PAI PAI-1 PAI-1 PAI-1 PAI-I PAI-1 MMP Leptin [5,6] FFA PAI-1 [9,10] 1 ZJN0504

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1 PAI-1 * erjinzhi@126.com -1 PAI-1 PAI-1 PAI-1 3 PAI-I PAI ECM ECM MMP ECM ECM [1] PA / PA PAI PAI 3 PAI PAI-1 PAI PAI-2 Nexin-1 [2] PA/ PAI-1 PAI-2 α 2 - α 2 -AP PN-I PAI-1 PAI [3,4] Leptin -α TNF-α -6 (IL-6) -1 PAI-1 [5,6] FFA PAI-1 [7] PA PAI-1 ECM PAI-l PA / ECM PAI-1 ECM ECM [8] PA/PAI-1 PAI-1 [9,10] 1 ZJN0504 1

2 O U.K PAI-1 FITC- Santa Cruz propidium iodide, PI TritonX-100 Sigma 2.3 D P L 5 I & O 80 8µm min 3 1,2-2 5min O 10 85% 50%1,2-1 2min 1 2 PBS min PBS min ph7.2~ TritonX mol/L PBS 4µg/ml 4µg/ml IgG 4 PBS 3 FITC min PBS mg/ml PI 5min PBS 3 DABCO laser scanning confocal microscopy, LSCM Image Pro Plus 5.0 / SPSS ± P < A D 2

3 1 2 1E F A D30 B D40 C D50 D D60 E P1 F P6 G P12 H P18 I L4 J L8 K L12 L L16 M L20 N L24 O L28 P L32 Q I24h R I48h S I72h T I96h O Fig1 Adipose tissue in mouse mammary gland ~21 1I 1J~L

4 M~P 72 1S 1T 3.2 PAI-1 PAI-1 3 2A PAI PAI PAI-1 72 PAI-1 2B A B D: P: L: I: n=15 P< PAI-1 Fig.2 Analyses of PAI-1 expression during mouse mammary gland development 3 PAI-1 40 PAI-1 PAI-1 PAI-1 6 PAI-1 24 PAI-1 PAI-1 PAI-1 PAI-1 4

5 PAI-1 48h PAI-1 72h 96h PAI-1 PAI-1 A D30 B D40 C D50 D D60 E P1 F P6 G P12 H P18 I L4 J L8 K L12 L L16 M L20 N L24 O L28 P L32 Q I24h R I48h S I72h T I96h PI = ~ 50µm 40 3 PAI-1 Fig3 Localization of PAI-1 in mouse mammary gland by LSCM 4 PAI-1 3 PAI-1 PAI-1 Leptin PAI-1 PAI-1 [11] 40 5

6 PAI-1 [12] PAI-1 PAI-1 PAI-1 PAI-1 [13,14] PAI-1 PAI-1 [15,16] IGFBP-5 αs 2 - PAI-1 IGFBP-5 IGFs αs 2 - PAI-1 tpa [13,14] AM Sorrell 2006 IGF-I PAI-1 IGFBP-5 HC11 PAI-1 tpa urokinase-type plasminogen activator, upa IGFBP-5 PAI-1 PAI-1 tpa IGFBP-5 PAI-1 IGFBP-5 tpa [17] IGFBP-5 [18] IGFBP-5 IGFBP-5 [19] IGFBP-5 feedback inhibitor of Lactation, FIL IGFBP-5 ECM [15,16] PAI-1 ECM [1] P.Mignatti. Extracellular matrix remodeling by metalloproteinases and plasminogen activators [J]. Kidney Int, 1995, 47(suppl 49):S12. [2] D.Collen. Report of the meeting of the subcommittee on fibrinolysis. San Diege, July 13, Thromb Haemost, 1985, 54:893. [3] J.D.Vassalli, Ap.Sappino and D.Belin. The plasminogen Activator/plasmin system [J]. J Clin Invest, 1991, 88:1067. [4] P.Trayhurn and J.H.Beattie. Physiological role of adipose tissue: white adipose tissue as an endocrine and secretory [J]. Proc Nutr Soc, 2001, 60 (3): [5] M.A.Lazar. How obesity causes diabetes: not a tall tale [J]. Science, 2005, 307(5708): [6] J (3): [7] K.Hara, M.Horikoshi, T.Yamauchi, et al. Measurement of the high-molecular weight form of adiponectin in plasma is useful for the prediction of insulin resistance and metabolic syndrome [J]. Diabetes Care, 2006, 29(6): [8] W.H.Baricos and S.L.Cortez. E1-Dharss et al. ECM degradation by cultured human mesangial cell is mediated by a PA/plasmin/MMP-2 cascade [J]. Kidney Int, 1995, 47:

7 [9] A.Festa, R Jr D'Agostino, R.P.Tracy, et al. Elevated levels of acute-phase proteins and plasminogen activator inhibitor-1 predict the development of type 2 diabetes: the insulin resistance atherosclerosis study [J]. Diabetes, 2002, 51(4): [10] A.Festa, R Jr D'Agostino, K.Williams, et al, Mayer-Davis EJ, Tracy RP, Haffner SM. The relation of body fat mass and distribution to markers of chronic inflammation [J]. Int J Obes Relat Metab Disord, 2001, 25(10): [11] D 2006 [12] G.B.Silberstein. Postnatal mammary gland morphogenesis [J]. Microsc Res Tech, 2001, 52(2): [13] T.J.Nam, J.R.Walker Busby and R.David. Clemmons. Insulin-Like Growth Factor Binding Protein-5 Binds to Plasminogen Activator Inhibitor-I [J]. Endocrinology, 1997, 38(7): [14] G.J.Allan, J.Beattie and D.J.Flint. The role of IGFBP-5 in mammary gland development and involution [J]. Domest Anim Endocrinol, 2004, 27(3): [15] J.Wiesen and Z.Werb. Proteinases, cell cycle regulation, and apoptosis during mammary gland involution [J]. Mol Reprod Dev, 2000, 56(4): [16] C.H.Knight, M.Peaker and C.J.Wilde. Local control of mammary development and function [J]. Rev Reprod, 1998, 3(2): [17] A.M.Sorrell, J.H.Shand, E.Tonner, et al. Insulin-like growth factor-binding protein-5 activates plasminogen by interaction with tissue plasminogen activator, independently of its ability to bind to plasminogen activator inhibitor-1, insulin-like growth factor-i, or heparin [J]. J Biol Chem, 2006, 281(16): [18] E.Marshman, K.A.Green, D.J.Flint, et al. Insulin-like growth factor binding protein 5 and apoptosis in mammary epithelial cells [J]. Journal of Cell Science, 2003, 116(4): [19] D 2006 The Expression of PAI-1 during Mouse Mammary Gland Development Zhao Feng, Li Qingzhang* Key Laboratory of Dairy Science of Ministry of Education, Northeast Agricultural University Harbin, PRC, (150030) Abstract In this research, some time points were set in all stages of mouse mammary gland development. Histology and immunofluorescence assay were used to detect plasminogen activator inhibitor-1(pai-1) by a laser scanning confocal microscope in mammary gland slices. The results of localization and quantification revealed that: PAI-1 distributed not only in adipose tissue but also epithelium and matrix.pai-1 expression level up-regulated three times during mammary gland development, which unrelated with the number of adipocytes, but related with the change of mammary gland structure. High level of PAI-1 expression inhibited extracellular matrix degrading by plasmin and limited adipocytes proliferating, therefore related with mammary gland devlopment. But enhanced PAI-1 expression couldn t reverse involution trend due to milk accumulation. Keywords: mouse, mammary gland, adipocyte, PAI-1, LSCM