Boysenberry polyphenol inhibits endothelial dysfunction and improves vascular health.
Endothelial cells have an important role in maintaining vascular homeostasis. Age-related disorders (including obesity, diabetes, and hypertension) or aging per se induce endothelial dysfunction that predisposes to the development of atherosclerosis. Polyphenols have been reported to suppress age-related endothelial cell disorders, but their role in vascular function is yet to be determined. We investigated the influence of boysenberry polyphenol on vascular health under metabolic stress in a murine model of dietary obesity. We found that administration of boysenberry polyphenol suppressed production of reactive oxygen species (ROS) and increased production of nitric oxide (NO) in the aorta. It has been reported that p53 induces cellular senescence and has a crucial role in age-related disorders, including heart failure and diabetes. Administration of boysenberry polyphenol significantly reduced the endothelial p53 level in the aorta and ameliorated endothelial cell dysfunction in iliac arteries under metabolic stress. Boysenberry polyphenol also reduced ROS and p53 levels in cultured human umbilical vein endothelial cells (HUVECs), while increasing NO production. Uncoupled endothelial nitric oxide synthase (eNOS monomer) is known to promote ROS production. We found that boysenberry polyphenol reduced eNOS monomer levels both in vivo and in vitro, along with an increase of eNOS dimerization. To investigate the components of boysenberry polyphenol mediating these favorable biological effects, we extracted the anthocyanin fractions. We found that anthocyanins contributed to suppression of ROS and p53, in association with increased NO production and eNOS dimerization. In an ex vivo study, anthocyanins promoted relaxation of iliac arteries from mice with dietary obesity. These findings indicate that boysenberry polyphenol and anthocyanins, a major component of this polyphenol, inhibit endothelial dysfunction and contribute to maintenance of vascular homeostasis.


Molecular Characterization of a Chrysovirus Isolated From the Citrus Pathogen Penicillium crustosum and Related Fungicide Resistance Analysis.
Penicillium sp. are damaging to a range of foods and fruits including citrus. To date, double-stranded (ds)RNA viruses have been reported in most Penicillium species but not in citrus pathogen P. crustosum. Here we report a novel dsRNA virus, designated as Penicillium crustosum chrysovirus 1 (PcCV1) and isolated from P. crustosum strain HS-CQ15. PcCV1 genome comprises four dsRNA segments, referred to as dsRNA1, dsRNA2, dsRNA3, and dsRNA4, which are 3600, 3177, 3078, and 2808 bp in length, respectively. Sequence analysis revealed the presence of four open reading frames (ORFs) in the PcCV1 genome. ORF1 in dsRNA1 encodes a putative RNA-dependent RNA polymerase (RdRp) and ORF2 in dsRNA2 encodes a putative coat protein (CP). The two remaining ORFs, ORF3 in dsRNA3 and ORF4 in dsRNA4, encode proteins of unknown function. Phylogenetic analysis based on RdRp sequences showed that PcCV1 clusters with other members of the genus Chrysovirus, family Chrysoviridae. Transmission electron microscope (TEM) analysis revealed that the PcCV1 visions are approximately 40 nm in diameter. Regarding biological effects of PcCV1, HS-CQ15 harboring the chrysovirus exhibited no obvious difference in colony morphology under fungicide-free conditions but decreased resistance to demethylation inhibitor (DMI)-fungicide prochloraz, as compared to PcCV1-cured strain. Here we provide the first evidence of a virus present in citrus pathogenic fungus P. crustosum and the chrysovirus-induced change in fungicide-resistance of its host fungus.


Telangiectases in scleroderma: a potential clinical marker of pulmonary arterial hypertension.
Clinical markers are needed to identify scleroderma patients at risk for pulmonary arterial hypertension (PAH) since early therapy may improve survival. We investigated whether increased numbers of telangiectases in scleroderma associate with measures of pulmonary vascular disease. One hundred forty-seven consecutive adult patients with scleroderma were enrolled in this cross-sectional study and scored for the presence of matted telangiectases on 11 body areas. Per body area, telangiectases were scored as 0 if none were present, 1 if there were fewer than 10 telangiectases, and 2 if 10 or more telangiectases were counted. Linear regression analysis was performed to assess the association between right ventricular systolic pressure (RVSP) and telangiectasia score, adjusted for age, race, smoking status, scleroderma subtype, disease duration, and autoantibody status. Logistic regression analysis was performed with PAH by right-heart catheterization (RHC) as the dependent variable. The mean telangiectasia score was 6.0 (SD 4.5, range 0-20). RVSP and telangiectasia score were positively correlated (r = 0.271, p = 0.001). The mean RVSP increased by 10.9 mm Hg for every 10-point increase in telangiectasia score (95% CI 3.6-18.3 mm Hg, p = 0.004), adjusted for potential confounders. The adjusted relative odds of PAH by RHC were 12.4 for patients with a 10-point increase in telangiectasia score (95% CI 1.78-85.9, p = 0.01). Increased numbers of telangiectases strongly associate with the presence of pulmonary vascular disease. Telangiectases may be a clinical marker of more widespread aberrant microvascular disease in scleroderma.


Repetitive behaviors in Tourette's syndrome and OCD with and without tics: what are the differences?
Gilles de la Tourette Syndrome (GTS) and obsessive-compulsive disorder (OCD) share obsessive-compulsive phenomena. The aims of this study were to compare the OC symptom distribution between GTS and OCD and to investigate whether a subdivision of these phenomena into obsessions, compulsions and 'impulsions' is useful in distinguishing GTS and OCD patients. Thirty-two GTS, 31 OCD (10 with tics, 21 without tics) and 29 control subjects were studied using the Leiden repetitive behaviors semi-structured interview to assess GTS as well as OCD-related behaviors. Each reported repetitive thought or action was evaluated on the presence of anxiety and on goal-directedness. This information was used to define whether the behavior was an obsession, compulsion, or 'impulsion'. Both the GTS and OCD study groups showed higher scores than control subjects on rating scales measuring depression, OC behavior and anxiety. In GTS, Y-BOCS severity scores and trait anxiety were lower than in the OCD groups. Furthermore, GTS patients differed from OCD patients in the distribution of symptoms. Aggressive repetitive thoughts, contamination worries and washing behaviors were reported more frequently by tic-free OCD, while mental play, echophenomena, touching and (self)-injurious behaviors were reported more frequently by GTS. OCD individuals with tics were intermediate, but closer to tic-free OCD. GTS individuals reported significantly more 'impulsions' and fewer obsessions and compulsions than OCD individuals with and without tics. Factor analysis revealed three factors accounting for 44% of the variance, resulting in an 'impulsive' factor related to GTS, a 'compulsive' factor related to OCD and an 'obsessive' factor related to tic-free OCD. In conclusion, OCD individuals reported more anxiety and goal-directedness associated with their behaviors than did GTS subjects. The distinction between obsessions, compulsions and impulsions is of importance in identifying Tourette-related vs. non-Tourette-related repetitions.


Effect of different antihypertensive treatments on Ras, MAPK and Akt activation in hypertension and diabetes.
Ras GTPases function as transducers of extracellular signals regulating many cell functions, and they appear to be involved in the development of hypertension. In the present study, we have investigated whether antihypertensive treatment with ARBs (angiotensin II receptor blockers), ACEi (angiotensin-converting enzyme inhibitors) and diuretics induce changes in Ras activation and in some of its effectors [ERK (extracellular-signal-regulated kinase) and Akt] in lymphocytes from patients with hypertension without or with diabetes. ACEi treatment transiently reduced Ras activation in the first month of treatment, but diuretics induced a sustained increase in Ras activation throughout the 3 months of the study. In patients with hypertension and diabetes, ARB, ACEi and diuretic treatment increased Ras activation only during the first week. ACEi treatment increased phospho-ERK expression during the first week and also in the last 2 months of the study; however, diuretic treatment reduced phospho-ERK expression during the last 2 months of the study. In patients with hypertension and diabetes, antihypertensive treatments did not induce changes in phospho-ERK expression in lymphocytes. ACEi treatment reduced phospho-Akt expression in patients with hypertension and diabetes only in the first month of treatment. In conclusion, these findings show that antihypertensive treatments with ACEi, and diuretics to a lesser extent, modify Ras activation and some of its signalling pathways, although in different directions, whereas ARBs do not appear to have any influence on Ras signalling pathways.


Factors affecting prognosis in male anorexics.
Twenty-seven male anorexics have been followed up over 2-20 yr (mean 8 yr) and their outcome assessed. This has been compared with outcome in female patients with similar pictures at presentation (Burns and Crisp, 1984). Features of the family and personal history and clinical aspects of the disease at presentation were compared with outcome and a number found to be significantly associated with it. Poor relationship with parents during childhood and the absence of normal adolescent sexual behavior and fantasy premorbidly were strongly predictive of a poor outcome. Long duration of illness, previous treatment and greater weight loss during illness were also associated with poor outcome, although no specific dietary behaviour was predictive. The remarkable similarity in outcome pattern between the male and female prompts re-evaluation of some theories of the nature of the role of sexual conflicts in anorexia nervosa.


His166 is critical for active-site proton transfer and phototaxis signaling by sensory rhodopsin I.
Photoinduced deprotonation of the retinylidene Schiff base in the sensory rhodopsin I transducer (SRI-Htrl) complex results in formation of the phototaxis signaling state S373. Here we report identification of a residue, His166, critical to this process, as well as to reprotonation of the Schiff base during the recovery phase of the SRI photocycle. Each of the residue substitutions A, D, G, L, S, V, or Y at position 166 reduces the flash yield of S373, to values ranging from 2% of wild type for H166Y to 23% for H166V. The yield of S373 is restored to wild-type levels in Htrl-free H166L by alkaline deprotonation of Asp76, a Schiff base proton acceptor normally not ionized in the SRI-Htrl complex, showing that proton transfer from the Schiff base in H166L occurs when an acceptor is made available. The flash yield and rate of decay of S373 of the mutants are pH dependent, even when complexed with Htrl, which confers pH insensitivity to wild-type SRI, suggesting that partial disruption of the complex has occurred. The rates of S373 reprotonation at neutral pH are also prolonged in all H166X mutants, with half-times from 5 s to 160 s (wild type, 1 s). All mutations of His166 tested disrupt phototaxis signaling. No response (H166D, H166L), dramatically reduced responses (H166V), or inverted responses to orange light (H166A, H166G, H166S, and H166Y) or to both orange and near-UV light (H166Y) are observed. Our conclusions are that His166 1) plays a role in the pathways of proton transfer both to and from the Schiff base in the SRI-Htrl complex, either as a structurally important residue or possibly as a participant in proton transfers; 2) is involved in the modulation of SRI photoreaction kinetics by Htrl; and 3) is important in phototaxis signaling. Consistent with the involvement of the His imidazole moiety, the addition of 10 mM imidazole to membrane suspensions containing H166A receptors accelerates S373 decay 10-fold at neutral pH, and a negligible effect is seen on wild-type SRI.


Interactions of Surface-Functionalized Starch Nanoparticles with Pepsin and Trypsin in Simulated Gastrointestinal Fluids.
Nanoparticles (NPs) can form a protein corona (PC) with proteins in biological fluids. We examined whether starch nanoparticles (SNPs) form a PC and interact with digestive enzymes in simulated gastric and intestinal fluids. We investigated the adsorption of pepsin and trypsin on unmodified, carboxyl-, and amino-modified SNPs (SNPs, COOH-SNPs, and NH2-SNPs, respectively). Quartz crystal microbalance data showed that a tight and irreversible pepsin corona formed on the NH2-SNPs, pepsin had little or no binding to the SNPs and COOH-SNPs, and trypsin had weak binding to all three kinds of NPs. Dynamic light scattering data showed that pepsin significantly increased the size of the NH2-SNPs from 120 ± 2.6 to 203 ± 12.2 nm and decreased their surface potential from 23.2 ± 1.0 to 12.7 ± 0.2 mV. NH2-SNPs could induce the fluorescence quenching of pepsin and change its secondary structures without affecting its activity.


Fe-induced crystalline-amorphous interface engineering of a NiMo-based heterostructure for enhanced water oxidation.
Engineering heterostructures with a unique surface/interface structure is one of the effective strategies to develop highly active noble-metal-free catalysts for the oxygen evolution reaction (OER), because the surface/interface of catalysts is the main site for the OER. Herein, we design a coralloid NiMo(Fe)-20 catalyst with a crystalline-amorphous interface through combining a hydrothermal method and an Fe-induced surface reconfiguration strategy. That is, after Fe3+ impregnation treatment, the Ni(OH)2-NiMoO4 pre-catalyst with a complete crystalline surface is restructured into a trimetallic heterostructure with a crystalline-amorphous interface, which facilitates mass diffusion and charge transfer during the OER. As expected, self-supported NiMo(Fe)-20 exhibits excellent electrocatalytic water oxidation performance (overpotential: η-10 = 220 mV, η-100 = 239 mV) in the alkaline electrolyte, and its electrocatalytic performance hardly changes after maintaining the current density of 50 mA cm-2 for 10 hours. Furthermore, nickel foam (NF) supported commercial Pt/C and self-supported NiMo(Fe)-20 served as the cathode and anode of the Pt/C‖NiMo(Fe)-20 electrolyzer, respectively, which exhibits a lower cell voltage (E-100 = 1.53 V) than that of the Pt/C‖RuO2 electrolyzer (E-100 = 1.58 V) assembled with noble metal-based catalysts. The enhanced electrocatalytic performance of the NiMo(Fe)-20 catalyst is mainly attributed to the synergistic effect between the crystalline-amorphous interface and the coralloid trimetallic heterostructure.


Diabetes and overexpression of proNGF cause retinal neurodegeneration via activation of RhoA pathway.
Our previous studies showed positive correlation between accumulation of proNGF, activation of RhoA and neuronal death in diabetic models. Here, we examined the neuroprotective effects of selective inhibition of RhoA kinase in the diabetic rat retina and in a model that stably overexpressed the cleavage-resistance proNGF plasmid in the retina. Male Sprague-Dawley rats were rendered diabetic using streptozotocin or stably express cleavage-resistant proNGF plasmid. The neuroprotective effects of the intravitreal injection of RhoA kinase inhibitor Y27632 were examined in vivo. Effects of proNGF were examined in freshly isolated primary retinal ganglion cell (RGC) cultures and RGC-5 cell line. Retinal neurodegeneration was assessed by counting TUNEL-positive and Brn-3a positive retinal ganglion cells. Expression of proNGF, p75(NTR), cleaved-PARP, caspase-3 and p38MAPK/JNK were examined by Western-blot. Activation of RhoA was assessed by pull-down assay and G-LISA. Diabetes and overexpression of proNGF resulted in retinal neurodegeneration as indicated by 9- and 6-fold increase in TUNEL-positive cells, respectively. In vitro, proNGF induced 5-fold cell death in RGC-5 cell line, and it induced >10-fold cell death in primary RGC cultures. These effects were associated with significant upregulation of p75(NTR) and activation of RhoA. While proNGF induced TNF-α expression in vivo, it selectively activated RhoA in primary RGC cultures and RGC-5 cell line. Inhibiting RhoA kinase with Y27632 significantly reduced diabetes- and proNGF-induced activation of proapoptotic p38MAPK/JNK, expression of cleaved-PARP and caspase-3 and prevented retinal neurodegeneration in vivo and in vitro. Taken together, these results provide compelling evidence for a causal role of proNGF in diabetes-induced retinal neurodegeneration through enhancing p75(NTR) expression and direct activation of RhoA and p38MAPK/JNK apoptotic pathways.


Increase of Autonomic Nerve Factors in Epicardial Ganglionated Plexi During Rapid Atrial Pacing Induced Acute Atrial Fibrillation.
BACKGROUND The cardiac autonomic nervous system plays an essential role in epicardial ganglionated plexi (GP) regulation of atrial fibrillation onset and progression. To date, the activity of GP and the function of the cardiac autonomic nervous system are not well understood. The aim of this study was to determine alterations in epicardial GP cholinergic nerve, adrenergic nerve, and nerve growth factor expression using rapid atrial pacing to induce atrial fibrillation in canines. MATERIAL AND METHODS Nine healthy adult beagles were divided into two groups: the pacing experimental group (n=6) and the sham-operation control group (n=3). For the pacing group, high frequency pacing of the left atrial appendage was performed for eight hours. In the control group, electrodes were implanted without rapid atrial pacing. Immunocytochemistry was used to identify neurons positively expressing tyrosine hydroxylase, choline acetyl transferase, nerve growth factor and neurturin. RESULTS After successfully establishing a rapid atrial pacing of the left atrial appendage induced atrial fibrillation model, we found that expression of choline acetyl transferase, tyrosine hydroxylase, nerve growth factor, and neurturin was significantly higher in the rapid atrial pacing group than the control group (p<0.05). CONCLUSIONS In our model, incremental excitability of both the adrenergic and cholinergic nerves led to frequent incidents of atrial fibrillation, which were possibly due to an imbalance of autonomic nerve factors in the epicardial GP during acute atrial fibrillation.


Mammary analogue secretory carcinoma of salivary glands: a clinicopathologic and molecular study including 2 cases harboring ETV6-X fusion.
Mammary analogue secretory carcinoma (MASC) is a recently described low-grade carcinoma with morphologic and genetic similarity, including ETV6-NTRK3 fusion, to secretory carcinoma of the breast. ETV6 is frequently involved in other epithelial and nonepithelial tumors, and many fusion partners of ETV6 have been reported. In the present study, 14 Japanese MASC cases were clinicopathologically and molecularly analyzed. The median age of the patients was 39 years, and the male:female ratio was 6:8. All cases showed histopathologic findings compatible with those previously described for MASC and harbored an ETV6 split as visualized by fluorescence in situ hybridization. Two cases showed thick fibrous septa and invasive features including vascular or perineural tumor involvement, findings that are rare in MASC. In addition, in these 2 cases, non-NTRK3 genes appeared to fuse with ETV6 (ETV6-X fusion). NTRK1 and NTRK2, both members of the NTRK family, were not involved. Of the 14 MASC cases, the ETV6-NTRK3 fusion transcript was positive in 6 cases, and the relative expression level of the ETV6-NTRK3 fusion transcript was variable, ranging from 1 to 5.8. Results of the present study of MASC suggest that (1) ETV6 occasionally fuses with unknown non-NTRK3 genes, (2) ETV6-X cases might have an invasive histology, (3) for molecular diagnosis of MASC, fluorescence in situ hybridization to detect ETV6 splits is the method of choice, and (4) the expression level of the ETV6-NTRK3 fusion transcript is considerably variable. These findings provide a novel insight into the oncogenesis, histopathology, diagnosis, treatment, and prognosis of this newly recognized carcinoma.


Fabricating pH-stable and swellable very thin hyperbranched poly(ethylene imine)-oligosaccharide films fabricated without precoating: first view on protein adsorption.
For using successful (ultra)thin dendritic macromolecule films in (bio)sensing and microfluidic devices and for obtaining reproducible film properties, alteration effects arising from precoatings have to be avoided. Here, oligosaccharide-modified hyperbranched poly(ethylene imine)s (PEI-OS) were used to fabricate very thin PEI-OS films (15-20 nm in dry state), cross-linked with citric acid under condensation, and vacuum condition. However, no reactive precoating is necessary to obtain stable films, which allows very simple film preparation and avoids alteration of the PEIS-OS film properties arising from precoating. Several methods [(in situ) ellipsometry, AFM, XPS, (in situ) ATR-IR, streaming potential measurements] were applied to characterize homogeneity, surface morphology, and stability of these PEI-OS films between pH 2 and pH 10, but also the low protein adsorption behavior.


Ice-age persistence and genetic isolation of the disjunct distribution of larch in Alaska.
Larix laricina (eastern larch, tamarack) is a transcontinental North American conifer with a prominent disjunction in the Yukon isolating the Alaskan distribution from the rest of its range. We investigate whether in situ persistence during the last glacial maximum (LGM) or long-distance postglacial migration from south of the ice sheets resulted in the modern-day Alaskan distribution. We analyzed variation in three chloroplast DNA regions of 840 trees from a total of 69 populations (24 new sampling sites situated on both sides of the Yukon range disjunction pooled with 45 populations from a published source) and conducted ensemble species distribution modeling (SDM) throughout Canada and United States to hindcast the potential range of L. laricina during the LGM. We uncovered the genetic signature of a long-term isolation of larch populations in Alaska, identifying three endemic chlorotypes and low levels of genetic diversity. Range-wide analysis across North America revealed the presence of a distinct Alaskan lineage. Postglacial gene flow across the Yukon divide was unidirectional, from Alaska toward previously glaciated Canadian regions, and with no evidence of immigration into Alaska. Hindcast SDM indicates one of the broadest areas of past climate suitability for L. laricina existed in central Alaska, suggesting possible in situ persistence of larch in Alaska during the LGM. Our results provide the first unambiguous evidence for the long-term isolation of L. laricina in Alaska that extends beyond the last glacial period and into the present interglacial period. The lack of gene flow into Alaska along with the overall probability of larch occurrence in Alaska being currently lower than during the LGM suggests that modern-day Alaskan larch populations are isolated climate relicts of broader glacial distributions, and so are particularly vulnerable to current warming trends.


Characterizing Dysarthria Diversity for Automatic Speech Recognition: A Tutorial from the Clinical Perspective.
Despite significant advancements in automatic speech recognition (ASR) technology, even the best performing ASR systems are inadequate for speakers with impaired speech. This inadequacy may be, in part, due to the challenges associated with acquiring a sufficiently diverse training sample of disordered speech. Speakers with dysarthria, which refers to a group of divergent speech disorders secondary to neurologic injury, exhibit highly variable speech patterns both within and across individuals. This diversity is currently poorly characterized and, consequently, difficult to adequately represent in disordered speech ASR corpora. In this paper, we consider the variable expressions of dysarthria within the context of established clinical taxonomies (e.g., Darley, Aronson, and Brown dysarthria subtypes). We also briefly consider past and recent efforts to capture this diversity quantitatively using speech analytics. Understanding dysarthria diversity from the clinical perspective and how this diversity may impact ASR performance could aid in (1) optimizing data collection strategies for minimizing bias; (2) ensuring representative ASR training sets; and (3) improving generalization of ASR across users and performance for difficult-to-recognize speakers. Our overarching goal is to facilitate the development of robust ASR systems for dysarthric speech using clinical knowledge.


The "de-escalation concept" and antibiotic de-escalation: a missed opportunity?
"De-escalation therapy" is a term that suggests the need to reduce the spectrum or the number of antibiotics formerly prescribed for critical patients, upon clinical improvement and/or microorganism recovery. The major goal of this concept is the use of broad-spectrum antibiotic agents as initial drugs of choice for severe patients, instead of "reserving" the most potent agents after an inadequate clinical response, or after the microorganism is recovered. Despite possible commercial concerns and an unproven but possible relationship with enhancing global antibiotic use, the concept was correct and in accordance with scientific evidence. However, the "de-escalation" component of the concept is very seldom reported, and no large clinical trial on this issue is available until today. To definitely put in practice this concept, comparative large trials must be designed and sponsored to insert this strategy at the same level of evidence of wide initial empiric antibiotic treatments.


The role of CYP2C19 in the metabolism of (+/-) bufuralol, the prototypic substrate of CYP2D6.
Upon characterization of baculovirus-expressed cytochrome P-450 (CYP) 2C19, it was observed that this enzyme metabolized (+/-) bufuralol to 1'hydroxybufuralol, a reaction previously understood to be selectively catalyzed by CYP2D6. The apparent K(m) for this reaction was 36 microM with recombinant CYP2C19, approximately 7-fold higher than for recombinant CYP2D6. The intrinsic clearance for this reaction was 37-fold higher with CYP2D6 than for CYP2C19. The involvement of human CYP1A2 in bufuralol 1'-hydroxylation was also confirmed using the recombinant enzyme. Using S-mephenytoin as an inhibitor, the K(i) for inhibition of recombinant CYP2C19-mediated bufuralol hydroxylation was 42 microM, which is the approximate K(m) for recombinant CYP2C19-mediated S-mephenytoin metabolism. The classic CYP2D6 inhibitors quinidine and quinine showed no inhibition of CYP2C19-catalyzed bufuralol metabolism at concentrations that abolished CYP2D6-mediated bufuralol metabolism. Ticlopidine, a potent inhibitor of CYP2C19 and CYP2D6, inhibited bufuralol 1'-hydroxylation by each of these enzymes equipotently. In human liver microsomes that are known to be deficient in CYP2D6 activity, it was shown that in the presence of quinidine, the K(m) shifted from 14 to 38 microM. This is consistent with the K(m) determination for recombinant CYP2C19 of 36 microM. In human liver microsomes that have high CYP2D6 and CYP2C19 activity, the K(m) shifted to 145 microM in the presence of S-mephenytoin and quinidine, consistent with the K(m) determined for CYP1A2. This data suggests that bufuralol, and possibly other CYP2D6 substrates, have the potential to be metabolized by CYP2C19.


Physiological properties, time of development, and central projection are correlated in the wing mechanoreceptors of Drosophila.
The wing of Drosophila contains 8 sensory structures (campaniform sensilla), which lie in specific locations and possess identical surface morphology. The axons of the campaniform neurons follow either a medial or a lateral tract within the CNS. Previous studies (Palka et al., 1986) indicate that choice of central pathway correlates with the time of birth and differentiation of the neurons rather than with their topographic distribution on the wing. On the basis of the response properties revealed by mechanical and electrical stimulation, these sensory cells also fall into 2 physiological categories, rapidly and slowly adapting, that correlate exactly with central projection and birthdate. Thus, within this discrete population of sensory neurons there exists a precise 3-way correlation between physiology, central projection, and time of development.


The isolation of human platelet factor V.
Human platelet factor V has been isolated using either a monoclonal or polyclonal antibody directed against human plasma factor V. The largest peptide observed upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of purified human platelet factor V comigrates with purified human plasma factor V. However, a significant portion of the isolated protein is represented by peptides of lower apparent molecular weight (Mr). These lower Mr species that copurify with platelet factor V have been shown to be platelet factor V components by their immunological cross-reactivity with monoclonal and polyclonal antibodies to purified human plasma factor V. Platelets isolated from whole blood drawn directly into inhibitors to prevent proteolysis and platelet activation demonstrate the pattern of fragmented platelet factor V. The components of purified platelet factor V demonstrate apparent Mr ranging between 115 K and 330 K and are detectably different from the intermediates and end products observed during the thrombin cleavage of single-chain plasma factor V. Upon treatment with thrombin the platelet factor V components are cleaved and the end products are indistinguishable from those obtained upon thrombin activation of plasma factor V to plasma factor Va. Examination of the components by immunoblotting demonstrates that some of the cleavages which have occurred in the platelet factor V molecule are within the 150-K activation peptide. Bioassay indicates that platelet factor V exists as a procofactor and cleavage by thrombin yields the active cofactor, platelet factor Va. These data suggest that human platelet factor V is stored in the platelet as a partially fragmented procofactor that can be activated by thrombin to yield human platelet factor Va, the active cofactor in the human prothrombinase complex.


Acidic glycosaminoglycans and gangliosides in the brains from four patients with genetic mucopolysaccharidosis.
Acidic glycosaminoglycans (GAGs) and gangliosides were analyzed in the brains from control fetuses, fetal Hurler syndrome, control children and three patients affected by genetic mucopolysaccharidosis (Hurler, Hunter and Morquio syndromes). GAGs contents in the brains from patients with Hurler and Hunter syndromes were approximately 1.5-3.0-fold greater as compared with those controls, and most of the GAGs were much more degraded than those from controls. Dermatan sulfate (DS), which was not identified in the control brains, comprised about 20--40% of the total GAGs. On the other hand, GAGs content and molecular weight distribution in the brain from the patient affected by Morquio syndrome were similar to those in the control brains. GAGs content in the brain from fetal Hurler syndrome was also 2.0-fold greater and DS, which was not detected in the control fetal brains, comprised 1.6% of the total GAGs. However, the molecular weight distribution of the GAGs was similar to those of the control fetal brains. The total amount of the brain gangliosides in all patients assayed here was similar to those in the control brains. However, a greater amount of GM1- and Gm2-gangliosides was observed in the brains from patients with Hurler and Hunter syndromes.


A pilot feasibility study of daily rTMS to modify corticospinal excitability during lower limb immobilization.
Short term immobilization of the lower limb is associated with increased corticospinal excitability at 24 hours post cast removal. We wondered whether daily stimulation of the motor cortex might decrease brain reorganization during casting. We tested the feasibility of this approach. Using transcranial magnetic stimulation (TMS), resting motor threshold and recruitment curves were obtained at baseline in 6 healthy participants who then had leg casts placed for 10 days. On 7 of the 10 days subjects received 20 minutes of 1 Hz repetitive TMS (rTMS). TMS measures were then recorded immediately after and 24 hours post cast removal. Four of 6 subjects completed the study. At the group level there were no changes in excitability following cast removal. At the individual level, two participants did not show any change, 1 participant had higher and one lower excitability 24 hours after cast removal. Daily rTMS over motor cortex is feasible during casting and may modify neuroplastic changes occurring during limb disuse. A prospective double blind study is warranted to test whether daily rTMS might improve outcome in subjects undergoing casting, and perhaps in other forms of limb disuse such as those following brain injury or weightlessness in space flight.


Left-sided congenital diaphragmatic hernia: can we improve survival while decreasing ECMO?
Mortality and ECMO rates for congenital diaphragmatic hernia (CDH) remain ~30%. In 2016, we changed our CDH guidelines to minimize stimulation while relying on preductal oxygen saturation, lower mean airway pressures, stricter criteria for nitric oxide (iNO), and inotrope use. We compared rates of ECMO, survival, and survival without ECMO between the two epochs. Retrospective review of left-sided CDH neonates at the University of Utah/Primary Children's Hospital NICUs during pre (2003-2015, n = 163) and post (2016-2019, n = 53) epochs was conducted. Regression analysis controlled for defect size and intra-thoracic liver. Following guideline changes, we identified a decrease in ECMO (37 to 13%; p = 0.001) and an increase in survival without ECMO (53 to 79%, p = 0.0001). Overall survival increased from 74 to 89% (p = 0.035). CDH management guideline changes focusing on minimizing stimulation, using preductal saturation and less aggressive ventilator/inotrope support were associated with decreased ECMO use and improved survival.


Identification of the VIL2 enhancer in human embryonic kidney cells.
We previously demonstrated that the VIL2 -87/+134 region exhibited promoter activity in some human cells, and a region further upstream of this promoter might contain an enhancer. However, the properties and location of this VIL2 enhancer remain unclear. In this study, we cloned the VIL2 -1541/-706 segment and investigated its transcriptional regulatory properties via luciferase assays in transiently transfected HEK-293 cells (human embryonic kidney cells). The VIL2 -1541/-706 was found to exhibit promoter activity. Furthermore, when this segment was located upstream of the VIL2 or SV40 (simian virus 40) promoters in the forward orientation, the expression levels of luciferase were dramatically enhanced. However, this transcriptional enhancement disappeared when this segment was located upstream of the promoter in the reverse orientation or downstream of the reporter gene in the forward or reverse orientation. In deletion experiments, we found several potential regulatory regions within the VIL2 -1541/-706. When these regions were separately located upstream of the VIL2 or SV40 promoters, only the -1297/-1186 considerably enhanced the activity of these promoters. Although the other regulatory regions exhibited significant transcriptional regulation in deletion experiments, they weakly enhanced VIL2 promoter activity and/or did not regulate SV40 promoter activity. These results suggest that the DNA sequence upstream of the VIL2 promoter functions as an enhancer in a position- and orientation-dependent manner, and the VIL2 -1297/-1186, which acts as a key enhancer, probably regulates VIL2 transcription in combination with other potential regulatory regions located upstream of the VIL2 promoter.


[Jan Roelof Prakken, editor-in-chief of the Dutch Journal of Medicine 1955-1970].
Jan Roelof Prakken (1897-1982) worked as a family physician before specializing in dermatology. In 1946, he was appointed professor of dermatology and venereal diseases at the University of Amsterdam. In 1955 he became editor-in-chief of the Nederlands Tijdschrift voor Geneeskunde (Dutch Journal of Medicine). He proved to be a progressive editor. Contrary to his predecessors and despite opposition from many readers, Prakken was of the opinion that sociomedical topics such as oral contraception, artificial insemination, abortion, and even sex change operations were of importance to physicians and had to be discussed in the Journal.


The influence of plate size on meal composition. Literature review and experiment.
Environmental factors, such as the size of containers, can influence our energy intake. Even though different sized food containers are often recommended to control portion sizes, the evidence to support this is contradictory. In the present study, we conducted a literature review and a controlled laboratory experiment to investigate whether plate size influences the composition of a meal and the total meal energy. The results of the review suggest that distraction factors, the type of container, the food-serving mode (self-service or being served) and the type of food offered all influenced the results observed in the various published studies. For the experiment in this study, eighty-three participants were individually invited to serve themselves a lunch from a buffet containing 55 replica food items. Either a standard size plate (27 cm) or a large plate (32 cm) was provided to the participants. The results of the experiment suggest that the plate size had no significant effect on the total energy of the meal (F(1,81) =0.782, P> .05). However, participants using a large plate served themselves significantly more vegetables (F(1,81) =4.786, P< .05), particularly vegetables generally eaten as side dishes (F(1,81) =6.804, P< .05). Therefore, reducing the plate size does not seem to be an appropriate intervention to reduce the total energy intake in order to promote weight loss. Rather, using a large plate might be a simple and inexpensive strategy to increase vegetable consumption.


Ultrafastly Interweaving Graphdiyne Nanochain on Arbitrary Substrates and Its Performance as a Supercapacitor Electrode.
A moderate method is first developed here for superfast (in seconds) growth of an ultrafine graphdiyne (GDY) nanochain on arbitrary substrates in the atmosphere. This is an environmentally friendly and metal-catalyst-free method, efficiently eliminating extraneous contaminations for the carbon materials. The seamless GDY coating on any substrates demonstrates that an all-carbon GDY possesses outstanding controllability and processability, perfectly compensating for the drawbacks of prevailing all-carbon materials. After the decoration of 3D GDY nanostructures, the substrates become superhydrophobic with contact angles high up to of 148° and can be used as outstanding frameworks for storing organic pollution. Because of the reasonable porous and 3D continuous features, the as-prepared samples can be applied as high-performance binder-free supercapacitor electrodes with high area capacitance of up to 53.66 mF cm -2, prominent power performance, and robust long-term retention (99% after 1300 cycles).


Study of cytokine response against panel of purified Mycobacterium leprae antigens by using whole blood assay in subjects residing in a resettlement village of cured leprosy patients.
Mycobacterium leprae being an intracellular pathogen, cell mediated immunity is very important in the clinical outcome of leprosy. Manifestation of the disease is correlated with the level and type of cell mediated immune response. The main objective of this study was to analyse TNF-alpha and IFN-gamma production by T-cells when challenged with different M. leprae purified antigens in subjects with known exposure. 50 subjects residing in resettlement village of cured leprosy patients were included in the study. Whole Blood assay studies were undertaken in which the blood was placed in culture and was challenged with 35-kDa antigen, whole M. leprae cells, M. leprae cell wall antigen and M. leprae soluble antigen minus LAM. T-cell derived cytokines TNF-alpha and IFN-gamma were measured by ELISA. It was observed that challenging the lymphocytes with 35-kDa antigen, the cell wall antigen and M. leprae soluble antigen minus LAM resulted in increased levels of IFN-gamma whereas challenge with 35-kDa antigen and M. leprae cell wall antigen resulted in increased levels of TNF-alpha.


Silence is Golden: Auditory Preferences in Zoo-housed Gorillas.
Enrichment is presented to improve the welfare of captive animals but sound is frequently presented with the assumption that it is enriching without assessing individuals' preferences. Typically, presented sounds are unnatural and animals are unable to choose which sounds they can listen to or escape them. We examined preferences of three zoo-housed gorillas for six categories of sound. The gorillas selected unique icons on a computer touchscreen that initiated brief samples of silence, white noise, nature, animal, percussion, and electronic instrumental sounds. Following training, gorillas selected each sound paired with silence (Phase 2), each sound paired with each other sound (Phase 3), and one sound among all other sound categories (Phase 4). Initially, a single sound was associated with each icon, but additional exemplars of the category were added in phases 5-8. Preferences were generally stable and one gorilla showed a strong preference for silence. Although there were individual differences, a surprising general preference for unnatural over natural sounds was revealed. These results indicate the importance of assessing preferences for individuals before introducing auditory stimulation in captive habitats.


Showing their true colors: a practical approach to volume rendering from serial sections.
In comparison to more modern imaging methods, conventional light microscopy still offers a range of substantial advantages with regard to contrast options, accessible specimen size, and resolution. Currently, tomographic image data in particular is most commonly visualized in three dimensions using volume rendering. To date, this method has only very rarely been applied to image stacks taken from serial sections, whereas surface rendering is still the most prevalent method for presenting such data sets three-dimensionally. The aim of this study was to develop standard protocols for volume rendering of image stacks of serial sections, while retaining the benefits of light microscopy such as resolution and color information. Here we provide a set of protocols for acquiring high-resolution 3D images of diverse microscopic samples through volume rendering based on serial light microscopical sections using the 3D reconstruction software Amira (Visage Imaging Inc.). We overcome several technical obstacles and show that these renderings are comparable in quality and resolution to 3D visualizations using other methods. This practical approach for visualizing 3D micro-morphology in full color takes advantage of both the sub-micron resolution of light microscopy and the specificity of histological stains, by combining conventional histological sectioning techniques, digital image acquisition, three-dimensional image filtering, and 3D image manipulation and visualization technologies. We show that this method can yield "true"-colored high-resolution 3D views of tissues as well as cellular and sub-cellular structures and thus represents a powerful tool for morphological, developmental, and comparative investigations. We conclude that the presented approach fills an important gap in the field of micro-anatomical 3D imaging and visualization methods by combining histological resolution and differentiation of details with 3D rendering of whole tissue samples. We demonstrate the method on selected invertebrate and vertebrate specimens, and propose that reinvestigation of historical serial section material may be regarded as a special benefit.


Lifetime body mass index and later atherosclerosis risk in young adults: examining causal links using Mendelian randomization in the Cardiovascular Risk in Young Finns study.
Mendelian randomization uses genetic variants related to environmentally modifiable risk factors in an attempt to improve causal inference from observational data. We examined the effect of lifetime body mass index (BMI) on adult carotid intima-media thickness (CIMT) and various atherosclerotic risk factors by using both Mendelian randomization and conventional analyses. A total of 2230 individuals (1218 women), aged 3-18 at study induction, took part in clinical examinations in 1980, 1983, 1986, and, most recently, 2001 when they were aged 24-39. In these analyses we utilized the known relation between FTO polymorphism rs9939609 and BMI. The dose-response association between the number of A alleles in FTO and higher mean BMI from childhood to adulthood was confirmed, but no associations with potential confounding factors were observed. In standard regression models, lifetime BMI was associated with adult CIMT, lifetime systolic blood pressure, adult fasting glucose, and adult HOMA-index. When variation in FTO was used as an instrument for unconfounded BMI levels, similar or larger effects of lifetime BMI on all these phenotypes were found, although with wider confidence intervals. Mutually supportive results from Mendelian randomization and standard regression models strengthen the evidence of the effect of lifetime BMI on atherosclerosis risk in young adults.


[Effect of forskolin eyedrops on intraocular pressure in healthy males].
Suspensions of Forskolin in concentrations of 0.3; 0.6; and 1.0% decreased effectively the intraocular pressure of healthy subjects when instillated in the conjunctival sac. The suspensions were compared with placebo in double blind studies. The maximum effect was reached 3 hours after application of the 0.3% (22.8%) and 0.6% (27.8%) suspensions and 4 hours after the 1.0% (26.5%) suspension. The higher concentrations decreased the intraocular pressure to the same extent as the lowest concentration but the effect lasted longer: 4 hours after instillation of the 0.3% suspension, 5 hours after instillation of the 0.6% suspension and 7 hours after the 1.0% suspension. The suspensions were well tolerated. Subjective sensations like burning, itching and augmentated lacrimation were observed only in a minor amount and for a short period of time.


[The therapeutic efficacy of interferon-alfa-2a (Roferon-A) in chronic hepatitis C].
Evaluation of 6-month treatment with roferon-A of patients with chronic hepatitis C (CHC) and comparison of the treatment regimens. 79 CHC patients received roferon-A for 3 months in a dose 6,000,000 IU 3 time a week. In case of the response to treatment was continued for the next 3 months (3,000,000 IU 3 times a week). Clinical-laboratory findings, results, of EIA and liver biopsy histology were examined. Primary remission was achieved in 71.8% of cases. 36.9% of patients developed recurrences including 14.1% recurrence arising in the course of therapy. Stable remission was obtained in 32.4% of patients. 28.2% patients were non-responders. Side effects were mild, discontinuation of the treatment was necessary only in 7% of cases. Roferon-A administration in a dose 6,000,000 IU for 24 weeks is optimal both as related to cost and efficacy of the treatment.


Human papillomavirus and salivary gland neoplasia: a p16INK4 immunohistochemical and in situ hybridisation study.
This study aimed to evaluate the association between human papillomavirus infection and salivary gland tumours in a Scottish cohort. Specimens from a range of salivary gland tumours operated on between 1997 and 2012 were studied. A tissue microarray constructed from tissue blocks was subjected to p16INK4 (cyclin-dependent kinase inhibitor 2A) immunohistochemistry and in situ hybridisation using probes specific for human papillomavirus, including types 16 and 18. A total of 61 tumours (benign and malignant) were deemed suitable for the study. p16INK4 staining yielded three (4.9 per cent) positive samples: one small cell carcinoma, one squamous cell carcinoma and one poorly differentiated carcinoma. Human papillomavirus in situ hybridisation demonstrated a positive signal in the latter sample only (1.6 per cent). This study demonstrated a very low human papillomavirus detection rate in salivary gland tumours. It can therefore be concluded that human papillomavirus infection is unlikely to play a role in salivary gland neoplasia. Rare human papillomavirus positive cases should be carefully evaluated to exclude the possibility of a metastatic lesion.


Radiation therapy in Peyronie's disease.
Peyronie's disease (PD) is characterized by fibrotic plaque formation within the tunica albuginea, leading to penile deformity, angulation, painful erections, and often, erectile dysfunction. There is limited knowledge of its etiology and pathophysiology. Currently, there is no standardization for reporting the evaluation and treatment outcomes. No consensus exists on the ideal management of PD. The aim of this study is to review the use of radiation therapy (RT) for the treatment of PD. This study used a review of the literature and informed expert opinion. The main outcome measures are the reported improvement with regard to pain, curvature, plaque size, and erectile function as well as a review of the safety profile of RT. Radiation has been used for the empirical treatment of PD with mixed results throughout the literature. Low-dose RT in the early stages of PD seems to be effective in patients with painful erections not improving with time or with the use of oral or intralesional therapies. Comparison with other current medical therapies is difficult due to the absence of criteria for therapeutic success and the absence of randomized, controlled trials. Multiple studies have demonstrated varied results in terms of curvature reduction, plaque volume, and sexual functioning. The absence of the use of validated means of measuring penile deformity and sexual function, the limited relevance of plaque and pain improvement combined with the potential negative consequences of radiation on erectile tissue structure and function lead us to make a recommendation that RT not be used for the management of PD. In clinical practice, mixed success for RT has led to the recommendation against its use as a treatment modality for PD.


CSFV induced mitochondrial fission and mitophagy to inhibit apoptosis.
Classical swine fever virus (CSFV), which causes typical clinical characteristics in piglets, including hemorrhagic syndrome and immunosuppression, is linked to hepatitis C and dengue virus. Oxidative stress and a reduced mitochondrial transmembrane potential are disturbed in CSFV-infected cells. The balance of mitochondrial dynamics is essential for cellular homeostasis. In this study, we offer the first evidence that CSFV induces mitochondrial fission and mitophagy to inhibit host cell apoptosis for persistent infection. The formation of mitophagosomes and decline in mitochondrial mass relevant to mitophagy were detected in CSFV-infected cells. CSFV infection increased the expression and mitochondrial translocation of Pink and Parkin. Upon activation of the PINK1 and Parkin pathways, Mitofusin 2 (MFN2), a mitochondrial fusion mediator, was ubiquitinated and degraded in CSFV-infected cells. Mitophagosomes and mitophagolysosomes induced by CSFV were, respectively, observed by the colocalization of LC3-associated mitochondria with Parkin or lysosomes. In addition, a sensitive dual fluorescence reporter (mito-mRFP-EGFP) was utilized to analyze the delivery of mitophagosomes to lysosomes. Mitochondrial fission caused by CSFV infection was further determined by mitochondrial fragmentation and Drp1 translocation into mitochondria using a confocal microscope. The preservation of mitochondrial proteins, upregulated apoptotic signals and decline of viral replication resulting from the silencing of Drp1 and Parkin in CSFV-infected cells suggested that CSFV induced mitochondrial fission and mitophagy to enhance cell survival and viral persistence. Our data for mitochondrial fission and selective mitophagy in CSFV-infected cells reveal a unique view of the pathogenesis of CSFV infection and provide new avenues for the development of antiviral strategies.


Retinoic acid receptors beta and gamma distinguish retinoid signals for growth inhibition and neuritogenesis in human neuroblastoma cells.
Retinoids induce marked growth inhibition and neuritic differentiation in human neuroblastoma cells. Expression patterns of nuclear retinoic acid receptors (RAR) in embryonic and adult tissues suggests that RAR subtypes alpha, beta and gamma have tissue-specific functions. We have transfected a human neuroblastoma tumor cell line with a vector expressing either human RAR alpha, beta or gamma cDNAs. In the absence of exogenous retinoid, RAR beta transfectants demonstrated marked growth inhibition without morphologic evidence of differentiation, whereas transfectant clones overexpressing RARs alpha and gamma had no significant reduction in cell growth rates. Although RAR gamma transfectants were sensitive to the growth inhibitory effects of exogenous retinoids, these cells demonstrated resistance to the neuritogenic retinoid effects. Only RAR beta transfectants exhibited increased sensitivity to retinoids added in vitro. These results suggest that distinct neuritogenic and growth inhibitory signalling pathways exist in neuroblastoma cells and that RAR beta expression may be necessary for the retinoid growth inhibitory pathway.


Stable carbon isotope fractionation of chloroethenes by dehalorespiring isolates.
Stable carbon isotope fractionation during the reductive dechlorination of chloroethenes by two bacterial strains that dechlorinate to ethene, Dehalococcoides ethenogenes 195 and Dehalococcoides sp. strain BAV1 as well as Sulfurospirillum multivorans and Dehalobacter restrictus strain PER-K23, isolates that do not dechlorinate past DCE, are reported. Fractionation by a Dehalococcoides-containing enrichment culture is also measured for comparison to the isolates. All data adequately fit the Rayleigh model and results are presented as enrichment factors. For strain 195, the measured enrichment factors were -9.6 +/- 0.4, -21.1 +/- 1.8, and -5.8 +/- 0.5 when degrading TCE, cDCE, and 1,1-DCE, respectively. Strain BAV1 exhibited enrichment factors of -16.9 +/- 1.4, -8.4 +/- 0.3, -21.4 +/- 0.9, and -24.0 +/- 2.0 for cDCE, 1,1-DCE, tDCE, and VC, respectively. The surprisingly large differences in enrichment factors caused by individual reductases (RDases) reducing different chloroethenes is likely the result of chemical structure differences among the chloroethenes. For TCE reduction, S. multivorans and D. restrictus strain PER-K23 exhibited enrichment factors of -16.4 +/- 1.5 and -3.3 +/- 0.3, respectively. While all of the organisms studied here utilize RDases that require corrinoid cofactors, the biotic TCE enrichment factors varied widely from those reported for the abiotic cobalamin-catalyzed reaction, indicating that additional factors affect the extent of fractionation in these biological systems. The enrichment factors measured for the Dehalococcoides-containing enrichment culture did not match well with those from any of the isolates, demonstrating the inherent difficulties in predicting fractionation factors of undefined communities. Although compound-specific isotope fractionation is a powerful tool for evaluating the progress of in situ bioremediation in the field, given the wide range of enrichment factors associated with functionally similar and phylogenetically diverse organisms, caution must be exercised when applying enrichment factors for the interpretation of dechlorination data.


Impact of the clinical frailty scale on clinical outcomes and bleeding events in patients with ST-segment elevation myocardial infarction.
The Clinical Frailty Scale (CFS) is a simple tool to assess patients' frailty and may help to predict adverse outcomes in elderly patients. The aim of the present study was to examine the impact of CFS on clinical outcomes and bleeding events after successful percutaneous coronary intervention (PCI) in ST-segment elevation myocardial infarction (STEMI). We enrolled 266 consecutive patients with STEMI who underwent primary PCI in between January 2015 and June 2018. Patients were categorized into two groups based on the CFS stages: CFS 1-3 and CFS ≥ 4. We collected the data and evaluated the relationship between the CFS grade and the incidence of major adverse cardiovascular events (MACE) and Bleeding Academic Research Consortium 3 or 5 bleeding events. Of these patients, CFS ≥ 4 was present in 59 (22.2%). During the follow-up, 37.3% in the CFS ≥ 4 group and 8.2% in the CFS 1-3 group experienced MACE. In Kaplan-Meier analysis, the proportion of MACE-free survival for 4 years was significantly lower in the CFS ≥ 4 group (log-rank P < 0.001). Additionally, the proportion of bleeding event-free survival was significantly lower in the CFS ≥ 4 group (log-rank P < 0.001). The CFS (per 1-grade increase) remained an independent significant predictor of MACE on multivariate Cox proportional hazard analysis [hazard ratio 1.39 (95% confidence interval: 1.08 to 1.79, P = 0.01)]. In conclusion, CFS was an independent predictor of future adverse cardiac events in patients with STEMI. Therefore, the assessment of CFS is crucial in this population.


Multiple sclerosis in gulf war era veterans. 2. Military deployment and risk of multiple sclerosis in the first gulf war.
Concern has been raised that US veterans of the 1990-1991 Gulf War (GW1) may be at increased risk to develop neurologic disease. An incident cohort of multiple sclerosis (MS) and other demyelinating disease (ODD) was assembled from the US military comprising the Gulf War era (1990-2007). Cases of MS and ODD meeting standard diagnostic criteria were matched to a database of all active duty personnel from the Department of Defense. Relative risk (RR) estimates for MS and all demyelinating disease based on onset, deployment status, and exposures were calculated. For GW1, a total of 1,841 incident cases of definite MS and ODD were identified, with 387 among 696,118 deployed and 1,454 among 1,786,215 nondeployed personnel. The RR for MS alone among those deployed compared to those nondeployed was 0.69 (confidence interval, CI: 0.61-0.78), with 0.72 (CI: 0.62-0.83) in men and 0.96 (CI: 0.75-1.22) in women. Deployment was also nonsignificant or protective as an MS risk factor across racial groups, all age groups, and each military service. RRs for MS by service were: Air Force 0.71 (CI: 0.53-0.96), Army 0.80 (CI: 0.67-0.96), Marines 0.96 (CI: 0.63-1.47), and Navy 0.56 (CI: 0.43-0.74). Military deployment to GW1 was not a risk factor for developing MS.


Towards an internet-scale overlay network for latency-aware decentralized workflows at the edge.
Small-scale data centers at the edge are becoming prominent in offering various services to the end-users following the cloud model while avoiding the high latency inherent to the classic cloud environments when accessed from remote Internet regions. However, we should address several challenges to facilitate the end-users finding and consuming the relevant services from the edge at the Internet scale. First, the scale and diversity of the edge hinder seamless access. Second, a framework where researchers openly share their services and data in a secured manner among themselves and with external consumers over the Internet does not exist. Third, the lack of a unified interface and trust across the service providers hinder their interchangeability in composing workflows by chaining the services. Thus, creating a workflow from the services deployed on the various edge nodes is presently impractical. This paper designs Viseu, a latency-aware blockchain framework to provide Virtual Internet Services at the Edge. Viseu aims to solve the puzzle of network service discovery at the edge, considering the peers' reputation and latency when choosing the service instances. Viseu enables peers to share their computational resources, services, and data among each other in an untrusted environment, rather than relying on a set of trusted service providers. By composing workflows from the peers' services, rather than confining them to the pre-established service provider and consumer roles, Viseu aims to facilitate scientific collaboration across the peers natively. Furthermore, by offering services from multiple peers close to the end-users, Viseu also minimizes end-to-end latency and data loss in the service execution at the Internet scale.


Positive or U-Shaped Association of Elevated Hemoglobin Concentration Levels with Metabolic Syndrome and Metabolic Components: Findings from Taiwan Biobank and UK Biobank.
Iron overnutrition has been implicated with a higher risk of developing metabolic and cardiovascular diseases, including metabolic syndrome (MetS), whereas iron deficiency anemia exacerbates many underlying chronic conditions. Hemoglobin (Hb) concentration in the blood, which reflects a major functional iron (i.e., heme iron) in the body, may serve as a surrogate of the nutritional status of iron. We conducted sex-specific observational association studies in which we carefully titrated the association between Hb deciles and MetS and its components among the Taiwanese Han Chinese (HC) from the Taiwan Biobank and Europeans of White ancestry from the UK Biobank, representing two large ethnicities. Our data show that at higher-than-normal levels of Hb, increasing deciles of Hb concentration were significantly associated with MetS across all sex subgroups in both ethnicities, with the highest deciles resulting in up to three times greater risk than the reference group [Taiwanese HC: OR = 3.17 (95% CI, 2.75-3.67) for Hb ≥ 16.5 g/dL in men, OR = 3.11 (2.78-3.47) for Hb ≥ 14.5 g/dL in women; European Whites: OR = 1.89 (1.80-1.98) for Hb ≥ 16.24 g/dL in men, OR = 2.35 (2.24-2.47) for Hb ≥ 14.68 g/dL in women]. The association between stronger risks and increasing Hb deciles was similarly observed with all metabolic components except diabetes. Here we found that both the highest Hb decile groups and contrarily the lowest ones, with respect to the reference, were associated with higher odds of diabetes in both ethnic groups [e.g., Taiwanese HC men: OR = 1.64 (1.33-2.02) for Hb ≥ 16.5 g/dL, OR = 1.71 (1.39-2.10) for Hb ≤ 13.5 g/dL; European Whites women: OR = 1.39 (1.26-1.45) for Hb ≥ 14.68 g/dL, OR = 1.81 (1.63-2.01) for Hb ≤ 12.39 g/dL]. These findings confirm that elevated Hb concentrations, a potential indicator of iron overnutrition, may play a role in the pathophysiology of MetS and metabolic components.


Artificial chromosome transgenesis in pigmentary research.
Pigmentary genes were among the first mammalian genes to be studied, mostly because of the obvious phenotypes associated with their mutations. In 1990, tyrosinase, encoding the limiting enzyme in the melanin synthesis pathway, was eventually assigned to the c (albino) locus by classical rescue experiments driven by functional constructs in transgenic mice. These pioneer reports triggered the study of the regulation of endogenous tyrosinase gene expression by combining different amounts of upstream regulatory and promoter regions and testing their function in vivo in transgenic animals. However, faithful and reproducible transgenic expression was not achieved until the entire tyrosinase expression domain was transferred to the germ-line of mice using artificial-chromosome-type transgenes. The use of these large tyrosinase transgenic constructs and the ease with which they could be manipulated in vitro enabled the discovery of previously unknown but fundamental regulatory regions, such as the tyrosinase locus control region (LCR), whose presence was required in order to guarantee position-independent and copy-number-dependent expression of tyrosinase transgenes, with an expression level, per copy, comparable to that of an endogenous wild-type allele. Subsequently, functional dissection of elements present within this LCR through the generation of new artificial-chromosome type tyrosinase transgenes has revealed the existence of different regulatory activities. The existence of some of these units had been suggested previously by standard-type transgenic analyses. In this review, we will discuss both independent approaches and conclude that optimal tyrosinase transgene expression requires the use of its complete expression domain.


To eat or not to eat: Reward delay impulsivity in children with loss of control eating, attention deficit / hyperactivity disorder, a double diagnosis, and healthy children.
Reward delay impulsivity is a feature of attention deficit/hyperactivity disorder (ADHD) and a likely feature of loss of control eating (LOC-E), which might explain the higher risk of children with ADHD or LOC-E to become obese. The goal of this study was to investigate reward delay impulsivity in children with LOC-E, ADHD, or a double diagnosis, in contrast to healthy children. Children (8 to 13 years) with LOC-E (n = 24), ADHD (n = 33), a double diagnosis (n = 9), and healthy children (n = 34) performed a computer game (door opening task [DOT]) and the delay of gratification task (DoGT) to assess food related facets of reward delay impulsivity. In addition, children reported whether they worried to lose control over eating during the DoGT. There were no group differences in the DOT. However, children with ADHD or a double diagnosis had a significantly higher risk to eat prematurely during the DoGT than children with LOC-E, who were not significantly different from healthy children. Children with a double diagnosis were most likely to worry about losing control over eating during the DoGT, followed by children with LOC-E, and both had a significantly higher probability to worry than healthy children. For children with a double diagnosis the probability to worry was significantly higher than for children with ADHD. If replicated, these findings point to a special relevance of reward delay impulsivity in children with ADHD or a double diagnosis, compared to children with LOC-E. ADHD should be regularly assessed in children with LOC-E.


Role of the M-loop and reactive center loop domains in the folding and bridging of nucleosome arrays by MENT.
MENT is a developmentally regulated heterochromatin-associated protein that condenses chromatin in terminally differentiated avian blood cells. Its homology to the serpin protein family suggests that the conserved serpin reactive center loop (RCL) and the unique M-loop are important for its function. To examine the role of these domains, we studied the interaction of wild-type and mutant MENT with naked DNA and biochemically defined nucleosome arrays reconstituted from 12-mer repeats containing nucleosome positioning sequences. Wild-type MENT folded the naked DNA duplexes into closely juxtaposed parallel structures ("tramlines"). Deletion of the M-loop, but not inactivation of the RCL, prevented tramline formation and the cooperative interaction of MENT with DNA. Reconstitution of wild-type MENT with nucleosome arrays caused their tight folding and self-association. M-loop deletion inhibited nucleosome array folding, whereas the inactive RCL mutant was competent to fold the nucleosome arrays, but had a significantly impaired ability to cause their self-association. Bifunctional chemical cross-linking of MENT revealed oligomerization of wild-type MENT in the presence of chromatin and DNA. This oligomerization was severely reduced in the RCL mutant. We propose that the mechanism of MENT-induced heterochromatin formation involves two independent events: bringing together nucleosome linkers within a chromatin fiber and formation of protein bridges between chromatin fibers. Ordered binding of MENT to linker DNA via its unique M-loop domain promotes the folding of chromatin, whereas bridging of chromatin fibers is facilitated by MENT oligomerization mediated by the RCL.


Azaphenalene phthalocyanines: phthalocyanine analogues with six-membered-ring units instead of five-membered-ring units.
Mixed-condensation reaction of 1,8-naphthalenedicarbonitrile and a 4,5-disubstituted phthalonitrile provided a series of phthalocyanine (Pc) analogues with azaphenalene (AP) moieties in place of the isoindole moieties. Monosubstituted species, APPc, and the two structural isomers of disubstituted species, adj-AP(2)Pc and opp-AP(2)Pc, were successfully isolated by gel-permeation chromatography on HPLC apparatus. Their structures were elucidated by (1)H NMR spectroscopy and X-ray crystallographic analysis. Replacement of the isoindole moieties with azaphenalene moieties created six-membered-ring units in the core and caused distortion of the molecular structures. The Q-band absorption shifted to the red upon an increase in the number of azaphenalene units; the shape of the absorption spectra depended on the molecular symmetries. APPc and opp-AP(2)Pc showed a large splitting of the Q band, whereas adj-AP(2)Pc exhibited a single broad Q band. These changes in the absorption spectra, as well as the unique electronic structures, are discussed in detail, based on magnetic circular dichroism spectra, electrochemical measurements, and density functional theory calculations.


Activation of unusual Mac-1+CD4-CD8- T cells bearing (alpha)(beta) antigen receptor in murine lungs during infection with Mycobacterium bovis BCG: regulation by interferon-gamma.
We have recently (Kawakami et al, Immunol. Lett. 1995;46: 143) demonstrated that unusual Mac-1+CD4-CD8- T cells bearing (alpha)(beta) antigen receptor (Mac-1+ (alpha)(beta) T cells) reside in a considerable proportion in murine lungs. The present study was performed to examine the dynamics of accumulation of these cells in the lungs following intravenous administration of Mycobacterium bovis BCG (BCG). Mac-1+ (alpha)(beta) T cells accumulated rapidly 24 hr after infection, followed by a gradual increase over the observation period of 15 days. Furthermore, the expression of Ia, ICAM-1 and FcgammaR II/III on their surface intensified dramatically after BCG infection. The kinetics of enhancement of Ia expression was slower than that of ICAM-1, with the maximum level attained in one day in the latter molecule but in two weeks in the former. Neutralization of endogenous IFN-gamma by specific mAb completely blocked the augmented expression of Ia on Mac-1+ (alpha)(beta) T cells after BCG infection, but did not have any significant effect on that of ICAM-1. In contrast, in vivo administration of IFN-gamma enhanced the expression of ICAM-1 as well as that of Ia. Our results indicate that accumulation of Mac-1+ (alpha)(beta) T cells within the lung is associated with a differential change in the expression of surface antigens, and suggest that these cells may play a role in the host defense against mycobacterial infection.


Chlamydia psittaci (psittacosis) as a cause of community-acquired pneumonia: a systematic review and meta-analysis.
Psittacosis is a zoonotic infectious disease caused by the transmission of the bacterium Chlamydia psittaci from birds to humans. Infections in humans mainly present as community-acquired pneumonia (CAP). However, most cases of CAP are treated without diagnostic testing, and the importance of C. psittaci infection as a cause of CAP is therefore unclear. In this meta-analysis of published CAP-aetiological studies, we estimate the proportion of CAP caused by C. psittaci infection. The databases MEDLINE and Embase were systematically searched for relevant studies published from 1986 onwards. Only studies that consisted of 100 patients or more were included. In total, 57 studies were selected for the meta-analysis. C. psittaci was the causative pathogen in 1·03% (95% CI 0·79-1·30) of all CAP cases from the included studies combined, with a range between studies from 0 to 6·7%. For burden of disease estimates, it is a reasonable assumption that 1% of incident cases of CAP are caused by psittacosis.


Does experience in creating a robot-assisted partial nephrectomy (RAPN) programme in an academic centre impact outcomes or complication rate?
To evaluate the impact of increased experience in robot-assisted partial nephrectomy (RAPN) on perioperative and oncological outcomes To detail the complications encountered in establishing a RAPN programme at a tertiary referral centre and the factors associated with these complications. The initial consecutive 233 patients undergoing RAPN between March 2008 and May 2012 at our institution were identified. Patients were divided into quartiles to evaluate outcomes, and uni- and multivariate predictors of complications were calculated. In progressive quartiles, patients tended to have more complex renal tumours as evaluated by R.E.N.A.L. nephrometry (P < 0.01) or preoperative aspects and dimensions used for an anatomical classification (PADUA) scores (P = 0.003), and percentage endophytic (P = 0.01). Developing technique increased the patients undergoing unclamped PN (P < 0.01). The mean time in the operating room (from skin incision to closure) decreased significantly from 225 to 183 min (P < 0.01) and warm ischaemia time decreased from 28 to 15 min, when clamping (P < 0.01). Clavien graded complication rate (P = 0.26) and positive margin rate (P = 0.32) was unchanged by quartile. We show that increasing experience allows more complex tumours to be removed with similar outcomes in patients undergoing RAPN. The complication rates and perioperative outcomes were similar in four successive quartiles of an initial experience of RAPN.


A Modulatory Effect of Brief Passive Exposure to Non-linguistic Sounds on Intrinsic Functional Connectivity: Relevance to Cognitive Performance.
A growing literature on resting-state fMRI (R-fMRI) has explored the impact of preceding sensory experience on intrinsic functional connectivity (iFC). However, it remains largely unknown how passive exposure to irrelevant auditory stimuli, which is a constant in everyday life, reconfigures iFC. Here, we directly compared pre- and post-exposure R-fMRI scans to examine: 1) modulatory effects of brief passive exposure to repeating non-linguistic sounds on subsequent iFC, and 2) associations between iFC modulations and cognitive abilities. We used an exploratory regional homogeneity (ReHo) approach that indexes local iFC, and performed a linear mixed-effects modeling analysis. A modulatory effect (increase) in ReHo was observed in the right superior parietal lobule (R.SPL) within the parietal attention network. Post hoc seed-based correlation analyses provided further evidence for increased parietal iFC (e.g., R.SPL with the right inferior parietal lobule). Notably, less iFC modulation was associated with better cognitive performance (e.g., word reading). These results suggest that: 1) the parietal attention network dynamically reconfigures its iFC in response to passive (thus irrelevant) non-linguistic sounds, but also 2) minimization of iFC modulation in the same network characterizes better cognitive performance. Our findings may open up new avenues for investigating cognitive disorders that involve impaired sensory processing.


Clinical effectiveness of chin cup treatment for the management of Class III malocclusion in pre-pubertal patients: a systematic review and meta-analysis.
Chin cup is regarded as the oldest orthodontic appliance for the management of Class III malocclusion. To assess its clinical effectiveness in pre-pubertal patients, a meta-analysis on specific cephalometric values is attempted. Detailed electronic and hand searches with no restrictions were performed up to July 2014. Only randomized controlled trials (RCTs) and cohort studies, i.e. prospective controlled trials (pCCTs) and (retrospective) observational studies (OS), were included. Analyses were performed by calculating the standard difference in means and the corresponding 95% confidence intervals, using the random effects model. Data heterogeneity and risk of bias assessment of the included studies were also performed. Study selection, data extraction and risk of bias assessment were performed twice. The level of significance was set at P ≤ 0.05 for all tests, except for heterogeneity (P ≤ 0.1). Seven treated groups from five studies (no RCTs, four pCCTs, one OS) were eligible for inclusion, assessing only the short-term occipital pull chin cup effects. In total, 120 treated patients (mean age: 8.5 to 11 years) compared with 64 untreated individuals (mean age: 7.3 to 9.89 years) were assessed by means of 13 cephalometric variables. The overall quality of these studies was low to medium. In comparison to untreated individuals, the SNB and gonial angles decreased significantly following chin cup use, whereas ANB, Wits appraisal, SN-ML, N-Me and overjet increased. For the rest of the variables, no statistically significant differences were detected. Although the occipital chin cup affects significantly a number of skeletal and dentoalveolar cephalometric variables, indicating an overall positive effect for the treatment of Class III malocclusion, data heterogeneity and between-studies variance impose precaution in the interpretation of the results.


Physical disturbance shapes vascular plant diversity more profoundly than fire in the sagebrush steppe of southeastern Idaho, U.S.A.
Fire is thought to profoundly change the ecology of the sagebrush steppe. The Idaho National Laboratory provides an ideal setting to compare the effects of fire and physical disturbance on plant diversity in high-native-cover sagebrush steppe. Seventy-eight 1-hectare transects were established along paved, green-striped, gravel, and two-track roads, in overgrazed rangeland, and within sagebrush steppe involving different fire histories. Transects were sampled for the diversity and abundance of all vascular plants. Alpha, beta, and phylogenetic beta diversity were analyzed as a response to fire and physical disturbance. Postfire vegetation readily rebounds to prefire levels of alpha plant diversity. Physical disturbance, in contrast, strongly shapes patterns of alpha, beta, and especially phylogenetic beta diversity much more profoundly than fire disturbance. If fire is a concern in the sagebrush steppe then the degree of physical-disturbance should be more so. This finding is probably not specific to the study area but applicable to the northern and eastern portions of the sagebrush biome, which is characterized by a pulse of spring moisture and cold mean minimum winter temperatures. The distinction of sagebrush steppe from Great Basin sagebrush should be revised especially with regard to reseeding efforts and the control of annual grasses.


Interaction mechanism between the focused ultrasound and lipid membrane at the molecular level.
Focused ultrasound (FUS) has a wide range of medical applications. Nowadays, the diagnostic and therapeutic ultrasound procedures are routinely used; effects of ultrasound on biological systems at the molecular level are, however, not fully understood. Experimental results on the interaction of the cell membrane, a simplest but important system component, with ultrasound are controversial. Molecular dynamics (MD) simulations could provide valuable insights, but there is no single study on the mechanism of the FUS induced structural changes in cell membranes. With this in mind, we develop a simple method to include FUS into a standard MD simulation. Adopting the 1,2-dioleoyl-sn-glycero-3-phosphocholine lipid membrane as a representative model described by the MARTINI coarse-grained force field, and using experimental values of the ultrasound frequency and intensity, we show that the heat and bubble cavitation are not the primary direct mechanisms that cause structural changes in the membrane. The spatial pressure gradients between the focused and free regions and between the parallel and perpendicular directions to the membrane are the origin of the mechanism. These gradients force lipids to move out of the focused region, forming a lipid flow along the membrane diagonal. Lipids in the free region move in the opposite direction due to the conservation of the total momentum. These opposite motions create wrinkles along the membrane diagonal at low FUS intensities and tear up the membrane at high FUS intensities. Once the membrane is torn up, it is not easy to reform. The implication of our findings in the FUS-induced drug delivery is discussed in some detail.


Rejoice in the hubris: useful things biologists could do for physicists.
Political correctness urges us to state how wonderful it is to work with biologists and how, just as the lion will someday lie down with the lamb, so will interdisciplinary work, where biologists and physicists are mixed together in light, airy buildings designed to force socialization, give rise to wonderful new science. But it has been said that the only drive in human nature stronger than the sex drive is the drive to censor and suppress, and so I claim that it is OK for physicists and biologists to maintain a wary distance from each other, so that neither one censors or suppresses the wild ideas of the other.


Pontocerebellar projection to the rabbit paramedian lobule by means of axonal collaterals: evidence for intralobular connections.
In this study, we utilized a double retrograde axonal tracing technique to investigate the possible existence of collateralized axonal projections from the pontine nuclei (PN) to the rostral (rPML) and caudal (cPML) parts of cerebellar paramedian lobule in the rabbit, known to be the forelimb and hindlimb regions, respectively. Following injections of fluorescent tracers Fast Blue (FB) and Diamidino Yellow (DY) within rPML and cPML, respectively, substantial numbers of FB and DY single labeled neurons were found in the dorsolateral, paramedian, lateral and peduncular pontine nuclei bilaterally with a very clearcut contralateral preponderance. No labeling was observed in the ventral pontine nucleus. Extensive areas of overlap of FB or DY labeled neurons indicated that no somatotopical relationship existed in projection from PN to the two functionally different PML target regions. In addition, a small number of double FB + DY labeled neurons was detected in the common areas of FB and DY single labeling in PN. These neurons give rise to pontocerebellar projections to rPML and cPML simultaneously by way of axonal collaterals and thus they may play a role in the coordination of unilateral forelimb and hindlimb movements.


Flavobacterium stagni sp. nov., isolated from a freshwater reservoir.
Strain WWJ-16T was isolated from a freshwater reservoir in Taiwan. Phylogenetic analyses based on 16S rRNA gene sequences and an up-to-date bacterial core gene set (92 protein clusters) indicated that strain WWJ-16T is affiliated with the species in the genus Flavobacterium. Strain WWJ-16T was most closely related to Flavobacterium fontis MIC3010T (97.9 % 16S rRNA gene sequence identity) and Flavobacterium squillarum CMJ-5T (97.6 %). Strain WWJ-16T showed 77.4 % average nucleotide identity and 20.6 % digital DNA-DNA hybridization identity with F. fontis MIC3010T. Cells of strain WWJ-16T were Gram-stain-negative, strictly aerobic, non-motile, rod-shaped and formed yellow colonies. Optimal growth occurred at 25 °C, pH 7 and in 0.5-1 % NaCl (w/v). Strain WWJ-16T contained iso-C15 : 1 G and iso-C15 : 0 as predominant fatty acids. The major hydroxyl fatty acids were iso-C15 : 0 3-OH, iso-C17 : 0 3-OH and iso-C16 : 0 3-OH. The polar lipid profile consisted of phosphatidylethanolamine, four uncharacterized aminophospholipids and four uncharacterized phospholipids. The major polyamine was homospermidine. The major isoprenoid quinone was MK-6. The genomic DNA G+C content of strain WWJ-16T was 39.4 mol%, as determined by genome sequencing. The genotypic and phenotypic characteristics indicate that strain WWJ-16T represents a novel species of the genus Flavobacterium, for which the name Flavobacterium stagni sp. nov. is proposed. The type strain is WWJ-16T (=BCRC 81125T =LMG 30600T=KCTC 62515T).


Birth defects in children conceived by ICSI compared with children conceived by other IVF-methods; a meta-analysis.
Intracytoplasmic sperm injection (ICSI) is a method of assisted reproductive technology that involves the selection of a single sperm cell and the manual injection of this cell into the egg. The lack of relevant experimental studies, the nature of the technology involving non-natural selection of the fertilizing sperm, and possible damage to the egg have caused concern that ICSI could increase the risk of birth defects. Data from available cohort studies comparing ICSI with standard in vitro fertilization (IVF) should be combined to evaluate the risks involved with ICSI. We reviewed more than 2500 titles and abstracts containing keywords related to ICSI and identified 22 scientific articles with data on birth defects among ICSI-births. A total of four peer-reviewed, non-overlapping prospective cohort studies provided reliable and comparable data on birth defects both for children conceived by ICSI and children conceived by standard IVF. These studies included a total of 5395 children born after ICSI. The pooled estimate of the risk of a major birth defect was a 1.12-fold increase after ICSI when compared with standard IVF (risk ratio = 1.12, 95% confidence interval (CI): 0.97-1.28, P = 0.12). There was no marked heterogeneity of risk ratios between these studies (P = 0.10). We found no significantly increased risks after ICSI for any of the categories cardiovascular defects, musculoskeletal defects, hypospadias, neural tube defects, or oral clefts. Our analysis does not indicate that the ICSI-procedure represents significant additional risks of major birth defects in addition to the risk involved in standard IVF. The data was limited, particularly on risks of specific categories of defects.


Inactivation of Rb and E2f8 synergizes to trigger stressed DNA replication during erythroid terminal differentiation.
Rb is critical for promoting cell cycle exit in cells undergoing terminal differentiation. Here we show that during erythroid terminal differentiation, Rb plays a previously unappreciated and unorthodox role in promoting DNA replication and cell cycle progression. Specifically, inactivation of Rb in erythroid cells led to stressed DNA replication, increased DNA damage, and impaired cell cycle progression, culminating in defective terminal differentiation and anemia. Importantly, all of these defects associated with Rb loss were exacerbated by the concomitant inactivation of E2f8. Gene expression profiling and chromatin immunoprecipitation (ChIP) revealed that Rb and E2F8 cosuppressed a large array of E2F target genes that are critical for DNA replication and cell cycle progression. Remarkably, inactivation of E2f2 rescued the erythropoietic defects resulting from Rb and E2f8 deficiencies. Interestingly, real-time quantitative PCR (qPCR) on E2F2 ChIPs indicated that inactivation of Rb and E2f8 synergizes to increase E2F2 binding to its target gene promoters. Taken together, we propose that Rb and E2F8 collaborate to promote DNA replication and erythroid terminal differentiation by preventing E2F2-mediated aberrant transcriptional activation through the ability of Rb to bind and sequester E2F2 and the ability of E2F8 to compete with E2F2 for E2f-binding sites on target gene promoters.


Maternal transmission disequilibrium of the glutamate receptor GRIK2 in schizophrenia.
Schizophrenia is characterized by thought disorders, hallucinations and delusions. Genetic studies have shown a high linkage at chromosome 6q16-21. Among the genes located in this region is the glutamate receptor ionotropic kainate 2 gene (GRIK2 or GLUR6), a functional candidate for susceptibility to schizophrenia. In this study, transmission of GRIK2 was evaluated in 356 schizophrenic patients from three different clinical centers. Whereas paternal transmission shows equilibrium, we observed maternal transmission disequilibrium of GRIK2 in the largest population (p=0.03), which was still significant when all populations were added (p=0.05). These results are similar to the maternal GRIK2 transmission disequilibrium previously reported for autism, and support the presence of a susceptibility gene for schizophrenia at 6q16.


[Irreversible changes in phage DNA after its protonation in solution and inside the virion detected by the transfection method].
The dependence of irreversible structural changes in phage lambda DNA on the degree of its protonation in a solution and inside the virion has been found by measuring the transfection activity of bacteriophage. The different effect of ionic strength on pH-dependence of the irreversible changes in the structure of DNA upon its protonation in a solution or in situ has been registered and explained. The insignificant shift of pH from neutral region value in 0.1 M NaCl has resulted in a damaging effect of H+ ions on compact DNA in situ as compared to the DNA in a solution. The effect of H+ ions on compact DNA in situ is mainly based on the formation of noncovalent intermolecular DNA-protein and DNA-DNA linkages.


Modulation of liver regeneration via myeloid PTEN deficiency.
Molecular mechanisms that modulate liver regeneration are of critical importance for a number of hepatic disorders. Kupffer cells and natural killer (NK) cells are two cell subsets indispensable for liver regeneration. We have focused on these two populations and, in particular, the interplay between them. Importantly, we demonstrate that deletion of the myeloid phosphatase and tensin homolog on chromosome 10 (PTEN) leading to an M2-like polarization of Kupffer cells, which results in decreased activation of NK cells. In addition, PTEN-deficient Kupffer cells secrete additional factors that facilitate the proliferation of hepatocytes. In conclusion, PTEN is critical for inhibiting M2-like polarization of Kupffer cells after partial hepatectomy, resulting in NK cell activation and thus the inhibition of liver regeneration. Furthermore, PTEN reduces growth factor secretion by Kupffer cells. Our results suggest that targeting PTEN on Kupffer cells may be useful in altering liver regeneration in patients undergoing liver resection.


Evidence for a direct reticulocyte origin of dense red cells in sickle cell anemia.
To explore our hypothesis of a direct reticulocyte origin of irreversibly sickled cells (ISCs), we fractionated light, reticulocyte-rich, and discocyte-rich sickle anemia red cells on Stractan gradients, and examined the effects of deoxygenation-induced sickling, external Ca2+, acidification, and replacing external Na+ by impermeant N-methyl-D-glucamine (NMG+). Sickling permeabilized light reticulocyte-rich cells to cations (Na+, K+, and Ca2+) more than discocytes; without external Ca2+, Na+ influx matched K+ efflux, with stable cell volume; with Ca2+, many light, low hemoglobin (Hb) F reticulocytes dehydrated rapidly (preventable by quinine, a Ca2(+)-dependent K+ channel inhibitor). Acidification of oxygenated discocytes (high mean Hb F) and reticulocyte-rich fractions yielded denser, reticulocyte-enriched cells with lower Hb F (as in light reticulocyte or dense ISC-rich fractions). Light cells shrank when NMG+ replaced Na+, supporting predictions of a Na(+)-dependent volume control system. Demonstration of sickling-induced, Ca2(+)-dependent dehydration of Hb F-free reticulocytes, and conservation of acid-stimulated K:Cl cotransport among low Hb F, reticulocyte-enriched cells in discocyte fractions support the hypothesis. Ancillary new findings included heparin stimulation of sickling-induced Na+ and K+ permeabilizations, and Ca2+ inhibition of the Na+ leak.


Biochemical characterization a digestive trypsin in the midgut of large cabbage white butterfly, Pieris brassicae L. (Lepidoptera: Pieridae).
A comprehensive study on digestive trypsin was undertaken in the larval midgut of Pieris brassicae L. Results of enzymatic compartmentalization showed a significantly higher activity of crude trypsin in the anterior larval midgut rather than posterior-midgut. Using Diethylaminoethyl cellulose fast flow column chromatography a purified trypsin was obtained by specific activity of 21 U mg-1 protein, recovery of 22%, purification fold of 28-fold and molecular weight of 25 kDa. This purified enzyme showed the highest activity at pH 8 and the corresponding temperature of 40°C. However, the specific inhibitors used including 4-(2-Aminoethyl) benzenesulfonyl fluroride hydrochloride, N-p-Tosyl-L-lysine methyl ester hydrochloride and Soybean Trypsin Inhibitor significantly lowered the activity of the purified enzyme in vitro. Moreover, the activity of trypsin and likewise the nutritional indices were significantly altered in the larval midgut feeding upon the leaves treated by 1 mM concentration of each inhibitor in comparison with control. Determination of enzymatic characteristics of insect trypsins is crucial in paving the path for controlling pests by potential natural compounds via transgenic plants.


Truncation of the carboxy-terminal domain of yeast beta-tubulin causes temperature-sensitive growth and hypersensitivity to antimitotic drugs.
beta-tubulin of budding yeast Saccharomyces cerevisiae is a polypeptide of 457 amino acids encoded by the unique gene TUB2. We investigated the function of the carboxy-terminal part of yeast beta-tubulin corresponding to the carboxy-terminal variable domain of mammalian and avian beta-tubulins. The GAA codon for Glu-431 of TUB2 was altered to TAA termination codon by using in vitro site-directed mutagenesis so that the 27-amino acid residues of the carboxyl terminus was truncated when expressed. The mutagenized TUB2 gene (tub2(T430)) was introduced into a haploid strain in which the original TUB2 gene had been disrupted. The tub2(T430) haploid strain grows normally less than 30 but not at 37 degrees C. The truncation of the carboxyl terminus caused hypersensitivity to antimitotic drugs and low spore viability at the permissive temperature for vegetative growth. Immunofluorescence labeling with antitubulin antibody and DNA staining with 4',6'-diamidino-2-phenylindole showed that in these cells at 37 degrees C, formation of spindle microtubules and nuclear division was inhibited and cytoplasmic microtubule distribution was aberrant. These results suggest that functions of the carboxy-terminal domain of yeast beta-tubulin are necessary for cells growing under suboptimal growth conditions although it is not essential for growth under the optimal growth conditions. Cells bearing tub2(411), a tub2 gene in which the GAA codon for Glu-412 was altered to TAA were no more viable at any temperature. In addition, a haploid strain carrying two functional beta-tubulin genes is not viable.


Utilization of public mental health services in a random community sample.
The aim of this study was to examine patterns of service utilization in the public mental health service in the Australian state of Victoria. Lifetime contact with the public mental health system will be used to approximate lifetime prevalence for schizophrenia in the community. A case-linkage design joined a statewide psychiatric register with a random sample of community members drawn from a statewide electoral roll (n = 4830). In cases where individuals had been in contact with public mental health services, their full contact history was extracted. Members of the community come into contact with public mental health services for a variety of reasons, including those beyond the scope of psychiatric diagnosis and treatment, with 23% of those who made contact not receiving a psychiatric diagnosis and/or ongoing treatment. Although only 0.7% of the sample had a lifetime diagnosis of schizophrenia, these persons accounted for a significant proportion of public mental health service use. Schizophrenia-spectrum disorders were particularly prevalent among psychiatric crisis and extended care and supervision services. The Australian public mental health system has undergone significant reform in recent years. As a result, there has been a shift towards community-based care and a marked reduction in inpatient facilities. The public mental health system is a service that primarily serves those with a psychotic illness, thereby dictating that persons with so-called high-prevalence disorders, including affective and/or substance use disorders, seek psychiatric treatment elsewhere.


Engineered resistance to Plasmodium falciparum development in transgenic Anopheles stephensi.
Transposon-mediated transformation was used to produce Anopheles stephensi that express single-chain antibodies (scFvs) designed to target the human malaria parasite, Plasmodium falciparum. The scFvs, m1C3, m4B7, and m2A10, are derived from mouse monoclonal antibodies that inhibit either ookinete invasion of the midgut or sporozoite invasion of salivary glands. The scFvs that target the parasite surface, m4B7 and m2A10, were fused to an Anopheles gambiae antimicrobial peptide, Cecropin A. Previously-characterized Anopheles cis-acting DNA regulatory elements were included in the transgenes to coordinate scFv production with parasite development. Gene amplification and immunoblot analyses showed promoter-specific increases in transgene expression in blood-fed females. Transgenic mosquito lines expressing each of the scFv genes had significantly lower infection levels than controls when challenged with P. falciparum.