Do cell surface trafficking impairments account for variable cell surface sodium iodide symporter levels in breast cancer?

The Na(+)/I(-) symporter (NIS) is a transmembrane glycoprotein that mediates iodide uptake into thyroid follicular cells and serves as the molecular basis of radioiodine imaging and therapy for thyroid cancer patients. The finding that NIS protein is present in 80-90% of breast tumors suggests that breast cancer patients may also benefit from NIS-mediated radionuclide imaging and targeted therapy. However, only 17-25% of NIS-positive breast tumors have detectable radionuclide uptake activity. The discrepancy between NIS expression and radionuclide uptake activity is most likely contributed by variable cell surface NIS protein levels. Apart from the prevalent view that NIS cell surface trafficking impairments account for the variability, our current study proposes that differential levels of NIS expression may also account for variable cell surface NIS levels among breast tumors. We address the need to confirm the identity of intracellular NIS staining to reveal the mechanisms underlying variable cell surface NIS levels. In addition, we warrant a quantitative correlation between cell surface NIS levels and radionuclide uptake activity in patients such that the cell surface NIS levels required for radionuclide imaging can be defined and the defects impairing NIS activity can be recognized.

Her-2/neu overexpression in muscle-invasive urothelial carcinoma of the bladder: prognostic significance and comparative analysis in primary and metastatic tumors.

PURPOSE: The prognostic significance of Her-2/neu overexpression in muscle-invasive urothelial carcinoma of the bladder is largely unknown. Accurate determination of Her-2/neu overexpression may have therapeutic importance. EXPERIMENTAL DESIGN: Eighty consecutive cases of muscle-invasive urothelial carcinoma of the bladder treated by radical cystectomy with available follow-up were analyzed. In each case, one representative section was stained with anti-Her-2/neu. Staining was graded as 1 = faint/equivocal, 2 = moderate, and 3 = strong and was considered positive if > or =2. In those cases with a metastasis, the stain was also performed in the metastatic tumor. Results were correlated with survival. RESULTS: Twenty-two (28%) cases were considered Her-2/neu-positive in the primary tumor, and 17 of 32 (53%) were considered Her-2/neu-positive in the lymph node metastasis. Median survival for Her-2/neu-positive primary tumors was 33 months, compared with 50 months for Her-2/neu-negative cases (P = 0.46). Similarly, Her-2/neu overexpression in the lymph node metastasis did not predict survival. Sixty metastatic urothelial carcinomas were further studied by comparing Her-2/neu expression in the primary tumor with that of the lymph node and/or distant metastasis. Forty-five percent of Her-2/neu-negative primary tumors had a Her-2/neu-positive lymph node metastasis, whereas only one case (8%) of Her-2/neu-positive primary tumors was Her-2/neu-negative in the lymph node metastasis (P = 0.009). Similarly, 67% of Her-2/neu-negative primary tumors had a Her-2/neu-positive distant metastasis, whereas no Her-2/neu-positive primary tumor was negative in the metastasis (P = 0.429). CONCLUSIONS: Her-2/neu overexpression in primary or metastatic tumor did not predict survival in this cohort of muscle-invasive tumors. Overexpression in the primary tumors consistently predicts overexpression in a distant or regional metastasis. However, some Her-2/neu-negative primary tumors may show overexpression in their corresponding metastasis. Her-2/neu analysis in a metastasis may be necessary to accurately determine Her-2/neu status in metastatic bladder urothelial carcinoma.

Centrally necrotizing carcinomas of the breast: a distinct histologic subtype with aggressive clinical behavior.

Most breast carcinomas exhibit ductal differentiation. However, recognition of less common histologic patterns provides clinically useful data. This report describes a distinctive subtype of breast carcinoma that we have termed "centrally necrotizing carcinoma" (CNC; in this study, N = 34), which is characterized by an unusual and aggressive natural history. Centrally necrotizing carcinomas are composed of well-circumscribed, unicentric nodules with extensive central necrosis that are surrounded by a narrow rim of viable high-grade tumor cells. These tumor cells show minimal ductal differentiation (i.e., tubule formation), but are usually associated with focal ductal carcinoma in situ. The mean age of the patients in this study was 57.5 +/- 11.6 years, and the mean tumor size was 2.5 +/- 1.2 cm. Twenty-eight percent of the patients had positive axillary lymph nodes (mean number of lymph nodes involved, 2.1 +/- 1.2). Ninety-four percent of cases were negative for estrogen and progesterone receptors. In 21 patients (62%), local and/or distant recurrences developed (median time to recurrence, 16.2 months), and, to date, 20 have died from breast cancer (median time to death, 22.5 months). Progression of disease (defined as the development of either a recurrence or death resulting from disease) occurred in 24 patients (71%). Comparison with a set of 26 poorly differentiated ductal carcinomas with (nonextensive, patchy) necrosis matched for age, tumor size, and lymph node status showed a significantly worse progression-free survival rate for the CNC group (p < 0.004). We conclude that CNC is an uncommon but readily identifiable subtype of breast carcinoma and is characterized by early systemic metastasis and an accelerated clinical course.

Clinicopathologic significance of ductal carcinoma in situ in breast core needle biopsies with invasive cancer.

To assess whether the presence and amount of intraductal component (IC) in diagnostic needle core biopsies (NCB) is predictive of an extensive IC (EIC), the authors evaluated 50 invasive ductal carcinomas diagnosed with NCB, and then excised via lumpectomy, with regard to the extent of IC in both the NCB and subsequent lumpectomy specimen. These parameters were compared with each other and with the lumpectomy margin status. Extent of IC in the NCB was evaluated by dividing the number of ducts that contained IC by the total number of tissue cores. A ratio of more than 0.5 was considered EIC (EICc). IC extent in the lumpectomy was established by estimating the percentage of the tumor corresponding to IC and was considered extensive (EIC(L)) if more than 25% and if there was presence of IC away from the invasive tumor. The mean size of resected tumors was 1.6 +/- 0.7 cm. In 29 cases (58%) there was no IC in the NCB (NegICc), 11 cases (22%) exhibited nonextensive IC (NEICc), and 10 cases (20%) demonstrated EICc. A total of 7%, 36%, and 70% of the NegICc, NEICc, and EICc cases respectively had EIC(L)(p < 0.0001). The presence of EIC(L) correlated significantly with close or positive margin status for in situ disease (EIC(L) positive, 12 of 13 [92%] vs EIC(L) negative, 11 of 37 [30%]; p = 0.004). None of the NegICc, 27% of NEICc, and 40% of EICc had a positive margin for in situ neoplasm in the lumpectomy specimen (p = 0.004), and 24%, 18%, and 50% had positive margins for invasive neoplasm (p = not significant). The authors conclude that EICc predicts EIC(L) and constitutes a risk factor for positive lumpectomy margin status-particularly for in situ tumor. EICc may thus be of clinical value in identifying a subset of patients that requires a wider local excision.

Immunohistochemical study of microphthalmia transcription factor and tyrosinase in angiomyolipoma of the kidney, renal cell carcinoma, and renal and retroperitoneal sarcomas: comparative evaluation with traditional diagnostic markers.

Angiomyolipoma has a unique immunophenotype with co-expression of muscle-specific actin and melanocytic markers such as HMB-45 and Melan-A. The most recently developed melanocytic markers, microphthalmia transcription factor and tyrosinase, have not been studied in the diagnosis of angiomyolipoma. We tested 29 renal angiomyolipomas (21 classic histology, 4 epithelioid variants, 2 lipomatous variants, and 2 leiomyomatous variants) with an immunohistochemical panel, including microphthalmia transcription factor, tyrosinase, HMB-45, Melan-A, and muscle-specific actin. Results were compared with 15 renal cell carcinomas (9 conventional types, 6 with sarcomatoid change), 2 leiomyosarcomas, 5 liposarcomas, and 1 unclassified high-grade sarcoma. Microphthalmia transcription factor expression was seen in 22 of 29 angiomyolipomas, one renal cell carcinoma, and one well-differentiated liposarcoma (that is, 2 of 23 non-angiomyolipomas; sensitivity 75%, specificity 91%). Tyrosinase expression was seen in 4 of 29 angiomyolipomas and 0 of 23 non-angiomyolipomas (sensitivity 14%, specificity 100%). HMB-45 was positive in 24 of 29 angiomyolipomas and 0 of 23 non-angiomyolipomas (sensitivity 83%, specificity 100%). Melan-A was expressed by 25 of 29 angiomyolipomas and 0 of 23 non-angiomyolipomas (sensitivity 86%, specificity 100%). Muscle-specific actin was expressed by 29 of 29 angiomyolipomas and 2 of 23 non-angiomyolipomas (both leiomyosarcomas; sensitivity 100%, specificity 91% [100% excluding leiomyosarcomas]). Microphthalmia transcription factor showed the most widespread staining in angiomyolipoma (50% of cases staining more than half of the tumor cells) followed by Melan-A (24% of cases staining more than 50%). Only three cases showed positivity for all four melanocytic markers, while in one case each only microphthalmia transcription factor and Melan-A were positive. We conclude that microphthalmia transcription factor, but not tyrosinase immunostaining, has a sensitivity and specificity that rivals those of the established markers, HMB-45 and Melan-A, in the diagnosis of angiomyolipoma. Our data supports the use of a panel in difficult cases that includes antibodies to microphthalmia transcription factor, either Melan-A or HMB-45, and muscle-specific actin to provide the best mix of high sensitivity, high specificity, nuclear and cytoplasmic immunolocalization, and widespread staining of cells within a given tumor.

Development of gastrointestinal beta2-microglobulin amyloidosis correlates with time on dialysis.

Dialysis-associated beta2-microglobulin (beta2m) amyloidosis affects predominantly musculoskeletal tissue, but visceral involvement also occurs. To evaluate the clinical significance and prevalence of gastrointestinal beta2m amyloidosis, we studied hemodialysis patients admitted for gastrointestinal-related complaints. Hemodialysis patients (excluding those with non-beta2m amyloidosis) who were admitted with gastrointestinal complaints from 1984 to 1994 were identified. Gastrointestinal tissues from patients with available autopsy or surgical specimens were examined using hematoxylin and eosin stain, Congo red stain, and beta2m immunostain. Each case was evaluated independently by two pathologists and scored for quantity and location of beta2m amyloid and associated pathology. Of 24 patients, eight (four men and 4 women) had beta2m amyloid deposits within the gastrointestinal tract. Acute clinical presentation ranged from abdominal pain to gastrointestinal bleeding and was not significantly different for patients with or without gastrointestinal beta2m amyloid deposits. However, the mean time on dialysis of 15.3 +/- 5.7 years (range 6-24 years) for patients with gastrointestinal beta2m amyloidosis was significantly greater than that of patients without gastrointestinal beta2m amyloidosis (10.5 +/- 7.0 years, range <1 to 22 years, p < 0.05). Vascular histopathology ranged from mild focal thickening of vessel walls to massive vascular beta2m amyloid deposition with thrombosis. Extravascular beta2m amyloid ranged from mild to severe with marked expansion of the submucosa. Mucosal pathology ranged from none to severe ulceration. The degree of beta2m amyloid and the associated pathology tended to increase in severity with time on dialysis. Gastrointestinal beta2m amyloid deposition is an underappreciated complication of chronic hemodialysis that is significantly associated with increased time on dialysis. Gastrointestinal beta2m amyloidosis should be considered in any patient on hemodialysis 10 years or more who has gastrointestinal symptoms and can be identified in resection specimens as well as some biopsy specimens. Congo red stain and beta2m immunostains may be necessary for sensitive histopathologic evaluation of gastrointestinal beta2m amyloidosis.

Grading the invasive component of urothelial carcinoma of the bladder and its relationship with progression-free survival.

Although grading is valuable prognostically in pTa and pT1 papillary urothelial carcinoma, it is unclear whether it provides any prognostic information when applied to the invasive component in muscle-invasive carcinoma. The authors analyzed 93 cases of muscle-invasive urothelial carcinoma of the bladder treated with radical cystectomy for which follow-up information was available. Each case was graded using the Malmström grading system for urothelial carcinoma, applied to the invasive component. Pathologic stage, lymph node status, and histologic invasion pattern were also recorded and correlated with progression-free survival. Thirty-four cases (37%) were pT2, 40 (43%) were pT3, and 19 (20%) were pT4. Of the 77 patients who had a lymph node dissection at the time of cystectomy, 34 (44%) had metastatic carcinoma to one or more lymph nodes. The median survival for pT2, pT3, and pT4 stages was 85, 24, and 29 months, respectively (p = 0.0001). Lymph node-negative and lymph node-positive patients had a median survival of 63 and 23 months, respectively (p = 0.0001). Fifteen patients (16%) were graded as 2b and 78 patients (84%) were graded as 3. Median survival of patients graded as 2b was 34 months compared with 31 months for patients graded as 3 (p value not significant). Three invasive patterns were recognized: nodular (n = 13, 14%), trabecular (n = 39, 42%), and infiltrative (n = 41, 44%). The presence of any infiltrative pattern in the tumor was associated with a median survival of 29 months, compared with 85 months in tumors without an infiltrative pattern (p = 0.06). Pathologic T stage and lymph node status remain the most powerful predictors of progression in muscle-invasive urothelial carcinoma. In this group of patients histologic grade, as defined by the Malmström system and as applied to the invasive component, provided no additional prognostic information. An infiltrative growth pattern may be associated with a more dismal prognosis.

Clinicopathologic analysis of microscopically invasive breast carcinoma.

Breast biopsy or mastectomy cases having diagnoses of carcinoma in situ with "microinvasion," "minimal invasion," "focal invasion," or "suggestive of invasion" were reviewed and all histologically identified foci of invasive disease from each case were measured using an ocular micrometer. Cases in which any single focus of invasion was greater than 5 mm or the added size of separate invasive foci exceeded 10 mm were excluded, resulting in a study group of 75 patients. Invasive neoplasm was present in the initial biopsy in 69 of 75 cases (92%); however, residual invasive neoplasm was found in the subsequent lumpectomy/mastectomy from 14 of these (20%). In 59% of cases, two or more histologically separate foci of invasion were identified. Invasive foci consisted of isolated cells or cell clusters, each less than 1 mm (microfocal invasion), in 33% of cases. In 12 cases, the sum of individual invasive foci was 5 to 10 mm. Axillary lymph nodes (LN) from 5 of 69 patients (7%) contained metastatic carcinoma (four cases, one LN positive; one case, two LN positive). The cumulative sizes of all invasive foci in the LN-positive group were microfocal invasion (one case), 0.6 mm (one case), 1.1 mm, 2.5 mm, and 5.8 mm. The difference in frequency of axillary node metastasis between tumors with microfocal and measurable invasion (4.3% v 8.6%) was not statistically significant. Follow-up data were available on 55 cases (mean interval, 66.1 months). One (node-negative) patient had duct carcinoma in situ recurrence in the same breast 4 years after initial treatment. Another (with unknown node status) developed an axillary lymph node metastasis 13 months after initial treatment (96% disease-free survival). We conclude that microscopic stromal invasion in breast carcinoma, at least in the setting of significant in situ component, is often initiated from multiple foci. Patients with microscopically invasive breast carcinoma have a small but significant risk of axillary metastases, although a highly favorable survival.

Histopathologic analysis of chromosome aneuploidy in ductal carcinoma in situ.

Formalin-fixed, paraffin-embedded sections from 28 cases of ductal carcinoma in situ (DCIS; 12 with coexisting invasive neoplasm) were analyzed for numerical alterations of chromosomes 7, 8, 16, and 17 by performing fluorescence in situ hybridization (FISH) using centromeric (alpha-satellite) probes. Based on signal counts in 200 to 300 nuclei, each hybridization was classified as disomic (copy loss in <40%, copy gain in < 10%), monosomic (copy loss in at least 50% of nuclei, partial if 40% to 49%) or trisomic/polysomic (copy gain in at least 20% of nuclei, partial if 10% to 19%). Grade I lesions were characterized by complete lack of significant chromosome gain, but 29% showed partial (focal) monosomy. Grade III lesions, in contrast, showed partial or complete trisomy/polysomy in 88% of hybridizations versus monosomy in only 4%. Grade II DCIS exhibited a mixed pattern of chromosome aneuploidy: 38% hybridizations were disomic, 36% trisomic/polysomic, and 26% monosomic (8 of 10 hybridizations showing complete monosomy occurred in grade II lesions). Disomic hybridizations exhibiting rare cells (5% to 10%) with copy gain were more frequent in tumors with coexisting invasive neoplasm (5 of 17 v 2 of 33, P = .02). In morphologically heterogeneous lesions, higher-grade foci were characterized by chromosome copy gain relative to corresponding lower-grade areas in 17 of 22 (77%) hybridizations. These results show the presence of multiple (at least 3) distinct chromosome aneuploidy patterns in DCIS, in keeping with divergent mechanisms of genetic alteration. Degree of chromosomal instability, moreover, may correlate with progression of DCIS to invasive growth, implying that genetic instability is a parameter that impacts the likelihood of early breast carcinoma progression.

Biological implications of tumor cells in blood and bone marrow of pancreatic cancer patients.

BACKGROUND: Patients with pancreatic cancer often have tumor recurrence despite curative resection. Cancer cells detected in blood or bone marrow at the time of diagnosis may relate to tumor stage and to prognosis. Recent research emphasis has centered on tumor cells in bone marrow aspirates, but whether these represent early micrometastases or blood-borne cells in transit is unknown. PATIENTS AND METHODS: We developed a specific immunocytochemical assay that evaluated more than 5.3 x 10(6) extracted mononuclear cells per sample of blood and bone marrow and that could identify a single tumor cell in that population. The assay was applied to samples of blood and bone marrow from 105 patients with pancreatic cancer and 66 controls. The prevalence of isolated tumor cells was compared with Union Internationale Contre le Cancer (UICC) stage. A multivariate Cox regression analysis for survival was performed. RESULTS: Pancreatic cancer cells were detected in 26% of blood samples and in 24% of bone marrow specimens. Specificity for cancer was 96%. The prevalence of isolated tumor cells in patients with proven resectable cancer was 9% in blood and 13% in bone marrow. The prevalence increased with UICC tumor stage in blood (P =.04) but not in bone marrow (P =.52) and correlated in blood with resectability (P =.02), progression of disease (P=.08), and peritoneal dissemination (P =.003). While survival correlated significantly with tumor stage (P <.001) and isolated tumor cells in blood correlated with tumor stage, the finding of cancer cells in blood or bone marrow, or both, was not independently associated with survival in patients with pancreatic cancer. CONCLUSIONS: Isolated tumor cells in blood but not bone marrow reflect the stage of growth and spread of pancreatic cancer, particularly in the peritoneal cavity. The findings are consistent with cells in bone marrow aspirates being in transit, not implanted. These disseminated cancer cells may be the consequence, rather than the cause, of progression.

Chemotactic peptide uptake in acute pancreatitis: correlation with tissue accumulation of leukocytes.

Chemotactic peptides bind specifically to receptors on leukocyte membranes. This property makes them prospective vehicles to evaluate inflammation and infection. We used two well-established models of acute pancreatitis to quantitate the binding of the chemotactic peptide N-formyl-methionyl-leucyl-phenylalanine-lysine (fMLFK) to leukocytes and its correlation to degree of organ inflammation. Uptake of the (99m)Tc-labeled nicotinyl hydrazine-derivatized chemotactic peptide analog fMLFK-HYNIC was measured in blood, pancreas, lung, and muscle specimens in rats with edematous or necrotizing pancreatitis and was compared with neutrophil sequestration assessed by myeloperoxidase activity and histology. Chemotactic peptide uptake in the pancreas was increased in mild and severe pancreatitis compared with controls, with higher levels in severe than in mild disease, and correlated with tissue myeloperoxidase activity (r = 0.7395, P < 0.001). Increased pulmonary uptake only in severe pancreatitis reflected pancreatitis-induced neutrophil sequestration in the lungs. Muscle uptake was unchanged compared with controls. Edema formation did not affect chemotactic peptide uptake. The data suggest that uptake of chemotactic peptides can contribute to quantitative assessment of neutrophils in localized inflammatory processes and is independent of associated edema formation or microcirculatory compromise.

Impact of laparoscopic staging in the treatment of pancreatic cancer.

HYPOTHESIS: Staging laparoscopy in patients with pancreatic cancer identifies unsuspected metastases, allows treatment selection, and helps predict survival. DESIGN: Inception cohort. SETTING: Tertiary referral center. PATIENTS: A total of 125 consecutive patients with radiographic stage II to III pancreatic ductal adenocarcinoma who underwent staging laparoscopy with peritoneal cytologic examination between July 1994 and November 1998. Seventy-eight proximal tumors and 47 distal tumors were localized. INTERVENTIONS: Based on the findings of spiral computed tomography (CT) and laparoscopy, patients were stratified into 3 groups. Group 1 patients had unsuspected metastases found at laparoscopy and were palliatedwithout further operation. Group 2 patients had no demonstrable metastases, but CT indicated unresectability due to vessel invasion. This group underwent external beam radiation with fluorouracil chemotherapy followed in selected cases by intraoperative radiation. Patients in group 3 had no metastases or definitive vessel invasion and were resection candidates. MAIN OUTCOME MEASURE: Survival. RESULTS: Staging laparoscopy revealed unsuspected metastases in 39 patients (31.2%), with 9 having positive cytologic test results as the only evidence of metastatic disease (group 1). Fifty-five patients (44.0%) had localized but unresectable carcinoma (group 2), of whom 2 (3.6%) did not tolerate treatment, 20 (36.4%) developed metastatic disease during treatment, and 21 (38.2%) received intraoperative radiation. Of 31 patients with potentially resectable tumors (group 3), resection for cure was performed in 23 (resectability rate, 74.2%). Median survival was 7.5 months for patients with metastatic disease, 10.5 months for those receiving chemoradiation, and 14.5 months for those who underwent tumor resection (P = .01 for group 2 vs. group 1; P<.001 for group 3 vs group 1). CONCLUSIONS: Staging laparoscopy, combined with spiral CT, allowed stratification of patients into 3 treatment groups that correlated with treatment opportunity and subsequent survival. Among the 125 patients, laparoscopy obviated 39 unnecessary operations and irradiation in patients with metastatic disease not detectable by CT. Laparoscopic staging can help focus aggressive treatment on patients with pancreatic cancer who might benefit.

Control of mammary tumor differentiation by SKI-606 (bosutinib).

C-Src is infrequently mutated in human cancers but it mediates oncogenic signals of many activated growth factor receptors and thus remains a key target for cancer therapy. However, the broad function of Src in many cell types and processes requires evaluation of Src-targeted therapeutics within a normal developmental and immune-competent environment. In an effort to understand the appropriate clinical use of Src inhibitors, we tested an Src inhibitor, SKI-606 (bosutinib), in the MMTV-PyVmT transgenic mouse model of breast cancer. Tumor formation in this model is dependent on the presence of Src, but the necessity of Src kinase activity for tumor formation has not been determined. Furthermore, Src inhibitors have not been examined in an autochthonous tumor model that permits assessment of effects on different stages of tumor progression. Here we show that oral administration of SKI-606 inhibited the phosphorylation of Src in mammary tumors and caused a rapid decrease in the Ezh2 Polycomb group histone H3K27 methyltransferase and an increase in epithelial organization. SKI-606 prevented the appearance of palpable tumors in over 50% of the animals and stopped tumor growth in older animals with pre-existing tumors. These antitumor effects were accompanied by decreased cellular proliferation, altered tumor blood vessel organization and dramatically increased differentiation to lactational and epidermal cell fates. SKI-606 controls the development of mammary tumors by inducing differentiation.

Reprogramming of the tumour microenvironment by stromal PTEN-regulated miR-320.

PTEN (Phosphatase and tensin homolog deleted on chromosome 10) expression in stromal fibroblasts suppresses epithelial mammary tumours, but the underlying molecular mechanisms remain unknown. Using proteomic and expression profiling, we show that Pten loss from mammary stromal fibroblasts activates an oncogenic secretome that orchestrates the transcriptional reprogramming of other cell types in the microenvironment. Downregulation of miR-320 and upregulation of one of its direct targets, ETS2 (v-ets erythroblastosis virus E26 oncogene homolog 2) are critical events in Pten-deleted stromal fibroblasts responsible for inducing this oncogenic secretome, which in turn promotes tumour angiogenesis and tumour-cell invasion. Expression of the Pten-miR-320-Ets2-regulated secretome distinguished human normal breast stroma from tumour stroma and robustly correlated with recurrence in breast cancer patients. This work reveals miR-320 as a critical component of the Pten tumour-suppressor axis that acts in stromal fibroblasts to reprogramme the tumour microenvironment and curtail tumour progression.

Role of brush border Na+/H+ exchange in canine ileal absorption.

Na+/H+ exchanger isoforms have been identified in mammalian intestinal enterocytes and cloned: NHE1 on the basolateral membrane regulating intracellular pH; and NHE2 and NHE3 on the brush border serving transcellular absorption in Na+. NHE1 and NHE2 are much more sensitive to inhibition by amiloride than NHE3, their in vitro IC50s for amiloride being 1 microM, 1 microM and 39 microM, respectively. This study tested the hypothesis that the brush border NHE3 isoform plays the predominant role in basal and meal-stimulated ileal absorption. Absorption studies (N = 72) were performed in dogs with 25-cm ileal Thiry-Vella fistulae. Six groups were studied over 4 hr. Perfusion with [14C]PEG and 140 mM Na+ was used to calculate absorption of water, ions, and glucose. Luminal amiloride was administered from the second to the fourth hours at doses of 20 microM in groups 3 and 4 to inhibit NHE1 and NHE2, and 1mM in groups 5 and 6 to also inhibit NHE3. A 480-kcal canine meal was ingested after the second hour in groups, 2, 4, and 6. Meal ingestion was followed by significant increases in water and electrolyte absorption. Amiloride (1 mM) caused significant reductions in basal and meal-stimulated ileal absorption, while the 20 microM dose had no effect on either. These data are consistent with the hypothesis that NHE3, but not NHE2, is involved in basal and meal-stimulated ileal water and Na+ absorption.

pT1 urothelial carcinoma of the bladder: criteria for diagnosis, pitfalls, and clinical implications.

One of the challenging areas in genitourinary pathology is the recognition of early invasion in urothelial neoplasia. Not uncommon, the patterns of invasion into lamina propria are subtle because a desmoplastic response is absent. Tangential sectioning due to inability to orient transurethral resection of bladder tumor specimens, crush and cautery artifacts further compound this problem. This review is presented to familiarize surgical pathologists with the criteria and different patterns of lamina propria invasion by urothelial carcinoma. Problems and pitfalls associated with the recognition of invasion and the clinicopathologic significance of lamina propria invasive urothelial cancer are also discussed.

Determination of Her-2/Neu status in breast carcinoma: comparative analysis of immunohistochemistry and fluorescent in situ hybridization.

Her-2/neu (H2N) status in breast carcinoma has been considered a prognostic factor that may have therapeutic implications; however, the correlation between H2N overexpression and gene amplification has not been completely defined. A consecutive series of ductal carcinomas (34 invasive and 7 in situ) were analyzed by fluorescent in situ hybridization for H2N gene and chromosome 17 copy number using touch preps of intact cells and by immunohistochemistry, using three different commercial antibodies to H2N protein (Zymed, clone 31G7; Ventana, clone CB11; and Dako, polyclonal) in corresponding formalin-fixed, paraffin-embedded tissue sections. Gene amplification was classified as unequivocal if more than five signals were present in more than 80% of the counted nuclei and absent if more than 80% of the nuclei counted contained two or fewer gene copies. Cases that did not fulfill the above criteria were considered equivocal for amplification. Immunostaining was classified as follows: 0 = no staining; 1+ = faint, incomplete membranous pattern; 2+ = moderate, complete membranous pattern; 3+ = strong membranous pattern. Of the 34 invasive tumors, 10 (29%) had unequivocal gene amplification. Furthermore, all had more than 10 copies of the gene in more than 60% of the counted nuclei. An additional nine cases (26%) had equivocal amplification, which was usually the result of chromosome 17 aneuploidy (seven of nine) or heterogeneity. With the Zymed and Dako antibodies, all tumors with 3+ staining had unequivocal gene amplification and all cases with 2+, 1+, or 0 staining were negative or equivocal for gene amplification. With the Ventana antibody, all cases with 3+ staining had unequivocal gene amplification, but two cases with unequivocal amplification by fluorescent in situ hybridization exhibited 1+ staining. Moderate (2+) H2N staining was observed in one case, three cases, and five cases with the Ventana, Dako, and Zymed reagents, respectively, and did not correlate with H2N gene copy number. Discordance between H2N and chromosome 17 copy number was not a useful means of defining amplification. Two cases of ductal carcinoma in situ with the Zymed antibody and two with the Dako antibody showed 3+ staining despite lack of unequivocal gene amplification. We conclude that (1) strong H2N immunostaining is highly associated with gene amplification, although there is minor variation in sensitivity between different antibodies; (2) a subset of breast carcinomas (3 to 15%) demonstrate moderate H2N staining without evidence of amplification, and it is unclear whether they represent highly sensitive staining or are a subset of cases that show overexpression without amplification; (3) gene amplification, as detected by fluorescent in situ hybridization, is associated with at least 10 gene copies per nucleus, and lower gene copy duplication (3 to 4/nucleus) is frequent, usually the result of chromosome 17 polysomy, and not associated with high-level overexpression; (5) overexpression of H2N without amplification may be more frequent in ductal carcinoma in situ, implying a different role in the biology of preinvasive versus invasive neoplasm.

Laparoscopy and peritoneal cytology in the staging of pancreatic cancer.

Staging laparoscopy in patients with pancreatic cancer allows identification of metastatic disease which is beyond the resolution of computed tomography. Laparoscopic ultrasound, dissection, and/or peritoneal cytology may be used to enhance the sensitivity of the staging procedure. Our experience at Massachusetts General Hospital with staging laparoscopy and peritoneal cytology over the past 8 years (N = 239) reveals that approximately 30% of patients without metastases by computed tomography harbor occult metastatic disease at laparoscopy. Additionally, published series demonstrate accurate determination of resectability in greater than 75% of patients after staging laparoscopy. Staging laparoscopy in patients with pancreatic cancer allows optimization of resources and avoidance of unnecessary surgery.

Glutathione S-transferase pi gene methylation: the search for a molecular marker of prostatic adenocarcinoma.

Glutathione S-transferase pi gene methylation has recently been described in prostatic adenocarcinoma. Aggregate data on 115 samples studied to date have found an 87% sensitivity and 92% specificity for prostate cancer diagnosis. The current literature about this new marker is herein summarized, and possible molecular mechanisms by which glutathione S-transferase pi may participate in prostatic carcinogenesis are reviewed. The possible clinical implications of this molecular alteration in the diagnosis of prostatic adenocarcinoma are also studied.