肝硬化患者骨髓间充质干细胞的体外培养及生长特性

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安方梅1 , 邓永东2 , 赵正斌2 ,陈　红2
" Z8 {  R- P1 y! J(1. 兰州大学临床医学院,甘肃兰州730000 ;2. 兰州大学第一医院感染科,甘肃兰州730000)( a' X) V. D" }7 H
摘　要:目的　研究肝硬化患者骨髓间充质干细胞(MSCs) 的体外扩增方法及其生长特性和特异性表面标记,为肝+ o2 U& K+ z  e/ G0 b$ c" y3 h
组织工程“种子细胞”的选择提供新的理论依据,为进一步研究自体骨髓间充质干细胞移植治疗肝衰竭时移植细胞
+ O8 k0 t" o0 N) a4 \. N4 [7 ^在体内的调控机制打下基础。方法　无菌抽取4 例肝硬化志愿者骨髓3～4 mL ,年龄30～40 岁,利用密度梯度离
0 u+ Y5 j& U7 w心法联合贴壁筛选法逐步筛选MSCs ,获取MSCs ,倒置显微镜下逐日观察细胞生长特性,细胞计数,绘制细胞生长! t$ t* O7 Y" e! m$ _
曲线,流式细胞仪检测细胞表面标志CD44 、CD45 。结果　原代培养细胞生长潜伏期0～9 d ,对数增殖期为10～193 }4 ]) b9 `7 {5 p: X2 @
d ,生长平台期为20～24 d ;传代细胞生长周期较原代细胞快,细胞生长潜伏期为4～24 h ,对数增殖期为3～12 d ,
, K9 d. f" R  x% x4 S/ g* w2 l13～15 d 进入生长平台期。肝硬化患者骨髓MSCs 高表达CD44 ,低表达CD45 。结论　肝硬化患者骨髓MSCs 体. m( T, E5 b( |5 @7 S0 N! j2 P
外生长良好,性质稳定,可以作为“种子细胞”用于自体骨髓干细胞移植和生物人工肝系统。- A6 R& u  N/ o, s- \/ u1 h$ v6 n0 f
关键词:肝硬化;骨髓间充质干细胞; 自体骨髓干细胞;移植
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- l! b, E' p! SI n vit ro culture and growth characteristics of bone marrow
8 j& }) d. D* smesenchymal stemcells from liver cirrhosis patients- H8 D7 P" [& R4 O% n& W
AN Fang2mei1 ,DEN G Yong2dong2 , ZHAO Zheng2bin2 , CHEN Hong2/ T! R8 @9 v4 `  V7 M
(1. S chool of Cl i nical Medici ne , L anz hou Uni versi t y , L anz hou , 730000 , Chi na;& o6 V. r( Y0 V2 T
2. Dep artment of I nf ect ion , T he Fi rst Hos pi tal of L anz hou Uni versi t y ,0 f+ B" S* k3 C$ G
L anz hou , 730000 , Chi na )& S5 ~" p) {# g+ g/ N% C& {
Abstract : Ob j ectives To st udy the growt h pat tern and surface markers of i n vi t ro cult ured bone2 H0 B. C3 f% ]5 a8 K! \
marrow mesenchymal stem cell s (MSCs) f rom liver cirrhosis patient s , provide new evidence for
, I0 n7 F- k+ W7 V1 Ut he choice of " seed cell s" for liver tissue engineer and lay basis for f urt her study of t he i n vi vo
* R. e8 p% N5 @! lregulatory mechanisms of t ransplanted autologous bone marrow MSCs in t he t reatment of liver
" [1 e; O2 d8 F5 J. |4 f  ^+ _failure. Met hods Bone marrow cell s were collected f rom 4 volunteer s aged f rom 30 to 40 wit h liver" k* H1 a0 K6 f& Y
cirrhosis , MSCs were separated by a density gradient cent rif ugation , biological features of cul2( `& e' l4 r5 a# C- _- p9 f
t ured MSCs were observed under inverted microscope and their growt h curves were investigated.' d' M; c! c1 e, @
Flow cytomet ry was used to detect surface antigenic markers of CD44 and CD45. Resul ts The la2
, q4 c6 H4 B+ K; V. o7 ltent p hase of the p rimary cultured cell s was 0 to 9 d followed by 10 to 19 d of logarithmical prolif2
! a9 b. b. s- eeration , and 20 to 24 d of platform growth. Passaged cells showed faster growt h t han primary
5 ^. w1 X, [7 q. Fcell s , and t heir latent p hase was 4 to 24 h , logarit hmical proliferation p hase was 3 to 12 d and
( v6 r: G8 @9 Rplatform growth was 13 to 15 d , MSCs expressed higher CD44 but lower CD45. ConclusionMSCs f rom liver cirrhosis patient s grow bet ter and have steady biological feat ures i n vi t ro , and
( J/ n" |6 ~4 S! ]may be used as " seed cell s" for autologous bone marrow MSCs t MSCs f rom liver cirrhosis patient s grow bet ter and have steady biological feat ures i n vi t ro , and" J7 e" i0 o. s3 J8 `8 w
may be used as " seed cell s" for autologous bone marrow MSCs t ransplantation and artificial liver .4 X2 N. _/ j1 |. F( o8 `4 J
Key words : cirrho sis ; bone marrow mesenchymal stem cell s ; autologous bone marrow mesenchy24 y9 A, c* t* y; R% b* i, h
mal stem cell s ; t ransplantationransplantation and artificial liver .
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: C& t2 P  X" u% xKey words : cirrho sis ; bone marrow mesenchymal stem cell s ; autologous bone marrow mesenchy2
) d' r4 w# k; Z3 I/ ?+ l. ?mal stem cell s ; t ransplantation  Q; |9 G7 B( G5 O) ~
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附件是pdf全文。感谢大家支持，回复本贴就可以下载了
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