半乳糖化白蛋白磁性纳米粒运载的阿霉素在大鼠体内分布的研究
摘要
目的：观察由半乳糖化白蛋白磁性纳米粒运载的阿霉素经舌静脉给药后在大鼠体内的的分布状况。
方法：全部大鼠随机分为四组，每组15只，每组设5个亚组，每个亚组设平行大鼠3只。四组为：A组：游离阿霉素组；B组：白蛋白阿霉素磁性纳米粒组；C组：半乳糖化白蛋白阿霉素磁性纳米粒组；D组：半乳糖化白蛋白阿霉素磁性纳米粒+外加磁场组。经舌静脉，按分组分别注射相应药物，剂量均为阿霉素2.5mg/kg体重。在注射药物前，大鼠沿腹正中线剪开腹壁，D组显露肝左叶，暴露于磁场中。按注射药物后5分钟、15分钟、30分钟、60分钟、120分钟断颈处死动物。取全血、心、肺、肝、脾、肾，D组分取靶区肝、非靶区肝。全血制成血浆，器官组织制成匀浆，盐酸乙醇法提取阿霉素，用荧光光度计测量。
结果： 静脉注射药物后，D组大鼠靶肝的ADM浓度最高，显著高于非靶肝区及A、B、C组(p＜0.05)；非靶肝区与C组肝中ADM浓度的差别无统计学意义(p＞0.05)，但均高于A、B两组(p＜0.05)；而血浆、心、肾、肺中ADM的浓度显著低于A、B组(p＜0.05)，但与C组相比较，其浓度差别无统计学意义(p＞0.05)。C组血浆、心、肾、肺中ADM浓度低于A、B组(p＜0.05)。B组肝ADM浓度高于A组，血浆、心、肾中ADM浓度低于A组(p＜0.05)，脾、肺中ADM浓度显著高于A、C、D组(p＜0.05)；而A、C、D组中脾ADM浓度差别无统计学意义(p＞0.05)。在各个测试的时间点上，随着时间
的延长，外加磁场和未加磁场的半乳糖磁性阿霉素白蛋白纳米粒组的
药物靶向指数和药物选择指数是逐渐增高的。
结论:白蛋白阿霉素磁性纳米粒经半乳糖化后，可显著增强阿霉
素对肝脏的靶向性，并显著降低心、肺、脾、’肾、血浆肝外器官的组
织阿霉素浓度。利用外加磁场，可提高阿霉素在肝脏特定部位蓄积的
能力。因此，静脉注射半乳糖化白蛋白阿霉素磁性纳米粒是可行的。
Object: to investigate distribution of adriamycin contained by galactosed albumin magnetic nanoparticle injected by lingual vein in rats.
Methods: All animals were divided into four groups which were A group: adriamycin; B group: adriamycin contained by magnetic albumin nanoparticle ; C group: adriamycin contained by galactosed albumin magnetic nanoparticle; D goup: adriamycin contained by galactosed albumin magnetic nanoparticle plus external magnetic field. Each group included 15animals, which were subdivided into 5 subgroups, thus each subgroup included 3 animals. The drugs were administrated into corresponding rats at a dosage of adriamycin 2.5mg/kg body weight. Before drugs were injected intravenously, animals were explored abdominal wall along middle line of abdomen , the left hepatic lobes of animals in D group were exposed ,which magnetite were placed on. Animal were sacrificed 5min 15 min 30min 60min 120min after drugs were injected. Blood heart lung liver kidney were harvested, targeted Liver or untargeted liver in D group was harvested. Blood samples were made into plasm, organic tissues were rinsed for homogenate. Adriamycin was pi
ck up by hydrochlore-ethanol method, which level was measured by fluorescence photometer.
Results: Adriamycin level of targeted liver tissue in D group was the highest in all groups, and not only singnificantly higher than that of untargeted in D group , but also that in A B C group(p<0.05); But
adriamycin level was not significantly different between targeted liver tissue in D group and liver tissue in C group(p>0.05) Adriamycin level of plasm heart lung kidney was not significantly different between C and D group(p>0.05), but significantly lower than that in A and B group(p<0.05) Adriamycin level of spleen in B group was the highest in all groups, but that was not significantly different in A C and D group(p>0.05) drug targeting index and drug selective index were increasing in C and D group as tested time went.
Conclusions: After adriamycin contained by galactosed albumin magnetic nanoparticle was administrated intravenously, adriamycin can be targetedly deliver to liver, and adriamycin level in plasm heart lung kidney spleen was significantly lowered in extra-hepatic organ, external magnetic field can significantly increase adriamycin level in specific site of liver tissue. So it was feasible that adriamycin contained by galactosed magnetic albumin nanoparticle was administrated intravenously.
方法：全部大鼠随机分为四组，每组15只，每组设5个亚组，每个亚组设平行大鼠3只。四组为：A组：游离阿霉素组；B组：白蛋白阿霉素磁性纳米粒组；C组：半乳糖化白蛋白阿霉素磁性纳米粒组；D组：半乳糖化白蛋白阿霉素磁性纳米粒+外加磁场组。经舌静脉，按分组分别注射相应药物，剂量均为阿霉素2.5mg/kg体重。在注射药物前，大鼠沿腹正中线剪开腹壁，D组显露肝左叶，暴露于磁场中。按注射药物后5分钟、15分钟、30分钟、60分钟、120分钟断颈处死动物。取全血、心、肺、肝、脾、肾，D组分取靶区肝、非靶区肝。全血制成血浆，器官组织制成匀浆，盐酸乙醇法提取阿霉素，用荧光光度计测量。
结果： 静脉注射药物后，D组大鼠靶肝的ADM浓度最高，显著高于非靶肝区及A、B、C组(p＜0.05)；非靶肝区与C组肝中ADM浓度的差别无统计学意义(p＞0.05)，但均高于A、B两组(p＜0.05)；而血浆、心、肾、肺中ADM的浓度显著低于A、B组(p＜0.05)，但与C组相比较，其浓度差别无统计学意义(p＞0.05)。C组血浆、心、肾、肺中ADM浓度低于A、B组(p＜0.05)。B组肝ADM浓度高于A组，血浆、心、肾中ADM浓度低于A组(p＜0.05)，脾、肺中ADM浓度显著高于A、C、D组(p＜0.05)；而A、C、D组中脾ADM浓度差别无统计学意义(p＞0.05)。在各个测试的时间点上，随着时间
的延长，外加磁场和未加磁场的半乳糖磁性阿霉素白蛋白纳米粒组的
药物靶向指数和药物选择指数是逐渐增高的。
结论:白蛋白阿霉素磁性纳米粒经半乳糖化后，可显著增强阿霉
素对肝脏的靶向性，并显著降低心、肺、脾、’肾、血浆肝外器官的组
织阿霉素浓度。利用外加磁场，可提高阿霉素在肝脏特定部位蓄积的
能力。因此，静脉注射半乳糖化白蛋白阿霉素磁性纳米粒是可行的。
Object: to investigate distribution of adriamycin contained by galactosed albumin magnetic nanoparticle injected by lingual vein in rats.
Methods: All animals were divided into four groups which were A group: adriamycin; B group: adriamycin contained by magnetic albumin nanoparticle ; C group: adriamycin contained by galactosed albumin magnetic nanoparticle; D goup: adriamycin contained by galactosed albumin magnetic nanoparticle plus external magnetic field. Each group included 15animals, which were subdivided into 5 subgroups, thus each subgroup included 3 animals. The drugs were administrated into corresponding rats at a dosage of adriamycin 2.5mg/kg body weight. Before drugs were injected intravenously, animals were explored abdominal wall along middle line of abdomen , the left hepatic lobes of animals in D group were exposed ,which magnetite were placed on. Animal were sacrificed 5min 15 min 30min 60min 120min after drugs were injected. Blood heart lung liver kidney were harvested, targeted Liver or untargeted liver in D group was harvested. Blood samples were made into plasm, organic tissues were rinsed for homogenate. Adriamycin was pi
ck up by hydrochlore-ethanol method, which level was measured by fluorescence photometer.
Results: Adriamycin level of targeted liver tissue in D group was the highest in all groups, and not only singnificantly higher than that of untargeted in D group , but also that in A B C group(p<0.05); But
adriamycin level was not significantly different between targeted liver tissue in D group and liver tissue in C group(p>0.05) Adriamycin level of plasm heart lung kidney was not significantly different between C and D group(p>0.05), but significantly lower than that in A and B group(p<0.05) Adriamycin level of spleen in B group was the highest in all groups, but that was not significantly different in A C and D group(p>0.05) drug targeting index and drug selective index were increasing in C and D group as tested time went.
Conclusions: After adriamycin contained by galactosed albumin magnetic nanoparticle was administrated intravenously, adriamycin can be targetedly deliver to liver, and adriamycin level in plasm heart lung kidney spleen was significantly lowered in extra-hepatic organ, external magnetic field can significantly increase adriamycin level in specific site of liver tissue. So it was feasible that adriamycin contained by galactosed magnetic albumin nanoparticle was administrated intravenously.
引文
[1] Johnson PJ. Hepatitis viruses, cirrhosis, and liver cancer[J]. J Surg Oncol, Suppl, 1993,3:28～33
[2] Rettinger S, Kennedy S, Wu X, et al. Liver-directed gene therapy: quantitative evaluation of promotor elemente by using in vivo retroviral transduction[J]. Proc Natl Acad Sci, 1994,USA91:1460～1464
[3] Berger G, Reszka R, Pohlen U, et al. Liposome encapsulation of cystostatic drugs and starch microspheres improve tumor targeting in locoregional therapy. An animal experiment study of CC 531 liver tumor[J]. Langenbecks Arch Chir Suppl Kongressbd. 1998;115(Suppl Ⅰ):693～697.
[4] Norihiro Kokudo, David R. Vera, Keiichiro Tada, M. D., et al. Predictors of successful hepatic resection: prognostic usefulness of hepatic asialoglyco-protein receptor analysis[J]. World J. Surg, 2002, 26:1342-1347
[5] Mitsuru Hashida, Ken Akamatsu, Makiya Nishikawa, et al. Design of polymeric prodrugs of prostaglandin E having galactose residue for heaptocyte targeting[J]. Journal of Control Release, 1999,62:253～262
[6] Ashwell G., Harford J. Carbohydrate-specific receptorsof the liver[J]. Annu. Rev. Biochem, 1982,51:531～554.
[7] Schwartz, A. L., Rup, D., Lodish, H. F. Difficulties in the quantification of asialy-glycoprotein receptors on the rat hepatocyte[J]. J. Biol. Chem, 1980,255: 9033～9036.
[8] 张钧寿。浅谈药物传输系统研究的几个热点。中国新药杂志，1999，8(3)：215
[9] 袁东锋，易以木。肝靶向纳米粒的研究进展[J]。医药导报，2003，22(2)：113-114
[10] 龚连生，张阳德，周少波。白蛋白阿霉素磁纳米粒在大鼠体内的生物分布[J]。中国现代医学杂志，2001，11(3)：8～9
[11] 徐敏，黄德华。单克隆抗体靶向给药系统[J]。中国药房，1998，9(2)：238～239
[12] Gupta PK, Hung CT Targeted delivery of low dose doxorubicin hydroch-
oride administered via magnetic albumin microspheres in rats[J]. J Microencapsul. 1990,7(1):85～94.
[13] 张阳德，毛先海，龚连生等．给药途径对磁性白蛋白纳米粒靶向分布的影响[J]。中国现代医学杂志，2001，11(3)：11～13
[14] Frantisek S, Makiya N, Yoshinobu T, et al. Liver uptake and hepato-biliary transfer of galactosylated proteins in rats are determined by the extent of galactosylation[J]. Biochem et Biophys Acta, 1999, 1427:183～192
[15] 温守明。肝细胞半乳糖(H-Gal)受体导向药物的研究进展[J]．药学进展，1992，16(1)：1～6
[16] Nishioka Y, Kyotani S, Okamura M, et al. A study of embolizing materials for chemo-embolization therapy of hepatocellular carcinoma: effects of particle size and dose on chitin-containing cis-diamminedichloroplatinum(Ⅱ) albumin microsphere antitumor activity in VX2 hepatic tumor model rabbits[J]. Biol Pharm Bull, 1994,17(9):1251～5.
[17] Iwao Matsuda, Hiroyuki Konno, Tatsuo Tanaka, et al. Antimetastatic Effect of Hepatotropic Liposomal Adriamycin on Human Metastatic Liver Tumors[J]. Surg Today (2001) 31:414～420
[18] James BB, Jacques UB, William RB, et al. Abnormal surface distribution of the human asialogycoprotein receptor in cirrhosis[J], hepatology, 1992, 15(4):702
[19] Wisse E, De Zeanger R, Jacobs R. Lobular gradients inendothelial fenestrate and sinusoidal diameter favour centroglobularexchange processes: a scanning EM study. In: Knook DL, Wisse E (eds) Sinusoidal liver cells[J]. Elsevier, Amsterdam, (1982) pp61～67
[20] Ciechanover A., Schwartz A. L., Lodish H. F. Sorting and recycling cell surface receptors and endocytosed ligands: the asialoglycoprotein and transferrin receptors[J]. J Cell Biochem, 1983,23: 107～130
[21] Ken-ichi Ogawara, Makiya Nishikawa, Yoshinobu Takakuru, et al. Parmacokinetic analysis of hepatic uptic of galactosylated bovine serum albumin in a perfused rat liver[J]. J Control release, 1998,50:309-317
[22] NISHIKZWAM, ISHIKZWAM, MIZAKIC, et al. Glactosylated proteins are recognized by the liver according to the surface denisity of galactose
moieties[J]. AM J phsiol, 1995, 268(1): G849-G856
[23] Fallon RJ., Schwartz AL. Asialoglycoprotein receptor phosphorylation and receptor-mediated endocytosis in hepatoma cells. Effect of phorbol esters[J]. J. Biol. Chem, 1988,263: 13159～13166.
[24] Sawamura T, Nakada H, Hazama H, et al. hyper asialogycoproteinemia in pateins with chronic liver diseases and/or liver cell cacinoma[J]. Gastroenterology, 1984, 87(5):1217
[25] Sussan G, Turaj E, Seyed M, et al. Dexamethasone-loaded magnetic albumin microsphere: preparation and in vitro release[J]. Int J Pharm, 130(1996):49～55
[26] Gupta PK, Hung CT. Albumin microsphere V. Evaluation of parameter s controlling the efficacy of magnetic microspheres in the targeted delivery of adriamycin in rats[J]. Int J pharm, 59(1990):57～67
[27] Drisooll CF, Morals RM, Senyei AE, et al. Magnetic targeting of microspheres in blood flow[J]. Microvas Res, 1984,27(2):363～369
[28] Widder KJ, Morino PA, Mortis PM, et al. Selective targeting of magnetic albumin microspheres to the Yoshida Sarcoma: Ultrastructural evaluation of microsphere disposition[J]. Eur J cancer Clin Oncol, 1983; 19(1):141～147
[29] 贾博琦，鲁云兰．现代临床使用药物守则。北京医科大学出版社，2001，697
[30] 陈江浩，王执民，吴道澄等。阿霉素毫微粒肝动脉灌注后在荷肝肿瘤体内的分布研究[]。中华放射学杂志，1999，33(3)：164～166
[31] 徐萌，张积仁，许少珍等．阿霉素心脏毒性的发生机制及其防治[J]。第一军医大学学报，2001，21(7)：532～534
[32] Okuda S, Oh Y, Tsuruda H, et al. Adriamycin -induced nephropathy as a model of chronic progressive glomerular disease[J]. Kidney Int, 1986, 29:502
[33] 李云春，管昌田，张辉敏等．乳糖化和半乳糖基人生长激素的肝靶向性比较研究[J]。华西医科大学学报，1999，30(2)：138～141
[2] Rettinger S, Kennedy S, Wu X, et al. Liver-directed gene therapy: quantitative evaluation of promotor elemente by using in vivo retroviral transduction[J]. Proc Natl Acad Sci, 1994,USA91:1460～1464
[3] Berger G, Reszka R, Pohlen U, et al. Liposome encapsulation of cystostatic drugs and starch microspheres improve tumor targeting in locoregional therapy. An animal experiment study of CC 531 liver tumor[J]. Langenbecks Arch Chir Suppl Kongressbd. 1998;115(Suppl Ⅰ):693～697.
[4] Norihiro Kokudo, David R. Vera, Keiichiro Tada, M. D., et al. Predictors of successful hepatic resection: prognostic usefulness of hepatic asialoglyco-protein receptor analysis[J]. World J. Surg, 2002, 26:1342-1347
[5] Mitsuru Hashida, Ken Akamatsu, Makiya Nishikawa, et al. Design of polymeric prodrugs of prostaglandin E having galactose residue for heaptocyte targeting[J]. Journal of Control Release, 1999,62:253～262
[6] Ashwell G., Harford J. Carbohydrate-specific receptorsof the liver[J]. Annu. Rev. Biochem, 1982,51:531～554.
[7] Schwartz, A. L., Rup, D., Lodish, H. F. Difficulties in the quantification of asialy-glycoprotein receptors on the rat hepatocyte[J]. J. Biol. Chem, 1980,255: 9033～9036.
[8] 张钧寿。浅谈药物传输系统研究的几个热点。中国新药杂志，1999，8(3)：215
[9] 袁东锋，易以木。肝靶向纳米粒的研究进展[J]。医药导报，2003，22(2)：113-114
[10] 龚连生，张阳德，周少波。白蛋白阿霉素磁纳米粒在大鼠体内的生物分布[J]。中国现代医学杂志，2001，11(3)：8～9
[11] 徐敏，黄德华。单克隆抗体靶向给药系统[J]。中国药房，1998，9(2)：238～239
[12] Gupta PK, Hung CT Targeted delivery of low dose doxorubicin hydroch-
oride administered via magnetic albumin microspheres in rats[J]. J Microencapsul. 1990,7(1):85～94.
[13] 张阳德，毛先海，龚连生等．给药途径对磁性白蛋白纳米粒靶向分布的影响[J]。中国现代医学杂志，2001，11(3)：11～13
[14] Frantisek S, Makiya N, Yoshinobu T, et al. Liver uptake and hepato-biliary transfer of galactosylated proteins in rats are determined by the extent of galactosylation[J]. Biochem et Biophys Acta, 1999, 1427:183～192
[15] 温守明。肝细胞半乳糖(H-Gal)受体导向药物的研究进展[J]．药学进展，1992，16(1)：1～6
[16] Nishioka Y, Kyotani S, Okamura M, et al. A study of embolizing materials for chemo-embolization therapy of hepatocellular carcinoma: effects of particle size and dose on chitin-containing cis-diamminedichloroplatinum(Ⅱ) albumin microsphere antitumor activity in VX2 hepatic tumor model rabbits[J]. Biol Pharm Bull, 1994,17(9):1251～5.
[17] Iwao Matsuda, Hiroyuki Konno, Tatsuo Tanaka, et al. Antimetastatic Effect of Hepatotropic Liposomal Adriamycin on Human Metastatic Liver Tumors[J]. Surg Today (2001) 31:414～420
[18] James BB, Jacques UB, William RB, et al. Abnormal surface distribution of the human asialogycoprotein receptor in cirrhosis[J], hepatology, 1992, 15(4):702
[19] Wisse E, De Zeanger R, Jacobs R. Lobular gradients inendothelial fenestrate and sinusoidal diameter favour centroglobularexchange processes: a scanning EM study. In: Knook DL, Wisse E (eds) Sinusoidal liver cells[J]. Elsevier, Amsterdam, (1982) pp61～67
[20] Ciechanover A., Schwartz A. L., Lodish H. F. Sorting and recycling cell surface receptors and endocytosed ligands: the asialoglycoprotein and transferrin receptors[J]. J Cell Biochem, 1983,23: 107～130
[21] Ken-ichi Ogawara, Makiya Nishikawa, Yoshinobu Takakuru, et al. Parmacokinetic analysis of hepatic uptic of galactosylated bovine serum albumin in a perfused rat liver[J]. J Control release, 1998,50:309-317
[22] NISHIKZWAM, ISHIKZWAM, MIZAKIC, et al. Glactosylated proteins are recognized by the liver according to the surface denisity of galactose
moieties[J]. AM J phsiol, 1995, 268(1): G849-G856
[23] Fallon RJ., Schwartz AL. Asialoglycoprotein receptor phosphorylation and receptor-mediated endocytosis in hepatoma cells. Effect of phorbol esters[J]. J. Biol. Chem, 1988,263: 13159～13166.
[24] Sawamura T, Nakada H, Hazama H, et al. hyper asialogycoproteinemia in pateins with chronic liver diseases and/or liver cell cacinoma[J]. Gastroenterology, 1984, 87(5):1217
[25] Sussan G, Turaj E, Seyed M, et al. Dexamethasone-loaded magnetic albumin microsphere: preparation and in vitro release[J]. Int J Pharm, 130(1996):49～55
[26] Gupta PK, Hung CT. Albumin microsphere V. Evaluation of parameter s controlling the efficacy of magnetic microspheres in the targeted delivery of adriamycin in rats[J]. Int J pharm, 59(1990):57～67
[27] Drisooll CF, Morals RM, Senyei AE, et al. Magnetic targeting of microspheres in blood flow[J]. Microvas Res, 1984,27(2):363～369
[28] Widder KJ, Morino PA, Mortis PM, et al. Selective targeting of magnetic albumin microspheres to the Yoshida Sarcoma: Ultrastructural evaluation of microsphere disposition[J]. Eur J cancer Clin Oncol, 1983; 19(1):141～147
[29] 贾博琦，鲁云兰．现代临床使用药物守则。北京医科大学出版社，2001，697
[30] 陈江浩，王执民，吴道澄等。阿霉素毫微粒肝动脉灌注后在荷肝肿瘤体内的分布研究[]。中华放射学杂志，1999，33(3)：164～166
[31] 徐萌，张积仁，许少珍等．阿霉素心脏毒性的发生机制及其防治[J]。第一军医大学学报，2001，21(7)：532～534
[32] Okuda S, Oh Y, Tsuruda H, et al. Adriamycin -induced nephropathy as a model of chronic progressive glomerular disease[J]. Kidney Int, 1986, 29:502
[33] 李云春，管昌田，张辉敏等．乳糖化和半乳糖基人生长激素的肝靶向性比较研究[J]。华西医科大学学报，1999，30(2)：138～141