Abstract

Purpose

   To investigate the therapeutic and gut microbiota-modulating effects of
   Qingreliangxuefang (QRLXF) on psoriasis.

Materials and Methods

   We used network pharmacology, a computational approach, to identify key
   bioactive compounds and biological targets, and explored the molecular
   mechanisms of QRLXF. The effects of QRLXF on keratinocyte proliferation
   and inflammation were evaluated using a mouse model of psoriasis.
   Changes in the gut microbiota were analyzed via 16SrDNA sequencing, and
   T cell subsets were assessed using flow cytometry.

Results

   Network pharmacology analysis suggested that QRLXF ameliorated
   psoriasis by modulating Th17 cell differentiation. Further experiments
   confirmed the anti-inflammatory effects and relief of psoriatic lesions
   in IMQ-induced mice. 16SrDNA sequencing revealed significant shifts in
   the gut microbiota, notably increases in Ligilactobacillus and
   Lactobacillus genera and decreases in Anaerotruncus, Negativibacillus,
   Bilophila, and Mucispirillum, suggesting a potential relationship
   between specific microbiota changes and Th17 cell differentiation.

Conclusion

   QRLXF alleviated psoriatic dermatitis by regulating Th17 cell responses
   and modifying gut microbiota profiles, highlighting its therapeutic
   potential for psoriasis treatment.

   Keywords: psoriasis, QRLXF, T cell differentiation, 16SrDNA sequencing,
   Th17 cell responses

Plain Language Summary

   Psoriasis affects millions globally. We studied a traditional herbal
   formula, Qingreliangxuefang (QRLXF), for its potential in treating
   psoriasis. Using computer tools, we found QRLXF may control Th17 cells,
   which play a role in psoriasis. Testing QRLXF in mice with
   psoriasis-like skin issues showed reduced inflammation and improved
   skin appearance. Gut bacteria analysis revealed more beneficial
   bacteria like Ligilactobacillus and Lactobacillus and fewer harmful
   ones. Our findings suggest QRLXF could be a promising new treatment
   option for psoriasis.

Graphical Abstract

   [44]graphic file with name DDDT-19-3269-g0001.jpg
   [45]Open in a new tab

Introduction

   Psoriasis is an immune-mediated skin disorder marked by erythema,
   scaling, and skin thickening, driven by an interaction of genetic
   factors and various external triggers.[46]1 The prevalence of psoriasis
   varies significantly across countries, but China has the largest number
   of affected adults globally due to its population size.[47]2,[48]3
   According to the 2021 Global Burden of Disease (GBD) study, the
   age-standardized incidence rate (ASIR) of psoriasis in China is
   59.70/100,000, while the age-standardized prevalence rate (ASPR) is
   474.02/100,000, imposing a substantial burden on healthcare systems and
   society.[49]4 Various psoriasis treatments exist, including topical
   corticosteroids, phototherapy, and systemic medications such as
   methotrexate and cyclosporine.^5 Biologic therapy targets precise
   inflammatory factors, demonstrating notable efficacy.[50]6,[51]7
   However, potential side effects persist during treatment, including
   liver toxicity, heightened infection risk, diminished efficacy, and
   possible immune drift.[52]8 Concurrently, the chronic nature of
   psoriasis imposes financial burdens on patients, eliciting additional
   concerns and anxieties.[53]9 Consequently, identifying safe,
   cost-effective, and efficacious treatments for psoriasis holds
   considerable importance.

   The pathogenesis of psoriasis is characterised by the persistent
   activation of various immune cells, resulting in
   inflammation.[54]10,[55]11 In the pathogenesis of psoriasis, a central
   role is played by Th17 cells, as the generation of IL-17 serves as a
   critical factor in sustaining chronic inflammatory
   responses.[56]12,[57]13 However, the precise mechanisms underlying
   psoriasis pathogenesis remain incompletely understood. Recent studies
   have demonstrated that psoriasis patients experience gut microbiota
   dysbiosis, with lower levels of probiotics and higher amounts of
   pro-inflammatory bacteria in their gut microbiota.[58]14–17 Moreover,
   findings indicate that those with psoriasis display an imbalance in the
   Firmicutes/Bacteroides (F/B) ratio, which impacts the production of
   intestinal short-chain fatty acids (SCFAs).[59]16,[60]18 Abnormal
   metabolites stimulate the proliferation of intestinal macrophages,
   thereby impacting gut immunity.[61]19

   The concept of the gut-microbiota-skin axis further elucidates the role
   of the microbiota in skin diseases from an immunological
   perspective.[62]20 The gastrointestinal tract serves as a crucial site
   for the regulation of Th17 cell proliferation and
   differentiation.[63]21 The gut microbiome can influence epithelial
   barrier function and immune cell differentiation, playing a pivotal
   role in various inter-organ communication processes.[64]22,[65]23
   Alterations in gut microbiota homeostasis and its metabolites drive the
   differentiation of Th17 cells in the colonic lamina propria,
   contributing to systemic inflammation in genetically predisposed
   individuals.[66]24–26 Studies have shown that segmented filamentous
   bacteria (SFB) can promote systemic Th17 cell proliferation upon
   intestinal colonization, exacerbating autoimmune diseases such as lupus
   nephritis[67]27 and autoimmune arthritis.[68]28 Conversely, antibiotics
   such as penicillin administered at appropriate doses can modulate
   bacterial populations, reduce Th17 cells in the small intestinal lamina
   propria, and reduce inflammatory bowel disease susceptibility in
   mice.[69]29 Similar effects have been observed in skin diseases.
   Studies have shown that germ-free and broad-spectrum antibiotic-treated
   mice exhibit reduced local and systemic Th17 activation when induced by
   imiquimod.[70]30,[71]31 Moreover,faecal transplantation experiments
   demonstrated that faeces from patients with severe psoriasis worsen
   skin inflammation and increase Th17 cell differentiation in mice.[72]32
   Thus suggests that the gut microbiota influences psoriasis development
   by modulating Th17 cell-mediated immune responses.

   Traditional Chinese Medicine (TCM) is frequently used in the clinical
   treatment of psoriasis because of its rapid efficacy, low cost, and
   capacity to prevent relapse. Blood-heat syndrome is the most common
   type of psoriasis observed during the active phase. Qingreliangxuefang
   (QRLXF), a Chinese medicine compound prescription, exerts therapeutic
   effects through heat clearing, blood cooling, detoxification, and
   dehumidification. However, the mechanisms underlying these effects
   require further investigations. Studies have confirmed that the
   constituents of QRLXF, including Scutellariae Radix,[73]33,[74]34
   Smilacis Glabrae Rhizoma,[75]35,[76]36 and Curcumae Radix,[77]37–39
   exhibit favourable therapeutic outcomes in psoriasis treatment and hold
   promise for modulating the gut microbiota. In clinical settings,
   compound formulations are extensively used in TCM, offering a holistic
   therapeutic impact, catering to a wide patient demographic range, and
   having significant practical utility. The intestinal microbiota is a
   crucial target for therapeutic intervention with TCM compounds.[78]40
   Nevertheless, the intricate nature of drug composition and structure
   presents challenges for advancing research in this area.

   The purpose of this study was to evaluate the efficacy of QRLXF in the
   treatment of psoriasis and explore its underlying mechanism. We used
   network pharmacology to make preliminary predictions and experimentally
   verify these mechanisms. In addition, we used 16S rDNA technology to
   investigate whether the therapeutic effects of QRLXF on psoriasis were
   related to the regulation of intestinal flora.

Materials and Methods

Preparation of Qingreliangxuefang Extract (QRLXF)

   Qingreliangxuefang includes 11 Chinese herbs: Bubali Cornu, Herba
   Solani Lyrati, Isatidis Folium, Curcumae Radix, Rehmanniae Radix,
   Moutan Cortex, Smilacis Glabrae Rhizoma, Scutellariae Radix, Sophorae
   Flos, Rhei Radix Et Rhizoma, and Arnebiae Radix. These herbs were
   supplied by Zhejiang Hospital of Traditional Chinese Medicine
   (Hangzhou, China). These herbs were soaked in purified water (1:10,
   w/v) for 30 min to allow the active ingredients to fully dissolve in
   the decoction. Bubali Cornu was removed and decocted separately for
   approximately 3 h. Subsequently, other herbs were added to the pots and
   boiled. After boiling, the mixture was simmered under low heat for 30
   min, and the decoction was collected. This process was repeated using
   the same volume of purified water as the first time. The two batches of
   decoction were then combined, concentrated under reduced pressure,
   freeze-dried into powder, and stored at –40 °C.

UHPLC-OE-MS and HPLC Analysis of QRLXF

   Sample 20mL was taken for Ultra High-Performance Liquid Chromatography
   - Orbitrap Exploris - Mass Spectrometry (UHPLC-OE-MS) to further
   clarify the composition of QRLXF. The analysis was conducted using a
   UHPLC system (Vanquish, Thermo Fisher Scientific) equipped with the
   Phenomenex Kinetex C18 (2.1 mm × 100 mm, 2.6 μm) and the Orbitrap
   Exploris 120 mass spectrometer (Orbitrap MS, Thermo). The mobile phase
   (A: 0.01% acetic acid in water; B: Isopropanol (IPA): Acetonitrile
   (ACN) (1:1, v/v)) was delivered at a speed of 0.3 mL/min: 0–1 min
   (1%B); 1–8 min (1%–99% B); 8–9 min (99%B) and 9–12 min (1% B). Under
   the control of the acquisition software (Xcalibur, Thermo), MS/MS
   spectra were acquired using the information-dependent acquisition (IDA)
   mode of an Orbitrap Exploris 120 mass spectrometer. The original data
   were processed and converted into the mzXML format using ProteoWizard.
   We then used the BiotreeDB (V3.0) and the R software to identify the
   metabolites.

   The specific contents of the active components were further quantified
   using HPLC, and standard references for “hypoxanthine, luteolin,