Protein profiling of non-malignant and malignant ascites by SELDI-TOF MS: proof of principle.

Ascites is a common clinical symptom in liver cirrhosis, inflammatory disorders of the abdomen and a major late manifestation of metastatic malignancies. Standard cytopathological techniques and immunocytochemistry have specificities and sensitivities of approximately 95 and 60%, respectively for the presence of tumor cells. Development of faster and more accurate screening methods would be of great clinical utility. In this work we examined differential analysis of the unbound proteins in the supernatant of ascites fluid by Protein-Chip SELDI mass spectrometry. There were 21 tumor cell-positive and 34 tumor cell-negative samples. We used principal component analysis coupled with linear regression applied to the mass spectra of the samples to distinguish between the sample groups. Two sample sets for statistical analysis were created after randomization, a training set with 37 samples and a validation set with 18 samples resulting in a specificity of 93% and a sensitivity of 83% on the training set. The validation set yielded a specificity and sensitivity of 75%. This study suggests that SELDI-TOF mass spectrometry appears to have great potential as a surrogate diagnostic tool to evaluate effusion specimens.

Pouchitis in ulcerative colitis: correlation between predictors from colectomy specimens and clinico-histological features.

Pouchitis after restorative proctocolectomy for ulcerative colitis is usually of ill-defined etiology and is encountered with sclerosing cholangitis, bacterial overgrowth, and ischemia. Recently, appendiceal involvement, ileitis, and fissures in the colectomy specimen have been associated with short- and long-term development of pouchitis. To corroborate these recent findings, the histology of 40 colectomies (70% males; mean age 46.3 years, age range 20-70 years; mean follow-up period 3.7 years, range 1-13 years) with yearly follow-up biopsies was correlated with pouchitis and clinical symptoms. Appendicitis, fissures, and ileitis were present in 47, 45 and 5% of the patients, respectively. Pouchitis in patients with appendicitis or with fissures was noted in 44 and 50% at first biopsy and in 70 and 58% during follow-up (p = NS). Of the patients without appendicitis or without fissures, 33 and 33% demonstrated pouchitis at the first biopsy and 30 and 55% during follow-up (p = NS). Clinico-histological correlation revealed normal/near-normal biopsies with the lowest clinical severity score in 77% and with the highest clinical score in 43% (p < 0.025). The histological findings of appendiceal involvement, fissuring ulcers, and ileitis in colectomies for ulcerative colitis do not correlate with the finding of pouchitis in early or late pouch biopsies. A high clinical suspicion score is frequently not correlated with significant inflammation of the pouch.

Laparoscopic colon resection for polyps: a good novice case?

PURPOSE: Laparoscopic colon resection of the endoscopically unresectable polyp has been considered an ideal case for the surgeon with moderate laparoscopic colectomy experience because tissues are not inflamed and the lymphadenectomy may not need to be as extensive compared with that required for cancer. To assess the appropriateness of this approach, we evaluated the incidence of invasive cancer in a series of laparoscopic colon resections for polyps. METHODS: A retrospective review was performed of 55 consecutive patients undergoing laparoscopic colon resection for endoscopically unresectable polyps during a 35-month period. Colonoscopy data, polyp characteristics, and final colon pathology were reviewed. RESULTS: On final pathologic examination, 18.2 percent of patients had invasive adenocarcinoma. Patient age, gender, indication for colonoscopy, polyp size, polyp location, polyp characteristics, and colonoscopic biopsy pathology were not predictive of adenocarcinoma on final pathology. CONCLUSIONS: A significant number of endoscopically unresectable polyps harbor adenocarcinoma, thereby requiring a formal lymphadenectomy at resection. Caution should be exercised when considering the laparoscopic resection of an endoscopically unresectable polyp as a "learning" case.

Observation of lymphatic vessels in orbital fat of patients with inflammatory conditions: a form fruste of lymphangiogenesis?

In the absence of antibodies specific for lymphatic vessels, analysis of lymphatic vessels within different tissues has been widely performed with light microscopic and, most importantly, electron microscopic techniques. In regard to lymphatic vessels in the ocular globe and the periocular structures, controversy remains about the specific distribution of lymphatic channels. It is postulated that bulbar and retrobulbar tissues are devoid of lymphatic vessels, but lymphatic vessels have been demonstrated in lacrimal gland and epibulbar conjunctiva. In this study, we analyzed orbital fat for the presence of lymphatic tissue using D2-40, a monoclonal antibody, specific for lymphatic vessels. We found lymphatic vessels present within bulbar conjunctiva extending to the level of the ciliary apparatus. No lymphatics were identified in healthy anterior orbital adipose tissue. In two cases of orbital mucor-mycosis and one case of panendophthalmitis, significant lymphovascular proliferation was present within granulation tissue associated with the acute inflammation. We conclude that lymph vessel proliferation may be induced in inflammatory conditions in tissues which are normally devoid of lymph channels.

Proliferation of D2-40-expressing intestinal lymphatic vessels in the lamina propria in inflammatory bowel disease.

Lymphatic vessels in the colon are normally distributed beneath the muscularis mucosae with rare branches reaching through the muscularis mucosae to the most basal aspect of the colonic crypts. In chronic inflammatory bowel disease demonstrating acute inflammation and architectural disarray, lymph vessel proliferation is seen within the lamina propria and within the submucosa. We analyzed the number and distribution of lymphatic vessels within the lamina propria and submucosa in chronic active and treated ulcerative colitis with restoration of architecture by immunostaining with D2-40, a specific monoclonal antibody against lymphatic vessels. We found significantly increased numbers of lymph vessels in chronic active ulcerative colitis both within the lamina propria and the submucosa as compared to normal mucosa. Numbers of lymph vessels in lamina propria were highest in severe chronic active ulcerative colitis and less in moderate and minimal residual disease with minimal architectural disarray (p<0.05). Lymph vessels in the submucosa were increased significantly above normal values in both severe, moderate and minimal residual disease. We conclude that lymph vessel distribution in chronic active ulcerative colitis extends into the lamina propria. With restoration of architectural morphology, the integrity of the lamina propria in regards to the distribution of lymph vessels is restored.

Identification of lymphatic vessels in malignant, adenomatous and normal colonic mucosa using the novel immunostain D2-40.

Distribution of lymphatic vessels in normal and neoplastic colon has been previously analyzed with electron microscopic techniques, as reliable antibodies have not been available for selective lymph vessel staining. A novel monoclonal antibody, D2-40, is recently available to differentiate lymphatic vessels from blood vessels. In this study, we analyzed the distribution of lymphatic vessels in normal colon, adenomas with and without superficial stalk invasion and invasive carcinomas without identifiable polypoid precursor lesions. In contrast to previous studies, we found lymphatic vessels in superficially misplaced stalk stroma in adenomas, and closely associated with early invasive epithelial nests in invasive lesions. Lymphatic vessels were identified within the lamina propria of the in situ aspect of in invasive tumors. We conclude that lymphatic vessel structures are seen more superficially in adenomas and invasive carcinomas than previously described. Since intramucosal carcinomas in adenomas do not metastasize, these lymph vessels may be immature or not communicate with deeper lymphatics. Proliferation and distribution of lymphatic vessels may be related to prognosis and early metastasis.

Sporadic adenomas in chronic ulcerative colitis.

Presence of polyps with intraepithelial neoplasia in patients with ulcerative colitis (UC) is associated with a very high risk of associated invasive carcinoma. Histologically, these lesions may not easily be differentiated from sporadic adenomas, which may occur in patients with UC. The treatment option for patients with polyps with intraepithelial neoplasia includes, at least in some centres, total colectomy which contrasts to polypectomy for patients with sporadic adenomas. Until recently, adenomatous polyps were not diagnosed in patients with UC because of the inability to differentiate adequately between UC-associated dysplastic polyps and adenomas. Multiple attempts have been made over the years to differentiate between UC-associated polyps with intraepithelial neoplasia and sporadic adenomas in patients with UC to ensure adequate treatment and follow-up for the correct diagnosis. These studies include morphologic, immunohistochemical and genetic analysis of lesions considered to be representative for each entity.

Intestinal spirochetosis: clinicopathological features with review of the literature.

OBJECTIVE: The purpose of this study is to evaluate the clinical presentation of patients with intestinal spirochetosis, as it is has not been well described in the literature. METHODS: We studied 15 patients with colonic biopsies that showed spirochetosis. The study group consisted of 11 males and 4 females, 10 of the males were heterosexual and one was homosexual. None of these patients were clinically immunocompromized. A colonoscopy was performed for evaluation of different symptoms. The histological diagnosis of spirochetosis was made on H&E examination and confirmed by Giemsa and/or Steiner stain. There was no evidence of associated active or any specific colitis in any of the cases. The control group consisted of 30 patients, 16 males and 14 females that were matched for age and clinical indications for obtaining a colonic biopsy. RESULTS: Colonic mucosa with prominent brush border-like surface colonized with large amounts of spirochetes was noted in all 15 cases in the study group and in none of 30 cases in the control group. The clinical presentation in patients with spirochetosis was compared to the control group. Of the spirochetosis patients 86% presented with some form of clinical symptoms compared with 13% of controls. These symptoms included chronic watery diarrhoea in 40% spirochetosis patients vs 5% in controls; a change in bowel habit was present in 33% of spirochetosis patients vs 3.3% in controls. The endoscopic appearance in spirochetosis patients was reported as normal in six patients, 'polyoid' in seven patients, erythematous in one patient and 'lesion' in one patient. CONCLUSION: These findings suggest that colonic spirochetosis affects a heterogenous group of patients, the majority of whom presented with gastrointestinal symptoms. The variable clinical findings may be related to the infecting organism and the condition may not be harmless in all patients.

Implication of the proliferation and apoptosis associated CSE1L/CAS gene for breast cancer development.

The CSEIL/CAS protein (CAS) is a Ran-binding protein with a function as a nuclear transport (export) factor. CSEIL/CAS, similar to Ran and other ran-binding proteins, plays at the same time an important role in the mitotic spindle checkpoint, which assures genomic stability during cell division. This checkpoint is frequently disturbed in neoplasms of various origin, including breast, hepatic and colonic tumors. CAS is located on chromosome 20ql3 and amplified in several cell lines, including breast, colon and bladder cancer. MEKl phosphorylation is known to be a reason for different CAS localization and activity. We evaluated the expression of CAS in 50 benign and malignant tumors of the breast by immunohistochemistry. Benign lesions of the breast (n=13) revealed a weak, predominantly cytoplasmatic CAS positivity. In ductal and lobular in situ carcinomas (n=17), 70-90% of the tumor cells were positive for anti-CAS staining which was predominantly cytoplasmatic. In invasive ductal and lobular carcinomas (n =20), 70-90% of the tumor cells stained positive with anti-CAS in a predominantly nuclear pattern. Different localization of CAS might affect its role not only for chromosome segregation, proliferation and apoptosis, but also its function in nuclear transport of proteins like retinoblastoma-gene-product, p53 and BRCAl. A different regulation in this checkpoint might contribute to the invasive potential in malignant carcinomas of the breast. Alteration of CAS-activity, possibly via MEKl-inhibition, might therefore be a possible option for breast cancer therapy.

Evaluation of the c-Met immunostain to detect malignant cells in body cavity effusions.

The cytologic diagnosis of malignant cells in serous effusions can be difficult. A wide variety of immunostains and other diagnostic techniques have been studied but without widespread acceptance of one staining panel or technique. Expression of the tyrosine kinase c-Met has been associated with several malignancies but not with benign mesothelium. We investigated the diagnostic value of the c-Met immunostain in serous effusions. Cell block material from 76 cases of unequivocally benign or malignant effusions were studied. Cases were stained with c-Met using the avidin-biotin complex method following antigen retrieval. The presence of strong granular cytoplasmic staining that was distinct from background staining was considered positive. Positive cells were identified in 38 of 42 (90%) malignant cases and in 18 of 34 (53%) benign cases. Typically, benign cases contained only individual positive cells, but positive cell clusters were also identified. The c-Met immunostain lacks sufficient specificity to be clinically useful in this cytologic setting. The expression of c-Met in benign mesothelium may reflect mesothelial proliferation.

Frequent hypermethylation of the 5' CpG island of E-cadherin in esophageal adenocarcinoma.

PURPOSE: E-cadherin, a M(r) 120,000 transmembrane glycoprotein, mediates calcium-dependent intercellular adhesion that is essential for normal tissue homeostasis. Loss of E-cadherin occurs in a variety of epithelial tumors and is correlated with invasion and metastasis. In esophageal adenocarcinoma, reduction of E-cadherin expression has been demonstrated previously, but mutations of the gene (CDH1) are rare. EXPERIMENTAL DESIGN: In this study, we used a nested PCR approach to examine the methylation status of the 5' CpG island of E-cadherin in esophageal specimens obtained from individuals with and without a history of esophageal cancer. RESULTS: In four individuals without esophageal cancer, E-cadherin was completely unmethylated in normal squamous cell-lined esophageal mucosa. In contrast, in patients with esophageal adenocarcinoma, E-cadherin was methylated in 26 of 31 (84%) tumor specimens. In the majority of cases, matched normal tissue (esophagus or stomach) from each patient was completely unmethylated. By immunostaining, methylated tumor samples demonstrated heterogeneously decreased membranous E-cadherin staining. CONCLUSIONS: These data suggest that epigenetic silencing via aberrant methylation of the E-cadherin promoter is a common cause of inactivation of this gene in esophageal adenocarcinoma.

Glucose transporter Glut-1 is of limited value for detecting breast carcinoma in serous effusions.

Diagnosing breast carcinoma that has metastasized to body cavity fluids can be difficult. Recently, immunostaining for the facultative glucose transporter Glut-1 has been described as a sensitive and specific means of detecting carcinomas in effusions. However, only five cases of breast carcinoma were studied. We examined Glut-1 specifically as a means of detecting breast carcinoma in effusion cytology. Using avidin-biotin immunocytochemistry, cell block material from 31 cases of breast carcinoma metastatic to body cavity effusions and 33 cases of benign effusions were studied. All cases were immunostained with the Glut-1 antibody. An additional set of slides from these same cases was stained for mucin using the Mayer's mucicarmine technique. Slides were graded for percentage of cells exhibiting immunoreactivity for Glut-1 or for the presence of mucin. Results of staining for both Glut-1 alone and in combination with mucicarmine were compared between the benign and malignant groups. Of the breast cancer cases, 19 of 31 (61%) were immunoreactive for Glut-1, and 25 of 31 (81%) were positive for either Glut-1 or mucicarmine. One of the 33 (3%) benign cases was immunoreactive for Glut-1, and none were positive for mucin. These data suggest that using Glut-1 as a single immunostain or in conjunction with mucicarmine is a specific but modestly sensitive means of detecting breast carcinoma in this cytologic setting.

Expression of survivin, YB-1, and KI-67 in sporadic adenomas and dysplasia-associated lesions or masses in ulcerative colitis.

Sporadic adenomas are said to exhibit an orderly growth pattern with a reversal of proliferative and apoptotic cell distribution as compared with normal colonic crypts. Dysplastic polyps of patients with ulcerative colitis (UC) may represent dysplasia-associated lesions or masses (DALM) with a high associated cancer risk, or, alternatively, may represent sporadic adenomas. Histologic criteria to differentiate between sporadic adenomas and DALM have not focused on the balance between cell renewal and cell loss. The expression of the novel anti-apoptosis gene product, survivin, and the proliferation markers, Ki-67 and Y-box binding protein (YB-1), were investigated by immunohistochemical localization in sporadic adenomas and DALM lesions of patients with UC. In adenomas, KI-67 was expressed preponderantly at the luminal aspect of the polyp, whereas its expression was diffuse in DALM. Survivin was detected diffusely in both adenomas and DALM. YB-1 showed positive staining in the deep aspect of adenomatous glands but only to a minor degree at the surface, whereas both deep and diffuse expression patterns of YB-1 were seen in DALM. The authors conclude that DALM and sporadic adenomas exhibit different patterns of cellular proliferation and that molecular markers of cell proliferation, Ki-67 and YB-1, may be useful to distinguish sporadic adenomas from DALM. However, the similar expression of survivin suggests that the underlying mechanisms that regulate apoptotic cell death are uniform in these lesions.

Lymphoid hyperplasia of the stomach: radiographic findings in five adult patients.

OBJECTIVE: The purpose of our study was to report the radiographic findings of biopsy-proven lymphoid hyperplasia of the stomach in five adult patients. CONCLUSION: Lymphoid hyperplasia of the stomach is characterized by distinctive findings on double-contrast upper gastrointestinal tract barium examinations; all five patients had innumerable tiny (1--3 mm in diameter) round frequently umbilicated nodules that carpeted the mucosa of the gastric antrum or antrum and body. Three of these five patients had associated Helicobacter pylori gastritis. The diagnosis of gastric lymphoid hyperplasia, therefore, can be suggested on the basis of the radiographic findings.

Glut-1 expression in dysplastic and regenerative lesions of the colon.

Monosaccaride transporter proteins are responsible for transmembrane transport of monosaccarides into cells. Glucose transporter protein 1 (Glut-1) is most prevalent in the cell membranes of erythrocytes and facilitates transport of glucose in tissues with barrier functions, i.e. blood brain barrier. Expression of Glut-1 in malignant tumors is increased due to increased metabolic need of the proliferating cell populations. In colorectal adenomas and carcinomas, membranous expression of Glut-1 has been associated with higher grade of tumors and decreased survival time. We studied the expression of Glut-1 in dysplastic proliferations of the colon which included sporadic adenomas and dysplasia associated lesions (DALM) in patients with ulcerative colitis and reactive/regenerative proliferations of the colon, including non-dysplastic chronic colitis, acute colitis and ischemia. Two patterns of Glut-1 expression were detected. Most adenomas and DALMs showed at least focal membranous expression of Glut-1. In addition a second staining pattern was recognized which consisted of prominent supranuclear dots. This pattern of staining was not only seen in adenomas and DALM but also in non-dysplastic areas immediately surrounding sporadic adenomas, in regenerative chronic colitis and in areas surrounding acute inflammation. Areas away from dysplasia did not show any positive staining for Glut-1. We conclude that two distinct patterns of Glut-1 expression may be found in colonic epithelial proliferation: membranous staining, associated with dysplasia, and, heretofore not described, supranuclear staining which may be related to Glut-1 expression secondary to expression of specific growth factors and not necessarily related to dysplasia.

Dieulafoy's lesion of the small bowel causing massive gastrointestinal bleeding: two case reports and literature review.

Dieulafoy's lesions are an often unrecognized cause of obscure, massive GI hemorrhage. Their diagnosis may elude conventional investigations, including upper and lower endoscopy, arteriography, and even laparotomy. In this paper, we report two cases of small-bowel Dieulafoy lesions. The first, a jejunal lesion, occurred in a young patient and was discovered at laparotomy. The second was an ileal Dieulafoy's malformation in an older patient. An intraoperative endoscopy with surgical guidance may be needed for definitive localization of this lesion.

An open-label trial of the PPAR-gamma ligand rosiglitazone for active ulcerative colitis.

OBJECTIVES: Previous research has demonstrated that ligands for the gamma subtype of peroxisome proliferator-activated receptors (PPARs) reduce inflammation in two different murine models of colitis. This study was designed to examine the potential efficacy of rosiglitazone, a ligand for the gamma subtype of PPARs, as a therapy for active ulcerative colitis. METHODS: Fifteen patients with mild to moderately active ulcerative colitis despite therapy with 5-aminosalicylic acid compounds were enrolled in an open-label study of rosiglitazone (4 mg b.i.d. p.o.) for 12 wk. Thirteen of 15 patients were receiving concomitant therapy with corticosteroids and/or immunomodulator medications. Disease activity was measured with the Disease Activity Index. RESULTS: After 12 wk of therapy, four patients (27%) had achieved clinical remission, of whom three (20%) also had an endoscopic remission. Four additional patients (27%) had a clinical response without achieving remission. Two patients were hospitalized with worsened disease activity, and one patient was withdrawn for nephrotic syndrome. CONCLUSIONS: These data suggest that ligands for the gamma subtype of PPARs may represent a novel therapy for ulcerative colitis. A double blind, placebo-controlled, randomized trial is warranted.

Diagnostic utility of BCA-225 in detecting adenocarcinoma in serous effusions.

OBJECTIVE: To determine the diagnostic value of the BCA-225 antibody in discriminating adenocarcinoma from benign mesothelium in body cavity effusions. STUDY DESIGN: One hundred four cases of unequivocally benign (34 cases) and malignant (70 cases) serous effusions with cell block material were immunostained for BCA-225 using the ABC method without antigen retrieval. The percentage of positively staining cells in each case was estimated in a blind fashion. RESULTS: BCA-225 stained at least 10% of morphologically malignant cells in 28 of 32 (88%) breast carcinomas and 58 of 67 (87%) adenocarcinomas overall. Neuroendocrine carcinomas (two cases) and one mesothelioma were positive in < or = 5% of their respective tumor cells. Of 34 benign cases, 6 (18%) exhibited positive staining, albeit in rare, morphologically benign cells. CONCLUSION: BCA-225 is able to discriminate adenocarcinoma from reactive mesothelium in cell block preparations and may prove useful as part of an antibody panel.