An ultrasonography-cytology protocol for the diagnostic management of regional nodes in a subset of patients with Merkel cell carcinoma of the skin.

BACKGROUND: The status of regional lymph nodes (LNs) is one of the most consistent predictors of survival in Merkel cell carcinoma (MCC). In cases of clinically localized disease, current practice involves sentinel lymph node (SLN) assessment. OBJECTIVES: To propose ultrasonography (US) followed by fine needle aspiration cytology (FNAC) and immunohistochemistry as a useful diagnostic tool in the pre-surgical management of patients with MCC. METHODS: US of LNs was performed in 75 patients with MCC (22 with stage III tumours; 53 with stage I-II). In patients with US suspected disease, US coupled with FNAC of the LN was performed. Smears were examined by routine cytological staining supplemented with immunohistochemical staining for cytokeratin 20. All patients underwent surgical removal of regional LNs. RESULTS: In all 22 patients with stage III tumours, US was indicative of tumour deposits and FNAC confirmed metastases to LNs. In 11 of 53 patients with localized MCC without clinical evidence of nodal disease, US revealed enlarged, equivocal nodes where FNAC was performed. Ten LNs were cytologically positive for metastases, and one was negative. Upon histological examination, the FNAC-negative case showed a metastasis 5 mm in diameter. In all the other 42 cases with no clinical or US evidence of LN involvement, only SLN biopsy was performed and in six cases small metastatic foci were detected. Ultimately, of the 53 stage I-II MCC, 17 had positive LN involvement. In 10 cases (59%) metastases were detected by FNAC, and in seven cases, were detected by SLN biopsy. CONCLUSIONS: In a selected subset (∼20%) of patients with MCC with clinically localized disease, US followed by FNAC in the suspect LN is a valid alternative to the classical protocol of SLN histological examination.

European guidelines for quality assurance in colorectal cancer screening and diagnosis. First Edition--Quality assurance in pathology in colorectal cancer screening and diagnosis.

Multidisciplinary, evidence-based guidelines for quality assurance in colorectal cancer screening and diagnosis have been developed by experts in a project coordinated by the International Agency for Research on Cancer. The full guideline document covers the entire process of population-based screening. It consists of 10 chapters and over 250 recommendations, graded according to the strength of the recommendation and the supporting evidence. The 450-page guidelines and the extensive evidence base have been published by the European Commission. The chapter on quality assurance in pathology in colorectal cancer screening and diagnosis includes 23 graded recommendations. The content of the chapter is presented here to promote international discussion and collaboration by making the principles and standards recommended in the new EU Guidelines known to a wider professional and scientific community. Following these recommendations has the potential to enhance the control of colorectal cancer through improvement in the quality and effectiveness of the screening process, including multi-disciplinary diagnosis and management of the disease.

European guidelines for quality assurance in colorectal cancer screening and diagnosis. First Edition--Annotations of colorectal lesions.

Multidisciplinary, evidence-based guidelines for quality assurance in colorectal cancer screening and diagnosis have been developed by experts in a project coordinated by the International Agency for Research on Cancer. The full guideline document covers the entire process of population-based screening. It consists of 10 chapters and over 250 recommendations, graded according to the strength of the recommendation and the supporting evidence. The 450-page guidelines and the extensive evidence base have been published by the European Commission. The chapter on quality assurance in pathology was supplemented by an annex describing in greater detail some issues raised in the chapter, particularly details of special interest to pathologists. The content of the annex is presented here to promote international discussion and collaboration by making the issues discussed in the guidelines known to a wider professional and scientific community.

The natural history of colorectal adenomas and early cancer.

Adenomas represent the morphological precursors of the vast majority of colorectal cancers: although every adenoma has the capacity of malignant evolution, most adenomas stabilize their progression or even regress. Pathological factors are predictive of the natural history of adenomas in terms of potential and time interval for becoming malignant. Regression of adenomas is histologically well established, but it is thought to be a dynamic process, with cycling phases of regression and growth. Colorectal carcinoma invading the submucosa but not the muscular layer represents the earliest form of clinically relevant colorectal cancer. Grade of differentiation of carcinoma, lymphovascular invasion, and state of the resection margin predict the risk of metastasis. Microstaging of invasive cancer together with tumuor budding allow the metastatic risk to be further stratified into minimal, low, and high. Two distinct profiles are identifiable in the natural history of cancerous adenomas: blocking the growth of early cancer and allowing its progression towards advanced cancer. Thus, biomarkers to distinguish between progressive and non-progressive pT1 neoplasia are needed.

CD31 and CD34 expression as immunohistochemical markers of endothelial transdifferentiation in human cutaneous melanoma.

INTRODUCTION: Vasculogenic mimicry, as previously described in aggressive melanoma, is characterized by the de novo generation of intratumoral patterned vascular channels, composed of PAS-positive basement membrane in the absence of endothelial cells, providing additional microcirculation, in support to the classic tumoral angiogenesis. METHODS: We investigated the immunohistochemical expression of two endothelial markers, CD31 and CD34, in tumoral cells of 60 melanomas (45 primary cutaneous and 15 metastatic) as possible evidence of vasculogenic mimicry. In addition we investigated the relationship between CD31 and CD34 expression and three pathological markers such as Clark's level, and skin ulceration, predictive of melanoma's aggressive behaviour, and mitotic index. RESULTS: No cases of common melanocytic nevi immunoreacted with CD31 or CD34. Random CD31 immunoreactivity was present in 6% of Clark's level I/II, 50% of Clark's level III and 80% Clark's level IV/V. CD34 was negative in Clark's level I/II but randomly stained the 20% and 55% of level III and IV/V respectively. 66% (10/15) of metastatic melanomas were CD31 positive showing a canalicular immunostaining pattern, conversely CD34 expression was never found. 7/8 cutaneous ulcerated melanomas immunostained for CD31 and 4/8 for CD34. CD31 immunostained 88% high/intermediate MI, and 53% of low MI melanomas. CD34 decorated the 29% of high/intermediate and 38% of low MI melanomas. CONCLUSIONS: CD31 and CD34 immunoreactivity closely parallel both the different morphologic steps of melanocytic tumor progression and the presence of histological parameters related to the aggressive behaviour. Their expression could be related to endothelial transdifferentiation of melanoma cells although a consequent functional role has not been demonstrated yet.

Correlations between rectal mucosa cell proliferation and the clinical and pathological features of nonfamilial neoplasia of the large intestine.

An in vitro study of proliferative activity as shown by immunohistochemical detection of the uptake of bromodeoxyuridine was run on rectal biopsies from 400 patients with nonfamilial large bowel neoplasia: 200 adenoma; 150 adenocarcinoma; 50 adenoma plus adenocarcinoma. The controls were 400 subjects with negative personal and family histories of colorectal neoplasia. The number and height distribution of bromodeoxyuridine positive cells were determined by dividing the crypt into five longitudinal compartments. The total labeling index and the labeling index of each compartment were higher in all three groups compared with the controls. In subjects with adenoma, total labeling index and labeling index values were correlated with tumor size and decreased in function of the duration of the polyp-free colon state. The major zone of DNA synthesis had shifted to the intermediate and surface crypt compartments in all three groups. This stage II abnormality was more marked in adenoma patients with a high degree of dysplasia and in those with adenoma plus adenocarcinoma. Hyperproliferation and the proliferative compartment shift are cytokinetic abnormalities that coexist in the flat rectal mucosa of patients with colorectal neoplasia. Nonetheless, they are independent, controlled by different factors, and are expressions of different biological aspects of large bowel carcinogenesis.

Microsatellite instability is associated with the histological features of the tumor in nonfamilial colorectal cancer.

Replication errors (RERs) at microsatellite loci were examined in 46 specimens of nonfamilial colorectal cancer. Somatic microsatellite alterations in at least two genetic loci, D11S904, D13S175, D2S123, and D10S197, consistent with a RER(+) phenotype were found in four cases (8.7%). Six additional cases (13%) showed alterations at a single locus. Mucinous differentiation was observed in 3 of 4 (75%) adenocarcinomas with a RER(+) phenotype and only in 19% (8 of 42) of RER(-) adenocarcinomas (P < 0.05). A distinct cap of inflammatory cells at the advancing edge of the tumor and Crohn's-like reaction in peritumoral stroma were histologically identified in 50 and 25% of RER(+) and in 5 and 0% of RER(-) tumors, respectively (P < 0.05). Also, the plexiform pattern of growth of carcinoma turned out to be significantly associated with the RER(+) phenotype (P < 0.05). Mucinous differentiation and stromal inflammatory reactions are frequent features of hereditary nonpolyposis colorectal cancer in which germ-line mutations of mismatch repair genes cause genetic instability. Our results indicate that a link exists between such histological features and somatic genetic instability consistent with a RER(+) phenotype also in nonfamilial colorectal cancer.

Apoptosis, cell replication, and Western-style diet-induced tumorigenesis in mouse colon.

In this study, feeding Western-style diets (WDs) to mice for a duration of two years without any chemical carcinogen led to the development of gross colonic lesions that were histologically classified as dysplastic crypts and focal hyperplasias with or without atypical nuclei. To better understand early biological events contributing to the development of colonic neoplasia, grossly normal colonic mucosa was investigated; mitotic and apoptotic colonic epithelial cells, atypical mitosis, and atypical nuclei were studied. A significant and transient increase of mitotic activity in the basal and intermediate portions of the colonic crypts was seen in young mice after feeding them the WDs. This was accompanied by diffuse activation of apoptosis of the colonic epithelial cells. In the middle of the rodents' life span, after administration of both the WDs and control diet, the rodents developed a marked depletion of apoptotic epithelial cells in the mid-region of the colonic crypts; this was followed by the expansion of an epithelial cell population containing atypical nuclei, and the emergence of the gross lesions noted above. With this sequence of events, prolonged feeding of WDs to mice produced single-crypt dysplastic lesions and focal hyperplasias indicative of tumorigenesis.

Serum bile acids, programmed cell death and cell proliferation in the mucosa of patients with colorectal adenomas.

BACKGROUND: Deoxycholic acid induced programmed cell death and an imbalance with cell proliferation may favour colorectal tumourigenesis according to 'in vitro' studies, but information is lacking on the relationships occurring 'in vivo' in humans. AIMS: To evaluate whether serum deoxycholic acid is associated with programmed cell death and cell proliferation in colonic mucosa. METHODS: In 10 patients with colorectal adenomas, we measured fasting serum levels of bile acids; and, in normal colonic mucosa, programmed cell death by the TUNEL technique and cell proliferation by immunohistochemical staining with anti-Ki67. Total and compartmental indices for both activities were calculated. RESULTS: Among serum bile acids, only total deoxycholic acid (median: 0.89 micromol/L +/- 0.54 95% CI), showed a significant positive correlation with the total and basal compartments PCD Index (r = 0.68, p < 0.05). Total proliferation index showed no correlation with either total PCD Index, or bile acids. Within the median compartment of the crypt, cell proliferation was negatively associated with all unconjugated bile acids. CONCLUSIONS: The positive association between deoxycholic acid and programmed cell death in the basal compartment of the crypt, and the negative association of cell proliferation and unconjugated bile acids in the median compartment, do not seem to support the co-carcinogenic effect of deoxycholic acid.

Bromodeoxyuridine uptake and proliferating cell nuclear antigen expression throughout the colorectal tumor sequence.

In vitro uptake of bromodeoxyuridine and expression of proliferating cell nuclear antigen (PCNA) were evaluated histochemically in rectal mucosa of control subjects and subjects with colorectal neoplasia in large intestine adenomas and adenocarcinomas. Both labeling indices progressively increased along the path of tumor progression, as did the difference between them (PCNA labeling indices were always greater than those of bromodeoxyuridine). The correlation between them was fairly close in the controls and in adenomas with low-grade dysplasia, whereas no significant linear relations were noted in adenomas with high-grade dysplasia or in adenocarcinomas. The progressive increase in PCNA would thus seem to be related to both hyperproliferation and neoplastic deregulation of PCNA synthesis. In the mucosa of subjects with colorectal neoplasia, PCNA labeling revealed hyperproliferation but not the surface-wards shift of the proliferative compartment detected by bromodeoxyuridine. PCNA expression, therefore, is not a sufficiently sensitive marker of the risk of tumor transformation in the intestinal mucosa.

DNA-ploidy analysis within selected regions of colorectal adenomas containing carcinoma.

In order to better understand the relationship of DNA ploidy, dysplasia, early cancer, and colorectal tumor progression, 11 colorectal adenomas containing carcinoma invading the submucosa were investigated using DNA flow cytometry. Multiple frozen samples were taken from the selected sectors corresponding to adenoma tissue with low-grade dysplasia, high grade dysplasia and early cancer. Sampling accuracy was performed under histologic examination by multiple cryostatic sections. Data were compared with previously reported results in non-cancerous adenomas and advanced carcinomas. Incidence of DNA aneuploidy among the dysplastic regions of the adenomas containing carcinomas resulted higher than that observed in non-cancerous adenomas (p=0.02). Furthermore, among the DNA aneuploid populations, the frequency of clones with high DNA Index (DI>1.3) was slightly higher in adenomas with cancer than in adenomas without cancer (p=0.07). We suggest that differences may exist in DNA aneuploidy evolution between these two types of lesions. In early cancer, the near-diploid clones were 57% with respect to 18% (p=0.01) in advanced cancer since in this latter case the majority of the DNA abnormal clones were in the near-hypertriploid region (82%). Thus, the acquisition of the invasive phenotype appears to be linked with the expansion and stabilization of high DNA aneuploid clones. Further analysis on a larger number of cases of adenomas containing carcinoma are necessary to validate these interpretations.

Preclinical mouse models for cancer chemoprevention studies.

To aid in identifying the ability of chemopreventive agents to inhibit tumor development, new preclinical in vivo rodent models have recently been developed. Some of the models contain targeted mutations capable of increasing the incidence and progression of neoplastic lesions, whereas in other models dietary nutrients induce preneoplastic lesions in normal mice. These new preclinical models are assisting the analysis of genetic and environmental factors leading to neoplasia, and clinical studies to evaluate the chemopreventive efficacy of specific nutrients and pharmacological agents.

Gastrointestinal stromal tumor, uncommitted type, with monosomies 14 and 22 as the only chromosomal abnormalities.

Karyotypic analysis of a gastric stromal tumor with the histologic and immunohistochemical features of a malignant, uncommitted lesion revealed clonal monosomies of chromosomes 14 and 22. Such changes, together with loss of chromosomes 15 and 18, as well as structural rearrangements involved chromosome 1, have been previously reported in gastrointestinal stromal tumors with smooth muscle differentiation. We suggest that monosomies of chromosomes 14 and 22 are early events in the malignant transformation of the mesenchymal cell-originating gastrointestinal stromal tumors.

Somatic alterations of the androgen receptor CAG repeat in human colon cancer delineate a novel mutation pathway independent of microsatellite instability.

The human androgen receptor gene contains a polymorphic CAG repeat region ranging from 8 to about 35 repeats in the normal human population. The repeat length is inversely related to the transactivation potential of the receptor. We have analyzed the repeat length in 50 sporadic colon cancer samples in comparison to surrounding healthy mucosa and have found somatic reductions of up to 10 repeats in 5 cases (10%), 3 of which were complex, probably involving both alleles. Alterations occurred in tumors with and without microsatellite instability indicating that they follow an independent mutation pathway. The similar repeat of the huntingtin gene did not show any somatic alterations in the same cases. No correlation to sex, tumor stage, location, or histology was evident. In the tumors that showed somatic reductions, the reduced allele was present in at least half of the cells and thus in most, if not all, of the tumor component of the sample. Somatic reductions of the androgen receptor CAG repeat thus occur frequently, through a pathway distinct from microsatellite instability and early during colon carcinogenesis. The receptor is expressed in most normal and neoplastic tissue samples analyzed. Apparent growth selection of cells bearing shortened AR alleles suggests that androgens contribute to colon carcinogenesis in a yet unknown way.

Cell proliferation in colorectal adenomas containing invasive carcinoma.

Immunohistochemical detection of the nuclear antigen recognised by the monoclonal antibody Ki67, DNA polymerase alpha, and the proliferating cell nuclear antigen (PCNA), and histochemical staining for the argyrophilic proteins associated with the nucleolar organizer regions (AgNOR) were carried out on histological sections from 107 colorectal adenomas containing invasive carcinoma (ACIC), including 7 with regional lymph node metastases. Separate evaluations were made for fields corresponding to adenoma with low-grade dysplasia, adenoma with high-grade dysplasia and early cancer. The same techniques were also employed in 20 cases of normal mucosa and 20 advanced carcinomas. The mean percentages of Ki67, DNA polymerase alpha, and PCNA-positive nuclei and the number of AgNOR per nucleus progressively increased along the sequence from normal mucosa via low-grade and high-grade dysplasia adenoma to advanced cancer, whereas the early cancer values were not significantly different from those in the low-grade dysplasia areas. No significant difference in PCNA positivity and number of AgNOR were noted in ACIC with and without lymph node metastases. It is suggested that the decrease in proliferative activity thus revealed in early cancer may be due to changes in the submucosa microenvironment caused by invasion, and that the metastatic potential of an early colorectal cancer cannot be correlated to such activity.

Small bowel tumors and polyposis syndromes.

Tumors of the small bowel are uncommon and seldom suspected on a clinical basis. Together with the relative inaccessibility of the small bowel to endoscopic investigation, the rarity of these tumors undoubtedly delays the diagnosis. Small bowel tumors may be an interesting field of application for enteroscopy, which now can be readily performed with dedicated enteroscopic evaluation in patients with suspected small bowel neoplasia could improve prognosis and treatment. Enteroscopy may also play an important role in the surveillance of inherited polyposis syndromes, as in other precancerous condition of the small bowel. In Peutz-Jeghers syndrome it may reduce polyp-induced complications and improve planning for surgery; in familial adenomatous polyposis it may contribute to preventing upper gastrointestinal tract cancer.

Histological aspects and healing rates of gastric ulcers treated with omeprazole 20 mg once daily or ranitidine 150 mg B.I.D.

The aim of this double blind trial was to compare omeprazole 20 mg once daily with ranitidine 150 mg b.i.d. in treatment of benign gastric ulcer, evaluating both rates and histological aspects of the ulcer healing process. Eighteen patients were randomized, 9 to each treatment; one patient (ranitidine group) was excluded from the analysis because of malignant ulcer. Omeprazole appeared to be more effective than ranitidine in healing gastric ulcer. A more rapid relief of symptoms was observed in the omeprazole group than in the ranitidine group. Both drugs reduced chronic atrophic gastritis (with a trend in favour of omeprazole), while omeprazole showed a prompter activity on the components of acute inflammation.

Sacrococcigeal and vertebral chordomas. Report of three cases and review of the literature.

Three cases of sacrococcygeal and vertebral chordoma are described. Histogenetic and anatomopathological aspects with particular reference to differential diagnosis from similarly distributed neoplasias are discussed. Anti-cytoskeleton monoclonal antibodies were used to this purpose. The clinical profile of sacrococcygeal and vertebral chordomas is characterized by a pronounced metastatic potential; radiation treatment can only partially counter their biological behaviour, while chemotherapy has proved little or no effectiveness. Where possible, radical surgery is currently the only treatment to guarantee long-term survival or complete cure. Palliative surgery, associate with efficient painkilling, offers a better quality of life and slows down the progress of the disease.

Intermediate biomarkers in the colorectal tumor progression.

Intermediate biomarkers are biological alterations in tissue which signal a stage of carcinogenesis between initiation and the development of a malignant tumor. Proliferation biomarkers are those that most satisfy the requisites for premorphological intermediate markers in the colorectal tumor progression. Cell proliferation changes in histologically normal intestinal mucosa are early events directly and closely associated with the morphogenesis of colorectal neoplasia. The transition from the morphological stage and the latter's further progression can be reliably monitored through the use of differentiation and genomic markers. In particular the incidence and the degree of DNA aneuploidy are indicators of the risk of malignant transformation in colorectal adenomas. New "regression-related" biomarkers should be investigated for the planning of measures designed to bring about the regression of premalignant lesions.