bluuebunny/crossref_metadata_2025 huggingface dataset:
A path recorder algorithm for Multiple Longest Common Subsequences (MLCS) problems
Authors: Wei Shiwei , Wang Yuping , Yang Yuanchao , Liu Sen
Published: February 2020

Motivation

Searching the Longest Common Subsequences of many sequences is called a Multiple Longest Common Subsequence (MLCS) problem which is a very fundamental and challenging problem in many fields of data mining. The existing algorithms cannot be applicable to problems with long and large-scale sequences due to their huge time and space consumption. To efficiently handle large-scale MLCS problems, a Path Recorder Directed Acyclic Graph (PRDAG) model and a novel Path Recorder Algorithm (PRA) are proposed.

Results

In PRDAG, we transform the MLCS problem into searching the longest path from the Directed Acyclic Graph (DAG), where each longest path in DAG corresponds to an MLCS. To tackle the problem efficiently, we eliminate all redundant and repeated nodes during the construction of DAG, and for each node, we only maintain the longest paths from the source node to it but ignore all non-longest paths. As a result, the size of the DAG becomes very small, and the memory space and search time will be greatly saved. Empirical experiments have been performed on a standard benchmark set of both DNA sequences and protein sequences. The experimental results demonstrate that our model and algorithm outperform the related leading algorithms, especially for large-scale MLCS problems.

Availability and implementation

This program code is written by the first author and can be available at https://www.ncbi.nlm.nih.gov/nuccore and https://blog.csdn.net/wswguilin.

Supplementary information

Supplementary data are available at Bioinformatics online.
Breeze: an integrated quality control and data analysis application for high-throughput drug screening
Authors: Potdar Swapnil , Ianevski Aleksandr , Mpindi John-Patrick , Bychkov Dmitrii , Fiere Clément , Ianevski Philipp , Yadav Bhagwan , Wennerberg Krister , Aittokallio Tero , Kallioniemi Olli , Saarela Jani , Östling Päivi
Published: February 2020

High-throughput screening (HTS) enables systematic testing of thousands of chemical compounds for potential use as investigational and therapeutic agents. HTS experiments are often conducted in multi-well plates that inherently bear technical and experimental sources of error. Thus, HTS data processing requires the use of robust quality control procedures before analysis and interpretation. Here, we have implemented an open-source analysis application, Breeze, an integrated quality control and data analysis application for HTS data. Furthermore, Breeze enables a reliable way to identify individual drug sensitivity and resistance patterns in cell lines or patient-derived samples for functional precision medicine applications. The Breeze application provides a complete solution for data quality assessment, dose–response curve fitting and quantification of the drug responses along with interactive visualization of the results.

Availability and implementation

The Breeze application with video tutorial and technical documentation is accessible at https://breeze.fimm.fi; the R source code is publicly available at https://github.com/potdarswapnil/Breeze under GNU General Public License v3.0.

Contact

swapnil.potdar@helsinki.fi

Supplementary information

Supplementary data are available at Bioinformatics online.
Trade-offs between burrowing and biting force in fossorial scincid lizards?
Authors: Le Guilloux Margot , Miralles Aurélien , Measey John , Vanhooydonck Bieke , O’Reilly James C , Lowie Aurélien , Herrel Anthony
Published: February 2020

Trade-offs are thought to be important in constraining evolutionary divergence as they may limit phenotypic diversification. The cranial system plays a vital role in many functions including defensive, territorial, predatory and feeding behaviours in addition to housing the brain and sensory systems. Consequently, the morphology of the cranial system is affected by a combination of selective pressures that may induce functional trade-offs. Limbless, head-first burrowers are thought to be constrained in their cranial morphology as narrow heads may provide a functional advantage for burrowing. However, having a wide and large head is likely beneficial in terms of bite performance. We used 15 skink species to test for the existence of trade-offs between maximal push and bite forces, and explored the patterns of covariation between external head and body morphology and performance. Our data show that there is no evidence of a trade-off between bite and burrowing in terms of maximal force. Species that generate high push forces also generate high bite forces. Our data also show that overall head size covaries with both performance traits. However, future studies exploring trade-offs between force and speed or the energetic cost of burrowing may reveal other trade-offs.
Platysace (Apiaceae) of south-western Australia: silent story tellers of an ancient human landscape
Authors: Lullfitz Alison , Byrne Margaret , Knapp Lynette , Hopper Stephen D
Published: February 2020

High gene flow and a population structure corresponding to human rather than geographical drivers are likely to be genetic patterns of human-dispersed plant taxa. We examined variation in geographical structure and gene flow estimates based on three non-coding regions of plastid DNA in three south-west Australian members of the Platysace genus to identify whether a human influence on dispersion of utilized taxa was detectable. Edible tubers of Platysace deflexa and Platysace trachymenioides have been harvested historically by Noongar traditional owners, whereas Platysace effusa has no known cultural significance. We found differences between utilized and non-utilized taxa, particularly when considered against the generally complex phylogeographical patterning in south-west Australian plant taxa. Platysace effusa showed a pattern of high population divergence, low gene flow and multiple refugia, consistent with a long evolutionary history, past climatic oscillations and persistence in a highly fragmented landscape. In contrast, higher gene flow estimates, less divergence between populations and common haplotypes in P. deflexa and in P. trachymenioides over the south-eastern part of its range are consistent with anthropogenic influences. This study contributes to the understanding of human influences on south-west Australian plant taxa that have been present since the late Pleistocene, but to date have received little scientific attention.
Hydrological and thermal responses of seeds from four co-occurring tree species from southwest Western Australia
Authors: Rajapakshe Rajapakshe P V G S W , Turner Shane R , Cross Adam T , Tomlinson Sean
Published: February 2020

Seed germination is a critical stage in the life cycle of most plants and is defined by specific tolerance thresholds beyond which rates and success of germination rapidly decline. Previous studies have demonstrated that widespread plant species commonly germinate over a broad range of temperatures and water stress levels, whereas range-restricted species often exhibit a narrower germination window in terms of temperature and moisture. We investigated the relationships of the key germination traits of maximum germination (Gmax) and time to 50% germination (t50) in response to temperature (5–35°C) and water stress (−1.5–0 MPa) in four co-occurring Western Australian native Eucalyptus species with widely varying biogeography. Eucalyptus caesia subsp. caesia and E. ornata exhibit a highly localized distribution and a narrow geographical range, being restricted either to granite outcrops or the upper slopes and tops of lateritic rises, respectively. These two species were compared with the two widespread and dominant congenerics E. salmonophloia and E. salubris. There was a distinctive hump-shaped response of t50 to temperature and an exponential response to water stress, characteristic of rate- and threshold-limited processes, but no consistent pattern in the response of Gmax. The four species were significantly different in their thermal performance of t50, with E. caesia and E. ornata displaying narrower thermal tolerance ranges than the two widespread species. In terms of mean final germination percentage, the two range-restricted endemic taxa exhibited higher lability in their response to thermal stress and drought stress compared to the two broadly distributed congenerics. These findings indicate a link between distributional extent, temperature and water stress tolerance and may have implications for identifying ecological filters of rarity and endemism.
Artificial intelligence deciphers codes for color and odor perceptions based on large-scale chemoinformatic data
Authors: Zhang Xiayin , Zhang Kai , Lin Duoru , Zhu Yi , Chen Chuan , He Lin , Guo Xusen , Chen Kexin , Wang Ruixin , Liu Zhenzhen , Wu Xiaohang , Long Erping , Huang Kai , He Zhiqiang , Liu Xiyang , Lin Haotian
Published: February 2020

Background

Color vision is the ability to detect, distinguish, and analyze the wavelength distributions of light independent of the total intensity. It mediates the interaction between an organism and its environment from multiple important aspects. However, the physicochemical basis of color coding has not been explored completely, and how color perception is integrated with other sensory input, typically odor, is unclear.

Results

Here, we developed an artificial intelligence platform to train algorithms for distinguishing color and odor based on the large-scale physicochemical features of 1,267 and 598 structurally diverse molecules, respectively. The predictive accuracies achieved using the random forest and deep belief network for the prediction of color were 100% and 95.23% ± 0.40% (mean ± SD), respectively. The predictive accuracies achieved using the random forest and deep belief network for the prediction of odor were 93.40% ± 0.31% and 94.75% ± 0.44% (mean ± SD), respectively. Twenty-four physicochemical features were sufficient for the accurate prediction of color, while 39 physicochemical features were sufficient for the accurate prediction of odor. A positive correlation between the color-coding and odor-coding properties of the molecules was predicted. A group of descriptors was found to interlink prominently in color and odor perceptions.

Conclusions

Our random forest model and deep belief network accurately predicted the colors and odors of structurally diverse molecules. These findings extend our understanding of the molecular and structural basis of color vision and reveal the interrelationship between color and odor perceptions in nature.
Back to the future: five forgotten lessons for the healthcare managers of today
Authors: Gibiino Giovanni , Rugo Michele , Maffoni Marina , Giardini Anna
Published: February 2020

Nowadays, political, economic and technological upheavals require the rediscovery of some elapsed soft skills by healthcare managers. These abilities are values grounded in the past and somewhat nebulous in the present. We suggest five forgotten lessons highlighting the importance of a stronger human and self-conscious approach in today’s and tomorrow’s healthcare managers.
A15 MICROBES MEDIATE FIBER-INDUCED INFLAMMATION IN IBD
Authors: Armstrong H , Dickner R , Rieger A , Mander I K , Jerasi J , Santer D , Valcheva R , Dijk S I , Petrova A , Dieleman L A , Carroll M W , Huynh H Q , Wine E
Published: February 2020

Background

The etiology of inflammatory bowel diseases (IBD) remains unknown, although gut microorganisms and diet have been implicated. Dietary fibers pass through the bowel undigested and are fermented within the intestine by microbes, promoting gut health. However, many IBD patients describe experiencing sensitivity to fibres. Interestingly, fiber receptors on immune cells are able to interact with fibers typically found on the surface of fungal cells (which share properties with dietary fibers), for example, resulting in a paradoxical pro-inflammatory response.

Aims

As an altered microbial composition is a hallmark of IBD, we hypothesized that the loss of fiber fermenting-microbes populating the IBD gut could lead to dietary fibers not being efficiently broken down into their beneficial biproducts, resulting in binding of intact fibers to pro-inflammatory host cell receptors. This can ultimately drive pro-inflammatory responses and a microenvironment that promotes continued dysbiosis and increased pathogenicity of select microbes, as observed in IBD.

Methods

Fiber receptor expression gut was examined using immunohistochemistry and flow cytometry and demonstrated elevated receptor expression due to increased presence of immune cells in IBD patient biopsies. Cytokine secretion, in response to fiber (5mg/mL) or pre-fermented fibers, cultured with microbes of interest, was measured by ELISAs in cell lines in vitro and biopsy tissues cultured ex vivo.

Results

Whole-fibers induced pro-inflammatory cytokine production in macrophage, monocytes, and neutrophils. Specific microbes were capable of fermenting fiber, measured by gas chromatography. Pre-fermentation of fibers by these microbes reduced inflammatory cytokine production. The fiber oligofructose increased IL-1β in pediatric CD (n=44) and UC (n=29) biopsies cultured ex vivo but not in non-IBD (n=25). The increase was greater in patients with more severe disease. Pre-fermentation of oligofructose by bacteria reduced this secretion of IL-1β. Whole-microbe intestinal washes from severe IBD patients were unable to ferment oligofructose or reduce fiber-associated inflammation in macrophage cells compared to remission or non-IBD children. Statistical analysis of food frequency questionnaire (FFQ) data on fiber consumption demonstrated that fiber-associated inflammation in patient biopsies cultured ex vivo (ELISA and qPCR) correlated with fiber avoidance (FFQ).

Conclusions

Comparing in vitro findings to our patient FFQs, intestinal washes (microbe abundance), and detailed patient history will better define the relationship between microbes, dietary fibers, and gut inflammation in IBD. This will allow for tailored dietary intervention through dietary recommendations, prebiotic, and/or probiotic therapies.

Funding Agencies

CCCWeston Foundation, WCHRI
A30 INHIBITION OF NF-KB SIGNALING IN DCLK1+ CELLS PROMOTES COLONIC INFLAMMATION AND COLITIS-ASSOCIATED CANCER
Authors: Good H , Shin A E , Zhang L , Asfaha S
Published: February 2020

Background

Colorectal cancer (CRC) is the 2nd leading cause of cancer death in Canada. A major risk factor for this disease is chronic inflammation. Despite the clear link between inflammation and cancer, the exact mechanism by which colitis leads to cancer is unknown. Our group has previously shown that a rare cell type in the gut marked by the expression of doublecortin-like kinase-1 (Dclk1) and known as a tuft cell, is quiescent, long-lived, and resistant to proliferation even upon mutation of the tumor suppressor APC. Interestingly, in the setting of colitis, these APC-mutated tuft cells become powerful cancer-initiating cells, but the mechanism by which this occurs is not known. NF-kB signaling is a major inflammatory pathway active in colitis and that has been linked to colorectal cancer. Inhibition of the NF-kB pathway in intestinal epithelial cells has also been shown to inhibit tumor initiation in a mouse model of colitis-associated cancer (Greten et al., 2004).

Aims

In the present study, we aim to examine the effect of NF-kB inhibition in tuft cells on colitis-associated cancer.

Methods

Dclk1CreERT2/APCf/f mice were crossed to IKK-β f/f mice and administered tamoxifen to conditionally knockout the function of both APC and IKK-β in tuft cells. Mice were then exposed to the colitis-inducing agent dextran sodium sulfate (DSS) to induce tumorigenesis. Approximately 16 weeks post-tamoxifen, colonic tumor number and size were analyzed to determine the effect of NF-kB pathway inhibition on tumor initiation and growth, respectively. Extent of inflammation was assessed by myeloperoxidase (MPO) activity and histological damage, and colonic tissue was collected for measurement of inflammatory mediators by qRT-PCR at both acute and long-term time points.

Results

Interestingly, at baseline we detected increased MPO activity in Dclk1CreERT2/APCf/f/IKK-β f/f mice compared to control mice, suggesting that inhibition of NF-kB in Dclk1+ cells may increase basal colonic inflammation. Consistent with this observation, inhibition of the NF-kB pathway also resulted in an increased number of tuft cell-derived tumors, with no observed change in tumor size. Acutely, we also observed an exacerbation of DSS-colitis in Dclk1CreERT2/APCf/f/IKK-β f/f mice, as detected by elevated MPO activity, increased histological damage, and reduced colon length compared to wildtype (IKK-β +/+) controls.

Conclusions

These data suggest that Dclk1+ cell-specific NF-kB signaling plays a key protective role against colitis and colitis-associated tumorigenesis. Targeting the NF-kB pathway may reduce the severity of colitis and the incidence of colitis-associated cancer.

Funding Agencies

CIHR
A32 THE REGULATION OF INTESTINAL SMOOTH MUSCLE CELL PROLIFERATION IN CROHN’S DISEASE – IS NR4A1 A NOVEL TARGET TO TREAT FIBROSTENOSIS?
Authors: Szczepanski H , Tsai Y , Venu V P , Alston L , Hirota S A
Published: February 2020

Background

Intestinal fibrosis and stricture formation are common complications of Crohn’s disease (CD). Recently, the stricturing phenotype has been recognized to be primarily attributable to hypertrophy/hyperplasia of smooth muscle, rather than an increase in fibrosis alone. Despite advances in treatment of CD, current therapies do little to prevent or reverse strictures. NR4A1 is an orphan nuclear receptor that has been reported as anti-fibrotic in non-intestinal systems and exhibits anti-proliferative effects in smooth muscle cells (SMCs). NR4A1 gene variants have been associated with increased risk of IBD, however, the mechanisms regulating NR4A1 expression and its role in intestinal SMC function has not been investigated.

Aims

To characterize and understand the role of NR4A1 activation in the regulation of mitogen-induced proliferation of intestinal SMCs.

Methods

Primary intestinal SMCs were isolated from Nr4a1+/+ and Nr4a1-/- mice. Furthermore, a commercially sourced human primary intestinal SMC line was used. To assess the response of SMCs to culture and identify growth differences, proliferation was measured via trypan blue exclusion and EdU incorporation. In mouse and human SMCs, proliferation was induced by platelet-derived growth factor-BB (PDGF-BB) and NR4A1 activation was assessed by pretreating with selective agonists, cytosporone B (Csn-B) and 6-mercaptopurine (6-MP), at various concentrations. Mass spectrometry was used to characterize proteomic differences between Nr4a1+/+ and Nr4a1-/- SMCs. Expression levels of NR4A1 were assessed by qPCR and western blot after mitogen exposure and Csn-B treatment.

Results

Nr4a1-/- cells exhibited a significantly higher rate of proliferation compared to Nr4a1+/+ cells, under both basal and mitogen-exposed conditions. Proteomic analysis showed that Nr4a1-/- SMCs exhibited increased expression of proteins related to the cell cycle and metabolism, compared to Nr4a1+/+ SMCs. Pretreating human intestinal SMCs with Csn-B and 6-MP significantly attenuated proliferation induced by PDGF-BB. Similar effects were observed in Nr4a1+/+ SMCs, however, the anti-proliferative effect of Csn-B was absent in Nr4a1-/- cells. Furthermore, NR4A1 expression was rapidly induced by Csn-B and PDGF-BB, the latter response suggesting the existence of a potential negative feedback mechanism to control mitogen-induced SMC proliferation.

Conclusions

Our results suggest that NR4A1 is a critical regulator of intestinal SMC proliferation and its induction by mitogens may contribute to a negative feedback loop to control smooth muscle growth. These data support targeting NR4A1 to treat excessive smooth muscle hypertrophy/hyperplasia that contributes to tissue remodelling observed in fibrostenotic CD.

Funding Agencies

CAG, CCC, CIHR
A37 ENTERIC MICROBIOTA CONTRIBUTE TO BEHAVIORAL ALTERATIONS OBSERVED IN MICE WITH COLITIS
Authors: Vicentini F , Griffin L , Keenan C , Cavin J , Nieves K , Hirota S A , Sharkey K A
Published: February 2020

Background

The enteric microbiota has been recognized as an essential regulator of both gut and brain physiology, a complex interaction generally termed the microbiota-gut-brain axis. Disturbances to gastrointestinal physiology lead to alterations in the composition of the enteric microbiota, whereas dysbiosis can also contribute to pathophysiology. Inflammatory bowel diseases (IBD) are chronic, relapsing inflammatory conditions of the gastrointestinal tract, associated with microbial dysbiosis. Interestingly, IBD patients exhibit an increased incidence of mental illness (i.e. anxiety and depression), often termed “sickness behavior”, even during the remitting phase of their disease. Nevertheless, it is unclear if alterations in the enteric microbiota associated with IBD are responsible for the observed modification in brain function and behavior. Here, we hypothesized that sickness behavior is driven by alterations in microbial composition, which occur in the context of intestinal inflammation.

Aims

We sought to determine whether transfer of the microbiota from colitic mice, exhibiting sickness behaviour, into healthy counterparts would induce behavioral changes.

Methods

Male mice (C57Bl/6J; 8 weeks old) were used in all experiments. Colitis was induced by administration of 2.5% dextran sodium sulfate (DSS) in the drinking water for 5 days. Colonic inflammation was assessed by measuring fecal lipocalin-2 and the expression of pro-inflammatory mediators via qPCR. Cecal matter from donor mice (control or DSS treated) were collected for fecal microbiota transplant (FMT). FMT was performed via oral gavage in antibiotic-treated recipient mice. Gut bacteria were evaluated by 16S rRNA sequencing in cecal samples. Anxiety- and depression-like behavior were assessed by elevated plus maze and tail suspension test, respectively.

Results

DSS-treated mice exhibited clinical disease, reflected by body weight loss, increased fecal lipocalin-2 and elevated colonic pro-inflammatory cytokine transcripts. An increase in anxiety-like behavior was observed in mice with colitis, although no alterations in depression-like behavior were detected. Colitic mice exhibited a unique microbial community. Transferring cecal material from colitic mice into recipient, antibiotic-treated mice, recapitulated alterations in behavior seen in colitic donors, as shown by increased anxiety-like behavior and unexpectedly, increased depression-like behavior. These behavioral changes occurred in the absence of colonic or brain inflammation in the recipient mice, but were associated with changes in stress-related gene expression (i.e. Crh).

Conclusions

Colitis-associated sickness behavior can be transmitted to antibiotic-treated recipient mice via FMT, which occurs in the absence of overt intestinal or brain inflammation.

Funding Agencies

CIHRNational Council for Scientific and Technological Development (CNPq-Brazil)
A41 LOSS OF SATB2 EXPRESSION IN COLORECTAL CANCER IS ASSOCIATED WITH DURATION OF INFLAMMATORY BOWEL DISEASE
Authors: Atin J , Hernandez-Rocha C , Borowski K , Stempak J , Smith M , Conner J R , Silverberg M
Published: February 2020

Background

Patients with inflammatory bowel disease (IBD) are at higher risk for developing colitis-associated colorectal cancer (CAC). Clinical and endoscopic features are used to stratify the risk of CAC, but new biomarkers are necessary to improve this stratification. Recent studies have shown that loss of expression of special AT-rich sequence binding protein 2 (SATB2) is frequent in CAC compared to sporadic colorectal cancer and this SATB2 status is found in pre-cancerous dysplastic lesions as well. However, the relationship of known clinical risk factors for CAC and loss of SATB2 has not been explored.

Aims

To assess the association of loss of SATB2 expression in CAC with clinical characteristics of IBD.

Methods

Patients with a known diagnosis of ileocolonic or colonic Crohn’s disease (CD), ulcerative colitis (UC), or IBD unclassified (IBDU) who underwent colectomy between October 2010 and December 2017 for CAC were included. SATB2 expression in neoplastic tissue was evaluated using immunohistochemistry (IHC), where less than 5% of tumor cells showing staining was considered loss of SATB2. Tumor grade, P53 and mismatch repair (MMR) status were assessed as well. Available clinical data such as sex, smoking status, IBD diagnosis (CD, UC or IBDU), age at IBD diagnosis, duration of IBD, extent of colitis and previous medications were collected. We used a generalized linear model to assess the association between these biomarkers and clinical data.

Results

A total of 58 patients with mean age at CAC diagnosis of 50.3 ±13 years, 27 (46%) females were analyzed. Mean IBD duration was 17.6 ±10 years and 22 (37.9%), 34 (58.6%) and 2 (3.4%) were CD, UC and IBDU, respectively. Thirty-two (55.2%) CACs had loss of SATB2 expression. There was no association between age at CAC diagnosis or grade of the tumor and loss of SATB2. However, longer duration of IBD (21.2 ± 9 years vs 13.7 ± 9 years, p = 0.01) was significantly associated with loss of SATB2. There was no association between SATB2 status and other explored clinical or endoscopic variables. Tumors with P53 mutation were associated with a younger age at diagnosis of CAC (47.2 ±13 vs 55.0 ±12 years, p = 0.03), but no other associations of this marker or MMR with clinical or endoscopic variables of IBD were found.

Conclusions

Loss of SATB2 expression is significantly associated with IBD duration, a well-known risk factor for CAC. This association with duration of IBD could denote an effect of longer chronic inflammation on SATB2 status. Given the previously reported association of loss of expression of SATB2 with pre-cancerous lesions in IBD patients, this could be a potential biomarker for risk of CAC.

Funding Agencies

National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
A44 NOVEL AND HIGHLY SENSITIVE FLOW CYTOMETRIC BASED METHOD FOR CONTINUOUS TRACKING OF INTESTINAL PERMEABILTIY
Authors: Tsai K , Ma C , Crowley S M , Priatel J , Vallance B
Published: February 2020

Background

Inflammatory bowel diseases (IBD) are chronic, inflammatory conditions of the intestinal tract. In addition to a complex mixture of genetic, and environmental factors, increased intestinal permeability is also thought to be involved. Despite its diagnostic importance, there is no reliable yet minimally invasive way to measure intestinal permeability in patients and animal models. Currently, most assays involve the detection of orally given sugar molecules in either the urine (MLR method) or the plasma (FITC-dextran). These methods offer only limited accuracy, and do not allow continual tracking of intestinal permeability within the same animal due to the requirement of euthanization. Herein, we describe a novel cytometric-based method using an ingested dietary protein (ovalbumin (OVA)) and flow cytometry. We show our method is reliable, highly sensitive, minimally invasive and allows for the continuous tracking of intestinal permeability within the same individual using small blood volumes.

Aims

To compare the utility of our bead-based method of intestinal permeability measurement to the FITC-Dextran method.

Methods

Wildtype (WT) C57BL/6 mice, and mucin 2-deficient (Muc2-/-) mice (known to suffer a leaky gut) at baseline, or mice given chemical or infection-induced colitis were gavaged with a solution containing OVA protein and FITC-Dextran. After 6 hours, mice were euthanized to collect plasma for spectrophotometry (FITC-Dextran), or for flow cytometry (bead-based ELISA) analyses.

Results

We found that both OVA and FITC-dextran were detectable in plasma samples of the Muc2-/- mice, but not the WT mice, 6 hours after oral co-administration, however the bead-based method produced more consistent readings. Immunostaining of tissue sections with the same antibodies also showed that OVA can readily diffuse through the gut epithelium of Muc2-/- mice but not the WT mice, validating the specificity of the antibodies. We also tested our assay on WT mice undergoing dextran sodium sulfate (DSS) colitis, or infected with Citrobacter rodentium. We found that our method could detect statically significant changes in intestinal permeability 2 to 3 days earlier than the FITC-dextran method, while providing greatly reduced variability between technical repeats. More importantly, we were able to use a small volume (5 μL) of whole blood collected via tail poke, to measure intestinal permeability without requiring euthanization of the mice. This allowed the continuous tracking of permeability changes within the same animal.

Conclusions

We conclude that the bead-based method is more sensitive and reliable than the FITC-Dextran method as tested in several murine colitis models. More importantly, we showed that the bead-based method allows continual tracking of intestinal permeability within the same animal, enabling time course measurements.

Funding Agencies

CAG, CCC, CIHR, NRCBC Children’s Research Institute
A46 THE INFLAMMATORY CASPASES COORDINATE MUCOSAL RESTRICTION OF SALMONELLA THROUGH THE EPITHELIAL-INTRINSIC INFLAMMASOME AND IL-22 DRIVEN MUCIN SECRETION
Authors: Crowley S M , Allaire J M , Han X , Graef F A , Stahl M , Knodler L , Vallance B
Published: February 2020

Background

Intestinal epithelial cells (IECs) are located at the interface between the gut lumen and the underlying mucosal defense system. Here, they play a central role in the coordination of intestinal homeostasis, tempering pro-inflammatory responses but remaining rapidly responsive to noxious stimuli such as enteric pathogens. One early response mechanism by which IECs engage in immune defense is through the activation of the inflammasome which mobilizes the inflammatory caspases; caspase-1 and -11.

Aims

Here, we investigated the role of the inflammasome in overall mucosal defense against the enteric pathogen Salmonella enterica serovar Typhimurium.

Methods

Streptomycin-pretreated C57BL/6 (wildtype), Casp1/11 deficient (−/−), Casp1−/− and Casp11−/− mice were orally infected with Salmonella and burdens determined in intestinal tissues at 18h post infection (p.i.).

Results

Increased pathogen loads were observed for all caspase-deficient mice compared to wildtype, which correlated with increased IEC intracellular Salmonella burdens. Interestingly, despite increased bacterial loads, pathology scores for all inflammatory caspase mice were decreased, especially with regard to ‘IEC damage’ and ‘goblet cell loss’.

To determine if the increased burdens were due to the loss of IEC-intrinsic inflammasomes, enteroid monolayers were derived and infected with Salmonella. This revealed significantly increased intracellular burdens in caspase-deficient monolayers as compared to wildtype, in concert with a marked decrease in IEC shedding and cell death. Peak inflammatory caspase activity was displayed in shedding wildtype IECs, suggesting the IEC-intrinsic inflammasome restricts Salmonella infection through infected IEC expulsion.

The role of inflammasome signaling in acute mucosal defense was also examined. Wildtype tissue demonstrated a dramatic increase in mucus thickness (as evaluated by Muc2 immunostaining) and antimicrobial Reg3γ and β lectin transcript levels compared to caspase-deficient mice. Mucin release and Reg3 induction has been previously linked to IL-22, therefore we measured IL-22 expression and observed increased secretion in infected wildtype mice compared to Casp1/11−/−. This correlated with increased cecal infiltration of IL-22 producing ILC3 and NK T-cells. When infected mice were treated with IL-22 neutralizing antibody, this increased Salmonella burdens and decreased infection-induced mucus secretion, while no differences were observed in Casp1/11−/− treated with neutralizing antibody or isotype control.

Conclusions

Therefore the intestinal epithelium utilizes inflammasome signaling to coordinate multiple layers of innate defense at the gut mucosal surface to ultimately restrict enteric pathogen infections and their systemic spread.

Funding Agencies

CCC, CIHR, NRCUBC
A54 DELTA OPIOID RECEPTOR SIGNALING MEDIATES OPIOID INDUCED TOLERANCE AND HYPERALGESIA IN COLONIC SENSORY NERVES
Authors: Jaramillo Polanco J O , Lopez Lopez C D , Yu Y , Reed D E , Lomax A E , Vanner S
Published: February 2020

Background

While opioid drugs are widely used for treating abdominal pain, prolonged exposure to opioids can induce tolerance and paradoxically increase pain. We previously showed in colonic afferent nerves that chronic morphine treatment causes tolerance and paradoxical hyperalgesia, however the mechanisms are unknown.

Aims

To evaluate the role of delta opioid receptor (DOR) signaling in opioid induced tolerance and hyperalgesia in colonic nerves during chronic morphine exposure.

Methods

C57BL/6 mice were injected (i.p.) twice daily for 7 days with escalating doses of morphine (10, 20, 30 mg/kg for the first 3 days, then 40mg/kg on 4th to 7th day); a subset of mice were also injected with 2.5 mg/kg naltrindole (NTI), a DOR antagonist. The analgesic response in vivo was monitored daily using the tail-flick test. To assess the effect of chronic morphine exposure, mice were euthanized on day 8, and the dorsal root ganglia (DRGs) and colons were harvested. Isolated DRG neurons were incubated with 1μM morphine (30 min) and neuronal excitability was determined by measuring the rheobase (amount of current required to elicit an action potential) using perforated patch clamp. To assess changes in colonic afferent nerve excitability, ex vivo afferent nerve recordings were obtained from flat-sheet colon preparations to determine the effects of 1μM morphine (10 min perfusion) on mechanosensitivity by probing with 1g von Frey hair.

Results

The tail flick test showed that the DOR antagonist NTI inhibited (30%) the maximal antinociceptive effect of morphine and reduced morphine tolerance. In patch clamp recordings, 1μM morphine paradoxically increased the excitability of small DRG neurons from morphine mice (rheobase decreased 28%; p<0.05, 2-way ANOVA), whereas this excitatory effect was absent on neurons from NTI+morphine mice. Acute application of 1μM morphine had no effect on afferent nerve responses to probing in morphine mice (14.9 vs 16.7 Hz; p=ns, paired t test, n=15) whereas it inhibited the afferent nerve response to probing in NTI+morphine mice (15.2 vs 9.9 Hz; p<0.001, paired t test, n=16). Furthermore, most single units from morphine mice had an increased response to probing following acute morphine application (7/15 vs 2/16 units excited), whereas most units form NTI-morphine mice were inhibited (14/16 vs. 3/15 units inhibited, P<0.001, Chi-square).

Conclusions

These findings suggest that DOR signaling plays a significant role in the development of opioid tolerance and hyperalgesia in colonic afferent nerves induced by chronic morphine treatment. Thus, antagonists of DOR may help to mitigate these side effects induced by opioids.

Funding Agencies

CCC
A57 VALPROATE INDUCED DRESS SYNDROME CAUSING MIXED LIVER INJURY
Authors: Schneider R , Verstegen R , Hulst J , Ito S , Ng V
Published: February 2020

Background

Drug reaction with eosinophilia and systemic symptoms (DRESS) syndrome occurs with 1 in every 1,000 to 10,000 drug exposures and has a mortality rate of up to 10%. It is one mechanism by which medications can induce liver injury with elevated liver enzymes seen in the majority of cases. In children, aromatic anticonvulsants are the drugs most commonly associated with DRESS syndrome. Valproate, a non-aromatic anti-epileptic, is not known to have a heightened risk of hypersensitivity syndromes and is often the anti-epileptic of choice in patients who develop hypersensitivity syndromes from other anti-epileptics. Valproate hepatotoxicity is normally caused by its inhibition of fatty acid transport and mitochondrial β-oxidation; vanishing bile duct syndrome is also reported.

Aims

We present a case, to the best of our knowledge the first in paediatrics, in which valproate causes DRESS syndrome and a secondary, predominantly cholestatic, liver injury.

Methods

Literature review and case report.

Results

A previously healthy 14-year-old girl was diagnosed with new-onset seizures and started on valproate. Three weeks later, she developed a pruritic exanthem. Despite discontinuing her valproate, the rash persisted and she developed fever and jaundice. She was admitted to the ICU at the Hospital for Sick Children with a diagnosis of DRESS syndrome (RegiSCAR DRESS score 7) for treatment with IV steroids. At admission, she had a skin eruption, fever, leukocytosis (22.86 x109/L), eosinophilia (1.03x109/L), atypical lymphocytes (1.62 x109/L), lymphadenopathy, and internal organ involvement (BiliC 138, GGT 501, INR 1.3, ALT 543, AST 370, Crt 109). Of note, her EBV PCR was positive. By discharge, her rash improved and kidney function normalized. Her cholestasis persisted, despite improvement in her transaminases and eosinophilia (BiliC 163, INR 1.0, GGT 338, ALT 506, AST 220, Eos 0.98x109/L). She was discharged home on an oral steroid wean, ursodeoxycholic acid, levocarnitine, and levetiracetam. Upon reaching a daily dose of 20mg of prednisone, her rash and pruritus worsened, she had lost 7kg, and she was readmitted for IV steroids. Her rash improved within 4 days and she was discharged on a slower steroid taper, with the addition of cholestyramine and insulin for steroid induced diabetes. At present (2 ½ months after diagnosis), she continues her oral steroids and has persistent liver injury and pruritus.

Conclusions

We report the first paediatric case of valproate induced DRESS syndrome causing a significant cholestatic presentation with otherwise preserved liver synthetic function. The prolonged cholestasis and pruritus may be a result of the natural course of DRESS syndrome, EBV reactivation/infection, or polypharmacy. Early recognition of DRESS syndrome as a mechanism of valproate induced liver injury in children is essential for earlier diagnosis and initiation of targeted therapy.

Funding Agencies

None
A70 ASSOCIATION BETWEEN PROTON PUMP INHIBITOR USE IN HOSPITALIZED CIRRHOTICS AND DEVELOPMENT OF HEPATIC ENCEPHALOPATHY AND SPONTANEOUS BACTERIAL PERITONITIS
Authors: Tan P , Ukos M , Miller C , Lombardi J , Kelly E M
Published: February 2020

Background

Proton pump inhibitors (PPI) are commonly prescribed medications which are indicated in various different gastrointestinal (GI) diseases, including peptic ulcer disease, gastroesophageal reflux disorder and upper GI bleeding. There is some evidence to suggest that PPI use in cirrhosis may predispose to the development of hepatic encephalopathy (HE) and spontaneous bacterial peritonitis (SBP), albeit with some controversy.

Aims

We aim to conduct a retrospective epidemiological analysis of the association between PPI use in hospitalized patients with cirrhosis, and prevalence of HE and SBP.

Methods

This was a retrospective cohort study of 600 adult patients (mean age 61.4 (SD=12.2)) admitted the Ottawa Hospital between January 1, 2011 and December 31, 2015 with cirrhosis. A chart review was conducted and relevant information extracted.

Results

Average MELD-Na on admission was 16.2 (sd=6.7). 14.8% of patients had a history of HE, 5.0% SBP, 7.8% with history of hepatocellular carcinoma and 1.0% with history of hepatorenal syndrome. 28.5% of patients had a history of varices, of which 11.0% had previous variceal bleeding. 69.3% of patients were on a PPI during their hospitalization. Not surprisingly, patients admitted with variceal bleeding were more likely to be exposed to PPI in hospital (97.2% vs 63.2%, p<0.01) Patients with a diagnosis of cirrhosis prior to index admission were more likely to be on a PPI in hospital (p=0.001) and on discharge (p=0.001). Patients with ascites were less likely to be on a PPI than those without ascites (64.1% vs 77.6%, p<0.01). There was no significant correlation between in hospital PPI use and MELD score (p=0.42). Amongst patients on PPI in hospital, 85.9% remained on a PPI at discharge. Although numerically greater, no statistically significant differences were observed in terms of prevalence of HE (21.3% in patients on PPI vs 8.3% in those not on PPI (p=0.37)), nor SBP (5.7% on PPI vs. 3.7% in those not on PPI (p=0.29)).

Conclusions

We did not observe a significant difference in HE and SBP among this cohort of cirrhotic patients by in-hospital PPI use. We did however note significantly higher PPI use in patients with previous diagnosis of cirrhosis as compared to those who were newly diagnosed, as well as those whose admissions were related to bleeding. Patients with ascites had lower prevalence of PPI use. Prescribing patterns for PPIs in patients with cirrhosis warrant further attention, including clinical utility and longer-term risks and benefits of this therapy.

Funding Agencies

None
A75 ON THE TRAIL OF ALTERNATIVE LIGANDS AND SIGNALING PATHWAYS FOR LGR5
Authors: Dagenais Bellefeuille S , Arguin G , Gendron F
Published: February 2020

Background

Leucine-rich G protein coupled receptor-5 (LGR5) is a GPCR originally identified as a marker for intestinal stem cells but is now associated to stemness in numerous other tissues. R-spondins (RSPO) are the only reported ligands of LGR5, which upon receptor binding, potentiates the canonical Wnt/b-catenin pathway. Surprisingly, despite the presence of classical GPCR features such as conserved DRY and NPXXY motifs RSPO binding to LGR5 does not induce classical GPCR behaviors such as coupling to heterotrimeric G-proteins or engagement of b-arrestins. For this reason, LGR5 is still considered to be an orphan receptor.

Aims

Aberrant expression of LGR5 is a common feature of many cancers, including gastrointestinal cancers, but it is still unclear why LGR5 is pro-tumorigenic in some cancers, while anti-tumorigenic in others. One potential explanation is that LGR5 does not only signals through the Wnt/b-catenin pathway, but also to alternative pathways. Thus, the goals of this study were 1), to elucidate LGR5 signaling networks and 2), to identify new LGR5 ligands and activity modulators. We suggested that the intestinal lumen content could be a source of bioactive molecules that could act as alternative LGR5 ligands

Methods

All experiments were performed on HEK293 cells expressing recombinant LGR5 (HEK293/LGR5) and compared to control cells expressing empty vector. Characterization of LGR5 signaling network was initiated with LC MS/MS spectrometric analyses using cells stimulated with Wnt3a and RSPO1. To identify new LGR5 modulators, commercially available bacteria-derived metabolites (BDM) were tested on HEK293/LGR5 cells stimulated with Wnt3a and RSPO1 and assessed using the TOP-Flash luciferase system. Finally, a mouse intestinal lumen extract (ILE) was investigated in whole-cell impedance assay to detect if it contained native BDM or potential host-derived LGR5 ligands.

Results

Mass spectrometric analyses revealed distinct protein expression and phosphorylation profiles in HEK293/LGR5 cells stimulated with Wnt3a and RSPO1. The TOP/Flash luciferase assays showed that the flavonoid derivative 3,4-dihydroxyphenylacetic acid (DOPAC) and lipopolysaccharide (LPS) acted as negative regulators of LGR5, while butyrate had no effect. Interestingly, stimulation of HEK293/LGR5 with ILE induced a Gαq-like response in the impedance assay, while control cells did not respond.

Conclusions

Stimulation of LGR5 with Wnt and RSPO triggered a complex signaling network. In agreement with our hypothesis, some BDM such as DOPAC and LPS were indeed activity modulators of LGR5. Moreover, a still unidentified compound present in the ILE triggered a selective GPCR-like response in HEK293/LGR5 cells. Future works aim at identifying this putative LGR5 ligand(s) and establish the LGR5 interactome using proximity labeling and mass spectrometry.

Funding Agencies

NSERC, Université de Sherbrooke, FRQS
A80 ADVANCED ENDOSCOPIC RESECTION OF LARGE POLYPS & EARLY NEOPLASIA: OUTCOMES OF ENDOSCOPIC MUCOSAL RESECTION IN BRITISH COLUMBIA
Authors: Zhao B , Chahal D , Lam E , Donnellan F
Published: February 2020

Background

Recent advances have resulted in a new technique termed endoscopic mucosal resection (EMR). This procedure has been successful at removing large or complex polyps and achieving remission rates comparable to surgery. EMR can also be used to remove early, non-metastatic cancer and they are less invasive than surgery. However, they have been associated with their own complications, most serious of which being perforation. This procedure has recently become available in British Columbia for resection of both complex polyps and early established cancers in the colon.

Aims

Here we present patient outcomes of EMR procedures for the resection of colorectal polyps in British Columbia.

Methods

Retrospective data were collected on all EMR procedures done in Vancouver General Hospital and St. Paul’s Hospital (Vancouver, B.C.) from October 2012 (when procedure became available) to July 2019. Inclusion criteria were all adults who had undergone EMR for resection of polyps in the colon. Exclusion criteria were patients younger than 18 or patients who had EMR that resected polyps in the upper GI tract. Patients were referred to one of two endoscopists when one or more polyps suitable for EMR were identified during colonoscopy by other gastroenterologists. Collected data included patient demographics, polyp characteristics, procedure outcome, and complications.

Results

There were 211 EMR procedures performed on 182 patients (48.9% male). Patient age ranged from 27 to 86 (mean = 67.1). A total of 244 colon polyps were removed with an average size of 2.91 cm and ranged from 0.8 cm to 15 cm. Resected polyps had the following distribution: ascending colon (63.5%), transverse colon (10.2%), descending colon (5.7%), sigmoid colon (15.2%), and rectum (5.3%). Of those that reported resection type, 84.2% were piecemeal and 15.8% were en bloc. 40.9% of polyps were tubulovillous adenoma, 33.2% were tubular, 16.2% were sessile serrated, 6.4% were villous, and 3.4% were adenocarcinoma. Patients from 11 of the 211 EMR cases (5.2%) experienced post-procedure bleed and 4 of these 11 patients (36.4%) had been on anti-platelet or anti-coagulants (discontinued before procedure). Overall, patients from 51 (24.2%) EMR cases were on anti-platelet or anti-coagulants. 33 cases (15.6%) had residual polyps at the resection site that required additional endoscopic resection during follow-up and 14 patients (6.6%) required surgery. None of the EMR procedures resulted in perforation.

Conclusions

EMR is an effective minimally-invasive procedures that can be used to remove large, complicated colonic polyps and achieve long-term remission rate. The procedure has an acceptable risk profile, with complication and re-intervention rate similar or less than other procedures used to remove large, complicated polyps.

Funding Agencies

None