Degenerative grade affects the responses of human nucleus pulposus cells to link-N, CTGF, and TGFβ3.
METHODS
Cells isolated from moderately and severely degenerated human intervertebral disks (IVDs) cultured in an alginate scaffold.


OBJECTIVE
To compare the regenerative potential of moderately versus severely degenerated cells using 3 proanabolic stimulants.


BACKGROUND
Injection of soluble cell signaling factors has potential to slow the progression of IVD degeneration. Although degenerative grade is thought to be an important factor in targeting therapeutic interventions it remains unknown whether cells in severely degenerated IVDs have impaired metabolic functions compared to lesser degenerative levels or if they are primarily influenced by the altered microenvironment.


METHODS
Nucleus pulposus (NP) cells were cultured in alginate for 21 days and treated with 3 different proanabolic stimulants: a growth factor/anti-inflammatory combination of transforming growth factor β3 (TGFβ3)+dexamethasone (Dex), or matricellular proteins connective tissue growth factor (CTGF) or Link-N. They were assayed for metabolic activity, DNA content, glycosaminoglycan, and qRT-PCR gene profiling.


RESULTS
Moderately degenerated cells responded to stimulation with increased proliferation, decreased IL-1β, MMP9, and COL1A1 expression, and upregulated HAS1 as compared with severely degenerated cells. TGFβR1 (ALK5) receptors were expressed at greater levels in moderately than severely degenerated cells. TGFβ3+Dex had a notable stimulatory effect on moderately degenerated NP cells with increased anabolic gene expression and decreased COL1A1 and ADAMTS5 gene expression. Link-N and CTGF had similar responses in all assays, and both treatments upregulated IL-1β expression and had a more catabolic response than TGFβ3+Dex, particularly in the more severely degenerated group. All groups, including different degenerative grades, produced similar amounts of glycosaminoglycan.


CONCLUSIONS
Proanabolic stimulants alone had limited capacity to overcome the catabolic and proinflammatory cytokine expression of severely degenerated NP cells and likely require additional anti-inflammatory treatments. Moderately degenerated NP cells had greater TGFβ receptor 1 expression and better responded to anabolic stimulation.
School of Engineering, University of Vermont, Burlington, VT, USA.
Rosalyn D
RD
Abbott
Devina
D
Purmessur
Robert D
RD
Monsey
David R
DR
Brigstock
Damien M
DM
Laudier
James C
JC
Iatridis
D000328:Adult; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism; D000090602:Collagen Type I, alpha 1 Chain; D055513:Connective Tissue Growth Factor / Q000494:pharmacology; D003907:Dexamethasone / Q000494:pharmacology; D005260:Female; D006801:Humans; D053583:Interleukin-1beta / Q000235:genetics / Q000378:metabolism; D007403:Intervertebral Disc / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D055959:Intervertebral Disc Degeneration / Q000378:metabolism / Q000473:pathology*; D008297:Male; D020780:Matrix Metalloproteinase 9 / Q000235:genetics / Q000378:metabolism; D008875:Middle Aged; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D000077293:Receptor, Transforming Growth Factor-beta Type I; D018125:Receptors, Transforming Growth Factor beta / Q000235:genetics / Q000378:metabolism; D012720:Severity of Illness Index; D053782:Transforming Growth Factor beta3 / Q000494:pharmacology
C526317:CCN2 protein, human; D024042:Collagen Type I; D000090602:Collagen Type I, alpha 1 Chain; D053583:Interleukin-1beta; D018125:Receptors, Transforming Growth Factor beta; D053782:Transforming Growth Factor beta3; D055513:Connective Tissue Growth Factor; D003907:Dexamethasone; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I; C000625841:TGFBR1 protein, human; D020780:Matrix Metalloproteinase 9