Confocal laser endomicroscopy in ulcerative colitis: beyond endoscopic assessment of disease activity.

BACKGROUND: The aim of this study was to investigate the role of confocal laser endomicroscopy (CLE) in the assessment of disease activity in ulcerative colitis (UC). METHODS: Consecutive patients with UC referred to our inflammatory bowel disease unit for colonoscopy were enrolled. Patients without UC were used as controls. UC activity was evaluated by white light endoscopy and classified according to the Mayo Ulcerative Colitis Endoscopic Score of Severity. Endoscopic biopsies were also taken for histological assessment of disease activity and then assessed with CLE. Three parameters were evaluated; crypt architecture (crypt diameter, inter-crypt distance, presence of fused crypts, crypts regularity), microvascular pattern (regular, dilated, irregular and deformed), fluorescein leakage. RESULTS: Fifty patients with UC and 10 controls were enrolled. At colonoscopy, 11 patients (22%), 19 patients (38%), 12 patients (24%) and 8 patients (16%) presented a Mayo score of 0, 1, 2 and 3, respectively. At CLE, fused crypts were present in all the patients with UC and absent in controls. Crypt diameter and inter-crypt distance showed a parallel increase with the Mayo score. Fluorescein leakage and irregular vessels were more frequently found in case of a high level of endoscopic severity, but were also identified in about 20% of UC patients with normal mucosa. Biopsies also demonstrated the presence of histological activity in 4 patients with endoscopically inactive colitis. CONCLUSIONS: CLE might be a useful tool to determine inflammatory activity in UC. Fused crypts appeared to be a CLE marker of UC, while other abnormalities, like microvascular alteration and fluorescein leakage, have also been identified in patients with mucosal healing at endoscopy. Larger series are required to validate these results and the advantages of a CLE-based assessment of UC activity.

In vivo assessment of tumour angiogenesis in colorectal cancer: the role of confocal laser endomicroscopy.

AIM: Tumour neoangiogenesis is a key factor in tumour progression and metastatic spread and the possibility to assess tumour angiogenesis might provide prognostic information. The aim of this study was to establish the role of probe-based confocal laser endomicroscopy (p-CLE) in the identification of vascular architecture and specific morphological patterns in normal colorectal mucosa and malignant lesions during routine endoscopy. METHOD: Fourteen consecutive patients with colorectal cancer were included. The following features were identified and then compared between normal and neoplastic mucosa on p-CLE images: vessel shape (straight vs irregular) vessel diameter the 'branching patterns' vessel permeability (fluorescein leakage) and blood flow (normal vs defective flux). Immunohistochemistry was used to confirm the presence and to study the morphology of vascular structures (CD-34 staining) and 'neo-vessels' (WT-1 staining) on tumour and normal mucosal sections. RESULTS: Tumour vessels appeared as irregular, ectatic and with a highly variable calibre and branching patterns on p-CLE images. The mean diameter of tumour vessels was significantly larger than those in normal mucosa (weighted mean difference 3.38, 95% CI 2.65-4.11, P = 0.01). Similarly, 'vessel branching' (OR 2.74, 95% CI 1.23-6.14, P = 0.01), fluorescent dye 'extravasation' (OR 3.46, 95% CI 1.39-8.57, P = 0.01) were significantly more frequent in colorectal cancer than in normal colorectal mucosa. Immunohistochemistry corroborated the p-CLE findings, showing higher vascularity in tumour sections due to neoformed vessels, presenting irregular patterns. CONCLUSION: Probe-based confocal laser endomicroscopy provides a noninvasive characterization of the microvascular architecture of colonic mucosa. Different morphological patterns have been described, discriminating normal and malignant microvascular networks in colorectal mucosa.

Efficacy of subcutaneous immunoglobulins in primary immunodeficiency with Crohn's-like phenotype: report of a case.

Common variable immune deficiency (CVID) is the most frequent primary immunodeficiency in adults. In CVID, the prevalence of gastrointestinal manifestations ranges between 2 and 50% with a complication-related morbidity second only to that of the respiratory tract. In some cases, clinical and endoscopic features are undistinguishable from those of inflammatory bowel disease (IBD). We describe the case of a 28-year-old man in which a diagnosis of Crohn's disease was firstly suspected. Subsequently, a diagnosis of Crohn's-like disease in a patient with CVID was made and a replacement therapy with human normal immunoglobulin intravenously was started. Unfortunately, serum IgG levels remained below 2.0 g/l in pre-infusional controls with persistence of gastrointestinal symptoms and malnutrition despite anti-inflammatory therapy (mesalazine, corticosteroids). Then, the patient began treatment with human normal immunoglobulins administered subcutaneously. The follow-up visits showed a progressive increase in serum IgG. Moreover, the patient reported improvement of gastrointestinal symptoms with reduction of diarrhoea, and laboratory tests showed a progressive and significant improvement. We confirm that therapy with subcutaneously administered immunoglobulins is safe and effective. In addition, our observations indicate that, for patients with CVID and enteropathic complications, replacement therapy with subcutaneous IgG may be the treatment of choice.

Increased mucosal nitric oxide production in ulcerative colitis is mediated in part by the enteroglial-derived S100B protein.

In the central nervous system glial-derived S100B protein has been associated with inflammation via nitric oxide (NO) production. As the role of enteroglial cells in inflammatory bowel disease has been poorly investigated in humans, we evaluated the association of S100B and NO production in ulcerative colitis (UC). S100B mRNA and protein expression, inducible NO synthase (iNOS) expression, and NO production were evaluated in rectal biopsies from 30 controls and 35 UC patients. To verify the correlation between S100B and NO production, biopsies were exposed to S100B, in the presence or absence of specific receptor for advanced glycation end-products (RAGE) blocking antibody, to measure iNOS expression and nitrite production. S100B and iNOS expression were evaluated after incubation of biopsies with lipopolysaccharides (LPS) + interferon-gamma (IFN-gamma) in the presence of anti-RAGE or anti-S100B antibodies or budesonide. S100B mRNA and protein expression, iNOS expression and NO production were significantly higher in the rectal mucosa of patients compared to that of controls. Exogenous S100B induced a significant increase in both iNOS expression and NO production in controls and UC patients; this increase was inhibited by specific anti-RAGE blocking antibody. Incubation with LPS + IFN-gamma induced a significant increase in S100B mRNA and protein expression, together with increased iNOS expression and NO production. LPS + IFN-gamma-induced S100B up-regulation was not affected by budesonide, while iNOS expression and NO production were significantly inhibited by both specific anti-RAGE and anti-S100B blocking antibodies. Enteroglial-derived S100B up-regulation in UC participates in NO production, involving RAGE in a steroid insensitive pathway.

Neo-adjuvant treatment of rectal cancer with capecitabine and oxaliplatin in combination with radiotherapy: a phase II study.

BACKGROUND: Preoperative chemoradiation is now standard treatment for stages II-III rectal cancer. Capecitabine (CAP) and oxaliplatin (OX) are synergistic with radiotherapy (RT) and active in colorectal neoplasms. PATIENTS AND METHODS: Two cycles of CAP 825 mg/m(2) b.i.d. (days 1-14) and OX 50 mg/m(2) (days 1 and 8) every 3 weeks were given concomitantly with pelvic conformal RT (45 Gy). Patients with a > or =T3 and/or node-positive rectal tumour were eligible. The pathologic tumour response was defined according to the tumour regression grade (TRG) scale. RESULTS: Forty-six patients were enrolled. Gastrointestinal adverse events were mostly G1-G2; only two patients experienced G3 vomiting and diarrhoea and six patients had G1 peripheral neuropathy. Haematological toxicity was rare. G2 proctitis and anal pain occurred in two patients. Pathological complete response (TRG1) was observed in nine patients (20.9%; 95% CI 8.7%-33.1%); TRG2 in 19 patients (44.2%); TRG3 in 12 patients (27.9%); and TRG4 in three patients (7%). Overall, nine patients recurred: five with distant metastases, one with local recurrence, and three with both local recurrence and distant metastases. CONCLUSIONS: CAP-OX-RT as preoperative treatment for rectal cancer induces a remarkable rate of complete or near-complete pathologically documented response and is well tolerated.

Effect of gestational hypercholesterolaemia on omental vasoreactivity, placental enzyme activity and transplacental passage of normal and oxidised fatty acids.

OBJECTIVE: Maternal hypercholesterolaemia during pregnancy increases lipid peroxidation in mothers and fetuses and programs increased susceptibility to atherosclerosis later in life. The objective of this study was to elucidate the role of the placenta in mediating oxidative stress from mother to offspring. DESIGN: Comparison between normo- and hypercholesterolaemic mothers (n = 36 each) and their children. SETTING: Obstetric wards, hospitals of the University of Naples and Regione Campania. POPULATION: Healthy primiparas delivering by caesarean section. METHODS: Biochemical measurements of oxidative stress and serum leptin in cord plasma and placenta, immunochemistry of placenta microvessels, and vasoreactivity studies were performed. MAIN OUTCOME MEASURES: Oxidative status (i.e. lipid composition and content of oxidised fatty acids, activity of pro- and antioxidant enzymes, immunohistochemical presence of oxidation-specific epitopes) in maternal and cord blood and in placental tissue, as well as vascular reactivity in omental arteries. RESULTS: Hypercholesterolaemia during pregnancy was associated with extensive changes in fatty acid composition of both maternal and cord blood lipids, sufficient to alter vasoreactivity of omental vessels. Results also indicated that the placenta is not only subject to substantial oxidative stress, but that it may further increase fetal oxidative stress through changes of pro- and antioxidant enzyme activities. CONCLUSIONS: The placenta plays an important role in both transmitting and enhancing pathogenic effects of gestational hypercholesterolaemia.

Expression of HECA-452 in parapsoriasis and mycosis fungoides.

We have investigated the HECA-452 expression in large plaque parapsoriasis (PP) and mycosis fungoides (MF) patients, evaluating the potential role of this biomarker in both cutaneous disorders. Skin specimens from 72 PP and 61 MF patients were selected in this study. We compared their actual histological diagnosis with their previous diagnosis and we found that all 72 PP patients had the same diagnosis as before (stable PP), while 26 out of 61 MF had a previous PP histological diagnosis (evolving PP). Our results show an increased expression of HECA-452 in MF compared to PP (p<0.01). Furthermore, evolving PP showed a significantly higher level of HECA-452 than stable PP (p<0.05). We conclude that HECA-452 expression increases during the natural history of Mycosis Fungoides. HECA-452 could be used as a biomarker for MF and predict which PP evolves to MF.

Neuroendocrine differentiation after neoadjuvant hormonal treatment in prostate cancer.

AIM: Neuroendocrine (NE) differentiation occurs in various degree in the majority of prostatic adenocarcinomas and it has been correlated with tumor progression and poor prognosis. There is little knowledge about the impact of NE differentiation on tumor response to neoadjuvant hormonal treatment (NHT). The role of NE differentiation as a marker of recurrence after radical prostatectomy (RP) is also unclear. We evaluated whether there is an increase in NE differentiation during the course of NHT and whether the tumor relapse after radical surgery correlates with the extent of NE differentiation. METHODS: RP specimens from 44 patients submitted to 3 months of NHT and RP specimens from 40 nonpretreated patients were histologically assessed. Staining for NE differentiated prostate tumor cells was carried out using a specific monoclonal antibody against chromogranin A (CgA). RESULTS: CgA positive cells were found in 4 of 40 patients (10%) in the RP group and in 4 of 44 patients (9%) of the NHT+RP group. At follow-up, we had 21 biochemically relapsed patients. Among them, 6 were CgA positive (75% of 8 patients), whereas is were CgA negative (20% of 76 patients). CONCLUSIONS: The NE differentiation doesn't increase after NHT. Although not statistically significant a trend to higher risk of relapse among the chromogranin positive samples was observed. The significance of NE differentiation in the progression of the disease and its relation to other known prognostic factors remains unclear.

Carotid endarterectomy in heart transplant patients.

AIM: The aim of this study was to determine the clinical outcome of carotid endarterectomy in heart transplant recipients and morphologic features of atherosclerotic plaques removed during operation. METHODS: Between April 1993 and October 2001 5 heart transplant patients with symptomatic carotid stenosis >70% underwent carotid endarterectomy with regional anesthesia, including a staged bilateral procedure in one patient. Cholesterol, triglycerides, HDL-cholesterol, LDL-cholesterol were evaluated in each patient. The plaques ( n=6) underwent histologic analysis after carotid endarterectomy. Carotid artery duplex imaging was added to the routine postoperative evaluation. RESULTS: Carotid plaques resulted to be echolucent on B-mode ultrasound examination. Cholesterol, triglycerides and LDL-cholesterol levels were found to be increased, while HDL-cholesterol were decreased. All patients underwent successful carotid endarterectomy; there were no perioperative deaths, major neurologic or cardiac events. The mean length of stay was 2.2 days. The mean follow-up was 44 months. In 1 case, an asymptomatic restenosis >50% occurred 9 months later and, in 2 other cases, a contralateral mild stenosis was found 12 and 36 months later. One patient had a progressive contralateral stenosis, requiring operation 18 months later. High lipid content and heterogeneous cellular infiltration were observed, including macrophages, T-lymphocytes, neutrophils, and also eosinophils in the rapidly progressing plaque. CONCLUSIONS: Heart transplant patients receiving immunosuppression may successfully undergo carotid endarterectomy, without increased risk, but progression of atherosclerotic disease in the carotid arteries seems to continue, despite lipid-lowering regimen and antiplatelet therapy.

Expression of biomarkers modulating prostate cancer progression: implications in the treatment of the disease.

OBJECTIVES: To determine whether COX-2, bcl-2 and neoangiogenesis are related to human prostate cancer relapse after definitive surgical treatment and progression toward androgen independence and to evaluate the association between the patterns of these tumoral biomarkers and other standard clinico-pathological parameters (such as Gleason score, PSA, TNM stage). MATERIALS AND METHODS: We retrospectively analyzed the records on 126 prostate cancer samples from patients treated at our University Hospital from 1995 to 2002. The 72 patients with clinically localized disease (group 1) had undergone radical prostatectomy. Another 54 patients (group 2) had metastatic androgen-independent disease. Archived material relating to the subjects was then immunostained for bcl-2, COX-2 and CD-31, using an anti-bcl-2 monoclonal primary antibody, an anti-COX-2 polyclonal rabbit antibody and an anti-CD-31 monoclonal mouse antibody to evaluate neoangiogenesis (MVD, microvessel density). RESULTS: We found that bcl-2, COX-2 and MVD expression increased from group 1 to group 2. The intergroup difference was significant only for high MVD (P < 0.05). On the other hand, high MVD, high bcl-2 and high COX-2 expression was correlated with a higher PSA level (P < 0.01), whereas only a high MVD was also related with Gleason score (P < 0.05). We used univariate analysis to evaluate the prognostic impact of biologic and clinico-pathologic parameters on the disease-free-survival of 72 patients treated by radical prostatectomy. A total of 30 patients (41.6%) experienced biochemical relapse; bcl-2, COX-2 and MVD significantly correlated with disease relapse in these patients. In fact, we observed disease relapse in 24/45 (53%) with high bcl-2 expression, in 15/21 (71%) with a high MVD count and finally, in 30/58 (52%) with high COX-2 expression. Finally, PSA value and Gleason score were the only two biologic markers significantly associated to disease relapse in a multivariate analysis. CONCLUSIONS: Our results strongly support a role for bcl-2, COX-2 and angiogenesis in the development and progression of prostate cancer. Of course, we are aware of the small sample size considered in our study. Further investigations would better clarify the prognostic and therapeutic implications of these findings.

Expression of proto-oncogene c-kit in high risk prostate cancer.

AIMS: We determined whether c-kit proto-oncogene is expressed in prostate cancer and whether its expression is related with biochemical relapse in high risk localized prostate cancer patients. METHODS: c-Kit expression was evaluated by immuno-histochemistry in 94 prostate cancer samples from patients treated by radical prostatectomy followed by adjuvant hormonal therapy because all patients had a pT3a stage (initially cT2 stage). All patients presented a >7 Gleason score and a >10 pre-operative PSA value. We evaluated association between c-kit positive staining and disease free survival. RESULTS: In 26 of 94 prostate cancer, we found an epithelial positive c-kit expression. The epithelial expression was found in the peripheral zone of prostate tissue. 13/94 relapsed and, although not statistically significant (p 0.055), a trend to a higher risk of relapse among the c-kit positive samples was observed in our series of prostate cancer patients. CONCLUSIONS: Our study is only an initial experience and it is necessary to consider a higher number of patients to clarify whether c-kit is really an independent predictor for disease recurrence. Further study in this area will help to understand whether anti c-kit drugs could become an effective complement to the armamentarium of prostate cancer therapies.

Cigarette smoking and appendectomy are risk factors for extraintestinal manifestations in ulcerative colitis.

OBJECTIVE: Two common factors, cigarette smoking and appendectomy, have been found to play a role in ulcerative colitis (UC). Data on their role in the development of extraintestinal manifestations (EIM) are scarce. METHODS: The relationship between cigarette smoking, appendectomy, and EIM was examined in a prospective study involving 535 (M/F = 319/216) consecutive UC patients followed up for 18 yr. We considered the major EIM: seronegative spondyloarthropathy, pyoderma gangrenosum/erythema nodosum, acute anterior uveitis, and primary sclerosing cholangitis. We excluded patients with a history of EIM or those colectomized before study entry, ex-smokers, and those who started to smoke during the course of UC. RESULTS: In UC patients, seronegative spondyloarthropathy and dermatologic complications were found increased in smokers (p < 0.0001; p = 0.001) or in subjects with appendectomy (p = 0.0003; p = 0.02), while acute anterior uveitis and primary sclerosing cholangitis did not differ. The Kaplan-Meier analysis showed 18-yr rates for EIM of 71% in smokers and 45% in nonsmokers (log-rank test, p = 0.0001), and of 85% in patients with appendectomy and 48% in those without (p = 0.0001). Cox proportional-hazard model showed that cigarette smoking and appendectomy are independent factors promoting EIM. In smokers with appendectomy the adjusted hazard ratio (3.197, 95% CI 1.529-6.684) was higher than in patients with appendectomy alone (2.617, 95% CI 1.542-4.442) or smoking alone (1.947, 95% CI 1.317-2.879). CONCLUSIONS: In UC patients, appendectomy and cigarette smoking are prognostic factors for the development of EIM. The unfavorable effect of cigarette smoking on EIM is additive to that of appendectomy.

Ulcerative colitis, seronegative spondyloarthropathies and allergic diseases: the search for a link.

BACKGROUND: Allergic diseases and seronegative spondyloarthropathies are frequently observed in ulcerative colitis (UC). In this report we have investigated possible relationships between IgE-mediated allergic disease (AD), allergic contact dermatitis (ACD) and seronegative spondyloarthropathy (SpA) in UC patients at different grades and extensions of mucosa inflammation. METHODS: Forty-five UC consecutive outpatients were graded according to clinical, endoscopic and histologic activity scores. SpA was diagnosed according to the European Spondyloarthropathy Study Group criteria. AD was detected by skin prick tests and confirmed by specific provocation tests, while ACD was diagnosed using the European standard series of patch tests. Thirty-seven patients' spouses or partners served as controls. RESULTS: Fourteen patients and 1 control subject showed SpA (P = 0.001). Diagnosis of rhinitis, conjunctivitis or asthma was made in 19 patients and in 5 controls (P = 0.004), while ACD was found in 10 and in 4 (P = 0.17), respectively. In UC, AD coexisted with SpA in 2 cases (P = 0.01), AD with ACD in 1 case (P = 0.03) and ACD with SpA in 5 (P = 0.24). CONCLUSIONS: Notwithstanding the high frequency of AD and SpA found in UC, the concurrence of AD with SpA or ACD is an unusual finding, while SpA and ACD may coexist. These data suggest that, in UC, atopy and seronegative arthritis, as well as atopy and delayed-type allergy, are strongly polarized conditions tending to mutual exclusion. In UC, the presence of AD without SpA or ACD, and of SpA or ACD without AD may indicate subgroups of patients in which T-helper-2 cell or T-helper-1 cell responses predominate.

Chronic treatment with sulfhydryl angiotensin-converting enzyme inhibitors reduce susceptibility of plasma LDL to in vitro oxidation, formation of oxidation-specific epitopes in the arterial wall, and atherogenesis in apolipoprotein E knockout mice.

The effects of chronic treatment with the new sulfhydryl angiotensin-converting enzyme (ACE)-inhibitor, zofenopril, in comparison with the classical sulfhydryl ACE-inhibitor captopril or enalapril or placebo on the development of atherosclerosis were determined in apolipoprotein-E knockout (apoE(-/-)) mice. Groups of 2-month-old male mice received either placebo (N=10), 0.05 mg/kg/day of zofenopril (N=10), 1 mg/kg/day of zofenopril (N=10), 5 mg/kg/day of captopril (N=10) or 0.5 mg/kg/day of enalapril (N=8). After 29 weeks of treatment, computer-assisted imaging analysis revealed that zofenopril reduced the aortic cumulative lesion area by 78% at 0.05 mg/kg/day and by 89% at 1 mg/ml/day of zofenopril compared to that of the placebo (P<0.0001). Captopril reduced by 52% aortic lesions compared to placebo (P<0.01 vs. placebo; P<0.05 vs. zofenopril at both doses). Enalapril did not reduce aortic lesions. Furthermore, 0.05 mg/kg/day of zofenopril reduced susceptibility of plasma LDL to in vitro oxidation compared to captopril, enalapril or placebo, as shown by significant reduction of malondialdehyde content (P<0.001 vs. placebo or enalapril; P<0.05 vs. captopril), as well as by the prolongation of lag-time (P<0.01 vs. placebo or enalapril P<0.05 vs. captopril). More importantly, mice treated with 1 mg/ml/day of zofenopril had a significant decrease in the intimal immunohistochemical presence of oxidation-specific epitopes on oxLDL (NA59 monoclonal antibody, P<0.01), macrophages derived foam cells (F4/80 monoclonal antibody, P<0.05) and native LDL (NP monoclonal antibody, P<0.01) compared to placebo, captopril or enalapril. Thus, chronic treatment with the new sulfhydryl ACE-inhibitor zofenopril has antiatherosclerotic and antioxidant effects in the arterial wall of hypercholesterolemic apoE(-/-) mice. This protection was significantly higher than that reached with captopril and at lower doses of the drug. Treatment with 0.5 mg/kg/day of enalapril did not provide any protective effect.

Maternal hypercholesterolemia and treatment during pregnancy influence the long-term progression of atherosclerosis in offspring of rabbits.

Maternal hypercholesterolemia during pregnancy is associated with enhanced fatty streak formation in human fetuses and faster progression of atherosclerosis during childhood even under normocholesterolemic conditions. A causal role of maternal hypercholesterolemia in lesion formation during fetal development has previously been established in rabbits. The same experimental model is now used to establish that maternal hypercholesterolemia or ensuing pathogenic events in fetal arteries enhance atherogenesis later in life. Five groups of rabbit mothers were fed chow, cholesterol-enriched chow, or cholesterol-enriched chow plus 1000 IU vitamin E, 3% cholestyramine, or both during pregnancy. Offspring of all groups (n=136) were fed a mildly hypercholesterolemic diet for up to a year and had similar cholesterol levels. Aortic lesion sizes and lipid peroxidation products in plasma and lesions in offspring were determined at birth, 6 months, or 12 months. Lesion progression in offspring of hypercholesterolemic mothers was greater than in all other groups. At each time point, offspring of hypercholesterolemic mothers had 1.5- to 3-fold larger lesions than offspring of normocholesterolemic mothers (P<0.01), with the greatest absolute differences at 12 months. Maternal treatment reduced lesions by 19% to 53%, compared with offspring of untreated hypercholesterolemic mothers (P<0.01), with the greatest effect in the vitamin E groups. At 12 months, lesions in offspring of all vitamin E and cholestyramine-treated mothers were similar to those of normocholesterolemic mothers. Lipid peroxidation end-products in lesions and plasma showed analogous differences between groups as lesions (P<0.01). Thus, pathogenic programming in utero increases the susceptibility to atherogenic risk factors later in life and maternal intervention with cholesterol-lowering drugs or antioxidants reduce postnatal lipid peroxidation and atherosclerosis in their offspring.