Bioactive Screening Library

As the L4F peptide was developed to mimic apoA-I function, and this appears to be borne out by our in vitro studies, we have compared apoA-I overexpression with L4F treatment in several pathophysiological endpoints in Ldlr male mice fed the HFHSC diet. The effect of apoA-I overexpression on insulin resistance in the Ldlr DIO model has not been reported on previously, and thus an important dataset for comparison to L4F treatment. Provided chronic inflammation which is associated with obesity related insulin resistance, our in vitro findings suggest that L4F and apoA-I overexpression could potentially improve GSK1120212 glucose tolerance in an in vivo model of DIO. In our previous study apoA-I overexpression relieved some of the adipose tissue inflammation. However, in the present study L4F treatment of 100 mg/day/ mouse had no impact on adipose tissue or systemic inflammatory markers. Further, neither the overexpression of apoA-I nor treatment with L4F had an impact on glucose tolerance nor glucose stimulated insulin levels. Note that neither apoAI overexpression nor L4F treatment had an impact on weight gain or adiposity in this model, confirmed in the peptide study by body composition analysis. ApoA-I and L4F mimetics are often cited for their anti-atherosclerotic properties. This effect of apoA-I overexpression in Ldlr mice fed a high fat diet has been documented and confirmed in the present study. Yet, L4F treatment had no such effect at a dosage of 100 mg/day/ mouse. It is striking that despite the dramatic effect of L4F on 3T3L1 cells, the peptide is without influence on the measured parameters studied in vivo at 100 mg/day/mouse. One limitation of this study is that the absence of any in vivo effect of peptide treatment raises questions about the efficacy of the dose and route of administration of the peptide. We have followed protocols and dosages that have been shown in apoE and ob/ob mice to be effective. The dose we employed, is equal to the effective dose of 4.5 mg/kg/day shown by Navab and colleagues to reduce plasma SAA levels. Importantly, they have found the dose was determining rather than the concentration in the plasma. This group has further suggested that the primary function of the peptides is to attenuate the oxidized lipid and unsaturated lysophosphatidic acid concentrations in the intestine. The current study utilized the L version of the peptide injected subcutaneously as it is not stable for oral delivery while other studies have used the D version of the peptide which is stable for oral delivery. However, data suggests that neither the route of administration nor the enantiomer version of the peptide determines effectiveness. One study directly compared L4F versus D4F delivered subcutaneously in rabbits and found similar reductions in atherosclerosis. While studies suggest the site of action is primarily in the intestine, Navab and colleagues have shown D4F is similarly effective when delivered subcutaneously or orally and equivalent amounts appear in the feces. L4F delivered orally with niclosamide was equally effective in reducing SAA in apoE mice as L4F delivered subcutaneously. This suggests that the effect of 4F peptide on inflammation and atherosclerosis is independent of route of administration and the enantiomer version of the peptide, and thus does not explain the lack of effect in the current study. Given the above discussion, it is possible that either L4F does not influence obesity, inflammation, or atherosclerosis in this model or that a higher dose of peptide is required to affect these outcomes in this model. In addition, we have no information on the role of oxidized lipids in the intestine in our model. Further work is needed to address the effectiveness of 4F p.

That said, systematic reviewers are usually familiar with their field and a priori aware of potentially eligible clinical trials and/or how the outcome in question is frequently measured. Complete prespecification also could improve efficiency in data abstraction and analysis during a systematic review. We assume that primary outcomes for both clinical trials and systematic reviews are chosen based on perceived clinical importance and/or importance to patients; and that they are usually measured and reported more thoroughly than nonprimary outcomes. Not surprisingly, in our study, primary outcomes were more completely specified than other outcome types. Our estimate of 94.7% protocols pre-specifying a primary outcome is somewhat higher than the 88% that has been reported as pre-specified in clinical trial protocols, and this could be related to the fact that we were examining protocols entered into software that requests the domain names of the pre-specified outcomes. In our study, the most incompletely pre-specified outcome was quality-of-life, a key patient-important outcome. This finding is concordant with other studies that have found that outcome reporting in clinical trials is a bigger problem for patient-important outcomes than other types of outcomes. Further, when patient-important outcomes are not primary outcomes in clinical trials, the likelihood that reporting is complete is further reduced. Our study aimed to evaluate the completeness and comparability of all outcomes, both patient-important and not. Our recommendation is that systematic reviewers should engage in discussion about and strongly consider pre-specifying all five elements of each outcome they wish to examine. When explicit pre-specification of all five elements of a given outcome is not possible, for example when all possible DAPT options for a given outcome element are not known or are too numerous, the systematic reviewers should enumerate all known acceptable options for each element and explicitly state that all options for that element would be accepted, or provide rationale for why it is impossible to completely pre-specify an element. The Preferred Reporting Items for Systematic Reviews and Meta-analyses Protocols is currently under development. We hope that the availability of reporting guidelines will improve the completeness of specification of outcomes. Assuming that the Cochrane Collaboration recognizes the importance of completeness of pre-specification, there are some possible ways to ensure that review authors are aware of the five elements of a completely specified outcome. First, editorial teams at Cochrane Review Groups should make all review authors aware of the five outcome elements early in the process. Second, peer reviewers should be directed to consider whether the outcomes are completely pre-specified and not likely to have been chosen based on the strength and direction of the findings for those outcomes. Third, the Cochrane Handbook and other systematic review guidance materials, in addition to training workshops and other educational avenues, should incorporate explicit descriptions of all five outcome elements. Other organizations producing guidance on systematic review methodology should also incorporate descriptions of the five outcome elements in their guidance materials. Organizations such as the Cochrane Collaboration suggest limiting the number of outcomes examined in a systematic review. However, in order to evaluate whether the effect of an intervention persists over time, an otherwise identical outcome is often measured at a number of time-points. For the purpose of counting the number of outcomes measured.

To evaluate the prognostic performance of the 13 miRNAs differentially expressed in grade II as compared with grade III+IV gliomas by RT-qPCR, we used the TCGA glioma datasets. This analysis demonstrated an association between increased expression and GBM prognosis for four miRNAs: miR155, miR-21, miR-210, and miR-22. Interestingly, these miRNAs belong to the subgroup of 9 miRNAs differentiating grade II gliomas from grade III+IV tumours in the Venn diagram generated by the analysis of our microarray data, further supporting their biological relevance. The examination of grade III tumours from the LGG dataset identified three miRNAs able to predict patients’ survival. Increased miR-155 expression was associated with worse OS in both grade III and grade IV gliomas. Whereas, increased miR-383 expression was associated with a lower risk of death only in grade III tumours. miR-1296 higher expression levels were also found associated with better prognosis in grade III gliomas but no information were available about its prognostic correlation with GBMs, since it was not included on the Agilent platform used to analyze the GBM dataset. As shown in Table S7, the 6 miRNAs associated with prognosis in this study are involved in all steps of cancer development and progression. We focused our attention on miR-21 and miR-210 because they were the only miRNAs associated with survival in both univariable and multivariable analyses in the GBM dataset and play a pivotal role in hypoxia-related pathways. Indeed, the main histopathological characteristics distinguishing GBMs from lower grade astrocytomas are the presence of foci of necrosis surrounded by hypercellular regions, and microvascular hyperplasia that are all hallmarks of hypoxia. This biological phenomenon has been related to aggressiveness and infiltrative behaviour in a wide variety of solid tumours. In GBM, gene expression profiles on microdissected cellular zones surrounding necrotic foci have revealed up regulation of HIF-1a, one of hypoxia-inducible factors mediating cancer cell adaptation to a hypoxic tumour environment. Overexpression of miR-21 in DU145 cells increases the expression of HIF-1a and VEGF and promotes tumor angiogenesis. Interestingly, Hermansen et al demonstrated by ISH analysis miR-21 expression in glioma and endothelial cells within the tumor, whereas no expression was detected in non-neoplastic blood vessels. These results are consistent with the hypothesis that miR-21 increased expression is related to tumor angiogenesis in gliomas. On the other side, miR-210 is ABT-263 robustly induced by hypoxia in many cell lines through HIF-1a. This activation determines downregulation of a number of genes which promote cancer cell survival and adaptation to adverse microenvironment, leading to the development of a more resistant and aggressive cell subpopulations. The downregulation of COX10 produces repression of electron transport chain to decrease oxygen-demand, and enhances anaerobic production of ATP via glicolysis enabling survival to hypoxic insults. Upregulation of miR-210 also causes repression of c-Myc inhibitor, MNT, promoting cell cycle and growth. It also contributes to maintenance of high HIF-1a levels during hypoxia, by suppressing of ISCU, SDHS, NUDFA4 proteins, in a feed-forward loop which sustains HIF-1a and its triggered processes. In addition hypoxia-induced miR-210 up-regulation seems to be able to directly enhance tumour angiogenesis by targeting EphrinA3 and ROD1.

Eukaryote chromosome integrity and stability since the telomeric DNA repeats and associated proteins prevent chromosome end-to-end TWS119 fusions and exonuclease degradation. The length of telomeres is a critical determinant of a cell’s replicative life span and telomere shortening has been linked to cell senescence, disease and ageing. Telomeres shorten with each cell division due to the so called end-replication problem, resulting in loss of telomeric repeats. The loss of telomeric repeats is compensated for by synthesis of new repeats. The enzyme responsible for synthesis of new repeats is the telomerase holoenzyme, which is an RNA-dependent DNA polymerase complex. This enzyme synthesises new tandem telomeric repeats de novo at the 39 chromosome strand end. Telomerase-negative cells experience telomere shortening and lose on average between 30 and 200 bp of telomeric sequence per cell division, a loss which is ultimately lethal. Telomerase consists of several subunits. The core subunit consists of Telomerase Reverse Transcriptase together with its conserved RNA component, which acts as the template for the synthesis of telomeric repeats. Telomere structure is dependent on multiple telomereassociated proteins and these proteins together with telomeric DNA form the so-called telosome. TAPs are part of the telomerase-mediated telomere maintenance and regulation mechanisms, including telomere loop formation. The gene encoding TERT has been deleted or mutated in a number of organisms which led to cell senescence and eventual cell death. The genome of malaria parasites is arranged into 14 linear chromosomes which contain telomeres consisting of 7-bp telomeric repeat sequences. The average telomere length per species varies, ranging from 850 bp to 6700 bp in the human parasites P. falciparum and P. vivax, respectively. Plasmodium telomere length appears to be kept constant during the erythrocytic cycle The genomes of different Plasmodium parasites contain a single copy tert gene. For TERT of P. falciparum it has been demonstrated that it is capable of de novo synthesis of telomeric repeats both to the 39 telomeric overhang and to non-telomeric 39 ends, thus contributing not only to telomere maintenance but also to adding new telomeric sequences to broken chromosomes. Telomerase activity in P. falciparum is detectable both in late stage trophozoites and schizonts, stages where DNA replication occurs. The RNA component of telomerase has been identified in silico in several Plasmodium species based on structural comparisons of conserved domains in TR domains from other organisms. The telomerase RNA is detectable in all erythrocytic stages and the ookinete stage of P. falciparum according to the PlasmoDB expression data. In order to analyse the importance of telomere metabolism/ dynamics for asexual blood stage multiplication of Plasmodium parasites we have attempted to generate mutants that lack expression of TERT. We have used the rodent malaria parasite P. berghei because of the high efficiency of transfection and rapid selection of gene-deletion mutants, which might be essential when TERT-deficient parasites show a delayed death phenotype as has been shown for other organisms. We found that we can target the P. berghei tert gene for gene deletion. However, our results also indicate that TERT is an essential enzyme for survival of P. berghei blood stages since we were unable to clone and propagate TERT-deficient parasites. This study reports an analysis of telomeres and telomerase of the rodent malaria parasite P. berghei and the unsuccessful attempts to isolate gene-deletion mutants lacking the gene encoding telomerase reverse transcriptase.

Therefore, a more reliable and sensitive molecular assay is needed to accurately diagnose mTBI. In this experiment, acute phase miRNA changes in the serum were measured to determine the feasibility of using miRNA as diagnostic markers of mTBI. Increasing grades of mTBI were induced by a weight drop device by increasing the height and weight of the falling rod. Injuries were characterized by the sensory motor responses designed to model clinical neurological examination and indicated the injuries to be within the mild spectrum of TBI. Consistent with mTBI, histological analysis of the brain showed no gross pathological changes following the injury. Behavior impairments such as depression and loss of interest have been described in patients with mTBI. Reduced escape behavior, measured in terms of vertical activity of the animal in an open field apparatus, can be a measure of depression-related behavior. Transient, but significant depression-related behavior in terms of reduced vertical activity was observed in the injured animals at day 1 post injury, which recovered over time. A general negative impact on the overall health of the animals was also observed at day 1 post injury, indicated by the reduction in horizontal activity of the injured animals. Sensory gating impairments have also been reported in mTBI. Increase in the OFL activity of the animals at the later time points of day 14 and day 30 as compared to day 1 activity was interesting. We hypothesize that the naı¨ve and sham control animals remember being in the chambers and that the other animals, due to injury, may not remember the chamber as well and spend more time exploring. ASR is a VE-821 structure complex behavior that engages higher brain functions and is related to information processing. A significantly reduced startle response was observed in animals with injury, such that the greater the injury severity, the less startle response was observed. The parietal lobe, where the injury was centered, is involved in the sensorimotor information processing. It is suggested that the right inferior parietal lobe plays a role in auditory signal processing and integration of sensory and motor functions. This observation indicates the diffuse nature of the injury that may also affect central brain functions. Many individuals, who suffer with mTBI do not exhibit neurobehavioral alterations and acute symptoms of mTBI, such as headaches, sleep disturbance and depression or recover spontaneously from the injury. A similar observation was made in the current experiment, where the percent of animals’ OFL activity in the mildest of mild injury group was similar to sham and naı¨ve control groups. Other groups, with a more severe form of the mild injury, however, showed greater reduction in the percent activity than the sham and naı¨ve groups. MiRNA changes in the serum have been suggested as a potential marker of disease and injury. MiRNA modulations in the serum of TBI patients have been reported.