RESUMO
This study aimed to investigate the effect of RNAi-mediated silencing of the Livin gene on biological properties of the colon cancer cell line LoVo. Interference vectors pSilencer4.1-Ll and pSilencer4.1-L2 targeting the Livin gene were constructed and transfected into LoVo cells. The expression of the Livin gene was determined by RT-PCR and Western blotting. The apoptosis, cell cycle, colony formation, proliferation of LoVo cells, as well as their sensitivity to cisplatin, were detected by flow cytometry, colony formation assay and MTT. Livin mRNA and protein expression in LoVo cells could be effectively silenced by pSilencer4.1-Ll but not pSilencer4.1-L2. In the pSilencer4.1-Ll transfection group, the apoptosis rate of LoVo cells was significantly higher than in the control group (24.2 ± 3.2 vs 8.1 ± 1.4%, P < 0.01), and after 72 h, cell proliferation was clearly decreased (about 70% inhibition). Compared with the control group, the colony formation rate in pSilencer4.1-Ll transfection group was obviously decreased (15 ± 4.6 vs 85 ± 5.8%, P < 0.01), with increased proportion of S phase cells (45.7 ± 4.9 vs 28.0 ± 3.0%, P < 0.01), decreased proportion of G1 phase cells (43.0 ± 5.2 vs 62.8 ± 5.1%, P < 0.01), and increased sensitivity to cisplatin (apoptosis rate increased from 43.4 ± 6.9 to 65.3 ± 6.2%, P < 0.01). pSilencer4.1-Ll can effectively silence Livin gene expression in LoVo colon cancer cells, inhibit cell proliferation and colony formation, induce apoptosis, and enhance sensitivity to cisplatin.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proliferação de Células/genética , Neoplasias do Colo/genética , Proteínas Inibidoras de Apoptose/genética , Proteínas de Neoplasias/genética , Interferência de RNA , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/biossíntese , Apoptose/genética , Ciclo Celular/efeitos dos fármacos , Cisplatino/administração & dosagem , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/patologia , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Proteínas Inibidoras de Apoptose/antagonistas & inibidores , Proteínas Inibidoras de Apoptose/biossíntese , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/biossíntese , RNA Mensageiro/biossínteseRESUMO
Livin, a novel member of the human inhibitors of apoptosis protein (IAP) family, plays an important role in tumor progression and occurrence by inhibiting cell apoptosis. It is selectively expressed in the most common human neoplasms and appears to be involved in tumor cell resistance to chemotherapeutic agents. To investigate its possibility as a therapeutic target for human malignancies, we established two genetically different stable tumor cell lines (LoVo and SPCA-1) and RNA interference (RNAi) technique was employed to downregulate Livin expression in two human tumor cell lines. The specific downregulation of Livin expression in tumor cell lines significantly inhibited in vitro cell proliferation and in vivo tumorigenicity. Furthermore, Livin knockdown led to cell arrest in the G(1)/G(0) phase of cell cycle, eventual apoptosis and chemosensitivity enhancement in tumor cells. All these results indicate that RNAi-mediated downregulation of Livin expression can lead to potent antitumor activity and chemosensitizing effects in human cancers.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Proliferação de Células , Inativação Gênica , Proteínas Inibidoras de Apoptose/fisiologia , Proteínas de Neoplasias/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Apoptose/genética , Apoptose/fisiologia , Western Blotting , Ciclo Celular/genética , Ciclo Celular/fisiologia , Linhagem Celular Tumoral , Feminino , Citometria de Fluxo , Vetores Genéticos/genética , Humanos , Proteínas Inibidoras de Apoptose/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/terapia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Proteínas de Neoplasias/genética , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The changes of plasma concentration of estradiol (E2), progesterone (P) and prostaglandins (PG) and decidual level of prostaglandins and progesterone receptor (PR) after injection of Yellow Daphne for induction of abortion were observed. The results showed that the treatment with Yellow Daphne could induce degeneration and necrosis of decidual tissues as well as increase of PGE2 and PGF2 alpha concentration with lowering of PR level in the tissue. On the other hand, plasma level of PGE2 and PGF2 alpha remained unchanged and the levels of E2 and P declined. The data suggested that the contragestive mechanism of Yellow Daphne was mainly the involvement of decidual degeneration and the release of PG thus interfered the intrauterine environment for maintenance of pregnancy which subsequently led to abortion.