2003-2004 > Academic research: refereed > Publication in refereed journalVersion of RecordPublishe

Du, XL

Katie, SKY

Li, RC

Qian, ZM

Sun, HZ

Wang, K

2003-2004 > Academic research: refereed > Publication in refereed journalVersion of RecordPublishedVoR allowe

PolyU Institutional Repository

　[ ????] 2002-04-23　　[ ????] 2002-07-07＊[ ????] ??????????(A-PC23 , A-PC98),????????????????????????(2000-1)[ ????] 　1000-4718(2003)07-0902-03?????????????? U-87MG???????＊王　夔1 , 　杜秀莲1 ,2 , 　常彦忠2 , 　李荣昌1 , 　司徒加怡3 , 　孙红哲3 , 　钱忠明2( 1???????????? , ?? 100083;2??????????????????????? ,??;3??????? , ??)　　[ ?　?] 　??:??????/??????????????????(Yb2Tf)??????????? U-87 MG?? , ??????????????????? U-87MG???????。??:????? ICP-MS????。??:? Yb2Tf???? , ????????。???? 2μmol/L? , ?????????????。?????????:apo Tf???????? , ?????? 1.5 ? ,?????????。0.4μmol/ L Yb2Tf????? U-87 MG ???? ,? Yb3+????10μmol??????????? 。??:????/?????????????????? U-87MG???????。?????? Yb3+??????? U-87 MG ?????。[ ???] 　Yb3+/ Yb2Tf;????/??????;?????;U-87 MG ??　　[ ?????] 　R363　　　　　[?????] 　A　　????(transferrin , Tf)????(Fe3+)?????。??????(transferrin receptor , TfR)?????(endocytosis)? Tf???????????????[ 1 ,2] 。??????????????????Tf/TfR????????[ 3 ,4] ,????????? Tf/TfR????????????????????????[ 3] 。? Tf/TfR????????????????????????? TfR????。TfR???? ,??????[ 1 ,2] 。??????????? ,?????????? TfR[ 5] 。???????Fe3+?? Tf?TfR????????????????????????。???????????????? G0/G1 ?? S ??? ,???? B16????????[ 6] 。?????????????? ,???????? Tf/TfR?????? ,??????????? 。???????? ,????? Tf/TfR??????????? U-87MG ???????????????。?　?　?　?　?1　??????????(U-87MG??)?? ATCC?? 。????? ,MEM ??? ,??????? ,10%????(Gibco)? Yb2O3 ?? Sigma ?? 。75cm2???? pronase ???? IWAKI ? Roche ??。????????? 。450 ????????????????(ICP -MS)??? PE ??(ELAN5000)? Bio-Rad?? 。2　???????????? U-87 MG??Yb3+?????　　U-87 MG?????? 10×104 U/L ??????? ,10%????(Gibco)? MEM ???? 。1∶3?? ,?3 d? 1? 。????? 80 %? ,????。????。???? ,???? 24 h。? 0.1mol/L Hepes(? 0.1 mol/L NaCl , pH=7.4)???????????。?? ,?? MEM ?????? ,?37℃?? 2 h ,?? 0.1 mol/L Hepes ?????? ,??????????[ 7 , 8] 。???? Yb2Tf?0.1 mol/L Hepes ,??? ,???????? 0.5 , 1 ,2 , 5? 10 μmol/L ,?? 37℃?? 30 min ?????。? 0.1 mol/L Hepes???? 3-4?? ,?? 2mL (1 g/L)pronase , ?????(4℃)1 h ,???????? 。? Pronase ?????????????TfR??????? ,?????????????????? Yb2Tf[ 9 ,10] 。???? 24 h? ,????? ,???????。? ICP-MS ?????Yb3+?[ 11] 。?????5?????。?????? ,???? pronase(1 g/L)??????????30 min ,???? Yb2Tf[ 12] ,???????? 。?????? Yb3+???????(apotransferrin , apo-Tf)????????????? 。????????????????? 1 μmol/L ,? 0 , 0.5∶1 , 1∶1 , 1.5∶1 , 2∶1???(the mole ratio)???????·902· ????????　Chinese Journal of Pathophysiology　2003 , 19(7):902-904Yb3+ ,37℃??30 min? ,?????? 。3　Yb3+ ,Yb2Tf ? U-87 MG????????? U-87 MG ??? 96 ???? ,?? 2×104??? 。24 h ? ,?? Yb3+? Yb2Tf????????? 0 , 0.5 , 2.5 , 5 , 10 μmol/L。? MTT ???(Roche ??)???????。? 450 ???? , 595 nm ??? 。?????? 8????。?????( x ±s)??。?　?　?　?　?1　???????????? U-87MG ??Yb3+?????　　? 1???? ,? Yb2Tf ????? ,????????? 。???? 2 μmol/L ? ,????????????? ,???????????????。?? ,??????????????? 。? 2? apoTf???????????????? ,???? apoTf???????????? 。?????? ,????????? 。????????????????????。????? 0.5 , 1.0?1.5? ,???????(3.76±0.80)ng/106 cells ,(6.37 ±1.10)ng/106 cells ?(9.67 ±2.10)ng/106cells。?????? 2.0??????(9.61±1.00)ng/106 cells???? 1.5??? 。????? ,??Tf???????? ,???????????????????? 。??????? ,? Tf????????? ,?Tf????????? ,?????????????????????????。???????????????? pronase???30 min???????? Yb2Tf ????? Tf/TfR??? U-87 MG ?????????????????。Fig 1　Effect of different concentrations of Yb2Tf on Yb3+uptake byU-87MG cells. x±s.n=5.? 1　Yb2Tf??? U-87 MG????????Fig 2　Effect of the mole ratio of Yb3+ to apoTf on Yb3+ uptake byU-87MG cells. x±s.n=5.?2　Yb3+? Tf??????U-87 MG????????Fig 3　Inhibition of Yb3+(★)and Yb2Tf(○)on the proliferationof U-87MG cells. x±s.n=8.＊P<0.05 , ＊＊P<0.01vs the control(0μmol/ L).?3　Yb3+(★)? Yb2Tf(○)??? U-87 MG???????2　Yb3+? Yb2Tf?????????????Yb3+? Yb2Tf? U-87 MG ?????????????(? 3), Yb3+?? 5 μmol/L ?????????????(P <0.05),? 10 μmol/L Yb3+?????????(P >0.05)。?? , Yb2Tf ?????? 0.4 μmol/L ? ,?????????????(P<0.05),??????? Yb2Tf??????????。???? 2.5 μmol/L ????????。????? Ga3+? In3+???????[ 12] 。Ga3+????????????? Ga2Tf ??????????????????????????? ,????????????[ 13] 。?????Yb2Tf???????????????。?? ,?Ga3+?? ,??????????????????。????????? Yb2Tf ? U-87 MG ???????????? U-87 MG ????????????? ,???????Yb2Tf??????????????。?????????????·903·?????????????。?? ,???????? Yb3+??????? ,??? Yb2Tf ?? ,Yb3+??????????? 。?????? ,????Tf???????????U-87MG??? ,??Tf???Yb?????????。??????? Yb2Tf????????????????????。　　??????????/?????????????????? U-87 MG????????。?????? Yb3+??????? U-87 MG?????。??????????。[ ?　?　?　?][ 1] 　Qian ZM , Tang PL.Mechanisms of iron uptake by mammaliancells[ J] .Biochim Biophys Acta , 1995 , 1269(3):205-214.[ 2] 　Qian ZM , Wang Q.Expression of iron transport proteins andexcessive iron accumulation in the brain in neurodegenerativedisorders[ J] .Brain Res Rev , 1998 , 27(3):257-267.[ 3] 　Li HY , Sun HZ , Qian ZM.The role of transferrin/ transferrinreceptor system in drug delivery and targeting[ J] .Trend Phar-macol Sci , 2001 , 23(5):206-209.[ 4] 　Roskams AJ , Connor JR.Aluminum access to the brain:arole for transferrin and its receptor[ J] .Proc Natl Acad SciUSA , 1990 , 87(22):9024-9027.[ 5] 　Testa EP , Testa U , Samoggia P , et al.Expression of transfer-rin receptors in human erythroleukemic lines:regulation in theplateau and exponential phase of growth[ J] .Cancer Res ,1986 , 46(10):5330-5334.[ 6] 　Sato T , Hashizume M , Hotta Y, et al.Morphology and prolif-eration of B16 melanoma cells in the presence of lanthanoidand Al3+ ions[ J] .Biometals , 1998 , 11(2):107-112.[ 7] 　Qian ZM , Morgan EH.Changes in transferrin-free and transfer-rin-bound iron uptake by cells during reticulocyte maturation invivo and in vitro[ J] .Biochim Biophys Acta , 1992 , 1135(1):35-43.[ 8] 　Qian ZM , Xiao DS , Tang PL , et al.The increased expressionof transferrin receptor on the membrane of erythroblast in stren-uous exercised rats[ J] .J Appl Physiol , 1999 , 87(2):523-529.[ 9] 　Qian ZM , Pu YM , Wang Q , et al.Cerebellar granule cellsacquire transferrin-free iron by a carrier-mediated process[ J] .Neuroscience , 1999 , 92(2):577-582.[ 10] Xie JX , Tsoi YK , Chang YZ , et al.Effects of ferroxidase ac-tivity and species on ceruloplasmin mediated-iron uptake byBT325 cells[ J] .Mol Brain Res , 2001 , 99(1):12-16.[ 11] ??? ,???.????????????????????????????[ J] .???????? ,2000 , 16(12):1317-1319.[ 12] Tallkvist J , Bowlus CL, Lonnerdal B.Functional and molecu-lar responses of human intestinal Caco-2 cells to iron treat-ment[ J] .Am J Clin Nutr , 2000 , 72(3):770-775.[ 13] Moran PL , Seligman PA.Effects of transferrin -indium oncellular proliferation of a human leukemia cell line[ J] .Can-cer Res , 1989 , 49(15):4237-4241.[ 14] Harris AW , Sephton RG.Transferrin promotion of 67Ga and59Fe uptake by cultured mouse myeloma cells[ J] .CancerRes , 1977 , 37(10):3634-3638.Transferrin-bound Yb2 uptake by U-87 MG cells andeffect of Yb on proliferation of the cellsWANG Kui1 , DU Xiu-lian1, 2 , CHANG Yan-zhong2 , LI Rong-chang1 ,SZETO Ke Yee Katie3 , SUN Hong-zhe3 , QIAN Zhong-ming2(1Department of Chem-biology , Institute of Pharmaceutics , Beijing University , Beijing 100083 , China;2Laboratory of Iron Metabolism , Department of Applied Biology &ChemicalTechnology , The Hong Kong Polytechnic University , Kowloon , HongKong , China;3Department of Chemistry , The University of Hong Kong , Hong Kong , China)　　[ ABSTRACT] 　AIM:To investigate the role of transferrin/ transferrin receptor system in transferrin-bound Yb2(Yb2Tf)uptake by U-87 MG cells and the effect of transferrin-bound and -free Yb2 on proliferation of U-87 MGcells.METHODS:Cell culture and ICP-MS measurement of Yb2.RESULTS:Yb2Tf uptake by U-87MG cells in-creased with the concentrations of Yb2Tf , and reached saturation as the concentration in the incubation medium was raisedto about 2μmol/L.Also , Yb2 uptake by the cells increased with increase of the mole ratio (Yb2:apoTf), reaching amaximum at 1.5 mole ratio.Yb2Tf in 0.4μmol/L significantly inhibited proliferation of U-87MG cells , however , 10μmol/L Yb3+ had no significant effect on proliferation of the cells.CONCLUSION:The uptake of Yb2 by U-87MGcells might be mediated by transferrin/ transferrin receptor system.Transferrin-bound but not transferrin-free Yb2 couldsignificantly inhibit proliferation of U-87MG cells.[ KEY WORDS] 　Yb3+/ Yb2Tf;Transferrin/ transferrin receptor;Glioma;U-87 MG cells·904·

Transferrin-bound Yb2 uptake by U-87 MG cells and effect of Yb on proliferation of the cells

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