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Cancers Apr 2024Colorectal tumorigenesis involves the development of aberrant crypt foci (ACF) or preneoplastic lesions, representing the earliest morphological lesion visible in colon...
Colorectal tumorigenesis involves the development of aberrant crypt foci (ACF) or preneoplastic lesions, representing the earliest morphological lesion visible in colon cancer. The purpose of this study was to determine changes in protein expression in carcinogen-induced ACF as they mature and transform into adenomas. Protein expression profiles of azoxymethane (AOM)-induced F344 rat colon ACF and adenomas were compared at four time points, 4 (control), 8, 16, and 24 weeks post AOM administration ( = 9/group), with time points correlating with induction and transformation events. At each time point, micro-dissected ACF and/or adenoma tissues were analyzed across multiple quantitative two-dimensional (2D-DIGE) gels using a Cy-dye labeling technique and a pooled internal standard to quantify expression changes with statistical confidence. Western blot and subsequent network pathway mapping were used to confirm and elucidate differentially expressed ( ≤ 0.05) proteins, including changes in vinculin (; = 0.007), scinderin (; = 0.02), and profilin (; = 0.01), By determining protein expression changes in ACF as they mature and transform into adenomas, a "baseline" of altered regulatory proteins associated with adenocarcinoma development in this model has been elucidated. These data will enable future studies aimed at biomarker identification and understanding the molecular biology of intestinal tumorigenesis and adenocarcinoma maturation under varying intestinal conditions.
PubMed: 38730628
DOI: 10.3390/cancers16091678 -
Frontiers in Immunology 2023The intestinal barrier plays a crucial role in distinguishing foods from toxins. Prostaglandin D (PGD) is one of the lipid-derived autacoids synthesized from cell...
INTRODUCTION
The intestinal barrier plays a crucial role in distinguishing foods from toxins. Prostaglandin D (PGD) is one of the lipid-derived autacoids synthesized from cell membrane-derived arachidonic acid. We previously reported that pharmacological stimulation of PGD receptor, D prostanoid 1 (DP1) attenuated the symptoms of azoxymethane/dextran sodium sulfate-induced colitis and ovalbumin-induced food allergy in mouse models. These observations suggested that DP1 stimulation protects the intestinal barrier. The present study aimed to uncover the effects of DP1 stimulation on intestinal barrier function and elucidate the underlying mechanisms.
MATERIALS AND METHODS
Intestinal permeability was assessed in mice by measuring the transfer of orally administered fluorescein isothiocyanate-dextran (40 kDa) into the blood. The DP1 agonist BW245C (1 mg/kg) was administered 10 min prior to dextran administration. The intestinal permeability was confirmed using the everted sac method. Tight junction integrity was evaluated by measuring the transepithelial electrical resistance (TER) in the human intestinal epithelial cell line Caco-2. Mucus secretion was assessed by observing Alcian Blue-stained intestinal sections.
RESULTS
Pharmacological DP1 stimulation reduced intestinal permeability both and . Immunohistochemical staining showed that DP1 was strongly expressed on the apical side of the epithelial cells. DP1 stimulation did not affect TER but induced mucus secretion from goblet cells. Mucus removal by a mucolytic agent N-acetyl-l-cysteine canceled the inhibition of intestinal permeability by DP1 stimulation.
CONCLUSION
These observations suggest that pharmacological DP1 stimulation decreases intestinal permeability by stimulating mucus secretion.
Topics: Humans; Animals; Mice; Prostaglandins; Dextrans; Prostaglandin D2; Caco-2 Cells; Mucus; Permeability
PubMed: 37942331
DOI: 10.3389/fimmu.2023.1276852 -
Arquivos de Gastroenterologia 2024Colorectal cancer is one of the most prevalent pathologies worldwide whose prognosis is linked to early detection. Colonoscopy is the gold standard for screening, and...
BACKGROUND
Colorectal cancer is one of the most prevalent pathologies worldwide whose prognosis is linked to early detection. Colonoscopy is the gold standard for screening, and diagnosis is usually made histologically from biopsies. Aiming to reduce the inspection and diagnostic time as well as the biopsies and resources involved, other techniques are being promoted to conduct accurate in vivo colonoscopy assessments. Optical biopsy aims to detect normal and neoplastic tissues analysing the autofluorescence spectrum based on the changes in the distribution and concentration of autofluorescent molecules caused by colorectal cancer. Therefore, the autofluorescence contribution analysed by image processing techniques could be an approach to a faster characterization of the target tissue.
OBJECTIVE
Quantify intensity parameters through digital processing of two data sets of three-dimensional widefield autofluorescence microscopy images, acquired by fresh colon tissue samples from a colorectal cancer murine model. Additionally, analyse the autofluorescence data to provide a characterization over a volume of approximately 50 µm of the colon mucosa for each image, at second (2nd), fourth (4th) and eighth (8th) weeks after colorectal cancer induction.
METHODS
Development of a colorectal cancer murine model using azoxymethane/dextran sodium sulphate induction, and data sets acquisition of Z-stack images by widefield autofluorescence microscopy, from control and colorectal cancer induced animals. Pre-processing steps of intensity value adjustments followed by quantification and characterization procedures using image processing workflow automation by Fiji's macros, and statistical data analysis.
RESULTS
The effectiveness of the colorectal cancer induction model was corroborated by a histological assessment to correlate and validate the link between histological and autofluorescence changes. The image digital processing methodology proposed was then performed on the three-dimensional images from control mice and from the 2nd, 4th, and 8th weeks after colorectal cancer chemical induction, for each data set. Statistical analyses found significant differences in the mean, standard deviation, and minimum parameters between control samples and those of the 2nd week after induction with respect to the 4th week of the first experimental study. This suggests that the characteristics of colorectal cancer can be detected after the 2nd week post-induction.
CONCLUSION
The use of autofluorescence still exhibits levels of variability that prevent greater systematization of the data obtained during the progression of colorectal cancer. However, these preliminary outcomes could be considered an approach to the three-dimensional characterization of the autofluorescence of colorectal tissue, describing the autofluorescence features of samples coming from dysplasia to colorectal cancer.
BACKGROUND
• A new digital image processing method was developed to measure intensity in 3D autofluorescence images of colorectal samples using a CRC mouse model.
BACKGROUND
• This method showed that autofluorescence intensity in colon mucosa is similar in healthy tissue but changes significantly in tumor development.
BACKGROUND
• Statistical analysis revealed CRC traits detectable from the second week post-induction, aiding in early CRC detection.
BACKGROUND
• The study provides a basis for 3D autofluorescence characterization in colorectal tissue from dysplasia to cancer, although variability in autofluorescence limits data systematization during cancer progression.
Topics: Animals; Mice; Disease Models, Animal; Microscopy; Azoxymethane; Biopsy; Colorectal Neoplasms
PubMed: 38451659
DOI: 10.1590/S0004-2803.246102023-62 -
Microorganisms May 2024The only reliable factor that reduces the risk of colorectal carcinogenesis is physical activity. However, the underlying mechanisms remain unclear. In this study, we...
The only reliable factor that reduces the risk of colorectal carcinogenesis is physical activity. However, the underlying mechanisms remain unclear. In this study, we examined the effects of physical activity against gut microbiota, including mucosa-associated microbiota (MAM) on azoxymethane-induced colorectal tumors in obese mice. We divided the subjects into four groups: normal diet (ND), high-fat diet (HFD), ND + exercise (Ex), and HFD + Ex groups. The Ex group performed treadmill exercise for 20 weeks. Thereafter, fecal and colonic mucus samples were extracted for microbiota analysis. DNA was collected from feces and colonic mucosa, and V3-V4 amplicon sequencing analysis of the 16SrRNA gene was performed using MiSeq. The HFD group had significantly more colonic polyps than the ND group (ND 6.5 ± 1.3, HFD 11.4 ± 1.5, < 0.001), and the addition of Ex suppressed the number of colonic polyps in ND and HFD groups (ND 6.5 ± 1.3, ND + Ex 2.8 ± 2.5, < 0.05). The HFD group showed significantly lower concentrations of succinic, acetic, butyric, and propionic acids (mg/g) in feces, compared with the ND group (succinic acid HFD 0.59, ND 0.17; acetic acid HFD 0.63, ND 2.41; propionic acid HFD 0.10, ND 0.47; and N-butyric acid HFD 0.31, ND 0.93). In the case of ND, succinic acid and butyric acid tended to decrease with Ex (succinic acid ND 0.17, ND + Ex 0.12; N-butyric acid ND 0.93, ND + Ex 0.74 0.74). Succinic acid, acetic acid, butyric acid, and propionic acid levels in feces were significantly lower in the HFD group than in the ND group; in both feces and mucus samples, and levels were significantly lower in the HFD group. was significantly increased in ND + Ex and HFD + Ex groups. Diet and exercise affected the number of colorectal tumors. Furthermore, diet and exercise alter intestinal MAM, which may be involved in colorectal tumor development.
PubMed: 38792787
DOI: 10.3390/microorganisms12050957 -
Cellular and Molecular Gastroenterology... May 2024Dysregulated colonic epithelial cell (CEC) proliferation is a critical feature in the development of colorectal cancer. We show that NF-κB-inducing kinase (NIK)...
BACKGROUND & AIMS
Dysregulated colonic epithelial cell (CEC) proliferation is a critical feature in the development of colorectal cancer. We show that NF-κB-inducing kinase (NIK) attenuates colorectal cancer through coordinating CEC regeneration/differentiation via noncanonical NF-κB signaling that is unique from canonical NF-kB signaling.
METHODS
Initial studies evaluated crypt morphology/functionality, organoid generation, transcriptome profiles, and the microbiome. Inflammation and inflammation-induced tumorigenesis were initiated in whole-body NIK knockout mice (Nik) and conditional-knockout mice following administration of azoxymethane and dextran sulfate sodium.
RESULTS
Human transcriptomic data revealed dysregulated noncanonical NF-kB signaling. In vitro studies evaluating Nik crypts and organoids derived from mature, nondividing CECs, and colonic stem cells exhibited increased accumulation and stunted growth, respectively. Transcriptomic analysis of Nik cells revealed gene expression signatures associated with altered differentiation-regeneration. When assessed in vivo, Nik mice exhibited more severe colitis with dextran sulfate sodium administration and an altered microbiome characterized by increased colitogenic microbiota. In the inflammation-induced tumorigenesis model, we observed both increased tumor burdens and inflammation in mice where NIK is knocked out in CECs (Nik). Interestingly, this was not recapitulated when NIK was conditionally knocked out in myeloid cells (Nik). Surprisingly, conditional knockout of the canonical pathway in myeloid cells (RelA) revealed decreased tumor burden and inflammation and no significant changes when conditionally knocked out in CECs (RelA).
CONCLUSIONS
Dysregulated noncanonical NF-κB signaling is associated with the development of colorectal cancer in a tissue-dependent manner and defines a critical role for NIK in regulating gastrointestinal inflammation and regeneration associated with colorectal cancer.
PubMed: 38750899
DOI: 10.1016/j.jcmgh.2024.05.004 -
Animal Models and Experimental Medicine Feb 2024Nuclear receptor-binding SET domain 2 (NSD2) is a histone methyltransferase, that catalyzes dimethylation of lysine 36 of histone 3 (H3K36me2) and is associated with...
BACKGROUND
Nuclear receptor-binding SET domain 2 (NSD2) is a histone methyltransferase, that catalyzes dimethylation of lysine 36 of histone 3 (H3K36me2) and is associated with active transcription of a series of genes. NSD2 is overexpressed in multiple types of solid human tumors and has been proven to be related to unfavorable prognosis in several types of tumors.
METHODS
We established a mouse model in which the NSD2 gene was conditionally knocked out in intestinal epithelial cells. We used azoxymethane and dextran sodium sulfate to chemically induce murine colorectal cancer. The development of colorectal tumors were investigated using post-necropsy quantification, immunohistochemistry, and enzyme-linked immunosorbent assay (ELISA).
RESULTS
Compared with wild-type (WT) control mice, NSD2 -Vil1-Cre mice exhibited significantly decreased tumor numbers, histopathological changes, and cytokine expression in colorectal tumors.
CONCLUSIONS
Conditional knockout of NSD2 in intestinal epithelial cells significantly inhibits colorectal cancer progression.
PubMed: 38400589
DOI: 10.1002/ame2.12392 -
Heliyon Sep 2023Chronic inflammation-induced oxidative stress is an important driving force for developing colitis-associated cancer (CAC). 4-hydroxynonenal (4-HNE) is a highly reactive...
Chronic inflammation-induced oxidative stress is an important driving force for developing colitis-associated cancer (CAC). 4-hydroxynonenal (4-HNE) is a highly reactive aldehyde derived from lipid peroxidation of ω-6 polyunsaturated fatty acids that contributes to colorectal carcinogenesis. Glutathione -transferase alpha 4 (Gsta4) specifically conjugates glutathione to 4-HNE and thereby detoxifies 4-HNE. The correlation of these oxidative biomarkers with the pathological changes in CAC is, however, unclear. In this study, we investigated the expression of Gsta4 and 4-HNE adducts in azoxymethane/dextran sulfate sodium (AOM/DSS)-induced murine CAC, and analyzed the correlations of 4-HNE and Gsta4 with inflammatory cytokines and the pathological scores in the colon biopsies. Real-time quantitative PCR showed that expression of IL6, TNFα, and Gsta4 sequentially increased in colon tissues for mice treated with DSS for 1, 2, and 3 cycles, respectively. Moreover, immunohistochemical staining showed remarkably increased expression of 4-HNE adducts, Gsta4, TNFα, and IL6 in the colon biopsies after 3 cycles of DSS treatment. Correlation analysis demonstrated that 4-HNE adducts in the colon biopsies were positively correlated with Gsta4 expression. Additionally, the expression of Gsta4 and 4-HNE adducts were strongly correlated with the pathological changes of colon, as well as the expression of TNFα and IL6 in colon tissues. These results provide evidence for the association of oxidative biomarkers Gsta4 and 4-HNE with the pathological changes of CAC and may help developing novel histopathological biomarkers and prevention targets for CAC.
PubMed: 37810110
DOI: 10.1016/j.heliyon.2023.e19815 -
Inflammatory Bowel Diseases Aug 2023Myeloid cells are critical for iron and immune homeostasis. Ferritin heavy chain (FTH1) is essential for intracellular iron storage. Myeloid FTH1 is important in the...
BACKGROUND
Myeloid cells are critical for iron and immune homeostasis. Ferritin heavy chain (FTH1) is essential for intracellular iron storage. Myeloid FTH1 is important in the pathogenesis of many inflammatory diseases. However, the role of myeloid FTH1 in colitis and colitis-associated cancer has not been determined.
METHODS
Myeloid FTH1 deficient and wild-type mice were treated with dextran sodium sulfate (DSS) or azoxymethane (AOM)-DSS to compare their susceptibility to acute colitis or colitis-associated cancer.
RESULTS
Myeloid FTH1-deficient mice fed with a high-iron diet were less susceptible to DSS-induced acute colitis than wild type mice. Mechanistic studies showed that myeloid FTH1 deficiency resulted in lower expression of an iron uptake protein divalent metal transporter 1 (DMT1) and active phosphorylated signal transducer and activator of transcription 3 (STAT3) in the colon tissues. Our studies also showed that pharmacological STAT3 reactivation restored the susceptibility of myeloid FTH1-deficient mice to DSS-induced acute colitis. Consistently, myeloid FTH1-deficient mice fed with a high-iron diet had reduced DMT1, phosphorylated STAT3 and inflammation in their colon tissues, and were less susceptible to colitis-associated colorectal cancer.
CONCLUSIONS
Our study demonstrated that myeloid FTH1 is required for colitis and colitis-associated colorectal cancer via maintaining of DMT1-iron-STAT3 signaling activation under excess iron condition.
Topics: Animals; Mice; Colitis-Associated Neoplasms; STAT3 Transcription Factor; Iron; Colitis; Azoxymethane; Dextran Sulfate; Mice, Inbred C57BL; Disease Models, Animal; Colorectal Neoplasms
PubMed: 36745026
DOI: 10.1093/ibd/izad009 -
Biomedicine & Pharmacotherapy =... Jun 2024Colitis-associated cancer (CAC) in inflammatory bowel diseases exhibits more aggressive behavior than sporadic colorectal cancer; however, the molecular mechanisms...
Colitis-associated cancer (CAC) in inflammatory bowel diseases exhibits more aggressive behavior than sporadic colorectal cancer; however, the molecular mechanisms remain unclear. No definitive preventative agent against CAC is currently established in the clinical setting. We investigated the molecular mechanisms of CAC in the azoxymethane/dextran sulfate sodium (AOM/DSS) mouse model and assessed the antitumor efficacy of erlotinib, a small molecule inhibitor of the epidermal growth factor receptor (EGFR). Erlotinib premixed with AIN-93 G diet at 70 or 140 parts per million (ppm) inhibited tumor multiplicity significantly by 96%, with ∼60% of the treated mice exhibiting zero polyps at 12 weeks. Bulk RNA-sequencing revealed more than a thousand significant gene alterations in the colons of AOM/DSS-treated mice, with KEGG enrichment analysis highlighting 46 signaling pathways in CAC development. Erlotinib altered several signaling pathways and rescued 40 key genes dysregulated in CAC, including those involved in the Hippo and Wnt signaling. These findings suggest that the clinically-used antitumor agent erlotinib might be repurposed for suppression of CAC, and that further studies are warranted on the crosstalk between dysregulated Wnt and EGFR signaling in the corresponding patient population.
Topics: Animals; Erlotinib Hydrochloride; Disease Models, Animal; Colitis-Associated Neoplasms; Mice; Azoxymethane; Dextran Sulfate; ErbB Receptors; Carcinogenesis; Mice, Inbred C57BL; Male; Signal Transduction; Wnt Signaling Pathway; Antineoplastic Agents; Gene Expression Regulation, Neoplastic; Colitis
PubMed: 38723513
DOI: 10.1016/j.biopha.2024.116580 -
Acta Cirurgica Brasileira 2023The aim of this study was to assess the effects of resistance and aerobic exercise on colorectal cancer (CRC) development in mice induced by azoxymethane (AOM) coupled...
PURPOSE
The aim of this study was to assess the effects of resistance and aerobic exercise on colorectal cancer (CRC) development in mice induced by azoxymethane (AOM) coupled with colitis.
METHODS
Forty animals induced with CRC were used, divided into five groups of eight animals each: sedentary; continuous aerobics; continuous anaerobic; aerobic PI; and anaerobic PI. AOM was administered to the animals in two doses of 10 mg/kg each over the course of two weeks, the first dose administered in the third week and the second administered in the fourth. For the colitis, three cycles of dextran sodium sulfate were administered for five days, separated by two weeks of water. The 14th week of the experiment saw the euthanasia, the removal of their colons, and the creation of microscopy slides for histological analysis.
RESULTS
Preneoplastic lesions developed in all five groups; there were no significant differences between them. However, in terms of inflammatory symptoms, mucosal ulceration was much more frequently in the exercise groups than in the sedentary group (p = 0.016). The number of polyps overall (p = 0.002), the distal region's polyp development (p = 0.003), and the proximal region's polyp development (p = 0.04) were all statistically different than sedentary group.
CONCLUSIONS
The study discovered no significant difference in disease activity index scores between groups, but there was a significant difference in the number of polyps and the presence of mucosal ulceration in the colon.
Topics: Humans; Animals; Mice; Colorectal Neoplasms; Resistance Training; Disease Models, Animal; Colitis; Azoxymethane; Mice, Inbred C57BL; Dextran Sulfate
PubMed: 37878986
DOI: 10.1590/acb384923