Location
Philadelphia, PA
Start Date
17-4-2026 1:30 PM
End Date
17-4-2026 2:00 PM
Description
Introduction: Growth plates, or epiphyseal plates, are cartilaginous regions at the end of long bones that allow skeletal elongation to occur. The epiphyseal plate can be divided into regions based on what the cells are doing in that region: the reserve zone where cells are stored, the proliferative zone where cells divide, the hypertrophic zone where cells enlarge, and the ossification zone where mature bone forms. Surrounding the growth plate is the perichondrium which forms the bone collar.
In the third metatarsal (MT3) only the distal end contains a growth plate. We used RNA sequencing to compare the growth plate forming and non-forming ends of the mouse MT3 to identify differentially expressed genes (DEGs) associated with growth plate formation. We compared the overlap of DEGs at postnatal day (P) 0, P4 and P9. Two genes that were upregulated during growth plate formation included Paired Related Homeobox Gene (Prrx1) and Fibroblast Growth Factor 10 (Fgf10).
Prrx1 is expressed in cardiac, skeletal, and smooth muscle tissue is involved in epithelial to mesenchymal transitions. Inactivation of Prrx1 leads to defects in the development of limbs, skull, mandible, and ear ossicles. Fgf10 is known to initiate limb development and is critical in the transition of epithelial to mesenchymal tissue, a key step in vertebrate limb formation. In the absence of Fgf10, mice fail to form limbs.
Objective: To further define the role of Prrx1 and Fgf10 during bone formation, we perform in situ hybridization to compare their spatial expression patterns in the growth plate forming and non-forming ends of neonatal MTs.
Methods: Histological specimens consisted of hind limbs collected at P0, P4, and P9 from FVB/NJ mice. Samples were prepared using fixation in an RNase-free preparation of 4% paraformaldehyde and decalcified in an RNase-free preparation of Morse’s solution (22.5% formic acid and 10% sodium citrate) for 24 hours. Limbs were embedded in paraffin following standard procedures. We performed in situ hybridization (ISH) on paraffin-embedded sections using RNAscope™ 2.5 HD Assay-RED (ACD Bio) for Fgf10 and Prrx1 following RNAscope™ standard protocol, except replacing standard antigen retrieval processes with the use of an RNAscope™ proprietary antigen retrieval enzyme for bone and cartilage tissue.
Results: RNA-seq demonstrated significant Prrx1 expression at the distal end of MT3 from P0-P9. In situ hybridization showed strong Prrx1 staining in the distal perichondrium across all ages. Staining was strongest in the bone collar region and continued to extend into the developing periosteum. While Fgf10 was significantly associated with growth plates by RNA-seq, staining was difficult to observe using in situ hybridization.
Conclusion: These findings support the role of Prrx1in mesenchymal transition, contributing to osteoblast differentiation and suggesting a role in long bone formation. Though not as prominent, Fgf10 is supported to have a role in skeletal development but with much lower expression levels.
Embargo Period
6-3-2027
Prrx1 and Fgf10 expression in the growth plate of developing metatarsals
Philadelphia, PA
Introduction: Growth plates, or epiphyseal plates, are cartilaginous regions at the end of long bones that allow skeletal elongation to occur. The epiphyseal plate can be divided into regions based on what the cells are doing in that region: the reserve zone where cells are stored, the proliferative zone where cells divide, the hypertrophic zone where cells enlarge, and the ossification zone where mature bone forms. Surrounding the growth plate is the perichondrium which forms the bone collar.
In the third metatarsal (MT3) only the distal end contains a growth plate. We used RNA sequencing to compare the growth plate forming and non-forming ends of the mouse MT3 to identify differentially expressed genes (DEGs) associated with growth plate formation. We compared the overlap of DEGs at postnatal day (P) 0, P4 and P9. Two genes that were upregulated during growth plate formation included Paired Related Homeobox Gene (Prrx1) and Fibroblast Growth Factor 10 (Fgf10).
Prrx1 is expressed in cardiac, skeletal, and smooth muscle tissue is involved in epithelial to mesenchymal transitions. Inactivation of Prrx1 leads to defects in the development of limbs, skull, mandible, and ear ossicles. Fgf10 is known to initiate limb development and is critical in the transition of epithelial to mesenchymal tissue, a key step in vertebrate limb formation. In the absence of Fgf10, mice fail to form limbs.
Objective: To further define the role of Prrx1 and Fgf10 during bone formation, we perform in situ hybridization to compare their spatial expression patterns in the growth plate forming and non-forming ends of neonatal MTs.
Methods: Histological specimens consisted of hind limbs collected at P0, P4, and P9 from FVB/NJ mice. Samples were prepared using fixation in an RNase-free preparation of 4% paraformaldehyde and decalcified in an RNase-free preparation of Morse’s solution (22.5% formic acid and 10% sodium citrate) for 24 hours. Limbs were embedded in paraffin following standard procedures. We performed in situ hybridization (ISH) on paraffin-embedded sections using RNAscope™ 2.5 HD Assay-RED (ACD Bio) for Fgf10 and Prrx1 following RNAscope™ standard protocol, except replacing standard antigen retrieval processes with the use of an RNAscope™ proprietary antigen retrieval enzyme for bone and cartilage tissue.
Results: RNA-seq demonstrated significant Prrx1 expression at the distal end of MT3 from P0-P9. In situ hybridization showed strong Prrx1 staining in the distal perichondrium across all ages. Staining was strongest in the bone collar region and continued to extend into the developing periosteum. While Fgf10 was significantly associated with growth plates by RNA-seq, staining was difficult to observe using in situ hybridization.
Conclusion: These findings support the role of Prrx1in mesenchymal transition, contributing to osteoblast differentiation and suggesting a role in long bone formation. Though not as prominent, Fgf10 is supported to have a role in skeletal development but with much lower expression levels.