Substantial evidence has highlighted the crucial role of sustained inflammation in the pathogenesis of Type 2 diabetes (T2D) during the past decades. It is well established that liver inflammation caused by high fat diet (HFD) and obesity impairs hepatic glucose and lipid metabolism, leading to local and systemic insulin resistance, ultimately contributing to the development of T2D [1
]. In particular, accumulating evidence suggests that inhibition of hepatic inflammatory response improves HFD and obesity-induced metabolic disorders, such as insulin resistance and hyperglycemia [3
]. Though the role of liver inflammation in the development of T2D has been widely investigated, the triggering mechanisms underlying HFD and obesity-induced hepatic inflammation remain to be elucidated.
Nicotinamide adenine dinucleotide (NAD+
), in addition to acting as a coenzyme, has been demonstrated to act as a key mediator in regulating a wide spectrum of physiological and pathological processes, such as metabolism, circadian rhythm, aging and cancer. Accumulating evidence has indicated that the level of NAD+
is declined due to the compromised NAD+
biosynthesis and the enhanced NAD+
breakdown in various tissues, including liver and fat, under HFD and obese conditions [5
]. Recently, it is well established that loss of NAD+
is involved in the development of HFD and obesity-induced metabolic disorders, such as steatosis, dyslipidemia and insulin resistance [7
]. Therefore, improving NAD+
metabolism has emerged as a potential therapeutic strategy for T2D and its complications. However, whether the dysregulation of NAD+
metabolism contributes to liver inflammation during T2D and the mechanisms underlying these effects remain to be investigated.
The NLR family, pyrin domain-containing 3 (NLRP3) inflammasome is an important member of the innate immune system, which is a cytoplasmic multi-protein complex composed of NLRP3, apoptosis-associated speck-like protein containing a CARD domain (ASC) and pro-cysteine aspartate-specific protease-1 (pro-caspase-1). Over the recent past, the importance of NLRP3 inflammasome-mediated caspase-1 activation and subsequent pro-inflammatory cytokines release as one of the contributing mechanisms to T2D has been recognized [9
]. Indeed, emerging evidence demonstrates that NLRP3 inflammasome activation leads to hepatic metaflammation, contributing to abnormal lipid and glucose metabolism, exacerbating T2D and its complications [11
]. Nevertheless, the mechanisms of NLRP3 inflammasome activation mediated by HFD and obesity are not fully understood.
Purple sweet potato color (PSPC) is a class of naturally occurring anthocyanins that is derived from purple sweet potato storage roots that reportedly has a variety of pharmacological properties including strong antioxidant, anti-inflammatory and neuroprotective effects [13
]. Our previous work revealed that PSPC treatment protected mouse liver from D-galactose-induced oxidative stress, inflammation and hepatocyte apoptosis [16
]. Recently, our studies indicated that PSPC treatment inhibited HFD-induced hepatic insulin resistance partly via repressing oxidative stress and endoplasmic reticulum stress (ER stress) [18
]. Thus, PSPC could be regarded as a suitable candidate for pharmacological intervention of T2D. However, whether PSPC attenuates HFD-induced hepatic inflammation has never been investigated.
In the present study, we hypothesized that PSPC treatment might inhibit NLRP3 inflammasome activation-mediated hepatic inflammation via improving NAD+ metabolism in HFD-fed mice. The improvement of NAD+ metabolism could be a novel protective mechanism of PSPC against T2D and its complications. This study is designed to address these issues.
The importance of NLRP3 inflammasome-mediated inflammation in the development of T2D has been well demonstrated over the past few years [9
]. However, the causal mechanisms underlying the activation of NLRP3 inflammasome during excess caloric consumption and obesity remain largely unknown. The present study provides novel mechanistic insights into NLRP3 inflammasome-mediated inflammation under HFD condition by revealing that NAD+
depletion-mediated ER stress contributed to NLRP3 inflammasome activation in the livers of HFD-treated mice. Furthermore, our findings revealed that PSPC protected against NLRP3 inflammasome-mediated inflammation via ameliorating oxidative stress-mediated NAD+
loss and consequent ER stress in the HFD-treated mouse livers, indicating that PSPC is a candidate for pharmacological intervention of obesity-related metabolic diseases.
There is increasing interest in the important role of NAD+
depletion in the pathogenesis of various injuries and diseases [5
]. Recently, accumulated evidence has established that NAD+
depletion plays a crucial role in the development of T2D and its complications, such as steatosis and insulin resistance [5
]. It has been well demonstrated that the protein expression of NAMPT is decreased, while the level of PARP1 is increased during HFD feeding and obesity, which leads to NAD+
]. In the present study, our results showed that HFD markedly diminished NAMPT protein expression and augmented the level of PARP1 protein in mouse livers, which is accompanied by a decreased level of NAD+
. Consistently, our findings suggested that HFD induced NAD+
depletion by abating NAD+
salvage biosynthesis and enhancing NAD+
consumption. Substantial evidences suggest that naturally occurring polyphenols exhibit beneficial effects on many aspects of T2D and its complications via strengthening NAD+
biosynthesis and restraining NAD+
breakdown pathways to boost NAD+
]. Consistent with these studies, our results revealed that PSPC dramatically restored NAD+
levels in the livers of HFD-treated mice by heightening NAMPT level and diminishing PARP1 level, which might be responsible for many aspects of its beneficial effects on T2D and its complications.
It is well established that intracellular NAD+
level is modulated by cellular stresses including oxidative stress [27
]. Oxidative stress is reported to depress NAMPT-mediated NAD+
biosynthesis during metabolic syndrome and aging [29
]. Accumulated evidence reveals that PARP1 is largely promoted by oxidative stress under various pathological conditions, resulting in NAD+
]. In the present study, our results showed that HFD induced a severe oxidative stress, which was attenuated by PSPC in mouse livers. As strong antioxidants, naturally occurring polyphenols are well demonstrated to boost intracellular NAD+
]. Therefore, our findings indicated that PSPC might improve NAD+
metabolism to restore hepatic NAD+
levels in HFD-treated mice via its antioxidant effect.
ER stress is the central feature in peripheral tissues during T2D, which is responsible for many aspects of the pathogenesis of T2D and its complications. It is well established that hepatic ER stress perturbs glucose and lipid homeostasis, contributing to the development of T2D [33
]. In this study, our results showed that HFD caused a severe ER stress in the mouse livers. Accumulated evidence indicates that naturally occurring polyphenols exhibit the strong inhibitory effects on ER stress during various pathological conditions, including T2D [18
]. Consistently, our findings revealed that PSPC markedly attenuated HFD-induced ER stress in mouse livers, indicating that PSPC exhibited beneficial effects on T2D partly via its inhibitory effects on ER stress. Recent studies have highlighted the association between NAD+
loss and ER stress in nonalcoholic fatty liver disease (NAFLD), which is the hepatic manifestation of T2D [36
]. It is suggested that NAD+
loss leads to the accumulation of aberrant proteins, which contributes to ER stress [38
]. Moreover, the activity of Sirt1, an NAD+
-dependent deacetylase, is compromised during NAD+
loss, which promotes ER stress [39
]. In particular, an accumulating body of evidence demonstrates that enhancing NAD+
salvage biosynthesis and consequently elevating NAD+
level alleviate ER stress [36
]. In the present study, the treatment of NR, a NAD+
precursor, largely increased the NAD+
levels and thereby abated ER stress in the livers of HFD-treated mice, confirming the ameliorative effects of NAD+
repletion on ER stress. Collectively, our findings revealed that PSPC might mitigate HFD-induced ER stress via restoring NAD+
levels in mouse livers.
In recent years, the important role of NLRP3 inflammasome implicated in the development of metabolic diseases including T2D received increasing attention [9
]. NLRP3 inflammasome activation leads to autocatalytic activation of caspase-1 and the cleavage of its substrates, including IL-1β, which plays an important role in the pathogenesis of T2D and its complications. In the present study, our results revealed that HFD treatment dramatically augmented NLRP3 inflammasome activation and the expressions of inflammation-related genes in mouse livers, indicating a critical role of NLRP3 inflammasome in HFD-induced hepatic inflammation. Increasing evidence points out that ER stress activates NLRP3 inflammasome under various pathological conditions via both unfolded protein response (UPR), including IRE1 signaling, -dependent and independent pathways [40
]. It is well established that NF-κB p65, the key transcription factor activated by ER stress, contributes to NLRP3 inflammasome activation [42
]. Moreover, recent evidence indicates that ER stress provokes inflammatory response through triggering NOD1 and NOD2 signaling, which plays a role in the activation of NLRP3 inflammasome [43
]. In this study, both NF-κB p65 nuclear translocation and NOD1/2 signaling were notably enhanced in the livers of HFD-treated mice. Our findings further suggested that HFD might induce NLRP3 inflammasome activation by promoting ER stress-mediated augment of IRE1 signaling, NF-κB p65 nuclear translocation and NOD1/2 signaling in mouse livers. It is well demonstrated that naturally occurring polyphenols effectively suppress NLRP3 inflammasome activation via their antioxidant activities [45
]. In the present study, PSPC significantly depressed NF-κB p65 nuclear translocation, NOD1/2 signaling and NLRP3 inflammasome activation in the livers of HFD-treated mice. Thus, our findings revealed that PSPC inhibited HFD-induced NLRP3 inflammasome activation by abating ER stress-mediated augment of IRE1 signaling, NF-κB p65 nuclear translocation and NOD1/2 signaling in mouse livers.
It is well established that obesity is an important risk factor for NAFLD, and is associated with many metabolic abnormalities in liver including inflammation [47
]. Substantial evidence suggests that obesity causes adipose tissue dysfunction, which leads to aberrant release of proinflammatory mediators, such as cytokines and specific fatty acids, contributing to the development of liver inflammation [47
]. Moreover, obesity alters gut microbiota and increases intestinal permeability, which elevates circulating bacterial endotoxins and absorption of dietary lipids, promoting liver inflammation [50
]. Accumulated evidence suggests that naturally occurring anthocyanins exhibit beneficial effects on metabolic abnormalities via their anti-obesity actions [52
]. In this study, PSPC markedly decreased epididymal adipose tissue masses, as well as body weight, without influencing food intake in HFD-fed mice. Consistent with these studies, our findings indicated that PSPC might attenuate HFD-induced liver inflammation by its anti-obesity effects, which requires further study.