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Effects of Interleukin-17A on Osteogenic Differentiation of Isolated Human Mesenchymal Stem Cells.

Osta B, Lavocat F, Eljaafari A, Miossec P - Front Immunol (2014)

Bottom Line: These levels decreased in combination with IL-17A at 6 h only.However, IL-17 decreased the TNF-α-induced BMP2 inhibition.Such increase of Schnurri-3 may in turn activate osteoclasts leading to bone destruction as in RA.

View Article: PubMed Central - PubMed

Affiliation: Immunogenomics and Inflammation Research Unit EA 4130, Department of Clinical Immunology and Rheumatology, Edouard Herriot Hospital, University of Lyon 1 , Lyon , France.

ABSTRACT

Objectives: Rheumatoid arthritis (RA) is characterized by defective bone repair and excessive destruction and ankylosing spondylitis (AS) by increased ectopic bone formation with syndesmophytes. Since TNF-α and IL-17A are involved in both diseases, this study investigated their effects on the osteogenic differentiation of isolated human bone marrow-derived mesenchymal stem cells (hMSCs).

Methods: Differentiation of hMSCs into osteoblasts was induced in the presence or absence of IL-17A and/or TNF-α. Matrix mineralization (MM) was evaluated by alizarin red staining and alkaline phosphatase (ALP) activity. mRNA expression was measured by qRT-PCR for bone morphogenetic protein (BMP)-2 and Runx2, genes associated with osteogenesis, DKK-1, a negative regulator of osteogenesis, Schnurri-3 and receptor activator of nuclear factor kappa B ligand (RANKL), associated with the cross talk with osteoclasts, and TNF-α receptor type I and TNF-α receptor type II (TNFRII).

Results: TNF-α alone increased both MM and ALP activity. IL-17A alone increased ALP but not MM. Their combination was more potent. TNF-α alone increased BMP2 mRNA expression at 6 and 12 h. These levels decreased in combination with IL-17A at 6 h only. DKK-1 mRNA expression was inhibited by TNF-α and IL-17A either alone or combined. Supporting an imbalance toward osteoblastogenesis, RANKL expression was inhibited by TNF-α and IL-17A. However, TNF-α but not IL-17 alone decreased Runx2 mRNA expression at 6 h. In parallel, TNF-α but not IL-17 alone increased Schnurri-3 expression with a synergistic effect with their combination. This may be related to an increase of TNFRII overexpression.

Conclusion: IL-17 increased the effects of TNF-α on bone matrix formation by hMSCs. However, IL-17 decreased the TNF-α-induced BMP2 inhibition. Synergistic interactions between TNF-α and IL-17 were seen for RANKL inhibition and Schnurri-3 induction. Such increase of Schnurri-3 may in turn activate osteoclasts leading to bone destruction as in RA. Conversely, in the absence of osteoclasts, this could promote ectopic bone formation as in AS.

No MeSH data available.


Related in: MedlinePlus

Effects of IL-17A and/or TNF-α on alkaline phosphatase activity are shown. hMSCs were plated at a density of 5 × 103 cell/cm2, cells were cultured in osteogenic medium for 14 days, in the absence (0) or presence of TNF-α 1 ng/ml and/or IL-17A 50 ng/ml. ALP activity was measured by fluorometry. Results were analyzed using the Wilcoxon test. *p  < 0.05; **p  < 0.005 vs. induction medium alone.
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Figure 2: Effects of IL-17A and/or TNF-α on alkaline phosphatase activity are shown. hMSCs were plated at a density of 5 × 103 cell/cm2, cells were cultured in osteogenic medium for 14 days, in the absence (0) or presence of TNF-α 1 ng/ml and/or IL-17A 50 ng/ml. ALP activity was measured by fluorometry. Results were analyzed using the Wilcoxon test. *p  < 0.05; **p  < 0.005 vs. induction medium alone.

Mentions: Since TNF-α and IL-17A increased mineralization of the extracellular matrix, we next investigated their effects on ALP, an enzyme, which is essential for bone mineralization. ALP activity was measured at days 3, 5, 7, and 14 (Figure 2). At day 3, no difference in the presence or absence of cytokine was detected. Increased levels of ALP activity were detected at day 5 when the two cytokines were combined (4.4 × 103 U/ml at day 3 vs. 7.4 × 103 U/ml at day 5, *p  < 0.05) or at day 7 in the presence of each cytokine alone (Figure 2). Moreover, the combination of the two cytokines resulted in a further increase of ALP activity, the maximum level being achieved at day 7 (1.8 × 103 U/ml without cytokine vs. 1.3 × 104 U/ml with both cytokines, **p  < 0.005). Day 14 shows almost the same results as day 7.


Effects of Interleukin-17A on Osteogenic Differentiation of Isolated Human Mesenchymal Stem Cells.

Osta B, Lavocat F, Eljaafari A, Miossec P - Front Immunol (2014)

Effects of IL-17A and/or TNF-α on alkaline phosphatase activity are shown. hMSCs were plated at a density of 5 × 103 cell/cm2, cells were cultured in osteogenic medium for 14 days, in the absence (0) or presence of TNF-α 1 ng/ml and/or IL-17A 50 ng/ml. ALP activity was measured by fluorometry. Results were analyzed using the Wilcoxon test. *p  < 0.05; **p  < 0.005 vs. induction medium alone.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4151036&req=5

Figure 2: Effects of IL-17A and/or TNF-α on alkaline phosphatase activity are shown. hMSCs were plated at a density of 5 × 103 cell/cm2, cells were cultured in osteogenic medium for 14 days, in the absence (0) or presence of TNF-α 1 ng/ml and/or IL-17A 50 ng/ml. ALP activity was measured by fluorometry. Results were analyzed using the Wilcoxon test. *p  < 0.05; **p  < 0.005 vs. induction medium alone.
Mentions: Since TNF-α and IL-17A increased mineralization of the extracellular matrix, we next investigated their effects on ALP, an enzyme, which is essential for bone mineralization. ALP activity was measured at days 3, 5, 7, and 14 (Figure 2). At day 3, no difference in the presence or absence of cytokine was detected. Increased levels of ALP activity were detected at day 5 when the two cytokines were combined (4.4 × 103 U/ml at day 3 vs. 7.4 × 103 U/ml at day 5, *p  < 0.05) or at day 7 in the presence of each cytokine alone (Figure 2). Moreover, the combination of the two cytokines resulted in a further increase of ALP activity, the maximum level being achieved at day 7 (1.8 × 103 U/ml without cytokine vs. 1.3 × 104 U/ml with both cytokines, **p  < 0.005). Day 14 shows almost the same results as day 7.

Bottom Line: These levels decreased in combination with IL-17A at 6 h only.However, IL-17 decreased the TNF-α-induced BMP2 inhibition.Such increase of Schnurri-3 may in turn activate osteoclasts leading to bone destruction as in RA.

View Article: PubMed Central - PubMed

Affiliation: Immunogenomics and Inflammation Research Unit EA 4130, Department of Clinical Immunology and Rheumatology, Edouard Herriot Hospital, University of Lyon 1 , Lyon , France.

ABSTRACT

Objectives: Rheumatoid arthritis (RA) is characterized by defective bone repair and excessive destruction and ankylosing spondylitis (AS) by increased ectopic bone formation with syndesmophytes. Since TNF-α and IL-17A are involved in both diseases, this study investigated their effects on the osteogenic differentiation of isolated human bone marrow-derived mesenchymal stem cells (hMSCs).

Methods: Differentiation of hMSCs into osteoblasts was induced in the presence or absence of IL-17A and/or TNF-α. Matrix mineralization (MM) was evaluated by alizarin red staining and alkaline phosphatase (ALP) activity. mRNA expression was measured by qRT-PCR for bone morphogenetic protein (BMP)-2 and Runx2, genes associated with osteogenesis, DKK-1, a negative regulator of osteogenesis, Schnurri-3 and receptor activator of nuclear factor kappa B ligand (RANKL), associated with the cross talk with osteoclasts, and TNF-α receptor type I and TNF-α receptor type II (TNFRII).

Results: TNF-α alone increased both MM and ALP activity. IL-17A alone increased ALP but not MM. Their combination was more potent. TNF-α alone increased BMP2 mRNA expression at 6 and 12 h. These levels decreased in combination with IL-17A at 6 h only. DKK-1 mRNA expression was inhibited by TNF-α and IL-17A either alone or combined. Supporting an imbalance toward osteoblastogenesis, RANKL expression was inhibited by TNF-α and IL-17A. However, TNF-α but not IL-17 alone decreased Runx2 mRNA expression at 6 h. In parallel, TNF-α but not IL-17 alone increased Schnurri-3 expression with a synergistic effect with their combination. This may be related to an increase of TNFRII overexpression.

Conclusion: IL-17 increased the effects of TNF-α on bone matrix formation by hMSCs. However, IL-17 decreased the TNF-α-induced BMP2 inhibition. Synergistic interactions between TNF-α and IL-17 were seen for RANKL inhibition and Schnurri-3 induction. Such increase of Schnurri-3 may in turn activate osteoclasts leading to bone destruction as in RA. Conversely, in the absence of osteoclasts, this could promote ectopic bone formation as in AS.

No MeSH data available.


Related in: MedlinePlus