Search

In an era marked by pervasive digital connectivity, cybersecurity concerns have escalated. The rapid evolution of technology has led to a spectrum of cyber threats, including sophisticated zero-day attacks. This research addresses the challenge of existing intrusion detection systems in identifying zero-day attacks using the CIC-MalMem-2022 dataset and autoencoders for anomaly detection. The trained autoencoder is integrated with XGBoost and Random Forest, resulting in the models XGBoost-AE and Random Forest-AE. The study demonstrates that incorporating an anomaly detector into traditional models significantly enhances performance. The Random Forest-AE model achieved 100% accuracy, precision, recall, F1 score, and Matthews Correlation Coefficient (MCC), outperforming the methods proposed by Balasubramanian et al., Khan, Mezina et al., Smith et al., and Dener et al. When tested on unseen data, the Random Forest-AE model achieved an accuracy of 99.9892%, precision of 100%, recall of 99.9803%, F1 score of 99.9901%, and MCC of 99.8313%. This research highlights the effectiveness of the proposed model in maintaining high accuracy even with previously unseen data., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Dai et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Motivation: Many real applications such as businesses and health generate large categorical datasets with uncertainty. A fundamental task is to efficiently discover hidden and non-trivial patterns from such large uncertain categorical datasets. Since the exact value of an attribute is often unknown in uncertain categorical datasets, conventional clustering analysis algorithms do not provide a suitable means for dealing with categorical data, uncertainty, and stability., Problem Statement: The ability of decision making in the presence of vagueness and uncertainty in data can be handled using Rough Set Theory. Though, recent categorical clustering techniques based on Rough Set Theory help but they suffer from low accuracy, high computational complexity, and generalizability especially on data sets where they sometimes fail or hardly select their best clustering attribute., Objectives: The main objective of this research is to propose a new information theoretic based Rough Purity Approach (RPA). Another objective of this work is to handle the problems of traditional Rough Set Theory based categorical clustering techniques. Hence, the ultimate goal is to cluster uncertain categorical datasets efficiently in terms of the performance, generalizability and computational complexity., Methods: The RPA takes into consideration information-theoretic attribute purity of the categorical-valued information systems. Several extensive experiments are conducted to evaluate the efficiency of RPA using a real Supplier Base Management (SBM) and six benchmark UCI datasets. The proposed RPA is also compared with several recent categorical data clustering techniques., Results: The experimental results show that RPA outperforms the baseline algorithms. The significant percentage improvement with respect to time (66.70%), iterations (83.13%), purity (10.53%), entropy (14%), and accuracy (12.15%) as well as Rough Accuracy of clusters show that RPA is suitable for practical usage., Conclusion: We conclude that as compared to other techniques, the attribute purity of categorical-valued information systems can better cluster the data. Hence, RPA technique can be recommended for large scale clustering in multiple domains and its performance can be enhanced for further research., Competing Interests: The authors have declared that no competing interests exist.

Background: It remains unclear as to whether polycystic ovary syndrome (PCOS) is an additional risk factor in the development of left ventricular (LV) hypertrophy in obese women. In the current study, we provide clarity on this issue by rigorously analysing patient LV geometry beyond the basic clinical measures currently used. Importantly, the cohort contained only normotensive patients that would normally be deemed low risk with no further intervention required., Methods: The study comprised 24 obese women with PCOS and 29 obese Control women. Transthoracic echocardiography was used to evaluate LV structure/function. Basic clinical and metabolic data were collected for each participant consisting of age, BMI, blood pressure, fasting glucose, LDL-C, HLD-C, cholesterol and triglyceride levels. Exclusion criteria; BMI < 30 g/m2, type 2 diabetes, hypertension., Results: Both groups exhibited concentric remodelling of the LV posterior wall at a prevalence of ~20%, this associated with grade 1 diastolic dysfunction. Estimated LV mass/height2.7 was increased patients with PCOS (45 ± 2.2 vs 37 ± 1.6) with 33% exhibiting LV mass/height2.7 above ASE guidelines, compared to 7% in Controls. Furthermore, 25% of patients with PCOS were characterised with concentric hypertrophy, an alteration in LV geometry that was not observed in the Control group., Conclusions: To our knowledge, this is the first study to assess LV geometric patterns in obese women with PCOS. The results suggest that obese women with PCOS are at greater risk of concentric hypertrophy than obese only women and provide justification for additional cardiovascular risk assessment in normotensive obese/PCOS women., Competing Interests: he authors have declared that no competing interests exist.

Background: Writing and digital storage have largely replaced organic memory for encoding and retrieval of information in the modern era, with a corresponding decrease in emphasis on memorization in Western education. In health professional training, however, there remains a large corpus of information for which memorization is the most efficient means of ensuring: A) that the trainee has the required information readily available; and B) that a foundation of knowledge is laid, upon which the medical trainee builds multiple, complex layers of detailed information during advanced training. The carefully staged progression in early- to late- years' medical training from broad concepts (e.g. gross anatomy and pharmacology) to in-depth, specialised disciplinary knowledge (e.g. surgical interventions and follow-on care post-operatively) has clear parallels to the progression of training and knowledge exposure that Australian Aboriginal youths undergo in their progression from childhood to adulthood to Tribal Elders., Methods: As part of the Rural Health curriculum and the undergraduate Nutrition and Dietetics program in the Monash University Faculty of Medicine, Nursing, and Health Sciences, we tested Australian Aboriginal techniques of memorization for acquisition and recall of novel word lists by first-year medical students (N = 76). We also examined undergraduate student evaluations (N = 49) of the use of the Australian Aboriginal memory technique for classroom study of foundational biomedical knowledge (the tricarboxylic acid cycle) using qualitative and quantitative analytic methods drawing from Bloom's taxonomy for orders of thinking and learning. Acquisition and recall of word lists were assessed without memory training, or after training in either the memory palace technique or the Australian Aboriginal narrative technique., Results: Both types of memory training improved the number of correctly recalled items and reduced the frequency of specific error types relative to untrained performance. The Australian Aboriginal method resulted in approximately a 3-fold greater probability of improvement to accurate recall of the entire word list (odds ratio = 2.82; 95% c.i. = 1.15-6.90), vs. the memory palace technique (odds ratio = 2.03; 95% c.i. = 0.81-5.06) or no training (odds ratio = 1.5; 95% c.i. = 0.54-4.59) among students who did not correctly recall all list items at baseline. Student responses to learning the Australian Aboriginal memory technique in the context of biomedical science education were overwhelmingly favourable, and students found both the training and the technique enjoyable, interesting, and more useful than rote memorization. Our data indicate that this method has genuine utility and efficacy for study of biomedical sciences and in the foundation years of medical training., Competing Interests: The authors have declared that no competing interests exist.

Neural spike sorting is prerequisite to deciphering useful information from electrophysiological data recorded from the brain, in vitro and/or in vivo. Significant advancements in nanotechnology and nanofabrication has enabled neuroscientists and engineers to capture the electrophysiological activities of the brain at very high resolution, data rate and fidelity. However, the evolution in spike sorting algorithms to deal with the aforementioned technological advancement and capability to quantify higher density data sets is somewhat limited. Both supervised and unsupervised clustering algorithms do perform well when the data to quantify is small, however, their efficiency degrades with the increase in the data size in terms of processing time and quality of spike clusters being formed. This makes neural spike sorting an inefficient process to deal with large and dense electrophysiological data recorded from brain. The presented work aims to address this challenge by providing a novel data pre-processing framework, which can enhance the efficiency of the conventional spike sorting algorithms significantly. The proposed framework is validated by applying on ten widely used algorithms and six large feature sets. Feature sets are calculated by employing PCA and Haar wavelet features on three widely adopted large electrophysiological datasets for consistency during the clustering process. A MATLAB software of the proposed mechanism is also developed and provided to assist the researchers, active in this domain., Competing Interests: The authors have declared that no competing interests exist.

Ceramics are quintessential indicators of human culture and its evolution across generations of social learners. Cultural transmission and evolution theory frequently emphasizes apprentices' need for accurate imitation (high-fidelity copying) of their mentors' actions. However, the ensuing prediction of standardized fashioning patterns within communities of practice has not been directly addressed in handicraft traditions such as pottery throwing. To fill this gap, we analysed variation in vessel morphogenesis amongst and within traditional potters from culturally different workshops producing for the same market. We demonstrate that, for each vessel type studied, individual potters reliably followed distinctive routes through morphological space towards a much-less-variable common final shape. Our results indicate that mastering the pottery handicraft does not result from accurately reproducing a particular model behaviour specific to the community's cultural tradition. We provide evidence that, at the level of the elementary clay-deforming gestures, individual learning rather than simple imitation is required for the acquisition of a complex motor skill such as throwing pottery., Competing Interests: The authors have declared that there are no competing interests.

In the last decade, house sparrow populations have shown a general decline, especially in cities. Avian malaria has been recently suggested as one of the potential causes of this decline, and its detrimental effects could be exacerbated in urban habitats. It was initially thought that avian malaria parasites would not have large negative effects on wild birds because of their long co-evolution with their hosts. However, it is now well-documented that they can have detrimental effects at both the primo- and chronical infection stages. In this study, we examined avian malaria infection and its physiological and morphological consequences in four populations of wild house sparrows (2 urban and 2 rural). We did not find any relationship between the proportions of infected individuals and the urbanisation score calculated for our populations. However, we observed that the proportion of infected individuals increased during the course of the season, and that juveniles were less infected than adults. We did not detect a strong effect of malaria infection on physiological, morphological and condition indexes. Complex parasite dynamics and the presence of confounding factors could have masked the potential effects of infection. Thus, longitudinal and experimental studies are needed to understand the evolutionary ecology of this very common, but still poorly understood, wild bird parasite., Competing Interests: The authors have declared that no competing interests exist.

Pathogens pose a major risk to wild host populations, especially in the face of ongoing biodiversity declines. Beak and feather disease virus (BFDV) can affect most if not all members of one of the largest and most threatened bird orders world-wide, the Psittaciformes. Signs of disease can be severe and mortality rates high. Its broad host range makes it a risk to threatened species in particular, because infection can occur via spill-over from abundant hosts. Despite these risks, surveillance of BFDV in locally abundant wild host species has been lacking. We used qPCR and haemagglutination assays to investigate BFDV prevalence, load and shedding in seven abundant host species in the wild in south-east Australia: Crimson Rosellas (Platycercus elegans), Eastern Rosellas (Platycercus eximius), Galahs (Eolophus roseicapillus), Sulphur-crested Cockatoos (Cacatua galerita), Blue-winged Parrots (Neophema chrysostoma), Rainbow Lorikeets (Trichoglossus moluccanus) and Red-rumped Parrots (Psephotus haematonotus). We found BFDV infection in clinically normal birds in six of the seven species sampled. We focused our analysis on the four most commonly caught species, namely Crimson Rosellas (BFDV prevalence in blood samples: 41.8%), Sulphur-crested Cockatoos (20.0%), Blue-winged Parrots (11.8%) and Galahs (8.8%). Species, but not sex, was a significant predictor for BFDV prevalence and load. 56.1% of BFDV positive individuals were excreting BFDV antigen into their feathers, indicative of active viral replication with shedding. Being BFDV positive in blood samples predicted shedding in Crimson Rosellas. Our study confirms that BFDV is endemic in our study region, and can inform targeted disease management by providing comparative data on interspecies variation in virus prevalence, load and shedding., Competing Interests: The authors have declared that no competing interests exist.

Background: Optical measurement techniques and recent advances in wearable technology have made heart rate (HR) sensing simpler and more affordable., Objectives: The Polar OH1 is an arm worn optical heart rate monitor. The objectives of this study are two-fold; 1) to validate the OH1 optical HR sensor with the gold standard of HR measurement, electrocardiography (ECG), over a range of moderate to high intensity physical activities, 2) to validate wearing the OH1 at the temple as an alternative location to its recommended wearing location around the forearm and upper arm., Methods: Twenty-four individuals participated in a physical exercise protocol, by walking on a treadmill and riding a stationary spin bike at different speeds while the criterion measure, ECG and Polar OH1 HR were recorded simultaneously at three different body locations; forearm, upper arm and the temple. Time synchronised HR data points were compared using Bland-Altman analyses and intraclass correlation., Results: The intraclass correlation between the ECG and Polar OH1, for the aggregated data, was 0.99 and the estimated mean bias ranged 0.27-0.33 bpm for the sensor locations. The three sensors exhibited a 95% limit of agreement (LoA: forearm 5.22, -4.68 bpm; upper arm 5.15, -4.49; temple 5.22, -4.66). The mean of the ECG HR for the aggregated data was 112.15 ± 24.52 bpm. The intraclass correlation of HR values below and above this mean were 0.98 and 0.99 respectively. The reported mean bias ranged 0.38-0.47 bpm (95% LoA: forearm 6.14, -5.38 bpm; upper arm 6.07, -5.13 bpm; temple 6.09, -5.31 bpm), and 0.15-0.16 bpm (95% LoA: forearm 3.99, -3.69 bpm; upper arm 3.90, -3.58 bpm; temple 4.06, -3.76 bpm) respectively. During different exercise intensities, the intraclass correlation ranged 0.95-0.99 for the three sensor locations. During the entire protocol, the estimated mean bias was in the range -0.15-0.55 bpm, 0.01-0.53 bpm and -0.37-0.48 bpm, for the forearm, upper arm and temple locations respectively. The corresponding upper limits of 95% LoA were 3.22-7.03 bpm, 3.25-6.82 bpm and 3.18-7.04 bpm while the lower limits of 95% LoA were -6.36-(-2.35) bpm, -6.46-(-2.30) bpm and -7.42-(-2.41) bpm., Conclusion: Polar OH1 demonstrates high level of agreement with the criterion measure ECG HR, thus can be used as a valid measure of HR in lab and field settings during moderate and high intensity physical activities., Competing Interests: The authors have declared that no competing interests exist.

Developing an optimal heterotrophic feeding regime has the potential to improve captive coral growth and health. This study evaluated the efficacy of three exogenous diets: Artemia nauplii (ART), a commercially available coral diet (Reef Roids) (RR), and a novel, micro-bound diet (ATF), against a comparatively natural, unfiltered seawater treatment (RAW), and an unfed, ultra-filtered seawater treatment (CTL), in adult Acropora millepora and Pocillopora acuta nubbins. After 90 days, both species showed significantly positive weight gain in response to one treatment (A. millepora-RAW, P. acuta-ART), and comparatively low growth in response to another (A. millepora-ATF, P. acuta-RR). The results highlighted substantial differences in the nutritional requirements between species. The nutritional composition of A. millepora in the best performing treatment was dominated by high-energy materials such as storage lipids and saturated and monounsaturated fatty acids. In contrast, the P. acuta nutritional profile in the superior treatment showed a predominance of structural materials, including protein, phospholipids, and polyunsaturated fatty acids. This study demonstrates that Artemia nauplii can successfully replace a natural feeding regime for captive P. acuta, yet highlights the considerable work still required to optimise supplementary feeding regimes for A. millepora., Competing Interests: The authors have declared that no competing interests exist.

Although Radio Frequency Identification (RFID) is poised to displace barcodes, security vulnerabilities pose serious challenges for global adoption of the RFID technology. Specifically, RFID tags are prone to basic cloning and counterfeiting security attacks. A successful cloning of the RFID tags in many commercial applications can lead to many serious problems such as financial losses, brand damage, safety and health of the public. With many industries such as pharmaceutical and businesses deploying RFID technology with a variety of products, it is important to tackle RFID tag cloning problem and improve the resistance of the RFID systems. To this end, we propose an approach for detecting cloned RFID tags in RFID systems with high detection accuracy and minimal overhead thus overcoming practical challenges in existing approaches. The proposed approach is based on consistency of dual hash collisions and modified count-min sketch vector. We evaluated the proposed approach through extensive experiments and compared it with existing baseline approaches in terms of execution time and detection accuracy under varying RFID tag cloning ratio. The results of the experiments show that the proposed approach outperforms the baseline approaches in cloned RFID tag detection accuracy.

The ability of Plasmodium parasites to egress from their host red blood cell is critical for the amplification of these parasites in the blood. Previous forward chemical genetic approaches have implicated the subtilisin-like protease (SUB1) and the cysteine protease dipeptidyl aminopeptidase 3 (DPAP3) as key players in egress, with the final step of SUB1 maturation thought to be due to the activity of DPAP3. In this study, we have utilized a reverse genetics approach to engineer transgenic Plasmodium falciparum parasites in which dpap3 expression can be conditionally regulated using the glmS ribozyme based RNA-degrading system. We show that DPAP3, which is expressed in schizont stages and merozoites and localizes to organelles distinct from the micronemes, rhoptries and dense granules, is not required for the trafficking of apical proteins or processing of SUB1 substrates, nor for parasite maturation and egress from red blood cells. Thus, our findings argue against a role for DPAP3 in parasite egress and indicate that the phenotypes observed with DPAP3 inhibitors are due to off-target effects.

The cerebellum processes pain inputs and is important for motor learning. Yet, how the cerebellum interacts with the motor cortex in individuals with recurrent pain is not clear. Functional connectivity between the cerebellum and motor cortex can be measured by a twin coil transcranial magnetic stimulation technique in which stimulation is applied to the cerebellum prior to stimulation over the motor cortex, which inhibits motor evoked potentials (MEPs) produced by motor cortex stimulation alone, called cerebellar inhibition (CBI). Healthy individuals without pain have been shown to demonstrate reduced CBI following motor acquisition. We hypothesized that CBI would not reduce to the same extent in those with mild-recurrent neck pain following the same motor acquisition task. We further hypothesized that a common treatment for neck pain (spinal manipulation) would restore reduced CBI following motor acquisition. Motor acquisition involved typing an eight-letter sequence of the letters Z,P,D,F with the right index finger. Twenty-seven neck pain participants received spinal manipulation (14 participants, 18-27 years) or sham control (13 participants, 19-24 years). Twelve healthy controls (20-27 years) also participated. Participants had CBI measured; they completed manipulation or sham control followed by motor acquisition; and then had CBI re-measured. Following motor acquisition, neck pain sham controls remained inhibited (58 ± 33% of test MEP) vs. healthy controls who disinhibited (98 ± 49% of test MEP, P<0.001), while the spinal manipulation group facilitated (146 ± 95% of test MEP, P<0.001). Greater inhibition in neck pain sham vs. healthy control groups suggests that neck pain may change cerebellar-motor cortex interaction. The change to facilitation suggests that spinal manipulation may reverse inhibitory effects of neck pain.

Maternal consumption of a high fat diet during early development has been shown to impact the formation of hypothalamic neurocircuitry, thereby contributing to imbalances in appetite and energy homeostasis and increasing the risk of obesity in subsequent generations. Early in postnatal life, the neuronal projections responsible for energy homeostasis develop in response to appetite-related peptides such as leptin. To date, no study characterises the genome-wide transcriptional changes that occur in response to exposure to high fat diet during this critical window. We explored the effects of maternal high fat diet consumption on hypothalamic gene expression in Sprague Dawley rat offspring at postnatal day 10. RNA-sequencing enabled discovery of differentially expressed genes between offspring of dams fed a high fat diet and offspring of control diet fed dams. Female high fat diet offspring displayed altered expression of 86 genes (adjusted P-value<0.05), including genes coding for proteins of the extra cellular matrix, particularly Collagen 1a1 (Col1a1), Col1a2, Col3a1, and the imprinted Insulin-like growth factor 2 (Igf2) gene. Male high fat diet offspring showed significant changes in collagen genes (Col1a1 and Col3a1) and significant upregulation of two genes involved in regulation of dopamine availability in the brain, tyrosine hydroxylase (Th) and dopamine reuptake transporter Slc6a3 (also known as Dat1). Transcriptional changes were accompanied by increased body weight, body fat and body length in the high fat diet offspring, as well as altered blood glucose and plasma leptin. Transcriptional changes identified in the hypothalamus of offspring of high fat diet mothers could alter neuronal projection formation during early development leading to abnormalities in the neuronal circuitry controlling appetite in later life, hence priming offspring to the development of obesity.

The establishment and early colonisation of the gastrointestinal (GI) tract has been recognised as a crucial stage in chick development, with pioneering microbial species responsible for influencing the development of the GI tract and influencing host health, fitness and disease status throughout life. Development of the microbiota in long lived seabirds is poorly understood. This study characterised the microbial composition of little penguin and short-tailed shearwater chicks throughout development, using Quantitative Real Time PCR (qPCR) and 16S rRNA sequencing. The results indicated that microbial development differed between the two seabird species with the short-tailed shearwater microbiota being relatively stable throughout development whilst significant fluctuations in the microbial composition and an upward trend in the abundance of Firmicutes and Bacteroidetes were observed in the little penguin. When the microbial composition of adults and chicks was compared, both species showed low similarity in microbial composition, indicating that the adult microbiota may have a negligible influence over the chick's microbiota.

Increasing consumption of a high fat 'Western' diet has led to a growing number of pregnancies complicated by maternal obesity. Maternal overnutrition and obesity have health implications for offspring, yet little is known about their effects on offspring kidney development and renal function. Female C57Bl6 mice were fed a high fat diet (HFD, 21% fat) or matched normal fat diet (NFD, 6% fat) for 6 weeks prior to pregnancy and throughout gestation and lactation. HFD dams were overweight and glucose intolerant prior to mating but not in late gestation. Offspring of NFD and HFD dams had similar body weights at embryonic day (E)15.5, E18.5 and at postnatal day (PN)21. HFD offspring had normal ureteric tree development and nephron number at E15.5. However, using unbiased stereology, kidneys of HFD offspring were found to have 20-25% more nephrons than offspring of NFD dams at E18.5 and PN21. Offspring of HFD dams with body weight and glucose profiles similar to NFD dams prior to pregnancy also had an elevated nephron endowment. At 9 months of age, adult offspring of HFD dams displayed mild fasting hyperglycaemia but similar body weights to NFD offspring. Renal function and morphology, measured by transcutaneous clearance of FITC-sinistrin and stereology respectively, were normal. This study demonstrates that maternal fat feeding augments offspring nephron endowment with no long-term consequences for offspring renal health. Future studies assessing the effects of a chronic stressor on adult mice with augmented nephron number are warranted, as are studies investigating the molecular mechanisms that result in high nephron endowment., Competing Interests: The authors have declared that no competing interests exist.

Background: The regulation of microRNAs (miRNAs) at different stages of the progression of type 2 diabetes mellitus (T2DM) and their role in glucose homeostasis was investigated., Methods: Microarrays were used to assess miRNA expression in skeletal muscle biopsies taken from healthy individuals and patients with pre-diabetes or T2DM, and insulin resistant offspring of rat dams fed a high fat diet during pregnancy., Results: Twenty-three miRNAs were differentially expressed in patients with T2DM, and 7 in the insulin resistant rat offspring compared to their controls. Among these, only one miRNA was similarly regulated: miR-194 expression was significantly reduced by 25 to 50% in both the rat model and in human with pre-diabetes and established diabetes. Knockdown of miR-194 in L6 skeletal muscle cells induced an increase in basal and insulin-stimulated glucose uptake and glycogen synthesis. This occurred in conjunction with an increased glycolysis, indicated by elevated lactate production. Moreover, oxidative capacity was also increased as we found an enhanced glucose oxidation in presence of the mitochondrial uncoupler FCCP. When miR-194 was down-regulated in vitro, western blot analysis showed an increased phosphorylation of AKT and GSK3β in response to insulin, and an increase in expression of proteins controlling mitochondrial oxidative phosphorylation., Conclusions: Type 2 diabetes mellitus is associated with regulation of several miRNAs in skeletal muscle. Interestingly, miR-194 was a unique miRNA that appeared regulated across different stages of the disease progression, from the early stages of insulin resistance to the development of T2DM. We have shown miR-194 is involved in multiple aspects of skeletal muscle glucose metabolism from uptake, through to glycolysis, glycogenesis and glucose oxidation, potentially via mechanisms involving AKT, GSK3 and oxidative phosphorylation. MiR-194 could be down-regulated in patients with early features of diabetes as an adaptive response to facilitate tissue glucose uptake and metabolism in the face of insulin resistance.

Pathogenesis of malaria infections is linked to remodeling of erythrocytes, a process dependent on the trafficking of hundreds of parasite-derived proteins into the host erythrocyte. Recent studies have demonstrated that the Plasmodium translocon of exported proteins (PTEX) serves as the central gateway for trafficking of these proteins, as inducible knockdown of the core PTEX constituents blocked the trafficking of all classes of cargo into the erythrocyte. However, the role of the auxiliary component PTEX88 in protein export remains less clear. Here we have used inducible knockdown technologies in P. falciparum and P. berghei to assess the role of PTEX88 in parasite development and protein export, which reveal that the in vivo growth of PTEX88-deficient parasites is hindered. Interestingly, we were unable to link this observation to a general defect in export of a variety of known parasite proteins, suggesting that PTEX88 functions in a different fashion to the core PTEX components. Strikingly, PTEX88-deficient P. berghei were incapable of causing cerebral malaria despite a robust pro-inflammatory response from the host. These parasites also exhibited a reduced ability to sequester in peripheral tissues and were removed more readily from the circulation by the spleen. In keeping with these findings, PTEX88-deficient P. falciparum-infected erythrocytes displayed reduced binding to the endothelial cell receptor, CD36. This suggests that PTEX88 likely plays a specific direct or indirect role in mediating parasite sequestration rather than making a universal contribution to the trafficking of all exported proteins.

Heart failure (HF) is an increasingly recognized complication of diabetes. Cardiac fibrosis is an important causative mechanism of HF associated with diabetes. Recent data indicate that inflammation may be particularly important in the pathogenesis of cardiovascular fibrosis. We sought to determine the mechanism by which cardiac fibrosis develops and to specifically investigate the role of the CXCR4 axis in this process. Animals with type I diabetes (streptozotocin treated mice) or type II diabetes (Israeli Sand-rats) and controls were randomized to treatment with a CXCR4 antagonist, candesartan or vehicle control. Additional groups of mice also underwent bone marrow transplantation (GFP+ donor marrow) to investigate the potential role of bone marrow derived cell mobilization in the pathogenesis of cardiac fibrosis. Both type I and II models of diabetes were accompanied by the development of significant cardiac fibrosis. CXCR4 antagonism markedly reduced cardiac fibrosis in both models of diabetes, similar in magnitude to that seen with candesartan. In contrast to candesartan, the anti-fibrotic actions of CXCR4 antagonism occurred in a blood pressure independent manner. Whilst the induction of diabetes did not increase the overall myocardial burden of GFP+ cells, it was accompanied by an increase in GFP+ cells expressing the fibroblast marker alpha-smooth muscle actin and this was attenuated by CXCR4 antagonism. CXCR4 antagonism was also accompanied by increased levels of circulating regulatory T cells. Taken together the current data indicate that pharmacological inhibition of CXCR4 significantly reduces diabetes induced cardiac fibrosis, providing a potentially important therapeutic approach.

The development of diabetic cardiomyopathy is a key contributor to heart failure and mortality in obesity and type 2 diabetes (T2D). Current therapeutic interventions for T2D have limited impact on the development of diabetic cardiomyopathy. Clearly, new therapies are urgently needed. A potential therapeutic target is protein kinase D (PKD), which is activated by metabolic insults and implicated in the regulation of cardiac metabolism, contractility and hypertrophy. We therefore hypothesised that PKD inhibition would enhance cardiac function in T2D mice. We first validated the obese and T2D db/db mouse as a model of early stage diabetic cardiomyopathy, which was characterised by both diastolic and systolic dysfunction, without overt alterations in left ventricular morphology. These functional characteristics were also associated with increased PKD2 phosphorylation in the fed state and a gene expression signature characteristic of PKD activation. Acute administration of the PKD inhibitor CID755673 to normal mice reduced both PKD1 and 2 phosphorylation in a time and dose-dependent manner. Chronic CID755673 administration to T2D db/db mice for two weeks reduced expression of the gene expression signature of PKD activation, enhanced indices of both diastolic and systolic left ventricular function and was associated with reduced heart weight. These alterations in cardiac function were independent of changes in glucose homeostasis, insulin action and body composition. These findings suggest that PKD inhibition could be an effective strategy to enhance heart function in obese and diabetic patients and provide an impetus for further mechanistic investigations into the role of PKD in diabetic cardiomyopathy.

Background: Over the course of ageing there is a natural and progressive loss of skeletal muscle mass. The onset and progression of age-related muscle wasting is associated with an attenuated activation of Akt-mTOR signalling and muscle protein synthesis in response to anabolic stimuli such as resistance exercise. MicroRNAs (miRNAs) are novel and important post-transcriptional regulators of numerous cellular processes. The role of miRNAs in the regulation of muscle protein synthesis following resistance exercise is poorly understood. This study investigated the changes in skeletal muscle miRNA expression following an acute bout of resistance exercise in young and old subjects with a focus on the miRNA species predicted to target Akt-mTOR signalling., Results: Ten young (24.2±0.9 years) and 10 old (66.6±1.1 years) males completed an acute resistance exercise bout known to maximise muscle protein synthesis, with muscle biopsies collected before and 2 hours after exercise. We screened the expression of 754 miRNAs in the muscle biopsies and found 26 miRNAs to be regulated with age, exercise or a combination of both factors. Nine of these miRNAs are highly predicted to regulate targets within the Akt-mTOR signalling pathway and 5 miRNAs have validated binding sites within the 3' UTRs of several members of the Akt-mTOR signalling pathway. The miR-99/100 family of miRNAs notably emerged as potentially important regulators of skeletal muscle mass in young and old subjects., Conclusion: This study has identified several miRNAs that were regulated with age or with a single bout of resistance exercise. Some of these miRNAs were predicted to influence Akt-mTOR signalling, and therefore potentially skeletal muscle mass. These miRNAs should be considered as candidate targets for in vivo modulation.

As a glycosphingolipid that can bind to several extracellular matrix proteins, sulfatide has the potential to become an effective targeting agent for tumors overexpressing tenasin-C in their microenvironment. To overcome the dose-limiting toxicity of doxorubicin (DOX), a sulfatide-containing nanoliposome (SCN) encapsulation approach was employed to improve treatment efficacy and reduce side effects of free DOX. This study analysed in vitro characteristics of sulfatide-containing nanoliposomal DOX (SCN-DOX) and assessed its cytotoxicity in vitro, as well as biodistribution, therapeutic efficacy, and systemic toxicity in a human glioblastoma U-118MG xenograft model. SCN-DOX was shown to achieve highest drug to lipid ratio (0.5∶1) and a remarkable in vitro stability. Moreover, DOX encapsulated in SCN was shown to be delivered into the nuclei and displayed prolonged retention over free DOX in U-118MG cells. This simple two-lipid SCN-DOX nanodrug has favourable pharmacokinetic attributes in terms of prolonged circulation time, reduced volume of distribution and enhanced bioavailability in healthy rats. As a result of the improved biodistribution, an enhanced treatment efficacy of SCN-DOX was found in glioma-bearing mice compared to the free drug. Finally, a reduction in the accumulation of DOX in the drug's principal toxicity organs achieved by SCN-DOX led to the diminished systemic toxicity as evident from the plasma biochemical analyses. Thus, SCN has the potential to be an effective and safer nano-carrier for targeted delivery of therapeutic agents to tumors with elevated expression of tenascin-C in their microenvironment.

The placenta plays a central role in determining the outcome of pregnancy. It undergoes changes during gestation as the fetus develops and as demands for energy substrate transfer and gas exchange increase. The molecular mechanisms that coordinate these changes have yet to be fully elucidated. The study performed a large scale screen of the transcriptome of the rat placenta throughout mid-late gestation (E14.25-E20) with emphasis on characterizing gestational age associated changes in the expression of genes involved in angiogenic pathways. Sprague Dawley dams were sacrificed at E14.25, E15.25, E17.25 and E20 (n = 6 per group) and RNA was isolated from one placenta per dam. Changes in placental gene expression were identified using Illumina Rat Ref-12 Expression BeadChip Microarrays. Differentially expressed genes (>2-fold change, <1% false discovery rate, FDR) were functionally categorised by gene ontology pathway analysis. A subset of differentially expressed genes identified by microarrays were confirmed using Real-Time qPCR. The expression of thirty one genes involved in the angiogenic pathway was shown to change over time, using microarray analysis (22 genes displayed increased and 9 gene decreased expression). Five genes (4 up regulated: Cd36, Mmp14, Rhob and Angpt4 and 1 down regulated: Foxm1) involved in angiogenesis and blood vessel morphogenesis were subjected to further validation. qPCR confirmed late gestational increased expression of Cd36, Mmp14, Rhob and Angpt4 and a decrease in expression of Foxm1 before labour onset (P<0.0001). The observed acute, pre-labour changes in the expression of the 31 genes during gestation warrant further investigation to elucidate their role in pregnancy.

Background: The aim of this work is to investigate the structure and function of enzymes immobilised on nanomaterials. This work will allow better understanding of enzyme-nanomaterial interactions, as well as designing functional protein-nanomaterial conjugates., Methodology/principal Findings: Multiwalled carbon nanotubes (MWNTs) were functionalised with amino groups to improve solubility and biocompatibility. The pristine and functionalised forms of MWNTs were characterised with Fourier-transform infrared spectroscopy. Thermogravimetric analysis was done to examine the degree of the functionalisation process. An immobilised biocatalyst was prepared on functionalised nanomaterial by covalent binding. Thermomyces lanuginosus lipase was used as a model enzyme. The structural change of the immobilised and free lipases were characterised with transmission electron Microscopy, X-ray photoelectron spectroscopy, Fourier-transform infrared spectroscopy and Circular dichroism spectroscopy. Biochemical characterisation of immobilised enzyme showed broader pH and thermal optima compared to soluble form. Reusability of the immobilised enzyme for hydrolysis of long chain esters was demonstrated up to ten cycles., Conclusion/significance: Lipase immobilised on MWNTs has exhibited significantly improved thermal stability. The exploration of advanced nanomaterial for enzyme immobilisation support using sophisticated techniques makes nanobiocatalyst of potential interest for biosensor applications.

Protein export into the host red blood cell is one of the key processes in the pathobiology of the malaria parasite Plasmodiumtrl falciparum, which extensively remodels the red blood cell to ensure its virulence and survival. In this study, we aimed to shed further light on the protein export mechanisms in the rodent malaria parasite P. berghei and provide further proof of the conserved nature of host cell remodeling in Plasmodium spp. Based on the presence of an export motif (R/KxLxE/Q/D) termed PEXEL (Plasmodium export element), we have generated transgenic P. berghei parasite lines expressing GFP chimera of putatively exported proteins and analysed one of the newly identified exported proteins in detail. This essential protein, termed PbCP1 (P. berghei Cleft-like Protein 1), harbours an atypical PEXEL motif (RxLxY) and is further characterised by two predicted transmembrane domains (2TMD) in the C-terminal end of the protein. We have functionally validated the unusual PEXEL motif in PbCP1 and analysed the role of the 2TMD region, which is required to recruit PbCP1 to discrete membranous structures in the red blood cell cytosol that have a convoluted, vesico-tubular morphology by electron microscopy. Importantly, this study reveals that rodent malaria species also induce modifications to their host red blood cell.

EpCAM is expressed at low levels in a variety of normal human epithelial tissues, but is overexpressed in 70-90% of carcinomas. From a clinico-pathological point of view, this has both prognostic and therapeutic significance. EpCAM was first suggested as a therapeutic target for the treatment of epithelial cancers in the 1990s. However, following several immunotherapy trials, the results have been mixed. It has been suggested that this is due, at least in part, to an unknown level of EpCAM expression in the tumors being targeted. Thus, selection of patients who would benefit from EpCAM immunotherapy by determining EpCAM status in the tumor biopsies is currently undergoing vigorous evaluation. However, current EpCAM antibodies are not robust enough to be able to detect EpCAM expression in all pathological tissues. Here we report a newly developed EpCAM RNA aptamer, also known as a chemical antibody, which is not only specific but also more sensitive than current antibodies for the detection of EpCAM in formalin-fixed paraffin-embedded primary breast cancers. This new aptamer, together with our previously described aptamer, showed no non-specific staining or cross-reactivity with tissues that do not express EpCAM. They were able to reliably detect target proteins in breast cancer xenograft where an anti-EpCAM antibody (323/A3) showed limited or no reactivity. Our results demonstrated a more robust detection of EpCAM using RNA aptamers over antibodies in clinical samples with chromogenic staining. This shows the potential of aptamers in the future of histopathological diagnosis and as a tool to guide targeted immunotherapy.

Sulfatide is a glycosphingolipid known to interact with several extracellular matrix proteins, such as tenascin-C which is overexpressed in many types of cancer including that of the colon. In view of the limited success of chemotherapy in colorectal cancer and high toxicity of doxorubicin (DOX), a sulfatide-containing liposome (SCL) encapsulation approach was taken to overcome these barriers. This study assessed the in vitro cytotoxicity, biodistribution, therapeutic efficacy and systemic toxicity in vivo of sulfatide-containing liposomal doxorubicin (SCL-DOX) using human colonic adenocarcinoma HT-29 xenograft as the experimental model. In vitro, SCL-DOX was shown to be delivered into the nuclei and displayed prolonged retention compared with the free DOX. The use of this nanodrug delivery system to deliver DOX for treatment of tumor-bearing mice produced a much improved therapeutic efficacy in terms of tumor growth suppression and extended survival in contrast to the free drug. Furthermore, treatment of tumor-bearing mice with SCL-DOX resulted in a lower DOX uptake in the principal sites of toxicity of the free drug, namely the heart and skin, as well as reduced myelosuppression and diminished cardiotoxicity. Such natural lipid-guided nanodrug delivery systems may represent a new strategy for the development of effective anticancer chemotherapeutics targeting the tumor microenvironment for both primary tumor and micrometastases.

Cancer cachexia is a wasting condition, driven by systemic inflammation and oxidative stress. This study investigated eicosapentaenoic acid (EPA) in combination with oxypurinol as a treatment in a mouse model of cancer cachexia. Mice with cancer cachexia were randomized into 4 treatment groups (EPA (0.4 g/kg/day), oxypurinol (1 mmol/L ad-lib), combination, or control), and euthanized after 29 days. Analysis of oxidative damage to DNA, mRNA analysis of pro-oxidant, antioxidant and proteolytic pathway components, along with enzyme activity of pro- and antioxidants were completed on gastrocnemius muscle. The control group displayed earlier onset of tumor compared to EPA and oxypurinol groups (P<0.001). The EPA group maintained body weight for an extended duration (20 days) compared to the oxypurinol (5 days) and combination (8 days) groups (P<0.05). EPA (18.2±3.2 pg/ml) and combination (18.4±3.7 pg/ml) groups had significantly higher 8-OH-dG levels than the control group (12.9±1.4 pg/ml, P≤0.05) indicating increased oxidative damage to DNA. mRNA levels of GPx1, MURF1 and MAFbx were higher following EPA treatment compared to control (P≤0.05). Whereas oxypurinol was associated with higher GPx1, MnSOD, CAT, XDH, MURF1, MAFbx and UbB mRNA compared to control (P≤0.05). Activity of total SOD was higher in the oxypurinol group (32.2±1.5 U/ml) compared to control (27.0±1.3 U/ml, P<0.01), GPx activity was lower in the EPA group (8.76±2.0 U/ml) compared to control (14.0±1.9 U/ml, P<0.05), and catalase activity was lower in the combination group (14.4±2.8 U/ml) compared to control (20.9±2.0 U/ml, P<0.01). There was no change in XO activity. The increased rate of weight decline in mice treated with oxypurinol indicates that XO may play a protective role during the progression of cancer cachexia, and its inhibition is detrimental to outcomes. In combination with EPA, there was little significant improvement from control, indicating oxypurinol is unlikely to be a viable treatment compound in cancer cachexia.

Oxidative stress is due to an imbalance of antioxidant/pro-oxidant homeostasis and is associated with the progression of several neurological diseases, including Parkinson's and Alzheimer's disease and amyotrophic lateral sclerosis. Furthermore, oxidative stress is responsible for the neuronal loss and dysfunction associated with disease pathogenesis. Survivin is a member of the inhibitors of the apoptosis (IAP) family of proteins, but its neuroprotective effects have not been studied. Here, we demonstrate that SurR9-C84A, a survivin mutant, has neuroprotective effects against H₂O₂-induced neurotoxicity. Our results show that H₂O₂ toxicity is associated with an increase in cell death, mitochondrial membrane depolarisation, and the expression of cyclin D1 and caspases 9 and 3. In addition, pre-treatment with SurR9-C84A reduces cell death by decreasing both the level of mitochondrial depolarisation and the expression of cyclin D1 and caspases 9 and 3. We further show that SurR9-C84A increases the antioxidant activity of GSH-peroxidase and catalase, and effectively counteracts oxidant activity following exposure to H₂O₂. These results suggest for the first time that SurR9-C84A is a promising treatment to protect neuronal cells against H₂O₂-induced neurotoxicity.

Background: Studies of mate choice in anuran amphibians have shown female preference for a wide range of male traits despite females gaining no direct resources from males (i.e. non-resource based mating system). Nevertheless, theoretical and empirical studies have shown that females may still gain indirect genetic benefits from choosing males of higher genetic quality and thereby increase their reproductive success., Methodology/principal Findings: We investigated two components of sexual selection in the Moor frog (Rana arvalis), pre-copulatory female choice between two males of different size ('large' vs. 'small'), and their fertilization success in sperm competition and in isolation. Females' showed no significant preference for male size (13 small and six large male preferences) but associated preferentially with the male that subsequently was the most successful at fertilizing her eggs in isolation. Siring success of males in competitive fertilizations was unrelated to genetic similarity with the female and we detected no effect of sperm viability on fertilization success. There was, however, a strong positive association between a male's innate fertilization ability with a female and his siring success in sperm competition. We also detected a strong negative effect of a male's thumb length on his competitive siring success., Conclusions/significance: Our results show that females show no preference for male size but are still able to choose males which have greater fertilization success. Genetic similarity and differences in the proportion of viable sperm within a males ejaculate do not appear to affect siring success. These results could be explained through pre- and/or postcopulatory choice for genetic benefits and suggest that females are able to perceive the genetic quality of males, possibly basing their choice on multiple phenotypic male traits.

Background: A large number of studies in postcopulatory sexual selection use paternity success as a proxy for fertilization success. However, selective mortality during embryonic development can lead to skews in paternity in situations of polyandry and sperm competition. Thus, when assessment of paternity fails to incorporate mortality skews during early ontogeny, this may interfere with correct interpretation of results and subsequent evolutionary inference. In a previous series of in vitro sperm competition experiments with amphibians (Litoria peronii), we showed skewed paternity patterns towards males more genetically similar to the female., Methodology/principal Findings: Here we use in vitro fertilizations and sperm competition trials to test if this pattern of paternity of fully developed tadpoles reflects patterns of paternity at fertilization and if paternity skews changes during embryonic development. We show that there is no selective mortality through ontogeny and that patterns of paternity of hatched tadpoles reflects success of competing males in sperm competition at fertilization., Conclusions/significance: While this study shows that previous inferences of fertilization success from paternity data are valid for this species, rigorous testing of these assumptions is required to ensure that differential embryonic mortality does not confound estimations of true fertilization success.