Anti-inflammatory effect of Dieffenbachia standleyi Croat extract on lipopolysaccharide-stimulated murine macrophages

ABSTRACT

Dieffenbachia standleyi Croat is a plant belonging to Araceae and is native to Central America, and South America. While there have been studies on the Dieffenbachia species, Dieffenbachia standleyi Croat has yet to be studied in depth. Dieffenbachia was used as a folk remedy to prevent inflammation and treat gout, erectile dysfunction, and coldness. Thus, this study investigates the pharmacological function of Dieffenbachia standleyi Croat's methanol extract (DSE). When RAW 264.7 macrophages were treated with DSE and stimulated with lipopolysaccharide, DSE reduced the expression of inducible nitric oxide (NO) synthase and cyclooxygenase-2, thereby decreasing NO and prostaglandin E2 production and pro-inflammatory cytokines including IL-1β, IL-18, IL-6, and TNF-α. By confirming DSE's anti-inflammatory mechanisms, we determined that DSE inhibits IκBα phosphorylation in the nuclear factor κappa B pathway, and inflammasome formation, without affecting MAPK pathways. This study suggests that Dieffenbachia standleyi Croat is a potential anti-inflammatory agent for inflammatory diseases.

KEYWORDS:

• Dieffenbachia standleyi
• inflammation
• NF-kB
• inflammasome

1. Introduction

Inflammation serves as a vital defense mechanism within the human body, orchestrating the elimination of harmful stimuli, including pathogens, toxins, and damaged cellular components (Chen et al., 2018; Pham et al., 2021). Inflammatory reactions are characterized by acute and chronic phases. Acute inflammation involves an upregulation of blood flow and vascular permeability, driven by the accumulation of inflammatory mediators. In contrast, chronic inflammation results from homeostatic imbalances, often occurring without the typical triggers of infection or wounds, and can contribute to various illnesses, including cardiovascular disease and Crohn's disease (Chen et al., 2018; Furman et al., 2019). Amid the pursuit of preventive and therapeutic measures against inflammation, numerous studies have explored the potential of herbal extracts and natural products (Bai et al., 2021; Hussain et al., 2020; Kim et al., 2017; Kim et al., 2021). However, this research is often conducted in response to the need for safer and more effective anti-inflammatory agents.

Dieffenbachia, a widely cultivated ornamental plant from the Araceae family, indigenous to tropical regions (Cumpston et al., 2003), presents a unique case. Despite its popularity and presence in many households, Dieffenbachia is among the most toxic plants known, posing significant risks to children, adults, and pets (Arditti & Rodriguez, 1982). Its toxicity is attributed to the presence of calcium oxalate crystals that cause tissue injury and release kinin involved in inflammation and pain (Arditti & Rodriguez, 1982; Dore, 1963). Additionally, histamine release has been implicated in Dieffenbachia toxicity (Barnes & Fox, 1955; Fochtman et al., 1969; Ladeira et al., 1975; Rizzini & Occhioni, 1957).

Historically, Dieffenbachia has found various applications, ranging from the creation of poisoned arrows by Amazon indigenous groups to contraceptive use among Caribbean natives and World War II-era Germans (Arditti & Rodriguez, 1982; Fochtman et al., 1969). It has also been explored for its potential in treating conditions such as gout, erectile dysfunction, and coldness (Barnes & Fox, 1955). However, while Dieffenbachia seguine has been studied for its anti-inflammatory properties, there is currently no research on Dieffenbachia standleyi Croat (DS), a species distributed in Honduras, Central America, and South America. In light of the reported anti-inflammatory effects of Dieffenbachia seguine, our study investigates the potential anti-inflammatory properties of Dieffenbachia standleyi Croat methanol extract (DSE) on RAW 264.7 murine macrophages.

2. Materials and methods

2.1. Cell culture

The murine macrophage RAW 264.7 cell line was obtained from the Korea Cell Line Bank (Seoul, Korea). Cells were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM; Welgene, Gyeongsan, Korea) supplemented with 10% heat-inactivated fetal bovine serum (Welgene), 100 U/mL penicillin, and 100 µg/mL streptomycin (P/S; Welgene) at 37 °C in a 5% CO₂ humidified incubator.

2.2. Plant material and extract construction

The leaves of Dieffenbachia standleyi Croat were collected in San Dionisio district, Matagalpa city, Nicaragua in November 2015 and identified by curator, Indiana Coronado, herbarium of National Autonomous University of Nicaragua at Leon. A voucher specimen (accession number KRIB 0073802) of the plant was deposited at the herbarium of KRIBB. The leaf powder of Dieffenbachia standleyi (100 g) were extracted with 1 L of 99.9%(v/v) methanol repeating sonication (15 min) and resting (2 h) for 3 days at a temperature lower than 45 °C to prevent any degradation of compounds caused by high temperature. The resultant product was filtered with non-fluorescence cottons, and concentrated by rotary evaporator (N-1000SWD, EYELA) under reduced pressure at a temperature lower than 45 °C. Finally, total 7.6 g of methanol extract of Dieffenbachia standleyi was obtained by freeze-drying. The extract was dissolved using a 160 mg/mL dimethyl sulfoxide concentration (DMSO; Duchefa Biochemie, Haarlem, The Netherlands) and used as a stock solution. In addition, 25, 50, 100, and 200 (µg/mL) concentrations were used as working solutions, and the final DMSO concentration in media was set to 0.5% (v/v).

2.3. Cell viability assay

RAW 264.7 cells were seeded onto a 96-well plate (5 × 10⁴ cells/100 µL per well) and incubated for 24 h. Cells were then treated with a 0-200 µg/mL DSE concentration for 4 h and stimulated with 1 µg/mL LPS for 20 h. 10 µL 3-(4,5-dimethylthiazol-2-yl)−2,5-diphenyltetrazolium bromide (MTT; Sigma-Aldrich, St. Louis, MO, USA) solution with a 5 mg/mL concentration in phosphate-buffered saline (PBS) was added to each well, and cells were incubated at 37 °C in 5% CO₂ humidified air for 4 h. Next, 100 µL of a 0.04 N HCl concentration in isopropanol was added to each well to dissolve the formazan crystals formed by the MTT solution. Lastly, absorbance was read at 540 nm by an EMax microplate reader (Molecular Devices, Sunnyvale, CA, USA) to determine cell viability.

2.4. NO assay

RAW 264.7 cells were seeded onto a 6-well plate (1 × 10⁶ cells/2 mL per well) and incubated for 24 h. Next, cells were treated with a 0-200 µg/mL DSE concentration for 4 h and stimulated with 1 µg/mL LPS for 20 h. After collecting the supernatant, 50 µL of supernatant and 50 µL of 40 mg/mL Griess agent were put into the 96-well plate. For a standard curve, NO level was measured using NaNO₂ (Sodium nitrite; Junsei Chemical Co., LTD., Chuo-ku, Tokyo, Japan) ranging from 0 µM to 100 µM. The absorbance was read at 540 nm by an EMax microplate to determine NO production.

2.5. Enzyme-linked immunosorbent assay (ELISA)

RAW 264.7 cells were seeded onto a 6-well plate (1 × 10⁶ cells/2 mL per well) and incubated for 24 h. Cells were treated with 0-200 µg/mL DSE for 4 h and stimulated with 1 µg/mL LPS for 20 h. After collecting the supernatant, the sandwich ELISA method was used to measure interleukin (IL)−1β, IL-6, IL-18, and tumor necrosis factor-α (TNF-α) secretion levels from RAW 264.7 cells. Briefly, primary antibodies were coated on a 96-well plate at 4 °C overnight. The next day, the plate was washed with PBS containing 0.05% (v/v) Tween 20 (VWR, Radnor, PA, USA) (0.05% PBST) and blocked by PBS containing 3% (w/v) BSA for 1 h and 30 min (MP Biomedicals, Irvine, CA, USA). Afterward, 50 µL supernatant was loaded into each well and incubated at 4 °C overnight. The plate was then washed with 0.05% PBST and incubated with biotinylated antibodies at room temperature for 30 min. After washing with 0.05% PBST, alkaline phosphatase-conjugated streptavidin (Jackson ImmunoResearch, West Grove, PA, USA) was added to each well and incubated at room temperature for 20 min. Finally, 20 mg/mL of 4-Nitrophenyl phosphate disodium salt hexahydrate (Sigma-Aldrich) was added to each well. Cytokine measurements were detected with an EMax microplate reader at 405 nm. For the standard curve, recombinant murine IL-1β, IL-6, TNF-α (Peprotech, Rocky Hill, NJ, USA), and IL-18 (BioLegend, San Diego, CA, USA) were used. The PGE₂ ELISA kit measured PGE₂ following the manufacturer's instructions. (Cayman Chemical, Ann Arbor, MI, USA). The list of antibodies used for ELISA is presented in Table 1.

Table 1. List of ELISA antibodies.

Manufacturer	Cytokine	Species Reactivity	Label	Source	Concentration used
MBL, MINATO-KU, TK, JP	IL-18	Mouse	Unconjugated	Monoclonal Rat IgG2A	4 µg/mL
MBL, MINATO-KU, TK, JP	IL-18	Mouse	Biotin	Monoclonal Rat IgG1	2 µg/mL
eBioscience, San Diego, CA, USA	IL-1β	Mouse	Unconjugated	Armenian hamster	3 µg/mL
eBioscience, San Diego, CA, USA	IL-1β	Mouse	Biotin	Rabbit	4 µg/mL
BD Biosciences, Franklin Lakes, NJ, USA	IL-6	Mouse	Unconjugated	Rat	3 µg/mL
BD Biosciences, Franklin Lakes, NJ, USA	IL-6	Mouse	Biotin	Rat	2 µg/mL
eBioscience, San Diego, CA, USA	TNF-α	Mouse	Unconjugated	Armenian hamster	2 µg/mL
eBioscience, San Diego, CA, USA	TNF-α	Mouse	Biotin	Rabbit	2 µg/mL

2.6. Western blotting

RAW 264.7 cells were pretreated with DSE for 4 h, followed by stimulation with LPS for 15 min. Cells were then dissolved in a RIPA Lysis and Extraction buffer (Thermo Fisher Scientific, Waltham, MA, USA) at 4 °C for 30 min and centrifuged at 4 °C for 15 min at 14000 × g. Proteins were quantified using the Pierce BCA Protein Assay Kit (Thermo Fisher Scientific) and loaded into 10% SDS-polyacrylamide gel electrophoresis gels. Separated proteins were transferred to the Amersham Hybrid P 0.2-μm PVDF membrane (Cytiva, Marlborough, MA, USA). The membrane was blocked at room temperature for 1 h with a blocking buffer made by adding 5% (w/v) skim milk (BD Biosciences) to Tris-buffer saline (TBS) containing 0.1% (v/v) Tween-20 (0.1% TBST) followed by washing with 0.5% (v/v) TBST. The membrane was then incubated at 4 °C overnight with primary Ab, washed with 0.5% (v/v) TBST three times, and incubated with secondary Ab for 2 h at room temperature. After washing with 0.5% (v/v) TBST three times, the membrane was reacted with a WESTSAVE Femto ECL western blot detection kit (AbFrontier, Seoul, Korea). Detected protein bands were measured using FUSION SOLOX and Evolution-Capt software (Vilber, Paris, France). All protein bands were normalized by GAPDH expression. For measuring iNOS and COX-2, RAW 264.7 cells were treated with DSE for 4 h and stimulated with LPS for 20 h. NOD-like receptor protein (NLRP) 3, apoptosis-associated speck-like protein containing CARD domain (ASC), and cleaved caspase-1 were treated with LPS for 5 h and ATP (5 mM) for 30 min. The subsequent experimental process is the same as above. The list of antibodies used for western blotting is represented in Table 2.

Table 2. List of Western Blot antibodies.

Target	Host	Dilution Ratio	Manufacturer	Cat. No
iNOS	Rabbit	1: 1000	Santa Cruz Biotechnology, Dallas, TX, USA	sc-651
COX-2	Rabbit	1: 1000	Bioss Antibodies, Woburn, MA, USA	bs-0732R
JNK	Mouse	1: 1000	Santa Cruz Biotechnology, Dallas, TX, USA	sc-7345
p-JNK	Rabbit	1: 1000	Cell Signaling, Danvers, MA, USA	4668
ERK1/2	Mouse	1: 1000	Santa Cruz Biotechnology, Dallas, TX, USA	sc-514302
p-ERK1/2	Rabbit	1: 2000	Cell Signaling, Danvers, MA, USA	4370
p38	Rabbit	1: 4000	BioLegend, San Diego, CA, USA	622403
p-p38	Rabbit	1: 1000	Cell Signaling, Danvers, MA, USA	4631
IκBα	Rabbit	1: 1000	Cell Signaling, Danvers, MA, USA	4812
p-IκBα	Mouse	1: 1000	Cell Signaling, Danvers, MA, USA	9246
GAPDH	Rabbit	1: 1000	Santa Cruz Biotechnology, Dallas, TX, USA	sc-25778
NLRP3	Rabbit	1: 1000	Cell Signaling, Danvers, MA, USA	15101
ASC	Rabbit	1: 1000	Cell Signaling, Danvers, MA, USA	67824
Cleaved caspase-1	Rabbit	1: 1000	Cell Signaling, Danvers, MA, USA	89332

Inducible Nitric Oxide Synthase (iNOS); Cyclooxygenase-2 (COX-2); c-Jun N-Terminal Kinase (JNK); Extracellular Signal-Related Kinase (ERK); inhibitor α protein (IκBα); Nod-Like Receptor Protein (NLRP); Assorted Speck-Like Protein Containing CARD Domain (ASC)

2.7. Statistical analyses

All data are presented as mean ± standard error of means (SEM). Statistical significance between groups was determined using the analysis of variance (ANOVA) and post hoc Tukey's test. p < 0.05 was statistically significant. All statistical analyses were conducted by Prism 5.0 (GraphPad Software, San Diego, CA, USA).

3. Results

3.1. DSE cytotoxicity in RAW 264.7 cells

MTT assay was performed to confirm DSE cytotoxicity in RAW 264.7 cells. Four different DSE concentrations (25, 50, 100, and 200 µg/mL) were prepared, and cytotoxicity was measured. The experimental data showed that DSE did not show cytotoxicity at every concentration level tested (Figure 1).

Figure 1. DSE’s effect on RAW 264.7 cell viability. RAW 264.7 cells were pretreated with DSE for 4 h and stimulated with LPS for 20 h. The MTT assays measured cell viability. The data are shown as the mean ± SEM of three MTT assays with similar results. Dieffenbachia standleyi Croat’s methanol extract (DSE); lipopolysaccharide (LPS); Standard Error of Means (SEM).

3.2. DSE’s effect on inhibiting inflammatory mediators and pro-inflammatory cytokines

After DSE treatment, inflammatory mediators NO and PGE₂ level differences were observed in RAW 264.7 cells. Compared with the control, DSE significantly decreased NO production at 100 and 200 µg/mL, and decreased iNOS expression at 200 µg/mL (Figure 2). DSE treatment also decreased PGE₂ production at all concentrations tested dose-dependently, while a significant COX-2 expression decrease was observed only at 200 µg/mL (Figure 2). This data suggests that DSE influences anti-inflammatory activity by regulating inflammatory mediators.

Figure 2. DSE’s effect on RAW 264.7 cells NO production and iNOS expression. RAW 264.7 cells were pretreated with DSE for 4 h and stimulated with LPS for 20 h. The culture supernatant and proteins were collected for (A) iNOS expression, (B) NO production, (C) COX-2 expression, and (D) PGE₂ production, respectively. The data are presented as mean ± SEMs of three different independent results. The data were analyzed through one-way ANOVA (P < 0.05) followed by Tukey’s multiple comparison test. Dieffenbachia standleyi Croat’s methanol extract (DSE); Nitric Oxide (NO); Inducible Nitric Oxide Synthase (iNOS); lipopolysaccharide (LPS); Cyclooxygenase-2 (COX-2); Prostaglandin E₂ (PGE₂); Standard Error of Means (SEM); Analysis of Variance (ANOVA).

Macrophages secrete pro-inflammatory cytokines such as TNF-α, IL-6, IL-1β, and IL-18 upon LPS stimulation, and ELISA measured production in DSE-treated macrophages. The data indicated that DSE significantly inhibited IL-6 and TNF-α production at 200 µg/mL (Figure 3A, B). Furthermore, DSE dose-dependently decreased IL-1β and IL-18 production (Figure 3C, D). Our results establish that DSE exerts anti-inflammatory effects by inhibiting pro-inflammatory cytokine production.

Figure 3. DSE’s effect on RAW 264.7 cells pro-inflammatory cytokine production. RAW 264.7 cells were pretreated with DSE for 4 h and stimulated with LPS for 20 h. The culture supernatants were collected, and (A) IL-6, (B) TNF-α, (C) IL-1β, and (D) IL-18 were measured by Sandwich-ELISA. The data are presented as mean ± SEMs of three different independent results. The data were analyzed through one-way ANOVA (P < 0.05) followed by Tukey’s multiple comparison test. Dieffenbachia standleyi Croat’s methanol extract (DSE); lipopolysaccharide (LPS); Interleukin (IL); Tumor Necrosis Factor (TNF); Enzyme-Linked Immunosorbent Assay (ELISA); Standard Error of Means (SEM); Analysis of Variance (ANOVA).

3.3. DSE’s effect on MAPK and NF-κB signaling pathways

To determine how DSE exerts an anti-inflammatory effect, we used Western blot analysis to examine protein factor levels related to MAPK pathways: c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK) ½, and p38 pathways (Figure 4). Total and phosphorylated forms of JNK, ERK1/2, and p38 were measured, and the phosphorylated form/total-form ratio was quantified. The data revealed that DSE treatment did not inhibit JNK, ERK1/2, and p38 phosphorylation, suggesting that DSE’s anti-inflammatory effect is independent of MAPK pathways.

Figure 4. DSE’s effect on MAPK and NF-κB pathways. RAW 264.7 cells were pretreated with DSE for 4 h and then stimulated with LPS for 15 min. (A) JNK, phosphor-JNK, ERK, phosphor-ERK, p38, and phosphor-p38 expressions were measured by Western blotting, (B) and JNK, ERK, and p38 phosphorylated form to total form ratio was calculated. (C) IκBα and phosphor-IκBα expressions were also measured by Western blotting and (D) quantified. The data are presented as mean ± SEMs of three different independent results. The data were analyzed through ANOVA (P < 0.05) and Tukey’s multiple comparison test. Dieffenbachia standleyi Croat’s methanol extract (DSE); mitogen-activated protein kinase (MAPK); Nuclear Factor κappa B (NFκB); lipopolysaccharide (LPS); c-Jun N-Terminal Kinase (JNK); Extracellular Signal-Related Kinase (ERK); inhibitor α protein (IκBα); Standard Error of Means (SEM); Analysis of Variance (ANOVA).

Inhibitor α protein (IκBα) is a regulatory protein that inhibits nuclear factors κappa B (NF-κB) by binding and keeping them inactive in the cytoplasm. Upon LPS stimulation, IκBα is phosphorylated, ubiquitinated, and decomposed, activating and translocating NF-κB into the nucleus. Therefore, IκBα and phosphor-IκBα levels were measured and quantified following treatment with four DSE concentrations in LPS-stimulated RAW 264.7 cells. The data showed that 100 and 200 µg/mL of DSE treatment inhibited IκBα phosphorylation, suggesting that DSE exerts anti-inflammatory effects by regulating the NF-κB pathway.

3.4. DSE’s effect on inflammasome

To further investigate how IL-1β and IL-18 were significantly reduced, DSE-treated RAW 264.7 cell inflammasome components were measured, including NLRP3 (NOD-, LRR- and pyrin domain-containing protein 3), ASC (apoptosis-associated speck-like protein containing a CARD), and cleaved caspase-1. As shown in Figure 5, DSE significantly reduced NLRP3, ASC, and cleaved caspase-1 levels. This implies that DSE interferes with caspase-1 activity in inflammasomes, resulting in IL-1β and IL-18 reduction.

Figure 5. DSE’s effect on inflammasomes. RAW 264.7 cells were pretreated with DSE for 4 h, stimulated with LPS for 5 h, then ATP (5 mM) for 30 min. NLRP3, ASC, and cleaved caspase-1 expressions were measured by (A) western blotting (B, C, D) and quantified. The data are presented as mean ± SEMs of three different independent results. The data were analyzed through ANOVA (P < 0.05) and Tukey’s multiple comparison test. Dieffenbachia standleyi Croat’s methanol extract (DSE); lipopolysaccharide (LPS); Nod-Like Receptor Protein (NLRP); Assorted Speck-Like Standard Error of Means (SEM); Analysis of Variance (ANOVA); Protein Containing CARD Domain (ASC).

4. Discussion

Dieffenbachia has long been recognized as a poisonous plant with toxicity attributed to calcium oxalate crystals. This was confirmed as non-inflammatory when rabbit eyes were treated with a 5% calcium oxalate suspension (Arditti & Rodriguez, 1982). However, the exact cause of its toxicity may be related to a proteolytic enzyme known as “dumbcane” or other labile protein-like substances (Walter, 1967; Walter & Khanna, 1972). Nevertheless, the role of proteins in Dieffenbachia's toxicity remains unconfirmed (Fochtman et al., 1969; Pohl, 1961; Rizzini & Occhioni, 1957). Additionally, some studies suggest that its toxicity could be linked to histamine release, leading to edema, vascular congestion, basement membrane degeneration, and inflammation (Barnes & Fox, 1955; Fochtman et al., 1969; Ladeira et al., 1975; Rizzini & Occhioni, 1957). Interestingly, Dieffenbachia seguine has been reported to possess anti-inflammatory properties (Barnes & Fox, 1955).

Macrophages play a pivotal role in immune responses by secreting various signaling molecules, including cytokines, growth factors, and chemokines (Ross et al., 2021; Zhang et al., 2021). Upon exposure to lipopolysaccharide (LPS), macrophages release pro-inflammatory cytokines such as IL-1β, IL-18, IL-6, and TNF-α, thereby activating other immune cells and amplifying the inflammatory response (Arango Duque & Descoteaux, 2014; Oishi & Manabe, 2018; Watanabe et al., 2019). In addition to these cytokines, other inflammatory mediators, such as nitric oxide (NO) and prostaglandin E2 (PGE2), are generated by inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), respectively (Król & Kepinska, 2021; Wautier & Wautier, 2023). NO plays a crucial role in regulating various physiological functions, including smooth muscle relaxation, neurotransmission, and host defense, while PGE2 is involved in vasodilation, fever, pain, and inflammation (Aoki & Narumiya, 2012; Ricciotti & FitzGerald, 2011; Sharma et al., 2007). Our experimental findings demonstrate that DSE exerts anti-inflammatory effects by downregulating the expression of iNOS and COX-2, leading to reduced levels of NO and PGE2. Furthermore, treatment with DSE resulted in a decrease in the production of pro-inflammatory cytokines. These observations suggest a potential therapeutic role for DSE in modulating inflammation.

Transcription factor NF-κB is paramount in controlling the innate and acquired immune systems (Dorrington & Fraser, 2019). NF-κB signaling pathway activation produces pro-inflammatory cytokine, cell survival, and leukocyte recruitment, the main contributors to an inflammatory response (Lawrence, 2009). The five monomers RelA (p65), NF-κB1 (p50), RelB, cRel, and NF-κB2 (p52) are NF-κB signal module members (Chen, 2005; Dorrington & Fraser, 2019). NF-κB’s primary pathway begins by stimulating TLR, IL-1R, or TNFR and activating the IKK complex. IKKβ phosphorylation subsequently causes IκB phosphorylation, ubiquitination, and decomposition. IκB disintegration releases p65/p50 (free NF-κB) for translocation into the nucleus, where free NF-κB binds to promoters, triggering inflammatory gene transcriptions (Biswas & Lewis, 2010). Thus, inflammatory reaction and disease severity extent depends on IκBα phosphorylation intensity in the NF-κB pathway.

Our data signified that DSE did not affect MAPK pathways but regulated the NF-κB pathway by reducing IκBα phosphorylation. As a result, DSE reduced inflammatory mediator and pro-inflammatory cytokine expressions. Notably, marked IL-1β and IL-18 decreases were observed through DSE treatment. Unlike other pro-inflammatory cytokines, IL-1β and IL-18 are produced from pro-IL-1β and pro-IL-18 enzymatic cleavage, respectively. In addition, the intracellular cysteine protease caspase-1 mediates the enzymatic process (Molla et al., 2020). Furthermore, NF-kB induces pro-inflammatory genes and regulates NLRP3 inflammasome formation consisting of NLRP3, ASC (apoptosis-associated speck-like protein containing CARD), and pro-caspase-1. An inflammasome complex’s precise assembly is critical for inflammatory responses against invading microorganisms. Our data substantiated that DSE interfered with inflammasome formation and caspase-1 activation while decreasing IL-1β and IL-18 expressions.

In summary, this study confirmed DSE’s anti-inflammatory ability in LPS-activated macrophages. DSE inhibited IκBα phosphorylation in the NF-κB pathway and inflammasome formation. As a result, inflammatory mediators NO and PGE₂ and pro-inflammatory cytokines IL-1β, IL-18, IL-6, and TNF-α secretion levels decreased. These results propose that DSE can be developed as an anti-inflammatory agent for inflammatory disease treatments.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Additional information

Funding

This work was supported by Ministry of Science and ICT, South Korea (H.M.): [grant no 2021R1F1A1061287]; Ministry of Food and Drug Safety of South Korea (B.K.K.): [grant no 22183MFDS366]; Chung-Ang University Graduate Research Scholarship in 2023 (E.L.): [grant no 2023].