RESUMO
Transforming growth factor (TGF)-beta1 is an important fibrogenic factor that is involved in the pathogenesis of diabetic nephropathy. We evaluated the effect of circular antisense TGF-beta1 oligodeoxynucleotides (ODNs) on the TGF-beta1 expression in the rat mesangial cell culture and in streptozotocin (STZ)-induced diabetic rats. Circular antisense TGF-beta1 ODNs were found to be stable in rat serum, significantly decreasing TGF-beta1 mRNA expression compared with linear antisense ODNs in the rat mesangial cell culture. Circular antisense TGF-beta1 ODNs were introduced into the tail vein of normal rats using hemagglutinating virus of Japan (HVJ)-liposome-mediated gene transfer method and were confirmed to be delivered effectively into the kidney, liver, lungs, and spleen. To inhibit the overexpression of TGF-beta1 in diabetic kidneys, we introduced circular antisense TGF-beta1 ODNs into the STZ-induced diabetic rats. On day 13 after circular antisense TGF-beta1 ODNs injection, TGF-beta1 mRNA and protein expression markedly decreased and urinary TGF-beta1 excretion rate also dropped in the circular antisense TGF-beta1 ODNs-treated diabetic rats. These results suggest that circular antisense TGF-beta1 ODNs may be a useful tool for developing new therapeutic application for progressive diabetic nephropathy.
Assuntos
Animais , Masculino , Ratos , Glicemia/metabolismo , Células Cultivadas , Diabetes Mellitus Experimental/terapia , Ensaio de Imunoadsorção Enzimática , Técnicas de Transferência de Genes , Imuno-Histoquímica , Lipossomos/química , Microscopia Confocal , Oligonucleotídeos Antissenso/metabolismo , RNA/metabolismo , RNA Mensageiro/metabolismo , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Estreptozocina , Fatores de Tempo , Transfecção , Fator de Crescimento Transformador beta/genéticaRESUMO
Bcr-abl antisense oligodeoxynucleotides (AS-ODNs) have provided evidence of an antileukemia effect when tested in vitro against Philadelphia-positive cells. In order to investigate the efficacy of AS-ODNs as purging agents in chronic myeloid leukemia (CML) patients, K562 cells, a human CML cell line, were treated in vitro with various types of AS-ODNs and interferon-alpha. Cells were treated in vitro for 0 and 36 hr with 40 microgram/mL of AS-ODNs, respectively, and incubated at 37 degrees C for 36 hr. Cytotoxic effects were measured by counting the number of viable cells as well as by MTT test. Clonogenic activities were evaluated by methylcellulose culture for 2 weeks. The effects of purging agents on the rearrangement of bcrabl gene were evaluated by RT-PCR. AS-ODNs inhibited the proliferation of K562 cells with time in cell count assay and MTT test. AS-ODNs were superior to INF-alpha in inhibiting clonogenic activity (recovery rate; 26.3% vs 64.0%). After incubation with bcr-abl AS-ODNs primers and mRNA isolated from K562 cells, positive bands were abolished, especially of b3a2 type and phosphorothioate type. Our results suggest that AS-ODNs mediated purging may be one of the efficient methods and that autograft may be an alternative treatment for allograft in high-risk group patients of CML if they do not have a stem cell donor.
Assuntos
Humanos , Purging da Medula Óssea , Ensaio de Unidades Formadoras de Colônias , Proteínas de Fusão bcr-abl/genética , Proteínas de Fusão bcr-abl/metabolismo , Células-Tronco Hematopoéticas/fisiologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/terapia , Neuroblastoma/terapia , Oligonucleotídeos Antissenso/metabolismo , Oligonucleotídeos Antissenso/uso terapêutico , Transplante Autólogo , Células Tumorais CultivadasRESUMO
Peroxiredoxin II (Prx II) is known not only to protect cells from oxidative damage caused by hydrogen peroxide (H2O2), but also to endow cancer cells with resistance to both H2O2 and cisplatin and to grant them radioresistance. In this study, we examined whether Prx II antisense could enhance cisplatin-induced cell death. When gastric cancer cells were transfected with various concentrations of Prx II antisense plasmid, pPrxII/AS, and then treated with the same concentrations of cisplatin, Prx II antisense enhanced cisplatin-induced cell death. The combination index (CI) at all doses of the combination was below 1, indicating that Prx II antisense sensitized cisplatin-induced cell death. This synergism was also observed in the cells transfected with a Prx II antisense oligomer. Our present results, therefore, suggest that Prx II antisense would be a very good sensitizer for cisplatin, and that Prx II as a target for chemosensitizers constitutes a promising avenue for future research.
Assuntos
Humanos , Antineoplásicos/farmacologia , Antioxidantes/metabolismo , Apoptose/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Cisplatino/farmacologia , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Vetores Genéticos , Oligonucleotídeos Antissenso/metabolismo , Peroxidases/metabolismo , Plasmídeos/genética , Neoplasias Gástricas/metabolismo , Células Tumorais CultivadasRESUMO
Peroxiredoxin II (Prx II) is known not only to protect cells from oxidative damage caused by hydrogen peroxide (H2O2), but also to endow cancer cells with resistance to both H2O2 and cisplatin and to grant them radioresistance. In this study, we examined whether Prx II antisense could enhance cisplatin-induced cell death. When gastric cancer cells were transfected with various concentrations of Prx II antisense plasmid, pPrxII/AS, and then treated with the same concentrations of cisplatin, Prx II antisense enhanced cisplatin-induced cell death. The combination index (CI) at all doses of the combination was below 1, indicating that Prx II antisense sensitized cisplatin-induced cell death. This synergism was also observed in the cells transfected with a Prx II antisense oligomer. Our present results, therefore, suggest that Prx II antisense would be a very good sensitizer for cisplatin, and that Prx II as a target for chemosensitizers constitutes a promising avenue for future research.