D2-40 negative pyogenic granuloma-like Kaposi's sarcoma: Diagnostic features and histogenetic hypothesis of an uncommon skin tumor in HIV-negative patients.

Pyogenic granuloma-like Kaposi's sarcoma (PGLKS) is a recently described skin tumor showing features both of pyogenic granuloma (PG) and Kaposi's sarcoma (KS). The differential diagnosis is often challenging. We reviewed a series of 50 PG and 23 Ks located on distal extremities with the aid of an immunohistochemical panel comprising CD34, CD31, FVIII, SMA, D2-40, HHV8. After revision, 6/50 PG lesions previously diagnosed as PG, showed positive immunostaining for LNA1-HHV8 and focal positivity for CD31 and FVIII in the endothelial cells of the proliferating vessels, with some SMA positive pericytes. D2-40, a marker of lymphatic endothelium positive in KS, stained negatively. These lesions were renamed PGLKS. Of note, in our series, PGLKS represented the only form of KS localized in the hand; all the patients were HIV-negative, older than PG patients, with a prevalence for male gender. PGLKS and PG need a different management and a follow-up is advisable for PGLKS, as for the other variants of KS. To date, D2-40 negative immunostaining has not yet been reported in PGLKS and should not lead to a misdiagnosis of PG. The morphological similarities with PG and the immunohistochemical findings, showing a defective phenotype of the neoplastic cells, suggest a histogenetic hypothesis in which D2-40 negative PGLKS could represent an early stage of HHV8 infection of a pre-existing PG, whose vessels loose progressively their blood vascular markers but have not still acquired the lymphatic ones.

The interleukin (IL)-31/IL-31R axis contributes to tumor growth in human follicular lymphoma.

Interleukin (IL)-31A binds to an heterodimer composed of IL-31 receptor A (IL-31RA) and Oncostatin M Receptor (OSMR). The IL-31/IL-31R complex is involved in the pathogenesis of various skin diseases, including cutaneous T-cell lymphoma. No information is available on the relations between the IL-31/IL-31R complex and B-cell lymphoma. Here we have addressed this issue in follicular lymphoma (FL), a prototypic germinal center(GC)-derived B-cell malignancy. IL-31 enhanced primary FL cell proliferation through IL-31R-driven signal transducer and activator of transcription factor 1/3 (STAT1/3), extracellular signal-regulated kinase 1/2 (ERK1/2) and Akt phosphorylation. In contrast, GC B cells did not signal to IL-31 in spite of IL-31R expression. GC B cells expressed predominantly the inhibitory short IL-31RA isoform, whereas FL cells expressed predominantly the long signaling isoform. Moreover, GC B cells lacked expression of other IL-31RA isoforms potentially involved in the signaling pathway. IL-31 protein expression was significantly higher in surface membrane than in cytosol of both FL and GC B cells. IL-31 was detected in plasma membrane microvesicles from both cell types but not released in soluble form in culture supernatants. IL-31 and IL-31RA expression was higher in lymph nodes from FL patients with grade IIIa compared with grade I/II, suggesting a paracrine and/or autocrine role of IL-31/IL-31RA complex in tumor progression through microvesicle shedding.

Immunohistochemical/histochemical double staining method in the study of the columnar metaplasia of the oesophagus.

Intestinal metaplasia in Barrett's oesophagus (BO) represents an important risk factor for oesophageal adenocarcinoma. Instead, few and controversial data are reported about the progression risk of columnar-lined oesophagus without intestinal metaplasia (CLO), posing an issue about its clinical management. The aim was to evaluate if some immunophenotypic changes were present in CLO independently of the presence of the goblet cells. We studied a series of oesophageal biopsies from patients with endoscopic finding of columnar metaplasia, by performing some immunohistochemical stainings (CK7, p53, AuroraA) combined with histochemistry (Alcian-blue and Alcian/PAS), with the aim of simultaneously assess the histochemical features in cells that shows an aberrant expression of such antigens. We evidenced a cytoplasmic expression of CK7 and a nuclear expression of Aurora A and p53,  both in goblet cells of BO and in non-goblet cells of CLO, some of which showing mild dysplasia. These findings suggest that some immunophenotypic changes are present in CLO and they can precede the appearance of the goblet cells or can be present independently of them, confirming the conception of BO as the condition characterized by any extention of columnar epithelium. This is the first study in which a combined immunohistochemical/histochemical method has been applied to Barrett pathology.

Human dental pulp cell apoptosis: immunohistochemical study after applying orthodontic traction.

The aim of this study was to compare human dental pulp stress and programmed cell death after 3 and 6 months of orthodontic treatments by assessing the degree of apoptosis and related proteins. Human dental pulps were collected from twenty young patients orthodontically treated by Straight Wire technique. Samples were fixed, paraffin-embedded and processed for histology and immunohistochemistry using anti-heat shock protein 60 kDa (Hsp60), -caspase 3, -caspase 9, and -PCNA antibodies, as well as TUNEL reactions. Moreover, we performed immunoprecipitation for Hsp60 and caspase 3, and for Hsp60 and caspase 9, from paraffin extracted tissues. Increased levels of both caspases and Hsp60 occurred in 6-months treated samples; at the same time, we found increased levels of proliferating cell nuclear antigen and terminal deoxynucleotidyl transferase dUTP nick end labeling positive cells. Immunoprecipitation showed that Hsp60 forms a complex with both Pro-caspase 3 and Caspase 3, and this may accelerate Pro-caspase 3 activation, especially in the 6-months treated group. On the contrary, no complex between Hsp60 and Pro-caspase 9 was detected. The orthodontic tractions may be a cause of stress, apoptosis and proliferation in pulp tissue. These results suggest the need of further studies about the effects of long term orthodontic treatments on the dental pulp.

P-selectin glycoprotein ligand-1 as a potential target for humoral immunotherapy of multiple myeloma.

Monoclonal antibodies (mAbs), successfully adopted in the treatment of several haematological malignancies, have proved almost ineffective in multiple myeloma (MM), because of the lack of an appropriate antigen for targeting and killing MM cells. Here, we demonstrate that PSGL1, the major ligand of P-Selectin, a marker of plasmacytic differentiation expressed at high levels on normal and neoplastic plasma cells, may represent a novel target for mAb-mediated MM immunotherapy. The primary effectors of mAb-induced cell-death, complement-mediated lysis (CDC) and antibody-dependent cell-mediated cytotoxicity (ADCC), were investigated using U266B1 and LP1 cell-lines as models. Along with immunological mechanisms, the induction of apoptosis by PSGL1 cross-linking was assessed. The anti-PSGL1 murine mAb KPL1 induced death of MM cells in a dose- and time-dependent fashion and mediated a significant amount of ADCC. KPL1 alone mediated C1q deposition on target cells but proved unable to induce CDC due to inhibition of the lytic activity of complement by membrane complement regulators (mCRP) expressed on the cell surface. Consistently, CDC was induced by KPL1 upon mCRP blockage. Our results suggest a role for PSGL1 in MM humoral immunotherapy and support further in vivo studies assessing the effects of anti-PSGL1 mAbs on MM growth and interaction with the bone marrow microenvironment.

Different immunophenotypical apoptotic profiles characterise megakaryocytes of essential thrombocythaemia and primary myelofibrosis.

AIMS: Essential thrombocythaemia (ET) and primary myelofibrosis (PMF) share some clinical and pathological features, but show different biological behaviour and prognosis. The latest contributions to understanding the nature of these disorders have focused on bone marrow microenvironment remodelling and proliferative stress, recognising megakaryocytes (MKCs) as "key-cells". The aim of this study was to investigate the apoptotic profile of ET and PMF MKCs in order to further characterise the biology of these disorders. METHODS: Bone marrow biopsy samples from 30 patients with ET, and 30 patients with PMF, were immunophenotypically studied for the expression of pro-apoptotic (Fas, Fas-L, Bax, Bad) and anti-apoptotic (Bcl-2, Bcl-XL, hTERT (human telomerase reverse transcriptase)) molecules and the "executioner" molecule caspase-3. The fraction of MKCs undergoing apoptosis was assessed by deoxynucleotidyl transferase-mediated dUTP nick-end labelling. RESULTS: Only the mitochondrial pathway seemed to be involved in MKC apoptosis. The anti-apoptotic molecule Bcl-XL was predominantly found in ET MKCs (50.5% of ET MKCs versus 35% of PMF MKCs; p = 0.036), while pro-apoptotic molecules Bax and Bad showed a prevalent expression in PMF MKCs (30.5% of ET MKCs versus 55% of PMF MKCs; 41% of ET MKCs versus 52% of PMF MKCs; p = 0.001 and p = 0.068, respectively). A significant fraction of PMF MKCs were committed to apoptosis according to caspase-3 expression and TUNEL, while only few ET cells were committed to apoptosis. hTERT was significantly more expressed in PMF (32% of ET MKCs versus 46% of PMF MKCs; p = 0.022), in agreement with the proliferative nature of this disease. CONCLUSIONS: It was found that ET and PMF MKCs, which barely differ in terms of morphology and aggregation, are characterised by markedly different apoptotic profiles. The rather high apoptotic fraction of PMF was able to support the fibrotic nature of this process, while the anti-apoptotic profile of ET cells fits well with their "steady" maturative state.