ABSTRACT
Preferentially Expressed Antigen in Melanoma (PRAME) and Ten-Eleven Translocation (TET) dioxygenase-mediated 5-hydroxymethylcytosine (5hmC) are emerging melanoma biomarkers. We observed an inverse correlation between PRAME expression and 5hmC levels in benign nevi, melanoma in situ, primary invasive melanoma, and metastatic melanomas via immunohistochemistry and multiplex immunofluorescence: nevi exhibited high 5hmC and low PRAME, whereas melanomas showed the opposite pattern. Single-cell multiplex imaging of melanoma precursors revealed that diminished 5hmC coincides with PRAME upregulation in premalignant cells. Analysis of TCGA and GTEx databases confirmed a negative relationship between TET2 and PRAME mRNA expression in melanoma. Additionally, 5hmC levels were reduced at the PRAME 5' promoter in melanoma compared to nevi, suggesting a role for 5hmC in PRAME transcription. Restoring 5hmC levels via TET2 overexpression notably reduced PRAME expression in melanoma cell lines. These findings establish a function of TET2-mediated DNA hydroxymethylation in regulating PRAME expression and demonstrate epigenetic reprogramming as pivotal in melanoma tumorigenesis. Teaser: Melanoma biomarker PRAME expression is negatively regulated epigenetically by TET2-mediated DNA hydroxymethylation.
ABSTRACT
Macrophages are critical for maintenance and repair of mucosal tissues. While functionally distinct subtypes of macrophage are known to have important roles in injury response and repair in the lungs, little is known about macrophages in the proximal conducting airways. Single-cell RNA sequencing and flow cytometry demonstrated murine tracheal macrophages are largely monocyte-derived and are phenotypically distinct from lung macrophages at homeostasis. Following sterile airway injury, monocyte-derived macrophages are recruited to the trachea and activate a pro-regenerative phenotype associated with wound healing. Animals lacking the chemokine receptor CCR2 have reduced numbers of circulating monocytes and tracheal macrophages, deficient pro-regenerative macrophage activation and defective epithelial repair. Together, these studies indicate that recruitment and activation of monocyte-derived tracheal macrophages is CCR2-dependent and is required for normal airway epithelial regeneration.
ABSTRACT
Differentiation of stem and progenitor cells is a highly regulated process that involves the coordinated action of multiple layers of regulation. Here we show how the post-transcriptional regulatory layer instructs the level of chromatin regulation via miR-144 and its targets to orchestrate chromatin condensation during erythropoiesis. The loss of miR-144 leads to impaired chromatin condensation during erythrocyte maturation. Among the several targets of miR-144 that influence chromatin organization, the miR-144-dependent regulation of Hmgn2 is conserved from fish to humans. Our genetic probing of the miR-144/Hmgn2 regulatory axis establish that intact miR-144 target sites in the Hmgn2 3'UTR are necessary for the proper maturation of erythrocytes in both zebrafish and human iPSC-derived erythroid cells while loss of Hmgn2 rescues in part the miR-144 null phenotype. Altogether, our results uncover miR-144 and its target Hmgn2 as the backbone of the genetic regulatory circuit that controls the terminal differentiation of erythrocytes in vertebrates.
Subject(s)
Chromatin , Erythropoiesis , MicroRNAs , Zebrafish , MicroRNAs/metabolism , MicroRNAs/genetics , Erythropoiesis/genetics , Zebrafish/genetics , Zebrafish/metabolism , Humans , Animals , Chromatin/metabolism , Chromatin/genetics , Erythrocytes/metabolism , 3' Untranslated Regions/genetics , Induced Pluripotent Stem Cells/metabolism , Induced Pluripotent Stem Cells/cytology , Cell Differentiation/geneticsABSTRACT
Background: Cardiac dysfunction in AL amyloidosis is thought to be partly related to the direct impact of AL LCs on cardiomyocyte function, with the degree of dysfunction at diagnosis as a major determinant of clinical outcomes. Nonetheless, mechanisms underlying LC-induced myocardial toxicity are not well understood. Methods: We identified gene expression changes correlating with human cardiac cells exposed to a cardiomyopathy-associated κAL LC. We then sought to confirm these findings in a clinical dataset by focusing on clinical parameters associated with the pathways dysregulated at the gene expression level. Results: Upon exposure to a cardiomyopathy-associated κAL LC, cardiac cells exhibited gene expression changes related to myocardial contractile function and inflammation, leading us to hypothesize that there could be clinically detectable changes in GLS on echocardiogram and serum inflammatory markers in patients. Thus, we identified 29 patients with normal IVSd but abnormal cardiac biomarkers suggestive of LC-induced cardiac dysfunction. These patients display early cardiac biomarker staging, abnormal GLS, and significantly reduced serum inflammatory markers compared to patients with clinically evident amyloid fibril deposition. Conclusion: Collectively, our findings highlight early molecular and functional signatures of cardiac AL amyloidosis, with potential impact for developing improved patient biomarkers and novel therapeutics.
ABSTRACT
Centenarians provide a unique lens through which to study longevity, healthy aging, and resiliency. Moreover, models of human aging and resilience to disease that allow for the testing of potential interventions are virtually non-existent. We obtained and characterized over 50 centenarian and offspring peripheral blood samples including those connected to functional independence data highlighting resistance to disability and cognitive impairment. Targeted methylation arrays were used in molecular aging clocks to compare and contrast differences between biological and chronological age in these specialized subjects. Isolated peripheral blood mononuclear cells (PBMCs) were then successfully reprogrammed into high-quality induced pluripotent stem cell (iPSC) lines which were functionally characterized for pluripotency, genomic stability, and the ability to undergo directed differentiation. The result of this work is a one-of-a-kind resource for studies of human longevity and resilience that can fuel the discovery and validation of novel therapeutics for aging-related disease.
ABSTRACT
Radial dysplasia (RD) is a congenital upper limb birth defect that presents with changes to the upper limb anatomy, including a shortened or absent radius, bowed ulna, thumb malformations, a radially deviated hand and a range of muscle and tendon malformations, including absent or abnormally shaped muscle bundles. Current treatments to address wrist instability caused by a shortened or absent radius frequently require an initial soft tissue distraction intervention followed by a wrist stabilisation procedure. Following these surgical interventions, however, recurrence of the wrist deviation remains a common, long-term problem following treatment. The impact of the abnormal soft connective tissue (muscle and tendon) anatomy on the clinical presentation of RD and the complications following surgery are not understood. To address this, we have examined the muscle, fascia and the fascial irregular connective tissue (ICT) fibroblasts found within soft connective tissues, from RD patients. We show that ICT fibroblasts isolated from RD patients are functionally abnormal when compared to the same cells isolated from control patients and secrete a relatively disordered extracellular matrix (ECM). Furthermore, we show that ICT fibroblast dysfunction is a unifying feature found in RD patients, even when the RD clinical presentation is caused by distinct genetic syndromes.
Subject(s)
Connective Tissue , Fibroblasts , Muscle, Skeletal , Humans , Fibroblasts/pathology , Connective Tissue/pathology , Muscle, Skeletal/abnormalities , Muscle, Skeletal/pathology , Male , Female , Radius/abnormalities , Radius/pathologyABSTRACT
BACKGROUND: Apart from the skin, little is known about the immunological processes in deeper tissues, which are typically not accessible to biopsy and inspection, of vascularized composite allografts (VCAs). Face transplant patients develop prominent adenopathy shortly after transplantation that resolves over time. The mechanisms underlying this process are not understood. MATERIALS AND METHODS: A retrospective cohort study was conducted on 9 patients who underwent 10 facial VCAs at the Brigham and Women's Hospital, Boston, MA, between April 2009 and July 2019. Clinical, radiological, and histological data related to lymphadenopathy of the head and neck were reviewed. RESULTS: Patients who received donor-derived lymph nodes (LNs) developed bilateral lymphadenopathy of the submental or submandibular superficial LNs. Median time of presentation was POD18 (range POD6-POM3). Notably, bilateral adenopathy of the neck was not observed in later stages of follow-up (mean follow-up, 115 months). Histology of 3 LNs showed increased histiocytes and apoptosis, with the features reminiscent of necrotizing histiocytic lymphadenitis, and B and T lymphocytes (mostly CD8 + T) admixed with CD163 + histiocytes and dendritic cells. Molecular chimerism analysis in one case showed the coexistence of donor (81%) and recipient (19%) derived lymphocytes. Granzyme B (GZMB) expression confirmed the presence of increased cytotoxic T cells in this LN sample. CONCLUSION: Our data suggested the involvement of an immunological process within the donor-derived LNs after facial allotransplantation between the recipient and donor cells. GZMB expression suggested LN rejection that can occurred independently of skin rejection. This finding supports the need to better define the role of donor-derived immune cells in the context of allograft rejection.
Subject(s)
Composite Tissue Allografts , Lymphadenopathy , Vascularized Composite Allotransplantation , Humans , Female , Retrospective Studies , Graft Survival , Graft Rejection , Vascularized Composite Allotransplantation/adverse effects , Lymph Nodes , Lymphadenopathy/pathologySubject(s)
Immune Checkpoint Inhibitors , Humans , Pilot Projects , Immune Checkpoint Inhibitors/adverse effects , Female , Male , Fluorescent Antibody Technique , Skin/immunology , Skin/pathology , Skin/drug effects , Middle Aged , Aged , Drug Eruptions/etiology , Drug Eruptions/immunology , Drug Eruptions/diagnosisABSTRACT
Merkel cell carcinoma (MCC) is a rare and aggressive skin cancer. Inhibitors targeting the programmed cell death 1 (PD-1) immune checkpoint have improved MCC patient outcomes by boosting antitumor T cell immunity. Here, we identify PD-1 as a growth-promoting receptor intrinsic to MCC cells. In human MCC lines and clinical tumors, RT-PCR-based sequencing, immunoblotting, flow cytometry, and immunofluorescence analyses demonstrated PD-1 gene and protein expression by MCC cells. MCC-PD-1 ligation enhanced, and its inhibition or silencing suppressed, in vitro proliferation and in vivo tumor xenograft growth. Consistently, MCC-PD-1 binding to PD-L1 or PD-L2 induced, while antibody-mediated PD-1 blockade inhibited, protumorigenic mTOR signaling, mitochondrial (mt) respiration, and ROS generation. Last, pharmacologic inhibition of mTOR or mtROS reversed MCC-PD-1:PD-L1-dependent proliferation and synergized with PD-1 checkpoint blockade in suppressing tumorigenesis. Our results identify an MCC-PD-1-mTOR-mtROS axis as a tumor growth-accelerating mechanism, the blockade of which might contribute to clinical response in patients with MCC.
Subject(s)
Carcinoma, Merkel Cell , Skin Neoplasms , Humans , B7-H1 Antigen , Carcinoma, Merkel Cell/drug therapy , Carcinoma, Merkel Cell/genetics , Programmed Cell Death 1 Receptor , Reactive Oxygen Species , Skin Neoplasms/drug therapy , Skin Neoplasms/genetics , TOR Serine-Threonine KinasesABSTRACT
BACKGROUND: Dedicator of cytokinesis 8 (DOCK8)-deficient patients have severe eczema, elevated IgE, and eosinophilia, features of atopic dermatitis (AD). OBJECTIVE: We sought to understand the mechanisms of eczema in DOCK8 deficiency. METHODS: Skin biopsy samples were characterized by histology, immunofluorescence microscopy, and gene expression. Skin barrier function was measured by transepidermal water loss. Allergic skin inflammation was elicited in mice by epicutaneous sensitization with ovalbumin (OVA) or cutaneous application of Staphylococcus aureus. RESULTS: Skin lesions of DOCK8-deficient patients exhibited type 2 inflammation, and the patients' skin was colonized by Saureus, as in AD. Unlike in AD, DOCK8-deficient patients had a reduced FOXP3:CD4 ratio in their skin lesions, and their skin barrier function was intrinsically intact. Dock8-/- mice exhibited reduced numbers of cutaneous T regulatory (Treg) cells and a normal skin barrier. Dock8-/- and mice with an inducible Dock8 deletion in Treg cells exhibited increased allergic skin inflammation after epicutaneous sensitization with OVA. DOCK8 was shown to be important for Treg cell stability at sites of allergic inflammation and for the generation, survival, and suppressive activity of inducible Treg cells. Adoptive transfer of wild-type, but not DOCK8-deficient, OVA-specific, inducible Treg cells suppressed allergic inflammation in OVA-sensitized skin of Dock8-/- mice. These mice developed severe allergic skin inflammation and elevated serum IgE levels after topical exposure to Saureus. Both were attenuated after adoptive transfer of WT but not DOCK8-deficient Treg cells. CONCLUSION: Treg cell dysfunction increases susceptibility to allergic skin inflammation in DOCK8 deficiency and synergizes with cutaneous exposure to Saureus to drive eczema in DOCK8 deficiency.
Subject(s)
Eczema , Guanine Nucleotide Exchange Factors , Mice, Knockout , Skin , Staphylococcus aureus , T-Lymphocytes, Regulatory , Animals , T-Lymphocytes, Regulatory/immunology , Guanine Nucleotide Exchange Factors/deficiency , Guanine Nucleotide Exchange Factors/genetics , Guanine Nucleotide Exchange Factors/immunology , Eczema/immunology , Staphylococcus aureus/immunology , Humans , Mice , Skin/immunology , Skin/pathology , Female , Male , Mice, Inbred C57BL , Dermatitis, Atopic/immunologyABSTRACT
Tissue homeostasis and the emergence of disease are controlled by changes in the proportions of resident and recruited cells, their organization into cellular neighbourhoods, and their interactions with acellular tissue components. Highly multiplexed tissue profiling (spatial omics) 1 makes it possible to study this microenvironment in situ , usually in 4-5 micron thick sections (the standard histopathology format) 2 . Microscopy-based tissue profiling is commonly performed at a resolution sufficient to determine cell types but not to detect subtle morphological features associated with cytoskeletal reorganisation, juxtracrine signalling, or membrane trafficking 3 . Here we describe a high-resolution 3D imaging approach able to characterize a wide variety of organelles and structures at sub-micron scale while simultaneously quantifying millimetre-scale spatial features. This approach combines cyclic immunofluorescence (CyCIF) imaging 4 of over 50 markers with confocal microscopy of archival human tissue thick enough (30-40 microns) to fully encompass two or more layers of intact cells. 3D imaging of entire cell volumes substantially improves the accuracy of cell phenotyping and allows cell proximity to be scored using plasma membrane apposition, not just nuclear position. In pre-invasive melanoma in situ 5 , precise phenotyping shows that adjacent melanocytic cells are plastic in state and participate in tightly localised niches of interferon signalling near sites of initial invasion into the underlying dermis. In this and metastatic melanoma, mature and precursor T cells engage in an unexpectedly diverse array of juxtracrine and membrane-membrane interactions as well as looser "neighbourhood" associations 6 whose morphologies reveal functional states. These data provide new insight into the transitions occurring during early tumour formation and immunoediting and demonstrate the potential for phenotyping of tissues at a level of detail previously restricted to cultured cells and organoids.
ABSTRACT
Differentiation of stem and progenitor cells is a highly regulated process that involves the coordinated action of multiple layers of regulation. Here we show how the post-transcriptional regulatory layer instructs the level of chromatin regulation via miR-144 and its targets to orchestrate chromatin condensation during erythropoiesis. The loss of miR-144 leads to impaired chromatin condensation during erythrocyte maturation. Among the several targets of miR-144 that influence chromatin organization, the miR-144-dependent regulation of Hmgn2 is conserved from fish to humans. Our genetic probing of the miR-144/Hmgn2 regulatory axis established that intact miR-144 target sites in the Hmgn2 3'UTR are necessary for the proper maturation of erythrocytes in both zebrafish and human iPSC-derived erythroid cells while loss of Hmgn2 rescues in part the miR-144 null phenotype. Altogether, our results uncover miR-144 and its target Hmgn2 as the backbone of the genetic regulatory circuit that controls the terminal differentiation of erythrocytes in vertebrates.
ABSTRACT
BACKGROUND: Oral squamous cell carcinoma (SCC) is associated with oral microbial dysbiosis. In this unique study, we compared pre- to post-treatment salivary microbiome in patients with SCC by 16S rRNA gene sequencing and examined how microbiome changes correlated with the expression of an anti-microbial protein. RESULTS: Treatment of SCC was associated with a reduction in overall bacterial richness and diversity. There were significant changes in the microbial community structure, including a decrease in the abundance of Porphyromonaceae and Prevotellaceae and an increase in Lactobacillaceae. There were also significant changes in the microbial community structure before and after treatment with chemoradiotherapy, but not with surgery alone. In patients treated with chemoradiotherapy alone, several bacterial populations were differentially abundant between responders and non-responders before and after therapy. Microbiome changes were associated with a change in the expression of DMBT1, an anti-microbial protein in human saliva. Additionally, we found that salivary DMBT1, which increases after treatment, could serve as a post-treatment salivary biomarker that links to microbial changes. Specifically, post-treatment increases in human salivary DMBT1 correlated with increased abundance of Gemella spp., Pasteurellaceae spp., Lactobacillus spp., and Oribacterium spp. This is the first longitudinal study to investigate treatment-associated changes (chemoradiotherapy and surgery) in the oral microbiome in patients with SCC along with changes in expression of an anti-microbial protein in saliva. CONCLUSIONS: The composition of the oral microbiota may predict treatment responses; salivary DMBT1 may have a role in modulating the oral microbiome in patients with SCC. After completion of treatment, 6 months after diagnosis, patients had a less diverse and less rich oral microbiome. Leptotrichia was a highly prevalent bacteria genus associated with disease. Expression of DMBT1 was higher after treatment and associated with microbiome changes, the most prominent genus being Gemella Video Abstract.
Subject(s)
Carcinoma, Squamous Cell , Microbiota , Mouth Neoplasms , Humans , Mouth Neoplasms/therapy , Longitudinal Studies , RNA, Ribosomal, 16S/genetics , Microbiota/genetics , Saliva/microbiology , Bacteria/genetics , Calcium-Binding Proteins , DNA-Binding Proteins , Tumor Suppressor ProteinsABSTRACT
Neuroendocrine tumors (NETs) are rare cancers that most often arise in the gastrointestinal tract and pancreas. The fundamental mechanisms driving gastroenteropancreatic (GEP)-NET growth remain incompletely elucidated; however, the heterogeneous clinical behavior of GEP-NETs suggests that both cellular lineage dynamics and tumor microenvironment influence tumor pathophysiology. Here, we investigated the single-cell transcriptomes of tumor and immune cells from patients with gastroenteropancreatic NETs. Malignant GEP-NET cells expressed genes and regulons associated with normal, gastrointestinal endocrine cell differentiation, and fate determination stages. Tumor and lymphoid compartments sparsely expressed immunosuppressive targets commonly investigated in clinical trials, such as the programmed cell death protein-1/programmed death ligand-1 axis. However, infiltrating myeloid cell types within both primary and metastatic GEP-NETs were enriched for genes encoding other immune checkpoints, including VSIR (VISTA), HAVCR2 (TIM3), LGALS9 (Gal-9), and SIGLEC10. Our findings highlight the transcriptomic heterogeneity that distinguishes the cellular landscapes of GEP-NET anatomic subtypes and reveal potential avenues for future precision medicine therapeutics.
Subject(s)
Intestinal Neoplasms , Neuroendocrine Tumors , Pancreatic Neoplasms , Stomach Neoplasms , Humans , Neuroendocrine Tumors/genetics , Intestinal Neoplasms/genetics , Stomach Neoplasms/genetics , Pancreatic Neoplasms/genetics , Tumor Microenvironment/geneticsABSTRACT
Hemogenic endothelial cells (HECs) are specialized cells that undergo endothelial-to-hematopoietic transition (EHT) to give rise to the earliest precursors of hematopoietic progenitors that will eventually sustain hematopoiesis throughout the lifetime of an organism. Although HECs are thought to be primarily limited to the aorta-gonad-mesonephros (AGM) during early development, EHT has been described in various other hematopoietic organs and embryonic vessels. Though not defined as a hematopoietic organ, the lung houses many resident hematopoietic cells, aids in platelet biogenesis, and is a reservoir for hematopoietic stem and progenitor cells (HSPCs). However, lung HECs have never been described. Here, we demonstrate that the fetal lung is a potential source of HECs that have the functional capacity to undergo EHT to produce de novo HSPCs and their resultant progeny. Explant cultures of murine and human fetal lungs display adherent endothelial cells transitioning into floating hematopoietic cells, accompanied by the gradual loss of an endothelial signature. Flow cytometric and functional assessment of fetal-lung explants showed the production of multipotent HSPCs that expressed the EHT and pre-HSPC markers EPCR, CD41, CD43, and CD44. scRNA-seq and small molecule modulation demonstrated that fetal lung HECs rely on canonical signaling pathways to undergo EHT, including TGFß/BMP, Notch, and YAP. Collectively, these data support the possibility that post-AGM development, functional HECs are present in the fetal lung, establishing this location as a potential extramedullary site of de novo hematopoiesis.
Subject(s)
Hemangioblasts , Hematopoiesis , Animals , Mice , Humans , Hematopoietic Stem Cells/metabolism , Cell Differentiation , Endothelium , Hemangioblasts/metabolismABSTRACT
Immune-related adverse events (irAEs) are a notable complication of PD-1 cancer immunotherapy. A better understanding of how these iatrogenic diseases compare with naturally arising autoimmune diseases is needed for treatment and monitoring of irAEs. We identified differences in anti-PD-1-induced type 1 diabetes (T1D) and spontaneous T1D in non-obese diabetic (NOD) mice by performing single-cell RNA-seq and TCR-seq on T cells from the pancreas, pancreas-draining lymph node (pLN), and blood of mice with PD-1-induced T1D or spontaneous T1D. In the pancreas, anti-PD-1 resulted in expansion of terminally exhausted/effector-like CD8+ T cells, an increase in T-bethi CD4+FoxP3- T cells, and a decrease in memory CD4+FoxP3- and CD8+ T cells in contrast to spontaneous T1D. Notably, anti-PD-1 caused increased TCR sharing between the pancreas and the periphery. Moreover, T cells in the blood of anti-PD-1-treated mice expressed markers that differed from spontaneous T1D, suggesting that the blood may provide a window to monitor irAEs rather than relying exclusively on the autoimmune target organ.
Subject(s)
Diabetes Mellitus, Type 1 , Animals , Mice , Mice, Inbred NOD , Pancreas , Forkhead Transcription Factors , Receptors, Antigen, T-CellABSTRACT
BACKGROUND: In ATTR amyloidosis, transthyretin (TTR) protein is secreted from the liver and deposited as toxic aggregates at downstream target tissues. Despite recent advancements in treatments for ATTR amyloidosis, the mechanisms underlying misfolded TTR-mediated cellular damage remain elusive. METHODS: In an effort to define early events of TTR-associated stress, we exposed neuronal (SH-SY5Y) and cardiac (AC16) cells to wild-type and destabilized TTR variants (TTRV122I (p.V142I) and TTRL55P (p.L70P)) and performed transcriptional (RNAseq) and epigenetic (ATACseq) profiling. We subsequently compared TTR-responsive signatures to cells exposed to destabilized antibody light chain protein associated with AL amyloidosis as well as ER stressors (thapsigargin, heat shock). RESULTS: In doing so, we observed overlapping, yet distinct cell type- and amyloidogenic protein-specific signatures, suggesting unique responses to each amyloidogenic variant. Moreover, we identified chromatin level changes in AC16 cells exposed to mutant TTR that resolved upon pre-incubation with kinetic stabilizer tafamidis. CONCLUSIONS: Collectively, these data provide insight into the mechanisms underlying destabilized protein-mediated cellular damage and provide a robust resource representing cellular responses to aggregation-prone proteins and ER stress.
Subject(s)
Amyloid Neuropathies, Familial , Amyloidosis , Neuroblastoma , Humans , Amyloid Neuropathies, Familial/complications , Amyloidogenic Proteins/genetics , Amyloidosis/metabolism , Neuroblastoma/complications , Neurons/metabolism , Prealbumin/genetics , Prealbumin/metabolismABSTRACT
Vascularized composite allotransplantation (VCA) is an evolving field of reconstructive surgery that has revolutionized the treatment of patients with devastating injuries, including those with limb losses or facial disfigurement. The transplanted units are typically comprised of different tissue types, including skin, mucosa, blood and lymphatic vasculature, muscle, and bone. It is widely accepted that the antigenicity of some VCA components, such as skin, is particularly potent in eliciting a strong recipient rejection response following transplantation. The fine line between tolerance and rejection of the graft is orchestrated by different cell types, including both donor and recipient-derived lymphocytes, macrophages, and other immune and donor-derived tissue cells (e.g., endothelium). Here, we delineate the role of different cell and tissue types during VCA rejection. Rejection of VCA grafts and the necessity of life-long multidrug immunosuppression remains one of the major challenges in this field. This review sheds light on recent developments in decoding the cellular signature of graft rejection in VCA and how these may, ultimately, influence the clinical management of VCA patients by way of novel therapies that target specific cellular processes.
Subject(s)
Vascularized Composite Allotransplantation , Humans , Vascularized Composite Allotransplantation/adverse effects , Immune Tolerance , Immunosuppression Therapy , Transplantation, Homologous , Graft RejectionABSTRACT
In this study, we demonstrate the utility of whole-slide CyCIF (tissue-based cyclic immunofluorescence) imaging for characterizing immune cell infiltrates in immune checkpoint inhibitor (ICI)-induced dermatologic adverse events (dAEs). We analyzed six cases of ICI-induced dAEs, including lichenoid, bullous pemphigoid, psoriasis, and eczematous eruptions, comparing immune profiling results obtained using both standard immunohistochemistry (IHC) and CyCIF. Our findings indicate that CyCIF provides more detailed and precise single-cell characterization of immune cell infiltrates than IHC, which relies on semi-quantitative scoring by pathologists. This pilot study highlights the potential of CyCIF to advance our understanding of the immune environment in dAEs by revealing tissue-level spatial patterns of immune cell infiltrates, allowing for more precise phenotypic distinctions and deeper exploration of disease mechanisms. By demonstrating that CyCIF can be performed on friable tissues, such as bullous pemphigoid, we provide a foundation for future studies to examine the drivers of specific dAEs using larger cohorts of phenotyped toxicity and suggest a broader role for highly multiplexed tissue imaging in phenotyping the immune mediated disease that they resemble.