The Correlation Between Brafv600E Gene Mutation And Expression Of Cyclind1,P27,NIS,TPO In PTC

Objective (1) To evaluate BRAF gene mutation of PTC in GuangXi region and examine the correlations with the clinicopathological parameters. To investigate the invasiveness,lymph node metastasis of BRAF MT PTC tissue, and postoperative recurrence. (2) To evaluate the expression of CyclinD1, P27, NIS and TPO in BRAF MT and WT PTC tissue and the correlations between BRAF gene mutation with the expression of CyclinD1, P27, NIS and TPO, to investigate the endopathic and mechanism of BRAF gene mutation for clinical pathological parameters of PTC.Methods To collect 115 cases of paraffin embed tissue of PTC and 30 cases of thyroid benign lesion as control (20 cases of nodular goiter and 10 cases of thyroid adenoma) from January 2004 to March 2010 in the first affiliated hospital of Guang Xi medical university. Genomic DNA was extracted from the paraffin-fixed slices by FFPE DNA Kit. Then analyzed DNA samples for mutation in BRAF exon 15 by PCR, and utilized SSCP(Single Strand Conformation Polymorphism, SSCP) analysis to identify abnormal staining bands, then the corresponding bands was sequenced directly. To investigated the protein expression of CyclinD1, P27, NIS and TPO in PTC and thyroid benign lesion by means of immunohistochemistry. Data obtained from the results of the application package SPSS13.0 statistical analysis to P<0.05 for the difference has statistical significance.Results 1. The ratio of BRAF gene mutation rate in PTC tissue was 48.7%(56/115). DNA bidirectional sequencing confirmed missense mutations, which is exon 15 single nucleotide missense mutation (T1799A). The ratio of BRAF gene mutation in the groups of sex and tumor size(<2cmor≥2cm) have no significant difference(χ2=0.352 , P>0.05 andχ2=0.235 , P >0.05); the difference was significant between the groups of age(≥45 or <45) (χ2=6.317,P<0.05), the different pathological subtype group(classic, follicular subtype, high-cell type)(χ2=20.325 , P < 0.05), clinical stage groups (Ⅰ/ⅡorⅢ/Ⅳ)(χ2=8.681 , P < 0.05), adjacent tissues violations and lymph node metastasis have no influence on BRAF gene mutation rate (χ2=20.892,P<0.05;χ2=13.349,P<0.05). 2. (1)The positive expression rate of CyclinD1 was 65.2%(75/115). The positive expression rate of CyclinD1 was 80.3%(45/56)and 50.8%(30/59)in BRAF MT and WT PTC. The two groups was significant difference byχ2 test (P<0.001). (2)The positive expression rate of P27 was 41.7%(48/115). The positive expression rate of P27 was 25.0%(14/56)and 57.6%(34/59)in BRAF MT and WT PTC. The two groups was significant difference byχ2 test (P<0.001). (3)The positive expression rate of NIS was 47.8%(55/115). The positive expression rate of NIS was 26.8%(15/56)and 67.8%(40/59)in BRAF MT and WT PTC. The two groups was significant difference byχ2 test (P<0.001). (4)The positive expression rate of TPO was 47.8%(55/115). The positive expression rate of TPO was 32.1%(18/56)and 62.7%(37/59)in BRAF MT and WT PTC. The two groups was significant difference byχ2 test (P<0.001). (5) Spearman correlation analysis of BRAF gene mutation and the expression of CyclinD1 have positive correlation (r=0.290, P<0.05); Spearman correlation analysis of BRAF gene mutation and the expression of P27, NIS, TPO have negative correlation (P<0.05), r=-0.352, -0.387, -0.298, respectively.Conclusions 1. BRAF gene mutation has a high detection rate in PTC in Guang Xi region, which indicated that BRAF mutation can be used as a molecular marker in differentiating thyroid benign and malignant tumor. 2. The correlation between BRAF gene mutation and the over expression of CyclinD1 and down expression of P27, NIS, TPO is very obvious. 3. The combined detection of BRAF gene mutation and the four proteins in PTC can be used to estimate the clinical situation of PTC, can also help formulating a reasonable treatment plan, assessment and prognosis of metastasis.