CAJ | 학술논문

目的：分离得到毛蚶蛋白活性组分并研究其体外抗肿瘤机制及对免疫细胞的作用。方法：低温水提及硫酸铵沉淀法获取毛蚶蛋白活性组分；MTT法检测50、100、200和400μg/mL的毛蚶蛋白活性组分分别作用24、48和72 h后对人结直肠癌细胞HT-29、HCT116，人肝癌细胞HepG-2，人肺癌细胞A549的细胞毒活性；倒置相差显微镜下观察细胞活性及Giemsa染色观察毛蚶蛋白活性组分对肿瘤细胞及核形态变化的影响，流式细胞术检测毛蚶蛋白活性组分对人结直肠癌HT-29细胞周期及凋亡的影响；MTT法检测毛蚶蛋白活性组分对小鼠巨噬细胞RAW264.7生长的影响，中性红法测定其对小鼠巨噬细胞RAW264.7吞噬能力的影响，Griess法检测其对小鼠巨噬细胞RAW264.7分泌NO的影响。结果：毛蚶蛋白活性组分对多种人肿瘤细胞系有抑制作用，且呈作用时间与剂量依赖性；流式细胞术结合Giemsa染色发现，其主要引起HT-29细胞G2~M期阻滞，多核细胞增加，并具一定的诱导细胞凋亡作用；在50～100μg/mL下能刺激小鼠巨噬细胞RAW264.7生长，增强其吞噬能力及NO分泌活性。结论：毛蚶蛋白活性组分具有一定抗肿瘤活性，主要机制为引起肿瘤细胞G2~M期阻滞及诱导其凋亡，且具一定免疫激活作用。
목적：분리득도모감단백활성조분병연구기체외항종류궤제급대면역세포적작용。방법：저온수제급류산안침정법획취모감단백활성조분；MTT법검측50、100、200화400μg/mL적모감단백활성조분분별작용24、48화72 h후대인결직장암세포HT-29、HCT116，인간암세포HepG-2，인폐암세포A549적세포독활성；도치상차현미경하관찰세포활성급Giemsa염색관찰모감단백활성조분대종류세포급핵형태변화적영향，류식세포술검측모감단백활성조분대인결직장암HT-29세포주기급조망적영향；MTT법검측모감단백활성조분대소서거서세포RAW264.7생장적영향，중성홍법측정기대소서거서세포RAW264.7탄서능력적영향，Griess법검측기대소서거서세포RAW264.7분비NO적영향。결과：모감단백활성조분대다충인종류세포계유억제작용，차정작용시간여제량의뢰성；류식세포술결합Giemsa염색발현，기주요인기HT-29세포G2~M기조체，다핵세포증가，병구일정적유도세포조망작용；재50～100μg/mL하능자격소서거서세포RAW264.7생장，증강기탄서능력급NO분비활성。결론：모감단백활성조분구유일정항종류활성，주요궤제위인기종류세포G2~M기조체급유도기조망，차구일정면역격활작용。
OBJECTIVE: To isolate the active component of Arca subcrenata protein,investigate the antitumor and immune activities in vitro. METHODS:The Arca subcrenata protein active component was extracted by ice water and ammonium sulfate precipitation. Cytotoxicity in human rectal concinoma HT-29 and other cell lines treated with Arca subcrenata protein active component of 50,100,200,400μg/mL for 24,48 and 72 h. Morphology changes in living cells were examined under the inverted phase contrast microscope. Cellular and nuclear morphological changes in human tumor cells treated with the Arca subcrenata protein active component was studied by Giemsa staining. Cell cycle and apoptosis in HT-29 cells were determined by flow cytometry. The growth of mice macrophage RAW264.7 treated the Arca subcrenata protein active component was detected by MTT assay. The phagocytosis of mice macrophage RAW264.7 treated with Arca subcrenata protein active component was determined by neutral red. The NO production by RAW264.7 was detected by Griess method. RESULTS:Arca subcrenata protein active component inhibited the survival of many tumor cell lines in concentration and time-dependent manners. Arca subcrenata protein active component induced cell were cycle arrest at G2-M cycle phases and apoptosis in HT-29 cells. It also showed that multinucleated cells were increased. Under the concentration of 50-100 μg/mL,Arca subcrenata protein active component could stimulate the growth of RAW264.7 cells,enhance phagocytosis and NO production activity. CONCLUSION:Arca subcrenata protein active component demonstrated anti-tumor activities due to its immune activation and the fact that it induced cell cycle arrest at G2-M cycle phases and apoptosis.