Amerigo Santoro

Defective dendritic cell migration and activation of adaptive immunity in PI3Kγ‐deficient mice

Gene‐targeted mice were used to evaluate the role of the gamma isoform of phosphoinositide 3‐kinase (PI3Kγ) in dendritic cell (DC) migration and induction of specific T‐cell‐mediated immune responses. DC obtained from PI3Kγ−/− mice showed a reduced ability to respond to chemokines in vitro and ex vivo and to travel to draining lymph nodes under inflammatory conditions. PI3Kγ−/− mice had a selective defect in the number of skin Langerhans cells and in lymph node CD8α− DC. Furthermore, PI3Kγ−/− mice showed a defective capacity to mount contact hypersensitivity and delayed‐type hypersensitivity reactions. This defect was directly related to the reduced ability of antigen‐loaded DC to migrate from the periphery to draining lymph nodes. Thus, PI3Kγ plays a nonredundant role in DC trafficking and in the activation of specific immunity. Therefore, PI3Kγ may be considered a new target to control exaggerated immune reactions.

Novel aspects of Sjögren’s syndrome in 2012

Sjögren’s syndrome (SS) is a systemic progressive autoimmune disease characterized by a complex pathogenesis requiring a predisposing genetic background and involving immune cell activation and autoantibody production. The immune response is directed to the exocrine glands, causing the typical ‘sicca syndrome’, but major organ involvement is also often seen. The etiology of the disease is unknown. Infections could play a pivotal role: compared to normal subjects, patients with SS displayed higher titers of anti-Epstein-Barr virus (EBV) early antigens, but lower titers of other infectious agent antibodies such as rubella and cytomegalovirus (CMV) suggest that some infections may have a protective role against the development of autoimmune disease. Recent findings seem to show that low vitamin D levels in patients with SS could be associated with severe complications such as lymphoma and peripheral neuropathy. This could open new insights into the disease etiology. The current treatments for SS range from symptomatic therapies to systemic immunosuppressive drugs, especially B cell-targeted drugs in cases of organ involvement. Vitamin D supplementation may be an additional tool for optimization of SS treatment.

Recruitment of dendritic cells in oral lichen planus

Using immunohistochemistry the presence of different dendritic cell (DC) subsets was analysed in 16 biopsies from patients with oral lichen planus (OLP). A significant increase of CD1a+/Langerin+ Langerhans cells, DC‐SIGN+ DC and CD123+/BDCA2+ plasmacytoid DCs (PDCs) was found in the epithelium and in the stroma of OLP biopsies compared to normal oral mucosa. A proportion of DCs were mature DC‐LAMP+ and expressed S100 or CD11c, typically found in the interdigitating DCs of nodal T‐cell areas. Double staining revealed that mature DCs co‐expressed CCR7, thus indicating the development of a nodal migratory phenotype upon maturation. Significant recruitment of PDCs producing IFN‐α was demonstrated by the expression of MxA within the lichenoid inflammatory infiltrate and close cell‐to‐cell contacts between PDCs and mature DCs were observed, with a significant correlation between the numbers of these two populations. Moreover, PDCs were also found to contain Granzyme‐B, an associated‐cytotoxic granule protein, inducing target cell apoptosis. Taken together, these results suggest that PDCs may promote maturation of DCs and amplify the cytotoxicity of lymphoid cells. Finally, the recruitment of different subtypes of DC, such as Langerhans cells, stromal DC‐SIGN+ DCs and PDCs, associated with a significant proportion of mature DCs, acquiring a CCR7+ ‘migratory’ phenotype, indicate that they may play a pivotal role in the development of the lichenoid inflammatory infiltrate that occurs typically in OLP. Copyright

Cutaneous distribution of plasmacytoid dendritic cells in lupus erythematosus. Selective tropism at the site of epithelial apoptotic damage

Recent evidences suggest a significant role of Plasmacytoid dendritic cells (PDC) role in the pathogenesis of lupus erythematosus (LE) via production of type I IFN. Taking advantage on the availability of multiple reagents (CD123, BDCA2, and CD2ap) specifically recognizing PDC on fixed tissues, we investigated the occurrence of PDC in a cohort of 74 LE patients. The large majority of LE biopsies (67/74; 90.5%) showed cutaneous infiltration of PDC. PDC were more frequently observed (96.4 vs 72.2) and numerous in cutaneous LE compared to systemic LE (SLE) and correlated with the density of the inflammatory infiltrate (r=0.40; p<0.001). PDC reduction in SLE might be related to a broader tissue distribution of this cellular population, as indicated by their occurrence in kidneys in 11 out of 24 (45.8%) cases studied. The distribution of cutaneous PDC showed two distinct patterns. More commonly, PDC were observed within perivascular inflammatory nodules in the dermis, associated with CD208+ mature DC and T-bet+ cells [D-PDC]. A second component was observed along the dermal-epithelial junction [J-PDC], in association with cytotoxic T-cells in areas of severe epithelial damage. Notably, chemerin reactivity was observed in 64% of LE biopsies on endothelial cells and in the granular layer keratinocytes. Cutaneous PDC in LE strongly produced type I IFN, as indicated by the diffuse MxA expression, and the cytotoxic molecule granzyme B. This study confirms cutaneous PDC infiltration as hallmark of LE. The topographical segregation in D-PDC and J-PDC suggests a novel view of the role of these cells in skin autoimmunity.

NF‐κB expression in oral and cutaneous lichen planus

Lichen planus (LP) is a chronic inflammatory disorder involving cutaneous and mucosal surfaces, characterized by a T‐cell‐mediated immune response against epithelial cells, with persistent accumulation of T lymphocytes and epithelial cell damage. The mechanisms involved in this chronic inflammatory disease are largely unknown. A pivotal role in the pathogenesis of long‐lasting inflammatory processes is played by the activation of nuclear factor kappa B (NF‐κB), a primary transcription factor which upon translocation to the nucleus, binds to promoter regions of different genes encoding immune and pro‐inflammatory mediators. Using immunohistochemistry, the present study analysed the expression of NF‐κB in 25 cases of cutaneous LP (CLP) and 28 cases of oral LP (OLP) and correlated this with the recruitment of cytotoxic T‐cells (expressing Tia‐1 or perforin) in the inflammatory infiltrate. Nuclear NF‐κB was expressed on basal and suprabasal keratinocytes in all cases of LP, while normal epithelium was consistently negative; OLP contained significantly higher numbers of NF‐κB‐positive keratinocytes than CLP (means: 89.32 versus 22.6; p < 0.05). Furthermore, nuclear NF‐κB expression by epithelial cells correlated with the amount of cytotoxic cell infiltration (p < 0.02). These data suggest that increased NF‐κB activity may represent the basis of maintenance of the inflammatory response. The differences observed between NF‐κB expression on epithelial cells in OLP and CLP and their correlation with the degree of cytotoxic inflammatory infiltrate might explain the different clinical courses of the two variants of the disease, since OLP is typically more recalcitrant than CLP. As proposed for other chronic inflammatory disorders associated with increased NF‐κB activity, the involvement of NF‐κB in the pathogenesis of LP could be considered for selective therapeutic inhibitory targeting. Copyright

Cytotoxic Molecule Expression and Epithelial Cell Apoptosis in Oral and Cutaneous Lichen Planus

We evaluated the expression of T cell-restricted intracellular antigen (Tia-1), granzyme B, and perforin by lymphocytes and the degree of epithelial apoptosis in oral and cutaneous lichen planus (LP) in 51 untreated cases, including 27 oral LP (OLP) and 24 cutaneous LP (CLP) cases. The number of total dermal-positive lymphocytes in OLP and CLP was similar, indicating similar activity of the inflammatory process. Intraepithelial Tia-1-positive, perforin-positive, and granzyme B-positive lymphoid cells were more numerous in OLP than in CLP (P < .05). The epithelial cell apoptotic index (AI) was increased significantly in OLP (P < .05), particularly in erosive-atrophic variants. A linear correlation between AI and the mean +/- SEM number of intraepithelial and dermal perforin+ cells (6.85 +/- 2.44 and 27.48 +/- 10.19, respectively), per 10 high-power fields for OLP and for CLP (1.17 +/- 0.88 and 10.42 +/- 5.74, respectively), was found (intraepithelial, r = 0.50; dermal, r = 0.51; P < .01). These data suggest a pivotal role for perforin in triggering epithelial cell apoptosis. The differences of infiltrating cytotoxic cells and related AI observed in OLP and CLP are in keeping with the clinical behaviors that distinguish these LP variants.

Usefulness of Claudin 4 in the cytological diagnosis of serosal effusions

The identification of metastatic cells in serous effusions has prognostic and therapeutic implications, thus leading to a continuous search for improvement of the existing diagnostic procedures, including immunocytochemistry. To evaluate the usefulness of an antibody recognizing the tight junction‐associated protein Claudin 4 in detecting metastatic tumor cells and in the differential with reactive and neoplastic mesothelium, we stained 345 cases of benign and neoplastic serous effusions obtained from pleura, peritoneum, and pericardium. Two‐hundred and twenty‐eight of 230 cases (99.1%) of epithelial metastasis of different origin were strongly stained by anti‐Claudin 4, whereas all cases of reactive mesothelitis (78) and malignant mesothelioma (37) were negative. With the exception of a single case of ovarian carcinoma hypercalcemic‐type, all tumors originating from the anatomical sites that most frequently metastasize to the serosae, including lung (61), breast (23), female genital tract (67), gastrointestinal tract (27), and peritoneum (6), were found to be positive. Claudin 4 was also extremely useful in detecting single‐tumor cells dispersed among heavy inflammatory reaction. Because of its high sensitivity (99.1%) and specificity (100%), Claudin 4 might be used as an ideal “single‐shot” marker for the identification of metastatic epithelial cells in serous effusions. Diagn. Cytopathol. 2011;39:313–317.

Neoplasms derived from plasmacytoid monocytes/interferon-producing cells: variability of CD56 and granzyme B expression.

To the Editor: We read with interest the paper by Petrella et al on the “agranular CD4+CD56+ hematodermic neoplasm,” which expands data from a previous study by the same group. In the latest WHO classification, this neoplasm is included as a variant of NK cell lymphoma, but the authors retain that it should be considered as a distinct clinicopathologic entity. Primary cutaneous infiltration, bone marrow involvement with or without leukemia, and very poor prognosis represent the cardinal clinical manifestations of the disease. Tumor cells display plasmacytoid morphology, express CD4, CD56, and CD123 (alpha chain of interleukin-3 receptor), but lack conventional pan-myeloid and panlymphoid cell markers. The origin of tumor cells remains uncertain; however, CD4+CD56+ cells have been identified in the peripheral blood of healthy volunteers treated with Flt3 and correspond to a limited fraction of total CD123+ cells, the latter referred to as plasmacytoid monocytes/interferonproducing cells (PM/IPCs). Therefore, Petrella et al concluded that the “agranular CD4+CD56+ hematodermic neoplasm” more likely represents the tumoral counterpart of a subset of PM/IPCs. Recently, Feuillard et al and Chaperot et al reported a series of hematologic neoplasms selected on the basis of their CD123 positivity, which showed many clinical, morphological, and phenotypical features in common with the cases reported by Petrella et al (Table 1); furthermore, tumor cells were found to produce high levels of interferonunder proper stimulation, corroborating their PM/IPC identity. The “agranular CD4+CD56+ hematodermic neoplasm” differs from tumoral proliferations of PM/IPCs originally reported as to “plasmacytoid T-cell lymphoma” and characterized by nodal proliferations of PM/IPCs associated with acute or chronic myeloid neo-

Reflectance confocal microscopy for the in vivo detection of Treponema pallidum in skin lesions of secondary syphilis.

BACKGROUND Secondary syphilis is a diagnostic challenge that relies on microscopic and laboratory tests. OBJECTIVE We sought to assess the usefulness of in vivo reflectance confocal microscopy (RCM) to detect Treponema pallidum in lesions suggestive of secondary syphilis. METHODS Macular and papular skin lesions from 3 patients clinically suggestive of secondary syphilis were imaged by RCM and confirmed by skin punch biopsy. RESULTS In all lesions RCM demonstrated elongated small bright particles with a spiral shape intermingled with the keratinocytes. These features corresponded with immunohistochemical findings that revealed several spirochetes infiltrating the epidermis. LIMITATIONS Unlike immunohistochemistry, RCM did not visualize T pallidum in the dermis and vascular walls because of limited imaging depth. The specificity and sensitivity of this technique need to be assessed. CONCLUSION RCM may be an effective diagnostic tool for in vivo real-time imaging of T pallidum in skin lesions of secondary syphilis, and seems to correlate well with immunohistochemistry.

Placental histological patterns and uterine artery Doppler velocimetry in pregnancies complicated by early or late pre‐eclampsia

To study placental patterns in pregnancies complicated by pre‐eclampsia (PE) and to verify whether the findings are related to gestational age (GA) at PE onset and second‐trimester uterine artery (UtA) Doppler.