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The immunopathogenesis of psoriasis, a common chronic inflammatory disease of the skin, is incompletely understood. Here we demonstrate, using a combination of single cell and spatial RNA sequencing, IL-36 dependent amplification of IL-17A and TNF inflammatory responses in the absence of neutrophil proteases, which primarily occur within the supraspinous layer of the psoriatic epidermis. We further show that a subset of SFRP2 + fibroblasts in psoriasis contribute to amplification of the immune network through transition to a pro-inflammatory state. The SFRP2 + fibroblast communication network involves production of CCL13 , CCL19 and CXCL12 , connected by ligand-receptor interactions to other spatially proximate cell types: CCR2 + myeloid cells, CCR7 + LAMP3 + dendritic cells, and CXCR4 expressed on both CD8 + Tc17 cells and keratinocytes, respectively. The SFRP2 + fibroblasts also express cathepsin S, further amplifying inflammatory responses by activating IL-36G in keratinocytes. These data provide an in-depth view of psoriasis pathogenesis, which expands our understanding of the critical cellular participants to include inflammatory fibroblasts and their cellular interactions. Changes in Psoriasis and other inflammatory skin diseases during severity stages can be investigated using single cell and spatial transcriptomics. Here the authors compare different inflammatory skin diseases to emphasise differences in immune cells and inflammatory markers particularly keratinocytes and fibroblasts.

The role of the vasculature in inflammatory skin disorders is an exciting area of investigation. Vascular endothelial cells (ECs) play instrumental roles in maintaining the vascular barrier and control of blood flow. Furthermore, ECs contribute to a variety of immune responses, such as targeting immune cells to specific areas of vascular damage, infection, or foreign material. However, mechanisms through which ECs participate in immune-mediated responses remain to be fully explored. In this issue of the JCI, Li, Shao, et al. report on vascular endothelial glycocalyx destruction and the mechanisms through which EC dysfunction contributes to the well-characterized immune-mediated features of psoriasis, a chronic inflammatory skin disease. Here, we discuss the implications of these findings and highlight some risks and benefits of existing therapies designed to target immune cell trafficking in a variety of inflammatory conditions.

Systemic sclerosis (SSc) is a devastating autoimmune disease characterized by excessive production and accumulation of extracellular matrix, leading to fibrosis of skin and other internal organs. However, the main cellular participants in SSc skin fibrosis remain incompletely understood. Here using differentiation trajectories at a single cell level, we demonstrate a dual source of extracellular matrix deposition in SSc skin from both myofibroblasts and endothelial-to-mesenchymal-transitioning cells (EndoMT). We further define a central role of Hippo pathway effectors in differentiation and homeostasis of myofibroblast and EndoMT, respectively, and show that myofibroblasts and EndoMTs function as central communication hubs that drive key pro-fibrotic signaling pathways in SSc. Together, our data help characterize myofibroblast differentiation and EndoMT phenotypes in SSc skin, and hint that modulation of the Hippo pathway may contribute in reversing the pro-fibrotic phenotypes in myofibroblasts and EndoMTs. Systemic sclerosis (SSc) is an autoimmune disease causing skin fibrosis and organ inflammation. Here the authors generate and analyze SSc skin single cell RNA sequencing data to propose contributions from both myofibroblasts and endothelial-to-mesenchymal -transitioning cells (EndoMT) to skin fibrosis, and to implicate the involvement of Hippo signaling pathways.

Hidradenitis suppurativa (HS) is a chronic inflammatory disease characterized by abscesses, nodules, dissecting/draining tunnels, and extensive fibrosis. Here, we integrate single-cell RNA sequencing, spatial transcriptomics, and immunostaining to provide an unprecedented view of the pathogenesis of chronic HS, characterizing the main cellular players and defining their interactions. We found a striking layering of the chronic HS infiltrate and identified the contribution of 2 fibroblast subtypes (SFRP4+ and CXCL13+) in orchestrating this compartmentalized immune response. We further demonstrated the central role of the Hippo pathway in promoting extensive fibrosis in HS and provided preclinical evidence that the profibrotic fibroblast response in HS can be modulated through inhibition of this pathway. These data provide insights into key aspects of HS pathogenesis with broad therapeutic implications.

Granulomas are complex cellular structures composed predominantly of macrophages and lymphocytes that function to contain and kill invading pathogens. Here, we investigated the single-cell phenotypes associated with antimicrobial responses in human leprosy granulomas by applying single-cell and spatial sequencing to leprosy biopsy specimens. We focused on reversal reactions (RRs), a dynamic process whereby some patients with disseminated lepromatous leprosy (L-lep) transition toward self-limiting tuberculoid leprosy (T-lep), mounting effective antimicrobial responses. We identified a set of genes encoding proteins involved in antimicrobial responses that are differentially expressed in RR versus L-lep lesions and regulated by interferon-γ and interleukin-1β. By integrating the spatial coordinates of the key cell types and antimicrobial gene expression in RR and T-lep lesions, we constructed a map revealing the organized architecture of granulomas depicting compositional and functional layers by which macrophages, T cells, keratinocytes and fibroblasts can each contribute to the antimicrobial response. Modlin and colleagues examined the skin lesions of human leprosy patients using single-cell RNA sequencing coupled to cellular spatial mapping. Their analysis maps the architecture of granulomas in leprosy lesions from patients with leprosy with localized disease (tuberculoid leprosy, reversal reaction) to those with progressive infection (lepromatous leprosy).

Generalized pustular psoriasis (GPP) is a severe subtype of psoriasis characterized by epidermal neutrophil infiltration, often presenting as acute, potentially life-threatening flares. However, the characterization of the immune micro-environment in GPP lesions remains largely unknown. Here, we use single-cell RNA profiling to interrogate the transcriptomes of 60,000 single cells from GPP lesional skin ( n  = 13) and healthy adult skin ( n  = 4), combined with spatial transcriptomics. We identify a neutrophil subset lacking CASP8 expression but exhibiting elevated levels of inflammatory pathway genes, including RIPK1 , NFKB1 , IL1B , CXCL1 , and CXCL8 in GPP flares, illustrating neutrophil transition from pre-inflammatory to a pro-inflammatory state, and activation of a communication network between IL36G + keratinocytes and neutrophils in GPP lesions, with TNFSF15 (TL1A) released from neutrophils exaggerating the inflammatory crosstalk. We further demonstrate that fibroblasts and capillary endothelial cells function as central communication hubs in GPP, through dynamic receptor-ligand interactions with several spatially proximate immune cells, including T cells, neutrophils, and macrophages. In this work, we provide an in-depth view of immune cell participation and highlight the role of neutrophil-keratinocyte crosstalk in GPP pathogenesis. Generalised pustular psoriasis (GPP) is a severe type of psoriasis characterized by epidermal neutrophil infiltration and potentially life-threatening flares. Here the authors use single cell and spatial transcriptomic analysis of skin samples implicating contact between IL36G+ keratinocytes and neutrophils as well as ligand-receptor interactions of fibroblasts with T cells, neutrophils or macrophages.

CRISPR/Cas9 has been proposed as a treatment for genetically inherited skin disorders. Here we report that CRISPR transfection activates STING-dependent antiviral responses in keratinocytes, resulting in heightened endogenous interferon (IFN) responses through induction of IFN-κ, leading to decreased plasmid stability secondary to induction of the cytidine deaminase gene APOBEC3G. Notably, CRISPR-generated KO keratinocytes had permanent suppression of IFN-κ and IFN-stimulated gene (ISG) expression, secondary to hypermethylation of the IFNK promoter region by the DNA methyltransferase DNMT3B. JAK inhibition via baricitinib prior to CRISPR transfection increased transfection efficiency, prevented IFNK promoter hypermethylation, and restored normal IFN-κ activity and ISG responses. This work shows that CRISPR-mediated gene correction alters antiviral responses in keratinocytes, has implications for future gene therapies for inherited skin diseases using CRISPR technology, and suggests pharmacologic JAK inhibition as a tool for facilitating and attenuating inadvertent selection effects in CRISPR/Cas9 therapeutic approaches.

BACKGROUNDCutaneous lichen planus (LP) is a recalcitrant, difficult-to-treat, inflammatory skin disease characterized by pruritic, flat-topped, violaceous papules on the skin. Baricitinib is an oral Janus kinase (JAK) 1/2 inhibitor that interrupts the signaling pathway of IFN-γ, a cytokine implicated in the pathogenesis of LP.METHODSIn this phase II trial, 12 patients with cutaneous LP received 2 mg daily baricitinib for 16 weeks, accompanied by in-depth spatial, single-cell, and bulk transcriptomic profiling of pre- and posttreatment samples.RESULTSAn early and sustained clinical response was seen, with 83.3% of patients responsive at week 16. Our molecular data identified a unique, oligoclonal IFN-γ, CD8+, and CXCL13+ cytotoxic T cell population in LP skin and demonstrated a rapid decrease in IFN signature within 2 weeks of treatment, most prominently in the basal layer of the epidermis.CONCLUSIONThis study demonstrates the efficacy and molecular mechanisms of JAK inhibition in LP.TRIAL REGISTRATIONNCT05188521FUNDINGEli Lilly, Appignani Benefactor Funds, 5P30AR075043, Mayo Clinic Clinical Trials Stimulus Funds.

Background The pathogenesis of preeclampsia superimposed on chronic hypertension (SI) is poorly understood relative to preeclampsia (PreE) occurring in pregnant people without chronic hypertension. Placental transcriptomes in pregnancies complicated by PreE and SI have not been previously compared. Methods We identified pregnant people in the University of Michigan Biorepository for Understanding Maternal and Pediatric Health with hypertensive disorders affecting singleton, euploid gestations (N = 36) along with non-hypertensive control subjects (N = 12). Subjects were grouped as: (1) normotensive (N = 12), (2) chronic hypertensive (N = 13), (3) preterm PreE with severe features (N = 5), (4) term PreE with severe features (N = 11), (5) preterm SI (N = 3), or (6) term SI (N = 4). Bulk RNA sequencing of paraffin-embedded placental tissue was performed. The primary analysis assessed differential gene expression relative to normotensive and chronic hypertensive placentas, where Wald adjusted P values < 0.05 were considered significant. Unsupervised clustering analyses and correlation analyses were performed between conditions of interest, and a gene ontology was constructed. Results Comparing samples from pregnant people with hypertensive diseases to non-hypertensive controls, there were 2290 differentially expressed genes. The log2-fold changes in genes differentially expressed in chronic hypertension correlated better with term (R = 0.59) and preterm (R = 0.63) PreE with severe features than with term (R = 0.21) and preterm (R = 0.22) SI. A relatively poor correlation was observed between preterm SI and preterm PreE with severe features (0.20) as well as term SI and term PreE with severe features (0.31). The majority of significant genes were downregulated in term and preterm SI versus normotensive controls (92.1%, N = 128). Conversely, most term and preterm PreE with severe features genes were upregulated compared to the normotensive group (91.8%, N = 97). Many of the upregulated genes in PreE with the lowest adjusted P values are known markers of abnormal placentation (e.g., PAAPA, KISS1, CLIC3), while the downregulated genes with the greatest adjusted P values in SI have fewer known pregnancy-specific functions. Conclusions We identified unique placental transcriptional profiles in clinically relevant subgroups of individuals with hypertension in pregnancy. Preeclampsia superimposed on chronic hypertension was molecularly distinct from preeclampsia in individuals without chronic hypertension, and chronic hypertension without preeclampsia, suggesting that preeclampsia superimposed on hypertension may represent a distinct entity.

Most anti-angiogenic preeclampsia models in rodents utilized the overexpression of a truncated soluble fms-like tyrosine kinase-1 (sFlt-1) not expressed in any species. Other limitations of mouse preeclampsia models included stressful blood pressure measurements and the lack of postpartum monitoring. We aimed to 1) develop a mouse model of preeclampsia by administering the most abundant human placental sFlt-1 isoform (hsFlt-1-e15a) in preeclampsia; 2) determine blood pressures in non-stressed conditions; and 3) develop a survival surgery that enables the collection of fetuses and placentas and postpartum (PP) monitoring. Pregnancy status of CD-1 mice was evaluated with high-frequency ultrasound on gestational days (GD) 6 and 7. Telemetry catheters were implanted in the carotid artery on GD7, and their positions were verified by ultrasound on GD13. Mice were injected through tail-vein with adenoviruses expressing hsFlt-1-e15a (n = 11) or green fluorescent protein (GFP; n = 9) on GD8/GD11. Placentas and pups were delivered by cesarean section on GD18 allowing PP monitoring. Urine samples were collected with cystocentesis on GD6/GD7, GD13, GD18, and PPD8, and albumin/creatinine ratios were determined. GFP and hsFlt-1-e15a expression profiles were determined by qRT-PCR. Aortic ring assays were performed to assess the effect of hsFlt-1-e15a on endothelia. Ultrasound predicted pregnancy on GD7 in 97% of cases. Cesarean section survival rate was 100%. Mean arterial blood pressure was higher in hsFlt-1-e15a-treated than in GFP-treated mice (∆MAP = 13.2 mmHg, p = 0.00107; GD18). Focal glomerular changes were found in hsFlt-1-e15a -treated mice, which had higher urine albumin/creatinine ratios than controls (109.3 ± 51.7 μg/mg vs. 19.3 ± 5.6 μg/mg, p = 4.4 x 10(-2); GD18). Aortic ring assays showed a 46% lesser microvessel outgrowth in hsFlt-1-e15a-treated than in GFP-treated mice (p = 1.2 x 10(-2)). Placental and fetal weights did not differ between the groups. One mouse with liver disease developed early-onset preeclampsia-like symptoms with intrauterine growth restriction (IUGR). A mouse model of late-onset preeclampsia was developed with the overexpression of hsFlt-1-e15a, verifying the in vivo pathologic effects of this primate-specific, predominant placental sFlt-1 isoform. HsFlt-1-e15a induced early-onset preeclampsia-like symptoms associated with IUGR in a mouse with a liver disease. Our findings support that hsFlt-1-e15a is central to the terminal pathway of preeclampsia, and it can induce the full spectrum of symptoms in this obstetrical syndrome.