Current views on in vivo models for breast cancer research and related drug development.

INTRODUCTION: Animal models play a crucial role in breast cancer research, in particular mice and rats, who develop mammary tumors that closely resemble their human counterparts. These models allow the study of mechanisms behind breast carcinogenesis, as well as the efficacy and safety of new, and potentially more effective and advantageous therapeutic approaches. Understanding the advantages and disadvantages of each model is crucial to select the most appropriate one for the research purpose. AREA COVERED: This review provides a concise overview of the animal models available for breast cancer research, discussing the advantages and disadvantages of each one for searching new and more effective approaches to treatments for this type of cancer. EXPERT OPINION: Rodent models provide valuable information on the genetic alterations of the disease, the tumor microenvironment, and allow the evaluation of the efficacy of chemotherapeutic agents. However, in vivo models have limitations, and one of them is the fact that they do not fully mimic human diseases. Choosing the most suitable model for the study purpose is crucial for the development of new therapeutic agents that provide better care for breast cancer patients.

Lifelong exercise training promotes the remodelling of the immune system and prostate signalome in a rat model of prostate carcinogenesis.

This work aimed to understand how lifelong exercise training promotes the remodelling of the immune system and prostate signalome in a rat model of PCa. Fifty-five male Wistar rats were divided into four groups: control sedentary, control exercised, induced PCa sedentary and induced PCa exercised. Exercised animals were trained in a treadmill for 53 weeks. Pca induction consisted on the sequential administration of flutamide, N-methyl-N-nitrosourea and testosterone propionate implants. Serum concentrations of C-reactive protein (CRP) and tumor necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK) were not different among groups. Peripheral levels of γδ T cells were higher in Pca exercised group than in the PCa sedentary group (p < 0.05). Exercise training also induced Oestrogen Receptor (ESR1) upregulation and Mitogen-activated Protein Kinase 13 (MAPK13) downregulation, changed the content of the phosphorylated (at Ser-104) form of this receptor (coded by the gene ESR1) and seemed to increase Erα phosphorylation and activity in exercised PCa rats when compared with sedentary PCa rats. Our data highlight the exercise-induced remodelling of peripheral lymphocyte subpopulations and lymphocyte infiltration in prostate tissue. Moreover, exercise training promotes the remodelling prostate signalome in this rat model of prostate carcinogenesis.

Klotho in Cancer: Potential Diagnostic and Prognostic Applications.

Klotho proteins, αKlotho, ßKlotho, and γKlotho, exert tumor-suppressive activities via the fibroblast growth factor receptors and multiple cell-signaling pathways. There is a growing interest in Klotho proteins as potential diagnostic and prognostic biomarkers for multiple diseases. However, recent advances regarding their roles and potential applications in cancer remain disperse and require an integrated analysis. The present review analyzed research articles published between 2012 and 2022 in the Cochrane and Scopus scientific databases to study the role of Klotho in cancer and their potential as tools for diagnosing specific cancer types, predicting tumor aggressiveness and prognosis. Twenty-six articles were selected, dealing with acute myeloid leukemia and with bladder, breast, colorectal, esophageal, gastric, hepatocellular, ovarian, pancreatic, prostatic, pulmonary, renal, and thyroid cancers. αKlotho was consistently associated with improved prognosis and may be useful in estimating patient survival. A single study reported the use of soluble αKlotho levels in blood serum as a tool to aid the diagnosis of esophageal cancer. γKlotho was associated with increased aggressiveness of bladder, breast, and prostate cancer, and ßKlotho showed mixed results. Further clinical development of Klotho-based assays will require careful identification of specific tumor subtypes where Klotho proteins may be most valuable as diagnostic or prognostic tools.

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.

Malignancy Associated with Low-Risk HPV6 and HPV11: A Systematic Review and Implications for Cancer Prevention.

High-risk human papillomavirus (HPV) is etiologically related to cervical cancer, other anogenital cancers and oropharyngeal carcinomas. Low-risk HPV, especially HPV6 and HPV11, cause genital warts and laryngeal papillomas. However, the accumulating data suggests that HPV6 and HPV11 may cause malignant lesions at non-cervical anatomic sites. This review aims to estimate the proportions of single and dual HPV6/11 infections in multiple cancers reported in the last 10 years in the Cochrane, Embasa and PubMed databases. Secondly, the genomes of HPV6/11 were compared with the most common high-risk genotype, HPV16, to determine the similarities and differences. A total of 11 articles were selected, including between one and 334 HPV+ cancer patients. The frequencies of single or dual HPV6/11 infections ranged between 0-5.5% for penile and 0-87.5% for laryngeal cancers and were null for vulvar, vaginal and oral cancers. The genomic similarities between HPV6/11 and HPV16 mainly involved the E7 gene, indicating a limited ability to block cell differentiation. The presence of single or dual HPV6/11 infections in variable proportions of penile and laryngeal cancers support the vaccination strategies that cover these genotypes, not only for preventing genital warts but also for cancer prevention. Other risk factors and co-carcinogens are likely to participate in epithelial carcinogenesis associated with low-risk HPV.

Realistic aspects behind the application of the rat model of chemically-induced mammary cancer: Practical guidelines to obtain the best results.

Cancer is one of the most important public health problems worldwide. Despite the great contribution of in-vitro studies for biomedical research, animals are essential to study diseases' biopathology and diagnosis, and searching for new preventive and therapeutic strategies. Breast cancer is currently the most common cancer globally, accounting for 12.5% of all new annual cancer cases worldwide. Although the rat model of mammary cancer chemically-induced is widely used to study this disease, there is a lack of standardization in procedures for cancer induction, sample collection, and analysis. Therefore, it is important to provide a practical guide for researchers aiming to work with this model to make the analysis of results more uniform. Thus, in this review, we provide the researchers with a detailed step-by-step guide to implement a rat model of mammary cancer, based on our wide experience in this field, to obtain the best results, maximum throughput of each experiment, and easy comparison among researches.

Quercus spp. extract as a promising preventive or therapeutic strategy for cancer: A systematic review.

Acorns have traditionally been used in the human diet and for the treatment of specific diseases. Therefore, the present study performed a systematic review of studies which investigated the effects of Quercus spp. extracts in cancer prevention and treatment. A systematic literature search was performed for original records which addressed the anticancer effects of Quercus spp. extract in in vitro and in vivo cancer models. Body composition, food consumption, tumor development and/or toxicity were evaluated in in vivo studies, while cytotoxicity was evaluated in in vitro studies. Few studies and low sample sizes presented a challenge in the drawing of solid conclusions. Overall, the results suggested a positive impact of Quercus spp. extract, by reducing cancer development. Therefore, more studies with different cancer cell lines and animal models to address the efficacy of the acorn extracts in several types of cancer are required. Furthermore, the effects of acorn flour, incorporated in the diet, in an animal model of mammary cancer should be evaluated.

Canine and feline papillomaviruses: an update.

Papillomaviruses are small viruses able to cause disease not only in mammalians, but also in birds and reptiles. In recent years, a rising number of papillomaviruses have been identified in dogs and cats, totaling 24 canine papillomavirus (CPV) and six feline papillomavirus (FcaPV). The canine and feline papillomaviruses (CPVs and FcaPVs, respectively) are responsible for multiple lesions in these domestic species but the potential pathological relevance of some recently identified types remains to be determined. CPVs are associated with oral papillomatosis, cutaneous papillomas and viral pigmented plaques, and have been rarely associated with the development of oral and cutaneous squamous cell carcinomas in their canine hosts. FcaPVs are associated with oral papillomas, viral plaques, and Bowenoid in situ carcinomas. The present review provides readers with the more recent advances on dog and cat papillomavirus research, bringing an update on this field to both veterinary practitioners and the virology community at large.

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.

The Impact of Resveratrol-Enriched Bread on Cardiac Remodeling in a Preclinical Model of Diabetes.

The World Health Organization aims to stop the rise of diabetes by 2025, and diet is one of the most efficient non-pharmacological strategies used to prevent it. Resveratrol (RSV) is a natural compound with anti-diabetic properties, and incorporating it into bread is a suitable way to make it more accessible to consumers as it can be included as part of their daily diet. This study aimed to evaluate the effect of RSV-enriched bread in preventing early type 2 diabetes cardiomyopathy in vivo. Male Sprague Dawley rats (3 weeks old) were divided into four groups: controls with plain bread (CB) and RSV bread (CBR), and diabetics with plain bread (DB) and RSV bread (DBR). Type 2 diabetes was induced by adding fructose to the drinking water for two weeks followed by an injection of streptozotocin (STZ) (40 mg/kg). Then, plain bread and RSV bread (10 mg RSV/kg body weight) were included in the rats' diet for four weeks. Cardiac function, anthropometric, and systemic biochemical parameters were monitored, as well as the histology of the heart and molecular markers of regeneration, metabolism, and oxidative stress. Data showed that an RSV bread diet decreased the polydipsia and body weight loss observed in the early stages of the disease. At the cardiac level, an RSV bread diet diminished fibrosis but did not counteract the dysfunction and metabolic changes seen in fructose-fed STZ-injected rats.

Studying and Analyzing Humane Endpoints in the Fructose-Fed and Streptozotocin-Injected Rat Model of Diabetes.

This work aimed to define a humane endpoint scoring system able to objectively identify signs of animal suffering in a rat model of type 2 diabetes. Sprague-Dawley male rats were divided into control and induced group. The induced animals drink a 10% fructose solution for 14 days. Then, received an administration of streptozotocin (40 mg/kg). Animals' body weight, water and food consumption were recorded weekly. To evaluate animal welfare, a score sheet with 14 parameters was employed. Blood glucose levels were measured at three time points. After seven weeks of initiating the protocol, the rats were euthanized. The induced animals showed weight loss, polyuria, polyphagia, and polydipsia. According to our humane endpoints table, changes in animal welfare became noticeable after the STZ administration. None of the animals hit the critical score limit (four). Data showed that the most effective parameters to assess welfare in this type 2 diabetes rat induction model were dehydration, grooming, posture, abdominal visualization, and stool appearance. The glycemia was significantly higher in the induced group when compared to the controls (p < 0.01). Induced animals' murinometric and nutritional parameters were significantly lower than the controls (p < 0.01). Our findings suggest that in this rat model of type 2 diabetes with STZ-induced following fructose consumption, our list of humane endpoints is suitable for monitoring the animals' welfare.

Editorial: Special Issue "Addressing New Therapeutic Strategies Using Models".

A disease model displays pathological processes observed in human or animal diseases [...].

Evolution of Models of Prostate Cancer: Their Contribution to Current Therapies.

Prostate cancer (PCa) is among the most frequent cancers worldwide. Nowadays, several therapeutic strategies are available for PCa treatment, namely chemotherapy, immunotherapy, radiotherapy, and hormonal therapy. Despite existing therapeutic approaches, in vitro and in vivo models are essential to better understand cancer development and search for more effective therapies, with a positive impact in cancer patient survival and quality of life. Among several models available, the rat model is the one most frequently used, since it shares anatomical, physiological, pathological, and behavioral features with humans. Animal models can be classified as: spontaneous, chemically-induced; hormonally-induced; implantation of cancer cell lines obtained from humans or from the same species, in the place of disease development or in a different place; and genetically-modified models. The chemically-induced models are among the most frequently used for PCa research. This manuscript provides a comprehensive overview of PCa models, presenting their application, advantages, and disadvantages, and their importance for the development of current therapies for prostate cancer.

Appraising Animal Models of Prostate Cancer for Translational Research: Future Directions.

The growing incidence of prostate cancer has prompted a great investment in basic biology and translational studies to develop new therapies. Multiple animal models have been established to study etiological factors, cancer-preventive strategies and the molecular determinants of aggressiveness and metastases. The rat model of prostate cancer induced by chemical carcinogen N-methyl-N-nitrosourea (MNU) and testosterone exposure has become an important tool to study prostatic carcinogenesis and chemopreventive approaches. Over prolonged treatment, this model develops prostatic lesions that closely mimic those observed in human patients. By modifying the experimental conditions, different research groups have been able to induce a vast spectrum of lesions, ranging from early prostatic intraepithelial neoplasia to metastatic cancer. These carefully tuned experimental settings allowed researchers to test lifestyle interventions, and different pharmacological and chemopreventive strategies. However, this model's great flexibility requires careful planning to ensure that the experimental conditions are adequate to obtain the spectrum of lesions intended. The present review addresses such issues, highlighting the value of the rat prostate cancer model and the multiple challenges and opportunities it offers to researchers worldwide.

Long-Term Exposure to Supraphysiological Levels of Testosterone Impacts Rat Submandibular Gland Proteome.

The salivary glands play a central role in the secretion of saliva, whose composition and volume affect oral and overall health. A lesser-explored dimension encompasses the possible changes in salivary gland proteomes in response to fluctuations in sex hormone levels. This study aimed to examine the effects of chronic exposure to testosterone on salivary gland remodeling, particularly focusing on proteomic adaptations. Therefore, male Wistar rats were implanted with subcutaneous testosterone-releasing devices at 14 weeks of age. Their submandibular glands were histologically and molecularly analyzed 47 weeks later. The results underscored a significant increase in gland mass after testosterone exposure, further supported by histologic evidence of granular duct enlargement. Despite increased circulating sex hormones, there was no detectable shift in the tissue levels of estrogen alpha and androgen receptors. GeLC-MS/MS and subsequent bioinformatics identified 308 proteins in the submandibular glands, 12 of which were modulated by testosterone. Of note was the pronounced upregulation of Klk3 and the downregulation of Klk6 and Klk7 after testosterone exposure. Protein-protein interaction analysis with the androgen receptor suggests that Klk3 is a potential target of androgenic signaling, paralleling previous findings in the prostate. This exploratory analysis sheds light on the response of salivary glands to testosterone exposure, providing proteome-level insights into the associated weight and histological changes.

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.

Validation of the Rat Model of Prostate Cancer: Correlating Seminal Vesicle Lesions With Dorsolateral Prostate Lesions.

BACKGROUND/AIM: Lesions in the seminal vesicle are described in the most used protocols for prostate cancer (PCa) induction. This study aimed to characterize the lesions of seminal vesicles associated with a protocol of PCa induction in rats to contribute to better characterization of this model. MATERIALS AND METHODS: Forty-five male Wistar Unilever rats were randomly divided into two control groups: CONT1 (n=10) and CONT2 (n=10); and two PCa-induced groups: IND1 (n=10) and IND2 (n=15), sacrificed at 35 and 61 weeks, respectively. Animals from the induced groups were exposed to a multistep protocol for PCa induction. Animals, seminal vesicles and dorsolateral prostate were weighed. Seminal vesicles and dorsolateral prostate were submitted to histopathological and immunohistochemical analysis. RESULTS: Animals in which PCa was induced had a lower mean body weight when compared with the control animals (p<0.05). The relative mean seminal vesicle weight was higher in groups with PCa when compared with control groups (p<0.05). Although the differences were not statistically significant, animals from the IND2 group developed more lesions than animals from the IND1 and CONT2 groups. It is worth noting that the animals from group IND2 developed papillary adenomas and carcinomas in situ, which were not observed in any other group. Similar to observations in seminal vesicles, animals from group IND2 developed more dorsolateral prostate lesions than animals from the IND1 group (p<0.05). CONCLUSION: We observed that the longer the exposure to testosterone was, the greater was the incidence of preneoplastic and neoplastic lesions in both the seminal vesicle and the prostate, suggesting that testosterone exposure affects the spectrum of developed lesions.

Effects of testosterone and exercise training on bone microstructure of rats.

Background and Aim: Male hypogonadism results from failure to produce physiological levels of testosterone. Testosterone in men is essential in masculine development, sperm production, and adult man's health. Osteoporosis is one of the consequences of hypogonadism. Regular physical exercise and exogenous testosterone administration are frequently used to prevent or treat this condition. This study aimed to understand the effects of lifelong exercise training and testosterone levels (isolated and together) in the main bone structure parameters. Materials and Methods: A total of 24 rats were used and randomly divided into four groups: Control group (CG; n=6), exercised group (EG, n=6), testosterone group (TG, n=6), and testosterone EG (TEG, n=6). A micro-computed tomography equipment was used to evaluate 15 bone parameters. Results: Both factors (exercise training and testosterone) seem to improve the bone resistance and microstructure, although in different bone characteristics. Testosterone influenced trabecular structure parameters, namely, connectivity density, trabecular number, and trabecular space. The exercise promoted alterations in bone structure as well, although, in most cases, in different bone structure parameters as bone mineral density and medullar mineral density. Conclusion: Overall, exercise and testosterone therapy seems to have a synergistic contribution to the general bone structure and resistance. Further studies are warranted, comparing different individual factors, as gender, lifestyle, or testosterone protocols, to constantly improve the medical management of hypogonadism (and osteoporosis).

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.