Cellular responses during kidney normothermic machine perfusion reflect graft outcomes

There are currently no molecular tools that can reliably predict post-transplant outcome of kidney grafts during normothermic machine perfusion (NMP). In this study we investigated whether the cellular responses of donor kidneys during NMP can predict post-transplant outcome via transcriptomic analyses. Snap-frozen biopsies of 28 kidneys were taken before and after 2-hour NMP. 6 kidneys were included for single nucleus RNA sequencing (snRNA-seq) and 22 were for quantitative real-time PCR (qPCR). snRNA-seq analyses revealed a total of 38,451 cells distributed in 11 distinct cell types. NMP induced an upregulation of genes for ATP production-related proteins, heat shock proteins (HSP), transporter proteins and proteins that prevent protein misfolding. These findings were confirmed by the qPCR analyses. We observed no significant gene differences between the delayed graft function (DGF) and non-DGF kidneys. However, after NMP, the expression of HSP was above average in 3 kidneys with primary non-function (PNF) and kidneys that experienced acute rejection in the first month after transplantation compared to well-functioning kidneys. Overall, this study demonstrates that 2-hour NMP impacts gene expression profiles of the majority of cell types of the kidney. It suggests that NMP may serve to predict kidney function post-transplantation by triggering differential gene expression patterns.