Intervertebral disc cells exhibit differences in gene expression in alginate and monolayer culture.
METHODS
The mRNA levels of aggrecan and collagen were quantified in intervertebral disc cells cultured under three conditions: primary alginate culture, monolayer culture, and re-encapsulation in alginate after monolayer culture.


OBJECTIVE
To compare the phenotype of intervertebral disc cells under different culture conditions and to investigate the reversibility of cell phenotype after re-encapsulation in alginate after monolayer culture.


BACKGROUND
The intervertebral disc contains heterogeneous populations of cells that vary with anatomic region. These cells possess significant differences in phenotype that can be preserved in vitro, although the effect of culture conditions on the phenotype of these cells is poorly understood.


METHODS
The intervertebral disc cells of 4-5-month-old pigs were isolated enzymatically from three anatomic zones: anulus fibrosus (AF), transition zone (TZ), and nucleus pulposus (NP). Gene expression levels of aggrecan and collagen Types I and II were measured using a quantitative reverse transcriptase--polymerase chain reaction.


RESULTS
Gene expression levels of anulus fibrosus and transition zone cells were shifted in monolayer compared with alginate, although the shift was partially reversed when re-encapsulated in alginate. However, NP cells appeared to be insensitive to culture conditions. Furthermore, characteristic patterns of gene expression among AF, TZ, and NP cells in primary alginate culture did not exist in monolayer culture, but they were also observed after re-encapsulation in alginate.


CONCLUSIONS
The findings of this study suggest that anulus fibrosus and transition zone cells undergo a reversible shift in phenotype when cultured in monolayer compared with alginate. These differences suggest that the culture system exerts a strong influence on cell phenotype and may play a role in the response of these cells to biophysical and biochemical stimuli in vitro.
Wang
J Y
JY
Department of Biomedical Engineering, Duke University, Durham, NC 27708, USA.
Baer
A E
AE
Kraus
V B
VB
Setton
L A
LA
D053674:Aggrecans; D000464:Alginates / Q000494:pharmacology*; D000818:Animals; D002478:Cells, Cultured; D003094:Collagen / Q000096:biosynthesis / Q000235:genetics*; D003470:Culture Media; D046508:Culture Techniques / Q000379:methods*; D017931:DNA Primers / Q000737:chemistry; D016326:Extracellular Matrix Proteins; D015870:Gene Expression / Q000187:drug effects*; D020723:Glucuronic Acid; D006603:Hexuronic Acids; D007403:Intervertebral Disc / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D037181:Lectins, C-Type; D008159:Lumbar Vertebrae; D010641:Phenotype; D011509:Proteoglycans / Q000096:biosynthesis / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013552:Swine
D053674:Aggrecans; D000464:Alginates; D003470:Culture Media; D017931:DNA Primers; D016326:Extracellular Matrix Proteins; D006603:Hexuronic Acids; D037181:Lectins, C-Type; D011509:Proteoglycans; D012333:RNA, Messenger; D020723:Glucuronic Acid; D003094:Collagen