Effect And Mechanism Of D-mannose On Psoriatic Dermatitis In Mice

Psoriasis is a complex and difficult-to-cure chronic inflammatory skin disease mediated by autoimmunity,causing significant physical and mental burdens on patients.The pathogenesis of psoriasis is not clear,involving various factors such as immunity,metabolism,genetics,and environment.The onset of psoriasis is related to various cells,including keratinocytes,T cells,macrophages,and dendritic cells.Among them,keratinocytes are the characteristic cells of pathological changes in psoriasis.They play an essential role in the onset of psoriasis by producing various pro-inflammatory cytokines,chemokines,and other immune molecules,including CCL20,and forming a vicious cycle that mutually reinforces with various immune cells.In addition,macrophages are activated and recruited to the lesion site in the skin,promoting the occurrence and progression of psoriatic lesions by activating inflammasomes and producing pro-inflammatory cytokines.Studies have shown that SPHK1regulates the activation of NLRP3 inflammasomes in macrophages.D-mannose is a monosaccharide that exists in nature and is widely distributed in the body.Its structure is similar to glucose,but its concentration in human blood is less than 1/50 of that of glucose.Studies have shown that D-mannose has good therapeutic effects on various metabolic and autoimmune diseases.However,its impact on psoriasis is still unclear.This study explored the therapeutic effect of D-mannose on psoriasis by investigating its regulation on chemokine CCL20 in keratinocytes,as well as its influence on the activation of the AIM2 inflammasome and pyroptosis in macrophages.The study preliminarily elucidated the potential mechanism of D-mannose in treating psoriasis and provided new insights for the mechanistic research and treatment strategies for psoriasis.Study on the mechanism of D-mannose regulating keratinocyte CCL20 to alleviate psoriasis-like dermatitis in miceObjective:（1）Clarify the therapeutic effect of D-mannose on psoriasis-like dermatitis in mice（2）From the perspective of keratinocytes,investigate the specific mechanism of D-mannose in regulating psoriasis skin inflammation,providing new insights for the exploration of the pathogenesis and treatment methods of psoriasis.Methods:（1）Establish an imiquimod（IMQ）-induced mouse psoriasis model using Balb/C mice and administer D-mannose orally or D-mannose hydrogel topically.Compare the skin lesion inflammation,spleen size,and weight changes between the treatment group and the model group.（2）Use flow cytometry to detect the changes in immune cell infiltration in mouse skin tissue due to D-mannose treatment.（3）Isolate primary na（?）ve CD4⁺T cells for in vitro differentiation induction,and use flow cytometry to detect the effect of D-mannose on Th17 cell differentiation.（4）Perform RNA-seq to detect changes in the mouse skin lesion transcriptome.（5）Use immunohistochemistry to assess the relative expression levels and localization of target proteins in tissues.（6）Utilize Western blotting to detect changes in protein levels,and real-time quantitative PCR to detect changes in m RNA levels.（7）Stimulate Ha Ca T cell line with IL-17a and TNF-αto mimic psoriasis keratinocyte inflammation in vitro and intervene with D-mannose.Evaluate the expression level changes of target proteins and target genes.（8）Employ CRISPR-Cas9 technology to create gene knockout cell lines.Results:（1）Both oral and topical administration of D-mannose effectively alleviated IMQ-induced psoriasis-like dermatitis in mice.Specifically,D-mannose treatment resulted in reduced PASI scores,thinner epidermal layers,and a decrease in infiltrating Th17 cells in the skin.（2）Transcriptome sequencing of mouse skin tissue suggests that the"Keratinocyte Pathway"score（KC Score）decreases in DM+IMQ group mice,and the expression levels of multiple cytokines and chemokines,including CCL20,are reduced.（3）Single-cell transcriptomic data analysis,bulk transcriptomic data from psoriatic patient skin tissues,and immunohistochemical staining indicate that CCL20 is highly expressed in keratinocytes of psoriasis patients and is positively correlated with PASI.（4）Immunohistochemical staining of mouse skin tissue and in vitro experiments using the Ha Ca T cell line demonstrate that D-mannose downregulates the expression of CCL20 in keratinocytes.（5）Analysis of the CCL20 promoter sequence suggests that HIF-1αmay regulate CCL20 transcription.（6）In vitro experiments with IL-17a and TNF-αstimulated Ha Ca T cell line show that D-mannose downregulates HIF-1αprotein levels in Ha Ca T cells.（7）Under stimulation with IL-17a and TNF-α,HIF-1αknockout Ha Ca T cells show a decrease in m RNA expression levels of CCL20..Conclusions:（1）Oral and topical administration of D-mannose can alleviate IMQ-induced psoriasis-like dermatitis in mice,suggesting that D-mannose has potential as both an oral and topical treatment for psoriasis.（2）In psoriasis patients,the expression of CCL20 in keratinocytes is elevated and positively correlates with PASI scores,indicating that it may be a potential therapeutic target for psoriasis.（3）D-mannose inhibits the HIF-1α/CCL20 axis in keratinocytes,blocking the interaction between keratinocytes and Th17 cells,thereby disrupting the vicious cycle and alleviating psoriasis.Study on the mechanism of D-mannose regulating macrophage pyroptosis through SPHK1Objective:（1）Investigate the mechanism of D-mannose in alleviating psoriasis from the perspective of macrophages.（2）Explore the relationship between D-mannose and the inflammatory regulatory molecule SPHK1.（3）Investigate the role and mechanism of SPHK1 in psoriasis.Objective:（1）Utilize transcriptomic data from mouse skin tissue to analyze the expression of SPHK1,NLRP3,AIM2,and pyroptosis-related genes.（2）Employ immunofluorescence staining to observe the impact of D-mannose treatment on macrophage infiltration in mouse skin.（3）Extract bone marrow-derived macrophages（BMDMs）from mice and validate the effect of D-mannose on SPHK1 in vitro.（4）Analyze public bulk transcriptomic data from psoriasis patient skin tissue and use immunohistochemistry staining to investigate SPHK1expression in psoriasis.（5）Analyze single-cell sequencing data from psoriasis patient skin tissue to screen for cell populations with differential SPHK1 expression.（6）Induce psoriasis-like dermatitis in SPHK1^-/-mice using IMQ to evaluate the role of SPHK1 in psoriasis.（7）Use immunofluorescence staining to observe changes in macrophage infiltration in SPHK1^-/-mouse skin.（8）Isolate BMDMs from mice,induce macrophage polarization,and detect using flow cytometry.（9）Employ Western Blot to detect changes in the expression of pyroptosis-related markers in SPHK1^-/-mouse skin.（10）Analyze public bulk transcriptomic data from psoriasis patient skin tissue to investigate the relationship between AIM2 inflammasomes and psoriasis.（11）Induce pyroptosis in mouse BMDM cells using LPS+poly（d A:d T）to explore the effects of D-mannose and SPHK1 on pyroptosis.Results:（1）Transcriptomics data from psoriasis mice showed that oral administration of D-mannose reduced SPHK1 expression in the skin of psoriasis mice,with no significant difference in SPHK2 expression.D-mannose also decreased the expression of AIM2,caspase-1,GSDMD,and IL-1β,genes associated with inflammasomes and pyroptosis.（2）D-mannose reduces macrophage infiltration in mouse skin.（3）SPHK1 is highly expressed in nucleated cells in the dermis of psoriasis patients.（4）In vitro experiments with mouse BMDMs show that D-mannose downregulates the LPS-induced high expression of SPHK1 in macrophages.（5）Single-cell sequencing data from patients show that SPHK1 is highly expressed in macrophages of psoriasis patients.（6）IMQ-induced psoriasiform dermatitis in SPHK1^-/-mice showed that knockout of SPHK1 alleviated skin inflammation in psoriasis and reduced macrophage infiltration.（7）Western Blot analysis of mouse skin tissue proteins revealed a decrease in pyroptosis-related markers in SPHK1^-/-mice.（8）Public transcriptomics data from patients show that AIM2 is highly expressed in patient skin lesions and positively correlates with PASI score.（9）Both D-mannose and PF-543（SPHK1 inhibitor）can inhibit AIM2inflammasome activation-induced macrophage pyroptosis.Conclusions:（1）D-mannose regulates the expression of SPHK1 in psoriasis.（2）SPHK1 is highly expressed in macrophages of psoriasis patients,making it a potential therapeutic target for the disease.（3）The inflammasome AIM2 is closely related to psoriasis.For macrophages,D-mannose inhibits activation of AIM2 inflammasome and pyroptosis by regulating the expression of SPHK1,thereby alleviating psoriasis.