Three-dimensional Expansion:In Suspension Culture of SD Rat's Osteoblasts in a Rotating Wall Vessel Bioreactor  

作　　者：KE-DONG SONG TIAN-QINGLIU XIANG-QIN LI ZHAN-FENG CUI XIANG-YU SUN AND XUE-HU MA 

机构地区：[1]Stem Cell and Tissue Engineering Laboratory, State Key Laboratory of Fine Chemicals, Dalian University of Technology, Dalian 116024, Liaoning, China [2]Department of Engineering Science, Oxford University,Parks Road, Oxford OX1 3PJ, U. K.

出　　处：《Biomedical and Environmental Sciences》2007年第2期91-98,共8页生物医学与环境科学（英文版）

基　　金：This work was supported by grants from the Int. Cooperation Project for National & Abroad Lab. of the National Natural Sciences Foundation of China (No. 2002008);The Science & Technology Foundation of Liaoning Province (No. 20022140).

摘　　要：Objective To study large-scale expansion of SD （Sprague-Dawley） rat＇s osteoblasts in suspension culture in a rotating wall vessel bioreactor （RWVB）. Methods The bioreactor rotation speeds were adjusted in the range of 0 to 20 rpm, which could provide low shear on the rnicrocarriers around 1 dyn/cm^2. The cells were isolated via sequential digestions of neonatal （less than 3 days old） SD rat calvaria. After the primary culture and several passages, the cells were seeded onto the microcarriers and cultivated in T-flask, spinner flask and RWVB respectively. During the culture period, the cells were counted and observed under the inverted microscope for morphology every 12 h. After 7 days, the cells were evaluated with scanning electron microscope （SEM） for histological examination of the aggregates. Also, the hematoxylin-eosin （HE） staining and alkaline phosphatase （ALP） staining were performed. Moreover, von-Kossa staining and Alizarin Red S staining were carded out for mineralized nodule formation. Results The results showed that in RWVB, the cells could be expanded by more than ten times and they presented better morphology and vitality and stronger ability to form bones. Conclusions The developed RWVB can provide the culture environment with a relatively low shear force and necessary three-dimensional （3D） interactions among cells and is suitable for osteopath expansion in vitro.Objective To study large-scale expansion of SD （Sprague-Dawley） rat＇s osteoblasts in suspension culture in a rotating wall vessel bioreactor （RWVB）. Methods The bioreactor rotation speeds were adjusted in the range of 0 to 20 rpm, which could provide low shear on the rnicrocarriers around 1 dyn/cm^2. The cells were isolated via sequential digestions of neonatal （less than 3 days old） SD rat calvaria. After the primary culture and several passages, the cells were seeded onto the microcarriers and cultivated in T-flask, spinner flask and RWVB respectively. During the culture period, the cells were counted and observed under the inverted microscope for morphology every 12 h. After 7 days, the cells were evaluated with scanning electron microscope （SEM） for histological examination of the aggregates. Also, the hematoxylin-eosin （HE） staining and alkaline phosphatase （ALP） staining were performed. Moreover, von-Kossa staining and Alizarin Red S staining were carded out for mineralized nodule formation. Results The results showed that in RWVB, the cells could be expanded by more than ten times and they presented better morphology and vitality and stronger ability to form bones. Conclusions The developed RWVB can provide the culture environment with a relatively low shear force and necessary three-dimensional （3D） interactions among cells and is suitable for osteopath expansion in vitro.

关 键 词：OSTEOBLAST BIOREACTOR MICROCARRIER Tissue engineering Laminar flow 

分 类 号：R336[医药卫生—人体生理学] Q593.6[医药卫生—基础医学]