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The growth kinetics of subcultured human synovial fibroblasts from 16 patients with inflammatory and noninflammatory arthropathies were studied in antibiotic free media. The experimental design allowed a clear distinction between the growth rates and final saturation densities achieved. The effects of refeeding and of the serum concentration were evaluated for each line. Inflammatory lines achieved significantly higher final saturation densities and growth rates than noninflammatory lines for most protocols, but the differences between rheumatoid and nonrheumatoid groups were less marked. Inflammatory fibroblasts demonstrated a greater independence to nutritional and growth stimulatory factors in their microenvironment than noninflammatory fibroblasts.The modulation of fibroblastic proliferation in culture is under complex control, and many of the factors involved are incompletely understood (1). Nevertheless, untransformed fibroblastic cells often exhibit greater or lesser degrees of density-dependent inhibition of growth (DDI). One measurement of DDI is reflected by the final cell saturation density (FSD) achieved by the fibroblastic cells under defined conditions of refeeding the cultures and of serum supplementation (2,3).The precise mechanisms involved in DDI are uncertain, but one possibility is that it is effected by the actual contact between cells. A mechanism for this contact inhibition of growth based on mutual modification of surface carbohydrate moieties by surface glycosyltransferases (4) has received both considerable support (5,6) and criticism (7,8).However, the FSD achieved by fibroblastic cells can be affected by different sets of circumstances in the microenvironment of the culture other than those affecting the growth rate of the cells. For example, the authors had previously demonstrated that although a change in culture pH of human skin fibroblasts does not affect growth rates, the FSD at the higher pH values was always greater than that achieved at lower pH at any serum concentration between 1% and 20% (9). Nutritional effects, operative mainly through diffusion gradients of the larger molecules in the extracellular microenvironment (3,10), may also modulate DDI under certain circumstances. However, we have demonstrated that such diffusion effects cannot entirely account for the phenomenon of DDI under various conditions of nutrient availability and serum supplementation (10).Previously published work on the growth of human rheumatoid and nonrheumatoid synovial cells in

Glycosaminoglycan (GAG) synthesis and glucose uptake by rheumatoid (R) and nonrheumatoid (NR) synovial cells were studied at the second subculture during four different sets of nutritional conditions and at sequential, defined intervals of the growth cycle. Synovial fibroblastic cells in monolayer cultures secrete both hyaluronic acid and sulfated GAGs in a ratio of about 8 : I. With increasing cell density the ability to sulfate GAGs appears to decrease. No significant differences in GAG synthesis between rheumatoid and nonrheumatoid lines on a per cell basis could be detected during any interval in the growth cycle. Similarly, no significant differences of glucose uptake per cell could be demonstrated between rheumatoid and nonrheumatoid lines under the various protocols applied throughout the growth cycle. The most important factor determining glucose uptake per cell is the availability of glucose in the medium which is in turn closely correlated with cell number.In a recent publication (1) we had demonstrated that synovial fibroblastic cells from inflammatory ar- thropathies in culture achieved higher final saturation densities and growth rates than synovial fibroblasts from noninflammatory arthropathies. These differences were significantly modified by nutritional factors in the microenvironment and were not specific for rheumatoid arthritis. Other studies have suggested that there are a number of persistent differences between rheumatoid and nonrheumatoid cells in long-term culture ( 2 4 which imply that there may be a specific or fundamental alteration in the rheumatoid fibroblastic cell. These metabolic alterations have included increased hyaluronic acid synthesis and glucose uptake by the rheumatoid cells, compared to controls.However, the published studies on the synthesis of glycosaminoglycans (GAG) by synovial fibroblastic cells in culture have not included a rigorous investigation of the nutritional effects of the cellular microenvironment or of the effects of cell growth. In some studies ( 3 3 ) concerned with hyaluronic acid (HA) secretion, methods have been used that would quantitate only the total GAG secreted by the synovial cells. Furthermore, while some authors have indicated that chondroitin sulfate (CS) is not released by synovial fibroblasts (2), other workers have identified CS in media from synovial cell cultures (6).In this study we have examined the GAG secretion of human synovial cells by a method (7) that allows clear separation of HA from CS. Experiments were also designed that take into account the nutritional effects of the microenvironment and the growth kinetics of the cells (1,8,9). This methodology has been extended to a number of fibroblastic cell lines derived from synovia of patients with inflammatory and noninflammatory arthropathies.

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