Optimizing Timing and Delivery of Hedgehog Signaling Activation to Improve Tendon-to-Bone Integration Jonathan Marcelin

Format:

Book

Thesis/Dissertation

Author/Creator:

Marcelin, Jonathan, author.

Contributor:

University of Pennsylvania. Bioengineering., degree granting institution.

Language:

English

Subjects (All):

0202.

0307.

0379.

0475.

Local Subjects:

0202.

0307.

0379.

0475.

Physical Description:

1 electronic resource (86 pages)

Contained In:

Dissertations Abstracts International 87-07B

Place of Publication:

Ann Arbor : ProQuest Dissertations and Theses, 2025

Language Note:

English

Summary:

Tendon and ligament injuries frequently occur at the tendon-to-bone insertion site, or enthesis. The enthesis is a specialized fibrocartilaginous interface composed of four distinct zones: tendon midsubstance, unmineralized fibrocartilage, mineralized fibrocartilage (MFC), and bone. These injuries often require surgical intervention, yet current repair techniques fail to reestablish the native, graded tendon-to-bone interface and typically result in disorganized scar tissue with poor mechanical integrity, contributing to high failure rates. Developmental studies have shown that the Hedgehog (Hh) signaling pathway plays a biphasic role in enthesis formation, with Sonic hedgehog (Shh) guiding early progenitor specification and Indian hedgehog (Ihh) promoting postnatal matrix maturation. Building on this foundation, our lab previously demonstrated that stimulation of the pathway genetically (with an over-expressing genetic mouse model) and pharmacologically (via systemic injections of a Hh agonist) enhances tendon-to-bone integration following anterior cruciate ligament reconstruction (ACLR) in mice. However, critical questions remain regarding the optimal timing of pathway activation and how to stimulate Hh signaling in a targeted manner without inducing off-target effects, given the pathway's vital role in multiple organ systems. This thesis investigates both the temporal and spatial dimensions of Hh pathway activation during adult enthesis healing. In Chapter 2, we used an αSMA-CreERT2 mouse model and temporally controlled delivery of the small-molecule Hh agonist SAG to dissect the pathway's biphasic role. We found that early delivery of the Hh agonist SAG increased proliferation of αSMA-lineage progenitor cells in both the healing tunnel and MFC, highlighting the important role of the Hh pathway in amplifying the progenitor pool. However, early delivery of SAG did not impact MFC formation, and late SAG delivery appeared to have a detrimental effect on MFC formation. Gene expression analysis at day 28 post-surgery revealed no significant changes in canonical Hh targets, extracellular matrix genes, or fibrocartilage markers. These results suggest that transcriptional changes induced by SAG delivery may be transient or may have occurred earlier in the healing process. Given that SAG was administered only during the first or third post-operative weeks and tissues were collected at day 28, it is likely that the transcriptional signature of pathway activation had already subsided by the time of analysis. A potential solution is to use a sustained-release system, such as electrospun scaffolds or hydrogels, to prolong exposure and achieve the sequential effect of progenitor expansion followed by a cumulative increase in MFC production. To that end, in chapter 3 we focused on localizing and prolonging Hh activation using SAG-loaded electrospun polycaprolactone (PCL) scaffolds. These scaffolds enabled dose- and time-dependent SAG release, effectively stimulating Hh signaling in a transverse tibial tunnel (TTT) in vivo model, and promoting mineral deposition in both the tendon graft and scaffold. By sustaining pathway activity, this approach increased fibrocartilage production and mineralization at the tendon-to-bone interface. Together, these findings suggest that Hh pathway activation is most therapeutically effective during the early proliferative phase of repair and that scaffold-based delivery systems offer a promising strategy to safely and precisely deliver SAG. Ultimately, this body of work advances our understanding of Hh signaling in enthesis repair and lays the groundwork for translational strategies to improve surgical outcomes in tendon and ligament reconstruction

Notes:

Advisors: Dyment, Nathaniel A. Committee members: Boerckel, Joel; Kuntz, Andrew F.; Gullbrand, Sarah; Tertuliano, Ottman

Source: Dissertations Abstracts International, Volume: 87-07, Section: B.

Ph.D. University of Pennsylvania 2025

Vendor supplied data

Local Notes:

School code: 0175

ISBN:

9798276001708

Access Restriction:

Restricted for use by site license