~(31)P-MRS无创量化肝细胞癌细胞代谢水平及其与血管生成的关系

目的 利用~(31)P-MRS对肝细胞癌(HCC)量化的代谢情况,探讨肿瘤血管生成及其成熟度对代谢的影响.方法 对经病理证实的肝细胞癌31例在手术前行~(31)P-MRS扫描,分析肿瘤和周围肝实质的细胞内pH值(pHi)、磷酸单脂(PME)、磷酸双脂(PDE)、无机磷(Pi)、γ-ATP、β-ATP、α-ATP、PME/ATP、Pi/ATP、PME/PDE、PME/Pi、PDE/Pi、PDE/ATP和低能磷酸盐(LEP)等.利用免疫组化技术检测肿瘤的VEGF、Flk-1表达情况及PCNA指数,利用病理图像自动分析系统统计微血管和成熟血管的数目、平均面积、总面积、周长、直径、异型指数、血管间距、表达部位、动脉数、静脉数、血管成熟指数和平均灌注分数等参数.利用HE染色计算细胞外间隙(ES)面积及细胞外血管外间隙(EES).将~(31)P-MRS技术测得的指标与血管参数进行对照分析.结果 HCC的pHi、PME/PDE以及Pi、β-ATP的峰下面积都高于肝脏(P＜0.05);PDE/ATP、PDE/Pi则相反(P＜0.05).Pi/ATP在VEGF阳性和阴性表达时差异有统计学意义(P=0.047,f=2.135),PDE/Pi在Flk-1阳性和阴性表达组也具统计学意义(P=0.001,t=2.13).pHi在CD34不同表达部位之间存在统计学意义(P=0.01,F=7.493).pHi与静脉数呈正相关(P=0.003,r=0.749);PME与EES负相关(P=0.029,r=-0.583);Pi与微血管总面积正相关(P=0,r=0.766);γ-ATP与动脉数目呈正相关(P=0.012,r=0.648).与β-ATP有关有ES、PI及微血管的直径、周长和平均面积等(P＜0.05).PME/Pi与PI正相关,与微血管异型指数负相关(P＜0.05).成熟血管数目、周长、直径及动脉数目与PME/ATP均相关(P＜0.05).PDE/Pi与成熟血管间距也为正相关(P=0.041,r=0.533),与LEP有关的因素包括平均灌注分数、EES和微血管总面积(P＜0.05).结论 HCC的pHi、Pi、β-ATP、PME/PDE都高于肝脏,PDE/ATP、PDE/Pi则相反;HCC血管生成的异质性影响~(31)P-MRS检测的各个代谢物的水平.

Correlation of microvessel and mature vessel in hepatocellular carcinoma with tumor metabolism quantized noninvasively by using in-vivo ~(31)P-MRS at 3.0 Tesla

Objective To probe the influence of angiogenesis and its maturation in hepatocellular carcinoma(HCC)over cell metabolism with in-vivo ~(31)P-MRS at 3.0 Tesla.Methods ~(31)P-MRS scanning with a single voxel on lesion and hepatic parenchyma respectively was carried out in 31 patients with pathologically proved HCC.Intracellular pH value(pHi),and some metabolites including phosphomonoester(PME),phosphodiester(PDE),inorganic phosphate(Pi),γ-,β-and α-ATP,lower energetic phosphate(LEP),and the ratios of PME/ATP,Pi/ATP,PME/PDE,PME/Pi,PDE/Pi and PDE/ATP were calculated.After surgery,the continuous slices of lesion were stained with hematoxylin-eosin(HE)and immunohistochemical staining of vascular endothelial growth factor(VEGF),Flk-1,PCNA,CD34 and alpha smooth muscle actin(SMA).Extracellular space(ES)and extracellular extravascular space(EES)by HE staining,and the expressions of the VEGF and the Flk-1,and PCNA index(PI)were evaluated.According to CD34 and SMA staining,some vascular parameters were counted in a computed analysis system including number,mean vessel area,total vessel area,circumference,diameter,distance between adjacent vessels(DAV),variety index and the expression position of microvessel and mature vessel.In addition,the number of arterioles and vein-lets,mature vessel index and mean perfused fraction(mPF)were also recorded.All vessel parameters were compared with the metabolites.Results ~(31)P-MRS showed an increase in Pi,β-ATP,pHi and PME/PDE,and a decrease in PDE/ATP and PDE/Pi in HCC when compared with normal hepatic parenchyma(P＜0.05).Pi/ATP was significantly increased in lesions with positive VEGF expression(P=0.047,t=2.135),and an increase in PDE/Pi was also detected in positive expression group of Flk-1(P=0.001,t=2.13).Among three patterns of CD34 expression position,the highest pHi was in type Ⅱ,whereas the lowest in type Ⅰ(P=0.01,F=7.493).pHi was correlated positively with the increased number of veinlets(P=0.003,r=0.749),and an inverse correlation of PME with EES was gotten(P = 0.029,r=-0.583).Positive correlations between microvessel total area and Pi,and between γ-ATP and the number of arterioles were showed(P＜0.05).Except ES,microvessel diameter,circumference,mean area and PI were positively correlated with β-ATP(P＜0.05).In addition,PME/Pi was inversely correlated with microvessel variety index,whereas positively with PI(P＜0.05).Some parameters including the number,diameter,and circumference of mature vessel,and the arteriolar number were correlated positively with PME/ATP(P＜0.05).There were positive correlations between PDE/Pi and mature DAV(P=0.041,r=0.533),and between mPF,microvessel total area and LEP,whereas an inverse correlation between EES and LEP was gotten(P＜0.05).Conclusion Pi,phi,β-ATP and PME/PDE in HCC are higher than those in liver.However,PDE/ATP,PDE/Pi are inverse.Metabolites in HCC detected with in-vivo ~(31)P-MRS may be influenced by the heterogeneity of angiogenesis.