Domenico Piscitelli

Clarithromycin-Resistant Genotypes and Eradication of Helicobacter Pylori

OBJECTIVE To compare the eradication rates among the different point mutations and the efficacy of triple therapy and a sequential regimen according to genotypic resistance. STUDY DESIGN Post hoc retrospective cohort study in a tertiary referral center for pediatric gastroenterology in southern Italy. All 168 children who were positive for Helicobacter pylori were enrolled. Patients had received clarithromycin-based 7-day triple therapy (73 children) or 10-day sequential therapy regimen (95 children). Real-time polymerase chain reaction for assessing clarithromycin resistance was performed on sections of paraffin-embedded gastric biopsy samples. RESULTS H pylori eradication was achieved in 16 of 32 (50%) children with the A2143G mutation, in 8 of 10 patients with either A2142G or A2142C strains (80%), and in 112 of 116 children with susceptible strains (88.9%). The presence of A2143G mutation was associated with a lower cure rate compared with the rate in the absence of this mutation (50% vs. 89%; P = .001). The sequential regimen achieved a higher cure rate than triple therapy in patients with A2143G mutant strains (80% vs nil; P < .001). CONCLUSIONS The A2143G mutation confers higher risk of treatment failure. Sequential regimen has higher efficacy than standard therapy, even in children with A2143G mutatant strains.

ERβ expression in normal, adenomatous and carcinomatous tissues of patients with familial adenomatous polyposis.

Abstract Objectives. The APC gene mutation triggers familial adenomatous polyposis (FAP) and approximately 80% of sporadic colorectal cancers. FAP summarizes the natural history of colorectal cancer because low- and high-grade dysplastic lesions and adenocarcinoma are simultaneously present in the same patients free from individual and environmental variability factors. Estrogen receptor beta (ERβ) has recently been suggested as the most likely mediator of estrogen-related anti-carcinogenic effects in ApcMin−/+ mice and humans. In this study we assessed the ERβ expression in the intestinal mucosa of FAP patients to verify its possible involvement in tumor progression in colorectal cancer. Material and methods. ERβ and ERα expression, cell proliferation (Ki-67) and apoptosis (TUNEL), were evaluated on archival biopsy material from six patients with FAP who underwent colectomy. Results. A progressive significant decrease of ERβ expression was observed in the different stages of the disease as compared to normal mucosa (p < 0.001). Interestingly, a decreased ERβ expression was directly correlated with apoptosis (r = 0.76, p < 0.001), and inversely correlated with cell proliferation (r = 0.54, p < 0.05). Conclusions. ERβ expression is related to the severity of the disease, supporting the role of ERβ as a relevant biomarker of tumor progression and possible chemopreventive target in patients at risk of colonic neoplasia.

Incidental metastases of well-differentiated thyroid carcinoma in lymph nodes of patients with squamous cell head and neck cancer: eight cases with a review of the literature

The examination of a large series of cervical lymph nodes in patients with head and neck cancer revealed the presence of incidental metastases of occult thyroid carcinoma in eight patients, of which six cases were squamous cell carcinoma of glottic and supraglottic sites of the larynx and two cases were pyriform sinus and tongue carcinomas. Three patients had two lymph nodes and the remaining patients had one lymph node each involved. The nodal chains affected were the jugular (n=5; level IV), Kuttner (level II), supraomohyoid (level III) and supraclavicular (level VI). In four cases, a subtotal thyroidectomy or unilateral lobectomy was performed during laryngectomy (for surgical reasons) or after histologic nodal examination; a minimal focus of thyroid papillary carcinoma was detected in one patient. Three of eight patients died from recurrence of the squamous cell carcinoma; no case presented clinical evidence of thyroid malignancy. The differential diagnosis from benign thyroid heterotopia was based on the presence of minimal nuclear atypia. The choice of treatment of patients with a coexisting neoplasm characterized by poor prognosis is difficult, and contrasting opinions exist regarding the use of radical thyroidectomy and the subsequent management. As reported in the literature (66 cases), the more aggressive squamous cell carcinoma will determine the prognosis of these patients; in fact, only one of the referred cases died of cerebellar metastases of the thyroid cancer. Our results emphasize the importance of an accurate re-evaluation and follow-up of patients with incidental occult metastases for detection of a primary thyroid tumor. In the general population, this incidental nodal involvement may be related to a minimal occult thyroid carcinoma.

phytoestrogens/insoluble fibers and colonic estrogen receptor β: randomized, double-blind, placebo-controlled study

AIM To assess the safety and effect of the supplementation of a patented blend of dietary phytoestrogens and insoluble fibers on estrogen receptor (ER)-β and biological parameters in sporadic colonic adenomas. METHODS A randomized, double-blind placebo-controlled trial was performed. Patients scheduled to undergo surveillance colonoscopy for previous sporadic colonic adenomas were identified, and 60 eligible patients were randomized to placebo or active dietary intervention (ADI) twice a day, for 60 d before surveillance colonoscopy. ADI was a mixture of 175 mg milk thistle extract, 20 mg secoisolariciresinol and 750 mg oat fiber extract. ER-β and ER-α expression, apoptosis and proliferation (Ki-67 LI) were assessed in colon samples. RESULTS No adverse event related to ADI was recorded. ADI administration showed a significant increases in ER-β protein (0.822 ± 0.08 vs 0.768 ± 0.10, P = 0.04) and a general trend to an increase in ER-β LI (39.222 ± 2.69 vs 37.708 ± 5.31, P = 0.06), ER-β/ER-α LI ratio (6.564 ± 10.04 vs 2.437 ± 1.53, P = 0.06), terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (35.592 ± 14.97 vs 31.541 ± 11.54, P = 0.07) and Ki-67 (53.923 ± 20.91 vs 44.833 ± 10.38, P = 0.07) approximating statistical significance. A significant increase of ER-β protein (0.805 ± 0.13 vs 0.773 ± 0.13, P = 0.04), mRNA (2.278 ± 1.19 vs 1.105 ± 1.07, P < 0.02) and LI (47.533 ± 15.47 vs 34.875 ± 16.67, P < 0.05) and a decrease of ER-α protein (0.423 ± 0.06 vs 0.532 ± 0.11, P < 0.02) as well as a trend to increase of ER-β/ER-α protein in ADI vs placebo group were observed in patients without polyps (1.734 ± 0.20 vs 1.571 ± 0.42, P = 0.07). CONCLUSION The role of ER-β on the control of apoptosis, and its amenability to dietary intervention, are supported in our study.

NEOPLASTIC DISEASE AND DELETION 22q11.2: A MULTICENTRIC STUDY AND REPORT OF TWO CASES

Deletion 22q11.2 is a chromosomal abnormality detected in young patients with clinical manifestations of the DiGeorge/velocardiofacial syndrome. Conotruncal heart defects are also associated with del22q11.2. An association of these cardiac malformations with neoplasias has been observed. Our series includes two cases of malignancies, a hepatoblastoma and a renal-cell carcinoma, arising in children with complex cardiac malformations. The aim of the study was to determine if the deletion at 22q11.2 was present and could be responsible for both pathological processes. Del22q11.2 was identified in both cases. Comparative genomic hybridization revealed terminal gains on chromosomes 1q and Xq and terminal loss on 1p in the hepatoblastoma, and gains in 1p, 12q, 16p, 20q, 22q, and whole chromosome 19 and loss of Xq in the renal-cell carcinoma. Our results confirm a common genetic basis for cardiac malformations, and del22q11.2 presents a risk factor for the development of pediatric tumours.

Evolution of nonspecific duodenal lymphocytosis over 2 years of follow-up.

AIM To assess the evolution of duodenal lymphocytosis (DL), a condition characterized by increased intraepithelial lymphocytes (IELs), over 2 years of follow-up. METHODS Consecutive patients undergoing upper endoscopy/histology for abdominal pain, diarrhea, weight loss, weakness or other extraintestinal features compatible with celiac disease (CD) were included. Evaluation of IELs infiltrate in duodenal biopsy samples was carried out by CD3-immunohistochemistry and expressed as number of positive cells/100 enterocytes. Diagnostic agreement on the IELs count was tested by calculating the weighted k coefficient. All patients underwent serological detection of autoantibodies associated with CD: IgG and IgA anti-tissue transglutaminase and endomysium. Each patient underwent further investigations to clarify the origin of DL at baseline and/or in the course of 2 years of follow-up every six months. Autoimmune thyroiditis, intestinal infections, parasitic diseases, bacterial intestinal overgrowth, hypolactasia and wheat allergy were detected. Colonoscopy and enteric magnetic resonance imaging were performed when necessary. Risk factors affecting the final diagnosis were detected by multinomial logistic regression and expressed as OR. RESULTS Eighty-five patients (16 males, 69 females, aged 34.1 ± 12.5 years) were followed up for a mean period of 21.7 ± 11.7 mo. At baseline, endoscopy/duodenal biopsy, CD3 immunohistochemistry revealed: > 25 IELs/100 enterocytes in 22 subjects, 15-25 IELs in 37 and < 15 IELs in 26. They all had negative serum anti-transglutaminase and anti-endomysium, whilst 5 showed IgG anti-gliadin positivity. In the course of follow-up, 23 developed CD seropositivity and gluten sensitivity (GS) was identified in 19. Other diagnoses were: 5 Helicobacter pylori infections, 4 jejunal Crohns disease, 1 lymphocytic colitis and 1 systemic sclerosis. The disease in the remaining 32 patients was classified as irritable bowel syndrome because of the lack of diagnostic evidence. At multivariate analysis, the evolution towards CD was associated with an IELs infiltrate > 25 (OR = 1640.4) or 15-25 (OR = 16.95), human leukocyte antigen (HLA) DQ2/8 (OR = 140.85) or DQA1*0501 (OR = 15.36), diarrhea (OR = 5.56) and weakness (OR = 11.57). GS was associated with IELs 15-25 (OR = 28.59), autoimmune thyroiditis (OR = 87.63), folate deficiency (OR = 48.53) and diarrhea (OR = 54.87). CONCLUSION DL may have a multifactorial origin but the IELs infiltrate and HLA are strong predictive factors for CD development and a clinical diagnosis of GS.

Augmenter of liver regeneration, a protective factor against ROS-induced oxidative damage in muscle tissue of mitochondrial myopathy affected patients

Mitochondria-related myopathies (MM) are a group of different diseases defined by a varying degree of dysfunctions of the mitochondrial respiratory chain which leads to reactive oxygen species (ROS) generation followed by oxidative stress and cellular damage. In mitochondrial myopathy muscle tissue an overexpression of antioxidant enzymes has been documented probably as an attempt to counteract the free radical generation. We previously documented, in human non-pathological muscle fibres, the expression of the augmenter of liver regeneration (ALR), a sulfhydryl oxidase enzyme, whose presence is related to the mitochondria; indeed it has been demonstrated that ALR mainly localizes in the mitochondrial inter-membrane space. Furthermore we reported, in different experimental models, in vivo and in vitro, the anti-apoptotic and anti-oxidative capacities of ALR, achieved by up-regulating Bcl-2 anti-apoptotic family factors and the anti-apoptotic/anti-oxidative secretory isoform of clusterin (sClu). With the present study we aimed to determine ALR, Bcl-2 protein, clusterin and ROS expression in muscle tissue biopsies from MM-affected patients. Non-pathological muscle tissue was used as control. Enzymatic, histochemical, immunohistochemical and immune electron microscopy techniques were performed. The data obtained revealed in MM-derived muscle tissue, compared to non-pathological tissue, the over-expression of ROS, ALR and Bcl-2 and the induction of the nuclear, pro-apoptotic, isoform of clusterin (nCLU).

Survey of KRAS, BRAF and PIK3CA mutational status in 209 consecutive Italian colorectal cancer patients

Molecular testing for KRAS and BRAF mutations in tumor tissue is a fundamental tool to identify patients with metastatic colorectal cancer (CRC) who are eligible for anti-EGFR monoclonal antibody therapy. We here report a molecular analysis by high-resolution melting analysis and direct sequencing of KRAS, BRAF and PIK3CA hot spot mutations in 209 Italian CRC patients. One hundred and ten patients (51%) were identified who were potentially nonresponders to anti-EGFR therapy: 90/209 patients (43%) harboring KRAS mutations, 13/117 (11.1%) with the V600E BRAF mutation, and 7/209 (3.3%) with mutations in PIK3CA exon 20. The prevalence of BRAF and PIK3CA mutations was significantly higher in patients older than 65 years (p=0.014 and p=0.018), while patients with triple-negative tumors were significantly younger than mutation carriers (p=0.000011). Patients with gene mutations also showed a trend towards preferential tumor location in the colon (p=0.026). Moreover, although involving a relatively small number of samples, we report the presence of a discordant mutational profile between primary tumors and secondary lesions (3/9 patients), suggesting that it is worthwhile to test other available tissues in order to better define the efficacy of targeted therapy. Further correlations of specific clinical features with tumor mutational profile could be helpful to predict the response of CRC patients to monoclonal antibody therapy.

Histology of micro polyps in chronic endometritis.

(P = 0.065) by univariate analysis (v) (Table 1). Patients with a negative MTA3 immunohistochemical staining reaction showed better progression-free (P = 0.008), cause-specific (P < 0.001) and overall (P = 0.012) survival by univariate survival analysis (Figure 3). Cox regression resulted in just one independent term that was predictive of progression-free survival, namely FIGO stage (P < 0.001). Independent prognostic factors for cause-specific survival were FIGO stage (P = 0.004), LVSI (P = 0.025), and MTA3 positivity (P = 0.001). Overall survival was also influenced only by FIGO stage (P < 0.001) (Table 2). The data obtained revealed that MTA3 can be upregulated in human cancer cells and might contribute to a more aggressive phenotype. However, as only a few target genes of MTA3 have been identified, the consequence of MTA3 overexpression in human cancer cells is far from being understood. We here demonstrate for the first time that MTA3 is upregulated in advanced stages of uterine non-endometrial cancer, and is associated with FIGO surgical stage, lymph node involvement, and LVSI. In addition, MTA3 staining was associated with progression-free, cause-specific and overall survival of patients with non-endometrioid carcinomas. Overall, immunolabelling of MTA3 represents an independent prognostic factor for causespecific survival, suggesting that this antigen could be used as a simple and efficient parameter with which to identify high-risk patients.

Seronegative Celiac Disease and Immunoglobulin Deficiency: Where to Look in the Submerged Iceberg?

In the present narrative review, we analyzed the relationship between seronegative celiac disease (SNCD) and immunoglobulin deficiencies. For this purpose, we conducted a literature search on the main medical databases. SNCD poses a diagnostic dilemma. Villous blunting, intraepithelial lymphocytes (IELs) count and gluten “challenge” are the most reliable markers. Immunohistochemistry/immunofluorescence tissue transglutaminase (tTG)-targeted mucosal immunoglobulin A (IgA) immune complexes in the intestinal mucosa of SNCD patients may be useful. In our experience, tTG-mRNA was similarly increased in seropositive celiac disease (CD) and suspected SNCD, and strongly correlated with the IELs count. This increase is found even in the IELs’ range of 15–25/100 enterocytes, suggesting that there may be a “grey zone” of gluten-related disorders. An immune deregulation (severely lacking B-cell differentiation) underlies the association of SNCD with immunoglobulin deficiencies. Therefore, CD may be linked to autoimmune disorders and immune deficits (common variable immunodeficiency (CVID)/IgA selective deficiency). CVID is a heterogeneous group of antibodies dysfunction, whose association with CD is demonstrated only by the response to a gluten-free diet (GFD). We hypothesized a familial inheritance between CD and CVID. Selective IgA deficiency, commonly associated with CD, accounts for IgA-tTG seronegativity. Selective IgM deficiency (sIgMD) is rare (<300 cases) and associated to CD in 5% of cases. We diagnosed SNCD in a patient affected by sIgMD using the tTG-mRNA assay. One-year GFD induced IgM restoration. This evidence, supporting a link between SNCD and immunoglobulin deficiencies, suggests that we should take a closer look at this association.