Performance of a deep neural network in teledermatology: a single-centre prospective diagnostic study.

BACKGROUND: The use of artificial intelligence (AI) algorithms for the diagnosis of skin diseases has shown promise in experimental settings but has not been yet tested in real-life conditions. OBJECTIVE: To assess the diagnostic performance and potential clinical utility of a 174-multiclass AI algorithm in a real-life telemedicine setting. METHODS: Prospective, diagnostic accuracy study including consecutive patients who submitted images for teledermatology evaluation. The treating dermatologist chose a single image to upload to a web application during teleconsultation. A follow-up reader study including nine healthcare providers (3 dermatologists, 3 dermatology residents and 3 general practitioners) was performed. RESULTS: A total of 340 cases from 281 patients met study inclusion criteria. The mean (SD) age of patients was 33.7 (17.5) years; 63% (n = 177) were female. Exposure to the AI algorithm results was considered useful in 11.8% of visits (n = 40) and the teledermatologist correctly modified the real-time diagnosis in 0.6% (n = 2) of cases. The overall top-1 accuracy of the algorithm (41.2%) was lower than that of the dermatologists (60.1%), residents (57.8%) and general practitioners (49.3%) (all comparisons P < 0.05, in the reader study). When the analysis was limited to the diagnoses on which the algorithm had been explicitly trained, the balanced top-1 accuracy of the algorithm (47.6%) was comparable to the dermatologists (49.7%) and residents (47.7%) but superior to the general practitioners (39.7%; P = 0.049). Algorithm performance was associated with patient skin type and image quality. CONCLUSIONS: A 174-disease class AI algorithm appears to be a promising tool in the triage and evaluation of lesions with patient-taken photographs via telemedicine.

Nitrosative stress by peroxynitrite impairs ATP production in human spermatozoa.

The most toxic species in live systems include reactive nitrogen species such as peroxynitrite, which at high levels induces nitrosative stress. In human spermatozoa, the negative effect of peroxynitrite on motility and mitochondrial membrane potential was recently demonstrated, and the hypothesis of this work is that impairment of ATP production could be one cause of the effect on motility. Therefore, the aim here was to evaluate ATP production by both glycolysis and oxidative phosphorylation (OXPHOS) in spermatozoa exposed to peroxynitrite in vitro. Human spermatozoa were incubated with SIN-1, a molecule which generates peroxynitrite, and the ATP level was evaluated. Then, to inactivate glycolysis or OXPHOS, spermatozoa were incubated with pharmacological inhibitors of these pathways. Spermatozoa treated for inactivating one or the other pathway were exposed to SIN-1, and the ATP level was compared to the control without SIN-1 in each condition. The ATP level fell after peroxynitrite exposure. The ATP in spermatozoa treated for inactivating one or the other metabolic pathway and subsequently exposed to peroxynitrite was reduced compared with the control. These results show for the first time that an important mechanism by which peroxynitrite reduces sperm function is the inhibition of ATP production, affecting both glycolysis and OXPHOS.

Use of the fluorescent dye tetramethylrhodamine methyl ester perchlorate for mitochondrial membrane potential assessment in human spermatozoa.

Mitochondrial membrane potential (ΔΨm) is an indicator of sperm quality and its evaluation complements the standard semen analysis. The fluorescent dye JC-1 has been widely used to assess sperm ΔΨm; however, some problems have been detected under certain experimental conditions. Another fluorescent compound, tetramethylrhodamine methyl ester perchlorate (TMRM), has been used in somatic cells and bovine spermatozoa but not in human spermatozoa. TMRM accumulates in hyperpolarised mitochondria and the fluorescence intensity of this compound correlates with ΔΨm. Thus, the aim of this study was to evaluate and validate the usefulness of the fluorescent dye TMRM for measuring sperm ΔΨm. The results showed that TMRM accurately detects sperm populations displaying either high or low ΔΨm. Moreover, TMRM was able to measure sperm ΔΨm under the experimental conditions in which JC-1 had previously presented difficulties. Differences in ΔΨm according to sperm and semen quality were properly detected and a positive correlation between ΔΨm and conventional semen parameters was observed. Finally, a positive correlation was found between the ΔΨm measurement by TMRM and by the widely used JC-1. In conclusion, TMRM is a simple, time-effective method, easy to set in laboratories equipped with flow cytometry technology, and can accurately detect changes in ΔΨm with efficiency comparable to JC-1 without its limitations.

Mitochondrial outer membrane permeabilization increases reactive oxygen species production and decreases mean sperm velocity but is not associated with DNA fragmentation in human sperm.

STUDY HYPOTHESIS: Does induction of mitochondrial outer membrane permeabilization (MOMP) in vitro affect specific functional parameters of human spermatozoa? STUDY FINDING: Our findings show that MOMP induction increases intracellular reactive oxygen species (ROS) and decreases mean sperm velocity but does not alter DNA integrity. WHAT IS KNOWN ALREADY: MOMP in somatic cells is related to a variety of apoptotic traits, such as alteration of mitochondrial membrane potential (ΔΨm), and increase in ROS production and DNA fragmentation. Although the presence of these apoptotic features has been reported in spermatozoa, to date the effects of MOMP on sperm function and DNA integrity have not been analysed. STUDY DESIGN, SAMPLES/MATERIALS, METHODS: The study included spermatozoa from fertile donors. Motile sperm were obtained using the swim-up method. The highly motile sperm were collected and diluted with human tubal fluid to a final cell concentration of 5 × 10(6) ml(-1). To induce MOMP, selected sperm were treated at 37°C for 4 h with a mimetic of a Bcl-2 pro-apoptotic protein, ABT-737. MOMP was evaluated by relocating of cytochrome c. In addition, the effect of ABT-737 on mitochondrial inner membrane permeabilization was assessed using the calcein-AM/cobalt chloride method. In turn, ΔΨm was evaluated with JC-1 staining, intracellular ROS production with dihydroethidium, sperm motility was analysed by computer-assisted sperm analysis and DNA fragmentation by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL) assay. Measurements were performed by flow cytometry. MAIN RESULTS AND THE ROLE OF CHANCE: MOMP was associated with ΔΨm dissipation (P < 0.05), increased ROS production (P < 0.05) and decreased mean sperm velocity (P < 0.05), but it was not associated with DNA fragmentation. MOMP did not induce a large increase in ROS, which could explain the negligible effect of MOMP on sperm DNA fragmentation under our experimental conditions. LIMITATIONS, REASONS FOR CAUTION: The study was carried out in vitro using highly motile sperm, selected by swim-up, from healthy donors. WIDER IMPLICATIONS OF THE FINDINGS: The results obtained in this study reveal that the alterations of sperm functions caused by MOMP are sufficiently relevant to justify its future study in male infertility. LARGE SCALE DATA: None. STUDY FUNDING AND COMPETING INTERESTS: The study was funded by grant DI12-0102 from the Universidad de La Frontera (J.V.V.) and a doctoral scholarship from CONICYT (F.T.). The authors declare no conflict of interest.

Peroxynitrite-mediated nitrosative stress decreases motility and mitochondrial membrane potential in human spermatozoa.

Nitrosative stress is produced by high levels of reactive nitrogen species (RNS). The RNS include peroxynitrite, a highly reactive free radical produced from a diffusion-controlled reaction between nitric oxide and superoxide anion. Peroxynitrite causes nitration and oxidation of lipids, proteins and DNA, and is thus considered an important pathogenic mechanism in various diseases. Although high levels of peroxynitrite are associated with astenozoospermia, few reports exist regarding the in vitro effect of high levels of this RNS on human sperm. The aim of this study was to evaluate the in vitro effect of nitrosative stress caused by peroxynitrite on the viability, motility and mitochondrial membrane potential of human spermatozoa. To do this, human spermatozoa from healthy donors were exposed in vitro to 3-morpholinosydnonimine (SIN-1), a molecule that generates peroxynitrite. Incubations were done at 37°C for up to 4 h with SIN-1 concentrations between 0.2 and 1.0 mmol/l. Generation of peroxynitrite was confirmed using dihydrorhodamine 123 (DHR) by spectrophotometry and flow cytometry. Sperm viability was assessed by propidium iodide staining; sperm motility was analyzed by CASA, and the state of mitochondrial membrane potential (ΔΨm) by JC-1 staining. Viability and ΔΨm were measured by flow cytometry. The results showed an increase in DHR oxidation, demonstrating the generation of peroxynitrite through SIN-1. Peroxynitrite decreased progressive and total motility, as well as some sperm kinetic parameters. Mitochondrial membrane potential also decreased. These alterations occurred with no decrease in sperm viability. In conclusion, peroxynitrite-induced nitrosative stress impairs vital functions in the male gamete, possibly contributing to male infertility.

Effect of mitochondrial calcium uniporter blocking on human spermatozoa.

Calcium (Ca(2+) ) regulates a number of essential processes in spermatozoa. Ca(2+) is taken up by mitochondria via the mitochondrial calcium uniporter (mCU). Oxygen-bridged dinuclear ruthenium amine complex (Ru360) has been used to study mCU because it is a potent and specific inhibitor of this channel. In bovine spermatozoa, it has been demonstrated that mitochondrial calcium uptake inhibition adversely affects the capacitation process. It has been demonstrated in human spermatozoa that mCU blocking, through Ru360, prevents apoptosis; however, the contribution of the mCU to normal human sperm function has not been studied. Therefore, the aim of this study was to evaluate the effect of mCU blocking on human sperm function. Spermatozoa obtained from apparently healthy donors were incubated with 5 and 10 µm Ru360 for 4 h at 37 °C. Viability was assessed using propidium iodide staining; motility was determined by computer-aided sperm analysis, adenosine triphosphate (ATP) levels using a luminescence-based method, mitochondrial membrane potential (ΔΨm) using JC-1 staining and reactive oxygen species (ROS) production using dihydroethidium dye. Our results show that mCU blocking significantly reduced total sperm motility and ATP levels without affecting sperm viability, ΔΨm and ROS production. In conclusion, mCU contributes to the maintenance of sperm motility and ATP levels in human spermatozoa.

Distinct isolates of uropathogenic Escherichia coli differentially affect human sperm parameters in vitro.

Sperm motility and vitality are decreased in male genital tract infection. Uropathogenic Escherichia coli (UPEC) are frequently associated with sperm parameter loss, but there are no reports to date regarding the effects of different E. coli isolates on human spermatozoa. The aim of this work was to compare the effect in vitro of different E. coli isolates on human sperm parameters. Normal spermatozoa were incubated with E. coli isolated from nine men with urinary tract infection. After 1 h of incubation, sperm motility, vitality and mitochondrial membrane potential (ΔΨm) were measured. The E. coli isolates were serotyped with specific antisera. Sperm motility was decreased with five of nine E. coli isolates. Two UPEC were typed as O6 strains, and they did not decrease sperm motility in the same experimental conditions as the other five isolates, despite the described high pathogenicity of the O6 strain in urogenital infections. Neither UPEC analysed affected vitality or ΔΨm. UPEC isolates were shown to be heterogeneous in their effects, suggesting the need to characterise the pattern defining the pathogenicity of E. coli on human spermatozoa.

Crystalline keratopathy secondary to the use of ciprofloxacin after cataract surgery with confirmation by histopathological study: A case report and review of the literature.

OBJECTIVE: To report the case of a 75-year-old patient who presented crystalline keratopathy secondary to the use of topical ciprofloxacin with histopathological verification, after cataract surgery without complications. METHOD: Case report with clinical and photographic follow-up, as well as slides with samples of epithelium and crystalline deposits. RESULTS: Corneal deposits resolved after drug suspension, topical lubricant change, and subsequent surgical debridement. The histopathological examination reported epithelial cells and basophilic particles compatible with drug precipitates. CONCLUSIONS: Crystalline keratopathy is a condition in which crystals of various kinds are deposited in the corneal epithelium and/or in the anterior stroma. It may have an infectious, pharmacological cause or, in rarer cases, corneal dystrophies. Certain factors such as a previous epithelial defect, systemic pathology with diabetes mellitus, ocular surgery and previous dry eye can favor the deposition of ciprofloxacin leading to the formation of a keratopathy.

Swine spermatozoa trigger aggregated neutrophil extracellular traps leading to adverse effects on sperm function.

In pigs, the number of PMN in uterus lumen increases within few hours after natural or artificial AI resulting in early PMN-derived innate immune reactions. Sperm-NETs formation was recently reported to occur in various mammalian species. Aim of this study was to investigate direct interactions of boar spermatozoa with swine PMN, the release of sperm-mediated NETs, and to assess NET-derived effects on sperm functionality. Sperm-triggered NETs were visualized by SEM- and immunofluorescence analyses. Sperm-mediated NETosis was confirmed by presence of extruded DNA with global histones and NE. Largest sizes of sperm-mediated aggNETs were detected after 5 h thereby resulting in effective massive sperm entrapment. The number of aggNETs increased from 3 h onwards. Kinetic studies of swine sperm-mediated NETosis showed to be a time-dependent cellular process. In addition, number of NETs-entrapped spermatozoa increased at 3 h of exposure whilst few free spermatozoa were detected after 3 h. Anchored NETs also increased from 3 h onwards. The cytotoxicity of NETs was confirmed by diminution of the total motility and the progressive motility. Spermatozoa membrane integrity and function loss exposed to NETs was confirmed from 3 h. Experiments revealed NETs-derived damaging effects on swine spermatozoa in membrane integrity, motility and functionality. We hypothesize that swine sperm-triggered aggNETs might play a critical role in reduced fertility potential in swine reproductive technique. Thus, aggNETs formation needs to be considered in future studies about uterine environment as well as advance of sperm in the porcine female reproductive tract.

Molecular Detection of Phytophthora ramorum by Real-Time Polymerase Chain Reaction Using TaqMan, SYBR Green, and Molecular Beacons.

ABSTRACT Sudden oak death, caused by Phytophthora ramorum, is a severe disease that affects many species of trees and shrubs. This pathogen is spreading rapidly and quarantine measures are currently in place to prevent dissemination to areas that were previously free of the pathogen. Molecular assays that rapidly detect and identify P. ramorum frequently fail to reliably distinguish between P. ramorum and closely related species. To overcome this problem and to provide additional assays to increase confidence, internal transcribed spacer (ITS), beta-tubulin, and elicitin gene regions were sequenced and searched for polymorphisms in a collection of Phytophthora spp. Three different reporter technologies were compared: molecular beacons, TaqMan, and SYBR Green. The assays differentiated P. ramorum from the 65 species of Phytophthora tested. The assays developed were also used with DNA extracts from 48 infected and uninfected plant samples. All environmental samples from which P. ramorum was isolated by PARP-V8 were detected using all three real-time PCR assays. However, 24% of the samples yielded positive real-time PCR assays but no P. ramorum cultures, but sequence analysis of the coxI and II spacer region confirmed the presence of the pathogen in most samples. The assays based on detection of the ITS and elicitin regions using TaqMan tended to have lower cycle threshold values than those using beta-tubulin and seemed to be more sensitive.

In vivo confocal microscopy for the oral cavity: Current state of the field and future potential.

Confocal microscopy (CM) has been shown to correlate with oral mucosal histopathology in vivo. The purposes of this review are to summarize what we know so far about in vivo CM applications for oral mucosal pathologies, to highlight some current developments with CM devices relevant for oral applications, and to formulate where in vivo CM could hold further application for oral mucosal diagnosis and management. Ovid Medline® and/or Google® searches were performed using the terms 'microscopy, confocal', 'mouth neoplasms', 'mouth mucosa', 'leukoplakia, oral', 'oral lichen planus', 'gingiva', 'cheilitis', 'taste', 'inflammatory oral confocal', 'mucosal confocal' and 'confocal squamous cell oral'. In summary, inclusion criteria were in vivo use of any type of CM for the human oral mucosa and studies on normal or pathological oral mucosa. Experimental studies attempting to identify proteins of interest and microorganisms were excluded. In total 25 relevant articles were found, covering 8 main topics, including normal oral mucosal features (n=15), oral dysplasia or neoplasia (n=7), inflamed oral mucosa (n=3), taste impairment (n=3), oral autoimmune conditions (n=2), pigmented oral pathology/melanoma (n=1), delayed type hypersensitivity (n=1), and cheilitis glandularis (n=1). The evidence for using in vivo CM in these conditions is poor, as it is limited to mainly small descriptive studies. Current device developments for oral CM include improved probe design. The authors propose that future applications for in vivo oral CM may include burning mouth syndrome, intra-operative mapping for cancer surgery, and monitoring and targeted biopsies within field cancerization.

Comparative studies of two-times-daily versus three-times-daily indinavir in combination with zidovudine and lamivudine.

OBJECTIVES: To compare the efficacy and safety of two-times-daily versus three-times-daily indinavir in combination with zidovudine and lamivudine. DESIGN: Two multicenter, open-label, randomized 24-week studies. METHODS: Adults HIV-1 infection, HIV-1 RNA greater than 10000 copies/ml, and no prior lamivudine or protease inhibitor therapy were eligible. In a pilot study (Study A), patients received indinavir at 800 mg every 8 h, 1000 mg every 12 h, or 1200 mg every 12 h. In a subsequent study (Study B), patients received indinavir at 800 mg every 8 h or 1200 mg every 12 h. All subjects received zidovudine (300 mg) and lamivudine (150 mg) every 12 h. An intent-to-treat analysis was used. RESULTS: In Study A, which enrolled 88 patients, neither HIV-1 RNA nor CD4 cell responses differed significantly between treatment groups at 24 weeks when corrected for multiple comparisons. Study B enrolled 433 patients, but was prematurely discontinued when interim analysis suggested greater efficacy of three-times-daily indinavir. Of the first 87 patients reaching week 24, HIV-1 RNA was less than 400 copies/ml in 91% receiving three-times-daily versus 64% receiving two-times-daily indinavir (P < 0.01). CONCLUSION: Three-times-daily indinavir appears more efficacious than two-times-daily dosing when administered with zidovudine and lamivudine. Two-times-daily indinavir dosing should only be considered in situations characterized by favorable pharmacokinetic drug-drug interactions.

Bacteriology of the gallbladder bile in normal subjects.

A bacteriologic analysis of the gallbladder bile in eighty patients who underwent operation was performed in a prospective study. In all twenty patients with a normal gallbladder as assessed by oral cholecystography and intraoperative palpation, cultured were negative. Among patients with chronic cholecystitis only 30 per cent had positive bile cultures and this rose to 47 per cent among patients with acute cholecystitis.

[New techniques in the study of asthma]. / Nuevas técnicas en el estudio del asma.

Asthma is a chronic inflammatory disease of the airways that is functionally characterized by variability of lung function and bronchial hyper-responsiveness. Its diagnosis and management is commonly hampered by the lack of objective parameters. This article reviews additional techniques for evaluating both, lung function (bronchial challenge with adenosin-monophosphate, lung resistance measurement by either the impulse oscillometry system or whole body pletismography) and bronchial immuno-inflammatory response (induced sputum, condensates of exhaled air, monitoring of nitric oxide in exhaled air), as well as our experience with them.

Barreras para la atención en salud del cáncer gástrico, Santander, Colombia. Etapa exploratoria / Barriers to Health Care for Gastric Cancer Patients in Santander, Colombia: Exploratory Stage

Resumen Introducción: el cáncer gástrico es un problema de salud pública; ocupa el quinto y tercer lugar en incidencia y mortalidad mundial, respectivamente. Objetivo: describir las barreras para la atención en salud percibidas por el adulto con cáncer gástrico, su cuidador y el médico tratante en el departamento de Santander, Colombia-etapa exploratoria en el período 2015-2016. Metodología: estudio cualitativo utilizando algunas técnicas del proceso de análisis de la teoría fundamentada a partir de la aplicación de entrevistas semiestructuradas que fueron codificadas y categorizadas con el software N-VIVO 10. Resultados: Emergieron 13 categorías, 182 códigos descritos en 6 ejes: el primero relacionado con el significado del cáncer y sus barreras; el segundo con las principales barreras para la atención en salud, cuyo orden de mayor a menor frecuencia fue: administrativas, económicas, culturales, de conocimiento, de comunicación e institucionales; el tercero es las estrategias para superar las barreras; el cuarto, las estrategias para disminuirlas; el quinto, sentimientos y el rol de la familia; y el sexto, necesidades del paciente. Conclusión: la fragmentación y segmentación del sistema de salud impone barreras que limitan el diagnóstico temprano del cáncer gástrico y su manejo oportuno. Además, amenazan la calidad de vida del adulto enfermo y su familia.

Abstract Introduction: Gastric cancer is a public health problem that ranks fifth in world incidence and third in mortality. Objective: The aim of the exploratory stage of this study was to describe the barriers to health care perceived by adults with gastric cancer, their caregivers, and their attending physicians in the department of Santander, Colombia in 2015 and 2016. Methodology: This is a qualitative study using process techniques and grounded theory analysis based on semi-structured interviews that were codified and categorized with N-VIVO 10. Results: Thirteen categories and 182 codes described along 6 axes emerged. The first axis is related to the meaning of cancer and its barriers. The second is related to the primary barriers to health care which are, in order from most frequent to least frequent: administrative, economic, cultural, knowledge, communication and institutional. The third axis consists of strategies to overcome barriers. The fourth consists of strategies to diminish barriers. The fifth is related to feelings and the role of the family, and the sixth contains the patient's needs. Conclusion: Fragmentation and segmentation of the health care system imposes barriers that limit early diagnosis of gastric cancer and timely management. In addition, they threaten the quality of life of the sick adult and her or his family.