Bruno Cogliati

University of São Paulo, San Paulo, São Paulo, Brazil

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Publications (40)57.06 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Hepatotoxicity, including drug-induced liver injury, is frequently accompanied by cell death. The latter is typically driven by apoptosis or necrosis, which substantially differ based upon biochemical and morpho- logical criteria. This chapter describes two commonly used methods to probe apoptotic and necrotic activi- ties in adherent monolayer cultures of primary hepatocytes. The apoptosis assay uses a prototypical substrate of caspase 3, the main executor of apoptotic cell death, which can be cleaved, yielding a product that can be measured fl uorimetrically. The second assay relies on the disruption of the cell plasma mem- brane, which typically occurs in necrotic cell death and that results in the extracellular release of cytoplas- mic enzymes, such as lactate dehydrogenase. The latter can be indirectly assessed by spectrophotometrically measuring the consumption of reduced nicotinamide adenine dinucleotide.
    Protocols in In Vitro Hepatocyte Research, 2015 edited by Mathieu Vinken, Vera Rogiers, 01/2015: chapter Measurement of Apoptotic and Necrotic Cell Death in Primary Hepatocyte Cultures: pages 349-362; Springer Protocols-Humana Press., ISBN: 978-1-4939-2073-0
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    ABSTRACT: Connexins are proteins that form gap junctions. Perturbations in the cell membrane reportedly promote changes in the expression profile of connexins. Electroporation promotes destabilization by applying electrical pulses, and this procedure is used in electrochemotherapy and gene therapy, among others. This in vitro work aimed to study the interference of electroporation on the expression profile of GJB2 (Cx26 gene) and Connexin 26 in melanoma cell line B16/BL6. The techniques of immunocytochemistry, Western blot, and real-time PCR were used. After electroporation, cells showed a transient decrease in GJB2 mRNA. The immunostaining of Cx26 showed no noticeable change after electroporation at different time points. However, Western blot showed a significant reduction in Cx26 30 min after electroporation. Our results showed that electroporation interferes transiently in the expression of Connexin 26 in melanoma and are consistent with the idea that electroporation is a process of intense stress that promotes cell homeostatic imbalance and results in disruption of cell physiological processes such as transcription and translation.
    Journal of Membrane Biology 10/2014; · 2.48 Impact Factor
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    ABSTRACT: Very few studies have evaluated the expression of homeobox A10 (HOXA10) and steroid (estrogen and progesterone) receptors exclusively in deep endometriosis. Conclusions drawn from studies evaluating peritoneal and ovarian endometriosis are usually generalized to explain the pathogenesis of the disease as a whole. We aimed to evaluate the expression of HOXA10, estrogen receptor α (ER-α), progesterone receptor (PR), and PR-B in rectosigmoid endometriosis (RE), a typical model of deep disease.
    Reproductive sciences (Thousand Oaks, Calif.) 09/2014; · 2.31 Impact Factor
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    ABSTRACT: Background: Melanoma is one of the most common skin neoplasms in humans and dogs. The tumor microenvironment in melanoma comprises cancer cells and stromal cells that interact to accelerate tumor progression. Several prognostic markers for melanomas have been studied in many human tumors, including fibroblast-specific protein 1 (S100A4). S100A4 is a member of the S100 family of calcium-binding proteins in stromal cells.Hypothesis/objectives: The objective of this study was to describe the immunohistochemical patterns of S100A4 in stroma and neoplastic cells of canine skin melanomas and correlate them with some histological parameters. Animals and Methods: Forty-eight samples (38 pigmented and 10 non-pigmented melanomas) were first selected and their nature confirmed using S100, Melan A and vimentin. All cases were examined by immunohistochemistry using S100A4 to correlate expression, histotype, and level of invasion.Results: All the tumors, including 10 non-pigmented, were positive for S100, Melan A, vimentin and negative for cytokeratin AE1/AE3 (consistent with melanomas). The 48 melanomas were classified as epithelioid (n = 21), spindle (n = 14), and mixed (n = 13). S100A4 was preferentially expressed in epithelioid and spindle cell types compared with mixed melanomas and S100A4 expression was not associated with level of invasion (Clark's levels IV to V).Conclusion: S100A4 expression in melanoma samples varied among histotypes but not between levels of invasion.
    The Veterinary quarterly. 07/2014;
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    ABSTRACT: Rationale: The excessive intake of vitamin A in the form of vitamin concentrate, supplement or vitamin-rich liver can result in hypervitaminosis A in man and animals. Although osteopathologies resulting from chronic vitamin A intoxication in cats are well characterized, no information is available concerning feline hypervitaminosis A-induced liver disease. Clinical summary: We report the first case of hepatic stellate cell lipidosis and hepatic fibrosis in a domestic cat that had been fed a diet based on raw beef liver. Radiographic examination revealed exostoses and ankylosis between vertebrae C1 and T7, compatible with deforming cervical spondylosis. Necropsy showed a slightly enlarged and light yellow to bronze liver. Microscopic and ultrastructural analyses of liver tissues revealed diffuse and severe liver fibrosis associated with hepatic stellate cell hyperplasia and hypertrophy. These cells showed immunopositive staining for α-smooth muscle actin and desmin markers. The necropsy findings of chronic liver disease coupled with osteopathology supported the diagnosis of hypervitaminosis A. Practical relevance: As in human hepatology, if there is dietary evidence to support increased intake of vitamin A, then hypervitaminosis A should be considered in the differential diagnosis of chronic liver disease in cats.
    Journal of feline medicine and surgery. 03/2014; 16(3):243-8.
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    ABSTRACT: Hair follicle tumours generally present as benign, solitary masses and have a good prognosis following surgical resection. This report describes a case of multiple trichoblastomas in a dog. A 2-year-old crossbred dog presented with multiple soft cutaneous periocular, perilabial, submandibular and nasal nodules, between 2 and 9 cm in diameter, located on the right side of the face. New nodules were observed on the same side of the face at a second consultation 3 weeks later. Surgical resection of all nodules was performed in two procedures. Three nodules were initially resected and submitted for histolopathology and immunohistochemistry. The diagnosis was trichoblastoma for all three. At the time of the second consultation, new and remaining nodules were biopsied and the diagnosis of trichoblastoma confirmed. The dog was treated with doxorubicin and piroxicam for 30 days prior to the second surgical procedure in an attempt to reduce new tumour growth and the size of present tumours. All nodules were resected and the defects closed using rotation flaps. No recurrence of the neoplasm was noted within 10 months after surgery. Trichoblastomas are generally benign but can present as multiple neoplasms that may require surgical resection and may respond to chemotherapy. To the authors' knowledge, this is the first report of multiple trichoblastomas in a dog.
    Veterinary Dermatology 02/2014; 25(1):48-e19. · 2.02 Impact Factor
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    ABSTRACT: The liver was among the first organs in which connexin proteins have been identified. Hepatocytes harbor connexin32 and connexin26, while non-parenchymal liver cells typically express connexin43. Connexins give rise to hemichannels, which dock with counterparts on adjacent cells to form gap junctions. Both hemichannels and gap junctions provide pathways for communication, via paracrine signaling or direct intercellular coupling, respectively. Over the years, hepatocellular gap junctions have been shown to regulate a number of liver-specific functions and to drive liver cell growth. In the last few years, it has become clear that connexin hemichannels are involved in liver cell death, particularly in hepatocyte apoptosis. This also holds true for hemichannels composed of pannexin1, a connexin-like protein recently identified in the liver. Moreover, pannexin1 hemichannels are key players in the regulation of hepatic inflammatory processes. The current paper provides a concise overview of the features of connexins, pannexins and their channels in the liver.
    Frontiers in Physiology 01/2014; 4:405.
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    ABSTRACT: Melanoma is a malignant neoplasm occurring in several animal species, and is the most frequently found tumor in the oral cavity in dogs. Melanomas are classified into two types: melanotic and amelanotic. Prior research suggests that human amelanotic melanomas are more aggressive than their melanotic counterparts. This study evaluates the behavior of canine melanotic and amelanotic oral cavity melanomas and quantifies cell proliferation and the expression of connexins. Twenty-five melanomas (16 melanotic and 9 amelanotic) were collected from dogs during clinical procedures at the Veterinary Hospital of the School of Veterinary Medicine and Animal Science of the University of São Paulo, Brazil. After diagnosis, dogs were followed until death or euthanasia. Histopathology confirmed the gross melanotic or amelanotic characteristics and tumors were classified according to the WHO. HMB45 or Melan A immunostainings were performed to confirm the diagnosis of amelanotic melanomas. Cell proliferation was quantified both by counting mitotic figures and PCNA positive nuclei. Expressions of connexins 26 and 43 were evaluated by immunohistochemistry, qRT-PCR and Western blot. Dogs bearing amelanotic melanomas presented a shorter lifespan in comparison to those with melanotic melanomas. Cell proliferation was significantly higher in amelanotic melanomas. Expressions of Connexins 26 and 43 were significantly reduced in amelanotic melanomas. The results presented here suggest that oral cavity melanotic and amelanotic melanomas differ regarding their behavior, cell proliferation and connexin expression in dogs, indicating a higher aggressiveness of amelanotic variants.
    Veterinary Research Communications 10/2013; · 1.08 Impact Factor
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    ABSTRACT: Apoptosis not only plays a key role in physiological demise of defunct hepatocytes, but is also associated with a plethora of acute and chronic liver diseases as well as with hepatotoxicity. The present paper focuses on the modelling of this mode of programmed cell death in primary hepatocyte cultures. Particular attention is paid to the activation of spontaneous apoptosis during the isolation of hepatocytes from the liver, its progressive manifestation upon the subsequent establishment of cell cultures and simultaneously to strategies to counteract this deleterious process. In addition, currently applied approaches to experimentally induce controlled apoptosis in this in vitro setting for mechanistic research purposes and thereby its detection using relevant biomarkers are reviewed.
    Archives of Toxicology 09/2013; · 5.22 Impact Factor
  • Journal of Feline Medicine & Surgery 09/2013; 15(9):818-28. · 1.08 Impact Factor
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    ABSTRACT: Background/objective The early detection of focal hepatic lesions using ultrasound scanning is challenging, and this challenge becomes even greater in the presence of diffuse parenchymal disease. This study aimed to evaluate the diagnostic performance of elastography and contrast-enhanced ultrasonography (CEUS) in the early detection of hepatocellular lesions in an experimental rat model of nonalcoholic steatohepatitis (NASH). Methods B-mode and Doppler ultrasonography was performed weekly in 30 rats divided into a NASH group (n = 20) and a group without liver disease (n = 10). The animals underwent elastography and CEUS and were then euthanized. Liver nodules were assessed by histopathology. Results Doppler mapping results of lesions with vascularization were considered indicative of malignancy, with a sensitivity of 29% before and 71% after contrast injection. The specificity was 71% before and 96% after CEUS. Elastograms of positive lesions showed areas of high stiffness, which were indicative of malignancy. This malignancy was confirmed by the histologic evaluation, with a sensitivity of 90% and a specificity of 60%. After CEUS analysis, 4 nodules were identified that were not observed on B-mode ultrasonography. Early wash-in was significantly associated with malignancy (sensitivity of 88% and specificity of 67%). Conclusions Both techniques allow for the correct diagnosis of well-differentiated to moderately differentiated hepatocellular carcinomas with good accuracy in an experimental rat model of NASH.
    Journal of Clinical and Experimental Hepatology. 06/2013; 3(2):96–101.
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    ABSTRACT: The liver plays a vital role in the organism, and thousands of patients suffer and even die from hepatic complications every year. Viral hepatitis is one of the most important causes of liver-related pathological processes. However, sterile liver diseases, such as drug-induced liver injury, cirrhosis and fibrosis, are still a worldwide concern and contribute significantly to liver transplantation statistics. During hepatocyte death, several genuine intracellular contents are released to the interstitium, where they will trigger inflammatory responses that may boost organ injury. Intracellular purines are key molecules to several metabolic pathways and regulate cell bioenergetics. However, seminal studies in early 70s revealed that purines may also participate in cell-to-cell communication, and more recent data have unequivocally demonstrated that the purinergic signalling plays a key role in the recognition of cell functionality by neighbouring cells and also by the immune system. This new body of knowledge has pointed out that several promising therapeutic opportunities may rely on the modulation of purine release and sensing during diseases. Here, we review the most recent data on the physiological roles of purinergic signalling and how its imbalance may contribute to injury progression during sterile liver injury.
    Liver international: official journal of the International Association for the Study of the Liver 03/2013; 33(3):353-61. · 3.87 Impact Factor
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    ABSTRACT: This study investigated the correlation between KIT gene expression determined by immunohistochemistry and real-time polymerase chain reaction (RT-PCR) and the rate of tumour recurrence and tumour-related deaths in dogs affected with mast cell tumour (MCT). Kaplan-Meier curves were constructed to compare tumour recurrence and tumour-related death between patients. The log-rank test was used to check for significant differences between curves. KIT-I, KIT-II and KIT-III staining patterns were observed in 9 (11.11%), 50 (61.73%) and 22 (27.16%) tumours, respectively. Tumour recurrence rates and tumour-related deaths were not associated with KIT staining patterns (P = 0278, P > 0.05), KIT (P = 0.289, P > 0.05) or KIT ligand (P = 0.106, P > 0.05) gene expression. Despite the lack of association between KIT staining pattern and patient survival time, the results suggest a correlation between aberrant KIT localization and increased proliferative activity of MCTs. RT-PCR seems to be a sensible method for quantitative detection of KIT gene expression in canine MCT, although expressions levels are not correlated with prognosis.
    Veterinary and Comparative Oncology 01/2013; · 1.56 Impact Factor
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    ABSTRACT: Gap junctions are communicating junctions which are important for tissue homeostasis, and their disruption is involved in carcinogenic processes. This study aimed to verify the influence of deletion of one allele of the Connexin 43 gene on cancer incidence in different organs. The 7, 12-dimethylbenzanthracene (DMBA) carcinogenic model, using hebdomadary doses by gavage of 9 mg per animal, was used to induce tumors in Connexin 43 heterozygous or wild-type mice. The experiment began in the eighth week of the mice life, and all of them were euthanized when reaching inadequate physical condition, or at the end of 53 weeks. No statistical differences occurred for weight gain and cancer survival time (P = 0.9853) between heterozygous and wild-type mice. Cx43(+/-) mice presented significantly higher susceptibility to lung cancer (P = 0.0200) which was not evidenced for benign neoplasms (P = 0.3449). In addition, incidence of ovarian neoplasms was 2.5-fold higher in Cx43(+/-) mice, although not statistically significant. Other organs showed a very similar cancer occurrence between Cx43 groups. The experiment strengthens the evidence of the relationship between Connexin 43 deficiency and carcinogenesis.
    BioMed research international. 01/2013; 2013:618475.
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    ABSTRACT: (Lam.) D.C. is a herb native to South America, and its inflorescences are popularly employed to treat inflammatory diseases. Here, the effects of the actions of the hydroalcoholic extract obtained from inflorescences of on neutrophil trafficking into inflamed tissue were investigated. Male Wistar rats were orally treated with extract, and inflammation was induced one hour later by lipopolysaccharide injection into the subcutaneous tissue. The number of leukocytes and the amount of chemotactic mediators were quantified in the inflammatory exudate, and adhesion molecule and toll-like receptor 4 (TLR-4) expressions and phorbol-myristate-acetate- (PMA-) stimulated oxidative burst were quantified in circulating neutrophils. Leukocyte-endothelial interactions were quantified in the mesentery tissue. Enzymes and tissue morphology of the liver and kidney were evaluated. Treatment with extract reduced neutrophil influx and secretion of leukotriene B4 and CINC-1 in the exudates, the number of rolling and adhered leukocytes in the mesentery postcapillary venules, neutrophil L-selectin, 2-integrin and TLR-4 expression, and oxidative burst, but did not cause an alteration in the morphology and activities of liver and kidney. Together, the data show that extract inhibits neutrophil functions related to the innate response and does not cause systemic toxicity.
    Evidence-based Complementary and Alternative Medicine 01/2013; 2013:787916. · 1.72 Impact Factor
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    ABSTRACT: S-Nitroso-N-acetylcysteine (SNAC) is a water soluble primary S-nitrosothiol capable of transferring and releasing nitric oxide and inducing several biochemical activities, including modulation of hepatic stellate cell activation. In this study, we evaluated the antifibrotic activity of SNAC in an animal model of nonalcoholic steatohepatitis (NASH) induced in Sprague-Dawley rats fed with a choline-deficient, high trans fat diet and exposed to diethylnitrosamine for 8 weeks. The rats were divided into three groups: SNAC, which received oral SNAC solution daily; NASH, which received the vehicle; and control, which received standard diet and vehicle. Genes related to fibrosis (matrix metalloproteinases [MMP]-13, -9, and -2), transforming growth factor β-1 [TGFβ-1], collagen-1α, and tissue inhibitors of metalloproteinase [TIMP-1 and -2] and oxidative stress (heat-shock proteins [HSP]-60 and -90) were evaluated. SNAC led to a 34.4% reduction in the collagen occupied area associated with upregulation of MMP-13 and -9 and downregulation of HSP-60, TIMP-2, TGFβ-1, and collagen-1α. These results indicate that oral SNAC administration may represent a potential antifibrotic treatment for NASH.
    Drug Design, Development and Therapy 01/2013; 7:553-563. · 3.49 Impact Factor
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    ABSTRACT: Studies of skin wound healing in crocodilians are necessary given the frequent occurrence of cannibalism in intensive farming systems. Air temperature affects tissue recovery because crocodilians are ectothermic. Therefore, the kinetics of skin wound healing in Caiman yacare were examined at temperatures of 33°C and 23°C. Sixteen caiman were selected and divided into two groups of eight maintained at 23°C or 33°C. The studied individuals' scars were photographed after 1, 2, 3, 7, 15 and 30 days of the experimental conditions, and samples were collected for histological processing after 3, 7, 15 and 30 days. Macroscopically, the blood clot (heterophilic granuloma) noticeably remained in place covering the wound longer for the caiman kept at 23°C. Microscopically, the temperature of 23°C slowed epidermal migration and skin repair. Comparatively, new blood vessels, labeled using von Willebrand factor (vWF) antibody staining, were more frequently found in the scars of the 33°C group. The collagen fibers in the dermis were denser in the 33°C treatment. Considering the delayed healing at 23°C, producers are recommended to keep wounded animals at 33°C, especially when tanks are cold, to enable rapid wound closure and better repair of collagen fibers because such lesions tend to compromise the use of their skin as leather.
    Biology open. 01/2013; 2(11):1171-8.
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    ABSTRACT: Freshly established cultures of primary hepatocytes progressively adopt a foetal-like phenotype and display increased production of connexin43. The latter is a multifaceted cellular entity with variable subcellular locations, including the mitochondrial compartment. Cx43 forms hemichannels and gap junctions that are involved in a plethora of physiological and pathological processes, such as apoptosis. The present study was conducted with the goal of shedding more light onto the role of connexin43 in primary hepatocyte cultures. Connexin43 expression was suppressed by means of RNA interference technology, and the overall outcome of this treatment on the hepatocellular proteome and metabolome was investigated using tandem mass tag-based differential protein profiling and (1)H NMR spectroscopy, respectively. Global protein profiling revealed a number of targets of the connexin43 knock-down procedure, including mitochondrial proteins (heat shock protein 60, glucose-regulated protein 75, thiosulphate sulphurtransferase and adenosine triphosphate synthase) and detoxifying enzymes (glutathione S-transferase μ 2 and cytochrome P450 2C70). At the metabolomic level, connexin43 silencing caused no overt changes, though there was some evidence for a subtle increase in intracellular glycine quantities. Collectively, these data could further substantiate the established existence of a mitochondrial connexin pool and could be reconciled with the previously reported involvement of connexin43 signalling in spontaneously occurring apoptosis in primary hepatocyte cultures.
    Archives of Toxicology 12/2012; · 5.22 Impact Factor
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    ABSTRACT: The thymus is a primary lymphatic organ that develops its activity in young organisms. But despite its function is fundamental mechanisms responsible for the acquisition and subsequent body defenses, it is not yet fully understood, neither the morphological basis accounting for such functions as the process of development and the organ involution. The objective was to analyze and characterize the morphology of the thymus, such as its size and volume, and histological aspects of the thymus in cats, correlating sex and age development. Twelve foetus of mongrel domestic cats (Felis domesticus), males and females, divided into three age groups. The thymus presented two parts with a pale pink color, the thoracic and cervical portion; each has a right lobe and a left lobe mostly. The thoracic portion was located in the region of cranial mediastinum, between the lungs and the heart base. And the cervical portion extended beyond the ribs cranially and is located ventral to the trachea. The thymus cellular structure was shown by the presence of organized concentric aggregates, named Hassall's corpuscles, formed by epithelial cells, supported by a connective tissue capsule that penetrated the parenchyma dividing it into lobules. Significant changes with number of lobes and thymus size between individuals of the same age, and between sex. The values for length, thickness and width, in general, showed an increase, according to the development of animals, with differences between sex.
    Pesquisa Veterinária Brasileira 12/2012; 32:41-46. · 0.54 Impact Factor
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    ABSTRACT: Lung cancer is the leading cause of cancer-related mortality in both men and women throughout the world. This disease is strongly associated with tobacco smoking. The aim of this manuscript was to establish an in vitro model that mimics the chronic exposures of alveolar epithelial type II cells to the tobacco-specific nitrosamine carcinogen, NNK. Immortalized non-neoplastic alveolar epithelial cells type II, (E10 cells), from BALB/c mice were exposed to low concentration of NNK (100 pM) during 5, 10, 15, and 20 cycles of 48 h. NNK-transformed cells showed an increase of proliferation rate and motility. Moreover, these cells underwent epithelial-to-mesenchymal transition (EMT). Increased migratory capacity and EMT were correlated to the time of exposure to NNK. NNK-transformed cells were tested for their growth and metastatic capacity in vivo. Subcutaneous injection of cells exposed to NNK for 20 cycles (E10-NNK20 clone) into BALB/c mice led to the formation of subcutaneous tumors that arose after 40 ± 17 d in all animals, which died 95 ± 18 d after cell inoculation, with lymph nodes and lung metastasis. The morphological characteristics of tumors were compatible with metastatic undifferentiated carcinoma. Cells exposed to NNK for 5-10 cycles did not display metastatic capacity, while those exposed for 15 cycles displayed low capacity. Our results show that prolonged exposures to NNK led the cells to increasingly acquire malignant properties. The cellular model presented in this study is suitable for studying the molecular events involved in the different stages of malignant transformation. © 2012 Wiley Periodicals, Inc.
    Molecular Carcinogenesis 11/2012; · 4.27 Impact Factor

Publication Stats

62 Citations
57.06 Total Impact Points

Institutions

  • 2008–2014
    • University of São Paulo
      • Department of Pathology (Sao Paulo)
      San Paulo, São Paulo, Brazil
  • 2012–2013
    • Free University of Brussels
      • • Faculty of Medicine and Pharmacy
      • • Department of Toxicology, Dermato-Cosmetology and Pharmacognosy (FAFY)
      Brussels, BRU, Belgium
  • 2010
    • Senac São Paulo
      San Paulo, São Paulo, Brazil
  • 2009
    • Experimental Pathology Laboratories, Inc.
      Sterling, Virginia, United States