Effects Of Unfolded Protein Response On Sensitivity Of Human Oral Squamous Cell Carcinoma To Chemothrapy

Aims:1. To investigate the protein GRP78expression in oral cancer tissue and therelationship between clinical pathological features and prognosis.2. To investigate the effects of the protein GRP78expression in the sensitivity of theoral cavity cancer cells to chemotherapy drugs.Methods:1. The statistical analysis of the relationship between GRP78expression and itsclinical pathological characteristics and prognosis;2. MTT method to analysis the effect of drug on cell proliferation inhibition;3. Colony clone formation method to detect the drug to cell colony clone formationof influence;4. The cell apoptosis was detected by flow cytometry with propidium iodidestaining.5. Caspase-3active detection kit detection drug effect within24h caspase-3activitychange;6. Western blot test caspase-3caspase-12protein expression and protein expressionof glucose regulation78.7. After transfect ion with a small interfering RNA-mediated GRP78(pSH1Si-GRP78), the apoptosis of the cells were detected with PI staining, theexpression of apoptotic proteins were detected by western blot.Results:1. The relationship between GRP78expression and its clinical pathologicalcharacteristics and prognosis 1) Patients with positive expression of GRP78has related with tumor size、 lymphnode metastasis、distant metastasis、clinical stage and pathological differentiation. Ithas no related with age, sex, smoking and drinking.2) The survival of patients with positive GRP78expression is lower than thenegative expression.3) The Cox model analysis that age, distant metastasis, GRP78positive expressionare risk factors that affects the survival in patients with oral squamous carcinoma.2. The different concentrations of tunicamycin and/or oxaliplatin on oralcancer KB cells proliferation and apoptosis1) Tunicamycin can enhance oxaliplatin into the oral cancer KB inhibition of cellproliferation.0.125μ mol·L-1tunicamycin handle oral cancer KB cells24,48,72h after thecell survival rate of90.37%、89.44%、and88.91%respectively.0.125μ mol·L-1tunicamycin combined with1μ mol·L-1oxaliplatin on oral cancer KB cells24,48,72h survival rate lower than tunicamycin, oxaliplatin into groups is50.78%、37.77%、23.24%,respectively.2) Tunicamycin can enhance oxaliplatin into oral cancer KB cell inhibition of cloneformation0.25μ mol·L-1tunicamycin can enhance on oxaliplatin on oral cancer cells KBcolony clone formation action.3) Tunicamycin can enhance oxaliplatin into induction effect of oral cancer KB cellapoptosis0.5μ mol·L-1tunicamycin induced KB cells48h apoptosis rate were8.3%.0.5μ mol·L-1tunicamycin and1μ mol·L-1oxaliplatin stimulate human oral cancer cellsKB48h apoptosis rate was50.3%, higher than only oxaliplatin which inducedapoptosis rate was24.6%.4) Tunicamycin can enhance oxaliplatin into activation of Caspase3in oral cancerKB cells.5) The effects of Tunicamycin and/or oxaliplatin can induce the expression ofGRP78, Caspase-12and Caspase-3. 3. Kockdown GRP78can increase the apoptosis induced by oxaliplatin into oralcancer cell.1) KB was treated with different sequences of recombinant plasmid, and after6h,cell lysates were examined by western blotting. β-actin was used as a loadingcontrol. Expression of GRP78was significantly reduced by GRP78-homo-659.2) KB was treated with GRP78-siRNA or control siRNA for6h, then treated withoxaliplatin24h and cell apoptosis rate was increased examined by PI staining.kockdown GRP78can increase the apoptosis induced by oxaliplatin into oral cancercell.Conslusion:1. The expression level of protein GRP78in tumor tissue is significantly higher thanthe expression in normal tissue2. The level of GRP78protein expression in tumor tissue has correlation whitclinical pathologic features and prognosis which can serve as a potential tumormarker to determine the malignant degree and predicting the prognosis of tumor.3. The excessive endoplasmic reticulum stress on the effect on promoting apoptosisof tumor cells is greater than the protection4. Kockdown GRP78can increase the apoptosis induced by oxaliplatin into oralcancer cell.