Cell engineering the osteoblast response to topography

Biomedical engineering

Ferris, Diana Marshall

University:University of Glasgow

关键词: Biomedical engineering;   

Others  :  http://theses.gla.ac.uk/71888/1/10394691.pdf

来源: University of Glasgow

【 摘 要 】

Grooved, rough, and porous surfaces are in use clinically to stabilise the bone/implant interface and encourage osseointegration for hip and knee replacements and dental implants. To date, there is no one topographical surface that is considered superior. No one has ever directly observed the process of bone formation by cells in three dimensional spaces. Also, little work on the response of cells to curved surfaces exists. The aim of this work was two-fold: to isolate and characterise primary osteoblasts from rat and human sources, and to study the response of those cells to different topographies. Fused silica grooved surfaces (width = 2 - 100 mum, depth =10 nm - 6.0 mum), polyurethane replicas, and fine quartz tubes (diameter = 150-700 mum, length = 5 mm - 2.5 cm) provided an array of topographical environments. Besides conventional techniques like video time lapse scanning cinemicrography. scanning electron microscopy, immunofluorescent (confocal scanning laser microscopy) and histological staining, two new techniques were used to assess osteoblast extracellular matrix production and orientation; polarised light microscopy and atomic force microscopy. Findings included rat and human osteoblast sensitivity to grooved features greater than 80 and 100 nm in depth respectively. Furthermore, preliminary results suggested that grooved surfaces (width: 5 mum, depth: 6 mum) influence extracellular matrix production, i.e. the alignment of collagen and mineral with the groove long axis. The ability to influence and control the orientation of new bone via topography is the first step towards tissue engineering organised bone. In addition, the ability to control new bone growth could have an impact in the acceleration and enhancement of the wound healing and repair process. Video time lapse cinemicrography revealed that within an hour of seeding, osteoblasts in tubes had extended towards each other and formed dynamic cord-like structures that spanned the tube diameter and along its length. Furthermore, after a few days, cells had formed nodule-like structures usually associated with two dimensional tissue culture despite the lack of ascorbic acid and ?-glycerophosphate. Examination of these tubular environments with polarised light revealed birefringent particles present in some of these nodules. Osteocalcin staining showed brightly stained globules produced along cell cords and suggested the inner wall of the tube was coated by small mineralised particles. In summary, the findings presented in this work demonstrate the ability of both material surfaces modified in a regular manner (grooves) and extended concave surfaces of small diameter (tubes) to influence osteoblast behaviour and mineralisation in vitro. There is some evidence to suggest that grooved surfaces influence extracellular matrix orientation, i.e. collagen and mineral alignment. Also, cells seeded into a three dimensional tubular environment behaved differently than similar' cells on flat surfaces in terms of overall activity, and extracellular matrix production. In conclusion, this work imparts new information on the response of osteoblasts to topographical surfaces and environments that could lead to the tissue engineering of bone and redesign of implant surfaces.