Proliferation assessment of primary human mesenchymal stem cells on collagen membranes for guided bone regeneration.
OBJECTIVE
Human mesenchymal stem cells (hMSCs) hold the potential for bone regeneration because of their self-renewing and multipotent character. The goal of this study was to evaluate the influence of collagen membranes on the proliferation of hMSCs derived from bone marrow. A special focus was set on short-term eluates derived from collagen membranes, as volatile toxic materials washed out from these membranes may influence cell behavior during the short time course of oral surgery.


METHODS
The proliferation of hMSCs seeded directly on a collagen membrane (BioGide) was evaluated quantitatively using the cell proliferation reagent WST-1 (4-3-[4-iodophenyl]-2-[4-nitrophenyl]-2H-[5-tetrazolio]-1, 3--benzol-disulfonate) and qualitatively by scanning electron microscopy. Two standard biocompatibility tests, namely the lactate dehydrogenase and MTT (3-[4, 5-dimethyl-2-thiazolyl]-2, 5-diphenyl-2H-tetrazoliumbromide) tests, were performed using hMSCs cultivated in eluates from membranes incubated for 10 minutes, 1 hour, or 24 hours in serum-free cell culture medium. The data were analyzed statistically.


RESULTS
Scanning electron microscopy showed large numbers of hMSCs with well-spread morphology on the collagen membranes after 7 days of culture. The WST test revealed significantly better proliferation of hMSCs on collagen membranes after 4 days of culture compared to cells cultured on a cover glass. Cytotoxicity levels were low, peaking in short-term eluates and decreasing with longer incubation times.


CONCLUSIONS
Porcine collagen membranes showed good biocompatibility in vitro for hMSCs. If maximum cell proliferation rates are required, a prewash of membranes prior to application may be useful.
Faculty of Health Sciences and Medicine, Bond University, Gold Coast, Queensland, Australia.
Qin
Q
Liu
Andreas
A
Humpe
Dimitris
D
Kletsas
Frauke
F
Warnke
Stephan T
ST
Becker
Timothy
T
Douglas
Sureshan
S
Sivananthan
Patrick H
PH
Warnke
D000368:Aged; D001672:Biocompatible Materials / Q000737:chemistry; D001854:Bone Marrow Cells / Q000502:physiology; D001861:Bone Regeneration / Q000502:physiology*; D018929:Cell Culture Techniques; D049109:Cell Proliferation; D048430:Cell Shape; D003094:Collagen / Q000737:chemistry; D024042:Collagen Type I / Q000737:chemistry; D024061:Collagen Type III / Q000737:chemistry; D004396:Coloring Agents; D005898:Glass / Q000737:chemistry; D048091:Guided Tissue Regeneration / Q000379:methods*; D006801:Humans; D007202:Indicators and Reagents; D007770:L-Lactate Dehydrogenase / Q000032:analysis; D008297:Male; D008567:Membranes, Artificial; D059630:Mesenchymal Stem Cells / Q000502:physiology*; D008855:Microscopy, Electron, Scanning; D013499:Surface Properties; D013778:Tetrazolium Salts; D013844:Thiazoles; D013997:Time Factors
C110656:2-(4-iodophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium; C106416:Bio-Gide; D001672:Biocompatible Materials; D024042:Collagen Type I; D024061:Collagen Type III; D004396:Coloring Agents; D007202:Indicators and Reagents; D008567:Membranes, Artificial; D013778:Tetrazolium Salts; D013844:Thiazoles; D003094:Collagen; D007770:L-Lactate Dehydrogenase; C022616:thiazolyl blue