RESUMO
Glucagon-like peptide 1 (GLP-1) is a promising candidate for the treatment of type II diabetes. However, the short in vivo half-life of GLP-1 has made peptide-based treatments challenging. Gene therapy aimed at achieving continuous GLP-1 expression presents one way to circumvent the rapid turnover of GLP-1. We have created a GLP-1 minigene that can direct the secretion of active GLP-1 (amino acids 7-37). Plasmid and adenoviral expression vectors encoding the 31-amino-acid peptide linked to leader sequences required for secretion of GLP-1 yielded sustained levels of active GLP-1 that were significantly greater than endogenous levels. Systemic administration of expression vectors to animals using two diabetic rodent models, db/db mice and Zucker Diabetic Fatty (ZDF) rats, yielded elevated GLP-1 levels that lowered both the fasting and random-fed hyperglycemia present in these animals. Because the insulinotropic actions of GLP-1 are glucose dependent, no evidence of hypoglycemia was observed. Improved glucose homeostasis was demonstrated by improvements in %HbA1c (glycated hemoglobin) and in glucose tolerance tests. GLP-1-treated animals had higher circulating insulin levels and increased insulin immunostaining of pancreatic sections. GLP-1-treated ZDF rats showed diminished food intake and, in the first few weeks following vector administration, a diminished weight gain. These results demonstrate the feasibility of gene therapy for type II diabetes using GLP-1 expression vectors.
Assuntos
Diabetes Mellitus Tipo 2/terapia , Terapia Genética/métodos , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Células Secretoras de Insulina/metabolismo , Adenoviridae/genética , Animais , Diabetes Mellitus Tipo 2/metabolismo , Feminino , Expressão Gênica , Engenharia Genética , Vetores Genéticos/administração & dosagem , Peptídeo 1 Semelhante ao Glucagon/análise , Peptídeo 1 Semelhante ao Glucagon/genética , Teste de Tolerância a Glucose , Hemoglobinas Glicadas/análise , Insulina/análise , Insulina/sangue , Células Secretoras de Insulina/química , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Obesos , Plasmídeos/administração & dosagem , Ratos , Ratos Zucker , Transdução Genética/métodos , Transfecção/métodosRESUMO
We have sequenced a cDNA clone for the Drosophila melanogaster gene Dsrc28C, a homolog of the vertebrate gene c-src. The cDNA contains a single open reading frame encoding a protein of 66 kilodaltons which contains features highly conserved within the src family of tyrosine protein kinases. Novel structural features of the Dsrc28C protein include a basic pI and a polyglycine domain near the amino terminus. Cell-free translation of in vitro-transcribed RNA yielded a protein of the predicted size which could be immunoprecipitated by anti-v-src antisera. RNA blot hybridization revealed that the gene is expressed predominantly during embryogenesis, in imaginal disks of third-instar larvae, and in adult females. In situ hybridization showed that expression in adult females is largely confined to nurse cells and developing oocytes.
