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6 `3 x7 N' n$ l1 ^ 【摘要】 目的:了解自体骨髓干细胞促进兔角膜苯酚烧伤后愈合的作用。9 p+ t* W( _8 S# T
8 P2 ?: Z# V# u& e Y方法:将10只家兔双眼角膜用300g/L的苯酚棉签烧伤后,随机一眼用自体骨髓干细胞滴眼,另一眼为对照眼,用无细胞的培养液滴眼,观察双侧角膜愈合时间,进行的统计学的配对分析t检验。结果:在自体骨髓干细胞滴眼的干预下,角膜烧伤的愈合时间平均为3.2d,对照眼的愈合时间平均为7.2d,根据统计学检验,两者具有显著性差异(P ) C& u* J0 D. A# x& S5 q: J1 c 【关键词】骨髓干细胞;角膜;化学伤;兔 Z/ W, D6 }* n) ]$ Z& B$ o* X1 } Research of autologous bone marrow stem cell treating the cornea burn with C6H5OH phenol9 c. Z: _+ r+ S9 V) C' @, f
9 z8 x+ I% l3 _& ]7 \2 W FangWei Ying, LiNa Huang, MingYing Lai, LiChun Wang, Jing He 0 d- B+ z; t% ` t& m. L ! ?) }; p' z4 v; ?/ { Foundation items: Medical Science Research Foundation of Guangdong Province, China (No. A2006591); Doctor Starting Project of Natural Science Foundation of Guangdong Province, China (No. 6300824)4 T$ E, S, u+ q6 F0 p
. k- y- Q# K ?8 u# m Department of Ophthalmology, Shenzhen Eye Hospital, Shenzhen 518001, Guangdong Province, China- p# K* c2 S/ B" _& U2 z. M! Z4 ^
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Correspondence to: LiNa Huang. Department of Ophthalmology, Shenzhen Eye Hospital, Shenzhen 518001, Guangdong Province, China. yingfangwei@yahoo.com.cn2 h! W* z0 c5 {2 S9 z
% z+ D/ u' G: h# O" R AbstractAIM: To understand the effect of autologous bone marrow stem cell in treating the cornea burn with C6H5OH phenol. 4 @4 T3 ^" F# P) A1 I7 [0 o
' H, e0 P/ G. f3 IMETHODS: After burned both of corneas of the 10 rabbits by 300g/L C6H5OH phenol, one of corneas was trickled of autologous bone marrow stem cells, the other trickled with acellular culture solution was as control group. The restoration time was calculated, and the results were statistically analyzed by paired ttest. RESULTS: With the autologous bone marrow stem cell treating, the restoration time of phenol burning eye was decreased significantly (mean 3.2 days), compared with 7.2 days in control group (P: c6 r+ ~9 M2 o) g+ f4 s. y# W6 b
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KEYWORDS: bone marrow stem cell; cornea; chemical injury; rabbit ) X+ A9 {" M: e, `) R0 r. v' O, N+ [6 l" x+ P
0引言 0 y- o5 C, D( ?* z8 z8 ~- I4 i' U0 G N/ O
随着工业化进程的不断发展,角膜化学性烧伤在临床上越来越常见,角膜化学性烧伤经常导致角膜上皮愈合不良,进而引起感染,甚至溃疡穿孔、眼内炎等并发症,这已经成为近年来角膜性失明的重要原因之一。本研究通过兔自体骨髓干细胞的干预,了解其对角膜的苯酚烧伤愈合的促进作用,希望能为化学性角膜烧伤的治疗开辟新的途径。 : B: J, H5 [! M: Z) l9 [ 5 m o1 ?# M1 C 1材料和方法 7 E3 `2 m0 k/ t8 \! U1 n! S& V3 u4 j$ d0 g" U. y3 n3 E7 l4 w
1.1材料 7 a2 `. h. ]2 Y1 i: n% J " V0 S) r$ |9 }7 m3 j# r 家兔10只(广东省动物中心),体质量2~2.5kg,苯酚,前段照相系统,10g/L荧光素钠(广州白云山明兴制药),淋巴细胞分离液(Ficoll)1.077g/mL,(天津),LDMEM培养基(ARP公司),胎牛血清(杭州四季青)等。 ; n! F8 }- j6 y S& U2 ?; {/ U4 v- a
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1.2.1制作苯酚烧伤角膜模型 " j9 M/ k% n! I% A4 E+ Z4 J# { o" h) Q- u, j' f# {, R( [
用30g/L戊巴比妥钠溶液静脉麻醉家兔,1mL/kg,用地卡因表面麻醉角膜,将浸透300g/L新鲜配制的苯酚溶液的棉签按压在兔双侧角膜中央区域5s,形成直径5mm的化学烧伤区。 * X4 K, L, F3 s0 L' q9 z 4 ^7 @5 n$ w, s j L 1.2.2收集骨髓干细胞[1] " G* {; D! S5 o$ F7 p+ r* z) u a) k% v2 ~9 O# Y! u( o
用16号骨髓穿刺针在家兔的胫骨上段内侧面抽取骨髓3mL,肝素抗凝,置于等体积的Ficoll分离液上,2000r/min离心20min后,吸取上方分离液,加入DHank’s液混合,1500r/min离心10min后,倒出上清液,再加入DHank’s液混合,1500r/min离心10min后,去除上清液后,用含150mL/L胎牛血清的LDMEM完全培养液重悬细胞,调整细胞的浓度为2106/mL,置于24孔板,在37℃,50mL/L CO2培养箱中培养。培养24h起每日收集悬浮的细胞,1500r/min离心10min后,去除上清液,加入完全培养液,调整细胞浓度至2106/mL,用于滴眼。 - Z/ }9 X) d# k( V! ~) x6 {5 G. P% }9 t6 q2 t: ^
1.2.3自体骨髓干细胞干预7 l& t: `) ~2 p8 W
# ~6 j5 f* p. I 随机选取10只家兔的一眼作为实验眼,另一眼则为对照眼。在烧伤后24h起,用新鲜收集的的骨髓干细胞悬液2滴滴于实验眼角膜表面,1次/d,对照眼则用无细胞的培养液滴眼,直至角膜愈合。裂隙灯下,结膜囊滴10g/L荧光素钠,每日观察双侧烧伤角膜的愈合情况,角膜荧光素染色情况,记录烧伤角膜愈合的时间(表1)。: y% K3 ?2 m# W3 l4 ]7 C