骨肉瘤热疗前后热休克蛋白70表达的变化

The changes of HSP70 expression in osteosarcoma pre and post hyperthemia therapy

目的 为明确骨肉瘤加热前后热休克蛋白(heat-shock protein,HSP)70 表达的异同,探讨骨肉瘤热疗与机体免疫功能变化的关系。方法1997 年4 月～1999 年6 月收集病理诊断为骨肉瘤的肿瘤标本25 例,患者均采用微波原位热疗进行保肢手术,术中将肿瘤加热50 ℃持续15 min,分别在微波热疗前后进行取材。选用OS- 732 和U2- OS人骨肉瘤细胞系,体外培养于6 mm ×24 m m 的盖玻片上,加热50 ℃15 min 和30 min。上述组织及细胞行HE和免疫组织化学染色,观察热疗前后HSP70 表达的变化。结果 微波热疗前骨肉瘤组织虽有HSP70 表达阳性的细胞,但表达的细胞数量极少(2%)。微波热疗后骨肉瘤组织内有大量HSP70 表达阳性的肿瘤细胞(30% ～40%),且表达的强度较热疗前明显增加。此外,肿瘤间质和瘤旁组织内可见有HSP70 表达阳性的细胞,且数量较多。OS- 732 和U2-OS骨肉瘤细胞系经50 ℃15 min 和30 min 加热后,采用台盼蓝染色,OS-732 和U2- OS细胞死亡率均为100% 。免疫组织化学染色发现,加热50 ℃15 min 后HSP70 的表达明显增多,阳性细胞占95% 左右。HSP70 表达多在细胞浆内,表达的程度较热疗前明显增强。加热50 ℃30 min 后全部的肿瘤细胞均有HSP70 表达,且HSP70 表达强度进一步增加。结论 微波热疗后骨肉瘤组织表达HSP70 的数量?

Objective To study the differences of heat shock protein(HSP)70 expression in osteosarcoma before and after hyperthemia therapy, and discuss the relationship between tumor hyperthemia and immune responses Methods Twenty five specimens of osteosarcoma diagnosed by histopathological examination were collected from April 1997 to June 1999 All of the patients were treated with limbsalvage by intraoperative microwave heat on tumor at 50 ℃for 15 minutes; a sample was obtained pre and post microwave heat respectively In addition, OS-732 and U2-OS human osteosarcoma cell lines were applied, 2×10 6 cells were cultured on the slides and heated at 50 ℃for 15 and 30 minutes in vitro After hyperthemia, the tissues and cells were observed after HE staining; immunohistochemical examination was also done with antibody against HSP70 Results Before microwave heat treatment, very few cells in osteosarcoma tissues showed HSP70 positive On the other hand, there were a lot of tumor cells expressing HSP70 (30%～40%) after microwave heat treatment, and the expression level (staining degree) of HSP70 were much higher than before microwave heat treatment Moreover, the cells with HSP70 expression were found in interstitial and tumor adjacent tissues abundantly After the two osteosarcoma cell lines were heated at 50 ℃for 15 and 30 minutes, the cell death rates of OS-732 and U2-OS were totally measured by Trypan blue staining Immunohistochemical examination showed that HSP70 expression increased following treatment with 50 ℃for 15 minutes, with 95%of the cells giving positive expression Most of HSP70 expression was located in cytoplasm Nearly all of the cells expressed HSP70, and the expression level increased after hyperthemia of 50 ℃for 30 minutes Conclusion Hyperthemia therapy could enhance HSP70 expression in osteosarcoma, which may present the antigens of sarcoma to T cell and stimulate host immune system, as well as improve the anti tumor immune activity