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BackgroundIn previous human skin single-cell data, inflammatory cells constituted only a small fraction of the overall cell population, such that functional subsets were difficult to ascertain.ObjectiveOur aims were to overcome the aforesaid limitation by applying single-cell transcriptomics to emigrating cells from skin and elucidate ex vivo gene expression profiles of pathogenic versus regulatory immune cell subsets in the skin of individuals with psoriasis.MethodsWe harvested emigrating cells from human psoriasis skin after incubation in culture medium without enzyme digestion or cell sorting and analyzed cells with single-cell RNA sequencing and flow cytometry simultaneously.ResultsUnsupervised clustering of harvested cells from psoriasis skin and control skin identified natural killer cells, T-cell subsets, dendritic cell subsets, melanocytes, and keratinocytes in different layers. Comparison between psoriasis cells and control cells within each cluster revealed that (1) cutaneous type 17 T cells display highly differing transcriptome profiles depending on IL-17A versus IL-17F expression and IFN-γ versus IL-10 expression; (2) semimature dendritic cells are regulatory dendritic cells with high IL-10 expression, but a subset of semimature dendritic cells expresses IL-23A and IL-36G in psoriasis; and (3) CCL27-CCR10 interaction is potentially impaired in psoriasis because of decreased CCL27 expression in basal keratinocytes.ConclusionWe propose that single-cell transcriptomics applied to emigrating cells from human skin provides an innovative study platform to compare gene expression profiles of heterogenous immune cells in various inflammatory skin diseases.

Introduction Pemphigus is a rare and life-threatening autoimmune disease that manifests as blisters and erosions on skin and/or mucosa. It is mediated by autoreactive B cells targeting desmoglein 1 (DSG1) [...], BACKGROUND. Pemphigus, a rare autoimmune bullous disease mediated by antidesmoglein autoantibodies, can be controlled with systemic medication like rituximab and high-dose systemic corticosteroids combined with immunosuppressants. However, some patients continue to experience chronically recurrent blisters in a specific area and require long-term maintenance systemic therapy. METHODS. Skin with chronic blisters was obtained from patients with pemphigus. Immunologic properties of the skin were analyzed by immunofluorescence staining, bulk and single-cell RNA and TCR sequencing, and a highly multiplex imaging technique known as CO-Detection by indEXing (CODEX). Functional analyses were performed by flow cytometry and bulk RNA-Seq using peripheral blood from healthy donors. Intralesional corticosteroid was injected into patient skin, and changes in chronically recurrent blisters were observed. RESULTS. We demonstrated the presence of skin tertiary lymphoid structures (TLSs) with desmoglein-specific B cells in chronic blisters from patients with pemphigus. In the skin TLSs, [CD4.sup.+] T cells predominantly produced CXCL13. These clonally expanded [CXCL13.sup.+][CD4.sup.+] T cells exhibited features of activated Th1-like cells and downregulated genes associated with T cell receptor-mediated signaling. Tregs are in direct contact with [CXCL13.sup.+][CD4.sup.+] memory T cells and increased CXCL13 production of [CD4.sup.+] T cells through IL-2 consumption and TGF-p stimulation. Finally, intralesional corticosteroid injection improved chronic blisters and reduced skin TLSs in patients with pemphigus. CONCLUSION. Through this study we conclude that skin TLSs are associated with the persistence of chronically recurrent blisters in patients with pemphigus, and the microenvironmental network involving [CXCL13.sup.+][CD4.sup.+] T cells and Tregs within these structures plays an important role in CXCL13 production. TRIAL REGISTRATION. ClinicalTrials.gov NCT04509570. FUNDING. This work was supported by National Research Foundation of South Korea (NRF-2021R1C1C1007179) and Korea Drug Development Fund, which is funded by Ministry of Science and ICT; Ministry of Trade, Industry, and Energy; and Ministry of Health and Welfare (grant RS-2022-00165917).

Background: Trials and real‐life studies demonstrated clinically meaningful improvements of disease activity in the majority of patients with moderate to severe atopic dermatitis (AD) treated with the anti‐IL‐4RA‐antibody dupilumab. However, misdiagnosis or confounding skin diseases in particular cutaneous T‐cell lymphoma (CTCL) may lead to inadequate response. Objective: To investigate the clinical and pathological features of patients with AD who showed insufficient response to dupilumab. Methods: We reviewed the medical records of 371 patients treated with dupilumab for severe AD. Insufficient response was defined as failure to achieve an improvement of the eczema area severity index (EASI) of at least 50% (EASI‐50) at Week 16 and of 75% (EASI‐75) at Week 52. Among 46 patients with insufficient response, 35 patients consented to a re‐evaluation including a full physical exam, biopsies and laboratory assessments including immunohistochemistry and T‐cell receptor gene rearrangement analysis to differentiate CTCL. Results: Of the 371 patients treated with dupilumab, 46 (12.3%) patients showed insufficient response to dupilumab. Of these, 35 underwent further evaluation, and 19 (54.2% of inadequate responders) were finally diagnosed with mycosis fungoides (MF). In these patients, transition to or addition of conventional MF treatment led to clinical improvements. Conclusions: Insufficient response to dupilumab treatment may help uncover early MF on an existing AD background. [ABSTRACT FROM AUTHOR]

This article discusses a study that used spatial transcriptomics to gain insights into the pathogenesis and therapy of pemphigus vulgaris (PV) and bullous pemphigoid (BP), which are autoimmune bullous skin diseases. The study compared the gene expression profiles of intact and blistered epidermis, as well as dermal immune cells, in PV and BP patients. The findings revealed distinct pathways and gene expression patterns associated with blister formation in each disease. The study provides a foundation for identifying potential therapeutic targets to regulate intracellular pathways involved in blister formation. [Extracted from the article]

Background: Studies on the interaction between tumour‐infiltrating immune cells (TIICs) and tumour cells in melanoma arising from congenital melanocytic nevus (CMN) are lacking. Objective: The aim of this study was to determine the intratumoral immune landscape of TIICs and tumour cells during invasion and metastasis. Methods: Tissue specimens were obtained from patients with melanoma originating from CMN. Differential gene expression in melanoma cells and TIICs during invasion and metastasis was determined using spatial transcriptomics. Results: As invasion depth increased, the expression of LGALS3, known to induce tumour‐driven immunosuppression, increased in melanoma cells. In T cells, the expression of genes that inhibit T‐cell activation increased with increasing invasion depth. In macrophages, the expression of genes related to the anti‐inflammatory M2 phenotype was upregulated with increasing invasion depth. Compared to primary tumour cells, melanoma cells in metastatic lesions showed upregulated expression of genes associated with cancer immune evasion, including AXL and EPHA2, which impede T‐cell recruitment, and BST2, associated with M2 polarization. Furthermore, T cells showed increased expression of genes related to immunosuppression, and macrophages exhibited increased expression of genes associated with the M2 phenotype. Conclusions: The interaction between melanomas arising from CMN and TIICs may be important for tumour progression and metastasis. [ABSTRACT FROM AUTHOR]

Background: Efforts to profile atopic dermatitis (AD) tissues have intensified, yet comprehensive analysis of systemic immune landscapes in severe AD remains crucial. Methods: Employing single‐cell RNA sequencing, we analyzed over 300,000 peripheral blood mononuclear cells from 12 severe AD patients (Eczema area and severity index (EASI) > 21) and six healthy controls. Results: Results revealed significant immune cell shifts in AD patients, including increased Th2 cell abundance, reduced NK cell clusters with compromised cytotoxicity, and correlated Type 2 innate lymphoid cell proportions with disease severity. Moreover, unique monocyte clusters reflecting activated innate immunity emerged in very severe AD (EASI > 30). While overall dendritic cells (DCs) counts decreased, a distinct Th2‐priming subset termed "Th2_DC" correlated strongly with disease severity, validated across skin tissue data, and flow cytometry with additional independent severe AD samples. Beyond the recognized role of Th2 adaptive immunity, our findings highlight significant innate immune cell alterations in severe AD, implicating their roles in disease pathogenesis and therapeutic potentials. Conclusion: Apart from the widely recognized role of Th2 adaptive immunity in AD pathogenesis, alterations in innate immune cells and impaired cytotoxic cells have also been observed in severe AD. The impact of these alterations on disease pathogenesis and the effectiveness of potential therapeutic targets requires further investigation. [ABSTRACT FROM AUTHOR]

Porcine islet xenotransplantation has been highlighted as an alternative to allo islet transplantation. Despite the remarkable progress that has been made in porcine‐islet pre‐clinical studies in nonhuman primates, immunological tolerance to porcine islets has not been achieved to date. Therefore, allo islet transplantation could be required after the failure of porcine islet xenotransplantation. Here, we report the long‐term control of diabetes by allogeneic pancreatic islet transplantation in diabetic rhesus monkeys that rejected previously transplanted porcine islets. Four diabetic male rhesus monkeys received the porcine islets and then allo islets (5700—19 000 IEQ/kg) were re‐transplanted for a short or long period after the first xeno islet rejection. The recipient monkeys were treated with an immunosuppressive regimen consisting of ATG, humira, and anakinra for induction, and sirolimus and tofacitinib for maintenance therapy. The graft survival days of allo islets in these monkeys were >440, 395, >273, and 127, respectively, similar to that in allo islet transplanted cynomolgus monkeys that received the same immunosuppressive regimen without xeno sensitization. Taken together, it is likely that prior islet xenotransplantation does not affect the survival of subsequent allo islets under clinically applicable immunosuppressants. [ABSTRACT FROM AUTHOR]

Here, we present a case of a 53-year-old female patient with chronic neck pain and systemic inflammation who was ultimately diagnosed with systemic lupus erythematosus. Notably, applying fat-suppressed T2-weighted MRI sequences was pivotal in detecting structural fascial changes commonly associated with systemic inflammatory diseases. PET-CT scans further revealed systemic inflammation around multiple joints, providing valuable insights into MRI signal alterations. This case underscores the importance of considering systemic autoimmune pathology as a potential underlying cause of chronic musculoskeletal pain. It also highlights the clinical utility of MRI with fat suppression sequences in identifying inflammation-related fascial changes. This case emphasizes the significance of a comprehensive evaluation, particularly in situations where clinical features overlap between autoimmune and degenerative skeletal pathologies. Fat-suppressed MRI can provide information about fascial pathology related to systemic inflammatory diseases. In this context, it is worth noting that PET-CT and fat suppression MRI complement each other by providing complementary information about inflammation and the underlying causes of a patient's pain. [ABSTRACT FROM AUTHOR]

Adult T-cell Leukemia/Lymphoma (ATLL) is a rare aggressive T-cell malignancy caused by human T-cell leukemia virus type 1 (HTLV-1) infection. However, little is known about the underlying activated molecular pathways at the single cell level. Moreover, the intercellular communications between the tumor microenvironment (TME) and tumor cells in this malignancy are currently unknown. Difficulties in harvesting fresh tissue in a clinical setting have hampered our deeper understanding of this malignancy. Herein, we examined ATLL using archived fresh frozen tissue after biopsy using single-cell RNA sequencing (scRNA-seq) with T-cell receptor (TCR) clonal analysis. Highly clonal tumor cells showed multiple activating pathways, suggesting dynamic evolution of the malignancy. By dissecting diverse cell types comprising the TME, we identified a novel subset of cancer-associated fibroblast, which showed enriched epidermal growth factor receptor (EGFR)-related transcripts including early growth response 1 and 2 (EGR1 and EGR2). Cancer associated fibroblasts (CAFs) of ATLL play an important role for CD4 T-cell proliferation via FGF7-FGF1 and PDGFA-PDGFRA/B signaling, and CAFs, particularly EGR-enriched, are also associated with CD8 and NKT expansion by EGFR. These findings suggest a potential targeted therapeutic pathway to better treat this neoplasm. [ABSTRACT FROM AUTHOR]

Background: Flow cytometry is a well‐accepted approach for immune profiling; however, its value is restricted by the limited number of markers that can be analyzed simultaneously. Mass cytometry/CyTOF offers broad‐scale immune characterization integrating large number of parameters. While partial blood phenotyping was reported in atopic dermatitis (AD), patients' comprehensive profiling, critical for leveraging new targeted treatments, is not available. IL‐21 may be involved in inflammatory skin diseases but its role in AD is not well established. Methods: We studied T‐cell polarization in the blood of 20 moderate‐to‐severe AD and 15 controls. Using CyTOF and an unsupervised analysis, we measured the frequencies and mean metal intensities of activated polar CD4+/CD8+ T‐cell subsets. Immunohistochemistry, immunofluorescence, and qRT‐PCR were used to analyze skin samples. Results: Examining 24 surface, intracellular markers, and transcription factors, we identified six CD4+ and five CD8+ T‐cell metaclusters. A CD4+ skin‐homing IL‐13+monocytokine and a novel IL‐13+IL‐21+ multicytokine metaclusters were increased in AD vs. controls (p <.01). While IL‐13 signature characterized both clusters, levels were significantly higher in the IL‐21+ group. Both clusters correlated with AD severity (r = 0.49, p =.029). Manual gating corroborated these results and identified additional multicytokine subsets in AD. Immunohistochemistry and immunofluorescence, validated by mRNA expression, displayed significantly increasedIL‐21 counts and colocalization with IL‐13/IL‐4R in AD skin. Conclusion: A multicytokine signature characterizes moderate‐to‐severe AD, possibly explaining partial therapeutic responses to one cytokine targeting, particularly in severe patients. Prominent IL‐21 signature in blood and skin hints for a potential pathogenic role of IL‐21 in AD. [ABSTRACT FROM AUTHOR]

Programmed death 1 (PD‐1)/programmed death ligand 1 (PD‐L1) inhibitors have demonstrated their efficacy in the treatment of various malignancies. Despite their benefits, their immunomodulatory activities can cause unpredictable cutaneous adverse events (CAE). This study aimed to identify characteristics of CAE in patients treated with PD‐1/PD‐L1 inhibitors through the medical records, photographs, and pathology reports. Fifty CAE occurred in 47 (2.75%) of 1711 patients treated with PD‐1/PD‐L1 inhibitors. Pruritic, psoriasiform, urticarial, and acneiform eruptions were the four most common types. Melanoma patients showed CAE more frequently than other malignancies. Acneiform eruption occurred more often at ages under 60 years. Urticarial eruption appeared earlier, while keratoacanthoma appeared later after immunotherapy. The overall survival times were not significantly different between the two groups with and without CAE by Kaplan–Meier analysis (p = 0.055). Studies on CAE may provide more information to understand these drugs and to help manage the patients. [ABSTRACT FROM AUTHOR]

Background: Acneiform eruption is the most common cutaneous adverse event associated with cetuximab. As it can affect quality of life and adversely affect chemotherapy schedule, additional medical care is required. Objectives: To investigate the adherence to and the duration of antibiotic administration to treat cetuximab-induced acneiform eruption. Methods: Medical data of patients who were referred to the Department of Dermatology were reviewed from January 2013 to June 2018. Dermatologists assessed the severity of acneiform eruption and prescribed tetracycline-class antibiotics according to the severity every 2 or 4 weeks. We investigated the duration and amount of oral antibiotic administration and analyzed the factors that may affect the control of acneiform eruption statistically. Results: A total of 207 of 267 patients referred to the Department of Dermatology showed acneiform eruption; 124 patients were treated with minocycline, 34 patients with doxycycline, 27 patients with both, and 22 patients with topical agents. The mean duration of oral antibiotic medication was 82.7 days. A statistical analysis of the factors that prolonged the use of antibiotics for more than 90 days showed that male and younger age were risk factors. Shorter time interval from starting cetuximab to starting antibiotics was associated with longer duration of antibiotic use, statistically. Conclusions: Cetuximab-induced acneiform eruption can be well controlled with tetracycline-class antibiotics in about 3 months. It can last longer in male and younger patients. The sooner and the more severe it appears, the longer it can last. [ABSTRACT FROM AUTHOR]

Background: Although pancreatic islet transplantation is becoming an effective therapeutic option for patients with type 1 diabetes (T1D) who suffer from a substantially impaired awareness of hypoglycemia, its application is limited due to the lack of donors. Thus, pig‐to‐human islet xenotransplantation has been regarded as a promising alternative due to the unlimited number of "donor organs." Long‐term xenogeneic islet graft survival in pig‐to‐non‐human primate (NHP) models has mainly been achieved by administering the anti‐CD154 mAb‐based immunosuppressant regimen. Since the anti‐CD154 mAb treatment has been associated with unexpected fatal thromboembolic complications in clinical trials, the establishment of a new immunosuppressant regimen that is able to be directly applied in clinical trials is an urgent need. Methods: We assessed an immunosuppressant regimen composed of clinically available agents at porcine islet transplantation in consecutive diabetic NHPs. Results: Porcine islet graft survival in consecutive diabetic NHPs (n = 7; >222, >200, 181, 89, 62, 55, and 34 days) without severe adverse events. Conclusion: We believe that our study could contribute greatly to the initiation of islet xenotransplantation clinical trials. [ABSTRACT FROM AUTHOR]

Background: Xenogeneic islet transplantation using porcine pancreata has been a promising option for substituting human islet transplantation. Moreover, recent advances in pre‐clinical results have put islet xenotransplantation closer to the possibility of clinical application. While preparing for the era of clinical xenotransplantation, developing non‐invasive immune monitoring method which could predict the graft fate could benefit the patient. However, there are few reports showing predictive immune parameters associated with the fate of the graft in islet xenotransplantation. Methods: The absolute number and ratio of T‐cell subsets have been measured via flow cytometry from the peripheral blood of 16 rhesus monkeys before and after porcine islet xenotransplantation. The correlation between the graft survival and the absolute number or ratio of T cells was retrospectively analyzed. Results: The ratio of CD4+ versus CD8+ T cells was significantly reduced due to CD8+ effector memory cells' increase. Correlation analyses revealed that CD4+/CD8+, CD4+/CD8+ naïve, CD4+ naïve/CD8+ naïve, and CD4+ central memory/CD8+ naïve cell ratios negatively correlated with the duration of graft survival. Conversely, further analyses discovered strong, positive correlation of CD4+/CD8+ cell ratios within the early graft‐rejected monkeys (≤60 days). Conclusions: This retrospective study demonstrated that CD4+/CD8+ ratios correlated with graft survival, especially in recipients which rejected the graft in early post‐transplantation periods. CD4+/CD8+ ratios could be used as a surrogate marker to predict the graft fate in pig‐to‐NHP islet xenotransplantation. [ABSTRACT FROM AUTHOR]

Anti-CD154 blockade-based regimens remain unequaled in prolonging graft survival in various organ transplantation models. Several studies have focused on transplantation tolerance with the anti-CD154 blockade, but none of these studies has investigated the mechanisms associated with its use as the sole treatment in animal models, delaying our understanding of anti-CD154 blockade-mediated immune tolerance. The purpose of this study was to investigate the mechanism underlying the anti-CD154 monoclonal antibody (mAb) blockade in inducing immune tolerance using an intrahepatic murine allogeneic islet transplantation model. Allogeneic BALB/c AnHsd (BALB/c) islets were infused into the liver of diabetic C57BL/6 (B6) mice via the cecal vein. Anti-CD154 mAb (MR1) was administered on −1, 0, 1, 3, 5, and 7 d posttransplantation at 0.5 mg per mouse. We showed that short-term MR1 monotherapy could prolong the allogeneic islet grafts to more than 250 d in the murine intrahepatic islet transplantation model. The second islet grafts transplanted under the kidney capsule of the recipients were protected from rejection. We also found that rejection of same-donor skin grafts transplanted to the tolerant mice was modestly delayed. Using a DEREG mouse model, FoxP3+ regulatory T (Treg) cells were shown to play important roles in transplantation tolerance. In mixed lymphocyte reactions, Treg cells from the tolerant mice showed more potency in suppressing BALB/c splenocyte-stimulated Teff cell proliferation than those from naïve mice. In this study, we demonstrated for the first time that a short-term anti-CD154 mAb single treatment could induce FoxP3+ Treg cell-mediated immune tolerance in the intrahepatic murine allogeneic islet transplantation model. [ABSTRACT FROM AUTHOR]

Clinical islet transplantation has recently been a promising treatment option for intractable type 1 diabetes patients. Although early graft loss has been well studied and controlled, the mechanisms of late graft loss largely remains obscure. Since long-term islet graft survival had not been achieved in islet xenotransplantation, it has been impossible to explore the mechanism of late islet graft loss. Fortunately, recent advances where consistent long-term survival (≥6 months) of adult porcine islet grafts was achieved in five independent, diabetic nonhuman primates (NHPs) enabled us to investigate on the late graft loss. Regardless of the conventional immune monitoring methods applied in the post-transplant period, the initiation of late graft loss could rarely be detected before the overt graft loss observed via uncontrolled blood glucose level. Thus, we retrospectively analyzed the gene expression profiles in 2 rhesus monkey recipients using peripheral blood RNA-sequencing (RNA-seq) data to find out the potential cause(s) of late graft loss. Bioinformatic analyses showed that highly relevant immunological pathways were activated in the animal which experienced late graft failure. Further connectivity analyses revealed that the activation of T cell signaling pathways was the most prominent, suggesting that T cell-mediated graft rejection could be the cause of the late-phase islet loss. Indeed, the porcine islets in the biopsied monkey liver samples were heavily infiltrated with CD3+ T cells. Furthermore, hypothesis test using a computational experiment reinforced our conclusion. Taken together, we suggest that bioinformatics analyses with peripheral blood RNA-seq could unveil the cause of insidious late islet graft loss. [ABSTRACT FROM AUTHOR]