RESUMO
In this work we extend the toxicological studies of hot aqueous extract of A. satureioides (As-HAE) evaluating cytotoxic and apoptotic effects on human peripheral blood mononuclear cells (PBMCs). We also determine genotoxic action of this extract in vivo. In addition, the extract was chemically characterized. Finally, we established a comparison with previous data of cold aqueous extract. The As-HAE induced cytotoxicity on PBMCs determined by trypan blue dye exclusion (CC50 = 653 µg/mL) and MTT (CC50 = 588 µg/mL) assays being more toxic than cold extract. However, As-HAE as well as cold extract did not induce apoptosis measured by Hoechst 33258 staining, TUNEL assay, and DNA fragmentation analysis. The in vivo micronucleus test showed that As-HAE exerted cytogenotoxic effects on bone marrow of mice, contrary to what was observed with cold extract. The chemical study of As-HAE allowed identifying the flavonoids found in cold extract: luteolin, quercetin, and 3-O-methylquercetin, but at higher concentrations. We suggest that toxic effects induced by As-HAE could be due to high concentrations of these flavonoids. Given that As-HAE is the most used in folkloric medicine, its administration should be controlled in order to prevent potential cell damage.
Assuntos
Flavonoides/farmacologia , Luteolina/farmacologia , Extratos Vegetais/farmacologia , Quercetina/análogos & derivados , Achyrocline/química , Animais , Apoptose/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Fragmentação do DNA/efeitos dos fármacos , Flavonoides/química , Humanos , Leucócitos Mononucleares/efeitos dos fármacos , Luteolina/isolamento & purificação , Camundongos , Extratos Vegetais/química , Quercetina/isolamento & purificação , Quercetina/farmacologiaRESUMO
Achyrocline satureioides is widely consumed as infusion or aperitif and shows important therapeutic properties. Previously, we reported absence of genotoxicity of cold aqueous extract (CAE) of A. satureioides by Allium test. However, one test cannot predict the genotoxic effects of a substance. Thus, the aim of this work was to investigate cytotoxicity, genotoxicity and apoptotic ability of CAE of A. satureioides. In addition, CAE was chemically characterized. The cytotoxicity was evaluated by Trypan blue and MTT assays. The apoptotic capacity was evaluated by Hoechst staining and DNA fragmentation-analysis. The genotoxicity was studied by comet assay (CA) and micronucleus test. The identification and quantification of flavonoids were performed by HPLC-ESI-MS/MS. The cytotoxicity studies indicated low toxicity of CAE. In addition, CAE did not induce apoptotic effects on human PBMCs. CAE did not show genotoxicity in vitro against Vero cells, at 10-50 µg/mL. CAE did not induce in vivo genotoxic effects, but it showed at high concentrations cytotoxicity by micronucleus assay. CAE presented flavonoids such as quercetin, 3-O-methylquercetin and luteolin. In conclusion, A. satureioides at popularly concentrations used, in aperitif or infusion, can be consumed safely because did not show any cytotoxic or genotoxic effects.
Assuntos
Achyrocline/química , Apoptose/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Extratos Vegetais/farmacologia , Animais , Chlorocebus aethiops , Cromatografia Líquida de Alta Pressão , Ensaio Cometa , Fragmentação do DNA , Avaliação Pré-Clínica de Medicamentos , Feminino , Humanos , Dose Letal Mediana , Leucócitos Mononucleares/efeitos dos fármacos , Luteolina/análise , Luteolina/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Testes para Micronúcleos , Extratos Vegetais/análise , Quercetina/análogos & derivados , Quercetina/análise , Quercetina/farmacologia , Espectrometria de Massas em Tandem , Células VeroRESUMO
The antiviral activity of alcoholic extracts of several species belonging to the Asteraceae, Labiatae, Plantaginaceae, Schizaceae, Umbelliferae, Usneaceae and Verbenaceae families has been studied. The tests were carried out in Vero celís-pseudorabies virus strain RC/79 (herpes suis virus) system. Eight plant extracts (Achyrocline satureioides, Ambrossia tenuifolia, Baccharis articulata, Eupatorium buniifolium, Mynthostachys verticillata, Plantago brasiliensis, Plantago mayor L and Verbascum thapsus) were able to inhibit at least 2 log, the viral infectivity.
Assuntos
Antivirais/isolamento & purificação , Herpesvirus Suídeo 1/efeitos dos fármacos , Extratos Vegetais/farmacologia , Plantas Medicinais/química , Animais , Antivirais/farmacologia , Argentina , Chlorocebus aethiops , Herpesvirus Suídeo 1/fisiologia , Células Vero , Replicação Viral/efeitos dos fármacosRESUMO
The RC/79 strain of the Aujeszky's disease virus was able to induce reproductive failure of pregnant gilts intranasally inoculated at different gestation periods. Four gilts 40-46 days pregnant (group A) and 6 gilts 70-73 days pregnant (group B) were instilled with 0.2 ml x 10(5) tissue culture infectious dose 50 (TCID50/0.2 ml) of the RC/79 strain into each nostril. Two gilts 70-73 days pregnant (group C) were used as non exposed controls. The three groups were kept in separated boxes and they were observed for clinical signs of infections and samples were collected for determination of viral shedding every day. Viral isolation was attempted in Vero cells (figure 1). From the 2nd to 7th day after inoculation, groups A and B showed fever anorexia, sneezing, coughing and depression; and viral isolation from nasal swabs was possible in 7 gilts at days 4 to 11, 9 gilts developed neutralizing antibodies. The virus caused fetal reabsorption in swine during the first period of pregnancy (group A), while infection during late pregnancy resulted in still birth or normal pigs and one mummification (group B). The entire a live litter was composed of no more than 8 suckling pigs in both groups. At necropsy virus from turbinates, ovary , placenta, spleen and lung could be isolated only from 3 gilts (group B, table 1). In 5 of 35 stillbirth and alive fetuses virus could be isolated from spleen (100%), lung (80%), liver (60%) and brain (40%) indicating that the virus has the ability to cross the placental barrier thus producing lesions in porcine fetuses and causing reproductive failure in sows (table 2). Tissue specimens from these 35 fetuses were fixed in 10% formalin, included in paraffin sectioned and stained with hematoxylin and eosin. In 13 fetuses microscopic lesions i.e. necrotic foci were found in lung (60%), liver (40%) and spleen (20%), these alterations were coincident with gross lesions in most of them. Inclusion bodies were absent. The gilts organs did not present gross lesions.
Assuntos
Morte Fetal/veterinária , Doenças Fetais/veterinária , Herpesvirus Suídeo 1/isolamento & purificação , Complicações Infecciosas na Gravidez/veterinária , Pseudorraiva/transmissão , Doenças dos Suínos/transmissão , Administração Intranasal , Animais , Anticorpos Antivirais/biossíntese , Encéfalo/embriologia , Encéfalo/microbiologia , Feminino , Morte Fetal/etiologia , Morte Fetal/microbiologia , Morte Fetal/patologia , Doenças Fetais/etiologia , Doenças Fetais/microbiologia , Doenças Fetais/patologia , Reabsorção do Feto/etiologia , Reabsorção do Feto/veterinária , Herpesvirus Suídeo 1/classificação , Herpesvirus Suídeo 1/imunologia , Troca Materno-Fetal , Gravidez , Complicações Infecciosas na Gravidez/microbiologia , Pseudorraiva/microbiologia , Suínos , Doenças dos Suínos/microbiologia , Células Vero , Vísceras/embriologia , Vísceras/microbiologiaRESUMO
The RC/79 strain of the Aujeszkys disease virus was able to induce reproductive failure of pregnant gilts intranasally inoculated at different gestation periods. Four gilts 40-46 days pregnant (group A) and 6 gilts 70-73 days pregnant (group B) were instilled with 0.2 ml x 10(5) tissue culture infectious dose 50 (TCID50/0.2 ml) of the RC/79 strain into each nostril. Two gilts 70-73 days pregnant (group C) were used as non exposed controls. The three groups were kept in separated boxes and they were observed for clinical signs of infections and samples were collected for determination of viral shedding every day. Viral isolation was attempted in Vero cells (figure 1). From the 2nd to 7th day after inoculation, groups A and B showed fever anorexia, sneezing, coughing and depression; and viral isolation from nasal swabs was possible in 7 gilts at days 4 to 11, 9 gilts developed neutralizing antibodies. The virus caused fetal reabsorption in swine during the first period of pregnancy (group A), while infection during late pregnancy resulted in still birth or normal pigs and one mummification (group B). The entire a live litter was composed of no more than 8 suckling pigs in both groups. At necropsy virus from turbinates, ovary , placenta, spleen and lung could be isolated only from 3 gilts (group B, table 1). In 5 of 35 stillbirth and alive fetuses virus could be isolated from spleen (100
), lung (80
), liver (60
) and brain (40
) indicating that the virus has the ability to cross the placental barrier thus producing lesions in porcine fetuses and causing reproductive failure in sows (table 2). Tissue specimens from these 35 fetuses were fixed in 10
formalin, included in paraffin sectioned and stained with hematoxylin and eosin. In 13 fetuses microscopic lesions i.e. necrotic foci were found in lung (60
), liver (40
) and spleen (20
), these alterations were coincident with gross lesions in most of them. Inclusion bodies were absent. The gilts organs did not present gross lesions.
RESUMO
The RC/79 strain of the Aujeszkys disease virus was able to induce reproductive failure of pregnant gilts intranasally inoculated at different gestation periods. Four gilts 40-46 days pregnant (group A) and 6 gilts 70-73 days pregnant (group B) were instilled with 0.2 ml x 10(5) tissue culture infectious dose 50 (TCID50/0.2 ml) of the RC/79 strain into each nostril. Two gilts 70-73 days pregnant (group C) were used as non exposed controls. The three groups were kept in separated boxes and they were observed for clinical signs of infections and samples were collected for determination of viral shedding every day. Viral isolation was attempted in Vero cells (figure 1). From the 2nd to 7th day after inoculation, groups A and B showed fever anorexia, sneezing, coughing and depression; and viral isolation from nasal swabs was possible in 7 gilts at days 4 to 11, 9 gilts developed neutralizing antibodies. The virus caused fetal reabsorption in swine during the first period of pregnancy (group A), while infection during late pregnancy resulted in still birth or normal pigs and one mummification (group B). The entire a live litter was composed of no more than 8 suckling pigs in both groups. At necropsy virus from turbinates, ovary , placenta, spleen and lung could be isolated only from 3 gilts (group B, table 1). In 5 of 35 stillbirth and alive fetuses virus could be isolated from spleen (100
), lung (80
), liver (60
) and brain (40
) indicating that the virus has the ability to cross the placental barrier thus producing lesions in porcine fetuses and causing reproductive failure in sows (table 2). Tissue specimens from these 35 fetuses were fixed in 10
formalin, included in paraffin sectioned and stained with hematoxylin and eosin. In 13 fetuses microscopic lesions i.e. necrotic foci were found in lung (60
), liver (40
) and spleen (20
), these alterations were coincident with gross lesions in most of them. Inclusion bodies were absent. The gilts organs did not present gross lesions.
RESUMO
The RC/79 strain of the Aujeszkys disease virus was able to induce reproductive failure of pregnant gilts intranasally inoculated at different gestation periods. Four gilts 40-46 days pregnant (group A) and 6 gilts 70-73 days pregnant (group B) were instilled with 0.2 ml x 10(5) tissue culture infectious dose 50 (TCID50/0.2 ml) of the RC/79 strain into each nostril. Two gilts 70-73 days pregnant (group C) were used as non exposed controls. The three groups were kept in separated boxes and they were observed for clinical signs of infections and samples were collected for determination of viral shedding every day. Viral isolation was attempted in Vero cells (figure 1). From the 2nd to 7th day after inoculation, groups A and B showed fever anorexia, sneezing, coughing and depression; and viral isolation from nasal swabs was possible in 7 gilts at days 4 to 11, 9 gilts developed neutralizing antibodies. The virus caused fetal reabsorption in swine during the first period of pregnancy (group A), while infection during late pregnancy resulted in still birth or normal pigs and one mummification (group B). The entire a live litter was composed of no more than 8 suckling pigs in both groups. At necropsy virus from turbinates, ovary , placenta, spleen and lung could be isolated only from 3 gilts (group B, table 1). In 5 of 35 stillbirth and alive fetuses virus could be isolated from spleen (100
) and brain (40
) indicating that the virus has the ability to cross the placental barrier thus producing lesions in porcine fetuses and causing reproductive failure in sows (table 2). Tissue specimens from these 35 fetuses were fixed in 10
formalin, included in paraffin sectioned and stained with hematoxylin and eosin. In 13 fetuses microscopic lesions i.e. necrotic foci were found in lung (60
) and spleen (20
), these alterations were coincident with gross lesions in most of them. Inclusion bodies were absent. The gilts organs did not present gross lesions.
RESUMO
The RC/79 strain of the Aujeszkys disease virus was able to induce reproductive failure of pregnant gilts intranasally inoculated at different gestation periods. Four gilts 40-46 days pregnant (group A) and 6 gilts 70-73 days pregnant (group B) were instilled with 0.2 ml x 10(5) tissue culture infectious dose 50 (TCID50/0.2 ml) of the RC/79 strain into each nostril. Two gilts 70-73 days pregnant (group C) were used as non exposed controls. The three groups were kept in separated boxes and they were observed for clinical signs of infections and samples were collected for determination of viral shedding every day. Viral isolation was attempted in Vero cells (figure 1). From the 2nd to 7th day after inoculation, groups A and B showed fever anorexia, sneezing, coughing and depression; and viral isolation from nasal swabs was possible in 7 gilts at days 4 to 11, 9 gilts developed neutralizing antibodies. The virus caused fetal reabsorption in swine during the first period of pregnancy (group A), while infection during late pregnancy resulted in still birth or normal pigs and one mummification (group B). The entire a live litter was composed of no more than 8 suckling pigs in both groups. At necropsy virus from turbinates, ovary , placenta, spleen and lung could be isolated only from 3 gilts (group B, table 1). In 5 of 35 stillbirth and alive fetuses virus could be isolated from spleen (100
) and brain (40
) indicating that the virus has the ability to cross the placental barrier thus producing lesions in porcine fetuses and causing reproductive failure in sows (table 2). Tissue specimens from these 35 fetuses were fixed in 10
formalin, included in paraffin sectioned and stained with hematoxylin and eosin. In 13 fetuses microscopic lesions i.e. necrotic foci were found in lung (60
) and spleen (20
), these alterations were coincident with gross lesions in most of them. Inclusion bodies were absent. The gilts organs did not present gross lesions.
