多巴胺D2受体正电子发射计算机断层显像示踪兔脊髓内移植的神经前体细胞

Tracking neural progenitor cells transplanted into rabbit spinal cord by detection of dopamine receptor 2 with positron emission computed tomography

目的探索脊髓内移植神经前体细胞的无创性活体示踪方法。方法培养人神经前体细胞系hNPC-TERT,采用逆转录聚合酶链反应(RT-PCR)、免疫荧光染色、放射性配基结合实验检测hNPC-TERT多巴胺D2受体在体内外的表达,移植于兔正常脊髓后,以D2受体拮抗剂“C- raclopride作为示踪剂,正电子发射计算机断层成像(PET)显像示踪兔活体脊髓和离体脊髓内移植的hNPC-TERT。HeLa细胞移植于脊髓作为对照组。结果体外培养和脊髓内移植第2天的hNPC- TERT表达D2受体。静脉注射11C-raclopride后PET显像,兔活体脊髓和离体脊髓内hNPC-TERT移植部位放射性积聚,对照组仅见本底影像。PET图像中,hNPC-TERT移植部位标准化摄取值明显高于Hela细胞移植部位(P<0．05)。离体脊髓放射性剖面曲线分析及脊髓切片免疫荧光染色进一步证实了PET显像结果。结论以11C-raclopride为示踪剂,PET显像可以示踪到脊髓内移植第2天的人神经前体细胞系HNPC-TERT。提供了一种以特异性表面标志为靶点进行PET显像的移植干细胞活体示踪方法。

Objective To explore the effect of in vivo positron emission computed tomography （PET） in tracking the stem cells transplanted into spinal cord. Methods Telomerase-immortalized human neural progenitor cells of the line hNPC-TERT were cultured. HeLa cells were used as control cells. RTPCR was used to detect the mRNA expression of dopamine receptor 2 （ D2 ） in both cell lines, ^3H-raclopride was added into the suspensions of these 2 cell lines. RT-PCR was used to detect the mRNA expression of D2, and immunofluorescent staining was conducted on the cells to detect the protein expression of D2. Twenty-two New Zealand rabbits were randomly divided into 4 groups to undergo transplantation of hNPC- TERT cell suspension into the spinal cord at the segment T10, or transplantation of HeLa cells. Two day after the transplantation some rabbits were killed to take out the spinal cord at the segment T10 to undergo immunofluorescent staining to examine the radioactivity in the spinal cord. Some rabbits were injected with n C-raclopride intravenously and then underwent PET imaging. Results RT-PCR showed that mRNA expression was positive in the hNPC-TERT cells but negative in the HeLa cells, and immunofluorescent staining showed protein expression of D2 in the in the hNPC-TERT cells and not in the HeLa cells. The spinal cords specimens taken 2 days after transplantation had human-specific nuclear （HN） antigen positive and D2 positive cells. Fluorescent microscopy showed that the hNPC-TERT cells in the injection site did not migrate remarkably; and showed that the D2 staining was negative and HN antigen was positive in the HeLa cells. ^11C-raclopride PET imaging of the live rabbits showed accumulation of radioactivity at the hNPC-TERT cell injection site with a standard uptake value significantly higher than that of the HeLa cell transplantation group （P 〈 0.01 ）. ^11C-raclopride PET imaging of the isolated spinal cords showed rounded focal image of increased radioactivity in the hNPC-TERT cell transplantation group and linear image of radioactivity without clear border in the HeLa cell transplantation group. Conclusion PET imaging with as radiotracer targeting at specific cellular marker is effective in tracking cells into the body and in vivo visual evaluation of stem cell transplantation.