Authors: Jun Chen Wei Yan Lori A Setton
Publish Date: 2006/03/18
Volume: 15, Issue: 3, Pages: 303-311
Abstract
The immature nucleus pulposus NP is populated by cells of notochordalorigin that are larger and contain an extensive cytoskeletal network and numerous vacuoles The disappearance of these cells with age is believed important in regulating metabolic shifts that may contribute to agerelated disc degeneration The precise biological function of these notochordal cells in the immature NP remains unclear however because of challenges in studying the mixed cell population in the NP In this study notochordallike cells were purified from immature NP cells using a new fluorescenceactivated cell sorting FACS protocol with autofluorescence and size analysis The unique molecular phenotypes of sorted notochordallike cells were characterized by the mRNA expression pattern for key matrix proteins and modulators and by the expression of cell–matrix receptor integrin subunits An FACS analysis showed that the immature NP contained a majority of cells that were larger than anulus fibrosus AF cells and with fluorescence higher than AF cells In comparison with the small NP cells separated by the FACS protocol sorted notochordallike cells expressed lower mRNA levels of type I collagen biglycan TIMP1 HSP70 and cfos and did not express detectable mRNA levels of decorin lumican multiple MMPs or IL1β via realtime quantitative RTPCR A greater number of these notochordallike cells also expressed the higher levels of α6 α1 and β1 integrin subunits as compared to small NP cells Together our results point towards a unique molecular phenotype for these notochordallike cells of NP characterized by the absence of gene expression for specific small proteoglycans and higher protein expression of integrin subunits that regulate interactions with collagens and laminin Future studies will be important for revealing if this unique molecular profile is coordinated with functional differences in pericellular matrix regions and/or integrinmediated cell–matrix interactions for these notochordallike cells within the NPWe gratefully acknowledge Dr Mike Cook for assistance with FACS Dr Huaxin Sheng and Steve Johnson for assistance with tissue harvesting and Maureen Upton Chris Gilchrist Dana Nettles and Larry Boyd for many helpful discussions This study was supported by funds from the NIH AR47442
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