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Recirculating aquaculture systems (RAS) have become more attractive due to reduced water consumption and effluent discharge. However, intensification of production increases the risk of introducing pathogens at farming sites. The emergence of uncultivable pathogens and RAS pathobiome diversity shifts the traditional disease paradigm from "one pathogen, one disease" to complex multiple-pathogen disease cases. Piscine orthoreovirus genotype 3 (PRV-3) is an excellent example, as it is capable of inducing anemia and heart pathology resembling heart and skeletal muscle inflammation under experimental conditions, and is associated with increased mortality in association with other pathogens in the field. The aim of this study was to develop a method for detection of multiple pathogens and putative pathogens, as co-infections are common in aquaculture. To do this, in the pilot study, we mapped the pathobiome of RAS-farmed rainbow trout (Oncorhynchus mykiss) (commercial RAS, farm A) using both standard diagnostic methods and metabarcording (16S rRNA) to investigate the gill microbiome. During this study, we observed infections with multiple pathogens, and detected two putative gill pathogens Candidatus Branchiomonas cysticola and Candidatus Piscichlamydia salmonis, both of which have been linked with complex gill disease in Atlantic salmon (Salmo salar). Based on the pilot study, we developed and tested a high throughput qPCR (HT-qPCR) chip targeting 22 viral and bacterial pathogens and putative pathogens, followed by a surveillance of a fish cohort in a commercial RAS farm during production (farm B). Co-infection with PRV-3 and Ca. B. cysticola combined with stress inducing management practices may explain the severe disease outbreak observed (37% mortality). The time course study sets the base for a future screening scheme for disease prediction and addresses limitations of the method when testing environmental DNA/RNA.
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Aquicultura , Coinfecção , Doenças dos Peixes , Oncorhynchus mykiss , Animais , Oncorhynchus mykiss/virologia , Oncorhynchus mykiss/microbiologia , Aquicultura/métodos , Coinfecção/microbiologia , Coinfecção/veterinária , Coinfecção/virologia , Doenças dos Peixes/virologia , Doenças dos Peixes/microbiologia , Reação em Cadeia da Polimerase em Tempo Real/métodos , RNA Ribossômico 16S/genética , Brânquias/virologia , Brânquias/microbiologia , Microbiota/genéticaRESUMO
PURPOSE: Ultrasound (US) is the primary imaging modality when a testicular tumor is suspected. Superb microvascular imaging (SMI) is a novel, highly sensitive Doppler technique that allows quantification of flow signals by determination of the Vascular Index (VI). The aim of the present study is to investigate the diagnostic significance of the SMI-derived VI in normal testicular tissue and testicular cancer. METHODS: This retrospective analysis included patients who underwent testicular US in our department from 2018 to 2022. Inclusion criteria were: i) sufficient image quality of the stored images, ii) US with standardized SMI-default setting (colour gain of 44 ± 5), iii) patient age ≥ 18 years, and iv) normal testicular findings or testicular tumor with histopathological workup. US examinations were performed as part of clinical routine using a high-end ultrasound system (Aplio i800/i900, Canon Medical Systems Corporation, Tochigi, Japan). Statistical analysis included Chi-square test and Mann-Whitney U tests and receiver operating characteristic (ROC) curve analysis. RESULTS: A total of 62 patients (31 each with normal findings and testicular tumors) were included. The VI differed statistically significantly (p < 0.001) between normal testis (median 2.5 %) and testicular tumors (median 17.4 %). Like vascular patterns (p < 0.001), the VI (p = 0.030) was shown to distinguish seminomas (median 14.8 %), non-seminomas (median 17.6 %) and lymphomas (median 34.5 %). CONCLUSIONS: In conclusion, our study has shown the VI to be a quantitative tool that can add information for differentiating testicular tumor entities. While further confirmation in larger study populations is desirable, our results suggest that the VI may be a useful quantitative parameter.
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Neoplasias Testiculares , Humanos , Masculino , Neoplasias Testiculares/diagnóstico por imagem , Estudos Retrospectivos , Adulto , Pessoa de Meia-Idade , Testículo/diagnóstico por imagem , Testículo/irrigação sanguínea , Idoso , Adulto Jovem , Ultrassonografia Doppler/métodosRESUMO
PURPOSE: Accurate surgical reconstruction of arterial vascular supply is a crucial part of living kidney transplantation (LDKT). The presence of multiple renal arteries (MRA) in grafts can be challenging. In the present study, we investigated the impact of ligation versus anastomosis of small accessory graft arteries on the perioperative outcome. METHODS: Clinical and radiological outcomes of 51 patients with MRA out of a total of 308 patients who underwent LDKT with MRA between 2011 and 2020 were stratified in two groups and analyzed. In group 1 (20 patients), ligation of accessory arteries (ARAs) and group 2 (31 patients) anastomosis of ARAs was performed. RESULTS: Significant differences were observed in the anastomosis-, surgery-, and warm ischemia time (WIT) in favor of group 1. Students t-test showed comparable serum creatinine levels of 2.33 (± 1.75) to 1.68 (± 0.83) mg/dL in group 1 and 2.63 (± 2.47) to 1.50 (± 0.41) mg/dL in group 2, were seen from 1 week to 1 year after transplant. No increased rates of Delayed graft function (DGF), primary transplant dysfunction and transplant rejection were seen, but graft loss and revision rates were slightly higher when the ARAs were ligated. Analysis of Doppler sonography revealed that segmental perfusion deficits tend to regenerate during the clinical course. CONCLUSION: Ligation of smaller accessory renal arteries may not affect the outcome of living kidney transplantation, except for a minor increase in the reoperation rate. Segmental perfusion deficits of the graft seem to regenerate in most cases as seen in Doppler sonography.
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Transplante de Rim , Humanos , Transplante de Rim/efeitos adversos , Artéria Renal/cirurgia , Doadores Vivos , Estudos Retrospectivos , Sobrevivência de Enxerto , Rim/diagnóstico por imagem , Rim/cirurgia , Rim/irrigação sanguínea , Resultado do TratamentoRESUMO
Ocean disposal of industrial waste from technical DDT [mainly 1,1'-(2,2,2-trichloroethane-1,1-diyl)bis(4-chlorobenzene), or 4,4'-DDT] manufacture occurred historically in the Southern California Bight. However, the paucity of historical records highlights uncertainties as to the mode, location, and timing of disposal or ongoing ecological effects of these wastes. This study combines sampling, chemical analysis, and numerical modeling of deep San Pedro Basin sediments revealing substantial DDT contamination that extends at least 25 km from the mainland. These findings narrate bulk DDT waste disposal to the offshore that peaked in the 1950s, prior to the onset of formal regulations; was agnostic to later-designated disposal sites; and has experienced sluggish transformation. Our findings further indicate an attenuating secondary source for the DDT daughter product, 1-chloro-4-[2,2-dichloro-1-(4-chlorophenyl)ethenyl]benzene (4,4'-DDE), which still deposits into deep San Pedro Basin sediments. While demonstrating the severity of DDT contamination to the region, these findings further define the burial potential of DDT wastes and inform the past, present, and future contamination potential that is needed to understand and predict ecological consequences. This work also points firmly to bulk, not containerized, disposal of DDT waste and to potential alternative contents of collocated waste.
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DDT , Resíduos Industriais , DDT/análise , Resíduos Industriais/análise , Monitoramento Ambiental , Oceanos e Mares , CaliforniaRESUMO
Carbon dioxide laser cutters are used to cut and engrave on various types of materials, including metals, wood, and plastics. Although many are equipped with fume extractors for removing airborne substances generated during laser cutting, gases and particulate matter can be released upon opening the lid after completion. This study focused on investigating laser cutting acrylic sheets and associated emissions. Real-time instruments were utilized to monitor both particulate concentrations and size distributions, while the patented Tsai diffusion sampler was used to collect particulate samples on a polycarbonate membrane and transmission electron microscopy (TEM) grid. Identification of released gases consisted of the use of gas sampling with Teflon gas bags followed by analysis using gas chromatography-mass spectrometry (GC-MS). A portable ambient infrared air analyzer was used to quantify the concentrations of the chemicals released by laser cutting activities. The results of the study found that a significant concentration of particulate matter, including nanoplastic particles ranging 15.4-86 nm in particle sizes, and microplastics with agglomerates were released each time the laser cutter lid was opened and were observed to gradually increase in concentration for a period of at least 20 min after the completion of a cut. The GC-MS gaseous samples primarily contained methyl methacrylate at a low level close to the detection limit of the infrared air analyzer.
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AbstractDeveloping organisms often plastically modify growth in response to environmental circumstances, which may be adaptive but is expected to entail long-term costs. However, the mechanisms that mediate these growth adjustments and any associated costs are less well understood. In vertebrates, one mechanism that may be important in this context is the highly conserved signaling factor insulin-like growth factor 1 (IGF-1), which is frequently positively related to postnatal growth and negatively related to longevity. To test this idea, we exposed captive Franklin's gulls (Leucophaeus pipixcan) to a physiologically relevant nutritional stressor by restricting food availability during postnatal development and examined the effects on growth, IGF-1, and two potential biomarkers of cellular and organismal aging (oxidative stress and telomeres). During food restriction, experimental chicks gained body mass more slowly and had lower IGF-1 levels than controls. Following food restriction, experimental chicks underwent compensatory growth, which was accompanied by an increase in IGF-1 levels. Interestingly, however, there were no significant effects of the experimental treatment or of variation in IGF-1 levels on oxidative stress or telomeres. These findings suggest that IGF-1 is responsive to changes in resource availability but is not associated with increased markers of cellular aging during development in this relatively long-lived species.
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Charadriiformes , Fator de Crescimento Insulin-Like I , Animais , Senescência Celular , Envelhecimento , AlimentosRESUMO
[This corrects the article DOI: 10.1039/D2SD00128D.].
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An amplification-free, nanopore-based nucleic acid detection platform has been demonstrated for rapid, 16S rRNA sequence-specific detection of Neisseria gonorrhoeae at 10-100 CFU mL-1 in human urine against background bacterial flora at 1000 CFU mL-1. Gonorrhea is a very common notifiable communicable disease, antibiotic resistant strains have emerged, and the rate of reported gonococcal infections continues to increase. Since rapid clinical identification of bacterial pathogens in clinical samples is needed to guide proper antibiotic treatment and to control disease spread, it is important to engineer rapid, sensitive, selective, and inexpensive point-of-care (POC) diagnostic devices for pathogens such as N. gonorrhoeae. Our detector technology is based on straightforward conductometric detection of sustained blockage of a glass nanopore. Charge neutral, complementary peptide nucleic acid probes are conjugated to polystyrene beads to capture N. gonorrhoeae 16S rRNA selectively. In the presence of an electric field applied externally through a glass nanopore, the PNA-microbead conjugates that acquire substantial negative charge upon target hybridization are driven to the smaller diameter nanopore. At least partial blockage of the nanopore results in a sustained drop in ionic current that can be measured easily with simple electronics. The ability to detect N. gonorrhoeae over the range of 10 to 100 CFU mL-1 spiked in human urine was demonstrated successfully with estimated sensitivity and specificity of â¼98% and â¼100%, respectively. No false positives were observed for the control group of representative background flora (E. coli, K. pneumoniae, and E. faecalis) at 1000 CFU mL-1. Also, N. gonorrhoeae at 50 CFU mL-1 was successfully detected against 1000 CFU mL-1 of background flora in urine. These results suggest that this amplification-free technology may serve as the basis for rapid, inexpensive, low-power detection of pathogens in clinical samples at the POC.
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In November 2016, infectious pancreatic necrosis virus (IPNV) was isolated from a broodstock female of landlocked sea trout (Salmo trutta) in Lake Vänern in Sweden. VP2 gene sequencing placed the IPNV isolate in genogroup 6, for which pathogenicity is largely unknown. Lake Vänern hosts landlocked sea trout and salmon populations that are endangered, and thus the introduction of new pathogens poses a major threat. In this study we characterized the novel isolate by conducting an infection trial on three salmonid species present in Lake Vänern, whole genome sequencing of the isolate, and prevalence studies in the wild sea trout and salmon in Lake Vänern. During the infection trial, the pathogenicity of the Swedish isolate was compared to that of a pathogenic genogroup 5 isolate. Dead or moribund fish were collected, pooled, and analyzed by cell culture to identify infected individuals. In the trial, the Swedish isolate was detected in fewer sample pools in all three species compared to the genogroup 5 isolate. In addition, the prevalence studies showed a low prevalence (0.2-0.5%) of the virus in the feral salmonids in Lake Vänern. Together the data suggest that the novel Swedish IPNV genogroup 6 isolate is only mildly pathogenic to salmonids.
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Significant strengthening of concrete structures can be obtained when using adhesively-bonded carbon fiber-reinforced polymer (CFRP) systems. Challenges related to such strengthening methods are; however, the brittle concrete delamination failure, reduced warning, and the consequent inefficient use of the CFRP. A novel ductile near-surface mounted reinforcement (NSMR) CFRP strengthening system with a high CFRP utilization is introduced in this paper. It is hypothesized that the tailored ductile enclosure wedge (EW) end anchors, in combination with low E-modulus and high elongation adhesive, can provide significant strengthening and ductility control. Five concrete T-beams were strengthened using the novel system with a CFRP rod activation stress of approximately 980 MPa. The beam responses were compared to identical epoxy-bonded NSMR strengthened and un-strengthened beams. The linear elastic response was identical to the epoxy-bonded NSMR strengthened beam. In addition, the average deflection and yielding regimes were improved by 220% and 300% (average values), respectively, with an ultimate capacity comparable to the epoxy-bonded NSMR strengthened beam. Reproducible and predictable strengthening effect seems obtainable, where a good correlation between the results and applied theory was reached. The brittle failure modes were prevented, where concrete compression failure and frontal overload anchor failure were experienced when failure was initiated.
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Accurate identification and quantification of proteins in solution using nanopores is technically challenging in part because of the large fraction of missed translocation events due to short event times and limitations of conventional current amplifiers. Previously, we have shown that a nanopore interfaced with a poly(ethylene glycol)-dimethacrylate hydrogel with an average mesh size of 3.1 nm significantly enhances the protein residence time within the pore, reducing the number of missed events. We used hydrogel-backed nanopores to sense unlabeled proteins as small as 5.5 kDa in size and 10 fM in concentration. We show that the frequency of protein translocation events linearly scales with bulk concentration over a wide range of concentrations and that unknown protein concentrations can be determined from an interpolation of the frequency-concentration curve with less than 10% error. Further, we show an iterative method to determine a protein volume accurately from measurement data for proteins with a diameter comparable to a nanopore diameter. Our measurements and analysis also suggest several competing mechanisms for the detection enhancement enabled by the presence of the hydrogel.
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Nanoporos , Hidrogéis , ProteínasRESUMO
Viral haemorrhagic septicaemia virus (VHSV) is a negative-sense single-stranded RNA virus that infects more than 140 different fish species. In this study, zebrafish larvae were employed as in vivo model organisms to investigate progression of disease, the correlation between propagation of the infection and irreversibility of disease, cell tropism and in situ neutrophil activity towards the VHSV-infected cells. A recombinant VHSV strain, encoding "tomato" fluorescence (rVHSV-Tomato), was used in zebrafish to be able to follow the progress of the infection in the live host in real-time. Two-day-old zebrafish larvae were injected into the yolk sac with the recombinant virus. The virus titre peaked 96 hr post-infection in zebrafish larvae kept at 18°C, and correlated with 33% mortality and high morbidity among the larvae. By utilizing the transgenic zebrafish line Tg(fli1:GFP)y1 with fluorescently tagged endothelial cells, we were able to demonstrate that the virus initially infected endothelial cells lining the blood vessels. By observing the rVHSV-Tomato infection in the neutrophil reporter zebrafish line Tg(MPX:eGFP)i114 , we inferred that only a subpopulation of the neutrophils responded to the virus infection. We conclude that the zebrafish larvae are suitable for real-time studies of VHS virus infections, allowing in vivo dissection of host-virus interactions at the whole organism level.
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Septicemia Hemorrágica Viral/virologia , Neutrófilos/metabolismo , Novirhabdovirus/fisiologia , Tropismo/fisiologia , Peixe-Zebra , Animais , Modelos Animais de DoençasRESUMO
Although resistive pulse sensing using solid-state nanopores is capable of single-molecule sensitivity, previous work has shown that nanoparticles, such as proteins, pass through nanopores too quickly for accurate detection with typical measurement apparatus. As a result, nanopore measurements of these particles significantly deviate from theoretically estimated current amplitudes and detection rates. Here, we show that a hydrogel placed on the distal side of a nanopore can increase the residence time of nanoparticles within the nanopore, significantly increasing the detection rate and allowing improved resolution of blockage currents. The method is simple and inexpensive to implement while being label-free and applicable to a wide range of nanoparticle targets. Using hydrogel-backed nanopores, we detected the protein IgG with event frequencies several orders of magnitude higher than those in the absence of the hydrogel and with larger measured currents that agree well with theoretical models. We also show that the improved measurement also enables discrimination of IgG and bovine serum albumin in a mixed solution. Finally, we show that measurements of IgG with the hydrogel-backed nanopores can also yield current amplitude distributions that can be analyzed to infer its approximate shape.
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Hidrogéis/química , Nanoporos , Proteínas/química , HumanosRESUMO
Chimeric inhibitors, which merge two drug pharmacophores in a single molecule have become a prominent approach for the design of novel anticancer compounds. Here, we examined animacroxam, which combines histone deacetylase (HDAC) inhibitory and cytoskeleton-interfering pharmacophores, in testicular germ cell tumors (TGCT). The effectiveness of animacroxam was compared to that of the commonly applied chemotherapeutic cisplatin as well as the clinically approved HDAC inhibitor vorinostat. The antineoplastic and antiangiogenic effects of animacroxam on TGCT in vivo were assessed through exploratory animal studies and a modified chorioallantoic membrane assay, revealing that animacroxam has significant antitumor activity in TGCT. A novel positron emission tomography/MR-imaging approach was applied to determine tumor volume and glucose [2-fluoro-2-deoxy-d-glucose (18F-FDG)] uptake in TGCT tumors, revealing reduced glucose uptake in animacroxam-treated TGCTs and showing a dose-dependent suppression of glycolytic enzymes, which led to a breakdown in glycolytic energy production. Furthermore, the observed antiangiogenic effects of animacroxam were related to its ability to inhibit endothelial cell-cell communication, as the expression of gap junction-forming connexin 43 was strongly suppressed, and gap-junctional intercellular mass transport was reduced. Our data suggest that the chimeric HDAC inhibitor animacroxam may become a promising candidate for the treatment of solid cancers and may serve as an interesting alternative to platinum-based therapies.
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Antineoplásicos/farmacologia , Cinamatos/farmacologia , Inibidores de Histona Desacetilases/farmacologia , Imidazóis/farmacologia , Neoplasias Embrionárias de Células Germinativas , Neoplasias Testiculares , Animais , Linhagem Celular Tumoral , Humanos , Masculino , Camundongos , Camundongos Nus , Neoplasias Embrionárias de Células Germinativas/tratamento farmacológico , Neoplasias Embrionárias de Células Germinativas/metabolismo , Neoplasias Embrionárias de Células Germinativas/patologia , Neoplasias Testiculares/tratamento farmacológico , Neoplasias Testiculares/metabolismo , Neoplasias Testiculares/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Red Mark Syndrome (RMS) is a skin disease reported from farmed rainbow trout. Since the turn of the millennium it has been spreading through Europe. RMS is probably a bacterial disease caused by a Midichloria-like organism (MLO). It is non-lethal and causes little obvious changes in appetite or behavior but results in red hyperaemic skin lesions, which may lead to economic losses due to downgrading. Here we transfer RMS to naïve specific pathogen free (SPF) fish by cohabitation with RMS-affected seeder fish. During disease development we characterize local cellular immune responses and regulations of immunologically relevant genes in skin of the cohabitants by immunohistochemistry and qPCR. Skin samples from SPF controls and cohabitants (areas with and without lesions) were taken at 18, 61, 82 and 97 days post-cohabitation. Gene expression results showed that lesions had a Th1-type profile, but with concurrent high expression levels of all three classes of immunoglobulins (IgD, IgM and IgT). The marked local infiltration of IgD + cells in the skin lesions as well as a highly up-regulated expression of the genes encoding sIgD and mIgD indicate that this immunoglobulin class plays an important role in skin immunity in general and in RMS pathology in particular. The co-occurrence of an apparent B cell dominated immune reaction with a Th1-type profile suggests that the local production of antibodies is independent of the classical Th2 pathway.
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Doenças dos Peixes/imunologia , Imunidade Humoral/imunologia , Oncorhynchus mykiss/imunologia , Pele/imunologia , Animais , Doenças dos Peixes/microbiologia , Expressão Gênica , Imunoglobulina D/imunologia , Oncorhynchus mykiss/microbiologia , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Pele/metabolismoRESUMO
A nucleic acid amplification-free, optics-free platform has been demonstrated for sequence-specific detection of Escherichia coli (E. coli) 16S rRNA at 1 aM (10-18 M) against a 106-fold (1 pM) background of Pseudomonas putida (P. putida) RNA. This work was driven by the need for simple, rapid, and low cost means for species-specific bacterial detection at low concentration. Our simple, conductometric sensing device functioned by detecting blockage of a nanopore fabricated in a sub-micron-thick glass membrane. Upon sequence-specific binding of target 16S rRNA, otherwise charge-neutral, PNA oligonucleotide probe-polystyrene bead conjugates become electrophoretically mobile and are driven to the glass nanopore of lesser diameter, which is blocked, thereby generating a large, sustained and readily observable step decrease in ionic current. No false positive signals were observed with P. putida RNA when this device was configured to detect E. coli 16S rRNA. Also, when a universal PNA probe complementary to the 16S rRNA of both E. coli and P. putida was conjugated to beads, a positive response to rRNA of both bacterial species was observed. Finally, the device readily detected E. coli at 10 CFU mL-1 in a 1 mL sample, also against a million-fold background of viable P. putida. These results suggest that this new device may serve as the basis for small, portable, low power, and low-cost systems for rapid detection of specific bacterial species in clinical samples, food, and water.
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Técnicas Biossensoriais/métodos , Escherichia coli/isolamento & purificação , Pseudomonas putida/isolamento & purificação , RNA Ribossômico 16S/análise , RNA Ribossômico 16S/genética , Sequência de Bases , Técnicas Biossensoriais/economia , Custos e Análise de Custo , Escherichia coli/genética , Limite de Detecção , Microesferas , Hibridização de Ácido Nucleico , Ácidos Nucleicos Peptídicos/química , Pseudomonas putida/genética , RNA Ribossômico 16S/química , Fatores de TempoRESUMO
Photodynamic therapy (PDT) has emerged as an effective and minimally invasive cancer treatment modality. In the present study, two novel phthalocyanines, tetratriethyleneoxysulfonyl substituted zinc phthalocyanine (ZnPc) and dihydroxy2,9(10),16(17),23(24)tetrakis(4,7,10trioxaundecan1sulfonyl) silicon phthalocyanine (Pc32), were investigated as photosensitizers (PS) for PDT of cholangiocarcinoma (CC). ZnPc showed a pronounced dosedependent and predominantly cytoplasmic accumulation in EGI1 and TFK1 CC cell lines. Pc32 also accumulated in the CC cells, but this was less pronounced. Without photoactivation, the PS did not exhibit any antiproliferative or cytotoxic effects. Upon photoactivation, ZnPc induced the formation of reactive oxygen species (ROS) and immediate phototoxicity, leading to a dosedependent decrease in cell proliferation, and an induction of mitochondriadriven apoptosis and cell cycle arrest of EGI1 and TFK1 cells. Although photoactivated Pc32 also induced ROS formation in the two cell lines, the extent was less marked, compared with that induced by ZnPcPDT, and pronounced antipoliferative effects occurred only in the less differentiated EGI1 cells, whereas the more differentiated TFK1 cells did not show sustained growth inhibition upon Pc32PDT induction. In vivo examinations on the antiangiogenic potency of the novel PS were performed using chorioallantoic membrane (CAM) assays, which revealed reduced angiogenic sprouting with a concomitant increase in nonperfused regions and degeneration of the vascular network of the CAM following induction with ZnPcPDT only. The study demonstrated the pronounced antiproliferative and antiangiogenic potency of ZnPc as a novel PS for PDT, meriting further elucidation as a promising PS for the photodynamic treatment of CC.
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Colangiocarcinoma/tratamento farmacológico , Indóis/uso terapêutico , Fotoquimioterapia , Fármacos Fotossensibilizantes/uso terapêutico , Silício/uso terapêutico , Zinco/uso terapêutico , Inibidores da Angiogênese/farmacologia , Animais , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Galinhas , Colangiocarcinoma/patologia , Membrana Corioalantoide/efeitos dos fármacos , Membrana Corioalantoide/metabolismo , Humanos , Indóis/farmacologia , Isoindóis , Fármacos Fotossensibilizantes/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Silício/farmacologia , Zinco/farmacologiaRESUMO
Vaccine development is important for sustainable fish farming and novel vaccines need to be efficacy tested before release to the market. Challenge of fish with the pathogen towards which the vaccine has been produced can be conducted either by external exposure though bathing or cohabitation, or by bypassing the mucosa through injection. The latter approach is often preferred since it is easier to control than the former. However, injection is not a very natural route of infection, and the bypass of the mucosa may result in a different efficacy profile of experimental fish compared to farmed fish, for which the vaccines are targeted. The zebrafish is by now a well established practical vertebrate model species due in part to its size and ease of maintenance and genetic manipulation. Here we use zebrafish as a model to visualize and compare the development of infection of Vibrio anguillarum on and in the fish following injection or bathing. Injection of 103 bacteria per fish resulted in approximately 50% mortality by day 4 post-injection. Similar mortality levels were reached in the other group by bathing in 1.25 × 109 bacteria for 1 min. The spreading of bacteria was followed for the first 24 h after injection/bathing by immunohistochemistry and optical projection tomography. The tissues and organs where bacteria were detected differed significantly as a result of time as well as treatment. In the bath group, bacteria were initially found on external surfaces including gut. After 24 h V. anguillarum still persisted in gut but had now also spread to the blood. In the injection group bacteria were found in the blood throughout all sampling times, as well as in the hypodermis and body cavity at most sampling times.
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Progressão da Doença , Doenças dos Peixes/microbiologia , Vibrioses/veterinária , Vibrio/fisiologia , Peixe-Zebra , Animais , Modelos Animais de Doenças , Doenças dos Peixes/imunologia , Tomografia Óptica/veterinária , Vibrioses/imunologia , Vibrioses/microbiologiaRESUMO
Photodynamic therapy (PDT) has gathered much attention in the field of cancer treatment and is increasingly used as an alternative solution for esophageal cancer therapy. However, there is a constant need for improving the effectiveness and tolerability of the applied photosensitizers (PS). Here, we propose tetra-triethyleneoxysulfonyl substituted zinc phthalocyanine (ZnPc) as a promising PS for photodynamic treatment of esophageal cancer. ZnPc-induced phototoxicity was studied in two human esophageal cancer cell lines: OE-33 (adenocarcinoma) and Kyse-140 (squamous cell carcinoma). In vitro studies focused on the uptake and intracellular distribution of the novel ZnPc as well as on its growth inhibitory potential, reactive oxygen species (ROS) formation and the induction of apoptosis. The chicken chorioallantoic membrane assay (CAM assay) and studies on native Wistar rats were employed to determine the antineoplastic and antiangiogenic activity of ZnPc-PDT as well as the tolerability and safety of non-photoactivated ZnPc in vivo. ZnPc was taken up by cancer cells in a dose- and time-dependent manner and showed a homogeneous cytoplasmic distribution. Photoactivation of ZnPc-loaded (1-10 µM) cells led to a dose-dependent growth inhibition of esophageal adenocarcinoma and squamous cell carcinoma cells of >90%. The antiproliferative effect was based on ROS-induced cytotoxicity and the induction of mitochondria-driven apoptosis. In vivo studies on esophageal tumor plaques grown on the CAM revealed pronounced antiangiogenic and antineoplastic effects. ZnPc-PDT caused long-lasting changes in the vascular architecture and a marked reduction of tumor feeding blood vessels. Animal studies confirmed the good tolerability and systemic safety of ZnPc, as no changes in immunological, behavioral and organic parameters could be detected upon treatment with the non-photoactivated ZnPc. Our findings show the extraordinary photoactive potential of the novel ZnPc as a photosensitizer for PDT of esophageal cancer.