurses which began in 1988 at the North Wales School of Nursing and which is currently under evaluation. It includes relaxation therapy, assertiveness training, and on-going group discussions which foster peer-group support and which explore the stressors and coping strategies relevant to different stages of training and ward specialisms.


Sexuality, HIV risk and potential acceptability of involving adolescent girls in microbicide research in Kisumu, Kenya.
Current microbicide clinical trials primarily enroll adult participants; however, females under the age of 18, because of their high rates of HIV acquisition, represent an important population for future microbicide clinical research. We sought to understand the individual, family and community-level factors that may influence the acceptability of microbicide use and research involving adolescent girls. We conducted 30 interviews with adolescent girls aged 14-17 and nine focus group discussions with adolescent girls, parents and community leaders in Kisumu, Kenya. Participants discussed adolescent sexuality, HIV prevention methods, perceptions about microbicide use and views about microbicide research involving adolescent girls. Adolescent sexual activity is stigmatised yet acknowledged to be a natural part of the 'adolescent stage.' Desperation to stop the spread of HIV among youth and support for female-initiated HIV prevention methods led to enthusiasm about microbicides and future microbicide research. Yet concerns about microbicides were numerous and included: difficulty using it in a timely manner due to the rushed, unplanned nature of adolescent sex; a fear of trying experimental products; concerns about microbicide efficacy; and parental worry that supporting microbicide use in youth would defy societal pressures that denounce adolescent sexual activity. Microbicide acceptability for youth in sub-Saharan Africa may be bolstered by desperation for new methods to stop the spread of HIV, yet hindered by misgivings about experimental HIV prevention methods for youth. Understanding and addressing the microbicide's perceived benefits and shortcomings, as well as the broader context of adolescent sexuality and HIV prevention, may facilitate future research and promotion of microbicides in this high-risk group.