RESUMEN
Mucopolysaccharidosis type I (MPS I) is a multisystemic disorder caused by the deficiency of alpha-L-iduronidase (IDUA) that leads to intracellular accumulation of glycosaminoglycans (GAG). In the present study we aimed to use cationic liposomes carrying the CRISPR/Cas9 plasmid and a donor vector for in vitro and in vivo MPS I gene editing, and compare to treatment with naked plasmids. The liposomal formulation was prepared by microfluidization. Complexes were obtained by the addition of DNA at +4/-1 charge ratio. The overall results showed complexes of about 110â¯nm, with positive zeta potential of +30â¯mV. The incubation of the complexes with fibroblasts from MPS I patients led to a significant increase in IDUA activity and reduction of lysosomal abnormalities. Hydrodynamic injection of the liposomal complex in newborn MPS I mice led to a significant increase in serum IDUA levels for up to six months. The biodistribution of complexes after hydrodynamic injection was markedly detected in the lungs and heart, corroborating the results of increased IDUA activity and decreased GAG storage especially in these tissues, while the group that received the naked plasmids presented increased enzyme activity especially in the liver. Furthermore, animals treated with the liposomal formulation presented improvement in cardiovascular parameters, one of the main causes of death observed in MPS I patients. We conclude that the IDUA production in multiple organs had a significant beneficial effect on the characteristics of MPS I disease, which may bring hope to gene therapy of Hurler patients.
Asunto(s)
Edición Génica , Terapia Genética , Mucopolisacaridosis I/genética , Mucopolisacaridosis I/terapia , Animales , Sistemas CRISPR-Cas , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Femenino , Fibroblastos/efectos de los fármacos , Humanos , Iduronidasa/metabolismo , Liposomas , Masculino , Ratones Endogámicos C57BL , Mucopolisacaridosis I/metabolismo , Distribución TisularRESUMEN
Background: Type 2 Diabetes Mellitus is a multifactorial syndrome with severe complications. Oxidative stress is accepted as a causal factor of chronic complications. Aim: To demonstrate alterations in oxidative stress after metabolic surgery. Methods: Twenty-four 2-day-old Wistar rats were used. In 16, Type 2 Diabetes Mellitus was induced by 100 mg/kg streptozotocin injection. The development of diabetes was confirmed after 10 weeks using an oral glucose tolerance test. Eight diabetic rats composed the diabetic surgical group; the remaining eight composed the diabetic group. Eight animals in which diabetes was not induced formed the clinical control group. The Marchesini technique was used in the diabetic surgical group. After 90 days, the rats were sacrificed, and the oxidative stress markers were measured. Results: Thiobarbituric acid reactive substances, superoxide dismutase and catalase were significantly reduced in the diabetic surgical group compared to the diabetic group. Conclusion: The duodenojejunostomy was effective in controlling the exacerbated oxidative stress present in diabetic rats.
Racional: Diabete melito tipo 2 é síndrome multifatorial com complicações graves. O estresse oxidativo é aceito como um fator causal de complicações crônicas. Objetivo: Demonstrar alterações no estresse oxidativo após a cirurgia metabólica. Métodos: Foram utilizados 24 ratos Wistar de dois dias de idade. Em 16, diabete melito tipo 2 foi induzida por 100 mg/kg de injeção de estreptozotocina. O desenvolvimento do diabete foi confirmado após 10 semanas, utilizando teste oral de tolerância à glucose. Oito ratos diabéticos compuseram o grupo cirúrgico diabético; os oito restantes constituíram o grupo diabético. Oito animais em que não foi induzido o diabete formaram o grupo controle clínico. A técnica de Marchesini foi utilizada no grupo cirúrgico diabético. Após 90 dias, os ratos foram sacrificados, e os marcadores de estresse oxidativo foram medidos. Resultados: Ácido tiobarbitúrico, superóxido dismutase e catalase foram significativamente reduzidos no grupo cirúrgico diabético quando comparado ao grupo diabético. Conclusão: O duodenojejunostomia foi eficaz no controle do estresse oxidativo exacerbado presente em ratos diabéticos.
RESUMEN
ABSTRACT Background: Type 2 Diabetes Mellitus is a multifactorial syndrome with severe complications. Oxidative stress is accepted as a causal factor of chronic complications Aim: To demonstrate alterations in oxidative stress after metabolic surgery. Methods: Twenty-four 2-day-old Wistar rats were used. In 16, Type 2 Diabetes Mellitus was induced by 100 mg/kg streptozotocin injection. The development of diabetes was confirmed after 10 weeks using an oral glucose tolerance test. Eight diabetic rats composed the diabetic surgical group; the remaining eight composed the diabetic group. Eight animals in which diabetes was not induced formed the clinical control group. The Marchesini technique was used in the diabetic surgical group. After 90 days, the rats were sacrificed, and the oxidative stress markers were measured. Results: Thiobarbituric acid reactive substances, superoxide dismutase and catalase were significantly reduced in the diabetic surgical group compared to the diabetic group. Conclusion: The duodenojejunostomy was effective in controlling the exacerbated oxidative stress present in diabetic rats.
RESUMO Racional: Diabete melito tipo 2 é síndrome multifatorial com complicações graves. O estresse oxidativo é aceito como um fator causal de complicações crônicas. Objetivo: Demonstrar alterações no estresse oxidativo após a cirurgia metabólica. Métodos: Foram utilizados 24 ratos Wistar de dois dias de idade. Em 16, diabete melito tipo 2 foi induzida por 100 mg/kg de injeção de estreptozotocina. O desenvolvimento do diabete foi confirmado após 10 semanas, utilizando teste oral de tolerância à glucose. Oito ratos diabéticos compuseram o grupo cirúrgico diabético; os oito restantes constituíram o grupo diabético. Oito animais em que não foi induzido o diabete formaram o grupo controle clínico. A técnica de Marchesini foi utilizada no grupo cirúrgico diabético. Após 90 dias, os ratos foram sacrificados, e os marcadores de estresse oxidativo foram medidos. Resultados: Ácido tiobarbitúrico, superóxido dismutase e catalase foram significativamente reduzidos no grupo cirúrgico diabético quando comparado ao grupo diabético. Conclusão: O duodenojejunostomia foi eficaz no controle do estresse oxidativo exacerbado presente em ratos diabéticos.