An in vitro organ culturing system for intervertebral disc explants with vertebral endplates: a feasibility study with ovine caudal discs.
METHODS
Whole ovine caudal intervertebral discs with vertebral endplates were cultured under uniaxial diurnal loading for 7 days.


OBJECTIVE
To establish and characterize an organ culture system for intervertebral discs, in which disc cells may be "maintained" in their native three-dimensional environment under load.


BACKGROUND
In vitro culturing of entire discs with preserved biologic and structural integrity would be a useful model to study the effects of nutrition and mechanical loading.


METHODS
To maintain endplate permeability, sheep were systemically anticoagulated before death and their caudal vasculature was evacuated with saline postmortem. The first 4 caudal discs were explanted with their adjacent endplates and cultured in bioreactors under uniaxial diurnal loading (0.2 MPa for 8 hours and 0.8 MPa for 16 hours) for 4 or 7 days. Solute transport into the center of the disc was measured after 4 days of culture using a low molecular weight fluorescent marker. Cell viability, glycosaminoglycan synthesis rate, and gene expression profile were measured after 7 days of culture and compared with fresh tissue.


RESULTS
Fluorescent images showed that solutes could diffuse into the disc under both static and diurnal loading, but penetration through the endplate increased with diurnal loading. Cell viability and glycosaminoglycan synthesis rates remained unchanged after 7 days of culture. Expression of catabolic genes was significantly up-regulated, whereas anabolic genes tended to be down-regulated after 7 days.


CONCLUSIONS
With this novel preparation and culturing technique, endplate permeability could be maintained, which allowed culturing of intact disc explants with endplates for up to 7 days.
Gantenbein
Benjamin
B
AO Research Institute, Davos, Switzerland.
Grünhagen
Thijs
T
Lee
Cynthia R
CR
van Donkelaar
Corrinus C
CC
Alini
Mauro
M
Ito
Keita
K
D000818:Animals; D001692:Biological Transport; D019149:Bioreactors; D002470:Cell Survival; D002940:Circadian Rhythm; D015536:Down-Regulation; D005240:Feasibility Studies; D005456:Fluorescent Dyes / Q000493:pharmacokinetics; D015870:Gene Expression; D006025:Glycosaminoglycans / Q000096:biosynthesis; D007403:Intervertebral Disc / Q000166:cytology / Q000378:metabolism / Q000502:physiology; D009924:Organ Culture Techniques; D010539:Permeability; D012756:Sheep; D013131:Spine / Q000378:metabolism / Q000502:physiology; D013997:Time Factors; D015854:Up-Regulation; D016474:Weight-Bearing
D005456:Fluorescent Dyes; D006025:Glycosaminoglycans