The reversed paradigm of chimerism induction: Donor conditioning with recipient‐derived bone marrow cells as a novel approach for tolerance induction in vascularized composite allotransplantation

Siemionow, Maria; Rampazzo, Antonio; Gharb, Bahar Bassiri; Cwykiel, Joanna; Klimczak, Aleksandra; Madajka, Maria; Nasır, Serdar; Bozkurt, Mehmet

doi:10.1002/micr.30041

Cited by 15 publications

(19 citation statements)

References 25 publications

(29 reference statements)

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“…Additionally, however, one may postulate that the vascularized donor BM (DBM) compartment of a VCA, such as the orthotopic hind limb transplant used in this study, plays a critical role in achieving long-term graft survival. As recently reported, the BM niche is a dynamic environment rich in a variety of cells and has shown a significant survival benefit in small and large animal models (25,32,39). In line with these findings, our results suggest that this VCA intrinsic feature -a viable BM component -allows for induction of stable multilineage mixed chimerism and long-term graft survival by TBI and CoB-based immunomodulation without the need for short-or long-term conventional maintenance immunosuppression.…”

Section: Discussionsupporting

confidence: 89%

Vascularized composite allotransplantation combined with costimulation blockade induces mixed chimerism and reveals intrinsic tolerogenic potential

Oh¹,

Furtmüller²,

Fryer³

et al. 2020

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Section: Discussionsupporting

confidence: 89%

Vascularized composite allotransplantation combined with costimulation blockade induces mixed chimerism and reveals intrinsic tolerogenic potential

Oh¹,

Furtmüller²,

Fryer³

et al. 2020

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“…Forty-eight male 7–8 weeks old ACI (RT1 a ) and 48 Lewis (RT1 l ) rats served as donors for the creation of DRCC therapy (Envigo, USA). BMC were isolated in a sterile manner, as previously described (Cwykiel et al 2021 ; Hivelin et al 2016 ; Siemionow et al 2016 ), from femurs and tibias using flashing technique. Harvested cells were filtered using 40 µm strainer, purified by Histopaque 1084 (Millipore–Sigma, USA) and counted with 0.4% Trypan Blue (Thermo Fisher Scientific, USA).…”

Section: Methodsmentioning

confidence: 99%

Donor Recipient Chimeric Cells Induce Chimerism and Extend Survival of Vascularized Composite Allografts

Cwykiel

Jundziłł

Klimczak

et al. 2021

Arch. Immunol. Ther. Exp.

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This study evaluated the efficacy of donor recipient chimeric cell (DRCC) therapy created by fusion of donor and recipient derived bone marrow cells (BMC) in chimerism and tolerance induction in a rat vascularized composite allograft (VCA) model. Twenty-four VCA (groin flaps) from MHC-mismatched ACI (RT1a) donors were transplanted to Lewis (RT1l) recipients. Rats were randomly divided into (n = 6/group): Group 1—untreated controls, Groups 2—7-day immunosuppression controls, Group 3—DRCC, and Group 4—DRCC with 7-day anti-αβTCR monoclonal antibody and cyclosporine A protocol. DRCC created by polyethylene glycol-mediated fusion of ACI and Lewis BMC were cultured and transplanted (2–4 × 106) to VCA recipients via intraosseous delivery route. Flow cytometry assessed peripheral blood chimerism while fluorescent microscopy and PCR tested the presence of DRCC in the recipient’s blood, bone marrow (BM), and lymphoid organs at the study endpoint (VCA rejection). No complications were observed after DRCC intraosseous delivery. Group 4 presented the longest average VCA survival (79.3 ± 30.9 days) followed by Group 2 (53.3 ± 13.6 days), Group 3 (18 ± 7.5 days), and Group 1 (8.5 ± 1 days). The highest chimerism level was detected in Group 4 (57.9 ± 6.2%) at day 7 post-transplant. The chimerism declined at day 21 post-transplant and remained at 10% level during the entire follow-up period. Single dose of DRCC therapy induced long-term multilineage chimerism and extended VCA survival. DRCC introduces a novel concept of customized donor-recipient cell-based therapy supporting solid organ and VCA transplants.

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“…All animals received humane care in compliance with the 'Principles of laboratory animal care' formulated by the National Society for Medical Research and the 'Guide for the care and use of laboratory animal resources' published by the US National Institute of Health. BMC were isolated from femurs and tibias harvested from randomly selected 7-8-week old male ACI (RT1 a , n=30) and Lewis (RT1 l , n=30) donors (Envigo, Indianapolis, IN, USA) in a sterile manner using flashing technique (22,23). Harvested cells were filtered using 40µm strainer, purified by Histopaque 1083 (MiliporeSigma, St. Louis, MO, USA) and counted with 0.4% Trypan Blue (ThermoFisher Scientific, Waltham, MA, USA).…”

Section: Bmc Isolationmentioning

confidence: 99%

“…BMC from ACI (RT1 a ) and Lewis (RT1 l ) donors were stained using PKH26 (ACI) or PKH67 (Lewis) fluorescent dyes (MiliporeSigma). Staining was performed as previously reported (22,24,25) with the adjusted staining time of 3 min.…”

Section: Fusion Proceduresmentioning

confidence: 99%

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Development of Donor Recipient Chimeric Cells of bone marrow origin as a novel approach for tolerance induction in transplantation

2021

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Background: Cell therapies and chimerism-based strategies are currently the most successful approach for tolerance induction in transplantation. This study aimed to establish and characterize novel Donor Recipient Chimeric Ccell (DRCC) therapy of bone marrow (BM) origin presenting donor-recipient phenotype to support tolerance induction. Methods: Ex vivo fusions of fully MHC-mismatched BM cells from ACI (RT1 a ) and Lewis (RT1 l ) rats were performed using polyethylene-glycol (PEG). The creation of rat DRCC was tested by flow cytometry (FC), confocal microscopy and PCR. FC characterized DRCC's phenotype (CD3, CD4, CD8, CD45, CD90, CD11b/c, CD45RA, OX-82, or CD4/CD25) and apoptosis, while mixed lymphocyte reaction assessed DRCC's immunogenicity and colony forming unit assay tested DRCC's differentiation and proliferation. DRCC's polyploidy was evaluated using Hoechst33342 staining and COMET assay tested genotoxicity of fusion procedure. ELISA analyzed the secretion of IL-2, IL-4, IL-10, TGFß1, IFNγ and TNFα by DRCC at day 1, 5 and 14 post-fusion. The DRCC's phenotype after long-term culturing was assessed by reversetranscription PCR.Results: The chimeric state of DRCC was confirmed. Fusion did not change the expression of hematopoietic markers compared to BM controls. Although an increased number of early and late apoptotic (Annexin V + /Sytox blueand Annexin V + /Sytox blue + , respectively) DRCC was detected at 24h postfusion, the number significantly decreased at day 5 (38.4%±3.1% and 22.6%±2.5%, vs. 28.3%±2.5% and 13.9%±2.6%, respectively, P<0.05). DRCC presented decreased immunogenicity, increased expression of IL-10 and TGFβ1 and proliferative potential comparable to BM controls. The average percentage of tetraploid DRCC was 3.1%±0.2% compared to 0.96%±0.1% in BM controls. The lack of damage to the DRCC's DNA content supported the DRCC's safety. In culture, DRCC maintained proliferation for up to 28 days while preserving hematopoietic profile.Conclusions: This study confirmed feasibility of DRCC creation via ex vivo PEG mediated fusion. The created DRCC revealed pro-tolerogenic properties indicating potential immunomodulatory effect of DRCC therapy when applied in vivo to support tolerance induction in solid organ and vascularized composite allograft transplantation.

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The reversed paradigm of chimerism induction: Donor conditioning with recipient‐derived bone marrow cells as a novel approach for tolerance induction in vascularized composite allotransplantation

Abstract: Donor preconditioning is a novel approach modifying recipient's responsiveness to donor allograft and prolonging the allograft survival under short-term immunosuppression. © 2015 Wiley Periodicals, Inc. Microsurgery 36:676-683, 2016.

Cited by 15 publications

References 25 publications

Vascularized composite allotransplantation combined with costimulation blockade induces mixed chimerism and reveals intrinsic tolerogenic potential

Vascularized composite allotransplantation combined with costimulation blockade induces mixed chimerism and reveals intrinsic tolerogenic potential

Donor Recipient Chimeric Cells Induce Chimerism and Extend Survival of Vascularized Composite Allografts

Development of Donor Recipient Chimeric Cells of bone marrow origin as a novel approach for tolerance induction in transplantation

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