ABSTRACT
@#[Abstract] Objective: To study the effect of Notch4 signaling pathway on the self-renewal capacity of cancer stem cells (CSCs) of oral squamous cell carcinoma PJ15 and PJ41 cells and its mechanism. Methods: The expression of Notch4 gene in head and neck tumors was analyzed using TCGA database. 2 μmol/L cisplatin was used to treat oral squamous cell carcinoma PJ15 and PJ41 cells for 48 h, following with the treatment of Notch pathway inhibitor DAPT for 24 h. Flow cytometry was used to detect the ratio of CSCs, Western blotting and qPCR were used to detect the expressions of CSCs markers (Sox2, Bmi-1, Oct4, Nanog) and downstream genes in Notch4 signaling pathway (JAG1, HEY1, HEY2, HES1, HES2, DLL4, etc.). Notch4 was knocked down by siRNA technology. Notch4 was silenced with siRNA technology. Western blotting and qPCR were used to detect the effect of si-Notch4 and cisplatin on the expression level of CSCs markers. The stem cell pelletingtestwasusedtodetecttheself-renewalabilityof CSCs. Results: Notch4 gene was highly expressed in head and neck tumor tissues (P=0.046). After cisplatin treatment, compared with the control group, expressions of NICD4 and CSCs markers (Sox2, Bmi-1, Oct4, Nanog) as well as downstream genes of Notch4 signaling pathway (JAG1, HEY1, HEY2, HES1,HES2, DLL4, etc.) increased significantly (all P<0.05), and the proportion of ALDH1 positive cells increased significantly (P<0.05); with the further addition of DAPT, the expressions of the above genes decreased significantly (P<0.05 or P<0.01). After knocking down Notch4, compared with the control group, the size and number of spheres of PJ15 cells [1.33±0.47] vs [8.00±0.82], P<0.01) and PJ41 cells ([1.00±0.82] vs [7.67±1.25], P<0.01) were significantly lower than that of the control group; after the addition of 2 μmol/L DDP for 24 h, there was no significant difference in the expressions of Nanog and Bmi-1 gene between si-Notch4 group and control group. Conclusion:Activation of Notch4 signaling pathway can enhance the self-renewal ability of CSCs in oral squamous cell carcinoma PJ15 and PJ41 cells.
ABSTRACT
Objective To investigate the effects of RhoC on biological behavior in the laryngeal squamous carcinoma.Methods By delivering exogenous gene into Hep2 laryngeal squamous carcinoma cell line,we alternatively repressed and strengthened the expression of RhoC.We tested the apoptosis of Hep2 tumor cell line with TUNEL,visualized tumor cells shape by staining cell skeleton with Alexa fluor phalloidin,measured the mRNA of CSC marker ALDH1A1 with QPCR.Results After repressing the expression of RhoC in Hep2 cell line,the apoptosis of cancer cells was elevated,the expression of CSC marker ALDH1A1 was significantly decreased.RhoC impacted the shapes of Hep2.Conclusion RhoC havd a positive role in LSCC metastasis.RhoC is a promising target of anti-metastases in LSC.
ABSTRACT
Objective To compare effect of chemotherapy agent DDP to MACS in sorting cancer stemcells (CSC) of laryngeal carcinoma cell line Hep-2. Methods CD133 magnetic beads were applied to sort Hep-2 cells. Different dosages of DDP were used to treat Hep-2 cells for 48 hours. Enrichment rate of CD133+ cells by MACS and after DDP treatment was detected by Flow Cytometer (FCM). Morphologic change was observed under inverse-phase microscope. Results FCM showed that the sorting rate of CD133+ cells through MACS was 64.33 %, while after DDP treatment for 48 hours, the rate of CD133+ cells was enriched significantly in each dosage of DDP, with the maximal rate was 50.7 %, in the dosage of 4 μg/ml. There was a significantly difference between MACS and each of DDP group (P <0.01). Cells treated with DDP were abnormal in morphology. Conclusion MACS and DDP sorting has respective advantages in enriching CSC in Hep-2 cell lines.