Parathyroid Hormone Receptor Type 1/Indian Hedgehog Expression Is Preserved in the Growth Plate of Human Fetuses Affected with Fibroblast Growth Factor Receptor Type 3 Activating Mutations

Abstract

The fibroblast growth factor receptor type 3 (FGFR3) and Indian hedgehog (IHH)/parathyroid hormone (PTH)/PTH-related peptide receptor type 1 (PTHR1) systems are both essential regulators of endochondral ossification. Based on mouse models, activation of the FGFR3 system is suggested to regulate the IHH/PTHR1 pathway. To challenge this possible interaction in humans, we analyzed the femoral growth plates from fetuses carrying activating FGFR3 mutations (9 achondroplasia, 21 and 8 thanatophoric dysplasia types 1 and 2, respectively) and 14 age-matched controls by histological techniques and in situ hybridization using riboprobes for human IHH, PTHR1, type 10 and type 1 collagen transcripts. We show that bone-perichondrial ring enlargement and growth plate increased vascularization in FGFR3-mutated fetuses correlate with the phenotypic severity of the disease. PTHR1 and IHH expression in growth plates, bone-perichondrial rings and vascular canals is not affected by FGFR3 mutations, irrespective of the mutant genotype and age, and is in keeping with cell phenotypes. These results indicate that in humans , FGFR3 signaling does not downregulate the main players of the IHH/PTHR1 pathway. Endochondral bone ossification is a highly regulated process involved in the formation and growth of cartilaginous templates of long bones, vertebrae, and some craniofacial bones. It requires the linear phenotypic progression of undifferentiated mesenchymal cells into hypertrophic chondrocytes and the subsequent replacement of mineralized cartilage matrix by bone.1-4 As part of this process, resting chondrocytes at the growth plate proliferate, then mature and differentiate into hypertrophic chondrocytes and finally undergo apoptosis after mineralization of the surrounding extracellular matrix. Lacunae formed in this way in the mineralized matrix are invaded by blood vessels and bone cells. At each stage, chondrocytes express specific markers including type 2 collagen in the resting, proliferative, and prehypertrophic zones, and type 10 collagen when hypertrophic.In addition to endochondral ossification, development and growth of bones formed via a cartilage model involve a distinct pattern of bone formation, namely perichondrial bone formation, 2,3,5 a process whereby bone is formed outside and around the cartilage anlage. When ossification starts, perichondrial ossification generates the primitive cortical bone (or bone collar) and its associated periosteum, and the bone-perichondrial ring that develops around the growth plate at the cartilage/bone junction. Bone-perichondrial ring (also called ossification groove of Ranvier) 5,6 comprises the bony ring (also called ring of Lacroix) 5,6 covered on the periosteal side by a loose connective tissue that acts as a germinative (cambium) layer, assuring growth in width. Its growth in length is insured by a germinative zone localized at the epiphyseal tip of the ring.Two main systems, namely the fibroblast growth factor receptor type 3 (FGFR3) and the Indian hedgehog (IHH)/

Development of the Endochondral Skeleton

“…Interestingly, in Fgfr3 -/ -mice, IHH expression and signaling in chondrocytes was increased relative to wild-type mice, whereas in the mice overexpressing activated FGFR3, IHH expression and signaling was inhibited (Naski et al 1998;Chen et al 1999;Li et al 1999). In contrast to phenotypes observed in mice, IHH and PTHR1 expression were not altered in human achondroplasia or thanatophoric dysplasia growth plate tissue (Cormier et al 2002). Thus, FGFR3 signaling may inhibit chondrocyte proliferation and hypertrophy in part through indirect modulation of the IHH/PTHrP axis, but additional mechanisms may also mediate the pathological consequences of FGFR3 activating mutations.…”

Sixteen years and counting: The current understanding of fibroblast growth factor receptor 3 (FGFR3) signaling in skeletal dysplasias

“…Similar phenotypes were found in the growth plates of S365C , G369C (corresponding to human TDI mutation p.G370C), or K644E‐Fgfr3 murine models for TD, involving, in addition to Ihh , the PTHrP receptor [Chen et al, 1999; 2001; Li et al, 1999]. Although these findings are at variance with the normal expression of Ihh and PTHrP , reported by others in human ACH and TD growth plates or those of the K644E‐Fgfr3 mouse [Cormier et al, 2002; Iwata et al, 2000], they open the possibility that inhibition of Ihh/PTHrP signaling contributes to disrupted chondrocyte differentiation, observed in TD.…”

Cell responses to FGFR3 signalling: growth, differentiation and apoptosis

“…Stem cells, which will differentiate into osteoblasts, are brought by blood vessels invading the cartilage after apoptosis of hypertrophic chondrocytes. Interestingly, the growth plates of ACH transgenic mice and human TD foetuses were shown to have abnormal vascularisation and this seems to correlate with the severity of the disease [54,68]. Moreover, it has been reported recently [69] that Fgfr3À/À mice show a down-regulation of vascular endothelial growth factor (VEGF), an angiogenic factor produced by hypertrophic chondrocytes that is required for their apoptosis [70].…”