Fluid-yielding Ducts Research Articles

Ductal Architecture: Corrosion Casting of Canine Mammary Glands Using an Intraductal Approach.

Detailed knowledge related to the morphology, anatomy, physiology, and pathology of the canine mammary gland is scarce. Mammary tissue undergoes massive changes instructed by hormones multiple times within the lifespan of every bitch, affecting its appearance. To address the ductal system's appearance and to present how different our findings regarding the canine mammary gland are in comparison with the available literature, we obtained cadaveric specimens after euthanasia and mastectomy. All bitches were euthanised due to poor prognosis for their recovery from maladies unrelated to mammae. Using intraductal cannulation ex vivo, milk- or fluid-yielding ducts were perfused using VasQtec (polyurethane resin), which revealed casts, i.e., imprints of ducts and glandular structures in situ. We observed primary, vertically positioned ducts that ascended throughout the teat and continued branching to secondary, tertiary, etc., horizontally positioned ducts, which drained mammary gland lobes under the skin located close to the abdominal wall. The ascendant teat canal could be split into two and could be connected to gland alveoli or end blind. Alveolar formations were located along ducts and ductules in bigger and/or smaller clusters. This study is the first to generate a 3D image of canine ducts and glandular tissue using an intraductal approach.

An Improved Breast Epithelial Sampling Method for Molecular Profiling and Biomarker Analysis in Women at Risk for Breast Cancer.

BACKGROUNDThere is a strong need to define the molecular changes in normal at-risk breast epithelium to identify biomarkers and new targets for breast cancer prevention and to develop a molecular signature for risk assessment. Improved methods of breast epithelial sampling are needed to promote whole-genome molecular profiling, increase ductal epithelial cell yield, and reduce sample cell heterogeneity.METHODSWe developed an improved method of breast ductal sampling with ductal lavage through a 22-gauge catheter and collection of ductal samples with a microaspirator. Women at normal risk or increased risk for breast cancer were studied. Ductal epithelial samples were analyzed for cytopathologic changes, cellular yield, epithelial cell purity, quality and quantity of DNA and RNA, and use in multiple downstream molecular applications.RESULTSWe studied 50 subjects, including 40 subjects at normal risk for breast cancer and 37 subjects with non-nipple aspirate fluid-yielding ducts. This method provided multiple 1.0 mL samples of high ductal epithelial cell content (median ≥8 samples per subject of ≥5,000 cells per sample) with 80%–100% epithelial cell purity. Extraction of a single intact ductal sample (fluid and cells) or the separate frozen cellular component provided DNA and RNA for multiple downstream studies, including quantitative reverse transcription- polymerase chain reaction (PCR) for microRNA, quantitative PCR for the human telomerase reverse transcriptase gene, whole-genome DNA amplification, and array comparative genomic hybridization analysis.CONCLUSIONAn improved breast epithelial sampling method has been developed, which should significantly expand the acquisition and biomarker analysis of breast ductal epithelium in women at risk for breast cancer.

Does ductal lavage assert its role as a noninvasive diagnostic modality to identify women at low risk of breast cancer development?

ObjectiveDuctal lavage (DL) involves evaluation of the ductal system of the breast for detection of intra-ductal carcinomas and precursor lesions by collecting breast epithelial cells using a small-gauge catheter inserted into a ductal orifice on the nipple. The aim of this survey was to analyze cytologic features of samples obtained from low-risk women with DL and to elucidate the efficacy of this diagnostic modality in evaluating fluid production, cannulating and determining atypical breast epithelial cells.MethodsInto this prospective study were consecutively registered 80 women between ages 28 to 67. Nipple aspiration was performed to identify all fluid-yielding ducts. According to the grading of specific features the interpretation of the sample included: normal/benign (category, 0), mild atypical (category, I), markedly atypical (category, II) or malignant (category, III) disorders.ResultsNinety five percent (316/334) of the nipple aspirate fluid samples were classified as category 0, 4.8% (16/334) as category I and 0.2% (2/334) as category II changes. Category III disorders were not detected. Therefore, in 80% of the women examined results were within normal limits while 17.5% of the participants presented mild atypical and 2.5% markedly atypical rates.ConclusionDL collection procedure proved to be rapid as well as acceptable by the women studied. It retains the advantage over other methods of nipple aspirate fluid in that it is easy to perform, thereby removing most clinician variability. It also helped low risk women to discriminate those with breast disorders that require additional investigation, further follow-up or administration of preventive medication.

The anatomy of fluid-yielding ducts in breast cancer

The concept of an intraductal approach to evaluate the breast microenvironment assumes direct access to the cancer-containing duct. Central duct access to the cancer-affected lobe is essential if cytology or cell markers are to be useful indicators of pre-malignant change. Access to the cancer-bearing lobe would be less important if field change effects of malignant change were predominantly supra-lobar. The aim of this study was to determine how often duct lavage fluid drains the breast cancer-affected segment. 58 patients undergoing mastectomy for breast cancer were recruited among which 47 had at least one fluid-yielding duct. Following duct lavage, fluid-yielding ducts were perfused ex vivo with Polyurethane Elastomer (PU4ii) resin. Specimens were sliced sagittally, and the extent of resin perfusion and anatomical relationship to the cancer-affected segment was recorded. Computed tomography (CT) scanning was performed on selected mastectomies before cut-up for a feasibility study of 3D duct reconstruction. The median number of fluid-yielding ducts cannulated per cancer-affected breast was 2 (range 1-4). 35/47 (74%) mastectomy specimens were successfully cannulated for resin perfusion. 29/35 (83%) showed tracing of the cancer-affected duct system, 6/35 resin perfusions traced duct systems unaffected by cancer and 12/35 perfusions extravasated. The proportion of sagittal breast slices perfused by resin was 13-68% (median 43%). Volume rendering CT showed it is feasible to produce a simulated image of the perfused ducts. Duct access to the cancer-containing segment is feasible in the majority of patients. Fluid-yielding ducts proportionately drain a significant volume of the breast. Large symptomatic cancers may cause obstruction with distal collapse. Further quantitative study of breast perfusion CT scans may be helpful for estimating the volume fraction of breast tissue perfused by fluid-yielding ducts. The intraductal approach is a valid concept for biomarker assessment of cancer-containing breast segments.

Anatomical association of fluid yielding ducts with location of the breast cancer affected segment in screen detected and symptomatic breast cancer

The concept of the intraductal approach to evaluating the breast microenvironment assumes direct access to the breast cancer containing duct. Previous studies on duct anatomy infusing a dye or contrast found fluid-yielding ducts to be associated with the cancer containing duct in approximately 50% of fluid-yielding breasts with cancer. A concordant anatomical relationship between accessible ductal sytems and the cancer-affected lobe is essential if cytology or other cell markers are to be successfully identified as indicators of cancer or of early cancerous change. The concordant relationship is less important if field change effects are considered to predictors of malignant change. The aim of this study was to determine how often duct lavage effluent drains the breast cancer affected segment.

Patient variables that predict atypical cytology by nipple duct lavage

Nipple duct lavage (NDL) may be useful for breast cancer risk stratification. Published guidelines recommend that women with a 5-year Gail risk >/=1.7% should consider undergoing lavage for any fluid-yielding ducts, but it is not known whether increased breast cancer risk or nipple fluid production predict lavage atypia. One hundred fifty women unselected for breast cancer risk underwent NDL with cannulation of all nipple aspirate fluid (NAF)-producing ducts and at least 1 dry duct. This resulted in 516 lavage samples. The rate of cytologic atypia was compared for NAF-positive ducts and NAF-negative ducts and for women with 5-year Gail risks >/=1.7% and <1.7%. Ducts from breasts with cancer (N = 113) were excluded from the analysis. The cytologic atypia rate was similar for the 240 NAF-producing ducts (19%) and the 163 dry ducts (15%; P = .36). No significant differences were observed when atypia was categorized as mild (13% vs 10%; P = .63) or marked (6% vs 4%; P = .53). Among the 83 patients who were unaffected by breast cancer, atypia was diagnosed in 15 of 44 patients (34%) with a 5-year Gail risk <1.7% and in 11 of 39 patients (28%) with a 5-year Gail risk >/=1.7% (P = .74). Neither NAF production nor 5-year Gail risk predicted lavage atypia. Limiting NDL to fluid-producing ducts in women with a 5-year Gail risk >/=1.7% significantly reduced the sensitivity of the test for population screening.

4–3 Ductal Lavage of Fluid-Yielding and Non–Fluid-Yielding Ducts in BRCA1 and BRCA2 Mutation Carriers and Other Women at High Inherited Breast Cancer Risk

4–3 Ductal Lavage of Fluid-Yielding and Non–Fluid-Yielding Ducts in BRCA1 and BRCA2 Mutation Carriers and Other Women at High Inherited Breast Cancer Risk

Nipple aspiration and ductal lavage in women with a germline BRCA1 or BRCA2 mutation

IntroductionThe aim of this study was to collect serial samples of nipple aspirate (NA) and ductal lavage (DL) fluid from women with germline BRCA1/2 mutations in order to create a biorepository for use in identifying biomarkers of breast cancer risk.MethodsBetween March 2003 and February 2005, 52 women with germline BRCA1 or BRCA2 mutations (median age 43 years, range 27 to 65 years) were scheduled for six-monthly NA, DL and venesection. DL was attempted for all NA fluid-yielding (FY) and any non-FY ducts that could be located at each visit.ResultsTwenty-seven (52%) women were postmenopausal, predominantly (19/27) from risk reducing bilateral salpingo-oophorectomy (BSO). FY ducts were identified in 60% of all women, 76% of premenopausal women versus 44% of postmenopausal (P = 0.026). Eighty-five percent of women had successful DL. Success was most likely in women with FY ducts (FY 94% versus non-FY 71% (P = 0.049). DL samples were more likely to be cellular if collected from FY ducts (FY 68% versus non-FY 43%; P = 0.037). Total cell counts were associated with FY status (FY median cell count 30,996, range 0 to >1,000,000 versus non-FY median cell count 0, range 0 to 173,577; P = 0.002). Four women (8%) had ducts with severe atypia with or without additional ducts with mild epithelial atypia; seven others had ducts with mild atypia alone (11/52 (21%) in total). Median total cell count was greater from ducts with atypia (105,870, range 1920 to >1,000,000) than those with no atypia (174, 0 to >1,000,000; P ≤ 0.001).ConclusionIt is feasible to collect serial NA and DL samples from women at high genetic risk of breast cancer, and we are creating a unique, prospective collection of ductal samples that have the potential to be used for discovery of biomarkers of breast cancer risk and evaluate the ongoing effects of risk reducing BSO. DL cellular atypia was not predictive of a current breast cancer and longer follow up is needed to determine whether atypia is an additional marker of future breast cancer risk in this population already at high genetic risk of breast cancer.

Patterns of reduced nipple aspirate fluid production and ductal lavage cellularity in women at high risk for breast cancer

IntroductionNipple aspiration is a noninvasive technique for obtaining breast fluids from the duct openings of the nipple for the evaluation of abnormalities associated with breast cancer. Nipple aspirate fluid (NAF) can be elicited from 48 to 94% of healthy women, and its production has been linked to an increased relative risk for breast cancer development. NAF production has been used in studies to guide the selection of ducts for ductal lavage, a procedure in which ducts are cannulated and flushed with saline to collect cells. In a previous multicenter trial to evaluate intraductal approaches in women at high-risk for breast cancer, NAF production was observed in 84% of the subjects. However, we observed a significantly lower proportion of fluid-yielding subjects in a similar series of high-risk women. The purpose of the present study was to identify variables associated with this reduction.MethodNipple aspiration was performed on 33 high-risk women (defined as having a 5-year Gail model index of more than 1.7, a personal or family history of breast cancer, and/or a BRCA1 or BRCA2 germline mutation) to identify ductal orifices for lavage procedures. Lavage was performed on all fluid-yielding ducts and on nine non-fluid-yielding ducts.ResultsFluid-yielding ducts were identified in 12 of 33 (36%) of the subjects in the present series, compared with 16 of 19 (84%) of the subjects undergoing identical procedures at our facility during a multicenter trial (P = 0.001). Reduced NAF yields were associated with postmenopausal status (P = 0.02), BRCA germline mutations (P = 0.004), and risk reduction therapies, including bilateral salpingo-oophorectomy (BSO) and/or selective estrogen receptor modulators (SERMs; P = 0.009). All nine (100%) of the ductal lavage specimens collected from non-fluidyielding ducts were acellular, in comparison with 3 of 13 specimens from fluid-yielding ducts (P < .001).ConclusionAnalysis of high-risk women in the present series revealed patterns of reduced NAF production and ductal lavage cellularity compared with a previous multicenter trial. The present series included more BRCA-positive women, many of whom had undergone BSO and/or were using SERMs. Our data suggest that endocrine mechanisms associated with these risk-reducing therapies may be related to patterns of diminished breast fluid production.

Ductal Lavage Findings in Women With Mammographic Microcalcifications Undergoing Biopsy

We designed a prospective study to assess the likelihood that early lesions presenting as mammographic calcifications could be accessed for cytological diagnosis by ductal lavage (DL). Consenting women with calcifications (Breast Imaging Reporting and Data System 4 or 5) underwent DL of fluid-yielding ducts (FYDs) before stereotactic core or excisional biopsy. The DL catheter was used to inject .2 to 1 mL of Isovue 300 into the duct to determine whether the FYD corresponded to the duct containing calcifications (designated overlap). Additional FYDs were injected, if possible, until overlap was identified. DL cytology was compared with histology. Twenty women were enrolled (mean age, 54.2 years); the mean size of the calcification-bearing area was 190 mm(2). The histological findings were as follows: 1 invasive cancer, 9 ductal carcinomas-in-situ (DCIS), 5 atypical hyperplasias, and 5 usual hyperplasias or fibrocystic changes. Four women had no FYD. In 15 women who underwent DL and ductography, overlap of dye and calcifications was seen in 4 (27%): 1 fibrocystic change, 1 hyperplasia, 1 atypical hyperplasia (cytological diagnosis mildly atypical), and 1 DCIS (cytological diagnosis benign). Of the remaining 8 DCIS lesions, 4 had no nipple aspiration fluid, 1 showed extravasation, and 3 were lavaged but the duct did not overlap. These results are consistent with data from women undergoing mastectomy for larger invasive cancer and DCIS and show that cancer-containing ducts do not yield nipple fluid in most cases.

Ductal Lavage of Fluid-Yielding and Non–Fluid-Yielding Ducts in BRCA1 and BRCA2 Mutation Carriers and Other Women at High Inherited Breast Cancer Risk

Nipple fluid production and atypical breast duct cells in women at high risk of breast cancer have been associated with further increased risk. Most publications on ductal lavage for cell collection report cannulating fluid-yielding ducts only. We report lavage of fluid-yielding and non-fluid-yielding ducts in women at high inherited breast cancer risk. A pilot breast cancer screening study including ductal lavage was conducted in 75 women at high inherited risk, 56 (74.7%) of whom had BRCA1/2 mutations. Ductal lavage was attempted in any duct identifiable with a catheter. Ducts were successfully catheterized in 60 of 75 patients (80%). Successfully catheterized patients were younger (median age 41 versus 53 years, P = 0.0003) and more often premenopausal (51.7% versus 20%, P = 0.041). Thirty-one successfully catheterized patients [51.6%, 95% confidence interval (39.4-63.9%)] had non-fluid-yielding ducts only. Seventeen patients [28.3% (18.5-40.9%)] had atypical cells. Twelve of seventeen [70.6% (46.8-87.2%)] samples with atypia were from non-fluid-yielding ducts. Patients with non-fluid-yielding ducts (versus fluid-yielding ducts) were more likely to have had prior cancer (48.4% versus 17.2%, P = 0.014) or chemotherapy (45.2% versus 17.2%, P = 0.027); this was also true in patients with atypia from non-fluid-yielding ducts. Successfully lavaged women were younger and more often premenopausal. Atypical cells can be found in non-fluid-yielding ducts in patients at high inherited breast cancer risk. Non-fluid-yielding ducts, and atypia from non-fluid-yielding ducts, are more common in patients with prior cancer and chemotherapy. Larger studies are needed to identify risk factors and prognostic significance associated with atypia and non-fluid-yielding ducts in high-risk populations, and define their role as biomarkers.

Ductal lavage of non-fluid yielding ducts in BRCA1 and BRCA2 mutation carriers and other women at high genetic risk for breast cancer

9535 Background: Atypical breast duct cells in patients at increased risk of breast cancer have been predictive of subsequent breast cancer. It has been reported that women whose breasts yield no fluid on nipple suction aspiration have lower risk for breast cancer than women whose breasts yield fluid; publications on ductal lavage (DL) have reported attempt of DL only on patients with fluid-yielding ducts (FY) on initial suction aspiration. We report preliminary results of a study of DL including evaluation of non-fluid yielding ducts (NFY) in women at increased genetic risk of breast cancer. Methods: A pilot study of screening with breast MRI, mammogram, and ductal lavage was conducted in 41 women at increased genetic risk of breast cancer; 26 (63%) had BRCA mutations, and 15 (37%) were included because family history conveyed greater than 10% breast cancer risk at 10 years via the Claus model. Whether or not fluid was obtained on suction aspiration, ducts which could be identified with a dilator underwent attempted lavage. Cytologic diagnostic categories were benign, atypical, or malignant. Fisher's exact test, 2 tailed, was used for statistical analysis. Results: DL was successful in 34 of 41 patients (82.9%); 17 (50%) had NFY. Ten patients (29.4%) had atypical cells; none had malignant cells. Seven of 17 patients (41.2%, 95% confidence interval 18.4–67.1%) with NFY had atypical cells. Patients with atypical cells from NFY were more likely than those with atypical cells from FY to have a BRCA mutation (57.1 versus 0%, p 0.20). Patients with NFY were more likely than those with FY to have had breast or ovarian cancer (58.8% versus 17.6%, p 0.03), and to have received chemotherapy (47.1% versus 17.6%, p 0.14). Conclusions: Atypical cells can be found in NFY patients at high risk for breast cancer, with trends toward higher incidence of atypical cells from NFY in BRCA mutation carriers, and of NFY in cancer survivors. Larger studies are needed to confirm this finding, and to identify risk factors and outcomes associated with atypical cells and NFY. No significant financial relationships to disclose.

Evaluation and management of the woman with an abnormal ductal lavage.

RATIONALE FOR DUCTAL LAVAGE The majority of breast cancers originate in the epithelial lining of the ductal system of the breast. Studies of ductal carcinoma in situ (DCIS), a lesion accepted as the precursor of invasive carcinoma, strongly suggest that the early changes of malignancy occur in a segmental fashion within a ductal system rather than diffusely throughout the breast. Multiple peripheral papillomas, a lesion associated with an increased risk of breast cancer development, also occur in a segmental distribution. This anatomic distribution, coupled with the prolonged time period from detection of low-grade DCIS to development of invasive carcinoma, suggests that examination of the ductal cells might be an effective method for identifying women at increased risk for developing breast carcinoma. Ductal lavage is a minimally invasive diagnostic tool that has been developed to identify evidence of cellular atypia in breast ducts. In the initial multicenter trial, 507 women at increased risk for breast cancer development on the basis of a personal history of breast cancer, or a 5-year Gail model risk of breast cancer development of 1.7% or more were studied. Fluid-yielding ducts were identified by nipple aspiration, and a microcatheter was inserted into each fluid-yielding duct and infused with 2 to 6 mL of normal saline. The effluent was collected, and the lavage procedure repeated until approximately 10 mL of saline were instilled into each duct. Cytologic diagnoses from nipple aspiration and ductal lavage were compared as the primary endpoint of the study. The procedures were well tolerated, and there were no serious adverse events. Fluid-yielding ducts were identified in 85% of subjects. Only 27% of specimens obtained by nipple aspiration contained an adequate number of epithelial cells for diagnosis, with a median of 120 epithelial cells per breast. In contrast, 78% of specimens from ductal lavage had adequate cells for diagnosis, with a median of 4,000 or 13,500 epithelial cells per duct, depending on the type of catheter used for lavage. Both these differences were highly statistically significant (p 0.001). A total of 24% of subjects had abnormal lavages, with 17% showing mild atypia, 6% moderate atypia, and in two cases ( 1%), frankly malignant cells. This study clearly demonstrates that ductal lavage is a safe and effective technique for collecting epithelial cells from the breast ductal system. This is an important finding for translational researchers because it provides an opportunity to test these cells for molecular markers that can predict development of the fully malignant phenotype, and to sample the same duct repeatedly over time. But this study has also raised many questions for clinicians and women at risk for developing breast cancer. In particular, these include the reproducibility of cytologic interpretations of lavage fluid, how to integrate the findings of ductal lavage into current clinical methods of risk assessment, the role of ductal lavage as a diagnostic tool for breast cancer, and the workup needed for the finding of atypia on ductal lavage. This article will review these subjects.

Detection of premalignant and malignant breast cells by ductal lavage

All ductal and lobular breast cancers originate in the single layer of epithelial cells that line the ductal/lobular systems of all milk ducts. Ductal lavage (DL) enables the retrieval of these cells using a microcatheter. The microcatheter is inserted into the milk ducts through the nipple surface orifices, and saline is flushed through the ducts to wash out cells for cytological evaluation. Methods: High-risk women (Gail index > 1.7, previous breast cancer or BRCA1/BRCA2–positive) with nonsuspicious mammograms and physical exams were aspirated to identify fluid-yielding ducts for subsequent lavage. Cytological diagnoses were rendered as either insufficient cellular material for diagnosis (ICMD), benign (B), mild atypia/premalignant cells (A/P), or suspicious for malignancy/unequivocally malignant cells (ASM/M). Results: DL was performed on 426 high-risk women (551 breasts). Cytological analysis of 470 eligible breasts has been completed. Of lavaged breasts, 15.1% and 5.1% yielded a cytological diagnosis of A/P and ASM/M, respectively, for a combined atypia prevalence of 20.2%. In two cases of ASM/M cytology, a 1.8-cm high-grade ductal carcinoma in situ (DCIS) and a 6-cm intermediate-grade DCIS were confirmed and resected with clear margins following exploration and partial resection of the afflicted ducts. In a third case, extensive papillomas with a small focus of cells resembling DCIS were confirmed following prophylactic mastectomy. Workup of two more cases with repeat mammograms, MRI, ultrasonography, and ductoscopy revealed papillomas but no high-grade intraductal lesions correlating with the lavage cytology, although ductoscopy is obviously limited in its ability to access and sample all ductal branches. The other ASM/M cases are still undergoing workup. Conclusions: Ductal lavage is a safe, well-tolerated, and relatively noninvasive procedure for collecting breast ductal epithelial cells to allow for the determination and differentiation of normal, premalignant, and malignant cytology. Ductal lavage may be a useful adjunct to mammography and other currently available imaging modalities for the early detection of intraductal breast pathology.