Batten disease through different in vivo and in vitro models: A review.

Batten disease consists of a family of primarily autosomal recessive, progressive neuropediatric disorders, also known as neuronal ceroid lipofuscinoses (NCLs). These pathologies are characterized by seizures and visual, cognitive and motor decline, and premature death. The pathophysiology of this rare disease is still unclear despite the years of trials and financial aids. This paper has reviewed advantages and limits of in vivo and in vitro models of Batten disease from murine and larger animal models to primitive unicellular models, until the most recently developed patient-derived induced pluripotent stem cells. For each model advantages, limits and applications were analyzed. The first prototypes investigated were murine models that due to their limits were replaced by larger animals. In vitro models gradually replaced animal models for practical, cost, and ethical reasons. Using induced pluripotent stem cells to study neurodegeneration is a new way of studying the disease, since they can be distinguished into differentiating elements like neurons, which are susceptible to neurodegeneration. In vivo and in vitro models have contributed to clarifying to some extent the pathophysiology of the disease. The collection and sharing of suitable human bio samples likely through biobanks can contribute to a better understanding, prevention, and to identify possible treatment strategies of Batten disease.

Metabolic, Apoptotic and Fibro-Inflammatory Profiles of the Heart Exposed to Environmental Electromagnetic Fields.

Environmental stress can disturb the integrative functioning of the cardiovascular system and trigger a number of adaptive and/or maladaptive cell responses. Concomitant with the expanding use of mobile communication systems, public exposure to electromagnetic fields (EMFs) raises the question of the impact of 900 MHz EMFs on cardiovascular health. Therefore, in this study, we experimentally investigated whether 915 MHz EMF exposure influenced cardiac metabolic, antioxidant, apoptotic, and fibro-inflammatory profiles in a mouse model. Healthy mice were sham-exposed or exposed to EMF for 14 days. Western blot analysis using whole cardiac tissue lysates demonstrated that there was no significant change in the expression of oxidative phosphorylation (OXPHOS) complexes between the control and EMF-exposed mice. In addition, the myocardial expression of fibro-inflammatory cytokines, antioxidant enzymes, and apoptosis-related markers remained unchanged in the EMF-challenged hearts. Finally, the structural integrity of the cardiac tissues was preserved among the groups. These findings suggest that the apoptotic, antioxidant, metabolic, and fibro-inflammatory profiles of the heart remained stable under conditions of EMF exposure in the analyzed mice.

Tart cherry (Prunus cerasus L.) dietary supplement modulates visceral adipose tissue CB1 mRNA levels along with other adipogenesis-related genes in rat models of diet-induced obesity.

PURPOSE: There is increasing evidence for the involvement of dietary bioactive compounds in the cross-talk modulation of endocannabinoid system and some of the key regulators of transcriptional control for adipogenesis. METHODS: We aimed to characterize the expression of cannabinoid CB1/CB2 receptors and fatty acid amide hydrolase (FAAH) along with selected adipogenesis-related genes (PPARγ, SREBP-1c and PREF-1), adipocyte-secreted factors (leptin and adiponectin), mitochondrial bioenergetic modulators (PGC-1A and UCP-2), and transient receptor potential vanilloid subtype 1 (TRPV1) and 2 (TRPV2) channels in visceral adipose tissue of rats fed with a high-fat diet (HFD) containing either tart cherry seeds alone or tart cherry seeds and juice for 17 weeks. The visceral adipose tissue was weighed and checked the expression of different markers by qRT-PCR, Western blot and immunohistochemistry. RESULTS: Tart cherry supplements were able to downregulate the HFD-induced mRNA expression of CB1 receptor, SREBP-1c, PPARγ, leptin, TRPV1 and TRPV2 resulting in potential anti-adipogenic effects. CONCLUSION: The present study points out that the intake of bioactive constituents of tart cherry may attenuate the effect of adipogenesis by acting directly on the adipose tissue and modulating the interplay between CB1, PPARγ and TRPV channel gene transcription.

Tart Cherry Juice and Seeds Affect Pro-Inflammatory Markers in Visceral Adipose Tissue of High-Fat Diet Obese Rats.

BACKGROUND: Tart cherries (Prunus cerasus L.) are a rich source of anthocyanins. They are phytochemical flavonoids found in red and blue fruits, and vegetables that can reduce hyperlipidemia. Visceral Adipose Tissue (VAT) has emerged as a major player in driving obesity-related inflammatory response. METHODS: This study has investigated the potential positive effects of tart cherries on rats with Diet-Induced Obesity (DIO). In particular, the inflammatory status in retroperitoneal (RPW) and perigonadal (PGW) adipose tissue were studied. Rats were fed ad libitum for 17 weeks with a hypercaloric diet with the supplementation of tart cherries seeds powder (DS) and seeds powder plus tart cherries juice containing 1mg of anthocyanins (DJS). In RPW and PGW, expression of CRP, IL-1 ß, TNF-α, CCL2 and CD36, were measured by qRT-PCR, Western blot and immunohistochemistry techniques. RESULTS: No differences in the weight of RPW and PGW animals were found between DS and DJS groups compared to DIO rats. However, an increase of inflammatory markers was observed in DIO group in comparison with control lean rats. A modulation of these markers was evident upon tart cherry supplementation. CONCLUSION: Study results suggest that tart cherry enriched-diet did not modify the accumulation of visceral fat, but it decreased inflammatory markers in both tissues. Therefore, this supplementation could be useful, in combination with healthy lifestyles, to modify adipose tissue cell metabolism limiting-obesity related organ damage.

Cardiovascular Changes Related to Metabolic Syndrome: Evidence in Obese Zucker Rats.

Metabolic syndrome (MetS) is a predictor of cardiovascular diseases, commonly associated with oxidative stress and inflammation. However, the pathogenic mechanisms are not yet fully elucidated. The aim of the study is to evaluate the oxidative status and inflammation in the heart of obese Zucker rats (OZRs) and lean Zucker rats (LZRs) at different ages. Morphological and morphometric analyses were performed in the heart. To study the oxidative status, the malondialdehyde (MDA), 4-hydroxynonenal (4-HNE), protein oxidation, and antioxidant enzymes were measured in plasma and heart. To elucidate the inflammatory markers involved, immunohistochemistry and Western blot were performed for cellular adhesion molecules and proinflammatory cytokines. OZRs were characterized by hypertension, hyperlipidemia, hyperglycemia, and insulin resistance. The obesity increased MDA and decreased the activities of superoxide dismutase (SOD) in plasma as well as in the heart, associated with cardiomyocytes hypertrophy. OxyBlot in plasma and in heart showed an increase of oxidativestate proteins in OZRs. Vascular cell adhesion molecule-1, interleukin-6, and tumor necrosis factor-α expressions in OZRs were higher than those of LZRs. However, these processes did not induce apoptosis or necrosis of cardiomyocytes. Thus, MetS induces the lipid peroxidation and decreased antioxidant defense that leads to heart tissue changes and coronary inflammation.

Differences in Mitochondrial Membrane Potential Identify Distinct Populations of Human Cardiac Mesenchymal Progenitor Cells.

Adult human cardiac mesenchymal progenitor cells (hCmPC) are multipotent resident populations involved in cardiac homeostasis and heart repair. Even if the mechanisms have not yet been fully elucidated, the stem cell differentiation is guided by the mitochondrial metabolism; however, mitochondrial approaches to identify hCmPC with enhanced stemness and/or differentiation capability for cellular therapy are not established. Here we demonstrated that hCmPCs sorted for low and high mitochondrial membrane potential (using a lipophilic cationic dye tetramethylrhodamine methyl ester, TMRM), presented differences in energy metabolism from preferential glycolysis to oxidative rates. TMRM-high cells are highly efficient in terms of oxygen consumption rate, basal and maximal respiration, and spare respiratory capacity compared to TMRM-low cells. TMRM-high cells showed characteristics of pre-committed cells and were associated with higher in vitro differentiation capacity through endothelial, cardiac-like, and, to a lesser extent, adipogenic and chondro/osteogenic cell lineage, when compared with TMRM-low cells. Conversely, TMRM-low showed higher self-renewal potential. To conclude, we identified two hCmPC populations with different metabolic profile, stemness maturity, and differentiation potential. Our findings suggest that metabolic sorting can isolate cells with higher regenerative capacity and/or long-term survival. This metabolism-based strategy to select cells may be broadly applicable to therapies.

Correction: Tayebati et al. Choline and Choline alphoscerate Do Not Modulate Inflammatory Processes in the Rat Brain. Nutrients 2017, 9, 1084.

For the original publication [...].

Protective effects of the R-(+)-thioctic acid treatment: possible anti-inflammatory activity on heart of hypertensive rats.

BACKGROUND: In cardiovascular disease, high blood pressure is associated with oxidative stress, promoting endothelial dysfunction, vascular remodeling, and inflammation. Clinical trials are discordant that the most effective treatment in the management of hypertension seems to be the administration of anti-hypertensive drugs with antioxidant properties. The study aims to evaluate the effects of the eutomer of thioctic acid on oxidative stress and inflammation in the heart of spontaneously hypertensive rats compared to normotensive Wistar Kyoto rats. METHODS: To study the oxidative status, the malondialdehyde and 4-hydroxynonenal concentration, protein oxidation were measured in the heart. Morphological analysis were performed. Immunohistochemistry and Western blot were done for alpha-smooth muscle actin and transforming growth factor beta to assess fibrosis; cytokines and nuclear factor kappaB to assess inflammatory processes. RESULTS: Spontaneously hypertensive rats were characterized by hypertension with increased malondialdehyde levels in the heart. OxyBlot in the heart of spontaneously hypertensive rats showed an increase in proteins' oxidative status. Cardiomyocyte hypertrophy and fibrosis in the ventricles were associated with an increased expression of alpha-smooth muscle actin and pro-inflammatory cytokines, reduced by the eutomer of thioctic acid supplementation. CONCLUSIONS: Based on this evidence, eutomer of thioctic acid could represent an appropriate antioxidant molecule to reduce oxidative stress and prevent inflammatory processes on the cardiomyocytes and cardiac vascular endothelium.

Correction: Micioni Di Bonaventura et al. Brain Alterations in High Fat Diet Induced Obesity: Effects of Tart Cherry Seeds and Juice. Nutrients 2020, 12, 623.

In the original publication [...].

Dysfunction of the Brown Adipose Organ in HFD-Obese Rats and Effect of Tart Cherry Supplementation.

Obesity has a great impact on adipose tissue biology, based on its function as a master regulator of energy balance. Brown adipose tissue (BAT) undergoes remodeling, and its activity declines in obese subjects due to a whitening process. The anti-obesity properties of fruit extracts have been reported. The effects of tart cherry against oxidative stress, inflammation, and the whitening process in the BAT of obese rats were investigated. Intrascapular BAT (iBAT) alterations and effects of Prunus cerasus L. were debated in rats fed for 17 weeks with a high-fat diet (DIO), in DIO supplemented with seed powder (DS), and with seed powder plus the juice (DJS) of tart cherry compared to CHOW rats fed with a normo-caloric diet. iBAT histologic observations revealed a whitening process in DIO rats that was reduced in the DS and DJS groups. A modulation of uncoupling protein-1 (UCP-1) protein and gene expression specifically were detected in the obese phenotype. An upregulation of UCP-1 and related thermogenic genes after tart cherry intake was detected compared to the DIO group. Metabolic adjustment, endoplasmic reticulum stress, protein carbonylation, and the inflammatory microenvironment in the iBAT were reported in DIO rats. The analysis demonstrated an iBAT modulation that tart cherry promoted. In addition to our previous results, these data confirm the protective impact of tart cherry consumption on obesity.

Myocardial capacity of mitochondrial oxidative phosphorylation in response to prolonged electromagnetic stress.

Introduction: Mitochondria are central energy generators for the heart, producing adenosine triphosphate (ATP) through the oxidative phosphorylation (OXPHOS) system. However, mitochondria also guide critical cell decisions and responses to the environmental stressors. Methods: This study evaluated whether prolonged electromagnetic stress affects the mitochondrial OXPHOS system and structural modifications of the myocardium. To induce prolonged electromagnetic stress, mice were exposed to 915 MHz electromagnetic fields (EMFs) for 28 days. Results: Analysis of mitochondrial OXPHOS capacity in EMF-exposed mice pointed to a significant increase in cardiac protein expression of the Complex I, II, III and IV subunits, while expression level of α-subunit of ATP synthase (Complex V) was stable among groups. Furthermore, measurement of respiratory function in isolated cardiac mitochondria using the Seahorse XF24 analyzer demonstrated that prolonged electromagnetic stress modifies the mitochondrial respiratory capacity. However, the plasma level of malondialdehyde, an indicator of oxidative stress, and myocardial expression of mitochondria-resident antioxidant enzyme superoxide dismutase 2 remained unchanged in EMF-exposed mice as compared to controls. At the structural and functional state of left ventricles, no abnormalities were identified in the heart of mice subjected to electromagnetic stress. Discussion: Taken together, these data suggest that prolonged exposure to EMFs could affect mitochondrial oxidative metabolism through modulating cardiac OXPHOS system.

Rodent Models of Huntington's Disease: An Overview.

Huntington's disease (HD) is an autosomal-dominant inherited neurological disorder caused by a genetic mutation in the IT15 gene. This neurodegenerative disorder is caused by a polyglutamine repeat expansion mutation in the widely expressed huntingtin (HTT) protein. HD is characterized by the degeneration of basal ganglia neurons and progressive cell death in intrinsic neurons of the striatum, accompanied by dementia and involuntary abnormal choreiform movements. Animal models have been extensively studied and have proven to be extremely valuable for therapeutic target evaluations. They reveal the hallmark of the age-dependent formation of aggregates or inclusions consisting of misfolded proteins. Animal models of HD have provided a therapeutic strategy to treat HD by suppressing mutant HTT (mHTT). Transgenic animal models have significantly increased our understanding of the molecular processes and pathophysiological mechanisms underlying the HD behavioral phenotype. Since effective therapies to cure or interrupt the course of the disease are not yet available, clinical research will have to make use of reliable animal models. This paper reviews the main studies of rodents as HD animal models, highlighting the neurological and behavioral differences between them. The choice of an animal model depends on the specific aspect of the disease to be investigated. Toxin-based models can still be useful, but most experimental hypotheses depend on success in a genetic model, whose choice is determined by the experimental question. There are many animal models showing similar HD symptoms or pathologies. They include chemical-induced HDs and genetic HDs, where cell-free and cell culture, lower organisms (such as yeast, Drosophila, C. elegans, zebrafish), rodents (mice, rats), and non-human primates are involved. These models provide accessible systems to study molecular pathogenesis and test potential treatments. For developing more effective pharmacological treatments, better animal models must be available and used to evaluate the efficacy of drugs.

Brain and Retinal Organoids for Disease Modeling: The Importance of In Vitro Blood-Brain and Retinal Barriers Studies.

Brain and retinal organoids are functional and dynamic in vitro three-dimensional (3D) structures derived from pluripotent stem cells that spontaneously organize themselves to their in vivo counterparts. Here, we review the main literature data of how these organoids have been developed through different protocols and how they have been technically analyzed. Moreover, this paper reviews recent advances in using organoids to model neurological and retinal diseases, considering their potential for translational applications but also pointing out their limitations. Since the blood-brain barrier (BBB) and blood-retinal barrier (BRB) are understood to play a fundamental role respectively in brain and eye functions, both in health and in disease, we provide an overview of the progress in the development techniques of in vitro models as reliable and predictive screening tools for BBB and BRB-penetrating compounds. Furthermore, we propose potential future directions for brain and retinal organoids, in which dedicated biobanks will represent a novel tool for neuroscience and ophthalmology research.

Cardiac Cell Exposure to Electromagnetic Fields: Focus on Oxdative Stress and Apoptosis.

Exposure to electromagnetic fields (EMFs) is a sensitive research topic. Despite extensive research, to date there is no evidence to conclude that exposure to EMFs influences the cardiovascular system. In the present study, we examined whether 915 MHz EMF exposure affects myocardial antioxidative and apoptotic status in vitro and in vivo. No statistically significant difference in the apoptotic cell profile and antioxidant capacity was observed between controls and short-term EMF-exposed mouse cardiomyocytes and H9C2 cardiomyoblasts. Compared with sham-exposed controls, mice subjected to a 915 MHz EMF for 48 h and 72 h had no significant effect on structural tissue integrity and myocardial expression of apoptosis and antioxidant genes. Therefore, these results indicate that short-term exposure to EMF in cardiac cells and tissues did not translate into a significant effect on the myocardial antioxidant defense system and apoptotic cell death.

Obesity-Related Brain Cholinergic System Impairment in High-Fat-Diet-Fed Rats.

A link between obesity and cerebral health is receiving growing recognition. Here, we investigate in the frontal cortex and hippocampus the potential involvement of cholinergic markers in brain alterations previously reported in rats with obesity induced by diet (DIO) after long-term exposure (17 weeks) to a high-fat diet (HFD) in comparison with animals fed with a standard diet (CHOW). The obesity developed after 5 weeks of HFD. Bodyweight, systolic blood pressure, glycemia, and insulin levels were increased in DIO rats compared to the CHOW group. Measurements of malondialdehyde (MDA) provided lipid peroxidation in HFD-fed rats. Western blot and immunohistochemical techniques were performed. Our results showed a higher expression of choline acetyltransferase (ChAT) and vesicular acetylcholine transporter (VAChT) in obese rats but not the VAChT expression in the frontal cortex after 17 weeks of HFD. Furthermore, the acetylcholinesterase (AChE) enzyme was downregulated in HFD both in the frontal cortex and hippocampus. In the brain regions analyzed, it was reported a modulation of certain cholinergic receptors expressed pre- and post-synaptically (alpha7 nicotinic receptor and muscarinic receptor subtype 1). Collectively, these findings point out precise changes of cholinergic markers that can be targeted to prevent cerebral injuries related to obesity.

Anti-Inflammatory and Antioxidant Properties of Tart Cherry Consumption in the Heart of Obese Rats.

Obesity is a risk factor for cardiovascular diseases, frequently related to oxidative stress and inflammation. Dietary antioxidant compounds improve heart health. Here, we estimate the oxidative grade and inflammation in the heart of dietary-induced obese (DIO) rats after exposure to a high-fat diet compared to a standard diet. The effects of tart cherry seed powder and seed powder plus tart cherries juice were explored. Morphological analysis and protein expressions were performed in the heart. The oxidative status was assessed by the measurement of protein oxidation and 4-hydroxynonenal in samples. Immunochemical and Western blot assays were performed to elucidate the involved inflammatory markers as proinflammatory cytokines and cellular adhesion molecules. In the obese rats, cardiomyocyte hypertrophy was accompanied by an increase in oxidative state proteins and lipid peroxidation. However, the intake of tart cherries significantly changed these parameters. An anti-inflammatory effect was raised from tart cherry consumption, as shown by the downregulation of analyzed endothelial cell adhesion molecules and cytokines compared to controls. Tart cherry intake should be recommended as a dietary supplement to prevent or counteract heart injury in obese conditions.

Altered Brain Cholinergic and Synaptic Markers in Obese Zucker Rats.

The association between obesity and loss of cognitive performance has been recognized. Although there are data regarding the metabolic alterations in obese conditions and the development of neuroinflammation, no clear evidence concerning obesity-related cholinergic and synaptic impairments in the frontal cortex and hippocampus has been reported yet. Here, we investigate different cholinergic and synaptic markers in 12-, 16-, and 20-week-old obese Zucker rats (OZRs) compared with lean littermate rats (LZRs), using immunochemical and immunohistochemical analysis. Consequently, OZRs showed body weight gain, hypertension, and dysmetabolism. In 20-week-old OZRs, the reduction of vesicular acetylcholine transporter (VAChT) and alpha7 nicotinic acetylcholine receptors (α7nAChR) occurred both in the frontal cortex and in the hippocampus, suggesting a cognitive dysfunction due to obesity and aging. Among the muscarinic receptors analyzed, the level of expression of type 1 (mAChR1) was lower in the hippocampus of the older OZRs. Finally, we showed synaptic dysfunctions in OZRs, with a reduction of synaptophysin (SYP) and synaptic vesicle glycoprotein 2B (SV2B) in 20-week-old OZRs, both in the frontal cortex and in the hippocampus. Taken together, our data suggest specific alterations of cholinergic and synaptic markers that can be targeted to prevent cognitive deficits related to obesity and aging.

Antioxidant Properties of Alpha-Lipoic (Thioctic) Acid Treatment on Renal and Heart Parenchyma in a Rat Model of Hypertension.

Renal and cardiac impairments are frequent events in the presence of hypertension. Organ damage is mainly linked to oxidative stress due to high blood pressure and may be reduced by antioxidant supplementation. Alpha-lipoic acid (ALA) is one of most effective antioxidants. It is widely used as a nutritional supplement in a racemic mixture (+/-), even though the (+)-enantiomer is biologically active. This study was designed to investigate the effect of treatment with (+/-)-ALA and its enantiomers on renal and heart parenchyma in spontaneously hypertensive rats (SHR), using immunochemical and immunohistochemical techniques. The results confirmed that the oxidative mechanisms of organ alterations, due to hypertension, and characterized by glomerular and tubular lesions, left ventricular hypertrophy, and fibrosis but not by apoptosis were accompanied by proteins' and nucleic acids' oxidation. We found greater effectiveness of (+)-ALA compared to (+/-)-ALA in reducing oxidative stress, cardiac and renal damages in SHR. To conclude, these data propose (+)-ALA as one of the more appropriate antioxidant molecules to prevent renal and cardiac alterations associated with hypertension.

Natural Antioxidant Application on Fat Accumulation: Preclinical Evidence.

Obesity represents one of the most important challenges in the contemporary world that must be overcome. Different pathological consequences of these physical conditions have been studied for more than 30 years. The most nagging effects were found early in the cardiovascular system. However, later, its negative impact was also investigated in several other organs. Damage at cellular structures due to overexpression of reactive oxygen species together with mechanisms that cause under-production of antioxidants leads to the development of obesity-related complications. In this view, the negative results of oxidant molecules due to obesity were studied in various districts of the body. In the last ten years, scientific literature has reported reasonable evidence regarding natural and synthetic compounds' supplementation, which showed benefits in reducing oxidative stress and inflammatory processes in animal models of obesity. This article attempts to clarify the role of oxidative stress due to obesity and the opposing role of antioxidants to counter it, reported in preclinical studies. This analysis aims to clear-up different mechanisms that lead to the build-up of pro-oxidants during obesity and how various molecules of different origins hinder this phenomenon, behaving as antioxidants.

Galanin promotes autophagy and alleviates apoptosis in the hypertrophied heart through FoxO1 pathway.

Autophagy and apoptosis are powerful regulators of multiple facets of cellular metabolism and homeostasis. Here, we uncover that galanin, a pleiotropic peptide, regulates cardiac autophagy and deactivates apoptotic cell death through the Forkhead box protein O1 (FoxO1) pathway. In hypertrophied heart, galanin promotes autophagy and metabolic shift from fatty acid (FA) to glucose oxidation and preserves mitochondrial integrity. In cardiomyoblasts, galanin triggers autophagosome formation and alleviates hypertrophy, apoptotic cell death, and mitochondrial stress. Mechanistically, galanin dictates cell autophagic and anti-apoptotic phenotypes through FoxO1 pathway. Together, these findings uncover a previously unknown role for galanin in the regulation of cardiac autophagy and provide new insights into the molecular mechanisms supporting cell survival in the hypertrophic reprogramming of the heart.