Mutant p53 reactivator SLMP53-2 hinders ultraviolet B radiation-induced skin carcinogenesis.

The growing incidence of skin cancer (SC) has prompted the search for additional preventive strategies to counteract this global health concern. Mutant p53 (mutp53), particularly with ultraviolet radiation (UVR) signature, has emerged as a promising target for SC prevention based on its key role in skin carcinogenesis. Herein, the preventive activity of our previously disclosed mutp53 reactivator SLMP53-2 against UVR-induced SC was investigated. The pre-treatment of keratinocyte HaCaT cells with SLMP53-2, before UVB exposure, depleted mutp53 protein levels with restoration of wild-type-like p53 DNA-binding ability and subsequent transcriptional activity. SLMP53-2 increased cell survival by promoting G1-phase cell cycle arrest, while reducing UVB-induced apoptosis through inhibition of c-Jun N-terminal kinase (JNK) activity. SLMP53-2 also protected cells from reactive oxygen species and oxidative damage induced by UVB. Moreover, it enhanced DNA repair through upregulation of nucleotide excision repair pathway and depletion of UVB-induced DNA damage, as evidenced by a reduction of DNA in comet tails, γH2AX staining and cyclobutane pyrimidine dimers (CPD) levels. SLMP53-2 further suppressed UVB-induced inflammation by inhibiting the nuclear translocation and DNA-binding ability of NF-κB, and promoted the expression of key players involved in keratinocytes differentiation. Consistently, the topical application of SLMP53-2 in mice skin, prior to UVB irradiation, reduced cell death and DNA damage. It also decreased the expression of inflammatory-related proteins and promoted cell differentiation, in UVB-exposed mice skin. Notably, SLMP53-2 did not show signs of skin toxicity for cumulative topical use. Overall, these results support a promising protective activity of SLMP53-2 against UVB-induced SC.

First report of Spirocerca vulpis in red foxes (Vulpes vulpes) in Portugal.

Recent studies have described Spirocerca lupi-like nematodes in the stomach of red foxes (Vulpes vulpes) in Europe. A phylogenetic analysis of those specimens using mitochondrial DNA and their morphological reexamination allowed their characterization as a different species, Spirocerca vulpis. Between the years of 2010 and 2017, roundworms were collected from seven red foxes of northeastern Portugal found at necropsy with nodular lesions on their stomach wall. Histopathological analysis of four foxes revealed granulomatous lesions of the gastric nodules. On morphological assessment, by light microscopy, nematodes revealed the presence of six triangular teeth-like buccal capsule structures, which are absent in S. lupi. Polymerase chain reaction was run to amplify a 551 bp partial fragment of the cytochrome c oxidase subunit 1 gene. Sequences were 99% similar to S. vulpis (85% coverage) of red foxes from Spain and Bosnia and Herzegovina, 99% similar (99% coverage) to sequences of Spirocerca sp. of red foxes from Denmark and 93% similar (99% coverage) to S. lupi from South Africa. This is the first report of S. vulpis in foxes or any other host from Portugal.

Canine Spindle Cell Mammary Tumor: A Retrospective Study of 67 Cases.

Canine spindle cell mammary tumor (CSCMT) is an infrequent canine mammary tumor (CMT) composed of spindle or fusiform cells, which represents a challenge for pathologists and clinicians. Mammary tumors submitted for histopathology from 1998 to 2013 and compatible with CSCMTs were retrospectively selected. The tumors were diagnosed based on the hematoxylin and eosin (HE)-stained section; malignant tumors were graded using a canine soft tissue sarcoma grading scheme and a canine mammary tumor grading scheme, and they were further assigned a diagnosis based on immunohistochemistry (IHC) for pancytokeratin, cytokeratin 14, p63, calponin, vimentin, Ki-67, CD31, desmin, myosin, smooth muscle actin, glial fibrillary acidic protein, and S-100. The origin of the tumors was assessed as mammary, skin, or unknown. The prevalence of CSCMT was 1% of all CMTs. CSCMTs included 3 benign tumors (1 angioma and 2 benign myoepitheliomas) and 67 malignant tumors that after IHC were diagnosed as malignant myoepithelioma (64%), carcinoma and malignant myoepithelioma (19%), hemangiosarcoma (8%), undifferentiated sarcoma (5%), peripheral nerve sheath tumor (3%), and fibrosarcoma (2%). The diagnosis based on the HE-stained section differed from the diagnosis after IHC in 75% of the malignant cases. The majority of malignant CSCMTs were solitary (57%) large tumors (6.42 ± 3.92 cm) with low metastatic potential and high survival rate (8% tumor-related mortality). Higher sarcoma grade was associated with older age (P = .034) and greater tumor size (P = .037). Malignant CSCMTs need to be evaluated by IHC to ensure the histotype and the relatively benign clinical behavior, despite their large size.

Canine invasive mammary carcinomas as models of human breast cancer. Part 1: natural history and prognostic factors.

PURPOSE: Dogs have been proposed as spontaneous animal models of human breast cancer, based on clinicopathologic similarities between canine and human mammary carcinomas. We hypothesized that a better knowledge of the natural history and prognostic factors of canine invasive mammary carcinomas would favor the design of preclinical trials using dogs as models of breast cancer. METHODS: The 2-year outcome of 350 female dogs with spontaneous invasive mammary carcinoma was studied. The investigated prognostic factors included age at diagnosis, pathologic tumor size, pathologic nodal stage, lymphovascular invasion, histological grade, and expression of Estrogen Receptor alpha (ERα), Progesterone Receptor, Ki-67, Human Epidermal Growth Factor Receptor 2, basal cytokeratins 5/6, and Epidermal Growth Factor Receptor. Multivariate survival analyses were performed using the Cox proportional hazards model. RESULTS: The overall survival after mastectomy was 11 months. Within 1 year post mastectomy, 41.5% of dogs (145/350) died from their mammary carcinoma. By multivariate analysis, the significant prognostic factors for overall survival included a pathologic tumor size larger than 20 mm [HR 1.47 (95% confidence interval 1.15-1.89)], a positive nodal stage [pN+, HR 1.89 (1.43-2.48)], a histological grade III [HR 1.32 (1.02-1.69)], ERα negativity [HR 1.39 (1.01-1.89)], a high Ki-67 proliferation index [HR 1.32 (1.04-1.67)], and EGFR absence [HR 1.33 (1.04-1.69)]. CONCLUSION: The short natural history of spontaneous canine invasive mammary carcinomas and high rate of cancer-related death allow for rapid termination of preclinical investigations. The prognostic factors of invasive mammary carcinomas are remarkably similar in dogs and humans, highlighting the similarities in cancer biology between both species.

Canine invasive mammary carcinomas as models of human breast cancer. Part 2: immunophenotypes and prognostic significance.

PURPOSE: Relevant animal models of human breast cancer are currently needed, especially for the aggressive triple-negative breast cancer subtype. Recent studies and our results (Part 1) indicate that spontaneous canine invasive mammary carcinomas (CMCs) resemble human breast cancer by clinics and pathology as well as behavior and prognostic indicators. We hypothesized that the current molecular classifications of human breast cancer, used for therapeutic decision, could be relevant to dogs. METHODS: Three hundred and fifty female dogs with spontaneous CMC and a 2-year follow-up were retrospectively included. By immunohistochemistry, CMCs were classified according to Nielsen (Clin Cancer Res 10:5367-5374, 2004) and Blows (PlosOne doi: 10.1371/journal.pmed.1000279, 2010) into the subtypes of human breast cancer. RESULTS: Four immunophenotypes were defined either according to Nielsen classification (luminal A 14.3%, luminal B 9.4%, triple-negative basal-like 58.6%, and triple-negative nonbasal-like 17.7% CMCs); or to Blows classification (luminal 1-: 11.4%, luminal 1+: 12.3%, Core basal phenotype: 58.6%, and five-negative phenotype: 17.7%). No HER2-overexpressing CMC as defined by a 3 + immunohistochemical score was observed in our cohort. By univariate and multivariate analyses, both immunophenotypical classifications applied to CMCs showed strong prognostic significance: luminal A or luminal 1+ CMCs showed a significantly longer disease-free interval (HR = 0.46), Overall (HR = 0.47), and Specific Survival (HR = 0.56) compared to triple-negative carcinomas, after adjustment for stage. CONCLUSIONS: In our cohort, triple-negative CMCs largely predominated (76%), were much more prevalent than in human beings, and showed an aggressive natural behavior after mastectomy. Dogs are thus potent valuable spontaneous models to test new therapeutic strategies for this particular subtype of breast cancer.

A Contrast-Enhanced Ultrasonographic Study About the Impact of Long-term Exercise Training on Mammary Tumor Vascularization.

OBJECTIVES: This study aimed to evaluate the impact of long-term exercise training on the vascularization of rat mammary tumors. METHODS: Female rats were divided into 4 groups: N-methyl-N-nitrosourea (MNU) treated sedentary, MNU treated exercised, control sedentary, and control exercised. Tumor development was induced in the MNU groups by MNU administration. Exercised groups were trained for 35 weeks. Tumor vascularization was evaluated by pulsed Doppler and contrast-enhanced ultrasonography. RESULTS: The pulsatility and resistive indices were slightly higher in the MNU sedentary group (P > .05). Mammary tumors mainly had centripetal and heterogeneous enhancement of the contrast, clear margins, and the presence of penetrating vessels. The MNU exercised group had a lower arrival time and time to peak and higher peak intensity, wash-in, and wash-out (P > .05). The area under the curve was similar between groups (P > .05). CONCLUSIONS: The contrast-enhanced ultrasonographic study did not detect differences in mammary tumor vascularization between MNU sedentary and MNU exercised groups previously detected by power Doppler imaging, B-flow imaging, and immunohistochemistry.

Histopathological and in vivo evidence of regucalcin as a protective molecule in mammary gland carcinogenesis.

Regucalcin (RGN) is a calcium-binding protein, which has been shown to be underexpressed in cancer cases. This study aimed to determine the association of RGN expression with clinicopathological parameters of human breast cancer. In addition, the role of RGN in malignancy of mammary gland using transgenic rats overexpressing the protein (Tg-RGN) was investigated. Wild-type (Wt) and Tg-RGN rats were treated with 7,12-dimethylbenz[α]anthracene (DMBA). Carcinogen-induced tumors were histologically classified and the Ki67 proliferation index was estimated. Immunohistochemistry analysis showed that RGN immunoreactivity was negatively correlated with the histological grade of breast infiltrating ductal carcinoma suggesting that progression of breast cancer is associated with loss of RGN. Tg-RGN rats displayed lower incidence of carcinogen-induced mammary gland tumors, as well as lower incidence of invasive forms. Moreover, higher proliferation was observed in non-invasive tumors of Wt animals comparatively with Tg-RGN. Overexpression of RGN was associated with diminished expression of cell-cycle inhibitors and increased expression of apoptosis inducers. Augmented activity of apoptosis effector caspase-3 was found in the mammary gland of Tg-RGN. RGN overexpression protected from carcinogen-induced mammary gland tumor development and was linked with reduced proliferation and increased apoptosis. These findings indicated the protective role of RGN in the carcinogenesis of mammary gland.

Electron Microscopy Findings in N-Methyl-N-Nitrosourea-Induced Mammary Tumors.

Although the rat model of mammary tumors chemically induced by N-methyl-N-nitrosourea (MNU) has been frequently used by several research teams, there is a lack of ultrastructural studies in this field. The main aim of this work was to perform an ultrastructural characterization of MNU-induced mammary tumors in female rats. Some alterations previously reported in human mammary tumors, such as nucleus size and shape, accumulation of heterochromatin in the perinuclear region, and interdigitating cytoplasmic processes between cancer cells were also observed in MNU-induced mammary tumors. Although a low number of samples were analyzed by transmission electron microscopy in the present study, we consider that it may contribute to a better understanding of MNU-induced mammary carcinogenesis in a rat model. The ultrastructural characteristics of the two most frequently diagnosed mammary carcinomas described in the present work can be useful to differentiate them from other histological patterns. In addition, the loss of cytoplasm in neoplastic cells and formation of vacuoles were described.

N-Methyl-N-nitrosourea as a mammary carcinogenic agent.

The administration of chemical carcinogens is one of the most commonly used methods to induce tumors in several organs in laboratory animals in order to study oncologic diseases of humans. The carcinogen agent N-methyl-N-nitrosourea (MNU) is the oldest member of the nitroso compounds that has the ability to alkylate DNA. MNU is classified as a complete, potent, and direct alkylating compound. Depending on the animals' species and strain, dose, route, and age at the administration, MNU may induce tumors' development in several organs. The aim of this manuscript was to review MNU as a carcinogenic agent, taking into account that this carcinogen agent has been frequently used in experimental protocols to study the carcinogenesis in several tissues, namely breast, ovary, uterus, prostate, liver, spleen, kidney, stomach, small intestine, colon, hematopoietic system, lung, skin, retina, and urinary bladder. In this paper, we also reviewed the experimental conditions to the chemical induction of tumors in different organs with this carcinogen agent, with a special emphasis in the mammary carcinogenesis.

Exploring the role of microRNAs as diagnostic and prognostic biomarkers in canine mammary tumors.

Canine mammary tumors (CMTs) represent a significant health concern in dogs, with a high incidence among intact female dogs. CMTs are a promising comparative model for human breast cancer, due to sharing several pathophysiological features. Additionally, CMTs have a strong genetic correlation with their human counterpart, including the expression of microRNAs (miRNAs). MiRNAs are a class of non-coding RNAs that play important roles in post-translational regulation of gene expression, being implicated in carcinogenesis, tumor progression, and metastasis. Moreover, miRNAs hold promise as diagnostic, prognostic, and metastatic biomarkers. Understanding the molecular mechanisms underlying CMTs is crucial for improving diagnosis, prognosis, and monitoring of treatments. Herein, we provide a comprehensive overview of the current knowledge on miRNAs in CMTs, highlighting their roles in carcinogenesis and their potential as biomarkers. Additionally, we highlight the current limitations and critically discuss the overarching challenges in this field, emphasizing the need for future research to translate miRNA findings into veterinary clinical practice.

Mammary Glands of Women, Female Dogs and Female Rats: Similarities and Differences to Be Considered in Breast Cancer Research.

Breast cancer is one of the most common and well-known types of cancer among women worldwide and is the most frequent neoplasm in intact female dogs. Female dogs are considered attractive models or studying spontaneous breast cancer, whereas female rats are currently the most widely used animal models for breast cancer research in the laboratory context. Both female dogs and female rats have contributed to the advancement of scientific knowledge in this field, and, in a "One Health" approach, they have allowed broad understanding of specific biopathological pathways, influence of environmental factors and screening/discovery of candidate therapies. This review aims to clearly showcase the similarities and differences among woman, female dog and female rat concerning to anatomical, physiological and histological features of the mammary gland and breast/mammary cancer epidemiology, in order to better portray breast tumorigenesis, and to ensure appropriate conclusions and extrapolation of results among species. We also discuss the major aspects that stand out in these species. The mammary glands of female dogs and women share structural similarities, especially with respect to the lactiferous ducts and lymphatic drainage. In contrast, female rats have only one lactiferous duct per nipple. A comprehensive comparison between humans and dogs is given a special focus, as these species share several aspects in terms of breast/mammary cancer epidemiology, such as age of onset, hormonal etiology, risk factors, and the clinical course of the disease. Holistically, it is clear that each species has advantages and limitations that researchers must consider during the development of experimental designs and data analysis.

Detection of Abnormal Prion Protein by Immunohistochemistry.

Abnormal prion proteins (PrPSc) are the disease-associated isoform of cellular prion protein and diagnostic markers of transmissible spongiform encephalopathies (TSEs). These neurodegenerative diseases affect humans and several animal species and include scrapie, zoonotic bovine spongiform encephalopathy (BSE), chronic wasting disease of cervids (CWD), and the newly identified camel prion disease (CPD). Diagnosis of TSEs relies on immunodetection of PrPSc by application of both immunohistochemistry (IHC) and western immunoblot methods (WB) on encephalon tissues, namely, the brainstem (obex level). IHC is a widely used method that uses primary antibodies (monoclonal or polyclonal) against antigens of interest in cells of a tissue section. The antibody-antigen binding can be visualized by a color reaction that remains localized in the area of the tissue or cell where the antibody was targeted. As such, in prion diseases, as in other fields of research, the immunohistochemistry techniques are not solely used for diagnostic purposes but also in pathogenesis studies. Such studies involve detecting the PrPSc patterns and types from those previously described to identify the new prion strains. As BSE can infect humans, it is recommended that biosafety laboratory level-3 (BSL-3) facilities and/or practices are used to handle cattle, small ruminants, and cervid samples included in the TSE surveillance. Additionally, containment and prion-dedicated equipment are recommended, whenever possible, to limit contamination. The PrPSc IHC procedure consists of a formic acid epitope-demasking step also acting as a prion inactivation measure, as formalin-fixed and paraffin-embedded tissues used in this technique remain infectious. When interpreting the results, care must be taken to distinguish non-specific immunolabeling from target labeling. For this purpose, it is important to recognize artifacts of immunolabeling obtained in known TSE-negative control animals to differentiate those from specific PrPSc immunolabeling types, which can vary between TSE strains, host species, and prnp genotype, further described herein.

A spontaneous ovarian teratoma in an FVB/n female mouse: Case report and literature review.

Background: Teratomas are rare types of germ cell neoplasms composed of various differentiated or undifferentiated tissues. Case Description: A 25-week-old female control FVB /n mouse in a 4-week toxicity study presented abdominal distension and poor body condition. It was euthanized, and the necropsy examination revealed a large mass connected to the tip of the right uterine horn, occupying the entire abdominal cavity. Microscopically, this mass showed areas of epidermal differentiation, with laminated keratin and sebaceous glands, differentiation into respiratory and digestive epithelium, cartilage, bone, and extensive areas of differentiation into the nervous tissue, being classified as an ovarian teratoma. Conclusion: As far as authors know, the occurrence of ovarian teratomas in the FVB/n mouse strain has never been previously described.

Estimating sequence diversity of prion protein gene (PRNP) in Portuguese populations of two cervid species: red deer and fallow deer.

Among the transmissible spongiform encephalopathies (TSEs), chronic wasting disease (CWD) in cervids is now a rising concern in wildlife within Europe, after the detection of the first case in Norway in 2016, in a wild reindeer and until June 2022 a total of 34 cases were described in Norway, Sweden and Finland. The definite diagnosis is post-mortem, performed in target areas of the brain and lymph nodes. Samples are first screened using a rapid test and, if positive, confirmed by immunohistochemistry and Western immunoblotting. The study of the genetics of the prion protein gene, PRNP, has been proved to be a valuable tool for determining the relative susceptibility to TSEs. In the present study, the exon 3 of PRNP gene of 143 samples from red deer (Cervus elaphus) and fallow deer (Dama dama) of Portugal was analysed. Three single nucleotide polymorphisms (SNPs) were found in red deer - codon A136A, codon T98A, codon Q226E - and no sequence variation was detected in fallow deer. The low genetic diversity found in our samples is compatible with previous studies in Europe. The comparison with results from North America suggests that the free-ranging deer from our study may present susceptibility to CWD, although lack of experimental data and the necessity of continuous survey are necessary to evaluate these populations.

Modelling the impact of nucleolin expression level on the activity of F3 peptide-targeted pH-sensitive pegylated liposomes containing doxorubicin.

Strategies targeting nucleolin have enabled a significant improvement in intracellular bioavailability of their encapsulated payloads. In this respect, assessment of the impact of target cell heterogeneity and nucleolin homology across species (structurally and functionally) is of major importance. This work also aimed at mathematically modelling the nucleolin expression levels at the cell membrane, binding and internalization of pH-sensitive pegylated liposomes encapsulating doxorubicin and functionalized with the nucleolin-binding F3 peptide (PEGASEMP), and resulting cytotoxicity against cancer cells from mouse, rat, canine, and human origin. Herein, it was shown that nucleolin expression levels were not a limitation on the continuous internalization of F3 peptide-targeted liposomes, despite the saturable nature of the binding mechanism. Modeling enabled the prediction of nucleolin-mediated total doxorubicin exposure provided by the experimental settings of the assessment of PEGASEMP's impact on cell death. The former increased proportionally with nucleolin-binding sites, a measure relevant for patient stratification. This pattern of variation was observed for the resulting cell death in nonsaturating conditions, depending on the cancer cell sensitivity to doxorubicin. This approach differs from standard determination of cytotoxic concentrations, which normally report values of incubation doses rather than the actual intracellular bioactive drug exposure. Importantly, in the context of development of nucleolin-based targeted drug delivery, the structural nucleolin homology (higher than 84%) and functional similarity across species presented herein, emphasized the potential to use toxicological data and other metrics from lower species to infer the dose for a first-in-human trial.

Assuring the Biofunctionalization of Silicone Covalently Bonded to Rhamnolipids: Antibiofilm Activity and Biocompatibility.

Silicone-based medical devices composed of polydimethylsiloxane (PDMS) are widely used all over the human body (e.g., urinary stents and catheters, central venous catheters stents) with extreme clinical success. Nevertheless, their abiotic surfaces, being prone to microorganism colonization, are often involved in infection occurrence. Improving PDMS antimicrobial properties by surface functionalization with biosurfactants to prevent related infections has been the goal of different works, but studies that mimic the clinical use of these novel surfaces are missing. This work aims at the biofunctional assessment of PDMS functionalized with rhamnolipids (RLs), using translational tests that more closely mimic the clinical microenvironment. Rhamnolipids were covalently bonded to PDMS, and the obtained surfaces were characterized by contact angle modification assessment, ATR-FTIR analysis and atomic force microscopy imaging. Moreover, a parallel flow chamber was used to assess the Staphylococcus aureus antibiofilm activity of the obtained surfaces under dynamic conditions, and an in vitro characterization with human dermal fibroblast cells in both direct and indirect characterization assays, along with an in vivo subcutaneous implantation assay in the translational rabbit model, was performed. A 1.2 log reduction in S. aureus biofilm was observed after 24 h under flow dynamic conditions. Additionally, functionalized PDMS lessened cell adhesion upon direct contact, while supporting a cytocompatible profile, within an indirect assay. The adequacy of the biological response was further validated upon in vivo subcutaneous tissue implantation. An important step was taken towards biofunctional assessment of RLs-functionalized PDMS, reinforcing their suitability for medical device usage and infection prevention.

Pulegone and Eugenol Oral Supplementation in Laboratory Animals: Results from Acute and Chronic Studies.

Essential oils are natural compounds used by humans for scientific purposes due to their wide range of properties. Eugenol is mostly present in clove oil, while pulegone is the main constituent of pennyroyal oil. To guarantee the safe use of eugenol and pulegone for both humans and animals, this study addressed, for the first time, the effects of these compounds, at low doses (chronic toxicity) and high doses (acute toxicity), in laboratory animals. Thirty-five FVB/n female mice were randomly assigned to seven groups (n = 5): group I (control, non-additive diet); group II (2.6 mg of eugenol + 2.6 mg of pulegone); group III (5.2 mg of eugenol + 5.2 mg of pulegone); group IV (7.8 mg of eugenol + 7.8 mg of pulegone); group V (7.8 mg of eugenol); group VI (7.8 mg of pulegone); and group VII (1000 mg of eugenol + 1000 mg of pulegone). The compounds were administered in the food. Groups I to VI were integrated into the chronic toxicity study, lasting 28 days, and group VII was used in the acute toxicity study, lasting 7 days. Animals were monitored to assess their general welfare. Water and food intake, as well as body weight, were recorded. On the 29th day, all animals were euthanized by an overdose of ketamine and xylazine, and a complete necropsy was performed. Blood samples were collected directly from the heart for microhematocrit and serum analysis, as well as for comet assay. Organs were collected, weighed, and fixed in formaldehyde for further histological analysis and enzymatic assay. Eugenol and pulegone induced behavioral changes in the animals, namely in the posture, hair appearance and grooming, and in mental status. These compounds also caused a decrease in the animals' body weight, as well as in the food and water consumption. A mortality rate of 20% was registered in the acute toxicity group. Both compounds modulated the serum levels of triglycerides and alanine aminotransferase. Eugenol and pulegone induced genetic damage in all animals. Eugenol increased the activity of the CAT enzyme. Both compounds increased the GR enzyme at the highest dose. Moreover, pulegone administered as a single compound increased the activity of the GST enzyme. Histopathological analysis revealed inflammatory infiltrates in the lungs of groups II, III, and IV. The results suggest that eugenol and pulegone may exert beneficial or harmful effects, depending on the dose, and if applied alone or in combination.

An Integrative Approach to Characterize the Early Phases of Dimethylhydrazine-Induced Colorectal Carcinogenesis in the Rat.

This study aimed to characterize an animal model of colorectal cancer (CRC) in the early stages of disease development. Twenty-nine male Wistar rats were divided into two control groups (CTRL1 and CTRL2), receiving EDTA-saline injections and two induced groups (CRC1 and CRC2), receiving 1,2-dimethylhydrazine (DMH) injections for seven consecutive weeks. CRC1 and CTRL1 were euthanized at the 11th week, while CRC2 and CTRL2 were euthanized at the 17th week. DMH treatment decreased microhematocrit values and IL-6, ghrelin, and myostatin serum levels. Histopathological analysis of intestinal sections showed that DMH-treated rats were characterized by moderate to severe epithelial dysplasia. An adenoma was observed in one animal (CRC2 group), and the presence of inflammatory infiltrate at the intestinal level was primarily observed in DMH-treated animals. DMH also induced Ki-67 immunoexpression. The gut microbiota analysis showed a higher abundance of Firmicutes, Clostridia, Clostridiales, Peptostreptococcaceae, Blautia, Romboutsia, and Clostridium sensu stricto in CRC than CTRL rats, whereas Prevotellaceae, Prevotella, Akkermansia, and Lactobacillus levels were more prevalent in CTRL animals. Our results suggest that this model could be helpful to investigate chemoprevention in the early stages of CRC.

Animal Models of Colorectal Cancer: From Spontaneous to Genetically Engineered Models and Their Applications.

Colorectal cancer is one of the most common gastrointestinal malignancies in humans, affecting approximately 1.8 million people worldwide. This disease has a major social impact and high treatment costs. Animal models allow us to understand and follow the colon cancer progression; thus, in vivo studies are essential to improve and discover new ways of prevention and treatment. Dietary natural products have been under investigation for better and natural prevention, envisioning to show their potential. This manuscript intends to provide the readers a review of rodent colorectal cancer models available in the literature, highlighting their advantages and disadvantages, as well as their potential in the evaluation of several drugs and natural compounds' effects on colorectal cancer.

Refinement of Animal Model of Colorectal Carcinogenesis through the Definition of Novel Humane Endpoints.

This study aimed to define appropriate humane endpoints (HEs) for an animal model of colorectal carcinogenesis (CRC). Twenty-nine male Wistar rats were divided into two control groups (CTRL1 and CTRL2) injected with ethylenediamine tetraacetic acid (EDTA)-saline solutions and two induced groups (CRC1 and CRC2) injected with 1,2-dimethylhydrazine (DMH) for seven weeks. A score sheet with 14 biological parameters was used to assess animal welfare. Groups CRC1 and CTRL1 and groups CRC2 and CTRL2 were euthanized 11 and 17 weeks after the first DMH administration, respectively. Five animals from the induced groups died unexpectedly during the protocol (survival rates of 75.0% and 66.7% for groups CRC1 and CRC2, respectively). The final mean body weight (BW) was smaller in the CRC groups when compared with that in the CTRL groups. A uniformity of characteristics preceding the premature animals' death was observed, namely an increase of 10% in mean BW, swollen abdomen, diarrhea, and priapism. The surface abdominal temperature of group CRC2 was significantly higher, when compared with that of group CTRL2. The parameters already described in other cancer models proved to be insufficient. For the CRC model, we considered assessing the abdominal temperature, priapism, and sudden increase in the BW.