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Circulating MicroRNAs Expression During Nerve Allografting In A Mouse Model
Ching-Hua Hsieh, MD, PhD; Johnson Chia-Shen Yang
Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan
Purpose: Nerve allografting with FK506 immunosuppression is able to gain some functional recovery in a short nerve defect in human. Regeneration through the nerve allograft is also the key factor for a successful composite tissue transfer; however, how to monitor the immunologic status or detect the rejection of the transplanted nerve is barely understood. Expression of the microRNAs (miRNAs) in the tissue and blood had been reported to reflect the progress of some diseases. A few studies had also demonstrated a correlation of circulating miRNAs expression with the status of acute and chronic rejection in the transplanted organs. This study was designed to profile the circulating miRNAs expression during the nerve allografting.
Methods: C57BL6 mice receive 0.5 cm right sciatic nerve resection and subsequent 1 cm nerve isograft from other C57BL6 or allograft from BALB/C donor mice with or without subcutaneous 1mg/kg/day FK506 from the time 3 days before the surgery and till sacrifice at 3, 7, 10 d after transplantation. miRNAs extracted from the grafts and the blood were analysed using an miRNA array based on miRBASE v.16 (Phalanx miRNA OneArray) with the identified target miRNAs being validated and quantified with real-time PCR using spiked-in cel-miR-39 as a control. Infiltration of the T-lymphocytes into the nerve allograft by CD3 immunohistochemical stain and distribution of circulating Th1, Th2, Th17 and Treg cells measured by flow cytometry was used to reflect the immunological status.
Results: Compared to the control group with nerve isografting, up-regulation of circulating miR-320, miR-423-5p, and miR-762 was found in the mice receiving allograft without FK-506 immunosuppression, but not in those with FK-506 immunosuppression. The up-regulated circulating miRNA targets is different from those (miR-125b-3p, miR-672, miR-467b*) expressed from the nerve allograft. Notably, discontinue of the FK-506 treatment for those mice receiving nerve allograft will result in rejection of the grafted nerve and sustained up-regulation of miR-320, miR-423-5p, and miR-762 in the circulation at 2, 4 and 6 d later.
Conclusion: Expression of circulating miR-320, miR-423-5p, and miR-762 may reflect the immunological status of the deep buried nerve allograft and presents a less invasive method for monitor of nerve allografting.
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