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ABSTRACTThis research was conducted to determine the effects of Andrographis paniculate extract on hybrid catfish. The fish were fed 0, 0.2, 0.4 and 0.6 g/kg extract to supplement their diets for 90 days. The highest weigh gain, average daily growth, specific growth rate, feed efficiency and protein efficiency ratio, as well as the lowest feed conversion rate, were found in the group fed 0.6 g/kg. RBC were at their highest in all treatment groups, while the highest levels of total WBC and haematocrit percentage were observed in the group fed 0.6 g/kg. ALT and triglyceride were at their lowest levels in all treatment groups, while the highest amounts of glucose were found in all treatment groups. The highest levels of high-density lipoprotein and the lowest cholesterol were observed in fish fed 0.6 g/kg. Lysozyme activity was initially observed at a significant difference on days 60–90, the highest activity was found in the group fed 0.6 g/kg. After bacterial challenge test, lysozyme activity and WBC were significantly increased in all treatment groups, the lowest cumulative mortality was found in the group fed 0.6 g/kg. These results suggest that 0.6 g/kg of the extract has positive effects on hybrid catfish.

Upon sensing DNA double-strand breaks (DSBs), eukaryotic cells either die or repair DSBs via one of the two competing pathways, i.e., non-homologous end-joining (NHEJ) or homologous recombination (HR). We show that cell fate after DSBs hinges on GIV/Girdin, a guanine nucleotide-exchange modulator of heterotrimeric Giα•βγ protein. GIV suppresses HR by binding and sequestering BRCA1, a key coordinator of multiple steps within the HR pathway, away from DSBs; it does so using a C-terminal motif that binds BRCA1's BRCT-modules via both phospho-dependent and -independent mechanisms. Using another non-overlapping C-terminal motif GIV binds and activates Gi and enhances the "free" Gβγ→PI-3-kinase→Akt pathway, which promotes survival and is known to suppress HR, favor NHEJ. Absence of GIV, or loss of either of its C-terminal motifs enhanced cell death upon genotoxic stress. Because GIV selectively binds other BRCT-containing proteins suggests that G-proteins may fine-tune sensing, repair, and survival after diverse types of DNA damage.

Neuroendocrine neoplasia (NENs) are a complex and heterogeneous group of cancers that can arise from neuroendocrine tissues throughout the body and differentiate them from other tumors. Their low incidence and high diversity make many of them orphan conditions characterized by a low incidence and few dedicated clinical trials. Study of the molecular and genetic nature of these diseases is limited in comparison to more common cancers and more dependent on preclinical models, including both in vitro models (such as cell lines and 3D models) and in vivo models (such as patient derived xenografts (PDXs) and genetically-engineered mouse models (GEMMs)). While preclinical models do not fully recapitulate the nature of these cancers in patients, they are useful tools in investigation of the basic biology and early-stage investigation for evaluation of treatments for these cancers. We review available preclinical models for each type of NEN and discuss their history as well as their current use and translation.

Background and Aims: Colorectal cancer (CRC) polygenic risk scores (PRS) may help personalize CRC prevention strategies. We investigated whether an existing PRS was associated with advanced neoplasia (AN) in a population undergoing screening and follow-up colonoscopy. Methods: We evaluated 10-year outcomes in the Cooperative Studies Program #380 screening colonoscopy cohort, which includes a biorepository of selected individuals with baseline AN (defined as CRC or adenoma ≥10 mm or villous histology, or high-grade dysplasia) and matched individuals without AN. A PRS was constructed from 136 prespecified CRC-risk single nucleotide polymorphisms. Multivariate logistic regression was used to evaluate the PRS for associations with AN prevalence at baseline screening colonoscopy or incident AN in participants with at least one follow-up colonoscopy. Results: The PRS was associated with AN risk at baseline screening colonoscopy (P = .004). Participants in the lowest PRS quintile had more than a 70% decreased risk of AN at baseline (odds ratio 0.29, 95% confidence interval 0.14–0.58; P < .001) compared to participants with a PRS in the middle quintile. Using a PRS cut-off of more than the first quintile to indicate need for colonoscopy as primary screening, the sensitivity for detecting AN at baseline is 91.8%. We did not observe a relationship between the PRS and incident AN during follow-up (P = .28). Conclusion: A PRS could identify individuals at low risk for prevalent AN. Ongoing work will determine whether this PRS can identify a subset of individuals at sufficiently low risk who could safely delay or be reassured about noninvasive screening. Otherwise, more research is needed to augment these genetic tools to predict incident AN during long-term follow-up.

Candida glabrata is an important pathogen causing invasive infection associated with a high mortality rate. One mechanism that causes the failure of Candida eradication is an increase in regulatory T cells (Treg), which play a major role in immune suppression and promoting Candida pathogenicity. To date, how C. glabrata induces a Treg response remains unclear. Dendritic cells (DCs) recognition of fungi provides the fundamental signal determining the fate of the T-cell response. This study investigated the interplay between C. glabrata and DCs and its effect on Treg induction. We found that C. glabrata β-glucan was a major component that interacted with DCs and consequently mediated the Treg response. Blocking the binding of C. glabrata β-glucan to dectin-1 and complement receptor 3 (CR3) showed that CR3 activation in DCs was crucial for the induction of Treg. Furthermore, a ligand–receptor binding assay showed the preferential binding of C. glabrata β-glucan to CR3. Our data suggest that C. glabrata β-glucan potentially mediates the Treg response, probably through CR3-dependent activation in DCs. This study contributes new insights into immune modulation by C. glabrata that may lead to a better design of novel immunotherapeutic strategies for invasive C. glabrata infection.

Heba Ahmed Osman,1 Hanaa Nafady-Hego,2 Khalid Ali Nasif,3,4 Heba A Ahmed,5 Ekram Abdel-Rahman Mahmoud,6 Noher Mohamad Abass,7 Amal Rayan,8 Marwa Ahmed Mahmoud,9 Asmaa Nafady10 1Department of Tropical Medicine and Gastroenterology, Qena Faculty of Medicine, South Valley University, Qena, Egypt; 2Department of Microbiology and Immunology, Faculty of Medicine, Assiut University, Assiut, Egypt; 3Department of Clinical Biochemistry, College of Medicine, King Khalid University, Abha, Kingdom of Saudi Arabia; 4Department of Biochemistry, Faculty of Medicine, Minia University, Minia, Egypt; 5Department of Clinical and Chemical Pathology Sohag Faculty of Medicine Sohag University, Sohag, Egypt; 6Department of Medical Microbiology & Immunology, Faculty of Medicine, Sohag University, Sohag, Egypt; 7Department of Internal Medicine, Faculty of Medicine, Sohag University, Sohag, Egypt; 8Department of Clinical Oncology, Faculty of Medicine, Assiut University, Assiut, Egypt; 9Department of Medical Physiology, Faculty of Medicine, Sohag University, Sohag, Egypt; 10Department of Clinical and Chemical Pathology, Qena Faculty of Medicine, South Valley University, Qena, EgyptCorrespondence: Heba Ahmed Osman, Tropical Medicine and Gastroenterology, Qena Faculty of Medicine, South Valley University, Qena, Egypt, Email drheba.saleh@med.svu.edu.eg; hebaahmed198098@yahoo.comIntroduction: HCC is frequently diagnosed late, when only palliative treatment is available. So, we try to use different immunological markers to identify early HCC in patients with unremarkable raised AFP.Methods: This study was conducted on 112 participants divided into two equal groups: Group I, 56 patients with liver cirrhosis and different stages of HCC; Group II, 56 patients with liver cirrhosis. The diagnosis of HCC was based on AASLD guidelines. TNM and BCLC classification systems are used for staging of HCC.Results: A significant reduction in the median percentage of lymphocyte subset (CD3+, CD4+, CD8+, CD19+) and NK cell percentage (CD56+) has been detected in HCC patients (all P < 0.001). In the HCC group the median monocyte subpopulations CD14+ CD16− Classical, CD14++ CD16+ Intermediate, and CD14−+ CD16++ Non-Classical were 11.7, 4.0, and 3.5, respectively, with marked reduction compared with liver cirrhosis group (all P < 0.001). Patients with advanced stages (BCLC C and D) were more likely to have significantly higher median CD33+ than patients with early stages (BCLC A and B) (P = 0.05); also, the median levels of HLA DR+ lymphocytes % in the HCC case group were 21.8 in patients with advanced disease (BCLC C and D) and 13.1 in patients with early stages of the disease (P = 0.04). Patients with late stage (TNM III) were more likely to have significantly higher median CD14+ CD16− Classical monocyte subset, CD36+ HLA DR+, and CD36+ CD16− than patients with early stages (TNM I and II).Conclusion: Patients with HCC with unremarkable raised AFP showed marked reduction in lymphocytes, natural killer cells, and all monocyte subpopulations. In addition, patients with advanced HCC showed increased CD33+ and HLA DR+ lymphocytes %, CD14+ CD16− Classical monocyte subset, CD36+ HLA DR+, and CD36+ CD16− compared with patients with early stages of HCC.Keywords: monocyte subpopulation, lymphocyte subset, hepatocellular carcinoma, Barcelona clinic liver cancer, tumor (T), nodes (N), metastases (M)

E-cigarette usage continues to rise, yet the safety of e-cigarette aerosols is questioned. Using murine models of acute and chronic e-cigarette aerosol inhalation, murine colon transcriptomics, and murine and human gut-derived organoids in co-culture models, we assessed the effects of e-cigarette use on the gut barrier. Histologic and transcriptome analyses revealed that chronic, but not acute, nicotine-free e-cigarette use increased inflammation and reduced expression of tight junction (TJ) markers. Exposure of murine and human enteroid-derived monolayers (EDMs) to nicotine-free e-cigarette aerosols alone or in co-culture with bacteria also causes barrier disruption, downregulation of TJ protein, and enhanced inflammation in response to infection. These data highlight the harmful effects of "non-nicotine" component of e-cigarettes on the gut barrier. Considering the importance of an intact gut barrier for host fitness and the impact of gut mucosal inflammation on a multitude of chronic diseases, these findings are broadly relevant to both medicine and public health.

PDZ domains are one of the most abundant protein domains in eukaryotes and are frequently found on junction-localized scaffold proteins. Various signaling molecules bind to PDZ proteins via PDZ-binding motifs (PBM) and fine-tune cellular signaling. However, how such interaction affects protein function is difficult to predict and must be solved empirically. Here we describe a long isoform of the guanine nucleotide exchange factor GIV/Girdin (CCDC88A) that we named GIV-L, which is conserved throughout evolution, from invertebrates to vertebrates, and contains a PBM. Unlike GIV, which lacks PBM and is cytosolic, GIV-L localizes onto cell junctions and has a PDZ interactome (as shown through annotating Human Cell Map and BioID-proximity labeling studies), which impacts GIV-L's ability to bind and activate trimeric G-protein, Gαi, through its guanine-nucleotide exchange modulator (GEM) module. This GEM module is found exclusively in vertebrates. We propose that the two functional modules in GIV may have evolved sequentially: the ability to bind PDZ proteins via the PBM evolved earlier in invertebrates, whereas G-protein binding and activation may have evolved later only among vertebrates. Phenotypic studies in Caco-2 cells revealed that GIV and GIV-L may have antagonistic effects on cell growth, proliferation (cell cycle), and survival. Immunohistochemical analysis in human colon tissues showed that GIV expression increases with a concomitant decrease in GIV-L during cancer initiation. Taken together, these findings reveal how regulation in GIV/CCDC88A transcript helps to achieve protein modularity, which allows the protein to play opposing roles either as a tumor suppressor (GIV-L) or as an oncogene (GIV).

ABSTRACT: E-cigarette and vaping device use continue to rise, particularly in adolescents and young adults, but the safety of inhaling the multitude of chemicals within e-cigarette aerosols has been questioned. While several studies have evaluated vaping effects on the lungs and heart; effects on the gastrointestinal tract remain unknown. Using established murine models of acute (1 week) and chronic (3 month) daily e-cigarette aerosol inhalation, both with nicotine-containing and vehicle control e-liquids, murine colon transcriptomics and organoid co-culture models, we assessed the effects of e-cigarette use on the gut barrier and mucosal health. Histologic analyses revealed that chronic exposure to nicotine-free e-cigarette aerosols induced mucosal inflammation. Transcriptome analyses revealed that chronic, but not acute, nicotine-free e-cigarette use significantly reduced expression of tight junction markers, including occluding, and drove expression of pro-inflammatory cytokines. Exposure of murine and human enteroid-derived monolayers (EDMs) to nicotine-free e-cigarette aerosols alone, or in co-culture with invasiveE. coli,confirmed that repetitive exposure was sufficient to recapitulate the key findings observedin vivo, i.e., barrier-disruption, downregulation of occludin, inflammation, and an accentuated risk of and response to bacterial infection. These data highlight an unexpected harmful effect of e-cigarette use on the gut barrier and pinpoint non-nicotine chemical components common across >90% of e-cigarette e-liquids as the source of harm. Given the ever-expanding importance of the integrity of the gut barrier for host fitness, and impact of gut mucosal inflammation on a multitude of chronic diseases, these findings are broadly relevant to medicine and public health. SIGNIFICANCE: The safety of electronic cigarettes has been questioned amidst emerging evidence that they may derail our immune system and increase our susceptibility to infections. Despite these insights, their impact on the most critical entity that separates trillions of microbes from the largest immune system in our body, i.e., the gut barrier, remains unexplored. Using a combination of mouse models, gut transcriptomics, and murine and human gut-derived organoids, here we show that chronic exposure to aerosols of electronic-cigarette disrupts the gut barrier, increases its susceptibility to bacterial infections and triggers inflammation. Given the importance of the gut barrier in the maintenance of immune homeostasis, these findings provide valuable insights into the potential long-term harmful effects of electronic cigarettes on health.

Besides being regulated by G-protein-coupled receptors, the activity of heterotrimeric G proteins is modulated by many cytoplasmic proteins. GIV/Girdin and DAPLE (Dvl-associating protein with a high frequency of leucine) are the best-characterized members of a group of cytoplasmic regulators that contain a Gα-binding and -activating (GBA) motif and whose dysregulation underlies human diseases, including cancer and birth defects. GBA motif-containing proteins were originally reported to modulate G proteins by binding Gα subunits of the Gi/o family (Gαi) over other families (such as Gs, Gq/11, or G12/13), and promoting nucleotide exchange in vitro However, some evidence suggests that this is not always the case, as phosphorylation of the GBA motif of GIV promotes its binding to Gαs and inhibits nucleotide exchange. The G-protein specificity of DAPLE and how it might affect nucleotide exchange on G proteins besides Gαi remain to be investigated. Here, we show that DAPLE's GBA motif, in addition to Gαi, binds efficiently to members of the Gs and Gq/11 families (Gαs and Gαq, respectively), but not of the G12/13 family (Gα12) in the absence of post-translational phosphorylation. We pinpointed Met-1669 as the residue in the GBA motif of DAPLE that diverges from that in GIV and enables better binding to Gαs and Gαq Unlike the nucleotide-exchange acceleration observed for Gαi, DAPLE inhibited nucleotide exchange on Gαs and Gαq These findings indicate that GBA motifs have versatility in their G-protein-modulating effect, i.e. they can bind to Gα subunits of different classes and either stimulate or inhibit nucleotide exchange depending on the G-protein subtype.

Polarized distribution of organelles and molecules inside a cell is vital for a range of cellular processes and its loss is frequently encountered in disease. Polarization during planar cell migration is a special condition in which cellular orientation is triggered by cell-cell contact. We demonstrate that the protein Daple (CCDC88C) is a component of cell junctions in epithelial cells which serves like a cellular "compass" for establishing and maintaining contact-triggered planar polarity. Furthermore, these processes may be mediated through interaction with the polarity regulator PARD3. This interaction, mediated by Daple's PDZ-binding motif (PBM) and the third PDZ domain of PARD3, is fine-tuned by tyrosine phosphorylation on Daple's PBM by receptor and non-receptor tyrosine kinases, such as Src. Hypophosphorylation strengthens the interaction, whereas hyperphosphorylation disrupts it, thereby revealing an unexpected role of Daple as a platform for signal integration and gradient sensing for tyrosine-based signals within the planar cell polarity pathway.

Previously, Aznar et al., showed that Daple/CCDC88C enables Wnt receptors to transactivate trimeric G-proteins during non-canonical Wnt signaling via a novel G-protein binding and activating (GBA) motif. By doing so, Daple serves two opposing roles; earlier during oncogenesis it suppresses neoplastic transformation and tumor growth, but later it triggers epithelial-to-mesenchymal-transition (EMT). We have identified and characterized two isoforms of the human Daple gene. While both isoforms cooperatively suppress tumor growth via their GBA motif, only the full-length transcript triggers EMT and invasion. Both isoforms are suppressed during colon cancer progression, and their reduced expression carries additive prognostic significance. These findings provide insights into the opposing roles of Daple during cancer progression and define the G-protein regulatory GBA motif as one of the minimal modules essential for Daple's role as a tumor suppressor.

The need for improvements in the water use efficiency by agricultural ecosystems requires a holistic assessment of the hydraulic functioning of cropped soils, taking into consideration the most relevant interactions and feedbacks that control the soil water budget. We implemented a mechanistic approach to isolate the effects of soil hydraulic properties (K-θ-h) of layered soils on water balance components and land and water productivity, adopting comprehensive scenarios of soil water availability and requirements. The agro-hydrological simulations were performed using the SWAP model integrated with the WOFOST crop growth module. The simulated scenarios included the rainfed crop growth of maize and soybean in three climate zones, evaluating the current climate scenarios as well as two future scenarios, a wetter and a drier one, totaling 108 scenarios simulated for 30 years each. Simulations were performed for six soils, grouped pairwise (3 × 2), where each pair represented the same soil group with two different long-term land uses: natural forest (proxy of a no-tillage system) and conventional agricultural use. The K-θ-h relationships were obtained simultaneously by inverse modeling for the full range of soil water contents commonly found in the domain of crop available water. The agro-hydrological simulations showed that the soil hydraulic properties affect dynamically water balance components and land productivity by relating soil hydraulic functioning to climate patterns and crop water requirements. In general, maize productivity was more sensitive to soil hydraulic properties under future climate scenarios than soybean. While land productivities of maize and soybean increased under the wetter climate scenario, water productivity of both crops was consistently reduced by both future climate scenarios. The K-θ-h of soils under conventional agricultural use over-performed their counterparts under long-term natural forest use, especially regarding land productivity during growing seasons with pronounced dry spells. Depending on the length and timing of drought stress during the growing season, the yield response is determined by soil-specific conditions strictly related to water availability. The long-term average revealed that the sampled loamy sand soils have more favorable hydraulic properties for crop growth; moreover, the reduced unproductive water losses, especially runoff, increased the dynamic water storage of those soils.

A loss of balance between G protein activation and deactivation has been implicated in the initiation of melanomas, and non-canonical Wnt signaling via the Wnt5A/Frizzled (FZD) pathway has been shown to be critical for the switch to an invasive phenotype. Daple [CCDC88C], a cytosolic guanine nucleotide exchange modulator (GEM) which enhances non-canonical Wnt5A/FZD signaling via activation of trimeric G protein, Gαi, has been shown to serve opposing roles-as an inducer of EMT and invasiveness and a potent tumor suppressor-via two isoforms, V1 (full-length) and V2 (short spliced isoform), respectively. Here we report that the relative abundance of these isoforms in the peripheral circulation, presumably largely from circulating tumor cells (CTCs), is a prognostic marker of cutaneous melanomas. Expression of V1 is increased in both the early and late clinical stages (p

Kirsty Sands,1,2 Owen B Spiller,2 Kathryn Thomson,1,2 Edward AR Portal,2 Kenneth C Iregbu,3 Timothy R Walsh1,2 1Ineos Institute of Antimicrobial Research, Department of Zoology, University of Oxford, Oxford, UK; 2Division of Infection and Immunity, Cardiff University, Cardiff, UK; 3National Hospital Abuja, Abuja, NigeriaCorrespondence: Kirsty Sands, Email kirsty.sands@zoo.ox.ac.ukAbstract: Neonatal sepsis is defined as a systemic infection within the first 28 days of life, with early-onset sepsis (EOS) occurring within the first 72h, although the definition of EOS varies in literature. Whilst the global incidence has dramatically reduced over the last decade, neonatal sepsis remains an important cause of neonatal mortality, highest in low- and middle-income countries (LMICs). Symptoms at the onset of neonatal sepsis can be subtle, and therefore EOS is often difficult to diagnose from clinical presentation and laboratory testing and blood cultures are not always conclusive or accessible, especially in resource limited countries. Although the World Health Organisation (WHO) currently advocates a ß-lactam, and gentamicin for first line treatment, availability and cost influence the empirical antibiotic therapy administered. Antibiotic treatment of neonatal sepsis in LMICs is highly variable, partially caused by factors such as cost of antibiotics (and who pays for them) and access to certain antibiotics. Antimicrobial resistance (AMR) has increased considerably over the past decade and this review discusses current microbiology data available in the context of the diagnosis, and treatment for EOS. Importantly, this review highlights a large variability in data availability, methodology, availability of diagnostics, and aetiology of sepsis pathogens.Keywords: early-onset neonatal sepsis, low- and middle-income countries, antimicrobial resistance, diagnosis, treatment

Summary: Upon sensing DNA double-strand breaks (DSBs), eukaryotic cells either die or repair DSBs via one of the two competing pathways, i.e., non-homologous end-joining (NHEJ) or homologous recombination (HR). We show that cell fate after DSBs hinges on GIV/Girdin, a guanine nucleotide-exchange modulator of heterotrimeric Giα•βγ protein. GIV suppresses HR by binding and sequestering BRCA1, a key coordinator of multiple steps within the HR pathway, away from DSBs; it does so using a C-terminal motif that binds BRCA1’s BRCT-modules via both phospho-dependent and -independent mechanisms. Using another non-overlapping C-terminal motif GIV binds and activates Gi and enhances the “free” Gβγ→PI-3-kinase→Akt pathway, which promotes survival and is known to suppress HR, favor NHEJ. Absence of GIV, or loss of either of its C-terminal motifs enhanced cell death upon genotoxic stress. Because GIV selectively binds other BRCT-containing proteins suggests that G-proteins may fine-tune sensing, repair, and survival after diverse types of DNA damage.

Cellular proliferation, differentiation, and morphogenesis are shaped by multiple signaling cascades, and their dysregulation plays an integral role in cancer progression. Three cascades that contribute to oncogenic potential are those mediated by Wnt proteins and the receptor Frizzled (FZD), growth factor receptor tyrosine kinases (RTKs), and heterotrimeric G proteins and associated GPCRs. Daple is a guanine nucleotide exchange factor (GEF) for the G protein Gαi Daple also binds to FZD and the Wnt/FZD mediator Dishevelled (Dvl), and it enhances β-catenin-independent Wnt signaling in response to Wnt5a-FZD7 signaling. We identified Daple as a substrate of multiple RTKs and non-RTKs and, hence, as a point of convergence for the three cascades. We found that phosphorylation near the Dvl-binding motif in Daple by both RTKs and non-RTKs caused Daple/Dvl complex dissociation and augmented the ability of Daple to bind to and activate Gαi, which potentiated β-catenin-independent Wnt signals and stimulated epithelial-mesenchymal transition (EMT) similarly to Wnt5a/FZD7 signaling. Although Daple acts as a tumor suppressor in the healthy colon, the concurrent increased abundance of Daple and epidermal growth factor receptor (EGFR) in colorectal tumors was associated with poor patient prognosis. Thus, the Daple-dependent activation of Gαi and the Daple-dependent enhancement of β-catenin-independent Wnt signals are not only stimulated by Wnt5a/FZD7 to suppress tumorigenesis but also hijacked by growth factor-activated RTKs to enhance tumor progression. These findings identify a cross-talk paradigm among growth factor RTKs, heterotrimeric G proteins, and the Wnt/FZD pathway in cancer.

Cellular proliferation is antagonistically regulated by canonical and noncanonical Wnt signals; their dysbalance triggers cancers. We previously showed that a multimodular signal transducer, Daple, enhances PI3-K→Akt signals within the noncanonical Wnt signaling pathway and antagonistically inhibits canonical Wnt responses. Here we demonstrate that the PI3-K→Akt pathway serves as a positive feedback loop that further enhances noncanonical Wnt signals by compartmentalizing β-catenin. By phosphorylating the phosphoinositide- (PI) binding domain of Daple, Akt abolishes Daple's ability to bind PI3-P-enriched endosomes that engage dynein motor complex for long-distance trafficking of β-catenin/E-cadherin complexes to pericentriolar recycling endosomes (PCREs). Phosphorylation compartmentalizes Daple/β-catenin/E-cadherin complexes to cell-cell contact sites, enhances noncanonical Wnt signals, and thereby suppresses colony growth. Dephosphorylation compartmentalizes β-catenin on PCREs, a specialized compartment for prolonged unopposed canonical Wnt signaling, and enhances colony growth. Cancer-associated Daple mutants that are insensitive to Akt mimic a constitutively dephosphorylated state. This work not only identifies Daple as a platform for cross-talk between Akt and the noncanonical Wnt pathway but also reveals the impact of such cross-talk on tumor cell phenotypes that are critical for cancer initiation and progression.

Canonical signal transduction via heterotrimeric G proteins is spatiotemporally restricted, i.e., triggered exclusively at the plasma membrane, only by agonist activation of G protein-coupled receptors via a finite process that is terminated within a few hundred milliseconds. Recently, a rapidly emerging paradigm has revealed a noncanonical pathway for activation of heterotrimeric G proteins via the nonreceptor guanidine-nucleotide exchange factor, GIV/Girdin. Biochemical, biophysical, and functional studies evaluating this pathway have unraveled its unique properties and distinctive spatiotemporal features. As in the case of any new pathway/paradigm, these studies first required an in-depth optimization of tools/techniques and protocols, governed by rationale and fundamentals unique to the pathway, and more specifically to the large multimodular GIV protein. Here we provide the most up-to-date overview of protocols that have generated most of what we know today about noncanonical G protein activation by GIV and its relevance in health and disease. © 2016 by John Wiley & Sons, Inc.

BackgroundExtensive literature support telehealth as a supplement or adjunct to in-person care for the management of chronic conditions such as congestive heart failure (CHF) and type 2 diabetes mellitus (T2DM). Evidence is needed to support the use of telehealth as an equivalent and equitable replacement for in-person care and to assess potential adverse effects. ObjectiveWe conducted a systematic review to address the following question: among adults, what is the effect of synchronous telehealth (real-time response among individuals via phone or phone and video) compared with in-person care (or compared with phone, if synchronous video care) for chronic management of CHF, chronic obstructive pulmonary disease, and T2DM on key disease-specific clinical outcomes and health care use? MethodsWe followed systematic review methodologies and searched two databases (MEDLINE and Embase). We included randomized or quasi-experimental studies that evaluated the effect of synchronously delivered telehealth for relevant chronic conditions that occurred over ≥2 encounters and in which some or all in-person care was supplanted by care delivered via phone or video. We assessed the bias using the Cochrane Effective Practice and Organization of Care risk of bias (ROB) tool and the certainty of evidence using the Grading of Recommendations Assessment, Development, and Evaluation. We described the findings narratively and did not conduct meta-analysis owing to the small number of studies and the conceptual heterogeneity of the identified interventions. ResultsWe identified 8662 studies, and 129 (1.49%) were reviewed at the full-text stage. In total, 3.9% (5/129) of the articles were retained for data extraction, all of which (5/5, 100%) were randomized controlled trials. The CHF study (1/5, 20%) was found to have high ROB and randomized patients (n=210) to receive quarterly automated asynchronous web-based review and follow-up of telemetry data versus synchronous personal follow-up (in-person vs phone-based) for 1 year. A 3-way comparison across study arms found no significant differences in clinical outcomes. Overall, 80% (4/5) of the studies (n=466) evaluated synchronous care for patients with T2DM (ROB was judged to be low for 2, 50% of studies and high for 2, 50% of studies). In total, 20% (1/5) of the studies were adequately powered to assess the difference in glycosylated hemoglobin level between groups; however, no significant difference was found. Intervention design varied greatly from remote monitoring of blood glucose combined with video versus in-person visits to an endocrinology clinic to a brief, 3-week remote intervention to stabilize uncontrolled diabetes. No articles were identified for chronic obstructive pulmonary disease. ConclusionsThis review found few studies with a variety of designs and interventions that used telehealth as a replacement for in-person care. Future research should consider including observational studies and studies on additional highly prevalent chronic diseases.

Objective: To evaluate the Impact of physical rehabilitation Interventions, supplemented with one or more adherence-enhancing components, on outcomes among adults with hip or knee osteoarthritis or chronic lower back pain. Design: Primary literature search from inception of each data-base to July 27, 2021, guided by relevant search terms and keywords to search titles and abstracts. All articles meeting eligibility criteria were included for data abstraction. Data Sources: MEDLINE, CINAHL Complete, and Embase. Eligibility Criteria for Selecting Studies: Randomized and nonrandomized trials evaluating adherence-focused intervention components conducted in addition to an index usual care or usual care-like physical rehabilitation program among adults with hip or knee osteoarthritis or chronic low back pain. Eligible studies included a comparator group of the same index physical rehabilitation intervention without the adjunctive adherence components. Included studies measured outcomes at least 3 months after the rehabilitation course. Results: Of the 10 studies meeting inclusion criteria, 6 interventions were delivered concurrent to an index rehabilitation program and 4 were delivered sequentially. Of the 3 studies that reported a positive effect on long-term adherence, only 1 was a low risk of bias study. There is very limited evidence of a beneficial treatment effect of adjunct adherence interventions on long-term physical function, self-efficacy, or adverse events. Conclusion: We found inadequate evidence evaluating adherence-enhancing interventions for the specific promotion of long-term adherence to home rehabilitation programs. Future studies should consider testing interventions specifically built to target behavioral maintenance of home rehabilitation programs. [ABSTRACT FROM AUTHOR]

Ibrahim M Sayed,1,* Mohamed A El-Mokhtar,1,* Mahmoud Abdel Rahman Mahmoud,2 Amal A Elkhawaga,1 Shereen Gaber,3 Nermien H Seddek,4 Lobna Abdel-Wahid,2 Ahmed M Ashmawy,2 Enas Ahmed Reda Alkareemy2 1Department of Medical Microbiology and Immunology, Faculty of Medicine, Assiut University, Assiut, Egypt; 2Department of Internal Medicine, Gastroenterology and Hepatology unit, Faculty of Medicine, Assiut University, Assiut, 71515, Egypt; 3Department of Biochemistry, Faculty of Medicine, Minia University, Minia, Egypt; 4Department of Respiratory Care, College of Applied Medical Sciences-Jubail 4030 (CAMSJ), Imam Abdulrahman Bin Faisal University, Al Jubail 35816, Saudi Arabia*These authors contributed equally to this workCorrespondence: Mohamed A El-Mokhtar; Ibrahim M SayedDepartment of Microbiology and Immunology, Faculty of Medicine, Assiut University, Assiut, EgyptEmail elmokhtarma@aun.edu.eg; Ibrahim.ibrahim@aun.edu.egBackground: Hepatitis E virus (HEV) is an emerging infectious agent that causes acute hepatitis in developing and developed countries. Diagnosis of HEV infection has not been routinely done in Egyptian hospitals, and clinicians do not prescribe ribavirin (RBV) for acute hepatitis cases of unknown etiology (AHUE). We aimed to screen patients with AHUE for the presence of HEV markers and to determine the complications associated with HEV infection.Patients and Methods: HEV markers (anti-HEV IgM, anti-HEV IgG, and HEV RNA) were assessed in patients with AHUE (n=300) admitted to Assiut University Hospitals. RT-qPCR was used to detect the viral load and sequencing analysis was carried out to determine the genotype of the detected viruses. Phylogenetic tree was constructed to evaluate the genetic relatedness between the isolates. Laboratory parameters and the outcomes of infection were determined.Results: Acute HEV infection (AHE) was detected in 30 out of 300 (10%) of AHUE patients. Anti-HEV IgM, HEV RNA, and anti-HEV IgG were reported in 83%, 50%, and 43% of the samples, respectively. HEV RNA load ranged from 5× 102 IU/mL to 1.1× 104 IU/mL. Sequencing of the isolated viruses revealed that five viruses belong to HEV-1 and one isolate belongs to HEV-3 with high homology to the virus recently isolated from the cow and goat milk in the Egyptian villages. Although previous reports showed that attenuated HEV isolates were circulating in Egypt, four out of 30 patients (13%) developed coagulopathy and hepatic encephalopathy and died due to fulminant hepatic failure (FHF) within 3– 6 weeks of hospitalization. Age, malignancy, and a history of pre-existing liver diseases were a risky factor for FHF development.Conclusion: AHE is common in Upper Egypt. Older patients with malignancy and/or a history of liver diseases are risky. HEV diagnosis and treatment become pivotal in Egyptian hospitals to reduce the fatality rate and they should start urgently and promptly.Keywords: acute hepatitis E, diagnosis, fulminant hepatitis failure, ribavirin, Egypt

We previously showed that guanine nucleotide-binding (G) protein α subunit (Gα)-interacting vesicle-associated protein (GIV), a guanine-nucleotide exchange factor (GEF), transactivates Gα activity-inhibiting polypeptide 1 (Gαi) proteins in response to growth factors, such as EGF, using a short C-terminal motif. Subsequent work demonstrated that GIV also binds Gαs and that inactive Gαs promotes maturation of endosomes and shuts down mitogenic MAPK-ERK1/2 signals from endosomes. However, the mechanism and consequences of dual coupling of GIV to two G proteins, Gαi and Gαs, remained unknown. Here we report that GIV is a bifunctional modulator of G proteins; it serves as a guanine nucleotide dissociation inhibitor (GDI) for Gαs using the same motif that allows it to serve as a GEF for Gαi. Upon EGF stimulation, GIV modulates Gαi and Gαs sequentially: first, a key phosphomodification favors the assembly of GIV-Gαi complexes and activates GIV's GEF function; then a second phosphomodification terminates GIV's GEF function, triggers the assembly of GIV-Gαs complexes, and activates GIV's GDI function. By comparing WT and GIV mutants, we demonstrate that GIV inhibits Gαs activity in cells responding to EGF. Consequently, the cAMP→PKA→cAMP response element-binding protein signaling axis is inhibited, the transit time of EGF receptor through early endosomes are accelerated, mitogenic MAPK-ERK1/2 signals are rapidly terminated, and proliferation is suppressed. These insights define a paradigm in G-protein signaling in which a pleiotropically acting modulator uses the same motif both to activate and to inhibit G proteins. Our findings also illuminate how such modulation of two opposing Gα proteins integrates downstream signals and cellular responses.

A long-standing issue in the field of signal transduction is to understand the cross-talk between receptor tyrosine kinases (RTKs) and heterotrimeric G proteins, two major and distinct signaling hubs that control eukaryotic cell behavior. Although stimulation of many RTKs leads to activation of trimeric G proteins, the molecular mechanisms behind this phenomenon remain elusive. We discovered a unifying mechanism that allows GIV/Girdin, a bona fide metastasis-related protein and a guanine-nucleotide exchange factor (GEF) for Gαi, to serve as a direct platform for multiple RTKs to activate Gαi proteins. Using a combination of homology modeling, protein-protein interaction, and kinase assays, we demonstrate that a stretch of ∼110 amino acids within GIV C-terminus displays structural plasticity that allows folding into a SH2-like domain in the presence of phosphotyrosine ligands. Using protein-protein interaction assays, we demonstrated that both SH2 and GEF domains of GIV are required for the formation of a ligand-activated ternary complex between GIV, Gαi, and growth factor receptors and for activation of Gαi after growth factor stimulation. Expression of a SH2-deficient GIV mutant (Arg 1745→Leu) that cannot bind RTKs impaired all previously demonstrated functions of GIV-Akt enhancement, actin remodeling, and cell migration. The mechanistic and structural insights gained here shed light on the long-standing questions surrounding RTK/G protein cross-talk, set a novel paradigm, and characterize a unique pharmacological target for uncoupling GIV-dependent signaling downstream of multiple oncogenic RTKs.

Gastroenteropancreatic neuroendocrine carcinomas are rare neoplasms with no effective treatments and poor prognosis. Few reliable preclinical models exist for the study of gastroenteropancreatic neuroendocrine carcinomas, limiting investigation of novel treatments. We used tumor spheroids from our recently established gastroenteropancreatic neuroendocrine carcinoma patient-derived xenograft models to systematically screen for compounds with diverse structures to identify potential new categories of therapeutic agents that can target gastroenteropancreatic neuroendocrine carcinomas. Tumor spheroids were derived from our NEC913 and NEC1452 gastroenteropancreatic neuroendocrine carcinoma patient-derived xenograft models. Gastroenteropancreatic neuroendocrine carcinoma spheroids were screened against a library of 885 compounds from the National Cancer Institute Diversity Set VII collection. Cell viability was measured via AlamarBlue assay. After identification of potential therapeutic compounds, synergy screening of a selected group with temozolomide and doxorubicin was performed, and these combinations were further analyzed for γH2AX and phosphorylated-ERK proteins. We identified 16 compounds that inhibit over 75% of gastroenteropancreatic neuroendocrine carcinoma spheroid survival. Seven are inhibitors of tyrosyl-DNA phosphodiesterase 1, a DNA repair enzyme working closely with the topoisomerase I complex. When combined with temozolomide or doxorubicin, the tyrosyl-DNA phosphodiesterase 1 inhibitor cytarabine increased the cytotoxic effects of these drugs on NEC1452 cells which was further evidenced by increasing γH2AX and decreasing phosphorylated-ERK in combination treatment compared to temozolomide alone. Both NEC913 and NEC1452 gastroenteropancreatic neuroendocrine carcinoma spheroid lines are useful preclinical models for drug testing. Our library screen revealed these gastroenteropancreatic neuroendocrine carcinoma spheroids are highly sensitive to a novel class of anti-cancer drugs that target nuclear genome stability. [ABSTRACT FROM AUTHOR]

Serotonin is emerging as a promising therapeutic target in tryptophan hydroxylase 1–positive tumors, but further mechanistic studies are needed to effectively target dysregulated serotonin metabolism. One challenge is a lack of methods for studying the dynamic nature of serotonin metabolism. Here, we report the development of a genetically encoded luminescent biosensor, termed iSero-Rluc, for the real-time detection of serotonin in live cells. The engineered serotonin binding domain (iSero) and Renilla luciferase (Rluc) reporter were cloned into yeast and mammalian expression vectors to create a fusion protein that could act as a biosensor to detect endogenous serotonin levels in live cells. The iSero-Rluc biosensor was stably expressed in the BON cell line and luciferase assays, mass spectroscopy, immunofluorescence, and Western blotting were used to study serotonin metabolism under different cell culture conditions. The iSero-Rluc sensor detected exogenous serotonin in a yeast model. When stably expressed in the BON cell line, iSero-Rluc revealed that serotonin biosynthesis is increased in an anchorage-independent growth state and is induced upon serum starvation. The iSero-Rluc biosensor is a powerful tool in the study of tumor serotonin metabolism. It enabled real-time detection of alterations in serotonin synthesis in living cells under various growth conditions and has the potential to provide greater insight into serotonin metabolism in different stages of tumor progression and to identify therapeutic strategies to target cancer metastases and carcinoid crises. [ABSTRACT FROM AUTHOR]

Background: Silver nanoparticle was developed to overcome the drawback of silver diamine fluoride. However, evidence is limited, especially in root caries. The aim of this study was to evaluate the remineralization effect of silver nanoparticles on root caries. Materials and Methods: Fifty‐five root human dentin slices size 5 × 5 mm2 from patients aged over 60 years old were immersed in demineralized solution to create artificial caries. Specimens were allocated into five groups according to the remineralizing agents: silver diamine fluoride (SDF), silver nanoparticles solution (AgNPs), silver nanoparticle solution followed by sodium fluoride varnish (AgNPs+NaF), sodium fluoride varnish (NaF), and tap water. After 8 days of pH‐cycling challenge, the microhardness test, lesion depth evaluation, dentin surface morphology, and elemental analysis were performed. Data was analysed using F‐test One‐way ANOVA followed by Tukey's post hoc test and paired T‐test. Results: All test groups demonstrated a significantly higher microhardness value and lower lesion depth compared with the control group. AgNPs+NaF and NaF‐treated groups showed lower efficacy than SDF. Crystal precipitation was presented in all groups composed of silver. Conclusion: Addition of fluoride varnish did not benefit for silver nanoparticles in preventing further demineralization. SDF provides the highest effectiveness in elderly root carious dentin. [ABSTRACT FROM AUTHOR]

Summary: E-cigarette usage continues to rise, yet the safety of e-cigarette aerosols is questioned. Using murine models of acute and chronic e-cigarette aerosol inhalation, murine colon transcriptomics, and murine and human gut-derived organoids in co-culture models, we assessed the effects of e-cigarette use on the gut barrier. Histologic and transcriptome analyses revealed that chronic, but not acute, nicotine-free e-cigarette use increased inflammation and reduced expression of tight junction (TJ) markers. Exposure of murine and human enteroid-derived monolayers (EDMs) to nicotine-free e-cigarette aerosols alone or in co-culture with bacteria also causes barrier disruption, downregulation of TJ protein, and enhanced inflammation in response to infection. These data highlight the harmful effects of “non-nicotine” component of e-cigarettes on the gut barrier. Considering the importance of an intact gut barrier for host fitness and the impact of gut mucosal inflammation on a multitude of chronic diseases, these findings are broadly relevant to both medicine and public health.

The synaptic scaffolding proteins CASK and Caskin1 are part of the fibrous mesh of proteins that organize the active zones of neural synapses. CASK binds to a region of Caskin1 called the CASK interaction domain (CID). Adjacent to the CID, Caskin1 contains two tandem sterile α motif (SAM) domains. Many SAM domains form polymers so they are good candidates for forming the fibrous structures seen in the active zone. We show here that the SAM domains of Caskin1 form a new type of SAM helical polymer. The Caskin1 polymer interface exhibits a remarkable segregation of charged residues, resulting in a high sensitivity to ionic strength in vitro. The Caskin1 polymers can be decorated with CASK proteins, illustrating how these proteins may work together to organize the cytomatrix in active zones.

Tidal turbines are entering an industrial phase and farms will soon be installed. In order to optimize the power output of tidal farms, a good understanding of the interactions between the ambient turbulence and a single turbine is crucial. Computational Fluid Dynamics, and more precisely Large Eddy Simulation, is one way of acquiring such knowledge. This study proposed a comparison between a Lattice Boltzmann Method LES approach and a Navier-Stokes LES approach to model the wake of a tidal turbine. Numerical results are compared with experimental results and a relatively good concordance is observed. Differences inherent to the approaches are then pointed out.

Summary: Polarized distribution of organelles and molecules inside a cell is vital for a range of cellular processes and its loss is frequently encountered in disease. Polarization during planar cell migration is a special condition in which cellular orientation is triggered by cell-cell contact. We demonstrate that the protein Daple (CCDC88C) is a component of cell junctions in epithelial cells which serves like a cellular “compass” for establishing and maintaining contact-triggered planar polarity. Furthermore, these processes may be mediated through interaction with the polarity regulator PARD3. This interaction, mediated by Daple's PDZ-binding motif (PBM) and the third PDZ domain of PARD3, is fine-tuned by tyrosine phosphorylation on Daple's PBM by receptor and non-receptor tyrosine kinases, such as Src. Hypophosphorylation strengthens the interaction, whereas hyperphosphorylation disrupts it, thereby revealing an unexpected role of Daple as a platform for signal integration and gradient sensing for tyrosine-based signals within the planar cell polarity pathway. : Biological Sciences; Cell Biology; Functional Aspects of Cell Biology Subject Areas: Biological Sciences, Cell Biology, Functional Aspects of Cell Biology

BackgroundNicotinamide phosphoribosyltransferase (NAMPT) plays a key role in the biosynthesis of nicotinamide adenine dinucleotide (NAD+), which is a vital cofactor in redox reactions and a substrate for NAD+ consuming enzymes including CD38, PARPs and sirtuins. NAMPT over-expression has been shown in various cancers and its inhibition decreases cancer cell growth, making it an attractive therapeutic target. Here we examine the NAMPT expression in a large cohort of resected stage I/II pancreatic ductal adenocarcinomas (PDAs) and correlate its expression with clinical outcomes and pathologic features.MethodsA retrospective review of patients with PDAs was conducted at a single institution. Tissue microarrays (TMAs) containing primary PDAs and their metastatic lymph nodes (mLNs) were constructed and stained for NAMPT expression. Each TMA core was evaluated for staining intensity of cancer cells (0 = no staining, 1+ = weak, 2+ = moderate, 3+ = strong) and a mean score was calculated for each case with at least two evaluable cores. NAMPT expression was correlated with clinicopathological variables using chi-squared or Fisher's exact test, and t-tests for categorical and continuous variables, respectively. Survival probabilities were estimated and plotted using the Kaplan-Meier method. Cox proportional hazards regression was used to assess the effects of NAMPT staining values on recurrence-free survival (RFS) and overall survival (OS). This study was conducted under an approved IRB protocol.Results173 primary PDAs had at least 2 TMA cores with identifiable cancer cells. The mean IHC score was 0.55 (range: 0 to 2.33). The mean IHC score of mLNs was 0.39 (range: 0-2), which was not significantly different from their primary tumors (mean IHC score = 0.47, P = 0.38). Sixty-four percent (111/173) of PDAs were positive for NAMPT staining. Stage II tumors were more likely to be positive (68% of 151 vs 41% of 22; P = 0.01). Non-obese non-diabetic patients were more likely to have NAMPT+ tumors (43.7% vs. 27.9%, P = 0.04). While RFS and OS were not statistically different between NAMPT+ vs. NAMPT- PDAs, patients with NAMPT- tumors tended to have a longer median OS (26.0 vs. 20.4 months, P = 0.34).ConclusionNAMPT expression was detected in 64% of stage I/II PDAs and up to 72% in non-obese non-diabetic patients. Frequency of NAMPT expression correlated with pathological stage, consistent with published literature regarding its role in cancer progression. While RFS and OS were not statistically significantly different, patients with NAMPT+ PDAs tended to have a shorter survival. Thus, NAMPT inhibition may prove beneficial in clinical trials.

Neutrophils play a critical role in innate immunity and also influence adaptive immune responses. This occurs in good part through their production of inflammatory and immunomodulatory cytokines, in conjunction with their prolonged survival at inflamed foci. While a picture of the signaling machinery underlying these neutrophil responses is now emerging, much remains to be uncovered. In this study, we report that neutrophils constitutively express various Src family isoforms (STKs), as well as Syk, and that inhibition of these protein tyrosine kinases selectively hinders inflammatory cytokine generation by acting posttranscriptionally. Accordingly, STK or Syk inhibition decreases the phosphorylation of signaling intermediates (e.g., eIF-4E, S6K, and MNK1) involved in translational control. By contrast, delayed apoptosis appears to be independent of either STKs or Syk. Our data therefore significantly extend our understanding of which neutrophil responses are governed by STKs and Syk and pinpoint some signaling intermediates that are likely involved. In view of the foremost role of neutrophils in several chronic inflammatory conditions, our findings identify potential molecular targets that could be exploited for future therapeutic intervention.