We present the first prospective test of Raman spectroscopy in diagnosing normal, benign, and malignant human breast tissues. Prospective testing of spectral diagnostic algorithms allows clinicians to accurately assess the diagnostic information contained in, and any bias of, the spectroscopic measurement. In previous work, we developed an accurate, internally validated algorithm for breast cancer diagnosis based on analysis of Raman spectra acquired from fresh-frozen in vitro tissue samples. We currently evaluate the performance of this algorithm prospectively on a large ex vivo clinical data set that closely mimics the in vivo environment. Spectroscopic data were collected from freshly excised surgical specimens, and 129 tissue sites from 21 patients were examined. Prospective application of the algorithm to the clinical data set resulted in a sensitivity of 83%, a specificity of 93%, a positive predictive value of 36%, and a negative predictive value of 99% for distinguishing cancerous from normal and benign tissues. The performance of the algorithm in different patient populations is discussed. Sources of bias in the in vitro calibration and ex vivo prospective data sets, including disease prevalence and disease spectrum, are examined and analytical methods for comparison provided.

Fibroadenoma (FA) and phyllodes tumor (PT) of the breast are biphasic tumors composed of variable proportions of epithelial and stromal cells. We identified a case of synchronous FA and PT in the same breast mass. Using laser capture microdissection, loss of heterozygosity analysis was performed on these components to gain potential insight into the histogenetic relationship between FA and PT. Genomic DNA was analyzed at 10 microsatellite loci by polymerase chain reaction. Both tumors showed allelic loss at D7S522. In addition, phylloid tumor showed allelic loss at TP53 and D22S264, not observed in FA. Our data suggest that FA and PT are clonally related and allelic loss at TP53 and D22S264 may be implicated in the progression of FA to phyllode tumor.

Using diffuse reflectance spectroscopy and intrinsic fluorescence spectroscopy, we have developed an algorithm that successfully classifies normal breast tissue, fibrocystic change, fibroadenoma, and infiltrating ductal carcinoma in terms of physically meaningful parameters. We acquire 202 spectra from 104 sites in freshly excised breast biopsies from 17 patients within 30 min of surgical excision. The broadband diffuse reflectance and fluorescence spectra are collected via a portable clinical spectrometer and specially designed optical fiber probe. The diffuse reflectance spectra are fit using modified diffusion theory to extract absorption and scattering tissue parameters. Intrinsic fluorescence spectra are extracted from the combined fluorescence and diffuse reflectance spectra and analyzed using multivariate curve resolution. Spectroscopy results are compared to pathology diagnoses, and diagnostic algorithms are developed based on parameters obtained via logistic regression with cross-validation. The sensitivity, specificity, positive predictive value, negative predictive value, and overall diagnostic accuracy (total efficiency) of the algorithm are 100, 96, 69, 100, and 91%, respectively. All invasive breast cancer specimens are correctly diagnosed. The combination of diffuse reflectance spectroscopy and intrinsic fluorescence spectroscopy yields promising results for discrimination of breast cancer from benign breast lesions and warrants a prospective clinical study.

The purpose of this study was to compare four cell block (CB) methods in the setting of nongynecologic ThinPrep (TP) specimens. 48 CBs were prepared from 12 nongynecologic TP specimens using the following CB methods: (1) Inverted filter sedimentation (IFS); (2) Thrombin method; (3) Albumin method; (4) Simple sedimentation. Each CB was assigned a cellularity score: 0 no cells, 1+ hypocellular, 2+ hypocellular with tissue fragments, 3+ cellular. A score of 2+ or 3+ was given for 11/12 of thrombin, 7/12 IFS, 5/12 albumin, and 2/12 simple sedimentation CBs. Thrombin CBs demonstrated a pale background clot with evenly distributed cells. Albumin CBs had a cracked uneven background. IFS CBs had a clear background, but were technically difficult and cells appeared artifactually crowded. In the setting of nongynecologic TP specimens, the thrombin CB was easily prepared and produced the best CB in regards to cellularity, cell distribution, and background quality. Diagn. Cytopathol. 2007;35:640–643. © 2007 Wiley‐Liss, Inc.

Currently breast cancer diagnosis is made clinically through triple assessment: annual clinical breast examination, x-ray mammography or breast ultrasound imaging, and biopsy. The majority of women with suspicious breast lesions undergo either stereotactic (needle) or surgical (excisional) biopsy. Due to a high incidence of "false positives" at clinical breast diagnosis and "false negatives" at surgery, a large number of women undergo unnecessary and costly breast surgery. We describe our program of development of techniques and instrumentation for clinical application of NIR Raman spectroscopy for improved breast cancer diagnosis.

BACKGROUND:That sweat gland type tumors occur occasionally in the breast is not surprising, as the breast and cutaneous sweat glands are embryologically related. Cylindromas present most commonly as solitary and sporadic dermal nodules on the face and scalp. Cases of multiple cylindromas are dominantly inherited, and the neoplasms are referred to as "turban tumors" when multiple lesions cover the scalp. Primary cylindroma of the breast has been reported once in the past. To the best of our knowledge, the fine needle aspiration cytology of primary breast cylindroma and its occurrence in the setting of familial cylindromatosis have not previously been reported.CASE:A 59-year-old woman presented with an ill-defined left breast mass. She had a personal and family history of dermal cylindromas on the head and face. Fine needle aspiration cytology demonstrated small, uniform cells with oval nuclei and finely granular cytoplasm, with some cells arranged around conspicuous cylinders of dense, acellular material. Excisional biopsy was recommended to exclude adenoid cystic carcinoma. Tissue biopsy revealed a benign cylindroma arising in breast parenchyma.CONCLUSION:Fine needle aspiration cytology of cylindroma very closely mimics that of adenoid cystic carcinoma. Although extremely rare, primary breast cylindroma is another entity to be included in the cytologic differential diagnosis of bland, basaloid cells associated with globular, extracellular material, a finding most commonly associated with adenoid cystic carcinoma.

Bcl-2 family members have been shown to be key mediators of apoptosis as either pro- or anti-apoptotic factors. It is thought that both classes of Bcl-2 family members act at the level of the mitochondria to regulate apoptosis, although the founding anti-apoptotic family member, Bcl-2 is localized to the endoplasmic reticulum (ER), mitochondrial, and nuclear membranes. In order to better understand the effect of Bcl-2 localization on its activity, we have utilized a Bcl-2 mutant that localizes only to the ER membrane, designated Bcl-2Cb5. Bcl-2Cb5 was expressed in MDA-MB-468 cells, which protected against apoptosis induced by the kinase inhibitor, staurosporine. Data presented here show that Bcl-2Cb5 inhibits this process by blocking Bax activation and cytochromec release. Furthermore, we show that Bcl-2Cb5 can inhibit the activation of a constitutively mitochondrial mutant of Bax, indicating that an intermediate between Bcl-2 on the ER and Bax on the mitochondria must exist. We demonstrate that this intermediate is likely a BH3-only subfamily member. Data presented here show that Bcl-2Cb5 can sequester a constitutively active form of Bad (Bad3A) from the mitochondria and prevent it from activating Bax. These data suggest that Bcl-2 indirectly protects mitochondrial membranes from Bax, via BH3-only proteins.

OBJECTIVE:To review the histologic findings in patients diagnosed with "atypical glandular cells of uncertain significance, endocervical cell type" (AGUS-EC) by ThinPrep Pap Test (TPPT) or conventional Pap smear (CPS) and to evaluate the clinical value of subclassifying AGUS-EC as "favor reactive" or "favor neoplastic."STUDY DESIGN:All TPPT and CPS diagnosed as AGUS-EC (favor reactive, unspecified and favor neoplastic) from January 1998 through December 1999 and all available histologic follow-up (defined as endocervical curettage, cervical biopsy, cervical conization or hysterectomy obtained within six months of the time of an AGUS-EC diagnosis) were obtained from a computerized database.RESULTS:AGUS-EC was diagnosed in 0.77% of CPS (683 of 88,825) and 0.59% of TPPT (183 of 30,968) (P = NS). There was no statistically significant difference in any of the follow-up histologic diagnoses between the CPS and TPPT groups. The majority of the follow-up biopsies demonstrated benign processes in both groups. Patients with a diagnosis of AGUS-EC "favor neoplastic" had a greater proportion of true glandular pathology as compared with AGUS-EC "unspecified" or "favor reactive" (P < .001). None of the patients with a diagnosis of AGUS-EC "favor reactive" were found to have true glandular pathology; however, a minority of them proved to have squamous pathology.CONCLUSION:In this study there was no difference in CPS and TPPT in regard to the specificity of a diagnosis of AGUS-EC for true glandular pathology. Subclassifying AGUS-EC as "favor reactive" or "favor neoplastic" may provide valuable information for directing patient follow-up.

Bcl-2 protein family members function either to promote or inhibit programmed cell death. Bcl-2, typically an inhibitor of apoptosis, has also been demonstrated to have pro-apoptotic activity (Cheng, E. H., Kirsch, D. G., Clem, R. J., et al. (1997) Science 278, 1966-1968). The pro-apoptotic activity has been attributed to the cleavage of Bcl-2 by caspase-3, which converts Bcl-2 to a proapoptotic molecule. Bcl-2 is a membrane protein that is localized in the endoplasmic reticulum (ER) membrane, the outer mitochondrial membrane, and the nuclear envelope. Here, we demonstrate that transient expression of Bcl-2 at levels comparable to those found in stably transfected cells induces apoptosis in human embryonic kidney 293 cells and in the human breast cell line MDA-MB-468 cells. Furthermore, we have targeted Bcl-2 specifically to either the ER or the outer mitochondrial membrane to test whether induction of apoptosis by Bcl-2 is dependent upon its localization within either of these membranes. Our findings indicate that Bcl-2 specifically targeted to the mitochondria induces cell death, whereas Bcl-2 that is targeted to the ER does not. The expression of Bcl-2 does result in its cleavage to a 20-kDa protein; however, mutation of the caspase-3 cleavage site (D34A) does not inhibit its ability to induce cell death. Additionally, we find that transiently expressed ER-targeted Bcl-2 inhibits cell death induced by Bax overexpression. In conclusion, the ability of Bcl-2 to promote apoptosis is associated with its localization at the mitochondria. Furthermore, the ability of ER-targeted Bcl-2 to protect against Bax-induced apoptosis suggests that the ER localization of Bcl-2 may play an important role in its protective function.

The classical (cWhipple) and the pylorus preserving Whippl e (ppWhipple) resection are both performed for the treatment of cancers of the pancreatic head or ampullary region.Since ppWh ipple resection is envisioned with improved postoperative weight gain and superior quality of life.we implemented preservation of the pylorus also in total pancreatectomy.Material and Methods: We analyzed early and late outcome after pylorus preserving total pancreatectomy (pptota l)in a non-selected, consecutive material.Patients demographics, intraopera tive and postoperative findings were recorded prospectively in a statistical database.After discharge.patients were seen at 6-month interva ls and quality of life, weight course, ability to work and survival were analyzed.For statistical evaluation Cruskal-Wallis and chi-square tests were used where appropri ate.For analysis of survival the Kaplan-Meyer Test was applied and differences were examine d with the log-rank test.Results: From 11/93 to 5/99.436