The essential findings of the present study are that hepatic I/R injury can induce remote kidney injury and that VNS treatment protects against AKI after hepatic I/R injury. Notably, we demonstrated for the first time that VNS significantly attenuated inflammation, oxidative stress, and apoptosis in the kidneys during the hepatic I/R process. Additionally, VNS was firstly noted to facilitate the activation of the Nrf2/HO-1 pathway in the kidneys after hepatic I/R injury.
AKI, also regarded as acute renal failure, is characterized by a sudden reduction in renal function or glomerular filtration rate, evolving from early injury to severe damage 6. It can result in chronic kidney disease (CKD) or even kidney failure, requiring kidney replacement therapy. Population data suggests that in liver recipients who experienced postoperative acute kidney failure, the risk of developing CKD increased by 100% 13. The kidneys are vulnerable to a series of challenges from remote hepatic I/R injury, owing to their abundant blood perfusion. Animal and clinical evidence of causality between hepatic I/R injury and AKI has been well documented. In rodents, hepatic I/R injury has been proven to cause AKI 14. In humans, a large single-center, case-controlled study has shown the same trend in increasing mortality and morbidity of AKI during the perioperative period 5. Previous findings support a strong relationship between hepatic I/R injury and AKI. Therefore, it is meaningful to progress an effective approach to arrest the progression of AKI.
From an anatomical point of view, except for the adrenal glands, the kidneys receive a more extensive nerve supply than any other abdominal organ 15. The kidney arteries were sympathetically innervated because of the numerous kidney sympathetic nerves close to the lumen of the kidney arteries 16. The pathogenesis of AKI was associated with the heightened activity of the sympathetic nervous system 17. This is consistent with a previous study where in the initial 24 h-period following hepatic I/R injury, the occurrence of AKI was attributed to intrarenal vasoconstriction resulting from splanchnic vasodilatation following portal hypertension 18. These observations indicate that activation of the sympathetic nervous system negatively influences kidney function. The activation of the vagus nerve system, which naturally offsets sympathetic vasoconstrictive activity, is an optional approach to restore autonomic regulatory function. Inoue et al. verified that 24 h before kidney ischemia, VNS markedly attenuated AKI and inhibited systemic inflammation through α7 nicotinic acetylcholine receptor (α7nAChR)-positive splenocytes 11. Hence, we originally proposed the hypothesis that VNS treatment might protect against AKI after hepatic I/R injury. It has already been revealed that mice subjected to severe hepatic I/R injury develop AKI characterized by immediate kidney peritubular capillary endothelial cell apoptosis with subsequent kidney proximal tubular necrosis and inflammation 18. Therefore, we explored whether VNS could block these pathways to exert protective effects against AKI induced after hepatic I/R injury.
In the development of hepatic I/R injury-induced AKI, a substantial systematic inflammatory response always bears the brunt. It is widely known that acute liver injury following hepatic I/R injury can cause a systemic inflammatory response, resulting in remote organ injury 19. Park et al. observed that hepatic I/R injury was attributed to a significant upregulation of proinflammatory cytokines in the kidneys 20. Furthermore, Koopman et al. reported that the approach for VNS to mitigate inflammatory diseases was to inhibit the production of proinflammatory cytokines 21. The VNS-mediated cholinergic anti-inflammatory pathway has been well described in the kidneys 9. A similar finding was reported by Okusa et al., indicating that the previously-described pathway has the potential to limit ischemic acute kidney injury 22. Recently, several researchers have applied vagal anti-inflammatory effects in remote organ injury after I/R injury. Lai et al. reported that VNS exhibited a protective effect in acute liver injury after kidney I/R injury by reducing the release of various inflammatory cytokines (e.g., TNF-α and IL-6) 12. Consistent with these previous studies, our data suggests that VNS greatly ameliorated inflammation in the kidneys after hepatic I/R injury.
Oxidative stress is another crucial contributor to the pathogenesis of AKI. During I/R, reactive oxygen species (ROS) are massively released and various ROS subsequently initiate lipid peroxidation (LP) reactions, contributing to inflammation activation and tissue damage. MDA serves as an oxidative stress marker and has been proven to increase in kidneys after hepatic I/R injury 18. A range of studies reveal that when AKI develops, systemic oxidative stress greatly increases, whether in humans or animals 23,24. In addition, a subsequent study further demonstrated that hepatic I/R injury can acutely induce a remote renal cortical oxidative stress response 25. Accumulating evidence from past studies suggested that VNS can reduce oxidative stress in multiple organs 9,26. Our study showed that VNS increased GSH levels and SOD activity, and decreased MDA levels and MPO activity in the kidneys after hepatic I/R injury. Based on the above, our results suggest that VNS protected against hepatic I/R injury-induced AKI, partly through its antioxidative properties.
Kidney apoptosis is a hallmark of AKI 27. Cellular apoptosis is a recognized pathological characteristic in the literature. A previous study revealed that VNS exerted protective effects on the liver after kidney I/R injury, probably by suppressing cellular apoptosis 12. Kidney is another important organ of the liver-kidney axis, compared to the previous study, our data showed that VNS exhibited the similar protective effect in the kidneys after hepatic I/R injury. Our study is the first to provide evidence and support the concept that VNS can alleviate apoptosis in the kidneys after hepatic I/R injury.
Crosstalk between organs affects each other’s functioning via various pathways, including endocrine, neural, and direct cell-cell signaling pathways 28. The pathophysiology of hepatic I/R injury-induced AKI is complex and multifactorial, with inflammation, oxidative stress, apoptosis, and activation of various signaling pathways. Nuclear factor erythroid 2-related factor 2 (Nrf2), a transcription factor, mediates cellular defense against oxidative stress in organs. Under physiological cellular conditions, Nrf2 is sequestered in the cytoplasm in association with its protein inhibitor, Kelth-like ECH-associated protein-1 (Keap-1). Under pathological cellular conditions, Nrf2 is transferred into the nucleus where it upregulates relevant cytoprotective enzymes, such as heme oxygenase-1 (HO-1) 29. Kudoh et al. reported that in a rat model of hepatic I/R injury, the depletion of Nrf2 aggravated inflammation and lead to oxidative stress and apoptosis 30. It is widely accepted that Nrf2 activation induces HO-1 transcription, which has been demonstrated to be closely involved in alleviating AKI by minimizing cellular oxidative stress in recent finding 31. α7nAChR is the target of the cholinergic anti-inflammatory pathway. Several studies have found that α7nAChR activation can directly induce the Nrf2/HO-1 pathway 32,33. Navarro et al. further demonstrated that the deletion of Nrf2 removed the protective effect of the α7nAChR agonist 33. As noted above, the Nrf2/HO-1 pathway may be an important signaling pathway for the protection of VNS in AKI. Consistent with previous findings, our data showed that VNS strongly enhanced the expression levels of Nrf2/HO-1. It is possible to conclude that the Nrf2/HO-1 pathway may serve as an important mediator of the protective effects of VNS in the setting of AKI following hepatic I/R injury.
Liver transplantation is the only definitive long-term treatment for end-stage liver disease and hepatic malignant tumors, with implementation beginning in the 1950s 34. AKI is a common clinical complication in liver recipients and is considered to be a lethal threat. The data from our study show that VNS may be a potential clinical treatment for hepatic I/R injury-induced AKI. Traditional VNS is always invasive and requires device implantation; therefore, the application of VNS is widely restricted. Auricular VNS, a non-invasive VNS, has been shown to obtain the similar effects as invasive VNS 35. Considering these observations together, non-invasive VNS might be a prospective clinical treatment for hepatic I/R injury-induced AKI.
There are limitations to our study which ought to be mentioned. First, we chose pentobarbital to anesthetize all experimental animals, which may affect the autonomic nervous system function. Second, different VNS frequencies, intensities, and durations have been shown to exert different therapeutic effects. However, only one stimulation parameter was used in the present study. It is unclear if there is an enhanced effect with a different parameter. Third, this experiment was tested in only one animal model, and extrapolation from rats to humans can be difficult.
In conclusion, our data suggest that VNS can protect against AKI after hepatic I/R injury. The potential mechanisms may involve inhibiting inflammation, suppressing oxidative stress, and reducing apoptosis. Additionally, VNS greatly activated the Nrf2/HO-1 pathway in the kidneys. Taken together, VNS may attenuate hepatic I/R injury-induced AKI by suppressing inflammation, oxidative stress, and apoptosis probably via the Nrf2/HO-1 pathway (see Fig. 8). With the development of non-invasive VNS, VNS might provide a novel clinical treatment for patients with hepatic I/R injury-induced AKI.