The in vitro and in vivo proliferative capacity of serum-free cultivated human conjunctival epithelial cells
The in vitro and in vivo proliferative capacity of serum-free cultivated human conjunctival epithelial cells
复制标题
无血清培养人结膜上皮细胞的体外和体内增殖能力
作者：
L. Ang;D. Tan;T. Phan;Jing Li;R. Beuerman;R. Lavker
目的。目的：研究无血清培养的人结膜上皮细胞的体外和体内增殖能力，并与目前使用含血清培养液和3T3饲养层的方法进行比较。方法：研究方法。人结膜上皮细胞分别在无血清培养液、3T3饲养层无血清培养液和3T3饲养层含血清培养液中培养。比较了它们的生长面积、集落形成效率和种群倍增数。通过分析培养细胞移植到裸鼠体内所产生的细胞数量来评估体内的增殖能力。培养细胞检测细胞角蛋白K3、K4、K12、K19以及凝胶杯状细胞粘蛋白MUC5AC的表达。结果。在无血清、无血清和饲养层细胞以及有血清和饲养层细胞中培养的细胞集落形成率分别为14.5±4.1%、10.1±3.1%和20.4±6.7%，群体倍增数分别为24.8±4.3、14.8±3.6和30.0±5.0。9日龄无血清培养的裸鼠结膜囊含1.29×106±0.46×106个细胞，含血清培养的囊壁含1.30×106±0.53×106个细胞。两种情况下的上皮复层程度相似。无血清培养的结膜细胞保持其体内特性，表达K4、K19和MUC5AC。RT-PCR证实这些细胞中存在MUC5AC基因。结论。无血清培养的结膜上皮细胞在体内表现出与含血清培养的3T3饲养层细胞相似的增殖能力。这具有重要的临床意义，因为用于临床移植的细胞的无血清体外扩增克服了使用动物血清和细胞的相关问题。
Purpose. To investigate the in vitro and in vivo proliferative capacity of human conjunctival epithelial cells cultured in serum-free media, and to compare this with current methods that utilize serum-containing media and 3T3 feeder layers. Methods. Human conjunctival epithelial cells were cultivated in serum-free media alone, serum-free media with a 3T3 feeder layer, and serum-containing media with a 3T3 feeder layer. The areas of outgrowth, colony-forming efficiencies and number of population doublings were compared. The in vivo proliferative potential was assessed by analyzing the number of cells generated by the implantation of cultured cells into athymic mice. Cultured cells were evaluated for the expression of cytokeratins K3, K4, K12, K19, as well as the gel-forming goblet cell mucin, MUC5AC. Results. Cells cultivated in serum-free media, serum-free media and feeder cells, and serum-containing media and feeder cells achieved colony-forming efficiencies of 14.5 ± 4.1%, 10.1 ± 3.1%, and 20.4 ± 6.7%, respectively, and number of population doublings of 24.8 ± 4.3, 14.8 ± 3.6, and 30.0 ± 5.0, respectively. Nine-day old athymic mice conjunctival cysts derived from serum-free cultures comprised 1.29 × 10 6 ± 0.46 × 10 6 cells, while cysts derived from serumcontaining cultures comprised 1.30 × 10 6 ± 0.53 × 10 6 cells. The degree of epithelial stratification was similar in both conditions. Serum-free cultivated conjunctival cells retained their in vivo characteristics and expressed K4, K19 and MUC5AC. The presence of MUC5AC mRNA in these cells was confirmed by RT-PCR. Conclusions. Conjunctival epithelial cells propagated in serum-free media demonstrated a similar in vivo proliferative capability, as compared to serum-containing media with 3T3 feeder cells. This has important clinical implications, as the serum-free ex vivo expansion of cells for clinical transplantation overcomes the problems associated with the use of animal serum and cells.
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影响因子：
4.4
作者：
P. Argüeso;M. Balaram;M. Balaram;S. Spurr-Michaud;H. Keutmann;R. Dana;R. Dana;I. Gipson
通讯作者：
P. Argüeso;M. Balaram;M. Balaram;S. Spurr-Michaud;H. Keutmann;R. Dana;R. Dana;I. Gipson
影响因子：
4.4
作者：
Z. Wei;R. Wu;R. Lavker;T. Sun
通讯作者：
Z. Wei;R. Wu;R. Lavker;T. Sun
影响因子：
8
作者：
Chen, N.;Eritja, N.;Lock, R.;Debnath, J.
通讯作者：
Debnath, J.
影响因子：
4.4
作者：
F. E. Kruse;Scheffer C. G. Tseng
通讯作者：
F. E. Kruse;Scheffer C. G. Tseng
在无血清培养基中培养的人角质形成细胞的体外生长和分化的控制。
影响因子：
--
作者：
Shipley,GD;Pittelkow,MR
通讯作者：
Pittelkow,MR