Chapter 12 Micromass Cultures of Limb and Other Mesenchyme

Abstract

This chapter discusses the micromass cultures of limb and other mesenchyme. The micromass culture technique was initially developed to study chondrogenesis and consists of high-density dot cultures of dissociated limb bud cells. The development of the micromass culture technique allowed a 20-to-25-fold increase in the number of replicate cultures that could be established from the same number of isolated limb bud cells. The high-density cultures form a three-dimensional, multilayered organization of mesenchyme with close cell associations and entrap nascent pericellular and extracellular matrix molecules (ECM) to potentiate differentiation. The sequence of chondrogenic events within micromass cultures recapitulates the sequence observed in vivo. In the limb, the first appearance of pattern is the formation of cellular aggregations or condensations in the core of the limb. The aggregates continue to enlarge mainly by assimilating surrounding mesenchyme followed by chondrogenic differentiation to form the cartilage template, establishing the basic pattern of the limb.