Potentially premalignant oral epithelial lesions (PPOELs) are a group of clinically suspicious conditions, of which a small percentage will undergo malignant transformation. PPOELs are suboptimally ...diagnosed and managed under the current standard of care. Dysplasia is the most well-established marker to distinguish high-risk PPOELs from low-risk PPOELs, and performing a biopsy to establish dysplasia is the diagnostic gold standard. However, a biopsy is limited by morbidity, resource requirements, and the potential for underdiagnosis. Diagnostic adjuncts may help clinicians better evaluate PPOELs before definitive biopsy, but existing adjuncts, such as toluidine blue, acetowhitening, and autofluorescence imaging, have poor accuracy and are not generally recommended. Recently, in vivo microscopy technologies, such as high-resolution microendoscopy, optical coherence tomography, reflectance confocal microscopy, and multiphoton imaging, have shown promise for improving PPOEL patient care. These technologies allow clinicians to visualize many of the same microscopic features used for histopathologic assessment at the point of care.
Histopathology plays a critical role in the diagnosis and surgical management of cancer. However, access to histopathology services, especially frozen section pathology during surgery, is limited in ...resource-constrained settings because preparing slides from resected tissue is time-consuming, labor-intensive, and requires expensive infrastructure. Here, we report a deep-learning-enabled microscope, named DeepDOF-SE, to rapidly scan intact tissue at cellular resolution without the need for physical sectioning. Three key features jointly make DeepDOF-SE practical. First, tissue specimens are stained directly with inexpensive vital fluorescent dyes and optically sectioned with ultra-violet excitation that localizes fluorescent emission to a thin surface layer. Second, a deep-learning algorithm extends the depth-of-field, allowing rapid acquisition of in-focus images from large areas of tissue even when the tissue surface is highly irregular. Finally, a semi-supervised generative adversarial network virtually stains DeepDOF-SE fluorescence images with hematoxylin-and-eosin appearance, facilitating image interpretation by pathologists without significant additional training. We developed the DeepDOF-SE platform using a data-driven approach and validated its performance by imaging surgical resections of suspected oral tumors. Our results show that DeepDOF-SE provides histological information of diagnostic importance, offering a rapid and affordable slide-free histology platform for intraoperative tumor margin assessment and in low-resource settings.
Microscopic evaluation of resected tissue plays a central role in the surgical management of cancer. Because optical microscopes have a limited depth-of-field (DOF), resected tissue is either frozen ...or preserved with chemical fixatives, sliced into thin sections placed on microscope slides, stained, and imaged to determine whether surgical margins are free of tumor cells—a costly and time- and labor-intensive procedure. Here, we introduce a deep-learning extended DOF (DeepDOF) microscope to quickly image large areas of freshly resected tissue to provide histologic-quality images of surgical margins without physical sectioning. The DeepDOF microscope consists of a conventional fluorescence microscope with the simple addition of an inexpensive (less than $10) phase mask inserted in the pupil plane to encode the light field and enhance the depth-invariance of the point-spread function. When used with a jointly optimized image-reconstruction algorithm, diffraction-limited optical performance to resolve subcellular features can be maintained while significantly extending the DOF (200 μm). Data from resected oral surgical specimens show that the DeepDOF microscope can consistently visualize nuclear morphology and other important diagnostic features across highly irregular resected tissue surfaces without serial refocusing. With the capability to quickly scan intact samples with subcellular detail, the DeepDOF microscope can improve tissue sampling during intraoperative tumor-margin assessment, while offering an affordable tool to provide histological information from resected tissue specimens in resource-limited settings.
Abstract Microparticle-based vaccine delivery systems are known to promote enhanced immune responses to protein antigens and can elicit TH 1-biased responses when used in combination with Toll-like ...receptor (TLR) agonists. It is important to understand the kinetics of the immune responses to microparticle-based protein vaccines in order to predict the duration of protective immunity and to optimize prime-boost vaccination regimens. We carried out a 10-week time course study to investigate the magnitude and kinetics of the antibody and cellular immune responses to poly(lactic- co -glycolic acid) (PLGA) microparticles containing 40 μg ovalbumin (OVA) protein and 16 μg CpG-ODN adjuvant (MP/OVA/CpG) in comparison to OVA-containing microparticles, soluble OVA plus CpG, or OVA formulated with Alhydrogel® aluminum adjuvant. Mice vaccinated with MP/OVA/CpG developed the highest TH 1-associated IgG2b and IgG2c antibody titers, while also eliciting TH 2-associated IgG1 antibody titers on par with Alhydrogel® -formulated OVA, with all IgG subtype titers peaking at day 56. The MP/OVA/CpG vaccine also induced the highest antigen-specific splenocyte IFN-γ responses, with high levels of IFN-γ responses persisting until day 42. Thus the MP/OVA/CpG formulation produced a sustained and heightened humoral and cellular immune response, with an overall TH 1 bias, while maintaining high levels of IgG1 antibody equivalent to that seen with Alhydrogel® adjuvant. The time course kinetics study provides a useful baseline for designing vaccination regimens for microparticle-based protein vaccines.
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•It is a challenge to assess risk of progression of oral lesions with mild dysplasia.•Optical imaging can aid in early detection of oral lesions with severe dysplasia.•We compared ...optical markers and gene expression for mild and severe oral dysplasia.•Optical marker positive mild dysplasia is genetically similar to severe dysplasia.
Optical imaging studies of oral premalignant lesions have shown that optical markers, including loss of autofluorescence and altered morphology of epithelial cell nuclei, are predictive of high-grade pathology. While these optical markers are consistently positive in lesions with moderate/severe dysplasia or cancer, they are positive only in a subset of lesions with mild dysplasia. This study compared the gene expression profiles of lesions with mild dysplasia (stratified by optical marker status) to lesions with severe dysplasia and without dysplasia.
Forty oral lesions imaged in patients undergoing oral surgery were analyzed: nine without dysplasia, nine with severe dysplasia, and 22 with mild dysplasia. Samples were submitted for high throughput gene expression analysis.
The analysis revealed 116 genes differentially expressed among sites without dysplasia and sites with severe dysplasia; 50 were correlated with an optical marker quantifying altered nuclear morphology. Ten of 11 sites with mild dysplasia and positive optical markers (91%) had gene expression similar to sites with severe dysplasia. Nine of 11 sites with mild dysplasia and negative optical markers (82%) had similar gene expression as sites without dysplasia.
This study suggests that optical imaging may help identify patients with mild dysplasia who require more intensive clinical follow-up. If validated, this would represent a significant advance in patient care for patients with oral premalignant lesions.
Summary
Oral cancer causes significant global mortality and has a five‐year survival rate of around 64%. Poor prognosis results from late‐stage diagnosis, highlighting an important need to develop ...better approaches to detect oral premalignant lesions (OPLs) and identify which OPLs are at highest risk of progression to oral squamous cell carcinoma (OSCC). An appropriate animal model that reflects the genetic, histologic, immunologic, molecular and gross visual features of human OSCC would aid in the development and evaluation of early detection and risk assessment strategies. Here, we present an experimental PIK3CA + 4NQO transgenic mouse model of oral carcinogenesis that combines the PIK3CA oncogene mutation with oral exposure to the chemical carcinogen 4NQO, an alternate experimental transgenic mouse model with PIK3CA as well as E6 and E7 mutations, and an existing wild‐type mouse model based on oral exposure to 4NQO alone. We compare changes in dorsal and ventral tongue gross visual appearance, histologic features and molecular biomarker expression over a time course of carcinogenesis. Both transgenic models exhibit cytological and architectural features of dysplasia that mimic human disease and exhibit slightly increased staining for Ki‐67, a cell proliferation marker. The PIK3CA + 4NQO model additionally exhibits consistent lymphocytic infiltration, presents with prominent dorsal and ventral tongue tumours, and develops cancer quickly relative to the other models. Thus, the PIK3CA + 4NQO model recapitulates the multistep genetic model of human oral carcinogenesis and host immune response in carcinogen‐induced tongue cancer, making it a useful resource for future OSCC studies.
PURPOSEThe purpose of this study was to report the clinical and refractive outcomes of eyes with long axial length (AL) and high myopia that underwent cataract surgery and compare the performance of ...intraocular lens (IOL) calculation formulae on these eyes. MATERIALS AND METHODSThis retrospective cohort included 183 eyes that underwent cataract surgery from January 2010 to December 2018. Demographics, AL, postoperative best-visual acuities, IOL power data, and postoperative complications were recorded. Refractive outcomes were analyzed and absolute predicted errors were compared between five IOL calculation formulas. RESULTSThe mean age included in the study was 65.4 ± 9.39 years with a mean AL of 26.76 ± 1.75 mm. Postoperatively, the mean sphere, cylinder, and manifest refraction spherical equivalent were 0.22 D ± 0.54, -0.78 D ± 0.50, and - 0.16 D ± 0.50, respectively. The average IOL power implanted was 11.12 D ± 4.59 D. No intraoperative complications were encountered, but there was one incidence of retinal tear with detachment reported postoperatively (0.55%). The Kane formula had the lowest mean absolute predicted error (MAE). A significant positive correlation between increasing AL and MAE was seen in the Sanders, Retzlaff and Kraft-Theoretical (SRK-T) and Ladas formulae but not statistically significant when the Kane, Barrett Universal II, and the Emmetropia Verifying Optical (EVO) formulae were used. CONCLUSIONCataract surgery in eyes with long ALs and high myopia is safe with a low incidence of intraoperative and postoperative complications. The Kane, Barrett, and EVO formulae were equally accurate in calculating the IOL power and achieved the least amount of residual error postoperatively.
Animal physiology, ecology and evolution are affected by temperature and it is expected that community structure will be strongly influenced by global warming. This is particularly relevant in the ...tropics, where organisms are already living close to their upper temperature limits and hence are highly vulnerable to rising temperature. Here we present data on upper temperature limits of 34 tropical marine ectotherm species from seven phyla living in intertidal and subtidal habitats. Short term thermal tolerances and vertical distributions were correlated, i.e., upper shore animals have higher thermal tolerance than lower shore and subtidal animals; however, animals, despite their respective tidal height, were susceptible to the same temperature in the long term. When temperatures were raised by 1°C hour(-1), the upper lethal temperature range of intertidal ectotherms was 41-52°C, but this range was narrower and reduced to 37-41°C in subtidal animals. The rate of temperature change, however, affected intertidal and subtidal animals differently. In chronic heating experiments when temperature was raised weekly or monthly instead of every hour, upper temperature limits of subtidal species decreased from 40°C to 35.4°C, while the decrease was more than 10°C in high shore organisms. Hence in the long term, activity and survival of tropical marine organisms could be compromised just 2-3°C above present seawater temperatures. Differences between animals from environments that experience different levels of temperature variability suggest that the physiological mechanisms underlying thermal sensitivity may vary at different rates of warming.
Chlamydia trachomatis is the most common cause of bacterial sexually transmitted infection. It produces an unusual intracellular infection in which a vegetative form, called the reticulate body (RB), ...replicates and then converts into an elementary body (EB), which is the infectious form. Here we use quantitative three-dimensional electron microscopy (3D EM) to show that C. trachomatis RBs divide by binary fission and undergo a sixfold reduction in size as the population expands. Conversion only occurs after at least six rounds of replication, and correlates with smaller RB size. These results suggest that RBs only convert into EBs below a size threshold, reached by repeatedly dividing before doubling in size. A stochastic mathematical model shows how replication-dependent RB size reduction produces delayed and asynchronous conversion, which are hallmarks of the Chlamydia developmental cycle. Our findings support a model in which RB size controls the timing of RB-to-EB conversion without the need for an external signal.
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