Fluorescence and Multiphoton Imaging for Tissue Characterization of a Model of Postmenopausal Ovarian Cancer.

BACKGROUND AND OBJECTIVES: To determine the efficacy of targeted fluorescent biomarkers and multiphoton imaging to characterize early changes in ovarian tissue with the onset of cancer. STUDY DESIGN/MATERIALS AND METHODS: A transgenic TgMISIIR-TAg mouse was used as an animal model for ovarian cancer. Mice were injected with fluorescent dyes to bind to the folate receptor α, matrix metalloproteinases, and integrins. Half of the mice were treated with 4-vinylcyclohexene diepoxide (VCD) to simulate menopause. Widefield fluorescence imaging (WFI) and multiphoton imaging of the ovaries and oviducts were conducted at 4 and 8 weeks of age. The fluorescence signal magnitude was quantified, and texture features were derived from multiphoton imaging. Linear discriminant analysis was then used to classify mouse groups. RESULTS: Imaging features from both fluorescence imaging and multiphoton imaging show significant changes (P < 0.01) with age, VCD treatment, and genotype. The classification model is able to classify different groups to accuracies of 75.53%, 69.53%, and 86.76%, for age, VCD treatment, and genotype, respectively. Building a classification model using features from multiple modalities shows marked improvement over individual modalities. CONCLUSIONS: This study demonstrates that using WFI with targeted biomarkers, and multiphoton imaging with endogenous contrast shows promise for detecting early changes in ovarian tissue with the onset of cancer. The results indicate that multimodal imaging can provide higher sensitivity for classifying tissue types than using single modalities alone. Lasers Surg. Med. © 2020 Wiley Periodicals, Inc.

In vivo molecular mapping of the tumor microenvironment in an azoxymethane-treated mouse model of colon carcinogenesis.

BACKGROUND AND OBJECTIVE: Development of miniaturized imaging systems with molecular probes enables examination of molecular changes leading to initiation and progression of colorectal cancer in an azoxymethane (AOM)-induced mouse model of the disease. Through improved and novel studies of animal disease models, more effective diagnostic and treatment strategies may be developed for clinical translation. We introduce use of a miniaturized multimodal endoscope with lavage-delivered fluorescent probes to examine dynamic microenvironment changes in an AOM-treated mouse model. STUDY DESIGN/MATERIALS AND METHODS: The endoscope is equipped with optical coherence tomography (OCT) and laser induced fluorescence (LIF) imaging modalities. It is used with Cy5.5-conjugated antibodies to create time-resolved molecular maps of colon carcinogenesis. We monitored in vivo changes in molecular expression over a five month period for four biomarkers: epithelial growth factor receptor (EGFR), transferrin receptor (TfR), transforming growth factor beta 1 (TGFß1), and chemokine (C-X-C motif) receptor 2 (CXCR2). In vivo OCT and LIF images were compared over multiple time points to correlate increases in biomarker expression with adenoma development. RESULTS: This system is uniquely capable of tracking in vivo changes in molecular expression over time. Increased expression of the biomarker panel corresponded to sites of disease and offered predictive utility in highlighting sites of disease prior to detectable structural changes. Biomarker expression also tended to increase with higher tumor burden and growth rate in the colon. CONCLUSION: We can use miniaturized dual modality endoscopes with fluorescent probes to study the tumor microenvironment in developmental animal models of cancer and supplement findings from biopsy and tissue harvesting.

Dual optical modality endoscopic imaging of cancer development in the mouse colon.

BACKGROUND AND OBJECTIVE: We utilize a miniature, dual-modality endoscope that combines fluorescence-based surface magnifying chromoendoscopy (SMC) and optical coherence tomography (OCT) to follow the anatomical changes that occur during adenoma development in the mouse colon. MATERIALS AND METHODS: Twenty-five mice were treated with the carcinogen azoxymethane (AOM) to induce tumor development in the distal colon, or were treated with saline as control, and were imaged over six months. OCT detects adenoma number with high sensitivity and specificity and can measure lesion size. In methylene blue-lavaged colons, SMC detects changes in the colonic crypts. SMC images of control mouse colons exhibit reticulated patterns of crypts of equal size, forming either a dot or honeycomb pattern. RESULTS: Images of AOM-treated colons show mild crypt irregularities even in grossly normal tissue. Images of small to medium adenoma exhibit larger crypts, more intense signal, and irregular spacing whereas those of large adenoma have heterogeneous, intense signal and loss of crypt structure. CONCLUSIONS: The combination of OCT and SMC permits the detection of neoplastic events from the earliest stages of crypt irregularities before gross tissue changes are noted, through to measuring the growth of protruding adenoma.

Two-photon excited fluorescence imaging of endogenous contrast in a mouse model of ovarian cancer.

BACKGROUND AND OBJECTIVE: Ovarian cancer has an extremely high mortality rate resulting from poor understanding of the disease. In order to aid understanding of disease etiology and progression, we identify the endogenous fluorophores present in a mouse model of ovarian cancer and describe changes in fluorophore abundance and distribution with age and disease. STUDY DESIGN/MATERIALS AND METHODS: A mouse model of ovarian cancer was created by dosing with 4-vinylcyclohexene diepoxide, which induces follicular apoptosis (simulating menopause), and 7,12-dimethylbenz[a]anthracene, a known carcinogen. Imaging of ovarian tissue was completed ex vivo with a multiphoton microscope using excitation wavelength of 780 nm and emission collection from 405 to 505 nm. Two-photon excited fluorescence images and corresponding histologic sections with selective stains were used to identify endogenous fluorophores. RESULTS: The majority of collected fluorescence emission was attributed to NADH and lipofuscin, with additional contributions from collagen and elastin. Dim cellular fluorescence from NADH did not show observable changes with age. Changes in ovarian morphology with disease development frequently caused increased fluorescence contributions from collagen and adipose tissue-associated NADH. Lipofuscin fluorescence was much brighter than NADH fluorescence and increased as a function of both age and disease. CONCLUSIONS: Our finding of NADH fluorescence patterns similar to that seen previously in human ovary, combined with the observation of lipofuscin accumulation with age and disease also seen in human organs, suggests that the findings from this model may be relevant to human ovarian disease. Increased lipofuscin fluorescence might be used as an indicator of disease in the ovary and this finding warrants further study.

Iterative prototyping based on lessons learned from the falloposcope in vivo pilot study experience.

Significance: High grade serous ovarian cancer is the most deadly gynecological cancer, and it is now believed that most cases originate in the fallopian tubes (FTs). Early detection of ovarian cancer could double the 5-year survival rate compared with late-stage diagnosis. Autofluorescence imaging can detect serous-origin precancerous and cancerous lesions in ex vivo FT and ovaries with good sensitivity and specificity. Multispectral fluorescence imaging (MFI) can differentiate healthy, benign, and malignant ovarian and FT tissues. Optical coherence tomography (OCT) reveals subsurface microstructural information and can distinguish normal and cancerous structure in ovaries and FTs. Aim: We developed an FT endoscope, the falloposcope, as a method for detecting ovarian cancer with MFI and OCT. The falloposcope clinical prototype was tested in a pilot study with 12 volunteers to date to evaluate the safety and feasibility of FT imaging prior to standard of care salpingectomy in normal-risk volunteers. In this manuscript, we describe the multiple modifications made to the falloposcope to enhance robustness, usability, and image quality based on lessons learned in the clinical setting. Approach: The ∼0.8 mm diameter falloposcope was introduced via a minimally invasive approach through a commercially available hysteroscope and introducing a catheter. A navigation video, MFI, and OCT of human FTs were obtained. Feedback from stakeholders on image quality and procedural difficulty was obtained. Results: The falloposcope successfully obtained images in vivo. Considerable feedback was obtained, motivating iterative improvements, including accommodating the operating room environment, modifying the hysteroscope accessories, decreasing endoscope fragility and fiber breaks, optimizing software, improving fiber bundle images, decreasing gradient-index lens stray light, optimizing the proximal imaging system, and improving the illumination. Conclusions: The initial clinical prototype falloposcope was able to image the FTs, and iterative prototyping has increased its robustness, functionality, and ease of use for future trials.

Study of fallopian tube anatomy and mechanical properties to determine pressure limits for endoscopic exploration.

Falloposcopy is the endoscopic examination of the fallopian tubes, which are challenging to access due to their deep body location, small opening from the uterus, and lumen filled with plicae. We and others have developed endoscopes that are inserted through the uterus guided by a hysteroscope into the tubal ostium. To better understand how to utilize these endoscopes either as standalone devices or in concert with everting delivery balloons, a preliminary study of anatomy and mechanical behavior was performed ex vivo on porcine and human fallopian tubes. Segments of fallopian tubes from the isthmus, ampulla and infundibulum were inflated with saline either to bursting or held at sub-burst pressures with saline or a saline-filled balloon. Formalin fixed, paraffin embedded tissue sections stained with Masson's trichrome were examined for damage to the mucosa and muscularis. Porcine fallopian tubes tolerated saline pressurization at 15 psi for 1 minute without morphological damage. Balloon inflation to 15 psi caused no apparent damage to the muscle layer or rupture of the fallopian tube, but balloon movement within the tube can denude the mucosal epithelial layer. Human fallopian tubes averaged higher burst pressure values than porcine tubes. Under pressurization, the external tube diameter expanded by minimal to moderate amounts. Human and porcine tissues were similar in histological appearance. These studies suggest that moderate pressurization is acceptable but will not appreciably expand the fallopian tube diameter. The results also indicate that pigs are a reasonable model to study damage from falloscopy as seen in human tissue.

Feasibility of optical coherence tomography imaging to characterize renal neoplasms: limitations in resolution and depth of penetration.

UNLABELLED: What's known on the subject? and What does the study add? Optical coherence tomography has been used for the diagnosis of retinal disease and has been used experimentally for imaging of vascular plaques, gastrointestinal pathology, bladder cancer, prostate cancer, and recently to examine benign kidney microanatomy. It has not been previously used to image kidney cancer. This study presents the first data on the utility of OCT in the imaging for renal neoplasms. It found that OCT was most successful in distinguishing AML and TCC from normal parenchyma. OCT had more limited success at differentiating oncocytoma. Clear cell tumors and other renal cancer subtypes had a more heterogenous appearance, precluding reliable identification using OCT. The study shows that higher resolution versions of OCT, such as OCM, will be needed to allow optical coherence imaging to reach clinical utility in the assessment of renal neoplasms. OBJECTIVES: • To determine the appearance of normal and neoplastic renal tissue when imaged with optical coherence tomography (OCT). • To preliminarily assess the feasibility of using OCT to differentiate normal and neoplastic renal tissue. PATIENTS AND METHODS: • After radical or partial nephrectomy in 20 subjects, normal renal parenchyma and neoplastic tissue samples were obtained. • The tissue was evaluated with light microscopy and using a bench-top laboratory OCT system with a lateral resolution of 10 µm. • OCT images were compared with histological slides to evaluate the ability of OCT to differentiate renal neoplasms. RESULTS: • Pathological subtypes included eight clear-cell, three papillary and two chromophobe renal carcinomas; two oncocytomas; one angiomyolipoma (AML); two transitional cell carcinomas (TCCs); and one haematoma. • Using OCT, benign renal parenchyma showed recognizable glomeruli and tubules. • TCC had a distinctive appearance on OCT whereas AML showed a unique identifiable signature because of its fat content. Oncocytomas had a lobulated appearance, which appeared subtly different from renal carcinoma. • Renal carcinoma lacked recognizable anatomical elements and had a heterogeneous appearance making differentiation from normal parenchyma at times difficult. • Subtypes of renal cancer appeared to vary on OCT imaging although discrimination was unreliable. CONCLUSIONS: • OCT imaging for renal neoplasms was most successful in distinguishing AML and TCC from normal parenchyma and malignant tumours. Oncocytoma differed subtly from renal carcinoma, making distinction more challenging. • Clear-cell tumours and other renal carcinoma subtypes had a heterogeneous appearance on OCT, which precluded reliable differentiation from normal parenchyma and between renal carcinoma subtypes. • Higher resolution versions of optical coherence imaging, such as optical coherence microscopy, will be necessary to achieve clinical utility.

Sub-millimeter endoscope demonstrates feasibility of in vivo reflectance imaging, fluorescence imaging, and cell collection in the fallopian tubes.

SIGNIFICANCE: Most cases of high-grade serous ovarian carcinoma originate as serous tubal intraepithelial carcinoma (STIC) lesions in the fallopian tube epithelium (FTE), enabling early endoscopic detection. AIM: The cell-acquiring fallopian endoscope (CAFE) was built to meet requirements for locating potentially pathological tissue indicated by an alteration in autofluorescence or presence of a targeted fluorophore. A channel was included for directed scrape biopsy of cells from regions of interest. APPROACH: Imaging resolution and fluorescence sensitivity were measured using a standard resolution target and fluorescence standards, respectively. A prototype was tested in ex vivo tissue, and collected cells were counted and processed. RESULTS: Measured imaging resolution was 88 µm at a 5-mm distance, and full field of view was ∼45 deg in air. Reflectance and fluorescence images in ex vivo porcine reproductive tracts were captured, and fit through human tracts was verified. Hemocytometry counts showed that on the order of 105 cells per scrape biopsy could be collected from ex vivo porcine tissue. CONCLUSIONS: All requirements for viewing STIC in the FTE were met, and collected cell counts exceeded input requirements for relevant analyses. Our benchtop findings suggest the potential utility of the CAFE device for in vivo imaging and cell collection in future clinical trials.

Evaluation of segmentation algorithms for optical coherence tomography images of ovarian tissue.

Ovarian cancer has the lowest survival rate among all gynecologic cancers predominantly due to late diagnosis. Early detection of ovarian cancer can increase 5-year survival rates from 40% up to 92%, yet no reliable early detection techniques exist. Optical coherence tomography (OCT) is an emerging technique that provides depth-resolved, high-resolution images of biological tissue in real-time and demonstrates great potential for imaging of ovarian tissue. Mouse models are crucial to quantitatively assess the diagnostic potential of OCT for ovarian cancer imaging; however, due to small organ size, the ovaries must first be separated from the image background using the process of segmentation. Manual segmentation is time-intensive, as OCT yields three-dimensional data. Furthermore, speckle noise complicates OCT images, frustrating many processing techniques. While much work has investigated noise-reduction and automated segmentation for retinal OCT imaging, little has considered the application to the ovaries, which exhibit higher variance and inhomogeneity than the retina. To address these challenges, we evaluate a set of algorithms to segment OCT images of mouse ovaries. We examine five preprocessing techniques and seven segmentation algorithms. While all preprocessing methods improve segmentation, Gaussian filtering is most effective, showing an improvement of 32 % ± 1.2 % . Of the segmentation algorithms, active contours performs best, segmenting with an accuracy of 94.8 % ± 1.2 % compared with manual segmentation. Even so, further optimization could lead to maximizing the performance for segmenting OCT images of the ovaries.

Comparison of Reproductive Function in Female TgMISIIR-TAg Transgenic and Wildtype C57BL/6 Mice.

Transgenic TgMISIIR-TAg (TAg) mice express the oncogenic virus SV40 in Mullerian epithelial cells. Female TAg mice spontaneously develop epithelial ovarian carcinoma, the most common type of ovarian cancer in women. Female TAg mice are infertile, but the reason has not been determined. We therefore investigated whether female TAg mice undergo puberty, demonstrate follicular development, maintain regular cycles, and ovulate. Ovarian cancers in women commonly develop after menopause. The occupational chemical 4-vinylcyclohexene diepoxide (VCD) accelerates follicle degeneration in the ovaries of rats and mice, causing early ovarian failure. We therefore used VCD dosing of mice to develop an animal model for menopause. The purpose of this study was to characterize reproductive parameters in female TAg mice and to investigate whether the onset of ovarian failure due VCD dosing differed between female TAg and WT C57BL/6 mice. As in WT female mice, TAg female mice underwent puberty (vaginal opening) and developed cyclicity in patterns that were similar between the groups. Vehicle-only TAg mice had fewer ovulations (numbers of corpora lutea) than WT animals. VCD exposure delayed the onset of puberty (day of first estrus) in TAg as compared with WT mice. Morphologic evaluation of ovaries revealed many more degenerating follicles in TAg mice than WT mice, and more VCD-dosed TAg mice were in ovarian failure than VCD-dosed WT mice. These results suggest that despite showing similar onset of sexual maturation, TAg mice have increased follicular degeneration and fewer ovulations than WT. These features may contribute to the inability of female TAg mice to reproduce.

Three-dimensional texture analysis of optical coherence tomography images of ovarian tissue.

Ovarian cancer has the lowest survival rate among all gynecologic cancers due to predominantly late diagnosis. Optical coherence tomography (OCT) has been applied successfully to experimentally image the ovaries in vivo; however, a robust method for analysis is still required to provide quantitative diagnostic information. Recently, texture analysis has proved to be a useful tool for tissue characterization; unfortunately, existing work in the scope of OCT ovarian imaging is limited to only analyzing 2D sub-regions of the image data, discarding information encoded in the full image area, as well as in the depth dimension. Here we address these challenges by testing three implementations of texture analysis for the ability to classify tissue type. First, we test the traditional case of extracted 2D regions of interest; then we extend this to include the entire image area by segmenting the organ from the background. Finally, we conduct a full volumetric analysis of the image volume using 3D segmented data. For each case, we compute features based on the Grey-Level Co-occurence Matrix and also by introducing a new approach that evaluates the frequency distribution in the image by computing the energy density. We test these methods on a mouse model of ovarian cancer to differentiate between age, genotype, and treatment. The results show that the 3D application of texture analysis is most effective for differentiating tissue types, yielding an average classification accuracy of 78.6%. This is followed by the analysis in 2D with the segmented image volume, yielding an average accuracy of 71.5%. Both of these improve on the traditional approach of extracting square regions of interest, which yield an average classification accuracy of 67.7%. Thus, applying texture analysis in 3D with a fully segmented image volume is the most robust approach to quantitatively characterizing ovarian tissue.

Does Mutated K-RAS Oncogene Attenuate the Effect of Sulindac in Colon Cancer Chemoprevention?

The NSAID sulindac has been successfully used alone or in combination with other agents to suppress colon tumorigenesis in patients with genetic predisposition and also showed its efficacy in prevention of sporadic colon adenomas. At the same time, some experimental and clinical reports suggest that a mutant K-RAS oncogene may negate sulindac antitumor efficacy. To directly assess sulindac activity at suppressing premalignant lesions carrying K-RAS mutation, we utilized a novel mouse model with an inducible colon-specific expression of the mutant K-ras oncogene (K-rasG12D ). Tumor development and treatment effects were monitored by minimally invasive endoscopic Optical coherence tomography. Expression of the mutant K-ras allele accelerated azoxymethane (AOM)-induced colon carcinogenesis in C57BL/6 mice, a strain otherwise resistant to this carcinogen. Sulindac completely prevented AOM-induced tumor formation in K-ras wild-type (K-ras wt) animals. In K-rasG12D -mutant mice, a 38% reduction in tumor number, an 83% reduction in tumor volume (P ≤ 0.01) and an increase in the number of adenoma-free mice (P = 0.04) were observed. The partial response of K-RasG12D animals to sulindac treatment was evident by the decrease in mucosal thickness (P < 0.01) and delay in progression of the precancerous aberrant crypt foci to adenomas. Molecular analyses showed significant induction in cyclooxygenase 2 (COX-2), cleaved caspase-3 (CC3), and Ki-67 expression by AOM, but not sulindac treatment, in all genotypes. Our data underscore the importance of screening for K-RAS mutations in individuals with colon polyps to provide more personalized interventions targeting mutant K-RAS signaling pathways. Cancer Prev Res; 11(1); 16-26. ©2017 AACR.

Volume holographic imaging endoscopic design and construction techniques.

A reflectance volume holographic imaging (VHI) endoscope has been designed for simultaneous in vivo imaging of surface and subsurface tissue structures. Prior utilization of VHI systems has been limited to ex vivo tissue imaging. The VHI system presented in this work is designed for laparoscopic use. It consists of a probe section that relays light from the tissue sample to a handheld unit that contains the VHI microscope. The probe section is constructed from gradient index (GRIN) lenses that form a 1:1 relay for image collection. The probe has an outer diameter of 3.8 mm and is capable of achieving 228.1 ?? lp / mm resolution with 660-nm Kohler illumination. The handheld optical section operates with a magnification of 13.9 and a field of view of 390 ?? ? m × 244 ?? ? m . System performance is assessed through imaging of 1951 USAF resolution targets and soft tissue samples. The system has also passed sterilization procedures required for surgical use and has been used in two laparoscopic surgical procedures.

Wavelength-coded volume holographic imaging endoscope for multidepth imaging.

A wavelength-coded volume holographic imaging (WC-VHI) endoscope system capable of simultaneous multifocal imaging is presented. The system images light from two depths separated by 100 µm in a tissue sample by using axial chromatic dispersion of a gradient index probe in combination with two light-emitting diode sources and a multiplexed volume hologram to separate the images. This system is different from previous VHI systems in that it uses planar multiplexed gratings and does not require curved holographic gratings. This results in improved lateral imaging resolution from 228.1 to 322.5 lp/mm. This letter describes the design and fabrication of the WC-VHI endoscope and experimental images of hard and soft resolution targets and biological tissue samples to illustrate the performance properties.

Intermittent Dosing with Sulindac Provides Effective Colorectal Cancer Chemoprevention in the Azoxymethane-Treated Mouse Model.

Sulindac is an NSAID that can provide effective chemoprevention for colorectal cancer. In this study, alternative dosing regimens of sulindac were evaluated for their chemoprevention effectiveness in the azoxymethane-treated A/J mouse model of colorectal cancer. High-resolution endoscopic optical coherence tomography was utilized to time-serially measure tumor number and tumor burden in the distal colon as the biological endpoints. Four treatment groups were studied: (i) daily for 20 weeks (sulindac-daily); (ii) for 2 weeks, then no sulindac for 2 weeks, cycle repeated 5 times (sulindac-2); (iii) for 10 weeks ("on"), then no sulindac for 10 weeks ("off"; sulindac-10); and (iv) no sulindac (sulindac-none). Sulindac-2 and sulindac-daily had statistically significantly lower final tumor counts and slopes (change in number of tumors per week) when compared with sulindac-none (P < 0.0001). All of the treatment groups had statistically significantly lower final tumor burdens and slopes when compared with sulindac-none (P < 0.001). There was a prolonged latency period in the sulindac-10 group, with no significant difference between the "off" portion of this treatment and sulindac-none. These results suggest that, although daily doses of sulindac provide the most optimal effects, intermittent doses of sulindac in a 50% duty cycle with an overall 4-week period (sulindac-2 model) can provide highly effective chemoprevention of colorectal cancer in this model. After cessation of sulindac treatment (sulindac-10 "off"), there is no evidence of either a persistent chemopreventive effect or a rebound effect. Cancer Prev Res; 10(8); 459-66. ©2017 AACR.

Imaging colon cancer development in mice: IL-6 deficiency prevents adenoma in azoxymethane-treated Smad3 knockouts.

The development of colorectal cancer in the azoxymethane-induced mouse model can be observed by using a miniaturized optical coherence tomography (OCT) imaging system. This system is uniquely capable of tracking disease development over time, allowing for the monitoring of morphological changes in the distal colon due to tumor development and the presence of lymphoid aggregates. By using genetically engineered mouse models deficient in Interleukin 6 (IL-6) and Smad family member 3 (Smad3), the role of inflammation on tumor development and the immune system can be elucidated. Smad3 knockout mice develop inflammatory response, wasting, and colitis associated cancer while deficiency of proinflammatory cytokine IL-6 confers resistance to tumorigenesis. We present pilot data showing that the Smad3 knockout group had the highest tumor burden, highest spleen weight, and lowest thymus weight. The IL-6 deficiency in Smad3 knockout mice prevented tumor development, splenomegaly, and thymic atrophy. This finding suggests that agents that inhibit IL-6 (e.g. anti-IL-6 antibody, non-steroidal anti-inflammatory drugs [NSAIDs], etc.) could be used as novel therapeutic agents to prevent disease progression and increase the efficacy of anti-cancer agents. OCT can also be useful for initiating early therapy and assessing the benefit of combination therapy targeting inflammation.

Multispectral fluorescence imaging of human ovarian and fallopian tube tissue for early-stage cancer detection.

With early detection, 5-year survival rates for ovarian cancer exceed 90%, yet no effective early screening method exists. Emerging consensus suggests over 50% of the most lethal form of the disease originates in the fallopian tube. Twenty-eight women undergoing oophorectomy or debulking surgery provided informed consent for the use of surgical discard tissue samples for multispectral fluorescence imaging. Using multiple ultraviolet and visible excitation wavelengths and emissions bands, 12 fluorescence and 6 reflectance images of 47 ovarian and 31 fallopian tube tissue samples were recorded. After imaging, each sample was fixed, sectioned, and stained for pathological evaluation. Univariate logistic regression showed cancerous tissue samples had significantly lower intensity than noncancerous tissue for 17 image types. The predictive power of multiple image types was evaluated using multivariate logistic regression (MLR) and quadratic discriminant analysis (QDA). Two MLR models each using two image types had receiver operating characteristic curves with area under the curve exceeding 0.9. QDA determined 56 image type combinations with perfect resubstituting using as few as five image types. Adaption of the system for future in vivo fallopian tube and ovary endoscopic imaging is possible, which may enable sensitive detection of ovarian cancer with no exogenous contrast agents.

Time-serial Assessment of Drug Combination Interventions in a Mouse Model of Colorectal Carcinogenesis Using Optical Coherence Tomography.

Optical coherence tomography (OCT) is a high-resolution, nondestructive imaging modality that enables time-serial assessment of adenoma development in the mouse model of colorectal cancer. In this study, OCT was utilized to evaluate the effectiveness of interventions with the experimental antitumor agent α-difluoromethylornithine (DFMO) and a nonsteroidal anti-inflammatory drug sulindac during early [chemoprevention (CP)] and late stages [chemotherapy (CT)] of colon tumorigenesis. Biological endpoints for drug interventions included OCT-generated tumor number and tumor burden. Immunochistochemistry was used to evaluate biochemical endpoints [Ki-67, cleaved caspase-3, cyclooxygenase (COX)-2, ß-catenin]. K-Ras codon 12 mutations were studied with polymerase chain reaction-based technique. We demonstrated that OCT imaging significantly correlated with histological analysis of both tumor number and tumor burden for all experimental groups (P < 0.0001), but allows more accurate and full characterization of tumor number and burden growth rate because of its time-serial, nondestructive nature. DFMO alone or in combination with sulindac suppressed both the tumor number and tumor burden growth rate in the CP setting because of DFMO-mediated decrease in cell proliferation (Ki-67, P < 0.001) and K-RAS mutations frequency (P = 0.04). In the CT setting, sulindac alone and DFMO/sulindac combination were effective in reducing tumor number, but not tumor burden growth rate. A decrease in COX-2 staining in DFMO/sulindac CT groups (COX-2, P < 0.01) confirmed the treatment effect. Use of nondestructive OCT enabled repeated, quantitative evaluation of tumor number and burden, allowing changes in these parameters to be measured during CP and as a result of CT. In conclusion, OCT is a robust minimally invasive method for monitoring colorectal cancer disease and effectiveness of therapies in mouse models.

In vivo time-serial multi-modality optical imaging in a mouse model of ovarian tumorigenesis.

Identification of the early microscopic changes associated with ovarian cancer may lead to development of a diagnostic test for high-risk women. In this study we use optical coherence tomography (OCT) and multiphoton microscopy (MPM) (collecting both two photon excited fluorescence [TPEF] and second harmonic generation [SHG]) to image mouse ovaries in vivo at multiple time points. We demonstrate the feasibility of imaging mouse ovaries in vivo during a long-term survival study and identify microscopic changes associated with early tumor development. These changes include alterations in tissue microstructure, as seen by OCT, alterations in cellular fluorescence and morphology, as seen by TPEF, and remodeling of collagen structure, as seen by SHG. These results suggest that a combined OCT-MPM system may be useful for early detection of ovarian cancer.

Diagnostic potential of multimodal imaging of ovarian tissue using optical coherence tomography and second-harmonic generation microscopy.

Ovarian cancer is particularly deadly because it is usually diagnosed after it has metastasized. We have previously identified features of ovarian cancer using optical coherence tomography (OCT) and second-harmonic generation (SHG) microscopy (targeting collagen). OCT provides an image of the ovarian microstructure while SHG provides a high-resolution map of collagen fiber bundle arrangement. Here we investigated the diagnostic potential of dual-modality OCT and SHG imaging. We conducted a fully crossed, multi-reader, multi-case study using seven human observers. Each observer classified 44 ex vivo mouse ovaries (16 normal and 28 abnormal) as normal or abnormal from OCT, SHG, and simultaneously viewed, co-registered OCT and SHG images and provided a confidence rating on a six-point scale. We determined the average receiver operating characteristic (ROC) curves, area under the ROC curves (AUC), and other quantitative figures of merit. The results show that OCT has diagnostic potential with an average AUC of 0.91 ± 0.06. The average AUC for SHG was less promising at 0.71 ± 0.13. The average AUC for simultaneous OCT and SHG was not significantly different from OCT alone, possibly due to the limited SHG field of view. The high performance of OCT and co-registered OCT and SHG warrants further investigation.