Background

Distal femoral fractures are 10 times less common than hip fractures. 12-month mortality has been reported as 25–30% but there is no longer-term data. In Northumbria hip fractures have a 5-year mortality of 68%.

Control of stem cell fate and function is critical for clinical and academic work. By combining surface chemistry-driven extracellular matrix (ECM) assembly with mesenchymal stem cells (MSCs) we are developing a system which can be used to regulate the behaviour of MSCs. The conformation of the ECM glycoprotein fibronectin (Fn) is different when adsorbed onto poly methylacrylate (PMA) where it is globular, and on poly ethylacrylate (PEA) where it forms a physiologically-similar network^[1] (Fig. 1). Using these polymers to govern Fn conformation, we are developing a 3D system incorporating MSC-responsive growth factors (GFs) and bone marrow MSCs capable of regulating MSC behaviour.

Toluene-dissolved PMA and PEA were spin coated onto glass coverslips before solvent extraction in vacuo and UV sterilisation. 20 mg ml⁻¹ human plasma FN was adsorbed onto the surfaces followed by 25 ng ml⁻¹ recombinant human BMP2/VEGF. FN conformations were characterised by atomic force microscopy (AFM). A collagen hydrogel was placed above the substrate. Adult human bone marrow STRO-1+ were cultured on the substrates for 3 weeks in supplemented DMEM. Expression of MSC stemness and HSC maintenance factors were analysed by In-Cell Western assay.

To establish the best combination of polymer/FN/GF, MSC stemness markers (ALCAM, NESTIN and STRO1), osteogenic differentiation markers (OCN and OPN) and bone marrow markers (SCF and VCAM1) were measured in MSCs cultured for 3-weeks on substrates. OCN, SCF, and VCAM1 expression was enhanced across all combinations compared to glass control, while for ALCAM/STRO1/NESTIN and OPN, PEA combinations enhanced their expression. PEA + FN + VEGF appeared to be system best suited to maintaining MSC stemness and supporting expression of osteogenesis markers and bone marrow markers.

We have shown that MSCs maintain their stem cells state and express high levels of SCF and VCAM-1 when cultured on PEA with adsorbed Fn and VEGF or BMP2. Next stages of this work will use PCR to verify results and analyse expression of other MSC markers to develop a role for these synthetic polymers as biomaterials.

Introduction

Cauda equina syndrome represents the constellation of symptoms and signs resulting from compression of lumbosacral nerve routes. Combined with subjective neurological findings, a reduction in anal tone is an important sign deeming further imaging necessary. Our main objective was to investigate the validity of DRE for assessment of anal tone.

Critical size defects in ovine tibiae, stabilised with intramedullary interlocking nails, were used to assess whether the addition of carboxymethylcellulose to the standard osteogenic protein-1 (OP-1/BMP-7) implant would affect the implant’s efficacy for bone regeneration. The biomaterial carriers were a ‘putty’ carrier of carboxymethylcellulose and bovine-derived type-I collagen (OPP) or the standard with collagen alone (OPC). These two treatments were also compared to “ungrafted” negative controls. Efficacy of regeneration was determined using radiological, biomechanical and histological evaluations after four months of healing. The defects, filled with OPP and OPC, demonstrated radiodense material spanning the defect after one month of healing, with radiographic evidence of recorticalisation and remodelling by two months. The OPP and OPC treatment groups had equivalent structural and material properties that were significantly greater than those in the ungrafted controls. The structural properties of the OPP- and OPC-treated limbs were equivalent to those of the contralateral untreated limb (p > 0.05), yet material properties were inferior (p < 0.05). Histopathology revealed no residual inflammatory response to the biomaterial carriers or OP-1. The OPP- and OPC-treated animals had 60% to 85% lamellar bone within the defect, and less than 25% of the regenerate was composed of fibrous tissue. The defects in the untreated control animals contained less than 40% lamellar bone and more than 60% was fibrous tissue, creating full cortical thickness defects. In our studies carboxymethylcellulose did not adversely affect the capacity of the standard OP-1 implant for regenerating bone.