The zebrafish model system for dyslipidemia and atherosclerosis research: Focus on environmental/exposome factors and genetic mechanisms.

Dyslipidemias and atherosclerosis play a pivotal role in cardiovascular risk and disease. Although some pathophysiological mechanisms underlying these conditions have been unveiled, several knowledge gaps still remain. Experimental models, both in vitro and in vivo, have been instrumental to our better understanding of such complex processes. The latter have often been based on rodent species, either wild-type or, in several instances, genetically modified. In this context, the zebrafish may represent an additional very useful in vivo experimental model for dyslipidemia and atherosclerosis. Interestingly, the lipid metabolism of zebrafish shares several features with that present in humans, recapitulating some molecular features and pathophysiological aspects in a better way than that of rodents. The zebrafish model may be of help to address questions related to exposome factors as well as to genetic features, aiming to dissect selected aspects of the more complex scenario observed in humans. Indeed, exposome-related dyslipidemia/atherosclerosis research in zebrafish may target different scientific questions, related to nutrition, microbiota, temperature, light exposure at the larval stage, exposure to chemicals and epigenetic consequences of such external factors. Addressing genetic features related to dyslipidemia/atherosclerosis using the zebrafish model is already a reality and active research is now ongoing in this promising area. Novel technologies (gene and genome editing) may help to identify new candidate genes involved in dyslipidemia and dyslipidemia-related diseases. Based on these considerations, the zebrafish experimental model appears highly suitable for the study of exposome factors, genes and molecules involved in the development of atherosclerosis-related disease as well as for the validation of novel potential treatment options.

Vascular region-specific changes in arterial tone in rats with type 2 diabetes mellitus: Opposite responses of mesenteric and femoral arteries to acetylcholine and 5-hydroxytryptamine.

AIMS: Type 2 diabetes mellitus (T2DM) ranks in the top 10 causes of mortality worldwide. The key factor of T2DM vascular complications is endothelial dysfunction. It is characterized by the vessels motor activity disruption and endothelium-derived factors imbalance. The blood vessels morphological and molecular heterogeneity greatly affects the changes occurring in T2DM. Therefore, we conducted a comparative study of vascular bed changes occurring in T2DM. MAIN METHODS: Male Wistar rats were fed a high-fat diet for 20 weeks, followed by a single streptozotocin injection (20 mg/kg). T2DM was confirmed with an oral glucose tolerance test. KEY FINDINGS: A dose-dependent contraction study showed an increase in third-order mesenteric arterioles response to serotonin but not to phenylephrine. These vessels also exhibited a decrease in acetylcholine-dependent relaxation and an increase in guanylate cyclase function. At the same time, the femoral arteries showed a tendency for increased acetylcholine-dependent relaxation. The blood plasma analysis revealed low bioavailable nitric oxide and high levels of endothelin-1 and ROS. SIGNIFICANCE: This knowledge, in conjunction with the features of the T2DM course, can allow further targeted approaches development for the prevention and treatment of vascular complications occurring in the disease.

Understanding complex dynamics of behavioral, neurochemical and transcriptomic changes induced by prolonged chronic unpredictable stress in zebrafish.

Stress-related neuropsychiatric disorders are widespread, debilitating and often treatment-resistant illnesses that represent an urgent unmet biomedical problem. Animal models of these disorders are widely used to study stress pathogenesis. A more recent and historically less utilized model organism, the zebrafish (Danio rerio), is a valuable tool in stress neuroscience research. Utilizing the 5-week chronic unpredictable stress (CUS) model, here we examined brain transcriptomic profiles and complex dynamic behavioral stress responses, as well as neurochemical alterations in adult zebrafish and their correction by chronic antidepressant, fluoxetine, treatment. Overall, CUS induced complex neurochemical and behavioral alterations in zebrafish, including stable anxiety-like behaviors and serotonin metabolism deficits. Chronic fluoxetine (0.1 mg/L for 11 days) rescued most of the observed behavioral and neurochemical responses. Finally, whole-genome brain transcriptomic analyses revealed altered expression of various CNS genes (partially rescued by chronic fluoxetine), including inflammation-, ubiquitin- and arrestin-related genes. Collectively, this supports zebrafish as a valuable translational tool to study stress-related pathogenesis, whose anxiety and serotonergic deficits parallel rodent and clinical studies, and genomic analyses implicate neuroinflammation, structural neuronal remodeling and arrestin/ubiquitin pathways in both stress pathogenesis and its potential therapy.

Biological Safety and Biodistribution of Chitosan Nanoparticles.

: The effect of unmodified chitosan nanoparticles with a size of ~100 nm and a weakly positive charge on blood coagulation, metabolic activity of cultured cardiomyocytes, general toxicity, biodistribution, and reactive changes in rat organs in response to their single intravenous administration at doses of 1, 2, and 4 mg/kg was studied. Chitosan nanoparticles (CNPs) have a small cytotoxic effect and have a weak antiplatelet and anticoagulant effect. Intravenous administration of CNPs does not cause significant hemodynamic changes, and 30 min after the CNPs administration, they mainly accumulate in the liver and lungs, without causing hemolysis and leukocytosis. The toxicity of chitosan nanoparticles was manifested in a dose-dependent short-term delay in weight gain with subsequent recovery, while in the 2-week observation period no signs of pain and distress were observed in rats. Granulomas found in the lungs and liver indicate slow biodegradation of chitosan nanoparticles. In general, the obtained results indicate a good tolerance of intravenous administration of an unmodified chitosan suspension in the studied dose range.

The zebrafish tail immobilization (ZTI) test as a new tool to assess stress-related behavior and a potential screen for drugs affecting despair-like states.

BACKGROUND: Affective disorders, especially depression and anxiety, are highly prevalent, debilitating mental illnesses. Animal experimental models are a valuable tool in translational affective neuroscience research. A hallmark phenotype of clinical and experimental depression, the learned helplessness, has become a key target for 'behavioral despair'-based animal models of depression. The zebrafish (Danio rerio) has recently emerged as a promising novel organism for affective disease modeling and CNS drug screening. Despite being widely used to assess stress and anxiety-like behaviors, there are presently no clear-cut despair-like models in zebrafish. NEW METHOD: Here, we introduce a novel behavioral paradigm, the zebrafish tail immobilization (ZTI) test, as a potential tool to assess zebrafish despair-like behavior. Conceptually similar to rodent 'despair' models, the ZTI protocol involves immobilizing the caudal half of the fish body for 5 min, leaving the cranial part to move freely, suspended vertically in a small beaker with water. RESULTS: To validate this model, we used exposure to low-voltage electric shock, alarm pheromone, selected antidepressants (sertraline and amitriptyline) and an anxiolytic drug benzodiazepine (phenazepam), assessing the number of mobility episodes, time spent 'moving', total distance moved and other activity measures of the cranial part of the body, using video-tracking. Both electric shock and alarm pheromone decreased zebrafish activity in this test, antidepressants increased it, and phenazepam was inactive. Furthermore, a 5-min ZTI exposure increased serotonin turnover, elevating the 5-hydroxyindoleacetic acid/serotonin ratio in zebrafish brain, while electric shock prior to ZTI elevated both this and the 3,4-dihydroxyphenylacetic acid/dopamine ratios. In contrast, preexposure to antidepressants sertraline and amitriptyline lowered both ratios, compared to the ZTI test-exposed fish. COMPARISON WITH EXISTINGMETHOD(S): The ZTI test is the first despair-like experimental model in zebrafish. CONCLUSIONS: Collectively, this study suggests the ZTI test as a potentially useful protocol to assess stress-/despair-related behaviors, potentially relevant to CNS drug screening and behavioral phenotyping of zebrafish.

Antibody against Na/K-ATPase Inhibitor Lowers Blood Pressure and Increases Vascular Fli1 in Experimental Preeclampsia.

BACKGROUND: Previous studies implicated cardiotonic steroids, including Na/K-ATPase inhibitor marinobufagenin (MBG), in the pathogenesis of preeclampsia (PE). We demonstrated that MBG induces fibrosis via mechanism involving inhibition of Fli1, a nuclear transcription factor and a negative regulator of collagen-1 synthesis. We hypothesized that PE blockade of increased MBG with antibody would lessen the fibrosis of umbilical arteries and lower the blood pressure in rats with PE. METHODS: We tested 36 pregnant Sprague-Dawley rats in which 12 were made hypertensive by 1.8% Na supplementation (days 6-19 of gestation), 12 pregnant rats served controls. At day 19, PE rats received one intraperitoneal injection of polyclonal anti-MBG-4 antibody (0.5 ug/ml) for 4 hours. RESULTS: PE was associated with higher blood pressure (117 ± 2 vs. 107 ± 2 mm Hg; P < 0.01), plasma MBG levels (1.54 ± 0.34 vs. 0.49 ± 0.11 nmol/L; P < 0.01), protein excretion (26 vs. 12 mg/24 hours), sFlt-1 (3-fold), decrease in Fli1 (7-fold) and increase in collagen-1 in aorta (4-fold) vs. control rats (all P < 0.01). In 12 rats treated with polyclonal anti-MBG-4 antibody blood pressure dropped (93 ± 3 mm Hg) and Fli1 was decreased much less (2-fold; P < 0.01 vs. nontreated rats). CONCLUSIONS: These results demonstrate that in experimental PE elevated MBG level is implicated in umbilical fibrosis via suppression of Fli1.

Silicon-containing nanocarriers for targeted drug delivery: synthesis, physicochemical properties and acute toxicity.

Silicon-containing nanoparticles (NPs) are considered promising drug carriers for targeted drug delivery. In this study, we investigated the physical and chemical properties of silicon-containing NPs, including silica and organomodified silica NPs (SiO2NPs and OrSiO2NPs, respectively), with different surface modifications, with the aim of increasing drug-loading efficiency. In addition, we described the original synthesis methods of different sizes of OrSiO2NPs, as well as new hybrid OrSiO2NPs with a silica core (SiO2 + OrSiO2NPs). Animal experiments revealed that the silicon-containing NPs investigated were non-toxic, as evidenced by a lack of hemodynamic response after intravenous administration. Bioelimination studies showed rapid renal excretion of OrSiO2NPs. In drug release kinetics studies, adenosine was immobilized on SiO2NPs using three different approaches: physical adsorption, ionic, and covalent bonding. We observed that the rate of adenosine desorption critically depended on the type of immobilization; therefore, adenosine release kinetics can be adjusted by SiO2NP surface modification technique. Adsorption of adenosine on SiO2 + OrSiO2NPs resulted in a significant attenuation of adenosine-induced hypotension and bradycardia.