MiR-145 May Influence The Progression Of Clear Cell Renal Cell Carcinoma And The Proliferation Of 786-O Cells By Targeting Negative Regulation Of SPOP

Objective:By exploring the regulatory relationship between miR-145 and SPOP in Renal Cell Carcinoma（RCC）,and their effects on the progression of RCC and the proliferation ability of786-o cells,explore the possible molecular mechanism of the progression of RCC,so as to provide a possible basis for seeking a new treatment of Renal Carcinoma.Methods:In this study,44 pairs of clear cell Renal Cell Carcinoma（cc RCC）tissues and matched adjacent tissues were collected.The expression levels of miR-145 and SPOP m RNA in 44 patients with cc RCC were measured by q RT-PCR,to determine the correlation between miR-145 and SPOP in cc RCC;By analyzing the relationship between the expression of miR-145 and SPOP m RNA and the clinicopathological characteristics of cc RCC,determine the effect of miR-145 and SPOP m RNA on the progression of cc RCC;The expression of SPOP in 786-O cells was observed to clarify the relationship between SPOP gene and miR-145 by interventing the expression of miR-145 in786-O cells;Finally,the level of miR-145 and SPOP in 786-O cells was artificially interfered,the A_R450nmR value of 786-O cells was detected by CCK8 test to reflect the effect of miR-145 and SPOP expression on 786-O cell proliferation.Results:1.The expression of miR-145 in clear cell Renal Cell Carcinoma was significantly lower than that in adjacent tissues（t=9.753,P<0.0001）,and the expression of SPOP m RNA in adjacent tissues was significantly higher than that in adjacent tissues（t=8.758,P<0.0001）,there was a negative correlation between the expression of SPOP m RNA and miR-145（R=-0.8121,P<0.0001）;Compared with HK-2 cells,miR-145 was lower in 786-o cells（t=9.659,P=0.0105）,while SPOP m RNA and protein were higher in 786-o cells（t=21.85,7.658;P=0.0021,0.0016）;2.The low expression rate of miR-145 and the high expression rate of SPOP m RNA in cc RCC patients with Tumor Diameter>4cm were higher than those in cc RCC patients with Tumor Diameter≤4cm（P=0.023,0.022）;the low expression rate of miR-145 and the high expression rate of SPOP m RNA in cc RCC patients with WHO/ISUP Grade 1,2 and 3 were different,and the difference was statistically significant（P=0.001,0.014）;But the low expression rate of miR-145 and the high expression rate of SPOP m RNA were not related to Age（P=0.968,0.539）,Gender（P=0.771,0.196）and Clinical Stage（P=0.501,0.963）;3.Quantitative analysis of miR-145 and SPOP m RNA expression showed that the expression of miR-145 in cc RCC with Tumor Diameter>4cm and Clinical StageⅡwas lower than that in cc RCC with Tumor Diameter≤4cm and Clinical StageⅠ（P=0.0047,0.0036）;The expression of SPOP m RNA in cc RCC with Tumor Diameter>4cm and Clinical StageⅡwas higher than that in cc RCC with Tumor Diameter≤4cm and Clinical StageⅠ（P=0.0066,0.0081）,The expression of miR-145 in the cc RCC with WHO/ISUP Grade 1,2 and 3decreased in turn（P≤0.05）,while the expression of SPOP m RNA in the cc RCC with WHO/ISUP Grade 1,2 and 3 increased in turn（P≤0.05）;4.When the level of miR-145 in 786-O cells was up-regulated,the expression of SPOP m RNA and SPOP protein in 786-O cells were lower than those in Control Group（t=8.522,P=0.0135;t=12.00,P<0.0001）,and when the level of miR-145 in 786-O cells was down-regulated,the expression of SPOP m RNA and SPOP protein were significantly higher than that of Control Group（t=31.49,P=0.0010,t=33.92,P<0.0001）;5.The results of CCK8 cell proliferation experiment showed that at 24H,48H and 72H,the A_R450nmR value of 786-O cells decreased after miR-145 was specifically up-regulated（P≤0.05）;On the contrary,the A_R450nmR value of 786-O cells increased after miR-145 was specifically down-regulated（P≤0.01）;The A_R450nmR value of 786-O cells decreased after knockdown SPOP gene（P≤0.05）.In view of the above experimental results,knockdown SPOP gene and up-regulate the expression of miR-145 at the same time,the A_R450nmR value of 786-O cells decreased significantly（P≤0.001）;knockdown SPOP gene and down-regulate the expression of miR-145 at the same time,the A_R450nmR value of 786-O cells was no significant change compared with the Control Group（P>0.05）.Conclusions:1.The expression of miR-145 was low in cc RCC,while the expression of SPOP m RNA was high in cc RCC,the expression of miR-145 and SPOP m RNA was negatively correlated in cc RCC;2.The larger the diameter of cc RCC,the higher the clinical stage and WHO/ISUP Grade,the lower the expression of miR-145 and the higher the expression of SPOP m RNA;3.miR-145 can negatively regulate the expression of SPOP gene in cc RCC;4.Up regulation of miR-145 and knockdown of SPOP have synergistic effect on reducing the proliferation of 786-O cells.