Expression Of PD-ECGF In Nephroblastoma And Its' Clinical Significance

PrefaceNephroblastoma is a kind of malignant and embryonic solid tumor commonly occurred in children aged from 6 months to 3 years.We have taken the synthesized therapeutics to nephroblastoma in recent years and the survival rate of two year and five year are 90.7% and 68% respectively .Angiogenesis is prerequisite to tumor growth and metastasis and is induced by various angiogenis factors produced by tumor cells and/or nonmalignant cells that infiltrate the tumor. Platelet - derived endothelial cell growth factor ( PD -ECGF) ,a member of the pyrimidine nucleoside phosphoralase family and is essential for DNA synthesis and can limite cell growth. PD - ECGF which indenti-fied as thymidine phosphorylase (TP) is a potent angiogenic factor and its' degradation products, thymine, 2 - deoxyribose - 1 - phosphate and 2 - deoxy - D - ribose, have angiogenic and antiapoptotic effects associating with tumor aggressiveness and poor survival. Thus , to clarify the role of PD - ECGF in nephroblastoma and the correlation with its' metastasis, clinical stage, pathology type and prognosis ,we compared the gene expression in 30 nephroblastoma and 8 paracancer renal tissues and 4 renal specimens of other chronic deseases by u-sing RT - PCR.Method30 nephroblastoma samples were collected and selected by operation in our department from 1999 to 2003 . These samples were staged according to NWTS₃,with 11 stage 1,8 stage 11,7 stage 111,4 stage IV;pathology type were confirmed after operation 10 with epithelium ,6 blastema , 6 stroma, 8 mixture. 8 samples from tumor - adjacent tissues and 4 samples from renal tissues with chronic diseases were collected after operation. All of samples were stored in - 80Tl. RT -PCR kit are obtained from Takara;Trizol from Huamei Co. Expression levels of PD - ECGF mRNA were determined by RT - PCR. Total RNA were refined according to kit direction, mRNA concentration and purity were tested by Ultraviolet spectrum. 2|xl of RNA were used for each 20 jxl RT cDNA reaction. The reaction was under the following condition: 65 X for 1 min ,30X1 for 5 min , 65T! for 30 min ,9SX. for 5 min, 5X. for 5 min.For PD - ECGF mRNA detection, 3 jxl of RT - cDNA product were ampli-field using the primers PD - ECGF - F 5'- CTG CCA CTC ATC AC A GCC TCC ATT -3'and PD -ECGF -R 5'-GAG CGT GGT GAC CAG GTC CCT TA -3' Each 25|xl reaction contained dd. H2O 0. 5jxl, 10 x buffer 2. 5 jxl,dNTPs 2ui, Taq - E 0.2 ui, PD - ECGF - F 0. 1 jjlI , PD - ECGF - R 0. 1 jxl. Each cycling condions were as followed;94CC for 3 min,35 cycles of 94Tl for 40 s, 62t for 1 min,and 72T1 1 min,followed by one cycle of 72X for 7 min. (B -actin which as an inside contrast took part in the reaction.Result1. Various degrees of PD - ECGF mRNA expression were presented in all samples. The PD -ECGF mRNA level was 1.22 -fold higher in tumour tissues than in paracancer renal tissues (t = 0. 855 p > 0. 05 ) and 2. 53 - fold higher than in renal tissue without tumour (t=2.05 p<0.05) . Paracancer renal tissues expressed 2. 07 - fold higher PD - ECGF mRNA level than renal tissue without tumour( t = 2.65 p < 0. 05 ) .2. We conclude that PD - ECGF mRNA level does not correlate with pathology type in nephroblastoma(F =0. 691 p>0.05 ).3. We conclude that PD - ECGF mRNA level does not correlate with clinical stages of nephroblastoma(F = 0. 05 p>0.05) .4. The most important result we got was that the PD - ECGF mRNA level innephroblastoma with metasis is 1.72 - fold higher than the tumour without me-tastasis(t =2.2766 p<0.05 ).5. The sensitivity and specificity for predicting tumour recurrence were 83.3% and 80. 0% respectively if use 0. 99 as a cutoff value of PD - ECGF mRNA level in this group of nephroblastoma.ConclusionsThere was over expression of PD - ECGF mRNA in nephroblastoma and the PD - ECGF mRNA level correlated with the tumour' s metastasis and recurrence. Analyzing the PD - ECGF mRNA level mighit contribute to the evaluation of nephroblastoma' s prognosis.