Gene Expression in HUMAN Osteoblastic Cells Under Mechanical Strain

Objectives:,

The mechanosensing mechanism by which the osteoblastic cells transduce mechanical stimuli into biochemical signals is still not clearly understood so the aims of this study were: 1. Identify gene expression of RunX2, C–fos and osteocalcin (OC) as specific markers of bone regeneration process in cultured human osteoblasts under mechanic deformation. 2. Quantify osteoblastic proliferation under mechanic deformation.

Methods:,

Human osteoblasts of American Type Culture Collection Ref. CRL-11372 cultured in 75cm2 flasks received deformation forces 500, 1.000 and 1.500 micro-strains (µS). Stimulated each 6 hours, during 6 hours for 4 days. Mechanotransductor was inside CO2 camera to maintain cell conditions in vitro. Gene expression was detected by real time RT-PCR using GAPDH (housekeeping). Control cells did not receive stimulation. Proliferation, morphology and cellular viability were controlled. Statistics by aleatory blocks.

Results:,

Gene Expression

The three genes had increased expression with stimulation forces, but each one presented a higher expression with different strains, as follows:

• C-fos - 500µS.

• RunX2 - 1.000µS.

• OC - 1.500µS.

Proliferation

Was increased in 400% with 500µS.

Viability

None of the strains affected cell viability.

Conclusion:,

RunX2 and C-fos are soon markers of bone formation, can be activated with low forces present in not mineralized tissue. OC expression appears in latest process of bone formation because it only can be activated with stronger forces. This suggests, that bone in vivo is not able to activate OC in earliest stadium because mineralized tissue is not ready to transfer enough strain to activate this gene.

Cell proliferation can be activated by mechanotransduction in earliest stadium of bone formation. But in mineralized tissue forces are very strong to activate it.