RESUMO
<p><b>OBJECTIVE</b>Isolation and expansion tumor spheres from colorectal cancer cell line Colo205 cultured in serum-free medium(SFM) supplemented with human recombinant EGF and bFGF.</p><p><b>METHODS</b>Colo205 cells were cultivated in SFM,while cells cultivated in serum-supplemented medium(SSM) served as the control. Cells morphology were observed by optical microscope, and expression of intestinal stem cells marker Musashi-1 was detected by immunocytochemical. To induce cell differentiation, tumour spheres were cultivated without EGF and bFGF in the presence of 10% serum. Then we analysed expressions of stem cell surface markers CD133 and CD44 among undifferentiated cell, post-differentiated cells and routine Colo205 cells under serum-supplemented culture condition by flow cytometry. At last we compared cell cycle and spectral karyotype between two groups.</p><p><b>RESULTS</b>In SFM consisting of EGF and bFGF, a minority of Colo205 cells could survive, proliferate and form the suspended tumor spheres. We detected high Musashi-1 expression in these cells. Compared with the SSM group and the post-differentiation SFM group, the expressions of CD133 and CD44 were significantly increased in the undifferentiated SFM group (P<0.05). There was no statistical difference in the expression of CD133 and CD44 between the post-differentiation SFM group and the SSM group (P>0.05). Cell cycle analysis indicated that tumor spheres were of a high proliferation state.We could not find any noticeable difference in the number of chromatosomes between the SFM group and the SSM group.</p><p><b>CONCLUSION</b>Tumor spheres in which enriched cancer stem cells can be generated under serum-free culture condition with EGF and bFGF.</p>
Assuntos
Humanos , Antígeno AC133 , Antígenos CD , Metabolismo , Técnicas de Cultura de Células , Métodos , Linhagem Celular Tumoral , Proliferação de Células , Meios de Cultura Livres de Soro , Glicoproteínas , Metabolismo , Receptores de Hialuronatos , Metabolismo , Células-Tronco Neoplásicas , Biologia Celular , Metabolismo , Proteínas do Tecido Nervoso , Metabolismo , Peptídeos , Metabolismo , Proteínas de Ligação a RNA , Metabolismo , Esferoides Celulares , Biologia CelularRESUMO
<p><b>OBJECTIVE</b>To evaluate the role of p53 gene mutation in colorectal carcinoma and assess the value of peripheral blood p53 single nucleotide polymorphism (SNP) in early diagnosis of colorectal carcinoma.</p><p><b>METHODS</b>NSP in axons 5-8 of p53 gene was detected using ligase detection reaction-polymerase chain reaction (LDR-PCR) in the peripheral blood of 100 patients with colorectal cancer and 100 healthy subjects.</p><p><b>RESULTS</b>The mutation rate of p53 gene was 24% (24/120) in colorectal carcinoma patients and 0% (0/120) in the healthy subjects (P<0.05).</p><p><b>CONCLUSION</b>p53 plays an important role in the carcinogenesis of colorectal carcinoma, and SNP analysis for p53 gene can be helpful in early diagnosis of colorectal carcinoma.</p>
Assuntos
Adulto , Feminino , Humanos , Masculino , Estudos de Casos e Controles , Neoplasias Colorretais , Diagnóstico , Genética , Análise Mutacional de DNA , DNA de Neoplasias , Genética , Detecção Precoce de Câncer , Mutação , Polimorfismo de Nucleotídeo Único , Proteína Supressora de Tumor p53 , GenéticaRESUMO
<p><b>OBJECTIVE</b>To determine whether the eukaryotic initiation factor-4E (eIF-4E) is involved in the cap-dependent translational regulation of heparanase and study the correlation between heparanase expression and metastatic potential of LS-174T cells.</p><p><b>METHODS</b>The protein and mRNA levels of inhibited eIF-4E were tested by Western blot and RT-PCR. Heparanase activity was defined as the ability to degrade high molecular weight (40-100 000) radiolabeled ((35)S) heparan sulfate (HS) substrate into low molecular weight (5-15 000) HS fragments. The invasive potential of tumor cells in vitro was observed by Matrigel invasion assay system.</p><p><b>RESULTS</b>The 20-mer antisense oligonucleotide (asODN) against eIF-4E specifically and significantly inhibited eIF-4E expression at both transcriptional and translational levels. The expression and the activity of heparanase were effectively lowered, which further decreased the invasive potential of LS-174T.</p><p><b>CONCLUSION</b>eIF-4E, probably being involved in translational regulation of heparanase in colon adenocarcinoma cell line LS-174T, can be a particularly interesting target for heparanase regulation, based on of its critical function.</p>