Engineering scarless repair of flexor tendon injuries

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Abstract. Flexor tendon injuries in zone II of the hand are prone to debilitating adhesions, a form of scar tissue that obstructs gliding of the flexor tendons, severely impairing hand function. There are presently no pharmacologic treatments for the prevention or resolution of tendon adhesions, which still occur in as high as 30% of flexor tendon repairs, despite advances in surgical techniques and post-operative rehabilitation. Therefore, there is an unmet need for a mechanistic understanding of scar etiology in flexor tendons that could lead to the identification of new therapies. In the previous funding period, we established that disruption of canonical TGF-beta signaling in Smad3 knockout mice reduced flexor tendon adhesions but also reduced the tensile strength of the repair tissue. However, the differential effects of TGF-beta on activating peritendinous and intratendinous fibrosis remain unknown, and could be key to identifying novel profibrotic cellular and molecular mechanisms. To address this gap in knowledge, we will first utilize tamoxifen inducible gene deletion mouse models to investigate the effects of loss of canonical TGF-beta signaling in peritendinous versus intratendinous fibroblasts on zone II flexor tendon adhesions (Aim1). Towards the identification of downstream signaling mediators of fibrosis, we demonstrated that TGF-beta upregulates the protease- suppressor, plasminogen activator inhibitor 1 (PAI-1), which inhibits plasmin-mediated MMP activation. Furthermore, we demonstrated that PAI-1 loss of function nullifies TGF-beta1 inhibition of protease (plasmin and MMP) activity, without reducing cell proliferation. Thus, we hypothesize that PAI-1 activity does not affect cell proliferation and flexor tendon healing, but inhibits protease (plasmin and MMP) activity leading to impaired remodeling and increased adhesions. This hypothesis will be tested in Aim 2 by examining flexor tendon healing in mouse models of PAI- 1 loss- and gain-of-function. Collectively, our data suggest that targeting TGF-beta directly might be therapeutically untenable in load-bearing tendons due to its dichotomous effects, which include the indispensible PAI-1-independent effect of activating cell proliferation and matrix synthesis necessary for healing, and the pathogenic effect of inhibiting protease activity via canonical induction of PAI-1. Thus, we hypothesize that localized therapeutic inhibition of PAI-1, irrespective of its cell source, will ameliorate tendon adhesions without adversely affecting repair strength. In Aim 3, we will test this hypothesis by optimizing and investigating the efficacy of an innovative nanoparticle-mediated siRNA delivery system against PAI-1 in zone II flexor tendon injuries in wild type mice. The proposed studies will elucidate the mechanisms by which canonical TGF-beta signaling differentially activates peritendinous fibroblasts leading to extrinsic fibrosis (Aim 1), demonstrate that canonical TGF-beta induction of PAI-1 precipitates fibrotic adhesions (Aim 2), and establish localized, transient nanoparticle-mediated delivery of siRNA to inhibit PAI-1 as a translational, strength-sustaining, anti-adhesion therapy for flexor tendon repair (Aim 3).