Histological Features of the Nasal Passage in Juvenile Japanese White Rabbits

Rabbits are sometimes used for intranasal toxicology studies. We investigated the postnatal development of the nasal passage in juvenile Japanese white rabbits from just after birth to 6-week-old to provide information for conducting intranasal toxicological evaluation using juvenile animals. On postnatal day (PND) 1, the nasal passage consisted of the septum with mostly cartilaginous nasal wall and turbinates. The lining squamous, transitional, respiratory, and olfactory epithelia were already distributed similar to adults and were still underdeveloped. The nasal passage gradually expanded with age, as did the nasal wall, including the turbinates formed by endochondral ossification. The maxilloturbinate elongated, during which it branched complexly. The respiratory epithelium takes the form of columnar epithelium together with a reduction in goblet cells. In addition, the olfactory epithelium had clear cytoplasm in the ethmoturbinate, the olfactory nerve bundles thickened, and Bowman’s gland acini increased in size and number. Other tissues, including the vomeronasal organ, nasal-associated lymphoid tissue, and nasolacrimal duct, also developed histologically with age. This investigation characterized the postnatal histological development of the nasal passage in Japanese white rabbits, providing basic knowledge regarding the histological examination and rationale for appropriate study design of intranasal toxicology studies in juvenile rabbits.

Exogenous PTH 1-34 Attenuates Impaired Fracture Healing in Endogenous PTH Deficiency Mice via Activating Indian Hedgehog Signaling Pathway and Accelerating Endochondral Ossification

Fracture healing is a complicated, long-term, and multistage repair process. Intermittent administration of parathyroid hormone (PTH) has been proven effective on intramembranous and endochondral bone formation during the fracture healing process, however, the mechanism is unclear. In this study, we investigated the role of exogenous PTH and endogenous PTH deficiency in bone fracture healing and explored the mechanism by using PTH knockout (PTH-/-) mice and ATDC5 cells. In a mouse femur fracture model, endogenous PTH deficiency could delay endochondral ossification whereas exogenous PTH promotes accumulation of endochondral bone, accelerates cartilaginous callus conversion to bony callus, enhances maturity of bony callus, and attenuates impaired fracture healing resulting from endogenous PTH deficiency. In fracture callus tissue, endogenous PTH deficiency could inhibit chondrocyte proliferation and differentiation whereas exogenous PTH could activate the IHH signaling pathway to accelerate endochondral ossification and rescue impaired fracture healing resulting from endogenous PTH deficiency. In vitro, exogenous PTH promotes cell proliferation by activating IHH signaling pathway on ATDC5 cells. In mechanistic studies, by using ChIP and luciferase reporter assays, we showed that PTH could phosphorylate CREB, and subsequently bind to the promoter of IHH, causing the activation of IHH gene expression. Therefore, results from this study support the concept that exogenous PTH 1-34 attenuates impaired fracture healing in endogenous PTH deficiency mice via activating the IHH pathway and accelerating endochondral ossification. Hence, the investigation of the mechanism underlying the effects of PTH treatment on fracture repair might guide the exploration of effective therapeutic targets for fracture.

Construction of developmentally inspired periosteum-like tissue for bone regeneration

The periosteum, a highly vascularized thin tissue, has excellent osteogenic and bone regenerative abilities. The generation of periosteum-mimicking tissue has become a novel strategy for bone defect repair and regeneration, especially in critical-sized bone defects caused by trauma and bone tumor resection. Here, we utilized a bone morphogenetic protein-2 (BMP-2)-loaded scaffold to create periosteum-like tissue (PT) in vivo, mimicking the mesenchymal condensation during native long bone development. We found that BMP-2-induced endochondral ossification plays an indispensable role in the construction of PTs. Moreover, we confirmed that BMP-2-induced PTs exhibit a similar architecture to the periosteum and harbor abundant functional periosteum-like tissue-derived cells (PTDCs), blood vessels, and osteochondral progenitor cells. Interestingly, we found that the addition of chondroitin sulfate (CS), an essential component of the extracellular matrix (ECM), could further increase the abundance and enhance the function of recruited PTDCs from the PTs and finally increase the regenerative capacity of the PTs in autologous transplantation assays, even in old mice. This novel biomimetic strategy for generating PT through in vivo endochondral ossification deserves further clinical translation.

A multi-stem cell basis for craniosynostosis and calvarial mineralization

Craniosynostosis is a group of disorders of premature calvarial sutural fusion. An incomplete understanding of the calvarial stem cells (CSCs) that produce fusion-driving osteoblasts has limited the development of non-surgical therapeutic approaches for craniosynostosis. Here we show that both physiologic calvarial mineralization and pathologic calvarial fusion in craniosynostosis reflect the interaction of two separate stem cell lineages; a recently reported CathepsinK (CTSK) lineage CSC (CTSK+ CSC)1 and a separate Discoidin domain-containing receptor 2 (DDR2) lineage stem cell (DDR2+ CSC) identified in this study. Deletion of Twist1, a gene associated with human craniosynostosis2,3, solely in CTSK+ CSCs is sufficient to drive craniosynostosis, however the sites destined to fuse surprisingly display a marked depletion of CTSK+ CSCs and a corresponding expansion of DDR2+ CSCs. This DDR2+ CSC expansion is a direct maladaptive response to CTSK+ CSC depletion, as partial suture fusion occurred after genetic ablation of CTSK+ CSCs. This DDR2+ CSC is a specific fraction of DDR2+ lineage cells that displayed full stemness features, establishing the presence of two distinct stem cell lineages in the sutures, with each population contributing to physiologic calvarial mineralization. DDR2+ CSCs mediate a distinct form of endochondral ossification where an initial cartilage template is formed but the recruitment of hematopoietic marrow is absent. Direct implantation of DDR2+ CSCs into suture sites was sufficient to induce fusion, and this phenotype was prevented by co-transplantation of CTSK+ CSCs. Lastly, the human counterparts of DDR2+ CSCs and CTSK+ CSCs are present in calvarial surgical specimens and display conserved functional properties in xenograft assays. The interaction between these two stem cell populations provides a new biologic interface to modulate calvarial mineralization and suture patency.

Over 400 million years of cooperation: Untangling the chondrocranium-dermatocranium connection

The cranial endo- and dermal skeletons, which comprise the vertebrate skull, evolved independently and form separately during embryogenesis. In mammals, the mostly cartilaginous cranial endoskeleton forms prior to the bony dermatocranium. Many features of the chondrocranium are transient, undergoing endochondral ossification or disappearing, so its role in skull morphogenesis is not understood The fibroblast growth factor (FGF) and receptor (FGFR) signaling pathway contributes significantly to the regulation of osteochondroprogenitor cell function. Mutations in FGFR genes are associated with diseases that impact the skull including dwarfing chondrodyplasia and craniosynostosis syndromes. We investigate the developing chondrocranium and dermatocranium using a mouse model for craniosynostosis carrying a gain of function mutation in Fgfr2 to assess development of these cranial skeleton systems. Dermatocrania and chondrocrania of Fgfr2cC342Y/+ mice and their Fgfr2c+/+ littermates were quantified in 3D from microcomputed tomography images of mouse embryos. Chondrocrania of embryonic mice carrying the Fgfr2 mutation are larger than their Fgfr2c+/+ littermates and include novel extensions of cartilage over the lateral and dorsal aspect of the brain. Like the forming chondrocranium, the embryonic dermatocranium is larger in Fgfr2cC342Y/+ mice throughout embryogenesis but after disappearance of much of the chondrocranium, the dermatocranium becomes progressively smaller relative to Fgfr2c+/+ littermates during postnatal growth. Results reveal the direct effects of this Fgfr2c mutation on embryonic cranial cartilage, the impact of chondrocranial structure on developing dermatocranial elements, the importance of the chondrocranium in decoding the impact of specific genetic variants on head morphogenesis, and the potential for harnessing these effects as therapeutic targets.

Bone Biomarkers in Mucopolysaccharidoses

The accumulation of glycosaminoglycans (GAGs) in bone and cartilage leads to progressive damage in cartilage that, in turn, reduces bone growth by the destruction of the growth plate, incomplete ossification, and growth imbalance. The mechanisms of pathophysiology related to bone metabolism in mucopolysaccharidoses (MPS) include impaired chondrocyte function and the failure of endochondral ossification, which leads to the release of inflammatory cytokines via the activation of Toll-like receptors by GAGs. Although improvements in the daily living of patients with MPS have been achieved with enzyme replacement, treatment for the bone disorder is limited. There is an increasing need to identify biomarkers related to bone and cartilage to evaluate the progressive status and to monitor the treatment of MPS. Recently, new analysis methods, such as proteomic analysis, have identified new biomarkers in MPS. This review summarizes advances in clinical bone metabolism and bone biomarkers.