Open Access Research

Innate immunity activation involved in unprotected porcine auto-transplant kidneys preserved by naked caspase-3 siRNA

Cheng Yang1, Long Li1, Yinjia Xue1, Zitong Zhao1, Tian Zhao1, Yichen Jia1, Ruiming Rong14, Ming Xu1, Michael L Nicholson2*, Tongyu Zhu15* and Bin Yang23*

Author Affiliations

1 Department of Urology, Zhongshan Hospital, Fudan University; Shanghai Key Laboratory of Organ Transplantation, 180 Fenglin Road, Shanghai 200032, PR China

2 Transplant Group, Department of Infection, Immunity and Inflammation, University of Leicester; Leicester General Hospital, University Hospitals of Leicester, Gwendolen Road, Leicester LE5 4PW, UK

3 Department of Nephrology, Affiliated Hospital of Nantong University, Nantong, PR China

4 Department of Transfusion, Zhongshan Hospital, Fudan University, Shanghai, PR China

5 Qingpu Branch Zhongshan Hospital, Fudan University, Shanghai, PR China

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Journal of Translational Medicine 2013, 11:210  doi:10.1186/1479-5876-11-210

Published: 13 September 2013

Abstract

Background

The naked caspase-3 small interfering RNA (siRNA) infused into the renal artery during cold preservation was effective, but did not protect auto-transplant porcine kidneys with increased inflammation and apoptosis in our previous study. The mechanisms involved, in particular, whether siRNA or complementary systemic feedback eliciting innate immune responses are worthy to be further investigated.

Methods

The protein and mRNA expression of innate immunity-related molecules were detected by western blotting and quantitative PCR in the tissues previously collected from 48 h auto-transplant kidneys. The donor kidneys were retrieved from mini pigs and cold preserved by University of Wisconsin solution with/without 0.3 mg caspase-3 siRNA for 24 h.

Results

The protein level of Toll like receptor (TLR) 3, TLR7, and their main adapters, TRIF and MyD88, was up-regulated in the siRNA preserved auto-transplant kidneys. The mRNA level of NF-κB and c-Jun was increased, as well as pro-inflammatory cytokines, including IL-1β, IL-6, TNF-α and interferon (IFN)-α, β and γ. In addition, the non-TLR RNA sensor PKR protein, but not RIG1, was also increased in the siRNA preserved auto-transplant kidneys.

Conclusions

The activation of innate immunity with amplified inflammatory responses in the caspase-3 siRNA preserved auto-transplant kidneys are associated with increased TLR3, TLR7 and PKR, which might be due to complementary systemic feedback, although persistent actions initiated by short-acting caspase-3 siRNA cannot be completely ruled out. These results provided valuable evidence to guide future siRNA design and pre-clinic studies.

Keywords:
Small interfering RNA; Innate immunity; Toll like receptor; PKR; Porcine kidney auto-transplantation; Ischemia reperfusion injury