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透明质酸钠(图1)又称玻璃酸钠,是由交替的N-乙酰葡糖胺(GlcNAc)和葡糖醛酸(GlcA)单元组成[1]的聚合物,聚合度最大的相对分子质量(Mr)可以达到107。具有高度黏弹性、可塑性、渗透性、独特的流变学特性以及良好的生物相容性等特点[2]。透明质酸钠广泛存在于人体,是构成人体细胞间质、眼玻璃体、关节滑液等结缔组织的主要成分,在体内具有保水、维持细胞外空间、调节渗透压、润滑、促进细胞修复等重要生理功能[3]。医用透明质酸钠凝胶(HA)为无色透明黏稠液体,主要成分为透明质酸钠。HA常用于肿瘤患者术后的抗粘连,其广泛的生理学活性有可能使得肿瘤细胞易于扩增和转移,文献报道内源性的透明质酸能促进肿瘤增殖[4-6]。因此,本研究考察外源性HA是否会促进肿瘤的生长和转移,以明确HA是否可用于肿瘤患者的术后防粘连。为HA用于肿瘤患者腹、盆腔手术术后防粘连提供实验依据。
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在体外实验中,肿瘤细胞Hela、HCT116和CT26在100% HA中均不能生长(图2)。Hela细胞在培养3 d和5 d时不同含量HA均不影响细胞的生长,7 d时25%HA和50%HA细胞生长率低于阴性对照组(P<0.001)。HCT116细胞培养7 d时,50%HA细胞生长率低于阴性对照组(P<0.01)。CT26细胞培养3 d时12.5%HA、25% HA和50%HA细胞生长率低于阴性对照组(P<0.001);培养5 d和7 d时,50%HA细胞生长率低于阴性对照组(P<0.001)。不同浓度5-Fu,培养3 d、5 d和7 d均抑制肿瘤细胞Hela、HCT116和CT26的生长,呈剂量相关性。
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Transwell实验结果显示,高浓度HA体外抑制肿瘤细胞迁移。50%HA、25%HA和12.5%HA组Hela细胞穿透数均低于阴性对照组(P<0.01),HA含量越高,Hela细胞穿透数越少;50%HA和12.5%HA组HCT116细胞穿透数均低于阴性对照组(P<0.05),25%HA组HCT116细胞穿透数低于阴性对照组,无统计学差异(图3)。
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裸鼠结肠原位接种HCT116瘤块后,HA组与模型组成瘤率均为100%,结果无差异。模型组裸鼠的瘤体体积达(339.1±258.0)mm3,5-Fu组裸鼠的瘤体体积为(128.5±111.9)mm3,小于模型组(P<0.01),HA组裸鼠的瘤体体积为(293.2±190.6)mm3,与模型组比较,差异无统计学意义;模型组裸鼠的瘤重为(0.359±0.219)g,5-Fu组裸鼠的瘤重为(0.149±0.105)g,小于模型组(P<0.001),HA组裸鼠的瘤重为(0.330±0.187)g,与模型组比较,差异无统计学意义。使用一定剂量的HA在裸鼠体内不影响HCT116的成瘤率,同时也不影响HCT116肿瘤的生长(表1)。
表 1 HA在裸鼠体内对HCT116肿瘤生长的影响(
$\bar{ x}\pm { s}$ )组别 动物数(只) 成瘤数(只) 成瘤率(%) 瘤体积(V/mm3) 瘤重(m/g) 假手术组 16 0 0 0.0 ± 0.0 0.000 ± 0.000 模型组 16 16 100 339.1 ± 258.0 0.359 ± 0.219 HA组 16 16 100 293.2 ± 190.6 0.330 ± 0.187 5-Fu组 16 14 87.5 128.5 ± 111.9** 0.149 ± 0.105*** **P<0.01、***P<0.001,与模型组比较 -
裸鼠结肠原位接种CT26瘤块,观察16 d后出现动物死亡,第18天处死存活裸鼠。模型组存活率为62.5%,HA组存活率68.8%,5-Fu组存活率100%,体内使用一定剂量的HA不影响裸鼠的存活率。模型组肝脏转移率90%,平均病灶数(2.2±1.7);5-Fu组肝脏转移率0.0%,低于模型组(P<0.001);平均病灶数(0.0±0.0),低于模型组(P<0.001)。HA组肝脏转移率36.4%,低于模型组(P<0.05);平均病灶数(0.6±1.0),低于模型组(P<0.01)。模型组肺转移率为40%,平均病灶数(0.5±0.7);5-Fu肺转移率6.3%,低于模型组(P<0.05);平均病灶数(0.1±0.3),低于模型组(P<0.05)。HA组肺转移率为18.2%,平均病灶数(0.2±0.4),与模型组均无统计学差异,见表2、图4和图5。
表 2 HA在裸鼠体内对CT26肿瘤转移的影响(
$\bar{ x}\pm { s}$ )组别 动物数(只) 成活率(%) 肝转移 肺转移 转移率(%) 病灶数 转移率(%) 病灶数 假手术组 15 100.0 0.0 0.0 ± 0.0 0.0 0.0 ± 0.0 模型组 16 62.5 90.0 2.2 ± 1.7 40.0 0.5 ± 0.7 HA 组 16 68.8 36.4* 0.6 ± 1.0** 18.2 0.2 ± 0.4 5-Fu 16 100.0 0.0*** 0.0 ± 0.0*** 6.3* 0.1 ± 0.3* *P<0.05、**P<0.01、***P<0.001,与模型组比较
The effects of medical sodium hyaluronate gel on the tumor proliferation and metastasis
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摘要:
目的 考察医用透明质酸钠凝胶(medical sodium hyaluronate gel,HA)在体外和裸鼠体内对腹、盆腔相关肿瘤生长和转移的影响。 方法 利用Hela、CT26和HCT116等3株肿瘤细胞,通过MTT法和Transwell实验分别考察不同浓度HA体外对肿瘤细胞生长和迁移的影响;建立裸鼠结肠原位种植瘤模型,比较瘤块接种4周后不同实验组的瘤块体积和瘤重,考察HA体内对结肠癌HCT116增殖的影响;通过比较接种3周后不同实验组裸鼠的肺和肝脏的肿瘤转移率和转移病灶数,考察HA对结肠癌CT26体内转移的影响。 结果 不同浓度HA在体外均未促进Hela、CT26和HCT116细胞的生长和转移,且在5 mg/ml HA浓度下,显示出一定的抑制作用。体内结果显示,HA未促进结肠癌HCT116肿瘤的生长,却显示出明显抑制CT26肿瘤的转移。 结论 在本实验条件下,HA在体内、外均未促进腹、盆腔相关肿瘤细胞Hela、CT26和HCT116的生长、迁移和转移。 Abstract:Objective To investigate the effect of medical sodium hyaluronate gel (HA) on the growth and metastasis of abdominal and pelvic tumor cells in vitro and in nude mice. Methods Three tumor cells, Hela, CT26 and HCT116, were used to investigate the effects of different HA concentrations on the growth and migration of tumor cells in vitro by MTT assay and Transwell assay. An orthotopic transplantation model of colonic tumor in nude mice was established to investigate the effect on the proliferation of cell HCT116 by comparing the tumor volume and tumor mass 4 weeks after inoculation. The effects on the metastasis of cell CT26 were investigated by comparing the tumor metastasis rate and the number of metastatic lesions of lung and liver in nude mice among the different experimental groups 3 weeks after inoculation. Results HA did not promote the growth and metastasis of Hela, CT26 and HCT116 cells in vitro at different concentrations. Actually, HA exhibited a certain inhibitory activity at the concentration of 5 mg/ml. In the orthotopic transplantation model of colonic tumor-HCT116, HA did not promote the growth of cell HCT116. In the orthotopic transplantation model of colonic tumor-CT26, HA inhibited CT26 tumor metastasis. Conclusion Under the experimental conditions, HA did not promote the growth, migration or metastasis of abdominal and pelvic related tumor cells including Hela, CT26 and HCT116 in vitro and in vivo. -
表 1 HA在裸鼠体内对HCT116肿瘤生长的影响(
$\bar{ x}\pm { s}$ )组别 动物数(只) 成瘤数(只) 成瘤率(%) 瘤体积(V/mm3) 瘤重(m/g) 假手术组 16 0 0 0.0 ± 0.0 0.000 ± 0.000 模型组 16 16 100 339.1 ± 258.0 0.359 ± 0.219 HA组 16 16 100 293.2 ± 190.6 0.330 ± 0.187 5-Fu组 16 14 87.5 128.5 ± 111.9** 0.149 ± 0.105*** **P<0.01、***P<0.001,与模型组比较 表 2 HA在裸鼠体内对CT26肿瘤转移的影响(
$\bar{ x}\pm { s}$ )组别 动物数(只) 成活率(%) 肝转移 肺转移 转移率(%) 病灶数 转移率(%) 病灶数 假手术组 15 100.0 0.0 0.0 ± 0.0 0.0 0.0 ± 0.0 模型组 16 62.5 90.0 2.2 ± 1.7 40.0 0.5 ± 0.7 HA 组 16 68.8 36.4* 0.6 ± 1.0** 18.2 0.2 ± 0.4 5-Fu 16 100.0 0.0*** 0.0 ± 0.0*** 6.3* 0.1 ± 0.3* *P<0.05、**P<0.01、***P<0.001,与模型组比较 -
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