The Loss of Phenotypic Traits by Differentiated Cells

Abstract

When mitotically quiescent chick chondroeytes are liberated from their polysaccharide matrix and cultured on a fibrin dot in the presence of embryo extract, the following is observed: (a) Within 18 hr the rounded chondrocytes transform into fibroblastic cells and cease synthesizing chondroitin sulfate. (b) Within 24 hr over 50% of the cells incorporate thymidine, and by 35 hr over 95% have completed at least one cell division. (c) The amount of chondroitin sulfate synthesized per cell in the log phase of growth is less than 5% that of the nondividing progenitor cells. (d) If after 7-10 days such fibroblastic chondrocytes contact their siblings, many resume a rounded shape and synthesize chondroitin sulfate in quantifies characteristic of their in vivo progenitors. (e) When cultured as fibroblastic cells for over 2 wk and then allowed, at high density to contact siblings, they neither round up nor deposit chondroitin sulfate. (/) Fibroblastic chondrocytes obtained by protracted culturing do not display the surface affinities of normal chondrocytes. Chondrocyte progeny which do not deposit chondroitin sulfate or aggregate with one another or with normal chondrocytes have been termed "dedifferentiated" or "altered" chondrocytes (1-5). Many of these observations have been confirmed by Kuroda (6) and Shuiman and Meyer (7) with chick chondrocytes, by Chiakulas (8) with frog chondrocytes and by Manning and Bonher (9) with human chondrocytes.Coon's (10) report on cloned chondrocytes both supports and conflicts with the above analysis. Coon made the important observation that many dedifferentiated chondrocytes which do not synthesize chondroitin sulfate in high density cultures, do yield progeny which synthesize the polysaccharide if cultured at low density. In addition, Coon and Calm (11) and Calm, Coon, and Calm (12) claim that a high molecular weight, heat-labile factor in embryo extract selectively inhibits the synthesis of chondroitin sulfate by chondrocytes and melanin by pigment ceils (see however, 13, 14). Medium containing this dedifferentiation factor was termed "nonpermissive"; medium Materials and MethodsIn general, culture procedures followed those recommended by Coon (10) and Cahn, Coon and Cahn (12).10-day vertebral cartilages were stripped of their adhering connective tissues (1) and incubated 2~'~ hr in a trypsin-collagenase mixture (0.1% trypsin, 1.4 mg/ml collagenase, 10% chick serum in calcium-magnesium-free Simm's balanced salt). After flushing the cartilages through a small bore pipette, the freshly liberated chondrocytes were counted in a hemocytometer and placed into 100 mm Petri dishes (Falcon tissue culture grade) at a density of 5 X 10 ~ cells/dish in 10 ml of medium. After 3-5 days, two distinct cellpopulations were apparent: a floating population and a population adherent to the substrate. The round, floating cells, referred to as "primary floaters," were found to be contaminated with less than 1% of nonchondrogenic ceils.Experiments were performed with cells derived from cultures initial...

Human chondrocytes in tridimensional culture

“…It is well established that chondrocytes, when liberated from their matrix by digestion and then cultivated in vitro in monolayers, usually rapidly lose their morphological and biochemical characteristics, assume a fibroblastic phenotype, and actively divide (1,2,8,10,40,47). When cultivated in a defined medium, in monolayers, rabbit The evolution of the PG amount in chondrocyte aggregates and in conditioned culture medium during cultivation can be explained as follows: During the first days of culture, chondrocytes progressively aggregated and constituted cellular masses ( -4-2 mm diameter), which before Day 9 were flaky but became solid and spherical during further cultivation (1 mo.).…”

Sulfate incorporation by articular chondrocytes in monolayer culture

“…and "dedifferentiated" (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13) . T h e usual measure of differentiation has been histochemical evidence of matrix production, particularly metachromasia, or autoradiographic demonstration of sulfate fixation.…”

Chondrogenic differentiation during midfacial development in the mouse: in vivo and in vitro studies

“…The morphogenetic phase is characterized by the migration and the condensation of mesenchymal cells under a strict genetic control carrying positional informations (Coelho and Kosher, 1991;Solursh, 1983;Francis-West et al, 1998). The differentiation of mesenchymal cells into chondrocytes takes place along a multistep pathway including condensation of mesenchymal cells followed by the differentiation of the condensed mesenchymal cells into chondrocytes, which surround themselves with an extracellular matrix mainly composed of type II collagen and aggrecan (Chacko et al, 1969). Furthermore, several transcription factors are known to be involved in early commitment of mesenchymal cells into the chondrogenic lineage.…”