RESUMO
BACKGROUND/AIMS: Angiogenesis is a feature of the atherogenic process, with intimal neovascularisation arising from vessels in the adventitia, adjacent to a plaque. Immature, leaky blood vessels from unstable plaques proliferate abnormally and, being poorly invested with smooth muscle cells, may contribute to instability of the plaque by facilitation of inflammatory cell infiltration and haemorrhagic complications. METHODS: We used laser-capture microdissection to isolate angiogenic areas of the extracellular matrix (containing CD105/flt-1-positive, fragile thin-walled vessels) and non-angiogenic vascular areas (CD105-negative, with smooth muscle cell covering) of complicated endarterectomy plaques, and specifically designed angiogenesis-TaqMan real-time PCR microarrays to identify gene expression. RESULTS: Important pro-angiogenic components, including Notch-3, delta-like-4 (DLL4), Tie-2, angiopoietin-1 (Angio-1) and receptor for advanced glycation end products (RAGE), and one anti-angiogenic factor, endostatin, were up-regulated in these regions. Immunohistochemistry demonstrated localisation within intimal, active (CD105-positive) microvessels and co-localisation of Notch-3 and DLL4/Tie-2 and Angio-1 in the same vessels indicating multiple/synergistic signalling mechanisms associated with vessel development. CONCLUSION: These data, although providing only a snapshot of information, demonstrate that plaque vascularisation occurs in the presence of multiple angiogenically active factors. Knowledge of their combined effects could help in the formulation of novel therapeutics designed to stabilise or prevent their formation in the treatment of atherosclerosis.
Assuntos
Proteínas Angiogênicas/genética , Estenose das Carótidas/genética , Dissecação/instrumentação , Perfilação da Expressão Gênica/métodos , Marcadores Genéticos , Lasers , Neovascularização Fisiológica/genética , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase , Proteínas Adaptadoras de Transdução de Sinal , Idoso , Proteínas Angiogênicas/análise , Angiopoietina-1/genética , Antígenos CD/análise , Proteínas de Ligação ao Cálcio , Artérias Carótidas/química , Artérias Carótidas/imunologia , Artérias Carótidas/cirurgia , Estenose das Carótidas/imunologia , Estenose das Carótidas/fisiopatologia , Estenose das Carótidas/cirurgia , Endarterectomia das Carótidas , Endoglina , Endostatinas/genética , Humanos , Imuno-Histoquímica , Peptídeos e Proteínas de Sinalização Intercelular/genética , Masculino , Pessoa de Meia-Idade , Receptor para Produtos Finais de Glicação Avançada , Receptor Notch3 , Receptor TIE-2/genética , Receptores de Superfície Celular/análise , Receptores Imunológicos/genética , Receptores Notch/genética , Ruptura , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/genéticaRESUMO
We have studied the expression of Na+-D-glucose cotransporter in brush-border membrane vesicles (BBMVs) of chicken enterocytes to correlate the changes in the apical Na+-dependent transport with the changes in the amounts of transporter determined by Western blot analysis. Two different rabbit polyclonal antibodies were used simultaneously. The antibody raised against amino acids 564-575 of the deduced amino acid sequence of rabbit intestinal SGLT-1 (antibody 1) specifically detects a single 75-kDa band in the three segments, and this band disappeared when the antibody was preabsorbed with the antigenic peptide. The antibody raised against the synthetic peptide corresponding to amino acids 402-420 of the same protein (antibody 2) only reacts with jejunal and ileal samples, but no signal is found in BBMVs of rectum. Only when antibody 1 was used was there a linear correlation between the maximal transport rates of hexoses in BBMVs and the relative protein amounts determined by Western blot. These results indicate that the Na+-D-glucose cotransport in the jejunum, the ileum, and the rectum of chickens is due to an SGLT-1 type protein.
Assuntos
Mucosa Intestinal/metabolismo , Glicoproteínas de Membrana/metabolismo , Proteínas de Transporte de Monossacarídeos/metabolismo , Animais , Anticorpos/farmacologia , Transporte Biológico/efeitos dos fármacos , Western Blotting , Immunoblotting , Masculino , Glicoproteínas de Membrana/imunologia , Metilglucosídeos/farmacocinética , Microvilosidades/metabolismo , Proteínas de Transporte de Monossacarídeos/imunologia , Coelhos , Transportador 1 de Glucose-SódioRESUMO
Na(+)-dependent D-glucose uptake was studied in jejunal brush-border membrane (BBM) vesicles of chickens at 2 days and 1, 2, 5-6, and 12-14 wk of age. Both initial rates and accumulation ratios of the Na(+)-dependent D-glucose transport were significantly higher during the 1st wk than at other ages. To explain the age-related changes observed in the transport of D-glucose, the phlorizin-specific binding, Na+ permeability, lipid composition, and fluidity were studied. Transporter site density was quantified using 50 mumol/l phlorizin and found to be higher during the 1st wk. During the 2nd wk it decreased and then remained constant. Permeability of Na+, studied using 22Na+, showed that fluxes were similar during the first 6 wk, and a significant decrease was observed in the oldest group. Furthermore, membrane fluidity results showed a significant age-dependent decrease that correlated well with both the increased molar ratio of cholesterol to phospholipid and the decreased ratio of lipid to protein found during development. In conclusion, changes in the density of Na(+)-dependent D-glucose transporter as well as in lipid content and fluidity might be involved in the changes observed in D-glucose uptake during the posthatching development.