Hepatocellular carcinoma with high and low metastatic potential before and after bone marrow mesenchymal stem cell transplantation: a MRI imaging study using Gd-RGD probe

doi:10.3969/j.issn.2095-4344.0899

Abstract

Abstract:

BACKGROUND: With the advancement of modern molecular imaging, possibilities to monitor the efficacy of bone marrow mesenchymal stem cells (BMSCs) against tumors have been achieved.
OBJECTIVE: To construct a novel metastasis-associated MRI molecular imaging probe, Gd-RGD, and to observe metastasis and proliferation of hepatocellular carcinoma (HCC) before and after intervention with BMSCs using MRI imaging
.METHODS: The metastasis-associated MR molecular imaging probe, integrin α_vβ₃ ligand cRGD-PEG-DGL-DTPA-Gd (Gd-RGD), was constructed, and the ingredient analysis and identification were implemented through ¹H-magnetic resonance spectroscopy (¹H-MRS) and inductively coupled plasma-atomic emission spectrometry (ICP-AES) methods. MHCC97-H and MHCC97-L animal models were established. After BMSCs intervention was administrated via the tail vein for 6 weeks, tumor weight inhibition rate was calculated. MRI imaging with a metastasis-associated integrin ligand RGD molecular probe was performed. Molecular imaging probe Gd-DTPA was taken as control. Signal noise ratio (SNR) and contrast to noise ratio (CNR) of MRI were taken as semi-quantitative indicators. MHCC97-H and MHCC97-L animal models before BMSCs intervention were co-cultured with BMSCs using Transwell method. Quantitative PCR method was used to detect proliferation and metastasis-associated indicators, tumor growth factor β1 (TGF-β1), osteopontin, integrin subunits α_v and β₃.
RESULTS AND CONCLUSION: (1) After BMSCs transplantation, the tumor tissue weight of HCC decreased obviously, and the tumor weight inhibition rate was peaked in the 3^rd week. (2) For high metastatic potential (MHCC97-H), both SNRs and CNRs of Gd-RGD MR imaging before and after BMSCs intervention were statistically different (P < 0.05). For low metastatic potential (MHCC97-L), SNRs of Gd-RGD MR imaging before and after BMSCs intervention were not statistically different (P > 0.05), while CNRs were statistically different (P < 0.05). (3) SNRs of Gd-RGD MRI were significant different between MHCC97-L and MHCC97-H after transplantation (P < 0.05), while SNRs and CNR of Gd-RGD MRI changed significantly as compared with those of Gd-DTPA after cell transplantation. (4) In MHCC97-H cells co-cultured with BMSCs, the osteopontin, integrin subunits α_v and β₃ and TGF-β1 expression significantly decreased (P < 0.05), but integrin subunit αv expression did not change obviously (P > 0.05). In MHCC97-L cells co-cultured with BMSCs, OPN, β₃, TGFβ1 and α_v expression significantly decreased (P < 0.05). These findings reveal that the CNR index of MR imaging in two kinds of tracers is a good indicator to distinguish high and low potential HCC tissues. After BMSCs transplantation, the SNR and CNR indexes of Gd-DTPA and Gd-RGD can both distinguish high and low potential HCC tissues, and Gd-RGD imaging can achieve a more accurate distinction compared with Gd-DTPA.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

Key words: Bone Marrow, Mesenchymal Stem Cell Transplantation, Liver Neoplasms, Integrin alphaVbeta3, Magnetic Resonance Imaging, Tissue Engineering

CLC Number:

R394.2

Li Tian-ran, Huang Xiao-bin, Yang Zhi-jie, Lu Guang-ming, Li Yan-jun, Yu Ming-hui. Hepatocellular carcinoma with high and low metastatic potential before and after bone marrow mesenchymal stem cell transplantation: a MRI imaging study using Gd-RGD probe[J]. Chinese Journal of Tissue Engineering Research, 2018, 22(21): 3341-3348.

Figures/Tables 3

2.1 实验动物数量分析 60只健康Balb/c裸鼠均进入结果分析。 2.2 BMSCs干预前后MHCC97-H肿瘤Gd-RGD和Gd-DTPA成像结果分析 MHCC97-H细胞接种于裸鼠腋窝处，MRI 增强采用T1WI成像序列，先T1WI平扫，后行CE-T1WI成像(图1，2)。由图可见，MHCC97-H肿瘤组织位于腋窝处，肿瘤组织分别行Gd-DTPA和Gd-RGD成像。肝癌组织在BMSCs干预前后Gd-RGD强化程度低于Gd-DTPA。计算定量指标SNR和CNR(图3，4)，结果显示，BSMCs干预前后Gd-DTPA成像SNR和CNR均高于Gd-RGD成像(P < 0.05)。 2.3 BMSCs干预前后MHCC97-L肿瘤模型Gd-RGD和Gd-DTPA成像结果分析 MHCC97-L细胞接种于裸鼠腋窝处，MRI 增强采用T1WI成像序列，先T1WI平扫，后行CE-T1WI成像(图5，6)，可见MHCC97-L肿瘤位于腋窝处，肿瘤组织分别行Gd-DTPA和Gd-RGD成像。肝癌组织在BMSCs干预前后，Gd-RGD强化程度低于Gd-DTPA。肝脏组织Gd-RGD强化程度也低于Gd-DTPA。计算定量指标SNR和CNR(图7，8)，结果显示，BSMCs干预前后Gd-DTPA对照组成像SNR和CNR均高于实验组Gd-RGD成像(P < 0.05)。 2.4 BMSCs干预肝癌组织前后MRI图像对比分析采用SNR和CNR作为半定量指标，分析肿瘤组织在BMSCs干预前后两种示踪剂对肿瘤成像结果(表3)。 BMSCs干预6周后，行Gd-RGD和Gd-DTPA成像。Gd-RGD成像SNR分析结果显示，MHCC97-H和MHCC97-L差异无显著性意义(P > 0.05)，而BMSCs干预后，MHCC97-L SNR高于MHCC97-H，差异有显著性意义(P < 0.05)；Gd-DTPA成像SNR结果与Gd-RGD结果相同。Gd-RGD成像CNR分析结果显示，MHCC97-L CNR高于MHCC97-H，差异有显著性意义(P < 0.05)，同样，BMSCs干预后，MHCC97-L CNR高于MHCC97-H，差异有显著性意义(P < 0.05)；Gd-DTPA成像CNR结果与Gd-RGD结果相同。对于高转移潜能肝癌(MHCC97-H)，BMSCs干预前后Gd-RGD成像SNR和CNR比较差异有显著性意义(P < 0.05)。而BMSCs干预前后Gd-DTPA成像SNR比较差异无显著性意义(P > 0.05)，CNR差异有显著性意义(P < 0.05)。对于低转移潜能肝癌(MHCC97-L)，BMSCs干预前后Gd-RGD成像SNR比较差异无显著性意义(P > 0.05)，CNR比较差异有显著性意义(P < 0.05)。而BMSC干预前后Gd- DTPA成像SNR和CNR比较差异均无显著性意义(P > 0.05)。 2.5 高、低转移潜能肝癌组织(MHCC97-H和MHCC97-L)在BMSCs干预前后肿瘤质量变化从BMSCs干预后第2周开始后观察(图9，10)。经BMSCs干预后，肿瘤质量降低，表明BMSCs对肝癌组织增殖能力具有抑制作用。实验结果显示MHCC97-H第2周肿瘤质量抑制率为22.71%，第3周为42.86%，第4周为34.48%，第5周为31.60%，第6周为29.17%。MHCC97-L第2周肿瘤质量抑制率为26.62%，第3周为52.00%，第4周为38.20%，第5周为31.98%，第6周为30.23%。结果显示，肿瘤在第3周时肿瘤质量抑制率最高，且BMSCs对MHCC97-L肿瘤的抑制率高于对MHCC97-H肿瘤的抑制率。 2.6 BMSCs与肝癌细胞共培养前后转移相关因子骨桥蛋白、αv、β3、转化生长因子β1基因表达见表4。高转移潜能细胞(MHCC97-H)与BMSCs共培养后骨桥蛋白、β3、转化生长因子β1表达明显下降(P < 0.05)，而αv表达无明显变化(P > 0.05)。低转移潜能细胞(MHCC97-L)与BMSCs共培养后骨桥蛋白、β3、转化生长因子β1、αv表达下降，差异有显著性意义(P < 0.05)。"

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