A Skin Rejection Grading System for Vascularized Composite Allotransplantation in a Preclinical Large Animal Model

Etra, Joanna W.; Grzelak, Michael; Fidder, Samuel A.J.; Kolegraff, Keli; Bonawitz, Steven C.; Shores, Jaimie T.; Oh, Byoungchol; Cooney, Damon S.; Beck, Sarah E.; Brandacher, Gerald

doi:10.1097/tp.0000000000002695

Cited by 24 publications

(18 citation statements)

References 25 publications

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“…Neutrophilic granulocytes showed a trend towards a higher infiltration after CHSA implantation on day 3. However, neutrophilic infiltrates are not considered specific to the pathogenesis of skin graft rejection and rather represent a generic acute inflammatory response …”

Section: Discussionmentioning

confidence: 99%

Cryopreserved human skin allografts promote angiogenesis and dermal regeneration in a murine model

Henn

Chen

Maan

et al. 2020

International Wound Journal

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Cryopreserved human skin allografts (CHSAs) are used for the coverage of major burns when donor sites for autografts are insufficiently available and have clinically shown beneficial effects on chronic non‐healing wounds. However, the biologic mechanisms behind the regenerative properties of CHSA remain elusive. Furthermore, the impact of cryopreservation on the immunogenicity of CHSA has not been thoroughly investigated and raised concerns with regard to their clinical application. To investigate the importance and fate of living cells, we compared cryopreserved CHSA with human acellular dermal matrix (ADM) grafts in which living cells had been removed by chemical processing. Both grafts were subcutaneously implanted into C57BL/6 mice and explanted after 1, 3, 7, and 28 days (n = 5 per group). A sham surgery where no graft was implanted served as a control. Transmission electron microscopy (TEM) and flow cytometry were used to characterise the ultrastructure and cells within CHSA before implantation. Immunofluorescent staining of tissue sections was used to determine the immune reaction against the implanted grafts, the rate of apoptotic cells, and vascularisation as well as collagen content of the overlaying murine dermis. Digital quantification of collagen fibre alignment on tissue sections was used to quantify the degree of fibrosis within the murine dermis. A substantial population of live human cells with intact organelles was identified in CHSA prior to implantation. Subcutaneous pockets with implanted xenografts or ADMs healed without clinically apparent rejection and with a similar cellular immune response. CHSA implantation largely preserved the cellularity of the overlying murine dermis, whereas ADM was associated with a significantly higher rate of cellular apoptosis, identified by cleaved caspase‐3 staining, and a stronger dendritic cell infiltration of the murine dermis. CHSA was found to induce a local angiogenic response, leading to significantly more vascularisation of the murine dermis compared with ADM and sham surgery on day 7. By day 28, aggregate collagen‐1 content within the murine dermis was greater following CHSA implantation compared with ADM. Collagen fibre alignment of the murine dermis, correlating with the degree of fibrosis, was significantly greater in the ADM group, whereas CHSA maintained the characteristic basket weave pattern of the native murine dermis. Our data indicate that CHSAs promote angiogenesis and collagen‐1 production without eliciting a significant fibrotic response in a xenograft model. These findings may provide insight into the beneficial effects clinically observed after treatment of chronic wounds and burns with CHSA.

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Section: Discussionmentioning

confidence: 99%

Cryopreserved human skin allografts promote angiogenesis and dermal regeneration in a murine model

Henn

Chen

Maan

et al. 2020

International Wound Journal

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“…Tape stripping was further performed on the back of naïve Massachusetts General Hospital (MGH) miniature swine (N S ; n = 8), on the native skin (back) (T S ; n = 6) or the skin component of tacrolimus‐treated nonrejecting heterotopic hind limb allografts (G S ; n = 5) in swine that had received an allograft, and on the skin component of untreated rejecting heterotopic hind limb allografts (R S ; n = 9) of MGH miniature swine (Table S2). Clinical rejection was scored based on a previously published scoring system [23] Swine hind limb transplantations were performed according to a prior published model [24]…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

“…Tape stripping was further performed on the back of na€ ıve Massachusetts General Hospital (MGH) miniature swine (N S ; n = 8), on the native skin (back) (T S ; n = 6) or the skin component of tacrolimus-treated nonrejecting heterotopic hind limb allografts (G S ; n = 5) in swine that had received an allograft, and on the skin component of untreated rejecting heterotopic hind limb allografts (R S ; n = 9) of MGH miniature swine (Table S2). Clinical rejection was scored based on a previously published scoring system [23] Swine hind limb transplantations were performed according to a prior published model [24] Protein extraction buffer For extraction of proteins from the D-Squame sampling disks, a protein extraction buffer containing phosphate buffered saline (1%; Sigma-Aldrich, St. Louis, MO, USA), 5% polypropylene glycol (LOT# 0731C130, Amrescoâ, Solon, OH, USA), 0.1% sodium dodecyl sulfate (SDS 20% solution, LOT# 1711C070, Amrescoâ, Solon, OH, USA), and one proteinase inhibitor cocktail tablet (Cat. No.…”

Section: Experimental Groupsmentioning

confidence: 99%

“…Fixed tissues were then embedded in paraffin, sectioned at 5µm, and stained with hematoxylin and eosin (H&E). All slides were reviewed and scored by an expert veterinary pathologist in a blinded fashion based on a previously published scoring system [23] Statistical analysis Prism GraphPad 7.0 (GraphPad Inc., San Diego, CA, USA) and the statistical software environment R 3.6.1 (The R Foundation for Statistical Computing, Vienna, Austria) were used for statistical analysis. Results are expressed as median and interquartile range (IQR).…”

Section: Histologymentioning

confidence: 99%

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Noninvasive evaluation of intragraft immune responses in upper extremity transplantation

et al. 2021

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In vascularized composite allotransplantation (VCA), invasive tissue biopsies remain the gold standard in diagnosing rejection carrying significant morbidity. We aimed to show feasibility of tape-stripping for noninvasive immune monitoring in VCA. Tape-stripping was performed on allografts and native skin of upper extremity transplant recipients. Healthy nontransplanted individuals served as controls. The technique was also used in swine on na€ ıve skin in nontransplanted animals, native skin of treated, transplanted swine, nonrejecting VCAs, and rejecting VCAs. Extracted protein was analyzed for differences in cytokine expression using Luminex technology. Significantly decreased levels of INFc and IL-1Ra were seen between human allograft samples and native skin. In swine, rejecting grafts had increased IL-1Ra compared to na€ ıve and native skin, decreased levels of GM-CSF compared to native skin, and decreased IL-10 compared to nonrejecting grafts. Unsupervised hierarchical clustering revealed rejecting grafts separated from the nonrejecting (P = 0.021). Variable importance in projection scores identified GM-CSF, IL-1Ra, and IL-2 as the most important profiles for group discrimination. Differences in cytokine expression are detectable in human VCA patient native skin and VCA graft skin using a noninvasive tape-stripping method. Swine studies suggest that differences in cytokines between rejecting and nonrejecting grafts are discernable.

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Heterotopic intestine and abdominal wall transplantation in a miniature swine: A model for graft immunology

Kuo

Chen

Yeh

et al. 2023

The Kaohsiung J of Med Scie

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In order to accurately monitor graft immunology, we have developed a method for performing intestine and abdominal wall transplantation heterotopically in miniature swine. The procedure consisted of simultaneous segmental terminal ileum and full‐thickness abdominal wall transplantation in Lanyu miniature swine, with the intestinal and the abdominal wall grafts being placed on the recipient's bilateral rear flank. Five transplantations were technically successful. One animal died on the first post‐transplant day due to anesthesia‐related issues, three abdominal wall and four intestinal grafts survived, while one abdominal wall graft failed due to vascular thrombosis. Acute cellular rejection (ACR) of the intestinal graft could occur preceding, simultaneously with or following ACR of the abdominal wall graft. Our experimental model demonstrates the technical feasibility of heterotopic intestine and abdominal wall transplantation in miniature swine without grafting in gastrointestinal continuity. This model could be suitable for further studies of graft immunology.

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A Skin Rejection Grading System for Vascularized Composite Allotransplantation in a Preclinical Large Animal Model

Abstract: Supplemental Digital Content is available in the text.

Cited by 24 publications

References 25 publications

Cryopreserved human skin allografts promote angiogenesis and dermal regeneration in a murine model

Cryopreserved human skin allografts promote angiogenesis and dermal regeneration in a murine model

Noninvasive evaluation of intragraft immune responses in upper extremity transplantation

Heterotopic intestine and abdominal wall transplantation in a miniature swine: A model for graft immunology

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