The overall of this research is to examine in cell culture the developmental potential of prechondroblasts from the mandibular condylar cartilage (MCC). This will allow a better understanding of the growth, differentiation and adaptation of this secondary cartilage which covers one of the articulating surfaces of the temporomandibular joint and is involved in the growth and development of the craniofacial complex. Three distinct cellular layers can be observed on histologic sections of this cartilage. Small prechondroblasts, normally sized functional chondroblasts and large hypertrophic chondroblasts. Studies in vivo and in organ culture have shown that during growth of the MCC, proliferation only occurs among non-differentiated prechondroblasts. These cells are surrounded by a non cartilaginous matrix and located directly under the fibrous articular layer. Prechondroblasts differentiate into chrondroblasts which do not proliferate, but synthesize a cartilaginous matrix and mature to a hypertrophic stage. Very little is known concerning the cellular mechanisms involved in this process. The research described here will develop methods for growing pure cultures of functioning, phenotypically authentic prechondroblasts from the MCC, in order to obtain a system in which their growth and differentiation may be studied.
The specific aims are (1) to isolate the different cell populations of the MCC using density gradient centrifugation techniques; (2) to establish for each isolated cell population its optimum growth culture system; and (3) to verify morphologically and biochemically in primary culture at confluence the phenotypic expression of each isolated cell population. Cells will be isolated from the MCC of 3 to 4 weeks old New Zealand white rabbit. Prechondroblasts will be separated from chondroblasts by isopycnic centrifugation and by rate zonal centrifugation using Percoll as a density gradient material. For each isolated cell population, growth curves are traced for initial plating densities varying from 2 x 104 cells/cm2 to 3 x 105 cells/cm2. Prechondroblasts are expected to proliferate in culture better and at a lower initial plating density than the cells from the other fractions. The morphologic phenotype of the culture is documented as function of time by light and electron microscopy. The presence of cartilage specific glycosaminoglycan in the extracellular matrix is tested by ELISA; collagen type is identified by SDS-PAGE of cyanogen bromide peptides from collagen alpha chains.
| Engel, F E; Swain, L D; Dean, D D et al. (1994) Nodule formation and calcification of mandibular condylar cartilage cell cultures mimic in vivo ultrastructure. Acta Anat (Basel) 150:136-49 |
