光动力疗法对输入的NK细胞在荷Heps肝癌鼠瘤内分布的影响

目的探讨NK细胞输入后在荷Heps肝癌小鼠体内的分布规律,光动力疗法（PDT）对输入的NK细胞在肿瘤组织内分布的影响。方法 96只KM小鼠建立Heps肝癌移植瘤模型后随机分为对照组、细胞组（NK组）和光免疫组（PIT组）,每组32只。对照组小鼠尾静脉输入生理盐水,NK组小鼠尾静脉输入4,＇6-二脒基-2-苯基吲哚（DAPI）标记的NK细胞,PIT组小鼠PDT处理后即刻输入DAPI标记的NK细胞。治疗后1、2、4、6和8 d摘取小鼠肝脏、脾脏、肺脏和肿瘤组织,在荧光显微镜下观察NK细胞在上述脏器的动态分布情况;治疗后2 d观察瘤组织的病理学变化;治疗后1、2、4、6和8 d摘除小鼠眼球取血以流式细胞仪检测外周血NK细胞百分率;治疗后6 d LDH释放法检测小鼠脾细胞杀伤活性。结果 （1）各治疗组,输入后1、2、4、6和8 d,NK细胞在小鼠肝脏、脾脏、肺脏和肿瘤组织中均有分布,其中以肿瘤组织内最多;NK细胞在瘤内浸润密度：以输入后2 d最高,而各时间点PIT组又显著高于NK组（P〈0.05）;（2）外周血中NK细胞含量：在治疗后1、2、4和6 d,PIT组较对照组明显增高（P〈0.01）;治疗后1、2和4 d,NK组较PIT组和对照组增高（与PIT组相比,P〈0.05;与对照组相比,P〈0.01）;（3）效靶比为10∶1、20∶1和50∶1时,NK组和PIT组脾细胞杀伤活性均显著高于对照组,其中PIT组又显著高于NK组（P〈0.01）。结论 NK细胞输入荷瘤小鼠体内后能选择性聚集于肿瘤组织内;PDT可增加输入的NK细胞在瘤内的浸润密度,增强小鼠脾细胞对Heps肝癌的杀伤活性。

Effects of Photodynamic Therapy on the Localization of Transferred Natrual Killer Cells in Transplanted Heps Hepatoma

Objective To investigate the distributional regularity of natural killer cells(NK) on Hepatoma-Heps-bearing mice in vivo,and whether photodynamic therapy(PDT) can promote the infiltration of NK cells in tumors. Methods The hepatoma model was established by subcutaneously inoculating Heps cells to 96 KM mice,who were then divided into the control,cell(NK) and photo immunotherapy(PIT) groups,each having 32 mice.The mice of the control group were only administered with NS.For the NK group,NK cells were labeled with 4,6-diamidino-2-phenylindole(DAPI) and then injected into the mice tail veins.As to the PIT group,labeled NK cells were injected into the mice tail veins immediately after PDT light exposure.The tumors,lungs,spleens and livers were removed on day 1,2,4,6 and 8 after the treatment and observed for the distributional regularity under fluorescence microscope.The histopathological changes of tumors were observed via HE staining on day 2.Blood samples from the mice eyes were taken on day 1,2,4,6 and 12 after the treatment and the percentage of NK cells in peripheral blood was tested with flow cytometry.The cytotoxicity of the mice spleen cells were determined through lactate dehydrogenase(LDH) release assay on day 6. Results(1) On day 1,2,4,6 and 8,labeled cells could be observed in all the said viscera organs of the treated mice.Labeled cells accumulated mostly in the tumors.On day 2 after the injection,much more labeled cells were seen in the tumors than at other time points(P<0.01).The infiltration density of labeled cells in the PIT group was higher than that of the NK group(P<0.01).(2) On day 1,2 and 4 after the treatment,the percentage of NK cells in mice peripheral blood of the NK group was larger than that of the PIT group(P<0.01).(3) When T ratio being 10 ∶1,20 ∶1 and 50 ∶1,the cytotoxicity of spleen cells in the PIT group and NK group were far higher than that in the control group(P<0.01),and that of the PIT group much higher than that of the NK group(P<0.01). Conclusions Adoptively transferred NK cells can accumulate selectively in tumors.The combination with PDT will promote the infiltration of NK cells in tumors and enhance the cytotoxicity of mice spleen cells.