Mammary fibromatosis in a female dog: case report.

Fibromatosis, or desmoid tumour, is characterized by excessive and infiltrative proliferation of connective tissue originating from aponeurotic muscle structures. Mammary fibromatosis is rare in humans and animals and its precise aetiology is unknown. A 10-year-old mixed-breed female dog developed a mass in the right cranial thoracic mammary gland (M1) and underwent lumpectomy. The mass was firm, with an irregular surface and distinct limits. Microscopically, it was a neoplastic proliferation of fusiform cells with low atypia, interspersed with abundant dense collagenous tissue, confirmed by histochemical staining with Gomori's trichrome and Masson's trichrome and immunopositivity for vimentin and smooth muscle actin, confirming mammary fibromatosis. Mammary fibromatosis in dogs needs further studies to elucidate its clinical, epidemiological and aetiopathogenic aspects.

Platelet-activating factor and protease-activated receptor 2 cooperate to promote neutrophil recruitment and lung inflammation through nuclear factor-kappa B transactivation.

Although it is well established that platelet-activated receptor (PAF) and protease-activated receptor 2 (PAR2) play a pivotal role in the pathophysiology of lung and airway inflammatory diseases, a role for a PAR2-PAFR cooperation in lung inflammation has not been investigated. Here, we investigated the role of PAR2 in PAF-induced lung inflammation and neutrophil recruitment in lungs of BALB/c mice. Mice were pretreated with the PAR2 antagonist ENMD1068, PAF receptor (PAFR) antagonist WEB2086, or aprotinin prior to intranasal instillation of carbamyl-PAF (C-PAF) or the PAR2 agonist peptide SLIGRL-NH2 (PAR2-AP). Leukocyte infiltration in bronchoalveolar lavage fluid (BALF), C-X-C motif ligand 1 (CXCL)1 and CXCL2 chemokines, myeloperoxidase (MPO), and N-acetyl-glycosaminidase (NAG) levels in BALF, or lung inflammation were evaluated. Intracellular calcium signaling, PAFR/PAR2 physical interaction, and the expression of PAR2 and nuclear factor-kappa B (NF-КB, p65) transcription factor were investigated in RAW 264.7 cells stimulated with C-PAF in the presence or absence of ENMD1068. C-PAF- or PAR2-AP-induced neutrophil recruitment into lungs was inhibited in mice pretreated with ENMD1068 and aprotinin or WEB2086, respectively. PAR2 blockade impaired C-PAF-induced neutrophil rolling and adhesion, lung inflammation, and production of MPO, NAG, CXCL1, and CXCL2 production in lungs of mice. PAFR activation reduced PAR2 expression and physical interaction of PAR2 and PAFR; co-activation is required for PAFR/PAR2 physical interaction. PAR2 blockade impaired C-PAF-induced calcium signal and NF-κB p65 translocation in RAW 264.7 murine macrophages. This study provides the first evidence for a cooperation between PAFR and PAR2 mediating neutrophil recruitment, lung inflammation, and macrophage activation.

Dietary lipids management: are there any benefits from the prevention and treatment of 4T1 murine breast carcinoma?

This study investigated the effect of vegetable and fish oils with different n-3 / n-6 PUFAS ratios on the lipoprotein profile and on the development of murine breast cancer 4T1. Female Balb/c mice (6-7 weeks) received diets containing 4.0% fat during seven weeks. On the fourth week, animals were inoculated into the posterior left flank with 2.5 × 106 4T1 cells. Body weight and food intake were registered and the profile serum lipoproteins was determined. Tumor volume, histopathological and immunohistochemical studies, myeloperoxidase and N-acetylglucosaminidase activities, TNF-α, hemoglobin and VEGF levels were analysed. The highest n-3 / n-6 ratio was found in fish oil (15.8:1), followed by linseed (2.4:1), canola (1:2.1) and soybean (1:9.4) oils. Body weight, food and caloric intake, lipoprotein profile, tumor weight, tumor evolution and histopathological analysis were not different. Canola oil increased cell proliferation when compared to soybean oil, and fish oil changed the inflammatory response and increased VEGF in tumors compared to other groups. The type of fatty acid and the high ratio of n-3 / n-6 PUFAs in the diet influenced cell proliferation and inflammation in the tumor differentially, highlighting the increase of neutrophils and VEGF levels in animals fed on fish oil.

Emetic Tartar-Loaded Liposomes as a New Strategy for Leishmaniasis Treatment.

Emetic tartar (ET), was used in the treatment of leishmaniasis but its use was discontinued due to its low therapeutic index. Liposomes have been shown to be a promising strategy for delivery of bioactive substances in the region of interest, in order to reduce and/or eliminate undesirable effects. In the present study, liposomes containing ET were prepared and characterized to evaluate acute toxicity as well as their leishmanicidal action using BALB/c mice with an inoculum of Leishmania (Leishmania) infantum. Liposomes were composed of egg phosphatidylcholine and 3ß-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol, with an average diameter of 200 nm, zeta potential of +18 mV, and ET encapsulated into liposomes at a concentration near 2 g/L. Healthy mice were treated with ET or liposome containing ET (Lip-ET) in a single dose of 16 mg/kg of Sb3+ intravenously and observed for 14 days. The death of two animals in the ET-treated group and no deaths in the Lip-ET-treated group was observed. Higher hepatic and cardiac toxicity were observed in animals treated with ET when compared to animals treated with Lip-ET, blank liposomes (Blank-Lip) and PBS. The study of antileishmanial efficacy was conducted by intraperitoneal administration of Lip-ET, for ten consecutive days. It was observed by limiting dilution that treatments with liposomal formulations containing ET, as well as Glucantime®, led to a significant reduction in parasitic load in spleen and liver (p < 0.05) when compared to the untreated control group.

High SARS-CoV-2 tropism and activation of immune cells in the testes of non-vaccinated deceased COVID-19 patients.

BACKGROUND: Cellular entry of SARS-CoV-2 has been shown to rely on angiotensin-converting enzyme 2 (ACE2) receptors, whose expression in the testis is among the highest in the body. Additionally, the risk of mortality seems higher among male COVID-19 patients, and though much has been published since the first cases of COVID-19, there remain unanswered questions regarding SARS-CoV-2 impact on testes and potential consequences for reproductive health. We investigated testicular alterations in non-vaccinated deceased COVID-19-patients, the precise location of the virus, its replicative activity, and the immune, vascular, and molecular fluctuations involved in the pathogenesis. RESULTS: We found that SARS-CoV-2 testicular tropism is higher than previously thought and that reliable viral detection in the testis requires sensitive nanosensors or RT-qPCR using a specific methodology. Through an in vitro experiment exposing VERO cells to testicular macerates, we observed viral content in all samples, and the subgenomic RNA's presence reinforced the replicative activity of SARS-CoV-2 in testes of the severe COVID-19 patients. The cellular structures and viral particles, observed by transmission electron microscopy, indicated that macrophages and spermatogonial cells are the main SARS-CoV-2 lodging sites, where new virions form inside the endoplasmic reticulum Golgi intermediate complex. Moreover, we showed infiltrative infected monocytes migrating into the testicular parenchyma. SARS-CoV-2 maintains its replicative and infective abilities long after the patient's infection. Further, we demonstrated high levels of angiotensin II and activated immune cells in the testes of deceased patients. The infected testes show thickening of the tunica propria, germ cell apoptosis, Sertoli cell barrier loss, evident hemorrhage, angiogenesis, Leydig cell inhibition, inflammation, and fibrosis. CONCLUSIONS: Our findings indicate that high angiotensin II levels and activation of mast cells and macrophages may be critical for testicular pathogenesis. Importantly, our findings suggest that patients who become critically ill may exhibit severe alterations and harbor the active virus in the testes.

Bifidobacterium longum subsp. longum 51A Attenuates Signs of Inflammation in a Murine Model of Food Allergy.

Food allergy is a pathological condition that can lead to hives, swelling, gastrointestinal distress, cardiovascular and respiratory compromise, and even anaphylaxis. The lack of treatment resources emphasizes the necessity for new therapeutic strategies, and in this way, probiotics has been pointed out as an alternative, especially because of its immunomodulatory properties. The goal of this study was to evaluate the probiotic effect of Bifidobacterium longum subsp. longum 51A (BL51A) in a murine model of ovalbumin (OVA) food allergy, as well as to investigate the effect of the dose and viability of the bacteria on the proposed model. For this purpose, the probiotic effect was assessed by clinical, immunological, and histological parameters in mice treated or not with the BL51A and sensitized or not with OVA. Oral administration of BL51A prevented weight loss and reduced serum levels of IgE anti-OVA and of sIgA in the intestinal fluid. Also, it reduced the intestinal permeability, proximal jejunum damage, recruitment of eosinophils and neutrophils, and levels of eotaxin-1, CXCL1/KC, IL4, IL5, IL6, IL13, and TNF. Furthermore, the treatment was able to increase the levels of IL10. Investigating different doses administered, the level of 108 CFU showed the best results in terms of protective effect. In addition, the administration of the inactivated bacteria did not present any beneficial effect. Results demonstrate that BL51A promotes a systemic immunomodulatory protective effect in a murine model of food allergy that depends on the dose and viability of the bacteria, suggesting its use as probiotic in such disease.

Acute toxicity and antitumor potential of 1,3,4-trisubstituted-1,2,3-triazole dhmtAc-loaded liposomes on a triple-negative breast cancer model.

For the first time, compounds developed from the 1,2,3-triazole scaffold were evaluated as novel drugs to treat triple-negative breast cancer (TNBC). Four organic salts were idealized as nonclassical bioisosteres of miltefosine, which is used in the topical treatment for skin metastasizing breast carcinoma. Among them, derivative dhmtAc displayed better solubility and higher cytotoxicity against the human breast adenocarcinoma cell line and mouse 4T1 cell lines, which are representatives of TNBC. In vitro assays revealed that dhmtAc interferes with cell integrity, confirmed by lactate dehydogenase leakage. Due to its human peripheral blood mononuclear cell (PBMC) toxicity, dhmtAc in vivo studies were carried out with the drug incorporated in a long-circulating and pH-sensitive liposome (SpHL-dhmtAc), and the acute toxicity in BALB/c mice was determined. Free dhmtAc displayed cardiac and pulmonary toxicity after the systemic administration of 5 mg/kg doses. On the other hand, SpHL-dhmtAc displayed no toxicity at 20 mg/kg. The in vivo antitumor effect of SpHL-dhmtAc was investigated using the 4T1 heterotopic murine model. Intravenous administration of SpHL-dhmtAc reduced the tumor volume and weight, without interfering with the body weight, compared with the control group and the dhmtAc free form. The incorporation of the triazole compound in the liposome allowed the demonstration of its anticancer potential. These findings evidenced 1,3,4-trisubstituted-1,2,3-triazole as a promising scaffold for the development of novel drugs with applicability for the treatment of patients with TNBC.

Solid Carcinoma of the Canine Mammary Gland: a Histological Type or Tumour Cell Arrangement?

Mammary neoplasms are the most frequently diagnosed tumours in female dogs and are classified into various histological types, including solid carcinomas. We proposed a subclassification of solid carcinomas based on morphological and immunohistochemical characteristics, and correlated the subtypes with prognostic factors. A total of 135 cases of solid mammary carcinoma were selected from 3,400 canine mammary neoplasms in the archives of the Laboratory of Comparative Pathology, Institute of Biological Sciences, Federal University of Minas Gerais, Brazil. Epidemiological and survival data were obtained, and immunolabelling for chromogranin A, pancytokeratin, cytokeratin 14, Ki67 and p63 was performed. Solid carcinomas were classified into six subgroups: malignant adenomyoepithelioma (68/135), carcinoma with solid pattern (22/135), malignant myoepithelioma (16/135), basaloid carcinoma (14/135), neuroendocrine carcinoma (10/135) and solid papillary carcinoma (5/135). Shorter survival time was associated with the presence of lymphatic invasion (P = 0.009) in the initial clinical staging (I-III). When considering all clinical stages (I-V), vascular invasion (P <0.001) and the presence of regional metastasis (P = 0.004) were important prognostic factors. Basaloid carcinoma and solid papillary carcinoma did not reach the median survival time for early-stage cases, and malignant myoepithelioma had the highest median survival in advanced stages. Carcinoma with a solid pattern was associated with a higher number of regional metastases. Distinguishing the various histological and immunophenotypic subtypes that exhibit a solid arrangement, using histological and immunohistochemical criteria, is essential for understanding the behaviour of these neoplasms and for the selection of more appropriate and specific therapies.

Inositol 1,4,5-trisphosphate receptor type 3 (ITPR3) is overexpressed in cholangiocarcinoma and its expression correlates with S100 calcium-binding protein A4 (S100A4).

Cholangiocarcinoma (CCA) is the second most malignant neoplasm in the liver that arises from the biliary tree. CCA is associated with a poor prognosis, and the key players involved in its pathogenesis are still not well understood. Receptor tyrosine kinases (RTKs), such as epidermal growth factor receptor (EGFR), can mediate intracellular calcium (Ca2+) signaling pathways via inositol 1,4,5-trisphosphate (InsP3), activating inositol 1,4,5-trisphosphate receptors (ITPRs) and regulating tumor growth. ITPR isoform 3 (ITPR3) is the main intracellular Ca2+ release channel in cholangiocytes. The effects of intracellular Ca2+ are mediated by calcium-binding proteins such as Calmodulin and S100 calcium-binding protein A4 (S100A4). However, the clinicopathological and biological significance of EGFR, ITPR3 and S100A4 in CCA remains unclear. Thus, the present work investigates the immunoexpression of these three proteins in 59 CCAs from patients who underwent curative surgical treatment and correlates the data with clinicopathological features and survival. High ITPR3 expression was correlated with CA 19-9 levels, TNM stage and lymph node metastasis (N). Furthermore, ITPR3 expression was increased in distal CCA compared to control bile ducts and intrahepatic and perihilar CCAs. These observations were confirmed by proteomic analysis. ITPR3 and S100A4 clinical scores were significantly correlated. Furthermore, it was demonstrated that EGF induces calcium signaling in a cholangiocarcinoma cell line and ITPR3 colocalizes with nonmuscle myosin IIA (NMIIA). In summary, ITPR3 overexpression could contribute to CCA progression and it may represent a potential therapeutic target.

C(3)1-TAg in C57BL/6 J background as a model to study mammary tumor development.

Diagnosis and prognosis of breast cancer is based on disease staging identified through histopathological and molecular biology techniques. Animal models are used to gain mechanistic insights into the development of breast cancer. C(3)1-TAg is a genetically engineered mouse model that develops mammary cancer. However, carcinogenesis caused by this transgene was characterized in the Friend Virus B (FVB) background. As most genetic studies are done in mice with C57BL/6 J background, we aimed to define the histological alterations in C3(1)-TAg C57BL/6 J animals. Our results showed that C3(1)-TAg animals with C57BL/6 J background develop solid-basaloid adenoid cystic carcinomas with increased fibrosis, decreased area of adipocytes, and a high proliferative index, which are triple-negative for progesterone, estrogen, and human epidermal growth factor receptor 2 (HER2) receptors. Our results also revealed that tumor development is slower in the C57BL/6 J background when compared with the FVB strain, providing a better model to study the different stages in breast cancer progression.

Characterization of neoplastic cells outlining the cystic space of invasive micropapillary carcinoma of the canine mammary gland.

BACKGROUND: Invasive micropapillary carcinoma (IMPC) is a rare malignant breast tumor and a variant form of invasive ductal carcinoma that is an aggressive neoplasm of the human breast and canine mammary gland. The importance of the tumor microenvironment in cancer development has gradually been recognized, but little is known about the cell types outlining the cystic space of canine IMPC. This study aimed to characterize the neoplastic cells outlining the cystic space of IMPC. RESULTS: Immunohistochemistry (IHC), immunofluorescence (IF), superresolution and transmission electron microscopy (TEM) were used to assess the cell types in the cystic areas of IMPCs. Cells expressing the mesenchymal markers alpha-smooth muscle actin (αSMA), Vimentin, and S100A4 outlined the cystic space of IMPC. Furthermore, loss of epithelial cell polarity in IMPC was shown by the localization of MUC1 at the stroma-facing surface. This protein modulates lumen formation and inhibits the cell-stroma interaction. Immunohistochemical and IF staining for the myoepithelial cell marker p63 were negative in IMPC samples. Furthermore, associated with peculiar morphology, such as thin cytoplasmic extensions outlining cystic spaces, was observed under TEM. These observations suggested cells with characteristics of myoepithelial-like cells. CONCLUSIONS: The cells outlining the cystic space of IMPC in the canine mammary gland were characterized using IHC, IF and TEM. The presence of cells expressing αSMA, Vimentin, and S100A4 in the IMPC stroma suggested a role for tumor-associated fibroblasts in the IMPC microenvironment. The reversal of cell polarity revealed by the limited basal localization of MUC1 may be an important factor contributing to the invasiveness of IMPC. For the first time, the cystic space of canine mammary gland IMPC was shown to be delimited by myoepithelial-like cells that had lost p63 expression. These findings may enhance our understanding of the cellular microenvironment of invasive tumors to improve cancer diagnosis and treatment.

Canine mammary cancer tumour behaviour and patient survival time are associated with collagen fibre characteristics.

Precise diagnosis and prognosis are key in prevention and reduction of morbidity and mortality in all types of cancers. Here we show that changes in the collagen fibres in the main histological subtypes of canine mammary gland carcinomas are directly associated with the tumour behaviour and the animal survival time and could become a useful tool in helping with diagnosis. Imaging by second harmonic generation and multiphoton excited fluorescence microscopy were performed to evaluate the collagen and cellular segment parameters in cancer biopsies. We present a retrospective study of 45 cases of canine mammary cancer analysing 836 biopsies regions including normal mammary gland tissue, benign mixed tumours, carcinoma in mixed tumour, carcinosarcoma, micropapillary carcinoma and solid carcinoma. The image analyses and the comparison between the tumour types allowed to assess the collagen fibre changes during tumour progression. We demonstrate that the collagen parameters correlate with the clinical and pathological data, the results show that in neoplastic tissues, the collagen fibres are more aligned and shorter as compared to the normal tissues. There is a clear association of the mean fibre length with the dogs survival times, the carcinomas presenting shorter collagen fibres indicate a worse survival rate.

NLRP6-associated host microbiota composition impacts in the intestinal barrier to systemic dissemination of Brucella abortus.

Brucella abortus is a Gram-negative bacterium responsible for a worldwide zoonotic infection-Brucellosis, which has been associated with high morbidity rate in humans and severe economic losses in infected livestock. The natural route of infection is through oral and nasal mucosa but the invasion process through host gut mucosa is yet to be understood. Studies have examined the role of NLRP6 (NOD-like receptor family pyrin domain-containing-6 protein) in gut homeostasis and defense against pathogens. Here, we investigated the impact of gut microbiota and NLRP6 in a murine model of Ba oral infection. Nlrp6-/- and wild-type (WT) mice were infected by oral gavage with Ba and tissues samples were collected at different time points. Our results suggest that Ba oral infection leads to significant alterations in gut microbiota. Moreover, Nlrp6-/- mice were more resistant to infection, with decreased CFU in the liver and reduction in gut permeability when compared to the control group. Fecal microbiota transplantation from WT and Nlrp6-/- into germ-free mice reflected the gut permeability phenotype from the donors. Additionally, depletion of gut microbiota by broad-spectrum-antibiotic treatment prevented Ba replication in WT while favoring bacterial growth in Nlrp6-/-. Finally, we observed higher eosinophils in the gut and leukocytes in the blood of infected Nlrp6-/- compared to WT-infected mice, which might be associated to the Nlrp6-/- resistance phenotype. Altogether, these results indicated that gut microbiota composition is the major factor involved in the initial stages of pathogen host replication and partially also by the resistance phenotype observed in Nlrp6 -/- mice regulating host inflammation against Ba infection.

Relevance of angiotensin-(1-7) and its receptor Mas in pneumonia caused by influenza virus and post-influenza pneumococcal infection.

Resolution failure of exacerbated inflammation triggered by Influenza A virus (IAV) prevents return of pulmonary homeostasis and survival, especially when associated with secondary pneumococcal infection. Therapeutic strategies based on pro-resolving molecules have great potential against acute inflammatory diseases. Angiotensin-(1-7) [Ang-(1-7)] is a pro-resolving mediator that acts on its Mas receptor (MasR) to promote resolution of inflammation. We investigated the effects of Ang-(1-7) and the role of MasR in the context of primary IAV infection and secondary pneumococcal infection and evaluated pulmonary inflammation, virus titers and bacteria counts, and pulmonary damage. Therapeutic treatment with Ang-(1-7) decreased neutrophil recruitment, lung injury, viral load and morbidity after a primary IAV infection. Ang-(1-7) induced apoptosis of neutrophils and efferocytosis of these cells by alveolar macrophages, but had no direct effect on IAV replication in vitro. MasR-deficient (MasR-/-) mice were highly susceptible to IAV infection, displaying uncontrolled inflammation, increased viral load and greater lethality rate, as compared to WT animals. Ang-(1-7) was not protective in MasR-/- mice. Interestingly, Ang-(1-7) given during a sublethal dose of IAV infection greatly reduced morbidity associated with a subsequent S. pneumoniae infection, as seen by decrease in the magnitude of neutrophil influx, number of bacteria in the blood leading to a lower lethality. Altogether, these results show that Ang-(1-7) is highly protective against severe primary IAV infection and protects against secondary bacterial infection of the lung. These effects are MasR-dependent. Mediators of resolution of inflammation, such as Ang-(1-7), should be considered for the treatment of pulmonary viral infections.

Hormone receptor expression, clinical and histopathological analysis in feline injection site sarcomas.

Feline injection site sarcomas (FISS) are aggressive, with high recurrence and rarely metastasising. The objective of this study was to evaluate, by immunohistochemistry, the expression of oestrogen (ER) and progesterone (PR) receptors in FISS and correlate them with clinical and histopathological aspects. This was a retrospective study with 51 cases of FISS. Immunohistochemistry was performed to detect vimentin, ER, PR and Ki67 expression. Clinical, histopathological and immunohistochemical characteristics were predictor variables and the expression of ER and PR were the dependent ones. Twenty-eight (55%) of the 51 FISS cases were female and 23 (45%) male with 10.7 ± 4.2 years and median tumour size of 3 cm (2.0-5.4). The trunk was the most affected site, with 38 cases (84%). Histological grade III was observed in 57% of the cases, considering differentiation score, necrosis and mitotic index. ER expression, positive in 64% of cases, was associated with the mitotic index (P = .05) and degree of pleomorphism (P = .04). PR was not associated with the variables and 63% of cases were negative for this receptor. Thus, ER expression can affect tumour growth. The knowledge on the FISS hormonal expression is important to clarify the pathophysiological mechanisms. Further studies are needed to predict the value of ER expression in the prognosis of FISS.

Analytical methods for assessing changes induced by gamma exposure in an animal model.

OBJECTIVE: Ionizing radiation can cause radio-induced changes in the cellular metabolome due to the breakdown of DNA bonds. Our goal was to find the early tissue response to radiation exposure supported by distinct analytical methods. METHODS: Histological analyses were performed on the organs extracted from rats to search for microscopic changes. The histological slides stained with hematoxyline-eosin (HE) were analyzed in magnification (40x). Subsequently, the tissues were subjected to mass spectrometry that allowed molecular analysis and DESI-MSI that generated the molecular image of lipids, assessing changes in intensities, especially in the brain. RESULTS: The histological analysis found nonspecific inflammatory changes; no areas of fibrosis, necrosis, or apoptosis were identified, suggesting non-morphological tissue alterations. However, the DESI-MSI images of brain lipids allowed the observation of many radio-induced changes in the lipid's intensities. CONCLUSIONS: No early radio induced histological or mass weight changes in the radiation exposed rats could be observed at 5 Gy. However, early changes in the molecular level were observed in the DESI-MSI images of the brain lipids. The DESI-MSI method proved to be efficient and relevant, allowing a regional molecular analysis of the tissues, expanding a new field of study that is still in its infancy: radiometabolomics.

Canine mammary cancer diagnosis from quantitative properties of nonlinear optical images.

We present nonlinear microscopy imaging results and analysis from canine mammary cancer biopsies. Second harmonic generation imaging allows information of the collagen structure in the extracellular matrix that together with the fluorescence of the cell regions of the biopsies form a base for comprehensive image analysis. We demonstrate an automated image analysis method to classify the histological type of canine mammary cancer using a range of parameters extracted from the images. The software developed for image processing and analysis allows for the extraction of the collagen fibre network and the cell regions of the images. Thus, the tissue properties are obtained after the segmentation of the image and the metrics are measured specifically for the collagen and the cell regions. A linear discriminant analysis including all the extracted metrics allowed to clearly separate between the healthy and cancerous tissue with a 91%-accuracy. Also, a 61%-accuracy was achieved for a comparison of healthy and three histological cancer subtypes studied.

Analytical methods for assessing changes induced by gamma exposure in an animal model

SUMMARY OBJECTIVE: Ionizing radiation can cause radio-induced changes in the cellular metabolome due to the breakdown of DNA bonds. Our goal was to find the early tissue response to radiation exposure supported by distinct analytical methods. METHODS: Histological analyses were performed on the organs extracted from rats to search for microscopic changes. The histological slides stained with hematoxyline-eosin (HE) were analyzed in magnification (40x). Subsequently, the tissues were subjected to mass spectrometry that allowed molecular analysis and DESI-MSI that generated the molecular image of lipids, assessing changes in intensities, especially in the brain. RESULTS: The histological analysis found nonspecific inflammatory changes; no areas of fibrosis, necrosis, or apoptosis were identified, suggesting non-morphological tissue alterations. However, the DESI-MSI images of brain lipids allowed the observation of many radio-induced changes in the lipid's intensities. CONCLUSIONS: No early radio induced histological or mass weight changes in the radiation exposed rats could be observed at 5 Gy. However, early changes in the molecular level were observed in the DESI-MSI images of the brain lipids. The DESI-MSI method proved to be efficient and relevant, allowing a regional molecular analysis of the tissues, expanding a new field of study that is still in its infancy: radiometabolomics.

RESUMO OBJETIVO: Radiação ionizante pode causar alterações no metaboloma celular devido à quebra de ligações no DNA. O objetivo deste trabalho foi evidenciar a resposta aguda tecidual induzida pela exposição da radiação ionizante. MÉTODOS: Análises histológicas foram realizadas nos órgãos extraídos de ratos para análise de alterações microscópicas. As lâminas histológicas coradas com hematoxilina eosina (HE) foram analisadas em aumento (40x). Posteriormente, os tecidos foram submetidos a espectrometria de massa, que permitiu análise molecular e o Desi-MSI que gerou imagem molecular de lipídios, identificando alterações na intensidade, principalmente no cérebro. RESULTADOS: As análises histológicas encontraram alterações inflamatórias inespecíficas, nem áreas de fibrose, necrose ou apoptose, sugerindo ausência de alterações morfológicas. As imagens de lipídios cerebrais obtidas por Desi-MSI permitiram observar as inúmeras alterações na intensidade nas seções teciduais do encéfalo. CONCLUSÕES: Alterações agudas radioinduzidas de massa do órgão e histológicas nos órgãos dos ratos expostos não puderam ser observadas a 5 Gy. Entretanto, mudanças em nível molecular foram observadas nas imagens de Desi-MSI dos lipídios cerebrais. O método Desi-MSI mostrou-se eficiente e relevante, permitindo a análise molecular regi-onal dos tecidos no SNC, expandindo um novo campo de estudo que ainda está em sua infância: a radiometaboloma.

Localization of angiotensin-(1-7) and Mas receptor in the rat ovary throughout the estrous cycle.

We have previously demonstrated the presence of Angiotensin (Ang)-(1-7) in rat ovary homogenates and its stimulatory effect on estradiol and progesterone production. The present study was undertaken to identify the cellular localization of Ang-(1-7) and its receptor Mas in the rat ovary in the different phases of the estrous cycle. Ang-(1-7) and Mas were localized by immunohistochemistry and Mas mRNA expression was assessed by RT-PCR. Immunostaining for both Ang-(1-7) and Mas was found in all phases of the estrous cycle, particularly in the thecal and interstitial cells, as well as in regressing corpora lutea. However, granulosa cells were positive only in antral and preovulatory follicles at proestrus and estrus phases. This pattern contrasted with the distribution of the octapeptide Ang II, which was abundant in granulosa but not in theca cells. In addition, the expression of Mas mRNA was demonstrated in all estrous cycle phases. Angiotensin-converting enzyme activity did not vary between estrous cycle phases, whereas prolyl endopeptidase activity was significantly higher in diestrus and neutral endopeptidase activity was significantly higher in metestrus. These data provide the first evidence that new RAS components are dynamically expressed in the ovary across the rat estrous cycle. Further functional studies should clarify the role of Ang-(1-7) signaling through Mas receptor in the regulation of ovarian physiology.

The Role of ST2 Receptor in the Regulation of Brucella abortus Oral Infection.

The ST2 receptor plays an important role in the gut such as permeability regulation, epithelium regeneration, and promoting intestinal immune modulation. Here, we studied the role of ST2 receptor in a murine model of oral infection with Brucella abortus, its influence on gut homeostasis and control of bacterial replication. Balb/c (wild-type, WT) and ST2 deficient mice (ST2-/-) were infected by oral gavage and the results were obtained at 3 and 14 days post infection (dpi). Our results suggest that ST2-/- are more resistant to B. abortus infection, as a lower bacterial colony-forming unit (CFU) was detected in the livers and spleens of knockout mice, when compared to WT. Additionally, we observed an increase in intestinal permeability in WT-infected mice, compared to ST2-/- animals. Breakage of the intestinal epithelial barrier and bacterial dissemination might be associated with the presence of the ST2 receptor; since, in the knockout mice no change in intestinal permeability was observed after infection. Together with enhanced resistance to infection, ST2-/- produced greater levels of IFN-γ and TNF-α in the small intestine, compared to WT mice. Nevertheless, in the systemic model of infection ST2 plays no role in controlling Brucella replication in vivo. Our results suggest that the ST2 receptor is involved in the invasion process of B. abortus by the mucosa in the oral infection model.