Peripheral nerves play vital roles in both motor and sensory nervous systems (22). PNI can cause a variety of disorders such as permanent denervation, neuropathic pain, sensory disruptions, and movement limitations (23, 24). In recent years, PNI treatment has developed rapidly with the development of nerve transplantation and gene-and cell-based therapies. Interest in SCs has led to growing attention in the study of nerve cell protection and reduction of cell death in the early stages of PNI.
SCs, the principal glial cells in the PNS, not only promote PNS integrity (25) but also maintain the function and survival of PNS neurons (26). Dysfunction of SCs can cause axon demyelination, disorders of axonal regeneration, directional guidance of axons (25, 27–30); and neurodegenerative diseases (26). In the acute stage of PNI, apoptosis and necrosis are the most common forms of SC death. Inhibition of apoptosis can partially protect SCs (31–34), and numerous studies have focused on the regulation of SC apoptosis. However, inhibition of necrosis may also protect SCs.
Few investigations have focused on the role of necrosis, which is an accidental cell death caused by extreme physiological, mechanical, or thermal stress (35). The number of necrotic cells is determined by the severity of the injury, ischemia, edema, secondary inflammatory response, and other factors. Necroptosis and necrosis have similar morphological features, but necroptosis is a programmed form of necrotic cell death that can be regulated (36). Using the same necroptosis and apoptosis pathways, necrotic cell death can be triggered through a complex consisting of RIPK1 and RIPK3 in the absence of caspase8 activity (37, 38). Nec-1 was discovered as an inhibitor of this pathway, which inhibits the function of RIP1 through the necrotic kinase, RIP1/RIPK1 (12). Several studies have disrupted necroptosis to investigate the neuroprotective effects of Nec-1 and have identified its protective potential in various models (13, 39–42). However, little is known regarding the effects of PNI on necroptosis. Whether Nec-1 can boost cell protection during PNI remains unclear.
SC necrosis was observed in both the PNI and Nec-1 + PNI groups in the present study. Necrosis may lead to further damage to the peripheral nerve tissues after PNI. Sections of injured sciatic nerves with reduced vacuolization and necrosis were observed in the Nec-1 + PNI group compared with those in the PNI group. The present results showed that Nec-1 inhibited necrosis and protected nerves by preventing vacuolar, axonal, and myelin degeneration after PNI. RIP1 and RIP3 were downregulated in the Nec-1 + PNI group 24 h after PNI, indicating inhibition of necroptosis. The present study took samples 24 h after PNI, according to the results that SCs begin to dramatically decrease at this time (21). To date, few studies have focused on the regulation of necroptosis to investigate the neuroprotective effect of Nec-1 in PNS. The present study explored the role of Nec-1 in PNS and found that inhibition of necroptosis may protect SCs and axons.
The functional recovery of peripheral nerves is obstructed by posttraumatic inflammation and oxidative stress (4–7). Posttraumatic inflammation after PNI is also driven by various inflammatory factors such as TNF-a, IL-1β and IL-6 involved in nerve injury (43–45). Detection of cytokines at the lesion site is more accurate and convenient than detection in the serum for the prediction of inflammation after PNI; therefore, these cytokines were detected in the sciatic nerve tissue using ELISA kits. The results showed that TNF-α, IL-1β, and IL-6 levels at 24 h after injury were the lowest in the sham group, and the levels in the PNI group were higher than those in the Nec-1 + PNI group. Nec-1 has a positive effect on relieving inflammation; however, the underlying mechanism remains unclear. Whether Nec-1 inhibits cytokines directly or through inhibition of necrosis requires further investigation. Oxidative stress may also contribute to secondary injuries after PNI. SCs and macrophages express pro-inflammatory factors, leading to the expression of ROS at the injury site (46–49). MDA is the main oxidative stress-related enzyme; therefore, the concentration of MDA was measured after PNI. MDA concentrations were higher in the PNI group than in the Nec-1 + PNI group, with the lowest levels observed in the sham group. These results show that Nec-1 modulates inflammatory factors through the inhibition of oxidative stress. Mitochondria are energy production and ATP synthesis organelles that provide the basis for oxidative stress formation and are also the main targets of oxidative insults, leading to disruption of the normal cell cycle (50–52). As such, it was speculated that maintaining mitochondrial function may provide new insights into the treatment of PNI with Nec-1. The effects of Nec-1 on myelin and axonal regeneration should be further investigated in future studies.
The major limitation of the present research is that the neural function (behavioral and electrophysiological observations) and time point results after the initial injury were not investigated. Detailed neural function studies and long-term follow-up studies should be conducted in the future.
The present study showed that Nec-1 could protect SCs and axons by inhibiting necroptosis, which is conducive to axonal regeneration and repair of peripheral nerve myelin. Therefore, Nec-1 may decrease the incidence of peripheral nerve lesions. The present findings also provide evidence that Nec-1 could reduce necroptosis by inhibiting RIP1 and RIP3 recruitment and effectively reduce inflammation and ROS production in the early stages of PNI. In conclusion, Nec-1 alleviated of necroptosis may provide new insights into early stage treatment of peripheral nerve repair after PNI.