Strontium-Substituted Bioceramics Particles: A New Way to Modulate MCP-1 and Gro-α Production by Human Primary Osteoblastic Cells

Abstract

Background: To avoid morbidity and limited availability associated with autografts, synthetic calcium phosphate (CaP) ceramics were extensively developed and used as bone filling materials. Controlling their induced-inflammatory response nevertheless remained a major concern. Strontium-containing CaP ceramics were recently demonstrated for impacting cytokines' secretion pattern of human primary monocytes. The present study focuses on the ability of strontium-containing CaP to control the human primary bone cell production of two major inflammatory and pro-osteoclastogenic mediators, namely MCP-1 and Gro-α, in response to ceramics particles.

Methods: This in vitro study was performed using human primary osteoblasts in which their response to ceramics was evaluated by PCR arrays, antibody arrays were used for screening and real-time PCR and ELISA for more focused analyses.

Results: Study of mRNA and protein expression highlights that human primary bone cells are able to produce these inflammatory mediators and reveal that the adjunction of CaP in the culture medium leads to their enhanced production. Importantly, the current work determines the down-regulating effect of strontium-substituted CaP on MCP-1 and Gro-α production.

Conclusion: Our findings point out a new capability of strontium to modulate human primary bone cells' communication with the immune system.

Keywords: calcium-phosphate; cytokines; human primary bone cells; strontium.

Figure 1

Gro-α and MCP-1 are produced by human primary bone cells in basal culture conditions. (A) Human primary bone cells’ cytokine expressions were measured using Real-Time Polymerase Chain Reaction (RT-PCR) after 7, 14, and 21 days of culture. The HPRT1 gene was used as the housekeeping gene (HKG); (B) Cytokine concentrations were measured in human primary bone cell culture supernatants after 7, 14, and 21 days. Both experiments were performed on 8 independent donors in duplicate. Red bars represent median values. Black points represent maximum and minimum values. Black bars represent first and ninth deciles and the limits of rectangles represent first and third quartiles.

Figure 2

Strontium effect on MCP-1 production by human primary bone cells. (A) CCL2 and mRNA expression was evaluated by RT-PCR analysis performed (as described in the Materials and Methods section) after 7, 14 and 21 days of stimulation by BCP, BCP_5%, and SrCl₂. Experiments were performed on 5 independent donors in duplicate. Data are shown as specific variation of target genes mRNA using the 2^−ΔΔCt method (HPRT1 was used as internal control); (B) MCP-1 concentration was measured in cell supernatants after 7, 14, and 21 days of stimulation by BCP, BCP_5%, and SrCl₂. * means p < 0.05 when compared with control and ^# p < 0.05 when compared with BCP. Red bars represent median values. Black points represent maximum and minimum values. Black bars represent first and ninth deciles and the limits of white rectangles represents first and third quartiles.

Figure 3

Strontium effect on Gro-α production by human primary bone cells. (A) CXCL1 and mRNA production were evaluated by RT-PCR analysis performed (as described in the Materials and Methods section) after 7, 14, and 21 days of stimulation by BCP, BCP_5%, and SrCl₂. Experiments were performed on 5 independent donors in duplicate. Data are shown as specific variation of target genes mRNA using the 2^−ΔΔCt method (HPRT1 was used as internal control); (B) Gro-α concentration was measured in cell supernatants after 7, 14, and 21 days of stimulation by BCP, BCP_5%, and SrCl₂. * means p < 0.05 when compared with control and ^# p < 0.05 when compared with BCP. Red bars represent median values. Black points represent maximum and minimum values. Black bars represent first and ninth deciles and the limits of white rectangle represents first and third quartiles.

Figure 4

Relative cytokine production in human primary bone cell supernatants in basal culture conditions, measured by antibody arrays. The minimal (bottom box), median (inside box line), and maximal (top box) production are shown from 3 independent donors.