| Objective:It shows that cancer stem cells (CSCs) are of self-renewing ability, and mostlyconsidered to be involved in recurrence, drug resistance, metastasis and root growthof malignant tumor. The purpose of this study is to dissociate gastric cancer stem cells(GCSCs) from human poor-differentiated gastric cancer cell line MGC803, and thenshow their characteristics of gastric cancer stem like cells (GCSLCs). We show thattreatment of GCSLCs with diallyl disulfide (DADS) inhibits proliferation, which maybe concerned with Shh and Gli1downregulation induced by DADS.Methods:1. MGC803cells were cultured in the so-called serum free media to form spheres.The purity of CD44and CD24were detected by flow cytometry. GCSLCs wereobserved under a fluorescence microscope. MTT and Soft agar colony formationassay was conducted to identify the propagation ability of GCSLCs. MGC803cellproliferation. Invasion ability of GCSLCs was performed by invasion assay. RT-PCRand Western blot were used to verify mRNA and protein expression of Shh and Gli1in MGC803cells and GCSLCs treated or untreated by DADS respectively.2. Treated by30mg/L DADS to MTT and soft agar colony formation assay wasconducted to identify the role of DADS on the propagation ability of GCSLCs.RT-PCR and Western blot were performed to analyse the effect of DADS on mRNAand protein expression of Shh and Gli1in MGC803cells and GCSLCs respectively.Results:1. Also only a subpopulation of MGC803cells could form floating spheres in serumfree media. The tumor cells were spherical growth, forming a typical tumor spheres.The percentage of CD24in GCLCs was91.21%, much higher than that in gastriccancer MGC803cells (45.56%); meanwhile, the percentage of CD44in GCLCs was 91.93%, higher than that in MGC803cells (82.99%).2. Under the fluorescence microscope, stronger green fluorescence of both CD44andCD24is showed on cell membrane of most GCLCs than that of MGC803cells.3. Soft agar colony formation assay shows that colony formation ability in GCLCswas better than that in MGC803cells (P<0.05). CLCs exceeded MGC803cells in thenumber and size of colony formation4. RT-PCR showed that Shh gene and Gli1gene expressions in GCLCs weresignificantly higher than those in gastric cancer MGC803cells (P<0.05). Shh andGli1protein expressions also increased in GCLCs, compared to MGC803cells(P<0.05).5. After treated with30mg/L DADS, OD value was significantly lower than theuntreated in GCLCs and MGC803cells respectively, which showed that DADSinhibited GCLCs proliferation. The number of colony formation is far less, and thesize of colony formation became small after induced by DADS (P<0.05).6. DADS decreased mRNA and protein expressions of Shh and Gli1in both MGC803cells and GCLCs (P<0.05)Conclusion:1．GCLCs were isolated and identified from human gastric cancer cell lineMGC803.2．DADS inhibited proliferation of MGC803cells and GCLCs, which was relatedto suppression of Shh and Gli1expression. |
PDF Full Text Request