Mechanisms of bone dynamics during fingertip regeneration in axolotl, mouse and human.
Final Report Abstract
The regeneration of extremities is studied in animal models that have the ability to reform these complex structures. The regeneration of a limb, for example, occurs relatively fast and faithfully in animals such as salamanders, and this potential extends to the whole limb. Some rodents and primates (including humans) are able to regenerate the distal end of the fingertip, while more proximal amputations fail to regenerate. If we have retained this regenerative ability, why does a more proximal amputation result in wound healing and scarring? The ultimate goal of regenerative research is to understand why humans have limited regeneration and how we can improve it. With recent advances in genomic technology, scientists have progressed in uncovering cellular and molecular elements involved in regeneration by studying animal models such as the Mexican salamander (axolotl). Still, it remains unclear how, after amputation, the mature tissue at the stump can at the same time produce progenitor cells to restore multiple tissues and integrate them with pre-existing tissue as one functional unit. Failure in this functional integration is detrimental to any naturally regenerating tissue or cell-based therapy. This proposal aimed to find unifying mechanisms, or alternatives, that drive successful regeneration in the fingertip of three different species: axolotl, mouse, and human. In particular, we used fingertip regeneration to understand bone regeneration, the extensive remodeling of the bone at the stump, and its interaction with newly forming bone. By studying this process in different species, we expand and accelerate the possibilities of experimentation and treatment validation. In the axolotl, we found that upon injury, extensive remodeling of the skeleton occurs within a short time window. This process is driven by multinucleated cells specialized in bone resorption called osteoclasts. Although these cells had previously been described by histological methods in the axolotl, we were able to identify them by their genetic markers. We followed their activity in vivo and observed their recruitment timing, localization along the skeleton, and clearance from the tissue. This allowed us to define the timing of resorption, which is consistent across different long bones of the limb. Furthermore, the timing of resorption in the axolotl parallels the timing of bone resorption in mouse digit tip regeneration. This fast mechanism, triggered by regeneration, contrasts with the long-lasting remodeling observed during fracture healing, suggesting regeneration-specific regulation. We also found that inhibiting skeletal resorption impairs the integration of the old skeleton with the regenerated one. The severity of this non-union regeneration inversely correlates with the amount of skeletal resorption. In human fingertip regeneration, we found that bone resorption also occurs when the injury affects the bone. In this project, we were also able to characterize skeletal development in the axolotl. We used an interdisciplinary approach combining mathematical modeling, in vivo experiments, mechanical measurements, and histology. We defined the timing of skeletal ossification and its progression until old age. We found that axolotls continue to grow throughout their lifetime and that their limb bones never fully ossify. In addition, we measured for the first time in vivo how the mechanical properties of the limbs change during development and regeneration. This thorough characterization provides a reference framework for future studies on salamander bone regeneration.
Publications
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Single-cell revolution unveils the mysteries of the regenerative mammalian digit tip. Developmental Biology, 461(2), 107-109.
Riquelme-Guzmán, Camilo & Contreras, Osvaldo
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Postembryonic development and aging of the appendicular skeleton in Ambystoma mexicanum. Developmental Dynamics, 251(6), 1015-1034.
Riquelme‐Guzmán, Camilo; Schuez, Maritta; Böhm, Alexander; Knapp, Dunja; Edwards‐Jorquera, Sandra; Ceccarelli, Alberto S.; Chara, Osvaldo; Rauner, Martina & Sandoval‐Guzmán, Tatiana
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In vivo assessment of mechanical properties during axolotl development and regeneration using confocal Brillouin microscopy. Open Biology, 12(6).
Riquelme-Guzmán, Camilo; Beck, Timon; Edwards-Jorquera, Sandra; Schlüßler, Raimund; Müller, Paul; Guck, Jochen; Möllmert, Stephanie & Sandoval-Guzmán, Tatiana
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Methods for Studying Appendicular Skeletal Biology in Axolotls. Methods in Molecular Biology, 155-163. Springer US.
Riquelme-Guzmán, Camilo & Sandoval-Guzmán, Tatiana
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Osteoclast-mediated resorption primes the skeleton for successful integration during axolotl limb regeneration. eLife, 11.
Riquelme-Guzmán, Camilo; Tsai, Stephanie L.; Carreon Paz, Karen; Nguyen, Congtin; Oriola, David; Schuez, Maritta; Brugués, Jan; Currie, Joshua D. & Sandoval-Guzmán, Tatiana
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The salamander limb: a perfect model to understand imperfect integration during skeletal regeneration. Biology Open, 13(2).
Riquelme-Guzmán, Camilo & Sandoval-Guzmán, Tatiana
