Electrochemical behavior of colchicine using graphite-based screen-printed electrodes.
Miniaturized electrochemical graphite based sensor for the analysis of colchicine, along with its detailed construction was described. The electrochemical behavior of colchicine, both by cyclic and differential pulse voltammetry was investigated, showing an irreversible reduction and oxidation mechanism. The effect of several different electrochemical mediators was studied in order to decrease the oxidation potential of colchicine, but none of them showed a favorable electron transfer between the alkaloid and the electrode's surface. The influences of different biomolecules (calf thymus DNA, bovine serum albumin and beta-tubulin) over colchicine's electrochemical behavior were also presented. For quantitative determinations the anodic differential pulse voltammetric method in H(3)PO(4)/HClO(4) 0.01 M (pH 2.05) was selected, evaluating one of the oxidation peaks of colchicine at +970 mV versus Ag pseudo-reference. The electrochemical scanning parameters were optimized by an experimental design using response surface modelling. The method was validated according to ICH guidelines in the concentration range 85-1200 ng mL(-1) (R(2)=0.9966, n=7) colchicine in terms of linearity, accuracy, limits of detection (LOD) (41 ng mL(-1)), limits of quantification (LOQ) and fidelity and it was successfully applied to the determination of colchicine in tablets, without the interference of the excipients. The studied method showed the same sensitivity as the more complex HPLC procedure and micellar electrokinetic chromatography with spectrophotometric detection.


[Cognitive impairment in multiple sclerosis].
In this article we present a review of problems associated with cognitive dysfunctions in multiple sclerosis (MS). Neuropsychological investigations demonstrated that cognitive dysfunctions are common in MS patients and affect 40-65% of them. Cognitive deficits were found mainly on measures of memory, attention, information-processing speed, executive functions and abstract reasoning. The differences and degree of cognitive dysfunctions in MS are highlighted and usually related to different clinical appearance (clinical course, duration, disability level, treatment type). Furthermore, we have reviewed published correlations between psychopathological dysfunctions and neuroimaging results (mainly MRI techniques and functional imaging). The results of these correlations showed that an important role in cognitive impairment is related to the total lesion area, the severity of the pathological damage of the normal-appearing brain tissue (NABT), and brain atrophy.


Ontogeny of atrial natriuretic peptide and its receptor in the lung: effects on perinatal surfactant release.
During the transition at birth to air breathing, regulation of surfactant release from alveolar type II (ATII) cells is critical. Atrial natriuretic peptide (ANP) stimulates natriuretic peptide receptor-A (NPR-A) and increases intracellular cGMP. We examined the changes in ANP and NPR-A in respiratory epithelium during the perinatal period using immunohistochemistry and studied the effect of ANP on surfactant release from ATII cells isolated from fetal and newborn lambs. NPR-A mRNA was detected in the fetal lung by Northern Blot and RT-PCR. At 100 d gestation (term 145 d), ANP staining was absent and NPR-A staining was weak in cuboidal epithelial cells. ANP and NPR-A staining was prominent in ATII cells at 136 d gestation and was undetectable postnatally. ANP stimulated (maximal effect at 10(-10)M) surfactant release from both late gestation fetal and neonatal ATII cells. Protein kinase G inhibition significantly blocked this release. We conclude that ANP stimulates surfactant release in isolated perinatal ATII cells by a cGMP-dependent mechanism. ANP and NPR-A expression in ATII cells is greatest in late gestation and declines sharply postnatally. We speculate that increased activity of the ANP/NPR-A pathway in late gestation may prime the surfactant system, preparing the lung for air breathing.


Ceftriaxone for the treatment of serious infections.
Ceftriaxone is an investigational cephalosporin with a half-life of five to eight hours. In an uncontrolled study, we evaluated its efficacy and safety in 30 pediatric and 12 young adult patients with serious bacterial infections. This agent was administered to children at a dosage of 50 to 75 mg/kg/day intravenously in two divided doses. Those with CNS infections received 100 mg/kg/day. In adults, the dosage was 1 g either once or twice daily. The diseases we treated included pneumonia (17), sepsis (eight), ventriculoperitoneal shunt infections (three), osteomyelitis (three), brain abscess (two), peritonitis (two), and miscellaneous (seven). Clinical cures were achieved in all cases, although one child with cystic fibrosis and Pseudomonas pneumonia had persistent colonization in his sputum. No serious side effects were observed. Although not the agent of choice for many of these pathogens, ceftriaxone appears to represent an important alternative to therapy.


Augmentation of Tissue Perfusion with Contrast Ultrasound: Influence of Three-Dimensional Beam Geometry and Conducted Vasodilation.
Cavitation of microbubble contrast agents with ultrasound produces shear-mediated vasodilation and an increase in tissue perfusion. We investigated the influence of the size of the cavitation volume by comparing flow augmentation produced by two-dimensional (2D) versus three-dimensional (3D) therapeutic ultrasound. We also hypothesized that cavitation could augment flow beyond the ultrasound field through release of vasodilators that are carried downstream. In 11 rhesus macaques, cavitation of intravenously administered lipid-shelled microbubbles was performed in the proximal forearm flexor muscles unilaterally for 10 min. Ultrasound cavitation (1.3 MHz, 1.5 MPa peak negative pressure) was performed with 2D or 3D transmission with beam elevations of 5 and 25 mm, respectively, and pulsing intervals (PIs) sufficient to allow complete postdestruction refill (5 and 12 sec for 2D and 3D, respectively). Contrast ultrasound perfusion imaging was performed before and after cavitation, using multiplane assessment within and beyond the cavitation field in 1.5-cm increments. Cavitation in the hindlimb of mice using 2D ultrasound at a PI of 1 or 5 sec was performed to examine microvascular flow changes from cavitation in only arteries versus the microcirculation. In primates, the degree of muscle flow augmentation in the center of the cavitation field was similar for 2D and 3D conditions (five- to sixfold increase for both, P < .01 vs baseline). The spatial extent of flow augmentation was only modestly greater for 3D cavitation because of an increase in perfusion with 2D transmission that was detected outside of the cavitation field. In mice, cavitation in the microvascular compartment (PI 5 sec) produced the greatest degree of flow augmentation, yet cavitation in the arterial compartment (PI 1 sec) still produced a three- to fourfold increase in flow (P < .001 vs control). The mechanism for flow augmentation beyond the cavitation zone was investigated by in vitro studies that demonstrated cavitation-related release of vasodilators, including adenosine triphosphate and nitric oxide, from erythrocytes and endothelial cells. Compared with 2D transmission, 3D cavitation of microbubbles generates a similar degree of muscle flow augmentation, possibly because of a trade-off between volume of cavitation and PI, and only modestly increases the spatial extent of flow augmentation because of the ability of cavitation to produce conducted effects beyond the ultrasound field.


Cell lipid alterations resulting from prolonged exposure to dimethylaminoethylmethacrylate.
Dimethylaminoethylmethacrylate (DMAEMA), a commonly-used component of visible-light polymerized dental resins, has the potential to elute and interact with tissue cells to cause cytotoxicity or sublethal metabolic changes. Short-term exposure of cultured oral epithelial cells to sublethal DMAEMA concentrations has been shown previously to affect cell neutral lipid and phospholipid metabolism, resulting in accumulation of significant quantities of dimethylphosphatidylethanolamine (DMPE). In non-treated cells, DMPE is a transient intermediate in phospholipid metabolism and is not detectable by standard methods. In the current study, the effects of prolonged exposure of cells to DMAEMA, and the mechanisms for formation of DMPE in the presence of DMAEMA were examined. Exposure of a keratinizing hamster buccal cheek pouch cell line (HCP cells) to 0.8 mM DMAEMA for 2, 3, 7, and 14 days resulted in reduced incorporation of [14C]acetate into several classes of phospholipids. DMPE was detectable at all time points in DMAEMA-exposed cultures and comprised between 12.48% and 18.33% of the total radiolabeled phospholipids. The results of short-term exchange experiments indicated that headgroup exchange was not the major reaction responsible for formation of DMPE in DMAEMA-treated cells; rather the formation appeared to occur through typical phospholipid metabolic pathways. The cells appeared able to re-establish and maintain homeostasis in the presence of this altered cell lipid composition.


The transcriptional coactivator peroxisome proliferator activated receptor (PPAR)gamma coactivator-1 alpha and the nuclear receptor PPAR alpha control the expression of glycerol kinase and metabolism genes independently of PPAR gamma activation in human white adipocytes.
The purpose of this work was to determine the pattern of genes regulated by peroxisome proliferator-activated receptor (PPAR) gamma coactivator 1 alpha (PGC-1 alpha) in human adipocytes and the involvement of PPARalpha and PPARgamma in PGC-1 alpha transcriptional action. Primary cultures of human adipocytes were transduced with a PGC-1 alpha adenovirus and treated with PPARgamma and PPARalpha agonists. Variation in gene expression was assessed using pangenomic microarrays and quantitative RT-PCR. To investigate glycerol kinase (GyK), a target of PGC-1 alpha, we measured enzymatic activity and glycerol incorporation into triglycerides. In vivo studies were performed on wild-type and PPARalpha(-/-) mice. The GyK promoter was studied using chromatin immunoprecipitation and promoter reporter gene assays. Among the large number of genes regulated by PGC-1 alpha independently of PPARgamma, new targets involved in metabolism included the gene encoding GyK. The induction of GyK by PGC-1 alpha was observed at the levels of mRNA, enzymatic activity, and glycerol incorporation into triglycerides. PPARalpha was also upregulated by PGC-1 alpha. Its activation led to an increase in GyK expression and activity. PPARalpha was shown to bind and activate the GyK promoter. Experiments in mice confirmed the role of PGC-1 alpha and PPARalpha in the regulation of GyK in vivo. This work uncovers novel pathways regulated by PGC-1 alpha and reveals that PPARalpha controls gene expression in human white adipocytes. The induction of GyK by PGC-1 alpha and PPARalpha may promote a futile cycle of triglyceride hydrolysis and fatty acid reesterification.


Structural variations of blood group A antigens in human normal colon and carcinomas.
The blood group A determinant is carried by four basic carrier carbohydrate chains. This paper reports the expression of blood group A variants, as determined by immunohistology with highly specific monoclonal anti-A antibodies, in 18 adenocarcinomas of the distal colon, 4 specimens of normal proximal mucosa from group A persons, and 5 specimens of fetal colonic mucosa. Monoclonal antibodies directed to type 1 chain A, type 1 chain ALeb, type 2 chain A, type 2 chain ALey, and type 3 chain A (repetitive A) were used. In normal mucosa, type 1 chain A and ALeb were expressed in proximal regions. Type 1 chain A was expressed in columnar cells, whereas type 1 chain ALeb was found in goblet cells. Type 2 and type 3 chain A structures were not found in normal adult mucosa. All types of A antigens were detected in adenocarcinomas from the distal colon as well as in normal fetal mucosa. In fetal mucosa, type 1 chain A and ALeb antigens and type 3 chain A antigens were expressed in columnar cells, whereas type 2 chain A and Ley and type 1 chain ALeb antigens were found in goblet cells. The results indicate that blood group A antigens with type 1, 2, and 3 carriers are present in fetal mucosa and adenocarcinomas of distal colon, while epithelial mucosa of normal adult colon is characterized by the exclusive expression of type 1 chain A antigens.


Concussions and Risk Within Cultural Contexts of Play.
Concussions are a type of traumatic injury caused by a jolting of the brain that disrupts normal brain function, and multiple concussions can lead to serious long-term health consequences. In this article, we examine the relationship between college students' understanding of concussions and their willingness to continue playing despite the possibility of sustaining multiple head injuries. We use a mixed-methods approach that includes participant observation, cultural domain analysis, and structured interviews. Our research finds that students hold a robust cognitive understanding of concussion yet discursively frame concussions as skeletomuscular injuries. More importantly, students affirm the importance of playing sports for themselves and others, so their decisions to risk multiple concussions must be understood within cultural and biocultural contexts of meaningful social play. We suggest that peoples' decision to risk multiple head injuries should be understood as a desire for meaningful social play rather than an uninformed health risk.


Antithrombin-heparin covalent complex reduces microemboli during cardiopulmonary bypass in a pig model.
Transcranial Doppler-detected high-intensity transient signals (HITS) during cardiopulmonary bypass (CPB) surgery have been associated with postoperative neurocognitive dysfunction, suggesting microemboli in the brain could be a contributing factor. HITS occur despite administration of unfractionated heparin (UFH). This study was done to determine whether antithrombin-heparin covalent complex (ATH), a more potent anticoagulant than heparin, can reduce HITS during CPB. In a pig CPB model, ATH, UFH, or UFH + antithrombin (AT) was intravenously administered to female Yorkshire pigs after sternotomy. Twenty minutes later, hypothermic CPB was initiated and continued for 1.25 hours, then normothermia was re-established for 45 minutes. Protamine sulfate was given to neutralize the anticoagulants, and pigs were allowed to recover. HITS were monitored using an arterial flow probe placed over the carotid artery. Compared with UFH (300 or 1000 U/kg), ATH reduced the number of HITS during CPB in a dose-dependent manner. AT (3 mg/kg) + UFH (300 U/kg) resulted in an intermediate HITS rate between UFH and ATH (2 mg/kg in terms of AT). Examination of brain sections for emboli formation confirmed that, similar to HITS, number of thrombi decreased in direct proportion to ATH dosage. These results support the hypotheses that the majority of HITS represent thromboemboli and that ATH reduces emboli formation during CPB.


A GATA factor mediates cell type-restricted induction of HLA-E gene transcription by gamma interferon.
The human major histocompatibility complex (MHC) class Ib gene, HLA-E, codes for the major ligand of the inhibitory receptor NK-G-2A, which is present on most natural killer (NK) cells and some CD8(+) cytotoxic T lymphocytes. We have previously shown that gamma interferon (IFN-gamma) induction of HLA-E gene transcription is mediated through a distinct IFN-gamma-responsive element, the IFN response region (IRR), in all cell types studied. We have now identified and characterized a cell type-restricted enhancer of IFN-gamma-mediated induction of HLA-E gene transcription, designated the upstream interferon response region (UIRR), which is located immediately upstream of the IRR. The UIRR mediates a three- to eightfold enhancement of IFN-gamma induction of HLA-E transcription in some cell lines but not in others, and it functions only in the presence of an adjacent IRR. The UIRR contains a variant GATA binding site (AGATAC) that is critical to both IFN-gamma responsiveness and to the formation of a specific binding complex containing GATA-1 in K562 cell nuclear extracts. The binding of GATA-1 to this site in response to IFN-gamma was confirmed in vivo in a chromatin immunoprecipitation assay. Forced expression of GATA-1 in nonexpressing U937 cells resulted in a four- to fivefold enhancement of the IFN-gamma response from HLA-E promoter constructs containing a wild-type but not a GATA-1 mutant UIRR sequence and increased the IFN-gamma response of the endogenous HLA-E gene. Knockdown of GATA-1 expression in K562 cells resulted in a approximately 4-fold decrease in the IFN-gamma response of the endogenous HLA-E gene, consistent with loss of the increase in IFN-gamma response of HLA-E promoter-driven constructs containing the UIRR in wild-type K562 cells. Coexpression of wild-type and mutant adenovirus E1a proteins that sequester p300/CBP eliminated IFN-gamma-mediated enhancement through the UIRR, but only partially reduced induction through the IRR, implicating p300/CBP binding to Stat-1alpha at the IRR in the recruitment of GATA-1 to mediate the cooperation between the UIRR and IRR. We propose that the GATA-1 transcription factor represents a cell type-restricted mediator of IFN-gamma induction of the HLA-E gene.


Arbuscular mycorrhizal influence on leaf water potential, solute accumulation, and oxidative stress in soybean plants subjected to drought stress.
This study investigated several aspects related to drought tolerance in arbuscular mycorrhizal (AM) soybean plants. The investigation included both shoot and root tissues in order to reveal the preferred target tissue for AM effects against drought stress. Non-AM and AM soybean plants were grown under well-watered or drought-stressed conditions, and leaf water status, solute accumulation, oxidative damage to lipids, and other parameters were determined. Results showed that AM plants were protected against drought, as shown by their significantly higher shoot-biomass production. The leaf water potential was also higher in stressed AM plants (-1.9 MPa) than in non-AM plants (-2.5 MPa). The AM roots had accumulated more proline than non-AM roots, while the opposite was observed in shoots. Lipid peroxides were 55% lower in shoots of droughted AM plants than in droughted non-AM plants. Since there was no correlation between the lower oxidative damage to lipids in AM plants and the activity of antioxidant enzymes, it seems that first the AM symbiosis enhanced osmotic adjustment in roots, which could contribute to maintaining a water potential gradient favourable to the water entrance from soil into the roots. This enabled higher leaf water potential in AM plants during drought and kept the plants protected against oxidative stress, and these cumulative effects increased the plant tolerance to drought.


Evaluation of glutathione-sensitive fluorescent dyes in cortical culture.
The sensitivity of six fluorophores to glutathione (GSH) was evaluated in living rat cortical neuronal/glial mixed cultures during the first 23 days in vitro (DIV). Four of the dyes require glutathione-S-transferase (GST) to form a fluorescent conjugate, potentially conferring specificity for GSH: these included t-butoxycarbonyl-Leu-Met-7-amino-4-chloromethylcoumarin (CMAC), 7-amino-4-chloromethylcoumarin (CMAC-blue), monochlorobimane (MCB), and 5-chloromethylfluorescein diacetate (CMFDA). The final two dyes examined, 2,3-naphthalenedicarboxaldehyde (NDA) and o-phthaldehyde (OPD), do not require GST for adduct formation with GSH. To examine the specificity of the dyes for GSH, cultures grown less than 6 DIV were pretreated with diethyl maleate or DL-buthionine-(S, R)-sulfoximine to deplete endogenous GSH. This resulted in a substantial loss of staining by CMAC, CMAC-blue, and MCB and partial loss of staining by OPD, indicating specificity for GSH, while staining by CMFDA or NDA was not altered, indicating a lack of specificity for GSH. Neurons experienced a dramatic decline in GSH levels relative to astrocytes between 5-6 DIV, as shown by a loss of neuronal staining with CMAC, CMAC-blue and MCB. This decrease in staining was not due to a decrease in GST activity, as neurons stained with the GST-insensitive OPD also exhibited a decline in GSH-sensitive staining. Immunolabeling experiments demonstrated that CMAC staining co-localized with GFAP-positive astrocytes, but not with MAP-2-positive neurons, in 18 DIV cultures. Finally, CMAC was exploited as a specific morphological marker of astrocytes in cultures aged >5 DIV. CMAC staining was employed to monitor astrocyte proliferation and to resolve astrocytes in living mixed cultures co-loaded with the Ca(2+)-sensitive dye, calcium green 5N-AM. GLIA 30:329-341, 2000. Published 2000 Wiley-Liss, Inc.


Loss of TIMP3 by promoter methylation of Sp1 binding site promotes oral cancer metastasis.
The tissue inhibitor of metalloproteinase-3 (TIMP3) is the only member of the TIMP family that binds to the extracellular matrix and suppresses cancer cell growth, angiogenesis, migration, and invasion. However, whether the abnormal expression and promoter methylation of TIMP3 facilitates oral cancer metastasis remain unclear. In this study, the DNA methylation levels of TIMP3 CpG islands were assessed through pyrosequencing. Artificial modulation of TIMP3 was performed to explore the role of TIMP3 in tumor metastasis in vitro and in vivo. Our results showed that the suppression of TIMP3 transcription by DNA methylation involves the inhibition of the binding of the transcription factor Sp1 to the TIMP3 promoter as well as the upregulation of DNMT1 and DNMT3B. Functional analyses revealed that TIMP3 overexpression reduced migration and invasion abilities in oral cancer cells and inhibited lymph node metastasis in vivo. Moreover, TIMP3 regulated epithelial-mesenchymal transition by increasing the expression of the epithelial markers and reducing the expression of the mesenchymal markers. In conclusion, our findings suggested that the suppression of TIMP3 by DNA methylation contributes to oral cancer metastasis.


Tau isoforms are differentially expressed across the hippocampus in chronic traumatic encephalopathy and Alzheimer's disease.
Chronic traumatic encephalopathy (CTE) is a progressive neurodegenerative disease, characterized by hyperphosphorylated tau, found in individuals with a history of exposure to repetitive head impacts. While the neuropathologic hallmark of CTE is found in the cortex, hippocampal tau has proven to be an important neuropathologic feature to examine the extent of disease severity. However, the hippocampus is also heavily affected in many other tauopathies, such as Alzheimer's disease (AD). How CTE and AD differentially affect the hippocampus is unclear. Using immunofluorescent analysis, a detailed histologic characterization of 3R and 4R tau isoforms and their differential accumulation in the temporal cortex in CTE and AD was performed. CTE and AD were both observed to contain mixed 3R and 4R tau isoforms, with 4R predominating in mild disease and 3R increasing proportionally as pathological severity increased. CTE demonstrated high levels of tau in hippocampal subfields CA2 and CA3 compared to CA1. There were also low levels of tau in the subiculum compared to CA1 in CTE. In contrast, AD had higher levels of tau in CA1 and subiculum compared to CA2/3. Direct comparison of the tau burden between AD and CTE demonstrated that CTE had higher tau densities in CA4 and CA2/3, while AD had elevated tau in the subiculum. Amyloid beta pathology did not contribute to tau isoform levels. Finally, it was demonstrated that higher levels of 3R tau correlated to more severe extracellular tau (ghost tangles) pathology. These findings suggest that mixed 3R/4R tauopathies begin as 4R predominant then transition to 3R predominant as pathological severity increases and ghost tangles develop. Overall, this work demonstrates that the relative deposition of tau isoforms among hippocampal subfields can aid in differential diagnosis of AD and CTE, and might help improve specificity of biomarkers for in vivo diagnosis.


Cardiorenal Syndrome: Role of Arginine Vasopressin and Vaptans in Heart Failure.
Heart and kidney failure continued to be of increasing prevalence in today's society, and their comorbidity has synergistic effect on the morbidity and mortality of patients. Cardiorenal syndrome (CRS) is a complex disease with multifactorial pathophysiology. Better understanding of this pathophysiological network is crucial for the successful intervention to prevent advancement of the disease process. One of the major factors in this process is neurohormonal activation, predominantly involving renin-angiotensin-aldosterone system (RAAS) and arginine vasopressin (AVP). Heart failure causes reduced cardiac output/cardiac index (CO/CI) and fall in renal perfusion pressures resulting in activation of baroreceptors and RAAS, respectively. Activated baroreceptors and RAAS stimulate the release of AVP (non-osmotic pathway), which acts on V2 receptors located in the renal collecting ducts, causing fluid retention and deterioration of heart failure. Effective blockade of AVP action on V2 receptors has emerged as a potential treatment option in volume overload conditions especially in the setting of hyponatremia. Vasopressin receptor antagonists (VRAs), such as vaptans, are potent aquaretics causing electrolyte-free water diuresis without significant electrolyte abnormalities. Vaptans are useful in hypervolemic hyponatremic conditions like heart failure and liver cirrhosis, and euvolemic hyponatremic conditions like syndrome of inappropriate anti-diuretic hormone secretion. Tolvaptan and conivaptan are pharmaceutical agents that are available for the treatment of these conditions.


Countering the Misincentivization of Cancer Medicine by Real-Time Personal Professional Education.
In the United States, public and private payer misincentivization of medical care and the invisibility of costs to the consumers of that care have conspired to create unsustainable growth in health care expenditure that undermines our economy, diminishes our productivity, and limits our international competitiveness. Cancer medicine provides a small yet salient example. On average, Medicare reimburses oncologists 6% above the average acquisition price for essential anticancer agents and supportive therapies. The costs of these agents vary across a stunning five orders of magnitude, from a few dollars to more than $400,000 per course of treatment. The profitability to providers varies across approximately four orders of magnitude, from cents to thousands of dollars per treatment. National guidelines (National Comprehensive Cancer Network [NCCN], American Society of Clinical Oncology [ASCO]) help providers select the most effective therapies without regard for cost. We created an oncologist-to-oncologist professional education program to help cancer physicians optimally use expensive long-acting white blood cell growth factors, in accordance with these national guidelines. We then compared their use across a population of approximately 97,000 Medicare members before and after our intervention. Baseline use was recorded over two consecutive quarters (2009 to 2010). In March 2010, our oncologists initiated real-time discussions with the oncologists of 22 separate groups if these agents were ordered for use with regimens that placed patients at less than 10% risk of febrile neutropenia, according to NCCN guidelines. Neither NCCN nor ASCO recommend the routine use of these agents in this low-risk group. The care of 82 such patients was thoroughly discussed in the following 6 months. The monthly costs for these agents decreased by more than 50% by the final month of our intervention, although savings began immediately, reducing costs by more than $150,000 per quarter. No episode of febrile neutropenia was recorded in any patient in the intervention group. These savings generalize to the entire Medicare population at $30 million each month. We conclude that personal, oncologist-to-oncologist, real-time professional education will favorably modify oncologic prescribing behavior and can do so with significant immediate savings at no risk to patients with cancer.


The microbial adhesive arsenal deciphered by atomic force microscopy.
Microbes employ a variety of strategies to adhere to abiotic and biotic surfaces, as well as host cells. In addition to their surface physicochemical properties (e.g. charge, hydrophobic balance), microbes produce appendages (e.g. pili, fimbriae, flagella) and express adhesion proteins embedded in the cell wall or cell membrane, with adhesive domains targeting specific ligands or chemical properties. Atomic force microscopy (AFM) is perfectly suited to deciphering the adhesive properties of microbial cells. Notably, AFM imaging has revealed the cell wall topographical organization of live cells at unprecedented resolution, and AFM has a dual capability to probe adhesion at the single-cell and single-molecule levels. AFM is thus a powerful tool for unravelling the molecular mechanisms of microbial adhesion at scales ranging from individual molecular interactions to the behaviours of entire cells. In this review, we cover some of the major breakthroughs facilitated by AFM in deciphering the microbial adhesive arsenal, including the exciting development of anti-adhesive strategies.


Junctional epidermolysis bullosa of late onset explained by mutations in COL17A1.
Junctional epidermolysis bullosa of late onset (JEB-lo) is a rare disease characterized by blistering of primarily the hands and feet starting in childhood. The pathogenesis remains unclear. To clarify the pathogenesis of JEB-lo. Two patients with JEB-lo, a brother and a sister, were examined using electron microscopy (EM), immunofluorescence (IF) antigen mapping and molecular analysis. We found subtle changes in IF antigen mapping and EM. The most remarkable changes were loss of the apical-lateral staining of monoclonal antibodies (mAbs) against type XVII collagen (Col17), and a broadened distribution of mAb staining against the ectodomain of Col17, laminin-332 and type VII collagen. Mutation analysis of COL17A1, encoding Col17, showed a compound heterozygosity for a novel mutation c.1992_1995delGGGT and the known mutation c.3908G>A in both patients. The deletion c.1992_1995delGGGT results in a premature termination codon and mRNA decay, leaving the patients functionally hemizygous for the missense mutation c.3908G>A (p.R1303Q) in the noncollagenous 4 domain of Col17. JEB-lo is an autosomal recessive disorder caused by mutations in COL17A1, and subtle aberrations in EM and IF antigen mapping are clues to diagnosis.


Decreased beta-adrenergic receptor density in mononuclear leukocytes from thyroidectomized patients.
Beta-adrenergic receptor characteristics were investigated in peripheral blood mononuclear leukocytes taken from patients before and after partial thyroidectomy. In order to discriminate the effect of surgical stress per se from that of thyroidectomy, the analysis was also performed on patients before and after cholecystectomy. Receptor characteristics were determined by using dihydroalprenolol as ligand in direct equilibrium binding experiments. The binding affinity showed no changes either when two different surgical treatments were compared or when the same patient was analysed before and after the operation. On the contrary, a significant decrease in receptor density was found in thyroidectomized patients when compared pre- and post-operatively. This fall in receptor number seems to be linked with thyroid function since no statistically significant changes were observed in cholecystectomized patients in relation to surgical operation. This view is further supported by data on T3 serum levels, which show a significant fall after thyroidectomy but no statistically significant modifications after cholecystectomy. It is concluded that beta-adrenoceptor modulation plays an important role in the relationship between thyroid and beta-adrenergic system.


Selenium and clinical trials: new therapeutic evidence for multiple diseases.
The understanding of the essential role of selenium (Se) in human health has increased substantially in recent decades. Micronutrient deficiencies are very common in the general population and may be even more common in patients with different pathologies due to genetic or environmental causes and prescription drug use. Selenium is used by people in the prevention and/or treatment of different disorders including cardiovascular disease, osteoarthritis, rheumatoid arthritis, hypothyroidism, stroke, atherosclerosis, cancer susceptibility and treatment, HIV, AIDS, neuronal diseases such as Alzheimer or amyotrophic lateral sclerosis, pancreatitis, depression, and diabetes amongst others. Several mechanisms have been suggested to mediate the biological effects of Se and these include antioxidant defence systems, synthesis and stability of metabolites that act as intermediates implicated in diverse selenoproteins expression pathways oxidative metabolism, immune system modulation, DNA intercalators, kinase regulation, enzymatic cofactor, and gene expression. A number of clinical trials in recent years have provided convincing evidence of the central role of this element, either alone or in combination with other micronutrients or antioxidants, in the prevention and treatment of multiple diseases. Based on these studies this review focuses on the advances made so far in the study of mechanisms and applications of selenium compounds that could be suitable for chronic diseases.


Growth and metabolism in the embryonic white-spotted bamboo shark, Chiloscyllium plagiosum: comparison with embryonic birds and reptiles.
Birds and reptiles have been important models for studying the energetics of embryonic development. Studies on these groups reveal three metabolic patterns: an exponential increase in metabolism with embryo age, a sigmoidal increase with age, or a sigmoidal increase followed by a decrease before hatching. Models developed to explain avian metabolic patterns and developmental costs partition total costs between growth and maintenance. To test the generality of these models, we examined embryonic energetics of the oviparous white-spotted bamboo shark Chiloscyllium plagiosum. Oviparous sharks must actively ventilate during development, which could increase their development costs relative to birds and reptiles. Our results demonstrated that bamboo shark embryos have a peaked metabolic pattern and sigmoidal increase in body mass similar to ratites, crocodilians, and some turtles. The total cost of development was higher in bamboo sharks than in reptiles and many birds. However, calculations reveal that the high cost of bamboo shark development can be explained by the relatively long incubation time rather than the additional cost of muscular movement. Finally, an avian model can reasonably describe shark embryonic metabolism, suggesting that movement costs do not significantly alter the metabolic pattern during development.


Contraceptive use by number of living children in Ghana: Evidence from the 2017 maternal health survey.
There is a significant literature describing the link between parity and contraceptive use. However, there is limited knowledge about the disaggregation by parity of the type of contraceptives. In this study, we describe the use of contraceptives by parity among women of reproductive age in Ghana, focusing on use of highly effective methods (injection, pill, intrauterine device, implant, and sterilization). Using the 2017 Ghana Maternal Health Survey, a nationally-representative cross-sectional household survey, we describe contraceptive method use by number of living children among sexually active women of reproductive age. We then estimated predictors of use of highly effective contraception in a multilevel logistic regression model. Most women in this survey are not using any method of contraception, although this varies by whether or not they have begun childbearing. Contraceptive method use varies by number of living children. Before having children, natural (periodic abstinence and withdrawal) and episodic (condoms) methods dominate. Once a woman has one living child, method preference changes to injectables and implants. Factors associated with using a highly effective method of contraception are: having >3 children, being in a relationship, having had an abortion, being younger than age 30, and having had sexual intercourse within days of answering the survey (p < .001 for all). In this analysis, the number of living children a woman has, her age, and timing of last intercourse are the most significant predictors of using a highly effective method of contraception. However, the majority of participants in this study report not using any method of contraception to avoid unwanted pregnancies. Future research that attempts to unpack the disconnect between not wanting to become pregnant and not using contraception is warranted.


SAXS models of TGFBIp reveal a trimeric structure and show that the overall shape is not affected by the Arg124His mutation.
Human transforming growth factor β induced protein (TGFBIp) is composed of 683 residues, including an N-terminal cysteine-rich (EMI) domain, four homologous fasciclin domains, and an Arg-Gly-Asp (RGD) motif near the C-terminus. The protein is of interest because mutations in the TGFBI gene encoding TGFBIp lead to corneal dystrophy (CD), a condition where protein aggregates within the cornea compromise transparency. The complete three-dimensional structure of TGFBIp is not yet available, with the exception of a partial X-ray structure of the archetype FAS1 domain derived from Drosophila fasciclin-1. In this study, small-angle X-ray scattering (SAXS) models of intact wild-type (WT) human TGFBIp and a mutant (R124H) are presented. The mutation R124H leads to a variant of granular CD. The deduced structure of the TGFBIp monomer consists of four FAS1 domains in a simple "beads-on-a-string" arrangement, constructed by the superimposition of four consecutive Drosophila fasciclin domains. The SAXS-based model of the TGFBIp R124H mutant displayed no structural differences from WT. Both WT TGFBIp and the R124H mutant formed trimers at higher protein concentrations. The similar association properties and three-dimensional shape of the two proteins suggest that the mutation does not induce any major structural rearrangements, but points towards the role of other corneal-specific factors in the formation of corneal R124H deposits.


Adolescent onset cognitive regression and neuropsychiatric symptoms associated with the A140V MECP2 mutation.
The phenotype attributed to MECP2 mutations continues to expand. In addition to classic and variant Rett syndrome, phenotypes include non-specific intellectual disability and autism spectrum disorder in females, and fatal neonatal encephalopathy in males. One particular phenotype of parkinsonism, pyramidal signs, and neuropsychiatric symptoms (PPM-X) has been described only in males. We report on the first female with the A140V MECP2 mutation presenting with late onset cognitive regression, pyramidal symptoms, parkinsonism, and bipolar symptoms. This finding emphasizes the need to consider MECP2 sequencing in females with non-classic Rett phenotypes, particularly those with intellectual disability and neuropsychiatric features.


Natural variation and selection in GmSWEET39 affect soybean seed oil content.
Soybean (Glycine max) is a major contributor to the world oilseed production. Its seed oil content has been increased through soybean domestication and improvement. However, the genes underlying the selection are largely unknown. The present contribution analyzed the expression patterns of genes in the seed oil quantitative trait loci with strong selective sweep signals, then used association, functional study and population genetics to reveal a sucrose efflux transporter gene, GmSWEET39, controlling soybean seed oil content and under selection. GmSWEET39 is highly expressed in soybean seeds and encodes a plasma membrane-localized protein. Its expression level is positively correlated with soybean seed oil content. The variation in its promoter and coding sequence leads to different natural alleles of this gene. The GmSWEET39 allelic effects on total oil content were confirmed in the seeds of soybean recombinant inbred lines, transgenic Arabidopsis, and transgenic soybean hairy roots. The frequencies of its superior alleles increased from wild soybean to cultivated soybean, and are much higher in released soybean cultivars. The findings herein suggest that the sequence variation in GmSWEET39 affects its relative expression and oil content in soybean seeds, and GmSWEET39 has been selected to increase seed oil content during soybean domestication and improvement.


Enhanced age-related cataract in copper-zinc superoxide dismutase null mice.
As the lens is constantly exposed to light and oxygen that generate harmful reactive oxygen species, the importance of the intracellular antioxidant enzyme copper-zinc superoxide dismutase for the protection against age-related cataract development was explored. The development of lens opacities and the lens oxidative status were studied in different age groups of mice lacking copper-zinc superoxide dismutase and in wild-type mice. The lens opacities were quantified from lens photographs using digital image analysis. Thereafter, the lenses were homogenized and analysed regarding their contents of reduced glutathione and protein carbonyls suggestive of protein oxidation. The 18-week-old mice of both genotypes had clear lenses. At 1 year of age, the copper-zinc superoxide dismutase null mice had developed cortical lens opacities, whereas the wild-type mice did not show equivalent changes until 2 years of age. The lens contents of glutathione decreased only in the 2-year-old wild-type mice, whereas the carbonyls increased over time without any differences between the two genotypes. This study indicates that the lack of copper-zinc superoxide dismutase may accelerate age-related lens opacity development and that intracellular superoxide-derived oxidative stress may be damaging to the lens during ageing. Participation of the anti-oxidant enzyme copper-zinc superoxide dismutase in the protection against age-related cataract was thus suggested.


Reporting and Implementing Interventions Involving Machine Learning and Artificial Intelligence.
Increasingly, interventions aimed at improving care are likely to use such technologies as machine learning and artificial intelligence. However, health care has been relatively late to adopt them. This article provides clinical examples in which machine learning and artificial intelligence are already in use in health care and appear to deliver benefit. Three key bottlenecks toward increasing the pace of diffusion and adoption are methodological issues in evaluation of artificial intelligence-based interventions, reporting standards to enable assessment of model performance, and issues that need to be addressed for an institution to adopt these interventions. Methodological best practices will include external validation, ideally at a different site; use of proactive learning algorithms to correct for site-specific biases and increase robustness as algorithms are deployed across multiple sites; addressing subgroup performance; and communicating to providers the uncertainty of predictions. Regarding reporting, especially important issues are the extent to which implementing standardized approaches for introducing clinical decision support has been followed, describing the data sources, reporting on data assumptions, and addressing biases. Although most health care organizations in the United States have adopted electronic health records, they may be ill prepared to adopt machine learning and artificial intelligence. Several steps can enable this: preparing data, developing tools to get suggestions to clinicians in useful ways, and getting clinicians engaged in the process. Open challenges and the role of regulation in this area are briefly discussed. Although these techniques have enormous potential to improve care and personalize recommendations for individuals, the hype regarding them is tremendous. Organizations will need to approach this domain carefully with knowledgeable partners to obtain the hoped-for benefits and avoid failures.


Occurrence and distribution of Amphidomataceae (Dinophyceae) in Danish coastal waters of the North Sea, the Limfjord and the Kattegat/Belt area.
Some species of the dinophytes Azadinium and Amphidoma (Amphidomataceae) produce azaspiracids (AZA), lipophilic polyether compounds responsible for Azaspiracid Shellfish Poisoning (AZP) in humans after consumption of contaminated seafood. Toxigenic Amphidomataceae are known to occur in the North Atlantic and the North Sea area, but little is known about their importance in Danish coastal waters. In 2016, 44 Stations were sampled on a survey along the Danish coastline, covering the German Bight, Limfjord, the Kattegat area, Great Belt and Kiel Bight. Samples were analysed by live microscopy, liquid chromatography-tandem mass spectrometry (LC-MS/MS) and quantitative polymerase-chain-reaction (qPCR) on the presence of Amphidomataceae and AZA. Amphidomataceae were widely distributed in the area, but were below detection limit on most of the inner Limfjord stations. Cell abundances of the three toxigenic species, determined with species-specific qPCR assays on Azadinium spinosum, Az. poporum and Amphidoma languida, were generally low and restricted to the North Sea and the northern Kattegat, which was in agreement with the distribution of the generally low AZA abundances in plankton samples. Among the toxigenic species, Amphidoma languida was dominant with highest cell densities up to 3 × 103 cells L-1 on North Sea stations and at the western entrance of the Limfjord. Azaspiracids detected in plankton samples include low levels of AZA-1 at one station of the North Sea, and higher levels of AZA-38 and -39 (up to 1.5 ng L-1) in the North Sea and the Limfjord entrance. Furthermore, one new AZA (named AZA-63) was discovered in plankton of two North Sea stations. Morphological, molecular, and toxinological characterisation of 26 newly established strains from the area confirmed the presence of four amphidomatacean species (Az. obesum, Az. dalianense, Az. poporum and Am. languida). The single new strain of Az. poporum turned out as a member of Ribotype A2, which was previously only known from the Mediterranean. Consistent with some of these Mediterranean A2 strains, but different to the previously established AZA-37 producing Az. poporum Ribotype A1 strains from Denmark, the new strain did not contain any AZA. Azaspiracids were also absent in all Az. obesum and Az. dalianense strains, but AZA-38 and -39 were found in all Am. languida strains with total AZA cell quotas ranging from 0.08 up to 94 fg cell-1. In conclusion, AZA-producing microalgae and their respective toxins were low in abundance but widely present in the area, and thus might be considered in local monitoring programs to preserve seafood safety in Danish coastal waters.


Etiology and therapeutic approach to elevated lactate levels.
Lactate levels are commonly evaluated in acutely ill patients. Although most often used in the context of evaluating shock, lactate levels can be elevated for many reasons. While tissue hypoperfusion may be the most common cause of elevation, many other etiologies or contributing factors exist. Clinicians need to be aware of the many potential causes of lactate level elevation as the clinical and prognostic importance of an elevated lactate level varies widely by disease state. Moreover, specific therapy may need to be tailored to the underlying cause of elevation. The present review is based on a comprehensive PubMed search between the dates of January 1, 1960, to April 30, 2013, using the search term lactate or lactic acidosis combined with known associations, such as shock, sepsis, cardiac arrest, trauma, seizure, ischemia, diabetic ketoacidosis, thiamine, malignancy, liver, toxins, overdose, and medication. We provide an overview of the pathogenesis of lactate level elevation followed by an in-depth look at the varied etiologies, including medication-related causes. The strengths and weaknesses of lactate as a diagnostic/prognostic tool and its potential use as a clinical end point of resuscitation are discussed. The review ends with some general recommendations on the management of patients with elevated lactate levels.


Development of dendritic tonic GABAergic inhibition regulates excitability and plasticity in CA1 pyramidal neurons.
Synaptic plasticity rules change during development: while hippocampal synapses can be potentiated by a single action potential pairing protocol in young neurons, mature neurons require burst firing to induce synaptic potentiation. An essential component for spike timing-dependent plasticity is the backpropagating action potential (BAP). BAP along the dendrites can be modulated by morphology and ion channel composition, both of which change during late postnatal development. However, it is unclear whether these dendritic changes can explain the developmental changes in synaptic plasticity induction rules. Here, we show that tonic GABAergic inhibition regulates dendritic action potential backpropagation in adolescent, but not preadolescent, CA1 pyramidal neurons. These developmental changes in tonic inhibition also altered the induction threshold for spike timing-dependent plasticity in adolescent neurons. This GABAergic regulatory effect on backpropagation is restricted to distal regions of apical dendrites (>200 μm) and mediated by α5-containing GABA(A) receptors. Direct dendritic recordings demonstrate α5-mediated tonic GABA(A) currents in adolescent neurons which can modulate BAPs. These developmental modulations in dendritic excitability could not be explained by concurrent changes in dendritic morphology. To explain our data, model simulations propose a distally increasing or localized distal expression of dendritic α5 tonic inhibition in mature neurons. Overall, our results demonstrate that dendritic integration and plasticity in more mature dendrites are significantly altered by tonic α5 inhibition in a dendritic region-specific and developmentally regulated manner.


SDF-1/CXCR4 and VLA-4 interaction regulates homing in Waldenstrom macroglobulinemia.
Waldenstrom macroglobulinemia (WM) is characterized by widespread involvement of the bone marrow at the time of diagnosis, implying continuous homing of WM cells into the marrow. The mechanisms by which trafficking of the malignant cells into the bone marrow has not been previously elucidated. In this study, we show that WM cells express high levels of chemokine and adhesion receptors, including CXCR4 and VLA-4. We showed that CXCR4 was essential for the migration and trans-endothelial migration of WM cells under static and dynamic shear flow conditions, with significant inhibition of migration using CXCR4 knockdown or the CXCR4 inhibitor AMD3100. Similarly, CXCR4 or VLA-4 inhibition led to significant inhibition of adhesion to fibronectin, stromal cells, and endothelial cells. Decreased adhesion of WM cells to stromal cells by AMD3100 led to increased sensitivity of these cells to cytotoxicity by bortezomib. To further investigate the mechanisms of CXCR4-dependent adhesion, we showed that CXCR4 and VLA-4 directly interact in response to SDF-1, we further investigated downstream signaling pathways regulating migration and adhesion in WM. Together, these studies demonstrate that the CXCR4/SDF-1 axis interacts with VLA-4 in regulating migration and adhesion of WM cells in the bone marrow microenvironment.


Acquired Hemophilia A following Pfizer-BioNTech SARS CoV-2 mRNA vaccine, successfully treated with prednisolone and rituximab.
Acquired haemophilia A (AHA) is a rare bleeding disorder, characterised by the presence of autoantibodies to clotting factor VIII (FVIII). AHA can be idiopathic or occur in the context of malignancy, autoimmune disease, drugs, or pregnancy. Recently, cases of AHA following both COVID-19 infection and vaccination have been reported. We report the case of a 95-year-old female who was immunised with the Pfizer-BioNTech SARS CoV-2 mRNA vaccine, with doses given three weeks apart. Spontaneous bruising over her extremities appeared one week after the initial dose, with hospital admission occurring three weeks after the second. Examination revealed a large haematoma on the dorsum of the right hand with resultant bleeding and widespread ecchymoses. Investigations confirmed a diagnosis of AHA. Initial management included high dose prednisolone, recombinant Factor VIII and tranexamic acid. There was no significant clinical improvement after three days, so intravenous rituximab 100 mg weekly for four weeks was commenced. The activated partial thromboplastin time (aPTT) normalised after two doses and Factor VIII level reached 0.68U/ml on day + 22. The patient was successfully discharged from hospital after 37 days. Four cases of AHA following administration of COVID mRNA vaccines (Pfizer and Moderna) have been documented. AHA should be a differential in patients presenting with bleeding following COVID-19 vaccination, in the presence of a normal platelet count. Rapid recognition, prompt initiation of immunosuppressive treatment and rigorous supportive cares are required to minimise morbidity and mortality.


Porcine collagen crosslinking, degradation and its capability for fibroblast adhesion and proliferation.
Porcine dermal collagen permanently crosslinked with hexamethylene diisocyanate was investigated for its suitability as a dermal tissue engineering matrix. It was found that the chemically crosslinked collagen had far fewer free lysine groups per collagen molecule than did the uncrosslinked matrix. The ability of the matrix to support human primary fibroblast outgrowth from explants was compared for matrices that had been presoaked in various solutions, including fibroblast media, cysteine and phosphate buffered saline (PBS). It was found that superior cell outgrowth was obtained after soaking with fibroblast media and PBS. The fibroblast attachment properties of the matrix were compared against tissue culture plastic and PET. The collagen matrix showed the least amount of cell retention compared to the other to matrices, however, the general trends were similar for all three scaffolds. Longer term cultures on the collagen showed fibroblasts covering the matrix stacking up on each other and bridging natural hair follicles. However, it was also observed that the fibroblasts were not able to penetrate into the matrix structure. This was believed to result from the chemical crosslinking, as shown by the resistance of the matrix to degradation by collagenases.


Fructose augments infection-impaired net hepatic glucose uptake during TPN administration.
During chronic total parenteral nutrition (TPN), net hepatic glucose uptake (NHGU) and net hepatic lactate release (NHLR) are markedly reduced (downward arrow approximately 45 and approximately 65%, respectively) with infection. Because small quantities of fructose are known to augment hepatic glucose uptake and lactate release in normal fasted animals, the aim of this work was to determine whether acute fructose infusion with TPN could correct the impairments in NHGU and NHLR during infection. Chronically catheterized conscious dogs received TPN for 5 days via the inferior vena cava at a rate designed to match daily basal energy requirements. On the third day of TPN administration, a sterile (SHAM, n = 12) or Escherichia coli-containing (INF, n = 11) fibrin clot was implanted in the peritoneal cavity. Forty-two hours later, somatostatin was infused with intraportal replacement of insulin (12 +/- 2 vs. 24 +/- 2 microU/ml, SHAM vs. INF, respectively) and glucagon (24 +/- 4 vs. 92 +/- 5 pg/ml) to match concentrations previously observed in sham and infected animals. After a 120-min basal period, animals received either saline (Sham+S, n = 6; Inf+S, n = 6) or intraportal fructose (0.7 mg x kg(-1) x min(-1); Sham+F, n = 6; Inf+F, n = 5) infusion for 180 min. Isoglycemia of 120 mg/dl was maintained with a variable glucose infusion. Combined tracer and arteriovenous difference techniques were used to assess hepatic glucose metabolism. Acute fructose infusion with TPN augmented NHGU by 2.9 +/- 0.4 and 2.5 +/- 0.3 mg x kg(-1) x min(-1) in Sham+F and Inf+F, respectively. The majority of liver glucose uptake was stored as glycogen, and NHLR did not increase substantially. Therefore, despite an infection-induced impairment in NHGU and different hormonal environments, small amounts of fructose enhanced NHGU similarly in sham and infected animals. Glycogen storage, not lactate release, was the preferential fate of the fructose-induced increase in hepatic glucose disposal in animals adapted to TPN.


Four New Pentacyclic Triterpene Glycosides Isolated from the Fruits of Cryptolepis buchananii R.Br. ex Roem. & Schult and Their Inhibition of NO Production in LPS-activated RAW264.7 Cells.
From the methanol extract of the Cryptolepis buchananii fruits, four undescribed pentacyclic triterpenene glycosides (1-4) and five known pentacyclic triterpenenes (5-9) were isolated. Their structures were determined to be uncargenin C 28-O-α-L-rhamnopyranosyl-(1→2)-β-D-glucopyranosyl ester (1), 3-O-β-D-glucopyranosyluncargenin C 28-O-α-L-rhamnopyranosyl-(1→2)-β-D-glucopyranosyl ester (2), 3-O-β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl-6β,23-dihydroxyursolic acid 28-O-α-L-rhamnopyranosyl-(1→2)-β-D-glucopyranosyl ester (3), 3-O-β-D-glucopyranosyl-(1→2)-β-D-glucopyranosylasiatic acid 28-O-α-L-rhamnopyranosyl-(1→2)-β-D-glucopyranosyl ester (4), asiatic acid (5), 2α,3β,23-trihydroxyoleana-11,13(18)-dien-28-oic acid (6), arjunolic acid (7), 6β-hydroxyarjunolic acid (8), and actinidic acid (9) based on analyses of their HR-ESI-MS, 1D and 2D NMR spectra. All the isolates showed significantly NO production inhibition in LPS-activated RAW264.7 cells with the IC50 values ranging from 18.79 to 37.57 μM, compared to that of the positive control compound, dexamethasone, which showed IC50 value of 14.05 μM.


Geographic song discrimination in relation to dispersal distances in song sparrows.
Whether geographic variation in signals actually affects communication between individuals depends on whether discriminable differences in signals occur over distances that individuals move in their lifetimes. We measure the ability of song sparrows (Melospiza melodia) to discriminate foreign from local songs using foreign songs recorded at a series of increasing distances and compare the results with previous measurements of dispersal distances. We test discrimination in males using playback of songs on territories and measuring approach and in females using playback to estradiol-treated captives and measuring courtship display. Females fail to discriminate against foreign songs recorded at 18 km but do discriminate against foreign songs recorded at 34, 68, 135, and 540 km. Males fail to discriminate against foreign songs recorded at 18, 34, 68, 135, and 270 km but do discriminate against foreign songs from 540 km. Females are more discriminating, but even they do not discriminate at a distance three times the root-mean-square dispersal distance, as estimated from mtDNA variation. We suggest that female preference for local songs benefits females not because it allows them to reject foreign males but because accurate production of local song serves as a test of song-learning ability.


Localization of seminal plasma proteinase inhibitors in human spermatozoa as revealed by the indirect immunofluorescence technique.
Acidic extracts of washed, ejaculated human spermatozoa contain, besides acrosin, two proteinase inhibitors, a trypsin-chymotrypsin (elastase) inhibitor (HUSI-I) and a trypsin-acrosin inhibitor (HUSI-II). Using the indirect immunofluorescence technique these inhibitors could be localized in the spermatozoa. Ejaculated spermatozoa were treated with monospecific antibodies raised in rabbits against HUSI-I and HUSI-II, respectively, and with fluorescein-labeled IgG from goat directed against the rabbit IgG. If acetone-fixed spermatozoa were used, fluorescence appeared only in a small ring near or at the equatorial segment of the spermatozoa. After prefixation of washed spermatozoa with 0.36% formaldehyde, however, distribution of both inhibitors in the region of the acrosomal caps could clearly be demonstrated. Present results suggest that they are attached at the plasma membrane. Obviously, in the case of human spermatozoa the inhibitors are relatively easily detached together with the membrane so that prefixation is necessary to achieve proper localization.


Influence of transdermal scopolamine on motion sickness during 7 days' exposure to heavy seas.
We conducted a double-blind, placebo-controlled study to evaluate the efficacy and tolerability of transdermal scopolamine in the prevention of motion sickness (MS) aboard a frigate during 7 days of continuously moderate or heavy seas. Forty-nine healthy sailors with a previous history of MS were randomly assigned to receive a transdermal therapeutic system of scopolamine (TTS-S) or transdermal placebo (TD-P). Patches were placed behind the ears at least 4 hours before departure and were removed 72 hours later. Subjects were observed on days 1 to 4 and 6. In the TTS-S group, both subjective feeling of MS and the incidence of nausea were reduced during the first 2 days. Because of adaptation, differences in signs and symptoms of MS between subjects receiving TTS-S and TD-P disappeared after the second day. During the first 3 days, vomiting occurred less often in the TTS-S group. On day 6, 3 days after removal of the patch, vomiting occurred in 23% of the TTS-S group, probably due to delay in adaptation, but none of the subjects in the TD-P group vomited. Concentration was not adversely influenced, since the ability to work increased in the TTS-S group. During prolonged continuous exposure to heavy and moderate seas, 2.5 cm2 TTS-S discs proved to be efficacious in preventing MS, with xerostomia as a tolerable side effect and no significant ocular side effects.


[Compulsory licensing of efavirenz in Brazil in 2007: contextualization].
The present article aims at contextualizing the first Brazilian experience with compulsory licensing, which functions as a defense mechanism to prevent excessive pricing by holders of patents. According to this mechanism, a government can authorize a third party to explore the patented object (in this case a drug) without previous consent from the patent holder. On May 4, 2007, Brazil officially issued compulsory licensing of the antiretroviral drug efavirenz for public, non-commercial use. Initially, generic versions of the drug were purchased from laboratories in India. The next step was the manufacture of efavirenz by Farmanguinhos, official pharmaceutical laboratory (Fundação Osvaldo Cruz). It is concluded that the decision made by the Brazilian government to issue compulsory licensing of efavirenz was correct, taking into account the projected savings of US$ 236.8 until 2012 and the guarantee of availability of efavirenz, the most usual free antiretroviral treatment provided in Brazil.


Radiographic evaluation of third molar development in 6- to 24-year-olds.
This study investigated the developmental stages of third molars in relation to chronological age and compared third molar development according to location and gender. A retrospective analysis of panoramic radiographs of 2490 patients aged between 6 and 24 years was conducted, and the developmental stages of the third molars were evaluated using the modified Demirjian's classification. The mean age, standard deviation, minimal and maximal age, and percentile distributions were recorded for each stage of development. A Mann-Whitney U test was performed to test the developmental differences in the third molars between the maxillary and mandibular arches and between genders. A linear regression analysis was used for assessing the correlation between the third molar development and chronological age. The developmental stages of the third molars were more advanced in the maxillary arch than the mandibular arch. Males reached the developmental stages earlier than females. The average age of the initial mineralization of the third molars was 8.57 years, and the average age at apex closure was 21.96 years. The mean age of crown completion was 14.52 and 15.04 years for the maxillary and the mandibular third molars, respectively. The developmental stages of the third molars clearly showed a strong correlation with age. The third molars developed earlier in the upper arch than the lower arch; further, they developed earlier in males than in females.


Community-based epidemiological study on diabetes in Pu-Li, Taiwan.
To determine the prevalence and possible risk factors associated with diabetes in Pu-Li, Taiwan. The project was a community-based cross-sectional study with stratified cluster sampling of residents greater than or equal to 30 yr of age. Medical students conducted home-visit interviews with public health nurses, drew fasting blood samples from 1738 of 2573 registered residents greater than or equal to 30 yr of age in Pu-Li, Taiwan. The prevalence of diabetes, including those with a history of diabetes and newly diagnosed cases determined by modified World Health Organization criteria, was 12.4%. With the standard world population of Segi, the age-adjusted prevalences were 6.9% for previous diabetes and 4.4% for new diabetes. The age-specific prevalence of diabetes increased significantly with age for new diabetes cases but not for previously diagnosed cases. To study factors associated with diabetes, we first applied univariate analysis. Stratified analyses by age and sex were used for interaction assessment. A significant interaction effect on diabetes was found between age and sex. Logistic regression was used for multivariate analysis. The final model showed that, after controlling for all other covariates, the significant factors associated with diabetes were family history of diabetes (father and/or mother, yes vs. no, odds ratio [OR] 3.13, 95% confidence interval [CI] 1.17-8.33), obesity (yes vs. no, OR 2.69, 95% CI 1.09-6.63), and age (continuous, OR 1.04, 95% CI 1.00-1.08). Locality and physical activity were not significantly correlated with diabetes. The age-adjusted prevalence of diabetes in Pu-Li were 6.9% for previous diabetes and 4.4% for new diabetes. Possible risk factors associated with diabetes with logistic regression were family history of diabetes, obesity, and age. Locality and physical activity were not significantly correlated with diabetes.


Cluster analysis in 975 patients with current cough identifies a phenotype with several cough triggers, many background disorders, and low quality of life.
Recognition of disorder phenotypes may help to estimate prognosis and to guide the clinical management. Current cough management guidelines classify patients according to the duration of the cough episode. However, this classification is not based on phenotype analyses. The present study aimed to identify cough phenotypes by clustering. An email survey among employed, working-age subjects identified 975 patients with current cough. All filled in a comprehensive 80-item questionnaire including the Leicester Cough Questionnaire. Phenotypes were identified utilizing K-means partitional clustering. A subgroup filled in a follow-up questionnaire 12 months later to investigate the possible differences in the prognosis between the phenotypes. Two clusters were found. The cluster A included 608 patients (62.4% of the population) and the cluster B 367 patients (37.6%). The three most important variables to separate the clusters were the number of the triggers of cough (mean 2.63 (SD 2.22) vs. 6.95 (2.30), respectively, p < 0.001), the number of the cough background disorders (chronic rhinosinusitis, current asthma, gastroesophageal reflux disease, 0.29 (0.50) vs. 1.28 (0.75), respectively, p < 0.001), and the Leicester Cough Questionnaire physical domain (5.33 (0.76) vs. 4.25 (0.84), respectively, p < 0.001). There were significant interrelationships between these three variables (each p < 0.001). Duration of the episode was not among the most important variables to separate the clusters. At 12 months, 27.0% of the patients of the cluster A and 46.1% of the patients of the cluster B suffered from cough that had continued without interruptions from the first survey (p < 0.001). Two cough phenotypes could be identified. Cluster A represents phenotype A, which includes the majority of patients and has a tendency to heal by itself. The authors propose that cluster B represents phenotype TBQ (Triggers, Background disorders, Quality of life impairment). Given the poor prognosis of this phenotype, it urges a prompt and comprehensive clinical evaluation regardless of the duration of the cough episode.


Assessment of a land-locked Atlantic salmon (Salmo salar L.) population as a potential genetic resource with a focus on long-chain polyunsaturated fatty acid biosynthesis.
The natural food for Atlantic salmon (Salmo salar) in freshwater has relatively lower levels of omega-3 (n-3) long-chain polyunsaturated fatty acids (LC-PUFA) than found in prey for post-smolt salmon in seawater. Land-locked salmon such as the Gullspång population feed exclusively on freshwater type lipids during its entire life cycle, a successful adaptation derived from divergent evolution. Studying land-locked populations may provide insights into the molecular and genetic control mechanisms that determine and regulate n-3 LC-PUFA biosynthesis and retention in Atlantic salmon. A two factorial study was performed comparing land-locked and farmed salmon parr fed diets formulated with fish or rapeseed oil for 8 weeks. The land-locked parr had higher capacity to synthesise n-3 LC-PUFA as indicated by higher expression and activity of desaturase and elongase enzymes. The data suggested that the land-locked salmon had reduced sensitivity to dietary fatty acid composition and that dietary docosahexaenoic acid (DHA) did not appear to suppress expression of LC-PUFA biosynthetic genes or activity of the biosynthesis pathway, probably an evolutionary adaptation to a natural diet lower in DHA. Increased biosynthetic activity did not translate to enhanced n-3 LC-PUFA contents in the flesh and diet was the only factor affecting this parameter. Additionally, high lipogenic and glycolytic potentials were found in land-locked salmon, together with decreased lipolysis which in turn could indicate increased use of carbohydrates as an energy source and a sparing of lipid.


Helicity discrimination in N,N'-dibenzoyl-1,2,3,4,7,8,9,10-octahydro-1,10-phenanthrolines and their thiono- and selenocarbonyl analogues by inclusion complexation with chiral diols.
X-ray crystallographic analysis of the title compounds revealed that they assume a folded helical conformation of an approximate C2 symmetry in the solid state. Dithioamide 5b, diselenoamide 5c and monoselenoamide 5d were resolved to enantiomers by inclusion crystallization with optically active diols (TADDOLs). The absolute configuration of the guest molecules in the complexes 5b·6a, 5c·6a and 5d·6a was assigned as P. The optical activity of the resolved compounds is manifested by their CD spectra showing relatively strong Cotton effects in the region of thionoamide and selenoamide n–π* transition. The optically active thiono- and selenoamides are configurationally labile compounds and gradually racemize in solution but they are stable in the form of the inclusion complexes. The first-order kinetics of the racemization in solution allowed us to assign the racemization barriers by the spectropolarimetric measurements.


The first 12 weeks following discharge from hospital: the experience of Gujarati South Asian survivors of acute myocardial infarction and their families.
The period following discharge from hospital after an acute myocardial infarction (MI) is associated with vulnerability and psychosocial and physical morbidity for many survivors and their families. It is reported that people experience interpersonal, family and financial problems, self-care obstacles, work and physical difficulties. Culture and ethnicity undeniably influence the illness experience and the process of recovery and adjustment. This study investigated the perceptions of Gujarati survivors of acute MI and their families in Leicester, United Kingdom in the first 12 weeks following discharge in order to develop a profile of their health seeking beliefs and needs. Thirty-one interviews with 19 Gujarati MI survivors and their families (representing approximately 31 hours of dialogue) were analysed using grounded theory. Qualitative data revealed a period of vulnerability not only for survivors but also their families as they processed recent events and faced the future. Data analysis revealed nine interrelated themes describing the post-discharge experience for Gujarati survivors and their families. Data revealed that normal life was often markedly changed by the MI experience. Reflection, contemplation and resignation characterize this period of vulnerability. Overwhelmingly, data analysis revealed that the Gujarati culture, beliefs and customs influenced the recovery experience.


LAMP-PCR detection of ochratoxigenic Aspergillus species collected from peanut kernel.
Over the last decade, ochratoxin A (OTA) has been widely described and is ubiquitous in several agricultural products. Ochratoxins represent the second-most important mycotoxin group after aflatoxins. A total of 34 samples were surveyed from 3 locations, including Mecca, Madina, and Riyadh, Saudi Arabia, during 2012. Fungal contamination frequency was determined for surface-sterilized peanut seeds, which were seeded onto malt extract agar media. Aspergillus niger (35%), Aspergillus ochraceus (30%), and Aspergillus carbonarius (25%) were the most frequently observed Aspergillius species, while Aspergillus flavus and Aspergillus phoenicis isolates were only infrequently recovered and in small numbers (10%). OTA production was evaluated on yeast extract sucrose medium, which revealed that 57% of the isolates were A. niger and 60% of A. carbonarius isolates were OTA producers; 100% belonged to A. ochraceus. Only one isolate, morphologically identified as A. carbonarius, and 3 A. niger isolates unstably produced OTA. A polymerase chain reaction (PCR)-based identification and detection assay was used to identify A. ochraceus isolates. Using the primer sets OCRA1/OCRA2, 400-base pair PCR fragments were produced only when genomic DNA from A. ochraceus isolates was used. Recently, the loop-mediated isothermal amplification assay using recombinase polymerase amplification chemistry was used for A. carbonarius and A. niger DNA identification. As a non-gel-based technique, the amplification product was directly visualized in the reaction tube after adding calcein for naked-eye examination.


Disrupting malaria parasite AMA1-RON2 interaction with a small molecule prevents erythrocyte invasion.
Plasmodium falciparum resistance to artemisinin derivatives, the first-line antimalarial drug, drives the search for new classes of chemotherapeutic agents. Current discovery is primarily directed against the intracellular forms of the parasite. However, late schizont-infected red blood cells (RBCs) may still rupture and cause disease by sequestration; consequently targeting invasion may reduce disease severity. Merozoite invasion of RBCs requires interaction between two parasite proteins AMA1 and RON2. Here we identify the first inhibitor of this interaction that also blocks merozoite invasion in genetically distinct parasites by screening a library of over 21,000 compounds. We demonstrate that this inhibition is mediated by the small molecule binding to AMA1 and blocking the formation of AMA1-RON complex. Electron microscopy confirms that the inhibitor prevents junction formation, a critical step in invasion that results from AMA1-RON2 binding. This study uncovers a strategy that will allow for highly effective combination therapies alongside existing antimalarial drugs.


Evaluation of resveratrol, green tea extract, curcumin, oxaloacetic acid, and medium-chain triglyceride oil on life span of genetically heterogeneous mice.
The National Institute on Aging Interventions Testing Program (ITP) was established to evaluate agents that are hypothesized to increase life span and/or health span in genetically heterogeneous mice. Each compound is tested in parallel at three test sites. It is the goal of the ITP to publish all results, negative or positive. We report here on the results of lifelong treatment of mice, beginning at 4 months of age, with each of five agents, that is, green tea extract (GTE), curcumin, oxaloacetic acid, medium-chain triglyceride oil, and resveratrol, on the life span of genetically heterogeneous mice. Each agent was administered beginning at 4 months of age. None of these five agents had a statistically significant effect on life span of male or female mice, by log-rank test, at the concentrations tested, although a secondary analysis suggested that GTE might diminish the risk of midlife deaths in females only.


Adherence intervention for HIV-infected persons who use drugs: adaptation, open trial, and pilot randomized hybrid type 1 trial protocol.
HIV-infected people who use drugs (PWUD) exhibit the highest rates of non-adherence to antiretroviral therapy (ART) among people living with HIV. This contributes to poor treatment outcomes, increased morbidity and mortality, and HIV transmission. However, current interventions fail to address the unique barriers to adherence faced by this population. Life Steps is a brief, single session intervention that demonstrated increased ART adherence among HIV-infected individuals. This study protocol seeks to improve clinical practice by adapting Life Steps for HIV-infected PWUD and adding a brief motivational intervention addressing drug use. This intervention will incorporate educational, motivational, and behavioral skills components specifically aimed at improving adherence and linkage to substance use treatment among HIV-infected PWUD. This project will consist of three phases using a mixed-methods approach. In Phase 1, qualitative interviews with HIV-infected PWUD and community providers, coupled with feedback from an expert review panel, will be used to enhance the existing Life Steps manual and interventionist training protocol. In Phase 2, the prototype will be pilot tested and qualitative exit interviews with HPWUD will identify the strengths and limitations of the intervention. Data regarding feasibility, acceptability, and barriers to delivery will guide modifications to finalize a modified Life Steps-Drug Use (LS-DU) protocol. In Phase 3, a pilot type 1 hybrid effectiveness-implementation trial will examine the effectiveness of LS-DU relative to a health education intervention control condition on ART adherence and viral load data at 1-, 3-, and 6-months. Data regarding clinic readiness for implementation and intervention sustainability potential will be collected. This protocol will adapt and evaluate an intervention to improve adherence among HIV-infected PWUD. Results of this study will provide significant data on the acceptability, initial effectiveness, and sustainability potential of an adherence intervention for a high risk and underserved population. Trial registration NCT02907697.


Chiroptical nature of two-exciton states of light-harvesting complex: Doubly resonant three-wave-mixing spectroscopy.
Photosynthetic light-harvesting complex is a coupled multichromophore system. Due to electronic couplings between neighboring chlorophylls in the complex, the one- and two-exciton states are delocalized and they can be written as linear combinations of singly and doubly excited configurations, respectively. Despite that the chiroptical properties of one-exciton states in such a multichromophore system have been investigated by using linear optical activity measurement techniques; those of two-exciton states have not been studied before due to a lack of appropriate measurement methods. Here, we present a theoretical description on chiroptical chi((2)) spectroscopy and show that it can be used to investigate such properties of a photosynthetic light-harvesting system, which is the Fenna-Matthews-Olson complex, consisting of seven bacteriochlorophylls in its protein subunit. To simulate the doubly resonant sum- and difference-frequency-generation spectra of the complex, one- and two-exciton transition dipoles were calculated. Carrying out quantum chemistry calculations of electronically excited states of a model bacteriochlorophyll system and taking into account the dipole-induced dipole electronic transition processes between the ground state and two-exciton states, we could calculate the two-dimensional sum-frequency-generation spectra revealing dominant second-order chiroptical transition pathways and involved one- and two-exciton states. It is believed that the present computational scheme and the theoretically proposed doubly resonant two-dimensional three-wave-mixing spectroscopy would be of use to shed light on the chiroptical natures of two-exciton states of arbitrary coupled multichromophore systems.


Reappraisal of Human HOG and MO3.13 Cell Lines as a Model to Study Oligodendrocyte Functioning.
Myelination of neuronal axons is essential for proper brain functioning and requires mature myelinating oligodendrocytes (myOLs). The human OL cell lines HOG and MO3.13 have been widely used as in vitro models to study OL (dys) functioning. Here we applied a number of protocols aimed at differentiating HOG and MO3.13 cells into myOLs. However, none of the differentiation protocols led to increased expression of terminal OL differentiation or myelin-sheath formation markers. Surprisingly, the applied protocols did cause changes in the expression of markers for early OLs, neurons, astrocytes and Schwann cells. Furthermore, we noticed that mRNA expression levels in HOG and MO3.13 cells may be affected by the density of the cultured cells. Finally, HOG and MO3.13 co-cultured with human neuronal SH-SY5Y cells did not show myelin formation under several pro-OL-differentiation and pro-myelinating conditions. Together, our results illustrate the difficulty of inducing maturation of HOG and MO3.13 cells into myOLs, implying that these oligodendrocytic cell lines may not represent an appropriate model to study the (dys)functioning of human (my)OLs and OL-linked disease mechanisms.


Anxiety-specific associations with substance use: Evidence of a protective factor in adolescence and a risk factor in adulthood.
Externalizing psychopathology is a strong risk factor for substance use, whereas the role of internalizing manifestations of distress, and anxiety in particular, in predicting substance use remains unclear. Studies have suggested that anxiety may be either a protective or risk factor for substance use. The present study aimed to clarify evidence for anxiety-specific associations with substance use, examining sex and developmental period (adolescence vs. adulthood) as potential moderators that may help explain conflicting results in the literature. In a longitudinal twin sample, cross-sectional associations of anxiety with substance use differed in adolescents and adults and in girls/women and boys/men. Controlling for externalizing psychopathology and depression, anxiety was associated with reduced substance use in adolescent girls and increased substance use in adult women. In contrast, anxiety-specific associations with substance use were not significant in boys and men. Possible explanations for these contrasting results across development and sex are discussed.


Prevalence and correlates of HIV infection among adolescents and young people living in fishing populations along Lake Victoria Fishing Communities in Uganda.
fishing communities in Uganda are key populations for HIV, with persistently higher prevalence and incidence than the general population. between March and August 2014, a cross sectional survey was conducted in 10 fishing communities of Lake Victoria in Uganda. Data was collected on socio-behavioural characteristics using interviewer administered questionnaires and venous blood collected for HIV testing. Prevalent HIV infections among adolescents and young people aged 13 to 24 years was estimated and the factors associated with those infections determined using multi variable logistic regression modelling. HIV prevalence was 10.8% among the 630 (96.5%) who provided a blood sample. Females were 3.5 times as likely to have HIV infection as males (aOR=3.52, 95% CI: 1.34-9.22). Young people aged 20-24 years were twice as likely to be HIV infected as those aged 13-19 years (aOR=1.77, 95% CI: 0.05-2.10), participants without formal education or those who had studied up to primary level were more likely to be HIV infected than those who had post primary education ((aOR=2.45, 95% CI: 1.19-5.07) or (5.29 (1.35-20.71) respectively). Reporting more than one sexual partner in the past 6 months was associated with HIV prevalent infection than those reporting no sexual partners (aOR=6.44, 95% CI: 1.27-32.83). adolescents and young people aged 13-24 years in fishing communities around Lake Victoria, Uganda, have a high HIV prevalence, with females having a three-fold higher level than males. These findings highlight-the need to improve HIV prevention among young females living in these fishing communities.


The GST domain of GDAP1 is a frequent target of mutations in the dominant form of axonal Charcot Marie Tooth type 2K.
Mutations in GDAP1 associate with demyelinating (CMT4A) and axonal (CMT2K) forms of CMT. While CMT4A shows recessive inheritance, CMT2K can present with either recessive (AR-CMT2K) or dominant segregation pattern (AD-CMT2K), the latter being characterised by milder phenotypes and later onset. The majority of the GDAP1 mutations are associated with CMT4A and AR-CMT2K, with only four heterozygous mutations identified in AD-CMT2K. We screened GDAP1 gene in a series of 43 index patients, 39 with CMT2 and 4 with intermediate CMT, with sporadic and familial occurrence of the disease. Three novel mutations were identified in three families with dominant segregation of the disease: two missense changes, p.Arg226Ser and p.Ser34Cys, affecting the GST domain of the GDAP1 protein and a novel deletion (c.23delAG) leading to early truncation of the protein upstream the GST domain. Wide variability in clinical presentation is shared by all three families mostly in terms of age at onset and disease severity. A rare variant p.Gly269Arg, located within the GST domain, apparently acts as phenotype modulator in the family carrying the deletion. The results obtained reveal a GDAP1 mutation frequency of 27% in the dominant families analysed, a figure still unreported for this gene, thus suggesting that GDAP1 involvement in dominant CMT2 might be higher than expected.


Tau oligomers and tau toxicity in neurodegenerative disease.
AD (Alzheimer's disease) is a progressive neurodegenerative disorder characterized by the extracellular accumulation of amyloid β-peptide and the intracellular accumulation of tau. Although there is much evidence linking tau to neurodegeneration, the precise mechanism of tau-mediated neurotoxicity remains elusive. The presence of tau-positive pre-tangle neurons lacking neurofibrillary tangles has been reported in AD brain tissue. In order to study this non-fibrillar tau, we generated a novel monoclonal antibody, named TOC1 (tau oligomeric complex 1), which selectively labels tau dimers and oligomers, but does not label filaments. Time-course analysis and antibody labelling indicates that oligomers appear as an early event in AD pathogenesis. Using a squid axoplasm assay, we have demonstrated that aggregated tau inhibits anterograde FAT (fast axonal transport), whereas monomeric tau has no effect. This inhibition requires a small stretch of N-terminal amino acids termed the PAD (phosphatase-activation domain). Using a PAD-specific antibody, TNT1 (tau N-terminal 1), we demonstrate that PAD exposure is increased in diseased neurons and this leads to an increase in FAT inhibition. Antibody co-labelling with the early-AD marker AT8 indicates that, similar to TOC1, TNT1 expression represents an early event in AD pathogenesis. Finally, the effects of the molecular chaperone Hsp70 (heat-shock protein 70) were also investigated within the squid axoplasm assay. We illustrate that Hsp70 preferentially binds to tau oligomers over filaments and prevents anterograde FAT inhibition observed with a mixture of both forms of aggregated tau. Together, these findings support the hypothesis that tau oligomers are the toxic form of tau in neurodegenerative disease.


Use of angiotensin-converting enzyme inhibitors and angiotensin receptor blockers in high-risk clinical and ethnic groups with diabetes.
Diabetes causes 45% of incident end-stage renal disease (ESRD). Risk of progression is higher in those with clinical risk factors (albuminuria and hypertension), and in ethnic minorities (including blacks, Asians, and Latinos). Angiotensin-converting enzyme inhibitors (ACE) and angiotensin receptor blockers (ARB) slow the progression of diabetic nephropathy, yet little is known about their use among patients at high risk for progression to ESRD. To examine the prevalence of ACE or ARB (ACE/ARB) use overall and within patients with high-risk clinical indications, and to assess for ethnic disparities in ACE/ARB use. Observational cohort study. Kaiser Permanente Northern California (KPNC) Diabetes Registry, a longitudinal registry that monitors quality and outcomes of care for all KPNC patients with diabetes. Individuals (N= 38887) with diabetes who were continuously enrolled with pharmacy benefits during the year 2000, and had self-reported ethnicity data on survey. Pharmacy dispensing of ACE/ARB. Forty-one percent of the cohort had both hypertension and albuminuria, 30% had hypertension alone, and 12% had albuminuria alone. Fourteen percent were black, 11% Latino, 13% Asian, and 63% non-Latino white. Overall, 61% of the cohort received an ACE/ARB. ACE/ARB was dispensed to 74% of patients with both hypertension and albuminuria, 64% of those with hypertension alone, and 54% of those with albuminuria alone. ACE/ARB was dispensed to 61% of whites, 63% of blacks, 59% of Latinos, and 60% of Asians. Among those with albuminuria alone, blacks were significantly (P =.0002) less likely than whites to receive ACE/ARB (47% vs 56%, respectively). No other ethnic disparities were found. In this cohort, the majority of eligible patients received indicated ACE/ARB therapy in 2000. However, up to 45% to 55% of high-risk clinical groups (most notably individuals with isolated albuminuria) were not receiving indicated therapy. Additional targeted efforts to increase use of ACE/ARB could improve quality of care and reduce ESRD incidence, both overall and in high-risk ethnic groups. Policymakers might consider use of ACE/ARB for inclusion in diabetes performance measurement sets.


Photosensitizing properties of palladium-tetraphenylporphycene on cultured tumour cells.
In this study we describe photokilling properties and effects on the mitotic index (MI) of cultured HeLa cells induced by palladium(II)-tetraphenylporphycene (PdTPP(0)). The drug was synthesized by refluxing tetraphenylporphycene (TPP(0)) and PdCl2 in dimethylformamide, followed by evaporation and purification by chromatography. Cells were treated with different concentrations of PdTPPo incorporated into dipalmitoylphosphatidylcholine liposomes, and red light irradiation (lambda > 600 nm) was performed at 21 mW/cm2. No dark toxicity was found when the drug was applied under our experimental conditions. Using lethal (LD(100)) treatments, cells showed the immediate occurrence of bubbles on the plasma membrane, whereas homogeneous nuclear condensation and loss of cytoplasm appeared 3-24 h later. An increased MI was found 6-8 h after sublethal LD(25) and LD(40) treatments as well as a high proportion of abnormal metaphases with altered spindle microtubules. Chromatin condensation and fragmentation were observed 8 h after LD(75) treatments. These results show that in comparison with TPP(0), the new sensitizer PdTPPo has more efficient photokilling properties which could be very valuable for the photodynamic therapy of cancer.


The molecular genetics of holoprosencephaly.
Holoprosencephaly (HPE) has captivated the imagination of Man for millennia because its most extreme manifestation, the single-eyed cyclopic newborn infant, brings to mind the fantastical creature Cyclops from Greek mythology. Attempting to understand this common malformation of the forebrain in modern medical terms requires a systematic synthesis of genetic, cytogenetic, and environmental information typical for studies of a complex disorder. However, even with the advances in our understanding of HPE in recent years, there are significant obstacles remaining to fully understand its heterogeneity and extensive variability in phenotype. General lessons learned from HPE will likely be applicable to other malformation syndromes. Here we outline the common, and rare, genetic and environmental influences on this conserved developmental program of forebrain development and illustrate the similarities and differences between these malformations in humans and those of animal models.


Differential effects of xanthine oxidase inhibition and exercise on albumin concentration in rat tissues.
Albumin is a protein present in almost all kinds of mammalian cells. It has binding sites for several molecules, and possesses antioxidant and other important properties. The aim of this study was to examine the effects of 2 different oxidative stress stimuli - exercise and allopurinol administration - and their combination on albumin concentration in several rat tissues. Samples of soleus, extensor digitorum longus (EDL), and gastrocnemius muscles, and the liver and heart were collected before, immediately after, and 5 h after exercise, and collected at the respective time points after allopurinol administration. Albumin dimmers, markers of oxidative stress, were also assessed in EDL muscle. Albumin concentration increased in the skeletal muscles examined, whereas it decreased in the heart and remained unaffected in the liver after exercise. Allopurinol alone did not affect albumin concentration in any of the tissues. Albumin concentration increased in soleus and EDL muscles, decreased in gastrocnemius muscle and the liver, and remained unaffected in the heart after exercise and allopurinol combination. Albumin dimmers also increased postexercise in EDL muscle. Our findings suggest that the increase in albumin concentration in skeletal muscles may be an antioxidant mechanism response, but may depend on the type of oxidative stress and be stimulation- and tissue-specific.


Correlates of medical and legal help seeking among women reporting intimate partner violence.
To examine the relationship between intimate partner violence (IPV) type, severity, and duration and abused women's use of medical and legal services. Participants were 1509 randomly sampled women from a large health plan who were interviewed by telephone to assess (1) self-reported lifetime exposure to IPV type (physical, sexual, and psychological), severity, and duration and (2) women's use of medical and legal services (civil protection orders). Compared with women who experienced psychological abuse only, sexually abused women were 1.3 times as likely to seek medical care, and women exposed to physical IPV or sexual IPV were 3.2 times and 1.6 times as likely, respectively, to seek legal services. Rates of medical and legal help seeking increased with increasing abuse severity among physically abused women, and rates of legal help seeking increased with abuse severity among sexually and psychologically abused women. Longer duration of physical and sexual IPV was also found to be associated with increased legal help seeking. Abused women who sought formal help were more likely to be exposed to physical or sexual IPV, severe psychological IPV, and severe and long-lasting physical and sexual IPV. Efforts should be considered to improve the healthcare and legal system's response to IPV.


Regional musculoskeletal conditions: pain in the forearm, wrist and hand.
Pain in the forearm, wrist or hand may arise from one of several discrete rheumatic disorders of soft tissues, such as tenosynovitis, or from a non-specific regional pain syndrome. Symptoms are prevalent in the general population and both patterns of illness are well represented. Many epidemiological investigations of prevalence, incidence, causal risk factors, management and prevention exist, although surveys have used a wide variety of case definitions, hampering comparisons. Improved standardized approaches to classification are in prospect and these are described. A synthesis is also attempted of the main findings of existing surveys. A growing body of evidence now links distal arm pain with physical risk factors in the workplace (e.g. repetition, force, duration, short cycle time and awkwardness of posture); some possible ergonomic solutions to occupational arm pain are discussed. But occupational and psychosocial factors are also linked with symptom reporting and disability, and their role in pathogenesis may be important in primary prevention and the management of recalcitrant cases. Some key research questions are proposed aimed at preventing chronicity and disablement from arm pain.