Publications by authors named "Jenna L Mueller"

17 Publications

  • Page 1 of 1

Intratumoral Photosensitizer Delivery and Photodynamic Therapy.

Nano Life 2021 Jun 9;11(2). Epub 2021 Jun 9.

Fischell Department of Bioengineering, University of Maryland, College Park, MD 20742, USA.

Photodynamic therapy (PDT) is a two-step procedure that involves the administration of special drugs, commonly called photosensitizers, followed by the application of certain wavelengths of light. The light activates these photosensitizers to produce reactive molecular species that induce cell death in tissues. There are numerous factors to consider when selecting the appropriate photosensitizer administration route, such as which part of the body is being targeted, the pharmacokinetics of photosensitizers, and the formulation of photosensitizers. While intravenous, topical, and oral administration of photosensitizers are widely used in preclinical and clinical applications of PDT, other administration routes, such as intraperitoneal, intra-arterial, and intratumoral injections, are gaining traction for their potential in treating advanced diseases and reducing off-target toxicities. With recent advances in targeted nanotechnology, biomaterials, and light delivery systems, the exciting possibilities of targeted photosensitizer delivery can be fully realized for preclinical and clinical applications. Further, in light of the growing burden of cancer mortality in low and middle-income countries and development of low-cost light sources and photosensitizers, PDT could be used to treat cancer patients in low-income settings. This short article introduces aspects of interfaces of intratumoral photosensitizer injections and nano-biomaterials for PDT applications in both high-income and low-income settings but does not present a comprehensive review due to space limitations.
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http://dx.doi.org/10.1142/s179398442130003xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8412167PMC
June 2021

Optimizing ethyl cellulose-ethanol delivery towards enabling ablation of cervical dysplasia.

Sci Rep 2021 Aug 19;11(1):16869. Epub 2021 Aug 19.

Department of Biomedical Engineering, Duke University, Durham, NC, USA.

In low-income countries, up to 80% of women diagnosed with cervical dysplasia do not return for follow-up care, primarily due to treatment being inaccessible. Here, we describe development of a low-cost, portable treatment suitable for such settings. It is based on injection of ethyl cellulose (EC)-ethanol to ablate the transformation zone around the os, the site most impacted by dysplasia. EC is a polymer that sequesters the ethanol within a prescribed volume when injected into tissue, and this is modulated by the injected volume and delivery parameters (needle gauge, bevel orientation, insertion rate, depth, and infusion rate). Salient injection-based delivery parameters were varied in excised swine cervices. The resulting injection distribution volume was imaged with a wide-field fluorescence imaging device or computed tomography. A 27G needle and insertion rate of 10 mm/s achieved the desired insertion depth in tissue. Orienting the needle bevel towards the outer edge of the cervix and keeping infusion volumes ≤ 500 µL minimized leakage into off-target tissue. These results guided development of a custom hand-held injector, which was used to locate and ablate the upper quadrant of a swine cervix in vivo with no adverse events or changes in host temperature or heart rate. After 24 h, a distinct region of necrosis was detected that covered a majority (> 75%) of the upper quadrant of the cervix, indicating four injections could effectively cover the full cervix. The work here informs follow up large animal in vivo studies, e.g. in swine, to further assess safety and efficacy of EC-ethanol ablation in the cervix.
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http://dx.doi.org/10.1038/s41598-021-96223-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8376953PMC
August 2021

The Evolving Landscape of Medical Device Regulation in East, Central, and Southern Africa.

Glob Health Sci Pract 2021 03 31;9(1):136-148. Epub 2021 Mar 31.

School of Medicine, Duke University, Durham, NC, USA.

Effective regulatory frameworks, harmonized to international standards, are critical to expanding access to quality medical devices in low- and middle-income countries. This review provides a summary of the state of medical device regulation in the 14 member countries of the College of Surgeons of East, Central, and Southern Africa (COSECSA) and South Africa. Countries were categorized according to level of regulatory establishment, which was found to be positively correlated to gross domestic product (GDP; r=0.90) and years of freedom from colonization (r=0.60), and less positively correlated to GDP per capita (r=0.40). Although most countries mandate medical device regulation in national legislation, few employ all the guidelines set forth by the World Health Organization. A streamlined regulatory process across African nations would simplify this process for innovators seeking to bring medical devices to the African market, thereby increasing patient access to safe medical devices.
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http://dx.doi.org/10.9745/GHSP-D-20-00578DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8087432PMC
March 2021

An Accessible Laparoscope for Surgery in Low- and Middle- Income Countries.

Ann Biomed Eng 2021 Jul 8;49(7):1657-1669. Epub 2021 Mar 8.

Duke Global Health Institute, Durham, NC, USA.

Laparoscopic surgery is the standard of care in high-income countries for many procedures in the chest and abdomen. It avoids large incisions by using a tiny camera and fine instruments manipulated through keyhole incisions, but it is generally unavailable in low- and middle-income countries (LMICs) due to the high cost of installment, lack of qualified maintenance personnel, unreliable electricity, and shortage of consumable items. Patients in LMICs would benefit from laparoscopic surgery, as advantages include decreased pain, improved recovery time, fewer wound infections, and shorter hospital stays. To address this need, we developed an accessible laparoscopic system, called the ReadyView laparoscope for use in LMICs. The device includes an integrated camera and LED light source that can be displayed on any monitor. The ReadyView laparoscope was evaluated with standard optical imaging targets to determine its performance against a state-of-the-art commercial laparoscope. The ReadyView laparoscope has a comparable resolving power, lens distortion, field of view, depth of field, and color reproduction accuracy to a commercially available endoscope, particularly at shorter, commonly-used working distances (3-5 cm). Additionally, the ReadyView has a cooler temperature profile, decreasing the risk for tissue injury and operating room fires. The ReadyView features a waterproof design, enabling sterilization by submersion, as commonly performed in LMICs. A custom desktop software was developed to view the video on a laptop computer with a frame rate greater than 30 frames per second and to white balance the image, which is critical for clinical use. The ReadyView laparoscope is capable of providing the image quality and overall performance needed for laparoscopic surgery. This portable low-cost system is well suited to increase access to laparoscopic surgery in LMICs.
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http://dx.doi.org/10.1007/s10439-020-02707-6DOI Listing
July 2021

Polymer-assisted intratumoral delivery of ethanol: Preclinical investigation of safety and efficacy in a murine breast cancer model.

PLoS One 2021 28;16(1):e0234535. Epub 2021 Jan 28.

Department of Biomedical Engineering, Duke University, Durham, North Carolina, United States of America.

Focal tumor ablation with ethanol could provide benefits in low-resource settings because of its low overall cost, minimal imaging technology requirements, and acceptable clinical outcomes. Unfortunately, ethanol ablation is not commonly utilized because of a lack of predictability of the ablation zone, caused by inefficient retention of ethanol at the injection site. To create a predictable zone of ablation, we have developed a polymer-assisted ablation method using ethyl cellulose (EC) mixed with ethanol. EC is ethanol-soluble and water-insoluble, allowing for EC-ethanol to be injected as a liquid and precipitate into a solid, occluding the leakage of ethanol upon contact with tissue. The aims of this study were to compare the 1) safety, 2) release kinetics, 3) spatial distribution, 4) necrotic volume, and 5) overall survival of EC-ethanol to conventional ethanol ablation in a murine breast tumor model. Non-target tissue damage was monitored through localized adverse events recording, ethanol release kinetics with Raman spectroscopy, injectate distribution with in vivo imaging, target-tissue necrosis with NADH-diaphorase staining, and overall survival by proxy of tumor growth. EC-ethanol exhibited decreased localized adverse events, a slowing of the release rate of ethanol, more compact injection zones, 5-fold increase in target-tissue necrosis, and longer overall survival rates compared to the same volume of pure ethanol. A single 150 μL dose of 6% EC-ethanol achieved a similar survival probability rates to six daily 50 μL doses of pure ethanol used to simulate a slow-release of ethanol over 6 days. Taken together, these results demonstrate that EC-ethanol is safer and more effective than ethanol alone for ablating tumors.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0234535PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7843014PMC
September 2021

Laparoscopic experience and attitudes toward a low-cost laparoscopic system among surgeons in East, Central, and Southern Africa: a survey study.

Surg Endosc 2020 Nov 17. Epub 2020 Nov 17.

Department of Surgery, DUMC, Duke University, Box 3815, Durham, NC, 27710, USA.

Background: Laparoscopic surgery has become standard of care in high-income countries but is rarely accessible in low- and middle-income countries (LMICs). This study assessed experience with laparoscopy and attitudes toward a low-cost laparoscopic system among surgeons in sub-Saharan Africa.

Methods: A survey assessing current laparoscopic practice and feedback on a low-cost laparoscopic system was administered to attendees of the College of Surgeons of East, Central, and Southern Africa (COSECSA) Scientific Conference between December 4 and December 6, 2019 in Kampala, Uganda.

Results: Fifty-six surgeons from 14 countries participated. A majority were male (n = 46, 82%) general surgeons (n = 37, 66%) from tertiary/teaching hospitals (n = 36, 64%). For those with training in laparoscopy (n = 33, 59%), 22 (67%) reported less than 1 year of training and over half (n = 17, 52%) reported 1 month or less. Overall, a minority (n = 21, 38%) used laparoscopy in current practice, with 57% (n = 12) of those performing laparoscopy less than once per week. The most common laparoscopic surgeries performed were cholecystectomy (n = 15), diagnostic laparoscopy (n = 14), and appendectomy (n = 12). Few surgeons were performing more complex cases (n = 5). Barriers to laparoscopy included poor access to training equipment (n = 34, 61%), mentors (n = 33, 59%), laparoscopic equipment (n = 31, 55%), equipment maintenance (n = 25, 45%), access to consumable supplies (n = 21, 38%), and cost (n = 31, 55%). Fifty-two participants (93%) were interested in increasing their use of laparoscopy; the majority felt that a low-cost laparoscope (n = 52, 93%) and lift retractor for gasless laparoscopy (n = 46, 82%) would serve an unmet need in their practice.

Conclusions: While the use of laparoscopy is currently limited in COSECSA countries, there is a significant interest among surgeons to increase implementation. A low-cost, durable laparoscopic system was viewed as a potential solution to the current barriers and could improve implementation in LMICs.
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http://dx.doi.org/10.1007/s00464-020-08151-wDOI Listing
November 2020

Understanding Factors Governing Distribution Volume of Ethyl Cellulose-Ethanol to Optimize Ablative Therapy in the Liver.

IEEE Trans Biomed Eng 2020 08 16;67(8):2337-2348. Epub 2019 Dec 16.

Objective: Ethanol ablation, the injection of ethanol to induce necrosis, was originally used to treat hepatocellular carcinoma, with survival rates comparable to surgery. However, efficacy is limited due to leakage into surrounding tissue. To reduce leakage, we previously reported incorporating ethyl cellulose (EC) with ethanol as this mixture forms a gel when injected into tissue. To further develop EC-ethanol injection as an ablative therapy, the present study evaluates the extent to which salient injection parameters govern the injected fluid distribution.

Methods: Utilizing ex vivo swine liver, injection parameters (infusion rate, EC%, infusion volume) were examined with fluorescein added to each solution. After injection, tissue samples were frozen, sectioned, and imaged.

Results: While leakage was higher for ethanol and 3%EC-ethanol at a rate of 10 mL/hr compared to 1 mL/hr, leakage remained low for 6%EC-ethanol regardless of infusion rate. The impact of infusion volume and pressure were also investigated first in tissue-mimicking surrogates and then in tissue. Results indicated that there is a critical infusion pressure beyond which crack formation occurs leading to fluid leakage. At a rate of 10 mL/hr, a volume of 50 μL remained below the critical pressure.

Conclusions: Although increasing the infusion rate increases stress on the tissue and the risk of crack formation, injections of 6%EC-ethanol were localized regardless of infusion rate. To further limit leakage, multiple low-volume infusions may be employed.

Significance: These results, and the experimental framework developed to obtain them, can inform optimizing EC-ethanol to treat a range of medical conditions.
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http://dx.doi.org/10.1109/TBME.2019.2960049DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7295656PMC
August 2020

Development of Algorithms for Automated Detection of Cervical Pre-Cancers With a Low-Cost, Point-of-Care, Pocket Colposcope.

IEEE Trans Biomed Eng 2019 08 18;66(8):2306-2318. Epub 2018 Dec 18.

Goal: In this paper, we propose methods for (1) automatic feature extraction and classification for acetic acid and Lugol's iodine cervigrams and (2) methods for combining features/diagnosis of different contrasts in cervigrams for improved performance.

Methods: We developed algorithms to pre-process pathology-labeled cervigrams and extract simple but powerful color and textural-based features. The features were used to train a support vector machine model to classify cervigrams based on corresponding pathology for visual inspection with acetic acid, visual inspection with Lugol's iodine, and a combination of the two contrasts.

Results: The proposed framework achieved a sensitivity, specificity, and accuracy of 81.3%, 78.6%, and 80.0%, respectively, when used to distinguish cervical intraepithelial neoplasia (CIN+) relative to normal and benign tissues. This is superior to the average values achieved by three expert physicians on the same data set for discriminating normal/benign cases from CIN+ (77% sensitivity, 51% specificity, and 63% accuracy).

Conclusion: The results suggest that utilizing simple color- and textural-based features from visual inspection with acetic acid and visual inspection with Lugol's iodine images may provide unbiased automation of cervigrams.

Significance: This would enable automated, expert-level diagnosis of cervical pre-cancer at the point of care.
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http://dx.doi.org/10.1109/TBME.2018.2887208DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6581620PMC
August 2019

Development of enhanced ethanol ablation as an alternative to surgery in treatment of superficial solid tumors.

Sci Rep 2017 08 18;7(1):8750. Epub 2017 Aug 18.

Department of Biomedical Engineering, Duke University, Durham, North Carolina, United States of America.

While surgery is at the foundation of cancer treatment, its access is limited in low-income countries. Here, we describe development of a low-cost alternative therapy based on intratumoral ethanol injection suitable for resource-limited settings. Although ethanol-based tumor ablation is successful in treating hepatocellular carcinomas, the necessity for multiple treatments, injection of large fluid volumes, and decreased efficacy in treatment of non-capsulated tumors limit its applicability. To address these limitations, we investigated an enhanced ethanol ablation strategy to retain ethanol within the tumor through the addition of ethyl cellulose. This increases the viscosity of injected ethanol and forms an ethanol-based gel-phase upon exposure to the aqueous tumor environment. This technique was first optimized to maximize distribution volume, using tissue-simulating phantoms. Then, chemically-induced epithelial tumors in the hamster cheek pouch were treated. As controls, pure ethanol injections of either four times or one-fourth the tumor volume induced complete regression of 33% and 0% of tumors, respectively. In contrast, ethyl cellulose-ethanol injections of one-fourth the tumor volume induced complete regression in 100% of tumors. These results contribute to proof-of-concept for enhanced ethanol ablation as a novel and effective alternative to surgery for tumor treatment, with relevance to resource-limited settings.
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http://dx.doi.org/10.1038/s41598-017-09371-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5562881PMC
August 2017

International Image Concordance Study to Compare a Point-of-Care Tampon Colposcope With a Standard-of-Care Colposcope.

J Low Genit Tract Dis 2017 Apr;21(2):112-119

1Department of Biomedical Engineering, Duke University, Durham, NC; 2Duke Global Health Institute, Duke University, Durham, NC; 3Department of Surgery, Duke University Medical Center, Durham, NC; 4Department of Biostatistics and Bioinformatics, Duke University Medical School, Durham, NC; 5Institute of Cytology and Preventative Oncology (ICMR), New Delhi, India; 6Cancer Institute (WIA), Chennai, India; 7Department of Obstetrics and Gynecology, Duke University Medical Center, Durham, NC; 8Kilimanjaro Christian Medical Centre, Moshi, Tanzania; 9Liga Contra el Cáncer, Pueblo Libre, Lima, Peru; and 10Department of Obstetrics and Gynecology, School of Medicine, Kenyatta University, Nairobi, Kenya.

Objective: Barriers to cervical cancer screening in low-resource settings include lack of accessible, high-quality services, high cost, and the need for multiple visits. To address these challenges, we developed a low-cost, intravaginal, optical cervical imaging device, the point-of-care tampon (POCkeT) colposcope and evaluated whether its performance is comparable with a standard-of-care colposcope.

Materials And Methods: There were 2 protocols, which included 44 and 18 patients. For the first protocol, white-light cervical images were collected in vivo, blinded by device, and sent electronically to 8 physicians from high-, middle-, and low-income countries. For the second protocol, green-light images were also collected and sent electronically to the highest performing physician from the first protocol who has experience in both a high- and low-income country. For each image, physicians completed a survey assessing cervix characteristics and severity of precancerous lesions. Corresponding pathology was obtained for all image pairs.

Results: For the first protocol, average percent agreement between devices was 70% across all physicians. The POCkeT and standard-of-care colposcope images had 37% and 51% agreement with pathology for high-grade squamous intraepithelial lesions (HSILs), respectively. Investigation of HSIL POCkeT images revealed decreased visibility of vascularization and lack of contrast in lesion margins. After changes were made for the second protocol, the 2 devices achieved similar agreement to pathology for HSIL lesions (55%).

Conclusions: Based on the exploratory study, physician interpretation of cervix images acquired using a portable, low-cost POCkeT colposcope was comparable to a standard-of-care colposcope.
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http://dx.doi.org/10.1097/LGT.0000000000000306DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5365351PMC
April 2017

Rapid staining and imaging of subnuclear features to differentiate between malignant and benign breast tissues at a point-of-care setting.

J Cancer Res Clin Oncol 2016 Jul 22;142(7):1475-86. Epub 2016 Apr 22.

Department of Biomedical Engineering, Duke University, 136 Hudson Hall Box 90281, Durham, NC, 27708, USA.

Purpose: Histopathology is the clinical standard for tissue diagnosis; however, it requires tissue processing, laboratory personnel and infrastructure, and a highly trained pathologist to diagnose the tissue. Optical microscopy can provide real-time diagnosis, which could be used to inform the management of breast cancer. The goal of this work is to obtain images of tissue morphology through fluorescence microscopy and vital fluorescent stains and to develop a strategy to segment and quantify breast tissue features in order to enable automated tissue diagnosis.

Methods: We combined acriflavine staining, fluorescence microscopy, and a technique called sparse component analysis to segment nuclei and nucleoli, which are collectively referred to as acriflavine positive features (APFs). A series of variables, which included the density, area fraction, diameter, and spacing of APFs, were quantified from images taken from clinical core needle breast biopsies and used to create a multivariate classification model. The model was developed using a training data set and validated using an independent testing data set.

Results: The top performing classification model included the density and area fraction of smaller APFs (those less than 7 µm in diameter, which likely correspond to stained nucleoli).When applied to the independent testing set composed of 25 biopsy panels, the model achieved a sensitivity of 82 %, a specificity of 79 %, and an overall accuracy of 80 %.

Conclusions: These results indicate that our quantitative microscopy toolbox is a potentially viable approach for detecting the presence of malignancy in clinical core needle breast biopsies.
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http://dx.doi.org/10.1007/s00432-016-2165-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4900949PMC
July 2016

A Fluorescence-Guided Laser Ablation System for Removal of Residual Cancer in a Mouse Model of Soft Tissue Sarcoma.

Theranostics 2016 1;6(2):155-66. Epub 2016 Jan 1.

6. Department of Orthopaedic Surgery, Duke University Medical Center, Durham, North Carolina.

The treatment of soft tissue sarcoma (STS) generally involves tumor excision with a wide margin. Although advances in fluorescence imaging make real-time detection of cancer possible, removal is limited by the precision of the human eye and hand. Here, we describe a novel pulsed Nd:YAG laser ablation system that, when used in conjunction with a previously described molecular imaging system, can identify and ablate cancer in vivo. Mice with primary STS were injected with the protease-activatable probe LUM015 to label tumors. Resected tissues from the mice were then imaged and treated with the laser using the paired fluorescence-imaging/ laser ablation device, generating ablation clefts with sub-millimeter precision and minimal underlying tissue damage. Laser ablation was guided by fluorescence to target tumor tissues, avoiding normal structures. The selective ablation of tumor implants in vivo improved recurrence-free survival after tumor resection in a cohort of 14 mice compared to 12 mice that received no ablative therapy. This prototype system has the potential to be modified so that it can be used during surgery to improve recurrence-free survival in patients with cancer.
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http://dx.doi.org/10.7150/thno.13536DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4729765PMC
January 2017

Structured Illumination Microscopy and a Quantitative Image Analysis for the Detection of Positive Margins in a Pre-Clinical Genetically Engineered Mouse Model of Sarcoma.

PLoS One 2016 22;11(1):e0147006. Epub 2016 Jan 22.

Department of Biomedical Engineering, Duke University, Durham, North Carolina, United States of America.

Intraoperative assessment of surgical margins is critical to ensuring residual tumor does not remain in a patient. Previously, we developed a fluorescence structured illumination microscope (SIM) system with a single-shot field of view (FOV) of 2.1 × 1.6 mm (3.4 mm2) and sub-cellular resolution (4.4 μm). The goal of this study was to test the utility of this technology for the detection of residual disease in a genetically engineered mouse model of sarcoma. Primary soft tissue sarcomas were generated in the hindlimb and after the tumor was surgically removed, the relevant margin was stained with acridine orange (AO), a vital stain that brightly stains cell nuclei and fibrous tissues. The tissues were imaged with the SIM system with the primary goal of visualizing fluorescent features from tumor nuclei. Given the heterogeneity of the background tissue (presence of adipose tissue and muscle), an algorithm known as maximally stable extremal regions (MSER) was optimized and applied to the images to specifically segment nuclear features. A logistic regression model was used to classify a tissue site as positive or negative by calculating area fraction and shape of the segmented features that were present and the resulting receiver operator curve (ROC) was generated by varying the probability threshold. Based on the ROC curves, the model was able to classify tumor and normal tissue with 77% sensitivity and 81% specificity (Youden's index). For an unbiased measure of the model performance, it was applied to a separate validation dataset that resulted in 73% sensitivity and 80% specificity. When this approach was applied to representative whole margins, for a tumor probability threshold of 50%, only 1.2% of all regions from the negative margin exceeded this threshold, while over 14.8% of all regions from the positive margin exceeded this threshold.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0147006PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4723137PMC
July 2016

Micro-anatomical quantitative optical imaging: toward automated assessment of breast tissues.

Breast Cancer Res 2015 Aug 20;17:105. Epub 2015 Aug 20.

Department of Bioengineering, Rice University, 6500 Main Street, BRC 502, Houston, TX, 77030, USA.

Introduction: Pathologists currently diagnose breast lesions through histologic assessment, which requires fixation and tissue preparation. The diagnostic criteria used to classify breast lesions are qualitative and subjective, and inter-observer discordance has been shown to be a significant challenge in the diagnosis of selected breast lesions, particularly for borderline proliferative lesions. Thus, there is an opportunity to develop tools to rapidly visualize and quantitatively interpret breast tissue morphology for a variety of clinical applications.

Methods: Toward this end, we acquired images of freshly excised breast tissue specimens from a total of 34 patients using confocal fluorescence microscopy and proflavine as a topical stain. We developed computerized algorithms to segment and quantify nuclear and ductal parameters that characterize breast architectural features. A total of 33 parameters were evaluated and used as input to develop a decision tree model to classify benign and malignant breast tissue. Benign features were classified in tissue specimens acquired from 30 patients and malignant features were classified in specimens from 22 patients.

Results: The decision tree model that achieved the highest accuracy for distinguishing between benign and malignant breast features used the following parameters: standard deviation of inter-nuclear distance and number of duct lumens. The model achieved 81 % sensitivity and 93 % specificity, corresponding to an area under the curve of 0.93 and an overall accuracy of 90 %. The model classified IDC and DCIS with 92 % and 96 % accuracy, respectively. The cross-validated model achieved 75 % sensitivity and 93 % specificity and an overall accuracy of 88 %.

Conclusions: These results suggest that proflavine staining and confocal fluorescence microscopy combined with image analysis strategies to segment morphological features could potentially be used to quantitatively diagnose freshly obtained breast tissue at the point of care without the need for tissue preparation.
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http://dx.doi.org/10.1186/s13058-015-0617-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4545917PMC
August 2015

A quantitative microscopic approach to predict local recurrence based on in vivo intraoperative imaging of sarcoma tumor margins.

Int J Cancer 2015 Nov 3;137(10):2403-12. Epub 2015 Jun 3.

Department of Biomedical Engineering, Duke University, Durham, North Carolina.

The goal of resection of soft tissue sarcomas located in the extremity is to preserve limb function while completely excising the tumor with a margin of normal tissue. With surgery alone, one-third of patients with soft tissue sarcoma of the extremity will have local recurrence due to microscopic residual disease in the tumor bed. Currently, a limited number of intraoperative pathology-based techniques are used to assess margin status; however, few have been widely adopted due to sampling error and time constraints. To aid in intraoperative diagnosis, we developed a quantitative optical microscopy toolbox, which includes acriflavine staining, fluorescence microscopy, and analytic techniques called sparse component analysis and circle transform to yield quantitative diagnosis of tumor margins. A series of variables were quantified from images of resected primary sarcomas and used to optimize a multivariate model. The sensitivity and specificity for differentiating positive from negative ex vivo resected tumor margins was 82 and 75%. The utility of this approach was tested by imaging the in vivo tumor cavities from 34 mice after resection of a sarcoma with local recurrence as a bench mark. When applied prospectively to images from the tumor cavity, the sensitivity and specificity for differentiating local recurrence was 78 and 82%. For comparison, if pathology was used to predict local recurrence in this data set, it would achieve a sensitivity of 29% and a specificity of 71%. These results indicate a robust approach for detecting microscopic residual disease, which is an effective predictor of local recurrence.
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http://dx.doi.org/10.1002/ijc.29611DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4575838PMC
November 2015

Optimization of a widefield structured illumination microscope for non-destructive assessment and quantification of nuclear features in tumor margins of a primary mouse model of sarcoma.

PLoS One 2013 23;8(7):e68868. Epub 2013 Jul 23.

Department of Biomedical Engineering, Duke University, Durham, North Carolina, USA.

Cancer is associated with specific cellular morphological changes, such as increased nuclear size and crowding from rapidly proliferating cells. In situ tissue imaging using fluorescent stains may be useful for intraoperative detection of residual cancer in surgical tumor margins. We developed a widefield fluorescence structured illumination microscope (SIM) system with a single-shot FOV of 2.1 × 1.6 mm (3.4 mm(2)) and sub-cellular resolution (4.4 µm). The objectives of this work were to measure the relationship between illumination pattern frequency and optical sectioning strength and signal-to-noise ratio in turbid (i.e. thick) samples for selection of the optimum frequency, and to determine feasibility for detecting residual cancer on tumor resection margins, using a genetically engineered primary mouse model of sarcoma. The SIM system was tested in tissue mimicking solid phantoms with various scattering levels to determine impact of both turbidity and illumination frequency on two SIM metrics, optical section thickness and modulation depth. To demonstrate preclinical feasibility, ex vivo 50 µm frozen sections and fresh intact thick tissue samples excised from a primary mouse model of sarcoma were stained with acridine orange, which stains cell nuclei, skeletal muscle, and collagenous stroma. The cell nuclei were segmented using a high-pass filter algorithm, which allowed quantification of nuclear density. The results showed that the optimal illumination frequency was 31.7 µm(-1) used in conjunction with a 4 × 0.1 NA objective (v=0.165). This yielded an optical section thickness of 128 µm and an 8.9 × contrast enhancement over uniform illumination. We successfully demonstrated the ability to resolve cell nuclei in situ achieved via SIM, which allowed segmentation of nuclei from heterogeneous tissues in the presence of considerable background fluorescence. Specifically, we demonstrate that optical sectioning of fresh intact thick tissues performed equivalently in regards to nuclear density quantification, to physical frozen sectioning and standard microscopy.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0068868PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3720887PMC
March 2014

Quantitative Segmentation of Fluorescence Microscopy Images of Heterogeneous Tissue: Application to the Detection of Residual Disease in Tumor Margins.

PLoS One 2013 18;8(6):e66198. Epub 2013 Jun 18.

Department of Biomedical Engineering, Duke University, Durham, North Carolina, United States of America.

Purpose: To develop a robust tool for quantitative in situ pathology that allows visualization of heterogeneous tissue morphology and segmentation and quantification of image features.

Materials And Methods: TISSUE EXCISED FROM A GENETICALLY ENGINEERED MOUSE MODEL OF SARCOMA WAS IMAGED USING A SUBCELLULAR RESOLUTION MICROENDOSCOPE AFTER TOPICAL APPLICATION OF A FLUORESCENT ANATOMICAL CONTRAST AGENT: acriflavine. An algorithm based on sparse component analysis (SCA) and the circle transform (CT) was developed for image segmentation and quantification of distinct tissue types. The accuracy of our approach was quantified through simulations of tumor and muscle images. Specifically, tumor, muscle, and tumor+muscle tissue images were simulated because these tissue types were most commonly observed in sarcoma margins. Simulations were based on tissue characteristics observed in pathology slides. The potential clinical utility of our approach was evaluated by imaging excised margins and the tumor bed in a cohort of mice after surgical resection of sarcoma.

Results: Simulation experiments revealed that SCA+CT achieved the lowest errors for larger nuclear sizes and for higher contrast ratios (nuclei intensity/background intensity). For imaging of tumor margins, SCA+CT effectively isolated nuclei from tumor, muscle, adipose, and tumor+muscle tissue types. Differences in density were correctly identified with SCA+CT in a cohort of ex vivo and in vivo images, thus illustrating the diagnostic potential of our approach.

Conclusion: The combination of a subcellular-resolution microendoscope, acriflavine staining, and SCA+CT can be used to accurately isolate nuclei and quantify their density in anatomical images of heterogeneous tissue.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0066198PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3688889PMC
October 2017
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