Publications by authors named "Patricia Friedrich"

66 Publications

Autogenous Arteriovenous Bundle Implantation Maintains Viability Without Increased Immune Response in Large Porcine Bone Allotransplants.

Transplant Proc 2021 Jan-Feb;53(1):417-426. Epub 2020 Sep 18.

Microvascular Research Laboratory, Department of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota, United States.

Background: Transplantation of living allogeneic bone segments may permit reconstruction of large defects, particularly if viability is maintained without immunosuppression. Development of a new autogenous osseous blood supply accomplishes this goal in rodent experimental models. This study evaluates potential systemic and local inflammatory responses to this angiogenesis in a large-animal model.

Methods: Vascularized allogeneic tibia segments were transplanted orthotopically into matched tibial defects in Yucatan minipigs. Microvascular anastomoses of bone nutrient artery and vein were supplemented by intramedullary placement of an autogenous arteriovenous (AV) bundle in group 1. Group 2 served as a no-angiogenesis control. A 3-drug immunosuppression regimen was withdrawn after 2 weeks. During the 20-week survival period, periodic leukocyte counts and inflammatory cytokine levels were measured. Thereafter, osteocyte survival was quantified and transplant rejection graded by histologic examination and quantitative real-time polymerase chain reaction of immunologic markers.

Results: Both groups developed an initial systemic response, which resolved after 4 to 6 weeks. No differences were seen in blood cytokine levels. Interleukin 2 expression was diminished in group 1 tibiae. As expected, nutrient pedicles had thrombosed without sustained immunosuppression, occluded by intimal hyperplasia. In group 1, angiogenesis from the autogenous AV bundle resulted in significantly less osteonecrosis (P = .04) and fibrosis (P = .02) than group 2 allotransplants.

Conclusions: Systemic immune responses to large-bone allotransplants were not increased by generation of an autogenous osseous blood supply within porcine tibial bone allotransplants. Implanted AV bundles diminished inflammation and fibrosis and improved bone viability when compared to no-angiogenesis controls.
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http://dx.doi.org/10.1016/j.transproceed.2020.07.020DOI Listing
May 2021

Transplant chimerism in porcine structural vascularized bone allotransplants.

Gene 2020 Jul 26;747:144627. Epub 2020 Mar 26.

Microvascular Research Laboratory, Department of Orthopedic Surgery, Mayo Clinic, Rochester, MN, USA. Electronic address:

Background: Bone allotransplant viability can be maintained long-term by implanting arteriovenous (AV) bundles and creating an autogenous neoangiogenic circulation. Only short-term immunosuppression is required. This study investigates the origin of viable osteocytes observed in areas of active bone remodeling in orthotopically transplanted tibiae in a Yucatan mini-pig model.

Methods: Segmental tibial defects created in female Yucatan minipigs (N = 14) were reconstructed with a matched vascularized composite allotransplant from a male donor. The circulation was microsurgically restored, with simultaneous autogenous AV-bundle implantation in group 1 (N = 7). A ligated AV-bundle was implanted as a no-angiogenesis control in group 2 (N = 7). After 20-weeks, repopulation of the allotransplant was assessed by real-time qPCR measurement of relative copy numbers of a Y chromosome-specific gene (SRY) and an autosomal housekeeping gene, ribosomal protein L4 (RPL4). A lower SRY/RPL4 ratio demonstrates replacement of male allogeneic cells with female, autogenous cells in the sample. Genomic DNA was extracted from cross-sections of the allotransplant, liver and spleen. Additionally, areas of new bone formation within the allotransplant were sampled by laser capture microdissection. A comparison was made between groups as well as male control samples. RNA was extracted from bone as well, as a measure of metabolically active cells.

Results: Laser-captured areas of new bone formation in animals with both normal and ligated AV-bundles were found to have significantly lower relative copy numbers of SRY (p = 0.03) than control specimens from male bone, indicating replacement by female (autogenous) bone-forming cells. Analysis of an entire segment of the allotransplant from Group 1 was similarly reduced (p = 0.04), unlike that from Group 2. RNA expression of SRY was observed in both groups. No chimerism could be found in non-bone tissues (liver and spleen).

Conclusion: We observed a significant level of transplant chimerism in areas of new bone formation sampled by laser capture microdissection. The migration of autogenous cells including osteocytes was seen in both groups. Survival of some allogeneic (male) cells was also demonstrable. No microchimerism was found in liver and spleen.
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http://dx.doi.org/10.1016/j.gene.2020.144627DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7298969PMC
July 2020

Functional Outcome after Reconstruction of a Long Nerve Gap in Rabbits Using Optimized Decellularized Nerve Allografts.

Plast Reconstr Surg 2020 06;145(6):1442-1450

From the Department of Orthopedic Surgery, Division of Hand Surgery, Mayo Clinic; the Department of Plastic, Reconstructive, and Hand Surgery, Erasmus Medical Center 's-Gravendijkwal; the Department of Plastic and Reconstructive Surgery, Radboudumc University Hospital; and the Xpert Clinic for Hand and Wrist Surgery.

Background: Processed nerve allografts are a promising alternative to nerve autografts, providing an unlimited, readily available supply and avoiding donor-site morbidity and the need for immunosuppression. Currently, clinically available nerve allografts do not provide satisfactory results for motor reconstruction. This study evaluated motor recovery after reconstruction of a long nerve gap using a processed nerve allograft and the influence of storage techniques.

Methods: Nerve allografts were decellularized using elastase and detergents and stored at either 4° or -80°C. In 36 New Zealand White rabbits, a 3-cm peroneal nerve gap was repaired with either an autograft (group 1, control) or a cold-stored (group 2) or frozen-stored (group 3) processed nerve allograft. Nerve recovery was evaluated using longitudinal ultrasound measurements, electrophysiology (compound muscle action potentials), isometric tetanic force, wet muscle weight, and histomorphometry after 24 weeks.

Results: Longitudinal ultrasound measurements showed that the cold-stored allograft provided earlier regeneration than the frozen-stored allograft. Furthermore, ultrasound showed significantly inferior recovery in group 3 than in both other groups (p < 0.05). Muscle weight and isometric tetanic force showed similar outcomes in the autograft and cold-stored allograft groups [p = 0.096 (muscle weight) and p = 0.286 (isometric tetanic force)], and confirmed the inferiority of the frozen-stored allograft to the autograft [p < 0.01 (muscle weight) and p = 0.02 (isometric tetanic force)].

Conclusions: Frozen storage of the nerve allograft significantly impairs functional recovery and should be avoided. The cold-stored optimized nerve allograft yields functional recovery similar to the gold standard autograft in the reconstruction of a 3-cm motor nerve defect. Future studies should focus on further improvement of the nerve allograft.
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http://dx.doi.org/10.1097/PRS.0000000000006818DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7255935PMC
June 2020

Gene expression and growth factor analysis in early nerve regeneration following segmental nerve defect reconstruction with a mesenchymal stromal cell-enhanced decellularized nerve allograft.

Plast Reconstr Surg Glob Open 2020 Jan 21;8(1):e2579. Epub 2020 Jan 21.

Department of Orthopedic Surgery, Mayo Clinic, Rochester, Minn.

The purpose of this study was to evaluate the molecular mechanisms underlying nerve repair by a decellularized nerve allograft seeded with adipose-derived mesenchymal stromal cells (MSCs) and compare it to the unseeded allograft and autograft nerve.

Methods: Undifferentiated MSCs were seeded onto decellularized nerve allografts and used to reconstruct a 10 mm gap in a rat sciatic nerve model. Gene expression profiles of genes essential for nerve regeneration and immunohistochemical staining (IHC) for PGP9.5, NGF, RECA-1, and S100 were obtained 2 weeks postoperatively.

Results: Semi-quantitative RT-PCR analysis showed that the angiogenic molecule was significantly increased in seeded allografts, and transcription factor was downregulated in seeded allografts. Seeded grafts showed a significant increase in immunohistochemical markers NGF and RECA-1, when compared with unseeded allografts.

Conclusions: MSCs contributed to the secretion of trophic factors. A beneficial effect of the MSCs on angiogenesis was found when compared with the unseeded nerve allograft, but implanted MSCs did not show evidence of differentiation into Schwann cell-like cells.
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http://dx.doi.org/10.1097/GOX.0000000000002579DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7015582PMC
January 2020

Revascularization patterns of nerve allografts in a rat sciatic nerve defect model.

J Plast Reconstr Aesthet Surg 2020 Mar 11;73(3):460-468. Epub 2019 Dec 11.

Department of Orthopedic Surgery, Mayo Clinic, 200 1st St. SW, Rochester, MN 55905, USA. Electronic address:

Introduction: The specific patterns of revascularization of allograft nerves after the addition of vascularization remain unknown. The aim of this study was to determine the revascularization patterns of optimized processed allografts (OPA) after surgically induced angiogenesis to the wound bed in a rat sciatic nerve model.

Materials And Methods: In 51 Lewis rats, sciatic nerve gaps were repaired with (i) autografts, (ii) OPA and (iii) OPA wrapped in a pedicled superficial inferior epigastric artery fascia flap (SIEF) to provide vascularization to the wound bed. At 2, 12, and 16 weeks, the vascular volume and vascular surface area in nerve samples were measured using micro CT and photography. Cross-sectional images were obtained and the number of vessels was quantified in the proximal, mid, and distal sections of the nerve samples.

Results: At 2 weeks, the vascular volume of SIEF nerves was comparable to control (P = 0.1). The vascular surface area in SIEF nerves was superior to other groups (P<0.05). At 12 weeks, vascularity in SIEF nerves was significantly higher than allografts (P<0.05) and superior compared to all other groups (P<0.0001) at 16 weeks. SIEF nerves had a significantly increased number of vessels compared to allografts alone in the proximal (P<0.05) and mid-section of the graft (P<0.05).

Conclusions: Addition of surgical angiogenesis to the wound bed greatly improves revascularization. It was demonstrated that revascularization occurs primarily from proximal to distal (proximal inosculation) and not from both ends as previously believed and confirms the theory of centripetal revascularization.
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http://dx.doi.org/10.1016/j.bjps.2019.11.048DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7770618PMC
March 2020

Neo-Angiogenesis, Transplant Viability, and Molecular Analyses of Vascularized Bone Allotransplantation Surgery in a Large Animal Model.

J Orthop Res 2020 02 6;38(2):288-296. Epub 2019 Oct 6.

Department of Orthopedic Surgery, Microvascular Research Laboratory, Mayo Clinic, 200 1st street SW55905, Building: MS-3-85, Rochester, Minnesota.

Vascularized composite allotransplantation of bone is a possible alternative treatment for large osseous defects but requires life-long immunosuppression. Surgical induction of autogenous neo-angiogenic circulation maintains transplant viability without this requirement, providing encouraging results in small animal models [1-3]. A preliminary feasibility study in a swine tibia model demonstrated similar findings [4, 5]. This study in swine tibial allotransplantation tests its applicability in a pre-clinical large animal model. Previously, we have demonstrated bone vascularized composite allotransplantation (VCA) survival was not the result of induction of tolerance nor an incompetent immune system [1]. Fourteen tibia vascularized bone allotransplants were microsurgically transplanted orthotopically to reconstruct size-matched tibial defects in Yucatan miniature swine. Two weeks of immunosuppression was used to maintain allotransplant pedicle patency during angiogenesis from a simultaneously implanted autogenous arteriovenous bundle. The implanted arteriovenous bundle was patent in group 1 and ligated in group 2 (a neo-angiogenesis control). At twenty weeks, we quantified the neo-angiogenesis and correlated it with transplant viability, bone remodeling, and gene expression. All patent arteriovenous bundles maintained patency throughout the survival period. Micro-angiographic, osteocyte cell count and bone remodeling parameters were significantly higher than controls due to the formation of a neo-angiogenic autogenous circulation. Analysis of gene expression found maintained osteoblastic and osteoclastic activity as well as a significant increase in expression of endothelial growth factor-like 6 (EGFL-6) in the patent arteriovenous bundle group. Vascularized composite allotransplants of swine tibia maintained viability and actively remodeled over 20 weeks when short-term immunosuppression is combined with simultaneous autogenous neo-angiogenesis. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:288-296, 2020.
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http://dx.doi.org/10.1002/jor.24481DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6980263PMC
February 2020

Outcomes of Vascularized Bone Allotransplantation with Surgically Induced Autogenous Angiogenesis in a Large Animal Model: Bone Healing, Remodeling, and Material Properties.

J Reconstr Microsurg 2020 Feb 30;36(2):82-92. Epub 2019 Aug 30.

Microvascular Research Laboratory, Department of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota.

Background:  Bone vascularized composite allotransplantation (VCA) is a possible alternative for the treatment of large bone defects. Clinical application of VCAs is limited by the need for life-long immunosuppression (IS). We report an alternative method to maintain bone allotransplant viability in a large animal model without the need for life-long IS by using autogenous vessel implantation.

Methods:  Fourteen bone only VCAs were transplanted in a porcine tibia defect model with short-term IS. Two groups were used to evaluate the effect of the implantation of an autogenous arteriovenous (AV)-bundle, therefore the only difference between the groups was the patency of the AV-bundle. We radiographically evaluated bone healing and allogenic pedicle patency. AV-bundle patency and union were evaluated with micro-CT. Bone remodeling was assessed with histomorphometry and material properties were evaluated with axial compression testing and cyclic reference point indentation.

Results:  Two subjects did not reach the final time point. Twelve tibiae healed proximally, and nine at the distal transplant-bone interface. Bone allotransplants showed their viability in the first 4 to 6 weeks by significant periosteal bridging arising from the transplant and maintained pedicle patency. Bone material properties were not affected by the implantation of an AV-bundle when compared with ligated AV-bundle controls, but diminished compared with normal bone. Significantly higher bone formation rates resulted from the implantation of a patent AV-bundle.

Conclusion:  New periosteal bone formation and subsequent bone healing result from blood flow through the microsurgically repaired nutrient blood supply, demonstrated by maintained allogenic pedicle patency. The implantation of a patent autogenous AV-bundle has no adverse effect on material properties, but a positive effect on bone remodeling of endosteal surfaces despite thrombosis of the allogenic pedicle. Bone material properties change after transplantation compared with normal bone, although 20-weeks survival time is relatively short for the final evaluation of bone material properties.
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http://dx.doi.org/10.1055/s-0039-1695052DOI Listing
February 2020

Brachial plexus nerve injury and repair in a rabbit model part II: Does middle trunk injury result in loss of biceps function while repair results in recovery of biceps function.

Microsurgery 2019 Oct 6;39(7):634-641. Epub 2019 Aug 6.

Department of Orthopedic Surgery and Division of Hand & Microvascular Surgery, Mayo Clinic, Rochester, Minnesota.

Introduction: There is conflicting anatomic and innervation data regarding the rabbit brachial plexus injury model. This study aims to validate a rabbit brachial plexus injury model. We hypothesize the middle trunk (C6, C7) is the primary innervation of the biceps, and when cut and unrepaired, would demonstrate lack of recovery and when repaired would demonstrate evidence of recovery.

Materials And Methods: Twenty two male New Zealand white rabbits (3-4 kg) underwent unilateral surgical division of the middle trunk. Five rabbits were randomly assigned to the "no-repair" group while the remaining 17 rabbits underwent direct coaptation ("repair" group). Rabbits were followed for 12 weeks, with ultrasound measurement of biceps cross-sectional area performed preoperatively, and at 4, 8, and 12 weeks postoperatively. At a euthanasia procedure, bilateral compound muscle action potential (CMAP) and isometric tetanic force (ITF) were measured. Bilateral biceps muscles were harvested and wet muscle weight was recorded. The operative side was expressed as a percentage of the non-operated side, and differences between the no repair and repair rabbits were statistically compared.

Results: The repair group demonstrated significantly higher CMA (23.3 vs. 0%, p < .05), ITF (25.6 vs. 0%, p < .05), and wet muscle weight (65.8 vs. 52.0%, p < .05) as compared to the unrepaired group. At 4 weeks postoperatively, ultrasound-measured cross-sectional area of the biceps demonstrated atrophy in both groups. At 12 weeks, the repair group had a significantly larger cross-sectional area as compared to the no-repair group (89.1 vs. 59.3%, p < .05).

Conclusions: This injury model demonstrated recovery with repair and lack of function without repair. Longer survival time is recommended for future investigations.
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http://dx.doi.org/10.1002/micr.30500DOI Listing
October 2019

The rabbit brachial plexus as a model for nerve injury and repair Part 1: Anatomic study of the biceps and triceps innervation.

Microsurgery 2020 Feb 27;40(2):183-188. Epub 2019 Jun 27.

Department of Orthopedic Surgery, Division of Hand and Microvascular Surgery, Mayo Clinic, Rochester, Minnesota.

Purpose: Animal models can be helpful in evaluating new surgical strategies for brachial plexus reconstruction. While several groups have already used the rabbit brachial plexus to model injury, reports conflict in anatomic detail and do not identify a nerve-muscle pair to measure motor function recovery after reconstruction. The purpose of the current study is to describe the innervations of the biceps and triceps muscles in rabbits, which are both amenable to study in brachial plexus injury models.

Materials And Methods: Thirteen rabbits weighing 2-2.5 kg were anesthetized. Six rabbits were sacrificed and dissected using loupe and microscope magnification to understand the overall morphology of the brachial plexus. Seven rabbits underwent electrophysiologic investigation. A bipolar nerve stimulator was used to systematically stimulate the roots, trunks and divisions, and nerve branches of the rabbit brachial plexus and compound muscle action potential was used to record muscle response. Nerve length and width measurements were not recorded.

Results: Roots contributing to the brachial plexus were C5, C6, C7, C8, and T1. In contrast to other anatomical studies, T2 did not contribute to the brachial plexus. The triceps was innervated by the radial nerve, which received contributions from C6 (1.6 mA), C7 (1.9 mA), C8, and T1 (12.2 mA).The biceps had dual innervation (proximally and distally). The proximal branch received contributions from C6 (3.5 mA) and C7 (5mA). The distal portion was innervated by a branch from the median nerve, which received innervation from C6, C7, C8, and T1.

Conclusions: The overall structure of rabbit brachial plexus is described and innervation of the biceps and triceps is described in detail. This anatomic investigation will form the basis of a future brachial plexus model of injury and repair.
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http://dx.doi.org/10.1002/micr.30482DOI Listing
February 2020

Effects of Surgical Angiogenesis on Segmental Bone Reconstruction With Cryopreserved Massive-Structural Allografts in a Porcine Tibia Model.

J Orthop Res 2019 08 17;37(8):1698-1708. Epub 2019 May 17.

Department of Orthopedic Surgery, Microvascular Research Laboratory, Mayo Clinic, 200 First Street SW, Rochester, Minnesota.

Cryopreserved bone allografts (CBA) used to reconstruct segmental bone defects provide immediate structural stability, but are vulnerable to infection, non-union and late stress fracture as the majority of the allograft remains largely avascular. We sought to improve the bone vascularity and bone formation of CBAs by surgical angiogenesis with an implanted arteriovenous (AV) bundle, using a porcine tibial defect model. Cryopreserved tibial bone allografts were transplanted in swine leukocyte antigen (SLA) mismatched Yucatan minipigs to reconstruct a 3.5 cm segmental tibial defect. A cranial tibial AV-bundle was placed within its intramedullary canal to induce angiogenesis. The AV bundle was patent in eight pigs and ligated in a control group of eight pigs. At 20 weeks neo-angiogenesis was evaluated by micro-angiography. Bone formation was measured by quantitative histomorphometry and micro-computed tomography. Seven of eight AV-bundles in the revascularized group were patent. One had thrombosed due to allograft displacement. Total vascular volume was higher in the revascularized allografts compared to the ligated group (p = 0.015). Revascularized allografts had increased levels of bone formation on the allograft endosteal surface compared to the ligated control group (p = 0.05). Surgical angiogenesis of porcine tibial CBAs by intramedullary implantation of an AV-bundle creates an enhanced autogenous neoangiogenic circulation and accelerates active bone formation on allograft endosteal surfaces. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:1698-1708, 2019.
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http://dx.doi.org/10.1002/jor.24318DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6824922PMC
August 2019

Comparable functional motor outcomes after repair of peripheral nerve injury with an elastase-processed allograft in a rat sciatic nerve model.

Microsurgery 2018 Oct 19;38(7):772-779. Epub 2018 Sep 19.

Department of Orthopedic Surgery, Microvascular Research Laboratory, Mayo Clinic, Rochester, Minnesota.

Background: A bridging nerve autograft is the gold standard for the repair of segmental nerve injury that cannot be repaired directly. However, limited availability and donor site morbidity remain major disadvantages of autografts. Here, a nerve allograft decellularized with elastase was compared with an autograft regarding functional motor outcome in a rat sciatic segmental nerve defect model. Furthermore, the effect of storage on this allograft was studied.

Methods: Sixty-six Lewis rats (250-300 g) underwent a 10-mm sciatic nerve reconstruction using either a cold- (n = 22) or frozen-stored (n = 22) decellularized nerve allograft or an autograft (n = 22). Sprague-Dawley rats (300-350 g) served as full major histocompatibility complex-mismatched donors. Functional motor outcome was evaluated after 12 and 16 weeks. Ankle angle, compound muscle action potential (CMAP), isometric tetanic force, wet muscle weight, and histomorphometry were tested bilaterally.

Results: For CMAP and isometric tetanic force, no significant differences were observed between groups. In contrast, for ankle angle, histomorphometry and muscle weight, the cold-stored allograft performed comparable to the autograft, while the frozen-stored allograft performed significantly inferior to the autograft. At week 16, ankle angle was 88.0 ± 3.1% in the cold-stored group, 77.4 ± 3.6% in the frozen-stored group, and 74.1 ± 3.1% in the autograft group (P < .001); At week 16, the muscle weight showed a recovery up to 71.1 ± 4.8% in the autograft group, 67.0 ± 6.6% in the cold-stored group, and 64.7 ± 3.7% in the frozen-stored group (P < .05).

Conclusions: A nerve allograft decellularized with elastase, if stored under the right conditions, results in comparable functional motor outcomes as the gold standard, the autograft.
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http://dx.doi.org/10.1002/micr.30371DOI Listing
October 2018

Bone vascularized composite allotransplantation model in swine tibial defect: Evaluation of surgical angiogenesis and transplant viability.

Microsurgery 2019 Feb 5;39(2):160-166. Epub 2018 Mar 5.

Microvascular Research Laboratory, Department of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota.

Introduction: In prior small animal studies, we maintained vascularized bone allotransplant viability without long-term immunotherapy. Instead, an autogenous neoangiogenic circulation is created from implanted vessels, sufficient to maintain bone viability with only 2 weeks immunosupression. Blood flow is maintained despite rejection of the allogeneic vascular pedicle thereafter. We have previously described a large animal (swine) pre-clinical model, reconstructing tibial defects with vascularized tibial allotransplants. In this manuscript, autologous angiogenesis is evaluated in this model and correlated with bone viability.

Materials And Methods: Allogeneic tibial segments were transplanted across a major swine leukocyte antigen mismatch. Microvascular repair of the bone VCA pedicle was combined with intraosseous implantation of an autogenous arteriovenous (AV) bundle. The bundle was ligated in group 1 (n = 4), and allowed to perfuse in group 2 (n = 4). Three-drug immunotherapy was given for 2 weeks. At 16 weeks micro-CT angiography quantified neoangiogenic vessel volume. Bone viability, rejection grade, and bone healing were analyzed.

Results: A substantial neoangiogenic circulation developed from the implanted AV-bundle in group 2, with vessel density superior to ligated AV-bundle controls (0.11 ± 0.05 vs. 0.01 ± 0.01, P = .029). Bone allotransplant viability was also significantly enhanced by neoangiogenesis (78.7 ± 4.4% vs. 27.7 ± 5.8%, P = .028) with higher bone healing scores (21.4 ± 2.9 vs. 12.5 ± 3.7, P = .029). Ligated control tibias demonstrated disorganized bone morphology and higher local inflammation (P = .143).

Conclusion: Implantation of autogenous AV bundles into vascularized bone allotransplants resulted in the rapid formation of a neoangiogenic autogenous blood supply in a swine tibia model that maintained bone viability, improved bone healing, and minimized rejection.
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http://dx.doi.org/10.1002/micr.30310DOI Listing
February 2019

Validation of Isometric Tetanic Force as a Measure of Muscle Recovery After Nerve Injury in the Rabbit Biceps.

J Hand Surg Am 2018 05 3;43(5):488.e1-488.e8. Epub 2018 Jan 3.

Department of Orthopaedic Surgery, Mayo Clinic, Rochester, MN. Electronic address:

Purpose: The purpose of this study was to describe and validate a technique for measurement of isometric tetanic force (ITF) in the rabbit biceps muscle.

Materials And Methods: Eighteen New Zealand White rabbits were randomized to test either the right side or the left side first. Under propofol anesthesia, the brachial plexus and biceps brachii were exposed. The middle trunk (C6, C7) was secured in a bipolar electrode. Compound muscle action potential (CMAP) was measured. The proximal, tendinous portion of the biceps was severed at the shoulder and clamped in a custom-made force transducer. Muscle preload and electrical stimulation variables were optimized to obtain the highest tetanic muscle contraction. Wet muscle weight (WMW) and nerve histomorphometry were analyzed. Statistical analysis was performed to determine side-to-side equivalence.

Results: The rabbit biceps muscle force demonstrated side-to-side equivalence with overlapping 95% confidence intervals (95% CI). The right side, expressed as a percentage of the left, averaged 99.69% (95% CI, 88.89%-110.5%). The WMW of the right expressed as a percentage of the left was 98.9% (95% CI, 95.8%-102%).

Conclusions: The ITF is equivalent from side to side in the rabbit as demonstrated by the high degree of overlap in the 95% CIs for each side. The width of the 95% CI implies that there is more variability in the rabbit upper extremity than for the lower extremity of the rabbit or rat models, and researchers should take this into account when performing sample size estimates in pre-experimental planning.

Clinical Relevance: The rabbit biceps muscle ITF measurements can be used to measure motor recovery in a rabbit model of brachial plexus injury and compared with the contralateral uninjured side.
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http://dx.doi.org/10.1016/j.jhsa.2017.10.037DOI Listing
May 2018

A new porcine vascularized tibial bone allotransplantation model. Anatomy and surgical technique.

Microsurgery 2018 Feb 13;38(2):195-202. Epub 2017 Nov 13.

Microvascular Research Laboratory, Department of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota.

Introduction: Vascularized bone allotransplantation may provide new options for reconstruction of segmental defects if problems of long-term immune modulation can be solved. The current literature lacks an orthotopic large animal model, limited to bone and without the confounding effects of other tissue types, permitting a multifaceted evaluation before new methods are used clinically. The purpose of this study was to develop a large animal model for vascularized bone allotransplantation.

Materials And Methods: Eight porcine hind limbs were dissected. Length, diameter, and location of all hindlimb vessels were measured and a single nutrient vessel supplying the tibial diaphysis identified enabling its use as a vascularized bone allotransplant. Four Yucatan minipigs were divided into two pairs with a major swine leukocyte antigen mismatch. A 3.5 cm tibial segment including its nutrient pedicle was raised simultaneously from each pig and transplanted into the matched defect of the other animal. Microarterial anastomosis of the pedicle and 3-drug immunosuppression maintained VCA viability. Bone healing and limb function were followed for 16 weeks.

Results: A consistent tibia diaphyseal nutrient artery arose from the caudal tibial artery to enter bone a mean 2.8 mm distal to the tibial tubercle with a pedicle length of 6.6 ± 3.3 mm and diameter of 1.6 ± 0.2 mm. Using this pedicle, we reconstructed a 3.5 cm tibial defect with a vascularized bone allotransplant in four animals. Immediate weightbearing as well as progressive bone healing was demonstrated.

Conclusion: We have developed a vascularized tibial bone allotranplantation large-animal model suitable for future bone-only allotranplantation research in mini-pigs.
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http://dx.doi.org/10.1002/micr.30255DOI Listing
February 2018

Surgical Angiogenesis in Porcine Tibial Allotransplantation: A New Large Animal Bone Vascularized Composite Allotransplantation Model.

J Vis Exp 2017 08 13(126). Epub 2017 Aug 13.

Microvascular Research Laboratory, Department of Orthopedic Surgery, Mayo Clinic;

Segmental bone loss resulting from trauma, infection malignancy and congenital anomaly remains a major reconstructive challenge. Current therapeutic options have significant risk of failure and substantial morbidity. Use of bone vascularized composite allotransplantation (VCA) would offer both a close match of resected bone size and shape and the healing and remodeling potential of living bone. At present, life-long drug immunosuppression (IS) is required. Organ toxicity, opportunistic infection and neoplasm risks are of concern to treat such non-lethal indications. We have previously demonstrated that bone and joint VCA viability may be maintained in rats and rabbits without the need of long-term-immunosuppression by implantation of recipient derived vessels within the VCA. It generates an autogenous, neoangiogenic circulation with measurable flow and active bone remodeling, requiring only 2 weeks of IS. As small animals differ from man substantially in anatomy, bone physiology and immunology, we have developed a porcine bone VCA model to evaluate this technique before clinical application is undertaken. Miniature swine are currently widely used for allotransplantation research, given their immunologic, anatomic, physiologic and size similarities to man. Here, we describe a new porcine orthotopic tibial bone VCA model to test the role of autogenous surgical angiogenesis to maintain VCA viability. The model reconstructs segmental tibial bone defects using size- and shape-matched allogeneic tibial bone segments, transplanted across a major swine leukocyte antigen (SLA) mismatch in Yucatan miniature swine. Nutrient vessel repair and implantation of recipient derived autogenous vessels into the medullary canal of allogeneic tibial bone segments is performed in combination with simultaneous short-term IS. This permits a neoangiogenic autogenous circulation to develop from the implanted tissue, maintaining flow through the allogeneic nutrient vessels for a short time. Once established, the new autogenous circulation maintains bone viability following cessation of drug therapy and subsequent nutrient vessel thrombosis.
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http://dx.doi.org/10.3791/55238DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5614272PMC
August 2017

The effect of full dose composite tissue allotransplantation immunosuppression on allograft motor nerve regeneration in a rat sciatic nerve model.

Microsurgery 2018 Jan 9;38(1):66-75. Epub 2017 Aug 9.

Microvascular Research Laboratory, Mayo Clinic, Rochester, Minnesota.

Background: The purpose of this study was to identify which triple immunosuppressive protocols, currently used for vascularized composite allotransplantation in the clinic, will have the best effect on motor function recovery following nerve allograft reconstruction.

Methods: Eighty-eight Lewis rats underwent a 1-cm sciatic nerve allograft transplantation and skin graft from 44 Brown-Norway rats. Group I received 0.9% isotonic saline (control); Group II, 2 mg/kg FK506; Group III, 1 mg/kg FK506 with 15 mg/kg mycophenolate mofetil (MMF); and Group IV, 2 mg/kg FK506 with 30 mg/kg MMF and prednisone. Each group consisted of 11 rats. After 12 weeks, motor function recovery was evaluated with isometric tetanic force, muscle mass, ankle contracture angle, electrophysiology, and nerve histomorphometry. Adequacy of immunosuppression was monitored with the transplanted skin graft. All data are expressed as a percentage of the contralateral side.

Results: Isometric tetanic force showed significantly better functional recovery in all groups treated with immunosuppression compared to control. Within the immunosuppression groups no significant difference was found: 42.1 ± 6.4% (Group I), 56.1 ± 12.4% (Group II), 58.4 ± 10.7% (Group III), and 61.3 ± 11.2% (Group IV). Group IV was superior to all other groups regarding ankle contracture (P < .05) and electrophysiology (P < .001). Skin graft rejection occurred in 41 and 0% (Groups III and IV, respectively).

Conclusions: FK506 significantly enhanced motor recovery after allograft reconstruction. This effect was comparable between combination treatment (low-dose FK506 and MMF) and triple therapy (high-dose FK506 and MMF plus prednisolone). However, triple therapy was more effective in suppressing skin rejection.
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http://dx.doi.org/10.1002/micr.30211DOI Listing
January 2018

Speaker Introductions at Internal Medicine Grand Rounds: Forms of Address Reveal Gender Bias.

J Womens Health (Larchmt) 2017 05 16;26(5):413-419. Epub 2017 Feb 16.

5 Department of Medicine, Mayo Clinic Rochester , Rochester, Minnesota.

Background: Gender bias has been identified as one of the drivers of gender disparity in academic medicine. Bias may be reinforced by gender subordinating language or differential use of formality in forms of address. Professional titles may influence the perceived expertise and authority of the referenced individual. The objective of this study is to examine how professional titles were used in the same and mixed-gender speaker introductions at Internal Medicine Grand Rounds (IMGR).

Methods: A retrospective observational study of video-archived speaker introductions at consecutive IMGR was conducted at two different locations (Arizona, Minnesota) of an academic medical center. Introducers and speakers at IMGR were physician and scientist peers holding MD, PhD, or MD/PhD degrees. The primary outcome was whether or not a speaker's professional title was used during the first form of address during speaker introductions at IMGR. As secondary outcomes, we evaluated whether or not the speakers professional title was used in any form of address during the introduction.

Results: Three hundred twenty-one forms of address were analyzed. Female introducers were more likely to use professional titles when introducing any speaker during the first form of address compared with male introducers (96.2% [102/106] vs. 65.6% [141/215]; p < 0.001). Female dyads utilized formal titles during the first form of address 97.8% (45/46) compared with male dyads who utilized a formal title 72.4% (110/152) of the time (p = 0.007). In mixed-gender dyads, where the introducer was female and speaker male, formal titles were used 95.0% (57/60) of the time. Male introducers of female speakers utilized professional titles 49.2% (31/63) of the time (p < 0.001).

Conclusion: In this study, women introduced by men at IMGR were less likely to be addressed by professional title than were men introduced by men. Differential formality in speaker introductions may amplify isolation, marginalization, and professional discomfiture expressed by women faculty in academic medicine.
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http://dx.doi.org/10.1089/jwh.2016.6044DOI Listing
May 2017

Optimizing decellularization techniques to create a new nerve allograft: an in vitro study using rodent nerve segments.

Neurosurg Focus 2017 Mar;42(3):E4

Department of Orthopedic Surgery, Microvascular Research Laboratory, Mayo Clinic, Rochester, Minnesota; and.

OBJECTIVE Commercially available processed nerve allografts have been shown to be inferior to autografts in previous animal studies. The authors hypothesized that combining different processing and storage techniques will result in improved nerve ultrastructure preservation, lower immunogenicity, and minimized cellular debris. Different processing protocols were evaluated using chemical detergents, enzymes, and irradiation, with the addition the of enzyme elastase, were used. Additionally, the difference between cold and frozen storage was investigated. The goal of this study was to create an optimized nerve allograft. METHODS Fifty rat nerves were decellularized with modifications of previous protocols and the addition of elastase. Subsequently, the nerve segments were stored at either 4°C or -80°C. Both processed and fresh control nerves were analyzed with confocal microscopy using immunohistochemical staining on the basal lamina (laminin γ-1), Schwann cells (S100 protein), and immunogenicity using major histocompatibility complex-I (MHCI) staining. Morphology of the ultrastructure and amount of cellular debris were analyzed on cross-sections of the nerves stained with toluidine blue and H & E, and by using electron microscopy. RESULTS Nerve ultrastructure was preserved with all decellularization protocols. Storage at -80°C severely altered nerve ultrastructure after any decellularization method. Elastase was found to significantly reduce the immunogenicity and amount of Schwann cells, while maintaining good structural properties. CONCLUSIONS Reduced immunogenicity, diminished cellular debris, and the elimination of Schwann cells was observed when elastase was added to the nerve processing while maintaining ultrastructure. Storage at -80°C after the decellularization process heavily damaged the nerve ultrastructure as compared with cold storage. Further in vivo studies are needed to prove the nerve regenerative capacity of these optimized allografts.
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http://dx.doi.org/10.3171/2017.1.FOCUS16462DOI Listing
March 2017

Recipient-derived angiogenesis with short term immunosuppression increases bone remodeling in bone vascularized composite allotransplantation: A pilot study in a swine tibial defect model.

J Orthop Res 2017 06 12;35(6):1242-1249. Epub 2016 Aug 12.

Microvascular Research Laboratory, Department of Orthopedic Surgery, Mayo Clinic, 200 First Street SW, Rochester, Minnesota.

Current vascularized composite allotransplantation (VCA) transplantation protocols rely upon life-long immune modulation to maintain tissue perfusion. Alternatively, bone-only VCA viability may be maintained in small animal models using surgical angiogenesis from implanted autogenous vessels to develop a neoangiogenic bone circulation that will not be rejected. This study tests the method's efficacy in a large animal model as a bridge to clinical practice, quantifying the remodeling and mechanical properties of porcine tibial VCAs. A segmental tibial defect was reconstructed in Yucatan miniature swine by transplantation of a matched tibia segment from an immunologically mismatched donor. Microsurgical repair of nutrient vessels was performed in all pigs, with simultaneous intramedullary placement of an autogenous arteriovenous (AV) bundle in Group 2. Group 1 served as a no-angiogenesis control. All received 2 weeks of immunosuppression. After 16 weeks, micro-CT and histomorphometric analyses were used to evaluate healing and remodeling. Axial compression and nanoindentation studies evaluated bone mechanical properties. Micro-CT analysis demonstrated significantly more new bone formation and bone remodeling at the distal allotransplant/recipient junction and on the endosteal surfaces of Group 2 tibias (p = 0.03). Elastic modulus and hardness were not adversely affected by angiogenesis. The combination of 2 weeks of immunosuppression and autogenous AV-bundle implantation within a microsurgically transplanted tibial allotransplant permitted long-term allotransplant survival over the study period of 16 weeks in this large animal model. Angiogenesis increased bone formation and remodeling without adverse mechanical effects. The method may allow future composite-tissue allotransplantation of bone without the risks associated with long-term immunosuppression. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1242-1249, 2017.
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http://dx.doi.org/10.1002/jor.23378DOI Listing
June 2017

Motor Nerve Recovery in a Rabbit Model: Description and Validation of a Noninvasive Ultrasound Technique.

J Hand Surg Am 2016 Jan;41(1):27-33

Department of Orthopedic Surgery, Microvascular Research Laboratory, Mayo Clinic, Rochester, MN. Electronic address:

Purpose: To develop and validate a noninvasive ultrasound technique for the longitudinal analysis of functional recovery after segmental peroneal nerve reconstruction in a rabbit model.

Methods: Twelve male New Zealand White rabbits underwent a 1-cm peroneal nerve autograft reconstruction. Ultrasound measurements were performed before surgery and at 1, 2, 4, 8, 12, and 16 weeks postoperatively. All rabbits were managed with manual restraint for the ultrasound procedure, avoiding the risks of anesthetics. At 12 and 16 weeks, we evaluated functional recovery using compound muscle action potential, isometric tetanic force measurements, wet muscle weight, and nerve histomorphometry. Data were compared with ultrasound measurements by calculating the Pearson correlation coefficient. We determined intra-rater and inter-rater reliability of the ultrasound measurements.

Results: Ultrasound demonstrated good correlation with isometric tetanic force measurements and wet muscle weight, good correlation with nerve histomorphometry, and moderate correlation with compound muscle action potential. Both intra-rater and inter-rater reliability of the ultrasound technique was excellent.

Conclusions: Ultrasound analysis of the tibialis anterior muscle provided a reliable method for analysis of functional recovery in a rabbit peroneal nerve reconstruction model. The noninvasive nature allowed for longitudinal follow-up within the same animal and measurement of early recovery without the use of anesthesia.

Clinical Relevance: Application of this noninvasive technique can reduce the variability and sample size necessary in peripheral nerve reconstruction studies and may provide an ideal tool for comparative studies in larger animal models.
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http://dx.doi.org/10.1016/j.jhsa.2015.11.002DOI Listing
January 2016

Effect of Vascular Endothelial Growth Factor Administration on Nerve Regeneration after Autologous Nerve Grafting.

J Reconstr Microsurg 2016 Mar 30;32(3):183-8. Epub 2015 Sep 30.

Department of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota.

Background: The aim of this study was to evaluate the effect of vascular endothelial growth factor (VEGF) administration around the autologous nerve graft on nerve recovery in a rat model.

Methods: A total of 69 rats were randomly divided into three experimental groups. A 10-mm sciatic nerve defect was made and reconstructed with the reversed nerve segment. Group I received an osmotic pump with saline, group II received an osmotic pump with VEGF, and group III added a silicone tube around the nerve graft to decrease the surrounding blood supply. Nine animals in each group were sacrificed on day 3 to evaluate improvement in new vessel formation. In each group 14 animals were sacrificed at 16 weeks after the initial procedure to evaluate the functional motor nerve regeneration using compound muscle action potential, isometric tetanic force, wet muscle weight, and nerve histomorphometry.

Results: The average vascular density on day 3 was 10.7% in group I, 21.4% in group II, and 0.9% in group III. These differences were significant. However, the average maximum isometric tetanic force at 16 weeks was 54.4% in group I, 57.5% in group II, and 47.6% in group III. No difference was found with or without VEGF administration. Histomorphometric analysis was also not significantly different between the groups.

Conclusions: New vessel formation on autologous nerve graft was enhanced by VEGF administration. However, the neovascularization effect of VEGF administration did not translate into better motor nerve recovery.
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http://dx.doi.org/10.1055/s-0035-1563709DOI Listing
March 2016

The influence of vascularization of transplanted processed allograft nerve on return of motor function in rats.

Microsurgery 2016 Feb 30;36(2):134-43. Epub 2014 Dec 30.

Department of Orthopedic Surgery, Mayo Clinic, Rochester, MN.

Processed nerve allografts have become an alternative to repair segmental nerve defects, with results comparable with autografts regarding sensory recovery; however, they have failed to reproduce comparable motor recovery. The purpose of this study was to determine how revascularizaton of processed nerve allograft would affect motor recovery. Eighty-eight rats were divided in four groups of 22 animals each. A unilateral 10-mm sciatic nerve defect was repaired with allograft (group I), allograft wrapped with silicone conduit (group II), allograft augmented with vascular endothelial growth factor (group III), or autograft (group IV). Eight animals from each group were sacrificed at 3 days, and the remaining animals at 16 weeks. Revascularization was evaluated by measuring the graft capillary density at 3 days and 16 weeks. Measurements of ankle contracture, compound muscle action potential, tibialis anterior muscle weight and force, and nerve histomorphometry were performed at 16 weeks. All results were normalized to the contralateral side. The results of capillary density at 3 days were 0.99% ± 1.3% for group I, 0.33% ± 0.6% for group II, 0.05% ± 0.1% for group III, and 75.6% ± 45.7% for group IV. At 16 weeks, the results were 69.9% ± 22.4% for group I, 37.0% ± 16.6% for group II, 84.6% ± 46.6% for group III, and 108.3% ± 46.8% for group IV. The results of muscle force were 47.5% ± 14.4% for group I, 21.7% ± 13.5% for group II, 47.1% ± 7.9% for group III, and 54.4% ± 10.6% for group IV. The use of vascular endothelial growth factor in the fashion used in this study improved neither the nerve allograft short-term revascularization nor the functional motor recovery after 16 weeks. Blocking allograft vascularization from surrounding tissues was detrimental for motor recovery. The processed nerve allografts used in this study showed similar functional motor recovery compared with that of the autograft.
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http://dx.doi.org/10.1002/micr.22371DOI Listing
February 2016

Vascularized bone transplant chimerism mediated by vascular endothelial growth factor.

Microsurgery 2015 Jan 30;35(1):45-51. Epub 2014 Jul 30.

Department of Orthopedic Surgery, Microvascular Research Laboratory, Mayo Clinic, Rochester, MN.

Background: Vascular endothelial growth factor (VEGF) induces angiogenesis and osteogenesis in bone allotransplants. We aim to determine whether bone remodeling in VEGF-treated bone allotransplants results from repopulation with circulation-derived autogenous cells or survival of allogenic transplant-derived cells.

Methods: Vascularized femoral bone transplants were transplanted from female Dark Agouti rats (DA;RT1(a) ) to male Piebald Viral Glaxo (PVG;RT1(c) ). Arteriovenous bundle implantation and short-term immunosuppression were used to maintain cellular viability. VEGF was encapsulated in biodegradable microspheres and delivered intramedullary in the experimental group (n = 22). In the control group (n = 22), no VEGF was delivered. Rats were sacrificed at 4 or 18 weeks. Laser capture microdissection of bone remodeling areas was performed at the inner and outer cortex. Sex-mismatched genes were quantified with reverse transcription-polymerase chain reaction to determine the amount of male cells to total cells, defined as the relative expression ratio (rER).

Results: At 4 weeks, rER was significantly higher at the inner cortex in VEGF-treated transplants as compared to untreated transplants (0.622 ± 0.225 vs. 0.362 ± 0.081, P = 0.043). At 4 weeks, the outer cortex in the control group had a significantly higher rER (P = 0.038), whereas in the VEGF group, the inner cortex had a higher rER (P = 0.015). Over time, in the outer cortex the rER significantly increased to 0.634 ± 0.106 at 18 weeks in VEGF-treated rats (P = 0.049). At 18 weeks, the rER was >0.5 at all cortical areas in both groups.

Conclusions: These in vivo findings suggest a chemotactic effect of intramedullary applied VEGF on recipient-derived bone and could imply that more rapid angiogenesis of vascularized allotransplants can be established with microencapsulated VEGF.
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http://dx.doi.org/10.1002/micr.22300DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4308546PMC
January 2015

Surgical revascularization in structural orthotopic bone allograft increases bone remodeling.

Clin Orthop Relat Res 2014 Sep 11;472(9):2870-7. Epub 2014 Jul 11.

Microvascular Research Laboratory, Mayo Clinic, 200 First Street, SW, Rochester, MN, 55905, USA.

Background: Osseous defects reconstructed with cryopreserved structural allografts are poorly revascularized and therefore are prone to nonunion, infection, deterioration of mechanical properties, and fracture. Whether this can be mitigated by specific interventions such as intramedullary surgical revascularization has been incompletely evaluated.

Questions/purposes: We aimed to study surgical revascularization as a means to improve bone remodeling in cryopreserved allograft. Second, we questioned whether spatial histomorphometric differences occur in cortical bone areas after intramedullary surgical revascularization. Third, biomechanical properties of the graft-recipient construct in surgically revascularized allograft were compared with those of conventional allografts.

Methods: Allografts were harvested from 10 Brown Norway rats, cryopreserved, and transplanted orthotopically in a 10-mm defect in two groups of 10 Lewis rats each (major histocompatibility mismatch). In the control group, no surgical revascularization was performed, whereas in the experimental group, a saphenous arteriovenous bundle was transposed in the bone marrow cavity. Bone remodeling was measured with histomorphometry, histology, and microcomputed tomography at 16 weeks. Spatial differences were analyzed with histomorphometry. To determine biomechanical properties, load at failure and structural stiffness in bending were evaluated by the three-point bend testing. In both groups, normal values of the contralateral femur also were analyzed.

Results: Surgically revascularized allografts had increased bone remodeling (bone formation rate to bone surface ratio: 130 ± 47 µm(3)/µm(2)/year versus 44 ± 43 µm(3)/µm(2)/year, p = 0.006) and higher cortical osteocyte counts (18.6% ± 12.7% versus 3.1% ± 2.8%, p = 0.002) than nonrevascularized grafts. In nonrevascularized grafts, the bone formation rate to bone surface ratio was 35% of the contralateral normal values, whereas in surgically revascularized grafts, the bone formation rate to bone surface ratio in the grafts exceeded the contralateral values (110%). Microcomputed tomography did not show differences in bone volume between groups, however in both groups, bone volume was less in grafts compared with the contralateral femurs. Inner cortical bone formation rate to bone surface ratio was greater in surgically revascularized grafts (65 ± 30 µm(3)/µm(2)/year versus 13 ± 16 µm(3)/µm(2)/year in the control group, p = 0.012). Outer cortical bone formation rate to bone surface ratio also increased in surgically revascularized grafts (49 ± 31 µm(3)/µm(2)/year versus 19 ± 21 µm(3)/µm(2)/year, p = 0.032). No differences were found in load at failure and structural stiffness between both groups. In the control group, load at failure and structural stiffness were lower in grafts than in the contralateral femurs (p = 0.004 and p = 0.02, respectively). In the experimental group, surgically revascularized grafts also had lower load at failure and structural stiffness than the contralateral femurs (p = 0.008 and p = 0.02, respectively).

Conclusions: Surgical revascularization of large segmental allografts improved bone remodeling and viability without an adverse effect on total bone volume or bending strength and stiffness in this short-term analysis.

Clinical Relevance: Cryopreserved allografts remain largely necrotic and are associated with a high rate of complications. Surgical revascularization increases graft healing which could contribute to graft survival with time.
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http://dx.doi.org/10.1007/s11999-014-3658-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4117914PMC
September 2014

Does the addition of a nerve wrap to a motor nerve repair affect motor outcomes?

Microsurgery 2014 Oct 14;34(7):562-7. Epub 2014 May 14.

Department of Orthopedic Surgery, St. Vincent's Hospital, The Catholic University of Korea, Seoul, Korea.

The purpose of this study was to evaluate the effect of wrapping bioabsorbable nerve conduit around primary suture repair on motor nerve regeneration in a rat model. Forty rats were randomly divided into two experimental groups according to the type of repair of the rat sciatic nerve: group I had primary suture repair; group II had primary suture repair and bioabsorbable collagen nerve conduit (NeuraGen® 1.5 mm, Integra LifeSciences Corp., Plainsboro, NJ) wrapped around the repair. At 12 weeks, no significant differences in the percentage of recovery between the two groups were observed with respect to compound muscle action potentials, isometric muscle force, and muscle weight (P = 0.816, P = 0.698, P = 0.861, respectively). Histomorphometric analysis as compared to the non-operative sites was also not significantly different between the two groups in terms of number of myelinated axons, myelinated fiber area, and nerve fiber density (P = 0.368, P = 0.968, P = 0.071, respectively). Perineural scar tissue formation was greater in primary suture repair group (0.36 ± 0.15) than in primary repair plus conduit wrapping group (0.17 ± 0.08). This difference was statistically significant (P < 0.001). Wrapping bioabsorbable nerve conduit around primary nerve repair can decrease perineural scar tissue formation. Although the scar-decreasing effect of bioabsorbable nerve wrap does not translate into better motor nerve recovery in this study, it might have an effect on the functional outcome in humans where scar formation is much more evident than in rats.
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http://dx.doi.org/10.1002/micr.22274DOI Listing
October 2014

An experimental study to determine and correlate choline acetyltransferase assay with functional muscle testing after nerve injury.

J Neurosurg 2014 May 21;120(5):1125-30. Epub 2014 Feb 21.

Microvascular Research Laboratory, and.

OBJECT Choline acetyltransferase (ChAT) is an enzyme synthesized within the body of a motor neuron whose role is to form the neurotransmitter acetylcholine. Quantification of ChAT levels in motor or mixed nerves has been proposed to provide information regarding the viability of a proximal nerve stump for motor neurotization following brachial plexus injury. To do so requires information regarding normal ChAT levels and those in injured nerves, as well as the correlation of ChAT level determined at surgery with eventual motor recovery. The purpose of this study was to determine ChAT activity in the normal and injured sciatic/peroneal nerve in a rat model, evaluate the correlation between ChAT and motor recovery, find the relationship between ChAT activity and isometric muscle force, and elucidate the parallel between ChAT activity and acetylcholinesterase (AChE) activity. METHODS Sixty animals were divided into 3 groups. The sciatic nerves in Group 1 were transected without repair. Nerves in Group 2 were transected and repaired. Nerves in Group 3 sustained a crush injury followed by transection and reconstruction. All animals were allowed 12 weeks of recovery followed by evaluation of ChAT levels in the peroneal nerve, correlated with measures of maximal isometric tibialis anterior muscle force and muscle weight (the operated side normalized to the control side). Karnovsky AChE staining of peroneal nerve segments was also compared with radiochemical assay of ChAT activity in the same nerve. RESULTS A significant difference in the tibialis anterior isometric tetanic force and the tibialis anterior muscle weight index (TAMI) was noted between Group 1 and Groups 2 and 3 (p < 0.0001); no significant difference was found comparing Group 2 with Group 3. The correlation between the force measurement and the TAMI was 0.382. Both AChE measurement and ChAT activity demonstrated significantly fewer fibers in the operated nerve compared with the contralateral nerve. Intergroup variability could also be illustrated using these tests. The correlation coefficient between the isometric tetanic force measurement and the ChAT analysis in Groups 1 and 2 was 0.468. The correlation for the AChE staining and the isometric tetanic force measurement was 0.111. The correlation between the TAMI and the ChAT levels was 0.773. The correlation between the TAMI and the AChE-stained fibers was 0.640. Correlating AChE staining to the ChAT analysis produced a correlation of 0.712. CONCLUSIONS The great variability in all groups and weak correlations to the functional muscle assessments and the ChAT radiochemical assay made this technique an unreliable method of determining motor nerve viability.
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http://dx.doi.org/10.3171/2014.1.JNS122241DOI Listing
May 2014

Fibroblast growth factor-2 and vascular endothelial growth factor mediated augmentation of angiogenesis and bone formation in vascularized bone allotransplants.

Microsurgery 2014 May 3;34(4):301-7. Epub 2014 Jan 3.

Department of Orthopedic Surgery, Microvascular Research Laboratory, Mayo Clinic, Rochester, MN.

We previously demonstrated recipient-derived neoangiogenesis to maintain viability of living bone allogeneic transplants without long-term immunosuppression. The effect of cytokine delivery to enhance this process is studied. Vascularized femur transplantation was performed from Dark Agouti to Piebald Virol Glaxo rats. Poly(d,l-lactide-co-glycolide) microspheres loaded with buffer (N = 11), basic fibroblast growth factor (FGF2) (N = 10), vascular endothelial growth factor (VEGF) (N = 11), or both (N = 11) were inserted intramedullarly alongside a recipient-derived arteriovenous bundle. FK-506 was administered for 2 weeks. At 18 weeks, bone blood flow, microangiography, histologic, histomorphometric, and alkaline phosphatase measurements were performed. Bone blood flow was greater in the combined group than control and VEGF groups (P = 0.04). Capillary density was greater in the FGF2 group than in the VEGF and combined groups (P < 0.05). Bone viability, growth, and alkaline phosphatase activity did not vary significantly between groups. Neoangiogenesis in vascularized bone allotransplants is enhanced by angiogenic cytokine delivery, with results using FGF2 that are comparable to isotransplant from previous studies. Further studies are needed to achieve bone formation similar to isotransplants.
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http://dx.doi.org/10.1002/micr.22221DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3976711PMC
May 2014

Return of motor function after repair of a 3-cm gap in a rabbit peroneal nerve: a comparison of autograft, collagen conduit, and conduit filled with collagen-GAG matrix.

J Bone Joint Surg Am 2013 Nov;95(21):1952-8

Department of Orthopedic Surgery, Mayo Clinic, 200 First Street S.W., Rochester, MN 55905. E-mail address for A.Y. Shin:

Background: The purpose of this study was to evaluate the motor nerve recovery in a rabbit model after repair of a 3-cm gap in the peroneal nerve with a conduit filled with a collagen-GAG (glycosaminoglycan) matrix and compare the results with those after reconstruction with an autograft or an empty collagen conduit.

Methods: Forty-two male New Zealand rabbits were divided into three experimental groups. In each group, a unilateral 3-cm peroneal nerve defect was repaired with a nerve autograft, an empty collagen conduit, or a conduit filled with a collagen-GAG matrix. At six months, nerve regeneration was evaluated on the basis of the compound muscle action potentials, maximum isometric tetanic force, and wet muscle weight of the tibialis anterior muscle as well as nerve histomorphometry.

Results: The autograft group had significantly better motor recovery than the conduit groups. The empty collagen conduits and conduits filled with the collagen-GAG matrix led to results that were similar to each other.

Conclusions: On the basis of this rabbit model, autologous nerve grafting remains the gold standard in the reconstruction of 3-cm segmental motor nerve defects.

Clinical Relevance: Segmental motor nerve defects should be reconstructed with autograft nerves. The use of a collagen conduit filled with a collagen-GAG matrix for motor nerve reconstruction should be limited until additional animal studies are performed.
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http://dx.doi.org/10.2106/JBJS.M.00215DOI Listing
November 2013

Functional evaluation in the rat sciatic nerve defect model: a comparison of the sciatic functional index, ankle angles, and isometric tetanic force.

Plast Reconstr Surg 2013 Nov;132(5):1173-1180

Rochester, Minn.; and Seoul, Republic of Korea From the Department of Orthopedic Surgery, The Catholic University of Korea; and the Microvascular Research Laboratory and the Departments of Neurosurgery and Orthopedic Surgery, Mayo Clinic.

Background: The sciatic functional index has long been the standard method of assessing motor recovery in the rat sciatic nerve model. The relative subjective nature of the assessment has led to development of newer methods, including video gait analysis and quantitative measurement of isometric tetanic muscle force.

Methods: Forty male Lewis rats, each with a 10-mm segmental defect in the sciatic nerve, were divided randomly into two groups: rats in group I underwent repair with reversed autograft, and those in group II received a collagen conduit. Video gait analyses were performed at 0, 4, 8, and 12 weeks, and sciatic functional index and ankle angles in four different walking phases were recorded. Isometric tetanic force of the tibialis anterior muscle was also measured at 12 weeks and correlated with sciatic functional index and video gait analysis data.

Results: The sciatic functional index results did not correlate with isometric tetanic force. Significantly, the sciatic functional index could not be measured in 26 percent of the rats at 8 weeks and 59 percent of the rats at 12 weeks secondary to toe contractures. Among various ankle angle measurements, only the ankle angle in toe-off phase correlated well with isometric tetanic force.

Conclusions: Toe contractures occurred more frequently in rats with better nerve recovery, and interfered with evaluation of the motor recovery using the sciatic functional index method. Ankle angle in toe-off phase measured from video gait analysis is a useful parameter that reflects functional recovery of the muscle force.
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http://dx.doi.org/10.1097/PRS.0b013e3182a3bfebDOI Listing
November 2013

The effect of long-term relocation on child and adolescent survivors of Hurricane Katrina.

J Trauma Stress 2013 Oct 24;26(5):613-20. Epub 2013 Sep 24.

Department of Psychiatry, Louisiana State University Health Sciences Center, New Orleans, Louisiana, USA.

The current study is designed to increase knowledge of the effects of relocation and its association with longer-term psychological symptoms following disaster. Following clinical observations and in discussions held with school officials expressing concerns about relocated students, it was hypothesized that students who relocated to a different city following Hurricane Katrina in 2005 would have more symptoms of posttraumatic stress compared to students who returned to New Orleans. The effect of Hurricane Katrina relocation was assessed on a sample of child and adolescent survivors in 5th through 12th grades (N = 795). Students with Orleans Parish zip codes prior to Hurricane Katrina were categorized into relocation groupings: (a) relocated to Baton Rouge, (b) returned to prior zip code, and (c) moved to a different zip code within Orleans Parish. Overall results revealed more trauma symptoms for relocated students. Results also revealed that younger relocated students had fewer symptoms compared to older students. The opposite was found for students who returned to their same zip code, with older students having fewer symptoms. This study supports the need for school-based services not only in disaster areas, but also in schools where survivors tend to migrate.
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http://dx.doi.org/10.1002/jts.21837DOI Listing
October 2013