Targeting pyroptosis in renal fibrosis: From molecular mechanisms to therapeutic horizons (Review)

Introduction

Renal fibrosis is a key pathological process driving the progression of chronic kidney disease (CKD) to end-stage renal disease, ultimately resulting in renal failure (1,2). The rate and severity of fibrotic progression critically influence patient survival and quality of life (3-5). However, no direct and effective treatment for renal fibrosis currently exists aside from kidney transplantation and renal dialysis (6-8). Therefore, elucidating the underlying mechanisms of renal fibrosis may offer novel insights for developing potential therapeutic strategies (9).

Pyroptosis is a highly inflammatory form of programmed cell death characterized primarily by inflammasome formation, activation of cysteine aspartic acid-specific proteases (caspases), gasdermin (GSDM) dependence, and the release of cellular contents and lactate dehydrogenase (10-13). Under physiological conditions, pyroptosis functions as an innate immune response that eliminates damaged cells and maintains in vivo homeostasis (14-16). Accumulating evidence indicates that pyroptosis is closely associated with organ fibrosis, including renal fibrosis, liver fibrosis and pulmonary fibrosis (17,18). A substantial body of research has demonstrated that sustained inflammation driven by pyroptosis is a pivotal contributor to the development of renal fibrosis (19). Mediators released by pyroptotic cells, such as interleukin (IL)-1β, IL-18 and damage-associated molecular patterns (DAMPs), not only recruit and activate immune cells such as M1 macrophages but also induce further pyroptosis in neighboring cells, thereby establishing a self-amplifying 'inflammation-pyroptosis' vicious cycle (20). In this context, pyroptosis acts not merely as an amplifier of inflammation but also as a critical initiator of the chronic pro-fibrotic inflammatory microenvironment (21-23). Notably, core effector molecules involved in pyroptosis have been robustly validated as promoters of renal fibrosis (24,25).

Inflammasomes serve as pyroptosis sentinels and are large multiprotein complexes; among them, the nod-like receptor protein 3 (NLRP3) inflammasome has garnered particular attention in the context of renal fibrosis due to its central role (17,26). Activation of NLRP3 promotes the release of pro-fibrotic factors (26,27). Gasdermin D (GSDMD) is the first GSDM family member identified as essential for pyroptosis (25,28). Upon disruption of its autoinhibitory conformation, GSDMD acquires pore-forming activity on the cell membrane, leading to cell swelling and rupture (29). As the principal executioner of pyroptosis, GSDMD holds considerable research potential in renal fibrosis (30). Genetic ablation of GSDMD has been shown to ameliorate renal fibrosis (31). Moreover, clinical studies have demonstrated that the severity of renal fibrosis in patients with diabetic nephropathy (DN) is positively correlated with the expression levels of both NLRP3 and GSDMD (32). Collectively, activation of these key molecules and the interconnectivity of pyroptosis signaling pathways coordinately regulate the progression of renal fibrosis (33,34).

In the pathological process of pyroptosis and renal fibrosis, crosstalk between pyroptosis and apoptosis, necroptosis, ferroptosis and autophagy promotes renal fibrosis and accelerates renal failure (35-38). For example, GSDMD can promote ferroptosis by increasing cellular iron accumulation (39). Ferroptosis inhibitors, such as Ferrostatin-1, indirectly attenuates the activity of the NLRP3 inflammasome (40). In addition, PANoptosis, a cross-regulation of pyroptosis, apoptosis and necroptosis (41,42), has been shown to be associated with the development of renal fibrosis (43,44). Despite these insights, the mechanisms underlying these interactions in renal fibrosis remain unclear. The pathological effects of the interactive network formed by pyroptosis and other forms of cell death undoubtedly exacerbate renal fibrosis. Crucially, this regulatory machinery extends beyond the kidney and is integrated into a complex network of inter-organ crosstalk (45). The gut-kidney axis acts as a pivotal remote switch, where gut-derived toxins or beneficial metabolites calibrate the activation threshold of the renal NLRP3 inflammasome (46,47). Therefore, elucidation of the mechanisms of multi-target synergistic regulation is becoming increasingly important in renal fibrosis (38,39) (Fig. 1).

Figure 1 Core regulatory molecules and interaction networks in pyroptosis. The core molecular machinery of pyroptosis involves upstream inflammasome sensors, NLRP1/3/6, the central Caspase family proteases, Caspase-1/4/5/11, and downstream executioner proteins, GSDMD/GSDME, culminating in the release of potent effector cytokines such as IL-1β and IL-18. Renal fibrosis does not occur in isolation. It may form a complex interaction network with other death pathways such as apoptosis, necroptosis, ferroptosis and autophagy through key molecules of pyroptosis. NLRP, nod-like receptor protein; GSDM, gasdermin.

At present, numerous studies have investigated the relationship between pyroptosis and renal fibrosis (5), but the regulatory mechanisms by which pyroptosis influences renal fibrosis remain under active investigation. Emerging evidence suggests that inhibiting pyroptosis may represent a potential therapeutic strategy to alleviate renal fibrosis (11). In the present review, the regulatory mechanisms of pyroptosis were summarized and its role in renal fibrosis was discussed.

Regulation mechanism of pyroptotic pathway in renal fibrosis

Caspase-1/GSDMD-mediated canonical pathway of pyroptosis

When cells or tissues are damaged, inflammasomes assemble upon sensing DAMPs or pathogen-associated molecular patterns (PAMPs) (48,49). The assembled inflammasomes recruit and activate procaspase-1, enabling its autocatalytic conversion into mature caspase-1 through autolysis. On the one hand, activated caspase-1 cleaves GSDMD to generate its N-terminal fragments (GSDMD-NT). Oligomerization of these fragments forms 10-14-nm-diameter plasma membrane pores, leading to rupture of renal tubular epithelial cells and the release of abundant DAMPs (26). On the other hand, caspase-1 mediates the maturation and secretion of pro-IL-1β and pro-IL-18 (50). Pyroptosis also recruits and polarizes macrophages toward the M1 phenotype through persistent pro-inflammatory signaling, amplifying inflammation and promoting the release of pro-fibrotic factors such as transforming growth factor-β (TGF-β) (20).

Among inflammasomes, NLRP3 is the most closely associated with renal fibrosis (51). Inhibition of NLRP3 activation has been shown to reduce the expression levels of both pyroptotic and fibrotic proteins (52). NLRP3 recognizes a broad spectrum of stimuli, triggering pyroptosis and promoting the production of fibrotic factors (5). Its activation is a key driver in the progression of renal fibrosis and occurs in two steps. First, NLRP3 expression is upregulated via Toll-like receptor/nuclear factor kappa-B (TLR/NF-κB) pathway-mediated priming. Subsequently, NLRP3 oligomerizes and recruits the adaptor protein apoptosis-associated speck-like protein (ASC) and procaspase-1 to form a complex, leading to caspase-1 self-cleavage. For instance, urate crystals activate the NLRP3 inflammasome through lysosomal rupture and release of cathepsin B/L (CTSB/L) (53,54), resulting in caspase-1-mediated cleavage of GSDMD and ultimately inducing fibrosis (55).

Upstream cellular events such as oxidative stress, mitochondrial damage, or defective autophagy can also prime NLRP3 inflammasome assembly (56-58). Liao et al (34) demonstrated that inhibiting the reactive oxygen species (ROS)/NLRP3 axis effectively alleviates renal fibrosis in 5/6 nephrectomy (5/6 NX) rats. Under high-glucose conditions, mitochondrial damage induces massive ROS production and triggers cytosolic release of mitochondrial DNA (mtDNA), which synergistically initiates NLRP3 assembly (59). Furthermore, impaired autophagic flux and defective autophagolysosomal degradation result in CTSB release, activating NLRP3 and pyroptosis, thereby facilitating fibrosis development (57). Although research in this field is extensive, current investigations remain largely phenomenological, primarily characterizing empirical associations between specific stimuli and pyroptotic phenotypes. The substantial heterogeneity among these upstream signals complicates identification of a convergent molecular node, which is critical for developing precision therapeutic targets capable of counteracting multiple pathogenic triggers across various etiologies.

Notably, epigenetic regulation also plays a pivotal role: N6-methyladenosine modification of NLRP3 enhances its stability and induces pyroptosis and inflammation in the kidney (60). Additionally, palmitate reinforces NLRP3 activation by enhancing its S-palmitoylation in synergy with lipopolysaccharide (LPS) stimulation (61). However, despite the substantial potential of these emerging regulatory mechanisms, current research remains at an early stage, with existing evidence primarily based on molecular expression correlations. Moreover, the scarcity of clinical-grade small-molecule inhibitors targeting these specific epigenetic and post-translational modifications continues to limit translational maturity and clinical applicability (Fig. 2).

Figure 2 Molecular mechanisms of pyroptotic signaling pathways. The pyroptotic machinery is primarily categorized into the Caspase-1-mediated canonical pathway, the Caspase-4/5/11-mediated non-canonical pathway, and alternative pathways driven by GSDME or GSDMC. The canonical pathway initiates with inflammasome assembly, triggered either by direct recognition of DAMPs or PAMPs via pattern recognition receptors, or through indirect mechanisms such as lysosomal rupture releasing cathepsins, which induce mitochondrial damage and mtDNA release to activate inflammasomes such as NLRP3. The assembled complex facilitates Caspase-1 activation; activated Caspase-1 subsequently cleaves GSDMD to generate an N-terminal pore-forming domain while simultaneously processing pro-IL-1β and pro-IL-18 into mature cytokines, which are released extracellularly to amplify the inflammatory response. By contrast, the non-canonical pathway is initiated by the direct binding of intracellular LPS to the CARD domains of human Caspase-4/5 or murine Caspase-11, leading to their oligomerization, auto-activation, and subsequent GSDMD cleavage. Furthermore, the heterogeneity of GSDM executioners enables the conversion of cell death modalities: Under apoptotic or chemotherapeutic stimuli, activated Caspase-3 or Caspase-8 can respectively cleave GSDME or GSDMC, effectively switching classical apoptotic signaling into an inflammatory pyroptotic program. GSDM, gasdermin; DAMPs, damage-associated molecular patterns; PAMPs, pathogen-associated molecular patterns; NLRP3, nod-like receptor protein 3; LPS, lipopolysaccharide; TLR4, Toll-like receptor 4; GSDMC-NT, gasdermin C N-terminal fragments; mtDNA, mitochondrial DNA.

The release of IL-1β and IL-18 during pyroptosis directly activates renal interstitial fibroblasts via the IL-1R/MyD88 and IL-18R/NF-κB signaling pathways (62), promoting collagen deposition and myofibroblast differentiation (63,64). Moreover, pyroptosis releases DAMPs, including mtDNA and adenosine triphosphate (ATP), which activate macrophages through TLR9/P2X purinoceptor 7 receptors and fibroblasts via purinergic signaling, thereby enhancing the TGF-β signaling pathway and driving extracellular matrix deposition through upregulation of fibronectin (FN) and α-smooth muscle actin (α-SMA) (65-67) (Fig. 3).

Figure 3 Regulatory role of pyroptosis in renal fibrosis. When renal resident cells and macrophages recognize DAMPs or PAMPs, the NF-κB pathway is activated, leading to the production of pro-IL-1β/18. Subsequently, the activated nod-like receptor protein 3 inflammasome induces auto-cleavage of pro-caspase-1, and the activated caspase-1 cleaves GSDMD to generate GSDMD-NT, which in turn induces the formation of plasma membrane pores. The release of inflammatory cytokines such as IL-1β and IL-18 stimulates the secretion of TGF-β1 and upregulates the expression of TGF-β receptors. Meanwhile, fibrotic factors such as TGF-β1 accelerate the differentiation of renal interstitial fibroblasts into myofibroblasts. The activation of myofibroblasts leads to excessive deposition of ECM components, such as α-SMA and collagen, ultimately resulting in renal fibrosis. DAMPs, damage-associated molecular patterns; PAMPs, pathogen-associated molecular patterns; NF-κB, nuclear factor kappa-B; GSDM D, gasdermin D; GSDMC-NT, GSDM D N-terminal fragments; ECM, extracellular matrix; α-SMA, α-smooth muscle actin; MMP, matrix metalloproteinase; TLR4, Toll-like receptor 4.

Caspase-4/5/11-mediated non-canonical pathway of pyroptosis

The non-canonical pathway involves the direct activation of caspase-4/5 (human) or caspase-11 (mouse) by LPS from Gram-negative bacteria, leading to GSDMD cleavage (26) and initiating a downstream cascade similar to that of the canonical pathway (28,68). GSDMD-mediated pyroptosis, executed through caspase-dependent cleavage, promotes inflammation and fibrosis (69,70). Surprisingly, research has shown that the conserved cysteine residue at position 191 (Cys191) in human GSDMD undergoes S-palmitoylation, which promotes GSDMD-mediated pyroptosis and cytokine release (71). Additionally, Cys191 mutations or treatment with palmitoyl-transferase inhibitors suppress GSDMD palmitoylation, its membrane localization and subsequent pyroptosis or IL-1β secretion (72).

However, from a critical perspective, the role of non-canonical pathways in renal fibrosis may be underestimated. Direct activation of caspase-11 by intracellular LPS triggers potassium (K+) efflux, which facilitates secondary NLRP3 inflammasome assembly and amplifies the pyroptotic signaling cascade. Crucially, this intracellular event is intricately linked to systemic regulation via the gut-kidney axis. Intestinal dysbiosis in the context of CKD compromises gut barrier integrity, leading to systemic accumulation and translocation of gut-derived LPS and uremic toxins into renal tissues. These remote pathogenic signals serve as pivotal triggers that initiate and sustain chronic renal inflammation and the pyroptotic cascade, representing a critical systemic mechanism driving the progression of renal fibrosis (45,73) (Fig. 4).

Figure 4 Crosstalk network of pyroptosis and its molecular mechanisms. (i) In the crosstalk between pyroptosis and apoptosis, caspase-8 plays a central role. Upon TNF-α signaling stimulation, it can simultaneously activate caspase-3 to induce apoptosis and cleave GSDMD/GSDME to trigger pyroptosis. Additionally, mitochondrial damage caused by pyroptosis releases cytochrome c, thereby activating caspase-3/9-dependent apoptosis, while apoptotic cells release ATP, which feedback-activates the NLRP3 inflammasome. (ii) PANoptosis, as a novel mode of programmed cell death, integrates key signaling pathways of apoptosis, necroptosis and pyroptosis. Death signals, particularly TNF-α, promote the formation of a PANoptosome complex containing components such as ZBP1, ASC, caspase-8 and RIPK1/3, which coordinately triggers large-scale inflammatory cell death. (iii) In terms of interaction with ferroptosis, cellular stress associated with pyroptosis promotes ferroptosis by disrupting intracellular redox homeostasis and impairing the GSH/GPX4 axis. Meanwhile, lipid peroxidation end-products 4-HNE accumulated during ferroptosis act as endogenous danger signals, directly exacerbating oxidative stress while also directly or indirectly enhancing NLRP3 inflammasome activation. (iv) Regarding crosstalk with autophagy, under pathological conditions, impaired autophagosome-lysosome fusion due to secondary lysosomal dysfunction may lead to the leakage of contents such as CSTB into the cytoplasm, thereby exacerbating NLRP3 inflammasome-mediated pyroptosis. Furthermore, potassium efflux and calcium influx induced by pyroptosis disrupt autophagosome-lysosome fusion and impair lysosomal function, ultimately inhibiting the completion of functional autophagic flux. TNF-α, tumor necrosis factor-α; GSDM, gasdermin; NLRP3, nod-like receptor protein 3; ASC, apoptosis-associated speck-like protein; GSH, reduced glutathione; GPX4, glutathione peroxidase 4; 4-HNE, 4-hydroxynonenal; HIF-1α, hypoxia-inducible factor 1α; ZBP1, Z-DNA binding protein 1; Cyt c, cytochrome c.

Caspase-3/GSDME-mediated pyroptosis

Chemotherapeutic agents such as cisplatin or granzyme B can induce caspase-3 activation, which cleaves GSDME to release the GSDME-NT fragment. GSDME-NT oligomerizes to form pores in the plasma membrane, thereby inducing pyroptosis. Li et al (74) found that caspase-3/GSDME-mediated pyroptosis of renal parenchymal cells contributes to fibrosis development following ureteral obstruction. Further investigations by Wu et al (75) team revealed that GSDME deficiency significantly attenuates renal fibrosis and dysfunction in both unilateral ureteral obstruction (UUO) and 5/6 NX models. Conversely, overexpression of GSDME-NT exacerbates fibrotic responses in UUO kidneys and TGF-β1-treated renal tubular epithelial cells (75). Notably, GSDME and GSDMD exhibit synergistic regulatory roles in the progression of pyroptosis and fibrosis (76). A previous study indicated that combined deletion of these two genes confers greater kidney protection than deletion of either gene alone (77).

Caspase-8/GSDMC-mediated pyroptosis

The metabolite α-ketoglutarate is upregulated by ROS, oxidizing death receptor 6 (DR6) and prompting its endocytosis and formation of the DR6 receptosome. Within this complex, caspase-8 undergoes autocleavage and activation, subsequently cleaving GSDMC to release GSDMC-NT, which forms membrane pores and induces pyroptosis (78). Moreover, under specific conditions such as activation of tumor necrosis factor-α (TNF-α) signaling, caspase-8 can cleave GSDMD to generate GSDMD-NT, leading to pyroptosis (29).

The central pathogenic hub through which pyroptosis drives renal fibrosis lies in GSDMD-mediated lytic rupture of the cell membrane. This pivotal execution step directly causes explosive release of pro-inflammatory factors and DAMPs, thereby initiating and perpetuating a pro-fibrotic tissue microenvironment. While single-target inhibition of the upstream sensor NLRP3 or the executioner GSDMD has shown therapeutic potential in preclinical models, inhibitors targeting GSDME, which may act as a compensatory executor, also present significant therapeutic prospects.

Overall, the core pathogenic hub of pyroptosis-driven renal fibrosis centers on GSDMD-mediated cytolytic rupture. This critical execution step directly triggers explosive release of pro-inflammatory cytokines and DAMPs, thereby initiating and sustaining a self-perpetuating pro-fibrotic tissue microenvironment (11). While single-target inhibition of the upstream sensor NLRP3 or the executioner protein GSDMD has demonstrated significant therapeutic potential in preclinical models, the existence of potential compensatory mechanisms involving other GSDMs, such as GSDME, suggests that inhibitors targeting these alternative executioners also hold substantial therapeutic promise.

Regulation of interaction network of pyroptosis and renal fibrosis

Recent evidence indicates that pyroptosis does not occur in isolation but rather engages in sophisticated crosstalk with other programmed cell death modalities, such as apoptosis and necroptosis. This complex regulatory network amplifies initial damage signals and dictates cellular fate, ultimately driving the progression of renal fibrosis (Fig. 4).

Crosstalk with apoptosis

Pyroptosis and apoptosis interact synergistically through the dual cleavage function of caspase-8 (79). Triggered by TNF-α signaling, caspase-8 functions as a molecular switch that initiates classical apoptosis via the caspase-3 cascade while simultaneously cleaving GSDMD or GSDME to execute inflammatory pyroptosis (29). This crosstalk is amplified under hypoxic conditions, where the GSDMC-mediated pyroptosis pathway accelerates tubular epithelial injury (80). Moreover, these pathways establish a self-perpetuating vicious cycle: Pyroptosis-induced mitochondrial dysfunction facilitates cytochrome c release and caspase-9/3-dependent apoptosis, whereas early apoptotic cells release ATP to feedback-activate the NLRP3 inflammasome, thereby intensifying the fibrotic response (79,81).

Crosstalk with PANoptosis

PANoptosis is an integrated programmed cell death pathway that coordinates signaling from pyroptosis, apoptosis and necroptosis through assembly of a multi-protein scaffold called the PANoptosome. Comprising key sensors and adaptors, including Z-DNA binding protein 1 (ZBP1), ASC, caspase-8 and receptor-interacting serine/threonine-protein kinase 1/3, this complex is primarily triggered by TNF-α to drive inflammatory tissue damage (43,82). Furthermore, RIPK3 facilitates renal fibrosis progression in UUO models by phosphorylating NLRP3, thereby enhancing inflammasome activity and inflammatory signaling (83,84). Single-cell RNA sequencing analysis by Zhuang et al (83) and Chen et al (85) confirmed that PANoptosis in proximal tubular epithelial cells significantly exacerbates renal injury. Consequently, targeting PANoptosis, particularly by inhibiting its activation in macrophages, represents a promising therapeutic strategy to mitigate systemic inflammation and chronic kidney damage (86,87).

Crosstalk with ferroptosis

Pyroptosis and ferroptosis synergistically drive kidney disease progression through a complex molecular interplay (39,40). Pyroptotic stress facilitates ferroptosis by perturbing intracellular redox homeostasis and impairing the glutathione/glutathione peroxidase 4 axis (39,88). Conversely, lipid peroxidation end-products such as 4-hydroxynonenal accumulated during ferroptosis serve as endogenous danger signals that directly aggravate oxidative stress and potentiate, either directly or indirectly, activation of the NLRP3 inflammasome, thereby further amplifying the inflammatory response (89). Ultimately, this reciprocal coupling establishes a self-amplifying loop of oxidative stress and inflammation, positioning this crosstalk as a central driver of renal fibrosis (90).

Crosstalk with autophagy

The crosstalk between autophagy and pyroptosis is complex, exhibiting bidirectional regulation in the pathological process. Selective mitophagy provides a protective mechanism by clearing damaged mitochondria and ROS, thereby suppressing NLRP3 inflammasome activation and negatively regulating pyroptosis (91). Conversely, pathological stress can induce lysosomal membrane permeabilization, leading to CTSB leakage into the cytoplasm and exacerbating pyroptotic cell death (92). Under persistent stimuli such as uric acid, the autophagic response may shift from an early adaptive defense to a late-stage dysregulated state (93). Consequently, resolving autophagic dysfunction and suppressing pyroptosis-driven inflammation are essential therapeutic strategies for mitigating renal fibrosis.

In conclusion, given the inherent complexity of pyroptosis and its interactive network, as well as the substantial functional redundancy among distinct programmed cell death modalities, targeting a single pathway may yield limited therapeutic efficacy due to compensatory cellular escape mechanisms. Consequently, multi-target synergistic intervention has emerged as a transformative therapeutic paradigm. Specifically, strategies targeting convergent terminal executioners (for example, dual GSDMD/E blockade) or upstream integrative hubs (for example, PANoptosome-targeted intervention) represent promising preclinical paradigms with high translational potential. Although challenges such as cell-type specificity, safety profiling and kidney-selective delivery remain to be addressed, these approaches constitute emerging therapeutic frameworks for interrupting the maladaptive acute-to-chronic kidney disease (AKI-to-CKD) transition and effectively halting the progression of renal fibrosis.

From pyroptotic mechanisms to clinical translation: A critical evaluation and limitations of experimental models of renal fibrosis

In exploring the pathological role of pyroptosis in driving renal fibrosis, the strategic selection of animal models is fundamental for assessing mechanistic reliability and clinical translational potential.

While LPS-induced models serve as the 'gold standard' for validating canonical and non-canonical pyroptotic pathways via rapid GSDMD cleavage, they exhibit inherent limitations in mimicking chronic fibrotic evolution, as they primarily simulate systemic sepsis or acute inflammatory responses rather than primary renal pathology, often resulting in transient functional shifts that fail to capture the sustained, progressive nature of clinical CKD. By contrast, the ischemia-reperfusion injury (IRI) model is more suited for capturing the pyroptotic cascade during the AKI-to-CKD transition, though its utility is frequently compromised by high technical sensitivity to factors such as core temperature and a narrow injury threshold that risks either rapid tissue repair or premature animal mortality. For mechanistic studies focused on interstitial fibrosis, UUO remains the core methodology for investigating the pressure-triggered NLRP3/Caspase-1 axis due to its superior reproducibility, albeit its purely mechanical etiology lacks clinical systemic comorbidities, and its compressed, irreversible pathological course limits its effectiveness in evaluating therapies aimed at regression of fibrosis. By comparison, the adenine-induced model provides an ideal setting for studying secondary pyroptosis triggered by crystalline-induced metabolic disturbances. Furthermore, the cisplatin model offers unique advantages in elucidating Caspase-3/GSDME-mediated pyroptosis, facilitating a deeper understanding of the cross-talk between apoptosis and pyroptosis, notwithstanding that traditional high-dose regimens typically induce severe acute necrosis and high lethality, thereby diverging from clinical repeated low-dose protocols and obscuring the window for longitudinal fibrotic assessment (94,95).

Despite these significant contributions, a gap persists between these experimental models and the complex pathophysiology of human CKD. Although5/6 NX and DN models offer higher translational value by simulating hemodynamic fluctuations and metabolic aberrations, their extended induction periods and complex background signals often complicate the precise quantification of specific contribution of pyroptosis (96). Consequently, cross-validating findings across multiple models to balance experimental stability with clinical relevance is an essential prerequisite for advancing pyroptosis research toward therapeutic intervention (Table I).

Table I Comparison of common animal models for renal pyroptosis.

Table I

Comparison of common animal models for renal pyroptosis.

Model name	Triggering factors	Fibrosis sensitivity	Translational value	Research challenges
Lipopolysaccharide	Endotoxin-mediated Toll-like receptor 4 activation; Damage-associated molecular patterns' release	Low (Primarily acute inflammation)	Moderate (Sepsis/AKI)	Rapid clearance; difficult to simulate chronic progressive fibrosis
Unilateral ureteral obstruction	Mechanical hydrostatic pressure; tubular stretch	High (Rapid and severe interstitial fibrosis)	High (Obstructive nephropathy)	Lacks metabolic background; non-reversible; cannot assess systemic renal function
IRI	Hypoxia/reoxygenation; reactive oxygen species generation; ATP depletion	Moderate (Dependent on injury severity)	High (Renal transplantation; surgery-related AKI)	High variability; sensitive to surgery duration and animal core temperature
Adenine diet	Crystalline-induced tubular injury; metabolic disturbance	High (Diffuse interstitial fibrosis)	High (Crystal-induced/ Metabolic CKD)	Significant systemic toxicity (anorexia, weight loss); high mortality if not strictly monitored.
Cisplatin-induced	Direct DNA damage; mitochondrial dysfunction	Moderate (Dose-dependent AKI-to-CKD)	High (Chemotherapy-induced nephrotoxicity)	Balancing acute mortality with chronic fibrotic progression; specific to GSDME pathway
5/6 NX	Hemodynamic overload; hyperfiltration; uremic toxins	High (Glomerulosclerosis and interstitial fibrosis)	Very High (Chronic renal insufficiency)	High surgical complexity; significant postoperative mortality; long induction period
Diabetic nephropathy	Chronic hyperglycemia; glycation end-products; oxidative stress	Moderate to High (Progressive thickening)	Very High (Diabetic kidney disease)	Longest induction period (weeks to months); complex crosstalk between metabolic and inflammatory pathways

[i] AKI, acute kidney injury; CKD, chronic kidney disease.

Anti-pyroptosis treatment and application prospects

Pharmacological interventions

While no single experimental model can fully encapsulate the intricate landscape of pyroptosis in human CKD, the integrated application of models such as UUO, IRI and DN has successfully identified the core pyroptotic signaling molecules driving renal fibrogenesis. Building upon the mechanistic breakthroughs derived from these experimental systems, a diverse array of targeted interventions, specifically those modulating inflammasome assembly, caspase activation and GSDM pore formation, has emerged, delineating a promising therapeutic blueprint for halting renal fibrosis progression and advancing clinical translation (Table II).

Table II Therapeutic agents targeting pyroptosis.

Table II

Therapeutic agents targeting pyroptosis.

Authors, year	Therapeutic agents	Targeting molecule	Potential mechanism	(Refs.)
NLRP3 inhibitors
Zahid et al, 2019; Coll et al, 2019; Zhang et al, 2019; Østergaard et al, 2022; Li et al, 2022	MCC950	NLRP3	NLRP3 ATPase activity, preventing inflammasome assembly	(97-101)
Huang et al, 2018; Kaneyama et al, 2010	Tranilast	NLRP3	Directly binds to the NACHT domain of NLRP3 and inhibits NLRP3 oligomerization	(102,103)
Mastrocola et al, 2016	BAY 11-7082	NLRP3	Inhibits NLRP3 ATPase activity and NF-κB activation	(104)
Jiang et al, 2017	CY-09	NLRP3	Directly binds the NLRP3 NACHT domain and inhibits NLRP3 ATPase activity	(106)
Youm et al, 2015; Aisyah et al, 2025	β-hydroxybutyrate	NLRP3	Prevents K+ efflux and reduces ASC oligomerization	(108,109)
Wang et al, 2020; Pan et al, 2019	Pterostilbene	NLRP3	Inhibits the activation of NLRP3	(111,112)
Li et al, 2017; Ma et al, 2022	Berberine	NLRP3	Inhibits the activation of NLRP3	(113,114)
Cui et al, 2020;	Phloretinin	NLRP3	Inhibits the activation of NLRP3	(115)
Wu et al, 2020; Yan et al, 2022	Butyrate	NLRP3	Inhibits the activation of NLRP3	(116,117)
Wang et al, 2022	Fucoidan	NLRP3	Inhibits the activation of NLRP3 and reduces GSDMD-N-terminal fragments' expression	(119)
Qu et al, 2022	Pyrroloquinoline quinone	NLRP3	Inhibits the NLRP3-caspase-1-GSDMD pathway and scavenges mitochondrial ROS	(121)
Bai et al, 2025; Elsayed et al, 2021	OLT1177	NLRP3	Inhibits the activation of NLRP3	(122,123)
Feng et al, 2023	Baicalein	NLRP3	Inhibits the activation of NLRP3	(22)
Zhou et al, 2023	Shizhifang	NLRP3	Suppresses the NLRP3-caspase-1-GSDMD pathway	(125)
Su et al, 2025	Qizhi Jiangtang Capsule	NLRP3	Suppresses the NLRP3-caspase-1-GSDMD pathway	(126)
Caspase-1 inhibitors
Flores et al, 2018; Wen et al, 2022	VX-765	Caspase-1	Directly binds to caspase-1	(127,128)
Yang et al, 2021	Ac-YYAD-cmk	Caspase-1	Inhibits the active site of caspase-1	(129)
Liu et al, 2025 GSDMD inhibitors	Quercetin	Caspase-1	Inhibits caspase-1	(130)
Hu et al, 2020; Silva et al, 2021	Disulfiram	GSDMD	Directly target the Cys191 of human GSDMD	(132,133)
Rathkey et al, 2018; Li et al, 2022	NSA	GSDMD	Directly inhibits GSDMD and oligomerization of p30-GSDMD	(136,137)
Zhou et al, 2023; Han et al, 2021	Metformin	GSDMD	Indirectly modulating the GSDMD pathway via AMPK activation	(138,139)
IL-1β inhibitors
Ling et al, 2017	Anakinra	IL-1β	Selective recombinant antagonist of the IL-1β receptor	(140)
Cherney et al, 2018	Canakinumab	IL-1β	IgGκ monoclonal antibody targeting IL-1β	(141)
Upstream inhibitors
Li et al, 2023	Syringaresinol	NRF2 -mediated antioxidant pathway	Reduces ROS, pyroptosis-related proteins	(142)
Liao et al, 2024	N-acetyl-L-cysteine	ROS	Inhibits oxidative stress-induced inflammasome activation and pyroptosis	(37)
Guo et al, 2025	Poria cocos	NRF2 and NF-kB/NOX4 axes	Regulating the NRF2 and NF-kB/NOX4 axis	(143)
Miao et al, 2025	Rhubarb	NF-κB-mediated pathway	Inhibiting NF-κB-mediated transcriptional priming	(144)
Feng et al, 2025	Bushen Huoxue	ROS/NLRP3	Suppresses oxidative stress and NLRP3 activation	(145)
Ruan et al, 2024	Qufeng Tongluo	Autophagy	Restoring impaired autophagic flux	(146)

[i] NF-κB, nuclear factor kappa-B; ROS, reactive oxygen species; NLRP3, nod-like receptor protein 3; GSDMD, gasdermin D.

NLRP3 inhibitors

Small-molecule inhibitors such as MCC950, a classic NLRP3 inhibitor, block NLRP3 ATPase activity, prevent inflammasome assembly (97,98), and reduce renal fibrosis (99,100). Although MCC950 demonstrated robust efficacy in animal studies, its clinical development was hindered by potential hepatotoxicity (101).

In the context of clinical translation, tranilast, an anti-fibrotic drug approved in Japan, has been shown to bind directly to the NACHT domain of NLRP3, thereby inhibiting its oligomerization, suppressing inflammasome assembly, and attenuating renal fibrosis (102,103).

Similarly, BAY 11-7082, a sulfonic acid derivative, suppresses NLRP3 activation by inhibiting both NLRP3 ATPase activity and NF-κB signaling (104). In diabetic rat models, it has demonstrated the potential to restore blood glucose and creatinine levels while ameliorating renal damage (105). However, due to its high electrophilicity, this compound is prone to broad non-specific off-target effects, and its systemic toxicity remains a significant barrier to clinical translation. CY-09, a novel NLRP3 inflammasome-specific inhibitor, directly binds the NLRP3 NACHT domain and inhibits NLRP3 ATPase activity (106). Nevertheless, current evidence for CY-09 is derived primarily from rodent studies, and it lacks essential human pharmacokinetic data and clinical validation regarding its long-term safety profile (107).

Natural compounds and metabolic derivatives offer diverse mechanistic pathways for intervening in pyroptosis. β-hydroxybutyrate, a ketone metabolite, downregulates NLRP3 by preventing K+ efflux and reducing ASC oligomerization and speck formation (108), thereby ameliorating renal fibrosis in a streptozotocin-induced diabetic model (109). However, its clinical utility is limited by a lack of significant lifespan extension in late-stage disease and a biphasic effect wherein concentrations >10 mM become cytotoxic (110). Pterostilbene (PT, trans-3,5-dimethoxy-4-hydroxystilbene) (111,112), berberine (BBR) (113,114), phloretin (115) and baicalein (22) attenuate renal fibrosis by inhibiting NLRP3 activation through various mechanisms, including inhibition of histone deacetylases (for example, HDAC3), induction of autophagy, or scavenging of mitochondrial ROS. Despite their therapeutic potential, the clinical translation of these natural compounds is hindered by unfavorable pharmacokinetic profiles. Specifically, BBR is limited by <1% bioavailability due to its quaternary ammonium structure and P-glycoprotein-mediated efflux, while PT, notwithstanding its lipophilicity, suffers from metabolic instability and poor renal targeting. Similarly, phloretin remains constrained by low bioavailability (~8.67%) and rapid systemic clearance. Furthermore, baicalein's efficacy is compromised by the mandatory intestinal hydrolysis of its glycoside form and subsequent rapid hepatic glucuronidation, which collectively limit effective drug accumulation in renal tissues.

Butyrate, a short-chain fatty acid (SCFA), suppresses NLRP3-mediated pyroptosis in an adenine-induced CKD mouse model and reduces pyroptosis-related proteins (IL-1β, caspase-1 and GSDMD), thereby delaying CKD progression (52). It also modulates epigenetic modifications through inhibition of histone deacetylases such as HDAC3 to inhibit pyroptosis (116,117). Clinical trials show that oral butyrate supplementation (4 g/day) reduces inflammatory markers in patients with CKD (118). Although butyrate has demonstrated potential in reducing inflammatory markers in small-scale clinical trials, these natural metabolites generally face significant translational bottlenecks, such as low bioavailability and short half-lives.

Fucoidan targets NLRP3-mediated podocyte pyroptosis through L-selectin binding, reducing GSDMD-NT expression and proteinuria in DN (119). However, its high molecular weight may limit renal bioavailability (120). Pyrroloquinoline quinone inhibits the NLRP3-caspase-1-GSDMD pathway and scavenges mitochondrial ROS, attenuating pyroptosis in high-glucose-treated HK-2 cells and DN mice and decreasing fibrosis markers (α-SMA and collagen IV) (121). However, its optimal dosage and safety profile remain to be established.

Dapansutrile (OLT1177) selectively inhibits the NLRP3 inflammasome by binding to its NACHT domain and blocking ATPase activity. In folic acid-induced models, it significantly lowered serum creatinine and reduced tubulointerstitial fibrosis. Its clinical efficacy is being evaluated in the DAPAN-DIA Phase II trial involving 300 diabetic patients with systemic inflammation. However, its clinical maturity is constrained by a lack of long-term human safety data, potential infection risks, and limited efficacy as a monotherapy during the NF-κB priming phase (122,123).

Traditional Chinese medicine (TCM) and its bioactive constituents offer a sophisticated, multi-targeted approach to alleviating renal fibrosis and facilitating tissue repair (124). For example, Shizhifang and Qizhi Jiangtang Capsule suppress pyroptosis via the NLRP3/caspase-1/GSDMD pathway, thereby improving renal injury (125,126).

Caspase-1 inhibitors

VX-765, a non-toxic small-molecule caspase-1 inhibitor, has shown promising results in the treatment of epilepsy but remains clinically unproven for nephropathy (127,128). Ac-YVAD-cmk (129), a caspase-1 inhibitor, reduces pyroptosis-related proteins and inflammatory factors by inhibiting the active site of caspase-1. Quercetin (130), a natural compound, displays potential to alleviate renal fibrosis by inhibiting the caspase-1-mediated canonical pyroptosis pathway. Similarly, both Ac-YVAD-cmk and quercetin show efficacy in animal models; however, the former lacks human data, and the latter is constrained by rapid metabolism and a bioavailability of <2% (131).

GSDMD inhibitors

GSDMD-mediated pyroptosis plays a critical role in the pathogenesis of renal fibrosis, positioning GSDMD inhibitors as a potential therapeutic strategy.

Disulfiram, an FDA-approved drug for alcoholism (132), blocks pore formation by covalently modifying the Cys191 residue of human GSDMD, significantly attenuating fibrosis in experimental UUO and diabetic nephropathy models (133). However, its long-term application is constrained by dose-dependent hepatotoxicity (134,135).

Although NSA inhibits GSDMD oligomerization, its clinical potential is limited by the complexity of its inhibitory mechanism (136,137). Additionally, metformin indirectly modulates the GSDMD pathway via AMP-activated protein kinase activation, reducing collagen deposition in animal models (138,139). Although these studies provide a robust mechanistic foundation, most inhibitors remain in the experimental stage, and their maturity and safety in the context of renal disease require careful validation through dedicated clinical trials.

IL-1β inhibitors

Direct IL-1β inhibitors, such as the IL-1 receptor antagonist anakinra and the monoclonal antibody canakinumab, have shown promise in reducing fibrosis. Anakinra reduces blood pressure and renal fibrosis in hypertensive mice but has minimal effects on inflammation (140). The clinical evidence for canakinumab was more extensively established in the landmark Canakinumab Anti-inflammatory Thrombosis Outcomes Study (CANTOS). A prespecified analysis of the CANTOS sub-cohort with moderate CKD [baseline estimated glomerular filtration rate (eGFR) <60 ml/min/1.73 m2] revealed that canakinumab reduced the risk of major adverse cardiovascular events by 18% (HR 0.82), with the most pronounced clinical benefits observed in 'biologic responders' who achieved on-treatment high-sensitivity C-reactive protein levels <2 mg/l. Although the trial did not demonstrate a statistically significant preservation of eGFR or reduction in albuminuria compared with placebo, the findings underscore the pivotal role of IL-1β in driving the 'residual inflammatory risk' prevalent in CKD populations and provide a safety-validated framework for anti-inflammatory strategies in cardiorenal syndromes (141).

Upstream signals and renal pyroptosis

Targeting upstream pathogenic signals, specifically oxidative stress and pro-inflammatory priming factors, represents a critical research frontier and therapeutic strategy for inhibiting pyroptosis and mitigating renal fibrosis. Syringaresinol (SYR) effectively clears ROS via the NRF2-mediated pathway, leading to reductions in pyroptosis-associated proteins and interstitial fibrosis in diabetic models (142). This effect is complemented by N-acetyl-L-cysteine, an established ROS scavenger that prevents oxidative stress-driven inflammasome activation and subsequent cell death (37).

Furthermore, TCM and its bioactive constituents offer a sophisticated, multi-targeted approach to modulating the upstream microenvironment. Poria cocos has demonstrated clinical potential as an adjunctive therapy for diabetic kidney disease and nephrotic syndrome by regulating the NRF2 and NF-κB/NOX4 axes via its triterpenoid and polysaccharide constituents to alleviate proteinuria and renal damage (143). Rhubarb (Rheum palmatum) (144) and its primary anthraquinone derivatives (145) (for example, rhein and emodin) exert potent effects by inhibiting NF-κB-mediated transcriptional priming while synergistically downregulating the TGF-β/Smad and Wnt/β-catenin pathways. These mechanisms are integrated within formulas such as the Bushen Huoxue decoction and compounds such as bicyclol, which collectively suppress inflammatory priming and NLRP3 activation (34). Concurrently, Qufeng Tongluo Decoction (146) has been shown to effectively alleviate podocyte injury by restoring impaired autophagic flux, thereby preserving the stability of cytoskeletal proteins such as nephrin. In summary, pharmacological modulation of these upstream triggers provides a robust framework for alleviating renal pathological injury at its metabolic and inflammatory origins.

Multi-target synergistic therapies

As aforementioned, the intricate crosstalk between pyroptosis and other regulated cell death (RCD) modes drives a pathological cascade that accelerates renal fibrosis, providing a mechanistic rationale for targeting multiple RCD processes.

Targeting PANoptosis to inhibit organ fibrosis is now a promising research direction. Licochalcone B and baicalin attenuate fibrosis by inhibiting ZBP1-dependent PANoptosome assembly, while the medicinal plant Achyranthes aspera has been shown to improve renal function and tubular injury by mitigating PANoptosis (87,147,148). Necrostatin-1, a RIPK1 inhibitor, suppresses overexpression of RIPK1-RIPK3-MLKL proteins and pyroptosis-related cytokines and decreases apoptotic cell death to reduce renal fibrosis (149). Moreover, the clinical relevance of this mechanism is underscored by the verified upregulation of ACSL4 in the renal tubules of patients with various CKDs, including IgA nephropathy and DN (150). Beyond mitigating lipid peroxidation, the ferroptosis inhibitor liproxstatin-1 also regulates PANoptosis, effectively inhibiting hepatic apoptosis, pyroptosis and necroptosis (151). At the transcriptional level, the ubiquitin-specific protease 11 inhibitor mitoxantrone prevents stabilization of the transcription factor Krüppel-like factor 4 (KLF4), thereby blocking the KLF4-induced dual pyroptotic pathways (Caspase-1/GSDMD and Caspase-3/GSDME) to alleviate renal fibrosis (152).

Genetic interventions and stem cell therapy

The rapid evolution of biotechnological interventions has positioned gene editing and stem cell-derived therapies as pioneering frontiers for disrupting the pyroptotic cascade and addressing renal fibrosis.

CRISPR-Cas9 technology is widely used to construct mouse models with gene knock-out or knock-in modifications to study the role of specific genes in renal fibrosis, such as GSDMD knock-out (153). CRISPR-Cas9 also facilitates genome-wide association studies for kidney function to identify risk genes associated with fibrotic kidney disease (154). A recent study found that an improved CRISPR-Cas9 system revives gene expression by specifically targeting methylated, silenced anti-fibrotic genes such as RASAL1 and Klotho, thereby improving renal fibrosis (155). However, translational challenges remain regarding the long-term safety and delivery efficiency of gene editing. Stem cell therapy, particularly exosome-based therapy, represents a novel therapeutic strategy against renal fibrosis. For example, overexpression of exosomal miR-342-3p suppresses high glucose-induced renal interstitial fibrosis (156). By elevating exosomal miR-342-3p levels, pyroptosis of renal tubular epithelial cells can be inhibited and renal function can be improved (157). Additionally, nanotechnology-based delivery platforms such as PEGylated liposomes could overcome the bioavailability limitations of natural compounds such as quercetin (158). Liposomal encapsulation of such compounds enhances therapeutic efficacy and reduces systemic toxicity in renal fibrosis (159). Nickel-cobalt alloy magnetic nanocrystals (an inorganic nanomaterial, exhibits broad-spectrum inflammasome inhibitory effects, simultaneously suppressing the activation of three inflammasomes, NLRP3, NLR family CARD domain containing 4 and absent in melanoma 2. This property renders them both anti-inflammatory agents and ideal delivery vehicles for anti-inflammatory drugs (160). In summary, this strategic direction is catalyzing a clinical paradigm shift from palliative symptom management toward targeted molecular repair.

Conclusion

In conclusion, GSDMD-mediated cytolytic rupture serves as the central pathogenic hub in renal fibrosis, acting as the terminal executioner that perpetuates the pro-fibrotic milieu. However, successful clinical translation requires moving beyond descriptive research to address several strategic gaps. Current investigations fail to elucidate how the renal microenvironment, specifically chronic hypoxia, electrolyte flux, and uremic toxins, differentially modulates pyroptosis activation thresholds across distinct nephron segments. Furthermore, the long-term safety of suppressing innate immune sentinels such as NLRP3 remains poorly defined; given its critical role in host defense, systemic inhibition may heighten susceptibility to opportunistic fungal or viral infections in immunocompromised patients with CKD. Standard experimental models also exhibit substantial translatability gaps, including functional compensation by the contralateral kidney in UUO models and strain-dependent resistance to fibrosis (for example, in C57BL/6 mice) in 5/6 NX models, factors that can obscure true therapeutic efficacy. Future research must prioritize multi-target synergistic interventions, particularly those directed at integrative molecular scaffolds such as the PANoptosome, to circumvent cellular escape mechanisms. The translational roadmap necessitates longitudinal functional validation using GFR-based metrics, identification of non-invasive urinary biomarkers for patient stratification, and development of renal-targeted precision delivery systems to maximize anti-fibrotic potency while minimizing systemic toxicity.

Availability of data and materials

Not applicable.

Authors' contributions

YT wrote the original draft, visualized data, and wrote, reviewed and edited the manuscript. QM wrote the original draft and visualized data. HF and HS wrote the original draft. JL and QZ wrote, reviewed and edited the manuscript, supervised the study and acquired funding. All authors read and approved the final version of the manuscript. Data authentication is not applicable.

Ethics approval and consent to participate

Not applicable.

Patient consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

Abbreviations:

Acknowledgments

Not applicable.

Funding

The present study was supported by the Sichuan Science and Technology Program (grant no. 2026NSFSC0641), the Southwest Medical University Technology Program (grant no. 2023QN019), the National Natural Science Foundation of China (grant no. 82205002), the Science and Technology Research Special Project of Sichuan Administration of Traditional Chinese Medicine (grant no. 2024MS524) and the Special project of integrated Chinese and Western medicine, Southwest Medical University (grant no. 2023ZYQJ04).

References