| Objective:By the in vitro release experiment and in vitro inhibition experiment of two sustained-release 5-fluorouracil (5-Fu) and cisplatin which are researched on th the theory of controlled-release drug delivery,we observe the situation of drug release in vitro,drug inhibition on human rhabdomyosarcoma RD cells and the effect on cell cycle.It is the basis for the study on the role of the slow-release agents on orbital rhabdomyosarcoma in the future.Methods:1. Sustained-release 5-Fu and cisplatin in vitro release experiment: we took slow-release agent with 5mg 5-Fu and placed in 10ml test tube, added 5ml 0.1mol /L PBS, soaked it on 37℃constant temperature, got leach fluid samples after immersing in PBS on the 1st,3rd,5th,8th day, used high performance liquid chromatography to measure drug concentration and calculate mass of drug release. The method of sustained-release cisplatin in vitro release was same with 5-Fu.2. The Rhabdomyosarcoma RD cells were reanimated, we observed the proliferative status and morphological characteristic by HE pigmentation,and made comparison between human rhabdomyosarcoma RD cells and primary orbital Rhabdomyosarcoma cells in morphology.3. Sustained-release 5-Fu and cisplatin anti-tumor in vitro experiments:The leaching fluid samples after soaking 1d, 3d, 5d, 8d of the sustained-release 5-Fu were applied to rhabdomyosarcoma RD cells, and we observed morphological changes of the rhabdomyosarcoma RD cells after 24 hours.The inhibition of RD cells proliferation was assayed by MTT. Method of slow-release agent of cisplatin was same with 5-Fu.4. To analyze the effect of different concentration of 5-Fu and cisplatin leaching fluid samples on the cell cycle of rhabdomyosarcoma RD cell by FCM.Results:1.5-Fu in vitro release experiment: the concentrations of 5-Fu were 138.84μg/ml, 42.08μg/ml,27.15μg/ml and 22.33μg/ml after immersing in PBS on the 1st,3rd,5th,8th day. That was, masses were respectively 649.20μg, 210.40μg, 135.75μg,111.65μg, release time could be up to more than 8 day.2. Rhabdomyosarcoma RD cells developed well and passed stably. The shape of cells were spindle-shaped or tadpole-shaped,the caryon is huge and round.It was same with the primary orbital rhabdomyosarcoma in morphology basically.3. Sustained-release 5-Fu and cisplatin anti-tumor in vitro experiments:The inhibition rates of leaching fluid samples after soaking 1d,3d,5d,8d of the sustained-release 5-Fu were 57.6%,49.6%,44.8%,34.5%. The inhibition rates of cisplatin were 87.8%,46.4%,42.5%,31.1%. OD values of 5-Fu and cisplatin measured by MTT method are lower than the control group, the difference was statistically significant (P<0.05). Blank release agent had no significant inhibitory effect on rhabdomyosarcoma RD cells and no statistically significant difference (P>0.05)compared with control group.4. The result of FCM indicated that the cell eycle of RD cells treated by sustained-release 5-Fu and cisplatin had changed. The proportion of cells in stage G1 increased, conversely, the proportion of cells in stage S and G2 decreased.PI% had statistically significant difference compared with control group(P<0.05).Conclusion:1. Sustained-released 5-Fu and cisplatin in vitro bursted first and then slow-released, sustained release stably in at least 8 day.Human rhabdomyosarcoma RD cells were continued to be inhibited,and cell cycle is interferred, that is the proportion of cells in stage G1 increased, and thereby the proliferation of cells is inhibited. 5-Fu can inhibit the proliferation of RD cell in dose-dependent manner.2. With the result of this study, we presume that sustained-released 5-Fu and cisplatin have the same effect on orbital rhabdomyosarcoma cell which has the same morphology with rhabdomyosarcoma RD cells. It is the basis for the study on the role of the slow-release agents on orbital rhabdomyosarcoma in the future.
|
PDF Full Text Request