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A Comparison of the Biological Activities of Human Osteoblast hFOB1.19 Between Iron Excess and Iron Deficiency

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Abstract

Bone metabolism has a close relationship with iron homeostasis. To examine the effects of iron excess and iron deficiency on the biological activities of osteoblast in vitro, human osteoblast cells (hFOB1.19) were incubated in a medium supplemented with 0–200 μmol/L ferric ammonium citrate and 0–20 μmol/L deferoxamine. The intracellular iron was measured by a confocal laser scanning microscope. Proliferation of osteoblasts was evaluated by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide assay. Apoptotic cells were detected using annexin intervention V/PI staining with a flow cytometry. Alkaline phosphatase (ALP) activity was measured using an ALP assay kit. The number of calcified nodules and mineral area was evaluated by von Kossa staining assay. The expressions of type I collagen and osteocalcin of cultured osteoblasts were detected by reverse transcriptase polymerase chain reaction and Western blot. Intracellular reactive oxygen species (ROS) was measured using the oxidation-sensitive dye 2,7-dichlorofluorescin diacetate by flow cytometry. The results indicated that excessive iron inhibited osteoblast activity in a concentration-dependent manner. Low iron concentrations, in contrast, produced a biphasic manner on osteoblasts: mild low iron promoted osteoblast activity, but serious low iron inhibited osteoblast activity. Osteogenesis was optimal in certain iron concentrations. The mechanism underlying biological activity invoked by excessive iron may be attributed to increased intracellular ROS levels.

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Abbreviations

FAC:

Ferric ammonium citrate

DFO:

Deferoxamine

ALP:

Alkaline phosphatase

COL-I:

Type I collagen

OC:

Osteocalcin

RT-PCR:

Reverse transcriptase polymerase chain reaction

ROS:

Reactive oxygen species

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Acknowledgments

This work was partially supported by the National Natural Science Foundation of China (no. 81273090), Jiangsu provincial grant (no. BK2012608), and science and technology projects of Suzhou (no. 510303).

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Authors and Affiliations

  1. Department of Orthopaedics, The Second Affiliated Hospital of Soochow University, Suzhou, 215004, China

    Guo-yang Zhao, Li-ping Zhao, Yin-feng He, Guang-Fei Li, Chao Gao & You-jia Xu

  2. Department of Molecular Diagnostics and Biopharmaceutics, College of Pharmacy, Soochow University, Suzhou, 215004, China

    Kai Li

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  1. Guo-yang Zhao
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Correspondence to You-jia Xu.

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Zhao, Gy., Zhao, Lp., He, Yf. et al. A Comparison of the Biological Activities of Human Osteoblast hFOB1.19 Between Iron Excess and Iron Deficiency. Biol Trace Elem Res 150, 487–495 (2012). https://doi.org/10.1007/s12011-012-9511-9

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