Ch in. J. B iochem. M o l. B io l. 2000, V o l. 16, N o. 4, cd NA

Transcript

1 ISSN CN gQ 16 4 Ch in. J. B iochem. M o l. B io l. 2000, V o l. 16, N o. 4, cd NA 3 (, ) (T PO ) T PO pcdna 3hT PO pcdna 3, EL ISA T PO ( IGF ) 2g g (PD GF), T PO, % 513% 411% 4138% (P < ) 4 16, EL ISA,, IGF s PD GF,, IGF2g, T PO,,, Q 527 Human Throm bopo ietin cd NA Tran sfer Prom otes M ice Growth A ssoc ia ted w ith an In sulin- like Growth Factor ZHAO J ian2zeng, CH EN L i2m in,m E I Y ing2jie, ZHAN G Guo2rong,W AN G X iao2p ing (L aboratory of Gene T herapy, Center f or C linical M olecu lar B iology, General H osp ital of A ir F orce, B eij ing , Ch ina) Abstract To study the effect of th rom bopo ietin (T PO ) gene delivery on the grow th of m ice, hum an T PO p lasm id DNA encap su lated w ith lipofectin w as in jected in to m ou se m u scle. A nd fem ale m ice of 8-12 w eek s o ld w ere found to have a sign ificen t body w eigh t gain com pared w ith the con t ro ls fo llow ing the 1st, 2nd, 3rd and 4th w eek ( P < 01005, 01002, 0101 and 0101 respectively), w h ich had a characteristic of rap id grow th (P < 0101) in liver and sp leen. How ever, the change of body w eigh t in such m ice in jected w ith T PO cdna as the m ales of the sam e age, the young ones of less than 4 w eek s o r the fem ales of m o re than 16 w eek s w ere no t sign ifican tly differen ṫ Sem i2quan titative EL ISA analysis dem on strated that the serum levels of p latelet derived grow th facto r (PD GF ), in su lin2like grow th facto rs ( IGF2) g and g in the delivered m ice ro se co rrespondingly. Serum gluco se, triglyceride, and alkaline pho sphatase w ere also increased. It is suggested that som e o ther grow th facto rs, especially IGF21, m ediate part of in v ivo activities of T PO. Key words T h rom bopo ietin, Grow th2p rom o tion, IGF2g, IGF2g, PD GF (T PO ), T PO [ 1, 2 ], T PO, 3 ( ) :,, ,, T el: (010) E2m ail: m ayhappy@public. easṫ cn. net : , :

2 4 : cdna 525 T PO, T PO (p latelet2 115 derived grow th facto r, PD GF) 2Β IGF2g g PD GF 4 Β2 T PO, [ 3 5 ] T PO EL ISA : 30 l PBS 100 l [ 6,, ], 4 96 :, T PO 3% 5% Tw een 20 PBS [ 7 Α2 ], 015 g 2 h; [ 8 ], T PO 1 h, 3, 3 5, 5 2 [ 9 ] [ 10 ] (Gibco BRL ), 015 m o lgl H 2SO nm, T PO cdna, T PO Balbgc 211 TPO cd NA [ 11 ] T PO, IFN 2Χ IL 22 TN F2Α g 12 T PO 1, [ 12 ],, F ig. 1 Balbgc T PO 4, T PO 413% 513% 411% 4138%,, ( IGF) 2g, g PD GF TPO cd NA T PO pcdna 3ghT PO, [ 11, ] pcdna ( ) Balbgc KM, ( 10 ), 60 g lipofectin (G IBCO BGL ) pcdna 3g ht PO DNA, pcdna 3, 113 ( in su lin2like grow th facto r, IGF ) 2g g PD GF2 AA (Pep re T ech) 2 Balbgc KM, 8 (P ) 20 (4 ) T PO, ( ) T PO F ig. 1 D ynam ic body w eigh t of ht PO cdna transferred 2 lgg , T PO,, m ice T he show n w as the body w eigh t of 8210 w eek s ages Balbgc m ice. Significant difference w as observed betw een the TPO transfer and the contro l group s(p < 01005, , and from the 7 d to 28 d, respectively)

3 (P 214 TPO < 0101), T PO, :, ;, ;, (P < 0105), T ab le 1 8 Χ2 Α2, d Table 1 T he o rgans w eigh t ( g ) in KM m ice transferred w ith ht PO cdna (x θ s) Group n N o rm al contro l pcdna 3 pcdna 3ghTPO T hym us Sp leen H eart L ung L iver Kidney P < 0101 compared w ith contro l 213, EL ISA T PO IGF2g g, PD GF F ig. 2 Balbgc T PO 14 d 3, PD GF IGF2g g ht PO T PO DNA, PD GF IGF s 7 d, T PO 14 d T PO,, IGF2g g,,,,, T PO T PO, PD GF IGF2 ( ) F ig. 2 Grow th facto r levels in circulation ( subjects from fem ale Balbgc m ice at the 14 th day) T PO T ab le 2 Table 2 T he blood biochem istry (mo lgl ) levels of the m ice transfected w ith pcdna 3ghT PO (x θ s) Contro l pcdna 3 pcdna 3ghTPO GLU CHOL TR IG ALB T P LDH A KP GGT (Χ2GT ) Α2HBDH AM Y P < 0105, compared w ith no rm al contro l, T PO g g,, IGF2g g PD GF T PO, IGF2g, IGF2g T PO PD GF, PD GF (GH ) GH, IGF

4 4 : cdna 527 T PO ligand ), GH 3, IGF2 4 O zak i S, Ko saka M, O zak i K, W akatsuk i S, H ash imo to T, g 6 8 [ ], T PO, 8 12, IGF2g IGF2g, T PO cdna T PO IGF2g, IGF2g IGF2g, IGF2g g [ ] IGF2g [ 17 ],, T PO 1) : 63a cdna, IGF ht PO cdna Balbgc T CD 4 + [ 21 ] IGF2g g CD 4 + T, IGF T, IGF s T, ht PO PD GF IGF2g g CD ,,,., T PO IGF s. (References) 1 de Sauvage F J, H ass P E, Spencer S D,M alloy B E, Gurney A L, Spencer S A, D arbonne W C, H enzel W J,W ong S C, Kuang W J,O les K J, H ultgren B, So lberg L A J r, Goeddel D V, Eaton D L. Stim ulation of m egakaryocytopo iesis and throm bopo iesis by the c2m p l ligand. N atu re, 1994, 369: L ok S, Kaushansky K, Ho lly R D, Kujiper J L, L ofton2d ay C E, Oo rt P J, Grant F J, H elpel M D, Burkhead S K, Kram er J M, Bell L A, Sp recher C A, B lum berg H, Johnson R, P runkard D, Ch ing A F T,M athew es S L, BalleyM C, Fo rstrom J W, Buddle M M, O sbo rn S G, Evans S J, Sheppard P O, P resnell S R, O H ara P J, H agen F S, Ro th G J, Fo ster D C. C loning and exp ression of the m urine th rom bopo ietin cdna and stim ulation of p latelet p roduction in v ivo. N atu re, 1994, 369: Yan X Q, L acey D, F letcher F, H artley C, M ce lroy P, Sun Y, X ia M, M u S, Saris C, H ill D, H aw ley R G, M cn iece I K. Chronic expo sure to retroviral vecto r encoded M GD F (mp l2 induces lineage2specific grow th and differentiation of m egakaryocytes in m ice. B lood, 1995, 86: M iyazaki H. Throm bopo ietin2responsible essential th rom bocythaem ia w ith m yelofibro sis. B r J H aem atol, 1997, 97: W ak ikaw a T, Sh io i A, H ino M, Inaba M, N ish izaw a Y, T atsum i N, M o rii H, O tani S. Throm bopo ietin inhibits in v itro o steoclastogenesis from m urine bone m arrow cellṡ E nd ocrinology, 1997, 138: Kubo ta Y, Kash im a M, Ko ish i M, Kaw a Y, M izoguch i M. Effects of throm bopo ietin on the p ro liferation and cell adhesion mo lecule exp ression of hum an endo thelial cells (abstract). E xp H em atol, 1996, 24: Fujimo to T, Katsutani S, Fujii T, Kim ura A, M iyazak i H, Fujim ura K. Throm bopo ietin induces the exp ression of one of the interferon2inducible gene fam ily, 9227: an analysis of the throm bopo ietin action ( abstract). B lood, 1996, 88 ( 10 supp l. 8 Bobik R, Hong Y, B reier G, M artin J F, E rusalim sky J D. Throm bopo ietin stim ulates V EGF release from c2m p l2 exp ressing cell lines and haem atopo ietic p rogenito rs. F EB S L ett, 1998, 423: Cardier J E, D emp sey J. T h rom bopo ietin and its recep to r, c2 mp l, are constitutively exp ressed by mouse liver endo thelial cells: evidence of throm bopo ietin as a grow th facto r fo r liver endo thelial cells. B lood, 1998, 91: M itjavila M T, F ilipp i M D, Cohen2So lal K, L e Pesteur F, V ainchenker W, Sainteny F. T he M p l2ligand is invo lved in the grow th2p romo ting activity of the m urine strom al cell linem S25 on ES cell2derived hem atopo iesis. E xp H em atol, 1998, 26: 124 (Zhao J Z, L iu L, Chen H R,M ei Y J. Exp ression of hum an th rom bopo ietin cdna in m ice and the effects on p ro liferation of hem atopo ietic p rogenito r cells. Chin J B iochem M ol B iol ), 1998, 14 (2) : ,,.. (Zhao J Z, L iu L,M ei Y J. Effects of throm bopo ietin gene delivery on the concentration of serum cytok ines in m ice. Ch in J Imm unol) 1999, 15: Jansen M, van Schaik F M A, R icker A T, Bullock B,W oods D E, Gabbay K H, N ussbaum A L, Sussenbach, V an den B rande J L. Sequence of cdna encoding hum an insulin2like grow th facto r g p recurso r. N atu re, 1983, 306: Stemp ien M M, Fong N F, Ball L B, Bell G İ Sequence of a p lacental cdna encoding the mouse insulin2like grow th facto r g p recurso ṙ DN A, 1986, 5: Palm iter R D, B rinster R L, H amm er R E, T rum bauer M E, Ro senfeld M G, B irnberg N C, Evans R M. D ram atic grow th of m ice that develop from eggs m icro injected w ith

5 m etallo thionein2grow th ho rmone fusion genes. N ature, 1982, 300: Palm iter R D, N o rstedt G, Gelinas R E, H amm er R E, B rinster R L. M etallo th ionein2hum an GH fusion genes stim ulate grow th of m ice. S cience, 1983, 222: M athew s L S, H amm er R E, Beh ringer R R, D erco le A J,Bell G I, B rinster R L, Palm iter R A. Grow th enhancem ent of transgenic m ice exp ression hum an insulin2like grow th facto r g. E nd ocrinology, 1988, 123: Koo ijm an R, W illem s M, R ijkers G T, B rinkm an A, van Buul O ffers S C, H eijnen C J, Zegers B J M. Effects of insulin2like grow th facto rs and grow th ho rmone on the in v itro p ro liferation of T lymphocytes. J N eu roimm unol, 1992, 38: Koo ijm an R, van Buul2O ffers S C, Scho ltens L E, Schuurm an H J, V an den B rande L J, Zegers B J M. T cell developm ent in insulin2like grow th facto r2g transgenic m ice. J Imm unol, 1995, 154: C lark R, Strasser J M ccabe S, Robbins K, Jardieu P. Insulin2 like grow th facto r21 stim ulation of lymphopo iesis. J C lin Invest, 1993, 92: ,,. T. ( Zhao J Z, Guo Z K, M ei Y J. T h rom bopo ietin cdna transfer p romo tes the p ro liferation and differentiation of T lymphocytes in m ice. J Imm unol) 1999, 15: , 1985, 1996,,,, 2000, :,,, 16 (A 4), 100, 8100 ( ), : : ( ) ; : (029) , E2m ail: imm uedit@fmm u. edu. cn