In vitro osteoblastic differentiation of human bone marrow cells in the presence of metal ions.
For periods up to 21 days human bone marrow was cultured in control conditions that favor the proliferation and differentiation of osteoblastic cells. The effect of AISI 316L corrosion products and the corresponding major separate metal ions (Fe, Cr, and Ni) were studied in three different phases of the culture period in order to investigate the effects of metal ions in cell populations representative of osteoblastic cells in different stages of differentiation. Toxicity consequences of the presence of metal ions in bone marrow cultures were evaluated by biochemical parameters (enzymatic reduction of MTT, alkaline phosphatase activity, and total protein content), histochemical assays (identification of ALP-positive cells and Ca and phosphates deposits), and observation of the cultures by light and scanning electron microscopy. Culture media were analyzed for total and ionized Ca and P and also for metal ions (Fe, Cr, and Ni). The presence of AISI 316L corrosion products and Ni salt in bone marrow cultures during the first and second weeks of culture significantly disturbs the normal behavior of these cultures, interfering in the lag phase and exponential phase of cell growth and ALP expression. However, the presence of these species during the third week of culture, when expression of osteoblastic functions occurs (mineralization process), did not result in any detectable effect. Fe salt also disturbs the behavior of bone marrow cell cultures when present during the lag phase and proliferation phase, and a somewhat compromised response between the normal pattern (control cultures) and intense inhibition (AISI 316L corrosion products and Ni salt-added cultures) was observed. Fe did not affect the progression of the mineralization phase. Osteogenic cultures exposed to Cr salt (Cr3+) presented a pattern similar to the controls, indicating that this element does not interfere, in the concentration studied, in the osteoblastic differentiation of bone marrow cells. Quantification of metal ions in the culture media showed that Cr (originated from AISI 316L corrosion products but from not Cr3+ salt) and Ni (originated from AISI 316L corrosion products and Ni salt) appear to be retained by the bone marrow cultures.
INEB-Instituto de Engenharia Biomédica, Departamento de Engenharia Química, Faculdade de Engenharia da Universidade do Porto, Praça do Coronel Pacheco, 1, 4050 Porto, Portugal.
S
S
Morais
N
N
Dias
J P
JP
Sousa
M H
MH
Fernandes
G S
GS
Carvalho
D000469:Alkaline Phosphatase / Q000378:metabolism; D000497:Alloys / Q000009:adverse effects; D001672:Biocompatible Materials / Q000009:adverse effects; D001854:Bone Marrow Cells / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects*; D002455:Cell Division; D002478:Cells, Cultured; D002857:Chromium / Q000009:adverse effects; D003343:Corrosion; D006801:Humans; D066298:In Vitro Techniques; D007501:Iron / Q000009:adverse effects; D008422:Materials Testing; D008670:Metals / Q000009:adverse effects*; D009532:Nickel / Q000009:adverse effects; D010006:Osteoblasts / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D010758:Phosphorus / Q000378:metabolism; D019736:Prostheses and Implants / Q000009:adverse effects; D011506:Proteins / Q000378:metabolism; D013193:Stainless Steel / Q000009:adverse effects; D013778:Tetrazolium Salts / Q000378:metabolism; D013844:Thiazoles / Q000378:metabolism
D000497:Alloys; D001672:Biocompatible Materials; D008670:Metals; D011506:Proteins; D013778:Tetrazolium Salts; D013844:Thiazoles; D002857:Chromium; D013193:Stainless Steel; D010758:Phosphorus; D009532:Nickel; D007501:Iron; D000469:Alkaline Phosphatase; C022616:thiazolyl blue; D002118:Calcium