Expression of a Drosophila mRNA is under circadian clock control during pupation.
Rhythmic eclosion of Drosophila adults requires per gene function. We have found that a previously identified 0.9 kb RNA transcribed from DNA adjacent to per becomes abundantly expressed during pupation, just prior to eclosion. The daily synchronized emergence of young adults, coupled with a subsequent rapid decay of the transcript, is responsible for what previously appeared to be cycling of the 0.9 kb RNA in adults. In situ hybridization analyses localize the 0.9 kb transcript to the epidermis of newly eclosed adults. Conceptual translation of genomic DNA and cDNA sequences predicts that the 0.9 kb transcript produces a 261 amino acid protein containing a putative signal sequence for membrane transport at its amino terminus. Pupae that reach the same stage of development at slightly different times of day show a subsequent synchronized rise in 0.9 kb RNA levels, indicating that the expression of this transcript is under circadian clock control.


Ontogenetic resource utilization and migration reconstruction with δ13C values of essential amino acids in the Cynoscion acoupa otolith.
With the increasing anthropogenic impacts on fish habitats, it has become more important to understand which primary resources sustain fish populations. This resource utilization can differ between fish life stages, and individuals can migrate between habitats in search of resources. Such lifetime information is difficult to obtain due to the large spatial and temporal scales of fish behavior. The otolith organic matrix has the potential to indicate this resource utilization and migration with δ13C values of essential amino acids (EAAs), which are a direct indication of the primary producers. In a proof-of-concept study, we selected the Acoupa weakfish, Cynoscion acoupa, as a model fish species with distinct ontogenetic migration patterns. While it inhabits the Brazilian mangrove estuaries during juvenile stages, it moves to the coastal shelf as an adult. Thus, we expected that lifetime resource utilization and migration would be reflected in δ13CEAA patterns and baseline values in C. acoupa otoliths. By analyzing the C. acoupa otolith edges across a size range of 12-119 cm, we found that baseline δ13CEAA values increased with size, which indicated an estuarine to coastal shelf distribution. This trend is highly correlated with inorganic δ13C values. The δ13CEAA patterns showed that estuarine algae rather than mangrove-derived resources supported the juvenile C. acoupa populations. Around the juvenile size of 40 cm, resource utilization overlapped with those of adults and mean baseline δ13CEAA values increased. This trend was confirmed by comparing otolith core and edges, although with some individuals potentially migrating over longer distances than others. Hence, δ13CEAA patterns and baseline values in otoliths have great potential to reconstruct ontogenetic shifts in resource use and habitats. The insight could aid in predictions on how environmental changes affect fish populations by identifying the controlling factors at the base of the food web.


Characteristics and clustering of human ribosomal protein genes.
The ribosome is a central player in the translation system, which in mammals consists of four RNA species and 79 ribosomal proteins (RPs). The control mechanisms of gene expression and the functions of RPs are believed to be identical. Most RP genes have common promoters and were therefore assumed to have a unified gene expression control mechanism. We systematically analyzed the homogeneity and heterogeneity of RP genes on the basis of their expression profiles, promoter structures, encoded amino acid compositions, and codon compositions. The results revealed that (1) most RP genes are coordinately expressed at the mRNA level, with higher signals in the spleen, lymph node dissection (LND), and fetal brain. However, 17 genes, including the P protein genes (RPLP0, RPLP1, RPLP2), are expressed in a tissue-specific manner. (2) Most promoters have GC boxes and possible binding sites for nuclear respiratory factor 2, Yin and Yang 1, and/or activator protein 1. However, they do not have canonical TATA boxes. (3) Analysis of the amino acid composition of the encoded proteins indicated a high lysine and arginine content. (4) The major RP genes exhibit a characteristic synonymous codon composition with high rates of G or C in the third-codon position and a high content of AAG, CAG, ATC, GAG, CAC, and CTG. Eleven of the RP genes are still identified as being unique and did not exhibit at least some of the above characteristics, indicating that they may have unknown functions not present in other RP genes. Furthermore, we found sequences conserved between human and mouse genes around the transcription start sites and in the intronic regions. This study suggests certain overall trends and characteristic features of human RP genes.


Enhancement of biogas production from food waste and sewage sludge - Environmental and economic life cycle performance.
Management of municipal solid waste is an efficient method to increase resource efficiency, as well as to replace fossil fuels with renewable energy sources due to that (1) waste to a large extent is renewable as it consists of food waste, paper, wood etc. and (2) when energy and materials are recovered from waste treatment, fossil fuels can be substituted. In this paper results from a comprehensive system study of future biological treatment of readily degradable waste in two Swedish regions are presented. Different collection and separation systems for food waste in households have been applied as well as technical improvements of the biogas process as to reduce environmental impact. The results show that central sorting of a mixed fraction into recyclables, combustibles, biowaste and inert is a competitive option compared to source separation. Use of pellets is beneficial compared to direct spreading as fertiliser. Fuel pellets seem to be the most favourable option, which to a large extent depends on the circumstances in the energy system. Separation and utilisation of nitrogen in the wet part of the digestion residue is made possible with a number of technologies which decreases environmental impact drastically, however to a substantial cost in some cases.


Activation of the West Nile virus NS3 protease: molecular dynamics evidence for a conformational selection mechanism.
The flaviviral nonstructural 3 protease (NS3pro) is essential for virus replication and is therefore a pharmaceutically relevant target to fight Dengue and West Nile virus (WNV). NS3pro is a chymotrypsin-like serine protease which requires a polypeptide cofactor (NS2B) for activation. Recent X-ray crystallography studies have led to the suggestion that the substrate binds to the two-component NS2B-NS3pro enzyme by an induced-fit mechanism. Here, multiple explicit water molecular dynamics simulations of the WNV NS2B-NS3pro enzyme show that the active conformation of the NS2B cofactor (in which its beta-loop is part of the substrate binding site) is stable over a 50-ns time scale even in the absence of the inhibitor. The partial and reversible opening of the NSB2 beta-loop and its correlated motion with an adjacent NS3pro loop, both observed in the simulations started from the active conformation, are likely to facilitate substrate binding and product release. Moreover, in five of eight simulations without inhibitor (started from two X-ray structures both with improperly formed oxyanion hole) the Thr132-Gly133 peptide bond flips spontaneously thereby promoting the formation of the catalytically competent oxyanion hole, which then stays stable until the end of the runs. The simulation results provide evidence at atomic level of detail that the substrate binds to the NS2B-NS3pro enzyme by conformational selection, rather than induced-fit mechanism.


A prospective study of the anatomic variations of the median nerve in the carpal tunnel in Asians.
A prospective study of anatomic variations of the median nerve and its relationship to the surrounding structures was performed of 354 consecutive operations in 192 patients with carpal tunnel syndrome at Korea University Anam Hospital from July 1995 to September 1997. A total of 184 patients were women and 8 patients were men. A total of 162 patients were bilateral and 30 were unilateral. Regarding the course of the motor branch, the extraligamentous type was most common (96.1%, 340 patients). A total of 81.1% of patients (N = 287) had the origin of the motor branch at the radial side of the median nerve (or radial one third), and 17.2% of patients (N = 61) had the origin of the motor branch at the anterior portion of the median nerve. Of these 61 patients, 30 patients (49.1%) were of the preligamentous type. Only one motor nerve branch was found in 89.5% of patients (N = 317), and multiple branches were found in 10.5% of patients. As a result of the comparison between two hands when both hands were operated, the origin and number of motor branches and their courses were identical in most patients (>60%). Hypertrophy of the flexor pollicis brevis was found in 8.5% of patients (N = 30), hypertrophy of the palmaris brevis was found in 4.2% of patients (N = 15), absence of the palmaris longus was discovered in 0.6% of patients (N = 2), existence of the median artery was found in 0.6% of patients (N = 2), and high division of the median nerve rejoining with the thenar motor nerve was found in 0.3% of patients (N = 1). Clearly, the anatomy of the carpal tunnel in Koreans is somewhat different, in part, from the results obtained from studies of whites, but overall results are not significantly different. These results can help obtain a better surgical outcome and complete decompression of the median nerve during operation while preventing inappropriate or inadvertent injury to the motor branch of the median nerve in Koreans.


Heredity, pet ownership, and confounding control in a population-based birth cohort.
The association between pet ownership in childhood and subsequent allergic disease is controversial. Bias related to selection of pet exposure has been suggested as a reason for contradictory study results. The purpose of this investigation was to elucidate how pet exposure depends on family history of allergic disease, smoking, and socioeconomic factors in a prospective birth cohort. Parents of 4089 two-month-old children answered a questionnaire that included detailed questions about family history of asthma (maternal, paternal, and sibling), rhinoconjunctivitis, atopic eczema/dermatitis syndrome, pollen and pet allergy, smoking habits, parental occupation, and family pet ownership (cat and dog). Dust samples collected from the mothers' beds were analyzed for Fel d 1 and Can f 1 in a subgroup of the cohort. Cats were less frequently kept in families with parental asthma, rhinoconjunctivitis, or pet or pollen allergy (3.5% to 5.8%) than in families without parental allergic disease (10.8% to 11.8%). Dogs were less common in families with (3.3%) than in families without (5.9%) parental atopic eczema/dermatitis syndrome. Families with smoking mothers and those with low socioeconomic index kept cats and dogs more frequently. Cat allergen levels were lower in homes with than in homes without maternal pet allergy, and this tended to hold true even for homes without a cat. Cat ownership decreased from birth to 2 years of age, especially in families with parental history of allergic diseases. There seems to be a selection of pet exposure based on parental history of allergy, maternal smoking, and socioeconomic factors. This has to be taken into consideration in evaluations of risk associations between pet exposure and allergic disease in childhood.


Impact of TREM2R47H variant on tau pathology-induced gliosis and neurodegeneration.
Alzheimer's disease (AD) is characterized by plaques containing amyloid-β (Aβ) and neurofibrillary tangles composed of aggregated, hyperphosphorylated tau. Beyond tau and Aβ, evidence suggests that microglia play an important role in AD pathogenesis. Rare variants in the microglia-expressed triggering receptor expressed on myeloid cells 2 (TREM2) gene increase AD risk 2- to 4-fold. It is likely that these TREM2 variants increase AD risk by decreasing the response of microglia to Aβ and its local toxicity. However, neocortical Aβ pathology occurs many years before neocortical tau pathology in AD. Thus, it will be important to understand the role of TREM2 in the context of tauopathy. We investigated the impact of the AD-associated TREM2 variant (R47H) on tau-mediated neuropathology in the PS19 mouse model of tauopathy. We assessed PS19 mice expressing human TREM2CV (common variant) or human TREM2R47H. PS19-TREM2R47H mice had significantly attenuated brain atrophy and synapse loss versus PS19-TREM2CV mice. Gene expression analyses and CD68 immunostaining revealed attenuated microglial reactivity in PS19-TREM2R47H versus PS19-TREM2CV mice. There was also a decrease in phagocytosis of postsynaptic elements by microglia expressing TREM2R47H in the PS19 mice and in human AD brains. These findings suggest that impaired TREM2 signaling reduces microglia-mediated neurodegeneration in the setting of tauopathy.


A randomized, double-blind trial of valaciclovir prophylaxis for cytomegalovirus disease in patients with advanced human immunodeficiency virus infection. AIDS Clinical Trials Group Protocol 204/Glaxo Wellcome 123-014 International CMV Prophylaxis Study Group.
Cytomegalovirus (CMV) disease is a common complication of advanced human immunodeficiency virus (HIV) infection. Administration of oral valaciclovir, a valine ester of acyclovir, achieves sufficient plasma acyclovir levels to inhibit many clinical isolates. Acyclovir has been associated with enhanced survival in AIDS but not with CMV disease prevention. CMV-seropositive patients (1227) with CD4 cell counts <100/mm3 were enrolled in a randomized, double-blind trial. Valaciclovir, 8 g/day, was compared with acyclovir, 3.2 or 0.8 g/day, for CMV prevention; all three arms were compared for survival. The confirmed CMV disease rate was 11.7% among valaciclovir recipients and 17.5% in the pooled acyclovir arms, a 33% reduction in risk. Time to confirmed CMV disease was significantly longer for the valaciclovir group (P = .03). A trend toward earlier mortality for valaciclovir recipients was seen (P = .06). Toxicity and earlier medication discontinuation were more common in this group. Valaciclovir significantly reduces the risk of CMV disease. Further exploration of a better-tolerated dose is warranted.


Plant mitochondria contain the protein translocase subunits TatB and TatC.
Twin-arginine translocation (Tat) pathways have been well-characterized in bacteria and chloroplasts. Genes encoding a TatC protein are found in almost all plant mitochondrial genomes but to date these have not been extensively investigated. For the first time it could be demonstrated that this mitochondrial-encoded TatC is a functional gene that is translated into a protein in the model plant Arabidopsis thaliana A TatB--like subunit localized to the inner membrane was also identified that is nuclear-encoded and is essential for plant growth and development, indicating that plants potentially require a Tat pathway for mitochondrial biogenesis.


Partial diazoxide responsiveness in a neonate with hyperinsulinism due to homozygous ABCC8 mutation.
We report a case of partial diazoxide responsiveness in a child with severe congenital hyperinsulinaemic hypoglycaemia (CHI) due to a homozygous ABCC8 mutation. A term baby, with birth weight 3.8 kg, born to consanguineous parents presented on day 1 of life with hypoglycaemia. Hypoglycaemia screen confirmed CHI. Diazoxide was commenced on day 7 due to ongoing elevated glucose requirements (15 mg/kg/min), but despite escalation to a maximum dose (15 mg/kg/day), intravenous (i.v.) glucose requirement remained high (13 mg/kg/min). Genetic testing demonstrated a homozygous ABCC8 splicing mutation (c.2041-1G>C), consistent with a diffuse form of CHI. Diazoxide treatment was therefore stopped and subcutaneous (s.c.) octreotide infusion commenced. Despite this, s.c. glucagon and i.v. glucose were required to prevent hypoglycaemia. A trial of sirolimus and near-total pancreatectomy were considered, however due to the significant morbidity potentially associated with these, a further trial of diazoxide was commenced at 1.5 months of age. At a dose of 10 mg/kg/day of diazoxide and 40 µg/kg/day of octreotide, both i.v. glucose and s.c. glucagon were stopped as normoglycaemia was achieved. CHI due to homozygous ABCC8 mutation poses management difficulties if the somatostatin analogue octreotide is insufficient to prevent hypoglycaemia. Diazoxide unresponsiveness is often thought to be a hallmark of recessively inherited ABCC8 mutations. This patient was initially thought to be non-responsive, but this case highlights that a further trial of diazoxide is warranted, where other available treatments are associated with significant risk of morbidity. Learning points: Homozygous ABCC8 mutations are commonly thought to cause diazoxide non-responsive hyperinsulinaemic hypoglycaemia. This case highlights that partial diazoxide responsiveness in homozygous ABCC8 mutations may be present. Trial of diazoxide treatment in combination with octreotide is warranted prior to considering alternative treatments, such as sirolimus or near-total pancreatectomy, which are associated with more significant side effects.


Auditory localization of ground-borne vibrations in snakes.
Interaural time differences allow many animals to perform azimuthal sound localization. Snakes lack a tympanic membrane, external ear openings, and any other superficial indication of an auditory mechanism. They do, however, possess an inner ear with functional cochlea. The oval window is connected through a loss-free osseous lever system to the two, de facto independent, sides of the lower jaw, which typically rest on the substrate. The footfall of prey generates small-amplitude, low propagation velocity, Rayleigh waves in the soil. This type of wave can be described as fluid motion. Accordingly we apply naval-engineering techniques to show that lower-jaw motion gives rise to a neuronal representation of the auditory world with realistic sensitivity and stereo precision.


Effect of amniotic membrane transplantation on the healing of bacterial keratitis.
To study, with the use of an animal model, the efficacy of amniotic membrane (AM) transplantation as adjunctive treatment in corneal healing after bacterial keratitis. Staphylococcus aureus keratitis was induced in 47 rats by injection of bacteria into the corneal stroma. Treatment was started 48 hours later with one of three randomly assigned protocols: cefazolin drops (50 mg/mL) and AM transplantation (n = 16); nonpreserved 0.9% saline drops and AM transplantation (n = 15); or cefazolin without AM transplantation (n = 16). Cefazolin and saline drops were administered every 30 minutes for 6 hours, then hourly for 6 hours. AM was transplanted 24 hours after termination of cefazolin or saline treatment. Results were clinically assessed 7 days after AM transplantation or at the corresponding time in the nontransplanted animals. The rats were then killed, and their corneas were removed for bacterial counts or histopathologic examination. The best clinical results were observed in the group treated with cefazolin and AM transplantation, manifested by the least corneal haze and neovascularization (P = 0.007 and P = 0.014, respectively) and minimal bacterial counts (28 colony-forming units [CFU]/mL compared with 160 CFU/mL and 240 CFU/mL, respectively). Histopathologic examination showed that the central corneal vessels from rats treated with cefazolin and AM were smaller and less congested than those from the other two groups. AM transplantation is a useful adjunctive treatment after bacterial keratitis in this rat model. The transplanted AM improved the healing process, resulting in decreased corneal haze and less neovascularization.


Programmed cell death of adult-generated hippocampal neurons is mediated by the proapoptotic gene Bax.
In the dentate gyrus (DG) of the adult mouse hippocampus, a substantial number of new cells are generated daily, but only a subset of these survive and differentiate into mature neurons, whereas the majority undergo programmed cell death (PCD). However, neither the intracellular machinery required for adult stem cell-derived neuronal death nor the biological implications of the significant loss of these newly generated cells have been examined. Several markers for apoptosis failed to reveal cell death in Bax-deficient mice, and this, together with a progressive increase in neuron number in the DG of the Bax knock-out, indicates that Bax is critical for the PCD of adult-generated hippocampal neurons. Whereas the proliferation of neural progenitor cells was not altered in the Bax-knock-out, there was an accumulation of doublecortin, calretinin+, and neuronal-specific nuclear protein+ postmitotic neurons, suggesting that Bax-mediated PCD of adult-generated neurons takes place during an early phase of differentiation. The absence of PCD in the adult also influenced the migration and maturation of adult-generated DG neurons. These results suggest that PCD in the adult brain plays a significant role in the regulation of multiple aspects of adult neurogenesis.


[The clinical report of 1,078 cases of hepatocellular carcinomas: National Cancer Center experience].
Hepatocellular carcinoma (HCC) is the third leading cause of cancer and the 5 year survival rate is 9.6% in Korea. To develop a strategy for surveillance and treatments, we studied the recent clinical characteristics of HCC diagnosed at single institution in Korea, where is in an endemic area of chronic hepatitis B. One thousand and seventy eight patients with HCC who visited the National Cancer Center between June 2001 and December 2003 were retrospectively studied. The male/female ratio was 4.5:1. The mean age of the patients was 56.3 years. 74.2% of patient had hepatitis B virus (HBV) infections, 8.6% had hepatitis C virus (HCV) infections, 6.9% of the patients abused alcohol and 10.3% of the patients had non-B non-C considered as the etiologic factors of their HCC. Only 10.0% of patients had a tumor sized 2 cm or less and 53.3% of patients had a large tumor over 5 cm in diameter. 33.2% of patients had a single tumor. At the time of diagnosis, the modified UICC staging was as follows: 6.5%, 20.1%, 30.9%, 25.2% and 17.3% in stages I, II, III, IVa and IVb, respectively. The initial treatment performed was transcatheter arterial chemoembolization (48.2%), radiofrequency ablation (1.5%), hepatic resection (11.2%), systemic chemotherapy (7.5%), radiotherapy (2.1%), and conservative medical treatments (29.5%). The mean number of treatments was 1.65. The response rates to the initial treatments were 27.9% (complete response), 23.6% (partial response), 7.5% (minimal response), 14.2% (stable disease), and 30.4% (progressive disease). HBV infection is a major etiologic factor for Korean HCC patients. Most cases are still in advanced stages and these cases responded poorly to any treatments. The national surveillance program and its guideline for HCC are expected to improve the survival of HCC patients.


Pregnancy restores the renal vasodilator response to glycine in Dahl salt-sensitive rats.
To determine if pregnancy alters the impaired renal vasodilator responses in hypertensive Dahl salt-sensitive (Dahl S) rats, we measured glomerular filtration rate and effective renal plasma flow and calculated renal vascular resistance before, during, and after renal vasodilation with glycine (0.17 mmol/kg per minute IV). Conscious, midterm (day 11 to 14) pregnant and age-matched virgin Dahl S and Dahl salt-resistant (Dahl R) rats were fed an 8% NaCl diet for 4 weeks (n = 6 per group). Mean arterial pressure was elevated (P < .05) in Dahl S compared with Dahl R rats, with no significant difference between pregnant and virgin animals in either group. Pregnancy resulted in significant increases in plasma volume, baseline glomerular filtration rate, and renal plasma flow and a significant decrease in renal vascular resistance. The glycine-induced increase in filtration rate in virgin Dahl S rats (27 +/- 4%) was less (P < or = .01) than pregnant Dahl S (60 +/- 4%) and either group of Dahl R (virgin, 43 +/- 3%; pregnant, 45 +/- 5%) rats. Similarly, the 21 +/- 4% increase in renal plasma flow in virgin Dahl S rats was less (P < or = .01) than the pregnant Dahl S (45 +/- 4%) and either pregnant (45 +/- 5%) or virgin (45 +/- 5%) Dahl R rats. Glycine decreased renal vascular resistance only 12 +/- 2% in the virgin Dahl S compared with 31 +/- 3% in the pregnant Dahl S rats and 30 +/- 4% and 28 +/- 3% in pregnant and virgin Dahl R rats.(ABSTRACT TRUNCATED AT 250 WORDS)


Gastrointestinal drug receptors.
Drug receptors consist of recognition sites coupled to transducer and cellular amplifier mechanisms. Slight differences in receptor recognition sites can be exploited pharmacologically to provide drugs with a high degree of selectivity for activating or blocking individual receptor subtypes. For example, it may now be possible to block, selectively, subtypes of muscarinic cholinergic receptors in the gastrointestinal tract with pirenzepine and other drugs that discriminate between subtypes of muscarinic cholinergic receptors. The recognition of subtypes of adrenergic receptors may allow highly selective pharmacological activation and blockage of gastrointestinal neural and smooth muscle receptors. The development of nonpeptide receptor antagonists of gastrointestinal hormones and peptide neurotransmitters also offers promise for improved therapy of digestive diseases. Tremendous progress has occurred in recent years in defining multiple types of opioid receptors that alter gastrointestinal secretory, absorptive and motility functions. These receptors are located in the mucosa, nerves and muscle of the intestine and in the brain and spinal cord.


Homing behavior of pigeons after telencephalic ablations.
In a first experiment, dorsomedial forebrain ablated birds showed similar homeward orientation when compared to untreated controls independent of whether the birds were released from a previous training site or a site they had never been before. However, although all control birds returned to the home loft, only 2 of 28 birds with lesions homed successfully. In a subsequent experiment, both sham operated control birds and birds with lesions of the visual Wulst homed successfully when released only 800 m from and in full view of their respective home lofts. Pigeons with dorsomedial forebrain lesions, however, failed to return to their respective home lofts. The results show that the avian dorsomedial forebrain plays a critical role in that step of the homing process by which a pigeon returns to its home loft once in its vicinity, and that the failure to reassociate with the home loft is a likely result of deficient recognition of the home loft and/or its surrounding area. In an additional experiment, pigeons with Wulst lesions were shown to orient as controls and to successfully return to the home loft when released from two distant sites. This experiment demonstrated that the avian Wulst plays no necessary role in the homing behavior of pigeons.


Cardiovascular abnormalities in children: a ten-year forensic pathologic study.
In order to obtain information on types, incidence, and significance of cardiovascular abnormalities in children, a total of 104 consecutive medicolegal autopsies of children aged 8 days to 16 years during a 10-year period from May 1974 to April 1984 were studied. Extensive histological examination of the hearts was performed in 92 out of 104 cases and complemented with chemical and microbiological analyses. In the natural death group consisting of 53 children, 26 (49%) showed abnormalities: 7 (13%) malformations, 11 (21%) cardiomyopathies, 5 (9%) idiopathic subaortic hypertrophy, and 3 (7%) a heart weight only 50% of the expected weight. In the violent death ("control") group, abnormalities were found in 8 of 39 cases (21%), all of which were cardiomyopathy. Only 5 of 34 cardiovascular abnormalities (every 7th case), all complex malformations, were clinically recognized. In 14 (15%) of the total 92 examined cases the cardiovascular abnormality was the only apparent cause of death, and in 12 (13%) a contributing cause of sudden unexpected natural death, while in 3 (3%) it was related to a fatal accidental injury. In 5 (13%) of the 39 cases of violent death, cardiomyopathy was an incidental finding without any connection to the circumstances or cause of death. The causes of cardiovascular abnormalities were associated with bacterial and viral infections, respiratory disorders, phenytoin sensitivity, or were unknown. Because of the differences in diagnostic criteria employed by previous investigators, it cannot be determined whether the incidence of the cardiovascular abnormalities and sudden cardiac death in children found in this material was higher than in other studies.


Natural Killer cell activation, reduced ACE2, TMPRSS2, cytokines G-CSF, M-CSF and SARS-CoV-2-S pseudovirus infectivity by MEK inhibitor treatment of human cells.
COVID-19 affects vulnerable populations including elderly individuals and patients with cancer. Natural Killer (NK) cells and innate-immune TRAIL suppress transformed and virally-infected cells. ACE2, and TMPRSS2 protease promote SARS-CoV-2 infectivity, while inflammatory cytokines IL-6, or G-CSF worsen COVID-19 severity. We show MEK inhibitors (MEKi) VS-6766, trametinib and selumetinib reduce ACE2 expression in human cells. In some human cells, remdesivir increases ACE2-promoter luciferase-reporter expression, ACE2 mRNA and protein, and ACE2 expression is attenuated by MEKi. In serum-deprived and stimulated cells treated with remdesivir and MEKi we observed correlations between pRB, pERK, and ACE2 expression further supporting role of proliferative state and MAPK pathway in ACE2 regulation. We show elevated cytokines in COVID-19-(+) patient plasma (N=9) versus control (N=11). TMPRSS2, inflammatory cytokines G-CSF, M-CSF, IL-1α, IL-6 and MCP-1 are suppressed by MEKi alone or with remdesivir. We observed MEKi stimulation of NK-cell killing of target-cells, without suppressing TRAIL-mediated cytotoxicity. Pseudotyped SARS-CoV-2 virus with a lentiviral core and SARS-CoV-2 D614 or G614 SPIKE (S) protein on its envelope infected human bronchial epithelial cells, small airway epithelial cells, or lung cancer cells and MEKi suppressed infectivity of the pseudovirus. We show a drug class-effect with MEKi to stimulate NK cells, inhibit inflammatory cytokines and block host-factors for SARS-CoV-2 infection leading also to suppression of SARS-CoV-2-S pseudovirus infection of human cells. MEKi may attenuate SARS-CoV-2 infection to allow immune responses and antiviral agents to control disease progression.


Peptide-based isolation of circulating tumor cells by magnetic nanoparticles.
Detection of rare circulating tumor cells (CTCs) in the peripheral blood of metastatic cancer patients has shown promise for improved diagnosis, staging and prognosis of cancers. The epithelial cell adhesion molecule (EpCAM) has been revealed to be over-expressed in CTCs while it is absent in normal blood cells and has been used as an efficient diagnosis and therapeutic target on CTCs, especially in CTC isolation and detection. Most of the CTC isolation techniques are based on nanomaterials or nanostructured surfaces functionalized with the EpCAM antibody. Herein, instead of anti-EpCAM, we report a new CTC isolation method with high efficiency by using the EpCAM recognition peptide functionalized iron oxide magnetic nanoparticles (MNPs) (Pep@MNPs). The de novo designed peptide, Pep10, with comparable binding affinity KD (1.98 × 10-9 mol L-1) to that of the anti-EpCAM (2.69 × 10-10 mol L-1) is attached onto MNPs via biotin-avidin interaction. We demonstrate that Pep10@MNPs (200 nm) have the comparable capture efficiency (reaching above 90%) and purity (reaching above 93%) to anti-EpCAM@MNPs for breast, prostate and liver cancers from spiked human blood. Furthermore, the captured cells still maintain viability for further molecular biological analysis with this method. The peptide-based CTC isolation method could be beneficial for cancer prognosis and metastasis prevention by increasing the stability and reproducibility.


IPSPs strongly inhibit climbing fiber-activated [Ca2+]i increases in the dendrites of cerebellar Purkinje neurons.
The interaction between the excitatory climbing fiber (CF) response and stellate cell inhibition was studied in guinea pig Purkinje cells in sagittal slices from the cerebellar vermis. Sharp microelectrode recordings from the soma or dendrites were combined with high-speed fluorescence imaging of intracellularly injected fura-2. In this way both the electrical responses and the associated [Ca2+]i changes could be monitored at the same time. Usually simultaneously activated inhibition caused almost no change to the somatically recorded CF response. However, the inhibition caused a strong reduction in the CF-associated [Ca2+]i increase which normally was widespread in the dendrites. This effect was graded; stronger inhibition caused a larger and more widespread reduction in the [Ca2+]i change that was greatest in the more distal dendrites. Sometimes the reduction was over 90% in the distal dendrites and occasionally it was localized to only a single dendritic branch. Both the inhibitory postsynaptic potential (IPSP) and the associated reduction in the CF-induced [Ca2+]i change were blocked by bicuculline, a GABAA receptor antagonist. Dendritic recordings showed that each CF response evoked a 2-3 msec wide action potential. The amplitude of this action potential was reduced in a graded manner by the IPSP in parallel with the reduction in the [Ca2+]i change. Varying the time between the activation of the IPSP and the CF response showed that both the reduction in the [Ca2+]i change and the action potential amplitude occurred in a narrow time window of about 8-10 msec, about the rise time of the IPSP. Together these results indicate that the CF response activates a fast dendritic Ca2+ spike that causes most of the [Ca2+]i increase, both of which can be blocked by an inhibitory shunting conductance. This interaction provides a means whereby Ca(2+)-dependent dendritic mechanisms can be modulated without affecting the immediate output of the Purkinje cell.


Quaternary ammonium block of mutant Na+ channels lacking inactivation: features of a transition-intermediate mechanism.
1. The quaternary ammonium (QA) lidocaine derivative QX-314 (2-(triethylamino)-N-(2,6-dimethylphenyl)-acetamide) induces internal pore blockade of single cardiac Na+ channels enzymatically modified (papain) to eliminate fast inactivation. The mechanism involves dual, interacting blocking modes (rapid and discrete) with binding domains deep in the pore from the cytoplasmic mouth, and where the rapid blocked configuration serves as a transition-intermediate for the development of discrete block. The primary goals of this study were to test for this mechanism in a recombinant Na+ channel genetically engineered to selectively lack fast inactivation, and if present, to explore the underlying structural features. 2. Fast inactivation was removed in rat skeletal muscle mu1 Na+ channels (RSkM1) with an IFM-QQQ mutation in the cytoplasmic III-IV interdomain (QQQ). QQQ was expressed in Xenopus oocytes and single-channel activity was studied in cell-free, inside-out membrane patches. Application of QX-314 (QX, 0-4 mM) to the cytoplasmic membrane surface caused two distinct modalities of single-channel blockade: reduction of unitary current and interruptions of current lasting tens of milliseconds. These are consistent with rapid and discrete pore block, respectively. The voltage and concentration dependence of block indicates that the modes interact and have binding sites that share a deep location in the pore, at approximately 65 % of the membrane electric field in from the cytoplasmic mouth. 3. Mutation of phenylalanine (F1579) in domain IV-S6, critical in local anaesthetic block, to alanine in QQQ (QQQ-F1579A) disabled discrete block but notably failed to alter rapid block, single-channel gating and slope conductance. 4. Amplitude distribution analysis was applied to long bursts (> 50 ms) of QQQ-F1579A activity to investigate the kinetics of rapid block. Computed rapid blocking and unblocking rate constants are 42 000 +/- 18 000 m-1 ms-1 and 82 +/- 22 ms-1, respectively (n = 3, -20 mV). 5. The results support a general transition-intermediate mechanism that governs internal QX and local anaesthetic pore block of voltage-gated Na+ channels and provide insight into underlying structural features.


Dry eye-induced CCR7+CD11b+ cell lymph node homing is induced by COX-2 activities.
We aimed to determine the role of CCR7+CD11b+ cell lymph node (LN) homing and T-cell differentiation in dry eye (DE)-induced immunopathogenesis and investigate the therapeutic effects of cyclooxygenase-2 (COX-2) and prostaglandin E2/eicosanoid-prostanoid (PGE2/EP) inhibitors against DE. Six-week-old female C57BL/6 mice were housed in a controlled-environment chamber and administered topical selective COX-2 inhibitors or EP2 antagonists. Expression of major histocompatibility complex (MHC)-IIhigh, CD11b+, CCR7+, IFN-γ+, IL-17+, and CD4+ in the corneas and draining LNs was evaluated using flow cytometry. Mixed lymphocyte reactions (MLRs) with carboxyfluorescein diacetate succinimidyl ester labeling and intracellular cytokine staining were used to verify DE-induced corneal dendritic cell function. mRNA expression of COX-2, EPs, and proinflammatory cytokines in ocular surface was evaluated using quantitative RT-PCR and immunohistochemical staining. Dry eye significantly increased MHC-IIhighCD11b+ and CCR7+CD11b+ cells in the cornea and LNs, and MLR revealed CCR7+CD11b+ cells from DE corneas stimulated IL-17+CD4+ cell proliferation. mRNA levels of COX-2, EP2, IFN-γ, TNF-α, IL-6, and IL-17 were significantly higher in DE ocular surface but were suppressed by topical COX-2 inhibitors and EP2-specific blockers. Immunohistochemical staining showed COX-2 and matrix metalloproteinase expression in DE corneal epithelia that was diminished by both topical treatments. Furthermore, both topical treatments significantly reduced frequencies of MHC-IIhigh, CD11b+, and CCR7+CD11b+ cells in the corneas and LNs, but also IL-17+CD4+ cells in LNs. Topical COX-2/EP2 treatment reduces CCR7+CD11b+ cells on the ocular surface with inhibition of cellular LN homing and suppresses Th17 immune response, suggesting the COX-2/PGE2/EP axis contributes to immuno-inflammatory pathogenesis on the ocular surface and may be a novel therapeutic target in DE.


Amygdala response to preattentive masked fear in children with conduct problems: the role of callous-unemotional traits.
In children with conduct problems, high levels of callous-unemotional traits are associated with amygdala hypoactivity to consciously perceived fear, while low levels of callous-unemotional traits may be associated with amygdala hyperactivity. Behavioral data suggest that fear processing deficits in children with high callous-unemotional traits may extend to stimuli presented below conscious awareness (preattentively). The authors investigated the neural basis of this effect. Amygdala involvement was predicted on the basis of its role in preattentive affective processing in healthy adults and its dysfunction in previous studies of conduct problems. Functional MRI was used to measure neural responses to fearful and calm faces presented preattentively (for 17 ms followed by backward masking) in boys with conduct problems and high callous-unemotional traits (N=15), conduct problems and low callous-unemotional traits (N=15), and typically developing comparison boys (N=16). Amygdala response to fearful and calm faces was predicted to differentiate groups, with the greatest response in boys with conduct problems and low callous-unemotional traits and the lowest in boys with conduct problems and high callous-unemotional traits. In the right amygdala, a greater amygdala response was seen in boys with conduct problems and low callous-unemotional traits than in those with high callous-unemotional traits. The findings were not explained by symptom levels of conduct disorder, attention-deficit hyperactivity disorder, anxiety, or depression. These data demonstrate differential amygdala activity to preattentively presented fear in children with conduct problems grouped by callous-unemotional traits, with high levels associated with lower amygdala reactivity. The study's findings complement increasing evidence suggesting that callous-unemotional traits are an important specifier in the classification of children with conduct problems.


Testing the link between population genetic differentiation and clade diversification in Costa Rican orchids.
Species population genetics could be an important factor explaining variation in clade species richness. Here, we use newly generated amplified fragment length polymorphism (AFLP) data to test whether five pairs of sister clades of Costa Rican orchids that differ greatly in species richness also differ in average neutral genetic differentiation within species, expecting that if the strength of processes promoting differentiation within species is phylogenetically heritable, then clades with greater genetic differentiation should diversify more. Contrary to expectation, neutral genetic differentiation does not correlate directly with total diversification in the clades studied. Neutral genetic differentiation varies greatly among species and shows no heritability within clades. Half of the variation in neutral genetic differentiation among populations can be explained by ecological variables, and species-level traits explain the most variation. Unexpectedly, we find no isolation by distance in any species, but genetic differentiation is greater between populations occupying different niches. This pattern corresponds with those observed for microscopic eukaryotes and could reflect effective widespread dispersal of tiny and numerous orchid seeds. Although not providing a definitive answer to whether population genetics processes affect clade diversification, this work highlights the potential for addressing new macroevolutionary questions using a comparative population genetic approach.


Oxidative Stress Facilitates IFN-γ-Induced Mimic Extracellular Trap Cell Death in A549 Lung Epithelial Cancer Cells.
We previously demonstrated that IFN-γ induces an autophagy-regulated mimic extracellular trap cell death (ETosis) in A549 human lung cancer cells. Regarding reactive oxygen species (ROS) are involved in ETosis, this study investigated the role of oxidative stress. After IFN-γ stimulation, a necrosis-like cell death mimic ETosis occurred accompanied by the inhibition of cell growth, aberrant nuclear staining, and nucleosome release. ROS were generated in a time-dependent manner with an increase in NADPH oxidase component protein expression. STAT1-mediated IFN regulatory factor-1 activation was essential for upregulating ROS production. By genetically silencing p47phox, IFN-γ-induced ROS and mimic ETosis were significantly attenuated. This mechanistic study indicated that ROS may mediate DNA damage followed by histone H3 citrullination. Furthermore, ROS promoted IFN-γ-induced mimic ETosis in cooperation with autophagy. These findings further demonstrate that ROS regulates IFN-γ-induced mimic ETosis in lung epithelial malignancy.


O-GlcNAc modification blocks the aggregation and toxicity of the protein α-synuclein associated with Parkinson's disease.
Several aggregation-prone proteins associated with neurodegenerative diseases can be modified by O-linked N-acetyl-glucosamine (O-GlcNAc) in vivo. One of these proteins, α-synuclein, is a toxic aggregating protein associated with synucleinopathies, including Parkinson's disease. However, the effect of O-GlcNAcylation on α-synuclein is not clear. Here, we use synthetic protein chemistry to generate both unmodified α-synuclein and α-synuclein bearing a site-specific O-GlcNAc modification at the physiologically relevant threonine residue 72. We show that this single modification has a notable and substoichiometric inhibitory effect on α-synuclein aggregation, while not affecting the membrane binding or bending properties of α-synuclein. O-GlcNAcylation is also shown to affect the phosphorylation of α-synuclein in vitro and block the toxicity of α-synuclein that was exogenously added to cells in culture. These results suggest that increasing O-GlcNAcylation may slow the progression of synucleinopathies and further support a general function for O-GlcNAc in preventing protein aggregation.


Beta blocker use in the treatment of community hospital patients discharged after myocardial infarction.
To explore the reasons for underutilization of beta blocker treatment after acute myocardial infarction. A retrospective chart review. Two large community hospitals in Milwaukee, Wisconsin. All subjects (n = 694) discharged alive from July 1, 1990, to June 30, 1991, who had a diagnosis of acute myocardial infarction were eligible. Of these, 250 had missing data, resulting in a final sample of 444. Twenty-nine percent of the 444 patients were prescribed beta blocker therapy on discharge. Characteristics of the patients and their treatment associated with receipt of beta blocker therapy were identified with a logistic regression model. The adjusted odds ratios were 0.52 for female gender, 0.34 for no health insurance, 0.21 for chronic obstructive pulmonary disease, 0.46 for congestive heart failure, 0.28 for atrioventricular block, 1.86 for hypertension, 1.93 for chest pain during acute myocardial infarction, and 4.65 for prehospital beta blocker use. Prescription of beta blocker therapy was also influenced by receipt of other treatment modalities. The adjusted odds ratios were 0.23 for receipt of beta blocker therapy associated with myocardial revascularization, 0.18 for prescription on discharge of calcium channel blockers, and 0.22 for receipt of angiotensin-converting enzyme inhibitors. A minority of patients discharged after acute myocardial infarction receive beta blocker therapy, and women are only half as likely as men to receive it, after controlling for other factors. Though there are no data relating to whether calcium channel blockers or angiotensin-converting enzyme inhibitors lessen the protective effect of beta blocker therapy post-acute myocardial infarction, it would appear that these agents are frequently being used in lieu of beta blocker therapy for post-acute myocardial infarction patients.


Hair cortisol levels track phylogenetic and age related differences in hypothalamic-pituitary-adrenal (HPA) axis activity in non-human primates.
Hair has been shown to archive a uniquely time averaged signal of endocrine activity, and holds attractive advantages for both laboratory and field research. Prior research has explored the potential of hair hormone analysis to examine hormone-behavior relationships. To date, no research has focused on the potential of the technique to investigate age-related changes or taxon differences in endocrine function. It is known that non-human primate infants of many taxa exhibit high cortisol levels after parturition, which rapidly decline with age. It has also been shown that hypercortisolism generally characterizes platyrrhine (New World monkey) endocrine function. These endocrine trends have been characterized using cortisol levels determined from serum, plasma, and feces. Here we test whether cortisol levels determined from hair recover similar phylogenetic and age related patterns in endocrine function in non-human primates. In order to test whether hair cortisol reflect infant hypercortisolism with significant age-related decline, hair cortisol levels are measured in samples from wild vervet monkeys (Chlorocebus aethiops) and captive Guinea baboons (Papio hamadryas papio), ranging in age from infants through juveniles. Further, in order to test whether platyrrhines exhibit significantly higher hair cortisol levels compared to strepsirrhines and catarrhines, and therefore faithfully recover similar signals as more traditionally used substrates (e.g. serum), hair cortisol levels are quantified in adult female hair samples collected from a broad range of non-human primate taxa. Results confirm that hair cortisol levels accurately reflect known phylogenetic and age related patterns of circulating cortisol levels. Therefore, these results suggest that hair may be an ideal hormone bearing substrate for research focused on the examination of population endocrine profiles, cross-sectional studies of endocrine function and taxon variation in hormone levels, as well as stable behavioral trends.


Synergistic Anticancer Effect of a Combination of Paclitaxel and 5-Demethylnobiletin Against Lung Cancer Cell Line In Vitro and In Vivo.
Lung cancer remains a highly prevalent disease and a leading cause of cancer-related deaths worldwide. Currently, exploring antitumor drugs derived from herbs used in traditional Chinese medicine is increasingly becoming an attractive area of research. Paclitaxel (PTX), a highly effective chemotherapeutic drug, is widely used for treating different cancers; however, the clinical use of PTX is dose limited because of its adverse side effects. Chemotherapeutic agents are being developed to enhance the anticancer activity of PTX, particularly for use in combination therapy. 5-Demethylnobiletin (5-DMN), a natural, active compound isolated from orange peel, has been reported to induce apoptosis in several cancer cell lines. In this study, we tested the synergistic anticancer antiproliferative effects of combinations of PTX and 5-DMN on CL1-5 lung cancer cells through the MTT and propidium iodide assays. After low-dose combination treatments (PTX and 5-DMN), a reduction in cell viability and a concomitant increase in apoptosis were observed in the CL1-5 cells. We propose that 5-DMN cooperates with PTX to induce apoptosis via the caspase pathway (by modulating caspase-3, caspase-8, and caspase-9 activities). Furthermore, we observed that the combination treatment significantly suppressed tumor growth in the nude mouse xenograft model. The results suggest that the synergistic effects of PTX and 5-DMN in lung cancer cells deserve particular attention and indicate the possibility of developing additional new strategies for treating lung cancer.


The detection and recovery of Erysipelothrix spp. in meat and abattoir samples in Western Australia.
To investigate the occurrence of Erysipelothrix rhusiopathiae and other Erysipelothrix spp. in abattoir and meat samples in Western Australia. Samples were collected from various parts of pig and sheep carcasses, as well as different sections of slaughtering line, pen soil and effluent. Previously evaluated culture methods were applied for the isolation of Erysipelothrix spp., in conjunction with phenotypic and genotypic detection and identification procedures. Of 109 samples from the two abattoirs, 35 (32.1%) were Erysipelothrix genus-specific PCR-positive. These came from swabs of animal exterior surfaces and joints, slaughtering areas, pig pen soil and abattoir effluent. Four samples (3.7%) from sheep arthritic joints and pig abattoir effluent were also E. rhusiopathiae species-specific PCR-positive. Of 123 carcass washing samples, 12 (9.8%) were genus-specific PCR-positive, and these came from all five kinds of meat samples tested, including beef, lamb, mutton, pork and chicken. Four of them (3.3%) were also species-specific PCR-positive. A total of 25 isolates was recovered from the samples, of which seven were identified as E. rhusiopathiae, seven were consistent with E. tonsillarum, and the remaining 11 were other species of Erysipelothrix. Erysipelothrix spp. can still be isolated and identified from specimens of animal origin with relative ease, provided that appropriate cultural and molecular procedures are used. Clinical microbiology laboratories may need to improve their diagnostic protocols. This study confirms that E. rhusiopathiae and other species of Erysipelothrix continue to colonize and contaminate farmed animals and animal products. Erysipelothrix infection still poses a potential threat to the economy of the farmed animal industry, as well as being a potential human public health hazard.


Metformin and Micronutrient Status in Type 2 Diabetes: Does Polypharmacy Involving Acid-Suppressing Medications Affect Vitamin B12 Levels?
Metformin is the first-choice drug in uncomplicated type 2 diabetes (T2DM) and is effective in improving glycaemic control. It is the most widely prescribed oral antidiabetic medicine and has a good safety profile. However, there is an abundance of evidence that metformin use is associated with decreased Vitamin B12 status, though the clinical implications of this in terms of increased risk of diabetic peripheral neuropathy are debated. There is growing evidence that other B vitamins, vitamin D and magnesium may also be impacted by metformin use in addition to alterations to the composition of the microbiome, depending on the dose and duration of therapy. Patients using metformin for prolonged periods may, therefore, need initial screening with intermittent follow-up, particularly since vitamin B12 deficiency has similar symptoms to diabetic neuropathy which itself affects 40-50% of patients with T2DM at some stage. Among patients with T2DM, 40% are reported to experience symptomatic gastroesophageal reflux disease (GORD), of whom 70% use oral antidiabetic medications. The most common medications used to treat GORD are proton pump inhibitors (PPIs) and antagonists of histamine selective H2 receptors (H2RAs), both of which independently affect vitamin B12 and magnesium status. Research indicates that co-prescribing metformin with either PPIs or H2RAs can have further deleterious effects on vitamin B12 status. Vitamin B12 deficiency related to metformin and polypharmacy is likely to contribute to the symptoms of diabetic neuropathy which may frequently be under-recognised. This review explores current knowledge surrounding these issues and suggests treatment strategies such as supplementation.


Predicting Anthracycline Benefit: TOP2A and CEP17-Not Only but Also.
Evidence supporting the clinical utility of predictive biomarkers of anthracycline activity is weak, with a recent meta-analysis failing to provide strong evidence for either HER2 or TOP2A. Having previously shown that duplication of chromosome 17 pericentromeric alpha satellite as measured with a centromere enumeration probe (CEP17) predicted sensitivity to anthracyclines, we report here an individual patient-level pooled analysis of data from five trials comparing anthracycline-based chemotherapy with CMF (cyclophosphamide, methotrexate, and fluorouracil) as adjuvant chemotherapy for early breast cancer. Fluorescent in situ hybridization for CEP17, HER2, and TOP2A was performed in three laboratories on samples from 3,846 of 4,864 eligible patients from five trials evaluating anthracycline-containing chemotherapy versus CMF. Methodologic differences did not affect HER2-to-CEP17 ratios but necessitated different definitions for CEP17 duplication: > 1.86 observed copies per cell for BR9601, NEAT, Belgian, and DBCG89D trials and > 2.25 for the MA.5 trial. Fluorescent in situ hybridization data were available in 89.3% (HER2), 83.9% (CEP17), and 80.6% (TOP2A) of 3,846 patient cases with available tissue. Both CEP17and TOP2A treatment-by-marker interactions remained significant in adjusted analyses for recurrence-free and overall survival, whereas HER2 did not. A combined CEP17 and TOP2A-adjusted model predicted anthracycline benefit across all five trials for both recurrence-free (hazard ratio, 0.64; 95% CI, 0.51 to 0.82; P = .001) and overall survival (hazard ratio, 0.66; 95% CI, 0.51 to 0.85; P = .005). This prospectively planned individual-patient pooled analysis of patient cases from five adjuvant trials confirms that patients whose tumors harbor either CEP17 duplication or TOP2A aberrations, but not HER2 amplification, benefit from adjuvant anthracycline chemotherapy.


Clinically significant cognitive dysfunction in OEF/OIF/OND veterans: Prevalence and clinical associations.
Cognitive performance in trauma-exposed populations, such as combat Veterans, has been shown to be worse than in nonexposed peers. However, cognitive performance has typically been within the normal range (within 1 SD of normative mean), and the prevalence of clinically significant cognitive dysfunction (i.e., performance more than 1 SD below the mean on multiple measures in a domain) in younger adults with trauma exposure remains unknown. The objective of our study was to measure this. We applied Diagnostic and Statistical Manual of Mental Disorders (5th ed.; DSM-5) cutoffs for clinically significant cognitive dysfunction (>1 SD below the mean in multiple measures within a domain) in the domains of memory, executive function, and attention to a sample of combat-exposed Operation Enduring Freedom/Operation Iraqi Freedom/Operation New Dawn (OEF/OIF/OND; N = 368, mean age = 31.7 years, 90% men) Veterans. We then compared psychiatric, physiological, and neural measures, as well as functional outcomes, between those with and without cognitive dysfunction. Veterans with cognitive dysfunction (n = 129, 35.1%) had lower premorbid reading ability and more severe psychological distress, including increased anxiety, depression, posttraumatic stress disorder (PTSD), sleep difficulties, pain, and alcohol consumption. Those with cognitive dysfunction also had worse functional outcomes, with mild but significant disability. In contrast, we found associations between outcome and age, traumatic brain injury, physiological and neural measures to be weak or not significant. Together, this suggests that premorbid abilities and trauma-related psychological symptoms contribute significantly to cognitive dysfunction in OEF/OIF/OND Veterans, and that neurological insult and aging may play less of a role. Cognitive dysfunction may be at least partially ameliorated by treating trauma-related symptoms. (PsycINFO Database Record (c) 2019 APA, all rights reserved).


Sources of off-target effects of vagus nerve stimulation using the helical clinical lead in domestic pigs.
Clinical data suggest that efficacious vagus nerve stimulation (VNS) is limited by side effects such as cough and dyspnea that have stimulation thresholds lower than those for therapeutic outcomes. VNS side effects are putatively caused by activation of nearby muscles within the neck, via direct muscle activation or activation of nerve fibers innervating those muscles. Our goal was to determine the thresholds at which various VNS-evoked effects occur in the domestic pig—an animal model with vagus anatomy similar to human—using the bipolar helical lead deployed clinically. Intrafascicular electrodes were placed within the vagus nerve to record electroneurographic (ENG) responses, and needle electrodes were placed in the vagal-innervated neck muscles to record electromyographic (EMG) responses. Contraction of the cricoarytenoid muscle occurred at low amplitudes (~0.3 mA) and resulted from activation of motor nerve fibers in the cervical vagus trunk within the electrode cuff which bifurcate into the recurrent laryngeal branch of the vagus. At higher amplitudes (~1.4 mA), contraction of the cricoarytenoid and cricothyroid muscles was generated by current leakage outside the cuff to activate motor nerve fibers running within the nearby superior laryngeal branch of the vagus. Activation of these muscles generated artifacts in the ENG recordings that may be mistaken for compound action potentials representing slowly conducting Aδ-, B-, and C-fibers. Our data resolve conflicting reports of the stimulation amplitudes required for C-fiber activation in large animal studies (>10 mA) and human studies (<250 μA). After removing muscle-generated artifacts, ENG signals with post-stimulus latencies consistent with Aδ- and B-fibers occurred in only a small subset of animals, and these signals had similar thresholds to those that caused bradycardia. By identifying specific neuroanatomical pathways that cause off-target effects and characterizing the stimulation dose-response curves for on- and off-target effects, we hope to guide interpretation and optimization of clinical VNS.


Cytogenetic studies of a human medullary thyroid carcinoma cell line.
Detailed karyotypic analyses were performed on early and late passages of the TT cell line derived from malignant cells of a patient with the sporadic form of medullary thyroid carcinoma. Most of the cells examined were hypodiploid with a modal chromosome number of 43. The cells have a complex karyotype with ten rearranged chromosomes found in early passages and 12 rearranged chromosomes in late passages. The karyotypic pattern was relatively stable with continued in vitro culture, and consistent alterations involved chromosomes X, #1, #3, #5, #7, #8, #9, #10, #11, #12, and #14. Rearrangements of chromosome #11 are of particular interest, because the gene for calcitonin, the polypeptide hormone marker for medullary thyroid carcinoma resides on this chromosome. In early passages, one #11 contained an insertion within band q13, and this abnormality was retained in late passages. In late passage cells the other #11 homolog developed a deletion of part of the short arm, which involves the calcitonin gene region. We were unable to detect any deletion of chromosome band 20p12, previously reported by other investigators to occur in the germ line of patients with the hereditary form of medullary thyroid carcinoma.


Comparison of surgical and nonsurgical treatment of bilateral condylar fractures based on maximal mouth opening.
This study presents a comparative analysis of the open surgical and nonsurgical treatment of patients with bilateral condylar fractures. Sixty-seven (67) patients were treated, and the completed data on 55 patients were reviewed to compare both therapeutic modalities, which consisted of nonsurgical and surgical treatment in 37 and 18 patients, respectively. In the nonsurgical group, 23 patients (23/37, 62%) had normal mouth opening. Functional success rate was 79% (15/19) and 44% (8/18) in young adult patients (-29yrs) and older patients (30+yrs), respectively, and there was a significant difference of outcome between the two groups. In nonsurgically treated young patients with disorders, bilateral dislocation and existence of concomitant mandibular fractures were commonly observed. In the open surgical group, seven (7/11, 64%) young adult and three (3/7, 43%) older patients gained normal mouth opening, and no significant difference was observed. Additionally, there was no difference between non-surgical and surgical treatment in any category. Patients undergoing rigid fixation benefited from restoring maximum mouth opening, although there was no significant difference between the rigid and non-rigid fixation groups. Based on these findings, nonrigid fixation should be avoided, and rigid fixation might improve outcome in young adult patients with severe fracture pattern, such as bilateral dislocation and concomitant mandibular fracture.


The Surveillance for Enteric Fever in Asia Project (SEAP), Severe Typhoid Fever Surveillance in Africa (SETA), Surveillance of Enteric Fever in India (SEFI), and Strategic Typhoid Alliance Across Africa and Asia (STRATAA) Population-based Enteric Fever Studies: A Review of Methodological Similarities and Differences.
Building on previous multicountry surveillance studies of typhoid and others salmonelloses such as the Diseases of the Most Impoverished program and the Typhoid Surveillance in Africa Project, several ongoing blood culture surveillance studies are generating important data about incidence, severity, transmission, and clinical features of invasive Salmonella infections in sub-Saharan Africa and South Asia. These studies are also characterizing drug resistance patterns in their respective study sites. Each study answers a different set of research questions and employs slightly different methodologies, and the geographies under surveillance differ in size, population density, physician practices, access to healthcare facilities, and access to microbiologically safe water and improved sanitation. These differences in part reflect the heterogeneity of the epidemiology of invasive salmonellosis globally, and thus enable generation of data that are useful to policymakers in decision-making for the introduction of typhoid conjugate vaccines (TCVs). Moreover, each study is evaluating the large-scale deployment of TCVs, and may ultimately be used to assess post-introduction vaccine impact. The data generated by these studies will also be used to refine global disease burden estimates. It is important to ensure that lessons learned from these studies not only inform vaccination policy, but also are incorporated into sustainable, low-cost, integrated vaccine-preventable disease surveillance systems.


A comparison of knowledge and attitude toward mental illness among secondary school students and teachers.
The current study aimed to assess and compare the level of knowledge and attitude towards mental illness between secondary school students and their teachers in Oman. An online survey was carried out to collect data from 400 students and 411 teachers about their knowledge and attitudes toward people with mental illness. Two independent case studies about depression and schizophrenia were also tested. Students have a poor knowledge of mental illness compared with their teachers, as more than half scored less than 60% compared with 16.5% of the teachers. More than two-thirds of the students (80%) and teachers (76.4%) have a low or minor positive attitude toward people with mental illness. The study identified significant differences in knowledge in favour of teachers, although the opposite was found regarding attitudes. Since students spend a significant amount of time in school, bridging the gap between teachers' and students' knowledge and attitudes toward mental illness is an essential part in enhancing the knowledge and attitudes of the students. In addition, knowledgeable teachers with positive attitude can assist in early identification of mental illnesses and help students when needed. In turn, students who possess knowledge and positive attitude toward mental illness can share their concerns with their teachers. In the presence of such accepting and cooperative environment, the stigma can be decreased and early detection of mental illness and help-seeking behaviour can be promoted.


Breast cancer risk in relation to type of estrogen contained in oral contraceptives.
We report analyses designed to address the recent hypothesis that women who use combination-type OCs containing ethinylestradiol (EE) are at increased risk of breast cancer before age 45, if use of such OCs occurs prior to first term pregnancy (FTP). Our findings, based on data from 1679 cases and 1689 controls, 20-44 years of age, from the population-based Cancer and Steroid Hormone Study, are against the hypothesis. The relative risk of breast cancer by duration of exclusive use prior to FTP of OCs containing EE is estimated to be 1.0 (1-12 months EE use), 1.2 (13-48 months EE use), and 0.9 (49+ months EE use). There was no evidence of a latent effect. Among parous women with 49+ months exclusive use prior to FTP of EE-containing OCs, the relative risk of breast cancer was estimated as 0.9 (0-4 years after FTP) and 0.6 (5-9 years after FTP). Among nulliparous women with 49+ months exclusive use of EE-containing OCs, the relative risk of breast cancer was estimated as 1.0 (0-4 years since first use), 0.7 (5-9 years since first use), and 1.1 (10-14 years since first use). With regard to exclusive use of OCs containing mestranol, parous women who used such preparations long-term before their FTP showed no alteration of breast cancer risk, even 15 years or more after pregnancy. Nulliparous women with exclusive use of ME-containing OCs did show elevations in breast cancer risk, but the magnitude of risk in relation to duration of use and latent interval shows no pattern that suggests a cause-effect relationship.


Vaccines against Ebola virus.
We have just witnessed the largest and most devastating outbreak of Ebola virus disease, which highlighted the urgent need for development of an efficacious vaccine that could be used to curtail future outbreaks. Prior to 2014, there had been limited impetus worldwide to develop a vaccine since the virus was first discovered in 1976. Though too many lives were lost during this outbreak, it resulted in the significantly accelerated clinical development of a number of candidate vaccines through an extraordinary collaborative global effort coordinated by the World Health Organisation (WHO) and involving a number of companies, trial centres, funders, global stakeholders and agencies. We have acquired substantial safety and immunogenicity data on a number of vaccines in Caucasian and African populations. The rapid pace of events led to the initiation of the landmark efficacy trial testing the rVSV-vectored vaccine, which showed high level efficacy in an outbreak setting when deployed using an innovative ring vaccination strategy. Though the Public Health Emergency of International Concern (PHEIC) declared by the WHO has now been lifted, the global scientific community faces numerous challenges ahead to ensure that there is a licensed, deployable vaccine available for use in future outbreaks for at least the Zaire and Sudan strains of Ebola virus. There remain several unanswered questions on the durability of protection, mechanistic immunological correlates and preferred deployment strategies. This review outlines a brief history of the development of Ebola vaccines, the significant progress made since the scale of the outbreak became apparent, some lessons learnt and how they could shape future development of vaccines and the management of similar outbreaks.


Ready for takeoff? A critical review of armodafinil and modafinil for the treatment of sleepiness associated with jet lag.
Jet lag syndrome (JLS) is a clinical syndrome of disrupted nocturnal sleep and daytime neurocognitive impairment which occurs in the context of rapid transmeridian travel. Many strategies for treatment of JLS exist, and include hypnotics to enhance nocturnal sleep, chronotherapeutic approaches (eg, light therapy, melatonin, or gradual schedule shifting), and alerting agents to counter daytime sleepiness. Safety concerns have prompted renewed interest in managing JLS-associated excessive daytime sleepiness (JLSAEDS). Off-label use of the newer alerting agents modafinil and armodafinil is increasing for this indication, often at the specific request of patients. In order to better evaluate the potential risks and benefits of these medications for the management of JLSAEDS, clinicians must be aware of what is known - and still not known. In this article, the pharmacology and pharmacokinetics of modafinil and armodafinil are reviewed, along with evidence for their efficacy in treating sleepiness associated with narcolepsy, obstructive sleep apnea and shift work sleep disorder. Clinical trial data for use of alerting agents in the management of JLSAEDS are limited to one three-day trial involving armodafinil, dosed in the morning to treat JLSAEDS in the setting of eastbound transmeridian travel. This study showed improvement in objective measures of daytime sleepiness at doses of 50 and 150 mg per day. However, global impression of clinical severity of symptom scores only improved on day 1 for those patients receiving 150 mg, and were otherwise not superior to placebo. Consideration for the use of modafinil or armodafinil for the treatment of sleepiness associated with JLS involves careful integration of patient-reported goals, a review of medical contraindications, and an awareness of rare adverse events. More research is needed in order to identify those who are most likely to benefit from this intervention and better define the risk-benefit ratio for this indication.


Expression of the AHPND Toxins PirAvp and PirBvp Is Regulated by Components of the Vibrio parahaemolyticus Quorum Sensing (QS) System.
Acute hepatopancreatic necrosis disease (AHPND) in shrimp is caused by Vibrio strains that harbor a pVA1-like plasmid containing the pirA and pirB genes. It is also known that the production of the PirA and PirB proteins, which are the key factors that drive the observed symptoms of AHPND, can be influenced by environmental conditions and that this leads to changes in the virulence of the bacteria. However, to our knowledge, the mechanisms involved in regulating the expression of the pirA/pirB genes have not previously been investigated. In this study, we show that in the AHPND-causing Vibrio parahaemolyticus 3HP strain, the pirAvp and pirBvp genes are highly expressed in the early log phase of the growth curve. Subsequently, the expression of the PirAvp and PirBvp proteins continues throughout the log phase. When we compared mutant strains with a deletion or substitution in two of the quorum sensing (QS) master regulators, luxO and/or opaR (luxOD47E, ΔopaR, ΔluxO, and ΔopaRΔluxO), our results suggested that expression of the pirAvp and pirBvp genes was related to the QS system, with luxO acting as a negative regulator of pirAvp and pirBvp without any mediation by opaRvp. In the promoter region of the pirAvp/pirBvp operon, we also identified a putative consensus binding site for the QS transcriptional regulator AphB. Real-time PCR further showed that aphBvp was negatively controlled by LuxOvp, and that its expression paralleled the expression patterns of pirAvp and pirBvp. An electrophoretic mobility shift assay (EMSA) showed that AphBvp could bind to this predicted region, even though another QS transcriptional regulator, AphAvp, could not. Taken together, these findings suggest that the QS system may regulate pirAvp/pirBvp expression through AphBvp.


Ventricular assist device support as a bridge to heart transplantation in patients with giant cell myocarditis.
Giant cell myocarditis (GCM) carries a poor prognosis and many patients require end-stage therapies. This study sought to determine the outcome of patients bridged with ventricular assist devices (VAD) to orthotopic heart transplantation (OHT). A retrospective data collection of all patients with GCM was performed. Diagnosis was determined by endomyocardial or explanted heart biopsy. Eight patients were found, but two of those patients went directly to OHT and were excluded. The remaining six patients received VADs, and these patients, aged 44 ± 18 years, were included. Five of the six patients were bridged with biventricular support and one patient was supported by left ventricular assist device (LVAD) alone. Two patients died on device support. Four patients were bridged to OHT 77 ± 42 days after device implantation. All four patients bridged with a VAD are alive, with a mean follow-up of 5.7 ± 4.1 years. Two patients were found to have recurrent GCM in the transplanted heart and were treated successfully with immunosuppression. Three patients had high grade (2R) rejection at 66 ± 52 days post-OHT. Cardiac function was preserved in all patients, and only one patient had cardiac allograft vasculopathy. Patients with end-stage GCM can be successfully bridged with VADs to OHT with very good post-OHT survival. The proper immunosuppressive regimen for this group needs further investigation given the frequency of rejection and GCM recurrence.


Structural and functional characterization of an orphan ATP-binding cassette ATPase involved in manganese utilization and tolerance in Leptospira spp.
Pathogenic Leptospira species are the etiological agents of the widespread zoonotic disease leptospirosis. Most organisms, including Leptospira, require divalent cations for proper growth, but because of their high reactivity, these metals are toxic at high concentrations. Therefore, bacteria have acquired strategies to maintain metal homeostasis, such as metal import and efflux. By screening Leptospira biflexa transposon mutants for their ability to use Mn(2+), we have identified a gene encoding a putative orphan ATP-binding cassette (ABC) ATPase of unknown function. Inactivation of this gene in both L. biflexa and L. interrogans strains led to mutants unable to grow in medium in which iron was replaced by Mn(2+), suggesting an involvement of this ABC ATPase in divalent cation uptake. A mutation in this ATPase-coding gene increased susceptibility to Mn(2+) toxicity. Recombinant ABC ATPase of the pathogen L. interrogans exhibited Mg(2+)-dependent ATPase activity involving a P-loop motif. The structure of this ATPase was solved from a crystal containing two monomers in the asymmetric unit. Each monomer adopted a canonical two-subdomain organization of the ABC ATPase fold with an α/β subdomain containing the Walker motifs and an α subdomain containing the ABC signature motif (LSSGE). The two monomers were arranged in a head-to-tail orientation, forming a V-shaped particle with all the conserved ABC motifs at the dimer interface, similar to functional ABC ATPases. These results provide the first structural and functional characterization of a leptospiral ABC ATPase.


Tanshinone IIA attenuates nerve structural and functional damage induced by nerve crush injury in rats.
After peripheral nerve crush injury, the fibers of distal nerve segments gradually disintegrate, and axons regrow from the proximal nerve segment, eventually reaching the target organ. However, the axon regeneration is generally not sufficient for the recovery of neurological function, so drug therapy is necessary. In the current study, we explored the effect of Tanshinone IIA in nerve regeneration in a sciatic nerve crush injury model using Sprague Dawley rats. The rats were administered 45 mg/kg of Tanshinone IIA once daily. Motor behavior and tibialis anterior muscle mass were assessed, and histological analysis of the sciatic nerve and lumbar spinal cord were conducted. The results showed that the administration of Tanshinone IIA improved nerve growth and motor function, and resulted in a marked decrease of neuronal death. The findings of this exploratory study suggest that Tanshinone IIA alleviates injury and boosts regeneration after nerve crush injury in a rat model of sciatic nerve injury.


The utility of childhood and adolescent obesity assessment in relation to adult health.
High childhood obesity prevalence has raised concerns about future adult health, generating calls for obesity screening of young children. To estimate how well childhood obesity predicts adult obesity and to forecast obesity-related health of future US adults. Longitudinal statistical analyses; microsimulations combining multiple data sets. National Longitudinal Survey of Youth, Population Study of Income Dynamics, and National Health and Nutrition Evaluation Surveys. The authors estimated test characteristics and predictive values of childhood body mass index to identify 2-, 5-, 10-, and 15 year-olds who will become obese adults. The authors constructed models relating childhood body mass index to obesity-related diseases through middle age stratified by sex and race. Twelve percent of 18-year-olds were obese. While screening at age 5 would miss 50% of those who become obese adults, screening at age 15 would miss 9%. The predictive value of obesity screening below age 10 was low even when maternal obesity was included as a predictor. Obesity at age 5 was a substantially worse predictor of health in middle age than was obesity at age 15. For example, the relative risk of developing diabetes as adults for obese white male 15-year-olds was 4.5 versus otherwise similar nonobese 15-year-olds. For obese 5-year-olds, the relative risk was 1.6. Main results do not include Hispanics due to sample size. Past relationships between childhood and adult obesity and health may change in the future. Early childhood obesity assessment adds limited information to later childhood assessment. Targeted later childhood approaches or universal strategies to prevent unhealthy weight gain should be considered.


Synthesis and Antiproliferative Activity of New Ruthenium Complexes with Ethacrynic-Acid-Modified Pyridine and Triphenylphosphine Ligands.
Pyridine- and phosphine-based ligands modified with ethacrynic acid (a broad acting glutathione transferase inhibitor) were prepared and coordinated to ruthenium(II)-arene complexes and to a ruthenium(III) NAMI-A type complex. All the compounds (ligands and complexes) were fully characterized by analytical and spectroscopic methods and, in one case, by single-crystal X-ray diffraction. The in vitro anticancer activity of the compounds was studied, with the compounds displaying moderate cytotoxicity toward the human ovarian cancer cell lines. All the complexes led to similar levels of residual GST activity in the different cell lines, irrespective of the stability of the Ru-ligand bond.


Brief Academic Review and Clinical Practice Guidelines for Pediatric Atopic Dermatitis.
In this clinical guidelines article, we first include a brief review of the epidemiology, pathogenesis, clinical diagnoses, and scoring-scales for pediatric atopic dermatitis (AD). We then offer a set of pharmacologic treatment guidelines for infants and toddlers (<2 years), children (2-12 years), and adolescents (>12 years). We recommend irritant avoidance and liberal emollient usage as the cornerstone of treatment in all age-groups. In infants <2 years, we recommend topical corticosteroids as first-line medication-based therapy. In infants as young as 3 months, pimecrolimus, a topical calcineurin inhibitor, may also be used. As a last resort in patients <2 years, non-traditional therapies, such as the Aron regime, may be a safer option for refractory or resistant AD before off- label medications are considered. In children and adolescents >2 years, topical corticosteroids are still considered first-line therapies, but there is sufficient safety data to utilize topical calcineurin inhibitors and topical PDE4 inhibitors as well. In children ages 2-12 years whose atopic dermatitis fails to respond to prior treatments, oral systemic immunosuppressants can be used. For adolescents >12, the biologic, dupilumab, is an additional therapeutic option. A trial of phototherapy may also be utilized in children, particularly in adolescents >12 years, if they have access to treatment. Although not currently approved for the treatment of AD, Janus-kinase (JAK) inhibitors represent a promising new class of biologics with recently completed phase III clinical trials (JADE-- MONO1/2).


Exploring the catalytic mechanism of dihydropteroate synthase: elucidating the differences between the substrate and inhibitor.
Dihydropteroate synthase (DHPS) catalyzes the condensation of 6-hydroxymethyl-7,8-dihydropterin pyrophosphate (DHPPP) with p-aminobenzoic acid (pABA) and is a well validated target for anti-malarial and anti-bacterial drugs. However, in recent years its utility as a therapeutic target has diminished considerably due to multiple mutations. As such, considerable structural biology and medicinal chemistry effort has been expended to understand and overcome this issue. To date no detailed computational analysis of the protein mechanism has been made despite the detailed crystal structures and multiple mechanistic proposals being made. In this study the mechanistic proposals for DHPS have been systematically investigated using a hybrid QM/MM method. We aimed to compare the energetics associated with SN1 and SN2 processes, whether the SN1 process involves a carbocation or neutral DHP intermediate, uncover the identity of the general base in the catalytic mechanism, and understand the differences in substrate vs. inhibitor reactivity. Our results suggest a reaction that follows an SN1 process with the rate determining step being C-O bond breaking to give a carbocation intermediate. Comparative studies on the inhibitor STZ confirm the experimental observations that it is also a DHPS substrate.


Cannabis and endocannabinoid modulators: Therapeutic promises and challenges.
The discovery that botanical cannabinoids such as delta-9 tetrahydrocannabinol exert some of their effect through binding specific cannabinoid receptor sites has led to the discovery of an endocannabinoid signaling system, which in turn has spurred research into the mechanisms of action and addiction potential of cannabis on the one hand, while opening the possibility of developing novel therapeutic agents on the other. This paper reviews current understanding of CB1, CB2, and other possible cannabinoid receptors, their arachidonic acid derived ligands (e.g. anandamide; 2 arachidonoyl glycerol), and their possible physiological roles. CB1 is heavily represented in the central nervous system, but is found in other tissues as well; CB2 tends to be localized to immune cells. Activation of the endocannabinoid system can result in enhanced or dampened activity in various neural circuits depending on their own state of activation. This suggests that one function of the endocannabinoid system may be to maintain steady state. The therapeutic action of botanical cannabis or of synthetic molecules that are agonists, antagonists, or which may otherwise modify endocannabinoid metabolism and activity indicates they may have promise as neuroprotectants, and may be of value in the treatment of certain types of pain, epilepsy, spasticity, eating disorders, inflammation, and possibly blood pressure control.


Viral cross talk: intracellular inactivation of the hepatitis B virus during an unrelated viral infection of the liver.
Hepatitis B virus (HBV) infection is thought to be controlled by virus-specific cytotoxic T lymphocytes (CTL). We have recently shown that HBV-specific CTL can abolish HBV replication noncytopathically in the liver of transgenic mice by secreting tumor necrosis factor alpha (TNF-alpha) and interferon gamma (IFN-gamma) after antigen recognition. We now demonstrate that hepatocellular HBV replication is also abolished noncytopathically during lymphocytic choriomeningitis virus (LCMV) infection, and we show that this process is mediated by TNF-alpha and IFN-alpha/beta produced by LCMV-infected hepatic macrophages. These results confirm the ability of these inflammatory cytokines to abolish HBV replication; they elucidate the mechanism likely to be responsible for clearance of HBV in chronically infected patients who become superinfected by other hepatotropic viruses; they suggest that pharmacological activation of intrahepatic macrophages may have therapeutic value in chronic HBV infection; and they raise the possibility that conceptually similar events may be operative in other viral infections as well.


Platelet-independent adhesion of calcium-loaded erythrocytes to von Willebrand factor.
Adhesion of erythrocytes to endothelial cells lining the vascular wall can cause vaso-occlusive events that impair blood flow which in turn may result in ischemia and tissue damage. Adhesion of erythrocytes to vascular endothelial cells has been described in multiple hemolytic disorders, especially in sickle cell disease, but the adhesion of normal erythrocytes to endothelial cells has hardly been described. It was shown that calcium-loaded erythrocytes can adhere to endothelial cells. Because sickle erythrocyte adhesion to ECs can be enhanced by ultra-large von Willebrand factor multimers, we investigated whether calcium loading of erythrocytes could promote binding to endothelial cells via ultra-large von Willebrand factor multimers. We used (immunofluorescent) live-cell imaging of washed erythrocytes perfused over primary endothelial cells at venular flow rate. Using this approach, we show that calcium-loaded erythrocytes strongly adhere to histamine-stimulated primary human endothelial cells. This adhesion is mediated by ultra-large von Willebrand factor multimers. Von Willebrand factor knockdown or ADAMTS13 cleavage abolished the binding of erythrocytes to activated endothelial cells under flow. Platelet depletion did not interfere with erythrocyte binding to von Willebrand factor. Our results reveal platelet-independent adhesion of calcium-loaded erythrocytes to endothelium-derived von Willebrand factor. Erythrocyte adhesion to von Willebrand factor may be particularly relevant for venous thrombosis, which is characterized by the formation of erythrocyte-rich thrombi.


Dynamic Computational Model of the Human Spinal Cord Connectome.
Connectomes abound, but few for the human spinal cord. Using anatomical data in the literature, we constructed a draft connectivity map of the human spinal cord connectome, providing a template for the many calibrations of specialized behavior to be overlaid on it and the basis for an initial computational model. A thorough literature review gleaned cell types, connectivity, and connection strength indications. Where human data were not available, we selected species that have been studied. Cadaveric spinal cord measurements, cross-sectional histology images, and cytoarchitectural data regarding cell size and density served as the starting point for estimating numbers of neurons. Simulations were run using neural circuitry simulation software. The model contains the neural circuitry in all ten Rexed laminae with intralaminar, interlaminar, and intersegmental connections, as well as ascending and descending brain connections and estimated neuron counts for various cell types in every lamina of all 31 segments. We noted the presence of highly interconnected complex networks exhibiting several orders of recurrence. The model was used to perform a detailed study of spinal cord stimulation for analgesia. This model is a starting point for workers to develop and test hypotheses across an array of biomedical applications focused on the spinal cord. Each such model requires additional calibrations to constrain its output to verifiable predictions. Future work will include simulating additional segments and expanding the research uses of the model.


Recent advances in the management of hepato-renal syndrome (HRS).
Hepato-renal syndrome (HRS) is a functional renal failure complicating end-stage liver disease. HRS is characterized by marked arterial vasodilation (mainly of the splanchnic bed) and severe renal vasoconstriction. HRS is classified into 2 types: type I HRS shows a rapid and progressive decline in renal function with a very poor prognosis (median survival of about 2 weeks); HRS type 2 has a more stable renal failure, with a median survival of about 6 months. The management of HRS is still a big challenge. The definitive therapy for HRS is liver transplantation (LT); however, the survival rate of HRS patients is poor, and important organ shortage exists. Various approaches have been used for HRS treatment including vasoconstrictor therapy. Recent evidence has shown that vasoconstrictor agents are effective and serve as a bridge to LT; the rationale for vasoconstrictors is to counteract the splanchnic arterial vasodilation and increase the effective arterial blood volume. Thus, renal perfusion and glomerular filtration rates improve. Terlipressin, a V1 vasopressin agonist, has been used frequently. A recent meta-analysis of clinical trials showed that the pooled rate of patients who reversed HRS after terlipressin therapy was 0.52 (95% CI, 0.42; 0.61; P = 0.0001, /2 = 24.6%). The pooled Odds Ratio (OR) for mortality rate in HRS patients who were not responders to terlipressin versus responders was 5.746 (95% CI, 1.5; 21.9; P = 0.0005). Prospective, controlled clinical trials are in progress to address the impact of vasoconstrictor use on survival in HRS patients. Alternative therapies such as transjugular intrahepatic portosystemic shunts (TIPS) and extracorporeal albumin dialysis (ECAD) have given encouraging results but experience is extremely limited.


The major symptom dimensions of obsessive-compulsive disorder are mediated by partially distinct neural systems.
Obsessive-compulsive disorder (OCD) is a clinically heterogeneous disorder characterized by multiple, temporally stable symptom dimensions. Preliminary functional neuroimaging studies suggest that these symptom dimensions may have distinct neural substrates. Whole-brain voxel-based morphometry was used to examine the common and distinct neuroanatomical (structural) substrates of the major symptom dimensions of OCD. First, we compared 55 medication-free patients with OCD and 50 age-matched healthy control subjects. Multiple regression analyses were then used to examine the relationship between global and regional grey matter (GM) and white matter (WM) volumes and symptom dimension scores within the patient group. OCD patients showed decreased GM volume in left lateral orbitofrontal (BA47), left inferior frontal (BA44/45), left dorsolateral prefrontal (BA9) and right medial prefrontal (BA10) cortices and decreased bilateral prefrontal WM volume. Scores on the 'symmetry/ordering' dimension were negatively correlated with 'global' GM and WM volumes. Scores on the 'contamination/washing' dimension were negatively correlated with 'regional' GM volume in bilateral caudate nucleus and WM volume in right parietal region. Scores on the 'harm/checking' dimension were negatively correlated with regional GM and WM volume in bilateral temporal lobes. Scores on the 'symmetry/ordering' dimension were negatively correlated with regional GM volume in right motor cortex, left insula and left parietal cortex and positively correlated with bilateral temporal GM and WM volume. The results remained significant after controlling for age, sex, educational level, overall illness severity, global WM and GM volumes and excluding patients with comorbid depression. The reported symptom dimension-specific GM and WM alterations support the hypothesis that OCD is an etiologically heterogeneous disorder, with both overlapping and distinct neural correlates across symptom dimensions. These results have clear implications for the current neuroanatomical model of OCD and call for a substantial revision of such model which takes into account the heterogeneity of the disorder.


Synthesis and Evaluation of Folate-Conjugated Phenanthraquinones for Tumor-Targeted Oxidative Chemotherapy.
Almost all cells are easily killed by exposure to potent oxidants. Indeed, major pathogen defense mechanisms in both animal and plant kingdoms involve production of an oxidative burst, where host defense cells show an invading pathogen with reactive oxygen species (ROS). Although cancer cells can be similarly killed by ROS, development of oxidant-producing chemotherapies has been limited by their inherent nonspecificity and potential toxicity to healthy cells. In this paper, we describe the targeting of an ROS-generating molecule selectively to tumor cells using folate as the tumor-targeting ligand. For this purpose, we exploit the ability of 9,10-phenanthraquinone (PHQ) to enhance the continuous generation of H2O2 in the presence of ascorbic acid to establish a constitutive source of ROS within the tumor mass. We report here that incubation of folate receptor-expressing KB cells in culture with folate-PHQ plus ascorbate results in the death of the cancer cells with an IC50 of ~10 nM (folate-PHQ). We also demonstrate that a cleavable spacer linking folate to PHQ is significantly inferior to a noncleavable spacer, in contrast to most other folate-targeted therapeutic agents. Unfortunately, no evidence for folate-PHQ mediated tumor regression in murine tumor models is obtained, suggesting that unanticipated impediments to generation of cytotoxic quantities of ROS in vivo are encountered. Possible mechanisms and potential solutions to these unanticipated results are offered.


Prevention effect of orally active heparin conjugate on cancer-associated thrombosis.
Thrombogenesis is a major cause of morbidity and mortality in cancer patients. Prophylaxis with low-molecular-weight heparin (LMWH) is recommended for cancer patients, but requires non-parenteral delivery methods for long-term treatments. In this study, we sought to generate a new oligomeric-bile acid conjugate of LMWH that can be used for oral delivery. We first synthesized a tetramer of deoxycholic acid (tetraDOCA), which was site-specifically conjugated at the end saccharide of LMWH. When LMWH-tetraDOCA conjugate (LHe-tetraD) was orally administered at a dose of 5 mg/kg in ICR mice, the maximum anti-factor Xa level was increased up to 0.62±0.05 IU/mL without any evidence of liver toxicity, gastrointestinal damage, or thrombocytopenia. The cancer-associated thrombosis was induced in tumor-bearing mice by local heat application, and the fibrin deposition in tumors was evaluated. The oral administration of LHe-tetraD (either a single dose or multiple daily doses for up to 10 days) in mice substantially abolished the coagulation-dependent tropism of fibrinogen in the heated tumors and significantly decreased hemorrhage, compared to the mice treated with saline or subcutaneous injection of LMWH. Thus, the anticoagulation effect of oral LHe-tetraD invokes the benefits of oral delivery and promises to provide an effective and convenient treatment for cancer patients at risk of thrombosis.