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Mechanical loading reduces inflammation-induced human osteocyte-to-osteoclast communication.

Pathak JL, Bravenboer N, Luyten FP, Verschueren P, Lems WF, Klein-Nulend J, Bakker AD - Calcif. Tissue Int. (2015)

Bottom Line: However, RA serum enhanced the RANKL/OPG expression ratio by 3.4-fold, while PFF ified this effect.Stat-CM from RA serum-pretreated osteocytes enhanced osteoclastogenesis compared with stat-CM from control serum-pretreated osteocytes, while PFF ified this effect.In conclusion, RA serum, containing inflammatory factors, did not alter the intrinsic capacity of osteocytes to sense mechanical stimuli, but upregulated osteocyte-to-osteoclast communication.

View Article: PubMed Central - PubMed

Affiliation: Department of Oral Cell Biology, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and VU University Amsterdam, MOVE Research Institute Amsterdam, Amsterdam, The Netherlands.

ABSTRACT
Multiple factors contribute to bone loss in inflammatory diseases such as rheumatoid arthritis (RA), but circulating inflammatory factors and immobilization play a crucial role. Mechanical loading prevents bone loss in the general population, but the effects of mechanical loading in patients with RA are less clear. Therefore, we aimed to investigate whether mechanical stimuli reverse the stimulatory effect of RA serum on osteocyte-to-osteoclast communication. Human primary osteocytes were pretreated with 10 % RA serum or healthy control serum for 7 days, followed by 1 h ± mechanical loading by pulsating fluid flow (PFF). Nitric oxide (NO) and prostaglandin E2 were measured in the medium. Receptor activator of nuclear factor-kappaB ligand (RANKL), osteoprotegerin (OPG), interleukin-6 (IL-6), cyclooxygenase-2 (COX2), matrix-extracellular phosphoglycoprotein (MEPE), cysteine-rich protein 61 (CYR61), and SOST gene expression was quantified by qPCR. Osteoclast precursors were cultured with PFF-conditioned medium (PFF-CM) or static-conditioned medium (stat-CM), and osteoclast formation was assessed. RA serum alone did not affect IL-6, CYR61, COX2, MEPE, or SOST gene expression in osteocytes. However, RA serum enhanced the RANKL/OPG expression ratio by 3.4-fold, while PFF ified this effect. PFF enhanced NO production to the same extent in control serum (2.6-3.5-fold) and RA serum-pretreated (2.7-3.6-fold) osteocytes. Stat-CM from RA serum-pretreated osteocytes enhanced osteoclastogenesis compared with stat-CM from control serum-pretreated osteocytes, while PFF ified this effect. In conclusion, RA serum, containing inflammatory factors, did not alter the intrinsic capacity of osteocytes to sense mechanical stimuli, but upregulated osteocyte-to-osteoclast communication. Mechanical loading ified this upregulation, suggesting that mechanical stimuli could contribute to the prevention of osteoporosis in inflammatory disease.

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Effect of Stat-CM from RA serum-pretreated osteocytes and PFF-CM from RA serum-pretreated osteocytes on osteoclast formation. CM was obtained from osteocytes cultured for 7 days with or without RA serum, followed by 1 h PFF or static control culture, and 1 h post-incubation without PFF. CM was added to osteoclast precursors. a Representative micrograph of TRACP-positive multinucleated cells (TRAP + MNC) in a culture of human PBMCs with stat-CM from control serum-pretreated osteocytes, PFF-CM from control serum-pretreated osteocytes, stat-CM from RA serum-pretreated osteocytes, and PFF-CM from RA serum-pretreated osteocytes. White arrows: TRACP + MNC with 3–5 nuclei; black arrows: TRACP + MNC with >5 nuclei. b PFF-CM from control serum-pretreated osteocytes inhibited the formation of osteoclasts with 3–5 nuclei. Stat-CM from RA serum-pretreated osteocytes enhanced the formation of osteoclasts with 3–5 nuclei, and PFF-CM from RA serum-pretreated osteocytes ified this effect. c PFF-CM from RA serum-pretreated osteocytes inhibited the formation of osteoclasts with >5 nuclei. Values are mean from three independent experiments, n = 9. Significant effect of Stat-CM from RA serum-pretreated osteocytes, *p < 0.05. Significant effect of PFF-CM, ##p < 0.01, ###p < 0.001
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Fig3: Effect of Stat-CM from RA serum-pretreated osteocytes and PFF-CM from RA serum-pretreated osteocytes on osteoclast formation. CM was obtained from osteocytes cultured for 7 days with or without RA serum, followed by 1 h PFF or static control culture, and 1 h post-incubation without PFF. CM was added to osteoclast precursors. a Representative micrograph of TRACP-positive multinucleated cells (TRAP + MNC) in a culture of human PBMCs with stat-CM from control serum-pretreated osteocytes, PFF-CM from control serum-pretreated osteocytes, stat-CM from RA serum-pretreated osteocytes, and PFF-CM from RA serum-pretreated osteocytes. White arrows: TRACP + MNC with 3–5 nuclei; black arrows: TRACP + MNC with >5 nuclei. b PFF-CM from control serum-pretreated osteocytes inhibited the formation of osteoclasts with 3–5 nuclei. Stat-CM from RA serum-pretreated osteocytes enhanced the formation of osteoclasts with 3–5 nuclei, and PFF-CM from RA serum-pretreated osteocytes ified this effect. c PFF-CM from RA serum-pretreated osteocytes inhibited the formation of osteoclasts with >5 nuclei. Values are mean from three independent experiments, n = 9. Significant effect of Stat-CM from RA serum-pretreated osteocytes, *p < 0.05. Significant effect of PFF-CM, ##p < 0.01, ###p < 0.001

Mentions: PFF-CM from control serum-pretreated osteocytes decreased the number of TRACP-positive osteoclasts with 3–5 nuclei by 1.6-fold (Fig. 3a, b). Stat-CM from RA serum-pretreated cell culture increased the number of TRACP-positive osteoclasts with 3–5 nuclei by 1.2-fold, while PFF ified this effect (Fig. 3a, b). PFF-CM from RA serum-pretreated osteocytes also inhibited formation of osteoclasts with >5 nuclei by 1.8-fold (Fig. 3a, c).Fig. 3


Mechanical loading reduces inflammation-induced human osteocyte-to-osteoclast communication.

Pathak JL, Bravenboer N, Luyten FP, Verschueren P, Lems WF, Klein-Nulend J, Bakker AD - Calcif. Tissue Int. (2015)

Effect of Stat-CM from RA serum-pretreated osteocytes and PFF-CM from RA serum-pretreated osteocytes on osteoclast formation. CM was obtained from osteocytes cultured for 7 days with or without RA serum, followed by 1 h PFF or static control culture, and 1 h post-incubation without PFF. CM was added to osteoclast precursors. a Representative micrograph of TRACP-positive multinucleated cells (TRAP + MNC) in a culture of human PBMCs with stat-CM from control serum-pretreated osteocytes, PFF-CM from control serum-pretreated osteocytes, stat-CM from RA serum-pretreated osteocytes, and PFF-CM from RA serum-pretreated osteocytes. White arrows: TRACP + MNC with 3–5 nuclei; black arrows: TRACP + MNC with >5 nuclei. b PFF-CM from control serum-pretreated osteocytes inhibited the formation of osteoclasts with 3–5 nuclei. Stat-CM from RA serum-pretreated osteocytes enhanced the formation of osteoclasts with 3–5 nuclei, and PFF-CM from RA serum-pretreated osteocytes ified this effect. c PFF-CM from RA serum-pretreated osteocytes inhibited the formation of osteoclasts with >5 nuclei. Values are mean from three independent experiments, n = 9. Significant effect of Stat-CM from RA serum-pretreated osteocytes, *p < 0.05. Significant effect of PFF-CM, ##p < 0.01, ###p < 0.001
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Fig3: Effect of Stat-CM from RA serum-pretreated osteocytes and PFF-CM from RA serum-pretreated osteocytes on osteoclast formation. CM was obtained from osteocytes cultured for 7 days with or without RA serum, followed by 1 h PFF or static control culture, and 1 h post-incubation without PFF. CM was added to osteoclast precursors. a Representative micrograph of TRACP-positive multinucleated cells (TRAP + MNC) in a culture of human PBMCs with stat-CM from control serum-pretreated osteocytes, PFF-CM from control serum-pretreated osteocytes, stat-CM from RA serum-pretreated osteocytes, and PFF-CM from RA serum-pretreated osteocytes. White arrows: TRACP + MNC with 3–5 nuclei; black arrows: TRACP + MNC with >5 nuclei. b PFF-CM from control serum-pretreated osteocytes inhibited the formation of osteoclasts with 3–5 nuclei. Stat-CM from RA serum-pretreated osteocytes enhanced the formation of osteoclasts with 3–5 nuclei, and PFF-CM from RA serum-pretreated osteocytes ified this effect. c PFF-CM from RA serum-pretreated osteocytes inhibited the formation of osteoclasts with >5 nuclei. Values are mean from three independent experiments, n = 9. Significant effect of Stat-CM from RA serum-pretreated osteocytes, *p < 0.05. Significant effect of PFF-CM, ##p < 0.01, ###p < 0.001
Mentions: PFF-CM from control serum-pretreated osteocytes decreased the number of TRACP-positive osteoclasts with 3–5 nuclei by 1.6-fold (Fig. 3a, b). Stat-CM from RA serum-pretreated cell culture increased the number of TRACP-positive osteoclasts with 3–5 nuclei by 1.2-fold, while PFF ified this effect (Fig. 3a, b). PFF-CM from RA serum-pretreated osteocytes also inhibited formation of osteoclasts with >5 nuclei by 1.8-fold (Fig. 3a, c).Fig. 3

Bottom Line: However, RA serum enhanced the RANKL/OPG expression ratio by 3.4-fold, while PFF ified this effect.Stat-CM from RA serum-pretreated osteocytes enhanced osteoclastogenesis compared with stat-CM from control serum-pretreated osteocytes, while PFF ified this effect.In conclusion, RA serum, containing inflammatory factors, did not alter the intrinsic capacity of osteocytes to sense mechanical stimuli, but upregulated osteocyte-to-osteoclast communication.

View Article: PubMed Central - PubMed

Affiliation: Department of Oral Cell Biology, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and VU University Amsterdam, MOVE Research Institute Amsterdam, Amsterdam, The Netherlands.

ABSTRACT
Multiple factors contribute to bone loss in inflammatory diseases such as rheumatoid arthritis (RA), but circulating inflammatory factors and immobilization play a crucial role. Mechanical loading prevents bone loss in the general population, but the effects of mechanical loading in patients with RA are less clear. Therefore, we aimed to investigate whether mechanical stimuli reverse the stimulatory effect of RA serum on osteocyte-to-osteoclast communication. Human primary osteocytes were pretreated with 10 % RA serum or healthy control serum for 7 days, followed by 1 h ± mechanical loading by pulsating fluid flow (PFF). Nitric oxide (NO) and prostaglandin E2 were measured in the medium. Receptor activator of nuclear factor-kappaB ligand (RANKL), osteoprotegerin (OPG), interleukin-6 (IL-6), cyclooxygenase-2 (COX2), matrix-extracellular phosphoglycoprotein (MEPE), cysteine-rich protein 61 (CYR61), and SOST gene expression was quantified by qPCR. Osteoclast precursors were cultured with PFF-conditioned medium (PFF-CM) or static-conditioned medium (stat-CM), and osteoclast formation was assessed. RA serum alone did not affect IL-6, CYR61, COX2, MEPE, or SOST gene expression in osteocytes. However, RA serum enhanced the RANKL/OPG expression ratio by 3.4-fold, while PFF ified this effect. PFF enhanced NO production to the same extent in control serum (2.6-3.5-fold) and RA serum-pretreated (2.7-3.6-fold) osteocytes. Stat-CM from RA serum-pretreated osteocytes enhanced osteoclastogenesis compared with stat-CM from control serum-pretreated osteocytes, while PFF ified this effect. In conclusion, RA serum, containing inflammatory factors, did not alter the intrinsic capacity of osteocytes to sense mechanical stimuli, but upregulated osteocyte-to-osteoclast communication. Mechanical loading ified this upregulation, suggesting that mechanical stimuli could contribute to the prevention of osteoporosis in inflammatory disease.

No MeSH data available.


Related in: MedlinePlus