Regulation of Childhood Growth
- Jeffrey Baron, MD, Head, Section on Growth and Development
- Kevin Barnes, PhD, Senior Research Assistant
- Julian Lui, PhD, Staff Scientist
- Youn Hee Jee, MD, Staff Clinician
- Joanna Courtis, BS, Postbaccalaureate Fellow
- Benjamin Hauser, BS, Postbaccalaureate Fellow
- Jacob Wagner, BS, Postbaccalaureate Fellow
- Elaine Zhou, BA, Postbaccalaureate Fellow
- Wei Wang, MSc, Biologist
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 this 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 increased 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. Genome-wide association studies and molecular studies of growth-plate biology suggest that there are hundreds of genes that control linear growth. Therefore, there are likely many genetic causes of linear growth disorders that remain to be discovered.
To discover new genetic causes of childhood growth disorders, we evaluated families with monogenic growth disorders using SNP (single-nucleotide polymorphism) arrays to detect deletions, duplications, mosaicism, and uniparental disomy, combined with exome sequencing to detect single-nucleotide variants and small insertions/deletions in coding regions and splice sites. When sequence variants that are likely to cause the disorder are identified, the variants and the genes in which they occur are 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 ACAN, the gene encoding aggrecan, 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 recently 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 (Figure 2). We found that the disorder was caused by a de novo mutation in SP7. SP7 encodes a transcription factor required for differentiation of osteoblasts, a cell type required for bone formation. We generated mice with a variant orthologous to that of our subject and found that the mice showed a similar complex skeletal phenotype, confirming that the variant was responsible for the disorder. The mutation shifted the DNA–binding specificity of SP7 from AT–rich motifs to a GC–consensus sequence (typical of other SP family members), resulting in an aberrant gene-expression profile and abnormal osteoblast differentiation. Thus, our study identifies a novel pathogenic mechanism in which a neomorphic mutation in a transcription factor shifts DNA–binding specificity to cause disease.
Figure 2.
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A child with a complex skeletal dysplasia caused by a neomorphic variant in SP7
We also studied disorders that causes excessive growth. We evaluated a patient with Weaver syndrome, a condition that includes overgrowth of many tissues, including the skeleton (Figure 3). The syndrome is caused by variants in EZH2. EZH2 encodes a histone methyltransferase that catalyzes the trimethylation of histone H3 at lysine 27 (H3K27), which serves as an epigenetic signal for chromatin condensation and transcriptional repression. We created a mouse model that showed mild overgrowth, recapitulating the Weaver phenotype. We found that the EZH2 variants responsible for Weaver syndrome cause a partial loss of enzymatic function. Interestingly, we also found that a more severe genetic lesion, loss of both EZH1 and EZH2, impairs bone growth in mice, and we explored the molecular mechanisms involved.
Figure 3.
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Growth chart of a child with Weaver syndrome
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 transdifferentiation 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 (Figure 4).
Figure 4.
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The pathogenic SP7 variant produces an aberrant gene expression profile when expressed in mesenchymal stem cells.
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, and which could possibly be used to treat such disorders. Targeting these 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. As a first test of this approach, we created fusion proteins combining these cartilage-targeting antibody fragments with IGF-1, an endocrine/paracrine factor that positively regulates chondrogenesis. The fusion proteins retained both cartilage binding and IGF-1 biological activity, and they were able to stimulate bone growth in an organ culture system. Using a GH–deficient mouse model, we found that subcutaneous injections of the 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 IGF-1 alone (Figure 5). Our findings provide proof of principle that targeting therapeutics to growth-plate cartilage can potentially improve treatment for childhood growth disorders.
Figure 5.
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Targeted IGF-1 treatment in growth hormone–deficient mice increases growth-plate height without increasing proliferation in the kidney.
We are currently working to optimize the efficacy of targeted IGF1 therapy. In other studies, we are using cartilage-tethering antibody fragments to target other chondrogenic endocrine and paracrine factors to the growth plate. We are exploring the utility of this 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, 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.
- Jee YH, Gangat M, Yeliosof O, Temnycky AG, Vanapruks S, Whalen P, Gourgari E, Bleach C, Yu CH, Marshall I, Yanovski JA, Link K, Ten S, Baron J, Radovick S. Evidence that the etiology of congenital hypopituitarism has a major genetic component but is infrequently monogenic. Front Genet 2021 12:697549.
- Jee YH, Won S, Lui JC, Jennings M, Whalen P, Yue S, Temnycky AG, Barnes KM, Cheetham T, Boden MG, Radovick S, Quinton R, Leschek EW, Aguilera G, Yanovski JA, Seminara SB, Crowley WF, Delaney A, Roche KW, Baron J. DLG2 variants in patients with pubertal disorders. Genet Med 2020 22:1329–1337.
- 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 2021 12:660731.
Collaborators
- Greti Aguilera, MD, Scientist Emeritus, NICHD, Bethesda, MD
- Angela Delaney Freedman, MD, St. Jude Children’s Research Hospital, Memphis, TN
- Lijin Dong, PhD, Genetic Engineering Core, NEI, Bethesda, MD
- Gabriele Häusler, MD, Medizinische Universität Wien, Vienna, Austria
- Ellen Leschek, MD, Division of Diabetes, Endocrinology, and Metabolic Diseases, NIDDK, Bethesda, MD
- Ola Nilsson, MD, PhD, Karolinska Institute, Stockholm, Sweden
- Sally Radovick, MD, Rutgers Biomedical and Health Sciences, Robert Wood Johnson Medical School, New Brunswick, NJ
- Gudrun Rappold, PhD, University of Heidelberg, Heidelberg, Germany
- Katherine W. Roche, PhD, Receptor Biology Section, NINDS, Bethesda, MD
- Jack Yanovski, MD, PhD, Section on Growth and Obesity, NICHD, Bethesda, MD
Contact
For more information, email jeffrey.baron@nih.gov or visit https://baron.nichd.nih.gov.
