ABSTRACT TNF inhibitors have been used to treat autoimmune and autoinflammatory diseases. Here we report an unexpected mechanism underlying the therapeutic effects of TNF inhibitors in

Behçet’s disease (BD), an autoimmune inflammatory disorder. Using serum metabolomics and peripheral immunocyte transcriptomics, we find that polymorphonuclear neutrophil (PMN) from patients

with BD (BD-PMN) has dysregulated mevalonate pathway and subsequently increased farnesyl pyrophosphate (FPP) levels. Mechanistically, FPP induces TRPM2-calcium signaling for neutrophil

extracellular trap (NET) and proinflammatory cytokine productions, leading to vascular endothelial inflammation and damage. TNF, but not IL-1β, IL-6, IL-18, or IFN-γ, upregulates TRPM2

aggravation, and novel insights for TNF inhibitor therapies on BD. SIMILAR CONTENT BEING VIEWED BY OTHERS EFFECTIVE XANTHINE OXIDASE INHIBITOR URATE LOWERING THERAPY IN GOUT IS LINKED TO AN

EMERGENT SERUM PROTEIN INTERACTOME OF COMPLEMENT AND INFLAMMATION MODULATORS Article Open access 19 October 2024 INFLAMMATORY DISEASE PROGRESSION SHAPES NANOPARTICLE BIOMOLECULAR

CORONA-MEDIATED IMMUNE ACTIVATION PROFILES Article Open access 22 January 2025 ANTIBODY-MEDIATED NEUTRALIZATION OF GALECTIN-3 AS A STRATEGY FOR THE TREATMENT OF SYSTEMIC SCLEROSIS Article

Open access 31 August 2023 INTRODUCTION Different from the well-characterized damage-associated molecular pattern (DAMP), metabolic intermediates, have recently been identified as a novel

class of endogenous danger signals that strongly elicit a variety of biological functions1,2,3,4. In response to external stimulations, polymorphonuclear neutrophil (PMN) tend to adopt

specific metabolic pathways5, for a tuned purpose to support specialized effector functions such as neutrophil extracellular trap (NET) formation (NETosis)6,7. Hence, considering the short

cell lifespan8, PMN is a potentially important source of serum danger signals during the shift of its metabolic patterns9. However, the immunometabolism of PMN in autoimmune and

autoinflammatory diseases remains to be investigated. Corresponding to the clinical features of autoinflammatory and autoimmune diseases, Behçet’s disease (BD) is a chronic systemic

vasculitis characterized by recurrent oral or genital ulcers, skin lesions, and involvement of vital organs, such as ocular, cardiovascular, gastrointestinal, and neurological

manifestations10. BD is sight-threatening and even life-threatening, imposing considerable financial burdens on society and individuals. BD is a representative type of immune disorder as it

exhibits aberrant and excessive activation of both innate and adaptive immunity, which is thus considered as a unique and crucial clinical condition linking both autoimmunity and

autoinflammation11. Although the etiology of BD is still unknown, it is well accepted that during the progression of BD, an interplay between endogenous danger signals and PMN is of

note12,13,14. In this regard, elevated serum DAMPs such as high mobility group box 1 (HMGB1)15 and S100 calcium-binding protein A12 (S100A12)16 have been reported in BD. These DAMPs promote

acute inflammation and recruitment of PMN to vascular lesions17, and thus BD was also recognized as a “neutrophilic vasculitis”18. Moreover, PMN is particularly prone to undergo cell

necrosis, mainly in the form of NETosis under inflammatory conditions19,20, which involves the release of various DAMPs, including DNA, histones19, HMGB1, and S100A8, etc., aggravating the

activation and inflammation of macrophages19 and vascular endothelium20,21 in BD. As an integrating mechanism, all these accounts for the production of proinflammatory cytokines, including

tumor necrosis factor (TNF), interferon-γ (IFN-γ), interleukin 6 (IL-6), and skewed T-helper (Th) 1 and Th17 cell activation22. Thus, BD is ideal for use as a representative disease to

investigate the mechanism of action underlying the therapeutic effects of TNF inhibitors in treating autoimmune and autoinflammatory diseases. Farnesyl pyrophosphate (FPP), a key metabolite

in the mevalonate (MVA) pathway, plays a crucial role in cholesterol biosynthesis, has been implicated in inflammatory responses, and potentially contributed to autoinflammatory and

autoimmune diseases23,24. However, its implications in BD pathogenesis remain largely to be elucidated. Here in this study, we conduct comprehensive analyses of serum metabolomics and

peripheral immune cell transcriptomics, revealing a dysregulated mevalonate pathway in PMN from BD patients (BD-PMN) and subsequently elevated FPP levels. Further investigations demonstrated

that FPP promotes BD-PMN hyperactivation via a calcium-TRPM2-dependent pathway. TNF upregulates TRPM2 expression on BD-PMN, while TNF inhibitors have the opposite effect. Our findings

highlight the potential pathogenic involvements of FPP in BD and uncover the immunometabolic mechanisms underlying disease progression. These insights provide novel therapeutic implications

for TNF inhibitors in BD and potentially other autoimmune and autoinflammatory disorders. RESULTS MULTI-OMIC ANALYSES HIGHLIGHT THE PROINFLAMMATORY CONTRIBUTIONS OF MVA-PATHWAY IN BD-PMN

HYPERACTIVATION To gain an unbiased understanding of immunometabolic profiles in BD, we first integrated multi-omic analyses in our previously published cohorts containing BD patients and

the sex- and age-matched HC, including serum metabolomics and lipidomics25, bulk RNA-sequencing of peripheral blood mononuclear cell (PBMC) (GSE19853311), PMN (GSE20586712), and single-cell

RNA-sequencing of PBMC (GSE19861611). We found that the steroid biosynthesis pathway is significantly upregulated in BD serum in comparison to that from HC (Fig. 1A, Supplementary Fig. 1A).

Multi-omic analyses of individual immune cell populations revealed that this metabolic alteration in serum was mainly displayed in the BD-PMN (Fig. 1B, and Supplementary Fig. 1B).

Considering that there are multiple downstream pathways in steroid synthesis, we then conducted an enrichment analysis of each pathway. We found that the cholesterol biosynthesis pathway,

but not the synthesis of bile acid and steroid hormone pathways, was significantly increased in BD-PMN in comparison to HC-PMN (Fig. 1C). It shall be noted that although our single-cell

sequencing analyses also indicated a slight upregulation of the steroid synthesis pathway in monocytes from BD patients, their cholesterol synthesis pathway was not significantly upregulated

as in the case of PMN (Supplementary Figs. 1C, D). Next, we examined the expression profiles of crucial enzymes in the cholesterol biosynthesis pathway from transcriptional datasets of

BD-PMN and HC-PMN (Fig. 1D), and further validated the transcription levels of these target genes by qRT-PCR in an independent cohort of twenty BD patients (Supplementary Fig. 1E, and

Supplementary Table 1). We found that those most differentially expressed enzymes, including _ACAT1, MVK, PMVK_, and _MVD_, were concentrated in the upstream pathway of cholesterol

biosynthesis, i.e., the MVA pathway (Fig. 1E). These results reveal an upregulated MVA pathway specifically in BD-PMN. To determine whether the upregulated MVA pathway is involved in the

hyperactivation of BD-PMN, we inhibited 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR) and farnesyl pyrophosphate synthase (FPPS, also known as farnesyl diphosphate synthase (FDPS)), which

control the start and end points of the MVA pathway, via the clinically-available drugs simvastatin and zoledronic acid, respectively (Fig. 1E). Notably, both inhibitors suppressed

proinflammatory cytokines in BD-PMN (Fig. 2A, B). Since FPP is the end product of the MVA pathway that is converted by FPPS enzyme from geranyl-pyrophosphate (GPP), we speculated that FPP

might be the primary component participating in the MVA pathway-induced BD-PMN hyperactivation. To confirm this hypothesis, we either inhibited FPP-metabolizing enzymes farnesyl-diphosphate

farnesyltransferase (FDFT1) (squalene synthase), via zaragozic acid (ZGA) and BPH-652, or activated FDFT1 function via ferroptosis inducing 56 (FIN56)26. The results demonstrated that

intracellular accumulation of FPP increased the production of proinflammatory cytokines in BD-PMN (Fig. 2C, D), and vice versa (Fig. 2E), both of which suggested a proinflammatory role of

FPP in PMN. Notably, none of the aforementioned inhibitors or agonists had any effect on PMN viability (Supplementary Figs. 2A, B). To conclude, these results suggested a proinflammatory

role of the MVA pathway, especially its metabolite FPP, in PMN activation and inflammation. FPP LEVELS WERE SIGNIFICANTLY HIGHER IN BD THAN IN HC AND CORRELATED WITH BD DISEASE ACTIVITY PMN

is particularly prone to undergo cell necrosis, mainly in the form of NETosis under inflammatory conditions19,20, which release DAMPs contributing to BD15,16,17,19. We further investigated

the abundance of FPP in both serum and total PMN lysis samples by targeted liquid chromatography-mass spectrometry (LC-MS). The relative levels of FPP were elevated in both types of samples

from active BD patients compared to those from HC (Fig. 2F). Notably, our longitudinal follow-up data showed a remarkable decrease in both PMN and serum FPP levels after BD patients achieved

remission (Fig. 2G, H). More importantly, we performed clinical evaluations and reviewed the medical records of all participants (Table 1). The results showed that FPP levels in BD-PMN were

positively correlated with C-reactive protein (CRP), and serum FPP levels in BD were positively correlated with CRP and erythrocyte sedimentation rate (ESR), indicators of BD disease

activity (Fig. 2I, J). In addition, PMN and serum FPP levels were markedly higher in patients with a Behçet’s Disease Current Activity Form (BDCAF) greater than 2 compared with those with a

BDCAF of 0 to 1 (Fig. 2K, L). We further analyzed receiver-operating characteristic (ROC) curves of serum FPP levels to investigate its diagnostic value and found that serum FPP level could

potentially differentiate BD from HC, with an area under the ROC curve (AUC) of 0.7467 (p value = 0.0041, cutoff = 1.400, sensitivity% = 65.22%, specificity% = 82.61%) (Supplementary Fig. 

3A). Moreover, we assessed the disease severity of treatment-naïve BD patients according to the well-established BD Disease Severity Score reported by Krause27,28,29. The median disease

severity score was 3 (range 2-6) for the BD-PMN cohort and 5 (range 2-7) for the BD serum cohort. Patients with a Disease Severity Score of less than or equal to 2 were characterized as mild

BD patients, while those with a score greater than 2 were defined as moderate-severe BD patients. Of note, serum FPP levels were significantly higher in moderate-severe BD than in mild BD

(Supplementary Fig. 3B). Remarkably, the serum levels of FPP were notably higher in BD patients with extracardiac vascular involvement (including deep vein thrombosis and aneurysms) than in

those with cardiac involvement (Supplementary Fig. 3C). A possible explanation may be that inflammatory indicators are particularly elevated in BD patients with extracardiac vascular

involvement. All these results drove us to examine the DAMPs-mediated direct effects of PMN inflammation and necrosis upon extracellular FPP stimulation in vitro. ENHANCED RESPONSIVENESS OF

BD-PMN TO EXTRACELLULAR FPP PROMOTES PMN HYPERACTIVATION AND VASCULAR ENDOTHELIAL INFLAMMATION AND DAMAGE We chemically synthesized FPP (method reported before24), and investigated the

expression of proinflammatory cytokines upon stimulation by FPP in PMN, lymphocytes, and monocytes from both active BD patients and HC. Notably, FPP significantly induced the production of

proinflammatory cytokines, including TNF, IL-6, IL-18, and IL-1β, in PMN but not in lymphocytes or monocytes, and BD-PMN exhibited a greater response than HC-PMN (Fig. 3A, D). FPP at

concentrations of 5 to 10 µg/mL stimulated the production of proinflammatory cytokines in PMN in a dosage-dependent manner (Fig. 3E), with no effect on their viability (Supplementary Fig. 

4A, B). When increasing the concentrations, we found that 60 μg/mL FPP induced the death of PMN, monocytes, and lymphocytes, but more cell death and a lower median lethal concentration

(LC50) of FPP were only observed in PMN from BD patients compared to HC (Supplementary Fig. 4C), especially in the form of NETosis (Fig. 3F), again with stronger responsiveness in BD-PMN

(Fig. 3G, H, and Supplementary Fig. 4D). Our published paper demonstrated a positive correlation between BD serum NETs-derived dsDNA (NETs-dsDNA) levels and CRP19. To further investigate the

involvement of FPP levels and PMN activity in the progression of BD, we measured NETs-dsDNA levels in the serum cohort and found a markedly increased NETs-dsDNA in active BD compared to HC

(Fig. 3I). Furthermore, our longitudinal follow-up data demonstrated a notable decrease in serum NETs-dsDNA levels in remission BD after appropriate treatment (Fig. 3J). Of noted, the

supernatants of BD-PMN stimulated by 60 μg/mL FPP induced a greater inflammation of vascular endothelial cell (VEC) than that of HC, as demonstrated by the upregulation of proinflammatory

cytokines and adhesion molecules (Fig. 3K). Immunofluorescence assays showed that FPP did not lead to the mortality of VECs (Supplementary Fig. 5A). Meanwhile, neither FPP alone nor

supernatants of unstimulated PMN contributed to the inflammation of VECs (Supplementary Fig. 5B, C). Taken together, FPP specifically triggers PMN hyperactivation and VEC inflammation, with

enhanced responsiveness of BD-PMN. CALCIUM AND TRP CHANNELS ARE ESSENTIAL FOR FPP-INDUCED PMN ACTIVATION FPP is known to trigger calcium influx by interacting with transient receptor

potential (TRP) channels24,30, which leads to gene transcription and cellular immune responses31. Thus, as critical controls, we examined the calcium influx in PMN by either removing

extracellular calcium or blocking the TRP channels with a universal inhibitor, ruthenium red (RR), in the presence of calcium (method reported before32). We found that both of these

conditions inhibited the FPP-induced calcium influx (Supplementary Fig. 6A), the production of cytokines (Fig. 4A), and NETosis of PMN (Fig. 4B, D, and Supplementary Fig. 6B), strongly

suggesting that calcium and TRP channels are essential for FPP to promote PMN hyperactivation. As a further validation, RR was found to significantly attenuate the effect of FPP on

NETosis-induced VEC activation (Fig. 4E). We also confirmed that NETs were essential components in the supernatants of FPP-stimulated PMN to induce VEC inflammation, as fully digesting the

NETs-dsDNA via deoxyribonuclease I (DNase I) resulted in significantly decreased expression of proinflammatory cytokines and adhesion molecules in VECs (Fig. 4C, E). Additionally, we

demonstrated that TRP channels mediated the enhanced response of BD-PMN to FPP stimulation, as RR more effectively suppressed the activation of BD-PMN in comparison to that of HC-PMN (Fig. 

4F, G). Further experiments by flow cytometry revealed a stronger calcium influx of BD-PMN than that of HC-PMN in response to FPP stimulation (Fig. 4H). Together, all these suggest that the

mechanism of FPP-induced PMN activation depends on the influx of calcium ions through TRP channels. TNF-TRPM2 AXIS ELICITS THE FPP-INDUCED INFLAMMATORY RESPONSE IN PMN To elucidate the

mechanism underlying the hyper-responsiveness nature of BD-PMN in response to FPP stimulation, we conducted a high-throughput transcriptional analysis of RR-sensitive calcium channels

including TRP channel, ryanodine receptors, cation channels sperm associated, two pore segment channels and MCU within in our RNA-sequencing database of BD patients. Remarkably elevated

expressions of four genes including _TRPM2, TRPC1, RYR1_, and _MCOLN2_ were identified (Fig. 5A). Among these, _TRPC1_ and _RYR1_ were expressed at extremely low levels in PMN (Supplementary

Table 2), and the regulation of cation flux and topology of MCOLN channels was mainly dependent on PH, rather than calcium33. In addition, the other two MCOLN family members, MCOLN1 and

MCOLN3, have been reported to be less sensitive to RR34. Consequently, we speculated that the increased FPP responsiveness in BD-PMN was mainly attributed to TRPM2, which was further

validated by qRT-PCR (Fig. 5B) and western blot (Fig. 5C). Notably, TRPM2 expression levels were significantly higher in active BD patients than those in remission (Fig. 5D), suggesting a

potential involvement of TRPM2 in BD progression. Importantly, silencing TRPM2 on BD-PMN significantly reduced the FPP-induced hyperactivation of BD-PMN, as evidenced by decreased production

of proinflammatory cytokines (Fig. 5E) and NETs (Fig. 5F–H, and Supplementary Fig. 6C), reducing the inflammation of VECs (Fig. 5I). As a critical control, the silencing efficiency of

TRPM2-specific siRNA was verified at both the transcript (Supplementary Fig. 6D) and protein levels (Supplementary Fig. 6E). To gain further insights into the molecular pathways of

FPP-induced PMN activation, we employed a series of inhibitors at graded concentrations to inhibit the downstream molecules activated by second messenger calcium ions, including PKC

inhibition with staurosporine (STS), NF-kB inhibition with PDTC, Pyk2 inhibition with PF-5662271, ERK inhibition with PD98059, cPLA2 inhibition with tanshinone I, and calcineurin inhibition

with cyclosporin A, respectively (Supplementary Fig. 7A-7F). Notably, only STS effectively reduces FPP-mediated PMN activation, suggesting a PKC-dependent molecular mechanism. Additionally,

the critical involvement of PKC molecules was further confirmed by other PKC inhibitors, including VTX27, ruboxistaurin, and Go 6983 (Supplementary Fig. 7G). Thus, all these results indicate

that FPP plays a crucial role in the opening of the TRPM2 channel, leading to the facilitated influx of calcium. Consequently, calcium ions act as pivotal second messengers, ultimately

resulting in the production of proinflammatory cytokines through the activation of PKC signaling pathway. Lastly, we investigated the mechanism driving the upregulated TRPM2 levels in BD-PMN

than HC-PMN, and in active BD patients than in those in remission. It is interesting to observe that HC-PMN cultured with active BD serum displayed significantly higher TRPM2 expression

levels compared to HC serum (Fig. 6A). To identify the serum factors responsible for enhanced TRPM2 levels in BD-PMN, we examined the main proinflammatory cytokines and found elevated TNF,

IL-6, and IFN-γ in BD serum than those in HC serum (Fig. 6B). These investigations revealed that only TNF significantly increased TRPM2 expression on PMN (Fig. 6C), but not the other

cytokines including IL-6, IL-18, IL-1β, or IFN-γ (Supplementary Figs. 8A, E). Notably, the effective reduction of TRPM2 expression by TNF-neutralizing antibodies further corroborated our

conclusion (Fig. 6D). To investigate whether TNF can enhance the TRPM2 expression in PMN in vivo, we examined the expression of TRPM2 in PMN from patients with BD treated with TNF inhibitors

(including adalimumab and infliximab). We found that TRPM2 expression was significantly reduced in BD-PMN after a median of 4 months (range 3-6) of TNF inhibitors treatment in vivo (Fig. 

6E). In addition, we performed long-term follow-up of these patients upon the treatment of TNF inhibitors with a median term of 10 months (range 7-24), of which 3 patients were lost to

follow-up. Consistently, the expression of TRPM2 in BD-PMN was significantly lower at the last visit than at baseline, although no significant difference was observed compared to the level

at the mid-term follow-up (Supplementary Fig. 8F). Furthermore, in vitro experiments showed that TNF blockade in BD serum successfully reduced FPP-induced PMN activation (Fig. 6F, G).

Therefore, the treatment with TNF inhibitors can reduce FPP-induced PMN activation by downregulating the expression of TRPM2 in BD-PMN. To explore the generalizability of our findings, we

comprehensively analyzed neutrophil RNA-seq or microarray data from publicly available databases for a variety of autoimmune and autoinflammatory diseases, including systemic lupus

erythematosus (SLE), rheumatoid arthritis (RA), inflammatory bowel disease (IBD), adult-onset Still’s disease (AOSD), and antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis

(AAV). These analyses were performed with a focus on the cholesterol synthesis pathway and TRPM2 expression levels. Notably, TRPM2 expression was significantly elevated in patients with IBD,

AOSD, AAV, and RA (Supplementary Fig. 9A). In these types of diseases, TNF inhibitors have been widely recommended by guidelines or have potential clinical benefits35,36,37,38,39,40,41,42.

In marked contrast, there was no upregulation of TRPM2 expression in patients with SLE (Supplementary Fig. 9A), in the case of which TNF inhibitors are contraindicated for clinical

usage43,44,45. Additionally, a remarkable upregulation of the cholesterol synthesis pathway was noted only in IBD patients (Supplementary Fig. 9B). Further analysis of the key enzymes of the

cholesterol synthesis pathway in IBD patients revealed that both upstream and downstream enzymes of FPP were elevated (Supplementary Fig. 9C). Notably, PMN hyperactivation is implicated in

the pathogenesis of RA and IBD, and TNF inhibitors are guideline-recommended biologics commonly used to treat patients with refractory/severe RA and IBD. We found that serum from patients

with active RA and IBD also promoted TRPM2 expression in the HC-PMN (Supplementary Fig. 10A), and further enhanced the response to FPP in the HC-PMN, as evidenced by increased production of

proinflammatory cytokines and NETs (Supplementary Fig. 10B, C). Furthermore, the blockade of TNF in RA and IBD sera effectively reduced TRPM2 expression in PMN (Supplementary Figs. 10D–E),

thereby decreasing its response to FPP (Supplementary Figs. 10F–I). The above results indicate that the TNF-TRPM2 axis is also potentially implicated in the pathogenesis of other autoimmune

and autoinflammatory diseases, thereby further extending the generality of our findings. THE FPP-TRPM2-PMN AXIS IS INVOLVED IN THE MOUSE MODEL OF VASCULITIS AND EXPERIMENTAL AUTOIMMUNE

UVEITIS To further confirm the critical role of FPP in vascular inflammation and damage, we induced vasculitis in myeloid cell-specific FPP synthetase (FPPS) knockout (FPPSflox/flox LysMcre)

mice and FPPSflox/flox mice (control group), according to the previously reported method46. Notably, FPPSflox/flox LysMcre mice demonstrated significantly reduced vascular inflammation and

damage compared with control mice, as evidenced by remarkably decreased neutrophil infiltration and vascular permeability of Evans-blue (Fig. 7A–D). In addition, a significantly lower

proportion of MPO-positive cells were noted in the skin tissues of FPPSflox/flox LysMcre mice than in control mice, as assayed by both immunohistochemistry and immunofluorescence (Fig. 

7E–H). Meanwhile, TRPM2 total knockout (TRPM2-KO) mice also demonstrated significantly reduced vascular damage compared with wild-type control mice, evidenced by reduced Evans-blue

permeability (Fig. 7I–K). All these layers of experiments revealed the critical involvement of FPP-TRPM2-PMN-axis in inducing vasculitis. In addition, after a combined consideration of the

clinical features of BD, the phenotypic features of available animal models, and the required experimental conditions, we chose a widely accepted experimental autoimmune uveitis (EAU) model

to investigate the involvement of FPP in BD pathogenesis. The EAU model is utilized because ocular inflammation, as characterized by uveitis, is reported to occur in 50–70% of BD patients47.

There are also multiple reports confirming the feasibility of inducing EAU in C57BL/6 J mice48,49,50. Of note, PMN hyperactivity has been implicated in EAU, as evidenced by excessive

production of NETs51. Based on the aforementioned thoughts and literature studies, we thus induced EAU in either FPPSflox/flox LysMcre or FPPSflox/flox control mice using interphotoreceptor

retinoid-binding protein (IRBP) peptide 1-20 according to the previously reported methods48,49,50. Induction of uveitis was assessed by ocular computerized tomography (OCT) and fundoscopy on

day 14 (Supplementary Fig. 11A), and a lower incidence of EAU was found in FPPSflox/flox LysMcre mice compared to the control mice (FPPSflox/flox LysMcre mice vs FPPSflox/flox mice: 25% vs.

66.67%). Shedding of CD62L is considered a marker of PMN activation52,53,54. On day 18, peripheral blood and eyes were collected from each type of mice, and a reduced percentage of CD11b+

Ly6G+ cells and higher CD62L expression were observed in the peripheral blood of FPPSflox/flox LysMcre mice compared with FPPSflox/flox mice (Supplementary Fig. 11B, C). Histological

assessments were also performed according to previously established criteria55. We found a reduction of PMN infiltration of ocular tissues and a lower pathological score in FPPSflox/flox

LysMcre mice compared to the control FPPSflox/flox mice (Supplementary Fig. 11D). These results provide additional support for the involvement of FPP in BD pathogenesis in disease model

studies. Overall, our data suggested the previously unrecognized mechanism by which TNF makes PMN more responsive to FPP by upregulating TRPM2 expression, which, together with the pro-TNF-

and NETs-producing effects of FPP, causes positive proinflammatory feedback loops in BD (Fig. 8). This mechanism may also contribute to other autoimmune and autoinflammatory diseases.

DISCUSSION Hyperactivation and infiltration of PMN are the predominant contributors to recurrent episodes of acute inflammation in BD14,20. Through integrated analyses, we revealed that the

MVA pathway metabolite FPP promoted BD-PMN hyperactivation in a calcium-TRPM2-dependent manner, ultimately exacerbating vascular endothelial inflammation. This report also highlighted that

the excessive levels of TNF, but not IL-6, IL-18, IL-1β, or IFN-γ, in BD serum triggered the upregulation of TRPM2, which further induced hypersensitivity to FPP in BD-PMN. All these led to

significantly increased serum levels of NETs and proinflammatory cytokines, including TNF, ultimately resulting in proinflammatory positive feedback loops. Thus, our findings uncover the

involvement of FPP in PMN hyperactivation and suggest that targeting FPP could be a potential strategy for treating BD. The MVA pathway plays a pivotal role in immune homeostasis, aberrant

of which contributes to the pathogenesis of autoimmune and autoinflammatory diseases56,57. Several studies have demonstrated that statins, inhibitors of HMGCR, a key enzyme in the initiation

of the MVA pathway, are effective in alleviating the clinical progression of patients with rheumatic diseases, including RA58,59,60, SLE61, Kawasaki’s disease62, and BD62,63. In addition,

simvastatin has been demonstrated to reduce neutrophil inflammation and NETosis in a mouse model of severe asthma, thereby alleviating lung inflammation and airway hyper-responsiveness64. As

none of the BD patients in our current cohort had a history of hyperlipidemia or statin usage, which made it challenging to find sufficient subjects for further studies. Moreover,

metabolites of the MVA pathway have been identified to enhance trained immunity in monocytes and macrophages65, function as a potent antigen for γδT cells, increasing the secretion of

proinflammatory cytokines, such as TNF66, and as a vital checkpoint to maintain T-reg functional and lineage stability57. Consequently, suppressive metabolites of the MVA pathway are

considered pivotal for the anti-inflammatory, anti-oxidant, and vascular repair properties of stains67. Here, by integrated analysis of serum metabolism and the peripheral immune cell

transcriptome, our study is the first to highlight the proinflammatory contributions of the MVA pathway in BD. Interestingly, endogenous accumulation of FPP, an end product of the MVA

pathway, mediated by ZGA treatment induces NRF2-mediated oxidative stress responses in keratinocytes68. Excessive extracellular FPP has been implicated in promoting epithelial eotaxin-3

production69, and even triggering cell necrosis as DAMP24. By inhibiting a series of key enzymes, our study proposed for the first time that FPP is a key metabolite in the MVA pathway that

contributes to the activation of BD-PMN, and confirmed the promotional effect of FPP on vascular and ocular inflammation and damage using FPPSflox/flox LysMcre mice. Mechanistically, it has

been proposed that FPP can regulate c-fos-directed DNA binding by reducing phosphorylation events of p38 and ERK, which ultimately affects Th1 cell differentiation70. However, the mechanism

by which FPP exerts danger signals is to activate the TRPM2 and TRPV3 channels, whereas the latter is mainly expressed in keratinocytes, not immune cells24,30. Although TRPM2-mediated

calcium influx has been reported to be responsible for 250 μM H2O2-induced chemokine production in monocytes71, neither monocytes nor lymphocytes were observed to respond to 5 to 10 µg/mL

FPP stimulation in our study. Additionally, the response of monocytes and lymphocytes to FPP-induced cell death was comparable in BD and HC. However, BD-PMN responded significantly higher to

FPP than HC, suggesting that PMN, but not monocytes in the peripheral blood, is the primary cell in which FPP contributes to the pathogenesis of BD. Our study is the first to propose a

TRPM2-calcium-dependent proinflammation mechanism of action of FPP in PMN. Notably, FPP specifically promoted PMN to produce more proinflammatory cytokines such as TNF, which in turn

upregulated PMN responsiveness to FPP. Moreover, FPP at the site of inflammation accelerated NETosis, leading to further release of FPP, and exacerbating the proinflammatory microenvironment

in BD. The TRPM2-dependent effect on vascular inflammation and damage was also confirmed using TRPM2-KO mice. Taken together, the generation of two types of proinflammatory feedback

centering on FPP highlights the theoretical rationale for developing metabolically-targeted drugs for BD but also complements the therapeutic mechanism of TNF inhibitors on BD from an

immunometabolism perspective. Our study has some limitations. First, we only focused on the effects of FPP on immune cells in peripheral blood, and further study of BD lesions would improve

our understanding of the role of FPP in BD pathogenesis. Second, due to the lack of specific antibodies against FPP, we merely measured FPP levels in BD serum and PMN using targeted LC-MS.

Specific antibodies are worth developing in the future for better quantification and localization of FPP in BD lesions, and for exploring the pathogenesis of BD. Third, this study provides

initial insights into the involvement of FPP with BD disease activity and severity. Further multi-center, larger long-term follow-up studies will be valuable in clarifying the significance

of FPP in BD pathogenesis. In summary, our study revealed the proinflammatory implications of FPP in fueling PMN hyperactivation, thus broadening the understanding of PMN

hyperactivation-associated diseases from an immunometabolism perspective. It also highlights a novel therapeutic mechanism of TNF inhibitors in BD and potentially other autoimmune and

autoinflammatory diseases. METHODS PATIENT ENROLLMENT Participants were recruited from the Peking Union Medical College Hospital (PUMCH) between November 2020 and February 2021. To perform

the qRT-PCR validation of differentially expressed MVA pathway key enzymes analyzed by GSEA, 20 BD and 20 HC participants were recruited between November 2020 and September 2022 from PUMCH.

Furthermore, a cohort of active BD patients was recruited between February 2021 and October 2023 and followed up until April 2024. A total of 39 BD-PMN samples and 37 BD sera were collected

for targeted mass spectrometry analysis of FPP. These included 24 active BD-PMN samples and 15 remission BD-PMN samples. Additionally, 23 active BD serum samples and 14 remission BD serum

samples were collected. All BD patients met the International Criteria for BD (ICBD), and their disease activity was assessed using ESR, CRP, and the BD Current Activity Form 2006 (BDCAF

2006)72. IBD serum samples were collected from SRRSH IBD Biobank in China (SRRSH-IBC). The PUMCH Ethical Committee approved this study (I-23PJ1123; I-24PJ0600), and all subjects provided

written informed consent. PBMC AND PMN ISOLATION Blood samples were collected from active BD patients and age- and gender-matched HCs with no personal or family history of autoimmune

diseases for mass spectrum analysis. PMN, monocytes, and lymphocytes were isolated using Ficoll-Hypaque density gradient centrifugation according to the manufacturer’s instructions at 500 x

g for 20 minutes at 24 °C. The PMN layer was harvested and red blood cell (RBC) lysis was performed using red blood cell lysis buffer (RBC lysis buffer, 10×, BioLegend) according to the 

manufacturer’s protocol. After washing with PBS, PMN was resuspended in RPMI-1640 growth medium (Gibco, USA), and their purity was confirmed by flow cytometry. Monocytes were isolated from

the PBMC layer using CD14 magnetic beads (Miltenyi, Germany, 130-050-201), and lymphocytes were defined as the remaining cells in PBMC after monocyte removal. The cells were washed with PBS

and resuspended for further experiments. CELL CULTURE AND TRANSFECTION Human microvascular endothelial cell line HMVEC was gifted from Qiong Wu lab in Tsinghua University. The cell line was

cultured in DMEM supplemented with 10% FBS, 1% non-essential amino acids, and 1% penicillin/streptomycin solution in a 5% CO2 incubator at 37 °C. To silence the expression of TRPM2, siRNAs

targeting TRPM2 were transfected into the cells according to the manufacturer’s instructions. MICE Adult C57BL/6 J mice were purchased from GemPharmatech Co., Ltd (Strain NO. N000013) in the

present study. Mice were maintained in separately ventilated cages in a specific pathogen-free (SPF) facility, in a room with standard ambient temperature and humidity, and the animals had

unrestricted access to food (Xietong Organism, 1010001) and water, with a 12-hour light/dark cycle. TRPM2-KO mice were generated with 4 nucleotide deletion mutation ACTT (ACGAGCAACACTTGGAGGT

 → ACGAGCAACGGAGGT) in exon 5 by CRISPR-Cas9 technique in C57BL/6 J background and were then backcrossed with wildtype C57BL/6 J for at least three generations before further functional

experiments. Myeloid cell-specific FPP synthase (FPPS) knockout mice (FPPSflox/floxLysMcre) and FPPSflox/flox mice were gifted from the laboratory of Prof. Yonghui Zhang26 at Tsinghua

University. Mice were euthanized by carbon dioxide (CO2) asphyxiation inhalation, and cervical dislocation was performed as a secondary euthanasia procedure, and then the tissues were

isolated. All animal experiments were approved by the Animal Research Ethics Committee and were carried out in accordance with the guidelines of the Laboratory Animal Research Center of

Tsinghua University, with an assurance identification number: 15-LWL3 and 19-LWL1 by the Institutional Animal Care and Use Committee (IACUC) of Tsinghua University. INHIBITORS Simvastatin (1

 μM, 10 μM), zoledronic acid (1 μM, 10 μM), FIN56 (1 μM, 5 μM), zaragozic acid (ZGA) (7.5 μM, 15 μM), and BPH-652 (40 μM, 80 μM) were purchased from MedChemExpress (MCE). All the inhibitors

used in this study for PKC screening including staurosporine (STS) (1 nM, 5 nM, 10 nM), PDTC (20 μM, 100 μM, 500 μM), PF-5662271 (5 nM, 10 nM, 20 nM), PD98059 (1 μM, 5 μM, 25 μM), tanshinone

I (1 μM, 5 μM, 25 μM), cyclosporin A (1 nM, 5 nM, 25 nM), VTX27 (10 nM), ruboxistaurin (10 nM), and Go 6983 (10 nM). These inhibitors were purchased from Tsinghua University active

screening platform. RNA ISOLATION AND QPCR Total RNA was extracted using an RNA-Quick Purification Kit (ES science, China, #RN001) according to the standard protocol. RNA was reverse

transcribed into cDNA using an M5 Super plus qPCR RT kit with gDNA remover (Mei5 Biotechnology, China, MF166-plus-01). Quantitative PCR (qPCR) was performed using 2X M5 HiPer SYBR Premix

EsTaq (with Tli RNaseH) (Mei5 Biotechnology, China, MF787-02). All primer sequences were presented in Supplementary Table 3. FPP STIMULATION ASSAY Cells were washed twice with PBS and

resuspended with normal saline to a final concentration of 106/mL. For the peripheral blood immune cells activation assay, the final concentrations of 0, 5, and 10 μg/mL FPP were incubated

with cells for 10 minutes, followed by adding 2 mM CaCl2 for 30 minutes, then cells were collected for follow-up assays. For VEC activation, PMN was incubated with 60 μg/mL FPP for 10 

minutes, followed by adding 2 mM CaCl2 for 30 minutes, the supernatant was collected and added to VECs at 80% density for 4 hours, the supernatant was removed and VECs were harvested for

subsequent assays. For all the experiments involving inhibitors, 30 minutes were allowed for the inhibitors to fully interact with cells before adding FPP. MASS SPECTRUM For cell samples,

equal numbers of PMN from HCs and BD were lysed using a solution of methanol and ammonium hydroxide (7:3, v/v). After sonication and deproteinization, samples were centrifuged, and the

resulting supernatant was dried and dissolved in acetonitrile and water (1:1, v/v) before injection into the LC-MS/MS system. For serum samples, 300 μL of serum from HCs and BDs was diluted

with 900 μL of 2% formic acid aqueous solution, vortexed, and loaded into solid phase extraction (SPE) cartridges pre-treated with 2% formic acid aqueous solution. After washing, elution was

performed using NH4OH:2-propanol:1-hexane (1:7:12, v/v/v). The eluted samples were evaporated and dissolved in acetonitrile and water (1:1, v/v) before injection into the LC-MS/MS system.

Qtrap 6500+ (AB Sciex, USA) coupled with H-Class UHPLC (Waters, USA) for FPP analysis. The “relative abundance” of FPP was determined based on the chromatographic area of the FPP signal. The

chromatographic area is directly proportional to the concentration of FPP in the sample. Therefore, by assessing the “relative abundance” of FPP extracted from equal serum or neutrophils,

we can determine whether FPP is up- or down-regulated. FLOW CYTOMETRY For cell death measurement after FPP stimulation by flow cytometry, cells were treated as indicated above, and the PI

(MCE, USA, HY-D0815, 20 μg/mL) signal was measured after 30 minutes of incubation. PI-positive cells were defined as dead cells. Peripheral blood was collected from the experimental

autoimmune uveitis (EAU)-induced mouse model on day 18. After washing and lysis of red blood cells, surface staining was performed for the neutrophil-specific marker Ly6G (1:100,

MultiScience Biotech, China, clone: RB6-8C5, 70-F21LY6G03-25) and the activation indicators CD11b (1:100, Biolegend, USA, clone: M1/70, 101205) and CD62L (1:100, Biolegend, USA, clone:

W18021D, 161204). Expression of these surface markers was measured by percentage of positive cells and mean fluorescence intensity (MFI). The stained cells were analyzed with a BD FACS Aria

II and FlowJo Software (Tree Star). DOUBLE-STRANDED DNA (DSDNA) QUANTIFICATION After isolating PMN from HC and BD, 6 × 105 cells were either stimulated with 0, 10, 30, or 60 μg/mL FPP for 10

 mins, followed by 2 mM CaCl2 for 30 mins at room temperature in 24-well plates. The resulting supernatant was collected after centrifugation at 500 × _g_ for 10 mins. Double-stranded DNA

(dsDNA) in the supernatants was quantified using a Quant-iT Pico-Green dsDNA assay kit (Invitrogen, USA, P7589) by incubating the samples with PicoGreen for 5 mins at room temperature and

measuring the fluorescence emission intensity at 520 nm after excitation at 485 nm. IMMUNOFLUORESCENCE AND IMMUNOHISTOCHEMICAL STAINING After isolating PMN from HC and BD, 6 × 105 cells were

seeded on poly-L-lysine-coated coverslips in 24-well plates and incubated with 60 μg/mL FPP for 10 mins, followed by 2 mM CaCl2, with or without 50 U/mL Dnase I for 60 mins. Cells were

fixed with 4% paraformaldehyde, permeabilized with 0.5% Triton X-100, and blocked with 2% BSA. Rabbit monoclonal to myeloperoxidase (MPO) (1:500, Abcam, UK, ab208670) were utilized as

primary antibodies. The paraffin-embedded skin sections of FPPSflox/floxLysMcre and FPPSflox/flox mice were stained with neutrophil-specific Ly6G (1:500, CST, USA, Clone: 1A8, 88876S) and

MPO (1:500, Abcam, UK, Clone: EPR20257, ab208670) antibodies. YF®594 Goat Anti-Rabbit IgG (1:1000, UElandy, China, Y6107L) were utilized as secondary antibodies. DAPI was used to stain the

nuclei. Confocal microscopy was used to capture images, and the percentage of PMN undergoing NETosis was calculated as the number of cells showing NETosis divided by the total number of

cells, multiplied by 100%. For immunohistochemical staining, paraffin-embedded skin sections of FPPSflox/flox LysMcre and FPPSflox/flox mice were stained with MPO antibody (1:1000, Abcam,

UK, Clone: EPR20257, ab208670) according to standard procedures and quantified by Image J software. ENZYME-LINKED IMMUNOSORBENT ASSAY (ELISA) Levels of proinflammatory cytokines, including

TNF, IL-6, and IFN-γ, in serum from BD patients and HC were measured using commercially available ELISA kits (human TNF ELISA Kit, MultiScience Biotech, China, EK182-96; human IL-6 precoated

ELISA Kit, Dakewei Biotech, China, 1110602; human IFN-γ ELISA Kit, MultiScience Biotech, China, EK180-96), following the manufacturer’s protocol. Levels of MPO-DNA complex, a specific

marker for NETs, in cell supernatant were measured by a commercially available human MPO-DNA complex ELISA Kit (MEIMIAN, China, MM-2467H1) according to the manufacturer’s instructions.

WESTERN BLOT Neutrophils were lysed on ice using RIPA buffer (Huaxingbio, China) supplemented with a protease inhibitor and phosphatase inhibitor cocktail (Thermo Scientific, USA, 78446).

The protein concentration of lysates was determined using the Pierce™ BCA Protein Assay Kits (Thermo Scientific, USA, 78446) according to the manufacturer’s instructions. Cell lysates were

subjected to 10% SDS–PAGE, and transferred onto polyvinylidene fluoride (PVDF) membranes (Bio-Rad, USA, 1620177). The membranes were blocked and then incubated with anti-TRPM2 (1:500,

SABbiotech, Clone: MGC133383, 22689) and anti-β-actin (1:1000, abcam, UK, Clone: 13E5, ab8227) overnight at 4 °C, respectively. The membranes were washed and incubated with anti-rabbit

IgG-HRP for 1 hour at room temperature. Blots were developed with chemiluminescence and detected by Tanon-5200 (Bio-Tanon, China). Gray value analysis was done by Image J (v.1.50 g, NIH)

software. PUBLIC DATA COLLECTION AND ANALYSIS Metabolomics data in BD was obtained from the National Genomics Data Center (NGDC) database (OMIX007402 and OMIX007403,

https://ngdc.cncb.ac.cn/bioproject/browse/PRJCA030253))25. Gene Set Enrichment Analysis (GSEA) was performed to identify significantly changed metabolic pathways in BD serum with the

threshold _p_-value < 0.05 and normalized enrichment score (NES) | > 1, analyzed by R package MetaboAnalystR. Bulk RNA sequencing (Bulk RNA-seq) data of BD PBMC and PMN were obtained

in the GEO dataset GSE198533 and GSE205867 from our published research, respectively11,12. The fold change, p-value, and adjusted p were calculated by R package _DESeq2_. Heatmap by R

package ComplexHeatmap was employed to demonstrate genes from cholesterol biosynthesis pathway in PMN bulk RNA-seq, normalized by rlog method in _DESeq2_. Genes with _p_ < 0.01, padj <

 0.1, and |log2Fold Change | >0.5 were considered significantly different. Single-cell RNA-sequencing (scRNA-seq) of PBMC was obtained in the GEO dataset GSE19861611 from our published

research. The fold change, _p_-value, and adjusted p were calculated by Function FindMarkers in the R package Seurat. Subgroups of immunocytes including T cells, B cells, monocytes, NK

cells, and dendritic cells were subset according to original research. GSEA of metabolic pathways in RNA-seq data was performed using the R package ClusterProfiler. Gene sets included

KEGG_STEROID_BIOSYNTHESIS, GOBP_VITAMIN_D_BIOSYNTHETIC_PROCESS, WP_CHOLESTEROL_ BIOSYNTHESIS_PATHWAY, KEGG_PRIMARY_BILE_ACID_BIOSYNTHESIS, and KEGG_STEROID_HORMONE_BIOSYNTHESIS from

Molecular Signatures Database (MSigDB). A gene set of calcium channels was obtained from the HUGO Gene Nomenclature Committee (HGNC), including Cation channels sperm associated (CATSPER),

Inositol 1,4,5-triphosphate receptors (ITPR), Ryanodine receptors (RYR), and Two pore segment channels (TPCN). We also investigated Mitochondrial Calcium Uniporter (MCU). Genes with p < 

0.01, padj <0.1, and |log2Fold Change | >0.5 were considered significantly different. INDUCTION AND EVALUATION OF VASCULITIS IN MICE The FPPSflox/floxLysMcre mice and FPPSflox/flox

mice (N = 6, 8 weeks old, 3 female and 3 male), TRPM2 knockout (KO) mice and wild-type (WT) mice (N = 4, 8 weeks old, 2 female and 2 male) were injected by intraperitoneal with 100 μL of PBS

containing 2% bovine serum albumin (BSA) (Solarbio, China, PC0001) and 1% Evans blue (MACKLIN, E808783), respectively. Then, the mice were intradermally injected with 40 μL of rabbit

anti-BSA antibody (Solarbio, China, SA263). Rabbit serum (Solarbio, China, T8570) was used as a control. 4 hours after injection, the mice were euthanized and dorsal skin samples were

collected and digitally photographed. Evans-blue dye was extracted from the dorsal skin of TRPM2 KO mice and WT mice in a vasculitis model,  and then dissolved in 50% trichloroacetic acid,

the absorbance of each group was then measured at 620 nm by spectrophotometry. Additionally, skin samples were fixed in 10% neutral buffered formalin and stained with hematoxylin for routine

microscopic examination. The severity of vasculitis was assessed by Image J analysis of the area of Evans blue permeation area and manual counting of perivascular neutrophils in HE-stained

sections under a 40x objective. INDUCTION AND EVALUATION OF EAU IN MICE The FPPSflox/flox LysMcre mice and FPPSflox/flox mice (N = 6, 8 weeks old, female) were injected subcutaneously with

an emulsion consisting of 200 μg of retinal antigen interphotoreceptor retinoid-binding protein (IRBP) peptide 1–20 (GPTHLFQPSLVLDMAKVLLD, Sangon, China) and complete Freund’s adjuvant

(sigma, USA, F5881) containing Mycobacterium tuberculosis strain H37Ra (BD Biosciences, USA, 231141) in a 1:1 volume ratio. Intraperitoneal injections of 250 μg Pertussis toxin (list lab,

USA, #180) were given on day 0 and day 2 postimmunization. The induction of uveitis was assessed by ocular computerized tomography (OCT) and fundoscopy on day 14 after immunization. On day

18, peripheral blood and eyes were collected from each type of mice. Neutrophil activation was measured by flow cytometry as indicated above. Histological assessments of eyes were performed

according to standard procedures and previously established criteria55. STATISTICS Experiments were repeated at least three times, with one representative dataset shown. Data are presented

as mean ± standard deviation (SD), median + quantile, or percentage. The Kolmogorov-Smirnov test was used to test for the normality of data distribution. Student’s t-test and non-parametric

test were used for comparisons between two groups, while paired t-test was used for comparisons before and after treatment. One-way ANOVA and two-way ANOVA tests were used with p-value

adjusted for multiple comparisons by FDR using the two-stage linear step-up procedure of Benjamini, Krieger and Yekutieli. Correlations were calculated using Pearson correlation analysis.

Statistical analyses were performed using SPSS V22.0 (IBM, USA) and GraphPad Prism V6.01 (GraphPad Software Inc, USA). A two-sided _p_-value < 0.05 was considered statistically

significant, with *_p_ < 0.05, **_p_ < 0.01, ***_p_ < 0.001, and ****_p_ < 0.0001 indicating significant differences. REPORTING SUMMARY Further information on research design is

available in the Nature Portfolio Reporting Summary linked to this article. DATA AVAILABILITY All the data supporting the findings of this study are present in the article and the

supplementary information files, or can be obtained from the corresponding author upon reasonable request. Source data are provided with this paper. CHANGE HISTORY * _ 03 JANUARY 2025 A

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and specificity of the new criteria. _J. Eur. Acad. Dermatol. Venereol._ 28, 338–347 (2014). Download references ACKNOWLEDGEMENTS This work was supported by the National Natural Science

Foundation of China (82371822-W.Z., 82171800-W.Z., 82302036-N.K., 32141004-W.L., 32430035-W.L.). This study was also supported by the Natural Science Foundation of Beijing (7232124-W.Z.,

Z230014-W.L.), Fundamental Research Funds for the Central Universities (3332023113-M.Z. and 3332023125-Z.W.), National High Level Hospital Clinical Research Funding (2022-PUMCH-C-008-W.Z.),

CAMS Innovation Fund for Medical Sciences (2023-I2M-C&T-B-049-W.Z.), Tsinghua University Spring Breeze Fund, Shenzhen Medical Research Fund (B2402012-W.L. and C2404002-W.L.), and grants

from Ministry of Science and Technology of China (2021YFC2300500-W.L. and 2021YFC2302403-C.L.). We thank the health professional staff from the Department of Rheumatology and Clinical

Immunology, Peking Union Medical College Hospital, and appreciate for the participation of all the patients and healthy volunteers in this study. AUTHOR INFORMATION Author notes * These

authors contributed equally: Menghao Zhang, Na Kang, Xin Yu. AUTHORS AND AFFILIATIONS * Department of Rheumatology and Clinical Immunology, Peking Union Medical College Hospital, Chinese

Academy of Medical Sciences & Peking Union Medical College, National Clinical Research Center for Dermatologic and Immunologic Diseases, The Ministry of Education Key Laboratory,

Beijing, China Menghao Zhang, Xin Yu, Zhimian Wang, Xiao’ou Wang, Yeling Liu, Lidan Zhao, Jinjing Liu, Hua Chen & Wenjie Zheng * State Key Laboratory of Membrane Biology, School of Life

Sciences, Institute for Immunology, China Ministry of Education Key Laboratory of Protein Sciences, Beijing Tsinghua Changgung Hospital, Tsinghua-Peking Center for Life Sciences, Tsinghua

University, Beijing, China Na Kang, Xiaoyang Zhang, Qinghui Duan, Yuxiao Zhang, Can Zhu, Ruiyu Gao, Xin Min, Cuifeng Li & Wanli Liu * School of Pharmaceutical Sciences, Beijing Advanced

Innovation Center for Structural Biology, MOE Key Laboratory of Bioorganic Phosphorus Chemistry & Chemical Biology, Tsinghua University, Beijing, China Xianqiang Ma, Qiancheng Zhao &

 Yonghui Zhang * Center for Neuroimmunology and Health Longevity, the Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, China Jin Jin * Department of gastroenterology &

Inflammatory bowel disease Center, Sir Run Run Shaw hospital, school of medicine, Zhejiang University, Hangzhou, China Qian Cao & Rongbei Liu * Department of Gastroenterology, Peking

Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China Xiaoyin Bai & Hong Yang Authors * Menghao Zhang View author

publications You can also search for this author inPubMed Google Scholar * Na Kang View author publications You can also search for this author inPubMed Google Scholar * Xin Yu View author

publications You can also search for this author inPubMed Google Scholar * Xiaoyang Zhang View author publications You can also search for this author inPubMed Google Scholar * Qinghui Duan

View author publications You can also search for this author inPubMed Google Scholar * Xianqiang Ma View author publications You can also search for this author inPubMed Google Scholar *

Qiancheng Zhao View author publications You can also search for this author inPubMed Google Scholar * Zhimian Wang View author publications You can also search for this author inPubMed 

Google Scholar * Xiao’ou Wang View author publications You can also search for this author inPubMed Google Scholar * Yeling Liu View author publications You can also search for this author

inPubMed Google Scholar * Yuxiao Zhang View author publications You can also search for this author inPubMed Google Scholar * Can Zhu View author publications You can also search for this

author inPubMed Google Scholar * Ruiyu Gao View author publications You can also search for this author inPubMed Google Scholar * Xin Min View author publications You can also search for

this author inPubMed Google Scholar * Cuifeng Li View author publications You can also search for this author inPubMed Google Scholar * Jin Jin View author publications You can also search

for this author inPubMed Google Scholar * Qian Cao View author publications You can also search for this author inPubMed Google Scholar * Rongbei Liu View author publications You can also

search for this author inPubMed Google Scholar * Xiaoyin Bai View author publications You can also search for this author inPubMed Google Scholar * Hong Yang View author publications You can

also search for this author inPubMed Google Scholar * Lidan Zhao View author publications You can also search for this author inPubMed Google Scholar * Jinjing Liu View author publications

You can also search for this author inPubMed Google Scholar * Hua Chen View author publications You can also search for this author inPubMed Google Scholar * Yonghui Zhang View author

publications You can also search for this author inPubMed Google Scholar * Wanli Liu View author publications You can also search for this author inPubMed Google Scholar * Wenjie Zheng View

author publications You can also search for this author inPubMed Google Scholar CONTRIBUTIONS W.Z. and W.L. conceptualized and designed the project and supervised the project. X.Y. conducted

the bioinformatics analysis with the help of M.Z. and Y.L. M.Z. and N.K. performed the experiments and verified the underlying data reported in the manuscript with the help of X.Z., Q.D.,

Y.Z., and C.Z. X.M. provided FPP and Q.Z. provided inhibitors and agonists of the MVA pathway. M.Z., Z.W., X.W, Y.L., Q.C., R.L., X.B., H.Y., L.Z., and J.L. participated in the

patient's enrollments, sample collection, and clinical analysis, with the help from J.J., R.G., X.M., and C.L. M.Z. and N.K. drafted the manuscript. W.Z., W.L., H.C., and Y.Z.

critically reviewed the manuscript and provided valuable input. All authors read and approved the manuscript. M.Z., N.K., and X.Y. are co-first authors, and the order of the co-first authors

was determined by workload. CORRESPONDING AUTHORS Correspondence to Wanli Liu or Wenjie Zheng. ETHICS DECLARATIONS COMPETING INTERESTS The authors declare no competing interests. PEER

REVIEW PEER REVIEW INFORMATION _Nature Communications_ thanks Yacine Boulaftali, Ricardo Silvestre and the other anonymous reviewer(s) for their contribution to the peer review of this work.

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mevalonate metabolite/TRPM2/calcium signaling axis in neutrophils to dampen vasculitis in Behçet’s disease. _Nat Commun_ 15, 9261 (2024). https://doi.org/10.1038/s41467-024-53528-3 Download

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