Navegando por Autor "HENRIQUES, Daniele Freitas"

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Imunopatologia hepática da infecção experimental do vírus dengue em Callithrix penicillata
(Universidade Federal do Pará, 2015-09-25) HENRIQUES, Daniele Freitas; VASCONCELOS, Pedro Fernando da Costa; http://lattes.cnpq.br/0973550817356564
Dengue is one of most important public health problem in the world, and one of the factors that contribute with this situation is the lack of an effective vaccine against the disease, and in part, this difficulty to obtain is due clinical the lack of an experimental model that mimic the infection and manifestation of the disease as those observed in humans. Then, the Evandro Chagas Institute showed the presence of dengue antigens using the Immunohistochemistry assay, in the liver of a non-human primate (NHP) of the species Callithrix jacchus, with a fatal outcome following dengue hemorrhagic fever. Based in the above considerations and in addition to the fact that the liver is the targetb organ for Dengue Virus (DENV) in humans, it is important to analyze the liver of primates of the genus Callithrix in order to study the pathogenesis and the immunopathology of sequential infection by DENV. A total of 26 NHP Callithrix penicillata were submitted to primary infection (PI) subcutaneously with DENV-3 (3.23 x 103 PFU/mL), and 13 of these animals anesthetized and sacrificed daily for seven days post-infection (dpi) (acute phase) and in random intervals until 60 dpi (convalescent phase); the secondary infection (SI) with DENV-2 (4.47 x 104 PFU/mL) was performed two months after the PI in the remaining 13 animals. Uninfected sentinels animals were reserved up ending of the experiment. The liver of the animals were processed for histopathology and Immunohistochemical assay using polyclonal antibodies to DENV and antibodies for analyses of the innate and cellular immune response as well as the cytokine expression . The NHP were susceptible to sequential infection by DENV-3 and DENV-2; in the liver viral antigens were expressed in hepatocytes, Kupffer cells and Councilman bodies; the histopathological changes in liver was characterized by the presence of apoptosis, focal lytic necrosis, steatosis, swelling cellular, inflammation (acinar, EP and HCV), hyperplasia/hypertrophy in Kupffer cells, hemosiderin in Kupffer cells and sinusoidal dilatation; the intensity of the liver damage was prominent in the acute phase of PI and SI resulting in acute hepatitis. In addition the apoptosis was the most frequent mechanism of death of hepatocytes; lytic necrosis and inflammatory infiltrate showed predominant distribution pattern in Z2. An increase of the acinar expression of activated macrophages, NK cells, S-100 protein and B-lymphocytes during the stages of PI and SI; as well as, increased acinar expression of TCD4 + lymphocytes during the acute phase of PI; as to the quantification of cytokines and molecules, was observed an increase in acinar expression of: IFN- during the stages of PI and SI, TNF-α and IL-8 with higher prevalence in SI, TGF-β and IL-10 with prevalence in acute phase of PI, Fas protein during the acute phase of PI and SI and VCAM in acute phase of SI, as well as increase in expression in the EP. These findings were similar to those observed in livers from fatal cases of dengue fever in humans, but with lower intensity and amplitude, thus indicating that NHP Callithrix penicillata species is a good experimental model for infection by DENV, involving immunopathologic studies.
Acesso aberto (Open Access)
Infecção persistente pelos flavivírus Ilhéus e Rocio em hamsters dourados jovens (Mesocricetus auratus)
(Universidade Federal do Pará, 2009-08-28) HENRIQUES, Daniele Freitas; VASCONCELOS, Pedro Fernando da Costa; http://lattes.cnpq.br/0973550817356564
Ilheus (ILHV) and Rocio (ROCV) are flaviviruses (family Flaviviridae, genus Flavivirus) of great importance to public health in Brazil because these viruses are associated to encephalitis cases in humans. Recent studies have reported persistence of experimental infections (in vivo and in vitro) and clinical reports. The purpose of this study was to investigate in vivo the possible occurrence of persistent infection caused by ILHV and ROCV using young golden hamsters (Mesocricetus auratus) as experimental model. Hamsters were inoculated intraperitoneally with a suspension of brains of newborn mice infected with titers of 9.8 and 9.6 DL50/ 0.02 mL of ROCV and ILHV respectively, and at pre-determined intervals, they were anesthetized and sacrificed for collection of blood samples, serum and urine and organ fragments during four months (120 days) post-inoculation (p.i.). Viral quantification was calculated in samples of brain, liver and blood, using the technique of Real Time RT-PCR (qRT-PCR). All collected specimens were inoculated into VERO cells for confirmation of viral replication; and viral antigens in the cell cultures were detected by indirect immunofluorescence test; the levels of antibodies were determined by hemagglutination-inhibition test. Histopathological examination by hematoxylin-eosin and detection of viral antigens by imunohistochemistry were assayed in viscera and central nervous tissue samples collected during the kinetics. The study showed that young golden hamsters are good experimental model for persistent infection by the flaviviruses ILHV and ROCV. Both viruses induced strong immune response, although the levels of antibodies to ILHV were greater than for ROCV. The ROCV has demonstrated to be more pathogenic in these animals, suggesting higher ability to cause neuronal invasiveness than ILHV. Infected viscera samples inoculated in VERO cells resulted in growth of both viruses from all infected organ, blood, serum and urine samples and were confirmed by indirect immunofluorescence assay. Regarding persistence of infection, ROCV was detected in the brain, liver and blood by qRT-PCR, for three months p.i., while ILHV persistence was observed only in the brain for 30 days p.i. by qRT-PCR. The ROCV was able to produce histopathological changes, and immuno-labeled cells expressing viral antigens in liver, kidney, lung and brain samples during four month were confirmed by imunohistochemistry. To the ILHV, the histopathological changes and expression of viral antigens in samples from the liver, kidney and lung were only confirmed up to 30 days p.i., but the brain was positive for four months p.i.; The findings obtained in this study showed that both viruses have capacity to cause persistent infection in hamsters intraperitoneally infected, studies additional are needed to determine the pathophysiology and pathogenesis of ILHV and ROCV persistent infections.
Acesso aberto (Open Access)
Persistence of experimental Rocio virus infection in the golden hamster (Mesocricetus auratus)
(2012-08) HENRIQUES, Daniele Freitas; QUARESMA, Juarez Antônio Simões; FUZII, Hellen Thais; NUNES, Márcio Roberto Teixeira; SILVA, Eliana Vieira Pinto da; CARVALHO, Valéria Lima; MARTINS, Lívia Carício; CASSEB, Samir Mansour Moraes; CHIANG, Jannifer Oliveira; VASCONCELOS, Pedro Fernando da Costa
Rocio virus (ROCV) is an encephalitic flavivirus endemic to Brazil. Experimental flavivirus infections have previously demonstrated a persistent infection and, in this study, we investigated the persistence of ROCV infection in golden hamsters (Mesocricetus auratus). The hamsters were infected intraperitoneally with 9.8 LD50/0.02 mL of ROCV and later anaesthetised and sacrificed at various time points over a 120-day period to collect of blood, urine and organ samples. The viral titres were quantified by real-time-polymerase chain reaction (qRT-PCR). The specimens were used to infect Vero cells and ROCV antigens in the cells were detected by immunefluorescence assay. The levels of antibodies were determined by the haemagglutination inhibition technique. A histopathological examination was performed on the tissues by staining with haematoxylin-eosin and detecting viral antigens by immunohistochemistry (IHC). ROCV induced a strong immune response and was pathogenic in hamsters through neuroinvasion. ROCV was recovered from Vero cells exposed to samples from the viscera, brain, blood, serum and urine and was detected by qRT-PCR in the brain, liver and blood for three months after infection. ROCV induced histopathological changes and the expression of viral antigens, which were detected by IHC in the liver, kidney, lung and brain up to four months after infection. These findings show that ROCV is pathogenic to golden hamsters and has the capacity to cause persistent infection in animals after intraperitoneal infection.