TCN2 promotes psoriatic skin inflammation and keratinocyte proliferation via the IL-1β-STAT3 pathway

TCN2 promotes psoriatic skin inflammation and keratinocyte proliferation via the IL-1β-STAT3 pathway | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article TCN2 promotes psoriatic skin inflammation and keratinocyte proliferation via the IL-1β-STAT3 pathway Jing-Kai Xu, Xin-Zhu Zhou, Ke Xue, Ang Li, Qing-Yue Xia, Zhou Zhuang, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7309098/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Psoriasis, a chronic immune-mediated inflammatory skin disorder, exerts systemic impacts across multiple organs. Although transcobalamin 2 (TCN2) has been reported to be associated with several autoimmune diseases, its role in psoriasis remains elusive. Hence, we investigated the role of TCN2 in psoriasis pathogenesis. Our results indicated that TCN2 was highly expressed in the skin tissues and peripheral blood mononuclear cells (PBMCs) of psoriatic patients, with downregulation following biologic therapy. Moreover, imiquimod (IMQ) - induced psoriasis in mice exhibited heightened TCN2 expression. To further explore the role of TCN2 in psoriasis, we generated Tcn2-deficient (Tcn2-/-) mice and established a psoriasis model using IMQ. IMQ-treated Tcn2-/- mice displayed milder psoriatic lesions and a lower level of inflammation. RNA-seq analysis of lesional skin revealed significant downregulation of inflammatory mediators (S100A7, S100A8, S100A9, IL-1β, IL-6) and suppression of STAT3 signaling in Tcn2-/- mice compared to WT-IMQ mice. Parallel in vitro experiments using TCN2-knockout HaCaT cells demonstrated cell cycle arrest. Collectively, our findings highlight TCN2 as a critical regulator of psoriatic inflammation, proposing it as a novel therapeutic target. TCN2 psoriasis skin inflammation Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 INTRODUCTION Psoriasis is a relapsing, immune-mediated, and chronic inflammatory skin disease that affects more than 125 million individuals worldwide, or about 2–3% of the global population [ 1 ]. Mechanistically, psoriatic lesions exhibit substantial infiltration of monocytes, neutrophils, and T cells. The inflammatory stimulation, abnormal differentiation and proliferation of keratinocytes, as well as dysregulated activation of stem cells work together to cause keratinocyte overgrowth and subsequent formation of thick, scaly plaques lesions [ 2 ]. Current understanding of psoriasis immunopathogenesis highlights the pivotal role of pro-inflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin-1 (IL-1), and interleukin-6 (IL-6), et.al. They can synergistically activate keratinocytes to produce chemokines and antimicrobial peptides [ 3 ]. In psoriasis lesions, chemokines such as C-C motif chemokine ligand 20 (CCL20), C-X-C motif chemokine ligand 1 (CXCL1), CXCL2, and CXCL8 play a key role in the recruitment and activation of immune cells, particularly interleukin 17A (IL-17A)-generated T cells and neutrophils [ 3 , 4 ]. At the same time, keratinocytes release antimicrobial peptides, including S100 calcium-binding protein A7 (S100A7), S100A8, S100A9, and Defensin β 4A/defensin β 2(DEFB4A/DEFB2). These peptides not only exert direct antimicrobial effects but also amplify local inflammatory response. This process contributes to the formation of a complex inflammatory network. [ 5 , 6 ] IL-1β, a classic proinflammatory cytokine, activates dermal γδT cells to produce IL-17, and stimulates chemokine release [ 7 ]. And, in psoriatic lesions, IL-1β is predominantly secreted by keratinocytes [ 8 ]. Transcobalamin II (TCN2) is a vitamin B12-binding protein that aids in the cellular uptake of vitamin B12, thus being a biomarker for active vitamin B12 [ 9 ]. Numerous studies have reported a close association between TCN2 and the development of autoimmune-related diseases, including multiple sclerosis, systemic lupus erythematosus and idiopathic recurrent spontaneous abortion [ 10 – 13 ]. It can participate in the occurrence of diseases by regulating a variety of biological processes, such as cell proliferation, apoptosis, and migration [ 9 ]. However, the role of TCN2 in psoriasis remains unexplored. In this study, we propose that TCN2 promotes aberrant proliferation of keratinocytes and exacerbates the inflammatory response in psoriasis by affecting IL-1β secretion. Mechanistically, TCN2 can inhibit the secretion of IL-17 and the activation of STAT3 pathway by affecting the expression of IL-1β and down-regulating the expression of related pro-inflammatory factors. Our study suggests the potential of TCN2 as a new therapeutic target for psoriasis. RESULT TCN2 expression is significantly elevated in psoriasis patients and IMQ - induced models To explore the expression of TCN2, we first conducted immunohistochemical staining on skin sections from healthy controls (HCs) and psoriasis patients. In HC skin TCN2 is primarily distributed in the epidermis basal layer. In psoriatic skin, TCN2 is widely expressed in the thickened epidermis, with increased expression in dermal vascular endothelial cells and inflammatory cells ( Fig. 1 a ) . The detailed information of the patients were shown in Supplementary Table 1 . Meanwhile, we measured the expression of TCN2 in PBMCs of 25 psoriasis patients and 25 HCs. It was found that the expression of TCN2 was significantly upregulated in psoriasis patients ( Fig. 1 b ) . These patients underwent clinical treatment with Secukinumab. Six months after treatment initiation, the peripheral blood of 14 patients was collected, and the expression of TCN2 was measured. Results demonstrated that, after treatment, not only was the psoriasis area and severity index (PASI) of these patients significantly decreased, but the expression of TCN2 was also significantly reduced ( Fig. 1 c ) . To further explore the expression of TCN2 in psoriasis, we employed IMQ-induced wide type (WT) mice to establish a psoriasis mouse model. qPCR, Western blotting and immunofluorescence staining were conducted to compare the expression of TCN2 in the skin tissues of WT and WT + IMQ mice. The results revealed that TCN2 levels were significantly upregulated in the WT + IMQ mice ( Fig. 1 d and 1 e ) . Taken together, these data suggest that TCN2 is associated with the progression of psoriasis and may contribute to its pathogenesis. Tcn2 deficiency attenuates IMQ-induced psoriasis phenotype Gene-edited mice have been extensively employed to explore the pathogenesis of numerous diseases. To clarify the role of TCN2 in psoriasis, we first generated a Tcn2-/- mouse model. Subsequently, we set up three distinct groups of psoriasis mouse models: WT + IMQ, Tcn2+/- + IMQ (T+/- + IMQ) and Tcn2-/- +IMQ (T-/- + IMQ) mice. Seven days–after IMQ induction, compared to the T-/- + IMQ mice, WT + IMQ mice exhibited more prominent scaling and roughness in the dorsal skin. Moreover, the severity of epidermal hyperplasia and spleen size decreased successively in the WT + IMQ, T+/- + IMQ and T-/- + IMQ mice ( Fig. 2 a and Supplementary Fig S1 a) . After that, HE staining was performed on the three groups, and results demonstrated that epidermal hyperplasia and inflammatory infiltration were more pronounced in the WT + IMQ mice compared to the T-/- + IMQ mice ( Fig. 2 b ) . Additionally, the epidermal thickness and spleen/body weight ratio were significantly decreased in the T-/- + IMQ mice ( Fig. 2 c and d) . Meanwhile, immunofluorescence and Western blotting revealed diminished IL17 and Cd45 expression in the T-/- + IMQ mice ( Fig. 2 e ) . We found that TCN2 deficiency could significantly reduce psoriasis-related inflammation. The findings demonstrate that TCN2 knockdown attenuates epidermal hyperplasia and skin inflammation, suggesting its potential as a therapeutic target for psoriasis. RNA-seq implicates TCN2 deficiency in inflammatory signaling To further elucidate the mechanism of TCN2 deficiency in psoriasis treatment, we employed bulk RNA-seq to analyze the expression profiles of WT, WT + IMQ and T-/- + IMQ mice. By comparing the genes enriched in WT mice and WT + IMQ mice, WT + IMQ mice and T-/- + IMQ mice, we identified 193 differentially expressed genes (DEGs) among the three groups ( Fig. 3 a, Supplementary Fig S1 b and Supplementary table2) . Figure 3 b showed the volcanic maps of DEGs changes in different groups. Compared to the WT + IMQ mice, 204 genes in the T-/- + IMQ mice were up-regulated and 410 genes down-regulated. (Fold change ≥ 2 and p < 0.05). And a lot of inflammation-related molecules (like Il1b, Il6, S100a7 and S100a8, et.al) were significantly reduced after TCN2 knockout (Fig. 3 c). Moreover, KEGG analysis showed that down-regulated DEGs were mainly involved in biological processes related to signal transduction and innate immune response, such as JAK-STAT, IL-17 pathway, Th17 cell differentiation, etc. ( Fig. 3 d ) . GSVA analysis was performed, revealing that TCN2 deficiency potently inhibited pivotal signaling pathways in psoriasis, and MAPK, NF-κB, and STAT3 pathways were significantly inhibited in the T-/- + IMQ mice ( Fig. 3 e ) . This observation implies that TCN2 deficiency might curtail the inflammatory response in psoriasis by impeding the activation of these pathways. TCN2 regulates the proliferation of epidermal cells in psoriasis Excessive proliferation of epidermal keratinocytes is a characteristic manifestation of psoriasis [ 14 ]. To explore the impact of TCN2 knockdown on the proliferation of epidermal cells in psoriasis, we constructed a TCN2-knockdown HaCat cell line and verified that TCN2 was knocked down by Western blotting ( Fig. 4 a and b) . The CCK-8 assay demonstrated that, in comparison to the control cells, the proliferation rate of the TCN2-knockdown cell lines was markedly diminished ( Fig. 4 c ) . Furthermore, through flow cytometry, we demonstrated that the knockdown of TCN2 modulated the cell cycle by inducing a G1 phase arrest in the cell lines ( Fig. 4 d and e) . Additionally, immunofluorescence techniques were employed to stain the skin tissues obtained from the three groups of mice, and it was ascertained that the silencing of TCN2 led to a decrease in Ki67 expression within the epidermis ( Fig. 4 f ) . Collectively, these results imply that the influence of TCN2 on cell proliferation and cycle might be associated with the excessive proliferation of epidermal cells. TCN2 regulates the progress of psoriasis through modulating IL-1β-STAT3 pathway To further validate our RNA-seq results and elucidate the mechanism of TCN2 in psoriasis, we investigated the expression levels of inflammatory factors in the skin tissues from the T-/- + IMQ and WT + IMQ mice, and found that the expression of Il-1β, S100a8, and S100a9 was significantly reduced ( Fig. 5 a ) . In addition, we studied the expression of Il17-a in the skin of the four-mouse-group and observed that TCN2 knockdown led to a substantial reduction in Il-17a expression ( Fig. 5 b ) . M5 stimulants (IL-1α, IL-17A, IL-22, TNF-α, and Oncostatin M) have been used to induce psoriasis-like inflammatory cells [ 15 ]. In our study, we treated the TCN2-knockdown (KO) HaCaT cells with M5 at a concentration of 10ng/ml for 24 hours. Notably, we found the mRNA expression of inflammatory factors, including IL-1β, IL-6, TNFα, S100A7, S100A8 and S100A9 , was significantly lower in the KO cells compared to the NC group under the same conditions ( Fig. 5 c ) . Additionally, the expression of IL-12R and LCN2 was also significantly reduced ( Fig. 5 c ) , which was consistent with the results of RNA-seq analysis. Based on these alterations of inflammatory factors and the results of data analysis, we proposed a potential correlation between TCN2 and the STAT signaling pathway as well as the IL-17 signaling pathway. This indicates that knockdown of TCN2 mitigates the inflammatory response. Subsequently, combining the RNA-seq results and existing studies, we suggest that TCN2 may be able to influence the activation of the STAT3 pathway by modulating IL-1β secretion [ 8 ]. Therefore, we extracted proteins from IMQ-modeled mice skin for Western blotting experiments and found that TCN2 knockdown significantly suppressed the activation of the Stat3 pathway ( Fig. 5 d ). Similarly, we came to the same conclusion in in vitro cellular experiments ( Fig. 5 e ) . DISCUSSION Psoriasis, a chronic inflammatory disease marked by recurrent episodes, is clinically characterized by hyperproliferation of keratinocytes, culminating in thickened plaques and scaly papules[ 16 ]. Currently, numerous studies have been dedicated to unraveling the etiological factors underlying psoriasis, yielding remarkable and far-reaching results [ 17 – 19 ]. These studies have not only built a solid theoretical foundation for clinical treatment but also provided crucial support for developing targeted drugs [ 20 – 22 ]. Vitamin B12, also known as cobalamin, plays a crucial role in DNA synthesis, fatty acid metabolism, and particularly the conversion of methyl tetrahydrofolate to tetrahydrofolate [ 23 ]. Approximately 20–25% of circulating cobalamin is bound to TCN2, and deficiencies may result in neurological, hematologic, and metabolic system-related disorders [ 23 – 25 ]. Currently, most studies on TCN2 are based on human genomic data, and the mechanisms by which TCN2 drives disease pathogenesis remain poorly characterized. In this study, we conducted an in-depth analysis of the molecular mechanisms underlying TCN2’s role in psoriasis. Initial observations revealed significantly elevated TCN2 expression in the skin tissues of psoriasis patients, with prominent localization in both the basal layer and the entire epidermis. These findings suggest a potential association between TCN2 and the proliferation of keratinized cells, a hallmark of psoriatic pathology. Meanwhile, TCN2 levels were markedly upregulated in the PBMC of psoriasis patients. Intriguingly, a six-month therapeutic regimen with Secukinumab resulted in substantial reduction in TCN2 expression, further corroborating its association with psoriasis. To validate our findings, we conducted comprehensive animal studies. Remarkably, TCN2-knockout mice showed significantly reduced dorsal skin thickness and scaling. Consistently, immunohistochemical analysis revealed a significant decrease in Ki-67 expression, a marker of cellular proliferation, in the epidermal layers of knockout mice. Subsequently, we performed well-designed in vitro cell assays. These assays demonstrated that TCN2 deficiency induces G1-phase cell cycle arrest, thereby suppressing keratinocyte proliferation. Collectively, these data establish TCN2 as a critical regulator of keratinocyte hyperproliferation in psoriasis. Subsequently, we compared the expression of skin inflammatory factors between the Tcn2 -/- mice and WT controls. To lend further credence to these initial observations, we employed a TCN2-knockdown HaCat cell line. The results revealed a strong correlation between TCN2 and IL-1β levels. RNA-seq data further suggested that Tcn2 deficiency might exert an influence on Il-1β secretion. Moreover, pathway enrichment analysis of DEGs pre- and post-psoriasis induction highlighted significant alterations in the STAT pathway. Crucially, both in vivo and in vitro models confirmed that TCN2 modulates STAT3 pathway activation. Additionally, under identical inflammatory stimulation, we noted a notably milder inflammatory response in the TCN2-knockout mice. Meanwhile, in vitro results showed that TCN2-depleted HaCat cells exhibited suppressed STAT3 pathway activation and reduced production of inflammatory factors, such as IL-1β. IL-17, a key pro-inflammatory cytokine in psoriasis, drives disease pathogenesis by targeting keratinocytes and amplifying inflammation. Immunofluorescence assay revealed a significant reduction in IL-17 expression within the lesional area of TCN2 -knockdown mice. To further explore the interplay between TCN2 and IL-17, we analyzed TCN expression in PBMC from patients with moderate-to-severe psoriasis undergoing IL-17-targeted monoclonal antibody therapy. Strikingly, TCN2 levels correlated inversely with therapeutic efficacy, underscoring a close link TCN2 and IL-17 signaling. At the same time, the expression of TCN2 in mice and knockdown cell lines was positively correlated with S100A8 and S100A9. Integrated analysis of murine transcriptomic data further implicated TCN2 in IL-17 pathway regulation. The specific receptor of LCN2, 24p3R, is upregulated in the lesional skin of psoriasis patients. In vitro studies demonstrate that LCN2 enhances the expression of inflammatory cytokines IL-1β, IL-23, CXCL1 and CXCL10 [ 26 ]. IL-12 plays a central role in T-cell-mediated inflammatory responses, promoting CD4 + T cell differentiation into interferon-gamma (IFN-γ) and helper T cells[ 27 ]. IL-12 shares the p40 subunit with IL-23, and IL-12 is also implicated in the pathogenesis of psoriasis [ 28 ]. Our findings revealed that TCN2 knockdown significantly reduced IL-12R and LCN2 expression. This not only highlights the relationship between TCN2 and psoriasis but also underscores TCN2’s role as an upstream regulator of inflammatory signaling. This regulatory mechanism may critically influence psoriatic inflammation. Intriguingly, TCN2 -knockout mice exhibited reduced body weight compared to the WT controls, suggesting an additional role of TCN2 in growth and development. In conclusion, this study identifies TCN2 as a previously unrecognized mediator of keratinocyte hyperproliferation and inflammatory response in psoriasis, positioning it as a promising therapeutic target. MATERIALS AND METHODS Human subjects: Peripheral blood samples were collected from 25 patients diagnosed with psoriasis, including 14 individuals with moderate-to-severe disease treated with Secukinumab for more than 6 months. Additionally, 25 peripheral blood samples were collected from healthy individuals recruited through the dermatology clinic. Skin biopsies were collected from psoriasis patients to confirm diagnosis. Healthy control skin samples were derived from individuals undergoing procedures requiring skin excision. PASI scale was used to quantify disease severity. Detailed demographic and clinical characteristics of participants are presented in Supplementary Table 1 . Peripheral blood samples were collected in EDTA- anticoagulated tubes and centrifuged at 1,500 rpm for 5 minutes. The supernatant serum was stored at -80°C; meanwhile, the precipitated layer was used to isolate peripheral blood mononuclear cells (PBMC). Cell culture and treatment: PBMCs were isolated by density gradient centrifugation utilizing Ficoll-Paque™ (GE Healthcare, USA). In short, a blood sample was diluted with sterile PBS, gently layered on a Ficoll-paque, then centrifuged at 1800 rpm for 20 minutes at 20°C (brakes disabled). Then, the PBMC was collected from the plasma-Ficoll interface. The HaCat cell line, derived from immortalized normal human keratinocytes, retains differentiation characteristics akin to primary human keratinocytes, and is widely used in cutaneous biology research. HaCat cellss were purchased from Procell Life Sciences Technology (Wuhan, China), and cultured in high-glucose Dulbecco’s Modified Eagle Medium (DMEM) (SH30243.01, HyClone, China), supplemented with 10% fetal bovine serum (FSP500, ExCell, Uruguay) and 1% penicillin-streptomycin. Cells were maintained in 6-well culture plates for different experiments. Short-hairpin RNA (shRNA) sequences were designed as listed in Supplementary Table 2 and subcloned into GV493 vectors. Transfection was performed using Lipofectamine™ 3000 transfection kit (L3000015; Invitrogen, Carlsbad, CA, USA). Animal experiments: The animal model utilized 8-week-old wild-type (WT) C57BL/6 mice purchased from SiBeiFu (Beijing, China) and TCN2-knockout (Tcn2-/-) mice (obtained from Cyagen Bioscience, Jiangsu, China). All mice were female with a C57BL/6 gene background, and maintained in a specific pathogen-free environment at the China-Japan Friendship Hospital (Beijing, China). To establish the psoriasis skin model, mice underwent back hair removal followed by daily tropical application of 62.5mg imiquimod cream daily for 7 consecutive days. Dynamic changes in erythema, scaling and skin thickening were monitored daily, and photographs of skin lesions, back skin thickness and body weight recordings were obtained every day. Staining: Skin tissue samples were fixed in 4% paraformaldehyde, embedded in OCT, and then sectioned into 5–10µm slices. Sections were subjected to hematoxylin and eosin (HE) staining and immunofluorescence analysis. Cell proliferation and cycle assays: The cells were inoculated in 96-well plates (5000 cells/well) and cultured for 0–5 days. To evaluate cell proliferation, 10µl of CCK-8 reagent (Yeasen, China) and 100µl of medium were added to each well, followed by incubation at 37 ° C for 1 hour. Optical density (OD) was measured at 450 nm using a microplate reader (Thermo Scientific, USA). To analyze the cell cycle, cells (1×106 cells/well) were stained in the dark with DNA marker solution (CYT-PIR-25, Cytognos, Spain) at room temperature for 10 minutes. Subsequently, cell cycle data were obtained using flow cytometry (BD Calibur, USA) under low flow rate settings. Quantitative polymerase chain reaction: Total RNA was extracted using TRizol® reagent (Invitrogen, USA) according to manufacturer's instructions. Subsequently, the RNA was reverse-transcribed into cDNA using Hifair® III 1st Strand cDNA Synthesis SuperMix (Yeasen, China). Gene expression levels were quantified using the Hieff UNICONT® Universal Blue qPCR SYBR Master Mix (Yeasen, China). ACTB served as the endogenous control for normalization. The primer sequences for qPCR are listed in Supplementary Table 2. Western blotting: Approximately 20 µg of protein was separated by 10% SDS-PAGE and transferred onto polyvinylidene fluoride membranes (Merck Millipore, USA). The membranes were blocked with 5% bovine serum albumin (Yeasen, China) for 1 hour at room temperature. Subsequently, membranes were incubated overnight at 4°C with primary antibodies, followed by 1-hour incubation with secondary antibodies. β-actin was used as the loading control. The bound antibodies were visualized using an enhanced chemiluminescence (ECL) detection system (Merck Millipore, USA). The information of primary antibodies is provided in Supplementary Table 3. Statistical analysis: The results are expressed as mean ± standard deviation. Student’s t test was used for comparisons between two groups, and one-way analysis of variance was applied for multi-group analysis. Statistical analyses were performed using Prism 9 software (GraphPad, La Jolla, CA, USA). A p value < 0.05 was considered statistically significant. Data availability statement The datasets presented in this study can be found in online repositories. RNA-seq data presented in the study are deposited in the GEO repository, accession number GSE295541. Further inquiries can be directed to the corresponding authors and first author upon reasonable request. Declarations Funding This work was supported by the National Key R&D Program of China (2022YFC3602002), the National Natural Science Foundation of China (Grant 82103100, 82404133, 82304013), the Elite Medical Professionals Project of China-Japan Friendship Hospital (ZRJY2023-QM27), and the National High-Level Hospital Clinical Research Funding (2024-NHLHCRF-TS-01). Competing interests The authors declare that they have no competing interests. Author Contributions All authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by [ Jingkai Xu], [Xinzhu Zhou] and [Ke Xue].Methodology,validation,formal analysis and visualization were performed by[Ang Li],[Qingyue Xia],[Zhou Zhuang] and [Xuejiao Song].Funding acquisition and supervision were performed by[Xianbo Zuo] and [Yong Cui].The first draft of the manuscript was written by [Xinzhu Zhou] and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript. Ethics approval All procedures involving human samples conformed to the ethical principles of the Declaration of Helsinki, and written informed consent was obtained from all participants prior to sample collection. This study was approved by the Ethics Committee of China-Japan Friendship Hospital (2023-KY-344-1). Consent to participate Informed consent was obtained from all individual participants included in the study. References R. Al-Horani, T. Chui, and B. Hamad. 2024. 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Chatthanawaree. 2011. Biomarkers of cobalamin (vitamin B12) deficiency and its application. Journal of Nutrition Health & Aging 15:227-231. A. Oussalah, J. Levy, P. Filhine-Trésarrieu, F. Namour, and J.L. Guéant. 2017. Association of TCN2 rs1801198 c.776G>C polymorphism with markers of one-carbon metabolism and related diseases: a systematic review and meta-analysis of genetic association studies. American Journal of Clinical Nutrition 106:1142-1156. J.Y. Ma, J.L. Chen, K. Xue, C. Yu, E.L. Dang, H.J. Qiao, H. Fang, B.Y. Pang, Q.Y. Li, Z.B. Sun, P. Qiao, L. Wang, G. Wang, and S. Shao. 2022. LCN2 Mediates Skin Inflammation in Psoriasis through the SREBP2-NLRC4 Axis. Journal of Investigative Dermatology 142:2194-2204.e11.. M. Reddy, C. Davis, J. Wong, P. Marsters, C. Pendley, and U. Prabhakar. 2007. Modulation of CLA, IL-12R, CD40L, and IL-2Rα expression and inhibition of IL-12- and IL-23-induced cytokine secretion by CNTO 1275. Cellular Immunology 247:1-11. M.W.L. Teng, E.P. Bowman, J.J. McElwee, M.J. Smyth, J.L. Casanova, A.M. Cooper, and D.J. Cua. 2015. IL-12 and IL-23 cytokines: from discovery to targeted therapies for immune-mediated inflammatory diseases. Nature Medicine 21:719-729. Additional Declarations No competing interests reported. Supplementary Files Supplementarytable.xlsx SupplementaryFigureS1.docx Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-7309098","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":501303448,"identity":"5333c1d3-efbe-404e-9062-a96cfc570f2d","order_by":0,"name":"Jing-Kai Xu","email":"","orcid":"","institution":"China-Japan Friendship Hospital","correspondingAuthor":false,"prefix":"","firstName":"Jing-Kai","middleName":"","lastName":"Xu","suffix":""},{"id":501303449,"identity":"8aa44228-80b5-40e9-8a7a-d7bbf742f93b","order_by":1,"name":"Xin-Zhu Zhou","email":"data:image/png;base64,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","orcid":"","institution":"Peking University China-Japan Friendship School of Clinical Medicine","correspondingAuthor":true,"prefix":"","firstName":"Xin-Zhu","middleName":"","lastName":"Zhou","suffix":""},{"id":501303450,"identity":"955131d0-be97-46b3-b415-aae1b3e230a3","order_by":2,"name":"Ke Xue","email":"","orcid":"","institution":"China-Japan Friendship Hospital","correspondingAuthor":false,"prefix":"","firstName":"Ke","middleName":"","lastName":"Xue","suffix":""},{"id":501303451,"identity":"5268d527-9a3d-452d-ada7-69e800671d4d","order_by":3,"name":"Ang Li","email":"","orcid":"","institution":"China-Japan Friendship Hospital, Chinese Academy of Medical Sciences \u0026 Peking Union Medical College","correspondingAuthor":false,"prefix":"","firstName":"Ang","middleName":"","lastName":"Li","suffix":""},{"id":501303452,"identity":"a123461f-5be1-4a4e-a9d8-d717979f7d9b","order_by":4,"name":"Qing-Yue Xia","email":"","orcid":"","institution":"China-Japan Friendship Hospital, Chinese Academy of Medical Sciences \u0026 Peking Union Medical College","correspondingAuthor":false,"prefix":"","firstName":"Qing-Yue","middleName":"","lastName":"Xia","suffix":""},{"id":501303453,"identity":"c97983d8-ee3f-4577-848c-45a038c0b1b2","order_by":5,"name":"Zhou Zhuang","email":"","orcid":"","institution":"Peking University China-Japan Friendship School of Clinical Medicine","correspondingAuthor":false,"prefix":"","firstName":"Zhou","middleName":"","lastName":"Zhuang","suffix":""},{"id":501303454,"identity":"4a3e1e13-b8a6-491b-bb63-e45065978526","order_by":6,"name":"Xue-Jiao Song","email":"","orcid":"","institution":"China-Japan Friendship Hospital","correspondingAuthor":false,"prefix":"","firstName":"Xue-Jiao","middleName":"","lastName":"Song","suffix":""},{"id":501303455,"identity":"48190f34-273d-4232-9753-65bbff7a8b23","order_by":7,"name":"Xian-Bo Zuo","email":"","orcid":"","institution":"China-Japan Friendship Hospital","correspondingAuthor":false,"prefix":"","firstName":"Xian-Bo","middleName":"","lastName":"Zuo","suffix":""},{"id":501303456,"identity":"3a769ee7-4f06-467d-bc76-94d78991e332","order_by":8,"name":"Yong Cui","email":"","orcid":"","institution":"China-Japan Friendship Hospital","correspondingAuthor":false,"prefix":"","firstName":"Yong","middleName":"","lastName":"Cui","suffix":""}],"badges":[],"createdAt":"2025-08-06 11:08:20","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-7309098/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7309098/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":89378647,"identity":"acfa029e-76ba-4f6d-b429-c964d07a4fff","added_by":"auto","created_at":"2025-08-19 11:33:18","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":503354,"visible":true,"origin":"","legend":"\u003cp\u003eTCN2 is significantly elevated in psoriasis patient samples and mouse models\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ea.\u003c/strong\u003e Representative immunohistochemical images of TCN2 in skin biopsy specimens from psoriasis patients and healthy donors N=3. Scale bar=100 μm \u003cstrong\u003eb.\u003c/strong\u003e TCN2 mRNA levels in PBMC from psoriasis patients (PS, N=25) and healthy donors (HC, N=25). \u003cstrong\u003ec.\u003c/strong\u003e TCN2 mRNA levels and PASI scores in PBMC from moderate-to-severe psoriasis patients (PS, N=30) treated with Secukinumab and healthy donors (HC, N=32). TCN2 mRNA levels and PASI scores in PBMCs of patients with moderate-to-severe psoriasis (N=14) treated for 6 months at baseline and after treatment. \u003cstrong\u003ed.\u003c/strong\u003e \u003cem\u003eTcn2\u003c/em\u003emRNA and protein levels in skin samples from wild-type mice (WT, N=5) and psoriasis model mice (N=5). \u003cstrong\u003ee.\u003c/strong\u003e Immunofluorescence staining showing TCN2 (green) and DAPI (blue). * p \u0026lt; 0.05, ** p \u0026lt; 0.01, *** p \u0026lt; 0.001, **** p \u0026lt; 0.0001).\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-7309098/v1/de39c23a9cb812584652d216.png"},{"id":89378650,"identity":"fb4b8500-6ceb-4147-80ac-f3224c8677e2","added_by":"auto","created_at":"2025-08-19 11:33:18","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":630829,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003eTcn2 \u003c/em\u003edeletion alleviates the psoriasis phenotype\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ea.\u003c/strong\u003e WT, \u003cem\u003eTCN2\u003c/em\u003e-knockdown heterozygous mice and \u003cem\u003eTCN2\u003c/em\u003e knockdown pure mice were treated with imiquimod for 7 consecutive days. \u003cstrong\u003eb.\u003c/strong\u003e Representative H\u0026amp;E-stained sections of skin tissues from the three mice groups o after IMQ treatment. \u003cstrong\u003ec. \u003c/strong\u003eMeasurement of skin thickness in lesional skin areas across the three groups. \u003cstrong\u003ed.\u003c/strong\u003e Spleen-to-body weight ratios of the three groups. \u003cstrong\u003ee.\u003c/strong\u003e Immunofluorescence images of the skin samples of modeled mice in the various groups. Il -17a (green) and Cd45 (red). (* p \u0026lt; 0.05, ** p \u0026lt; 0.01, *** p \u0026lt; 0.001).\u003c/p\u003e","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-7309098/v1/12d33d5bae22fd803651b4ef.png"},{"id":89380094,"identity":"4c19807b-18d7-44d4-a2d8-b41d3dce9c03","added_by":"auto","created_at":"2025-08-19 11:41:18","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":313488,"visible":true,"origin":"","legend":"\u003cp\u003eRNA-seq reveals the mechanism by which TCN2 ameliorates IMQ-induced psoriasis\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ea. \u003c/strong\u003eVenn diagram of differential genes in Ctr group and Pso group, Pso group and T2ho group. \u003cstrong\u003eb.\u003c/strong\u003eVolcanic maps of downregulated (blue) and upregulated (red) DEGs in Ctr and Pso groups, Pso and T2ho groups. \u003cstrong\u003ec.\u003c/strong\u003e Heatmap of downregulated DEGs associated with inflammatory responses. \u003cstrong\u003ed.\u003c/strong\u003e Downregulated DEGs classified based on their gene ontology (GO) terms for biological process (BP), cellular component (CC), and molecular function (MF). \u003cstrong\u003ee.\u003c/strong\u003e Heatmap comparing the enrichment scores of key pathways across the three groups.\u003c/p\u003e","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-7309098/v1/e8d119ec13db4000740d02fe.png"},{"id":89380095,"identity":"05e268cd-8b3b-459a-913a-440d024c1abe","added_by":"auto","created_at":"2025-08-19 11:41:18","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":465666,"visible":true,"origin":"","legend":"\u003cp\u003ePro-proliferative effects of TCN2 in psoriasis\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ea, b. \u003c/strong\u003eWestern blotting to verify the efficiency of TCN2 knockdown. \u003cstrong\u003ec. \u003c/strong\u003eCCK-8 to detect the proliferation rate of control (NC) and TCN2 knockdown strain (TKO) cells. \u003cstrong\u003ed. \u003c/strong\u003eFlow cytometry to detect the cell cycle of NC and TKO groups. \u003cstrong\u003ee.\u003c/strong\u003eComparison of the percentage of cells at each stage of the cell cycle in NC and TKO groups. \u003cstrong\u003ef.\u003c/strong\u003e Representative immunofluorescence images of the skin samples of modeled mice in the various groups summarizing the Ki67 (red) expression in representative immunofluorescence images. (* p \u0026lt; 0.05, ** p \u0026lt; 0.01, *** p \u0026lt; 0.001)\u003c/p\u003e","description":"","filename":"floatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-7309098/v1/73893fd01a914be7779b54e1.png"},{"id":89378654,"identity":"34cdb90f-973c-4bf9-9079-1ca1556e2cd9","added_by":"auto","created_at":"2025-08-19 11:33:18","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":396145,"visible":true,"origin":"","legend":"\u003cp\u003ePro-inflammatory effects of TCN2 in psoriasis\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ea\u003c/strong\u003e. qPCR to compare the changes in inflammatory factors in the skin of WT versus \u003cem\u003eTcn2\u003c/em\u003e-/- mice. \u003cstrong\u003eb. \u003c/strong\u003eWestern blotting to validate the changes in Il-17a expression in the skin of WT versus \u003cem\u003eTcn2\u003c/em\u003e-/- mice. \u003cstrong\u003ec.\u003c/strong\u003e qPCR to compare the changes in inflammatory factors in the NC group versus the TCN2 KO group after the use of M5 stimulant.\u003cstrong\u003e d.\u003c/strong\u003e Western blotting to validate the changes in Stat3 pathway activation in the skin of WT versus \u003cem\u003eTcn2\u003c/em\u003e-/- mice. \u003cstrong\u003ee.\u003c/strong\u003eWestern blotting to verify the changes in STAT3 pathway activation in NC group and TCN2 KO group after the use of M5 stimulant. (* p \u0026lt; 0.05, ** p \u0026lt; 0.01, *** p \u0026lt; 0.001, **** p \u0026lt; 0.0001)\u003c/p\u003e","description":"","filename":"floatimage5.png","url":"https://assets-eu.researchsquare.com/files/rs-7309098/v1/a34f9c2eb68d05eea2523f33.png"},{"id":97136101,"identity":"e8f25d54-c6eb-46fb-bc00-233c553765fe","added_by":"auto","created_at":"2025-12-01 09:55:32","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":3111072,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7309098/v1/26afb010-8c09-4eb2-8a59-456c36f2625c.pdf"},{"id":89378651,"identity":"63541dc4-c350-417d-8d59-45f8d7036c7e","added_by":"auto","created_at":"2025-08-19 11:33:18","extension":"xlsx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":47509,"visible":true,"origin":"","legend":"","description":"","filename":"Supplementarytable.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-7309098/v1/6ac9cf741185801222e24dea.xlsx"},{"id":89378653,"identity":"d6fe2d54-8153-44f5-9d75-9857e3335cfa","added_by":"auto","created_at":"2025-08-19 11:33:18","extension":"docx","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":728653,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryFigureS1.docx","url":"https://assets-eu.researchsquare.com/files/rs-7309098/v1/ac7a31f5ea8a2272a8330c4a.docx"}],"financialInterests":"No competing interests reported.","formattedTitle":"TCN2 promotes psoriatic skin inflammation and keratinocyte proliferation via the IL-1β-STAT3 pathway","fulltext":[{"header":"INTRODUCTION","content":"\u003cp\u003ePsoriasis is a relapsing, immune-mediated, and chronic inflammatory skin disease that affects more than 125\u0026nbsp;million individuals worldwide, or about 2\u0026ndash;3% of the global population [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. Mechanistically, psoriatic lesions exhibit substantial infiltration of monocytes, neutrophils, and T cells. The inflammatory stimulation, abnormal differentiation and proliferation of keratinocytes, as well as dysregulated activation of stem cells work together to cause keratinocyte overgrowth and subsequent formation of thick, scaly plaques lesions [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eCurrent understanding of psoriasis immunopathogenesis highlights the pivotal role of pro-inflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin-1 (IL-1), and interleukin-6 (IL-6), et.al. They can synergistically activate keratinocytes to produce chemokines and antimicrobial peptides [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. In psoriasis lesions, chemokines such as C-C motif chemokine ligand 20 (CCL20), C-X-C motif chemokine ligand 1 (CXCL1), CXCL2, and CXCL8 play a key role in the recruitment and activation of immune cells, particularly interleukin 17A (IL-17A)-generated T cells and neutrophils [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. At the same time, keratinocytes release antimicrobial peptides, including S100 calcium-binding protein A7 (S100A7), S100A8, S100A9, and Defensin β 4A/defensin β 2(DEFB4A/DEFB2). These peptides not only exert direct antimicrobial effects but also amplify local inflammatory response. This process contributes to the formation of a complex inflammatory network. [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e, \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]\u003c/p\u003e\u003cp\u003eIL-1β, a classic proinflammatory cytokine, activates dermal γδT cells to produce IL-17, and stimulates chemokine release [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. And, in psoriatic lesions, IL-1β is predominantly secreted by keratinocytes [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eTranscobalamin II (TCN2) is a vitamin B12-binding protein that aids in the cellular uptake of vitamin B12, thus being a biomarker for active vitamin B12 [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Numerous studies have reported a close association between TCN2 and the development of autoimmune-related diseases, including multiple sclerosis, systemic lupus erythematosus and idiopathic recurrent spontaneous abortion [\u003cspan additionalcitationids=\"CR11 CR12\" citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. It can participate in the occurrence of diseases by regulating a variety of biological processes, such as cell proliferation, apoptosis, and migration [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. However, the role of TCN2 in psoriasis remains unexplored.\u003c/p\u003e\u003cp\u003eIn this study, we propose that TCN2 promotes aberrant proliferation of keratinocytes and exacerbates the inflammatory response in psoriasis by affecting IL-1β secretion. Mechanistically, TCN2 can inhibit the secretion of IL-17 and the activation of STAT3 pathway by affecting the expression of IL-1β and down-regulating the expression of related pro-inflammatory factors. Our study suggests the potential of TCN2 as a new therapeutic target for psoriasis.\u003c/p\u003e"},{"header":"RESULT","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003eTCN2 expression is significantly elevated in psoriasis patients and IMQ - induced models\u003c/h2\u003e\u003cp\u003eTo explore the expression of TCN2, we first conducted immunohistochemical staining on skin sections from healthy controls (HCs) and psoriasis patients. In HC skin TCN2 is primarily distributed in the epidermis basal layer. In psoriatic skin, TCN2 is widely expressed in the thickened epidermis, with increased expression in dermal vascular endothelial cells and inflammatory cells \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ea\u003cb\u003e)\u003c/b\u003e. The detailed information of the patients were shown in \u003cb\u003eSupplementary Table\u0026nbsp;1\u003c/b\u003e. Meanwhile, we measured the expression of TCN2 in PBMCs of 25 psoriasis patients and 25 HCs. It was found that the expression of TCN2 was significantly upregulated in psoriasis patients \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb\u003cb\u003e)\u003c/b\u003e. These patients underwent clinical treatment with Secukinumab. Six months after treatment initiation, the peripheral blood of 14 patients was collected, and the expression of TCN2 was measured. Results demonstrated that, after treatment, not only was the psoriasis area and severity index (PASI) of these patients significantly decreased, but the expression of TCN2 was also significantly reduced \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ec\u003cb\u003e)\u003c/b\u003e.\u003c/p\u003e\u003cp\u003eTo further explore the expression of TCN2 in psoriasis, we employed IMQ-induced wide type (WT) mice to establish a psoriasis mouse model. qPCR, Western blotting and immunofluorescence staining were conducted to compare the expression of TCN2 in the skin tissues of WT and WT\u0026thinsp;+\u0026thinsp;IMQ mice. The results revealed that TCN2 levels were significantly upregulated in the WT\u0026thinsp;+\u0026thinsp;IMQ mice \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ed and \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ee\u003cb\u003e)\u003c/b\u003e. Taken together, these data suggest that TCN2 is associated with the progression of psoriasis and may contribute to its pathogenesis.\u003c/p\u003e\u003cp\u003e\u003cb\u003eTcn2\u003c/b\u003e \u003cb\u003edeficiency attenuates IMQ-induced psoriasis phenotype\u003c/b\u003e\u003c/p\u003e\u003cp\u003eGene-edited mice have been extensively employed to explore the pathogenesis of numerous diseases. To clarify the role of TCN2 in psoriasis, we first generated a Tcn2-/- mouse model. Subsequently, we set up three distinct groups of psoriasis mouse models: WT\u0026thinsp;+\u0026thinsp;IMQ, Tcn2+/- + IMQ (T+/- + IMQ) and Tcn2-/- +IMQ (T-/- + IMQ) mice.\u003c/p\u003e\u003cp\u003eSeven days\u0026ndash;after IMQ induction, compared to the T-/- + IMQ mice, WT\u0026thinsp;+\u0026thinsp;IMQ mice exhibited more prominent scaling and roughness in the dorsal skin. Moreover, the severity of epidermal hyperplasia and spleen size decreased successively in the WT\u0026thinsp;+\u0026thinsp;IMQ, T+/- + IMQ and T-/- + IMQ mice \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ea \u003cb\u003eand Supplementary Fig \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003ea)\u003c/b\u003e. After that, HE staining was performed on the three groups, and results demonstrated that epidermal hyperplasia and inflammatory infiltration were more pronounced in the WT\u0026thinsp;+\u0026thinsp;IMQ mice compared to the T-/- + IMQ mice \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eb\u003cb\u003e)\u003c/b\u003e. Additionally, the epidermal thickness and spleen/body weight ratio were significantly decreased in the T-/- + IMQ mice \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ec \u003cb\u003eand d)\u003c/b\u003e. Meanwhile, immunofluorescence and Western blotting revealed diminished IL17 and Cd45 expression in the T-/- + IMQ mice \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ee\u003cb\u003e)\u003c/b\u003e. We found that TCN2 deficiency could significantly reduce psoriasis-related inflammation. The findings demonstrate that TCN2 knockdown attenuates epidermal hyperplasia and skin inflammation, suggesting its potential as a therapeutic target for psoriasis.\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eRNA-seq implicates TCN2 deficiency in inflammatory signaling\u003c/h3\u003e\n\u003cp\u003eTo further elucidate the mechanism of TCN2 deficiency in psoriasis treatment, we employed bulk RNA-seq to analyze the expression profiles of WT, WT\u0026thinsp;+\u0026thinsp;IMQ and T-/- + IMQ mice. By comparing the genes enriched in WT mice and WT\u0026thinsp;+\u0026thinsp;IMQ mice, WT\u0026thinsp;+\u0026thinsp;IMQ mice and T-/- + IMQ mice, we identified 193 differentially expressed genes (DEGs) among the three groups \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ea, \u003cb\u003eSupplementary Fig \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003eb and Supplementary table2)\u003c/b\u003e. Figure\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eb showed the volcanic maps of DEGs changes in different groups. Compared to the WT\u0026thinsp;+\u0026thinsp;IMQ mice, 204 genes in the T-/- + IMQ mice were up-regulated and 410 genes down-regulated. (Fold change\u0026thinsp;\u0026ge;\u0026thinsp;2 and \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05). And a lot of inflammation-related molecules (like Il1b, Il6, S100a7 and S100a8, et.al) were significantly reduced after TCN2 knockout (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ec).\u003c/p\u003e\u003cp\u003eMoreover, KEGG analysis showed that down-regulated DEGs were mainly involved in biological processes related to signal transduction and innate immune response, such as JAK-STAT, IL-17 pathway, Th17 cell differentiation, etc. \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ed\u003cb\u003e)\u003c/b\u003e. GSVA analysis was performed, revealing that TCN2 deficiency potently inhibited pivotal signaling pathways in psoriasis, and MAPK, NF-κB, and STAT3 pathways were significantly inhibited in the T-/- + IMQ mice \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ee\u003cb\u003e)\u003c/b\u003e. This observation implies that TCN2 deficiency might curtail the inflammatory response in psoriasis by impeding the activation of these pathways.\u003c/p\u003e\n\u003ch3\u003eTCN2 regulates the proliferation of epidermal cells in psoriasis\u003c/h3\u003e\n\u003cp\u003eExcessive proliferation of epidermal keratinocytes is a characteristic manifestation of psoriasis [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. To explore the impact of TCN2 knockdown on the proliferation of epidermal cells in psoriasis, we constructed a TCN2-knockdown HaCat cell line and verified that TCN2 was knocked down by Western blotting \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea \u003cb\u003eand b)\u003c/b\u003e. The CCK-8 assay demonstrated that, in comparison to the control cells, the proliferation rate of the TCN2-knockdown cell lines was markedly diminished \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ec\u003cb\u003e)\u003c/b\u003e. Furthermore, through flow cytometry, we demonstrated that the knockdown of TCN2 modulated the cell cycle by inducing a G1 phase arrest in the cell lines \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ed \u003cb\u003eand e)\u003c/b\u003e. Additionally, immunofluorescence techniques were employed to stain the skin tissues obtained from the three groups of mice, and it was ascertained that the silencing of TCN2 led to a decrease in Ki67 expression within the epidermis \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ef\u003cb\u003e)\u003c/b\u003e. Collectively, these results imply that the influence of TCN2 on cell proliferation and cycle might be associated with the excessive proliferation of epidermal cells.\u003c/p\u003e\n\u003ch3\u003eTCN2 regulates the progress of psoriasis through modulating IL-1β-STAT3 pathway\u003c/h3\u003e\n\u003cp\u003eTo further validate our RNA-seq results and elucidate the mechanism of TCN2 in psoriasis, we investigated the expression levels of inflammatory factors in the skin tissues from the T-/- + IMQ and WT\u0026thinsp;+\u0026thinsp;IMQ mice, and found that the expression of Il-1β, S100a8, and S100a9 was significantly reduced \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ea\u003cb\u003e)\u003c/b\u003e. In addition, we studied the expression of Il17-a in the skin of the four-mouse-group and observed that \u003cem\u003eTCN2\u003c/em\u003e knockdown led to a substantial reduction in Il-17a expression \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eb\u003cb\u003e)\u003c/b\u003e.\u003c/p\u003e\u003cp\u003eM5 stimulants (IL-1α, IL-17A, IL-22, TNF-α, and Oncostatin M) have been used to induce psoriasis-like inflammatory cells [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. In our study, we treated the TCN2-knockdown (KO) HaCaT cells with M5 at a concentration of 10ng/ml for 24 hours. Notably, we found the mRNA expression of inflammatory factors, including \u003cem\u003eIL-1β, IL-6, TNFα, S100A7, S100A8\u003c/em\u003e and \u003cem\u003eS100A9\u003c/em\u003e, was significantly lower in the KO cells compared to the NC group under the same conditions \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ec\u003cb\u003e)\u003c/b\u003e. Additionally, the expression of \u003cem\u003eIL-12R\u003c/em\u003e and \u003cem\u003eLCN2\u003c/em\u003e was also significantly reduced \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ec\u003cb\u003e)\u003c/b\u003e, which was consistent with the results of RNA-seq analysis. Based on these alterations of inflammatory factors and the results of data analysis, we proposed a potential correlation between TCN2 and the STAT signaling pathway as well as the IL-17 signaling pathway. This indicates that knockdown of TCN2 mitigates the inflammatory response.\u003c/p\u003e\u003cp\u003eSubsequently, combining the RNA-seq results and existing studies, we suggest that TCN2 may be able to influence the activation of the STAT3 pathway by modulating IL-1β secretion [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. Therefore, we extracted proteins from IMQ-modeled mice skin for Western blotting experiments and found that \u003cem\u003eTCN2\u003c/em\u003e knockdown significantly suppressed the activation of the Stat3 pathway \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ed\u003cb\u003e).\u003c/b\u003e Similarly, we came to the same conclusion in in vitro cellular experiments \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ee\u003cb\u003e)\u003c/b\u003e.\u003c/p\u003e"},{"header":"DISCUSSION","content":"\u003cp\u003ePsoriasis, a chronic inflammatory disease marked by recurrent episodes, is clinically characterized by hyperproliferation of keratinocytes, culminating in thickened plaques and scaly papules[\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Currently, numerous studies have been dedicated to unraveling the etiological factors underlying psoriasis, yielding remarkable and far-reaching results [\u003cspan additionalcitationids=\"CR18\" citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. These studies have not only built a solid theoretical foundation for clinical treatment but also provided crucial support for developing targeted drugs [\u003cspan additionalcitationids=\"CR21\" citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eVitamin B12, also known as cobalamin, plays a crucial role in DNA synthesis, fatty acid metabolism, and particularly the conversion of methyl tetrahydrofolate to tetrahydrofolate [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e]. Approximately 20\u0026ndash;25% of circulating cobalamin is bound to TCN2, and deficiencies may result in neurological, hematologic, and metabolic system-related disorders [\u003cspan additionalcitationids=\"CR24\" citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. Currently, most studies on TCN2 are based on human genomic data, and the mechanisms by which TCN2 drives disease pathogenesis remain poorly characterized. In this study, we conducted an in-depth analysis of the molecular mechanisms underlying TCN2\u0026rsquo;s role in psoriasis. Initial observations revealed significantly elevated TCN2 expression in the skin tissues of psoriasis patients, with prominent localization in both the basal layer and the entire epidermis. These findings suggest a potential association between TCN2 and the proliferation of keratinized cells, a hallmark of psoriatic pathology. Meanwhile, TCN2 levels were markedly upregulated in the PBMC of psoriasis patients. Intriguingly, a six-month therapeutic regimen with Secukinumab resulted in substantial reduction in TCN2 expression, further corroborating its association with psoriasis.\u003c/p\u003e\u003cp\u003eTo validate our findings, we conducted comprehensive animal studies. Remarkably, TCN2-knockout mice showed significantly reduced dorsal skin thickness and scaling. Consistently, immunohistochemical analysis revealed a significant decrease in Ki-67 expression, a marker of cellular proliferation, in the epidermal layers of knockout mice. Subsequently, we performed well-designed in vitro cell assays. These assays demonstrated that TCN2 deficiency induces G1-phase cell cycle arrest, thereby suppressing keratinocyte proliferation. Collectively, these data establish TCN2 as a critical regulator of keratinocyte hyperproliferation in psoriasis.\u003c/p\u003e\u003cp\u003eSubsequently, we compared the expression of skin inflammatory factors between the \u003cem\u003eTcn2\u003c/em\u003e-/- mice and WT controls. To lend further credence to these initial observations, we employed a TCN2-knockdown HaCat cell line. The results revealed a strong correlation between TCN2 and IL-1β levels. RNA-seq data further suggested that Tcn2 deficiency might exert an influence on Il-1β secretion. Moreover, pathway enrichment analysis of DEGs pre- and post-psoriasis induction highlighted significant alterations in the STAT pathway. Crucially, both in vivo and in vitro models confirmed that TCN2 modulates STAT3 pathway activation.\u003c/p\u003e\u003cp\u003eAdditionally, under identical inflammatory stimulation, we noted a notably milder inflammatory response in the TCN2-knockout mice. Meanwhile, in vitro results showed that TCN2-depleted HaCat cells exhibited suppressed STAT3 pathway activation and reduced production of inflammatory factors, such as IL-1β.\u003c/p\u003e\u003cp\u003eIL-17, a key pro-inflammatory cytokine in psoriasis, drives disease pathogenesis by targeting keratinocytes and amplifying inflammation. Immunofluorescence assay revealed a significant reduction in IL-17 expression within the lesional area of \u003cem\u003eTCN2\u003c/em\u003e-knockdown mice. To further explore the interplay between TCN2 and IL-17, we analyzed TCN expression in PBMC from patients with moderate-to-severe psoriasis undergoing IL-17-targeted monoclonal antibody therapy. Strikingly, TCN2 levels correlated inversely with therapeutic efficacy, underscoring a close link TCN2 and IL-17 signaling.\u003c/p\u003e\u003cp\u003eAt the same time, the expression of TCN2 in mice and knockdown cell lines was positively correlated with S100A8 and S100A9. Integrated analysis of murine transcriptomic data further implicated TCN2 in IL-17 pathway regulation.\u003c/p\u003e\u003cp\u003eThe specific receptor of LCN2, 24p3R, is upregulated in the lesional skin of psoriasis patients. In vitro studies demonstrate that LCN2 enhances the expression of inflammatory cytokines IL-1β, IL-23, CXCL1 and CXCL10 [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e]. IL-12 plays a central role in T-cell-mediated inflammatory responses, promoting CD4\u0026thinsp;+\u0026thinsp;T cell differentiation into interferon-gamma (IFN-γ) and helper T cells[\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]. IL-12 shares the p40 subunit with IL-23, and IL-12 is also implicated in the pathogenesis of psoriasis [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e]. Our findings revealed that TCN2 knockdown significantly reduced IL-12R and LCN2 expression. This not only highlights the relationship between TCN2 and psoriasis but also underscores TCN2\u0026rsquo;s role as an upstream regulator of inflammatory signaling. This regulatory mechanism may critically influence psoriatic inflammation. Intriguingly, \u003cem\u003eTCN2\u003c/em\u003e-knockout mice exhibited reduced body weight compared to the WT controls, suggesting an additional role of TCN2 in growth and development.\u003c/p\u003e\u003cp\u003eIn conclusion, this study identifies TCN2 as a previously unrecognized mediator of keratinocyte hyperproliferation and inflammatory response in psoriasis, positioning it as a promising therapeutic target.\u003c/p\u003e"},{"header":"MATERIALS AND METHODS","content":"\u003cdiv id=\"Sec9\" class=\"Section2\"\u003e\u003ch2\u003eHuman subjects:\u003c/h2\u003e\u003cp\u003ePeripheral blood samples were collected from 25 patients diagnosed with psoriasis, including 14 individuals with moderate-to-severe disease treated with Secukinumab for more than 6 months. Additionally, 25 peripheral blood samples were collected from healthy individuals recruited through the dermatology clinic.\u003c/p\u003e\u003cp\u003eSkin biopsies were collected from psoriasis patients to confirm diagnosis. Healthy control skin samples were derived from individuals undergoing procedures requiring skin excision. PASI scale was used to quantify disease severity. Detailed demographic and clinical characteristics of participants are presented in \u003cb\u003eSupplementary Table\u0026nbsp;1\u003c/b\u003e.\u003c/p\u003e\u003cp\u003ePeripheral blood samples were collected in EDTA- anticoagulated tubes and centrifuged at 1,500 rpm for 5 minutes. The supernatant serum was stored at -80\u0026deg;C; meanwhile, the precipitated layer was used to isolate peripheral blood mononuclear cells (PBMC).\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eCell culture and treatment:\u003c/h3\u003e\n\u003cp\u003ePBMCs were isolated by density gradient centrifugation utilizing Ficoll-Paque\u0026trade; (GE Healthcare, USA). In short, a blood sample was diluted with sterile PBS, gently layered on a Ficoll-paque, then centrifuged at 1800 rpm for 20 minutes at 20\u0026deg;C (brakes disabled). Then, the PBMC was collected from the plasma-Ficoll interface.\u003c/p\u003e\u003cp\u003eThe HaCat cell line, derived from immortalized normal human keratinocytes, retains differentiation characteristics akin to primary human keratinocytes, and is widely used in cutaneous biology research. HaCat cellss were purchased from Procell Life Sciences Technology (Wuhan, China), and cultured in high-glucose Dulbecco\u0026rsquo;s Modified Eagle Medium (DMEM) (SH30243.01, HyClone, China), supplemented with 10% fetal bovine serum (FSP500, ExCell, Uruguay) and 1% penicillin-streptomycin. Cells were maintained in 6-well culture plates for different experiments.\u003c/p\u003e\u003cp\u003eShort-hairpin RNA (shRNA) sequences were designed as listed in Supplementary Table\u0026nbsp;2 and subcloned into GV493 vectors. Transfection was performed using Lipofectamine\u0026trade; 3000 transfection kit (L3000015; Invitrogen, Carlsbad, CA, USA).\u003c/p\u003e\u003cdiv id=\"Sec11\" class=\"Section2\"\u003e\u003ch2\u003eAnimal experiments:\u003c/h2\u003e\u003cp\u003eThe animal model utilized 8-week-old wild-type (WT) C57BL/6 mice purchased from SiBeiFu (Beijing, China) and TCN2-knockout (Tcn2-/-) mice (obtained from Cyagen Bioscience, Jiangsu, China). All mice were female with a C57BL/6 gene background, and maintained in a specific pathogen-free environment at the China-Japan Friendship Hospital (Beijing, China). To establish the psoriasis skin model, mice underwent back hair removal followed by daily tropical application of 62.5mg imiquimod cream daily for 7 consecutive days. Dynamic changes in erythema, scaling and skin thickening were monitored daily, and photographs of skin lesions, back skin thickness and body weight recordings were obtained every day.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec12\" class=\"Section2\"\u003e\u003ch2\u003eStaining:\u003c/h2\u003e\u003cp\u003eSkin tissue samples were fixed in 4% paraformaldehyde, embedded in OCT, and then sectioned into 5\u0026ndash;10\u0026micro;m slices. Sections were subjected to hematoxylin and eosin (HE) staining and immunofluorescence analysis.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec13\" class=\"Section2\"\u003e\u003ch2\u003eCell proliferation and cycle assays:\u003c/h2\u003e\u003cp\u003eThe cells were inoculated in 96-well plates (5000 cells/well) and cultured for 0\u0026ndash;5 days. To evaluate cell proliferation, 10\u0026micro;l of CCK-8 reagent (Yeasen, China) and 100\u0026micro;l of medium were added to each well, followed by incubation at 37 \u0026deg; C for 1 hour. Optical density (OD) was measured at 450 nm using a microplate reader (Thermo Scientific, USA). To analyze the cell cycle, cells (1\u0026times;106 cells/well) were stained in the dark with DNA marker solution (CYT-PIR-25, Cytognos, Spain) at room temperature for 10 minutes. Subsequently, cell cycle data were obtained using flow cytometry (BD Calibur, USA) under low flow rate settings.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec14\" class=\"Section2\"\u003e\u003ch2\u003eQuantitative polymerase chain reaction:\u003c/h2\u003e\u003cp\u003eTotal RNA was extracted using TRizol\u0026reg; reagent (Invitrogen, USA) according to manufacturer's instructions. Subsequently, the RNA was reverse-transcribed into cDNA using Hifair\u0026reg; III 1st Strand cDNA Synthesis SuperMix (Yeasen, China). Gene expression levels were quantified using the Hieff UNICONT\u0026reg; Universal Blue qPCR SYBR Master Mix (Yeasen, China). ACTB served as the endogenous control for normalization. The primer sequences for qPCR are listed in Supplementary Table\u0026nbsp;2.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec15\" class=\"Section2\"\u003e\u003ch2\u003eWestern blotting:\u003c/h2\u003e\u003cp\u003eApproximately 20 \u0026micro;g of protein was separated by 10% SDS-PAGE and transferred onto polyvinylidene fluoride membranes (Merck Millipore, USA). The membranes were blocked with 5% bovine serum albumin (Yeasen, China) for 1 hour at room temperature. Subsequently, membranes were incubated overnight at 4\u0026deg;C with primary antibodies, followed by 1-hour incubation with secondary antibodies. β-actin was used as the loading control. The bound antibodies were visualized using an enhanced chemiluminescence (ECL) detection system (Merck Millipore, USA). The information of primary antibodies is provided in Supplementary Table\u0026nbsp;3.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec16\" class=\"Section2\"\u003e\u003ch2\u003eStatistical analysis:\u003c/h2\u003e\u003cp\u003eThe results are expressed as mean\u0026thinsp;\u0026plusmn;\u0026thinsp;standard deviation. Student\u0026rsquo;s t test was used for comparisons between two groups, and one-way analysis of variance was applied for multi-group analysis. Statistical analyses were performed using Prism 9 software (GraphPad, La Jolla, CA, USA). A p value\u0026thinsp;\u0026lt;\u0026thinsp;0.05 was considered statistically significant.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec17\" class=\"Section2\"\u003e\u003ch2\u003eData availability statement\u003c/h2\u003e\u003cp\u003eThe datasets presented in this study can be found in online repositories. RNA-seq data presented in the study are deposited in the GEO repository, accession number GSE295541. Further inquiries can be directed to the corresponding authors and first author upon reasonable request.\u003c/p\u003e\u003c/div\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis work was supported by the National Key R\u0026amp;D Program of China (2022YFC3602002), the National Natural Science Foundation of China (Grant 82103100, 82404133, 82304013), the Elite Medical Professionals Project of China-Japan Friendship Hospital (ZRJY2023-QM27), and the National High-Level Hospital Clinical Research Funding (2024-NHLHCRF-TS-01).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that they have no competing interests.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor Contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by [ Jingkai Xu], [Xinzhu Zhou] and [Ke Xue].Methodology,validation,formal analysis and visualization were performed by[Ang Li],[Qingyue Xia],[Zhou Zhuang] and [Xuejiao Song].Funding acquisition and supervision were performed by[Xianbo Zuo] and [Yong Cui].The first draft of the manuscript was written by [Xinzhu Zhou] and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll procedures involving human samples conformed to the ethical principles of the Declaration of Helsinki, and written informed consent was obtained from all participants prior to sample collection. This study was approved by the Ethics Committee of China-Japan Friendship Hospital (2023-KY-344-1).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eInformed consent was obtained from all individual participants included in the study.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eR. Al-Horani, T. Chui, and B. Hamad. 2024. The pipeline and market for psoriasis drugs. \u003cem\u003eNat Rev Drug Discov \u003c/em\u003e\u003c/li\u003e\n\u003cli\u003eJ. Mokry and R. Pisal. 2020. Development and Maintenance of Epidermal Stem Cells in Skin Adnexa. \u003cem\u003eInternational Journal of Molecular Sciences \u003c/em\u003e21\u003c/li\u003e\n\u003cli\u003eR. Singh, S. Koppu, P.O. Perche, and S.R. Feldman. 2021. 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IL-12 and IL-23 cytokines: from discovery to targeted therapies for immune-mediated inflammatory diseases. \u003cem\u003eNature Medicine\u003c/em\u003e 21:719-729.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"TCN2, psoriasis, skin inflammation","lastPublishedDoi":"10.21203/rs.3.rs-7309098/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7309098/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003ePsoriasis, a chronic immune-mediated inflammatory skin disorder, exerts systemic impacts across multiple organs. Although transcobalamin 2 (TCN2) has been reported to be associated with several autoimmune diseases, its role in psoriasis remains elusive. Hence, we investigated the role of TCN2 in psoriasis pathogenesis. Our results indicated that TCN2 was highly expressed in the skin tissues and peripheral blood mononuclear cells (PBMCs) of psoriatic patients, with downregulation following biologic therapy. Moreover, imiquimod (IMQ) - induced psoriasis in mice exhibited heightened TCN2 expression. To further explore the role of TCN2 in psoriasis, we generated Tcn2-deficient (Tcn2-/-) mice and established a psoriasis model using IMQ. IMQ-treated Tcn2-/- mice displayed milder psoriatic lesions and a lower level of inflammation. RNA-seq analysis of lesional skin revealed significant downregulation of inflammatory mediators (S100A7, S100A8, S100A9, IL-1β, IL-6) and suppression of STAT3 signaling in Tcn2-/- mice compared to WT-IMQ mice. Parallel in vitro experiments using TCN2-knockout HaCaT cells demonstrated cell cycle arrest. Collectively, our findings highlight TCN2 as a critical regulator of psoriatic inflammation, proposing it as a novel therapeutic target.\u003c/p\u003e","manuscriptTitle":"TCN2 promotes psoriatic skin inflammation and keratinocyte proliferation via the IL-1β-STAT3 pathway","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-08-19 11:33:13","doi":"10.21203/rs.3.rs-7309098/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"b608c774-7689-4951-83e3-b7a8e9fb1f0b","owner":[],"postedDate":"August 19th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-11-26T05:39:08+00:00","versionOfRecord":[],"versionCreatedAt":"2025-08-19 11:33:13","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7309098","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7309098","identity":"rs-7309098","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}