RESUMO
BACKGROUND: Mesenchymal stem cells (MSCs) have been considered to be the attractive vehicles for delivering therapeutic agents toward various tumor diseases. This study was to explore the distribution pattern, kinetic delivery of adenovirus, and therapeutic efficacy of the MSC loading of E1A mutant conditionally replicative adenovirus Adv-Stat3(-) which selectively replicated and expressed high levels of anti-sense Stat3 complementary DNA in breast cancer and melanoma cells. METHODS: We assessed the release ability of conditionally replicative adenovirus (CRAd) from MSC using crystal violet staining, TCID(50) assay, and quantitative PCR. In vitro killing competence of MSCs carrying Adv-Stat3(-) toward breast cancer and melanoma was performed using co-culture system of transwell plates. We examined tumor tropism of MSC by Prussian blue staining and immunofluorescence. In vivo killing competence of MSCs carrying Adv-Stat3(-) toward breast tumor was analyzed by comparison of tumor volumes and survival periods. RESULTS: Adv-Stat3(-) amplified in MSCs and were released 4 days after infection. MSCs carrying Adv-Stat3(-) caused viral amplification, depletion of Stat3 and its downstream proteins, and led to significant apoptosis in breast cancer and melanoma cell lines. In vivo experiments confirmed the preferential localization of MSCs in the tumor periphery 24 hours after tail vein injection, and this localization was mainly detected in the tumor parenchyma after 72 hours. Intravenous injection of MSCs carrying Adv-Stat3(-) suppressed the Stat3 pathway, down-regulated Ki67 expression, and recruited CD11b-positive cells in the local tumor, inhibiting tumor growth and increasing the survival of tumor-bearing mice. CONCLUSIONS: These results indicate that MSCs migrate to the tumor site in a time-dependent manner and could be an effective platform for the targeted delivery of CRAd and the amplification of tumor killing effects.
Assuntos
Adenoviridae/genética , Neoplasias da Mama/patologia , Melanoma/patologia , Células-Tronco Mesenquimais/citologia , Animais , Apoptose , Neoplasias da Mama/terapia , Linhagem Celular Tumoral , Sobrevivência Celular , Técnicas de Cocultura , Feminino , Imunofluorescência , Humanos , Antígeno Ki-67/genética , Antígeno Ki-67/metabolismo , Melanoma/terapia , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos Nus , Neoplasias/patologia , Neoplasias/terapia , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Replicação ViralRESUMO
Membrane-type 1 matrix metalloproteinase (MT1-MMP/MMP-14) is a key enzyme involved in degradation of extracellular matrix (ECM) and various surface-associated proteins that control cell growth, differentiation and survival, plays crucial roles in molecular carcinogenesis, tumor cell growth, invasion, and angiogenesis. We tested the inhibitory effect of antisense MT1-MMP on the ability of metastatic human ovarian carcinoma cell line SW626 in proliferation and invasion. RT-PCR was used to amplify MT1-MMP cDNA fragments with two different restriction sites at its 5'-end. Antisense MT1-MMP cloned in eukaryotic expression vector pMMP14as was transfected into SW626 cells. MT1-MMP protein expression, activities of MMP-2 and MMP-9, changes of cell proliferation, and cell invasion ability were detected by Western blot, optimized gelatin zymography, MTT assay and matrigel in vitro invasion assay, respectively. After 48 h transfection, decreased expression of endogenous MT1-MMP protein was detected in pMMP14as-transfected SW626 cells and showed significantly lower proliferation level when compared with control cells. The activation of proMMP-2 was inhibited markedly, and the mean percentage of invasive cells was 63.30+/-5.80% in pMMP14as-transfected cells, which was less than that (97.60+/-7.50%) in control cells (P<0.05). Both cell proliferation and invasion in SW626 cells were inhibited effectively by antisense MT1-MMP transfection, suggesting that MT1-MMP may be a proper target molecule for anti-invasion therapy for human ovarian cancers.
Assuntos
Metaloproteinases da Matriz/metabolismo , Invasividade Neoplásica/patologia , Neoplasias Ovarianas/enzimologia , Neoplasias Ovarianas/patologia , Western Blotting , Proliferação de Células , Regulação para Baixo , Feminino , Humanos , Metaloproteinases da Matriz Associadas à Membrana , Reação em Cadeia da Polimerase Via Transcriptase Reversa , TransfecçãoRESUMO
The purpose of this study was to clarify the direct effect of hypoxia-inducible factor-1α (HIF-1α) on tumor growth, apoptosis and migration in vitro. To achieve this aim, a comparison was made of the differences in growth rates, apoptotic indices and cell invasive ability in the human cervical cancer cell line HeLa and the HIF-1α-blocked counterpart in a three-dimensional spheroid culture. A significant decrease in cell proliferation and invasion, and an increase in cell apoptosis were observed in HIF-1α-blocked cells in the three-dimensional culture. The data indicated that a multicellular spheroid culture is an ideal model of hypoxia in vitro and that HIF-1α is a significant regulator of adaptive processes that promote tumor cell malignant phenotypes, such as proliferation, anti-apoptosis and invasive ability.