RESUMO
AIM: To evaluate the antioxidant effect of N-acetylcysteine (NAC) on the stomach of rats with portal hypertension. METHODS: Twenty-four male Wistar rats weighing ± 250 g were divided into four experimental groups (n = 6 each): Sham-operated (SO), SO + NAC, partial portal vein ligation (PPVL), and PPVL + NAC. Treatment with NAC in a dose of 10 mg/kg (i.p.) diluted in 0.6 mL of saline solution was administered daily for 7 d starting 8 d after the surgery. Animals from the PPVL and SO group received saline solution (0.6 mL) for the same period of time as the PPVL + NAC and SO + NAC group. On the 15(th) day the animals were anesthetized and we evaluated portal pressure by cannulating mesenteric artery. After, we removed the stomach for further analysis. We performed immunohistochemical analysis for endothelial nitric oxide synthase (eNOS), vascular endothelial growth factor (VEGF), and nitrotirosine (NTT) proteins in stomach. We also evaluated eNOS and VEGF by Western blot analysis and assessed DNA damage in blood samples by the comet assay. RESULTS: The portal hypertension group exhibited increases in portal pressure when compared to SO group (29.8 ± 1.8 vs 12.0 ± 0.3 mmHg) (P < 0.001). The same was observed when we compared the eNOS (56.8 ± 3.7 vs 13.46 ± 2.8 pixels) (P < 0.001), VEGF (34.9 ± 4.7 vs 17.46 ± 2.6 pixels) (P < 0.05), and NTT (39.01 ± 4.0 vs 12.77 ± 2.3 pixels) (P < 0.05) expression by immunohistochemistry of the PPVL animals with the SO group. The expression of eNOS (0.39 ± 0.03 vs 0.25 ± 0.03 a.µ) (P < 0.01) and VEGF (0.38 ± 0.04 vs 0.26 ± 0.04 a.µ) (P < 0.01) were also evaluated by Western blot analysis, and we observed an increase of both proteins on PPVL animals. We also evaluated the DNA damage by comet assay, and observed an increase on damage index and damage frequency on those animals. NAC decreased portal pressure values in PPVL + NAC animals (16.46 ± 2 vs 29.8 ± 1.8 mmHg) (P < 0.001) when compared to PPVL. The expression of eNOS (14.60 ± 4.1 vs 56.8 ± 3.7 pixels) (P < 0.001), VEGF (19.53 ± 3.2 vs 34.9 ± 4.7 pixels) (P < 0.05) and NTT (21.84 ± 0.7 vs 39.01 ± 4.0 pixels) (P < 0.05) evaluated by immunohistochemistry were also reduced in PPVL + NAC animals. Also, when evaluated by Western blot eNOS expression (0.32 ± 0.03 vs 0.39 ± 0.03 a.µ) (P < 0.05) and VEGF expression (0.31 ± 0.09 vs 0.38 ± 0.04 a.µ) (P < 0.01). Furthermore, NAC modulated DNA damage in PPVL + NAC animals. CONCLUSION: In view of these results, we believe NAC is able to protect the stomach from the alterations induced by the PPVL procedure.
Assuntos
Acetilcisteína/farmacologia , Antioxidantes/farmacologia , Dano ao DNA/efeitos dos fármacos , Hipertensão Portal/tratamento farmacológico , Neovascularização Patológica , Estômago/irrigação sanguínea , Estômago/efeitos dos fármacos , Vasodilatação/efeitos dos fármacos , Animais , Western Blotting , Ensaio Cometa , Modelos Animais de Doenças , Mucosa Gástrica/metabolismo , Hipertensão Portal/sangue , Hipertensão Portal/genética , Hipertensão Portal/fisiopatologia , Imuno-Histoquímica , Masculino , Óxido Nítrico Sintase Tipo III/metabolismo , Pressão na Veia Porta/efeitos dos fármacos , Ratos Wistar , Tirosina/análogos & derivados , Tirosina/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Schistosomiasis is a major cause of portal hypertension worldwide. It associates with portal fibrosis that develops during chronic infection. The mechanisms by which the pathogen evokes these host responses remain unclear. We evaluated the hypothesis that schistosome eggs release factors that directly stimulate liver cells to produce osteopontin (OPN), a pro-fibrogenic protein that stimulates hepatic stellate cells to become myofibroblasts. We also investigated the utility of OPN as a biomarker of fibrosis and/or severity of portal hypertension. Cultured cholangiocytes, Kupffer cells and hepatic stellate cells were treated with soluble egg antigen (SEA); OPN production was quantified by quantitative reverse transcriptase polymerase chain reaction (qRTPCR) and ELISA; cell proliferation was assessed by BrdU (5-bromo-2'-deoxyuridine). Mice were infected with Schistosoma mansoni for 6 or 16 weeks to cause early or advanced fibrosis. Liver OPN was evaluated by qRTPCR and immunohistochemistry (IHC) and correlated with liver fibrosis and serum OPN. Livers from patients with schistosomiasis mansoni (early fibrosis n=15; advanced fibrosis n=72) or healthy adults (n=22) were immunostained for OPN and fibrosis markers. Results were correlated with plasma OPN levels and splenic vein pressures. SEA-induced cholangiocyte proliferation and OPN secretion (P<0.001 compared with controls). Cholangiocytes were OPN (+) in Schistosoma-infected mice and humans. Liver and serum OPN levels correlated with fibrosis stage (mice: r=0.861; human r=0.672, P=0.0001) and myofibroblast accumulation (mice: r=0.800; human: r=0.761, P=0.0001). Numbers of OPN (+) bile ductules strongly correlated with splenic vein pressure (r=0.778; P=0.001). S. mansoni egg antigens stimulate cholangiocyte proliferation and OPN secretion. OPN levels in liver and blood correlate with fibrosis stage and portal hypertension severity.
Assuntos
Proliferação de Células , Hipertensão Portal/metabolismo , Cirrose Hepática/metabolismo , Osteopontina/metabolismo , Esquistossomose mansoni/metabolismo , Adolescente , Adulto , Animais , Antígenos de Helmintos/farmacologia , Ductos Biliares/citologia , Ductos Biliares/efeitos dos fármacos , Ductos Biliares/metabolismo , Linhagem Celular , Células Cultivadas , Feminino , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/metabolismo , Interações Hospedeiro-Parasita , Humanos , Hipertensão Portal/genética , Hipertensão Portal/parasitologia , Imuno-Histoquímica , Células de Kupffer/efeitos dos fármacos , Células de Kupffer/metabolismo , Cirrose Hepática/genética , Cirrose Hepática/parasitologia , Masculino , Camundongos , Pessoa de Meia-Idade , Osteopontina/sangue , Osteopontina/genética , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Schistosoma/fisiologia , Esquistossomose mansoni/genética , Esquistossomose mansoni/parasitologia , Adulto JovemRESUMO
Lysosomal acid lipase (LAL) deficiency is an under-recognized lysosomal disease caused by deficient enzymatic activity of LAL. In this report we describe two affected female Mexican siblings with early hepatic complications. At two months of age, the first sibling presented with alternating episodes of diarrhea and constipation, and later with hepatomegaly, elevated transaminases, high levels of total and low-density lipoprotein cholesterol, and low levels of high-density lipoprotein. Portal hypertension and grade 2 esophageal varices were detected at four years of age. The second sibling presented with hepatomegaly, elevated transaminases and mildly elevated low-density lipoprotein and low high-density lipoprotein at six months of age. LAL activity was deficient in both patients. Sequencing of LIPA revealed two previously unreported heterozygous mutations in exon 4: c.253C>A and c.294C>G. These cases highlight the clinical continuum between the so-called Wolman disease and cholesteryl ester storage disease, and underscore that LAL deficiency represents a single disease with a degree of clinical heterogeneity.