Expression And Significance Of VEGF-A And Its Receptor VEGFR2 In Hepatocellular Carcinoma

Objective:To detect the levels of VEGF-A protein and HBV-DNA in the peripheral blood of patients with HCC as well as the expression of VEGF-A and VEGFR2 mRNrA in HCC and normal liver tissues,analyze the relationship between VEGF-A and the clinical pathological factors such as viral load,TNM stage,pathological stage,vascular invasion and lymphatic metastasis,and explore the progress and prognosis of VEGF-A and VEGFR2 in HCC.Methods:Based on the standard diagnosis and treatment of primary liver cancer in China(2017/2019),patients with hepatitis B-related HCC were divided into groups according to tumor stage.The expression level of VEGF-A protein in peripheral blood serum was detected by ELISA;HBV-DNA load in peripheral blood serum was detected by qPCR;total RNA was extracted by Trizol after grinding frozen cancer tissues and VEGF-A and VEGFR2 were detected by PCR after reverse transcription.With GAPDH as the internal reference,SPSS 20.0 was used to analyze the differential expression of various factors in the patients with liver cancer and the normal controls,and SABC immunohistochemistry was used to detect the protein content of VEGF-A and VEGFR2.The factors of liver cancer patients in different TNM stages and pathological stages were analyzed,and the relationship between the expression of VEGF-A and its receptors and clinical pathological factors such as vascular invasion and lymphatic metastasis and the correlation between liver elasticity and VEGF-A.Results:VEGF-A and VEGFR2 in peripheral blood of patients with liver cancer were higher than those in the normal control group(P<0.05).Serum VEGF-A content was significantly different among patients with different TNM stages(P=0.040),and there was no statistical significance between different pathological grades;there was no statistical difference between HBV-DNA load and the content in different pathological grades and TNM stages(P>0.05),and the correlation between VEGF-A and HBV-DNA expression was not statistically significant,but it was positively correlated with HBV-DNA expression in HBV-DNA(+)patients;serum VEGF-A in tumor diameter(d>5cm,d<5cm)(t=3.639,P=0.001),vascular infiltration(+/-)(t=11.754,P=0.000),lymph node metastasis(+/-)(t=3.537,P=0.001),HBV-DNA(+/-)(t=2.213,P=0.032)groups have different expression levels,the former serum VEGF-A are higher than the latter and the difference is statistically significant(P<0.05);There was no significant difference in the expression of the gender group(P>0.05).The optimal cutoff value of serum VEGF-A was 213ng/L and divided into high VEGF-A and low VEGF-A groups to draw the survival curve.It was found that the high VEGF-A group had PFS(Progression free survival)(x2=4.263,P=0.039)and overall survival(OS,Overall survival)(x2=3.993,P=0.046)were lower than those in the low VEGF-A group;and those in the high VEGF-A group.The recurrence rate is high,the risk of vascular invasion is high,there are many TN-M stages III and IV,and the differentiation is poor.The levels of VEGF-A in the serum of patients with liver cancer and normal controls were(297.30±239.30)ng/L and(167.00±119.41)ng/L.The serum HBV-DNA content of HCC patients was(1.79±4.64).There was statistical significance(t=4.346,P=0.041)(t=4.442,P=0.000).The levels of VEGF-A and HBV-DNA in different TNM stages were:stage ?((193.25±96.88)ng/L,1.01 ± 2.48),stage ?((279.69 ±105.75ng/L,1.05±2.82),stage ?((352.88±201.17)ng/L,2.95 ± 3.20),stage IV((429.89±280.25)ng/L,1,86±2.84),the difference in VEGF-A protein expression levels between different TNM stages(F=5.010,P=0.040)had statistical significance,but no HBV-DNA(F=1.523,P=0.221).The levels of VEGF-A and HBV-DNA in different levels of pathological differentiation were:well-differentiated((142.00±94.99)ng/L,1.57±2.70),Moderate differentiation((316.81±198.73)ng/L,1.93 ±2.96),low differentiation((323.67±212.38)ng/L,1.57±2.86)and there was no significant difference in the expression levels between VEGF-A(F=2.404,P=0.101)and HBV-DNA(F=0.090,P=0.914)in different pathological stages(P>0.05).There was a weak correlation between serum VEGF-A and HBV-DNA expression levels in patients with liver cancer(r=-0.161,P=0.264),but there was no statistical significance(P>0.05),and in HCC patients with HBV-DNA(+)only The expression of serum VEGF-A and HBV-DNA was still moderately positive(r=0.559,P=0.030)and statistically significant(P<0.05).VEGF-A was weakly correlated with HBV-DNA in TNM-I and III stages(r=0.137,P=0.575;r=0.030,P=0.911),and moderately correlated in TNM-II and IV stages(r=-0.624,P=0.185;r=-0.489,P=0.181),but the differences were not statistically significant(P>0.05);VEGF-A and HBV-DNA were weakly correlated at different stages of differentiation(r=-0.177,P=0.703;r=-0.063,P=0.736;r=0.149,P=0.643),but the differences were not statistically significant(P>0.05).The serum VEGF-A of HCC patients were grouped according to radiological,pathological and other features.It was found that VEGF-A was grouped by tumor diameter(t=3.399,P=0.000)and vascular infiltration(+/-)group(t=11.754,P=0.000),lymphatic metastasis(+/-)group(t=3.537,P=0.001)group,HBV-DNA(+/-)group(t=2.213,P=0.032)have different expression levels,and it was statistically significant(P<0.05),but the difference between the gender groups was not statistically significant(P>0.05).The ratio of serum total protein to white ball in the tumor diameter>5cm group(t=3.745,P=0.000)(t=2.590,P=0.013),vascular infiltration group(r=4.055,P=0.000)(t=2.734,P=0.009),HBV-DNA positive group(t=2.068,P=0.044)(t=2.434,P=0.019)Were lower than the other group,and there was no significant difference in lymphatic metastasis and gender grouping.Tissue VEGF-A and VEGFR2 cDNA gel scan results showed that the fluorescence intensity of VEGF-A(t=6.061,P=0.000)and VEGFR2(t=14.809,P=0.000)bands in the HCC patient group(18 cases)were all Compared with the normal healthy control group(3 cases),the difference was statistically significant(P<0.05).The expression levels of VEGF-A(F=4.751,P=0.017)and VEGFR2(F=3.514,P=0.044)are different in different TNM staging liver cancer tissues,and the difference is statistically significant(P<0.05);There was no significant difference in the expression level in differently differentiated HCC tissues(F=2.274,P=0.137)(F=2.192,P=0.146).The expression of VEGF-A and VEGFR2 in HCC tissues was highly positively correlated(r=0.844,P=0.000),which was statistically significant(P<0.05).The expression levels of VEGF and VEGFR2 in the HBV-DNA(+)group were(1.13±0.15)(0.95±0.13),higher than the HBV-DNA(-)group(0.88±0.07)(0.72±0.06),and the difference was statistically significant(t=4.464,P=0.000),(t=3.555,P=0.003).The results of immunohistochemistry showed that VEGF-A and VEGFR2 were mainly distributed in the cytoplasm of hepatoma cells,and VEGFR2 was also found in vascular endothelial cells.VEGF-A staining granules were abundant in the cytoplasm of moderately differentiated hepatoma cells,but not significantly enhanced in the cytoplasm of poorly differentiated hepatoma cells.VEGFR2 is not only highly expressed in the vascular endothelial cells of liver cancer,but also has the highest staining intensity in the cell membrane and cytoplasm of moderately differentiated liver cancer cells,suggesting that the ability of the membrane inversion of moderately differentiated liver cancer cells is still strong,and VEGFR2 could be enriched on the surface of the cell membrane.Immunohistochemistry revealed that the expressions of VEGF-A and VEGFR2 protein in liver cancer tissues(18 cases)were higher than those in normal controls(3 cases)(P=0.045)(P=0.019).The expressions of the two were different in groups with different degrees of differentiation,but the difference was not statistically significant(P>0.05).Transient elastography(TE)analysis found that liver stiffness(15.41±3.96)kpa in HCC patients was significantly higher than normal liver(4.76±1.32)kpa(P<0.05);tissue VEGF-A(r=0.488,P=0.040)and VEGFR2(r=0.401,P=0.099)expression was positively correlated with liver stiffness measurement(LSM),but the correlation between VEGFR2 and LSM was not statistically significant.Conclusion:VEGF-A in peripheral blood of patients with hepatocellular carcinoma,VEGF-A and VEGFR2 in cancer tissues were significantly increased.Those with poor TNM staging have higher VEGF-A levels.Vascular infiltration(+),HBV-DNA(+)liver cancer patients with VEGF-A levels increased more significantly;HBV-DNA can promote liver cancer patients with VEGF-A overexpression.The expression of VEGF-A and its receptor VEGFR2 are positively correlated.The high expression of the two promotes the growth of liver cancer;the overall survival rate and disease-free survival rate of those with high serum VEGF-A are lower,and the recurrence rate is higher.Liver cancer patients with high expression of VEGF-A and VEGFR2 have significantly increased LSM values,which are moderately positively correlated with each other,and indicate worsening of the condition and poor prognosis.Figure 26;Table 27;Reference 78.