GSDMD is the first gasdermin to be associated with pyroptosis (He et al., 2015; Broz et al., 2020). Activated Caspase-1/4/5/11 cleaves GSDMD at the ₂₇₂FLTD₂₇₅ site (equivalent to ₂₇₃LLSD₂₇₆ in mouse GSDMD), inducing pyroptosis (Wang K. et al., 2020). In the canonical pathway, DAMPs and PAMPs trigger the activation of pattern recognition receptors (PRRs) such as the NOD-like receptors (NLRs) and the absent in melanoma 2 (AIM2)-like receptors (ALRs). Recognition of the agonists by their receptors normally lead to the assembly of inflammasome, which recruits and activates the corresponding caspases, leading to the processing of pro-cytokines and gasdermins, followed by pore formation on the cell membrane and lytic cell death.

The NLRP3 inflammasome is the most extensively studied pathway that function through GSDMD and have been extensively reviewed (Xu and Nunez, 2023; Li LR. et al., 2024). The NLRC4 inflammasome activation is triggered by selected PAMPs such as bacterial flagellin and the type 3 secretion system proteins, through its function partner NAIPs (Andrade and Zamboni, 2020). Cytosolic dsDNA, regardless of its sequence and origin (from foreign bacteria, fungi, viruses, or damaged cells), triggers AIM2 activation and formation of the inflammasome complex, which lead to Caspase-1 activation and the initiation of pyroptosis (Sharma et al., 2019). Activation of the pyrin inflammasome, as a result of reduced pyrin phosphorylation, leads to the recruitment and activation of Caspase-1 (Aubert et al., 2016; Ng et al., 2010; Kamanova et al., 2008; Xu et al., 2014). Bacillus anthracis activates NLRP1b inflammasome through anthrax lethal toxins, which facilitate ubiquitination and degradation of the N-terminal domain of NLRP1b. The remaining C-termini of NLRP1b recruit and activate Caspase-1 (Sandstrom et al., 2019; Chavarría-Smith and Vance, 2013). Rotavirus infection activates NLRP9b inflammasome via double-stranded RNA (dsRNA) in intestinal epithelial cells. The dsRNA interacts with DEAH-box helicase 9 (DHX9), which then binds to NLRP9b. Subsequently, NLRP9b forms inflammasome complexes with the adaptor protein ASC and activates Caspase-1 (Liu and Gack, 2020; Zhu et al., 2017).

In the non-canonical pyroptosis pathway, cytosolic LPS binds directly to pro-Caspase-4/5 in humans and pro-Caspase-11 in mice, resulting in autophosphorylation and activation (Yu P. et al., 2021). The active forms of Caspase-4/5 and Caspase-11 then trigger cleavage of GSDMD, ultimately leading to pyroptosis. LPS-induced pyroptosis allows potassium (K⁺) efflux through pores formed by GSDMD. This K⁺ efflux subsequently stimulates the activation of NLRP3 inflammasomes, leading to canonical pyroptosis mediated by Caspase-1. This connection suggests a coordination between Caspase-1-mediated canonical pyroptosis and Caspase-11-mediated non-canonical pyroptosis through the involvement of NLRP3 inflammasomes.

The involvement of Caspase-8 in the cleavage of GSDMD and initiation of pyroptosis was discovered more recently (Gram et al., 2019; Chen and Brodsky, 2023). Mouse GSDMD was cleaved by active caspase‐8 at two sites, D276 and D88, and the cleavage at D276 generated the active P30 fragment as in the case of caspase-1 cleavage (Orning et al., 2018; Sarhan et al., 2018; Chen KW. et al., 2019). Activation of caspase-8 was RIPK1-dependent, mediated through its interaction with FAS Associated Death Domain Protein (FADD). Interestingly, GSDMD cleavage and pyroptosis occur upstream of NLRP3 activation, triggered caspase-1 activation and additional cleavage of GSDMD similarly as in the case of non-cononical activation (Chen and Brodsky, 2023).

GSDME can be cleaved by Caspase-3 and Caspase-8 downstream of the NLRP3 inflammasome (Wang C. et al., 2021). Caspase-3 was reported to cleave human GSDME at ²⁶⁷DMPD²⁷⁰, mouse GSDME at ²⁶⁷DMLD²⁷⁰, and zebrafish GSDME1 at ²⁵³SEVD²⁵⁶ (Wang et al., 2017). Previous studies revealed that Caspase-1 and/or Caspase-8 are critical for activating Caspase-3/GSDME-dependent cell death in the absence of GSDMD expression (Wei et al., 2023). In mice with the gain-of-function mutations in NLRP3, deficiency of GSDMD alone could not prevent secretion of IL-1β and IL-18 from macrophages in response to LPS or TNF-α stimulation (Wang C. et al., 2021). Sustained NLRP3 inflammasome activation induced Caspase-8/3 activation, GSDME cleavage, and IL-1β maturation in Gsdmd ^−/− macrophages. Aizawa et al. reported that Caspase-8 induced GSDME-dependent pyroptosis in Caspase-1/11-deficient macrophages (Aizawa et al., 2020). NLRP3 inflammasome activation recruited ACS, and subsequently Caspase-8. Activated Caspase-8 cleaved GSDME. Recently, Caspase-3/GSDME-mediated pyroptosis has been reported in human lung epithelial cells infected by SARS-CoV-2 (Planès et al., 2022). Interestingly, multiple SARS-CoV-2 3CL-(3C-like) proteases cleaved human NLRP1 at the Q333 site, triggering inflammasome assembly and cell death. At the same time, 3CL proteases also inactivate GSDMD, and alternatively, cells died via Caspase-3/GSDME mediated pyroptosis. The 3C protease from the EV71 enterovirus has been shown to cleave and inactivate GSDMD at the Q193 site (Lei et al., 2017).

The mechanisms of GSDME-mediated pyroptosis in tumor cells have been well studied (Hu et al., 2023). A variety of chemotherapeutic drugs kill tumor cells through both apoptosis and GSDME-mediated pyroptosis via Caspase-3 or Caspase-8 activation. For example, Zhang et al. reported doxorubicin (DOX)-induced pyroptosis via the Caspase-3/GSDME pathway in two breast cancer cell lines (MDA‐MB‐231 and T47D) (Zhang Z. et al., 2021). The GSDME-dependent pyroptosis is activated by the ROS/JNK signaling pathway. DOX treatment induced ROS production, which promoted the phosphorylation of JNK into p‐JNK that initiated Caspase-3 activation. In parallel, Caspase-8 was also activated by ROS, which promoted the cleavage/activation of Caspase‐3. Similarly, Shen et al. reported that DOX‐induced pyroptosis occured through the ROS‐JNK‐Caspase 3‐GSDME signaling in human tubular epithelial cells (Shen et al., 2021).

Other gasdermins, including GSDMA, GSDMB and GSDMC are previously considered lacking the inflammatory Caspase (Caspase-1 and Caspase-4/5/11) cleavage site (Shi et al., 2017). However, recent study reveals that GSDMA in many bird, amphibians, and reptiles species contain caspase-1 cleavage sites such as YVAD or FASD in the linker and were cleaved by activated caspase-1 (Billman et al., 2024). Another protease that cleaves GSDMA is Streptococcal pyrogenic exotoxin B (SpeB), the virulence factor of group A Streptococcus (GAS), which directly cleaves GSDMA after Gln246 in keratinocytes (Deng et al., 2022; LaRock et al., 2022). In addition, recent studies have unveiled a correlation between the degradation of GSDMA and cisplatin resistance in tumor cells, suggesting that platinum-based drugs may induce GSDMA-mediated cell death in tumors through some unknown mechanism (Wang H. et al., 2024).

In airway epithelial cells, GSDMB has been shown to be cleaved by activated Caspase-1 after D236, releasing N-terminal fragments capable of inducing pyroptosis (Panganiban et al., 2018). Pyrotosis mediated by Granzyme A (GZMA)/GSDMB has been reported in tumor cells. GZMA released from cytotoxic lymphocytes and natural killer (NK) cells enter tumor cells via perforin. GSDMB is cleaved by cytosolic GZMA at Lys244 to release the N-terminal domain, which perforates the plasma membrane and leads to pyroptosis (Zhou et al., 2020). Research on granzymes has spanned over 30 years, and granzymes are involved in the elimination of virus-infected cells by immune cells. The discovery of GSDMB cleavage by GZMA offered a new mechanism for GZMA-mediated cell death. Interestingly, full-length GSDMB can promote oligomerization of Caspase-4 by a direct binding to its CARD domain, thereby facilitating cleavage of GSDMD and triggering non-canonical pyroptosis (Chen Q. et al., 2019).

GSDMC is expressed in multiple tissues, including the trachea, small intestine, colon, esophagus, skin, spleen, and vagina (Zhu et al., 2024). Both artificially truncated GSDMC-NT and the GSDMC-NT fragment that intracellularly cleaved by Caspase-8 have been demonstrated to provoke pyroptosis (Zhang JY. et al., 2021). A recent study by Hou et al. demonstrated that PD-L1 nuclear translocation in hypoxic tumor cells increases the production of GSDMC, which converted apoptosis to pyroptosis (Hou J. et al., 2020). In a hypoxic environment, phosphorylated STAT3 (Signal transducer and activator of transcription 3) interacts with PD-L1, leading to its nuclear translocation and subsequent upregulation of GSDMC transcription. As GSDMC levels increase, TNF promotes the cleavage of GSDMC by Caspase-8, producing the GSDMC-N-terminal fragment that drives pyroptosis. Additionally, the cellular metabolite α-ketoglutarate (α-KG) facilitates the assembly of the DR6 receptosome in tumor cells, forming a molecular platform that enables the efficient proteolysis of GSDMC by activated Caspase-8, thereby inducing pyroptosis.

NF-κB, IκB, and IKK are the main signaling nexus/hubs of the NF-κB signaling pathway. The NF-κB family consists of five main members: p65 (RelA), RelB, c-Rel, p105/p50 (NF-κB1) and p100/p52 (NF-κB2) (Figure 2). They share a conserved Rel Homology Domain (RHD) of over 300 amino acids at the N-terminus, which endows the ability of dimerization, nuclear translocation, and binding to specific sequences on DNA (Dorrington and Fraser, 2019; Yu et al., 2020; Zinatizadeh et al., 2021). Based on their structural characteristics at the C-terminus, the NF-κB family can be divided into NF-κB subfamily and Rel subfamily. NF-κB subfamily members, p105 and p100, possess long repetitive sequences, ankyrin repeats, at their C-terminus. While the Rel subfamily members, p65, RelB, and c-Rel, have a transcription-activation domain (TAD) at their C-terminus. The TAD is considered essential for the nuclear translocation of NF-κB, hence NF-κB subfamily homodimers are regarded as inhibitory regulators due to the lack of TAD. NF-κB subfamily could only activate gene transcription when forming heterodimers with the Rel subfamily members (Yu H. et al., 2021). In recent years, a sixth member of the NF-κB family has been discovered: RelAp43, a new splicing variant of RelA, lacking the TAD at its C-terminus. It is primarily activated by the rabies virus matrix protein, although its ability to interact with other NF-κB family members has been demonstrated in vitro, this perspective has not been widely accepted (Struzik and Szulc-Dąbrowska, 2019; Ben Khalifa et al., 2016; Luco et al., 2012).

Inhibitor of κB (IκB) serves as a key regulator in the NF-κB signaling pathway, as its name suggests, by inhibiting NF-κB. Its members include IκBα, IκBβ, IκBε, IκBγ, IκBz, Bcl-3, as well as the precursors of NF-κB subfamily, p105, and p100 (Zhang J. et al., 2023). Within 2 years of discovering NF-κB, Baltimore proposed the concept of IκB and identified IκBα, which remains the most extensively studied IκB to date (Baeuerle and Baltimore, 1988). IκB sequesters NF-κB in the cytoplasm, forming an inactive complex. Phosphorylation and degradation of IκB, which is typically mediated by the ubiquitin-proteasome system, are required for activating NF-κB, after that the free NF-κB, with the assistance of the nuclear transport receptor importin α/β, translocate through the nuclear pore and activates transcription (Hinz et al., 2012; Wang X. et al., 2020). The classical IκB proteins, including IκBα, IκBβ, and IκBε, are typically distributed in the cytoplasm. They contain six ankyrin repeats and are capable of binding to and covering the Rel Homology Domain (RHD), thereby exerting inhibitory effects. Non-classical IκB primarily resides in the nucleus (aka nuclear IκB) and is induced to increase as a target gene of NF-κB, which contains more than six ankyrin repeats (Wang X. et al., 2020).

IκB kinase (IKK) is primarily responsible for the phosphorylation of IκB, which was discovered nearly a decade after NF-κB. The IKK complex consists of active IKKα and IKKβ, and two molecules of the non-active IKKγ (NEMO) to form a tetramer (DiDonato et al., 1997; Rothwarf et al., 1998), while whether other components are involved is still under considerable controversy. IKKα and IKKβ are highly similar in sequence and structure, their N-terminal kinase domains need to be activated to phosphorylate IκB (Zhang J. et al., 2023). This activation mechanism may arise from autophosphorylation or upstream signals from kinases such as TAK1, MEKK3, and others (Wang et al., 2001; Mulero et al., 2019). In vitro recombination experiments have shown that IKKγ can only directly bind to IKKβ, while the binding of IKKγ to IKKα depends on IKKβ. Therefore, it is inferred that the non-enzymatic IKKγ plays a scaffold and regulatory role (Zandi and Karin, 2023). Although IKK possesses two enzymatically active subunits (α and β), studies showed that in specific pathways, the enzymatic activity of only one subunit is required (Israël, 2010). Other proteins that may participate in the IKK complex include NIK, MEKK1, IKAP (Cohen et al., 1998). It is worth noting that IKK does not always act as a positive regulator of NF-κB: in immune cells (T and B lymphocytes), IKK phosphorylates Bcl10, leading to inhibition of the NF-κB pathway (Abd-Ellah et al., 2018; Zhou et al., 2004).

During infection and oxidative stress, NF-κB regulates the transcription of inflammation-related genes (Capece et al., 2022; Xu G. et al., 2024). During the resting state, NF-κB is sequestered in the cytoplasm by IκB, preventing excessive gene transcription activity. Upon receiving specific stimuli, such as inflammatory cytokines or cellular stress, IKK phosphorylates and cleaves IκB, releasing NF-κB to enable nucleus translocation that initiates transcription of target genes (Anilkumar and Wright-Jin, 2024).

The NF-κB activation may proceed through the canonical and non-canonical pathways. Initially signaling mediated by IKKβ/NEMO/RelA was regarded as the canonical pathway. However, with the discovery that IKKβ can be partially replaced by IKKα and the involvement of RelA in the non-canonical pathway, this viewpoint has been challenged. It has been suggested that NEMO dependency to be used to define the canonical pathway (Liu T. et al., 2017). Despite this, research on the canonical pathway primarily focuses on IKKβ. Similarly, studies of RelA predominantly focused on the canonical pathway (Shih et al., 2011). Upon stimulation, ligand-loaded receptors recruit adaptor proteins such as TRADD, TRAF2, and RIK1 to activate IKKβ (Ma et al., 2024). The IκB proteins are then phosphorylated by IKK followed by ubiquitination and degradation, releasing the NF-κB dimer for translocation into the nucleus. The dimers corresponding to the classical pathway mainly consist of four types: RelA:RelA, RelA:p50, cRel:cRel, and cRel:p50.

The non-canonical pathway is mediated by IKKα and the NF-κB-inducing kinase (NIK) (Rodriguez et al., 2024). NIK (also known as MAP3K14) is the first identified component in the non-canonical pathway. Currently identified inducers of the non-canonical NF-κB pathway all converge at NIK. Therefore, the activation of NIK is defined as a key event in the non-canonical pathway, which normally lead to the RelB:p52 dimer. Under resting conditions, NIK remains at relatively low levels due to its degradation by TRAF3. Degradation of TRAF3 triggered by the stimulation signals results in the accumulation of NIK within the cell. However, since NIK requires de novo synthesis, it takes some time to reach the threshold required to trigger the non-canonical pathway, this may explain why the signaling response of this pathway is relatively slow. In the steady state, p52 primarily exists in the form of its precursor, p100. Both p100 and IκBα are phosphorylated by NIK (Xiao et al., 2001b). Subsequently, p100 is processed into p52, forming the RelB:p52 dimer. Phosphorylation of IKKα activates it. While p100 has been proposed to be a substrate for IKKα, IKKα did not induce the direct processing of p100. A potential explanation is the binding between IKKα and p100 requiring NIK (Yu et al., 2020; García-García et al., 2024). Although the role of IKKα has not been directly demonstrated, studies on transgenic mice have shown that mice with IKKα deficiency and NIK knockout have similar phenotypes, indicating the critical role of IKKα in the non-canonical NF-κB pathway. On the other hand, IKKα also functions as a negatively feedback regulator of non-canonical NF-κB activity by phosphorylating and degrading NIK, these contradictory results may be related to signal crosstalk (Razani et al., 2010; Gray et al., 2014). Other regulatory mechanisms mainly involve modulating the levels of NIK, such as TNAP, Zfp91, TBK1, and NLRP12 (Sun, 2011; Jin et al., 2012), but the specific regulatory mechanisms are not yet fully understood. OTUD1b has been reported to deubiquitinate TRAF3 to prevent its degradation, thus being regarded as a negative regulatory factor (Hu et al., 2013).

It is almost impossible to have completely independent signaling pathways in complex cells. Crosstalk within the NF-κB signaling pathways has been nicely-reviewed (Oeckinghaus et al., 2011). Here, we summarize the reports of NF-κB signaling pathway activation in inflammation over the past decade. The activation of NF-κB is thought to be part of a stress response as it is activated through the recognition of a variety of stimuli by receptors. These include, but are not limited to, the pattern recognition receptors like TLRs, cytokine receptors such as TNFR and IL-1R, and antigen receptors TCR (T cell receptor) and BCR (B cell Receptor) (Barnabei et al., 2021). Additionally, the cGAS-STING pathway has been identified as a central regulator of type I interferon (IFN) and NF-κB. In this review, we will focus on NF-κB signaling pathways mediated by TLRs, IL-1R,TNFR and cGAS-STING (Figure 3).

NLRP3 inflammasome activation normally occurs in two stages, involving a priming signal (signal I) and an activation signal (signal II) (Figure 4). In most cell types, including macrophages, minimal levels of NLRP3 were observed before inflammasome activation, and pro-IL-1β was not constitutively expressed (He Y. et al., 2016). Signal I upregulated the transcription of NLRP3 and pro-IL-1 (Garlanda et al., 2013; McKee and Coll, 2020). NLRP3 is regulated at the transcriptional level, by TLR and cytokine stimulation through activating the NF-κB pathway (Bauernfeind et al., 2009). It has been reported that NF-κB regulates NLRP3 transcription by binding directly to the NLRP3 promoter region (Qiao et al., 2012). Recent research has added Sphingosine 1-phosphate (S1P)/S1P Receptor (S1PR) signaling as an additional inducer for NLRP3 priming (Hou L. et al., 2020). While the precise mechanism behind S1P/S1PR signaling mediated NLRP3 inflammasome priming remains unclear, S1PR has been shown to activate the NF-κB pathway under acute pancreatitis conditions (Yang et al., 2022).

Several other factors regulate NLRP3 activation through modulating the NF-κB pathway. Leucine rich repeat protein ribonuclease inhibitor (RNH1) plays a role in attenuating NLRP3 inflammasome activation and is involved in reducing the priming signal. RNH1 knockout cells exhibited reduced levels of IκBα and increased levels of phospho-IκBα, indicating enhanced NF-κB activation (Bombaci et al., 2022). Inhibition of bromodomain-containing protein 4 (BRD4), which reduces NF-κB signaling through the suppression of IκBα phosphorylation and degradation, results in the upregulation of NLRP3 at the transcriptional level by enhancing the activity of the NLRP3 promoter (Tan et al., 2020). NEK7 has been identified as a mediator of increased expression of Caspase-1, NLRP3, and GSDMD in inflammatory bowel disease (IBD). In NEK7 knockdown intestinal epithelial cells, ATP/LPS-induced activation of the NLRP3 inflammasome was abolished. NEK7 itself is regulated by the NF-κB signaling pathway, with p65 promoting the transcription of NEK7 through binding to its corresponding binding site within the NEK7 promoter, while p65 inhibition blocks this effect (Chen X. et al., 2019).

FAS-associated death domain protein (FADD) and Caspase-8 were found to contribute to NLRP3 inflammasome priming (Lemmers et al., 2007). Caspase-8 was recruited to the complex containing IKKαβ in response to TLR4 stimulation (Lemmers et al., 2007), This interaction is pivotal for the initiation of NF-κB signaling. Macrophages derived from mice deficient in Caspase-8 exhibited reduced pro-IL-β and TNF levels upon priming and decreased NLRP3 activation (Allam et al., 2014). FADD has been identified as an upstream regulator of Caspase-8, as supported by the observation that FADD-depleted macrophages exhibit significantly reduced processing of mature Caspase-8 when subjected to canonical NLRP3 inflammasome stimuli. Accordingly, FADD deficiency impaired NLRP3 activation by LPS or C. rodential infection (Gurung et al., 2014). Both FADD and Caspase-8 were required for the transcriptional upregulation of NLRP3 and pro-IL-1β in an NF-κB dependent manner. Moreover, deletion of both Caspase-8 and NEMO, the NF-κB essential modulator, resulted in the transcriptional downregulation of NLRP3, TNF-α and IL-1β expression after LPS treatment in hepatocytes (Boaru et al., 2015).

The role of IKKβ in inflammasome activation is controversial. Firstly, Nanda and co-workers have reported a pivotal role for IKKβ in the transcriptional upregulation of inflammasome components (Nanda et al., 2021). Inhibition of IKKβ resulted in decreased inflammasome activation, which is consistent with other studies (Bauernfeind et al., 2009; Boaru et al., 2015; Unterreiner et al., 2021; Schroder and Tschopp, 2010). Intriguingly, their research also unveiled the involvement of IKKβ in the assembly of the NLRP3 inflammasome through recruiting the NLRP3 inflammasome to the trans-Golgi network, where the assembly occurs. Furthermore, IKKβ directly binds to NLRP3, facilitating NLRP3 binding to phosphatidylinositol-4-phosphate (PI4P) on the trans-Golgi network, thereby promoting oligomerization and inflammasome assembly (Asare et al., 2022; Schmacke et al., 2022). Contrary to the observations mentioned above, Greten et al. reported that inhibition of IKKβ, either pharmacologically or genetically, enhanced Caspase-1 dependent IL-1β production in macrophages and related inflammation upon LPS challenge (Greten et al., 2007). Despite a decrease in pro-IL-1β mRNA synthesis under these conditions, efficient pro-IL-1β processing by casaspe-1 leads to even higher IL-1β secretion. It is speculated that during LPS challenge, NF-κB contributed to the upregulation of transcriptions of several genes encoding inhibitors of Caspase-1 activity, thus reducing IL-1β processing. The use of prolonged LPS treatment alone in the latter study, in contrast to the combined use of LPS and NLRP3 stimuli to rapidly activate the NLRP3 inflammasome, might explain the contradictory observation mentioned above.

Additionally, inhibitors targeting TBK1/IKKε have been found to amplify NLRP3 inflammasome responses, including IL-1β and IL-18 secretion, Caspase-1 cleavage, and ASC speck formation. Interestingly, the inhibition of TBK1/IKKε on NLRP3 inflammasome responses does not seem to rely on transcriptional mechanisms but rather through posttranslational regulation, mediated by the phosphorylation of the serine 3 (S5 in humans) on NLRP3 (Fischer et al., 2021). Conversely, a more recent study highlightes an alternative role for TBK1, demonstrating that TBK1 activation enhances NF-κB signling, as evidenced by increased phosphorylation of NF-κB p65. This activation is associated with elevated protein levels of key NLRP3 inflammasome components (NLRP3, ASC, and caspase-1) and increased production of cleaved IL-1β in microglia (Liao et al., 2024).

Moreover, previous research has demonstrated that IKK/NF-κB pathway plays a role in promoting autophagy by upregulating the expression of the autophagy receptor p62, which enhanced the clearance of damaged mitochondria through mitophagy via a Parkin-ubiquitin-dependent mechanism (Zhong et al., 2016). Deletion of p62 specifically in myeloid cells lead to the abnormal accumulation of damaged mitochondria and excessive production of IL-1β, resulting in increased sensitivity to endotoxin-induced shock. A truncating variant of p50 (NFKB1^R157*/R157*) has been identified in patients with necrotizing fasciitis or severe soft tissue inflammations (Nurmi et al., 2024). Autophagy was impaired in patients’ cells, causing increased release of mtROS and mtDNA from damaged mitochondria, which enhances NLRP3 inflammasome activation. These findings collectively suggest that while NF-κB may mediate the priming signal of NLRP3 inflammasome activation, the induction of mitophagy or autophage by NF-κB may function as an autoregulatory mechanism to restrain the pro-inflammatory function.

Recent studies have increasingly highlighted the activation of the NF-κB/NLRP3 pathway in various disease models. Molecules and chemicals that modulate this pathway are being investigated for their therapeutic potential (Li Y. et al., 2024; Xiong et al., 2025; Zhao P. et al., 2024; Hao et al., 2024; Alad et al., 2024; Wei et al., 2024). One notable study examined the role of acetyl-CoA synthetase 2 (ACSS2) in the pathogenesis of acute kidney injury (AKI). Mice deficient in ACSS2 showed reduced NLRP3 inflammasome activation in renal tubular epithelial cells. These effects are regulated by the transcription factor KLF5, which mediates NF-κB activation (Lu et al., 2024). In cardiovascular diseases, the NF-κB/NLRP3 pathway is frequently implicated, especially upon activation by oxidative stress or ROS (Xu Z. et al., 2024; Antar et al., 2024). However, the detailed mechanisms remain poorly understood. In a cadmium (Cd) induced chronic kidney injury model, ROS was found to promote NLRP3 inflammasome activation by inhibiting SIRT1 expression and deacetylase activity. This led to decreased SIRT1–p65 interactions, increased acetylated p65 levels, and subsequent NF-κB activation (Dong et al., 2024a). A similar regulatory role of SIRT1 has been observed in shortwave blue light (SWBL)-induced cataracts. SWBL irradiation suppressed SIRT1 expression, promoted nuclear translocation of NF-κB and activation of the NLRP3 inflammasome, leading to lens epithelial cells pyroptosis and cataract formation (Ji et al., 2024). Furthermore, under hypoxic conditions, NF-κB signaling is regulated by HIF-1α, which acts as a positive regulator of NLRP3 inflammasome actuvation and ASC oligomerization in human dental pulp fibroblasts (Wang D. et al., 2024).

Much less is known about the role of NF-κB in the activation of the NLRC4 inflammasome. In intestinal epithelial cells, the activation of the NLRC4 inflammasome and subsequent pyroptosis induced by flagellin from the gram-positive bacterium Clostridioides difficile is mediated by the NF-κB pathway (Chebly et al., 2022; Keestra-Gounder and Nagao, 2023). Unlike in macrophages, inhibition of IKKα in intestinal epithelial cells leads to decreased expression of pro-casapase-1 and interleukin genes, suggesting a role of NF-κB in NLRC4 activation in this context (Chebly et al., 2022). Further evidence supporting a role of the NF-κB pathway in NLRC4 inflammasome activation is provided by findings that TLR4 inhibition in HK-2 cells (an immortalized proximal tubule epithelial cell line) reduced NLRC4 inflammasome activation, as indicated by decreased protein levels of NLRC4, Caspase-1, ASC, and IL-1β (Dong et al., 2024b). Moreover, NF-κB signaling has been demonstrated to negatively regulate NLRC4-induced cell death. Pretreatment of macrophages with NF-κB activators blocks the NLRC4/Caspase-8-dependent cell death, likely through the initiation of transcriptional anti-apoptotic responses (Lee et al., 2018). Anaplasma Phagocytophilum has been reported to activate the NLRC4 inflammasome in macrophages, mediated by RIPK2 through activating NF-κB signaling upon infection, which promoted COX2 expression. COX2 contributed to the production of PGE2, which activated the NLRC4 inflammasome. (Wang et al., 2016). However, we found the evidence provided by the authors on the involvement of NLRC4 activation is not convincing. Experiments using cells deficient in NLRC4 should be included as controls.

A few studies reported a role of the NF-κB pathway in regulating the AIM2 inflammasome activity, however, the exact mechanism was not clearly defined. A correlation, rather thatn causation, was normally reported. In human papilloma virus (HPV)-infected cervical cancer cells, Sirtuin 1 (SIRT1) enabled HPV-infected cervical cancer cells to continue growing by nullifying AIM2 inflammasome-mediated immunity. Mechanistically, SIRT1 repressed the NF-κB-driven transcription of the AIM2 gene by destabilizing the RELB mRNA (So et al., 2018). In an LPS induced acute lung injury model, HMGB1 functioned through the TLR2/TLR4 dependent NF-κB signaling pathways to regulate AIM2 inflammasome activation (Wang J. et al., 2020). The mRNA levels of AIM2, ASC and Caspase-1 are upregulated upon administration of recombinant HMGB1. Additionally, ozanimod, a sphingosine 1-phosphate (S1P) receptor modulator, has been shown to suppress intracerebral hemorrhage (ICH) and neuroinflammation by inhibiting the SIRT3/NF-κB/AIM2 pathway. This inhibition is evidenced by the downregulation of AIM2 expression level, less cleaved Caspase-1, and reduced levels of NF-κB signaling pathways, as indicated by decreased p65 levels (Li et al., 2023). Similarly, Nicorandil, an ATP-sensitive potassium channel opener, was reported to protect against neuroinflammation in ischemic stroke by suppressing the NF-κB/AIM2/GSDMD pathway (Zhao C. et al., 2024). Treatment with channel moculators was found to affect inflammatory cytokine secretion and pyroptosis-related protein expression, including AIM2, while at the same time lead to changes of the level of p-NF-κB p65, NF-κB p65, and p-IκBα. But a direct regulation of NF-kB on AIM2 is lacking. Moreover, Pseudorabies virus (PRV) infection enhances mRNA levels and protein expression of pro-IL-1β by targeting the TLR-NF-κB axis. The AIM2 inflammasome, activated by PRV genomic DNA, is essential for mature IL-1β secretion both in vivo and in vitro. AIM2-deficient mice displayed heightened susceptibility to PRV infection, indicating a critical role of AIM2-mediated inflammatory responses in host defense against PRV infection. Notably, multiple TLR receptors (including TLR2,3,4 and 5), participate in pro-IL-1β upregulation, all capable of activating NF-κB upon PRV infection (Zhou et al., 2023). While a direct role of NF-κB on AIM2 was not reported, the two pathways are clearly integrated function collaboratively to fend off virus infection.