The effects of patient age on human osteoblasts' response to Ti–6Al–4V implants in vitro

Abstract

Osseointegrated implants are a common therapy for the elderly population as lifespan increases. Understanding the effects of age and sex on osseointegration is important for successful implant therapy. Therefore, the response of primary human osteoblasts (HOB) to implant materials was studied. HOBs were obtained by outgrowth of cells from bone from orthopaedic procedures and categorized as Young (Y), <15; Middle (M), 30-50; and Old (0), >60 years old. Initially the HOB phenotype was determined on tissue culture plastic. Alkaline phosphatase (ALP) staining and activity were significantly increased in HOBs from older patients. Message levels of type I collagen (COL), bone sialoprotein (BSP) and ALP were significantly higher (from 2.3-to 3.8-fold) in Y subjects compared to M and 0 patients at 2 weeks. Studies of the response of HOBs to implant materials were undertaken using Ti-6A14V disks prepared in a manner similar to orthopaedic implants. A I . '!-fold (p < 0.05) increase in cell attachment was found inHOBs from Y compared with 0 in female subjects but not in male subjects. Cell proliferation at 24 h was not significantly different by age or sex, nor was DNA content different at 2 and 4 weeks. Mineralization in HOB-implant cultures was 2.3-fold higher in Y than in 0, and 1.7-fold higher in Y compared to M HOBs from female but not male subjects at 4 weeks. Northern blot and RT-PCR analysis at 2 weeks of culture showed significantly higher levels (1.6-2.3-fold) of COL, BSP, and osteocalcin (OC) mRNAs in Y HOBs compared to M and 0 HOBs from female subjects. We conclude that human osteoblasts from older female patients have a decreased ability to form bone on implants.

Porous tantalum stimulates the proliferation and osteogenesis of osteoblasts from elderly female patients

“…7,11,21 The resulting material is composed of approximately 99% tantalum and 1% vitreous carbon (by weight), has a pore size between 400 and 600 mm and porosity of approximately 80%. 14,22 TFM was prepared as discs 20 mm  1.7 mm (diameter  thickness) die-punched from sintered cpTi. TFM fabrication involves intertwining multiple cpTi fibers, sintering at high temperatures to weld fibers into a mesh structure.…”

“…The first three passages were used for experiments. We have previously shown that more than 90% of these cells express alkaline phosphatase (ALP), an early marker of osteoblast progenitors, 14 and also over 80% of them express activated leukocyte cell adhesion molecule (ALCAM) which has been found in osteogenic precursors. 26 For mineralization, quantitative PCR and scanning electron microscopy (SEM), cells were cultured in the proliferation medium for 1 week, and for 2 more weeks in the differentiation medium of a-MEM, 10% FBS, 1% antibiotics, and 50 mg/ml of fresh ascorbic acid added daily.…”

“…14 To eliminate any radioactive thymidine being retained inside the substrates, the discs were cultured without cells and levels of radioactivity were measured and corrected. To normalize [ 3 H]-thymidine incorporation to DNA content, additional wells of cells cultured for 48 h were scraped off in 1 N NaOH and digested overnight at 48C, a technique that has routinely been used during DNA isolation to selectively degrade RNA in the DNA samples.…”

“…14 Quantitative PCR Analysis Total RNA was isolated from the cells cultured for 3 weeks by using TRI Reagent (Molecular Research Center, Cincinnati, OH). Two micrograms of RNA was reverse transcribed into cDNA followed by quantitative PCR for type I collagen, ALP, and osteocalcin (OCN), as previously described and normalized to GAPDH.…”

“…7,[11][12][13] Previous in vitro studies in our lab have demonstrated that the response of human bone cells to orthopedic and dental materials varies depending on the sex and age of the patient. 14 In cell culture, cells grown out of bone from female patients older than 60 years old demonstrated decreased attachment to implant materials, mineralization, and expression levels of bone matrix proteins compared to those cells from bone samples of younger females and males of all ages. These isolated cells had previously been shown to have the potential to become either osteoblasts, adipocytes or chondrocytes and, therefore, have been named ''multipotential mesenchymal stem cells from adult bone''.…”