Cytocentrifugation: a convenient and efficient method for seeding tendon-derived cells into monolayer cultures or 3-D tissue engineering scaffolds

Abstract

Tendon and ligament injuries are very common, requiring some 200,000 reconstructions per year in the USA. Autografting can be used to repair these but donor tissue is limited and harvesting leads to morbidity at the graft sites. Tissue engineering has been used to grow simple tissues such as skin, cartilage and bone and due to their low vascularity and simple structure, tendons should be ideal candidates for such an approach. Scaffolds are essential for tissue engineering as they provide structure and signals that regulate growth. However, they present a physical barrier to cell seeding with the majority of the cells congregating at the scaffold surface. To address this we used centrifugation to enhance penetration of tendon-derived cells to the centres of 3-D scaffolds. The process had no apparent deleterious effects on the cells and both plating efficiency and cell distribution improved. After attachment the cells continued to proliferate and deposit a collagenous matrix. Scaffold penetration was investigated using layers of Azowipes allowing the separation and examination of individual leaves. At relatively low g-forces, cells penetrated a stack of 6 Azowipes leaving cells attached to each leaf. These data suggest that cytocentrifugation improves the penetration and homogeneity of tendon derived cells in 3-D and monolayer cultures.

Role of the ERK1/2 Signaling Pathway in Osteogenesis of Rat Tendon-Derived Stem Cells in Normoxic and Hypoxic Cultures

“…Tendinopathy is a common painful tendon condition caused by overuse, mechanical injury or intrinsic degeneration 1 - 3 . Histologically, calcification is usually reported in some tendinopathies 4 , 5 , which leads to a failed self-healing and predisposes the diseased tendon to rupture 6 .…”

Direct contact-mediated non-viral gene therapy using thermo-sensitive hydrogel-coated dressings

“…As hydrogels have been used to generate moisturizing drug delivery platforms, we hypothesized that our GET-laden hydrogel system could be used on a fabric substrate to form a bandage that could be used to transfer therapeutics. Other groups have employed a biologically inert non-hydrolysable scaffold (commercially available as the product Azo-wipe™, Synergy Healthcare) for cell culture [54,58], so we used this substrate coated with GET-laden hydrogels with the aim to deliver transfection to the immediate environment (Fig. 5).…”

A Sample Preparation Protocol for High Throughput Immunofluorescence of Suspension Cells on an Adherent Surface

“…Some protocols and equipment have been developed to mechanically attach cells to surfaces. 10,11 However, even if suspension cells initially sediment and manage to attach to the plates or glass slides, the shear stress and flow from pipetting during the sample preparation can be sufficient to detach cells from the surface. A typical IF protocol consists of multiple steps including fixation, permeabilization, and primary and secondary antibody incubation along with several washing steps.…”