Introduction
Atopic dermatitis (AD) represents one of the most prevalent chronic inflammatory skin diseases, with an astonishing incidence rate affecting approximately 3% of adults and escalating to 20% among children, indicating its widespread impact across diverse age groups and ethnic backgrounds.1 The global prevalence of AD has shown a concerning upward trend, emphasizing the urgent need for effective management strategies.2 Patients suffering from AD experience debilitating symptoms, such as intense itching, with around 91% of individuals reporting chronic pruritus lasting longer than six weeks.3 This distressing condition frequently disrupts sleep patterns, consequently leading to a marked decline in overall quality of life. Alarmingly, AD is ranked highest in global disease burden attributable to skin disorders.4 Nonetheless, the intricate mechanisms underlying the pathogenesis of AD remain incompletely understood, highlighting the interplay of genetic predisposition, epidermal dysfunction, and multiple immune pathways dominated by T-cell inflammation.2 Traditional treatment options for AD predominantly focus on topical agents, notably corticosteroids and calcineurin inhibitors, which have been in use for decades.5 However, sustained use of these treatments may lead to adverse effects, including localized skin atrophy linked to corticosteroid use and potential long-term cancer risks associated with topical calcineurin inhibitors.6 Although newer agents like Dupilumab, which targets the IL-4 receptor, as well as Janus kinase (JAK) inhibitors—such as upadacitinib and baricitinib—show promising therapeutic efficacy, the high costs and requirement for sustained evaluations remain significant barriers.7 Therefore, the exploration of innovative pharmacological agents with potent symptom relief, long-term applicability, and minimal side effects is of paramount importance.
Arctii (Fructus arctii), derived from the mature fruit of Arctium lappa L of the Compositae family, has gained recognition in traditional medicine as a commonly used herbal remedy, demonstrating therapeutic efficacy for a variety of ailments including pyrexia, dizziness, diabetes, toothache, furuncle, and alopecia.11 The compound Arctiin (ARC), characterized as a lignan, is recognized as the principal active and marker component of Arctii, as noted within the Chinese pharmacopoeia, with its availability in over 38 plant species.12 ARC exhibits a spectrum of biological activities, notably anti-inflammatory, anti-allergic, anti-cancer, antibacterial, antioxidant, and anti-platelet properties, also acting as a calcium antagonist.13 Remarkably, ARC has been shown to alleviate IgE-mediated passive skin allergic reactions and anaphylactic shock triggered by compound 48/80.14 Research indicates potent anti-inflammatory effects stemming from its inhibition of the NF-κB transport pathway while concurrently reducing levels of inflammatory mediators such as IL-6, IL-1β, and TNF-α.15
However, the specific effects of ARC on AD remain ambiguous. Thus, our study sought to delineate the impact of ARC on AD in Balb/c mice induced by DNCB, alongside evaluations in HaCat cells stimulated with TNF-α and IFN-γ.
Materials and Methods
Arctiin
Arctiin was procured from Beijing Solarbio Science & Technology Co., Ltd., (Beijing, China), ensuring high purity with High Performance Liquid Chromatography (HPLC) results of ≥98%, batch number: SA8440.
Animals
Female Balb/c mice aged 6–8 weeks, weighing approximately 20±2g, were acquired from Huafukang Biotechnology Co. Ltd. (Beijing, China, RRID: IMSR_APB:4790). All mice were kept in pathogen-free stainless steel cages in a strictly controlled environment (12h light/12h dark cycle; temperature: 22 ± 1 °C; humidity: 55 ± 5%). They were provided food and water ad libitum. All experimental procedures followed the NIH Guidelines for the Care and Use of Laboratory Animals and received approval from the Ethics Committee of China Medical University (2022PS1149K).
Induction of AD and Treatment
Prior to experimentation, all mice underwent a one-week acclimatization period within the laboratory environment. The 24 mice were allocated into four groups (n = 6 each): control group, model group, arctiin group (25 mg/kg, ARC group), and positive control group (1 mg/kg, dexamethasone, DEX group), utilizing random number generation for distribution. The induced AD-like skin lesions in mice were provoked by applying a 1% DNCB solution (dissolved in a mixture of acetone and olive oil) on the shaved dorsal surface, targeting a skin area of approximately 2 × 2 cm2. Mice were sensitized on days one and two before receiving a 0.5% DNCB solution bi-daily for a total of nine applications. At the conclusion of day 21 of the experiment, the mice were euthanized, and blood samples, along with dorsal skin tissue, were harvested for further analysis.
Figure 1 Schematic diagram of the animal experiment.
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Evaluation of the Severity of Skin Dermatitis and Dorsal Skin Thickness
Skin dermatitis severity scores were systematically recorded weekly throughout the experiment, with evaluations based on four distinct symptoms: erythema, edema, dryness, and ulceration, using a defined scoring system to classify the severity of each symptom.
Measurement of Serum IgE
Blood samples were taken at the conclusion of the experimental phase, and serum was extracted through microcentrifugation. Total serum IgE levels were evaluated through an ELISA assay, abiding by the manufacturer’s protocols.
Histological Analysis
Post-euthanasia, the dorsal skin tissue underwent fixation in 4% paraformaldehyde, followed by embedding in paraffin. Tissue blocks were sectioned at 3.5 μm, stained using hematoxylin-eosin (H&E) to observe histological alterations, and assessed for epidermal thickness and mast cell infiltration using toluidine blue staining.
Immunohistochemistry
The sections underwent standardized deparaffinization followed by antigen retrieval protocols and were incubated overnight with specified primary antibodies to assess protein expression related to TLR4 and NF-κB signaling.
Cell Culture
HaCaT cells were cultured in Minimum Essential Medium supplemented with fetal bovine serum and penicillin-streptomycin in a controlled 37 °C environment maintained with 5% CO2.
Cytotoxicity Assay
The Cell Counting Kit-8 (CCK-8) was employed to evaluate cell viability post-incubation with varying concentrations of ARC, following manufacturers’ guidelines.
Molecular Docking
Molecular docking analysis was performed using AutoDock Vina 4.0 to examine interactions between arctiin and TLR4, utilizing molecular models obtained from database repositories.
Statistical Analysis
Statistical analysis was performed employing GraphPad Prism 10.0 with results expressed as mean ± standard deviation (SD). Normality assessments incorporated the Shapiro-Wilk test for determining appropriate analytical methodologies.
Results
Effect of ARC on Atopic Dermatitis in Balb/c Mice Induced with DNCB
Effect of ARC on Atopic Skin Lesions in Balb/c Mice Induced with DNCB
A comprehensive investigation into ARC’s effects on AD involved inducing dorsal skin inflammation in Balb/c mice with DNCB, utilizing dexamethasone as a positive control treatment. Notably, the intervention with ARC and dexamethasone significantly alleviated clinical symptoms associated with AD.Figure 2A
Furthermore, the dermatitis score index was significantly elevated in the DNCB group compared to the control mice, highlighting the ameliorative effects of ARC and dexamethasone.pTable 2
Table 2 Dermatitis Score at Different Time (Mean ± SD, n = 6)
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Histological assessments through H&E staining revealed marked increases in epidermal thickness in DNCB-treated mice, however, ARC and dexamethasone treatment significantly mitigated this epidermal change.Figure 2D
Effect of ARC on Serum IgE and Cytokines
With hypersecretion of IgE recognized as a core issue in AD,19 the examination of ARC’s impact on serum IgE levels revealed substantial reductions following treatment. Real-time PCR data indicated that inflammatory cytokines (IFN-γ, TSLP) produced within the DNCB group were significantly abated by ARC and DEX treatments.
Effect of ARC on TLR4/MyD88/NF-κB Signalling Pathway in Balb/c Mice Induced with DNCB
Evidence underscored the pivotal roles that TLR4 engages in immune responses and inflammation regulation. Results indicated ARC notably inhibited the expression levels of TLR4 and its downstream signaling pathway components, thereby attenuating immune response elevation induced by DNCB.Figure 3A
Effect of ARC on TLR4/MyD88/NF-κB Signalling Pathway in TNF-α/IFN-γ-Stimulated HaCaT Cells
Our studies involving TNF-α/IFN-γ stimulation in HaCaT cells demonstrated ARC’s cytotoxicity profile was favorable at concentrations up to 400 μg/mL.Figure 4A
In summary, ARC exhibited significant suppression of TLR4/MyD88/NF-κB pathway activation in HaCaT cells stimulated by TNF-α/IFN-γ, indicating its potential as a therapeutic agent.Figure 4D
ARC Demonstrated Anti-Pyroptotic Effects
In our investigations of protein expression associated with pyroptosis in dorsal skin, increases in expression levels were observed in the DNCB group, whereas treatment with ARC and DEX effectively reduced these elevated levels,pFigure 5A.
Figure 5 Analysis of the pyroptotic pathway in skin tissues: (A) Protein expression levels of NLRP3 and IL-1β in DNCB-induced mice. (B) Transcriptional levels of IL-18 following treatment. |
Conclusion
This study provides compelling evidence that ARC serves as an effective mitigator of AD symptoms in DNCB-induced mice, significantly reducing skin thickness, mast cell infiltration, and serum IgE levels. Both in vivo and in vitro analyses confirm that ARC impedes activation of critical signaling pathways. Collectively, these findings position ARC as a promising alternative therapy for AD, establishing a new molecular target and enhancing the quality of life for affected individuals.
Introduction
Ah, atopic dermatitis (AD), the skin condition that’s making more people itch than a bad Tinder date! A staggering 3% of adults and a whopping 20% of children are reportedly afflicted, which really makes you wonder how many of them are just attacking with impromptu scratching sessions instead of seeking therapy. But fear not, folks, we delve into the itchiness of AD, a condition that ranks at the top of the global burden ladder for skin diseases. It’s like the valedictorian of skin troubles; nobody wants that title, yet here we are.
The pathogenesis of AD? Oh, it’s complex and layered like an onion—and no, we’re not crying just yet. Genetics, epidermal dysfunction, and a gaggle of immune pathways, particularly T-cell inflammation, are all in the mix. It’s like the latest hot chocolate recipe: serve it up with some irritation and a sprinkle of expensive medications like Dupilumab or JAK inhibitors, which are great—unless you check your bank balance afterward. Hence, the quest for a new miracle drug continues…
Enter Arctii, also known as Fructus arctii—that’s a mouthful, isn’t it? It’s not just a plant you’d find on a quirky botanist’s coffee table. This bold little guy has a storied history in traditional medicine for treating various ailments, ranging from headaches to hair loss—so, basically the Swiss Army knife of plants. Our star compound, arctiin (ARC), has decided to join the party, packing some seriously powerful punches against inflammation, allergies, and even cancer. I mean, stop it already; who does this fruit think it is?
This study aims to find out whether ARC can indeed calm the raging, itchy storm that is atopic dermatitis in model mice. Because if it works on mice, let’s face it, there’s hope for all of us humans swiping in the realm of skin disorders. Buckle up!
Materials and Methods
Arctiin
They got Arcitn. 98% purity! Apparently, our buddies at Beijing Solarbio Science and Technology are running a tight ship—all chemical dreams wrapped up in a shiny package. Nice.
Animals
Welcome to the mouse realm, where the Balb/c mice are prepped with the utmost care! These furry subjects are treated better than your average college student during finals week—complete with a specially controlled environment, gourmet food, and no 2 AM pizza deliveries. What more could a mouse ask for?
Induction of AD and Treatment
A little bit of DNCB (that’s short for dinitrochlorobenzene, and no, it’s not a trendy new drink) was the weapon of choice to induce AD symptoms in our mice guests. They were split into four groups—sounds like a reality show waiting to happen! The control group, the model group for misery, and two treatment groups featuring our hero, Arctiin, and the reigning champ, dexamethasone. Let’s see which one makes these mice feel better—game on!
Evaluation of the Severity of Skin Dermatitis and Dorsal Skin Thickness
If you thought mouse beauty standards were relaxed, think again. Each week, their dermatitis severity scores were carefully monitored, ensuring only the most pristine specimens enough to be bronzed for the ’gram. Spoiler: it wasn’t pretty in the DNCB group.
Measurement of Serum IgE
Blood was drawn (don’t worry folks, it’s all in the name of science!) to check the total serum level of IgE, a key indicator, as high levels are synonymous with our friend AD. You might say they’re like that annoying person who shows up uninvited—unpleasant!
Histological Analysis
Skin samples were turned into glamour shots, something straight out of a magazine—but for mice. H&E staining was performed, leaving no puffiness unnoticed, and watch out for those mast cells; they might just throw quite the party!
Immunohistochemistry
On to the grand finale of antibody staining! Cell sections received the royal treatment, resulting in a showcase of positivity. You know, like a TikTok influencer’s reveal.
Cell Culture
Meanwhile, HaCaT cells were living their best life in the lab; with all sorts of conditions designed to keep them pumping out the data we desperately need. These cultured keratinocytes were treated like royalty, ensuring no food poisoning here!
Results
Effect of ARC on Atopic Dermatitis in Balb/c Mice Induced with DNCB
So, what’s the verdict? Did our herbal wonder save the day? ARC was applied gallantly to those poor mice with an itchy, miserable life. The result? A noticeable decrease in itchiness, skin thickness, and all things nasty related to AD! Dexamethasone, the traditional treatment, moved aside a bit as well—much to the surprise of no one.
Effect of ARC on Serum IgE and Cytokines
Here comes the big news: serum IgE levels back down from “sky-high” levels to something more manageable. Yeah, smart move, ARC! And the cytokine levels? Let’s say they got the memo too—both IFN-γ and TSLP were reduced significantly. Hallelujah!
Effect of ARC on the TLR4/MyD88/NF-κB Signalling Pathway
But don’t just take our word for it; we took a deep dive into the cellular signaling pathways. ARC strolled right in and dampened that TLR4/MyD88/NF-κB signaling pathway from sending out inflammatory invites. Those pesky inflammatory cytokines were left waiting by the phone, wondering why nobody’s returning their calls!
Discussion
Now that we’ve covered all the nitty-gritty details, let’s put our cards on the table. Atopic dermatitis remains complex and evasive—much like your elusive Tinder match who never responds. But let’s keep it real; ARC isn’t just a pretty face in the herbal world; it’s packing some serious anti-inflammatory heat. So open the door to possible new treatments for those with itchy hopes and dreams of relief from their persistent woes!
Conclusion
This study marks the beginning of hope for atopic dermatitis sufferers. With ARC leading the charge, we’re onto a potential new therapeutic direction, offering new pathways to managing this wide-reaching skin condition. No more scrounging through old cabinets for last-resort remedies; it’s time the world recognized the prowess of this unassuming herb!
Acknowledgments
All this research was generously funded by a scientific project focused on restoring skin barrier functions. So, big thanks to whoever keeps the grant money flowing—you’re the real MVP!
Disclosure
None of our authors have conflicts of interest—no hidden agendas here. Just a bunch of mice, a fruitful herb, and a serious desire to tackle atopic dermatitis.
References
If you survived this far, hats off to you! And if you want to get into the nitty-gritty stats and details that tested mice have everyone talking, dive right into those citations!
JAK-STAT pathway in atopic dermatitis
For it! The deeper look into the signaling mechanisms revealed that ARC not only eased the symptoms of atopic dermatitis but also inhibited the TLR4/MyD88/NF-κB pathway, which is known to be crucial in the inflammatory response. This further emphasizes ARC’s role as a modulator of systemic inflammation, providing a multi-faceted approach to treatment.
### Implications and Future Directions
The promising results observed in both in vivo (mouse model) and in vitro (HaCaT keratinocytes) studies indicate that ARC could be positioned as a potential therapeutic agent in the management of atopic dermatitis. Given its ability to affect key signaling pathways and cytokine production, future investigations could explore clinically relevant applications and the underlying mechanisms in greater detail. A broader scope of experiments, including various dosages, combinations with other therapies, and long-term studies, may further elucidate ARC’s efficacy and safety for human patients.
Furthermore, exploring other related skin conditions or inflammatory diseases could expand the therapeutic potential of ARC beyond just atopic dermatitis. Ultimately, the goal remains to offer better, affordable, and accessible treatments that can improve the quality of life for individuals suffering from chronic skin conditions.
### Closing Thoughts
the investigation into the role of arctiin as a potential treatment for atopic dermatitis showcases not just a glimmer of hope but a robust scientific inquiry into herbal medicine’s capabilities in tackling modern health issues. The effectiveness of ARC represents an encouraging stride toward integrating natural remedies into mainstream treatment regimens for skin health, paving the way for innovative approaches to managing chronic inflammatory conditions. So, if you’ve been itching for a solution that doesn’t break the bank, ARC may just be the herbal miracle you’ve been waiting for!