Introduction
Acute intestinal ischemia is a life-threatening clinical condition with a high mortality rate due to the difficulty of diagnosis and lack of treatment. Animal studies demonstrated that the innate immune response, in particular the complement system, is a key mediator in intestinal ischemia reperfusion (IR)-induced tissue injury but this has not yet been investigated in humans. Therefore, this study aims to gain insight into the underlying mechanisms of complement-mediated injury in human using in vivo and in vitro models of intestinal IR.

Methods
In vivo intestinal IR was induced in a segment of jejunum (to be routinely resected) in patients (n=10) undergoing pancreaticoduodenectomy. RNAseq analysis was performed on intestinal crypt epithelium isolated from frozen tissue sections using laser capture microdissection and additional tissues sections were stained by immunohistochemistry. Intestinal IR was mimicked in human small intestinal organoids which were exposed to normoxia or hypoxia followed by reoxygenation (HR). Gene expression of complement factors (CFs) and regulators (CRs) was analyzed by qPCR.

Results
Pathway analysis of human intestinal crypt epithelium revealed substantial regulation of the complement pathway during IR. Gene expression of CFs CFD, C5AR1 and C5AR2 and CRs CD55, CD59 and Clusterin were significantly upregulated. This was accompanied by increased CD55 protein expression in the intestinal crypts. In hSIOs, significant upregulation of CD55 was detected after HR compared to control.

Conclusion
Our findings demonstrate substantial regulation of the complement pathway during intestinal IR in humans in vivo. Next to upregulation of crucial CFs, the prominent increase in CD55 expression in the intestinal crypt is anticipated to protect stem cells from autologous complement attack during injury. In hSIOs, we can recapitulate relevant findings of the human in vivo model, hence, this organoid model can be used to further study the protective effect of CD55 in complement-mediated injury in human intestinal IR.