Evidence for a role of autoinflammation in early-phase psoriasis.

Psoriasis is a common, inflammatory immune-mediated skin disease mainly presenting with plaques whose pathogenesis is based on the central role of the interleukin (IL)-23/IL-17 axis. However, the mechanisms acting in papular lesions of early-phase psoriasis are not fully understood. The aim of this study was to assess the involvement of autoinflammation, a state of sterile inflammation mainly driven by IL-1 over-production that has been recently hypothesized to act in the early phase of disease. Lesional skin of 10 patients with recent onset, untreated psoriasis has been investigated for expression of IL-1ß, IL-17, IL-23 and other cytokines involved in the disease in comparison with normal skin of 10 healthy controls using a protein array method. Immunohistochemical phenotyping of inflammatory infiltrate and co-localization experiments with immunofluorescence confocal microscopy were conducted. IL-1ß was significantly more expressed in psoriasis than in normal skin (P < 0·0001). The chemokine IL-8 was also over-expressed in psoriasis (P = 0·03) while IL-12, IL-17, IL-23, tumour necrosis factor-α and interferon-γ were only slightly more expressed in psoriasis than in normal skin, without reaching statistical significance. The inflammatory infiltrate consisted mainly of neutrophils with a relevant number of macrophages and dendritic cells and only scattered, predominantly T helper 1 lymphocytes. IL-1ß co-localized mainly with CD66b, a neutrophil marker, suggesting that neutrophils were the major source of this cytokine. IL-1ß over-expression in combination with low expression of cytokines that are predominant in late-phase plaque psoriasis may support the role of autoinflammation in early-phase disease, possibly paving the way to randomized trials with IL-1 antagonists.

A novel oncogenic BTK isoform is overexpressed in colon cancers and required for RAS-mediated transformation.

Bruton's tyrosine kinase (BTK) is essential for B-cell proliferation/differentiation and it is generally believed that its expression and function are limited to bone marrow-derived cells. Here, we report the identification and characterization of p65BTK, a novel isoform abundantly expressed in colon carcinoma cell lines and tumour tissue samples. p65BTK protein is expressed, through heterogeneous nuclear ribonucleoprotein K (hnRNPK)-dependent and internal ribosome entry site-driven translation, from a transcript containing an alternative first exon in the 5'-untranslated region, and is post-transcriptionally regulated, via hnRNPK, by the mitogen-activated protein kinase (MAPK) pathway. p65BTK is endowed with strong transforming activity that depends on active signal-regulated protein kinases-1/2 (ERK1/2) and its inhibition abolishes RAS transforming activity. Accordingly, p65BTK overexpression in colon cancer tissues correlates with ERK1/2 activation. Moreover, p65BTK inhibition affects growth and survival of colon cancer cells. Our data reveal that BTK, via p65BTK expression, is a novel and powerful oncogene acting downstream of the RAS/MAPK pathway and suggest that its targeting may be a promising therapeutic approach.

'Indeterminate for malignancy' (Tir3/Thy3 in the Italian and British systems for classification) thyroid fine needle aspiration (FNA) cytology reporting: morphological criteria and clinical impact.

BACKGROUND: The British system (Thy1-5), the Bethesda system for reporting thyroid cytopathology (BSRTC) and the Italian Society of Anatomic Pathology and Cytology (SIAPEC) classification represent the most important international classifications for thyroid cytopathology. Irrespective of the system used, the 'indeterminate' categories are still debated among cytopathologists, particularly with regard to diagnostic criteria, clinical impact of subclassification and role of molecular techniques. AIM: We aimed to find answers to the following questions: Are there shared criteria in cytological preparations that allow the division of indeterminate follicular lesions into subcategories? What is the true clinical impact of this possible subclassification? METHODS: Among 1150 consecutive thyroid fine needle aspiration (FNA) specimens, 80 patients had nodules with a final cytological report of Tir3 (SIAPEC)/Thy3. These 80 cases were re-evaluated and subclassified according to morphological criteria into three groups: pure follicular proliferations, Hürthle cell follicular lesions and atypical proliferations. RESULTS: Sixteen (20%) cases were categorized as pure follicular proliferations, 40 (50%) as Hürthle cell follicular lesions and 24 (30%) as atypical proliferations. Surgery was performed in 57 cases (71%). Cyto-histological correlation showed that follicular adenoma was the most frequent final diagnosis in the cases treated by surgery (24/57, 42%). The overall malignancy rate in the Tir3 category was 28% (16/57). Atypical proliferations were more often malignant than either of the follicular groups (53% versus 19%, P = 0.019). CONCLUSIONS: A five-tiered classification, subdividing the 'indeterminate for malignancy' class into 'follicular proliferations' and 'atypical lesions' could be adopted. As a result of their higher risk of malignancy, surgical management of the atypical lesions would be justified. In future, the introduction of a genetic panel might contribute to their stratification, to the determination of a more accurate risk of malignancy of the atypical lesions and to the verification of follicular proliferations that are benign.

Immunohistochemical expression of Annexin A5 in preeclamptic placentas.

OBJECTIVE: To study the expression of Annexin A5 (A5) in relation to preeclampsia using immunohistochemical Tissue Microarray (TMA) technique. STUDY DESIGN: Case-control study of 66 singleton preeclamptic (PE) patients matched for gestational age (GA) at delivery with 63 normotensive controls with normally grown fetuses. Immunohistochemical expression of A5 and other population characteristics were compared between the two groups using Chi-square, One-way ANOVA, Spearman's Correlation, and Linear Regression. RESULTS: The two groups were similar for maternal age and rate of corticosteroid administration, but differed for nulliparity, Body Mass Index (BMI), blood pressure, presence of placental histological lesions, and placental weight. Expression of A5 was similar in PE and controls (p = 0.10); however it was found to be lower in PE cases complicated by fetal growth restriction (FGR, n = 34) compared with matched controls (n = 55) (p = 0.04). An inverse correlation was found between A5 and GA in cases but not in controls (p = 0.04 vs p = 0.71). The association was even more significant in the subgroup of PE complicated by FGR (p = 0.02). A5 expression was not influenced by blood pressure, proteinuria, or placental weight. CONCLUSIONS: Annexin A5 expression seems to be related only to FGR and not to PE or its clinical severity.

Extensive sclerosing mesenteritis of the rectosigmoid colon associated with erosive colitis.

Sclerosing mesenteritis is a rare, idiopatic, usually benign, inflammatory process of the mesenteric adipose tissue. The most common site of involvement is the small bowel mesentery. We present a case of sclerosing mesenteritis of the rectosigmoid colon as a cause of severe abdominal pain, abdominal obstruction, and ischemic colic mucosal lesions. Contrast enema, colonoscopy, angiography, and CT were the imaging modalities used. A 20 cm diameter, fibrotic mass causing extensive compression of rectosigmoid colon was found at laparotomy. Histological examination showed extended fibrosis, inflammatory cells infiltration, lipophages, and granulomas within the mesenteric adipose tissue associated with erosive colitis. Clinical presentation and treatment are discussed.

Frequency and clinical significance of human beta-herpesviruses in cervical samples from Italian women.

Human papillomaviruses (HPVs) are necessary, but not sufficient, for the development of cervical cancer (CC). Human beta-herpesviruses (beta-HHVs) have been suggested as possible cofactors in the oncogenesis of CC. In this cross-sectional study, the prevalence and possible association of cytomegalovirus (CMV), HHV-6 and -7 with HPV presence was investigated by quantitative real-time PCR assays in cervical samples obtained from 208 italian women. The two most common high-risk HPV types found were 31 and 16. Overall, the positive rates for CMV, HHV-6 and HHV-7 were 66%, 25%, and 6%, respectively. In particular, the prevalence of CMV was found to be extremely high irrespective of either the cytological category or HPV positivity. The prevalence of HHV-6 DNA was significantly higher in high-grade squamous intraepithelial lesions (HSIL) respect to normal women (P < 0.017); by contrast, the prevalence HHV-7 DNA was generally low and not associated with SIL. Copresence of CMV and HHV-6 DNA was found to be significantly higher in patients with SIL respect to normal women (P < 0.05). No correlation was demonstrated between the viral load of all three beta-HHVs and the different cytological stages or with the HPV presence. A few patients with severe disease however showed very high viral loads which for HHV-6 may be indicative of viral integration. In conclusion, this study suggests that CMV and HHV-7 alone are probably not implicated in the oncogenesis of CC whilst HHV-6 alone or together with CMV may contribute to the development of CC.

Prediction of nodal spread of breast cancer by using artificial neural network-based analyses of S100A4, nm23 and steroid receptor expression.

The expression of tumour promoter gene S100A4, metastasis suppressor gene nm23, oestrogen and progesterone receptors, and tumour grade and size have been investigated for their potential to predict breast cancer progression. The molecular and cellular data have been analysed using artificial neural networks to determine the potential of these markers to predict the presence of metastatic tumour in the regional lymph nodes. This study shows that tumour grade and size are poor predictors. The relative expression of S100A4 and nm23 genes is the single most effective predictor of nodal status. Inclusion of oestrogen- and progesterone-receptor status with tumour grade and size markers improves prediction; however, there may be some overlap between steroid receptors and molecular markers. This study also underscores the power of artificial neural network techniques to predict the potential of primary breast cancers to spread to axillary lymph nodes. This could aid the clinician in determining whether invasive procedures of axially node dissection can be obviated and whether conservative forms of treatment might be appropriate in the management of the patient.

Engagement of the leukocyte-associated Ig-like receptor-1 induces programmed cell death and prevents NF-kappaB nuclear translocation in human myeloid leukemias.

Leukocyte-associated Ig-like receptor-1 (LAIR-1) is a surface molecule that functions as an inhibitory receptor on natural killer cells, T lymphocytes and monocytes. Here, we provide evidence that occupancy of LAIR-1 on human myelomonocytic leukemic cell lines inhibits proliferation and leads to programmed cell death (PCD), evaluated by propidium iodide staining and transmission electron microscopy. Interestingly, PCD elicited via LAIR-1 was not blocked by different caspase inhibitors, at variance with apoptosis induced via CD95/Fas, which was prevented by the caspase-1 and caspase-8 specific inhibitors. In addition, we show that the p65 subunit of the nuclear factor kappaB (NF-kappaB), constitutively expressed in the nucleus of these cell lines, was retained in the cytoplasm upon engagement of LAIR-1. This was evident already 8 h after LAIR-1 occupancy, when apoptosis was not yet detectable by fluorometric or ultrastructural analysis. Moreover, a reduction in inhibitor kappaBalpha phosphorylation was observed after LAIR-1 engagement. As blocking of NF-kappaB activation has been shown to rescue sensitivity to anti-cancer drugs in solid tumors, we suggest that LAIR-1 may represent a possible target for pharmacological approaches aimed to potentiate anti-leukemic therapy.

Factors affecting survival and long-term outcome in the cirrhotic patient undergoing hepatic resection for hepatocellular carcinoma.

AIMS: Prognostic analysis of hepatocellular carcinoma (HCC) in the cirrhotic patient undergoing hepatic resection is necessary in order to determine the clinical effect of hepatectomy on prognosis. PATIENTS AND METHODS: Univariate and multivariate retrospective analyses were performed in 51 cirrhotic patients (38 men, 13 women; mean age 65 years, range 43-81 years) with supervening HCC undergoing hepatic resection between January 1993 and December 1997. RESULTS: Segmental liver resection was performed in 39 patients (76%) with non-anatomical (wedge) resections in the remainder of cases. The post-operative mortality rate was 8%. The tumours recurred in 23 patients (45%), with 12 patients (52% of recurrences) recurring within 1 year of surgery and 22 patients (96% of recurrences) within 3 years. Recurrent disease was most frequently intrahepatic (22 patients). Significant risk factors for recurrence were micro/macro vascular invasion, and symptoms. CONCLUSIONS: The recurrence rate of hepatocellular carcinoma in patients with cirrhosis undergoing surgical resection alone is high and actuarial survival at 4 years is low. Other approaches to the treatment of hepatocellular carcinoma in patients with cirrhosis require consideration.

Vitamin D3 affects differentiation, maturation, and function of human monocyte-derived dendritic cells.

We studied the effects of 1alpha,25-dihydroxyvitamin D3 (1alpha, 25-(OH)2D3) on differentiation, maturation, and functions of dendritic cells (DC) differentiated from human monocytes in vitro in the presence of GM-CSF and IL-4 for 7 days. Recovery and morphology were not affected by 1alpha,25-(OH)2D3 up to 100 nM. DC differentiated in the presence of 10 nM 1alpha,25-(OH)2D3 (D3-DC) showed a marked decrease in the expression of CD1a, while CD14 remained elevated. Mannose receptor and CD32 were significantly increased, and this correlated with an enhancement of endocytic activity. Costimulatory molecules such as CD40 and CD86 were slightly decreased or nonsignificantly affected (CD80 and MHC II). However, after induction of DC maturation with LPS or incubation with CD40 ligand-transfected cells, D3-DC showed marginal increases in MHC I, MHC II, CD80, CD86, CD40, and CD83. The accessory cell function of D3-DC in classical MLR was also inhibited. Moreover, allogeneic T cells stimulated with D3-DC were poor responders in a second MLR to untreated DC from the same or an unrelated donor, thus indicating the onset of a nonspecific hyporesponsivity. In conclusion, our data suggest that 1alpha,25-(OH)2D3 may modulate the immune system, acting at the very first step of the immune response through the inhibition of DC differentiation and maturation into potent APC.

Effects of cryopreservation on in vitro and in vivo long-term function of human islets.

BACKGROUND: The possibility of performing transplantation several days after explant seems to be a peculiarity of islet grafts, and the opportunity to cryopreserve human islets may permit an indefinite period for modulating the recipient immune system. The aim of the present study was the evaluation of in vitro and in vivo functional properties of cryopreserved human islets. METHODS: We used six consecutive human islet preparations not suitable for an immediate transplantation in diabetic patients because the limited islet mass separated. The in vitro function of cryo and fresh islets was studied by determination of insulin and glucagon secretion in response to such classical stimuli as glucose (16.7 mM), glucose (16.7 mM) + 3-isobutyl-1-methylxanthine (0.1 mM), arginine (10 mM), and tolbutamide (100 microM). In vivo islet function was assessed through intravenous glucose tolerance tests performed at 15, 30, 60, and 90 days after transplantation of 1000 hand-picked fresh or cryopreserved islets in nude mice. RESULTS: Basal secretion of true insulin was significantly higher in cryopreserved islets than in fresh ones. The response of cryopreserved islets to arginine and glucose + isobutyl-1-methylxanthine seemed partially impaired. Proinsulin-like molecule secretion seemed higher in cryopreserved than in fresh islets in response to all secretagogues used, and the difference was statistically significant for arginine. The capacity of human cryopreserved islets to maintain a correct metabolic control in diabetic nude mice was progressively lost in 3 months. CONCLUSIONS: These findings showed that cryopreservation affects the function of isolated human islets, maintaining in vivo function for a limited period of time.

Glucocorticoids increase the endocytic activity of human dendritic cells.

We have investigated the effect of glucocorticoids (GC) on antigen uptake molecule expression and on endocytic activity of human dendritic cells (DC). Human monocyte-derived DC were differentiated in vitro for 7 days with granulocyte macrophage colony stimulating factor and IL-4 in the presence or absence of dexamethasone 10(-8) M (Dex). Dex-treated DC showed an enhancement of mannose receptor (MR)-mediated endocytosis (measured as uptake of FITC-dextran) and of fluid-phase endocytosis [measured as uptake of Lucifer yellow (LY)] The effect was dose dependent and correlated with the length of exposure to Dex. The expression of receptors involved in antigen capture was investigated by FACS analysis. Dex up-regulates MR, CD16 and CD32 expression on DC. After maturation with tumor necrosis factor-alpha or CD40 ligand in Dex-treated DC, despite a reduction induced by maturation the endocytic activity of FITC-dextran and LY, the expression of MR, CD16 and CD32 remained higher than in control DC. In view of the fact that antigen capture was increased in cells cultured with Dex, we evaluated the ability to present soluble antigen that needs to be taken up and processed. Cells differentiated in the presence of Dex showed much lower efficiency in presenting tetanus toxin to specific autologous T cell lines. In conclusion our data suggest a new mechanism by which GC may influence immune responses. In fact with the increase in endocytic activity, Dex favors the scavenging of antigen from the external milieu, decreasing antigen concentration and availability, and simultaneously inhibiting the capacity to stimulate T cells.

Glucocorticoids affect human dendritic cell differentiation and maturation.

Because dendritic cells (DC) play a major role in the initiation of T cell-mediated immunity, we studied the effects of glucocorticoids, well-known inhibitors of the immune and inflammatory response, on the differentiation and maturation of human DC. DC were differentiated from human monocytes by culture with GM-CSF and IL-4 for 7 days with and without dexamethasone (Dex). Cells treated with Dex (10-8 M) (Dex-DC) developed a characteristic dendritic morphology; however, membrane phenotype analysis demonstrated that they were not fully differentiated. Dex-DC expressed low levels of CD1a and, unlike untreated cells, high levels of CD14 and CD16. Molecules involved in Ag presentation (CD40, CD86, CD54) were also impaired. In contrast, molecules involved in Ag uptake (mannose receptor, CD32) and cell adhesion (CD11/CD18, CD54) were up-regulated. After exposure to TNF-alpha or CD40 ligand, Dex-DC expressed lower levels of CD83 and CD86 than untreated cells. Dex-DC showed a higher endocytic activity, a lower APC function, and a lower capacity to secrete cytokines than untreated cells. Overall, these results indicate that DC differentiated in the presence of Dex are at a more immature stage. Moreover, Dex also partially blocked terminal maturation of already differentiated DC. In conclusion, our data suggest that glucocorticoids may act at the very first step of the immune response by modulating DC differentiation, maturation, and function.

Human proinsulin production in primary rat hepatocytes after retroviral vector gene transfer.

The development of autologous somatic cells, engineered for the synthesis and release of human insulin under physiological stimuli, would certainly represent a major breakthrough in the therapy of insulin-dependent diabetes mellitus. We generated a retroviral vector containing the human proinsulin cDNA and the gene coding for the human nerve growth factor receptor for quantitative analysis of transduced cells. Primary rat hepatocytes were selected as target cells because of the constitutive expression of the pancreatic beta-cell glucose transporter GLUT-2 and the glycolitic enzyme glucokinase. Appropriate conditions for culture and retroviral transduction are described. The highest transduction efficiency, evaluated as percentage of LNGFr expressing cells was obtained by repeated infection cycles (40+/-10%). Human proinsulin accumulated in the culture medium of transduced rat hepatocytes (mean+/-SD): 18.1+/-7.9 (range 8.7-36.4) ng/24h/10(6) cells. Primary rat hepatocytes can be efficiently transduced by a retroviral vector and the de novo synthesis of human proinsulin can be induced. Primary cultured hepatocytes represent an useful model to test retroviral constructs engineered for the glucose-inducible expression of insulin under the control of liver-specific promoters.

[Hemangiopericytoma-like tumor (HPCLT) of the nasal cavity: morphological, immunohistochemical, and ultrastructural features]. / Tumore simil emangioperitocitoma (HPCLT) della cavità nasale: aspetti morfologici, immunoistochimici ed ultrastrutturali.

A case of hemangiopericytoma-like tumour (HPCLT) of the nasal cavity is described in a 81-year-old woman. The tumour, accompanied by symptoms of nasal obstruction and epistaxis, presented as a polypoid lesion filling the left middle meatus region. Histological, immunohistochemical and electron microscope studies revealed a uniform sheet-like proliferation of spindle to oval vimentin-positive cells growing around a rich vascular network. The favourable outcome sixteen months after endoscopic sphenoethmoidectomy would seem to confirm the diagnosis of HPCLT. The differential diagnosis of this rare lesion is discussed.

Clinical presentation, diagnosis and survival of resected distal bile duct cancer.

The aim of the study was the definition of the clinical features and survival of 27 resected cases of distal bile duct carcinoma. This neoplasm accounted for 14% of all periampullary malignancies treated by pancreaticoduodenectomy between 1990 and 1996. Jaundice was present in 96% of patients, but was the first symptom only in 78%. Preoperative investigations allowed to recognize distal bile duct cancer in a minority of patients (41%). Operative mortality and morbidity were 3.7 and 44%, respectively. Most patients (88%) were assigned to UICC stage IV-A. Postoperative survival was not significantly better than survival of 101 patients undergoing pancreaticoduodenectomy for pancreatic ductal carcinoma; median survival was 22 months, with a 13% 5-year survival rate. Determinants of a better prognosis were UICC stage

Expression of metastasis-associated genes h-mts1 (S100A4) and nm23 in carcinoma of breast is related to disease progression.

The murine 18A2/mts1 and its human homolog h-mts1 (S100A4), encoding a Ca2+-binding protein belonging to the S-100 family, are associated with high invasive and metastatic potentials of murine tumors, human tumor cell lines in vitro, and human tumors growing as xenografts. The nm23 is a putative metastasis-suppressor gene whose expression has been found to correlate inversely with the metastatic potential of some forms of human cancer. The products of both human genes alter cytoskeletal dynamics, with antagonistic effects. In view of the equivocal association of nm23 with the metastatic potential of human cancer, we suspected that the relative expression of h-mts1 and nm23 might reflect tumor progression more accurately than either of them alone. We describe here the expression of these genes in infiltrating ductal carcinomas of the breast and show that high h-mts1 expression is associated with metastatic spread to the regional lymph nodes. The expression of nm23 on its own did not show a statistically significant inverse correlation with nodal spread. However, the expression status of the two genes, taken together, correlated strongly with the occurrence of nodal metastases. Breast cancers with no detectable expression of h-mts1 were found to be estrogen and progesterone receptor positive. Expression of h-mts1 was not related to tumor differentiation. The clinical data, together with the state of expression of steroid receptors and the expression levels of h-mts1 and nm23 genes, were analyzed using artificial neural networks for accuracy in predicting nodal spread of the carcinomas. These analyses support the conclusion that, overall, h-mts1 expression appears to be associated with and indicative of more aggressive disease. Complemented with nm23, h-mts1 could provide a powerful marker of breast cancer prognosis.