A non-canonical type 2 immune response coordinates tuberculous granuloma formation and epithelialization.

The central pathogen-immune interface in tuberculosis is the granuloma, a complex host immune structure that dictates infection trajectory and physiology. Granuloma macrophages undergo a dramatic transition in which entire epithelial modules are induced and define granuloma architecture. In tuberculosis, relatively little is known about the host signals that trigger this transition. Using the zebrafish-Mycobacterium marinum model, we identify the basis of granuloma macrophage transformation. Single-cell RNA-sequencing analysis of zebrafish granulomas and analysis of Mycobacterium tuberculosis-infected macaques reveal that, even in the presence of robust type 1 immune responses, countervailing type 2 signals associate with macrophage epithelialization. We find that type 2 immune signaling, mediated via stat6, is absolutely required for epithelialization and granuloma formation. In mixed chimeras, stat6 acts cell autonomously within macrophages, where it is required for epithelioid transformation and incorporation into necrotic granulomas. These findings establish the signaling pathway that produces the hallmark structure of mycobacterial infection.

The In Vitro M1/M2 Polarization of Macrophages of BCG-Infected Mice.

Cultured peritoneal macrophages from intact (control) and BCG-infected (experiment) male BALB/c mice were studied 90 days after infection. Polarization of macrophages by M1 (expression of GM-CSF, IFNγ, and CD16/32) and M2 (expression of bFGF and CD36) differentiation pathways was studied with consideration for their the nuclearity class. Mononuclear cells predominated (90% and higher) in macrophage cultures of both groups and presumably, were presented by mainly epithelioid cells. The results indicated polarization of mononuclear and multinuclear macrophages in the M2 direction under conditions of BCG granulomatosis and a higher initial M2 polarization of binuclear macrophages. In control cultures, the ratio of M2 to M1 macrophages was 0.57, in experimental cultures this ratio was 1.6. It seems that long persistence of Mycobacterium tuberculosis in macrophages served as a factor stimulating the plastic processes and transformation of macrophages into epithelioid cells that form the "core" of granulomas and their enlargement upon incorporation of macrophages.

HEG1 Is a Highly Specific and Sensitive Marker of Epithelioid Malignant Mesothelioma.

Malignant mesothelioma can be difficult to distinguish from other malignancies, particularly non-small cell lung carcinomas (NSCLCs), without immunohistochemistry. However, conventional markers of mesothelial lineage all have variable degrees of cross-reactivity with other neoplasms, including NSCLCs, necessitating the use of multiple mesothelioma and carcinoma markers in every case for accurate diagnosis. A recently described monoclonal HEG homolog 1 (HEG1) antibody was proposed to be a specific marker for mesothelioma. Here we performed a large scale assessment of the SKM9-2 HEG1 antibody using tissue microarrays containing 69 epithelioid mesotheliomas, 32 sarcomatoid mesotheliomas, 167 NSCLCs, and 17 ovarian high-grade serous carcinomas. Strong membrane staining, usually diffuse, for HEG1 was seen in 65/69 (94%) epithelioid mesotheliomas, 0/60 pulmonary squamous cell carcinomas, 0/73 pulmonary adenocarcinomas, and 0/13 pulmonary large cell carcinomas. HEG1 showed staining in 14/32 (44%) sarcomatoid mesotheliomas compared with 0/21 sarcomatoid pulmonary carcinomas. Three of 17 (18%) high-grade serous carcinomas demonstrated membrane staining. Ten B3 thymoma whole sections were negative. On the microarrays, the conventional mesothelial markers calretinin, WT1, D2-40, and CK5/6 had sensitivities for epithelioid mesothelioma of 94%, 90%, 96%, and 91%, respectively. We conclude that HEG1 SKM9-2 antibody offers sensitivity comparable to conventional markers for epithelioid mesotheliomas, but provides considerably better specificity, such that the diagnosis of epithelioid mesothelioma versus NSCLC potentially could be confirmed with a combination of HEG1 and a suitable broad spectrum carcinoma marker such as claudin-4. HEG1 is specific but insensitive for separating sarcomatoid mesotheliomas from sarcomatoid lung carcinomas.

V-domain Ig-containing suppressor of T-cell activation (VISTA), a potentially targetable immune checkpoint molecule, is highly expressed in epithelioid malignant pleural mesothelioma.

V-domain Ig-containing suppressor of T-cell activation (VISTA) is an immune checkpoint gene that inhibits anti-tumor immune responses. Since most malignant pleural mesotheliomas do not respond to anti-programmed cell death(-ligand)1 (PD-(L)1)/cytotoxic T-lymphocyte-associated protein 4 (CTLA4) therapy and given the recent finding of The Cancer Genome Atlas Study that pleural mesothelioma displays the highest expression of VISTA among all cancers studied, we examined VISTA expression in a large pleural mesothelioma cohort. VISTA and PD-L1 immunohistochemistry were performed on tissue microarray of immunotherapy-naive pleural mesotheliomas (254 epithelioid, 24 biphasic and 41 sarcomatoid) and ten whole-tissue sections of benign pleura (VISTA only). Percentages of tumor and inflammatory cells with positive staining were assessed. Optimal prognostic cutoff percentages were determined using maximally selected rank statistics. Overall survival was evaluated using Kaplan-Meier methods and Cox proportional hazard analysis. All benign mesothelium expressed VISTA. Eighty-five percent of 319 and 38% of 304 mesotheliomas expressed VISTA and PD-L1 (88% and 33% of epithelioid, 90% and 43% of biphasic, and 42% and 75% of sarcomatoid), respectively. Median VISTA score was significantly higher in epithelioid (50%) (vs. biphasic [20%] and sarcomatoid [0]) (p < 0.001), while median PD-L1 score was significantly higher in sarcomatoid tumors (20%) (vs. biphasic and epithelioid [both 0%]) (p < 0.001). VISTA and PD-L1 were expressed in inflammatory cells in 94% (n = 317) and 24% (n = 303) of mesothelioma, respectively. Optimal prognostic cutoffs for VISTA and PD-L1 were 40% and 30%, respectively. On multivariable analysis, VISTA and PD-L1 expression in mesothelioma were associated with better and worse overall survival (p = 0.001 and p = 0.002), respectively, independent of histology. In a large cohort of mesothelioma, we report frequent expression of VISTA and infrequent expression of PD-L1 with favorable and unfavorable survival correlations, respectively. These findings may explain poor responses to anti-PD-(L)1 immunotherapy and suggest VISTA as a potential novel target in pleural mesothelioma.

Cutaneous epithelioid angiomatous nodule: a report of a series including a case with moderate cytologic atypia and immunosuppression.

BACKGROUND: Cutaneous epithelioid angiomatous nodule (CEAN) is a very rare and relatively recently recognized vascular proliferation characterized usually by minimal cytological atypia and accompanying mitotic activity. As such, CEAN represents an important diagnostic pitfall, which could lead to significant misdiagnosis and unnecessary treatment. METHODS: The clinicopathologic findings of 5 cases of CEAN were reviewed including a unique case with typical findings but also moderate cytologic atypia and brisk mitotic activity in a patient on immunosuppression. RESULTS: The cases were in 3 women and 2 men ranging in age from 18 to 61 years with lesions in the neck (2 cases), upper arm, back and shoulder. In 4 of the cases, the patients did not have any relevant potentially contributory clinical history, and in 1 case the patient was on immunosuppressive treatment. All 5 cases were superficially located within the dermis, well-circumscribed and similarly composed of epithelioid cells displaying minimal (in 4 cases) and moderate (1 case) atypia. The mitotic count ranged from 1 to 3 per 10 high power fields (HPF) in 4 cases and up to 9 per 10 HPF in the immunosuppressed patient. Atypical mitoses were not encountered in any of the cases. Two lesions that were incompletely excised recurred, but none of the patients showed distant metastases. CONCLUSION: While cytologically alarming, CEAN has a characteristic microscopic appearance and if completely excised follows an indolent course.

Glypican-1 immunohistochemistry is a novel marker to differentiate epithelioid mesothelioma from lung adenocarcinoma.

Histological morphology alone is not sufficient for the pathological diagnosis of malignant mesothelioma. Positive and negative immunohistochemical markers are necessary to differentiate it from lung adenocarcinoma. As calretinin and D2-40, the recognized positive markers of mesothelioma, are expressed in lung adenocarcinoma to some extent, novel markers with high specificity are desirable. In this study, we investigated the applicability of glypican-1 immunohistochemistry to differentiate epithelioid mesothelioma from lung adenocarcinoma. We investigated 82 cases of epithelioid mesothelioma and 97 cases of lung adenocarcinoma for glypican-1 expression by immunohistochemistry using a commercially available antibody. All 82 cases of epithelioid mesothelioma showed glypican-1 expression, most with diffuse and strong reactivity. In contrast, only three cases of lung adenocarcinoma showed focal glypican-1 expression. Glypican-1 expression showed 100 sensitivity, 97% specificity, and a 98% accuracy rate to differentiate epithelioid mesothelioma from lung adenocarcinoma. The sensitivity of glypican -1 immunohistochemistry is as high as that of calretinin and D2-40, and its specificity is far better than that of calretinin and D2-40. Therefore, we recommend including glypican -1 immunohistochemistry as a positive marker of epithelioid mesothelioma.

Incidental simultaneous finding of intravascular histiocytosis and reactive angioendotheliomatosis: a case report.

Reactive angioendotheliomatosis (RAE) is a rare cutaneous vascular disorder characterized by intravascular hyperplasia of endothelial cells, sometimes with a vascular proliferation. Intravascular histiocytosis (IH) is a similar vascular disorder characterized by the presence of dilated vessels containing aggregates of mononuclear histiocytes (macrophages) within their lumina. Although their pathogenesis remains uncertain, there has been speculation about the possible relationship between IH and RAE. We report a case of coexistence of RAE and IH in a patient who underwent a wide reexcision of a metastatic malignant melanoma. The excision specimen did not show any residual melanoma but exhibited an intravascular collection of CD-68-positive histiocytes admixed with CD-31-positive endothelial cells and fibrin surrounded by D2-40-positive vascular wall. The presence of intravascular cells initially raised concern of intravascular invasion by melanoma. As there was no clinical lesion and immunohistochemical stains for melanocytic makers were negative, we interpret this as an incidental finding. Knowledge of this benign vascular disorder is important because the histologic changes may be mistaken for intravascular invasion of a malignant neoplasm.

BRAFV600E immunopositive melanomas show low frequency of heterogeneity and association with epithelioid tumor cells: a STROBE-compliant article.

Treatment of BRAFV600E-mutant melanoma by small molecule inhibitors that target BRAFV600E or MEK kinases is increasingly used in clinical practice and significantly improve patient outcome. However, patients eventually become resistant and therapeutic improvement is required. Molecular diversity within individual tumors (intratumor heterogeneity) and between tumors within a single patient (intrapatient heterogeneity) poses a significant challenge to precision medicine. Using immunohistochemistry, we determined the extent of BRAFV600E intratumor and intrapatient heterogeneity and the influence of morphological heterogeneity in a large series of 171 melanomas of 81 patients. The BRAFV600E mutation rate found in our melanoma series is 44%, with none of 22 (0%) melanoma in situ, 23 of 56 (41%) primary tumors, 28 of 59 (48%) regional metastases, and 24 of 34 (71%) distant metastases harboring the mutation. In general, a diffuse homogeneous immunostaining was seen, even in tumors consisting of more than one cell type, that is, epithelioid, spindle, and/or small cell types. Nevertheless, BRAFV600E-mutant melanomas more often had a purely epithelioid cell population (P=0.063), that is more evident among distant metastases (P=0.014). Only two of 75 (3%) mutated specimens (one primary and one metastasis) displayed heterogeneous BRAFV600E expression. The primary tumor was also morphologically heterogeneous and exclusively displayed BRAFV600E in the epithelioid component, confirming an association between BRAFV600E and epithelioid cells. Twenty-eight of 30 patients (93%) had concordant BRAFV600E mutation status between their tumors. Taken together, BRAFV600E intratumor and intrapatient heterogeneity in melanoma is diminutive, nevertheless, the identified exceptions will have important implications for the clinical management of this disease.

Immunohistochemical detection of possible cellular origin of hepatic histiocytic sarcoma in mice.

Histiocytic sarcoma (HS) spontaneously arises in the liver in mice ; however, the cellular origins of hepatic HS have not been fully clarified. In this study, we immunohistochemically analyzed 18 cases of hepatic HS from the archives of our previous experiments. In all cases, the tumor cells showed positive reactions for the macrophage-specific markers F4/80 and CD68. The cells were negative for mesenchymal cell and lymphoid cell markers, suggesting that germ cell tumor or lymphoma components do not coexist in the neoplasm. We detected scattered Ly6C(+)F4/80(-) macrophage precursors in the extramedullary hematopoietic foci and liver tissue around the HS lesions. We also showed that certain populations of HS cells express the Ly-6C antigen. These findings suggest that Ly-6C(+) macrophage progenitor cells are a possible cellular origin of murine hepatic HS. Our study identified a novel phenotype of murine HS in two of 18 cases. These cases showed the nodular accumulations of tumor cells with cohesive cytoplasm mimicking the features of epithelioid granuloma. In agreement with the expression of CD204 in epithelioid cells in granulomatous diseases, these HS cells hardly expressed CD204, although the common type HS cells were strongly positive for this antigen. These data suggest that hepatic HS may stem from Ly-6C(+) macrophage precursors. Furthermore, a subset of hepatic HS cases can possibly differentiate into epithelioid cell-like phenotypes.

Microscopic endometrial perivascular epithelioid cell nodules: a case report with the earliest presentation of a uterine perivascular epithelioid cell tumor.

Perivascular epithelioid cell (PEC) tumors (PEComas) are a family of related mesenchymal tumors composed of PECs which co-express melanocytic and smooth muscle markers. Although their distinctive histologic, immunohistochemical, ultrastructural, and genetic features have been clearly demonstrated, their histogenesis and normal counterpart remain largely unknown. Precursor lesions of PEComas have rarely been reported. We herein describe a tuberous sclerosis patient with microscopic PEC nodules in the endometrium of adenomyosis, pelvic endometriosis, an ovarian endometriotic cyst, and the endometrium of the uterine cavity. The nodules showed a mixture of spindle-shaped and epithelioid cells concentrically arranged around small arteries. The cells exhibited uniform nuclei, light eosinophilic cytoplasm, and immunoreactivity with HMB-45 and CD10. Some nodules revealed continuity with a PEComa in the myometrium. These findings support microscopic endometrial PEC nodules possibly being precursor lesions of uterine PEComas. The wide distribution of the nodules in the pelvis may be related to the multicentricity of PEComas in tuberous sclerosis patients. Owing to the immunoreactivity with CD10, microscopic endometrial PEC nodules may be misinterpreted as endothelial stromal cells unless melanocytic markers are stained. To the best of our knowledge, this is a case with the earliest manifestation of PEC lesions occurring in the endometrium. Virtual slides: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/9658280017862643.

Granular cell variant of epithelioid cell histiocytoma.

Classic granular cell tumors (GCTs) stain strongly and uniformly positive for S100 protein and are believed to show Schwann cell derivation. Polypoid cutaneous tumors composed of cells with large nuclei and abundant granular cytoplasm that do not stain for S100 protein or show apparent Schwannian differentiation have been reported by several groups under names including "primitive polypoid granular cell tumors," "dermal nonneural granular cell tumor," and "primitive nonneural granular cell tumors of skin." We report a polypoid tumor composed of S100-negative epithelioid cells with abundant eosinophilic granular cytoplasm that meets diagnostic criteria for (primitive polypoid dermal) nonneural GCT but also meets criteria for a granular cell variant of epithelioid cell histiocytoma. We have identified a single previous report of a similar lesion. We report the immunohistochemical characteristics of these lesions and address how they are best classified.

A misleading hepatic tumour: epithelioid angiomyolipoma.

Hepatic angiomyolipoma (HAML) is a rare, benign mesenchymal neoplasm composed of varying amounts of smooth muscle cells, adipose tissue, and vessels. Its morphological diversity often poses diagnostic problems. In this paper, the authors report a peculiar case of epithelioid HAML mimicking histologically hepatocellular carcinoma with focal areas resembling inflammatory pseudotumour. A 57 year-old male patient presented with abdominal pain and discomfort. Non enhanced CT scan demonstrated a heterogeneous hypodense mass located in segment II and IV of the liver. Hepatocellular carcinoma was suspected and the patient underwent left lobectomy. Histologically, the tumour was mainly composed of epithelioid cells arranged in trabeculae and sheets (50% of the tumour surface) admixed with mature fat cells (20%) and thick-walled blood vessels. Lymphocytic aggregates and clusters of foamy histiocytes were focally found in the stroma (30%). Most of the epithelioid tumour cells were immunoreactive to homatropine methylbromide 45 (HMB-45) and smooth muscle actin. Morphological pattern and immunophenotype were consistent with epithelioid HAML.

Experimental leprosy: a model of epithelioid cell granuloma.

An animal model of granulomatous hypersensitivity has been developed, which reproduces some features of the pathologies of important chronic granulomatous disorders, including tuberculosis, tuberculoid leprosy, sarcoidosis, berylliosis, Crohn's disease, and sensitivity to zirconium. The lesions consist of focal collections of epithelioid cells surrounded by lymphocytes to form tubercles. The epithelioid cell has a secretory function and is not phagocytic. Plasmacytoid dendritic cells are precursors of epithelioid cells, which are therefore part of the innate immune system. Subplasmalemmal linear densities are also present in these cells. This autoimmune model has been induced in rabbits using a non-myelin sensory peripheral antigen to reproduce the features of tuberculoid leprosy. The antigen is probably present only in human tissue. A granuloma antigen, which is tissue specific similar to that in peripheral nerves, could be present in sarcoidosis and Crohn's disease. In multiple sclerosis, mononuclear cells in the brain parenchyma are not phagocytic and are therefore similar to epithelioid cells. The induction of tolerance leading to the development of a vaccine to prevent the lesions in multiple sclerosis, sarcoidosis, and Crohn's disease is possible after purification of the granuloma antigen.

Persistent lymphoedema in Morbihan disease: formation of perilymphatic epithelioid cell granulomas as a possible pathogenesis.

Morbihan disease is a rare complication of rosacea, characterized by persistent lymphoedema on the upper half of the face, occurring during the chronic clinical course of rosacea. This refractory condition has been also designated as 'rosacea lymphoedema' and 'solid persistent facial oedema of rosacea'. We report a patient with Morbihan disease showing persistent lymphoedema on the upper half of the face accompanied by unique histological findings of striking dermal dilated lymphatics and damage of the lymphatics at the site of the adjacent epithelioid cell granulomas, with histiocytes bulging into the lymphatic lumen. The marked epithelioid cell granulomas forming around dermal lymphatic vessels with subsequent lymphatic damage and luminal obstruction by histiocytic infiltration may account for the development of lymphoedema in this patient.

Evaluation of CD10 and procollagen 1 expression in atypical fibroxanthoma and dermatofibroma.

Atypical fibroxanthoma (AFX) (dermal pleomorphic sarcoma) remains a somewhat controversial entity. Some authors have averred that AFX is a fiction, suggesting that such lesions merely represent misclassified examples of spindled squamous cell carcinoma. In addition, the immunoperoxidase confirmation of AFX has been less than straightforward and has historically been approached as a diagnosis of exclusion because of the lack of sensitivity and specificity of available "positive" reagents. Procollagen 1 (PC1) and CD10 represent recently developed immunoperoxidase reagents that have been forwarded as useful in this setting, and we sought to characterize our experience, both to confirm the utility of these antibodies and to compare them. Our investigation included 3 separate data sets. Group 1 consisted of a retrospective review of 98 consecutive cases in which PC1 was used in the evaluation of dermatopathology specimens in routine practice during a 13-month interval. Group 2 consisted of a direct comparison of 11 AFX, 11 dermatofibroma (DF), and 7 epithelioid dermatofibroma (EDF) using the CD10 reagent on cases identified by database search. Group 3 consisted of a retrospective review of 47 cases in which CD10 was used in routine practice during a 10-month interval. Group 1 included 47 AFX, 13 carcinomas, and 6 melanomas. PC1 expression was observed in 45 of 47 AFX (96%), with a strong reaction in 78% of cases. Among a comparison group of carcinomas, 13 of 13 displayed strong keratin immunopositivity and 11 of 13 (85%) lacked PC1 expression whereas 2 showed focal weak labeling. Six of six melanomas exhibited avid S100 expression and none labeled with PC1. In group 2, strong CD10 immunoreactivity was present in 11 of 11 AFX. Similarly, 11 of 11 DFs were also positive. In contrast, 6 of 7 cases of EDF lacked CD10 expression. Group 3 included 38 AFX and 9 miscellaneous spindle cell proliferations. Of the 38 AFX, 37 (97%) labeled with CD10 and in 34 (92%) the reaction was strong. PC1 immunostaining was also completed in 34 of 38 AFX from group 3 and 27 (79%) cases showed positive labeling. Our results confirm that both PC1 and CD10 can be used as positive markers of AFX. We believe that CD10 and PC1 immunostaining can be used as a useful adjunct to supplement the diagnosis of AFX, within the context of an immunoperoxidase panel. Not surprisingly, CD10 expression is also common in DF, a benign analog of AFX, with the exception of its epithelioid variant. In direct head-to-head comparison, our experience indicates that the staining of AFX with CD10 is more avid than that observed with PC1. Lastly, out data includes over 80 examples of AFX, <5% of which showed keratin labeling. Given a general lack of keratin expression, it seems unlikely that AFX merely represents poorly differentiated squamous carcinoma.

Perivascular epithelioid cell tumour of the liver.

Perivascular epithelioid cell tumour is not uncommon in the liver but seldom malignant.

Sclerosing PEComa: clinicopathologic analysis of a distinctive variant with a predilection for the retroperitoneum.

PEComas (tumors showing perivascular epithelioid cell differentiation) are a family of mesenchymal neoplasms that include angiomyolipoma, clear cell "sugar" tumor of the lung, lymphangiomyomatosis, and a group of uncommon lesions that arise in soft tissue, visceral organs, and skin. We describe a distinctive variant of PEComa that shows extensive stromal hyalinization, a feature not previously described in these tumors. Thirteen PEComas with extensive stromal hyalinization were identified from a total of 70 cases of PEComa received between 1996 and 2006 (19%). All patients were women, with a mean age of 49 years (range, 34 to 73y). One patient had tuberous sclerosis. Ten tumors (77%) arose in the retroperitoneum (8 pararenal), and 1 each in the pelvis, uterus, and abdominal wall. Median tumor size was 9.5 cm (range, 4.5 to 28 cm). All except 2 were grossly well-circumscribed. The tumors were composed of cords and trabeculae of cytologically uniform bland epithelioid cells with palely eosinophilic, granular to clear cytoplasm and round nuclei with small nucleoli, embedded in abundant densely sclerotic stroma. Five tumors contained a spindle cell component, and 6 showed focally sheetlike areas. In all cases the tumor cells were focally arranged around blood vessels. All tumors lacked the delicate nesting vascular pattern typical of other PEComas. Mitoses ranged from 0 to 3/50 high-power field (mean 1) in all cases except 1. One tumor showed abrupt transition to areas with strikingly pleomorphic morphology, marked nuclear atypia, frequent mitoses (22/10 high-power field), and fascicular and nested architecture. This was the only case with necrosis. All tumors were immunopositive for desmin (usually diffusely) and HMB-45 (generally in scattered cells); 12/13 (92%) expressed smooth muscle actin, 11/12 (92%) caldesmon, 11/12 (92%) microphthalmia transcription factor (D5), and 3/13 (23%) melan-A. Only 1 (8%) was focally S-100 positive. All tumors were negative for epithelial membrane antigen, PAN-K, and KIT (CD117). Follow-up was available for 9 patients, ranging from 10 to 64 months (median, 33). One patient (whose tumor showed transition to high-grade malignant morphology) developed metastases to lung, liver, and abdominal wall. No other tumor has recurred or metastasized thus far. Sclerosing PEComa is a distinctive variant with a predilection for the pararenal retroperitoneum of middle-aged women. Sclerosing PEComas seem to pursue an indolent clinical course, unless associated with a frankly malignant component. Long-term follow-up will be required to confirm these findings.