H1受体拮抗剂异丙嗪对顺铂治疗骨肉瘤的影响及机制

Efficacy and mechanism of H1 receptor antagonist promethazine on osteosarcoma treated with cisplatin

目的观察异丙嗪在骨肉瘤细胞对顺铂化疗敏感性的影响,探讨其用于替代地塞米松(Dex)作为化疗辅助药物的可行性。方法构建荷人骨肉瘤裸鼠模型,随机分成6组,每组6只。分别腹腔注射200μL的生理盐水、化疗药物顺铂(3mg/kg)、Dex(4mg/kg)、异丙嗪(20mg/kg)、Dex+顺铂、异丙嗪+顺铂治疗。每隔3天给药1次,共4次。定期测量肿瘤体积大小,至第15天麻醉处死小鼠取肿瘤组织,称量瘤体重量。免疫荧光染色法检测肿瘤组织中血管内皮细胞标志物CD31的表达和分布;观察上述不同药物处理对骨肉瘤细胞株MG-63细胞血管内皮细胞生长因子VEGF表达和分泌的影响;MTT法检测MG63的增殖及顺铂诱导的增殖抑制作用;用Annexin V-FITC/PI双染法检测异丙嗪和顺铂对MG-63细胞凋亡的影响。结果与单用顺铂治疗组相比,异丙嗪+顺铂组与Dex+顺铂组的肿瘤体积和离体瘤重均显著增大。Dex+顺铂组和异丙嗪+顺铂组肿瘤间质组织中CD31的表达和分布明显多于单用顺铂组。单用Dex或异丙嗪处理24h或36h并不改变VEGF mRNA和蛋白分泌水平,单用顺铂处理则可明显降低VEGF的表达和分泌,而异丙嗪与顺铂联用则可显著逆转顺铂的作用。10nM~10μM异丙嗪处理MG63细胞72h,仅10nM异丙嗪对细胞增殖抑制作用,抑制率约为25.7%;与2μg/mL顺铂联用时,异丙嗪也不能逆转顺铂对细胞增殖抑制作用。联用Dex可以部分逆转顺铂导致的细胞凋亡,凋亡率为12.7%,而联用异丙嗪并不能减少顺铂对细胞的凋亡诱导作用。结论骨肉瘤化疗时使用H1受体拮抗剂异丙嗪会降低化疗药的效果,机制可能与异丙嗪促进肿瘤血管生成有关。

Objective To observe the effect of promethazine on the sensitivity of cisplatin chemotherapy in osteosarcoma cells,and to explore the feasibility of using it as a substitute for dexamethasone(Dex)as an adjuvant chemotherapy.Methods A nude mouse model bearing human osteosarcoma was constructed and randomly divided into 6 groups with 6 mice in each group.Inject 200μL of normal saline,chemotherapy drugs cisplatin(3 mg/kg),Dex(4 mg/kg),promethazine(20 mg/kg),Dex+cisplatin,promethazine+cisplatin respectively.It was administered once every 3 days for a total of 4 times.The tumor volume was measured regularly,until the 15 th day the mice were anesthetized and the tumor tissues were taken and the tumor weight was weighed.lmmunofluorescence staining method was used to detect the expression and distribution of vascular endothelial cell marker CD31 in tumor tissues.The effects of the above-mentioned different drug treatments on the expression and secretion of vascular endothelial growth factor VEGF in MG-63 cells were observed.MTT method was used to detect osteosarcoma cell line MG63 Proliferation and cisplatin-induced proliferation inhibition.Annexin V-FITC/PI double staining method was used to detect the effect of promethazine and cisplatin on the apoptosis of MG-63 cells.Results Compared with the cisplatin treatment group alone,the tumor volume and isolated tumor weight of the promethazine+cisplatin group and the Dex+cisplatin group were significantly increased.The expression and distribution of CD31 in tumor interstitial tissues of Dex+cisplatin group and promethazine+cisplatin group were significantly higher than that of cisplatin alone group.Treatment with Dex or promethazine alone for 24 hours or 36 hours does not change the levels of VEGF mRNA and protein secretion.Treatment with cisplatin alone can significantly reduce the expression and secretion of VEGF,while the combination of promethazine and cisplatin can significantly reverse cisplatin.The role of platinum.When MG63 cells were treated with 10 nM-10μM promethazine for 72 hours,only 10 nM promethazine inhibited cell proliferation,and the inhibition rate was about 25.7%;when combined with 2μg/mL cisplatin,promethazine could not reverse the inhibition of cisplatin on cell proliferation.effect.Combined use of Dex can partially reverse the apoptosis caused by cisplatin,with an apoptotic rate of 12.7%,while combined use of promethazine cannot reduce the apoptosis-inducing effect of cisplatin on cells.Conclusion The use of H1 receptor antagonist promethazine during chemotherapy of osteosarcoma may reduce the effect of chemotherapy drugs,and the mechanism may be related to the promotion of tumor angiogenesis by promethazine.