The Impact of Metal Ion Exposure on the Cellular Behavior of Human Osteoblasts and PBMCs: In Vitro Analyses of Osteolytic Processes
Abstract
:1. Introduction
2. Results
2.1. Effects of CoCr29Mo6 Ions in Single Cultures of Human Osteoblasts and PBMCs
2.1.1. Viability
2.1.2. Apoptosis and Necrosis
2.1.3. Osteoblastic Differentiation and Induction of Osteoclastic Differentiation Markers
2.1.4. Gene Expression of Pro-Osteolytic Mediators
2.2. Influence of CoCr29Mo6 Ions in Co-Cultures
2.2.1. Adherent and Non-Adherent PBMCs
2.2.2. PBMCs and Osteoblasts
3. Discussion
4. Conclusions
5. Materials and Methods
5.1. Isolation of Human Osteoblastic Cells and PBMCs
5.2. Cell Viability
5.3. Determination of Apoptosis and Necrosis
5.4. Gene Expression Analyses
5.5. Pro-Collagen Type 1 Synthesis Rate
5.6. Experimental Design
5.7. Data Illustration and Statistical Analysis
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Differentiation Marker | Osteoblasts | Adherent PBMCs | ||
---|---|---|---|---|
48 h | 96 h | 48 h | 96 h | |
Col1A1 | 95 (57; 119) | 214 ** (104; 411) | - | - |
C1CP (protein) | 80 (66; 102) | 136 (85; 179) | - | - |
ALP | 91 (42; 124) | 116 (76; 141) | - | - |
OC | 110 (82; 248) | 90 (72; 193) | - | - |
OPG | 95 (36; 162) | 107 (19; 139) | - | - |
RANKL | 286 (5; 2026) | 46 (14; 837) | - | - |
RANK | - | - | 56 *,# (50; 76) | 36 *,# (35; 49) |
TRAP5b | - | - | 5 * (2; 9) | 7 * (2; 11) |
Pro-Osteolytic Mediators | Osteoblasts | Adherent PBMCs | ||
---|---|---|---|---|
48 h | 96 h | 48 h | 96 h | |
MMP1 | 95 (36; 162) | 52 (36; 144) | 234 (92; 338) | 161 (23; 778) |
TIMP1 | 140 * (110; 299) | 110 (66; 269) | 91 (30; 337) | 122 (89; 675) |
IL-6 | 276 ** (117; 546) | 99 (75; 204) | 120 (14; 659) | 94 (56; 166) |
IL-8 | 143 (48; 247) | 58 * (50; 73) | 404 * (127; 1594) | 246 (34; 4104) |
TNF-α | 104 (51; 165) | 198 (51; 435) | 129 (96; 312) | 83 (76; 130) |
MCP-1 | 120 (42; 149) | 101 (68; 130) | 29 * (19; 41) | 703 (13; 1628) |
Differentiaton or Pro-Osteolytic Mediators | Osteoblasts | Adherent PBMCs |
---|---|---|
Col1A1 | 31 * (24; 49) | - |
RANKL | 2 (1; 5) | - |
OPG | 155 * (118; 206) | - |
RANK | - | 85 (47; 108) |
TRAP5b | - | 10,649 * (9263; 12,099) |
MMP1 | 128 (111; 307) | 285 * (212; 361) |
TIMP1 | 2950 * (307; 8706) | 2573 * (978; 3490) |
IL-6 | 5,306,604 * (3,684,037; 5,701,287) | 1093 * (700; 1315) |
IL-8 | 4,089,451 * (1,769,434; 5,611,116) | 31,637 * (21,602; 46,144) |
TNF-α | 489,124 * (194,021; 741,895) | 603 * (258; 887) |
MCP-1 | 207,580 * (169,374; 267,626) | 654 (98; 1277) |
Primer | Forward (Sequence 5′-3′) | Reverse (Sequence 5′-3′) |
---|---|---|
Alkaline phosphatase (ALP) | cattgtgaccaccacgagag | ccatgatcacgtcaatgtcc |
Collagen 1 (Col1A1) | acgaagacatcccaccaatc | agatcacgtcatcgcacaac |
Hypoxanthine-guanine phosphoribosyltransferase (HPRT) | ccctggcgtcgtgattagtg | tcgagcaagacgttcagtcc |
Interleukin 6 (IL-6) | tggattcaatgaggagacttgcc | ctggcatttgtggttgggtc |
Interleukin 8 (IL-8) | tctgtgtgaaggtgcagttttg | atttctgtgttggcgcagtg |
Matrix metalloproteinase 1 (MMP1) | agagcagatgtggacatgc | tcccgatgatctcccctgac |
Monocyte chemotactic protein 1 (MCP-1) | ccgagaggctgagactaacc | ggcattgattgcatctggctg |
Osteocalcin (OC) | ggtgcagcctttgtgtcc | tcagccaactcgtcacagtc |
Osteoprotegerin (OPG) | tggattcaatgaggagacttgcc | ctggcatttgtggttgggtc |
Receptor activator of nuclear κ-b (RANK) | agaaaaccaccaaatgaacccc | gccacaagcctcattgatcc |
Receptor activator of nuclear κ-b-Ligand (RANKL) | agaagccaccaaagaattgcag | accatcgctttctctgctctg |
Tartrate-resistant acid phosphatase 5b (TRAP5b) | gggagatctgtgagccagtg | gtccacatgtccatccaggg |
Tissue inhibitor of metallo-proteinase 1 (TIMP1) | attgctggaaaactgcaggatg | gtccacaagcaatgagtgcc |
Tumour necrosis factor α (TNF-α) | gttgtagcaaaccctcaagctg | gaggtacaggccctctgatg |
Content in | Co | Cr | Mo | Ni |
---|---|---|---|---|
stock solution | 12.0 ± 2.4 mg/L | 3.9 ± 0.6 mg/L | 0.9 ± 0.1 mg/L | 1.3 ± 0.6 mg/L |
experimental solution | 120 ± 24 µg/L | 39 ± 5.7 µg/L | 8.8 ± 1.1 µg/L | 12.8 ± 6.0 µg/L |
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Jonitz-Heincke, A.; Tillmann, J.; Klinder, A.; Krueger, S.; Kretzer, J.P.; Høl, P.J.; Paulus, A.C.; Bader, R. The Impact of Metal Ion Exposure on the Cellular Behavior of Human Osteoblasts and PBMCs: In Vitro Analyses of Osteolytic Processes. Materials 2017, 10, 734. https://doi.org/10.3390/ma10070734
Jonitz-Heincke A, Tillmann J, Klinder A, Krueger S, Kretzer JP, Høl PJ, Paulus AC, Bader R. The Impact of Metal Ion Exposure on the Cellular Behavior of Human Osteoblasts and PBMCs: In Vitro Analyses of Osteolytic Processes. Materials. 2017; 10(7):734. https://doi.org/10.3390/ma10070734
Chicago/Turabian StyleJonitz-Heincke, Anika, Jenny Tillmann, Annett Klinder, Simone Krueger, Jan Philippe Kretzer, Paul Johan Høl, Alexander C. Paulus, and Rainer Bader. 2017. "The Impact of Metal Ion Exposure on the Cellular Behavior of Human Osteoblasts and PBMCs: In Vitro Analyses of Osteolytic Processes" Materials 10, no. 7: 734. https://doi.org/10.3390/ma10070734
APA StyleJonitz-Heincke, A., Tillmann, J., Klinder, A., Krueger, S., Kretzer, J. P., Høl, P. J., Paulus, A. C., & Bader, R. (2017). The Impact of Metal Ion Exposure on the Cellular Behavior of Human Osteoblasts and PBMCs: In Vitro Analyses of Osteolytic Processes. Materials, 10(7), 734. https://doi.org/10.3390/ma10070734