Domenico Vitolo

Probiotic administration in patients with ileal pouch–anal anastomosis for ulcerative colitis is associated with expansion of mucosal regulatory cells

Background: Probiotics have anti‐inflammatory effects in patients with inflammatory bowel disease and appear to regulate mucosal immune response through reductions in proinflammatory cytokines. The probiotic VSL#3 prevents pouchitis if started within a week of ileostomy closure and maintains remission following antibacterial treatment in patients with refractory or recurrent pouchitis. However, the efficacy of probiotics and their effects on regulatory cells if started at a greater time after surgery in patients undergoing ileal pouch anal anastomosis (IPAA) for ulcerative colitis are unknown. Methods: We conducted an open‐label study in which 31 patients at different periods from surgery without signs and symptoms of pouchitis were randomized to 2 sachets of VSL#3 once daily or no treatment for 12 months. Pouchitis disease activity index (PDAI) was evaluated at baseline and after 3, 6, and 12 months. The percentage of CD4+ T lymphocytes expressing CD25 and the inactive form of transforming growth factor‐&bgr; [latency‐associated peptide (LAP)] were evaluated at baseline and after 3 and 6 months in peripheral‐blood mononuclear cells and mucosal biopsies. Variation in tissue interleukin‐1&bgr; and Foxp3 mRNA expression was also evaluated. Results: During the study period, VSL#3‐treated patients showed a significant reduction in PDAI score and a significant increase in the percentage of mucosal CD4+CD25high and CD4+ LAP‐positive cells compared with baseline values. Tissue samples at different points showed a significant reduction in IL‐1&bgr; mRNA expression, and a significant increase in Foxp3 mRNA expression. Conclusions: We conclude that VSL#3 administration in patients with IPAA modulates the PDAI and expands the number of mucosal regulatory T cells.

Frequent overexpression of multiple ErbB receptors by head and neck squamous cell carcinoma contrasts with rare antibody immunity in patients

In an effort to elucidate the role of ErbB receptors in human head and neck squamous cell carcinoma (HNSCC), expression abnormalities and subcellular localization of epidermal growth factor receptor (EGFR), ErbB2, ErbB3, and ErbB4 were investigated along with EGF and tenascin by immunohistochemistry in 38 carcinomas as compared to adjacent normal mucosa of 24 cases. Although tumour‐specific overexpression affected each ErbB receptor (EGFR 47%, ErbB2 29%, ErbB3 21%, ErbB4 26%), EGFR abnormalities were most prevalent. The latter, and overexpression of more than two ErbB receptors in the same tumour, which always included EGFR, correlated with metastatic disease. ErbB products were specifically detected on the cell membrane and in the cytoplasm. In contrast, ErbB4 was uniquely localized to the nucleus in 7 carcinomas and a tumour‐derived cell line, indicating a role for regulated intramembrane proteolysis resulting in nuclear ErbB4 translocation in HNSCC. Expression of prototype ligand EGF or low‐affinity stromal activator tenascin correlated significantly with EGFR overexpression, implying chronic EGFR activation. Simultaneous overexpression of additional ErbB receptors in most of these cases suggested recurrent involvement of receptor heterodimers. In spite of frequent ErbB receptor alterations, autologous ErbB serum antibodies were rare, with only 1 of 38 tumour patients exhibiting an ErbB2‐specific immune response. Based on upregulation of several known immunosuppressive molecules, scarcity of ErbB‐specific antibodies is consistent with attenuation of natural tumour‐specific immune responses in HNSCC. Copyright

Co-localization of multiple ErbB receptors in stratified epithelium of oral squamous cell carcinoma.

The expression of all four ErbB receptors was compared by immunohistochemistry, using receptor‐specific polyclonal antisera, in 32 invasive, 11 in situ carcinomas, six benign lesions, and 22 samples of histologically normal mucosa adjacent to specimens of carcinoma originating from oral cavity epithelium. Among invasive and in situ carcinoma, EGFR expression was the most prevalent (in 29/32 and 8/11 cases, respectively) followed by ErbB2 (17/32 and 2/11) and ErbB4 (9/32 and 1/10), while ErbB3 was only detected in invasive tumours (12/32). Specific patterns included invasive tumours with expression of EGFR (8/32) or ErbB4 (1/32) alone, as well as different receptor combinations (EGFR+ErbB2, EGFR+ErbB4, EGFR+ErbB2+ErbB3, EGFR+ErbB2+ErbB4, and all four receptors). Simultaneous expression of three or four ErbB receptors correlated with tumour invasion (p=2.2×10−4) and localized in the intermediate epithelial cell layer of well and moderately differentiated tumours. No other significant correlation with clinico‐pathological features was noticed. Some benign lesions and histologically normal mucosa adjacent to carcinomas showed weak immunostaining of EGFR (10/28), ErbB2 (4/28) or ErbB4 (3/28). By comparison, overexpression, as indicated by increased staining intensity, was observed in invasive tumours for EGFR (18/32), ErbB2 (8/32), ErbB4 (3/32), and ErbB3 (3/32). Statistical evaluation demonstrated a significant association of EGFR or ErbB2 overexpression with invasive carcinoma when compared with benign lesions and apparently normal epithelium (p=5.2×10−7 and p=5×10−3, respectively). Tumour‐specific overexpression of ErbB receptors and their co‐expression, most frequently involving EGFR and ErbB2, in the same cell layer of neoplastic epithelium, implicate receptor heterodimers in the pathogenesis of oral squamous carcinoma. Copyright

Expression of adhesion molecules and extracellular matrix proteins in glioblastomas: relation to angiogenesis and spread

We studied the immunohistochemical expression of inducible adhesion molecules, integrins and extracellular matrix proteins in 10 cases of glioblastoma multiforme in order to investigate their angiogenesis, local invasiveness, poor metastasizing properties and their lack of tumour infiltrating leukocytes. In glioblastomas endothelial proliferations represent the majority of vascular structures; they were positive for endothelial markers (vWF, CD31, VE‐cadherin) and negative for macrophage markers (CD68, PAM‐1). Immunohistologically, they were subtyped into: 1 solid‐glomeruloid ICAM‐1, α2β1, α3β1, α5β1 negative; 2 channelled‐branching ICAM‐1 negative and α2β1, α3β1, α5β1 positive; 3 channelled‐telangiectatic ICAM‐1, α2β1, α3β1, α5β1 positive. In channelled proliferations, the expression and distribution of tenascin and merosin in the basal membrane was similar to that of normal brain vessels. The expression of all these molecules might indicate different steps of maturation of endothelial proliferations. The majority of endothelial proliferations may be immunohistologically considered as incomplete vascular structures; this might account for the low metastasizing tendency and low recruitment of leukocytes by these tumours. Neoplastic astrocytes were GFAP‐1, ICAM‐1, VCAM‐1, α2β1, α3β1 and α5β1 immunoreactive and α6β4 negative; this allows them to interact with extracellular matrix proteins and might, in part, explain the tendency of glioblastomas to infiltrate locally.

Thymoma and thymic carcinoma

Thymic tumors comprise a heterogeneous group of neoplasms with a wide spectrum of clinical presentations. The evolution of the disease is often unpredictable, ranging from an indolent attitude to the possibility of intra- and extrathoracic spread. From the histological point of view, thymoma and thymic carcinoma are the most frequent subtypes and arise only from thymic epithelial cells. Other histological types are even more rare and are usually considered separately. A number of prognostic factors have been validated as predictors of outcome: staging, World Health Organization histological classification, diameter of the tumor, associated paraneoplastic syndromes, completeness of resection, and early onset of recurrence. Complete surgical resection is the key factor for cure and should be considered the gold standard at any stage. Especially for more aggressive lesions, surgery should be considered with a multimodality approach, involving induction and adjuvant therapy according to the stage. Multimodality therapy protocols have been designed based on the integration of clinical staging and histology. Neoadjuvant therapy is now administered before surgical resection in patients with tumors considered inoperable as it improves resectability and survival and reduces the risk of recurrence. Adjuvant treatment has been extensively reported after both complete or partial resection. New targeted therapies are in the developmental stage, and in the future they will be part of the standard protocols. Integrated treatment modalities require strict cooperation between medical and radiation oncologists, thoracic surgeons, and pathologists.

Caveolin‐1 overexpression is associated with simultaneous abnormal expression of the E‐cadherin/α–β catenins complex and multiple erbb receptors and with lymph nodes metastasis in head and neck squamous cell carcinomas

The presence of lymph node metastases is one of the most important prognostic indicators in head and neck squamous cell carcinomas (HNSCCs). An alteration of the E‐cadherin–catenins complex and EGFR is essential for the invasiveness of cancer cells. Caveolin‐1, the major structural protein of the caveolae, represents a scaffolding molecule for several signaling proteins including EGFR. Although caveolin‐1 has been shown to play a role in inducing the invasive phenotype of cancer cells, its role appears to be cell‐type specific and for some tumors it has not been defined yet. In this study we used 57 HNSCC specimens to investigate whether the abnormal expression of caveolin‐1 was associated with the derangement of the E‐cadherin–catenins complex and with the overexpression of ErbB receptors. We demonstrate that in HNSCCs caveolin‐1 overexpression is associated with the simultaneous abnormal expression of at least one member of the E‐cadherin/α–β catenins complex and multiple ErbB receptors as well as with lymph node metastases. We also demonstrate that chronic stimulation of a human hypopharyngeal carcinoma cell line (FaDu) with EGF induced the internalization of β‐catenin and caveolin‐1 and their co‐localization with EGFR. Moreover, EGF treatment induced an increased physical interaction between EGFR/β‐catenin/caveolin‐1 and between E‐cadherin/β‐catenin/caveolin‐1. These molecular events were associated with an increased directional motility of FaDu cells in vitro. These findings may provide new insight into the biology of HNSCC progression and help to identify subgroups of primary HNSCCs with a more aggressive behavior. J. Cell. Physiol. 227: 3344–3353, 2012.

Clinical and immunological findings in four infants with Omenn's syndrome: a form of severe combined immunodeficiency with phenotypically normal T cells, elevated IgE, and eosinophilia.

We report four cases of Omenns syndrome (OS), an autosomal recessive disease characterized by early erythrodermia, protracted diarrhea, severe infections, lymphadenopathy, hepatosplenomegaly, failure to thrive, and leukocytosis with marked eosinophilia. The immunological investigations revealed B lymphopenia with increased levels of serum IgE and marked depression of T-cell activation, not restored by the addition of exogenous interleukin 2 (IL-2). IL-2 and interferon-gamma (IFN-gamma) production in vitro were very low or absent. One patient was treated with HLA-identical bone marrow transplant with a complete remission of the clinical picture and the immunological defect. The infant died of graft versus host disease 4 months after the graft. For the remaining three infants the outcome was also fatal within the first year of life. In conclusion, OS should be considered a severe combined immunodeficiency disease with peculiar clinical, immunological, and histological findings.

Immunoreactivity for S-100 protein in dendritic and in lymphocyte-like cells in human lymphoid tissues*

SummaryS-100 protein is an immunohistochemical marker for a subset of dendritic cells, the interdigitating reticulum cells (IDRCs), which are mainly located in T-dependent areas of lymphoid tissues. In the present study we have investigated the distribution of S-100-positive cells in lymph nodes, spleen, thymus and peripheral blood of normal subjects. Immunoreactivity for S-100 protein was demonstrated in large cells with dendritic morphology and in small lymphocyte-like cells present in the lymph node paracortex, thymic medulla, splenic periarterial lymphatic sheaths (PALS) and in peripheral blood. S-100-positive lymphocyte-like cells were frequently detected around high endothelial venules (HEV) and were present in numbers comparable to those of S-100-positive IDRCs. Immunoelectron microscopy confirmed the existence of positive cells with lymphoid morphology and revealed that the intracellular distribution of the immunoreaction product was similar in lymphoid and dendritic cells. Further characterization of S-100-positive cells demonstrated that both lymphoid and dendritic cells were unreactive with a large panel of monocytic and macrophage markers.

Management of abdominal lymphoproliferative diseases in the era of laparoscopy

BACKGROUND The treatment of lymphoproliferative diseases requires extensive histological, immunohistochemical, and cytogenetic diagnosis. The aim of this study was to analyze the results of 66 laparoscopic procedures in the diagnosis, staging, and restaging of hematological malignancies localized in the abdominal cavity. METHODS Between July 1993 and March 1998, 64 consecutive patients (28 male and 36 female; mean age 46.6 years, range 7 to 69) with diagnosed or suspected lymphoproliferative diseases were referred for primary diagnosis or reassessment and for staging/restaging. RESULTS Sixty-two out of 66 procedures (93.9%) were completed laparoscopically. CONCLUSIONS The minimally invasive approach, in the management of lymphoproliferative diseases, is able not only to provide an adequate specimen for proper diagnostic techniques, but also, when compared with open surgery, to offer a large number of advantages.

Usefulness of the organ culture system when villous height/crypt depth ratio, intraepithelial lymphocyte count, or serum antibody tests are not diagnostic for celiac disease

The existence of mild forms of celiac disease (CD) can make the histology-based diagnosis difficult to reach. Since anti-endomysium (EMA) and anti-tissue transglutaminase (anti-tTG) are detectable in culture supernatants of duodenal biopsies from CD patients, our aim was to assess if this system can support the histology in the diagnostic work-up. A total of 559 suspected CD patients underwent serum EMA/anti-tTG detection, upper endoscopy with duodenal biopsy sampling, histologic analysis, and organ culture to detect EMA/anti-tTG in supernatants. A subgroup of 30 patients with organ culture positive results were put on a gluten-free diet (GFD). Their gluten-dependency was evaluated by the psychological general well-being and beck depression inventory indexes. Statistical analysis was performed by Cohen k inter-test, Friedman test, and Dunn multiple comparison. Two hundred forty-one out of 559 (43.1%) patients showed intestinal villous atrophy, whereas serum and organ culture EMA/anti-tTG were positive in 293/559 (52.4%) and 334/559 (59.7%) patients, respectively. The strength of agreement resulted good for serology vs histology (k = 0.730), good for organ culture vs histology (k = 0.662), and very good for serology vs organ culture (k = 0.852). After 12 months of GFD, psychological general well-being index significantly increased, and beck depression inventory index significantly decreased (P < 0.001 for each one). Data highlight the organ culture system as a useful tool to assist the histology in diagnosing CD, mainly in cases without villous atrophy or in seronegative patients. The marked improvement in quality of life after a GFD further supports the reliability of this system in diagnosing CD.