Cell populations of tendon: a simplified method for isolation of synovial cells and internal fibroblasts: confirmation of origin and biologic properties

Abstract

Tendons transmit the force of muscle contraction to bone to effect limb movement. Special structural and biological properties of tendon have developed to facilitate force transmission. The tendon has a complex organization of cells surrounding the collagen bundles inside tendon as well as at the tendon surface. Internal cells may act to maintain the bulk of the collagen in tendon. External cells in the epitenon may provide lubrication for tendon gliding. To develop better understanding of these processes and the roles the cell populations play, we isolated cells from the surface and interior of tendon and studied them in vitro. Flexor tendons from 8-week-old white Leghorn chickens were separated into two distinct cell populations: the outer synovial cells and the fibroblasts more internal in tendon. These cell populations were discernible by their locations in the intact tendon, determined by sequential enzymatic and physical release from their substrata. Initially, some cells eluted in Hanks' salt solution (HSS) (population 1); then synovial cells were released after a 2-min treatment with 0.5% collagenase (population 2). Next, a population of synovial cells was released in high yield by treatment with 0.25% trypsin (step III, population 3). Step III, population 3 cells were used as synovial cells (SCs). Next, a population of SCs and fibroblasts were released by scraping with a rubber policeman (population 4). Subsequently, fibroblasts were released after incubation with 0.5% collagenase (population 5). A more direct procedure (procedure 2) to isolate the synovial and internal tendon cells involved treatment in 0.5% collagenase followed by sedimentation at 900 g. Cells that sedimented were largely fibroblasts, whereas the cells that remained at the top of the tube were largely SCs. Cells designated as SCs, isolated by procedure 2, most likely contained surface cells from epitenon and internal interfascicular cells from endotenon and paratenon. Surface tendon cells separated by sequential enzymatic and physical release from their substrata (by procedure 1) had all the following characteristics: distinct subpopulations of cells based on morphology; presence of cytoplasmic, lipid-containing vesicles; decreased sensitivity to trypsin; and reduced generation time as compared with that of internal fibroblasts. Conversely, the internal fibroblasts (IFs) appeared to represent a more uniform population based on morphological characteristics.