Methods: Bone marrow derived hMSCs (cryopreserved) were rapidly thawed and then plated (30,000 cells/disk) onto 12 mm cpTitanium disks prepared by machining (mTi) or by grit-blasting and acid etching. (eTi). HMSCs were cultured for 7 days in DMEM containing 10% fetal bovine serum. These loading conditions were shown to provide viable hMSCs that differentiate into osteoblastic cells in a SCID mouse model. Total RNA was isolated from mTi-and eTi-adherent cell layers using TriSol and quantified by UV spectrophotometry. 32P-labelled probes were synthesized for mTi and eTi derived RNAs (Atlas Pure Toltal RNA Labering System, Clontech) and human Atlas 1.2 gene arrays were hybridized overnight at 65oC, washed at high stringency and exposed using phosphoimaging screens. Digitally captured images were analyzed using Atlas Image 2.0(Clontech) software.
Results: The comparison of eTi- and mTi-adherent hMSC mRNA levels revealed that 45 gene levels were 2 fold greater on eTi compared to mTI. 104 mRNA levels were 2 fold lower on eTi. Of potential interest, ICAM-1 was elevated 34-fold, erbB2 was elevated 9.8 fold and CTGF was elevated 9.6 fold.
Conclusion: Cell adhesion and associated signal transduction related to surface characteristics are attractive targets for continued endosseous implant engineering. The use of gene array technology to examine global transcriptional changes that result from cell adhesion to different endosseous implants is possible. Ongoing studies seek to challenge this technology using linear amplification of small amounts of RNA derived from in vivo samples to probe arrays of 8,300 – 18,000 genes.
This research was supported by AADR Fellowship.