Carla Rabitti

Dilated Intercellular Spaces of Esophageal Epithelium in Nonerosive Reflux Disease Patients with Physiological Esophageal Acid Exposure

OBJECTIVES:It has been demonstrated that dilation of intercellular spaces of esophageal epithelium is a marker of tissue injury in GERD patients with a pathological esophageal acid exposure time. To evaluate the relationship among ultrastructural changes, acid esophageal exposure, and GERD symptoms, intercellular space diameters have been assessed in nonerosive reflux disease (NERD) patients with/without abnormal acid exposure time.METHODS:Following a pharmacological wash-out, 20 NERD patients underwent upper endoscopy, esophageal manometry, and 24-h pH monitoring. Biopsies were taken at 5 cm above the lower esophageal sphincter and intercellular space diameters were measured on transmission electron microscopy photomicrographs. Seven asymptomatic controls underwent the same protocol.RESULTS:Acid exposure time was in the normal range in all controls and in 11 patients (NERD pH-negative); it was abnormal in 9 patients (NERD pH-positive). Mean intercellular space diameter in NERD pH-negative and in NERD pH-positive patients was three times greater than in controls (1.45 and 1.49 μm vs 0.45, p < 0.001). Mean values of maximum intercellular spaces in all NERD patients were greater, two-fold or more, than those in controls (p < 0.001). No difference in mean and maximal space diameters was observed between NERD pH-positive and pH-negative patients.CONCLUSIONS:Dilation of intercellular spaces is a feature of NERD patients, irrespective of esophageal acid exposure, and can be considered an objective, structural marker of GERD symptoms. Impaired esophageal mucosal resistance, even to small amounts of acid refluxate, plays a key role in the pathophysiology of NERD.

Histopathology of the Supraspinatus Tendon in Rotator Cuff Tears

Background Causes of rotator cuff pathology are poorly understood. Hypothesis Macroscopically intact supraspinatus tendon may show profound light microscopy changes. These changes may be the pathogenic precursor to a subsequent rotator cuff tear. Study Design Comparative laboratory study. Methods Tendon samples were harvested from 88 individuals (49 men, 39 women; mean age, 58.2 years) who had sustained a rotator cuff tear and underwent arthroscopic repair of the lesion, and from 5 male patients who died of cardiovascular events (mean age, 69. 6 years). A full-thickness supraspinatus tendon biopsy specimen was harvested en bloc within the arthroscopically intact middle portion of the tendon between the lateral edge of the tendon tear and the muscle-tendon junction. Slides stained with hematoxylin and eosin were interpreted twice by the same observer using a semiquantitative grading scale assessing fiber structure and arrangement, rounding of the nuclei, regional variations in cellularity, increased vascularity, decreased colagen stainability, and hyalinization. Intraobserver reliability of the subscore readings was calculated. Results The mean pathologic sum-score of ruptured tendons was significantly greater than the mean pathologic score of control tendons (15.66 ± 1.82 vs 3.7 ± 2.31, P = .001). Within each specific category of tendon abnormalities, the control and ruptured tendons were significantly different <χ2 test); all variables were significantly different (Mann-Whitney U test <0.05; P = .001). The agreement between the 2 readings ranged from 0.56 to 0.86 (kappa statistics). Conclusion Nonruptured supraspinatus tendons, even at an advanced age, and ruptured supraspinatus tendons are clearly part of 2 distinct populations. Clinical Relevance During cuff repair, it is not necessary to excessively freshen the torn tendon to bleeding tissue: the macroscopically intact supraspinatus tendon is degenerated as well, and the failed healing response is not limited to the ends of the torn tendon.

Prognostic impact of VEGF, CD31, CD34, and CD105 expression and tumour vessel invasion after radical surgery for IB–IIA non-small cell lung cancer

Aims: To evaluate the prognostic impact of tumour angiogenesis assessed by vascular endothelial growth factor (VEGF), microvessel density (MVD), and tumour vessel invasion in patients who had undergone radical resection for stage IB–IIA non-small cell lung cancer (NSCLC). Methods: Fifty one patients (42 men, nine women; mean age, 62.3 years; SD, 6.9) undergoing complete surgical resection (35 lobectomy, 16 pneumonectomy) of pathological stage IB (n  =  43) and IIA (n  =  8) NSCLC were evaluated retrospectively. No patient underwent postoperative chemotherapy or neoadjuvant treatment. Tumour specimens were stained for VEGF and specific MVD markers: CD31, CD34, and CD105. Results: VEGF expression significantly correlated with high CD105 expression (p < 0.0001) and tumour vessel invasion (p  =  0.04). Univariate analysis showed that those patients with VEGF overexpression (p  =  0.0029), high MVD by CD34 (p  =  0.0081), high MVD by CD105 (p  =  0.0261), and tumour vessel invasion (p  =  0.0245) have a shorter overall survival. Furthermore, multivariate Cox regression analysis showed that MVD by CD34 (p  =  0.007), tumour vessel invasion (p  =  0.024), and VEGF expression (p  =  0.042) were significant predictive factors for overall survival. Finally, the presence of both risk factors, tumour vessel invasion and MVD by CD34, was highly predictive of poor outcome (odds ratio, 3.4; 95% confidence interval, 1.7 to 6.5; p  =  0.0002). Conclusions: High MVD by CD34 and tumour vessel invasion are more closely related to poor survival than the other neoangiogenetic factors in stage IB–IIA NSCLC. This may be because these factors are more closely related to the metastatic process.

Early cervical carcinoma

Although parametrectomy is the most difficult step in the surgical treatment of cervical carcinoma and is the main cause of postoperative complications, little attention has been given to the patterns of parametrial spread.

Nonrandom Distribution of Aberrant Promoter Methylation of Cancer-Related Genes in Sporadic Breast Tumors

Purpose: In an effort to additionally determine the global patterns of CpG island hypermethylation in sporadic breast cancer, we searched for aberrant promoter methylation at 10 gene loci in 54 primary breast cancer and 10 breast benign lesions. Experimental Design: Genomic DNA sodium bisulfate converted from benign and malignant tissues was used as template in methyl-specific PCR for BRCA1, p16, ESR1, GSTP1, TRβ1, RARβ2, HIC1, APC, CCND2, and CDH1 genes. Results: The majority of the breast cancer (85%) showed aberrant methylation in at least 1 of the loci tested with half of them displaying 3 or more methylated genes. The highest frequency of aberrant promoter methylation was found for HIC1 (48%) followed by ESR1 (46%), and CDH1 (39%). Similar methylation frequencies were detected for breast benign lesions with the exception of the CDH1 gene (P = 0.02). The analysis of methylation distribution indicates a statistically significant association between methylation of the ESR1 promoter, and methylation at CDH1, TRβ1, GSTP1, and CCND2 loci (P < 0.03). Methylated status of the BRCA1 promoter was inversely correlated with methylation at the RARβ2 locus (P < 0.03). Conclusions: Our results suggest a nonrandom distribution for promoter hypermethylation in sporadic breast cancer, with tumor subsets characterized by aberrant methylation of specific cancer-related genes. These breast cancer subgroups may represent separate biological entities with potential differences in sensitivity to therapy, occurrence of metastasis, and overall prognosis.

Intratumoural FOXP3-positive regulatory T cells are associated with adverse prognosis in radically resected gastric cancer.

We investigated the clinical significance of tumour-infiltrating FOXP3-positive regulatory T cells (Tregs) in radically resected (R0) gastric cancer. From a single-institution database, tumors of 110 patients who underwent R0 resection for stage II-III disease were studied for FOXP3-positive Tregs by immunohistochemistry. The observed median number of FOXP3-positive Tregs was used as the cut-point in analyses (<6 versus >or=6 count). Tregs were significantly higher in gastric carcinomas than in normal tissue (P = 0.0001). Tregs count >or=6 was significantly associated with vascular/lymphatic/perineural invasion (VELIPI) in the tumour (P = 0.03). Multivariate analysis showed association between adverse relapse-free survival and grading 3, stage III, VELIPI and Tregs count >or=6 (P = 0.02). Adverse overall survival was associated with grading 3, stage III, VELIPI and Tregs count >or=6 (P = 0.006). FOXP3-positive Tregs may be a novel marker for identifying high-risk gastric cancer patients. Present findings deserve additional investigation as Tregs may also represent an innovative therapeutic target.

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.

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 χ2 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 κ 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.

Prognostic Role of Thymidylate Synthase Polymorphisms in Gastric Cancer Patients Treated with Surgery and Adjuvant Chemotherapy

Purpose: To investigate the prognostic role of thymidylate synthase (TS) polymorphisms in gastric cancer patients treated with radical surgery and fluorouracil-based adjuvant chemotherapy. Experimental Design: Ninety gastric cancer cases were identified among 187 patients previously enrolled in prospective case-control studies for disease susceptibility. Patients were genotyped for a G/C nucleotide change within a triple 28 bp variable number of tandem repeat sequence in the TS 5′-untranslated region (5′-UTR) and a 6 bp deletion in the TS 3′-untranslated region (3′-UTR). According to available functional data, patients with 5′-UTR 2R/2R, 2R/3C, 3C/3C genotypes were classified as low TS producers (5′-UTRlow) and patients with 5′-UTR 3G/3G, 3G/3C, 2R/3G genotypes as high TS producers (5′UTRhigh). Patients with 3′-UTR del6/del6 and del6/ins6 genotypes were classified as low TS producers (3′-UTRlow) and patients with 3′-UTR ins6/ins6 genotype as high TS producers (3′-UTRhigh). The prognostic analysis was based on 5′-UTR/3′-UTR combined genotypes. Results: Ten patients (11%) were 5′-UTRhigh/3′-UTRhigh, 36 patients were 5′-UTRhigh/3′-UTRlow, 19 patients were 5′-UTRlow/3′-UTRhigh, and 25 patients were 5′-UTRlow/3′-UTRlow. 5′-UTRlow/3′-UTRlow patients showed the best outcome and the threshold of statistical significance was achieved in the comparison of disease-free survival and overall survival with 5′-UTRhigh/3′-UTRlow patients and 5′-UTRhigh/3′-UTRhigh patients. The presence of at least one high TS expression genotype showed independent adverse prognostic role in multivariate analysis. Conclusions: The prognostic role of TS polymorphisms in gastric cancer deserves further investigation because the adverse effect of high TS expression genotypes may be a relevant information to improve adjuvant chemotherapeutic strategies.

Mutations of the D310 mitochondrial mononucleotide repeat in primary tumors and cytological specimens

A mononucleotide repeat (D310) in mitochondrial DNA has been recently identified as a mutational hot spot in primary tumors. We analyzed 56 tumors for insertion/deletion mutations in the D310 repeat. A total of 13 mutations were detected. The highest frequency of mutations was found for cervical cancer, followed by bladder tumors, breast cancer and endometrial neoplasia. No alterations were observed in four patients suspected of malignancy but without evidence of malignant tumor. We detected identical changes in four of four urine sediments from patients with bladder cancer and in three of three fine needle aspirates of patients with breast cancer. Our results indicate that D310 abnormalities are detectable in cytology specimens from patients with cancer and support the notion that D310 analysis may represent a new molecular tool for cancer detection.