A transcriptional atlas of the pubertal human growth plate reveals two populations of stem cells and direct effect of growth hormone

The cartilaginous growth plate is a critical organ responsible for longitudinal bone growth. It closes after puberty in humans but remains open throughout life in mice. Although cartilage stem cells have been identified in murine growth plates, their existence in humans and their regulation by growth hormone (GH), the most widely used therapy for growth retardation, remain unknown. Here, we characterized the cellular and molecular organization of early pubertal human growth plates using unique surgical specimens from growth-restricting procedures and examined their direct responsiveness to GH. Single-cell and spatial analyses revealed two distinct stemlike populations in the resting zone, differing in proliferative activity, molecular identity, and regulatory cues. The root stem cells express multiple skeletal stem cell markers but not parathyroid hormone–related peptide and reside in a specialized microenvironment low in WNT and TGF-β growth factors. A similar population was identified in transcriptionally profiled unsorted murine growth plates, and clonal lineage tracing demonstrated that these root cells, marked by expression of the Prrx1 gene, generate extensive chondrocyte clones and differentiate into stromal and osteoblastic lineages, confirming their stem cell properties. Human growth plate explant cultures showed that GH directly activates JAK/STAT, TGF-β, and ERK intracellular signaling pathways, inhibits AKT signaling, and stimulates cartilage growth and proliferation of cartilage stem cells and chondrocytes in the proliferative zone. Together, these findings uncover a conserved dual stem cell organization in human and mouse growth plates and define direct mechanisms of GH action, providing a framework for optimizing growth-promoting therapies.