The Diagnostic Value of microRNA Expression Analysis in Detecting Intraductal Papillomas in Patients with Pathological Nipple Discharge.

Patients with pathological nipple discharge (PND) often undergo local surgical procedures because standard radiologic imaging fails to identify the underlying cause. MicroRNA (MiRNA) expression analysis of nipple fluid holds potential for distinguishing between breast diseases. This study aimed to compare miRNA expression levels between nipple fluids from patients with PND to identify possible relevant miRNAs that could differentiate between intraductal papillomas and no abnormalities in the breast tissue. Nipple fluid samples from patients with PND without radiological and pathological suspicion for malignancy who underwent a ductoscopy procedure were analyzed. We used univariate and multivariate regression analyses to identify nipple fluid miRNAs differing between pathologically confirmed papillomas and breast tissue without abnormalities. A total of 27 nipple fluid samples from patients with PND were included for miRNA expression analysis. Out of the 22 miRNAs examined, only miR-145-5p was significantly differentially expressed (upregulated) in nipple fluid from patients with an intraductal papilloma compared to patients showing no breast abnormalities (OR 4.76, p = 0.046), with a diagnostic accuracy of 92%. miR-145-5p expression in nipple fluid differs for intraductal papillomas and breast tissue without abnormalities and, therefore, has potential as a diagnostic marker to signal presence of papillomas in PND patients. However, further refinement and validation in clinical trials are necessary to establish its clinical applicability.

Clinical Significance of Contrast-Enhanced Ultrasound Galactography in Pre-operative Diagnosis of Patients With Pathologic Nipple Discharge.

OBJECTIVE: The aim of the work described here was to investigate the feasibility and diagnostic value of using contrast-enhanced ultrasound (CEUS) galactography with SonoVue in patients with pathologic nipple discharge (PND). METHODS: Twenty-eight patients who underwent breast surgery for PND from May 2019 to August 2021 were included. Routine ultrasound, ductoscopy and CEUS galactography were performed successively. Lesions were diagnosed and localized. The sensitivity, specificity and pre-operative localization value of each examination method were evaluated on post-operative pathology. RESULTS: CEUS galactography was successfully conducted in all 28 patients and revealed negative ductal ectasia, filling stop and filling defect. Ductoscopy revealed positive nodules in 21 cases and negative nodules in 7 cases. A total of 18 nodules were found by routine ultrasound, and the relationship between all nodules and the discharge duct was confirmed after CEUS galactography. Compared with the other two methods, CEUS galactography had higher sensitivity, positive predictive value and negative predictive value (100%, 81.82% and 100%, respectively), while it has the same specificity as routine ultrasound (both 60%). The pre-operative location of the nipple duct was consistent with the intra-operative findings in 28 patients after CEUS galactography. CONCLUSION: The ultrasound contrast agent SonoVue can be used for CEUS galactography in patients with PND. CEUS galactography can improve the detection of ductal nodules and locate the nipple discharge duct pre-operatively. As the technique does not emit radiation and SonoVue is easily metabolized and safe, CEUS galactography is better than conventional imaging for PND patients.

KCTD1 and Scalp-Ear-Nipple ('Finlay-Marks') syndrome may be associated with myopia and Thin basement membrane nephropathy through an effect on the collagen IV α3 and α4 chains.

INTRODUCTION: Scalp-Ear-Nipple syndrome is caused by pathogenic KCTD1 variants and characterised by a scalp defect, prominent ears, and rudimentary breasts. We describe here further clinical associations in the eye and kidney. METHODS: Fifteen affected members from two unrelated families with p.(Ala30Glu) or p.(Pro31Leu) in KCTD1 were examined for ocular and renal abnormalities. The relevant proteins were studied in the eye and kidney, and the mutation consequences determined from mouse knockout models. RESULTS: Five males and 10 females with a median age of 40 years (range 1-70) with pathogenic variants p.(Ala30Glu) (n = 12) or p.(Pro31Leu) (n = 3) in KCTD1 were studied. Of the 6 who underwent detailed ophthalmic examination, 5 (83%) had low myopic astigmatism, the mean spherical equivalent of 10 eyes was 2.38D, and one (17%) had hypermetropic astigmatism. One female had a divergent strabismus.Five individuals had renal cysts (5/15, 33%), with renal biopsy in one demonstrating a thinned glomerular basement membrane identical to that seen in Thin basement membrane nephropathy (AD Alport syndrome).In the eye, KCTD1 and its downstream targets, TFAP2, and the collagen IV α3 and α4 chains localised to the cornea and near the retinal amacrine cells. In the kidney, all these proteins except TFAP2 were expressed in the podocytes and distal tubules. TFAP2B and COL4A4 knockout mice also had kidney cysts, and COL4A3 and COL4A4 knockout mice had myopia. CONCLUSION: Individuals with a pathogenic KCTD1 variant may have low myopic astigmatism and represent a further rare genetic cause for a thinned glomerular basement membrane.

Nipple fluid for breast cancer diagnosis using the nanopore of Phi29 DNA-packaging motor.

Detection of cancer in its early stage is a challenging task for oncologists. Inflammatory breast cancer has symptoms that are similar to mastitis and can be mistaken for microbial infection. Currently, the differential diagnosis between mastitis and Inflammatory breast cancer via nipple aspirate fluid (NAF) is difficult. Here, we report a label-free and amplification-free detection platform using an engineered nanopore of the phi29 DNA-packaging motor with biomarker Galectin3 (GAL3), Thomsen-Friedenreich (TF) binding peptide as the probe fused at its C-terminus. The binding of the biomarker in NAF samples from breast cancer patients to the probe results in the connector's conformational change with a current blockage of 32 %. Utilization of dwell time, blockage ratio, and peak signature enable us to detect basal levels of biomarkers from patient NAF samples at the single-molecule level. This platform will allow for breast cancers to be resolved at an early stage with accuracy and thoroughness.

KCTD1 mutants in scalp­ear­nipple syndrome and AP­2α P59A in Char syndrome reciprocally abrogate their interactions, but can regulate Wnt/ß­catenin signaling.

Potassium­channel tetramerization-domain-containing 1 (KCTD1) mutations are reported to result in scalp­ear­nipple syndrome. These mutations occur in the conserved broad­complex, tramtrack and bric a brac domain, which is associated with inhibited transcriptional activity. However, the mechanisms of KCTD1 mutants have not previously been elucidated; thus, the present study aimed to investigate whether KCTD1 mutants affect their interaction with transcription factor AP­2α and their regulation of the Wnt pathway. Results from the present study demonstrated that none of the ten KCTD1 mutants had an inhibitory effect on the transcriptional activity of AP­2α. Co­immunoprecipitation assays demonstrated that certain mutants exhibited changeable localization compared with the nuclear localization of wild­type KCTD1, but no KCTD1 mutant interacted with AP­2α. Almost all KCTD1 mutants, except KCTD1 A30E and H33Q, exhibited differential inhibitory effects on regulating TOPFLASH luciferase reporter activity. In addition, the interaction region of KCTD1 to the PY motif (amino acids 59­62) in AP­2α was identified. KCTD1 exhibited no suppressive effects on the transcriptional activity of the AP­2α P59A mutant, resulting in Char syndrome, a genetic disorder characterized by a distinctive facial appearance, heart defect and hand abnormalities, by altered protein cellular localization that abolished protein interactions. However, the P59A, P60A, P61R and 4A AP­2α mutants inhibited TOPFLASH reporter activity. Moreover, AP­2α and KCTD1 inhibited ß­catenin expression levels and SW480 cell viability. The present study thus identified a putative mechanism of disease­related KCTD1 mutants and AP­2α mutants by disrupting their interaction with the wildtype proteins AP­2α and KCTD1 and influencing the regulation of the Wnt/ß­catenin pathway.

Correlation of Ductoscopic and Histopathological Findings and Their Relevance as Predictors for Malignancy: A German Multicenter Study.

BACKGROUND/AIM: The study aimed at investigating the correlation between ductoscopic and histopathological findings and clarify whether the former allow for accurate prediction of malignancy. PATIENTS AND METHODS: The prospective national multi-center study covered a sample of 224 patients with pathologic nipple discharge. A total of 214 patients underwent ductoscopy with subsequent extirpation of the mammary duct. The ductoscopic findings were categorized according to shape, number, color and surface structure of lesions and vascularity and compared to the histological results and analyses. RESULTS: Ductoscopy revealed lesions in 134 of 214 patients (62.2%). The criteria "multiple versus solitary lesion" differed significantly between malignant and benign lesions. All other criteria were not statistically significant. Malignant tumors were more frequently presented as multiple lesions, benign lesions or masses as solitary lesions (80% vs. 24.8%; p=0.018). CONCLUSION: The ductoscopic criterion "solitary vs. multiple lesion" appears to have a low diagnostic prediction of malignancy or benignity.

Application of Nipple Aspirate Fluid miRNA Profiles for Early Breast Cancer Detection and Management.

The authors of the recently published review, "Why the Gold Standard Approach by Mammography Demands Extension by Multiomics [...].

Nanoemulsions Enhance in vitro Transpapillary Diffusion of Model Fluorescent Dye Nile Red.

While the feasibility of transpapillary drug delivery has previously been established, localized transport via the mammary ducts may be improved with tailored drug delivery formulations. The objective of this study was to investigate the impact of nanoemulsion encapsulation on transpapillary delivery in vitro. Nanoemulsion formulations composed of isopropyl myristate and Tween 80 encapsulating a fluorescent dye were applied topically on porcine nipples using a Franz diffusion cell. A combination of dye extraction and fluorescence image analysis was used to quantify the total amount of dye retained within the nipple and to characterize the penetration routes. After diffusion for 6 hours, the amount of dye deposited in the nipple was proportional to the formulation's water concentration. The 90% water formulation deposited significantly more dye via both the stratum corneum and mammary ducts, while the 80% and 70% water formulations moderately increased ductal penetration, but minimally altered stratum corneum penetration as compared to the control solution. Similar trends were found after diffusion for 48 hours; however, the overall impact was diminished, likely due to the nanoemulsion's topical instability. This study indicates that drug delivery vehicles, nanoemulsions specifically, enhance delivery of encapsulated molecules via the stratum corneum and mammary ducts in a formulation-dependent basis.

Asymmetric Ductal Ectasia: An Often Overlooked Sign of Malignancy.

OBJECTIVE. The objective of this article is to define the clinical significance of asymmetric ductal ectasia by a review of literature and to describe the imaging findings. CONCLUSION. Asymmetric ductal ectasia has a significant risk for malignancy and high-risk lesions. The findings on conventional imaging may be subtle and easily overlooked. Asymmetric ductal ectasia should be included in the search pattern during image interpretation. Tissue sampling is usually warranted. Ultrasound is critical in identifying ductal abnormalities to guide biopsy.

Liposomes Enhance Dye Localization within the Mammary Ducts of Porcine Nipples.

Transductal and transepidermal diffusion are two distinct penetration routes of molecules administered via the nipple. To improve the therapeutic potential of this drug administration technique, drug penetration into the mammary ducts should be maximized, which may be accomplished through design optimization of drug delivery vehicles. In this study, we evaluated liposomes, ranging in size from 100 to 3000 nm, to improve ductal penetration of model fluorescent dyes using fluorescence microscopy and image analysis. Liposomes encapsulating a model fluorescent lipophilic dye, nile red, or hydrophilic dye, sulforhodamine B, were applied topically on porcine nipples for 6 h in vitro. Liposome encapsulation of sulforhodamine B significantly reduced the total amount of dye penetrating the nipple, while penetration of liposome-encapsulated nile red varied depending on vesicle size, as compared to their solution controls. However, the fluorescence intensity localized at the ductal epithelium was higher at extended nipple depths in tissues treated with liposomes versus dye solutions, suggesting a higher concentration of dye penetrating the nipple via the ducts. In contrast, the fluorescence intensity measured at the stratum corneum was reduced (sulforhodamine B) or unchanged (nile red) in nipples treated with liposomes versus dye solutions, suggesting a decrease or no change in dye penetration of the nipple via the stratum corneum. Furthermore, the limited penetration distance into the connective tissue beyond the ductal epithelium for both liposome-encapsulated nile red and sulforhodamine B suggests that liposomes remain intact over the 6 h duration of this study when penetrating through the ducts and enhance retention within the ductal lumen. However, the varied penetration profiles into the connective tissue beyond the stratum corneum between liposome-encapsulated nile red and sulforhodamine B suggests that the liposomes destabilize when penetrating the outer tissues layers of the nipple. Overall, liposomes, regardless of size, improved penetration into and retention within the mammary ducts, while limiting penetration into the stratum corneum, indicating their capacity to target the mammary ductal network.

Nipple leiomyoma: A rare neoplasm with a broad spectrum of histologic appearances.

Cutaneous leiomyomas are rare benign smooth-muscle tumors. These lesions are distinguished based on their cell of origin and are subclassified as pilar leiomyoma, angioleiomyoma, and genital-type leiomyoma. Nipple leiomyoma is the least common genital-type leiomyoma, arising from the dartoic muscle cell of the nipple. Histologic examination of the lesion is necessary for definitive diagnosis, and these uncommon tumors can pose a diagnostic challenge. We describe herein a series of six nipple leiomyomas with a spectrum of histologic appearances.

Factors associated to breast implants and breastfeeding. / Fatores associados à mamoplastia de aumento e o aleitamento materno.

OBJECTIVE: To analyze the association between the surgical characteristics of breast implants, time elapsed since surgery, access route, implant placement and implanted volume and variables related to breastfeeding, type, first 'milk let-down', breast engorgement, pain, lesion, milk production and use of galactagogues. METHOD: A prospective cohort carried out during the hospital stay (12 to 72 hours after delivery), home care (5thto 7thday after delivery) and telephone contact (between the 30thand 32ndday postpartum) of 115 postpartum women with breast implants between 2015 and 2017. RESULTS: The first evaluation identified more frequent use of oral galactagogues (p=0.029) by puerperal women with prepectoral implants, and of oxytocin spray by those with implants up to 270 ml (p=0.040). The second evaluation showed a higher pain score among those with prepectoral implants (p=0.046). Around the 30thday postpartum, the presence of nipple lesion (p=0.021), pain (p=0.025) and a higher pain score (p=0.039) was more frequent among those with mammoplasty performed less than 10 years ago. CONCLUSION: The presence of pain and a higher pain score, the occurrence of lesion and the use of oral and nasal galactagogues were associated with implant placement, implant size and time elapsed since surgery.

Short communication: Teat-end shape and udder-level milking characteristics and their associations with machine milking-induced changes in teat tissue condition.

Machine milking-induced changes in teat tissue condition, such as congestion and edema, have been associated with teat canal openness, penetrability, and thus susceptibility to new intramammary infections and diminished animal well-being. The primary objective of this study was to investigate the association of teat-end shape and machine milking-induced short-term changes in teat tissue condition. Our secondary objective was to study the association of udder-level milking characteristics and short-term changes. Data from 125 Holstein cows were analyzed in a longitudinal prospective cohort study. Cows were housed in freestall pens with sand bedding, fed a total mixed ration, and milked 3 times per day. Teat-end shape was classified into 3 categories: pointed, flat, and round. Udder-level milking characteristics were obtained from electronic on-farm milk meters. Short-term changes in teat tissue condition after machine milking were assessed visually. Multivariable generalized mixed model analysis showed an association between teat-end shape and short-term changes. Compared with teats with round teat-end shape, the odds ratio (95% confidence interval) for short-term changes was 1.68 (0.53-5.31) and 0.03 (0.004-0.19) in teats with pointed and flat teat-end shape, respectively. There was an association between milking characteristics and short-term changes such that higher milk flow rate during the first 15 s of milking decreased the likelihood of short-term changes. The adjusted probability of short-term changes for a milking observation of a mid-lactation cow and an average first 15-s milk flow rate of 0.5 and 1.5 kg/min was 53.0% (42.8-63.8) and 32.9% (15.2-57.3), respectively. Our results suggest that teat-end shape may be one of the risk factors that contribute to machine milking-induced short-term changes. Milking characteristics (e.g., first 15-s milk flow rate) may have the potential as a measure to indirectly monitor teat tissue changes associated with machine milking on a daily basis, though further research is needed to validate this hypothesis and to establish thresholds that could serve as on-farm guidelines.

Determination of permeation pathways of hydrophilic or hydrophobic dyes through the mammary papilla.

The transport pathways and permeation kinetics of lipophilic and hydrophilic fluorescent dyes through porcine mammary papillae were visualized and quantified. Porcine mammary papillae, removed from full-thickness abdominal tissue, were positioned in a Franz diffusion cell for passive diffusion studies. Solutions containing the fluorescent dyes were applied topically for time periods ranging from 30 min to 48 h. Dye concentrations in tissue and Franz diffusion compartments were analyzed using fluorescence microscopy and fluorimetry. Fluorescence micrographs elucidated two permeation pathways, transepidermal and transductal. Hydrophilic sulforhodamine B predominantly penetrated via the transepidermal route, while lipophilic nile red diffused mainly by the transductal route. An almost 4-fold higher amount of sulforhodamine B was retained within the nipple over time compared to nile red, despite both dyes permeating through the tissue at similar rates. Diffusion through the porcine nipple was 500-fold higher than through adjacent skin for both dyes, likely attributable to the two mammary ducts which provide an entry point and transport route through the tissue. These results, generated from both qualitative and quantitative evidence at a micro and macro scale, demonstrate that the mammary ducts provide a direct pathway that contributes significantly to passive transport through the nipple, particularly for lipophilic molecules.

Chemoprevention of Breast Cancer by Transdermal Delivery of α-Santalol through Breast Skin and Mammary Papilla (Nipple).

PURPOSE: Almost all breast cancers originate from epithelial cells lining the milk ducts in the breast. To this end, the study investigated the feasibility of localized transdermal delivery of α-santalol, a natural chemopreventive agent to the breast. METHODS: Different α-santalol formulations (cream, solution and microemulsion) were developed and the in vitro permeability was studied using excised animal (porcine and rat) and human breast skin/mammary papilla (nipple). The in vivo biodistribution and efficacy studies were conducted in female rats. A chemical carcinogenesis model of breast cancer was used for the efficacy studies. RESULTS: Phospholipid based α-santalol microemulsion showed the highest penetration through the nipple and breast skin. Delivery of α-santalol through the entire breast (breast skin and nipple) in vivo in rats resulted in significantly higher concentration in the mammary gland compared to transdermal delivery through the breast skin or nipple. There was no measurable α-santalol concentration in the blood. Transdermal delivery of α-santalol reduced the tumor incidence and tumor multiplicity. Furthermore, the tumor size was significantly reduced with α-santalol treatment. CONCLUSIONS: The findings from this study demonstrate the feasibility of localized transdermal delivery of α-santalol for chemoprevention of breast cancer.

Diagnosis of Nipple Discharge: Value of Magnetic Resonance Imaging and Ultrasonography in Comparison with Ductoscopy.

BACKGROUND: Pathologic nipple discharge, which is a common reason for referral to the breast imaging service, refers to spontaneous or bloody nipple discharge that arises from a single duct. The most common cause of nipple discharge is benign breast lesions, such as solitary intraductal papilloma and papillomatosis. Nevertheless, in rare cases, a malignant cause of nipple discharge can be found. AIMS: To study the diagnostic value of ultrasonography, magnetic resonance imaging, and ductoscopy in patients with pathologic nipple discharge, compare their efficacy, and investigate the importance of magnetic resonance imaging in the diagnosis of intraductal pathologies. STUDY DESIGN: Diagnostic accuracy study. METHODS: Fifty patients with pathologic nipple discharge were evaluated by ultrasonography and magnetic resonance imaging. Of these, 44 ductoscopic investigations were made. The patients were classified according to magnetic resonance imaging, ultrasonography, and ductoscopy findings. A total of 25 patients, whose findings were reported as intraductal masses, underwent surgery oincluding endoscopic excision for two endoscopic excision. Findings were compared with the pathology results that were accepted as the gold standard in the description of the aetiology of nipple discharge. In addition, magnetic resonance imaging, ultrasonography and ductoscopy findings were analysed comparatively in patients who had no surgery. RESULTS: Intraductal masses were reported in 26 patients, 20 of whom operated and established accurate diagnosis of 18 patients on magnetic resonance imaging. According to the ultrasonography, intraductal masses were identified in 22 patients, 17 of whom underwent surgery. Ultrasonography established accurate diagnoses in 15 patients. Intraductal mass was identified in 22 patients and ductoscopy established accurate diagnoses based on histopathologic results in 16 patients. The sensitivities of methods were 75% in ultrasonography, 90% in magnetic resonance imaging, and 94.6% in ductoscopy. The specificities were 66.7% in ultrasonography, 66.7% in magnetic resonance imaging, and 40% in ductoscopy. Intraductal papillomas were mostly observed as oval nodules with well-circumscribed smooth margins within dilated ducts and persistant in the dynamic analysis. Lesions that protruded into the lumen of the ducts, either solitary or multiple, were characteristic ductoscopy findings of our patients who were diagnosed as having papilloma/papillomatosis. CONCLUSION: Magnetic resonance imaging and ductoscopy had no statistical superiority over each other, however they were superior to ultrasonography in the diagnosis of pathologic nipple discharge. Magnetic resonance imaging may be highly sensitive for diagnosing nipple discharge with new techniques and sequences and a non-invasive method that more advantageous for showing ductal tree visualization and is able to detect completely obstructed intraductal lesions.

Influence of Hydroalcoholic Vehicle on In Vitro Transport of 4-Hydroxy Tamoxifen Through the Mammary Papilla (Nipple).

Majority of breast cancers originate from epithelial cells in the duct and lobules in the breast. Current systemic treatments for breast cancer are associated with significant systemic side effects, thus warranting localized drug delivery approaches. The aim of this study was to investigate the influence of hydroalcoholic vehicle on topical delivery of 4-hydroxy tamoxifen (4-HT) through the mammary papilla (nipple). The in vitro permeability of 4-HT through porcine mammary papilla was studied using different hydroalcoholic vehicles (0, 33.33, and 66.66% alcohol). Nile red was used as a model lipophilic dye to characterize the drug transport pathway in the mammary papilla. The penetration of 4-HT through the mammary papilla increased with increase in alcohol concentration in the vehicle. The solubility of 4-HT was enhanced by increasing alcohol concentration in the vehicle. On the other hand, the epidermis/vehicle partition coefficient decreased with increase in alcohol concentration. The mammary papilla served as a depot and slowly released 4-HT into the receptor medium. Highest drug penetration was observed with saturated drug solution in 66.66% alcohol, and 4-HT levels were comparable to IC50 value of 4-HT. Results from this study demonstrate the possibility of using mammary papilla as a potential route for direct delivery of 4-HT to the breast.

Transpapillary (Nipple) Delivery of Macromolecules to the Breast: Proof of Concept Study.

Localized drug delivery to the breast can maximize drug concentration at the target site and minimize systemic drug distribution. To this end, the study explored the feasibility of delivering macromolecules to the breast through mammary papilla (nipple). The in vitro penetration of model macromolecules (inulin, dextran, ovalbumin, and bovine serum albumin) varying in molecular weight from 5 to 67 kDa was studied using excised porcine and human mammary papilla. The penetration of macromolecules decreased with increase in molecular weight. The penetration of the macromolecules was significantly higher through the mammary papilla in comparison to breast skin. In vitro penetration of the macromolecules was similar in human and porcine mammary papilla. Iontophoresis was used to enhance the transport of bovine serum albumin (BSA) through the mammary papilla. The flux and cumulative amount permeated was increased by 2- to 4-fold by iontophoresis. The macromolecules were transported through the ducts and the surrounding connective tissue in the mammary papilla. Overall, the results from this study for the first time demonstrate the feasibility of delivering macromolecules through the mammary papilla. These findings have implications for developing safe and effective localized therapeutic approaches for breast cancer.

Cytokeratin 7-negative mammary Paget's disease: A diagnostic pitfall.

Pathologists should be aware of the existence of a rare CK7-negative variant of breast carcinoma in general, and of Paget's disease in particular. Cytokeratin 7-negative Paget's disease and CK7-negative ductal intraepithelial neoplasia (ductal carcinoma in situ) present a major diagnostic challenge for pathologists since there is limited awareness of their existence. When there is classic Paget's morphology on H&E sections, GATA3 positivity should resolve any doubts about the diagnosis in the setting of a CK7-negative neoplastic cell population.