Histochemistry and Cell Biology

, Volume 148, Issue 6, pp 625–638 | Cite as

Co-culture of chondrons and mesenchymal stromal cells reduces the loss of collagen VI and improves extracellular matrix production

  • H. A. Owida
  • T. De Las Heras Ruiz
  • A. Dhillon
  • Y. Yang
  • N. J. Kuiper
Original Paper
  • 215 Downloads

Abstract

Adult articular chondrocytes are surrounded by a pericellular matrix (PCM) to form a chondron. The PCM is rich in hyaluronan, proteoglycans, and collagen II, and it is the exclusive location of collagen VI in articular cartilage. Collagen VI anchors the chondrocyte to the PCM. It has been suggested that co-culture of chondrons with mesenchymal stromal cells (MSCs) might enhance extracellular matrix (ECM) production. This co-culture study investigates whether MSCs help to preserve the PCM and increase ECM production. Primary bovine chondrons or chondrocytes or rat MSCs were cultured alone to establish a baseline level for ECM production. A xenogeneic co-culture monolayer model using rat MSCs (20, 50, and 80%) was established. PCM maintenance and ECM production were assessed by biochemical assays, immunofluorescence, and histological staining. Co-culture of MSCs with chondrons enhanced ECM matrix production, as compared to chondrocyte or chondron only cultures. The ratio 50:50 co-culture of MSCs and chondrons resulted in the highest increase in GAG production (18.5 ± 0.54 pg/cell at day 1 and 11 ± 0.38 pg/cell at day 7 in 50:50 co-culture versus 16.8 ± 0.61 pg/cell at day 1 and 10 ± 0.45 pg/cell at day 7 in chondron monoculture). The co-culture of MSCs with chondrons appeared to decelerate the loss of the PCM as determined by collagen VI expression, whilst the expression of high-temperature requirement serine protease A1 (HtrA1) demonstrated an inverse relationship to that of the collagen VI. Together, this implies that MSCs directly or indirectly inhibited HtrA1 activity and the co-culture of MSCs with chondrons enhanced ECM synthesis and the preservation of the PCM.

Keywords

Chondron Chondrocyte Mesenchymal stromal cells (MSCs) Co-culture Collagen VI HtrA1 

Notes

Acknowledgements

The authors thank Miss. Katy Cressy and Mr. John Misra in GHRC, Keele University for their technical support.

Compliance with ethical standards

Conflict of interest

The authors declare that there is no conflict of interest.

Supplementary material

418_2017_1602_MOESM1_ESM.tif (166 kb)
Figure SI: Immunofluorescence staining images of positive and negative controls. Positive control: freshly dissected bovine cartilage; negative control: staining without primary antibodies. (A) Collagen VI (green); (B) Collagen II (red); (C) HtrA1 (red). The cells were counter-stained by DAPI (blue). Scale bars represent 20 μm (TIFF 165 kb)
418_2017_1602_MOESM2_ESM.tif (245 kb)
Figure SII: Illustration of freshly dissected bovine cartilage stained by alcian blue. Scale bars represent 150 μm (TIFF 245 kb)
418_2017_1602_MOESM3_ESM.tif (59 kb)
Figure SIII sGAG production in media and in cells for monoculture [chondron (CN) and chondrocyte (CY)] and co-culture with 20, 50, 80% of MSC ratio at days 1, 3, 5, and 7 at P0 cells (TIFF 59 kb)

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Copyright information

© Springer-Verlag GmbH Germany 2017

Authors and Affiliations

  • H. A. Owida
    • 1
  • T. De Las Heras Ruiz
    • 1
  • A. Dhillon
    • 1
  • Y. Yang
    • 1
  • N. J. Kuiper
    • 1
  1. 1.Institute of Science and Technology in MedicineUniversity of KeeleStoke-on-TrentUK

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