# Ex-vivo limb perfusion in military and civilian medicine: inspired by ex-vivo organ perfusion, pioneered for traumatic limb amputation and peripheral nerve regeneration

## TL;DR

A new ex-vivo limb perfusion system extends limb viability and supports nerve regeneration after traumatic amputation.

## Contribution

The first ex-vivo limb perfusion system (EVEP) that investigates peripheral nerve regeneration and mitigates ischemia damage.

## Key findings

- EVEP reduces serum injury markers and preserves joint mobility when medication is used.
- Medication group showed lower pro-regenerative cytokine levels linked to Wallerian degeneration.
- EVEP creates a pro-regenerative environment for nerve recovery by omitting anti-inflammatory drugs.

## Abstract

Traumatic amputations have increased worldwide over the past two decades and are expected to increase by 72% by 2050. Surgical replantation provides superior functional recovery and patient satisfaction but is limited to specialized centers and restricted by short ischemia times, due to life-over-limb prioritization in patient care. To overcome these limitations, we developed an ex vivo limb perfusion system (EVEP) to extend limb viability and, for the first time, investigate its impact on peripheral nerve regeneration, a key prerequisite for functional recovery following replantation.

Hind limbs of 6 healthy pigs were amputated, and after 2 h of warm ischemia, limbs were either perfused normothermally for 6 h with PerfadexPlus® ± medication using in-house developed EVEP or stored statically (4 °C vs. room temperature). Perfusion parameters, blood gas analysis, serum markers, cytokine levels, thermal imaging, colloid oncotic pressure, weight gain, joint mobility, peripheral nerve histomorphometric and stereological analyses were performed.

Data confirm a valid and reliable EVEP with an optimized perfusion protocol. Comparison of perfusion groups revealed lower serum injury markers in the medication group, which included methylprednisolone treatment. Additionally, the medication group exhibited reduced weight gain and preserved unrestricted joint mobility, but concurrently led to a significant decrease in pro-regenerative cytokine levels associated with Wallerian degeneration (WD).

In general, EVEP mitigates ischemia-related damage and facilitates ex vivo induction of WD, a critical prerequisite for nerve regeneration, functional recovery, and prevention of neuroma formation with subsequent phantom pain, by establishing the pro-regenerative environment for WD, which is further amplified by omitting the anti-inflammatory methylprednisolone.

The online version contains supplementary material available at 10.1186/s40779-025-00656-6.