Tania O. Crisan

Interferon Regulatory Factor 5 plays an important role in the regulation of innate immune responses by amplifying the Nuclear Factor κB response, which is critical in gout inflammation. Furthermore, the rs4728141 polymorphism C allele was associated with both increased IRF5 expression and susceptibility to gout. We examine the association between rs4728141 and cytokine production in response to various Toll-Like Receptor ligands and describe the transcriptomic and proteomic changes observed in patients with gout and controls in relation to this polymorphism. We examine the transcriptome of freshly isolated peripheral blood mononuclear cells (PBMCs) from 93 normouricemic donors and 63 gout patients as well as serum inflammatory proteome in 197 control and 195 gout samples. Stimulation experiments of freshly isolated human PBMCs were performed over 24 h, followed by RNA-sequencing in gout patients and cytokine production measurement by ELISA in normouricemic donors and gout patients. The rs4728141 C allele was associated with increased IL-1β expression in unstimulated PBMCs of controls, but not in gout. No association between the polymorphism and serum inflammatory proteome was found. As expected, an increased IRF5 expression was observed in stimulated PBMCs of rs4728141 C allele carriers in response to several stimulations. Interestingly, IL-1β production was specifically enhanced in association to the rs4728141 C allele when cells were stimulated with palmitate with or without monosodium urate crystals. This pattern of cytokine production shows a functional impact of rs4728141 in gout through altered IL-1β production.

Background Interferon Regulatory Factor 5 plays an important role in the regulation of innate immune responses by amplifying the Nuclear Factor κB response, which is critical in gout inflammation. Furthermore, the rs4728141 polymorphism C allele was associated with both increased IRF5 expression and susceptibility to gout. We examine the association between rs4728141 and cytokine production in response to various Toll-Like Receptor ligands and describe the transcriptomic and proteomic changes observed in patients with gout and controls in relation to this polymorphism. Materials and Methods We examine the transcriptome of freshly isolated peripheral blood mononuclear cells (PBMCs) from 93 healthy donors and 63 gout patients as well as serum inflammatory proteome in 197 control and 195 gout samples. 24-hour stimulation experiments of freshly isolated human PBMCs were performed, followed by RNA-sequencing in gout patients and cytokine production measurement by ELISA in healthy donors and gout patients. Results The rs4728141 C allele was associated with increased IL-1β expression in unstimulated PBMCs of controls, but not in gout. No association between the polymorphism and serum inflammatory proteome was found. As expected, an increased IRF5 expression was observed in stimulated PBMCs of rs4728141 C allele carriers in response to several stimulations. Interestingly, IL-1β production was specifically enhanced in association to the rs4728141 C allele when cells were stimulated with palmitate with or without monosodium urate crystals. Conclusions The recently identified gout risk allele C rs4728141, which maps to the vicinity to the IRF5 gene is associated with elevated proinflammatory responses and this was specifically observed in response to gout-relevant stimulations such as palmitate in presence or absence of monosodium urate crystals. This pattern of cytokine production shows a functional impact of rs4728141 in gout through altered IL-1β production, a main driver of inflammation in gout.

Introduction: Monosodium urate (MSU) crystals are the primary cause of gout, however the knowledge on MSU crystals exposure and inflammatory changes found in immune cells is diverse, with reports ranging from very limited inflammatory effects to substantial reprogramming of the cells induced by MSU crystals alone. We examine the IL-1β production patterns and the transcriptomic signature in response to MSU crystals alone or in combination with TLR ligands in freshly isolated primary human peripheral blood mononuclear cells (PBMCs). Materials and Methods: PBMCs were isolated by density gradient centrifugation and were stimulated for 24 hours with palmitate in the presence or absence of MSU crystals, followed by cytokine production measurement by ELISA. Two bulk RNA-sequencing analyses were performed independently following the same experimental conditions using PBMCs of patients with gout stimulated with medium control, palmitate and LPS in the presence or absence of MSU crystals. Results: MSU crystals alone induced a small but significant increase in IL-1β production in human PBMCs. IL-1β production was significantly increased when PBMCs were stimulated with palmitate and this was further amplified by the palmitate-MSU combination. Of high interest, MSU crystals alone or in combination with other stimuli caused no significant transcriptomic alterations. Conclusions: We confirm the synergistic effect of MSU crystals with palmitate that leads to higher IL-1β production. Transcriptomic analysis shows that MSU crystal exposure is not associated with major transcriptional changes in PBMCs. This suggests that the production of IL-1β in response to MSU crystals may largely be regulated at the post-transcriptional level and additional synergistic stimuli are likely required to fully explain the inflammatory response observed clinically in gout. Moreover, this could also bear relevance for other metabolic disorders associated to hyperuricemia where asymptomatic MSU crystal deposition may be present.

Gout is a chronic inflammatory arthritis driven by monosodium urate crystal deposition and the NACHT, LRR, and PYD domains-containing protein 3 (NLRP3) inflammasome activation, leading to interleukin-1β (IL-1β)-mediated inflammation. Recent studies reveal that hyperuricemia induces a state of immunological memory in innate immune cells through persistent epigenetic and metabolic reprogramming of monocytes and macrophages. These alterations enhance the responsiveness of innate immune cells, leading to exaggerated inflammatory reactions upon subsequent stimulation. This review synthesizes recent studies that elucidate how metabolic shifts (e.g., increased glycolysis and fumarate accumulation) and epigenetic changes (e.g., altered histone methylation and DNA methylation) reinforce this pathogenic memory. Importantly, these mechanistic insights provide the rationale for emerging therapeutic strategies: IL-1β inhibitors aim to interrupt the central inflammatory axis; metabolic modulators (e.g., metformin, statins) seek to reverse the trained metabolic state; and epigenetic therapies [e.g., histone deacetylase (HDAC) or DNA methyltransferase (DNMT) inhibitors] hold potential to reset dysregulated immune programming. Collectively, this review argues that multi-layered intervention, such as cytokine blockade for acute control, coupled with metabolic or epigenetic remodeling for long-term reprogramming, could yield sustained disease suppression and reduced flare frequency in gout.

Gout is caused by the response of the innate immune system to monosodium urate (MSU) crystals. A recent gout GWAS identified a signal of genetic association at a locus encompassing IL1RN-IL1F10. Colocalisation analysis using Genotype Tissue Expression Database (GTEx) eQTL data showed that the signal overlaps with genetic control of IL1RN/IL1F10 gene expression. We assess the functional implications of IL1RN rs9973741, the lead gout-associated variant. We conducted functional validation of IL1RN rs9973741 in patients with gout and controls. The transcription level of IL1RN/IL1F10 was investigated in unstimulated or MSU-crystal co-stimulated PBMCs. Ex vivo functional assays were performed in PBMCs stimulated with C16 + MSU crystals or LPS for 24 h. Cytokine levels were assessed by ELISA. In unstimulated PBMCs, no association of IL1RN rs9973741 (gout risk allele G) to IL1RN expression was observed while IL-1F10 was hindered by low expression at both transcriptional and protein levels. However, G allele carriers showed lower IL1RN expression in PBMCs stimulated with C16/MSU crystal and lower concentrations of circulating IL-1Ra in both controls and gout patients. PBMCs depicted less spontaneous IL-1Ra release in GG homozygous controls and lower IL-1Ra production in response to C16 + MSU crystal costimulation in patients with gout. The G allele was associated with elevated IL-1β cytokine production in response to C16 + MSU crystal stimulation in controls. The gout risk allele G associates with lower circulating IL-1Ra, lower IL-1Ra production in PBMC assays and elevated IL-1β production in PBMCs challenged with C16 + MSU crystals but not in unchallenged cells. Our data indicate that the genetic signal that associates with gout at IL1RN-IL1F10 region functions to alter expression of IL-1Ra when stimulated by MSU crystals.

Gout is a crystal-induced arthropathy in which monosodium urate (MSU) crystals precipitate within joints as a result of persistent hyperuricemia and elicit an inflammatory response. An intriguing aspect is the occurrence of gout in only 10–15% of hyperuricemic individuals, suggesting the presence of additional risk factors. Although MSU crystal deposition is widely recognized as the cause of gout flares, the variability in initiating the inflammatory response to hyperuricemia and MSU deposition is not well understood. Several studies bring up-to-date information about the environmental and genetic influences on the progression towards clinical gout. Elevated urate concentrations and exposure to different external factors precipitate gout flares, highlighting the potential involvement of epigenetic mechanisms in gouty inflammation. A better understanding of the alteration of the epigenetic landscape in gout may provide new perspectives on the dysregulated inflammatory response. In this review, we focus on understanding the current view of the role of epigenomic reprogramming in gout and the mechanistic pathways of action.

Type I interferons (IFN1s) mediate innate responses to microbial stimuli and regulate interleukin (IL)‐1 and IL‐1 receptor antagonist (Ra) production in human cells. This study explores interferon‐stimulated gene (ISG) alterations in the transcriptome of patients with gout and stimulated human primary cells in vitro in relation to serum urate concentrations. Peripheral blood mononuclear cells (PBMCs) and monocytes of patients with gout were primed in vitro with soluble urate, followed by lipopolysaccharide (LPS) stimulation. Separately, PBMCs were stimulated with various toll‐like receptor (TLR) ligands. RNA sequencing and IL‐1Ra cytokine measurement were performed. STAT1 phosphorylation was assessed in urate‐treated monocytes. Cytokine responses to IFN‐β were evaluated in PBMCs cultured with or without urate and restimulated with LPS and monosodium urate (MSU) crystals. Transcriptomics revealed suppressed IFN‐related signalling pathways in urate‐exposed PBMCs or monocytes which was supported by diminishment of phosphorylated STAT1. The stimulation of PBMCs with IFN‐β did not modify the urate‐induced inflammation. Interestingly, in vivo, serum urate concentrations were inversely correlated to in vitro ISG expression upon stimulations with TLR ligands. These findings support a deficient IFN1 signalling in the presence of elevated serum urate concentrations, which could translate to increased susceptibility to infections.

Gout is a prevalent form of inflammatory arthritis caused by the crystallization of uric acid in the joints and soft tissues, leading to acute, painful attacks. Activation of the NLRP3 inflammasome in mononuclear cells, along with inflammasome-independent pathways, is responsible for the inflammatory phenotype in gout. Research into the different aspects of gout pathophysiology and potential treatment options is ongoing. This review highlights some of the basic research published in the 12 months following the 2022 Gout, Hyperuricemia, and Crystal-Associated Disease Network (G-CAN) conference and focuses on mechanisms of inflammation, encompassing pro- and anti-inflammatory pathways, as well as the exploration of various biological systems, such as single-cell transcriptomics, proteomics, metabolomics, and microbiome analyses.

The mRNA vaccine against COVID-19 protects against severe disease by the induction of robust humoral and cellular responses. Recent studies have shown the capacity of some vaccines to induce enduring non-specific innate immune responses by the induction of trained immunity, augmenting protection against unrelated pathogens. This study aimed to assess whether the mRNA vaccine BNT162b2 can induce lasting non-specific immune responses in myeloid cells following a three-dose vaccination scheme. In a sample size consisting of 20 healthy individuals from Romania, we assessed inflammatory proteins using the Olink^® Target 96 Inflammation panel, as well as ex vivo cytokine responses following stimulations with unrelated PRR ligands. We assessed the vaccine-induced non-specific systemic inflammation and functional adaptations of myeloid cells. Our results revealed the induction of a stimulus- and cytokine-dependent innate immune memory phenotype that became apparent after the booster dose and was maintained eight months later in the absence of systemic inflammation.

Gout, an inflammatory disease orchestrated by interleukin-1β activation and release, is more prevalent in men. The clinical profiles of patients with gout report differences by sex. This study aims to investigate sex-specific cytokine profiles in circulation and in stimulated peripheral blood mononuclear cells (PBMCs) of patients with gout and controls. Participants included in the gout group met the criteria of the American College of Rheumatology/European League Against Rheumatism (ACR/EULAR). The control group included individuals with varying levels of serum urate and absence of gout. PBMCs were stimulated in vitro for 24 h with various TLR ligands. Cytokines were determined in culture supernatants and plasma. Plasma IL-1Ra and high-sensitivity C-reactive protein (hsCRP) were higher in men with gout compared to men without gout whereas no significant differences in circulating cytokines were observed in women. PBMCs of patients with gout showed higher cytokine production of IL-1β, IL-1Ra, and TNF-α following 24 h stimulation, predominantly observed in women. We identified sex-specific cytokine production in gout in response to in vitro stimulation. While men with gout had higher levels of circulating cytokines, stimulated PBMCs of women with gout show an enhanced capacity for cytokine production. These data may suggest potentially different regulatory mechanisms of inflammation in men and women with gout.