Regulation of Childhood Growth
- Jeffrey Baron, MD, Head, Section on Growth and Development
- Lesley Brown, PhD, Staff Scientist
- Julian Lui, PhD, Staff Scientist
- Krishma Tailor, PhD, Biologist
- Wei Wang, MSc, Biologist
- Kirtal Hansdah, PhD, Visiting Fellow
- Joanna Courtis, BS, Postbaccalaureate Fellow
- Fatima Elzamzami, BS, Postbaccalaureate Fellow
- Connor Sisk, BS, Postbaccalaureate Fellow
- Jacob Wagner, BS, Postbaccalaureate Fellow
Children grow taller because their bones grow longer. Bone elongation occurs at the growth plate, a thin layer of cartilage found near the ends of juvenile bones (Figure 1). In the growth plates, new cartilage is produced through chondrocyte proliferation, hypertrophy, and cartilage matrix synthesis, and then the newly formed cartilage is remodeled into bone. The process, termed endochondral ossification, results in bone elongation, which causes children to grow in height (linear growth).
We investigate the cellular and molecular mechanisms governing childhood growth and development. We focus particularly on growth at the growth plate, which drives bone elongation and therefore determines height. One goal of this work is to gain insight into the many human genetic disorders that cause childhood growth failure or overgrowth. A second goal is to develop new treatments for children with severe growth disorders.
Figure 1.
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Histological image of a growth plate, showing the three principal zones
Novel genetic causes of childhood growth disorders
Mutations in genes that regulate growth-plate chondrogenesis cause abnormal bone growth and short stature in children. Depending on the severity and nature of the genetic abnormality, the phenotype can range from chondrodysplasias with short, malformed bones, to severe, often disproportionate, short stature, to mild proportionate short stature. If the genetic defect affects tissues other than the growth plate cartilage, the child may present with a more complex syndrome that includes other clinical abnormalities. Less commonly, mutations in these genes cause excessive growth-plate chondrogenesis and therefore abnormally tall stature. Often the increased proliferation occurs in many tissues, producing a generalized overgrowth syndrome, which can include other medical problems such as developmental delay and elevated cancer risk.
For many children who are brought to medical attention for linear growth disorders, clinical, laboratory, and genetic evaluation fails to identify the underlying etiology. To discover new genetic causes of childhood growth disorders, we evaluated families with monogenic growth disorders using exome sequencing. When sequence variants that are likely to cause the disorder were identified, the variants and the genes in which they occur were studied in the laboratory to confirm that the variant is pathogenic, to elucidate the pathogenesis of the disorder, and to explore the role of the gene in normal growth.
Using this approach, we identified new causes of childhood growth disorders. We previously found that variants in QRICH1, a gene of unknown function, cause a chondrodysplasia attributable to impaired growth-plate chondrocyte hypertrophic differentiation. We also found evidence that heterozygous deletion of CYP26A1 and CYP26C1, which encode enzymes that metabolize retinoic acid (RA), cause elevated RA concentrations, which accelerate bone and dental maturation in humans and cause developmental defects involving the eye and central nervous system. Our group also discovered that variants in aggrecan (ACAN), a component of cartilage extracellular matrix, cause autosomal-dominant short stature with advanced skeletal maturation and that such patients tend to develop early-onset osteoarthritis.
We also studied a child with a complex skeletal dysplasia who presented with severe scoliosis, thickened calvarium, craniosynostosis, osteosclerosis of the clavicles and spine, and recurring fractures in the lower extremities. We discovered that the disorder was caused by a de novo, neomorphic mutation in SP7, which encodes a transcription factor required for differentiation of osteoblasts.
Recently, we have focused on disorders that cause excessive growth. We evaluated a patient with Weaver syndrome, an overgrowth disorder caused by variants in EZH2. EZH2 encodes a histone methyltransferase and thereby acts as an epigenetic writer. We found that the EZH2 variants responsible for Weaver syndrome cause a partial loss of enzymatic function.
Figure 2.
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Growth chart of a child with overgrowth caused by a variant in SPIN4
We also studied a child with generalized overgrowth of prenatal onset (Figure 2). Exome sequencing identified a hemizygous frameshift variant in Spindlin 4 (SPIN4), with X-linked inheritance. Ablation of Spin4 in mice recapitulated the human phenotype with generalized overgrowth, including increased longitudinal bone growth (Figure 3). We found evidence that SPIN4 binds specific histone modifications (Figure 4), promotes canonical WNT signaling, and inhibits cell proliferation in vitro and that the identified frameshift variant had lost all these functions. Growth-plate analysis revealed increased cell proliferation in the proliferative zone and an elevated number of progenitor chondrocytes in the resting zone. We also found evidence of decreased canonical Wnt signaling in growth-plate chondrocytes, providing a potential explanation for the elevated number of resting-zone chondrocytes. Taken together, our findings provide strong evidence that SPIN4 is an epigenetic reader that negatively regulates mammalian body growth, and that loss of SPIN4 causes an overgrowth syndrome in humans, expanding our knowledge of the epigenetic regulation of human growth.
Figure 3.
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Body weight (left) and sizes of several organs (right) are increased in mice lacking SPIN4.
Figure 4. Spin4 binds to specific modified histones.
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Histone peptide arrays were used to assess histone-binding properties of wild-type (WT) and mutant (Mut) SPIN4. Each spot on the array contains a peptide portion of a histone that has undergone specific post-translational modifications. Darker color indicates greater binding of the indicated Spin protein to that modified peptide. WT SPIN4 binds to specific modified histone peptides, while Mut SPIN4 showed substantially diminished binding. The box-and-whisker plot shows these data quantitatively.
Molecular and cellular mechanisms by which specific genes and pathways regulate childhood growth
Our group also studies the fundamental mechanisms governing childhood growth. Much of our work has focused on the growth plate. Growth at the growth plate is controlled by several interacting regulatory systems, involving endocrine, paracrine, extracellular matrix–related, and intracellular pathways. Previously, our group studied growth-plate regulation by FGFs, BMPs, C-type natriuretic peptide, retinoids, WNTs, PTHrP/IHH, IGFs, estrogens, glucocorticoids, and microRNAs. We also found evidence that SOX9, a transcription factor, regulates the trans-differentiation of growth-plate chondrocytes into osteoblasts.
We also investigated the mechanisms that cause bone growth to occur rapidly in early life but then to slow progressively with age and eventually cease. We found evidence that the developmental program responsible for the decline in growth-plate function plays out more slowly in larger bones than in smaller bones and that such differential aging contributes to the disparities in bone length and therefore to establishing normal mammalian skeletal proportions.
New treatment approaches for growth-plate disorders
Recombinant human growth hormone (GH) is commonly used to treat short stature in children. However, GH treatment has limited efficacy, particularly in severe, non-GH–deficient conditions such as chondrodysplasias, and has off-target effects. Systemic insulin-like growth factor-1 (IGF-1) treatment has similar deficiencies. There are many endocrine and paracrine factors that promote chondrogenesis at the growth plate, which could potentially be used to treat these disorders. Targeting such growth factors specifically to the growth plate might augment the therapeutic skeletal effect while diminishing undesirable effects on non-target tissues. To develop growth plate–targeted therapy, we previously used yeast display to identify single-chain human antibody fragments that bind to cartilage with high affinity and specificity. We then created fusion proteins combining these cartilage-targeting antibody fragments with IGF-1, an endocrine/paracrine factor that positively regulates chondrogenesis. Using a GH–deficient mouse model, we found that subcutaneous injections of such fusion proteins increased growth plate height without increasing proliferation in kidney cortical cells, demonstrating greater on-target efficacy at the growth plate and less off-target effect on the kidney than with IGF-1 alone. Our findings provide proof of principle that targeting therapeutics to growth-plate cartilage can potentially improve treatment for childhood growth disorders.
We are currently working to optimize the efficacy of targeted IGF1 therapy. In other studies, we are applying this approach to target other chondrogenic endocrine and paracrine factors to the growth plate. We are exploring the utility of the approach both to stimulate growth-plate chondrogenesis non-specifically and also to reverse specific genetic defects in growth-plate function by modulating the abnormal molecular pathway responsible for the growth failure.
Publications
- Lui JC, Wagner J, Zhou E, Dong L, Barnes KM, Jee YH, Baron J. Loss of function variant in SPIN4 causes an X-linked overgrowth syndrome. JCI Insight 2023 8:8(9):e167074.
- Lui JC, Raimann A, Hojo H, Dong L, Roschger P, Kikani B, Wintergerst U, Fratzl-Zelman N, Jee YH, Haeusler G, Baron J. A neomorphic variant in the transcription factor SP7 alters sequence specificity and causes a high-turnover bone disorder. Nat Commun 2022 13:700.
- Lui JC, Baron J. Epigenetic causes of overgrowth syndromes. J Clin Endocrinol Metab 2023 online ahead of print.
- Lui JC, Baron J. CNP-related short and tall stature: a close-knit family of growth disorders. J Endocr Soc 2022 6:bvac064.
- Weiss B, Eberle B, Roeth R, de Bruin C, Lui JC, Paramasivam N, Hinderhofer K, van Duyvenvoorde HA, Baron J, Wit JM, Rappold GA. Evidence that non-syndromic familial tall stature has an oligogenic origin including ciliary genes. Front Endocrinol (Lausanne) 2021 12:660731.
Collaborators
- Lijin Dong, PhD, Genetic Engineering Core, NEI, Bethesda, MD
- Ola Nilsson, MD, PhD, Karolinska Institute, Stockholm, Sweden
- Gudrun Rappold, PhD, University of Heidelberg, Heidelberg, Germany
Contact
For more information, email jeffrey.baron@nih.gov or visit https://www.nichd.nih.gov/research/atNICHD/Investigators/baron.
