Browsing by Subject "Tacrolimus (FK506)"

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    Mesenchymal stem cells and local tacrolimus delivery synergistically enhance neurite extension
    (Wiley, 2021) Saffari, Sara; Saffari, Tiam M.; Chan, Katelyn; Borschel, Gregory H.; Shin, Alexander Y.; Surgery, School of Medicine
    Background: The aim of this study was to investigate the combined effect of mesenchymal stem cells (MSC) and local delivery of tacrolimus (FK506) on nerve regeneration when applied to nerve autografts and decellularized allografts. Methods: A three-dimensional in vitro compartmented cell culture system consisting of a neonatal dorsal root ganglion adjacent to a nerve graft was used to evaluate the regenerating neurites into the peripheral nerve scaffold. Nerve autografts and allografts were treated with (i) undifferentiated MSCs, (ii) FK506 (100 ng/mL) or (iii) both (N = 9/group). After 48 hours, neurite extension was measured to quantify nerve regeneration and stem cell viability was evaluated. Results: Stem cell viability was confirmed in all MSC-treated grafts. Neurite extension was superior in autografts treated with FK506, and MSCs and FK506 combined (p < 0.001 and p = 0.0001, respectively), and autografts treated with MSCs (p = 0.12) were comparable to untreated autografts. In allografts, FK506 treatment and combined treatment were superior to controls (p < 0.001 and p = 0.0001, respectively), and treatment with MSCs (p = 0.09) was comparable to controls. All autograft groups were superior compared to their respective allograft treatment group (p < 0.05) in neurite extension. Conclusions: Alone, either MSC or FK506 treatment improved neurite outgrowth, and combined they further enhanced neurite extension in both autografts and allografts.
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    QS5: The Effect of Stem Cells and Local Tacrolimus on Neurite Extension
    (Wolters Kluwer, 2021) Saffari, Sara; Saffari, Tiam M.; Chan, Katelyn; Borschel, Gregory H.; Shin, Alexander Y.; Surgery, School of Medicine
    Purpose: Application of mesenchymal stem cells (MSCs) or tacrolimus (FK506), an FDA approved immunosuppressant, to nerve grafts has been a topic of interest to enhance peripheral nerve regeneration. The aim of this study was to investigate the combined effect of MSCs and local delivery of FK506 on nerve regeneration when applied to nerve autografts and decellularized nerve allografts. Methods: A three-dimensional (3D) in vitro compartmented cell culture system, validated by Tajdaran et al (2019), consisted of rat neonatal dorsal root ganglion (DRG) adjacent to rat nerve autograft or decellularized allograft. This model was used to evaluate regenerating neurites from the DRG into the peripheral nerve scaffold. Nerve autografts and decellularized allografts were augmented with (i) dynamic undifferentiated MSC seeding, (ii) local application of FK506 (100 ng/mL) or (iii) both (N=9/group). Local application was ensured by isolating the central system (i.e. DRG side) from the peripheral system (i.e. nerve graft side), where treatment was applied. After 48-hours of incubation, DRG-nerve graft constructs were collected, fixed, sectioned and stained against neurofilament-160 to measure neurite extension. CD90 staining was used to confirm stem cell characterization. Results: All grafts treated with MSCs confirmed CD90 expression. Compared to untreated autografts, neurite extension in autografts treated with FK506 and autografts treated with MSCs and FK506 combined were found superior (P<0.001 and P=0.0001, respectively), and comparable to autografts treated with MSCs (P=0.12). Compared to untreated allografts, allografts treated with FK506, and allografts treated with MSCs and FK506 were found superior (P<0.001 and P=0.0001, respectively), and allografts treated with MSCs were found comparable (P=0.09). All autograft groups were found superior compared to their respective allograft treatment group (P<0.05). Solely allografts receiving combined treatment were found superior to untreated autografts (P<0.05). Conclusion: MSCs or FK506 treatment improved neurite outgrowth and when combined, this resulted in significant synergistic neurite extension in both autografts and allografts in comparable patterns. Schematic overview of 3D compartmented cell culture system for isolated evaluation of treatment with MSCs and local FK506 in vitro. A 3.5 mm autograft or allograft with or without undifferentiated MSC seeding is attached to a DRG. DRG-nerve graft constructs are placed through a silicone isolator in the middle of a 24-wells plate to isolate the DRG from the nerve graft. FK506 containing media was added to the nerve graft side.
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    VP1: Combined Local Delivery of Tacrolimus and Stem Cells in Fibrin Gel is a Viable Potential Treatment for Enhancing Peripheral Nerve Regeneration
    (Wolters Kluwer, 2022) Saffari, Tiam M.; Chan, K.; Saffari, S.; Zuo, K. J.; Borschel, G. H.; Shin, A. Y.; Surgery, School of Medicine
    INTRODUCTION: The immunosuppressive and neuroregenerative potential of tacrolimus (FK506) may overcome the rejection of stem cells to enhance nerve regeneration. The aim of this study was to determine the feasibility and effectiveness of combining local, sustained delivery of tacrolimus (FK506) with stem cells. MATERIALS AND METHODS: The drug release profile of local FK506 over the course of 35 days was evaluated in phosphate-buffered saline (PBS). FK506 was incorporated into fibrin gel in poly(lactic-co-glycolic) acid (PLGA) microspheres in concentrations of 0.01 ng/mL and 100 ng/mL and incubated in phosphate-buffered saline (PBS) at physiological temperature. Microspheres were prepared and characterized according to validated protocols by Tajdaran et al. (2015). Adipose derived mesenchymal stem cells (MSCs) were cultured in collected PBS to mimic systemic exposure representing released concentrations at day 7, 15 and 28 from 0.01 ng/mL and 100 ng/mL microspheres. MSCs were cultured in the following groups: (i) hydrogel only, (ii) PLGA empty microspheres in hydrogel, (iii) Hydrogel Infused with Stem cells and Tacrolimus (HIST) 0.01 ng/mL, and (iv) HIST 100 ng/mL. Staining against CD90 was conducted to confirm stem cell characterization. MSC viability was evaluated at 24h, 48h, 72h, and seven days using Live/Dead staining and MTS assays. RESULTS: Microspheres containing 0.01 ng/mL showed depletion of FK506 by 13 days. FK506 microspheres containing 100 ng/mL revealed a sustained release up to 35 days. An inflation point was observed at day 15 due to erosion of gel, resulting in an increase of FK506 release. CD90 staining confirmed stem cell characterization. MSCs were viable up to seven days and no significant differences in viability were found between groups. CONCLUSION: Encapsulation of 100 ng/mL FK506 microspheres and MSCs in hydrogel is feasible and has strong potential to enhance survival of transplanted cells, which may ultimately lead to improved nerve regeneration.
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    VP2: Mesenchymal Stem Cells and Local Tacrolimus Delivery Synergistically Enhance Neurite Extension
    (Wolters Kluwer, 2022) Saffari, Tara Sara; Saffari, Tiam M.; Chan, Katelyn; Borschel, Gregory H.; Shin, Alexander Y.; Surgery, School of Medicine
    INTRODUCTION: Application of mesenchymal stem cells (MSCs) or tacrolimus (FK506), an FDA approved immunosuppressant, to nerve grafts has been a topic of interest to enhance peripheral nerve regeneration. The aim of this study was to investigate the combined effect of MSCs and local delivery of FK506 on nerve regeneration when applied to nerve autografts and decellularized nerve allografts. MATERIALS AND METHODS: A three-dimensional in vitro compartmented cell culture system consisted of rat neonatal dorsal root ganglion (DRG) adjacent to rat nerve autograft or decellularized allograft. This model was used to evaluate regenerating neurites from the DRG into the peripheral nerve scaffold. Nerve autografts and decellularized allografts were augmented with (i) dynamic undifferentiated MSC seeding, (ii) local application of FK506 (100 ng/mL) or (iii) both (N=9/group). Local application was ensured by isolating the central system (i.e. DRG side) from the peripheral system (i.e. nerve graft side), where treatment was applied. After 48-hours of incubation, DRG-nerve graft constructs were collected, fixed, sectioned and stained against neurofilament-160 to measure neurite extension. CD90 staining was used to confirm stem cell characterization. RESULTS: All grafts treated with MSCs confirmed CD90 expression. Compared to untreated autografts, neurite extension in autografts treated with FK506 and autografts treated with MSCs and FK506 combined were found superior (P<0.001 and P=0.0001, respectively), and comparable to autografts treated with MSCs (P=0.12). Compared to untreated allografts, allografts treated with FK506, and allografts treated with MSCs and FK506 were found superior (P<0.001 and P=0.0001, respectively), and allografts treated with MSCs were found comparable (P=0.09). All autograft groups were found superior compared to their respective allograft treatment group (P<0.05). Solely allografts receiving combined treatment were found superior to untreated autografts (P<0.05). CONCLUSION: MSCs or FK506 treatment improved neurite outgrowth and when combined, this resulted in significant synergistic neurite extension in both autografts and allografts in comparable patterns.