Avaliação da segurança reprodutiva e da transmissão passiva de imunidade após processo de imunização com proteína peroxidoxina recombinante de Leishmania braziliensis durante a prenhez de ratas

Exportado OPUSMade available in DSpace on 2019-08-14T01:52:20Z (GMT). No. of bitstreams: 1 tese_apbr_corrigida.pdf: 1837035 bytes, checksum: ca478ba6874733512a5148f89c90db4a (MD5) Previous issue date: 27A proteína peroxidoxina recombinante de Leishmania braziliensis conjugada ao adjuvante MPL (Monophosphoryl Lipid A) tem se mostrado um promissor candidato a vacina antileishmaniose. Apesar do potencial imunogênico do processo vacinal ser amplamente demonstrado para diferentes vacinas antiparasitárias, dados sobre a avaliação da segurança de proteínas recombinantes em formulações vacinais administradas durante o período gestacional, assim como informações sobre a transferência passiva de imunidade em animais vacinados durante a prenhez, são ainda escassos. O objetivo desse trabalho foi avaliar a segurança vacinal quando da administração no período gestacional, com possíveis causas de prejuízos maternos ou fetais e avaliar a transferência passiva de imunidade para o recém-nascido de anticorpos reativos com a referida proteína, após processo de imunização em modelo experimental com ratos Wistar. Ratas em idade reprodutiva foram acasaladas e distribuídas em três grupos: Controle - ratas que receberam salina; Adjuvante - ratas que receberam adjuvante MPL, e Vacina - ratas que receberam a composição adjuvante mais a proteína peroxidoxina recombinante. A administração ocorreu por injeção subcutânea, na região dorsal, em três momentos (dias zero, sete e 14 de gestação). No 21º dia de prenhez, os animais foram anestesiados e mortos. Amostras de sangue materno e fetal foram coletadas para dosagem de anticorpos IgG anti-peroxidoxina por ELISA. O útero foi removido para se obter dados do desempenho reprodutivo e os fetos foram submetidos à analise de anomalias externas, esqueléticas e viscerais. Para avaliar a transferência de anticorpos via aleitamento, foi realizado um experimento de cross-fostering, onde ratas imunizadas e ratas controle tiveram seus filhotes por parto vaginal e as ninhadas foram trocadas para que recebessem amamentação adotiva. No 21º dia de lactação, os animais foram anestesiados e o sangue coletado para análise sorológica por ELISA. Houve um aumento na perda pós-implantação no grupo vacina (14,7%) em relação aos grupos controle (5,0%) e adjuvante (4,4%). Foi verificado um maior índice de anomalias viscerais em fetos provenientes do grupo Vacina. Com relação à sorologia, verificou-se que os níveis de imunoglobulina IgG anti-peroxidoxina foram superiores no grupo Vacina. Filhotes de ratas vacinadas também apresentaram níveis de IgG superiores àqueles provindos de ratas controle e adjuvante. A transferência de IgG anti-peroxidoxina permanece no período pós-natal, via aleitamento materno, e os índices de transferência de anticorpos via placenta ou amamentação são semelhantes. Esses dados permitem concluir que a imunização estimulou a produção da imunoglobulina IgG anti-peroxidoxina, sendo essa imunização transferida para feto via placenta e leite, mas a vacina aumentou as perdas pós-implantação e as anomalias fetais, mostrando que sua utilização durante a gravidez exige cuidado e maiores estudos.The peroxidoxin recombinant protein of Leishmania braziliensis associated with adjuvant MPL (Monophosphoryl Lipid A) has been shown to be a promising candidate for antileishmaniose vaccine. Although the immunogenic potential of the vaccine process is extensively shown for different anti-parasitic vaccines, data on the safety assessment of recombinant proteins in vaccine formulations administered during pregnancy, as well as information about the passive transfer of immunity in animals vaccinated during pregnancy are remain scarce. The objective of this study was to evaluate vaccine safety when the administration during pregnancy, with possible causes of maternal or fetal losses and evaluate the passive transfer of immunity to the newborn after immunization process in experimental model with Wistar rats. Rats in reproductive age were mated and allocated in three groups: Control - rats received saline; Adjuvant - rats received adjuvant MPL, and Vaccine - rats that received the composition adjuvant and peroxidoxin. The administration was by subcutaneous injection at the dorsal region, three times (days zero, seven and 14 of pregnancy). At day 21 of pregnancy the animals were anesthetized and blood collected for serological analysis by ELISA. There was an increase in post-implantation loss in the vaccine group (14.7%) compared to the control group (5.0%) and adjuvant (4.4%). It was verified a higher rate of visceral anomalies in fetuses from the vaccine group. Concerning to serology, it was found that levels of anti- peroxidoxin IgG were higher in the vaccine group. Offspring of vaccinated rats also showed higher IgG levels compared the offspring of rats control and adjuvant. The anti-IgG peroxidoxina transfer remains in the postnatal period, via breastfeeding, and transfer rates of antibodies through the placenta or breast-feeding are similar. These data showed that the immunization stimulated the production of IgG anti-peroxidoxin immunoglobulin, and this immunization transferred to the fetus through the placenta and milk, but the vaccine increased post-implantation loss and fetal anomalies, showing that its use during pregnancy requires care and further study

Brazilian Propolis: A Natural Product That Improved the Fungicidal Activity by Blood Phagocytes

Natural product incorporation into microcarriers increases the bioavailability of these compounds, consequently improving their therapeutic properties. Natural products, particularly those from bees such as propolis, are widely used in popular medicine. Propolis is a powerful treatment for several diseases. In this context, the present study evaluated the effect of propolis Scaptotrigona sp. and its fractions, alone or adsorbed to polyethylene glycol (PEG) microspheres, on the activity of human phagocytes against Candida albicans. The results show that propolis exerts a stimulatory effect on these cells to assist in combating the fungus, especially as the crude extract is compared with the fractions. However, when incorporated into microspheres, these properties were significantly potentiated. These results suggest that propolis adsorbed onto PEG microspheres has immunostimulatory effects on phagocytes in human blood. Therefore, propolis may potentially be an additional natural product that can be used for a variety of therapies

Safety evaluation of a vaccine: Effect in maternal reproductive outcome and fetal anomaly frequency in rats using a leishmanial vaccine as a model.

While the immunogenic potential of the vaccination against infectious diseases was extensively shown, data on the safety assessment of recombinant proteins in vaccine formulations administered during pregnancy are still scarce. In the current study, the antigenicity of a vaccine against leishmaniasis (based on Leishmania braziliensis recombinant protein peroxidoxin) during pregnancy and possible maternal reproductive outcomes and fetal anomalies after immunization with a leishmanial vaccine or adjuvant alone (Bordetella pertussis derived MPLA adjuvant) were assessed. Rats were mated and allocated in three groups: Control-rats received saline; Adjuvant-rats received the adjuvant MPLA, and Vaccine-rats received the combination of MPLA and peroxidoxin. The administration was subcutaneously at the dorsal region, three times (days 0, 7, 14 of pregnancy). On day 21 of pregnancy, all rats were bled for biochemical and immunological measurements. The gravid uterus was weighed with its contents, and the fetuses were analyzed. The immunization with peroxidoxin induced a significant production of circulating IgG levels compared to other groups but caused a significant in post-implantation loss (14.7%) when compared to Control (5.0%) and Adjuvant (4.4%) groups. Furthermore, a significantly high rate of fetal visceral anomalies, such as hydronephrosis and convoluted ureter, was also observed in animals that received vaccine when compared to Control or Adjuvant groups. These data indicate the importance of safety evaluation of vaccines during pregnancy and the limited use of peroxidoxin administration during pregnancy. More importantly, the safety monitoring of immunization with MPLA derived from Bordetella pertussis demonstrated no reproductive outcomes associated with adjuvant administration, suggesting its safe use during pregnancy

Reproductive outcome from rats treated with saline (Control), MPLA (Adjuvant) or MPLA + recombinant peroxidoxin of <i>Leishmania braziliensis</i> (Vaccine) during pregnancy.

<p>Reproductive outcome from rats treated with saline (Control), MPLA (Adjuvant) or MPLA + recombinant peroxidoxin of <i>Leishmania braziliensis</i> (Vaccine) during pregnancy.</p

Ossification sites of rats treated with saline (Control), MPLA (Adjuvant) or MPLA + recombinant peroxidoxin of <i>Leishmania braziliensis</i> (Vaccine) during pregnancy.

<p>Ossification sites of rats treated with saline (Control), MPLA (Adjuvant) or MPLA + recombinant peroxidoxin of <i>Leishmania braziliensis</i> (Vaccine) during pregnancy.</p

Body weight, water intake and food consumption of rats treated with saline (Control), MPLA (Adjuvant) or MPLA + recombinant peroxidoxin of <i>Leishmania braziliensis</i> (Vaccine) during pregnancy.

<p>Body weight, water intake and food consumption of rats treated with saline (Control), MPLA (Adjuvant) or MPLA + recombinant peroxidoxin of <i>Leishmania braziliensis</i> (Vaccine) during pregnancy.</p

Frequency of fetal anomalies of rats treated with saline (Control), MPLA (Adjuvant) or MPLA + recombinant peroxidoxin of <i>Leishmania braziliensis</i> (Vaccine) during pregnancy.

<p>Frequency of fetal anomalies of rats treated with saline (Control), MPLA (Adjuvant) or MPLA + recombinant peroxidoxin of <i>Leishmania braziliensis</i> (Vaccine) during pregnancy.</p

Biochemical parameters of rats treated with saline (Control), MPLA (Adjuvant) or MPLA + recombinant peroxidoxin of <i>Leishmania braziliensis</i> (Vaccine) during pregnancy.

<p>Biochemical parameters of rats treated with saline (Control), MPLA (Adjuvant) or MPLA + recombinant peroxidoxin of <i>Leishmania braziliensis</i> (Vaccine) during pregnancy.</p

Quantification of maternal IgG anti-peroxidoxin of rats immunized with saline (Control), MPLA (Adjuvant) or MPLA + recombinant peroxidoxin of <i>Leishmania braziliensis</i> (Vaccine) during pregnancy.

<p><b>Data is shown as mean ± Standard Deviation (SD)</b>. <i>*p<0</i>.<i>001 compared to Control group (ANOVA followed Tukey’s Multiple Comparison test)</i>. ^<i>#</i><i>p<0</i>.<i>001 compared to Adjuvant group (ANOVA followed by Tukey’s Multiple Comparison test)</i></p