scholarly journals Development of a Multiplex Immunohistochemistry Workflow to Investigate the Immune Microenvironment in Mouse Models of Inflammatory Bowel Disease and Colon Cancer

2021 ◽  
Vol 22 (20) ◽  
pp. 11001
Author(s):  
Lokman Pang ◽  
Matthias Ernst ◽  
Jennifer Huynh
Keyword(s):  
Colon Cancer ◽  
Mouse Model ◽  
Mouse Models ◽  
Colon Adenocarcinoma ◽  
Tumour Microenvironment ◽  
Human Patient ◽  
Mouse Tissues ◽  
Formalin Fixed Paraffin Embedded ◽  
Ffpe Tissues ◽  
Inflammatory Bowel

Multiplex immunohistochemistry (mIHC) enables simultaneous staining of multiple immune markers on a single tissue section. Mounting studies have demonstrated the versatility of mIHC in evaluating immune infiltrates in different diseases and the tumour microenvironment (TME). However, the majority of published studies are limited to the analysis of human patient samples. Performing mIHC on formalin-fixed paraffin-embedded (FFPE) mouse tissues, particularly with sensitive antigens, remain challenging. The aim of our study was to develop a robust and reproducible protocol to uncover the immune landscape in mouse FFPE tissues. Effective antibody stripping while maintaining sensitivity to antigens and tissue adhesion to the glass slide is critical in developing an mIHC panel to allow successive rounds of staining. Thus, we identified a highly efficient stripping method that preserves signal intensity and antigenicity to allow multiple rounds of staining. We subsequently optimised an mIHC workflow with antibodies specific against CD4, CD8α, FOXP3 and B220 to identify distinct T and B cell populations on mouse FFPE tissues. Lastly, the application of this mIHC panel was validated in a mouse model of inflammatory bowel cancer, two allograft mouse models of spontaneous colon adenocarcinoma and a sporadic mouse model of colon cancer. Together, these demonstrate the utility of the aforementioned protocol in establishing the quantity and spatial localisation of immune cells in different pathological tissues.

scholarly journals Activation of the GPR35 pathway drives angiogenesis in the tumour microenvironment

Gut ◽  
2021 ◽  
pp. gutjnl-2020-323363
Author(s):  
Ester Pagano ◽  
Joshua E Elias ◽  
Georg Schneditz ◽  
Svetlana Saveljeva ◽  
Lorraine M Holland ◽  
...  
Keyword(s):  
Colon Cancer ◽  
Tumour Growth ◽  
Tumour Microenvironment ◽  
Tube Formation ◽  
Tissue Remodelling ◽  
Colon Cancers ◽  
Inflammatory Bowel ◽  
Ion Pumping ◽  
G Protein Coupled

ObjectivePrimary sclerosing cholangitis (PSC) is in 70% of cases associated with inflammatory bowel disease. The hypermorphic T108M variant of the orphan G protein-coupled receptor GPR35 increases risk for PSC and ulcerative colitis (UC), conditions strongly predisposing for inflammation-associated liver and colon cancer. Lack of GPR35 reduces tumour numbers in mouse models of spontaneous and colitis associated cancer. The tumour microenvironment substantially determines tumour growth, and tumour-associated macrophages are crucial for neovascularisation. We aim to understand the role of the GPR35 pathway in the tumour microenvironment of spontaneous and colitis-associated colon cancers.DesignMice lacking GPR35 on their macrophages underwent models of spontaneous colon cancer or colitis-associated cancer. The role of tumour-associated macrophages was then assessed in biochemical and functional assays.ResultsHere, we show that GPR35 on macrophages is a potent amplifier of tumour growth by stimulating neoangiogenesis and tumour tissue remodelling. Deletion of Gpr35 in macrophages profoundly reduces tumour growth in inflammation-associated and spontaneous tumour models caused by mutant tumour suppressor adenomatous polyposis coli. Neoangiogenesis and matrix metalloproteinase activity is promoted by GPR35 via Na/K-ATPase-dependent ion pumping and Src activation, and is selectively inhibited by a GPR35-specific pepducin. Supernatants from human inducible-pluripotent-stem-cell derived macrophages carrying the UC and PSC risk variant stimulate tube formation by enhancing the release of angiogenic factors.ConclusionsActivation of the GPR35 pathway promotes tumour growth via two separate routes, by directly augmenting proliferation in epithelial cells that express the receptor, and by coordinating macrophages’ ability to create a tumour-permissive environment.

scholarly journals Impact of dietary amino acids and polyamines on intestinal carcinogenesis and chemoprevention in mouse models

2007 ◽  
Vol 35 (2) ◽  
pp. 322-325 ◽  
Author(s):  
E.W. Gerner
Keyword(s):  
Colon Cancer ◽  
Mouse Model ◽  
Mouse Models ◽  
Specific Inhibitor ◽  
Colon Carcinogenesis ◽  
Tumour Suppressor Gene ◽  
Polyamine Metabolism ◽  
Adenomatous Polyposis ◽  
Polyamine Synthesis ◽  
Intestinal Carcinogenesis

Colon cancer in humans is influenced by both genetic and dietary risk factors. The majority of colon cancers have somatic mutations in the APC (adenomatous polyposis coli) tumour-suppressor gene. Dietary arginine enhances the risk of APC-dependent colon carcinogenesis in mouse models by a mechanism involving NOS2 (nitric oxide synthase 2), as elimination of NOS2 alleles suppresses this phenotype. DFMO (difluoromethylornithine), a specific inhibitor of polyamine synthesis, also inhibits dietary arginine-induced colon carcinogenesis in C57BL/6J-ApcMin/J mice. The primary consequence of dietary arginine is to increase the adenoma grade in these mice. Either loss of NOS2 alleles or inhibition of polyamine synthesis suppresses the arginine-induced increase in adenoma grade. In addition to promoting intestinal carcinogenesis, polyamines can also reduce the efficacy of certain intestinal cancer chemopreventive agents. The NSAID (non-steroidal anti-inflammatory drug) sulindac is a potent inhibitor of intestinal carcinogenesis in the C57BL/6J-ApcMin/J mouse model and is used to treat humans with FAP (familial adenomatous polyposis). Dietary putrescine reduces the ability of sulindac to suppress intestinal tumorigenesis in the mouse model. These data suggest that reducing polyamine metabolism and dietary polyamine levels may enhance strategies for colon cancer chemoprevention.

scholarly journals Computational Interspecies Translation Between Alzheimer’s Disease Mouse Models and Human Subjects Identifies Innate Immune Complement, TYROBP, and TAM Receptor Agonist Signatures, Distinct From Influences of Aging

2021 ◽  
Vol 15 ◽  
Author(s):  
Meelim J. Lee ◽  
Chuangqi Wang ◽  
Molly J. Carroll ◽  
Douglas K. Brubaker ◽  
Bradley T. Hyman ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Mouse Model ◽  
Mouse Models ◽  
Late Onset ◽  
Innate Immune ◽  
Cytokine Signaling ◽  
Preclinical Research ◽  
Human Patient ◽  
Receptor Family

Mouse models are vital for preclinical research on Alzheimer’s disease (AD) pathobiology. Many traditional models are driven by autosomal dominant mutations identified from early onset AD genetics whereas late onset and sporadic forms of the disease are predominant among human patients. Alongside ongoing experimental efforts to improve fidelity of mouse model representation of late onset AD, a computational framework termed Translatable Components Regression (TransComp-R) offers a complementary approach to leverage human and mouse datasets concurrently to enhance translation capabilities. We employ TransComp-R to integratively analyze transcriptomic data from human postmortem and traditional amyloid mouse model hippocampi to identify pathway-level signatures present in human patient samples yet predictive of mouse model disease status. This method allows concomitant evaluation of datasets across different species beyond observational seeking of direct commonalities between the species. Additional linear modeling focuses on decoupling disease signatures from effects of aging. Our results elucidated mouse-to-human translatable signatures associated with disease: excitatory synapses, inflammatory cytokine signaling, and complement cascade- and TYROBP-based innate immune activity; these signatures all find validation in previous literature. Additionally, we identified agonists of the Tyro3 / Axl / MerTK (TAM) receptor family as significant contributors to the cross-species innate immune signature; the mechanistic roles of the TAM receptor family in AD merit further dedicated study. We have demonstrated that TransComp-R can enhance translational understanding of relationships between AD mouse model data and human data, thus aiding generation of biological hypotheses concerning AD progression and holding promise for improved preclinical evaluation of therapies.

scholarly journals A novel mouse model of sporadic colon cancer induced by combination of conditional Apc genes and chemical carcinogen in the absence of Cre recombinase

2019 ◽  
Vol 40 (11) ◽  
pp. 1376-1386 ◽  
Author(s):  
Jeffrey S Souris ◽  
Hannah J Zhang ◽  
Urszula Dougherty ◽  
Nai-Tzu Chen ◽  
Joseph V Waller ◽  
...  
Keyword(s):  
Colon Cancer ◽  
Small Intestine ◽  
Mouse Model ◽  
Mouse Models ◽  
Cancer Biology ◽  
Human Colon ◽  
Cre Recombinase ◽  
Tumor Incidence ◽  
Human Colon Cancer ◽  
Colon Tumors

AbstractAlthough valuable insights into colon cancer biology have been garnered from human colon cancer cell lines and primary colonic tissues, and animal studies using human colon cancer xenografts, immunocompetent mouse models of spontaneous or chemically induced colon cancer better phenocopy human disease. As most sporadic human colon tumors present adenomatous polyposis coli (APC) gene mutations, considerable effort has gone into developing mice that express mutant Apc alleles that mimic human colon cancer pathogenesis. A serious limitation of many of these Apc-mutant murine models, however, is that these mice develop numerous tumors in the small intestine but few, if any, in the colon. In this work, we examined three spontaneous mouse models of colon tumorigenesis based upon the widely used multiple intestinal neoplasia (Min) mouse: mice with either constitutive or conditional Apc mutations alone or in combination with caudal-related homeobox transcription factor CDX2P-Cre transgene — either with or without exposure to the potent colon carcinogen azoxymethane. Using the CDX2 promoter to drive Cre recombinase transgene expression effectively inactivated Apc in colonocytes, creating a model with earlier tumor onset and increased tumor incidence/burden, but without the Min mouse model’s small intestine tumorigenesis and susceptibility to intestinal perforation/ulceration/hemorrhage. Most significantly, azoxymethane-treated mice with conditional Apc expression, but absent the Cre recombinase gene, demonstrated nearly 50% tumor incidence with two or more large colon tumors per mouse of human-like histology, but no small intestine tumors — unlike the azoxymethane-resistant C57BL/6J-background Min mouse model. As such this model provides a robust platform for chemoprevention studies.

scholarly journals Novel gut pathobionts confound results in a widely used mouse model of human inflammatory disease

2021 ◽  
Author(s):  
Samuel C. Forster ◽  
Simon Clare ◽  
Benjamin S. Beresford-Jones ◽  
Katherine Harcourt ◽  
George Notley ◽  
...  
Keyword(s):  
Mouse Model ◽  
Mouse Models ◽  
Large Scale ◽  
Bacterial Species ◽  
The Novel ◽  
Host Immune Responses ◽  
Causes Of Disease ◽  
Health And Disease ◽  
Inflammatory Bowel ◽  
Scale Experiment

The mammalian gut microbiota consists of hundreds of anaerobic bacterial species that shape intestinal homeostasis and influence host immune responses. Although the causal roles of specific human gut bacterial species in health and disease are emerging, the role of indigenous gut bacteria in driving immunophenotypic variability in mouse models of human disease remains poorly understood. We performed a large-scale experiment using 579 laboratory mice designed to identify and validate the causes of disease variability in the widely used dextran sulphate sodium (DSS) mouse model of inflammatory bowel disease. Using microbiome analysis, coupled with machine learning and targeted anaerobic culturing, we identified and isolated the novel gut pathobiont species Duncaniella muricolitica and Alistipes okayasuensis and fulfilled Koch’s postulates in mice to show that each pathobiont exerts dominant effects in the DSS model leading to variable treatment responses. We show these pathobiont species are common, but not ubiquitous, in mouse facilities around the world, raising experimental design opportunities for improved mouse models of human intestinal diseases.

Role of MicroRNAs in Colon Cancer Progression and Metastasis

2020 ◽  
Vol 20 ◽  
Author(s):  
Shruthi Sanjitha Sampath ◽  
Sivaramakrishnan Venkatabalsubramanian ◽  
Satish Ramalingam
Keyword(s):  
Colon Cancer ◽  
Cancer Progression ◽  
Target Genes ◽  
Tumour Progression ◽  
Intestinal Barrier ◽  
Colon Cancer Progression ◽  
Bowel Diseases ◽  
Inflammatory Bowel ◽  
Novel Therapeutic Strategies

: MicroRNAs regulate gene expression at the posttranscriptional level by binding to the mRNA of their target genes. The dysfunction of miRNAs is strongly associated with the inflammation of the colon. Besides, some microRNAs are shown to suppress tumours while others promote tumour progression and metastasis. Inflammatory bowel diseases include Crohn’s disease and Ulcerative colitis which increase the risk factor for inflammation-associated colon cancer. MicroRNAs are shown to be involved in gastrointestinal pathologies, by targeting the transcripts encoding proteins of the intestinal barrier and their regulators that are associated with inflammation and colon cancer. Detection of these microRNAs in the blood, serum, tissues, faecal matter, etc will enable us to use these microRNAs as biomarkers for early detection of the associated malignancies and design novel therapeutic strategies to overcome the same. Information on MicroRNAs can be applied for the development of targeted therapies against inflammation-mediated colon cancer.

scholarly journals Intranasal oxytocin administration ameliorates social behavioral deficits in a POGZWT/Q1038R mouse model of autism spectrum disorder

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Kohei Kitagawa ◽  
Kensuke Matsumura ◽  
Masayuki Baba ◽  
Momoka Kondo ◽  
Tomoya Takemoto ◽  
...  
Keyword(s):  
Autism Spectrum Disorder ◽  
Social Behavior ◽  
Mouse Model ◽  
Mouse Models ◽  
De Novo ◽  
Neurodevelopmental Disorder ◽  
Autism Spectrum ◽  
Spectrum Disorder ◽  
De Novo Mutation ◽  
Behavioral Deficits

AbstractAutism spectrum disorder (ASD) is a highly prevalent neurodevelopmental disorder characterized by core symptoms of impaired social behavior and communication. Recent studies have suggested that the oxytocin system, which regulates social behavior in mammals, is potentially involved in ASD. Mouse models of ASD provide a useful system for understanding the associations between an impaired oxytocin system and social behavior deficits. However, limited studies have shown the involvement of the oxytocin system in the behavioral phenotypes in mouse models of ASD. We have previously demonstrated that a mouse model that carries the ASD patient-derived de novo mutation in the pogo transposable element derived with zinc finger domain (POGZWT/Q1038R mice), showed ASD-like social behavioral deficits. Here, we have explored whether oxytocin (OXT) administration improves impaired social behavior in POGZWT/Q1038R mice and found that intranasal oxytocin administration effectively restored the impaired social behavior in POGZWT/Q1038R mice. We also found that the expression level of the oxytocin receptor gene (OXTR) was low in POGZWT/Q1038R mice. However, we did not detect significant changes in the number of OXT-expressing neurons between the paraventricular nucleus of POGZWT/Q1038R mice and that of WT mice. A chromatin immunoprecipitation assay revealed that POGZ binds to the promoter region of OXTR and is involved in the transcriptional regulation of OXTR. In summary, our study demonstrate that the pathogenic mutation in the POGZ, a high-confidence ASD gene, impairs the oxytocin system and social behavior in mice, providing insights into the development of oxytocin-based therapeutics for ASD.

scholarly journals The Function of Selenium in Central Nervous System: Lessons from MsrB1 Knockout Mouse Models

Molecules ◽  
2021 ◽  
Vol 26 (5) ◽  
pp. 1372
Author(s):  
Tengrui Shi ◽  
Jianxi Song ◽  
Guanying You ◽  
Yujie Yang ◽  
Qiong Liu ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Synaptic Plasticity ◽  
Mouse Model ◽  
Mouse Models ◽  
Knockout Mouse ◽  
Methionine Sulfoxide ◽  
Methionine Sulfoxide Reductase ◽  
Knockout Mouse Model ◽  
The Central Nervous System

MsrB1 used to be named selenoprotein R, for it was first identified as a selenocysteine containing protein by searching for the selenocysteine insert sequence (SECIS) in the human genome. Later, it was found that MsrB1 is homologous to PilB in Neisseria gonorrhoeae, which is a methionine sulfoxide reductase (Msr), specifically reducing L-methionine sulfoxide (L-Met-O) in proteins. In humans and mice, four members constitute the Msr family, which are MsrA, MsrB1, MsrB2, and MsrB3. MsrA can reduce free or protein-containing L-Met-O (S), whereas MsrBs can only function on the L-Met-O (R) epimer in proteins. Though there are isomerases existent that could transfer L-Met-O (S) to L-Met-O (R) and vice-versa, the loss of Msr individually results in different phenotypes in mice models. These observations indicate that the function of one Msr cannot be totally complemented by another. Among the mammalian Msrs, MsrB1 is the only selenocysteine-containing protein, and we recently found that loss of MsrB1 perturbs the synaptic plasticity in mice, along with the astrogliosis in their brains. In this review, we summarized the effects resulting from Msr deficiency and the bioactivity of selenium in the central nervous system, especially those that we learned from the MsrB1 knockout mouse model. We hope it will be helpful in better understanding how the trace element selenium participates in the reduction of L-Met-O and becomes involved in neurobiology.

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