Environmental regulation of notochordal gene expression in nucleus pulposus cells.
Cells of the nucleus pulposus (NP) in the intervertebral disc are derived directly from the embryonic notochord. In humans, a shift in NP cell population coincides with the beginning of age-related changes in the extracellular matrix that can lead to spinal disorders. To begin identifying the bases of these changes, the manner by which relevant environmental factors impact cell function must be understood. This study investigated the roles of biochemical, nutritional, and physical factors in regulating immature NP cells. Specifically, we examined cell morphology, attachment, proliferation, and expression of genes associated with the notochord and immature NP (Sox9, CD24, and type IIA procollagen). Primary cells isolated from rat caudal discs were exposed to different media formulations and physical culture configurations either in 21% (ambient) or 2% (hypoxic) O2. As expected, cells in alginate beads retained a vacuolated morphology similar to chordocytes, with little change in gene expression. Interestingly, NP tissues not enzymatically digested were more profoundly influenced by oxygen. In monolayer, alpha-MEM preserved vacuolated morphology, produced the highest efficiency of attachment, and best maintained gene expression. DMEM and Opti-MEM cultures resulted in high levels of proliferation, but these appeared to involve small non-vacuolated cells. Gene expression patterns for cells in DMEM monolayer cultures were consistent with chondrocyte de-differentiation, with the response being delayed by hypoxia. Overall, results indicate that certain environmental conditions induce cellular changes that compromise the notochordal phenotype in immature NP. These results form the foundation on which the mechanisms of such changes can be elucidated.
Rastogi
Anshu
A
Department of Bioengineering, University of Maryland, College Park, Maryland 20742, USA.
Thakore
Pratiksha
P
Leung
Aileen
A
Benavides
Magda
M
Machado
Monica
M
Morschauser
Michael A
MA
Hsieh
Adam H
AH
D000464:Alginates / Q000378:metabolism; D000818:Animals; D051927:CD24 Antigen / Q000235:genetics*; D002448:Cell Adhesion / Q000235:genetics; D015687:Cell Hypoxia; D049109:Cell Proliferation; D048430:Cell Shape / Q000235:genetics; D002478:Cells, Cultured; D024043:Collagen Type II / Q000235:genetics*; D003470:Culture Media / Q000378:metabolism; D005786:Gene Expression Regulation; D020723:Glucuronic Acid / Q000378:metabolism; D006603:Hexuronic Acids / Q000378:metabolism; D007403:Intervertebral Disc / Q000166:cytology / Q000378:metabolism*; D008297:Male; D009672:Notochord / Q000166:cytology / Q000378:metabolism*; D010641:Phenotype; D051381:Rats; D017207:Rats, Sprague-Dawley; D055758:SOX9 Transcription Factor / Q000235:genetics*; D013997:Time Factors
D000464:Alginates; D051927:CD24 Antigen; D024043:Collagen Type II; D003470:Culture Media; D006603:Hexuronic Acids; D055758:SOX9 Transcription Factor; D020723:Glucuronic Acid