Necrotic Keratinocytes Are Common in Psoriasis and Have a Predilection to the Upper Epidermis: A Quantitative and Comparative Analysis.

BACKGROUND: The most salient histopathological features of psoriasis are epidermal hyperplasia, hypogranulosis, parakeratosis, dilated capillaries in dermal papillae, and intraepidermal and intracorneal neutrophils. Several additional "nonclassic" features of psoriasis have recently been reported, including necrotic keratinocytes (NK). To determine the diagnostic utility of NK, we characterized NK in a large cohort of psoriasis cases compared with psoriasiform spongiotic dermatitis (PSD) and normal skin. METHODS: NK were quantified in 101 cases of psoriasis, 20 cases of PSD, and 20 cases of normal skin. The location of NK within the lower, middle, or upper thirds of the epidermis was recorded. RESULTS: NK were identified in 77/101 (76%) of psoriasis cases. By comparison, NK were seen in 8/20 (40%) cases of PSD and 4/20 (20%) cases of normal skin. The linear concentration of NK was significantly higher in psoriasis (0.36 NK/mm) compared with PSD (0.12 NK/mm) and normal skin (0.03 NK/mm) (P = 0.0009). NK were preferentially located in the upper (58%) and middle (31%) epidermis in psoriasis. CONCLUSIONS: NK are a common feature of psoriasis and have a predilection to the upper layers of epidermis. Superficial NK may provide additional diagnostic support for psoriasis in challenging or borderline cases.

Histopathology-guided mass spectrometry differentiates benign nevi from malignant melanoma.

PURPOSE: Distinguishing benign nevi from malignant melanoma using current histopathological criteria may be very challenging and is one the most difficult areas in dermatopathology. The goal of this study was to identify proteomic differences, which would more reliably differentiate between benign and malignant melanocytic lesions. METHODS: We performed histolpathology - guided mass spectrometry (HGMS) profiling analysis on formalin-fixed, paraffin embedded tissue samples to identify differences at the proteomic level between different types of benign nevi and melanomas. A total of 756 cases, of which 357 cases of melanoma and 399 benign nevi, were included in the study. The specimens originated from both biopsies (376 samples) and tissue microarray (TMA) cores (380 samples). After obtaining mass spectra from each sample, classification models were built using a training set of biopsy specimens from 111 nevi and 100 melanomas. The classification algorithm developed on the training data set was validated on an independent set of 288 nevi and 257 melanomas from both biopsies and TMA cores. RESULTS: In the melanoma cohort, 239/257 (93%) cases classified correctly in the validation set, 3/257 (1.2%) classified incorrectly, and 15/257 (5.8%) classified as indeterminate. In the cohort of nevi, 282/288 (98%) cases classified correctly, 1/288 (0.3%) classified incorrectly, and 5/288 (1.7%) were indeterminate. HGMS showed a sensitivity of 98.76% and specificity of 99.65% in determining benign vs malignant. CONCLUSION: HGMS proteomic analysis is an objective and reliable test with minimal tissue requirements, which can be a helpful ancillary test in the diagnosis of challenging melanocytic lesions.

Subcutaneous Panniculitis-Like T-Cell Lymphoma With Granulomas as the Predominant Feature.

Subcutaneous panniculitis-like T-cell lymphoma (SPTCL) is a rare primary cutaneous lymphoma preferentially localized in the subcutaneous adipose tissue and composed of cytotoxic T cells with an α/ß immunophenotype. The neoplastic T cells can be variably admixed with other inflammatory cells, including histiocytes, which can rarely form noncaseating granulomas. We present a case of SPTCL in which granulomas are the predominant feature, composing 75%-80% of the inflammatory infiltrate. The top differential diagnoses included infectious and autoimmune etiologies. However, special stains for microorganisms were negative, and immunohistochemical analysis of the atypical lymphocytes showed a CD3, CD8, TIA-1+, T-cell receptor (TCR) beta+, and CD4 infiltrate with a high Ki67 proliferation index of approximately 30%. TCR gene rearrangement studies by polymerase chain reaction with confirmation by high-throughput sequencing were necessary to exclude an autoimmune etiology, specifically lupus erythematosus panniculitis. To the best of our knowledge, only 1 other case of SPTCL with prominent granulomas has been reported in the literature. It is important for dermatopathologists to recognize this presentation of SPTCL. SPTCL with predominant granulomas should be included in the differential diagnosis of granulomatous panniculitis along with infectious and autoimmune panniculitides as well as other granulomatous lymphomas.

Macrophage Migration Inhibitory Factor Regulates U1 Small Nuclear RNP Immune Complex-Mediated Activation of the NLRP3 Inflammasome.

OBJECTIVE: High-expression alleles of macrophage migration inhibitory factor (MIF) are linked genetically to the severity of systemic lupus erythematosus (SLE). The U1 small nuclear RNP (snRNP) immune complex containing U1 snRNP and anti-U1 snRNP antibodies, which are found in patients with SLE, activates the NLRP3 inflammasome, comprising NLRP3, ASC, and procaspase 1, in human monocytes, leading to the production of interleukin-1ß (IL-1ß). This study was undertaken to investigate the role of the snRNP immune complex in up-regulating the expression of MIF and its interface with the NLRP3 inflammasome. METHODS: MIF, IL-1ß, NLRP3, caspase 1, ASC, and MIF receptors were analyzed by enzyme-linked immunosorbent assay, Western blotting, quantitative polymerase chain reaction, and cytometry by time-of-flight mass spectrometry (CytoF) in human monocytes incubated with or without the snRNP immune complex. MIF pathway responses were probed with the novel small molecule antagonist MIF098. RESULTS: The snRNP immune complex induced the production of MIF and IL-1ß from human monocytes. High-dimensional, single-cell CytoF analysis established that MIF regulates activation of the NLRP3 inflammasome, including findings of a quantitative relationship between MIF and its receptors and IL-1ß levels in the monocytes. MIF098, which blocks MIF binding to its cognate receptor, suppressed the production of IL-1ß, the up-regulation of NLRP3, which is a rate-limiting step in NLRP3 inflammasome activation, and the activation of caspase 1 in snRNP immune complex-stimulated human monocytes. CONCLUSION: The U1 snRNP immune complex is a specific stimulus of MIF production in human monocytes, with MIF having an upstream role in defining the inflammatory characteristics of activated monocytes by regulating NLRP3 inflammasome activation and downstream IL-1ß production. These findings provide mechanistic insight and a therapeutic rationale for targeting MIF in subgroups of lupus patients, such as those classified as high genotypic MIF expressers or those with anti-snRNP antibodies.

Quantitative assessment of neuropilin-2 as a simple and sensitive diagnostic assay for spitzoid melanocytic lesions.

There is a significant need for the development of diagnostic tools that can precisely distinguish Spitz nevi and spitzoid melanomas. Here, we report the development of a PCR-based quantitative diagnostic assay for spitzoid melanocytic lesions utilizing the expression ratio of neuropilin-2 and melan-A genes in primary tumor specimens. We find that the expression ratio of neuropilin-2/melan-A is significantly increased in spitzoid melanomas compared with Spitz nevi. The diagnostic potential of this quantitative assay was validated in two independent sets of patient samples as demonstrated in a receiver operating characteristic curve analysis showing an area under the curve value of 91.8%. Furthermore, the assay was found to quantitatively distinguish the clinical nature of atypical spitzoid melanocytic lesions that were diagnostically undetermined using histopathologic criteria alone. Our data indicate that this quantitative assay may be used as a tool in determining the diagnostic classification of histologically challenging spitzoid tumors.

Proteomic Mass Spectrometry Imaging for Skin Cancer Diagnosis.

Mass spectrometry imaging can be successfully used for skin cancer diagnosis, particularly for the diagnosis of challenging melanocytic lesions. This method analyzes proteins within benign and malignant melanocytic tumor cells and, based on their differences, which constitute a unique molecular signature of 5 to 20 proteins, can render a diagnosis of benign nevus versus malignant melanoma. Mass spectrometry imaging may assist in the differentiation between metastases and nevi as well as between proliferative nodules in nevi and melanoma arising in a nevus. In the difficult area of atypical Spitzoid neoplasms, mass spectrometry diagnosis can predict clinical outcome better than histopathology.

Mass Spectrometry Imaging Can Distinguish on a Proteomic Level Between Proliferative Nodules Within a Benign Congenital Nevus and Malignant Melanoma.

Histopathological interpretation of proliferative nodules occurring in association with congenital melanocytic nevi can be very challenging due to their similarities with congenital malignant melanoma and malignant melanoma arising in association with congenital nevi. We hereby report a diagnostically challenging case of congenital melanocytic nevus with proliferative nodules and ulcerations, which was originally misdiagnosed as congenital malignant melanoma. Subsequent histopathological examination in consultation by one of the authors (R.L.) and mass spectrometry imaging analysis rendered a diagnosis of congenital melanocytic nevus with proliferative nodules. In this case, mass spectrometry imaging, a novel method capable of distinguishing benign from malignant melanocytic lesions on a proteomic level, was instrumental in making the diagnosis of a benign nevus. We emphasize the importance of this method as an ancillary tool in the diagnosis of difficult melanocytic lesions.

Spitz nevi and Spitzoid melanomas: exome sequencing and comparison with conventional melanocytic nevi and melanomas.

We performed exome sequencing of 77 melanocytic specimens composed of Spitz nevi (n=29), Spitzoid melanomas (n=27), and benign melanocytic nevi (n=21), and compared the results with published melanoma sequencing data. Our study highlights the prominent similarity between Spitzoid and conventional melanomas with similar copy number changes and high and equal numbers of ultraviolet-induced coding mutations affecting similar driver genes. Mutations in MEN1, PRKAR1A, and DNMT3A in Spitzoid melanomas may indicate involvement of the protein kinase A pathway, or a role of DNA methylation in the disease. Other than activating HRAS variants, there were few additional mutations in Spitz nevi, and few copy number changes other than 11p amplification and chromosome 9 deletions. Similarly, there were no large-scale copy number alterations and few somatic alterations other than activating BRAF or NRAS mutations in conventional nevi. A presumed melanoma driver mutation (IDH1Arg132Cys) was revealed in one of the benign nevi. In conclusion, our exome data show significantly lower somatic mutation burden in both Spitz and conventional nevi compared with their malignant counterparts, and high genetic similarity between Spitzoid and conventional melanoma.

A 65-Year-Old Female from Connecticut with Orf Infection.

The virus, which causes orf and induces acute pustular skin lesions in sheep and goats, is transmissible to humans yet is rarely observed in North America. We present a case of a 65-year-old female farmer from Connecticut who contracted orf from her sheep. The clinical and histopathologic features, important to arrive at the correct diagnosis of this uncommon yet important infection, are described. We also discuss the benign nature of this condition and emphasize that treatment is not required.

Imaging mass spectrometry assists in the classification of diagnostically challenging atypical Spitzoid neoplasms.

BACKGROUND: Previously, using imaging mass spectrometry (IMS), we discovered proteomic differences between Spitz nevi and Spitzoid melanomas. OBJECTIVE: We sought to determine whether IMS can assist in the classification of diagnostically challenging atypical Spitzoid neoplasms (ASN), to compare and correlate the IMS and histopathological diagnoses with clinical behavior. METHODS: We conducted a retrospective collaborative study involving centers from 11 countries and 11 US institutions analyzing 102 ASNs by IMS. Patients were divided into clinical groups 1 to 4 representing best to worst clinical behavior. The association among IMS findings, histopathological diagnoses, and clinical groups was assessed. RESULTS: There was a strong association between a diagnosis of Spitzoid melanoma by IMS and lesions categorized as clinical groups 2, 3, and 4 (recurrence of disease, metastases, or death) compared with clinical group 1 (no recurrence or metastasis beyond a sentinel node) (P < .0001). Older age and greater tumor thickness were strongly associated with poorer outcome (P = .01). CONCLUSIONS: IMS diagnosis of ASN better predicted clinical outcome than histopathology. Diagnosis of Spitzoid melanoma by IMS was strongly associated with aggressive clinical behavior. IMS analysis using a proteomic signature may improve the diagnosis and prediction of outcome/risk stratification for patients with ASN.

Report of two cases of vacuolar interface dermatitis initially suspected as melanoma in situ and review of the literature / Relato de dois casos de dermatite de interface vacuolar, inicialmente diagnosticados como melanoma in situ, e revisão da literatura

ABSTRACT The diagnosis of melanocytic lesions can be challenging and immunohistochemical study is a valuable tool for dermatopathologists. We report two cases initially simulating melanoma in situ, reviewing the histopathological and immunohistochemical findings and the cases published in the literature with similar findings/results. We emphasize the importance of clinicopathological correlation in the evaluation of lesions with interface changes and in the "pseudomelanocytic nests", which may simulate melanoma in situ. We also highlight the importance of using a immunohistochemistry panel in addition to Melan-A, in the study of these lesions.

RESUMO O diagnóstico de lesões melanocíticas pode ser desafiador, e o estudo imuno-histoquímico é uma ferramenta valiosa para os dermatopatologistas. Relatamos dois casos inicialmente diagnosticados como melanoma in situ, avaliando os achados histopatológicos e imuno-histoquímicos e os casos publicados na literatura com resultados semelhantes. Ressaltamos a importância da correlação clinicopatológica na avaliação das lesões com danos na interface e nos "ninhos pseudomelanocíticos", que podem simular melanoma in situ. Destacamos também a importância da utilização de um painel de imuno-histoquímica, além do Melan-A, no estudo dessas lesões.

Somatic Mosaicism in Blaschkolinear Inflammatory Disorders.

Linear lichen planus (LP) is a rare dermatologic disease in which lichenoid lesions conform to a blaschkolinear distribution, most commonly on the extremities. Linear discoid lupus erythematosus (DLE) is a cutaneous manifestation of lupus that also conforms to Blaschko's lines. Blaschkolinear disorders have been shown to result from somatic mosaicism, most recently in nevus sebaceus, epidermal nevi, and syringocystadenoma papilliferum. In linear LP and DLE, presentation of papules along Blaschko's lines suggests that these disorders can result from keratinocytic genetic mosaicism. Their onset later in life suggests that a secondary trigger is necessary to drive inflammatory reactions to these linear lesions. To date causative mutations have not be identified. We report 2 cases of linear lichen planus and 1 case of linear discoid lupus erythematosus, all histologically confirmed. Both LP patients have experienced episodic regression and recurrence of their lesions in precisely the same distribution, with moderate symptomatic benefit from topical steroids and non-steroidal anti-inflammatories. The DLE patient showed gradual response to hydroxychloroquine over the course of 14 months. These cases highlight linear inflammatory diseases which represent localized variants of disorders for which there are few efficacious therapies. Their linear presentations suggest that they result from somatic mosaicism and genetic investigation of such disorders may reveal relevant therapeutic targets.

Evidence for glycosylation as a regulator of the pigmentary system: key roles of sialyl(α2-6)gal/GalNAc-terminated glycans in melanin synthesis and transfer.

UNLABELLED: The major regulators of melanogenesis are glycoproteins, however no role for glycosylation in the pathway has yet been described. We stained skin biopsies and melanocyte-keratinocyte co-cultures with a panel of 20 lectins as oligosaccharide markers. Notably, the Elderberry Bark Lectin (EBL/SNA) stained melanocytes in both systems. EBL binds the sequence Neu5Ac(α(2-6)Gal/GalNAc)- at the termini of some oligosaccharide antennae. We used inhibitors of synthesis and/or binding of this sequence to assess effects on pigmentation. METHODS: Cell culture, lectin histochemistry, siRNA transfection, and assays for dopa oxidase and melanin were carried out by standard techniques. RESULTS: 6'-sialyllactose, a short homolog of the sequence in question, anti-sialyltransferase 6 (ST6) siRNA, and cytidine, a sialyltransferase (ST) inhibitor, each inhibited EBL binding, melanogenesis and melanosome transfer. Unexpectedly, 3'-sialyllactose and siRNA for ST3, chosen as a negative controls, also inhibited these processes. Though strong inhibitors of melanization, none of the agents affected tyrosinase/dopa oxidase activity, indicating previously unrecognized post-tyrosinase regulation of melanization. CONCLUSIONS: We report for the first time that Neu5Ac (α(2-6)Gal/GalNAc)- and possibly Neu5Ac(α(2-3)Gal/GalNAc)-terminated oligosaccharides play multiple roles in melanin synthesis and transfer.

Congenital nevi versus metastatic melanoma in a newborn to a mother with malignant melanoma - diagnosis supported by sex chromosome analysis and Imaging Mass Spectrometry.

A 37-year-old pregnant woman presented with a 2-cm irregular reddish nodule on her left upper arm during pregnancy. A biopsy from the lesion showed a 2.2-mm thick malignant melanoma with intravascular invasion, 25 mitosis/mm(2) and no ulceration. Following induction of labor, the patient underwent re-excision with sentinel lymph node biopsy. This showed no residual melanoma and no lymph node metastasis. The newborn boy had multiple pigmented lesions on the trunk, some of which were large and irregular. Two were biopsied and histologic examination showed dense dermal proliferation of medium sized melanocytes with multiple mitotic figures and no maturation with their descent into the dermis, raising suspicion of transplacental metastases. Examination of the placenta failed to show metastatic lesions. Multiplex polymerase chain reaction (PCR)-based genotyping, including testing for amelogenin locus for sex chromosome determination, demonstrated the presence of Y chromosome material in the melanocytes of the newborn's lesions excluding maternal origin. A diagnosis of congenital nevi was rendered. Subsequently, Imaging Mass Spectrometric analysis of the mother's lesion showed proteomic signature expression indicative of malignant melanoma, whereas the two lesions in the newborn showed changes indicative of nevi. This case demonstrates the utility of genotyping and Mass Spectrometry analysis in this challenging clinical scenario.

Staphylococcal Purpura Fulminans: Report of a Case.

Purpura fulminans (PF) is associated with several infections and most commonly with meningococcemia. However, there are only a few reports of this entity in association with toxic shock syndrome toxin-1-producing Staphylococcus aureus. We report a 53-year-old man who presented with fever, progressive hemodynamic instability, multiorgan failure, and thrombocytopenia following lobectomy for a solitary lung metastasis from rectal adenocarcinoma. He developed progressive generalized eruption of nonblanching red, purple, and black macules, papules, and plaques on the trunk and extremities consistent with PF. He died on postadmission day 3. Autopsy examination revealed purulent pleural exudate, which grew toxic shock syndrome toxin-1-producing S. aureus. Premortem and autopsy skin biopsies demonstrated epidermal necrosis, subepidermal bullae, and fibrin thrombi within small cutaneous vessels with minimal perivascular lymphocytic inflammation and without accompanying vasculitis. With this case report, we would like to draw attention to the fact that staphylococcal toxic shock syndrome-associated PF may be highly underrecognized and much more common than reflected in the literature.

Comparison of the expression of vimentin and actin in spitz nevi and spitzoid malignant melanomas.

INTRODUCTION: The differentiation between Spitz nevi (SN) and Spitzoid malignant melanomas (SMM) represents a challenge to dermatopathologists. We recently demonstrated differential expression of vimentin and Actin in SN and SMM by mass spectrometry (MS). We sought to investigate whether this differential expression could be detected using conventional immunohistochemistry or automated quantitative analysis (AQUA) of histological sections. METHODS: Cases of SN and SMM, which were previously studied by MS and have readily available blocks and enough material in the block, were selected from the Yale Spitzoid Neoplasm Repository. The cases were stained for vimentin and muscle-specific actin using standard protocols. H-scores were calculated by multiplying the percentage of cells staining and the intensity of staining. Selected cases were also studied for quantitative immunofluorescent staining using the AQUA method. RESULTS: All 21 cases of SN showed strong and diffuse staining for vimentin; 19 of 21 (91%) cases had an H-score of 300 (average, 294). Similar staining results were observed in SMM; 13 of 14 (93%) cases had an H-score of 300 (average, 297). Muscle-specific actin was weakly and focally positive in 5 of 21 (24%) SN (H-score = 3.3) and 5 of 14 (39%) SMM (H-score = 3.5). The AQUA method showed no significant difference in the staining intensity of SN and SMM for both vimentin and actin. CONCLUSIONS: There was no difference in the expression of vimentin and actin in SN and SMM shown by conventional immunohistochemistry or the AQUA method. This study shows that MS has much grater sensitivity in detecting the differential expression of these proteins.