Gynura segetum induces hepatic sinusoidal obstruction syndrome in mice by impairing autophagy

ABSTRACT Purpose: To investigate the underlying mechanism of hepatic sinusoidal obstruction syndrome (HSOS) induced by Gynura segetum by measuring autophagy in mouse models. Methods: The model group was administered G. segetum (30 g/kg/d) by gavage, while the normal control group was administered an equal volume of saline daily for five weeks. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), hepatic histopathological examinations, and Masson staining were performed to evaluate liver injury. Liver intercellular adhesion molecule-1 (ICAM-1) and P-selectin were evaluated by immunohistochemistry. Hepatocellular apoptosis was assessed using the terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay. Protein expression levels of autophagy markers were measured using Western blot analysis. Results: Gynura segetum was found to significantly induce liver injury compared with control mice, as evidenced by the increase of serum transaminases, a decrease in triglyceride levels, and histopathological changes in mice. Gynura segetum remarkably induced hepatocellular apoptosis and upregulated the expressions of ICAM-1 and P-selectin and also downregulated the protein expression levels of LC3, Atg12 and cytoplasmic polyadenylation element binding protein. Conclusions: Our results suggested that G. segetum induced liver injury with HSOS, and it was partly due to its ability to impair the autophagy pathway.

Suppresses of Astragalus Polysaccharide on E. coli-Induced Injured Intestinal Microvascular through TLR4-NF-B Signal Pathways in Chickens

To investigate the hypothesis that APS can attenuate E. coli-induced microvascular dysfunction in chicken intestine, 60 0-day old male chickens were divided into three groups with 5 replications of 4 chicks. Chicken in the APS group were treated with 15 mg APS daily while the others were given 0 mg APS for 6 days. Then all 7-day old chicken were injected intraperitoneally by E. coli, except for the chicken in the control group. After 4 h, all chickens intestinal samples were collected to detect gene expressions involved in inflammatory factors and adhesion molecules. Results showed that APS attenuated the signs of edema and hemorrhage in 7-day old chicken intestinal mucosa which were induced by E. coli injection. Consistently, APS significantly reduced the increasing mRNA levels of inflammatory factors such as Tumor necrosis factor-a (TNF-a), interleukin (IL) -1 and inducible nitric oxide synthase (iNOS) (p 0.05), the same results were observed in vascular adhesion molecules such as E-selectin, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). Moreover, we observed that APS supplementation in water suppressed significantly (p 0.05) the decline of reparative factors such as epithelial growth factor (EGF) and basic fibroblast growth factor (bFGF) in E. coli injected group. Furthermore, supplementation with APS substantially blocked (p 0.05) the increase in Toll-like receptor-4 (TLR4) and Nuclear factor (NF)-B mRNA abundance (p=0.087) induced by E. coli infection. This study indicated that microvascular injured chicken intestine induced by E. coli would be attenuated with feeding APS, and the mechanism of repairing were probably mediated through TLR4-NF-B signal pathways.(AU)

Suppresses of Astragalus Polysaccharide on E. coli-Induced Injured Intestinal Microvascular through TLR4-NF-B Signal Pathways in Chickens

To investigate the hypothesis that APS can attenuate E. coli-induced microvascular dysfunction in chicken intestine, 60 0-day old male chickens were divided into three groups with 5 replications of 4 chicks. Chicken in the APS group were treated with 15 mg APS daily while the others were given 0 mg APS for 6 days. Then all 7-day old chicken were injected intraperitoneally by E. coli, except for the chicken in the control group. After 4 h, all chickens intestinal samples were collected to detect gene expressions involved in inflammatory factors and adhesion molecules. Results showed that APS attenuated the signs of edema and hemorrhage in 7-day old chicken intestinal mucosa which were induced by E. coli injection. Consistently, APS significantly reduced the increasing mRNA levels of inflammatory factors such as Tumor necrosis factor-a (TNF-a), interleukin (IL) -1 and inducible nitric oxide synthase (iNOS) (p 0.05), the same results were observed in vascular adhesion molecules such as E-selectin, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). Moreover, we observed that APS supplementation in water suppressed significantly (p 0.05) the decline of reparative factors such as epithelial growth factor (EGF) and basic fibroblast growth factor (bFGF) in E. coli injected group. Furthermore, supplementation with APS substantially blocked (p 0.05) the increase in Toll-like receptor-4 (TLR4) and Nuclear factor (NF)-B mRNA abundance (p=0.087) induced by E. coli infection. This study indicated that microvascular injured chicken intestine induced by E. coli would be attenuated with feeding APS, and the mechanism of repairing were probably mediated through TLR4-NF-B signal pathways.

Exogenous normal lymph alleviates lipopolysaccharide-induced acute lung injury through lessening the adhesion molecules

To evaluate the role of exogenous normal lymph (ENL) on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in rats. ALI was induced by the jugular vein injection of LPS (iv, 15 mg/kg) in rats of the LPS and LPS+ENL groups within 15 min, then, ENL without cell components (5 ml/kg) was infused at the speed of 0.5 ml per minute in the LPS+ENL group, the same amount of saline was administered in the LPS group. The rats in the sham group received the same surgical procedure and saline. The histomorphology and the levels of P-selectin, intercellular adhesion molecule-1 (ICAM-1), myeloperoxidase (MPO) in pulmonary tissue were assessed. LPS induced pulmonary injury as well as increased the wet/dry weight ratio (W/D) and the levels of P-selectin, ICAM-1, and MPO in pulmonary tissues. These deleterious effects of LPS were significantly ameliorated by ENL treatment. Exogenous normal lymph could markedly alleviate the acute lung injury induced by lipopolysaccharide, and its effects might be related to lessening the adhesion molecules.(AU)

Contribuição ao estudo da ação in vitro do veneno da aranha marrom, Loxosceles gaucho, sobre plaquetas humanas e de coelho. Participação das plaquetas na dermonecrose induzida experimentalmente pelo veneno loxoscélico em coelhos / Contribution to the study of the in vitro action induced by the venom of the brown spider, Loxosceles gaucho, on human and rabbit platelets. Platelet participation in the dermonecrotic lesion experimentally induced in rabbits by the loxoscelic venom

Os venenos de aranhas do gênero Loxosceles possuem uma ampla gama de atividades em diferentes células, tecidos e sistemas, e é notória a sua capacidade de causar a formação da lesão dermonecrótica. Em um trabalho prévio, relatamos uma intensa trombocitopenia induzida pelo veneno loxoscélico em coelhos. Objetivou-se no presente estudo avaliar primeiramente a interação in vitro entre as plaquetas, humanas e de coelhos, com o veneno de Loxosceles gaucho e seu principal componente, a fração esfingomielinásica. Posteriormente, investigou-se o papel da plaqueta no desenvolvimento da lesão dermonecrótica através de um modelo de trombocitopenia em coelhos. Nos estudos in vitro, os resultados de agregação em plasma rico em plaquetas evidenciou que tanto o veneno total quanto a fração esfingomielinásica induziram um aumento desta resposta nas plaquetas das duas espécies, que foi mais intensa nas plaquetas humanas. Uma vez que em plaquetas lavadas a agregação não foi constatada, confirmou-se a necessidade de componente(s) plasmático(s) para a indução desta resposta. Ainda, os dados de LDH indicaram que o veneno total, nas concentrações aqui utilizadas, não foi capaz de causar à lise plaquetária. Com relação à ativação, o veneno total, foi capaz de aumentar a expressão da LIBS1 nas plaquetas de ambas as espécies e de P-selectina na plaqueta de coelho. Sob as nossas condições experimentais, a fração não promoveu o aumento dos marcadores de ativação. Finalizando os estudos in vitro, o veneno total e a fração provocaram um aumento da adesão nas plaquetas das duas espécies, sendo que a resposta à fração esfingomielinásica mostrou-se dose-dependente. O estudo experimental, por vez, necessitou de uma etapa preliminar para a obtenção de um anticorpo anti-plaqueta, produzido em cabras, que mostrou-se eficaz em induzir um intenso quadro trombocitopênico em coelhos. Análises histopatológicas indicam que a ausência das plaquetas não diminuiu a formação de trombos intravasculares na área da derme sob a ação do veneno. O desenvolvimento da lesão dermonecrótica em coelhos injetados com o veneno loxoscélico mostrou-se mais intenso naqueles depletados de plaquetas do que nos animais com número normal de plaquetas, constatável pelo desenvolvimento de maiores áreas de equimose e das escaras necróticas. Os resultados semelhantes dos níveis plasmáticos do fator de von Willembrand, dos tempos de protrombina e de tromboplastina parcial ativada, assim como dos testes de fagocitose por polimorfonucleares isolados do sangue periférico, entre os coelhos trombocitopênicos e aqueles com contagem normal de plaquetas, evidenciam que não houve alterações expressivas nas células endoteliais, no sistema da coagulação e na capacidade fagocítica via C3b dos polimorfonucleares do sangue periférico. Os presentes resultados mostram que o veneno loxoscélico in vitro é capaz de elicitar em plaquetas humanas e de coelhos as respostas de agregação, de adesão e de ativação. Por fim, as diferenças no desenvolvimento da lesão, constatadas entre o animal trombocitopênico e o animal normal, evidenciam que a plaqueta é um fator importante para controlar a expansão e a gravidade da lesão dermonecrótica induiza em coelhos pelo veneno loxoscélico

Venoms from Loxosceles spiders exhibit a wide range of activities on different cells, tissues and systems, being specially evident their ability to cause a dermonecrotic lesion. In a previous paper, we showed that Loxosceles gaucho spider venom induces intense thrombocytopenia in rabbits. Thus, in the present study, we firstly aimed to evaluate the in vitro interaction between human and rabbits platelets and L. gaucho venom and its major component, the sphingomyelinase fraction. Secondly, we investigated the platelet role in the dermonecrotic lesion formation using an experimental model of thrombocytopenia in rabbits. Both total venom and its sphingomyelinase fraction stimulated in vitro aggregation per se in platelet-rich plasma from both species, but a more intense response was obtained with human platelets. Taking into consideration that neither total venom nor its sphingomyelinase fraction induced aggregation of washed platelets, it can be deduced that one or more plasma components are necessary to induce this response. Moreover, LDH data indicated that total venom, at least at the concentration levels used in this study, cannot induce platelet lysis. Regarding platelet activation, total venom was able to increase the expression of LIBS1 on both human and rabbit platelet surface and P-selectin on only rabbit platelet surface. However, the sphingomyelinase fraction did not increase the expression of any marker of platelet activation, at least in the experimental conditions used in this study. Another in vitro study showed that both total venom and its sphingomyelinase fraction induce an increase in human and rabbit platelet adhesion, with the sphingomyelinase fraction exhibiting a dose-dependent response. For the study using an animal model of thrombocytopenia, it was necessary to previously produce goat antibodies against rabbit platelets, which showed to be effective to induce an intense thrombocytopenia in rabbits. Histopathological analysis of venom-induced dermic tissue lesion indicated that platelet absence does not decrease the formation of intravascular thrombi in the injured area. Nonetheless, bigger areas of equimosis and necrotic scabs could be observed in the dermonecrotic lesion of rabbits injected with loxoscelic venom and depleted of platelets. In the different groups, plasma levels of von Willembrand factor, prothrombin and activated partial thromboplastin times, and data from phagocytosis tests using polymorphonuclear cells isolated from rabbit periferical blood were similar comparing thrombocitopenic rabbits with those exhibiting normal platelet count; therefore, we can deduce that there is not any expressive alteration in endotelial cells, coagulation system and phagocytic ability of polymorphonuclear leukocytes via their complement receptor C3b. In conclusion, the loxoscelic venom is able to unchain in vitro different mechanisms leading to adhesion, activation and aggregation of both human and rabbit platelets. Furthermore, due to differences in the dermic lesion patterns between thrombocitopenic and normal animals, we can conclude that platelets must play a key role in controlling the expansion and severity of the dermonecrotic lesion induced in rabbits by L. gaucho venom