透明质酸酶增强5-FU对结肠癌抗癌作用的动物实验研究

Enhancement of hyaluronidase on the therapeutic effect of fluorouracil in colon carcinoma of BALB/c nude mice

目的:观察透明质酸酶(Hyaluronidase,Hyase)在氟尿嘧啶(5-FU)对裸小鼠结肠癌移植瘤生长影响中的作用,并探讨其作用的可能机制。方法:18只雄性裸小鼠左侧腹壁皮下移植三维培养人结肠癌HT-29细胞,随机分为6组,每组3只:Ⅰ组(对照组)、Ⅱ组[Hyase200U/(kg.d)]、Ⅲ组[5-FU30mg/(kg.d)]、Ⅳ组[Hyase50U/(kg.d)+5-FU30mg/(kg.d)]、Ⅴ组[Hyase100U/(kg.d)+5-FU30mg/(kg.d)]和Ⅵ组[Hyase200U/(kg.d)+5-FU30mg/(kg.d)],Hyase于肿瘤细胞移植处第1、5天皮下注射,5-FU于第1～5天右侧腹腔注射。每周以游标卡尺测定瘤体最长径和最短径,计算肿瘤体积和抑瘤率,第6周杀鼠称瘤重,取瘤组织行HE染色,Westernblotting法检测瘤组织中E-钙黏素(E-cadherin)、细胞间黏附分子-1(ICAM-1)和NF-κBp65蛋白的表达并对蛋白条带的光密度(OD)进行半定量分析。结果:裸小鼠均成功移植皮下肿瘤,与对照组(Ⅰ组)比较,单纯Hyase治疗组(Ⅱ组)肿瘤体积减小不明显(P>0.05),单纯5-FU治疗组(Ⅲ组)自第3周开始肿瘤体积明显减小(P<0.05),而不同剂量Hyase联合5-FU治疗组(Ⅳ、Ⅴ、Ⅵ组)瘤体积减小更明显(P<0.05);与单纯5-FU治疗组(Ⅲ组)比较,第3周开始联合治疗组(Ⅳ、Ⅴ、Ⅵ组)瘤体积明显减小(P<0.05);第6周Ⅱ～Ⅵ组抑瘤率分别为6.8%、17.5%、34.1%、46.3%和56.6%;Ⅲ～Ⅵ组瘤重均低于Ⅰ组(P<0.05或P<0.01);Ⅱ、Ⅳ、Ⅴ、Ⅵ组瘤组织内E-cadherin和ICAM-1蛋白条带OD值均较Ⅰ组有不同程度减低,而NF-κBp65蛋白条带OD值在各组间无明显变化(P>0.05)。结论:透明质酸酶可能通过抗细胞黏附在结肠癌化疗中具有增敏作用。

Objective：To observe hyaluronidase as a chemosensitizer in nude mice colon carcinoma and to explore its mechanisms. Methods：Eighteen male BALB/c nude mice were subcutaneously implanted with HT-29 spheroids, they were treated by hyaluronidase or/ and 5-FU through intraperitoneal injection. We measured the length and width of tumor using sliding caliper weekly. The tumor volumes and inhibiting rate were calculated. The tumor mass were excised 6 weeks later, and the expression of E-cadherin, ICAM-1 and NF-κB p65 was detected by western blotting and analyzed by semiquantitative method. Results：All nude mice were implanted HT-29 spheroid cells successfully. Treatment of hyaluronidase alone couldn＇t reduce the tumor volume, bat the tumor volume by using 5-FU alone or byaluronidase ＋ 5-FU was decreased to some extent. The effect of high dose hyaluronidase ＋ 5-FU was most significant. Comparing with 5-FU alone, combination of byaluronidase and 5-FU could significantly reduce the tumor volume （ P 〈 0. 05 ）. The expression of E-cadherin and ICAM-1 was decreased by treatment of hyaluronidase, but NF-κB p65 was no difference in all groups. Conclusion：Hyaluronidase can be used as a chemosensitizer in colon carcinoma and the mechanisms may be involving its antiadhesion effect.