Partial nephrectomy margin imaging using structured illumination microscopy.

Partial nephrectomy (PN) is the recommended procedure over radical nephrectomy (RN) for patients with renal masses less than 4 cm in diameter (Stage T1a). Patients with less than 4 cm renal masses can also be treated with PN, but have a higher risk for positive surgical margins (PSM). PSM, when present, are indicative of poor clinical outcomes. The current gold-standard histopathology method is not well-suited for the identification of PSM intraoperatively due to processing time and destructive nature. Here, video-rate structured illumination microscopy (VR-SIM) was investigated as a potential tool for PSM detection during PN. A clinical image atlas assembled from ex vivo renal biopsies provided diagnostically useful images of benign and malignant kidney, similar to permanent histopathology. VR-SIM was then used to image entire parenchymal margins of tumor resection covering up to >1800× more margin surface area than standard histology. Aided by the image atlas, the study pathologist correctly classified all parenchymal margins as negative for PSM with VR-SIM, compared to standard postoperative pathology. The ability to evaluate large surgical margins in a short time frame with VR-SIM may allow it to be used intraoperatively as a "safety net" for PSM detection, allowing more patients to undergo PN over RN.

DRAQ5 and Eosin ('D&E') as an Analog to Hematoxylin and Eosin for Rapid Fluorescence Histology of Fresh Tissues.

Real-time on-site histopathology review of biopsy tissues at the point-of-procedure has great potential for significant clinical value and improved patient care. For instance, on-site review can aid in rapid screening of diagnostic biopsies to reduce false-negative results, or in quantitative assessment of biospecimen quality to increase the efficacy of downstream laboratory and histopathology analysis. However, the only currently available rapid pathology method, frozen section analysis (FSA), is too time- and labor-intensive for use in screening large quantities of biopsy tissues and is too destructive for maximum tissue conservation in multiple small needle core biopsies. In this work we demonstrate the spectrally-compatible combination of the nuclear stain DRAQ5 and the anionic counterstain eosin as a dual-component fluorescent staining analog to hematoxylin and eosin intended for use on fresh, unsectioned tissues. Combined with optical sectioning fluorescence microscopy and pseudo-coloring algorithms, DRAQ5 and eosin ("D&E") enables very fast, non-destructive psuedohistological imaging of tissues at the point-of-acquisition with minimal tissue handling and processing. D&E was validated against H&E on a one-to-one basis on formalin-fixed paraffin-embedded and frozen section tissues of various human organs using standard epi-fluorescence microscopy, demonstrating high fidelity of the staining mechanism as an H&E analog. The method was then applied to fresh, whole 18G renal needle core biopsies and large needle core prostate biospecimen biopsies using fluorescence structured illumination optical sectioning microscopy. We demonstrate the ability to obtain high-resolution histology-like images of unsectioned, fresh tissues similar to subsequent H&E staining of the tissue. The application of D&E does not interfere with subsequent standard-of-care H&E staining and imaging, preserving the integrity of the tissue for thorough downstream analysis. These results indicate that this dual-stain pseudocoloring method could provide a real-time histology-like image at the time of acquisition and valuable objective tissue analysis for the clinician at the time of service.

Nondestructive Diagnosis of Kidney Cancer on 18-gauge Core Needle Renal Biopsy Using Dual-color Fluorescence Structured Illumination Microscopy.

OBJECTIVE: To present a novel imaging technique used for rapid, nondestructive histological assessment of renal neoplasias using a dual-component fluorescence stain and structured illumination microscopy (SIM). MATERIALS AND METHODS: After Institutional Review Board approval, 65 total biopsies were obtained from 19 patients undergoing partial or radical nephrectomy. Biopsies were stained with a dual-component fluorescent, and optically sectioned SIM images were obtained from the surface of the intact biopsies. Specimens were subsequently fixed and analyzed using hematoxylin and eosin (H&E) histopathologic methods and compared with SIM images. A single, board-certified pathologist blinded to specimens reviewed all SIM images and H&E slides, and determined the presence or absence of neoplasias. Results of blinded diagnosis of SIM were validated against traditional pathology. RESULTS: Of the 19 patients, 15 underwent robotic partial nephrectomies and 4 underwent laparoscopic nephrectomies. Indications included clinical suspicion of renal cell carcinoma. In total, 65 biopsy specimens were available for review. Twenty-one specimens were determined to be neoplastic on H&E, whereas 41 represented benign renal tissue. The final sensitivity and specificity of our study were 79.2% and 95.1%, respectively. CONCLUSION: SIM is a promising technology for rapid, near-patient, ex vivo renal biopsy assessment. By improving the ability to rapidly assess sufficiency of biopsy specimens and enabling immediate diagnostic capability, SIM aids in more effective biopsy performance, tissue triage, and patient counseling regarding management options. Additionally, because tissue is preserved, effective utilization of downstream diagnostic tests and molecular assessments are possible.

Gigapixel surface imaging of radical prostatectomy specimens for comprehensive detection of cancer-positive surgical margins using structured illumination microscopy.

Achieving cancer-free surgical margins in oncologic surgery is critical to reduce the need for additional adjuvant treatments and minimize tumor recurrence; however, there is a delicate balance between completeness of tumor removal and preservation of adjacent tissues critical for normal post-operative function. We sought to establish the feasibility of video-rate structured illumination microscopy (VR-SIM) of the intact removed tumor surface as a practical and non-destructive alternative to intra-operative frozen section pathology, using prostate cancer as an initial target. We present the first images of the intact human prostate surface obtained with pathologically-relevant contrast and subcellular detail, obtained in 24 radical prostatectomy specimens immediately after excision. We demonstrate that it is feasible to routinely image the full prostate circumference, generating gigapixel panorama images of the surface that are readily interpreted by pathologists. VR-SIM confirmed detection of positive surgical margins in 3 out of 4 prostates with pathology-confirmed adenocarcinoma at the circumferential surgical margin, and furthermore detected extensive residual cancer at the circumferential margin in a case post-operatively classified by histopathology as having negative surgical margins. Our results suggest that the increased surface coverage of VR-SIM could also provide added value for detection and characterization of positive surgical margins over traditional histopathology.

Intralesional Injection of Interferon-α2b Improves Penile Curvature in Men with Peyronie's Disease Independent of Plaque Location.

PURPOSE: Collagenase clostridium histolyticum is the only FDA (Food and Drug Administration) approved treatment for Peyronie's disease. However, to our knowledge collagenase clostridium histolyticum has not been studied in men with ventral plaques. Given this limitation and the paucity of literature on ventral plaque outcomes, we compared the results of Peyronie's disease treatment in men with different plaque locations treated with intralesional interferon-α2b. MATERIALS AND METHODS: We retrospectively analyzed the records of men treated with intralesional interferon-α2b for Peyronie's disease at 1 institution from 2001 to 2014. The men received 2 million U interferon-α2b injected every 2 weeks for 6 to 24 treatments. All men underwent penile duplex Doppler ultrasound before and after interferon-α2b treatment. Patient characteristics, penile duplex Doppler ultrasound and objective measurements were reviewed. Patients were stratified into ventral and dorsal/lateral plaque cohorts with a positive response defined as a 20% or greater reduction in curvature. RESULTS: A total of 131 patients with a mean±SD age of 53.8±9.5 years underwent a median of 12 intralesional interferon-α2b injections (range 6 to 24). Mean pretreatment dorsal curvature was 42.5±18.6 degrees in group 1 of 111 men and mean ventral curvature was 44.5±21.5 degrees in group 2 of 21 men (p=0.66). Overall 91% of patients responded to therapy. No significant difference was noted between the 2 groups in response rate (54% vs 52%, p=0.92) or absolute change in curvature (mean 8.7±12.6 vs 9.3±17.7 degrees, p=0.84). CONCLUSIONS: Treatment with intralesional interferon-α2b provided a greater than 20% reduction in curvature in the majority of men with Peyronie's disease. This improvement was independent of plaque location.

Effect of botulinum-A toxin injection into bulbospongiosus muscle on ejaculation latency in male rats.

INTRODUCTION: Premature ejaculation (PE) is the most common male sexual dysfunction. A variety of pharmacotherapeutic strategies have been employed to treat men suffering with lifelong PE. However, there are currently no pharmaceuticals approved by the U.S. Food and Drug Administration specifically designed for PE treatment. AIM: Given that the bulbospongiosus muscle is involved in the ejaculatory reflex in both humans and rodents and that local administration of botulinum-A can abolish muscle contractions, the current study examined the effect of injection of botulinum-A toxin into the bulbospongiosus muscle on the ejaculatory latency of male rats. METHODS: After screening for normal sexual activity with sexually receptive female rats, 33 sexually experienced male Long-Evans rats (Harlan Laboratories, Indianapolis, IN, USA) underwent an additional four pretreatment sexual exposures over the course of the following week, during which all components of sexual behavior were video recorded by trained observers. On the day after their fourth experience, rats were anesthetized and received a single injection of either 0.5 unit (n = 11) or 1 unit (n = 11) of botulinum-A toxin or saline vehicle (n = 11). Botulinum-A toxin was dissolved in 0.1 mL of saline vehicle and injected bilaterally into the bulbospongiosus muscle by the percutaneous route. Beginning 2 days after treatment, sexual behaviors were reexamined over the course of the following week on four separate occasions. MAIN OUTCOME MEASURES: The latency to achieve ejaculation, and the frequencies and latencies of mounts and intromissions were video recorded by trained observers in a blinded fashion. RESULTS: Relative to pretreatment measurements, bilateral injection of saline vehicle into the bulbospongiosus muscle did not affect ejaculation latencies. However, rats treated with either 0.5 or 1 unit of botulinum-A toxin exhibited significantly longer latencies to achieve ejaculation relative to pretreatment performance. Of note, botulinum-A toxin did not affect the ability to achieve mounts, intromissions, or ejaculation. CONCLUSIONS: These results demonstrate that botulinum-A toxin injection into the bulbospongiosus muscle is a safe and effective treatment that extends ejaculatory latency in rats without affecting the ability to engage in sexual activity or achieve ejaculation. Further studies are required to evaluate this therapeutic concept in PE patients.

A critical analysis of the surgical outcomes for the treatment of Peyronie's disease.

Peyronie's disease (PD) is a relatively common condition, which can impair sexual function and result in emotional and psychological distress. Despite an abundance of minimally invasive treatments, few have confirmed efficacy for improving penile curvature and function. Surgical therapies include many different techniques and are reserved for patients with stable disease of ⩾12 months' duration. We searched PubMed for all articles from 1990 to the present relating to the surgical management of PD. Preference was given to recent articles, larger series, and those comparing various techniques and/or materials. Outcomes were subsequently analysed and organised by surgical technique and the graft material used. Available surgical techniques include plication/corporoplasty procedures, incision and grafting (I&G), and placing a penile prosthesis with or without adjunctive procedures. Although several surgical algorithms have been reported, in general, plication/corporoplasty procedures are reserved for patients with adequate erectile function, simple curvatures of <60°, and with no deformities (hour-glass, hinge). I&G are reserved for complex curvatures of >60° and those with deformities. Penile prostheses are indicated for combined erectile dysfunction and PD. Overall outcomes show high rates of improved curvature and patient satisfaction, with mildly decreased erectile function with both plication and the I&G procedure (I&G >plication) and decreases in penile length (plication >I&G). Surgical management of PD remains an excellent treatment option for patients with penile curvature precluding or impairing sexual activity. Surgical algorithms are available to assist treating clinicians in appropriately stratifying surgical candidates. Additional research is needed to identify optimal surgical techniques and materials based on patient and disease characteristics.