Psoraleae Fructus (PF, Psoralea corylifolia L.), a traditional medicine with a long history of application, is widely used clinically for the treatment of various diseases. However, the reports of PF-related adverse reactions, such as hepatotoxicity, phototoxic dermatitis, and allergy, are increasing year by year, with liver injury being the mostly common. Our previous studies have demonstrated that PF and its preparations can cause liver injury in lipopolysaccharide (LPS)-mediated susceptibility mouse model, but the mechanism of PF-related liver injury is unclear. In this study, we showed that PF and bavachinin, a major component of PF, can directly induce the expression of caspase-1 and interleukin-1β (IL-1β), indicating that PF and bavachinin can directly triggered the activation of inflammasome. Furthermore, pretreatment with NLR family pyrin domain-containing 3 (NLRP3), NLR family CARD domain containing 4 (NLRC4) or absent in melanoma 2 (AIM2) inflammasome inhibitors, containing MCC950, ODN TTAGGG (ODN) and carnosol, all significantly reversed bavachinin-induced inflammasome activation. Mechanistically, bavachinin dose-dependently promote Gasdermin D (GSDMD) post-shear activation and then induce mitochondrial reactive oxygen species (mtROS) production and this effect is markedly inhibited by pretreatment with N-Acetylcysteine amide (NAC). In addition, combination treatment of LPS and bavachinin significantly induced liver injury in mice, but not LPS or bavachinin alone, and transcriptome analysis further validated these results. Thus, PF and bavachinin can induce the activation of inflammasome by promoting GSDMD cleavage and cause hepatotoxicity in mice. Therefore, PF, bavachinin, and PF-related preparations should be avoided in patients with inflammasome activation-associated diseases.

Traumatic spinal cord injury (SCI) is a serious disease of the central nervous system. Aside from the limited intrinsic regenerative capacity of neurons, complex microenvironmental disturbances can also lead to further cellular damage and growth inhibition. Programmed cell death regulated by pyroptosis has an important role in the pathogenesis of SCI. While there has been a wealth of new knowledge regarding cellular pyroptosis, a detailed understanding of its role in SCI and possible therapeutic strategies is still lacking. This review summarizes current advances in the regulatory role of pyroptosis-regulated cell death and inflammasome components in the inhibitory microenvironment following SCI, as well as recent therapeutic advances.

Abdominal aortic aneurysm (AAA) is a chronic aortic disease that lacks effective pharmacological therapies. This study was performed to determine the influence of treatment with the gasdermin D inhibitor necrosulfonamide on experimental AAAs. AAAs were induced in male apolipoprotein E-deficient mice by subcutaneous angiotensin II infusion (1000 ng/kg body weight/min), with daily administration of necrosulfonamide (5 mg/kg body weight) or vehicle starting 3 days prior to angiotensin II infusion for 30 days. Necrosulfonamide treatment remarkably suppressed AAA enlargement, as indicated by reduced suprarenal maximal external diameter and surface area, and lowered the incidence and reduced the severity of experimental AAAs. Histologically, necrosulfonamide treatment attenuated medial elastin breaks, smooth muscle cell depletion, and aortic wall collagen deposition. Macrophages, CD4+ T cells, CD8+ T cells, and neovessels were reduced in the aneurysmal aortas of necrosulfonamide- as compared to vehicle-treated angiotensin II-infused mice. Atherosclerosis and intimal macrophages were also substantially reduced in suprarenal aortas from angiotensin II-infused mice following necrosulfonamide treatment. Additionally, the levels of serum interleukin-1β and interleukin-18 were significantly lower in necrosulfonamide- than in vehicle-treated mice without affecting body weight gain, lipid levels, or blood pressure. Our findings indicate that necrosulfonamide reduced experimental AAAs by preserving aortic structural integrity as well as reducing mural leukocyte accumulation, neovessel formation, and systemic levels of interleukin-1β and interleukin-18. Thus, pharmacologically inhibiting gasdermin D activity may lead to the establishment of nonsurgical therapies for clinical AAA disease.

Noncanonical pyroptosis is triggered by Caspase 4/5/11, which cleaves Gasdermin D (GSDMD), leading to cell lysis. While GSDMD has been studied previously in systemic lupus erythematosus (SLE), the role of pyroptosis in SLE pathogenesis remains unclear and contentious, with limited understanding of Caspase 11-mediated pyroptosis in this condition. In this study, we explored the level of Caspase 11-mediated pyroptosis in SLE, identifying both the upstream pathways and the interaction between pyroptosis and adaptive immune responses. We observed increased Caspase 5/11 and GSDMD-dependent pyroptosis in the macrophages/monocytes of both lupus patients and mice. We identified serum lipopolysaccharide (LPS), released from the gut due to a compromised gut barrier, as the signal that triggers Caspase 11 activation in MRL/lpr mice. We further discovered that pyroptotic macrophages promote the differentiation of mature B cells independently of T cells. Additionally, inhibiting Caspase 11 and preventing LPS leakage proved effective in improving lupus symptoms in MRL/lpr mice. These findings suggest that elevated serum LPS, resulting from a damaged gut barrier, induces Caspase 11/GSDMD-mediated pyroptosis, which in turn promotes B cell differentiation and enhances autoimmune responses in SLE. Thus, targeting Caspase 11 could be a viable therapeutic strategy for SLE.

Gasdermin D (GSDMD) is emerging as an important player in autoimmune diseases, but its exact role in lupus nephritis (LN) remains controversial. Here, we identified markedly elevated GSDMD in human and mouse LN kidneys, predominantly in CD11b+ myeloid cells. Global or myeloid-conditional deletion of GSDMD was shown to exacerbate systemic autoimmunity and renal injury in lupus mice with both chronic graft-versus-host (cGVH) disease and nephrotoxic serum (NTS) nephritis. Interestingly, RNA sequencing and flow cytometry revealed that myeloid GSDMD deficiency enhanced granulopoiesis at the hematopoietic sites in LN mice, exhibiting remarkable enrichment of neutrophil-related genes, significant increases in total and immature neutrophils as well as granulocyte/macrophage progenitors (GMPs). GSDMD-deficient GMPs and all-trans-retinoic acid (ATRA)-stimulated human promyelocytes NB4 were further demonstrated to possess enhanced clonogenic and differentiation abilities compared with controls. Mechanistically, GSDMD knockdown promoted self-renewal and granulocyte differentiation by restricting calcium influx, contributing to granulopoiesis. Functionally, GSDMD deficiency led to increased pathogenic neutrophil extracellular traps (NETs) in lupus peripheral blood and bone marrow-derived neutrophils. Taken together, our data establish that GSDMD deletion accelerates LN development by promoting granulopoiesis in a calcium influx-regulated manner, unraveling its unrecognized critical role in LN pathogenesis.

Cardiac dysfunction, an early complication of endotoxemia, is the major cause of death in intensive care units. No specific therapy is available at present for this cardiac dysfunction. Here, we show that the N-terminal gasdermin D (GSDMD-N) initiates mitochondrial apoptotic pore and cardiac dysfunction by directly interacting with cardiolipin oxidized by complex II-generated reactive oxygen species (ROS) during endotoxemia. Caspase-4/11 initiates GSDMD-N pores that are subsequently amplified by the upregulation and activation of NLRP3 inflammation through further generation of ROS. GSDMD-N pores form prior to BAX and VDAC1 apoptotic pores and further incorporate into BAX and VDAC1 oligomers within mitochondria membranes to exacerbate the apoptotic process. Our findings identify oxidized cardiolipin as the definitive target of GSDMD-N in mitochondria of cardiomyocytes during endotoxin-induced myocardial dysfunction (EIMD), and modulation of cardiolipin oxidation could be a therapeutic target early in the disease process to prevent EIMD.

Pyroptosis is a programmed cell death process that frequently occurs in many diseases, including hyperuricemic nephropathy (HN). In HN, a range of stimuli mediates inflammation, leading to the activation of inflammasomes and the production of gasdermin D (GSDMD). Baicalin (BA), a natural flavonoid renowned for its antioxidant and anti-inflammatory properties, was investigated for its role in HN in this study. Initially, HN-like inflammation and pyroptosis were induced in HK-2 cells with treatment of monosodium urate (MSU), followed by the BA treatment. The expression of pyroptosis-associated genes, Panx-1 and P2X7, at both mRNA and protein levels was assessed through real-time polymerase chain reaction (RT-qPCR) and Western blotting (WB) without or with BA treatment. The results showed that expression of Panx-1 and P2X7 at mRNA and protein levels was increased in MSU-treated HK-2 cells, which subsequently decreased upon the BA treatment. Further experiments showed that BA could combine NLRP3 inflammasome and GSDMD, destabilizing GSDMD protein. Moreover, BA protected the cell membrane from MSU-induced damage, as evidenced by Hoechst 33342 and PI double staining, lactate dehydrogenase (LDH) assays, and electron microscopy observations. These results suggest that BA is involved in the regulating Panx-1/P2X7 pathways and thus inhibits pyroptosis, highlighting its potential therapeutic effect for HN.

Acute liver failure (ALF) is a life-threatening disease with high mortality. Given excessive inflammation is one of the major pathogenesis of ALF, candidates targeting inflammation could be beneficial in the condition. Now the effect of hyperactivated succinate dehydrogenase (SDH) on promoting inflammation in lipopolysaccharide (LPS)-treated macrophages has been studied. However, its role and mechanism in ALF is not well understood. Here intraperitoneal injection of D-galactosamine and LPS was conducted in male C57BL/6 J mice to induce the ALF model. Dimethyl malonate (DMM), which inhibited SDH activity, was injected intraperitoneally 30 min before ALF induction. Macrophage pyroptosis was induced by LPS plus adenosine triphosphate (ATP). Pyroptosis-related molecules and proteins including GSDMD oligomer were examined by ELISA and western blot techniques, respectively. ROS production was assessed by fluorescence staining. The study demonstrated SDH activity was increased in liver macrophages from ALF mice. Importantly, DMM administration inhibited ROS, IL-1β, and pyroptosis-associated proteins levels (NLRP3, cleaved caspase-1, GSDMD-N, and GSDMD oligomers) both in the ALF model and in macrophages stimulated with LPS plus ATP. In vitro, ROS promoted pyroptosis by facilitating GSDMD oligomerization. Additionally, when ROS levels were increased through the addition of H2O2 to the DMM group, the levels of GSDMD oligomers were reverted. In conclusion, SDH hyperactivation promotes macrophage pyroptosis by ROS-mediated GSDMD oligomerization, suggesting that targeting this pathway holds promise as a strategy for treating ALF and other inflammatory diseases.

Migraine is a common and disabling primary headache disorder and inflammation is a proposed factor in the complex ethiology of the disease. Gasdermin D (GSDMD) is a membrane pore-forming protein acting through the caspase system. End result is cell death caused by leakage of intracellular components to extracellular space which also results in inflammation. Stemming from this knowledge, the potential role of GSDMD in migraine was investigated in this prospective study. This prospective study was conducted between September 2022 to April 2023. 47 patients with migraine were designated as the patient group, whereas 47 healthy volunteers were designated as the control group. Serum GSDMD levels of both groups were compared, with an additional comparison between migraine patients during symptom-free and attack periods. Migraine related characteristics of the patients were also included in the study. Median GSDMD levels of the patient and control group did not reveal a significant difference. Nausea, vomiting and severity of headache were found to be correlated with GSDMD levels in migraine patients. Patients with nausea revealed a higher GSDMD level compared to patients without nausea during both symptom-free and attack periods (p = 0.021 and p = 0.01, respectively). Nausea was correlated to higher GSDMD levels in the patient population during symptom-free period (p = 0.030). The severity of pain was positively correlated with GSDMD levels during the attack period (p < 0.001). Gasdermin family and GSDMD in particular are promising prospects for therapy in a wide spectrum of disorders. Gasdermin proteins are candidates to be the focus for future studies both related to pathogenesis and drug therapy in migraine and varying inflammatory-driven clinical pictures.

During infections, host cells are exposed to pathogen-associated molecular patterns (PAMPs) and virulence factors that stimulate multiple signaling pathways that interact additively, synergistically, or antagonistically. The net effect of such higher-order interactions is a vital determinant of the outcome of host-pathogen interactions. Here, we demonstrate one such complex interplay between bacterial exotoxin- and PAMP-induced innate immune pathways. We show that two caspases activated during enterohemorrhagic Escherichia coli (EHEC) infection by lipopolysaccharide (LPS) and Shiga toxin (Stx) interact in a functionally antagonistic manner; cytosolic LPS-activated caspase-11 cleaves full-length gasdermin D (GSDMD), generating an active pore-forming N-terminal fragment (NT-GSDMD); subsequently, caspase-3 activated by EHEC Stx cleaves the caspase-11-generated NT-GSDMD to render it nonfunctional, thereby inhibiting pyroptosis and interleukin-1β maturation. Bacteria typically subvert inflammasomes by targeting upstream components such as NLR sensors or full-length GSDMD but not active NT-GSDMD. Thus, our findings uncover a distinct immune evasion strategy where a bacterial toxin disables active NT-GSDMD by co-opting caspase-3.