Inés de Torres

Diarrhoea-predominant IBS patients show mast cell activation and hyperplasia in the jejunum

Background: Increased numbers of mast cells and mast cell activation in distal gut segments are associated with symptom onset and severity in irritable bowel syndrome (IBS). Although upper gut symptoms are common, mast cells have not been thoroughly evaluated in proximal gut in IBS patients. Methods: Jejunal biopsies obtained by Watson’s capsule, aspiration of intestinal fluid and one blood sample were obtained in 20 diarrhoea-predominant patients with IBS (D-IBS) and 14 healthy volunteers (H). Psychological stress (Holmes-Rahe Scale) and depression (Beck’s Depression Inventory) were evaluated at baseline and food and respiratory allergy excluded. Biopsies were processed for H&E staining and microscopic inflammation assessed by counting intraepithelial lymphocytes. Mast cells in lamina propria were counted by immunohistochemistry with CD117 (c-kit). Tryptase concentration was measured in intestinal fluid and serum. Results: D-IBS patients showed higher psychological stress than healthy volunteers (D-IBS: 203 (SD 114) v H: 112 (SD 99); p = 0.019). Immunohistochemical staining of jejunal mucosa revealed mild increase in intraepithelial CD3+ cells in D-IBS patients (D-IBS: 15.3 (SD 5.5; 95% CI 12.7 to 17.9) v H: 10.3 (SD 3.9; 95% CI 8.0 to 12.5); p = 0.006). Moreover, D-IBS patients showed marked increase in mast cells numbers (D-IBS: 34 (SD 9.3); H: 15.3 (SD 4.4) mast cells/hpf; p<0.001) and higher tryptase concentration in jejunal fluid (D-IBS: 0.45 (SD 0.38); H: 0.09 (SD 0.10) µg/l; p = 0.005). Upper gut symptoms were not associated with gender, mast cell counts, jejunal tryptase or basal stress. Conclusion: This jejunal mucosal inflammatory profile may help identify diarrhoea-predominant IBS, a stress-related disorder.

Diarrhoea-predominant irritable bowel syndrome: an organic disorder with structural abnormalities in the jejunal epithelial barrier

Objective Recently, the authors demonstrated altered gene expression in the jejunal mucosa of diarrhoea-predominant irritable bowel syndrome patients (IBS-D); specifically, the authors showed that genes related to mast cells and the intercellular apical junction complex (AJC) were expressed differently than in healthy subjects. The aim of the authors here was to determine whether these alterations are associated with structural abnormalities in AJC and their relationship with mast cell activation and IBS-D clinical manifestations. Design A clinical assessment and a jejunal biopsy were obtained in IBS-D patients (n=45) and healthy subjects (n=30). Mucosal mast cell number and activation were determined by quantifying CD117+ cells/hpf and tryptase expression, respectively. Expression and distribution of AJC specific proteins were evaluated by western blot and confocal microscopy. AJC ultrastructure was assessed by transmission electron microscopy. Results Compared with healthy subjects, IBS-D patients exhibited: (a) increased mast cell counts and activation; (b) increased protein expression of claudin-2, reduced occludin phosphorylation and enhanced redistribution from the membrane to the cytoplasm; and (c) increased myosin kinase expression, reduced myosin phosphatase and, consequently, enhanced phosphorylation of myosin. These molecular alterations were associated with ultrastructural abnormalities at the AJC, specifically, perijunctional cytoskeleton condensation and enlarged apical intercellular distance. Moreover, AJC structural alterations positively correlated both with mast cell activation and clinical symptoms. Conclusion The jejunal mucosa of IBS-D patients displays disrupted apical junctional complex integrity associated with mast cell activation and clinical manifestations. These results provide evidence for the organic nature of IBS-D, a heretofore model disease of functional gastrointestinal disorders.

Increased expression of angiogenic factors in human colonic angiodysplasia

OBJECTIVE:Angiodysplasia of the colon is a distinct vascular abnormality characterized by focal accumulation of ectatic vessels in the mucosa and submucosa. To investigate whether angiogenesis contributes to the pathogenesis of human colonic angiodysplasia, we examined the expression of basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF), and its endothelial cell receptors flt-1 and KDR.METHODS:Immunohistochemistry was performed in sections of specimens obtained from 18 patients with colonic angiodysplasia and from eight patients with colon cancer and its adjacent, histologically normal margins of resection. We used affinity-purified rabbit polyclonal antibodies and a streptoavidin–biotin peroxidase method.RESULTS:We detected strong immunoreactivity for vascular endothelial growth factor, homogeneously distributed in the endothelial lining of blood vessels of all sizes in 16 (89%) specimens of colonic angiodysplasia and in seven (88%) patients with colon cancer. In contrast, very limited immunoreactivity was found in normal colon. Vascular staining for flt-1 was observed in eight (44%) and one (12.5%) of the colonic angiodysplasia or colon cancer specimens, respectively, but not in normal colon. Vascular immunoreactivity for basic fibroblast growth factor was observed in seven (39%) specimens from patients with colonic angiodysplasia, whereas either very limited or no immunostaining was found in sections from specimens of patients with colon cancer and its normal margins.CONCLUSIONS:In human colonic angiodysplasia, increased expression of angiogenic factors is likely to play a pathogenic role.

Prognostic value of immunohistochemical expression of the c-erbB-2 oncoprotein in metastasic prostate cancer.

It is well established that the activation of proto‐oncogenes could trigger uncontrolled cell growth and cancer development. Although this correlation has already been evidenced in several human tumors, no conclusive studies have related oncogene activation with the development of prostatic neoplasia. Nevertheless, some reports suggest that c‐erbB‐2, which is a prognostic marker in breast cancer, could be implicated in the development of prostatic adenocarcinoma. We have studied the expression of the c‐erbB‐2 oncoprotein in primary prostatic tissue in a series of 70 patients with metastasic disease, by means of immunohistochemistry. The NCL‐B 11 anti‐c‐erbB‐2 monoclonal antibody was used, and the immunoreactivity was quantified by image analysis. The overall rate of prostatic‐tissue sections presenting positive c‐erbB‐2 immunostaining was 64.3%. No significant relation was observed between histological grade and c‐erbB‐2 over‐expression or severity of the disease, based on the extent of metastases. The average specific survival in patients with c‐erbB‐2 over‐expression was 33 months, while it was 54 in patients with c‐erbB‐2 negativity; p < 0.034. These results, as well as the logistic‐regression analysis, suggest that expression of c‐erbB‐2 oncoprotein would be considered as an independent prognostic factor of metastatic prostate cancer. Moreover, it could discriminate between the prognosis of patients with Gleason score 2 to 7 and those with score 8 to 10. Our results suggest that the expression of c‐erbB‐2 oncoprotein in primary prostatic tissue could have a prognostic value in patients with metastatic prostate cancer. Int. J. Cancer (Pred. Oncol.) 84:421–425, 1999.

Increased humoral immunity in the jejunum of diarrhoea-predominant irritable bowel syndrome associated with clinical manifestations

Background and aims Altered intestinal barrier is associated with immune activation and clinical symptoms in diarrhoea-predominant IBS (IBS-D). Increased mucosal antigen load may induce specific responses; however, local antibody production and its contribution to IBS aetiopathogenesis remain undefined. This study evaluated the role of humoral activity in IBS-D. Methods A single mucosal jejunal biopsy, luminal content and blood were obtained from healthy volunteers (H; n=30) and IBS-D (n=49; Rome III criteria) participants. Intraepithelial lymphocytes, mast cells, B lymphocytes and plasma cells were studied by imaging techniques. Differential gene expression and pathway analysis were assessed by microarray and PCR techniques. Blood and luminal immunoglobulins (Igs) were quantified. Gastrointestinal symptoms, respiratory atopy and stress and depression were also recorded. Results Patients with IBS-D showed a higher number and activation of mucosal B lymphocytes and plasma cells (p<0.05). Mast cell density was increased in patients with IBS-D (non-atopic) and in close proximity to plasma cells (p<0.05). Microarray profiling identified differential humoral activity in IBS-D, involving proliferation and activation of B lymphocytes and Igs production (p<0.001). Mucosal humoral activity was higher in IBS-D, with upregulation of germline transcripts and Ig genes (1.3-fold–1.7-fold increase; p<0.05), and increased IgG+ cells and luminal IgG compared with H (p<0.05), with no differences in blood. Biological markers of humoral activity correlated positively with bowel movements, stool form and depression. Conclusions Enhanced small bowel humoral immunity is a distinctive feature of IBS-D. Mucosal Ig production contributes to local inflammation and clinical manifestations in IBS-D.

PTOV1, a novel protein overexpressed in prostate cancer containing a new class of protein homology blocks

In a search for molecular markers of progression in prostate cancer by means of differential display, we have identified a new gene, which we have designated PTOV1. Semiquantitative RT–PCR has established that nine out of 11 tumors overexpress PTOV1 at levels significantly higher than benign prostatic hyperplasia or normal prostate tissue. The human PTOV1 protein consists almost entirely of two repeated blocks of homology of 151 and 147 amino acids, joined by a short linker peptide, and is encoded by a 12-exon gene localized in chromosome 19q13.3. A Drosophila melanogaster PTOV1 homolog also contains two tandemly arranged PTOV blocks. A second gene, PTOV2, was identified in humans and Drosophila, coding for proteins with a single PTOV homology block and unrelated amino- and carboxyl-terminal extensions. A 1.8-Kb PTOV1 transcript was detected abundantly in normal human brain, heart, skeletal muscle, kidney and liver, and at low levels in normal prostate. Immunocytochemical analysis and expression of chimeric GFP-PTOV1 proteins in cultured cells showed a predominantly perinuclear localization of PTOV1. In normal prostate tissue and in prostate adenomas, PTOV1 was undetectable or expressed at low levels, whereas nine out of 11 prostate adenocarcinomas showed a strong immunoreactivity, with a focal distribution in areas of carcinoma and prostatic intraepithelial neoplasia. Therefore, PTOV1 is a previously unknown gene, overexpressed in early and late stages of prostate cancer. The PTOV homology block represents a new class of conserved sequence blocks present in human, rodent and fly proteins.

Metabolic syndrome increases the risk of aggressive prostate cancer detection

Metabolic syndrome can identify patients at high risk of cardiovascular disease. The prevalence of metabolic syndrome is increasing worldwide and is associated with increased age, obesity and hypogonadism. The association between metabolic syndrome and prostate cancer development has not been studied comprehensively, and published studies report divergent results. This study indicates that tumours detected in men with metabolic syndrome are more aggressive than those detected in men without this condition.

Identification and genotyping of human papillomavirus in a Spanish cohort of penile squamous cell carcinomas: Correlation with pathologic subtypes, p16INK4a expression, and prognosis

BACKGROUND Penile squamous cell carcinoma (PSCC) is a tumor with a high metastatic potential. In PSCC the attributable fraction to human papillomavirus (HPV) is not well established. OBJECTIVE We sought to provide novel data about the prevalence of HPV in a large series of penile intraepithelial neoplasia (PeIN) and invasive PSCC, correlating the results with the histologic subtype, p16(INK4a) immunostaining, and prognosis. METHODS A total of 82 PSCC were included in the study, 69 invasive and 13 PeIN. HPV detection was performed by polymerase chain reaction with SPF-10 broad-spectrum primers followed by DNA enzyme immunoassay and genotyping with a reverse hybridization line probe assay. P16(INK4a) immunohistochemical expression on tissue microarrays was also analyzed. RESULTS HPV DNA was identified in 31 of 77 (40.2%) PSCC (22 of 67 invasive and 9 of 10 PeIN). In 25 of 31 (80.6%) cases HPV-16 was identified. HPV detection was significantly associated with some histologic subtypes: most basaloid and warty tumors were high-risk HPV (hrHPV) positive, whereas only 15% of usual PSCC were hr-HPV positive. All hrHPV-positive PSCC had an adjacent undifferentiated PeIN. Strong p16(INK4a) immunostaining correlated with hrHPV infection. Most undifferentiated PeIN showed p16(INK4a) immunohistochemical overexpression. Both hrHPV-positive and p16(INK4a)-positive tumors showed a better overall survival without reaching statistical significance. LIMITATIONS This was a retrospective study. CONCLUSIONS Our results suggest that most hrHPV-positive PSCC develop from undifferentiated hrHPV-positive PeIN. P16(INK4a) immunostaining may be useful in identifying both etiologically related hrHPV-positive tumors and those with better outcome. The routine use of p16(INK4a) staining should be incorporated in histologic evaluation of PSCC.

Increased glyceraldehyde-3-phosphate dehydrogenase expression in renal cell carcinoma identified by RNA-based, arbitrarily primed polymerase chain reaction

Renal cell carcinoma (RCC) comprises 85% of renal tumors and displays a great capacity to metastasize. The lack of diagnostic and prognostic markers complicates its early detection and in the majority of cases metastases are present at the time of diagnosis.

Prognostic Significance of Vascular Endothelial Growth Factor and Cyclooxygenase-2 in Patients with Rectal Cancer Treated with Preoperative Radiotherapy

Purpose: To analyze the prognostic value of cyclooxygenase-2 (COX-2) and vascular endothelial growth factor (VEGF) in patients with locally advanced rectal cancer treated with preoperative radiotherapy. Methods: Eighty-one patients with locally advanced rectal cancer were studied. All patients received preoperative pelvic radiotherapy. Forty-seven patients received concomitant chemotherapy. Surgical resection was performed 4–8 weeks later in all patients. Immunohistochemical examination of COX-2 and VEGF was performed on the preirradiation diagnostic biopsies. An immunohistochemical score established from the extension and intensity of the markers was used for analysis. The log-rank test and proportional hazards regression analysis were used to calculate the probability that the biomarkers were associated with patient outcome. Results: COX-2 expression was positive in 38 tumors (51%) while VEGF expression was positive in 43 (57%). The only clinicopathological parameter significantly associated with COX-2 or VEGF expression was performance status. None of the 2 markers were found to predict treatment response. There was no statistically significant correlation between COX-2 and VEGF. Univariate analysis identified pathological stage (pT, pN) as prognostic for disease-free survival. When VEGF expression was analyzed, disease-free survival was reduced among patients with VEGF-positive tumors (p = 0.047). This was specifically related to metastases-free survival (p = 0.016). These results were not observed for COX-2. After multivariate analysis, the pT and pN stage remained as independent prognostic factors. Conclusions: VEGF-positive expression is an indicator of poor disease-free survival, specifically linked to distant metastasis. More aggressive treatment strategies are warranted in pT3–4 and pN1–2 rectal cancer patients.