Human equilibrative nucleoside transporter 1 and carcinoma of the ampulla of Vater: expression differences in tumour histotypes.

The human equilibrative nucleoside transporter 1 (hENT1) is the major means by which gemcitabine enters human cells; recent evidence exists that hENT1 is expressed in carcinoma of the ampulla of Vater and that it should be considered as a molecular prognostic marker for patients with resected ampullary cancer. Aim of the present study is to evaluate the variations of hENT1 expression in ampullary carcinomas and to correlate such variations with histological subtypes and clinicopathological parameters. Forty-one ampullary carcinomas were histologically classified into intestinal, pancreaticobiliary and unusual types. hENT1 and Ki67 expression were evaluated by immunohistochemistry, and apoptotic cells were identified by the terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate biotin nick end labelling (TUNEL) method. hENT1 overexpression was detected in 63.4% ampullary carcinomas. A significant difference in terms of hENT1 and Ki67 expression was found between intestinal vs. pancreaticobiliary types (P=0.03 and P=0.009 respectively). Moreover, a significant statistical positive correlation was found between apoptotic and proliferative Index (P=0.036), while no significant correlation was found between hENT1 and apoptosis. Our results on hENT1 expression suggest that classification of ampullary carcinoma by morphological subtypes may represent an additional tool in prospective clinical trials aimed at examining treatment efficacy; in addition, data obtained from Ki67 and TUNEL suggest a key role of hENT1 in tumour growth of ampullary carcinoma.

Effect of ursodeoxycholic acid on inflammatory infiltrate in gallbladder muscle of cholesterol gallstone patients.

BACKGROUND: Reduced gallbladder (GB) contractility and chronic inflammatory changes in the mucosa have been reported in patients with cholesterol gallstones (GS). Ursodeoxycholic acid (UDCA) restores GB contractility and antagonises liver macrophage activation. In the colon, hydrophobic bile acid, not hydrophilic UDCA, induces mast cell degranulation. We studied the presence of monocyte/macrophage infiltrate, cyclooxygenase (COX)-2, inducible nitric oxide synthase (iNOS) expression, the number of total and degranulated mast cells in the GB muscle layer of cholesterol GS patients, and the effect of UDCA administration. METHODS: Gallbladder tissue was obtained from cholesterol GS patients, either treated or untreated with UDCA (10 mg kg(-1) day(-1)) for 30 days prior to surgery. Gallbladders removed for neoplastic diseases, not involving GB, were evaluated for control purposes. The presence of monocytes/macrophages (CD68 positive), granulocytes, and mast cells, and the COX-2 and iNOS expression, was determined immunohistochemically. KEY RESULTS: The number of CD68, granulocytes, mast cells, COX-2 and iNOS positive cells was significantly higher in the muscle layer of GS patients than in controls. Compared to untreated patients, those treated with UDCA showed significantly lower levels of CD68, COX-2 positive cells and degranulated mast cells and a lesser number of iNOS positive cells and granulocytes. CONCLUSIONS & INFERENCES: An inflammatory monocyte/macrophage, mast cell and granulocyte infiltrate is present in the GB muscle layer of GS patients. Ursodeoxycholic acid decreases macrophages, degranulated mast cells and COX-2 expression. These results suggest that monocytes/macrophages and degranulating mast cells contribute to muscle cell dysfunction in cholesterol GS patients and support the anti-inflammatory effect of UDCA.

Characteristics at haematoxylin and eosin staining of ruptures of the long head of the biceps tendon.

OBJECTIVE: To examine the relative prevalence of histological changes that have been found to be associated with the process of tendinopathy in lesions of the tendon of the long head of the biceps brachii and to evaluate the reliability of histopathological evaluation of tendon tissue in lesions of the tendon of the long head of the biceps. DESIGN: Tendon samples were taken from 51 patients (31 men, 20 women; mean age 63.2 years) who underwent arthroscopic release of the long head of the biceps tendon because of refractory biceps tendinopathy and from 5 male patients who died of cardiovascular events (mean age 69.6 years). Histological examination was performed using haematoxylin and eosin staining of sections, which were interpreted using a semiquantitative grading scale assessing fibre structure and arrangement, rounding of the nuclei, regional variations in cellularity, increased vascularity, decreased collagen staining and hyalinisation. RESULTS: The mean (SD) pathological sum score of ruptured tendons was greater than that of control tendons (15.76 (3.11) vs 3.4 (1.9), p<0.001). Within each specific category of tendon abnormalities, the chi(2) test showed significant differences between the control and ruptured tendons; all the variables were significantly different (Mann-Whitney U test 0.05, p<0.001). Using the kappa statistic, the agreement between the two readings ranged from 0.53 to 0.85. CONCLUSIONS: Unruptured tendons of the long head of the biceps, even at an advanced age and ruptured tendons of the long head of the biceps are clearly part of two distinct populations.

Promyelocytic leukemia (PML) gene expression is a prognostic factor in ampullary cancer patients.

BACKGROUND: Promyelocytic leukemia (PML) tumor suppressor gene plays a key role in acute PML pathogenesis but its involvement in pathogenesis and prognosis of solid cancers has not been defined yet. PATIENTS AND METHODS: In all, 62 ampullary adenocarcinoma patients who underwent curative surgery between 1996 and 2005 were included. Expression analysis of PML was carried out by immunohistochemical staining and correlated with disease-free survival (DFS) and overall survival (OS). RESULTS: In 24 tumor specimens (38.7%), PML was classified as absent, in 16 (25.8%) as focally expressed and in 22 (35.5%) as diffusely expressed. By univariate analysis, DFS was significantly influenced by pathological T stage (P=0.03), lymph nodal involvement (P=0.002), and PML expression (P=0.001). DFS in patients without PML expression was 28.0 months versus 45.1 and 75.5 for patients with focal and diffuse expression, respectively. OS in the group of patients without PML expression, with focal expression, and with diffuse expression was 40, 48, and 77 months, respectively (P=0.002). By a multivariate analysis, PML expression was the strongest prognostic factor for DFS (P=0.003) and the only statically significant prognostic factor for OS (P=0.009). CONCLUSIONS: Our preliminary data suggest PML as a novel prognostic tool for ampullary cancer patients.

Human equilibrative nucleoside transporter 1 (hENT1) protein is associated with short survival in resected ampullary cancer.

BACKGROUND: Gemcitabine is an acceptable alternative to best supportive care in the treatment of advanced biliary tract cancers. The human equilibrative nucleoside transporter 1 (hENT1) is a ubiquitous protein and is the major means by which gemcitabine enters human cells. Moreover, recent reports indicate a significant correlation between immunohistochemical variations of hENT1 in tumor samples and survival after gemcitabine therapy in patients with solid tumors. MATERIALS AND METHODS: We used immunohistochemistry to assess the abundance and distribution of hENT1 in tumor samples from radically resected cancer of the ampulla, and sought correlations between immunohistochemical results and clinical parameters including disease outcomes. RESULTS: In the 41 individual tumors studied, 12 (29.3%) had uniformly high hENT1 immunostaining. Statistical analysis showed a significant correlation between hENT1 and Ki-67 (P = 0.04). No statistical significant differences were found between immunohistochemical findings and patient characteristics (sex, age, and tumor-node-metastasis). On univariate analysis, hENT1 and Ki-67 expression were associated with overall survival (OS). Specifically, those patients with overexpression of hENT1 showed a shorter OS (P = 0.022) and those with high Ki-67 staining showed a shorter survival (P = 0.05). CONCLUSIONS: hENT1 expression is a molecular prognostic marker for patients with resected ampullary cancer and holds promise as a predictive factor to assist in chemotherapy decisions.

COX-2 expression in lobular in situ neoplasia of the breast: correlation with histopathological grading system according to the Tavassoli classification.

AIMS: There is considerable evidence to link cyclooxygenase (COX)-2 to the development of cancer. The aim of this study was to assess COX-2 expression and its subcellular localization in lobular in situ neoplasia (LIN) of the breast and to verify differences in COX-2 expression between different grades of lesions according to the Tavassoli classification. METHODS AND RESULTS: We analysed the expression of COX-2 protein by immunohistochemistry in tissue samples of 51 LIN lesions classified into three grades according to the Tavassoli classification. COX-2 immunostaining was observed in 78.4% of LIN samples and showed a prevalent membranous rather than cytoplasmic pattern. COX-2 was expressed in 16/17 (94.1%) LIN1, 22/25 (88%) LIN2 and 2/9 (22.2%) LIN3. As regards COX-2 expression, a statistically significant difference was found between LIN1 and LIN3 (P = 0.001) and between LIN2 and LIN3 (P =0.001). No difference was found between LIN1 and LIN2. Moreover, a significant negative correlation was found between LIN grade and COX-2 expression (P < 0.0001). CONCLUSIONS: COX-2 is highly expressed in LIN, supporting a role for this protein in the early stage of breast carcinogenesis, representing the rationale for using COX-2 selective inhibitors in the earliest stages of breast cancer.

Tumor regression in mesorectal lymphnodes after neoadjuvant chemoradiation for rectal cancer.

AIMS: The histological modification produced by neoadjuvant chemoradiation on primary rectal cancer has been investigated by many authors, and a prognostic value of tumor regression grade (TRG) has been identified. Tumor regression grade on metastatic mesorectal lymphnodes has been never evaluated. The purpose of this study is to analyse the TRG on mesorectal lymphnodes (lymphnode regression grade, LRG) after preoperative chemoradiation in rectal cancer patients and to determine the correlation with TRG of primary tumor. METHODS: Surgical specimens from 35 patients who underwent chemoradiation were included. LRG on mesorectal lymphnodes was assessed by immunohistochemistry. Response to treatment was evaluated by a 5-point LRG based on the ratio of residual tumor to fibrosis. RESULTS: Complete pathologic response (LRG 1) was observed in 18 patients (51%). In 4 patients (11%) no regression was observed (LRG 5). In 4 cases only reactive lymphnodes were found. LRG on lymphnodes significantly correlated with TRG on primary tumor (p<0.05). CONCLUSIONS: Neoadjuvant chemoradiation determines a tumor regression on mesorectal lymphnodes as on primary tumor; further studies are needed to evaluate the prognostic value of LRG.

Dilated intercellular spaces and acid reflux at the distal and proximal oesophagus in patients with non-erosive gastro-oesophageal reflux disease.

BACKGROUND: Acid exposure of proximal oesophagus and dilated intercellular space diameters of oesophageal epithelium are relevant in the perception of gastro-oesophageal reflux. AIM: To explain the relationship between gastro-oesophageal reflux disease symptoms, acid exposure and intercellular space diameter along the oesophageal epithelium and to assess time-related variability of intercellular space diameter. METHODS: Thirty-three non-erosive reflux disease (NERD), six erosive oesophagitis patients and 12 asymptomatic controls underwent oesophageal manometry and 24-h dual-channel oesophageal pH-monitoring following endoscopy. Biopsies were taken 5 cm above the LES and 10 cm below the UES, at comparable levels, as pH sensors. A total of 100 intercellular space diameters per patient/control were measured blindly at transmission electron microscopy. In 15 patients, the investigation was repeated after 1 year. RESULTS: In all NERD patients, acid exposure was higher at mid-proximal oesophagus (P < 0.01) and mean intercellular space diameters, at distal and mid-proximal oesophagus, was three- and twofold higher (1.5 and 0.82 micro m, respectively) compared with controls. Intra-patient intercellular space diameter values were stable over time, not overlapping with those of controls. CONCLUSIONS: Dilation of intercellular space diameter occurs along the distal and proximal oesophageal epithelium in NERD patients and could be responsible for the enhanced perception of proximal acid reflux. This finding appears to be time-reproducible and to represent a sensitive, histopathological marker of NERD.

Complementary use of local excision and transanal endoscopic microsurgery for rectal cancer after neoadjuvant chemoradiation.

BACKGROUND: Neoadjuvant therapies have significantly improved local control and survival of patients with rectal cancer. Nevertheless, although a complete pathologic response can be achieved in 30% of cases, a transabdominal surgical resection is always required. This study aimed, for the first time, to test in the literature the feasibility of local excision combined with transanal endoscopic microsurgery (TEM) as a surgical option for patients treated with neoadjuvant chemoradiation. METHODS: Between July 1997 and December 2002, 30 patients with rectal cancer affected by an extraperitoneal tumor entered a protocol consisting of neoadjuvant chemoradiation followed by surgery. The surgical treatment, consisting of open surgery, local excision, or TEM, was planned according to the patient's clinical response after chemoradiation and distance from the anal verge. RESULTS: A significant clinical downstaging was observed in eight patients. Five of these patients underwent TEM, and three had local excision. Consequently, open surgery was performed for 22 patients. Histology showed six cases of complete pathologic response: three in the open surgery group and three in the transanal excision group. After a mean follow-up period of 47 months, the disease-free survival rate was 77% in the open surgery group and 100% in TEM or local excision group. CONCLUSIONS: The findings suggest the complementary feasibility of TEM and local excision after neoadjuvant chemoradiation. However, randomized trials are needed to confirm the oncologic safety of this approach.

Subcellular shift of the hepatic growth hormone receptor with progression of hepatitis C virus-related chronic liver disease.

AIMS: To evaluate the cytoplasmic and nuclear expression of hepatic growth hormone receptor (GHR) in different stages (S0, S1, S3 and S4, according to Knodell's classification) of chronic liver disease (CLD) and in hepatocellular carcinoma (HCC). METHODS AND RESULTS: Liver specimens from 31 patients with hepatitis C virus-related CLD, five patients with HCC and nine controls were examined for expression of hepatic GHR by immunohistochemistry with MAb 263. Cytoplasmic and nuclear staining were evaluated as a percentage of positively stained cells. The cytoplasmic expression of GHR was comparable between normal liver and S0 hepatitis, while it progressively decreased in S1, S3 and S4 CLD (P < 0.01). Conversely, nuclear GHR showed increased expression in S3 and S4 CLD (P < 0.05). No differences were observed between HCC and normal liver in terms of GHR immunoreactivity. CONCLUSIONS: This is the first study to show that the subcellular expression of hepatic GHR changes with the progression of CLD. The increase in nuclear expression of GHR with advanced stages of CLD suggests that GH may act directly at the nuclear level to promote hepatocyte proliferation/regeneration.

Cetuximab and irinotecan as third-line therapy in advanced colorectal cancer patients: a single centre phase II trial.

The epidermal growth factor receptor (EGFR), which participates in signalling pathways that are deregulated in cancer cells, is frequently mutated in colorectal-cancer cells. Cetuximab is a monoclonal antibody that specifically blocks the EGFR. We evaluated the efficacy of cetuximab in weekly combination with irinotecan in metastatic colorectal cancer patients refractory to previous treatments based on oxaliplatin or irinotecan. We included 55 heavily pretreated patients (colon/rectum: 34/11, M/F: 16/29, median age 63 years, range: 27-79) whose disease had progressed during or within an oxaliplatin-based first-line chemotherapy and a irinotecan-based second-line regimen. Patients were followed for tumour response and were also evaluated for the time to tumour progression, and safety of treatment. Cetuximab was given at an initial dose of 400 mg m(-2), followed by weekly infusions of 250 mg m(-2). Irinotecan was administered weekly at the dose of 90 mg m(-2). All patients were assessable for treatment efficacy and safety response rate was 25.4% (95% CI: 21.7-39.6%); 38.2% (95 CI: 18.6-39.8%) of patients showed a disease stability as the best response. As a consequence, the overall tumour control rate was 63.6% (95% CI: 46.4-70.6%). The median time to progression was 4.7 months (95% CI: 2.5-7.1 months) and the median survival time was 9.8 months (95% CI: 3.9-10.1 months). The most common G3-4 noncutaneous side toxicities were: diarrhoea (16.4%), fatigue (12.7%) and stomatitis (7.3%). 89.1% of patients developed skin toxicity and 32.6% of cases was of grade 3-4. No allergic reactions were identified at any courses in any patients. Fever was documented in 27.3% of patients and was most commonly recorded after the first administration. Cetuximab has clinically significant activity even in heavily pretreated colorectal cancer patients progressed after both oxaliplatin and irinotecan-based chemotherapy regimens.

Expression and heterodimer-binding activity of Ku70 and Ku80 in human non-melanoma skin cancer.

BACKGROUND: Experimental data suggest that exposure to ultraviolet radiation may indirectly induce DNA double-strand breaks. AIM: To investigate the contribution of the non-homologous end-joining repair pathway in basal and squamous cell carcinomas. METHODS: Levels of Ku70 and Ku80 proteins were determined by immunohistochemical analysis and Ku70-Ku80 heterodimer-binding activity by electrophoretic mobility shift assay. Matched pathological normal margins and skin from healthy people were used as controls. RESULTS: A significant increase in Ku70 and Ku80 protein levels was found for both tumour types as compared with normal skin (p<0.001). Squamous cell carcinoma showed increased immunostaining as compared with basal cell tumours (p<0.02). A direct correlation was found between Ku70 and Ku80 protein levels and expression of the proliferation markers Ki-67/MIB-1 (p<0.02 and p<0.002, respectively) in basal cell carcinoma. DNA binding activity was increased in basal cell carcinoma samples as compared with matched skin histopathologically negative for cancer (p<0.006). In squamous cell carcinomas, however, the difference was significant only with normal skin (p<0.02) and not with matched pathologically normal margins. CONCLUSIONS: Overall, an up regulation of the Ku70 and Ku80 protein levels seems to correlate only with tumour proliferation rate. As non-homologous end joining is an error-prone mechanism, its up regulation may ultimately increase genomic instability, contributing to tumour progression.

COX-2 expression in ampullary carcinoma: correlation with angiogenesis process and clinicopathological variables.

BACKGROUND: There is evidence that the anti-neoplastic effect of non-steroidal anti-inflammatory drugs is attributable to cyclooxygenase-2 (COX-2) inhibition, but the exact mechanisms whereby COX-2 can promote tumour cell growth remain unclear. One hypothesis is the stimulation of tumour angiogenesis by the products of COX-2 activity. To data, there have been few clinicopathological studies on COX-2 expression in human ampullary carcinoma and no data have been reported about its relation with tumour angiogenesis. OBJECTIVE: To investigate by immunohistochemistry the expression of COX-2 and the angiogenesis process in a series of primary untreated ampullary carcinomas. METHODS: Tissue samples from 40 archival ampullary carcinomas were analysed for COX-2, vascular endothelial growth factor (VEGF), and an endothelial cell marker von Willebrand factor (vWF) by immunohistochemistry, using specific antibodies. RESULTS: COX-2 expression was detected in 39 tissue samples (97.5%), of which two (5%) were graded as weak, 26 (65%) as moderate, and 11 (27.5%) as strong. Only one lesion (2.5%) was negative for COX-2 expression. VEGF expression was detected in 36 tissue samples (90%). A significant positive correlation was found between COX-2 and VEGF expression. No statistic correlation was found between COX-2 expression and microvessel density. CONCLUSIONS: COX-2 is highly expressed in ampullary carcinomas. This suggests an involvement of the COX-2 pathway in ampullary tumour associated angiogenesis, providing a rationale for targeting COX-2 in the treatment of ampullary cancer.

Cutaneous lymphoadenoma: a rare clinicopathological entity.

Cutaneous Lymphadenoma (Benign Lymphoepithelial tumour of the skin) is a rare tumour, with distinctive clinical and histological features. To date, very few cases of this entity have been reported. We present a case of cutaneous lymphoadenoma in a 52-year-old man and a short review of the literature, summarizing the principal clinical and morphological characteristics of this rare tumour.

Tumour proliferation, angiogenesis, and ploidy status in human colon cancer.

AIMS: Tumour angiogenesis is essential for carcinogenesis and facilitates the process of tumour development and metastasis. Vascular endothelial growth factor (VEGF) is a well characterised angiogenetic factor and is known to play a crucial role in new vessel development. To gain further insight into the effects of microvessel density and VEGF expression in colon cancer, their relation with tumour proliferation, ploidy status, and p53 expression was investigated in colon cancer. METHODS: Tissue samples of 50 archived colon cancers were analysed by immunohistochemistry for VEGF, p53, and the endothelial cell marker, von Willebrand factor (VWF), using specific antibodies. The same samples were re-cut for flow cytometric studies to obtain S phase fraction (SPF) and ploidy status. RESULTS: A positive significant correlation was found between SPF and angiogenesis. The median microvessel count in high SPF tumours was significantly higher than in low SPF ones. No association was found between VEGF expression and SPF. A positive correlation was found between ploidy status and p53 expression and microvessel count. Furthermore, a positive correlation was established between DNA ploidy, VEGF expression, and microvessel count. CONCLUSION: This study provides evidence that in colon cancer, tumour growth may be stimulated by vascular supply, and the lack of a correlation between tumour cell proliferation and VEGF expression indicates that these two parameters may be regulated by separate mechanisms. Furthermore, the positive correlation between microvessel density, VEGF expression, and ploidy status provides more evidence that genetic alterations are involved in tumour angiogenesis.

A rare case of solitary fibrous tumour of the pre-sacral space: morphological and immunohistochemical features.

A 28-year-old woman presented with abdominal pain. Ultrasonograhic examination showed a pre-sacral mass, with complex structure and well delimitated cystss with thick walls. The resected specimen was 7.5 x 6 x 4 cm in size, well circumscribed and yellow in colour, with cysstic change containing mucoid-like material. Histologically, the lesion was composed of spindle cells with high cellularity and rich vascularization with a haemangiopericytoma-like pattern. The diagnosis of solitary fibrous tumour (SFT) was made. The differential diagnosis for SFT of the pre-sacral spaace involves haemangiopericytoma, GIST, malignant mesothelioma, synovial sarcoma, leiomyomatous tumours and granulosa cell tumour. Immunohistochemical studies revealed reactivity for CD34, CD99 and Bcl-2, but no staining for desmin, inhibin, c-kit, EMA, CK, SMA, S-100 and CD31, confirming a diagnosis of SFT. Although SFT is usually associated with a favourable prognosis, close follow-up is recommended because of the limited information on its long-term behaviour.