Preparation and in vitro studies of microencapsulated cells releasing human tissue inhibitor of metalloproteinase-2.
OBJECTIVE
To prepare microencapsulated cells releasing human tissue inhibitor of metalloproteinase-2 (TIMP-2), and investigate their biological characteristics in vitro.


METHODS
Chinese hamster ovary (CHO) cells were stably transfected with a human TIMP-2 expression vector, encapsulated in barium alginate microcapsules and cultured in vitro. Morphological appearance of the microcapsules was observed under a light microscope. Cell viability was assessed using MTT (3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyltetrazolium bromide) assay. Enzyme linked immunosorbent assay (ELISA) and reverse zymography were used to confirm the release of biologically active TIMP-2 from the microcapsules. Cryopreservation study of the microencapsulated cells was carried out using dimethyl sulfoxide (DMSO) as preservative agent.


RESULTS
The microcapsules appeared like a sphere with diameter of 300 - approximately 600 microm. The surface of the capsule wall was clearly smooth. The microencapsulated cells survived well and kept proliferating over the 6 weeks observed. No significant difference in TIMP-2 secretion was found between encapsulated and unencapsulated cells. Reverse zymography confirmed the bioactivity of MMP (matrix metalloproteinase) inhibition of TIMP-2. The cryopreservation process did not damage the microcapsule morphology nor the viability of the cells inside.


CONCLUSIONS
Microencapsulated engineered CHO cells survive at least 6 weeks after preparation in vitro, and secrete bioactive TIMP-2 freely from the microcapsules.
Jiang
Qiang
Q
Cancer Institute, Second Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou 310009, China. jiangtiger@zju.edu.cn
Zhang
Su-Zhan
SZ
Peng
Jia-Ping
JP
Wang
Xu-Lin
XL
D000818:Animals; D016466:CHO Cells; D018914:Cells, Immobilized; D006224:Cricetinae; D015925:Cryopreservation; D006801:Humans; D008863:Microspheres; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D023822:Tissue Engineering; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000096:biosynthesis* / Q000235:genetics; D014162:Transfection
D011994:Recombinant Proteins; D019716:Tissue Inhibitor of Metalloproteinase-2