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Transcriptomic analysis of Kidney Machine normothermic perfusion and comparision to cold storage

ABSTRACT: Transcriptomic analysis of Kidney Machine normothermic perfusion and comparision to cold storage

PROVIDER: PRJNA497370 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR8074340_1.fastq.gz	Fastqsanger.gz
	SRR8074340_2.fastq.gz	Fastqsanger.gz
	SRR8074350_1.fastq.gz	Fastqsanger.gz
	SRR8074350_2.fastq.gz	Fastqsanger.gz
	SRR8074360_1.fastq.gz	Fastqsanger.gz

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Transcriptomic analysis of Kidney Machine normothermic perfusion and comparision to cold storage

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Mechanisms and treatment of cold-storage-induced microvascular obstructions in normothermic machine perfusion of human kidney

Project description:Human kidneys which were accepted for transplantation and subsequently not used were subjected to ex vivo normothermic machine perfusion. During perfusion it was found the tubular epithelia synthesized fibrinogen which led to red blood cell aggregation and pathologic plugging of renal microvasculature. Sequential ex vivo delivery of plasminogen and tissue plasminogen activator (tPA) during normothermic machine perfusion effectively lyses these fibrinogen-mediated plugs.

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Pan-caspase inhibition during normothermic machine perfusion of discarded livers mitigates ischemia-reperfusion injury

Project description:Access to liver transplantation is limited by a significant organ shortage. The recent introduction of machine perfusion technology allows surgeons to monitor and assess ex situ liver function prior to transplantation. However, many donated organs are of inadequate quality for transplant, though opportunities exist to rehabilitate organ function with adjunct therapeutics during normothermic machine perfusion. In this preclinical study, we targeted the apoptosis pathway as a potential method of improving hepatocellular function. Treatment of discarded human livers during normothermic perfusion with an irreversible pan-caspase inhibitor, emricasan, resulted in significant mitigation of ischemia-reperfusion injury at both the transcriptional and protein level. This was evidenced by significantly decreased circulating levels of the pro-inflammatory cytokines, interleukin-6, interleukin-8, and interferon-gamma, compared to control livers. Untreated livers also demonstrated transcriptional changes notable for enrichment in pathways involved in innate immunity, leukocyte migration, and cytokine-mediated signaling. Targeting of unregulated apoptosis may represent a viable therapeutic intervention for rehabilitating liver hepatocellular function during machine perfusion.

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Gene expression profiling of human livers during normothermic machine perfusion

Project description:Normothermic machine perfusion (NMP) has been successfully implemented in clinical routine of liver transplantation over the past years. However, little is known about the mechanisms how NMP impacts on the transcriptome of a human donor liver. We herein examined gene expression profiles in transplanted and non-transplanted livers over NMP time. 50 livers subjected to NMP were included in this study. 30 were transplanted after a maximum of 20 hours (h) perfusion, while 15 were discarded due to poor performance. Biopsies were collected befor eNP (PRE), 1h, 6h, 12h, 20h of NMP and after reperfusion. Next-generation sequencing was applied in liver biopsies to assess differential gene expression over perfusion time. Perfusate samples were collected regularly to monitor liver function. Comparison in differential gene expression between PRE and 20h NMP showed 415 upregulated and 727 downregulated genes. Most significantly upregulated genes were associated with extra cellular matrix organization, cell growth/differentiation processes and cytokine signaling. A set of genes were identified which were significantly differentially expressed and important for classification of non-transplanted vs transplanted biopsies, especially at 12 and 20h of NMP. A 7-gene-signature showed good separation already at 6H NMP, thereby CD274 (PD-L1) expression was ponted out as most important.

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Project description: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.

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