Francesco Cetta

Genetic Alterations in Thyroid Carcinoma Associated with Familial Adenomatous Polyposis: Clinical Implications and Suggestions for Early Detection

AbstractGerm-line mutations of the adenomatous polyposis ( APC) gene, responsible for familial adenomatous polyposis (FAP) were analyzed in 15 patients with FAP-associated papillary thyroid carcinomas: 13 had the mutation between codons 778 and 1309 (exon 15), 1 at codon 593 (exon 14), and 1 at codon 140 (exon 3). Therefore APC gene mutations clustered in the genomic area associated with congenital hypertrophy of the retinal pigment epithelium (CHRPE) (codons 463–1387). Ocular patches were documented in 12 patients. In particular, 4 of the 15 patients, all women with a mean age of 23.5 (range 20–32), were found during the study of 15 consecutive kindreds who had undergone systematic screening for extracolonic manifestations. Three of them belonged to the same kindred and were asymptomatic. These four patients were also screened for loss of heterozygosity of APC in the thyroid tumoral tissue. No biallelic inactivation of the APC gene was found. In contrast, three of these four patients had activation of theret-PTC oncogene. In particular, there was activation of the ret-PTC1 isoform, a chimeric gene resulting from fusion of a gene named H4 with the RET gene. On histologic examination, three of the four patients showed Hashimoto-like lymphocytic infiltration. Present data suggest that: (1) the incidence of FAP-associated thyroid cancer probably has been underestimated in the past; (2) intensive screening could detect a larger than expected number of thyroid carcinomas; (3) systematic screening is recommended in patients with ocular patches and genetic mutation in exon 15; (4) Hashimoto-like findings do not exclude carcinoma but are a frequent accompanying finding; (5) despite frequent multicentricity and early lymph node involvement, FAP-associated thyroid tumors seem to have an excellent prognosis, in particular those showing ret-PTC activation.

Evidence of digenic inheritance in Alport syndrome

Background Alport syndrome is a clinically heterogeneous, progressive nephropathy caused by mutations in collagen IV genes, namely COL4A3 and COL4A4 on chromosome 2 and COL4A5 on chromosome X. The wide phenotypic variability and the presence of incomplete penetrance suggest that a simple Mendelian model cannot completely explain the genetic control of this disease. Therefore, we explored the possibility that Alport syndrome is under digenic control. Methods Using massively parallel sequencing, we identified 11 patients who had pathogenic mutations in two collagen IV genes. For each proband, we ascertained the presence of the same mutations in up to 12 members of the extended family for a total of 56 persons studied. Results Overall, 23 mutations were found. Individuals with two pathogenic mutations in different genes had a mean age of renal function deterioration intermediate with respect to the autosomal-dominant form and the autosomal-recessive one, in line with molecule stoichiometry of the disruption of the type IV collagen triple helix. Conclusions Segregation analysis indicated three possible digenic segregation models: (i) autosomal inheritance with mutations on different chromosomes, resembling recessive inheritance (five families); (ii) autosomal inheritance with mutations on the same chromosome resembling dominant inheritance (two families) and (iii) unlinked autosomal and X-linked inheritance having a peculiar segregation (four families). This pedigree analysis provides evidence for digenic inheritance of Alport syndrome. Clinical geneticists and nephrologists should be aware of this possibility in order to more accurately assess inheritance probabilities, predict prognosis and identify other family members at risk.

Sporadic childhood hepatoblastomas show activation of β-catenin , mismatch repair defects and p53 mutations

Hepatoblastoma, a rare embryonic tumor that may arise sporadically or in the context of hereditary syndromes (familial adenomatous polyposis and Beckwith–Wiedemanns) is the most frequent liver cancer of childhood. Deregulation of the APC/β-catenin pathway occurs in a consistent fraction of hepatoblastomas, with mutations in the APC and β-catenin genes implicated in familial adenomatous polyposis-associated and sporadic hepatoblastomas, respectively. Alterations in other cancer-related molecular pathways have not been reported. We investigated a series of 21 sporadic paraffin-embedded hepatoblastoma cases for mutations in the p53 (exons 5–8) and β-catenin (exon 3) genes, loss of heterozygosity at APC, microsatellite instability and immunohistochemical expression of β-catenin and of the two main mismatch repair proteins, MLH1 and MSH2. No loss of heterozygosity at APC was detected. We found mutations in β-catenin and p53 in 4/21 (19%) and 5/21 (24%) cases respectively, β-catenin protein accumulation in 14/21 cases (67%), microsatellite instability in 17/21 cases (81%), of which eight resulted positive for high-level of microsatellite instability (in four cases associated with loss of MLH1/MSH2 immunostaining). No correlations between involved molecular pathway(s) and hepatoblastoma histotype(s) emerged. This study confirms that β-catenin deregulation is involved in sporadic hepatoblastoma and also suggests that mismatch repair defects and p53 mutations contribute to this rare liver cancer. Sporadic hepatoblastoma appears to be molecularly and phenotypically heterogeneous and may reflect different pathways of liver carcinogenesis.

Rokitansky-Aschoff Sinuses of the Gallbladder are Associated with Black Pigment Gallstone Formation: A Scanning Electron Microscopy Study

Rokitansky-Aschoff sinuses are the result of hyperplasia and herniation of epithelial cells through the fibromuscular layer of the gallbladder wall and are usually referred to as adenomyomatosis. The role of this study is to demonstrate that Rokitansky-Aschoff sinuses of the gallbladder are a risk factor for the formation of black pigment gallstones. A total of 179 removed gallbladders, were hystologically examinated. Sixty-four of the 179 consecutive cholecystomized patients had typical adenomyomatosis. Thirty-eight of the 64 patients with adenomyomatosis had black pigment gallstones, alone ( n =22) or in association with single ( n =12) or multiple ( n =4) cholesterol gallstones in the same gallbladder. Twelve of these patients did not have the typical risk factors for black stones (hemolysis, cirrhoses, gastrectomy, etc). Gallstones were examined by infrared spectroscopy and X-ray diffractometry. In addition, in a subset of 14 patients, the gallstones and the gallbladder wall were examined by scanning electron microscopy. At least in the initial phases of formation, Rokitansky-Aschoff sinuses were found close to small intraparietal vessels and sometimes they contained black pigment microstones. After the fourth to fifth decades of life, black gallstones can be found in the Rokitansky-Aschoff sinuses and in the main gallbladder lumen. Black pigment gallstones can form in Rokitansky-Aschoff sinuses of the gallbladder in absence of the typical risk factors for bilirubin suprasaturation of bile.

Mutational screening of the RB1 gene in Italian patients with retinoblastoma reveals 11 novel mutations

AbstractRetinoblastoma (RB, OMIM#180200) is the most common intraocular tumour in infancy and early childhood. Constituent mutations in the RB1 gene predispose individuals to RB development. We performed a mutational screening of the RB1 gene in Italian patients affected by RB referred to the Medical Genetics of the University of Siena. In 35 unrelated patients, we identified germline RB1 mutations in 6 out of 9 familial cases (66%) and in 7 out of 26 with no family history of RB (27%). Using the single-strand conformational polymorphism (SSCP) technique, 11 novel mutations were detected, including 3 nonsense, 5 frameshift and 4 splice-site mutations. Only two of these mutations (1 splice site and 1 missense) were previously reported. The mutation spectrum reflects the published literature, encompassing predominately nonsense or frameshift and splicing mutations. RB1 germline mutation was detected in 37% of our cases. Gross rearrangements outside the investigated region, altered DNA methylation, or mutations in non-coding regions, may be the cause of disease in the remainder of the patients. Some cases, e.g. a case of incomplete penetrance, or variable expressivity ranging from retinoma to multiple tumours, are discussed in detail. In addition, a case of pre-conception genetic counselling resolved by rescue of banked cordonal blood of the affected deceased child is described.

Gallbladder cancers: associated conditions, histological types, prognosis, and prevention.

Introduction Gallbladder cancer has a poor prognosis, with a reported 5-year survival of 5%. The prognosis improves when an R0 resection is feasible, but an early diagnosis is rare. The aim of the present study is to analyze the different conditions associated with gallbladder carcinomas and to report the main prognostic factors for these tumors to enable prevention. Materials and methods From 1986 to 2012, 75 patients were found to have gallbladder cancer during the study of 2942 patients affected by biliary tract diseases; 34 of these patients had gallbladder and gallstones, and had been subjected to bile analysis. Pancreatobiliary reflux was studied by biliary trypsin and C-Ki-ras genes were analyzed in 11 cases. Results Gallstones were found in 72 of 75 gallbladder cancer patients; in particular, large gallstones were associated with 88.88% of squamous-cell carcinoma, 68.2% of adenocarcinoma, and never with papillary adenocarcinoma. Pancreatobiliary reflux was associated with papillary adenocarcinoma in 100% of cases, but seldom with squamous cell carcinoma. C-Ki-ras mutations were found in 100% of patients with papillary carcinoma. Discussion and conclusion R0 resection in in-situ cancer has the best prognosis. Preventive cholecystectomy should be performed in high-risk patients (gallstones larger 3 cm; adenomatous polyps>1 cm; pancreatobiliary reflux, porcelain gallbladder, segmental adenomyomatosis, xanthogranulomatous cholecystitis). The histological stratification of gallbladder cancer should be carried out before starting further studies because squamous-cell carcinoma, adenocarcinoma, and papillary carcinoma are associated with different risk factors and genetic mutations and have different responsiveness to chemotherapies.

Short-term effects of neoadjuvant chemoradiotherapy on internal anal sphincter function: a human in vitro study.

BACKGROUND: Neoadjuvant chemoradiotherapy is recommended in the treatment of locally advanced rectal cancer. Studies have suggested that chemoradiotherapy adversely affects anorectal function. However, the functional implication and the underlying neuromyogenic changes involved in radiation-induced damage are poorly understood. OBJECTIVE: This study evaluated the functional changes following chemoradiotherapy on the internal anal sphincter. DESIGN AND PATIENTS: This article describes an in vitro study on the internal anal sphincter collected from patients undergoing abdominoperineal resection or proctectomy. Five patients were treated by surgery alone (control group), and 6 received preoperative chemoradiotherapy (treatment group). Sphincter strips were mounted in organ bath, and the responses to electrical field stimulation and drugs were monitored. SETTINGS: The study was performed at the University of Oxford. MAIN OUTCOME MEASURES: The end points of this study were to investigate whether chemoradiotherapy has any significant effects on internal anal sphincter function and, subsequently, to establish the type of injury induced. RESULTS: Chemoradiotherapy strips developed similar tone, but significantly lower spontaneous activity (p = 0.001) than controls. Electrical field stimulation induced relaxation, followed by contraction. At 50 Hz, electrical field stimulation produced 25.6 ± 4.9% (mean ± SE) of maximum relaxation followed by a contraction of 5.5 ± 0.9% of basal tone in chemoradiotherapy strips i9n comparison with 47.0 ± 6.2% (p = 0.009) and 17.7 ± 4.0% (p = 0.007) in controls. Relaxation was significantly attenuated by N&ohgr;-nitro-L-arginine. Significant differences were found in responses to carbachol (p = 0.018) and phenylephrine (p = 0.022), but not to sodium nitroprusside. LIMITATIONS: This work was limited by the relatively small number of patients enrolled, because of the difficulty of finding human tissue for laboratory studies, and the lack of long-term results. CONCLUSIONS: Chemoradiotherapy significantly impairs internal anal sphincter function and intrinsic nerves seem more susceptible than smooth muscle. The exclusion of anal canal from the radiation field is recommended, when oncologically safe.

Large Foreign Body as a Nidus for a Common Duct Stone in a Patient Without Spontaneous Biliary Enteric Fistula or Previous Abdominal Surgery

We report a case of a brown pigment gallstone, which formed around a phytobezoar in the common bile duct, in a patient without spontaneous biliary enteric fistula or previous abdominal surgery. A brief comment on the possible origin of the phytobezoar in this case and on the pattern of deposition of brown material over a pre-existent nidus is also presented.

Black pigment gallstones with cholesterol gallstones in the same gallbladder - 13 Cases in a surgical series of 1226 patients with gallbladder stones

We studied 1312 consecutive patients who underwent surgery for gallstones in the biliary tract at one university hospital in Siena, Italy, with a systematic classification of gallstones found within the gallbladder. Of these patients, 1226 were found to have gallbladder stones; 94 of these had black pigment gallstones. Of these, 13 patients were found to have black pigment gallstones and cholesterol gallstones within their gallbladder. They all had multiple black pigment gallstones, usually very small (all <6 mm diameter), in association with larger cholesterol stones in the gallbladder lumen. The cholesterol gallstones were single in seven cases, double in two cases, and multiple in four cases. All 13 of these patients with black pigment stones in association with cholesterol stones had histologic evidence of either adenomyomatosis or Rokitansky-Aschoff sinuses in the gallbladder wall. In nine of the 13 patients, the black pigment stones were located both in the gallbladder lumen and in close association with the gallbladder wall (in areas of adenomyomatosis or in Rokitanski-Aschoff sinuses). In the other four patients, the stones were found in close association with the gallbladder wall alone and not freely mobile within the gallbladder lumen. It is concluded that cholesterol stones and black pigment stones may be found in the same gallbladder. This association is infrequent with an incidence of 13 of 1226 (1.06%) in our series. There appears to be some relationship between the formation of the black pigment stones and the presence of adenomyomatosis or Rokitanski-Aschoff sinuses. However, the pathogenesis of these two compositionally distinctly different types of stones within the same gallbladder is not understood and deserves further study.

New biliary endoprosthesis less liable to block in biliary infections: description and in vitro studies.

OBJECTIVE To test in vitro stents made from a new biomaterial that is less liable to encourage adhesion of bacteria that may lead to blockage of the stent. DESIGN Laboratory experiment. SETTING University hospital, Italy. MATERIAL 15 polyethylene biliary endoprostheses that had been removed endoscopically a mean of 151 days (range 55416) after insertion. PUPA, a biomaterial that contains polyamidoamine cross-linked to polyurethane chains. This can bind large quantities of heparin and HyalSx (hyaluronic acid at a different stage of sulphation) in a stable fashion. MAIN OUTCOME MEASURES Incidence of blockage and growth of pathogens in the polyethylene biliary prostheses. Adhesion of pathogens to PUPA in vitro on electron microscopy. RESULTS 12 of 15 polyethylene prostheses were blocked by brown concretions composed of calcium bilirubinate, palmitate, and little cholesterol. All concretions grew more than one pathogen, and the growth always included Escherichia coli. Of the 5 PUPA stents tested, only 1 had bacteria sticking to their surfaces. CONCLUSION These results confirm previous studies that showed that HyalSx appreciably inhibited the adhesion of bacteria and is therefore a suitable material for the manufacture of biliary stents.