Human meniscal proteoglycan metabolism in long-term tissue culture

Abstract

For the purpose of human meniscal allografting, menisci have been maintained viable in in vitro culture. The influence of long-term tissue culture on the extracellular matrix metabolism of the meniscus has been studied. Fetal calf serum (FCS) was used as a supplement for the growth factors necessary to maintain optimal meniscal cell metabolism. A series of semilunar cartilage samples was cultured under serum-free conditions since foreign proteins could be responsible for immunological problems after eventual allografting. The proteoglycan metabolism in human menisci cultured in FCS-supplemented and in serum-free culture media was compared. To rule out any influence of topographical variations in glycosaminoglycan (GAG) content on proteoglycan (PG) metabolism, GAG concentrations within the tissue were determined, and sulphate (³⁵S) incorporation was studied in tissue samples with a comparable biochemical composition. Sulphate incorporation was preserved when 20% FCS was added to the nutrient medium. The meniscal tissue fibroblasts continued to produce³⁵S-PG during 4 weeks of culture. The PG molecules were shown to consist of PG-aggregates, monomers and a low molecular-weight PG population. Newly synthesized GAG consisted of approximately 55% chondroitin 4- and 6-sulphate and 33% dermatan sulphate. In the presence of serum.³⁵S incorporation in PG and in the PG-aggregate fraction significantly increased during the first 2 weeks and then decreased during the following 2 weeks of in vitro culture. Newly synthesized PG-aggregates were almost entirely accumulated in the tissue during these weeks. In the 3rd week the values for this parameter decreased slightly.³⁵S-PG synthesis dramatically declined after 4 weeks of in vitro culture. Catabolism probably resulted in increased proportions of³⁵S-PG in the incubation media. In the absence of serum,³⁵S-PG production also increased in the 2nd week of culture. However,³⁵S activity was almost exclusively found in small PG, and this material apparently diffused to the incubation media. Consequently, catabolism is higher, and the immobilization of³⁵S-PG is poor when FCS is not added to the culture media. Our findings suggest that menisci are maintained in viable condition and may serve for allografting at least during 2 weeks of tissue culture.