The Correlation Research Between Lung Cancer And The Expression Of E-cadherin MRNA And Protein

Objective: To explore the role of E-cadherin in lung cancer tumorigenesis, tumorprogress and clinical pathological value from transcriptional and translational level and to reveal whether the transcriptional level or translational level was inhibited that causes the E-cadherin expression abnormality, and to provide scientifical evidence for lung cancer tumorigenesis and tumorprogress.Methods: By adjusted the reaction conditions of nested reverse transcription-polymerase chain reaction(nested RT-PCR) and selected the optimized reaction conditions, the relative quantitative method for E-cadherin mRNA expression has been established. Then this method and Western Blot method were used to detect the expression E-cad mRNA and its protein in all 30 cases of lung cancer tissues, paracancer lung tissues and farcancer lung tissues, 5 cases of non-cancer lung tissues were also detected. The two-step immunohistochemical method was used to confirm some results of Western Blot.Results:1．The optimized RT-PCR conditions were: 1×PCR buffer, dNTP200μmol/L, Mgcl₂ 1．5mmol/L, primers 600nmol/L, Taq DNA polymerase 0．1U/μl, templates 50ng, the total volum was 25μl。The cycling conditions were as follows: After 10 min at 95℃, reactions were cycled through 1 min denaturation at 95℃, 1 min annealing at 66℃,and 1 min extension at 72℃for 25(the first round PCR)or 35(the second round PCR andβ-actin)repeats; followed 72℃for 10 minutes.2．The means of the relative rate of E-cad mRNA andβ-actin mRNA in lung cancer tissues, paracancer tissues, farcancer tissues and non-cancer lung tissues were 1．33±0．48,1．61+0．55,1．93±0．45,2．09±0．09．Statistic analysis showed that there was significantly different between lung cancer tissue and paracancer lung tissue and between lung cancer tissue and farcancer lung tissue (P<0．05) . There was also significantly different between paracancer lung tissue and farcancer lung tissue and between paracancer and non-cancer lung tissues (P<0．05) .But there was no difference between farcancer tissues and non-cancer lung tissues (P>0．05) .3．In the 30 cases of lung cancer tissues, the realitive rate of E-cad mRNA andβ-actin mRNA was 1．21±0．53 in squmaous cell carcinoma (SCC), 1．40±0．46 in adenocarcinoma(Ad) and 1．51±0．27 in adenosquamous carcinoma(AdCa). In the 30 cases of paracancer tissues, the realitive rate of E-cad mRNA andβ-actin mRNA was 1．59±0．45 in SCC, 1．58±0．67in Ad and 1．80±0．26in AdCa. In the 30 cases of paracancer tissues, the realitive rate of E-cad mRNA andβ-actin mRNA was 1．85±0．41 in SCC, 1．90±0．44 in Ad and 2．37±0．53 in AdCa. Statistic analysis showed that there was no significantly different between SCC and Ad in the group of lung cancer tissue, paracancer lung tissue and farcancer lung tissue.4．The result of Western Blot:The positive rate of E-cad was 36．7%(11/30) in lung cancer tissue,70．0%(21/30) in paracancer tissue, and 96．7%(29/30) in farcancer lung tissue. Statistic analysis showed that there was a significant difference between lung cancer tissue and paracancer tissue (x²=6．7, P<0．017) ; and also a significant difference between lung cancer tissue and farcancer tissue was found (x²=24．3, P<0．017) . There was also a significant different between paracancer lung tissue and farcancer lung tissue (x²=7．68, P<0．017) . The positive rate of E-cad in 5 cases of non-cancer lung tissue were 100%(5/5) . All the samples displayed clear bands of GAPDH.5．In the 30 cases of lung cancer tissue, the positive expression rate of E-Cadherin in different histology type was 33．3%(4/12) in SCC,40%(6/15) in Ad, and 33．3%(1/3) in AdCa. In the 30 cases of paracancerlung tissue, the positive expression rate of E-Cadherin in differenthistology type was 66．7%(8/12) in SCC, 73．3%(11/15) in Ad, and 66．7%(2/3) in AdCa. In the 30 cases of farcancer lung tissue, the positiveexpression rate of E-Cadherin in different histology type was 91．7%(11/12) in SCC, 100%(15/15) in Ad, and 100%(3/3) in AdCa. Statisticanalysis showed that the expression of E-cad in all the three groupshad no significant difference in SCC, Ad and AdCa(x²=0．343, x²=0．464, x²=2．14, P>0．05) .6．We selected 5 patients that the results of Western Blot displayednegative in their lung cancer tissue to confirm the results of Westernblot by immunohistochemistry. 5 cases of farcancer lung tissue andpositive control showed typical membranous expression of E-cad ,suggesting there were normal gap junction and cell adhesion among cells.Only one(18) in 5 cases of paracancer lung tissues showed negativeexpression. However, all the 5 lung cancer tissue and negative controlshowed negative expression.The expressed intensity of E-cad reducedobviously, some of them were mainly expressed in cytoplasm, a few ofthem were expressed in nucleus or/and membrane. The results wereidentified with the Western Blot results.Conclusions:1．E-cad mRNA and its protein expression in lung cancer group were lower than paracancer or farcancer group, which meant that the loss of E-cad helped in tumorgenesis and tumorgenesis. It might be an early accident since there were differences between paracancer and farcancer group or paracancer group and non-cancer lung tissues.2．There was no evident relationship of E-cad mRNA and its protein among different pathological types. This showed that the expression abnormal of E-cad mRNA and its protein might be the common course in the tumorgenesis and tumorgenesis of SCC and Ad.3．The results of Western blot were identifed with nested RT-PCR.This suggest that the downregulation of E-cadherin results from the transcriptional repression.