LDL Exposure Disrupts Mitochondrial Function and Dynamics in a Hippocampal Neuronal Cell Line.

Hypercholesterolemia has been associated with cognitive dysfunction and neurodegenerative diseases. Moreover, this metabolic condition disrupts the blood-brain barrier, allowing low-density lipoprotein (LDL) to enter the central nervous system. Thus, we investigated the effects of LDL exposure on mitochondrial function in a mouse hippocampal neuronal cell line (HT-22). HT-22 cells were exposed to human LDL (50 and 300 µg/mL) for 24 h. After this, intracellular lipid droplet (LD) content, cell viability, cell death, and mitochondrial parameters were assessed. We found that the higher LDL concentration increases LD content compared with control. Both concentrations increased the number of Annexin V-positive cells, indicating apoptosis. Moreover, in mitochondrial parameters, the LDL exposure on hippocampal neuronal cell line leads to a decrease in mitochondrial complexes I and II activities in both concentrations tested and a reduction in Mitotracker™ Red fluorescence and Mitotracker™ Red and Mitotracker™ Green ratio in the higher concentration, indicating mitochondrial impairment. The LDL incubation induces mitochondrial superoxide production and decreases superoxide dismutase activity in the lower concentration in HT-22 cells. Finally, LDL exposure increases the expression of genes associated with mitochondrial fusion (OPA1 and mitofusin 2) in the lower concentration. In conclusion, our findings suggest that LDL exposure induces mitochondrial dysfunction and modulates mitochondrial dynamics in the hippocampal neuronal cells.

Oral glutamine supplementation relieves muscle loss in immobilized rats, altering p38MAPK and FOXO3a signaling pathways.

BACKGROUND: Skeletal muscle synthesizes, stores, and releases body L-glutamine (GLN). Muscle atrophy due to disabling diseases triggers the activation of proteolytic and pro-apoptotic cell signaling, thus impairing the body's capacity to manage GLN content. This situation has a poor therapeutic prognosis. OBJECTIVE: Evaluating if oral GLN supplementation can attenuate muscle wasting mediated by elevated plasma cortisol and activation of caspase-3, p38MAPK, and FOXO3a signaling pathways in soleus and gastrocnemius muscles of rats submitted to 14-day bilateral hindlimbs immobilization. METHODS: Animals were randomly distributed into six groups: non-immobilized rats (Control), control orally supplemented with GLN (1 g kg-1) in solution with L-alanine (ALA: 0.61 g kg-1; GLN+ALA), control orally supplemented with dipeptide L-alanyl-L-glutamine (DIP; 1.49 g kg-1), hindlimbs immobilized rats (IMOB), IMOB orally GLN+ALA supplemented (GLN+ALA-IMOB), and IMOB orally DIP supplemented (DIP-IMOB). Plasma and muscle GLN concentration, plasma cortisol level, muscle caspase-3 activity, muscle p38MAPK and FOXO3a protein content (total and phosphorylated forms), and muscle cross-sectional area (CSA) were measured. RESULTS: Compared to controls, IMOB rats presented: a) increased plasma cortisol levels; b) decreased plasma and muscle GLN concentration; c) increased muscle caspase-3 activity; d) increased total and phosphorylated p38MAPK protein content; e) increased FOXO3a and decreased phosphorylated FOXO3a protein content; f) reduced muscle weight and CSA befitting to atrophy. Oral supplementation with GLN+ALA and DIP was able to significantly attenuate these effects. CONCLUSIONS: These findings attest that oral GLN supplementation in GLN+ALA solution or DIP forms attenuates rats' skeletal muscle mass wasting caused by disuse-mediated muscle atrophy.

Scientific production and most researched diseases in the Biological Sciences postgraduate programs in Brazil / Produção científica e doenças mais pesquisadas nos programas de pós-graduação em Ciências Biológicas no Brasil

This macro-level scientometrics study aimed to analyze the similarities and differences in the scientific communication patterns of the Brazilian postgraduate programs (BPPs) belonging to the Biological Sciences II field (BS2), as defined by Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES). Also, it was identified the most researched diseases and it was discussed their relationship with the needs of Brazilian public health considering the burden of disease (Disability-Adjusted Life Year - DALY, Brazil) estimated by the World Health Organization (WHO). Thus, the scientific production of the BS2's sub-areas Biophysics, Biochemistry, Pharmacology, Physiology, and Morphology was evaluated from 2013 to 2016, through considering the citation impact, Impact Factor (Journal Citation Reports), and scientific collaboration. Data collected included formal information provided to CAPES by all BPPs through the Plataforma Sucupira as well as metadata from Web of Science documents. In addition, were employed the standardized Medical Subject Headings (PubMed) for the analysis of researched diseases. We concluded that the patterns of scientific communication in Biophysics, Biochemistry, Pharmacology, Physiology, and Morphology were predominantly different. Thus, there is a need to consider specificities among the five sub-areas in the evaluation process performed by CAPES. Different approaches are revealed by identifying the most frequently researched diseases and explaining the contributions of each sub-area for Brazilian public health.

Este estudo cientométrico de nível macro teve como objetivo analisar as semelhanças e as diferenças nos padrões de comunicação científica dos programas de pós-graduação brasileiros (PPGs) da área de Ciências Biológicas II, avaliados pela Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES). Além disso, foram identificadas as doenças mais pesquisadas e foi discutido sua relação com as necessidades de saúde pública brasileira, considerando a carga de doenças (Disability-Adjusted Life Year - DALY, Brasil) estimada pela Organização Mundial da Saúde (OMS). Assim, a produção científica das subáreas Biofísica, Bioquímica, Farmacologia, Fisiologia e Morfologia da área de Ciências Biológicas II foi avaliada de 2013 a 2016, considerando o impacto de citações, o Fator de Impacto (Journal Citation Reports) e a colaboração científica. Os dados coletados incluíram informações declaradas à CAPES por todos os PPGs por meio da Plataforma Sucupira, bem como metadados de documentos da Web of Science. Além disso, foram utilizados os cabeçalhos de Medical Subject Headings (PubMed) para a análise das doenças pesquisadas. Concluímos que os padrões de comunicação científica entre as subáreas Biofísica, Bioquímica, Farmacologia, Fisiologia e Morfologia foram predominantemente diferentes. Assim, é necessário considerar as especificidades entre as cinco subáreas no processo de avaliação realizado pela CAPES. Diferentes abordagens são reveladas a partir da identificação das doenças mais pesquisadas e da explicação das contribuições de cada subárea para a saúde pública brasileira.

Static Magnetic Stimulation Induces Changes in the Oxidative Status and Cell Viability Parameters in a Primary Culture Model of Astrocytes.

Astrocytes play an important role in the central nervous system function and may contribute to brain plasticity response during static magnetic fields (SMF) brain therapy. However, most studies evaluate SMF stimulation in brain plasticity while few studies evaluate the consequences of SMF at the cellular level. Thus, we here evaluate the effects of SMF at 305 mT (medium-intensity) in a primary culture of healthy/normal cortical astrocytes obtained from neonatal (1 to 2-day-old) Wistar rats. After reaching confluence, cells were daily subjected to SMF stimulation for 5 min, 15 min, 30 min, and 40 min during 7 consecutive days. Oxidative stress parameters, cell cycle, cell viability, and mitochondrial function were analyzed. The antioxidant capacity was reduced in groups stimulated for 5 and 40 min. Although no difference was observed in the enzymatic activity of superoxide dismutase and catalase or the total thiol content, lipid peroxidation was increased in all stimulated groups. The cell cycle was changed after 40 min of SMF stimulation while 15, 30, and 40 min led cells to death by necrosis. Mitochondrial function was reduced after SMF stimulation, although imaging analysis did not reveal substantial changes in the mitochondrial network. Results mainly revealed that SMF compromised healthy astrocytes' oxidative status and viability. This finding reveals how important is to understand the SMF stimulation at the cellular level since this therapeutic approach has been largely used against neurological and psychiatric diseases.

Resveratrol increases the activation markers and changes the release of inflammatory cytokines of hepatic stellate cells.

The phytoalexin Resveratrol (3,5,4'-trihydroxystilbene; RSV) has been related to numerous beneficial effects on health by its cytoprotection and chemoprevention activities. Liver fibrosis is characterized by the extracellular matrix accumulation after hepatic injury and can lead to cirrhosis. Hepatic stellate cells (HSC) play a crucial role during fibrogenesis and liver wound healing by changing their quiescent phenotype to an activated phenotype for protecting healthy areas from damaged areas. Strategies on promoting the activated HSC death, the quiescence return or the cellular activation stimuli decrease play an important role on reducing liver fibrosis. Here, we evaluated the RSV effects on some markers of activation in GRX, an HSC model. We further evaluated the RSV influence in the ability of GRX on releasing inflammatory mediators. RSV at 1 and 10 µM did not alter the protein content of α-SMA, collagen I and GFAP; but 50 µM increased the content of these activation-related proteins. Also, RSV did not change the myofibroblast-like morphology of GRX. Interestingly, RSV at 10 and 50 µM decreased the GRX migration and collagen-I gel contraction. Finally, we showed that RSV triggered the increase in the TNF-α and IL-10 content in culture media of GRX while the opposite occurred for the IL-6 content. Altogether, these results suggested that RSV did not decrease the activation state of GRX and oppositely, triggered a pro-activation effect at the 50 µM concentration. However, despite the increase of TNF- α in culture media, these results on IL-6 and IL-10 secretion were in accordance with the anti-inflammatory role of RSV in our model.

Oral glutamine supplementation attenuates inflammation and oxidative stress-mediated skeletal muscle protein content degradation in immobilized rats: Role of 70 kDa heat shock protein.

Skeletal muscle disuse results in myofibrillar atrophy and protein degradation, via inflammatory and oxidative stress-mediated NF-kB signaling pathway activation. Nutritional interventions, such as l-glutamine (GLN) supplementation have shown antioxidant properties and cytoprotective effects through the modulation on the 70-kDa heat shock protein (HSP70) expression. However, these GLN-mediated effects on cell signaling pathways and biochemical mechanisms that control the myofibrillar protein content degradation in muscle disuse situations are poorly known yet. This study investigated the effects of oral GLN plus l-alanine (ALA; GLN â€‹+ â€‹ALA-solution) supplementation, either in their free or dipeptide (L-alanyl-l-glutamine-DIP) form, on GLN-glutathione (GSH) axis and cytoprotection mediated by HSP70 protein expression in the slow-twitch soleus and fast-twitch gastrocnemius skeletal muscle of rats submitted to 14-days of hindlimb immobilization-induced disuse muscle atrophy. Forty-eight Wistar rats were distributed into 6 groups: hindlimb immobilized (IMOB group) and hindlimb immobilized orally supplemented with either GLN (1 g kg-1) plus ALA (0.61 g kg-1) â€‹(GLN â€‹+ â€‹ALA-IMOB group) or 1.49 â€‹g â€‹kg-1 of DIP (DIP-IMOB group) and; no-immobilized (CTRL) and no-immobilized supplemented GLN â€‹+ â€‹ALA and DIP baselines groups. All animals, including CTRL and IMOB rats (water), were supplemented via intragastric gavage for 14 days, concomitantly to immobilization period. Plasma and muscle GLN levels, lipid (thiobarbituric acid reactive substances-TBARS) and protein (carbonyl) peroxidation, erythrocyte concentration of reduced GSH and GSH disulfide (GSSG), plasma and muscle pro-inflammatory TNF-α levels, muscle IKKα/ß-NF-kB signaling pathway and, the myofibrillar protein content (MPC) were measured. The MPC was significantly lower in IMOB rats, compared to CTRL, GLN â€‹+ â€‹ALA, and DIP animals (p â€‹< â€‹0.05). This finding was associated with reduced plasma and muscle GLN concentration, equally in IMOB animals. Conversely, both GLN â€‹+ â€‹ALA and DIP supplementation restored plasma and muscle GLN levels, which equilibrated GSH and intracellular redox status (GSSG/GSH ratio) in erythrocytes and skeletal muscle even as, increased muscle HSP70 protein expression; attenuating oxidative stress and TNF-α-mediated NF-kB pathway activation, fact that reverberated on reduction of MPC degradation in GLN â€‹+ â€‹ALA-IMOB and DIP-IMOB animals (p â€‹< â€‹0.05). In conclusion, the findings shown herein support the oral GLN â€‹+ â€‹ALA and DIP supplementations as a therapeutic and effective nutritional alternative to attenuate the deleterious effects of the skeletal muscle protein degradation induced by muscle disuse.

Exogenous expression of caveolin-1 is sufficient for hepatic stellate cell activation.

Caveolin-1 (Cav-1) expression is increased in hepatic stellate cells (HSC) upon liver cirrhosis and it functions as an integral membrane protein of lipid rafts and caveolae that regulates and integrates multiple signals as a platform. This study aimed to evaluate the role of Cav-1 in HSC. Thus, the effects of exogenous expression of Cav-1 in GRX cells, a model of activated HSC, were determined. Here, we demonstrated through evaluating well-known HSC activation markers - such as α-smooth muscle actin, collagen I, and glial fibrillary acidic protein - that up regulation of Cav-1 induced GRX to a more activated phenotype. GRXEGFP-Cav1 presented an increased migration, an altered adhesion pattern, a reorganization f-actin cytoskeleton, an arrested cell cycle, a modified cellular ultrastructure, and a raised endocytic flux. Based on this, GRX EGFP-Cav1 represents a new cellular model that can be an important tool for understanding of events related to HSC activation. Furthermore, our results reinforce the role of Cav-1 as a molecular marker of HSC activation.

Neuroprotective Effects of Guanosine Administration on In Vivo Cortical Focal Ischemia in Female and Male Wistar Rats.

Guanosine (GUO) has neuroprotective effects in experimental models of brain diseases involving glutamatergic excitotoxicity in male animals; however, its effects in female animals are poorly understood. Thus, we investigated the influence of gender and GUO treatment in adult male and female Wistar rats submitted to focal permanent cerebral ischemia in the motor cortex brain. Female rats were subdivided into non-estrogenic and estrogenic phase groups by estrous cycle verification. Immediately after surgeries, the ischemic animals were treated with GUO or a saline solution. Open field and elevated plus maze tasks were conducted with ischemic and naïve animals. Cylinder task, immunohistochemistry and infarct volume analyses were conducted only with ischemic animals. Female GUO groups achieved a full recovery of the forelimb symmetry at 28-35 days after the insult, while male GUO groups only partially recovered at 42 days, in the final evaluation. The ischemic insult affected long-term memory habituation to novelty only in female groups. Anxiety-like behavior, astrocyte morphology and infarct volume were not affected. Regardless the estrous cycle, the ischemic injury affected differently female and male animals. Thus, this study points that GUO is a potential neuroprotective compound in experimental stroke and that more studies, considering the estrous cycle, with both genders are recommended in future investigation concerning brain diseases.

Glioprotective Effect of Resveratrol: an Emerging Therapeutic Role for Oligodendroglial Cells.

Resveratrol is a natural polyphenol compound highly found in red wine that displays several beneficial effects on the central nervous system (CNS), preventing or slowing the progression of a wide variety of neurological diseases. Its neuroprotective role is particularly associated to modulation of antioxidant and anti-inflammatory responses in glial cells in a mechanism dependent of heme oxygenase 1 (HO-1) signaling pathway. Oligodendrocyte progenitor cells (OPC), primarily known for giving rise to mature oligodendrocytes, have emerged as dynamic cells that are also important to maintain the CNS homeostasis. In this sense, we have demonstrated that resveratrol has a protective effect on oligodendroglial functionality against lipopolysaccharide (LPS)-mediated cytotoxicity and that its glioprotective mechanism involves the nuclear factor erythroid 2-related factor 2 (Nrf2) and HO-1 pathways. LPS, through toll-like receptor 4 (TLR4), affected the release of trophic factors by OPC, including transforming growth factor beta (TGF-ß), brain-derived neurotrophic factor (BDNF), and glial cell-derived neurotrophic factor (GDNF), and resveratrol reestablished the trophic factor release to control levels. Additionally, resveratrol prevented the LPS-induced increase in the intracellular reactive oxygen species (ROS) as well as the decrease in glutathione (GSH) levels and in glutamate cysteine ligase (GCL) activity, through Nrf2/HO-1 signaling pathways. Resveratrol also prevented the increase of the transcriptional activities of nuclear factor κB (NFκB) and hypoxia-inducible factor 1 alpha (HIF-1α) after LPS challenge. In summary, this is the first study showing the glioprotective effect of resveratrol on oligodendroglial cells.

Octyl gallate reduces ATP levels and Ki67 expression leading HepG2 cells to cell cycle arrest and mitochondria-mediated apoptosis.

Octyl gallate (OG) is an antioxidant that has shown anti-tumor, anti-diabetic and anti-amyloidogenic activities. Mitochondria play an important role in hepatocellular carcinoma, mainly by maintaining accelerated cellular proliferation through the production of ATP. Thus, the mitochondria may be a target for antitumor therapies. Here, we investigated the effects of OG in the hepatocarcinoma cell line (HepG2) and the mechanisms involved. We report, for the first time, that treatment with OG for 24h inhibited HepG2 cell growth by decreasing mitochondrial activity and mass, which led to the reduction of ATP levels. This reduction in the energy supply triggered a decrease in Ki67 protein expression, leading cells to cycle arrest. In addition, treatment with two doses of OG for 48h induced loss of mitochondrial functionality, mitochondrial swelling and apoptosis. Finally, we report that HepG2 cells had no resistance to treatment after multiple doses. Collectively, our findings indicate that metabolic dysregulation and Ki67 protein reduction are key events in the initial anti-proliferative action of OG, whereas mitochondrial swelling and apoptosis induction are involved in the action mechanism of OG after prolonged exposure. This suggests that OG targets mitochondria, thus representing a candidate for further research on therapies for hepatocarcinoma.

Dissociation between dopaminergic response and motor behavior following intrastriatal, but not intravenous, transplant of bone marrow mononuclear stem cells in a mouse model of Parkinson's disease.

Parkinson's disease is characterized by the progressive loss of dopaminergic neurons from the substantia nigra, a process that leads to a dopamine deficiency in the striatum. This deficiency is responsible for the development of motor symptoms, including resting tremor, bradykinesia, rigidity and postural instability. Based on the observation of substantial neuronal death, alternatives to Parkinson's disease treatment have been studied, including cell-based therapies. The present study aimed to assess the therapeutic potential of intravenous and intrastriatal transplant of bone marrow mononuclear cells in a mouse model of Parkinson's disease. Animals underwent stereotaxic surgery and received an injection of 6-hydroxydopamine into their medial forebrain bundle. Three weeks later, mice were injected with bone marrow mononuclear cells or saline through the caudal vein or directly into their right striatum. Motor function was assessed using the rotarod and apomorphine-induced rotation tests. Our results showed that intrastriatal bone marrow mononuclear cells, but not intravenous, have a short-term therapeutic effect on dopaminergic response in this mice model of parkinsonism assessed by the apomorphine-induced rotation test. This phenomenon was not identified on the rotarod test, showing dissociation between dopaminergic response and motor behavior. Further experiments are needed to elucidate the precise mechanisms involved in these effects.

Resveratrol Regulates the Quiescence-Like Induction of Activated Stellate Cells by Modulating the PPARγ/SIRT1 Ratio.

The activation of hepatic stellate cell (HSC), from a quiescent cell featuring cytoplasmic lipid droplets to a proliferative myofibroblast, plays an important role in liver fibrosis development. The GRX line is an activated HSC model that can be induced by all-trans-retinol to accumulate lipid droplets. Resveratrol is known for activating Sirtuin1 (SIRT1), a NAD(+)-dependent deacetylase that suppresses the activity of peroxisome proliferator-activated receptor gamma (PPARγ), an important adipogenic transcription factor involved in the quiescence maintenance of HSC. We evaluated the effects of 0.1 µM of resveratrol in retinol-induced GRX quiescence by investigating the interference of SIRT1 and PPARγ on cell lipogenesis. GRX lipid accumulation was evaluated through Oil-red O staining, triacylglycerides quantification, and [(14)C] acetate incorporation into lipids. mRNA expression and protein content of SIRT1 and PPARγ were measured by RT-PCR and immunoblotting, respectively. Resveratrol-mediated SIRT1 stimuli did not induce lipogenesis and reduced the retinol-mediated fat-storing capacity in GRX. In order to support our results, we established a cell culture model of transgenic super expression of PPARγ in GRX cells (GRXPγ). Resveratrol reduced lipid droplets accumulation in GRXPγ cells. These results suggest that the PPARγ/SIRT1 ratio plays an important role in the fate of HSC. Thus, whenever the PPARγ activity is greater than SIRT1 activity the lipogenesis is enabled.