Studies Of Mannose-binding Lectin On Proliferation, Apoptosis And The Expression Of Relative Protein Of Thyroid Carcinoma Cells

Ojectives:To cultivate thyroid papillary cancer cells(K1),thyroid follicular cancer cells (FTC-133), with different concentrations of mannose binding lectin.To investigate the cells proliferation and choose proper concentration and effect time we used the MTT.We also wanted to know whether MBL can promote the cells apoptosis by FCM.At the same time,we used Western blot to detect the protein expression of Bcl-2 and p53,and discussed the relationship between MBL and cells proliferation and apoptosis.To provide theoretical basis for preventing thyroid tumors.Methods:1 Cell culture:The thyroid papillary carcinoma (K1) cells were cultivated in containing 10% fetal bovine serum, 100u/ml penicillin and 100ug/ml streptomycin,DMEM:F12:MCDB105(2:1:1);and thyroid follicular carcinoma (FTC-133) cells in DMEM-F12(1:1),also containing 10% fetal bovine serum, 100u/ml penicillin and 100ug/ml streptomycin;both K1 and FTC-133 were cultured in the incubator with 37℃,5%CO2,change culture solution each 2 or 3 days.Digestive with 0.25% trypsogen,used the growth phase cells to the experiments.2 To detect cell proliferation with MTT:Papillary thyroid cancer cells(K1), thyroid follicular cancer cells(FTC-133)were cultivated with different concentrations of rhMBL(concentrations were 0,0.01,0.1,1,10ug/ml)after 12,24 and 48h,respectively,we used the mehtythiazolyltertazolium(MTT)to assay the cell proliferation .3 To demonstrate the apoptosis with FCM:Two cells were were cultivated with different concentrations of rhMBL(concentration is 0,1ug/ml)after 48h,demonstrated the cells apoptosis with Annexin V-FITC Apoptosis Detection Kit by Flow Cytometry.4 The expression of Bcl-2 and p53 gene protein:Extract total protein of two cells which cultivated with different concentrations of rhMBL (concentration is 0,1ug/ml) after 48h,β-actin as the inner parameter,to detect the relatively expression of Bcl-2 and p53 gene protein after incubation with rhMBL.Results:1 The results of the effect of rhMBL on K1 and FTC-133:Both cells were restrained by rhMBL,with a dose,time effect relations.2 The results of the cell apoptosis:After treated with rhMBL,both cells can be detected obvious apoptosis peak.The apoptosis of K1 (9.36±0.35) %,compared with control group(3.24±0.71)% significantly different(p<0.05); The apoptosis of FTC-133(12.87±0.47)%,compared with control group (5.15±0.52)% significantly different(p<0.05).3 The expression of Bcl-2 and p53 gene protein3.1 The expression of Bcl-2 gene protein3.1.1 K1:Western blot results showed that it was a strip at 26 KD place,after analysis the density scans,the result of experimental group was (0.59±0.04) obviously down regulated compared with control group(1.20±0.07).(p<0.05)3.1.2 FTC– 133:Western blot results showed that it was a strip at 26 KD place, after analysis the density scans,the result of experimental group was(0.33±0.06)obviously down regulated compared with control group(1.07±0.11). (p<0.05)3.2 The expression of p53 gene protein3.2.1 K1:Western blot results showed that it was a strip at 53 KD place,after analysis the density scans,the result of experimental group was(0.74±0.07) obviously down regulated compared with control group(1.51±0.29).(p<0.05)3.2.2 FTC-133:Western blot results showed that it was a strip at 53 KD place, after analysis the density scans,the result of experimental group was(0.35±0.08)obviously down regulated compared with control group(0.88±0.14). (p<0.05) Conclusions:1 In vitro,rhMBL can inhibit the proliferation of thyroid cancer cells with a dose,time effect relations.2 In vitro,rhMBL can promote thyroid cancer cells apoptosis,which provided the experimental data for prventing thyroid tumor.3 rhMBL could down regulate the experssion of Bcl-2 and p53 gene protein,which could promote the thyroid cancer cells'apoptosis,Bcl-2 and p53 play an important role in treatment for thyroid tumors.