Dystrophin myonuclear domain restoration governs treatment efficacy in dystrophic muscle.

Dystrophin is essential for muscle health: its sarcolemmal absence causes the fatal, X-linked condition, Duchenne muscular dystrophy (DMD). However, its normal, spatial organization remains poorly understood, which hinders the interpretation of efficacy of its therapeutic restoration. Using female reporter mice heterozygous for fluorescently tagged dystrophin (DmdEGFP), we here reveal that dystrophin distribution is unexpectedly compartmentalized, being restricted to myonuclear-defined sarcolemmal territories extending ~80 µm, which we called "basal sarcolemmal dystrophin units (BSDUs)." These territories were further specialized at myotendinous junctions, where both Dmd transcripts and dystrophin protein were enriched. Genome-level correction in X-linked muscular dystrophy mice via CRISPR/Cas9 gene editing restored a mosaic of separated dystrophin domains, whereas transcript-level Dmd correction, following treatment with tricyclo-DNA antisense oligonucleotides, restored dystrophin initially at junctions before extending along the entire fiber-with levels ~2% sufficient to moderate the dystrophic process. We conclude that widespread restoration of fiber dystrophin is likely critical for therapeutic success in DMD, perhaps most importantly, at muscle-tendon junctions.

Axillary reverse mapping using near-infrared fluorescence imaging in invasive breast cancer (ARMONIC study).

BACKGROUND: Axillary lymph node dissection (ALND) in patients with breast cancer has potential side effects, including upper-limb lymphedema. Axillary reverse mapping (ARM) is a technique that enables discrimination of the lymphatic drainage of the upper limb in the axillary lymph node basin from that of the breast. We aimed to evaluate ARM node identification by near-infrared (NIR) fluorescence imaging during total mastectomy with ALND and then to analyze potential predictive factors of ARM node involvement. METHODS: The study enrolled 119 patients diagnosed with invasive breast cancer with an indication for ALND. NIR imaging using indocyanine green dye was performed in 109 patients during standard ALND to identify ARM nodes and their corresponding lymphatic ducts. RESULTS: 94.5% of patients had ARM nodes identified (95%CI = [88.4-98.0]). The ARM nodes were localized in zone D in 63.4% of cases. Metastatic axillary lymph nodes were found in 55% in the whole cohort, and 19.4% also had metastasis in ARM nodes. Two patients had metastatic ARM nodes but not in the remaining axillary lymph nodes. No serious adverse events were observed. Only the amount of mitosis was significantly associated with ARM node metastasis. CONCLUSIONS: ARM by NIR fluorescence imaging could be a reliable technique to identify ARM nodes in real-time when ALND is performed. The clinical data compared with ARM node histological diagnosis showed only the amount of mitosis in the diagnostic biopsy is a potential predictive factor of ARM node involvement. CLINICAL TRIAL REGISTRATION: NCT02994225.

Near-Infrared Fluorescence Axillary Reverse Mapping (ARM) Procedure in Invasive Breast Cancer: Relationship between Fluorescence Signal in ARM Lymph Nodes and Clinical Outcomes.

The near-infrared (NIR) fluorescence axillary reverse mapping (ARM) procedure is a promising tool to identify and preserve arm lymphatic drainage during axillary lymph node dissection (ALND). The ARMONIC clinical trial was conducted to validate the technique on a large cohort of patients and to analyze the predictive clinical factors for ARM lymph node metastasis. For the first time, the fluorescence signal intensity from the ARM lymph nodes was measured and correlated with clinical findings. A total of 109 patients with invasive breast cancer and indications of mastectomy and ALND underwent the NIR fluorescence ARM procedure. Indocyanine green was administered by intradermal injection followed by intraoperative identification and resection of the ARM lymph nodes with NIR fluorescence camera guidance. The fluorescence signal intensity and signal distribution were then measured ex vivo and compared with clinical outcomes. ARM lymph nodes were successfully identified by fluorescence in 94.5% of cases. The mean normalized fluorescence signal intensity value was 0.47 with no significant signal difference between metastatic and non-metastatic ARM lymph nodes (p = 0.3728). At the microscopic level, the fluorescence signal distribution was focally intense in lymphoid tissue areas. Only the preoperative diagnosis of metastasis in the axillary nodes of patients was significantly associated with a higher ARM node fluorescence signal intensity (p = 0.0253), though it was not significantly associated with the pathological nodal (pN) status (p = 0.8081). Based on an optimal cut-off fluorescence value, the final sensitivity and specificity of the NIR fluorescence ARM procedure for ARM lymph node metastatic involvement were 64.7% and 47.3%, respectively. Although our preliminary results did not show that fluorescence signal intensity is a reliable diagnostic tool, the NIR fluorescence ARM procedure may be useful for ARM lymph node identification. Clinical trial registration: NCT02994225.

Confocal laser endomicroscopy and confocal microscopy for head and neck cancer imaging: Recent updates and future perspectives.

Confocal laser endomicroscopy (CLE) is a promising technology that provides real-time histological visualization of living tissue, described as "optical biopsy". It has been used successfully in dermatology, gastroenterology and ophthalmology for clinical diagnosis and tumor margin assessment. In addition to in vivo tissue assessment, confocal microscopy can also be used ex vivo on freshly excised or fixed tissue for intraoperative tumor margin assessment. While CLE and ex vivo confocal microscopy (EVCM) have shown promising results for the identification of head and neck cancers at cell level, several methodological issues need to be addressed before they can be widely used in routine clinical use. The present review will focus on recent advances in CLE and EVCM in the diagnosis and management of head and neck cancer and discuss remaining challenges for their clinical application.

Diagnostic accuracy of in vivo early tumor imaging from probe-based confocal laser endomicroscopy versus histologic examination in head and neck squamous cell carcinoma.

OBJECTIVES: Probe-based confocal laser endomicroscopy (pCLE) is a noninvasive and real-time imaging technique allowing acquisition of in situ images of the tissue microarchitecture during oral surgery. We aimed to assess the diagnostic performance of pCLE combined with patent blue V (PB) in improving the management of early oral cavity, oro/hypopharyngeal, and laryngeal cancer by imaging squamous cell carcinoma in vivo. MATERIALS AND METHODS: The prospective study enrolled 44 patients with early head and neck lesions. All patients underwent white-light inspection or panendoscopy depending on the lesion's location, followed by pCLE imaging of the tumor core and its margins after topical application of PB. Each zone imaged by pCLE was interpreted at distance of the exam by three pathologists blinded to final histology. RESULTS: Most imaged zones could be presented to pathologists; the final sensitivity and specificity of pCLE imaging in head and neck cancers was 73.2-75% and 30-57.4%, respectively. During imaging, head and neck surgeons encountered some challenges that required resolving, such as accessing lesions with the flexible optical probe, achieving sufficiently precise imaging on the targeted tissues, and heterogeneous tissue staining by fluorescent dye. CONCLUSION: Final sensitivity scores were reasonable but final specificity scores were low. pCLE zones used to calculate specificity were acquired in areas of tumor margins, and the poor quality of the images acquired in these areas explains the final low specificity scores. CLINICAL RELEVANCE: Practical adjustments and technical training are needed to analyze head and neck lesions in various anatomical sites in real-time by pCLE.

Cord blood-endothelial colony forming cells are immunotolerated and participate at post-ischemic angiogenesis in an original dorsal chamber immunocompetent mouse model.

BACKGROUND: Cardiovascular diseases are the main cause of morbidity and mortality worldwide. Restoring blood supply to ischemic tissues is an essential goal for the successful treatment of these diseases. Growth factor or gene therapy efficacy remains controversial, but stem cell transplantation is emerging as an interesting approach to stimulate angiogenesis. Among the different stem cell populations, cord blood-endothelial progenitor cells (CB-EPCs) and more particularly cord blood-endothelial progenitor cell-derived endothelial colony forming cells (CB-ECFCs) have a great proliferative potential without exhibiting signs of senescence. Even if it was already described that CB-ECFCs were able to restore blood perfusion in hind-limb ischemia in an immunodeficient mouse model, until now, the immunogenic potential of allogenic CB-ECFCs remains controversial. Therefore, our objectives were to evaluate the immune tolerance potency of CB-ECFCs and their capacity to restore a functional vascular network under ischemic condition in immunocompetent mice. METHODS: In vitro, the expression and secretion of immunoregulatory markers (HLA-G, IL-10, and TGF-ß1) were evaluated on CB-ECFCs. Moreover, CB-ECFCs were co-cultured with activated peripheral blood mononuclear cells (PBMCs) for 6 days. PBMC proliferation was evaluated by [3H]-thymidine incorporation on the last 18 h. In vivo, CB-ECFCs were administered in the spleen and muscle of immunocompetent mice. Tissues were collected at day 14 after surgery. Finally, CB-ECFCs were injected intradermally in C57BL/6JRj mice close to ischemic macrovessel induced by thermal cauterization. Mice recovered until day 5 and were imaged, twice a week until day 30. RESULTS: Firstly, we demonstrated that CB-ECFCs expressed HLA-G, IL-10, and TGF-ß1 and secreted IL-10 and TGF-ß1 and that they could display immunosuppressive properties in vitro. Secondly, we showed that CB-ECFCs could be tolerated until 14 days in immunocompetent mice. Thirdly, we revealed in an original ischemic model of dorsal chamber that CB-ECFCs were integrated in a new functional vascular network. CONCLUSION: These results open up new perspectives about using CB-ECFCs as an allogeneic cell therapy product and gives new impulse to the treatment of cardiovascular diseases.

Value of Full-Field Optical Coherence Tomography Imaging for the Histological Assessment of Head and Neck Cancer.

BACKGROUND AND OBJECTIVES: In head and neck surgery, intraoperative and postoperative evaluation of tumor margins is achieved by histopathological assessment, which is a multistep process. Intraoperative analysis of tumor margins to obtain a preliminary diagnosis is usually carried out on frozen sections. Analysis of frozen sections is challenging due to technical difficulties in processing. Full-field optical coherence tomography (FFOCT) provides ex vivo images of fresh tissue samples at a microscopic scale without tissue processing. The objectives of our study were to define the diagnostic criteria required to interpret head and neck FFOCT images and to evaluate the reliability of a histological diagnosis made on an "optical biopsy" produced by head and neck FFOCT imaging compared with conventional histology. STUDY DESIGN/MATERIALS AND METHODS: First, we established an atlas of comparative images (FFOCT/standard histology) and defined the diagnostic criteria based on FFOCT images. Two pathologists subsequently performed a blinded review on 57 FFOCT images (32 patients). Specificity and sensitivity were measured by comparison with the standard histological diagnosis. The primary endpoint was major concordance, defined as two classifications leading to the same therapeutic decision (treatment/no treatment). RESULTS: Pathologists identified four main criteria for tissue diagnosis on FFOCT images: heterogeneous cell distribution, stromal reaction, coiling, and keratinization abnormalities. The correlation study showed good results, with sensitivity from 88% to 90% and specificity from 81% to 87%, regardless of whether the FFOCT image review was performed by a pathologist with or without previous experience in optical imaging. CONCLUSIONS: Our results demonstrate that FFOCT images can be used by pathologists for differential diagnosis, and that high-resolution FFOCT imaging can provide an assessment of microscopic architecture in head and neck tissues without tissue processing requirements. Lasers Surg. Med. © 2020 Wiley Periodicals, Inc.

Multimodal imaging for tumour characterization from micro- to macroscopic level using a newly developed dorsal chamber designed for long-term follow-up.

Optical imaging of living animals is a unique method of studying the dynamics of physiological and pathological processes at a subcellular level. One-shot acquisitions at high resolution can be achieved on exteriorized organs before animal euthanasia. For longitudinal follow-up, intravital imaging can be used and involves imaging windows implanted in cranial, thoracic or dorsal regions. Several imaging window models exist, but none have proven to be applicable for long-term monitoring and most biological processes take place over several weeks. Moreover, none are compatible with multiple imaging modalities, meaning that different biological parameters cannot be assessed in an individual animal. We developed a new dorsal chamber that was well tolerated by mice (over several months) and allowed individual and collective cell tracking and behaviour analysis by optical imaging, ultrasound and magnetic resonance tomography. This new model broadens potential applications to areas requiring study of long-term biological processes, as in cancer research.

An Accessible and Unique Insight into Metastasis Mutational Content Through Whole-exome Sequencing of Circulating Tumor Cells in Metastatic Prostate Cancer.

BACKGROUND: Genomic analysis of circulating tumor cells (CTCs) could provide a unique and accessible representation of tumor diversity but remains hindered by technical challenges associated with CTC rarity and heterogeneity. OBJECTIVE: To evaluate CTCs as surrogate samples for genomic analyses in metastatic castration-resistant prostate cancer (mCRPC). DESIGN, SETTING, AND PARTICIPANTS: Three isolation strategies (filter laser-capture microdissection, self-seeding microwell chips, and fluorescence-activated cell sorting) were developed to capture CTCs with various epithelial and mesenchymal phenotypes and isolate them at the single-cell level. Whole-genome amplification (WGA) and WGA quality control were performed on 179 CTC samples, matched metastasis biopsies, and negative controls from 11 patients. All patients but one were pretreated with enzalutamide or abiraterone. Whole-exome sequencing (WES) of 34 CTC samples, metastasis biopsies, and negative controls were performed for seven patients. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: WES of CTCs was rigorously qualified in terms of percentage coverage at 10× depth, allelic dropout, and uncovered regions. Shared somatic mutations between CTCs and matched metastasis biopsies were identified. A customized approach based on determination of mutation rates for CTC samples was developed for identification of CTC-exclusive mutations. RESULTS AND LIMITATIONS: Shared mutations were mostly detected in epithelial CTCs and were recurrent. For two patients for whom a deeper analysis was performed, a few CTCs were sufficient to represent half to one-third of the mutations in the matched metastasis biopsy. CTC-exclusive mutations were identified in both epithelial and nonepithelial CTCs and affected cytoskeleton, invasion, DNA repair, and cancer-driver genes. Some 41% of CTC-exclusive mutations had a predicted deleterious impact on protein function. Phylogenic relationships between CTCs with distinct phenotypes were evidenced. CONCLUSIONS: CTCs can provide unique insight into metastasis mutational diversity and reveal undiagnosed genomic aberrations in matched metastasis biopsies. PATIENT SUMMARY: Our results demonstrate the clinical potential of circulating tumor cells to provide insight into metastatic events that could be critical to target using precision medicine.

Near-infrared fluorescence imaging for the prevention and management of breast cancer-related lymphedema: A systematic review.

Sentinel lymph node identification by near infrared (NIR) fluorescence with indocyanine green (ICG) is recognized in the literature as a useful technique. NIR fluorescence technology could become key in the prevention and management of lymphedema after axillary dissection for breast cancer. Here, we conducted a systematic review focusing on ICG imaging to improve lymphedema prevention and treatment after axillary surgery. A systematic literature review was performed using MEDLINE and Embase to identify articles focused on ICG imaging for breast-cancer-related lymphedema (BCRL). Qualitative analysis was performed to summarize the characteristics of reported ICG procedures. In situ tissue identification and functionality assessment based on fluorescence signal were evaluated. Clinical outcomes were appraised when reported. Studies relating to axillary reverse mapping, lymphography and upper limb supermicrosurgery combined with ICG imaging were identified. We included a total of 33 relevant articles with a total of 2016 patients enrolled. ICG imaging for axillary reverse mapping was safe for all 951 included patients, with identification of arm nodes in 80%-88% of patients with axillary lymph nodes dissection. However, the papers discuss the oncologic safety of the approach and how - regardless of the contrast agent - concerns limit its adoption. ICG lymphography is openly supported in BCRL management, with 1065 patients undergoing this procedure in 26 articles. The technique is reported for lymphedema diagnosis, with high sensitivity and specificity, staging, intraoperative mapping and patency control in lymphaticovenular anastomosis. The substantial advantages/disadvantages of ICG imaging procedures are finally described.

Parathyroid gland management using optical technologies during thyroidectomy or parathyroidectomy: A systematic review.

New optical technologies enhancing localization or assessing viability of parathyroid glands (PG) during endocrine surgery have been reported in clinical studies. These technologies could become complementary to the surgeon's eyes and may improve surgical outcomes in thyroidectomy and parathyroidectomy. Here, we conducted a systematic review focusing on PG identification and functional assessment using optical methods to enhance surgery. A systematic literature review was performed using MEDLINE and Embase database. Two authors selected studies and extracted data; qualitative analysis was performed to summarize the characteristics of reported optical tools for thyroidectomy or parathyroidectomy. Identification and vascularisation of PG during surgery were evaluated. Clinical and biochemical outcomes were appraised when reported. Studies relating to parathyroidectomy or thyroidectomy combined with autofluorescence, fluorescent methylene blue, 5-aminolevulinic acid, indocyanine green (ICG), optical coherence tomography, laser speckle contrast imaging, dynamic optical contrast imaging and Raman spectroscopy were identified with MEDLINE and Embase. We included a total of 47 relevant articles with a total of 1615 patients enrolled. Each optical technique is described and appreciated related to its surgical purpose. Autofluorescence and ICG imaging of PG are the most widely reported optical technologies for identification and assessment of vascularisation of PG. Results are mainly based on observational studies and argue for the feasibility of both techniques in endocrine surgery but prospective randomized studies have not been performed. In vivo applications are still limited for the other methods and further investigations correlating these techniques with post-operative parathormone measurements are still needed before considering these technologies in clinical practice.

Prospective evaluation of the limitations of near-infrared imaging in detecting axillary sentinel lymph nodes in primary breast cancer.

We compared the performance of near-infrared imaging using indocyanine green (ICG) with the radioisotope (ISO) method to detect sentinel lymph nodes (SLNs) in breast cancer, to analyze predictive factors for negative ICG identification. The study included 122 patients who underwent sentinel lymph node biopsy (SLNB) using the combined ISO and ICG technique for primary breast cancer. We assessed the putative association between pathologic/clinical variables and ICG failure to detect SLNs. The ISO identification rate was 96.7% and ICG identification 81.9%. Overweight patients or presence of macrometastasis in SLNB were associated with the risk of ICG failing to detect SLNs (P = 0.02).

Method for semi-automated microscopy of filtration-enriched circulating tumor cells.

BACKGROUND: Circulating tumor cell (CTC)-filtration methods capture high numbers of CTCs in non-small-cell lung cancer (NSCLC) and metastatic prostate cancer (mPCa) patients, and hold promise as a non-invasive technique for treatment selection and disease monitoring. However filters have drawbacks that make the automation of microscopy challenging. We report the semi-automated microscopy method we developed to analyze filtration-enriched CTCs from NSCLC and mPCa patients. METHODS: Spiked cell lines in normal blood and CTCs were enriched by ISET (isolation by size of epithelial tumor cells). Fluorescent staining was carried out using epithelial (pan-cytokeratins, EpCAM), mesenchymal (vimentin, N-cadherin), leukocyte (CD45) markers and DAPI. Cytomorphological staining was carried out with Mayer-Hemalun or Diff-Quik. ALK-, ROS1-, ERG-rearrangement were detected by filter-adapted-FISH (FA-FISH). Microscopy was carried out using an Ariol scanner. RESULTS: Two combined assays were developed. The first assay sequentially combined four-color fluorescent staining, scanning, automated selection of CD45(-) cells, cytomorphological staining, then scanning and analysis of CD45(-) cell phenotypical and cytomorphological characteristics. CD45(-) cell selection was based on DAPI and CD45 intensity, and a nuclear area >55 µm(2). The second assay sequentially combined fluorescent staining, automated selection of CD45(-) cells, FISH scanning on CD45(-) cells, then analysis of CD45(-) cell FISH signals. Specific scanning parameters were developed to deal with the uneven surface of filters and CTC characteristics. Thirty z-stacks spaced 0.6 µm apart were defined as the optimal setting, scanning 82 %, 91 %, and 95 % of CTCs in ALK-, ROS1-, and ERG-rearranged patients respectively. A multi-exposure protocol consisting of three separate exposure times for green and red fluorochromes was optimized to analyze the intensity, size and thickness of FISH signals. CONCLUSIONS: The semi-automated microscopy method reported here increases the feasibility and reliability of filtration-enriched CTC assays and can help progress towards their validation and translation to the clinic.

Intraoperative Near-infrared Imaging for Parathyroid Gland Identification by Auto-fluorescence: A Feasibility Study.

BACKGROUND: Parathyroid glands (PGs) can be particularly hard to distinguish from surrounding tissue and thus can be damaged or removed during thyroidectomy. Postoperative hypoparathyroidism is the most common complication after thyroidectomy. Very recently, it has been found that the parathyroid tissue shows near-infrared (NIR) auto-fluorescence which could be used for intraoperative detection, without any use of contrast agents. The work described here presents a histological validation ex vivo of the NIR imaging procedure and evaluates intraoperative PG detection by NIR auto-fluorescence using for the first time to our knowledge a commercially available clinical NIR imaging device. METHODS: Ex vivo study on resected operative specimens combined with a prospective in vivo study of consecutive patients who underwent total or partial thyroid, or parathyroid surgery at a comprehensive cancer center. During surgery, any tissue suspected to be a potential PG by the surgeon was imaged with the Fluobeam 800 (®) system. NIR imaging was compared to conventional histology (ex vivo) and/or visual identification by the surgeon (in vivo). RESULTS: We have validated NIR auto-fluorescence with an ex vivo study including 28 specimens. Sensitivity and specificity were 94.1 and 80 %, respectively. Intraoperative NIR imaging was performed in 35 patients and 81 parathyroids were identified. In 80/81 cases, the fluorescence signal was subjectively obvious on real-time visualization. We determined that PG fluorescence is 2.93 ± 1.59 times greater than thyroid fluorescence in vivo. CONCLUSIONS: Real-time NIR imaging based on parathyroid auto-fluorescence is fast, safe, and non-invasive and shows very encouraging results, for intraoperative parathyroid identification.

Patent blue V and indocyanine green for fluorescence microimaging of human peritoneal carcinomatosis using probe-based confocal laser endomicroscopy.

BACKGROUND: Peritoneal carcinomatosis is a metastatic stage aggravating abdominal and pelvic cancer dissemination. The preoperative evaluation of lesions remains difficult today. Probe-based confocal laser endomicroscopy (pCLE) provides dynamic images of tissue architecture and cellular details. This technology allows in vivo histological interpretation of tissue. The main limitation of pCLE for adoption in the clinic is the unavailability of fluorescent contrast agents. The aim of our study was to evaluate the staining performance of indocyanine green and patent blue V for histological diagnosis of pCLE images of pathological and non-pathological peritoneal tissue. METHODS: We performed a correlative study with the histological gold standard on ex vivo human specimens from 25 patients operated for peritoneal carcinomatosis; 70 specimens were stained by topical application with ICG or patent blue V and then imaged with a probe-based confocal laser endomicroscope. A total of 350 pCLE images and 70 corresponding histological sections were randomly and blindly interpreted by two pathologists (PT1 and PT2). The images were first classified into two categories, tumoral versus non-tumoral, and a refined histological diagnosis was then given. RESULTS: All presented images were interpreted by PT1 (who received prior training on PCLE image reading) and PT2 (no training). 100 % sensitivity for PT1 and PT2 was noticed with tissues stained with ICG to differentiate tumoral and non-tumoral tissue. Global scores were always better for PT1 (major concordance between 86 and 94 %) than for PT2 (major concordance between 77 and 89 %) independently of the fluorescent dye when histological diagnosis was done on pCLE images. CONCLUSION: In conclusion, the pair ICG-pCLE offers the best combination for a non-trained pathologist for the interpretation of pCLE images from peritoneum.

Polo-like Kinase Inhibitor Volasertib Exhibits Antitumor Activity and Synergy with Vincristine in Pediatric Malignancies.

BACKGROUND: Polo-like kinase 1 (PLK1) controls the main cell-cycle checkpoints, suggesting utility of its inhibition for cancer treatment, including of highly proliferative pediatric cancer. This preclinical study explored the selective PLK1 inhibitor volasertib (BI 6727) alone and combined with chemotherapy in pediatric malignancies. MATERIALS AND METHODS: Inhibition of proliferation was explored in vitro using dimethylthiazol carboxymethoxyphenyl sulfophenyl tetrazolium (MTS) assay. Mice bearing human xenografts were treated with weekly intravenous injections of volasertib. RESULTS: Volasertib inhibited proliferation in all 40 cell lines tested, with a mean half-maximal growth inhibitory concentration of 313 nmol/l (range: 4-5000 nmol/l). Volasertib was highly active against RMS-1 alveolar rhabdomyosarcoma xenografts, resulting in 100% tumor regression. Activity was associated with complete and prolonged G2/M arrest and subsequent apoptotic cell death. Volasertib showed synergistic activity with vincristine but antagonistic effects with etoposide. CONCLUSION: These findings support the further exploration of volasertib for pediatric malignancies, particularly alveolar rhabdomyosarcoma, and its combination with mitotic spindle poison.

Red and far-red fluorescent dyes for the characterization of head and neck cancer at the cellular level.

BACKGROUND: Primary upper aerodigestive tract malignancy remains a cancer having a poor prognosis, despite current progress in treatment, due to a generally late diagnosis. OBJECTIVES: We conducted a preliminary assessment of five dyes approved for human use for the imaging of head and neck tissues at the cellular level, which could be considered for clinical examination. METHODS: We investigated fluorescence endomicroscopic images on fresh samples obtained from head and neck surgeries after staining with hypericin, methylene blue, toluidine blue, patent blue or indocyanine green to provide a preliminary consideration as to whether these images contain enough information for identification of non-pathologic and pathologic tissues. The distribution pattern of dye has been examined using probe-based confocal laser endomicroscopy (pCLE) in ex vivo specimens and compared with corresponding histology. RESULTS: In most samples, the image quality provided by pCLE with both dyes allowed pathologists to recognize histological characteristics to identify the tissues. CONCLUSION: The combination of pCLE imaging with these dyes provides interpretable images close to conventional histology; a promising clinical tool to assist physicians in examination of upper aerodigestive tract, as long as depth imaging issues can be overcome.

Confocal laser endomicroscopy for non-invasive head and neck cancer imaging: a comprehensive review.

Histological assessment is an essential tool in the diagnosis and guidance of the treatment of various diseases, in particular cancer, of the head and neck. Recent major advances in optical imaging techniques have made it possible to acquire high-resolution in vivo images at the cellular scale. Confocal endomicroscopy is a non-invasive technique, which can be highly useful whenever meaningful in situ histological information is required. The technical aspects of confocal endomicroscopy are introduced, followed by an overview of major clinical studies in the field of head and neck cancer. Ongoing technical developments, contributing to improvements in imaging of the upper aero-digestive tract, are also discussed. Finally, the potential complementarities of functional and molecular imaging, as compared to morphological endomicroscopy, are highlighted.