Multiple Phases of Chondrocyte Enlargement Underlie Differences in Skeletal Proportions

Even a casual pass through the great halls of mammals in the world’s natural history museums highlights the wide diversity of skeletal proportions that allow us to distinguish between species even when reduced to their calcified components. Similarly each individual is comprised of a variety of bones of differing lengths. The largest contribution to the lengthening of a skeletal element, and to the differential elongation of elements, comes from a dramatic increase in the volume of hypertrophic chondrocytes in the growth plate as they undergo terminal differentiation(1–7). Despite this recognized importance, the mechanisms of chondrocyte volume enlargement have remained a mystery(8–11). Here we use quantitative phase microscopy(12) to show that chondrocytes undergo three distinct phases of volume increase, including a phase of massive cell swelling in which the cellular dry mass is significantly diluted. In light of the tight fluid regulatory mechanisms known to control volume in many cell types(13), this stands as a remarkable mechanism for increasing cell size and regulating growth rate. It is, however, the duration of the final phase of volume enlargement by proportional dry mass increase at low density that varies most between rapidly and slowly elongating growth plates. Moreover, we find that this third phase is locally regulated through an Insulin-like Growth Factor-dependent mechanism. This study provides a framework for understanding how skeletal size is regulated and for exploring how cells sense, modify, and establish a volume set point.