目的 探究全反式维甲酸（ATRA）对多房棘球蚴原头节体外活性及生长的影响。 方法 从保种沙鼠体内无菌剖取包囊，经过研磨、过滤、清洗获取活性良好的多房棘球蚴原头节。将原头节分为不同浓度ATRA组（10、25、50、100 μmol/L组，添加对应终浓度的ATRA）、二甲基亚砜（DMSO）组（DMSO终浓度为0.1％）和空白组（加入等量完全培养基），共干预9 d，伊红染色后于显微镜下观察其活力和形态变化，计算原头节的存活率并绘制存活率曲线。将原头节与大鼠肝癌RH35细胞共培养5~6周，获得多房棘球蚴微囊泡后，使用不同终浓度ATRA（10、100 μmol/L）干预微囊泡9 d，显微镜下观察其活力和形态变化，同时设置DMSO组和空白组。干预原头节48 h后，分别使用EdU细胞成像检测原头节EdU阳性率，使用半胱天冬氨酸蛋白酶-3（Caspase-3）试剂盒检测原头节中Caspase-3相对表达量；干预原头节24 h后，活性氧（ROS）检测试剂盒测定各组原头节ROS水平。两组样品的比较采用独立样本t检验，不同浓度ATRA组与DMSO组差异比较采用单因素方差分析（ANOVA）。 结果 10、25、50、100 μmol/L组中的死亡原头节体积明显缩小，可被伊红染液染成红色，随着时间延长和药物浓度升高出现轮廓模糊、透光性减弱、小钩脱落增多等。第4天时，10、25、50、100 μmol/L组原头节存活率分别为（87.33 ± 4.90）%、（74.00 ± 2.08）%、（64.33 ± 2.03）%、（53.33 ± 1.86）%，均低于DMSO组[（95.67 ± 1.20）%]（F = 98.41，P < 0.05）；第9天时，10、25、50、100 μmol/L组原头节存活率仅为（62.00 ± 2.64）%、（36.33 ± 2.52）%、（7.67 ± 1.53）%、0，均低于DMSO组[（85.67 ± 2.08）%]（F = 1 154.34，P < 0.05）。不同浓度的ATRA与多房棘球蚴微囊泡共培养9 d时，随ATRA浓度的升高，囊泡生长缓慢，囊内结构逐渐浑浊；空白组和DMSO组的微囊泡生发层和角质层结构完整。EdU细胞成像可见不同浓度ATRA组均呈现红色及蓝色荧光，10、25、50、100 μmol/L组原头节的EdU阳性率分别为（51.63 ± 3.09）%、（42.09 ± 1.36）%、（38.46 ± 0.65）%、（25.23 ± 1.32）%，均低于DMSO组[（58.32 ± 0.91）%]（F = 168.59，P < 0.05）。10、25、50、100 μmol/L组的Caspase-3活性分别为（19.23 ± 2.27）、（26.27 ± 3.45）、（43.29 ± 2.10）、（72.80 ± 1.40）U/L，呈浓度依赖性升高，均高于DMSO组[（14.22 ± 0.52）U/L]（F = 404.08，P < 0.05）。不同浓度ATRA组ROS均可见绿色荧光，强于DMSO组，10、25、50、100 μmol/L组荧光强度分别为260.96 ± 2.52、282.10 ± 7.40、330.30 ± 12.46、346.10 ± 6.39，均高于DMSO组（236.03 ± 6.89）（F = 80.53，P < 0.05）。 结论 ATRA对多房棘球蚴原头节体外活性及生长具有抑制作用，可诱导原头节内ROS显著积累，抑制原头节增殖活性、促进凋亡进程。

Objective To investigate the effect of all-trans retinoic acid (ATRA) on the activity and growth of Echinococcus multilocularis protoscoleces in vitro. Methods The cysts were dissected aseptically from the gerbils and the active E. multilocularis protoscolices were obtained by grinding, filtering, and cleaning. The protoscolices were divided into different concentrations of the ATRA groups (10, 25, 50, and 100 μmol/L groups with the corresponding final concentration of ATRA), the dimethyl sulfoxide (DMSO) group (the final concentration of DMSO was 0.1%), and the blank group (the same amount of complete medium was added). After 9 days of intervention, eosin staining was used to observe the activity and morphological changes of the groups under the microscope. The survival rate of the protoscolices was calculated, and the survival curve was drawn. E. multilocularis microcysts were obtained by co-culturing the protoscolices with rat hepatocellular carcinoma RH35 cells for 5-6 weeks, and the microcysts were treated with different final concentrations of ATRA (10, 100 μmol/L) for 9 days. The activity and morphological changes of the microcysts were observed under a microscope, and the DMSO group and blank group were set up. After 48 h of intervention, the EdU positive rate was detected by EdU cell imaging and the relative expression of Caspase-3 in protoscolices was detected using the Caspase-3 kit. After 24 hours of intervention, a ROS detection kit was used to quantify the levels of reactive oxygen species (ROS) in each group. An independent sample t-test was used to compare the two groups of samples, and a one-way analysis of variance (ANOVA) was used to compare the difference between the ATRA and DMSO groups of different concentrations. Results The volume of dead protoscoleces in the 10, 25, 50, and 100 μmol/L groups significantly decreased and could be stained by eosin dye. With the increase in time and drug concentration, the outline blurred, light transmission weakened, and small hook detachment increased. On day 4, the survival rates of the protoscoleces in the 10, 25, 50, and 100 μmol/L groups were (87.33 ± 4.90) %, (74.00 ± 2.08) %, (64.33 ± 2.03) % and (53.33 ± 1.86) %, respectively. They were all lower than those in the DMSO group [(95.67 ± 1.20) %] (F = 98.41, P < 0.05). On day 9, the survival rates of protoscoleces in the 10, 25, and 50 μmol/L groups were (62.00 ± 2.64)%, (36.33 ± 2.52)% and (7.67 ± 1.53)%, which were lower than those in the DMSO group [(85.67 ± 2.08)%] (F = 1154.34, P < 0.05). When different concentrations of ATRA were co-cultured with multilocular echinococcus microcysts for 9 days, with the increase in ATRA concentration, the vesicles grew slowly, and the intracapsular structure became cloudy. The germinal layer and stratum corneum of microcysts were intact in the blank group and the DMSO group. EdU cell imaging showed red and blue fluorescence in ATRA groups with different concentrations. The EdU positive rates in the 10, 25, 50, and 100 μmol/L groups were (51.63 ± 3.09) %, (42.09 ± 1.36) %, (38.46 ± 0.65) %, and (25.23 ± 1.32) %, respectively. All of them were lower than those in the DMSO group [(58.32 ± 0.91)%] (F = 168.59, P < 0.05). The caspase-3 activity of the 10, 25, 50, and 100 μmol/L groups was (19.23 ± 2.27), (26.27 ± 3.45), (43.29 ± 2.10) and (72.80 ± 1.40) U/L, respectively, showing a dose-dependent increase. All of them were higher than the DMSO group [(14.22 ± 0.52) U/L] (F = 404.08, P < 0.05). The green fluorescence of ROS in the ATRA group was stronger than that in the DMSO group. The fluorescence intensity of the 10, 25, 50, and 100 μmol/L groups was 260.96 ± 2.52, 282.10 ± 7.40, 330.30 ± 12.46 and 346.10 ± 6.39, respectively. All of them were higher than the DMSO group (236.03 ± 6.89) (F = 80.53, P < 0.05). Conclusion ATRA can inhibit the activity and growth of E. multilocularis protoscoleces in vitro, induce significant ROS accumulation in protoscolices, inhibit protoscolices proliferation, and promote apoptosis.