Background

Peripheral nerve injury (PNI) represents a major health problem that often leads to significant functional impairment and permanent disability, resulting in annual costs of billions of dollars (Noble et al., 1998; Robinson, 2000; Foster et al., 2019; Karsy et al., 2019). PNI can range from simple compression to injuries with an intervening gap, where the nerve is completely separated (Campbell, 2008; Robinson, 2004; Caillaud et al., 2019). While optimal functional recovery after crush injury (axonotmesis, loss of axon continuity with preservation of nerve sheath) occurs over a period of 4–6 weeks (Hare et al., 1992; Moore et al., 2012; Caillaud et al., 2019), functional recovery after a severed nerve injury (neurotmesis, where the nerve itself is transected) with microsurgical repair and grafting is poor (Zochodne, 2012; Faroni et al., 2015), and full functional recovery is rare.

The available treatment for a completely severed nerve is end-to-end repair, nerve grafting, nerve conduits, or nerve transfer (Siemionow and Brzezicki, 2009; Isaacs, 2013). Poor functional recovery after these highly advanced microsurgical and reconstructive techniques could be partly due to the failure of these techniques to address the complex cellular and molecular events associated with PNI and repair (Zochodne, 2012; Faroni et al., 2015). To improve functional results for patients with PNIs, it is important to investigate multiple critical factors that are intimately related to the success of nerve regeneration and target innervation. While experimental studies to uncover mechanistic insights of PNI are impossible and unethical in humans, we recently established a novel standardized peripheral nerve transection technique in mice, which we refer to as Stepwise Transection and fibrin Glue (STG) (Lee et al., 2020), to investigate the mechanisms underlying poor functional outcomes. In the STG method, the nerve is first incompletely cut to 80% of its width, to prevent gap formation. Then, fibrin glue is applied around the laceration site, and the remaining 20% of the nerve is completely transected before the complete clotting of the glue. The STG method demonstrated post-injury pathophysiological changes comparable with the gold-standard suture repair, in terms of nerve histology and functional recovery (Lee et al., 2020). The STG method decreased surgical difficulties and variability by avoiding microsurgical manipulations on the nerve, thus ensuring the reproducibility and reliability of this animal model. Our novel transection method has several advantages as an experimental model when compared with current gold standard end-to-end neurorrhaphy, including: (i) standardized gap distance without microsurgical suturing, (ii) simplicity and reproducibility with faster procedure time, and (iii) lack of confounding factors associated with multiple suturing sites.

The sciatic nerve crush injury in rodents is a century-old model, widely used to study peripheral nerve regeneration and the potential beneficial effects of novel therapeutic strategies. Although crush injury can recover well within weeks in animal models (Hare et al., 1992; Moore et al., 2012), severe crush injury in humans with neuroma formation requires surgical intervention (Tos et al., 2012; Caillaud et al., 2019). Different tools (jeweler’s forceps, forceps with custom jig, malleus nipper, or needle driver), forces (~5 MPa to ~190 MPa), and duration (3 sec to 10 min) are used to create crush injuries (Chen et al., 1992; Bridge et al., 1994; Alant et al., 2012; Savastano et al., 2014; Tseng et al., 2016; Geary et al., 2017; Yue et al., 2019; Wandling et al., 2021), and there are concerns regarding the severity of crush injury pressure and the reproducibility of the method used. There have been limited attempts to quantify the pressure applied during crush injury in rodents, using a thin cell load for digital pressure display (Alant et al., 2012; Chen et al., 1992), and pressure-sensitive films for later analysis (Geary et al., 2017). However, there is no documentation of a pressure monitoring device that can be readily used for real-time pressure measurements. Importantly, operator precision and inter-device reliability may limit the standardization of the pressure applied and reproducibly of the findings. To address this gap, we constructed a novel portable device comprised of an Arduino Uno microcontroller board and force sensitive resistor (Figure 1), capable of reporting the real-time pressure applied to a nerve during crush injury (Wandling et al., 2021). We believe that the ability to deliver a more precise and quantifiable crush injury pressure using our novel device will increase standardization of crush injury and its reliability, leading to increased reproducibility of findings in the literature.

The objective of this article is two-fold: (A) To present a novel standardized peripheral nerve transection method in mice, that uses fibrin glue for modeling peripheral nerve transection injury with a reproducible gap distance between the severed nerve ends (Figures 2–3), and (B) to document the step-wise description of constructing a pressure sensor device (Figures 4–17) for crush injury pressure measurements.