The importance of the expression profile of claudins in the molecular classification of breast cancer (BC) is currently under investigation. Claudins, together with cadherins, serve an important role in the epithelial-mesenchymal transition and influence the chemosensitivity of cancer cells. Adjuvant chemotherapy is administered following surgical resection in selected cases of BC. Previous neoadjuvant chemotherapy may change the molecular profile of a tumour and subsequently also its chemosensitivity. In the current study, the expression of claudin-1, -3 and -4, E- and N-cadherin and the standard BC biomarkers [oestrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) and marker of proliferation Ki-67 (Ki-67)] in formalin-fixed, paraffin-embedded sections from 62 patients with invasive BC was analysed using immunohistochemistry prior to and following neoadjuvant chemotherapy. The results revealed increased expression of claudin-1 (P=0.03) and decreased expression of claudin-3 (P=0.005), PR (P<0.001) and Ki-67 (P=0.01) following the neoadjuvant therapy. No significant changes in the expression of ER, claudin-4 or E- and N-cadherin were observed following therapy. Furthermore, an association between the expression of claudin-1 and the standard BC markers (P<0.05) was identified. A high expression of claudin-1 was more frequently observed in the triple-negative BC cohort than in the cohort with positive ER, PR and/or HER2 before (P=0.04) and after chemotherapy (P=0.02). The expression of N-cadherin was associated with the expression of ER, PR, HER2 and tumour grade (P<0.05). A positive association between the expression of claudin-3 and E-cadherin (P=0.005) was observed. No association was found between the expression of E- and N-cadherin. In conclusion, significant changes in the expression of claudin-1 and -3 but not in the expression of claudin-4, E- and N-cadherin were observed in samples taken from patients with BC following chemotherapy. These findings indicate that claudins-1 and -3 serve a role in the response of BC to chemotherapy.

In 2002, Colantuoni et al described the possibility of dependency in rats after intermittent, excessive consumption of a 25% glucose solution over a one-week period. We hypothesized that the intermittent consumption of any tasty solution can lead to endogenous opioid dependency. Another aim was to determine whether dependency is connected to the taste of the consumed substance or with its physiological significance. Rats were maintained on chow and cyclic glucose, NaCl, or monosodium glutamate (MSG) solution for 8 days. On day 9, after a 12-hr deprivation period and administration of intraperitoneally (IP) naloxone, the general withdrawal index was calculated as the sum of teeth chattering, head shaking, forepaw tremors, and wet dog shakes. Motor activity was also documented. After the intermittent consumption of any tasty solution, rats were found to demonstrate signs of endogenous opioid dependence. The development of dependence was not related to taste preferences or the amount of solution consumed. Intermittent use of essential substance with a pleasant taste, as glucose and sodium chloride, leads to the rapid development of endogenous opioid dependence. Withdrawal syndrome following the consumption of MSG solution is associated with the presence of sodium ions in the MSG molecule.

Arterial hypertension (HT) is common in the Russian adult population, with half of affected individuals inadequately controlled. Low adherence to medication seems likely to be a factor. We report a scoping review of studies on adherence to antihypertensive therapy (AHT) in Russia to determine the extent of research undertaken, the frequency of adherence among adults diagnosed with HT, methodologies used in the studies, and their ability to describe determinants of adherence.

Morphometric analysis is a common approach for comparing and categorizing botanical samples; however, completing a suite of analyses using existing tools may require a multi-stage, multi-program process. To facilitate streamlined analysis within a single program, Morphological Analysis of Size and Shape (MASS) for leaves was developed. Its utility is demonstrated using exemplar leaf samples from Acer saccharum, Malus domestica, and Lithospermum.

Polymorphic nuclear simple sequence repeat (nSSR) markers were developed for Sanguinaria canadensis (Papaveraceae), a spring ephemeral native to eastern North America.

Platelet-rich plasma (PRP) and hyaluronic acid have been shown to be useful in the treatment of knee osteoarthritis. However, investigations comparing the efficacy of these two drugs together are insufficient.

The long non-coding RNA (LncRNA) antisense RNA in the INK4 locus (ANRIL) is overexpressed in several cancers including breast cancer. To better understand the role of ANRIL in breast cancer this study investigated where ANRIL was expressed in breast tumors using in situ hybridization by RNAscope. Additional RNAscope assays for IL6, CCL2, and POSTN were used to establish whether ANRIL correlated with increased tumor promoting cytokines. Breast tumors with ANRIL over expressed from real-time quantitative (RT-q) PCR assays were selected for analysis using RNAscope. All tumors showed ANRIL expression in malignant cells, but amongst tumors ANRIL showed different subcellular locations with 56% of tumors with ANRIL only in the nucleus, 16% with ANRIL only in the cytoplasm and 28% with ANRIL in both the nucleus and cytoplasm. Cases with nuclear ANRIL were positively correlated with POSTN expression in malignant cells (ρ = 0.57, P = 0.0086), and no correlation was found between ANRIL and IL6 or CCL2. Reduced POSTN was also found using siRNA to ANRIL in MDA-MB-231 and MCF7 breast cancer cells. These data indicate that ANRIL is expressed in malignant breast cells, and suggest its subcellular location may indicate its function in cancer progression.

Oral squamous cell carcinoma (OSCC) is a deadly disease that comprises 60% of all head and neck squamous cell cancers. The leaves of the Neem tree (Azadirachta indica) have been used in traditional Ayurvedic medicine for centuries to treat numerous oral maladies and are known to have significant anti-inflammatory properties. We hypothesize that a highly pure super critical CO2 Neem leaf extract (SCNE) prevents initiation and progression of OSCC via downregulation of intra-tumor pro-inflammatory pathways, which promote tumorigenesis. Hence, we investigated the anticancer effects of SCNE using in vitro and in vivo platforms. OSCC cell lines (SCC4, Cal27, and HSC3) were treated with SCNE while inflammation, proliferation, and migration were analyzed over time. SCNE treatment significantly inhibited OSCC cell proliferation and migration and reduced MMP activity in vitro, suggesting its potential to inhibit tumor growth and metastasis. The preventive effects of SCNE in ectopic xenograft and 4NQO-1 (4-Nitroquinoline-1-oxide) carcinogen-induced mouse models of OSCC were also evaluated. Indeed, xenografted nude mice showed significant reduction of OSCC tumor volumes. Likewise, SCNE significantly reduced the incidence of tongue dysplasia in the 4NQO-1 OSCC initiation model. In both OSCC animal models, SCNE significantly depressed circulating pro-cancer inflammatory cytokines (host and tumor-secreted) including NFkB, COX2, IL-1, IL-6, TNFα, and IFNγ. In addition, we demonstrate that SCNE downregulates STAT3 and AKT expression and activity in vitro. We also demonstrate that the primary active component, nimbolide (NIM), has significant anticancer activity in established OSCC xenografts. Lastly, we show that SCNE induces an M1 phenotype in tumor associated macrophages (TAMS) in vivo. Taken together, these data strongly support SCNE as means of preventing OSCC via downregulation of pro-cancer inflammatory cascades and NIM as a potential new therapy for existing OSCC.

Toxoplasma gondii is capable of actively invading almost any mammalian cell type including phagocytes. Early events in phagocytic cells such as dendritic cells are not only key to establishing parasite infection, but conversely play a pivotal role in initiating host immunity. It is now recognized that in addition to changes in canonical immune markers and mediators, alteration in metabolism occurs upon activation of phagocytic cells. These metabolic changes are important for supporting the developing immune response, but can affect the availability of nutrients for intracellular pathogens including T. gondii. However, the interaction of T. gondii with these cells and particularly how infection changes their metabolism has not been extensively investigated. Herein, we use a multi-omics approach comprising transcriptomics and metabolomics validated with functional assays to better understand early events in these cells following infection. Analysis of the transcriptome of T. gondii infected bone marrow derived dendritic cells (BMDCs) revealed significant alterations in transcripts associated with cellular metabolism, activation of T cells, inflammation mediated chemokine and cytokine signaling pathways. Multivariant analysis of metabolomic data sets acquired through non-targeted liquid chromatography mass spectroscopy (LCMS) identified metabolites associated with glycolysis, the TCA cycle, oxidative phosphorylation and arginine metabolism as major discriminants between control uninfected and T. gondii infected cells. Consistent with these observations, glucose uptake and lactate dehydrogenase activity were upregulated in T. gondii infected BMDC cultures compared with control BMDCs. Conversely, BMDC mitochondrial membrane potential was reduced in T. gondii-infected cells relative to mitochondria of control BMDCs. These changes to energy metabolism, similar to what has been described following LPS stimulation of BMDCs and macrophages are often termed the Warburg effect. This metabolic reprogramming of cells has been suggested to be an important adaption that provides energy and precursors to facilitate phagocytosis, antigen processing and cytokine production. Other changes to BMDC metabolism are evident following T. gondii infection and include upregulation of arginine degradation concomitant with increased arginase-1 activity and ornithine and proline production. As T. gondii is an arginine auxotroph the resultant reduced cellular arginine levels are likely to curtail parasite multiplication. These results highlight the complex interplay of BMDCs and parasite metabolism within the developing immune response and the consequences for adaptive immunity and pathogen clearance.

Miniaturized culture systems of hepatic cells are emerging as a strong tool facilitating studies related to liver diseases and drug discovery. However, the experimental optimization of various parameters involved in the operation of these systems is time-consuming and expensive. Hence, developing numerical tools predicting the function of such systems can significantly reduce the associated cost. In this paper, a perfusion-based three dimensional (3D) bioreactor comprising encapsulated human liver hepatocellular carcinoma (HepG2) spheroids are analyzed. The flow and mass transfer equations for oxygen as well as different metabolites such as albumin, glucose, glutamine, ammonia, and urea were solved in three different domains, i.e., free flow, hydrogel, and spheroid porous media sections. Since the spheroids were encapsulated inside the hydrogel, shear stress imposed on them were found to be less than tolerable thresholds. The predicted cumulative albumin concentration over the 7 days of culture period showed a good agreement with the experimental data. Based on the critical role of oxygen supply to the hepatocytes, a parametric study was performed and the effect of various parameters was investigated. Results illustrated that convection mechanism was the dominant transport mechanism in the main-stream section contrary to the intra spheroids parts where the diffusion was the prevailing transport mechanism. In the hydrogel parts, the rate of diffusion and convection mechanisms were almost identical. As expected, higher perfusion rate would provide high oxygen level for the cells and, smaller spheroids with a diameter of 100 μm were at the low risk of hypoxic conditions due to short diffusive oxygen penetration depth. Numerical results evidenced that spheroids with diameter size >200 μm at low porosities (ε = 0.2-0.3) were at risk of oxygen depletion, especially at locations near the core center. Therefore, these results could be beneficial in preventing hypoxic conditions during in vitro experiments. The presented numerical model provides a numerical platform which can help researchers to design and optimize complex bioreactors and obtain numerical indexes of the main metabolites in a very short time prior to any fabrications. Such numerical indexes can be helpful in certifying the outcomes of forensic investigations.

We used T2 relaxation, chemical exchange saturation transfer (CEST), and dynamic contrast-enhanced (DCE) magnetic resonance imaging (MRI) to assess whether bacterial infection can be differentiated from inflammation in a myositis-induced mouse model. We measured the T2 relaxation time constants, %CEST at 5 saturation frequencies, and area under the curve (AUC) from DCE-MRI after maltose injection from infected, inflamed, and normal muscle tissue models. We applied principal component analysis (PCA) to reduce dimensionality of entire CEST spectra and DCE signal evolutions, which were analyzed using standard classification methods. We extracted features from dimensional reduction as predictors for machine learning classifier algorithms. Normal, inflamed, and infected tissues were evaluated with H&E and gram-staining histological studies, and bacterial-burden studies. The T2 relaxation time constants and AUC of DCE-MRI after injection of maltose differentiated infected, inflamed, and normal tissues. %CEST amplitudes at -1.6 and -3.5 ppm differentiated infected tissues from other tissues, but these did not differentiate inflamed tissue from normal tissue. %CEST amplitudes at 3.5, 3.0, and 2.5 ppm, AUC of DCE-MRI for shorter time periods, and relative Ktrans and kep values from DCE-MRI could not differentiate tissues. PCA and machine learning of CEST-MRI and DCE-MRI did not improve tissue classifications relative to traditional analysis methods. Similarly, PCA and machine learning did not further improve tissue classifications relative to T2 MRI. Therefore, future MRI studies of infection models should focus on T2-weighted MRI and analysis of T2 relaxation times.

We investigated a projection interpolation method for reconstructing dynamic contrast-enhanced (DCE) heart images from undersampled x-ray projections with filtered backprojecton (FBP). This method may facilitate the application of sparse-view dynamic acquisition for ultralow-dose quantitative computed tomography (CT) myocardial perfusion (MP) imaging. We conducted CT perfusion studies on 5 pigs with a standard full-view acquisition protocol (984 projections). We reconstructed DCE heart images with FBP from all and a quarter of the measured projections evenly distributed over 360°. We interpolated the sparse-view (quarter) projections to a full-view setting using a cubic-spline interpolation method before applying FBP to reconstruct the DCE heart images (synthesized full-view). To generate MP maps, we used 3 sets of DCE heart images, and compared mean MP values and biases among the 3 protocols. Compared with synthesized full-view DCE images, sparse-view DCE images were more affected by streak artifacts arising from projection undersampling. Relative to the full-view protocol, mean bias in MP measurement associated with the sparse-view protocol was 10.0 mL/min/100 g (95%CI: -8.9 to 28.9), which was >3 times higher than that associated with the synthesized full-view protocol (3.3 mL/min/100 g, 95% CI: -6.7 to 13.2). The cubic-spline-view interpolation method improved MP measurement from DCE heart images reconstructed from only a quarter of the full projection set. This method can be used with the industry-standard FBP algorithm to reconstruct DCE images of the heart, and it can reduce the radiation dose of a whole-heart quantitative CT MP study to <2 mSv (at 8-cm coverage).

Preclinical imaging is critical in the development of translational strategies to detect diseases and monitor response to therapy. The National Cancer Institute Co-Clinical Imaging Resource Program was launched, in part, to develop best practices in preclinical imaging. In this context, the objective of this work was to develop a 1-hour, multiparametric magnetic resonance image-acquisition pipeline with triple-negative breast cancer patient-derived xenografts (PDXs). The 1-hour, image-acquisition pipeline includes T1- and T2-weighted scans, quantitative T1, T2, and apparent diffusion coefficient (ADC) parameter maps, and dynamic contrast-enhanced (DCE) time-course images. Quality-control measures used phantoms. The triple-negative breast cancer PDXs used for this study averaged 174 ± 73 μL in volume, with region of interest-averaged T1, T2, and ADC values of 1.9 ± 0.2 seconds, 62 ± 3 milliseconds, and 0.71 ± 0.06 μm2/ms (mean ± SD), respectively. Specific focus was on assessing the within-subject test-retest coefficient-of-variation (CVWS) for each of the magnetic resonance imaging metrics. Determination of PDX volume via manually drawn regions of interest is highly robust, with ∼1% CVWS. Determination of T2 is also robust with a ∼3% CVWS. Measurements of T1 and ADC are less robust with CVWS values in the 6%-11% range. Preliminary DCE test-retest time-course determinations, as quantified by area under the curve and Ktrans from 2-compartment exchange (extended Tofts) modeling, suggest that DCE is the least robust protocol, with ∼30%-40% CVWS.

The aim of this study was to evaluate potential protective effects of propolis on furan-induced hepatic damage by assessing the levels of malondialdehyde (MDA) and reduced glutathione (GSH), antioxidant enzyme activities, and histopathological changes in the liver.

Over the past 30 years, there has been a dramatic rise in the number of infections caused by multidrug-resistant bacteria, which have proliferated due to the misuse and overuse of antibiotics. Over this same time period, however, there has also been a decline in the number of antibiotics with novel mechanisms of action coming to market. Therefore, there is a growing need for an increase in the speed at which new antibiotics are discovered and developed. Natural products produced by bacteria have been and continue to be a robust source of novel antibiotics; however, new and complementary methods for screening large bacterial libraries for novel antibiotic production are needed due to the current agar methods being limited in scope, time consuming, and prone to error. Herein, we describe a rapid, robust, and quantitative high-throughput liquid culture screening method for antibiotic production by bacteria. This method has the ability to screen both mono- and coculture mixtures of bacteria in vitro and be adapted to other phenotypic natural product analyses. Over 260 bacterial species were screened in monoculture, and 38 and 34% were found to produce antibiotics capable of inhibition of Staphylococcus aureus or Escherichia coli, respectively, with 8 and 4% being classified as strong producers (≥30% growth inhibition), respectively. Bacteria found to not produce antibiotics in monoculture were also screened in coculture using an adaptation of this method. Of the more than 270 cocultures screened, 14 and 30% were found to produce antibiotics capable of inhibition of S. aureus or E. coli, respectively. Of those bacteria found to produce antibiotics in monoculture, 43 bacteria were subjected to 16S rRNA sequencing and found to be majority Pseudomonas (37%), Serratia (19%), and Bacillus (14%) bacteria, but two novel producers, Herbaspirillum and Kluyvera, were also found.

The United Nations Office on Drugs and Crime has designated several "legal highs" as "plants of concern" because of the dangers associated with their increasing recreational abuse. Routine identification of these products is hampered by the difficulty in distinguishing them from innocuous plant materials such as foods, herbs, and spices. It is demonstrated here that several of these products have unique but consistent headspace chemical profiles and that multivariate statistical analysis processing of their chemical signatures can be used to accurately identify the species of plants from which the materials are derived. For this study, the headspace volatiles of several species were analyzed by direct analysis in real-time high-resolution mass spectrometry (DART-HRMS). These species include Althaea officinalis, Calea zacatechichi, Cannabis indica, Cannabis sativa, Echinopsis pachanoi, Lactuca virosa, Leonotis leonurus, Mimosa hositlis, Mitragyna speciosa, Ocimum basilicum, Origanum vulgare, Piper methysticum, Salvia divinorum, Turnera diffusa, and Voacanga africana. The results of the DART-HRMS analysis revealed intraspecies similarities and interspecies differences. Exploratory statistical analysis of the data using principal component analysis and global t-distributed stochastic neighbor embedding showed clustering of like species and separation of different species. This led to the use of supervised random forest (RF), which resulted in a model with 99% accuracy. A conformal predictor based on the RF classifier was created and proved to be valid for a significance level of 8% with an efficiency of 0.1, an observed fuzziness of 0, and an error rate of 0. The variables used for the statistical analysis processing were ranked in terms of the ability to enable clustering and discrimination between species using principal component analysis-variable importance of projection scores and RF variable importance indices. The variables that ranked the highest were then identified as m/z values consistent with molecules previously identified in plant material. This technique therefore shows proof-of-concept for the creation of a database for the detection and identification of plant-based legal highs through headspace analysis.

Pyrazoles, thiazoles and fused thiazoles have been reported to possess many biological activities. 3-Methyl-5-oxo-4-(2-arylhydrazono)-4,5-dihydro-1H-pyrazole-1-carbothioamides 3a,b (obtained from the reaction of ethyl 3-oxo-2-(2-arylhydrazono)butanoates 1a,b with thiosemicarbazide) could be transformed into a variety of thiazolyl-pyrazole derivatives 6a-h, 10a-c, 15a-c, 17, 19 and 21 via their reaction with a diversity hydrazonoyl chlorides as well as bromoacetyl derivatives. Moreover, the computational studies were carried out for all new compounds. The results indicated that five compounds showed promising binding affinities (10a: - 3.4 kcal/mol, 6d: - 3.0 kcal/mol, 15a: - 2.2 kcal/mol, 3a: - 1.6 kcal/mol, and 21: - 1.3 kcal/mol) against the active site of the epidermal growth factor receptor kinase (EGFR). The cytotoxicity of the potent products 3a, 6d, 10a, 15a, and 21 was examined against human liver carcinoma cell line (HepG-2) and revealed activities close to Doxorubicin standard drug. There was an understanding between the benefits of restricting affinities and the data obtained from the practical anticancer screening of the tested compounds.

The transcription factor Nuclear factor erythroid-2-related factor 2 (NRF2) and its principal repressive regulator, Kelch-like ECH-associated protein 1 (KEAP1), are perilous in the regulation of inflammation, as well as maintenance of homeostasis. Thus, NRF2 activation is involved in cytoprotection against many inflammatory disorders. N'-Nicotinoylquinoxaline-2-carbohdyrazide (NQC) was structurally designed by the combination of important pharmacophoric features of bioactive compounds reported in the literature.

Objective: To describe the development of structure, process and outcome indicators that will advance the quality of walking rehabilitation for Canadians with spinal cord injury or disease (SCI/D) by 2020. Method: A framework for the evaluation of the quality of walking rehabilitation was developed by experts in walking after SCI/D. A systematic literature review identified factors influencing walking outcomes and potential walking indicators. A Driver diagram analysis summarized the factors affecting walking outcomes and subsequently informed the selection of structure and process indicators. Psychometric properties and clinical utility of potential walking indicators were considered during the selection of outcome indicators. Results: The structure indicator is the number of physical therapists using evidence-based walking interventions per number of ambulatory individuals with SCI/D. The process indicator is the number of received hours of walking interventions during inpatient rehabilitation per number of ambulatory individuals with SCI/D. The intermediary outcome indicator, which is collected at discharge from inpatient rehabilitation, is either the modified Timed Up and Go or the 10-Meter Walk Test, the choice of measure is dictated by the stage of walking recovery, as defined by the Standing and Walking Assessment Tool. The final outcome indicator, collected at 18 months post-discharge, is the Spinal Cord Independence Measure III-Mobility subscale. Conclusion: The selected indicators align with current clinical practice in Canada. The indicators will direct the timing and enhance the volume of walking therapy delivered, to ultimately increase the proportion of patients who achieve their walking potential by 18 months post-rehabilitation.

Ascidians are the closest living relatives of vertebrates, and their study is important for understanding the evolutionary processes of oocyte maturation and ovulation. In this study, we first examined the ovulation of Ciona intestinalis Type A by monitoring follicle rupture in vitro, identifying a novel mechanism of neuropeptidergic regulation of oocyte maturation and ovulation. Ciona vasopressin family peptide (CiVP) directly upregulated the phosphorylation of extracellular signal-regulated kinase (CiErk1/2) via its receptor. CiVP ultimately activated a maturation-promoting factor, leading to oocyte maturation via germinal vesicle breakdown. CiErk1/2 also induced expression of matrix metalloproteinase (CiMMP2/9/13) in the oocyte, resulting in collagen degradation in the outer follicular cell layer and liberation of fertile oocytes from the ovary. This is the first demonstration of essential pathways regulating oocyte maturation and ovulation in ascidians and will facilitate investigations of the evolutionary process of peptidergic regulation of oocyte maturation and ovulation throughout the phylum Chordata.