Ultrastructural Damage Research Articles

Cadmium induces ferroptosis mediated inflammation by activating Gpx4/Ager/p65 axis in pancreatic β-cells

Cadmium (Cd) is a widely distributed endocrine disruptor and has been reported to be closely correlated to the pathogenesis of diabetes. Since pancreatic β-cells loss and dysfunction are central to pathogenesis of diabetes, studying Cd toxicity on pancreatic β-cells and its molecular mechanism is an important scientific issue. However, less attention has been payed to study how Cd induces pancreatic β-cells death and dysfunction in recent years. Thus, our study aims to explore the toxic mechanism of Cd treatment on pancreatic β-cells using both cellular and animal models. Firstly, it was confirmed that Cd induced decreased cell viability and insulin secretion in a dose-and time-dependent manner in MIN6 cells. To explore the underlying mechanism, transcriptomic analysis was employed to screen the differentially expressed genes and disturbed metabolic pathways. Go and KEGG analysis showed that Cd exposure triggered ferroptosis process in MIN6 cells. We further validated that Cd led to GSH depletion, Gpx4 reduction, lipid peroxidation, mitochondrial membrane potential loss and ultrastructural damage at mitochondrial level. Since immune system process was also perturbed based on GO analysis, we found that Cd activated Ager/Pkc/p65 inflammatory process. Moreover, ferroptosis inhibitor Fer-1 could effectively antagonized the activation of Ager-mediated immune process. It was also revealed that Cd induced iron accumulation as well as decreased Gpx4 expression in mice islets. We also uncovered that Cd led to systemic and pancreatic inflammation as early as third week after Cd exposure. Our study emphasizes the importance of ferroptotic cell death on Cd-induced systemic chronic inflammation. A novel target is provided to prevent Cd-induced pancreatic β-cells dysfunction and improve the chronic inflammatory state for prediabetes prevention.

Tear film instability is associated with weakened colocalization between occludin and MUC5AC in scopolamine-induced dry eye disease (DED) rats.

Dry eye disease (DED) is a disease with tear film instability because of multiple factors. This study was conducted to explore roles of occludin and MUC5AC in tear film instability in DED rat model. A total of 20 SD rats were divided into DED group (n = 10) and normal control (NC) group (n = 10). DED rat model was established by subcutaneously injecting with scopolamine hydrobromide. Clinical examinations, including tear breakup time (tBUT), Schirmer's test and corneal fluorescein staining, were conducted to determine corneal functions. Transmission electron microscopy was used to measure the ultrastructures of corneal epithelial cells. Western blotting assay was used to identify occludin expression in corneal tissues of DED rats. Real-time PCR (RT-PCR) was performed to verify gene transcription of occludin and MUC5AC. Colocalization between occludin and MUC5AC was identified with confocal fluorescence microscopy. Tear breakup time was significantly shorter, and corneal fluorescein staining score was predominantly higher in DED rats compared to those in normal rats (P < 0.05). Normal rats showed a steady tear secretion throughout the whole experiments, while DED rats showed a dramatic reduction on day 14. DED rats demonstrated ultrastructural damage of Golgi apparatus and endoplasmic reticulum in corneal epithelial cells. Occludin and MUC5AC expressions were significantly downregulated in corneal tissue of DED rats compared with those of normal rats (P < 0.05). Percentage of occludin-MUC5AC-colocalized corneal epithelial cells in DED rats was significantly less compared with those in normal rats (P < 0.01). Tear film stability was damaged in scopolamine-induced DED rats because of the weakened colocalization between occludin and MUC5AC molecule. This study would provide a potential clue for the pathogenesis and a promising theoretical basis for clinical work of DED.

Sirtuin 3 Deficiency Aggravates Kidney Disease in Response to High-Fat Diet through Lipotoxicity-Induced Mitochondrial Damage.

Sirtuin 3 (SIRT3) is the primary mitochondrial deacetylase that controls the antioxidant pathway and energy metabolism. We previously found that renal Sirt3 expression and activity were reduced in mice with type 2 diabetic nephropathy associated with oxidative stress and mitochondrial abnormalities and that a specific SIRT3 activator improved renal damage. SIRT3 is modulated by diet, and to assess whether Sirt3 deficiency aggravates mitochondrial damage and accelerates kidney disease in response to nutrient overloads, wild-type (WT) and Sirt3−/− mice were fed a high-fat-diet (HFD) or standard diet for 8 months. Sirt3−/− mice on HFD exhibited earlier and more severe albuminuria compared to WT mice, accompanied by podocyte dysfunction and glomerular capillary rarefaction. Mesangial matrix expansion, tubular vacuolization and inflammation, associated with enhanced lipid accumulation, were more evident in Sirt3−/− mice. After HFD, kidneys from Sirt3−/− mice showed more oxidative stress than WT mice, mitochondria ultrastructural damage in tubular cells, and a reduction in mitochondrial mass and energy production. Our data demonstrate that Sirt3 deficiency renders mice more prone to developing oxidative stress and mitochondrial abnormalities in response to HFD, resulting in more severe kidney diseases, and this suggests that mitochondria protection may be a method to prevent HFD-induced renal injury.

MiR-301b-5p and its target gene nfatc2ip regulate inflammatory responses in the liver of rainbow trout (Oncorhynchus mykiss) under high temperature stress

Rainbow trout (Oncorhynchus mykiss) is a typical cold-water aquaculture fish and a high-end aquatic product. When water temperature exceeds its optimal range of 12–18 °C, the immune system of rainbow trout becomes weakened and unbalanced. High temperature in summer and global warming severely impact rainbow trout industry. The focus of this study was to explore the mechanisms regulating the immune response of rainbow trout under high temperature stress and identify molecular elements that account for resistance to high temperature. In this study, individual fish were screened in a high temperature stress experiment and divided into resistant (R) and sensitive (S) groups. The hepatic transcriptome sequencing and analysis of mRNAs and microRNAs of the R, S, and control groups showed that the number of the differentially expressed genes (DEGs) in the S group (9259) was higher than that in the R group (5313). Furthermore, the 1233 genes differentially expressed between S and R groups were mainly enriched in immune-related pathways, including cytokine-cytokine receptor interaction, TNF signaling and IL-17 signaling. Among these DEGs were miR-301b-5p and its target gene that encodes nuclear factor of activated T cells two interacting protein (nfatc2ip). The dual-luciferase reporter system and immunofluorescence experiments verified the relationship between miR-301b-5p and nfatc2ip. We also showed that expression levels of miR-301b-5p and nfatc2ip significantly negatively correlated in the liver of rainbow trout under high temperature stress. By performing functional experiments, we showed that activation of miR-301b-5p expression or inhibition of nfatc2ip expression stimulated the phosphorylation of p65, p38, and JNK in the classical nuclear factor kappa-B and mitogen-activated protein kinase pathways under high temperature stress. These manipulations initially promoted the secretion of the pro-inflammatory factor IL-1β and then increased the levels of IL-6, IL-12, and TNF-α. In addition, activation of miR-301b-5p expression or inhibition of nfatc2ip expression stimulated the repair of the hepatic ultrastructural damage caused by high temperature stress by activating the inflammatory response in rainbow trout liver.

Sodium para-aminosalicylic acid ameliorates lead-induced hippocampal neuronal apoptosis by suppressing the activation of the IP3R-Ca2+-ASK1-p38 signaling pathway

Lead (Pb) is a naturally occurring heavy metal, which can damage the brain and affect learning and memory. Sodium para-aminosalicylic acid (PAS-Na), a non-steroidal anti-inflammatory drug, can readily cross the blood-brain barrier. Our previous studies have found that PAS-Na alleviated Pb-induced hippocampal ultrastructural damage and neurodegeneration, but the mechanism has yet to be defined. Here, we investigated the molecular mechanisms that mediate Pb-induced apoptosis in hippocampal neurons, and the efficacy of PAS-Na in alleviating its effects. This work showed that juvenile developmental Pb exposure impaired rats cognitive ability by inducing apoptotic cell death in hippocampal neurons. Pb-induced neuronal apoptosis was accompanied by increased inositol 1,4,5-trisphosphate receptor (IP3R) expression and enhanced intracellular calcium [Ca2+]i levels, which resulted in increased phosphorylation of neuronal apoptosis signal-regulating kinase 1 (ASK1) and p38. Activation of ASK1 and p38 was blocked by IP3R inhibitor and a Ca2+ chelator. Importantly, PAS-Na treatment improved the Pb-induced effects on cognitive deficits in rats, concomitant with rescued neuronal apoptosis. In addition, PAS-Na reduced the expression of IP3R and the ensuing increase in intracellular Ca2+ and decreased the phosphorylation of ASK1 and p38 in Pb-exposed neurons. Taken together, this study demonstrates that the IP3R-Ca2+-ASK1-p38 signaling pathway mediates Pb-induced apoptosis in hippocampal neurons, and that PAS-Na, at a specific dose-range, ameliorates these changes. Collectively, this study sheds novel light on the cellular mechanisms that mediate PAS-Na efficacy, laying the groundwork for future research to examine the treatment potential of PAS-Na upon Pb poisoning.

Liuwei Dihuang Decoction Alleviates Cognitive Dysfunction in Mice With D-Galactose-Induced Aging by Regulating Lipid Metabolism and Oxidative Stress via the Microbiota-Gut-Brain Axis.

BackgroundAging is an important cause of cognitive dysfunction. Liuwei Dihuang decoction (LW), a commonly applied Chinese medicine formula, is widely used for the treatment of aging-related diseases in China. Previously, LW was confirmed to be effective in prolonging life span and reducing oxidative stress in aged mice. Unfortunately, the underlying mechanism of LW remains unclear. The aim of this study was to interpret the mechanism by which LW alleviates cognitive dysfunction related to aging from the perspective of the microbiota-gut-brain axis.MethodAll C57BL/6 mice (n = 60) were randomly divided into five groups: the control, model, vitamin E (positive control group), low-dose LW and high-dose LW groups (n = 12 in each group). Except for those in the control group, D-galactose was subcutaneously injected into mice in the other groups to induce the aging model. The antiaging effect of LW was evaluated by the water maze test, electron microscopy, 16S rRNA sequencing, combined LC–MS and GC–MS metabolomics, and ELISA.ResultsLiuwei Dihuang decoction ameliorated cognitive dysfunction and hippocampal synaptic ultrastructure damage in aging mice. Moreover, LW decreased Proteobacteria abundance and increased gut microbiota diversity in aging mice. Metabolomic analysis showed that LW treatment was associated with the significantly differential abundance of 14 metabolites, which were mainly enriched in apelin signaling, sphingolipid metabolism, glycerophospholipid and other metabolic pathways. Additionally, LW affected lipid metabolism and oxidative stress in aging mice. Finally, we also found that LW-regulated microbial species such as Proteobacteria and Fibrobacterota had potential relationships with lipid metabolism, oxidative stress and hippocampal metabolites.ConclusionIn brief, LW improved cognitive function in aging mice by regulating lipid metabolism and oxidative stress through restoration of the homeostasis of the microbiota-gut-brain axis.

Naltrexone-Induced Cardiac Function Improvement is Associated With an Attenuated Inflammatory Response and Lipid Perioxidation in Volume Overloaded Rats.

In previous studies, upregulation of myocardial opioid receptors as well as the precursors of their endogenous ligands were detected in the failing heart due to chronic volume overload. Moreover, opioid receptor blockade by naltrexone improved left ventricular function. In parallel, inflammatory processes through cytokines have been confirmed to play an important role in the pathogenesis of different forms of heart failure. Thus, the present study examined the systemic and myocardial inflammatory response to chronic volume overload and its modulation by chronic naltrexone therapy. Chronic volume overload was induced in male Wistar rats by applying an infrarenal aortocaval fistula (ACF) for 28 days during which the selective opioid receptor antagonist naltrexone (n = 6) or vehicle (n = 6) were administered via a subcutaneously implanted Alzet minipump. The ultrastructural, morphometric and hemodynamic characterization of ACF animals were performed using an intraventricular conductance catheter in vivo and electron microscopy in vitro. Co-localization of mu-, delta- and kappa-opioid receptor subtypes (MOR, DOR, and KOR respectively) with the voltage gated L-type Ca2+ channel (Cav1.2), the ryanodine receptor (RyR), and mitochondria in cardiomyocytes as well as IL-6, IL-12, TNF-alpha, and Malondialdehyde (MDA) were determined using double immunofluorescence confocal microscopy, RT-PCR and ELISA, respectively. In rat left ventricular myocardium, three opioid receptor subtypes MOR, DOR, and KOR colocalized with Cav1.2, RyR and mitochondria suggesting a modulatory role of the excitation-contraction coupling. In rats with ACF-induced volume overload, signs of heart failure and myocardial ultrastructural damage, chronic naltrexone therapy improved cardiac function and reversed the systemic and myocardial inflammatory cytokine expression as well as lipid peroxidation. In conclusion, antagonism of the cardiodepressive effects of the myocardial opioid system does not only improve left ventricular function but also blunts the inflammatory response and lipid peroxidation.

Humic acids alleviate the toxicity of reduced graphene oxide modified by nanosized palladium in microalgae

The use of graphene-family materials modified by nanosized palladium (Pd/GFMs) has intensified rapidly in various fields; however, the effects of environmental factors (e.g., natural organic matter (NOM)) on the transformation and ecotoxicity of Pd/GFMs remain largely unknown. In this study, reduced graphene oxide modified by nanosized Pd (Pd/rGO) was incubated with humic acid (HA) under light irradiation for 56 d to explore the effects of NOM on the physicochemical transformations (e.g., defects, surface charges and dispersity) and biological toxicity (e.g., growth inhibition, oxidative stress and ultrastructural damage on algae cells) of Pd/GFMs. The results revealed that HA increased the defects and dispersity of Pd/rGO. Growth inhibition, damage to cellular ultrastructures, and oxidative stress in microalgae cells were induced by Pd/rGO, and corresponding defense responses (e.g., superoxide dismutase, peroxidase and glutathione) were activated. HA diminished the above defense responses in microalgae triggered by Pd/rGO by regulating GSH metabolism and the alanine biosynthesis pathway. In the presence of HA, cell wall damage (i.e., hole formation) caused by exposure to Pd/rGO was restored, and the plasmolysis area was reduced by 28.6 %. In addition, growth inhibition, lipid peroxidation, loss of mitochondrial membrane potential and ROS formation induced by 1.0 mg/L MoS2NPs were decreased by 1.4–65.6 %, 13.9–26.1 %, 21.8–58.3 % and 9.6–16.1 %, respectively. These findings highlight the need to consider the effects of HA on the environmental transformation and biological toxicity of Pd/GFMs, which presents significant implications for the management of Pd/GFMs.

PHARMACOLOGICAL AND MECHANISTIC STUDIES OF UREQUINONA, A MOLECULE FROM ROOTS OF PENTALINON ANDRIEUXII MUELL-ARG THAT HEALS MURINE LEISHMANIA MEXICANA INFECTION.

In this work we tested both the in vitro and in vivo anti-Leishmania mexicana activity of a molecule we originally identified in the root of Pentalinon andrieuxii Muell-Arg, a plant that is widely used in Mayan traditional medicine. The chemical name of this molecule is 24-methylcholesta-4-24(28)-dien-3-one, but for simplicity's sake, we assigned the short and trivial name of urequinona that will be used throughout this work. It induces necrosis and apoptosis of promastigotes cultured in vitro and extensive ultrastructural damage of amastigotes. It also induces production of Interleukin (IL)-2 and interferon (IFN)-γ by splenic cells from infected and urequinona treated mice stimulated in vitro with parasite antigen (Ag) but inhibits the production of IL-6 and IL-12p70 by bone-marrow-derived macrophages (BMM) infected in vitro and then treated with urequinona. It also induces activation of transcription factors such as NFkB and AP-1 (NFkB/AP-1) in RAW reporter cells. We also developed a novel pharmaceutical preparation of urequinona encapsulated in hydroxyethyl cellulose for dermal application that significantly reduced (P < 0.05) experimentally induced ear lesions of C57BL/6 mice. We conclude the preparation containing this molecule is a good candidate for a novel anti-leishmanial drug's preparation.

Ultrastructural Damages to H1N1 Influenza Virus Caused by Vapor Essential Oils.

Influenza viruses are transmitted from human to human via airborne droplets and can be transferred through contaminated environmental surfaces. Some works have demonstrated the efficacy of essential oils (EOs) as antimicrobial and antiviral agents, but most of them examined the liquid phases, which are generally toxic for oral applications. In our study, we describe the antiviral activity of Citrus bergamia, Melaleuca alternifolia, Illicium verum and Eucalyptus globulus vapor EOs against influenza virus type A. In the vapor phase, C. bergamia and M. alternifolia strongly reduced viral cytopathic effect without exerting any cytotoxicity. The E. globulus vapor EO reduced viral infection by 78% with no cytotoxicity, while I. verum was not effective. Furthermore, we characterized the EOs and their vapor phase by the head-space gas chromatography–mass spectrometry technique, observing that the major component found in each liquid EO is the same one of the corresponding vapor phases, with the exception of M. alternifolia. To deepen the mechanism of action, the morphological integrity of virus particles was checked by negative staining transmission electron microscopy, showing that they interfere with the lipid bilayer of the viral envelope, leading to the decomposition of membranes. We speculated that the most abundant components of the vapor EOs might directly interfere with influenza virus envelope structures or mask viral structures important for early steps of viral infection.

Approach to optimal assessment of right ventricular remodelling in heart transplant recipients: insights from myocardial work index, T1 mapping, and endomyocardial biopsy.

Right ventricular (RV) dysfunction is an important cause of graft failure after heart transplantation (HTx). We sought to investigate relative merits of echocardiographic tools and cardiac magnetic resonance (CMR) with T1 mapping for the assessment of functional adaptation and remodelling of the RV in HTx recipients. Sixty-one complete data set of echocardiography, CMR, right heart catheterization, and biopsy were obtained. Myocardial work index (MWI) was quantified by integrating longitudinal strain (LS) with invasively measured pulmonary artery pressure. CMR derived RV volumes, T1 time, and extracellular volume (ECV) were quantified. Endomyocardial biopsy findings were used as the reference standard for myocardial microstructural changes. In HTx recipients who never had a previous allograft rejection, longitudinal function parameters were lower than healthy organ donors, while ejection fraction (EF) (52.0 ± 8.7%) and MWI (403.2 ± 77.2 mmHg%) were preserved. Rejection was characterized by significantly reduced LS, MWI, longer T1 time, and increased ECV that improved after recovery, whereas RV volumes and EF did not change MWI was the strongest determinant of rejection related myocardial damage (area under curve: 0.812, P < 0.0001, 95% CI: 0.69-0.94) with good specificity (77%), albeit modest sensitivity. In contrast, T1 time and ECV were sensitive (84%, both) but not specific to detect subclinical RV damage. Subclinical adaptive RV remodelling is characterized by preserved RV EF despite longitudinal function abnormalities, except for MWI. While ultrastructural damage is reflected by MWI, ECV, and T1 time, only MWI has the capability to discriminate functional adaptation from transition to subclinical structural damage.

Intermittent hypoxia promotes the recovery of motor function in rats with cerebral ischemia by regulating mitochondrial function.

Hypoxia preconditioning is neuroprotective, but the therapeutic effects of intermittent hypoxia were not fully considered. The present study investigated the neuroprotective effect and mechanism of intermittent hypoxia on motor function after cerebral ischemia and explored alternative clinical treatment options. In total, 36 8-week-old male Sprague-Dawley rats were subjected to 60 min of transient middle cerebral artery occlusion (tMCAO) and then randomly divided into a sham-operated group (SHAM), tMCAO-sedentary group (SED), and tMCAO-intermittent hypoxia group (IH). The intervention was performed 1 week after tMCAO and lasted 4 weeks. Rats in the IH group were placed in an animal hypoxic chamber (altitude 5000 m and oxygen concentration of 13%) for 4 h/day and 7 days/week, and rats in the SED group were placed in a normoxic environment for 4 weeks. Body weights, neurological deficit scores, cerebral infarction volume ratios, gait analyses, mitochondrial structure, adenosine triphosphate (ATP) content and AMO-activated protein kinase (AMPK), peroxisome proliferator-activated receptor γ co-activator-1α (PGC-1α), and silencing regulatory protein 3 (Sirt3) expression in the peri-ischemic region brain tissues were detected during the intervention. Compared with the SED group, the body weight of the IH group gradually recovered, and the neurological deficit scores were significantly reduced (P < 0.05). The gait analysis results showed that the pressure of the affected paw and the maximum content area, swing speed, stride length, and other parameters were significantly restored (P < 0.05). The cerebral infarction volume ratio was significantly reduced (P < 0.01). Mitochondrial morphological structure damage in the peri-ischemic region brain tissues recovered, the number was significantly increased (P < 0.05), and the expression of AMPK, PGC-1α, and Sirt3 proteins (P < 0.05), and ATP content were significantly increased (P < 0.05). Intermittent hypoxia may activate the AMPK-PGC-1α-Sirt3 signaling pathway, promote mitochondrial biogenesis, repair mitochondrial ultrastructural damage, and improve mitochondrial function to reduce brain damage and promote motor function recovery in rats with cerebral ischemia.

ELECTRON MICROSCOPIC STUDY OF THE LUNGS OF DECEASED PATIENTS WITH COVID-19

TYPE: Late Breaking Abstract TOPIC: Imaging PURPOSE: To visualize the ultrastructural pulmonary damage in patients deceased from SARS-CoV-2 infection. METHODS: An observational study was performed from March 15th 2020 to December 31st 2021 at the Fundación Jiménez Díaz (Spain). Epidemiological and analytical data were obtained. Ultrasound-guided post-mortem percutaneous biopsies were performed with a 33-mm 14G automatic gun, for their histological examination, and for electron microscopy observation. RESULTS: Five patients (80% women) were included. Mean age was 86 years (84-88); 60% had two medical conditions (HT, DM, obesity). The median length of hospitalization was three days (range 1-16). Percutaneous biopsies were performed at the bedside in the first hour after death. Diffuse alveolar damage was present in all patients, with increased alveolar wall thickness. Loss of lung structure and cell lysis and vacuolization were observed in endothelial cells and in pneumocytes of both types. In type II pneumocytes, images consistent with coronavirus viral particles included inside vesicles as well as extracellular particles close to membrane remnants, were identified. Although every patient received at least one prophylactic dose of heparin, 66% had a DD above 3,000 μg/L. Fibrin microthrombi, mainly in alveolar capillaries, were identified in all patients, with no signs of vasculitis. CONCLUSIONS: Pulmonary microthrombosis was observed in all patients, in spite of anticoagulant treatment. A more detailed study under EM showed frequent formation of hyaline membranes as well as severe cellular damage in both type II pneumocytes and endothelial cells. CLINICAL IMPLICATIONS: Microthrombosis seems to be present from the onset even in patients treated with anticoagulation. DISCLOSURE: No significant relationships. KEYWORD: Electron microscopic

Mitigation Effects and Associated Mechanisms of Environmentally Relevant Thiols on the Phytotoxicity of Molybdenum Disulfide Nanosheets.

Thorough investigations of the environmental fate and risks are necessary for the safe application of engineered nanomaterials. Nevertheless, the current understanding of potential transformations of MoS2 (an intensively studied two-dimensional nanosheet) upon interactions with ubiquitous environmentally relevant thiols (ERTs) in water is limited. This study revealed that two ERTs, l-cysteine and mercaptoacetic acid, could modify MoS2 by covalently grafting thiol groups on S atoms of 1T phases, improving the colloidal persistence and chemical stability of MoS2. Compared with the pristine form, MoS2-thiols with higher dispersity exhibited significantly mitigated envelopment and ultrastructural damage to microalgae. MoS2-triggered growth inhibition, upregulation of reactive oxygen species, photosynthetic injury, and metabolic perturbation in algae were remarkably attenuated by ERTs. The diminished capability for MoS2 to generate reactive intermediates and glutathione oxidation driven by ERTs caused the weakness of oxidative stress and negative effects. Additionally, molecular dynamics simulations demonstrated that ERTs altered the extent of the influence of MoS2 on the secondary structures and functions of adsorbed intracellular proteins, which also contributed to the lower phytotoxicity of MoS2. Our findings provide evidence for the crucial role of specific organic ligands in the risk of MoS2 in aquatic environments.

Ethanol Extract of Artemisia Annua Prevents LPS-Induced Inflammation and Blood-Milk Barrier Disruption in Bovine Mammary Epithelial Cells.

This experiment evaluated the pre-protective effect of AAE on inflammatory injury and tight junction disturbance in bMECs induced by LPS. The bMECs were treated with AAE (3, 6, 12 μg/mL) for 3 h and then incubated with 10 μg/mL lipopolysaccharide (LPS) for 12 h. Our results showed that LPS significantly increased the mRNA and protein expression of CD36, induced the phosphorylation of IκBα and p65 and elevated the levels of TNF-α, IL-1β and IL-6 mRNA, which further resulted in ultrastructural damage, disrupted the expression of tight junction proteins (occludin, zonula occludens (ZO-1) and claudin-1) and decreased the viability of bMECs (p &lt; 0.05). More importantly, AAE pretreatment attenuated the expression of CD36, suppressed the activity of the NF-κB signaling pathway and down-regulated the levels of inflammatory factors in LPS-stimulated bMECs (p &lt; 0.05). Therefore, AAE can effectively protect bMECs against inflammatory injury and tight junction dysfunction, which has important research value for the prevention of bovine mastitis.

Responses of Growth, Oxidative Injury and Chloroplast Ultrastructure in Leaves of Lolium perenne and Festuca arundinacea to Elevated O3 Concentrations.

The effects of increasing atmospheric ozone (O3) concentrations on cool-season plant species have been well studied, but little is known about the physiological responses of cool-season turfgrass species such as Lolium perenne and Festuca arundinacea exposed to short-term acute pollution with elevated O3 concentrations (80 ppb and 160 ppb, 9 h d−1) for 14 days, which are widely planted in urban areas of Northern China. The current study aimed to investigate and compare O3 sensitivity and differential changes in growth, oxidative injury, antioxidative enzyme activities, and chloroplast ultrastructure between the two turf-type plant species. The results showed that O3 decreased significantly biomass regardless of plant species. Under 160 ppb O3, total biomass of L. perenne and F. arundinacea significantly decreased by 55.3% and 47.8% (p < 0.05), respectively. No significant changes were found in visible injury and photosynthetic pigment contents in leaves of the two grass species exposed to 80 ppb O3, except for 160 ppb O3. However, both 80 ppb and 160 ppb O3 exposure induced heavily oxidative stress by high accumulation of malondialdehyde and reactive oxygen species in leaves and damage in chloroplast ultrastructure regardless of plant species. Elevated O3 concentration (80 ppb) increased significantly the activities of superoxide dismutase, catalase and peroxidaseby 77.8%, 1.14-foil and 34.3% in L. perenne leaves, and 19.2%, 78.4% and 1.72-fold in F. arundinacea leaves, respectively. These results showed that F. arundinacea showed higher O3 tolerance than L. perenne. The damage extent by elevated O3 concentrations could be underestimated only by evaluating foliar injury or chlorophyll content without considering the internal physiological changes, especially in chloroplast ultrastructure and ROS accumulation.

Fluoride Induced Neurobehavioral Impairments in Experimental Animals: a Brief Review.

Fluoride is one of the major toxicants in the environment and is often found in drinking water at higher concentrations. Living organisms including humans exposed to high fluoride levels are found to develop mild-to-severe detrimental pathological conditions called fluorosis. Fluoride can cross the hematoencephalic barrier and settle in various brain regions. This accumulation affects the structure and function of both the central and peripheral nervous systems. The neural ultrastructure damages are reflected in metabolic and cognitive activities. Hindrances in synaptic plasticity and signal transmission, early neuronal apoptosis, functional alterations of the intercellular signaling pathway components, improper protein synthesis, dyshomeostasis of the transcriptional and neurotrophic factors, oxidative stress, and inflammatory responses are accounted for the fluoride neurotoxicity. Fluoride causes a decline in brain functions that directly influence the overall quality of life in both humans and animals. Animal studies are widely used to explore the etiology of fluoride-induced neurotoxicity. A good number of these studies support a positive correlation between fluoride intake and toxicity phenotypes closely associated with neurotoxicity. However, the experimental dosages highly surpass the normal environmental concentrations and are difficult to compare with human exposures. The treatment procedures are highly dependent on the dosage, duration of exposure, sex, and age of specimens among other factors which make it difficult to arrive at general conclusions. Our review aims to explore fluoride-induced neuronal damage along with associated histopathological, behavioral, and cognitive effects in experimental models. Furthermore, the correlation of various molecular mechanisms upon fluoride intoxication and associated neurobehavioral deficits has been discussed. Since there is no well-established mechanism to prevent fluorosis, phytochemical-based alleviation of its characteristic indications has been proposed as a possible remedial measure.

Synaptic loss in a mouse model of euthyroid Hashimoto\u2019s thyroiditis: possible involvement of the microglia

BackgroundHashimoto’s thyroiditis (HT) is an autoimmune illness that renders individuals vulnerable to neuropsychopathology even in the euthyroid state, the mechanisms involved remain unclear. We hypothesized that activated microglia might disrupt synapses, resulting in cognitive disturbance in the context of euthyroid HT, and designed the present study to test this hypothesis.MethodsExperimental HT model was induced by immunizing NOD mice with thyroglobulin and adjuvant twice. Morris Water Maze was measured to determine mice spatial learning and memory. The synaptic parameters such as the synaptic density, synaptic ultrastructure and synaptic-markers (SYN and PSD95) as well as the interactions of microglia with synapses were also determined.ResultsHT mice had poorer performance in Morris Water Maze than controls. Concurrently, HT resulted in a significant reduction in synapse density and ultrastructure damage, along with decreased synaptic puncta visualized by immunostaining with synaptophysin and PSD-95. In parallel, frontal activated microglia in euthyroid HT mice showed increased engulfment of PSD95 and EM revealed that the synaptic structures were visible within the microglia. These functional alterations in microglia corresponded to structural increases in their attachment to neuronal perikarya and a reduction in presynaptic terminals covering the neurons.ConclusionOur results provide initial evidence that HT can induce synaptic loss in the euthyroid state with deficits might be attributable to activated microglia, which may underlie the deleterious effects of HT on spatial learning and memory.

Effects of electroacupuncture on ROS-NLRP3 inflammatory pathway and autophagy related proteins in hippocampus of vascular dementia rats

To observe the effect of electroacupuncture (EA) on learning-memory ability, ultrastructural changes of hippocampal CA1 neurons, reactive oxygen species (ROS) level, Nod-like receptor protein 3 (NLRP3) and auto-phagy-related proteins expression in the hippocampus of vascular dementia (VD) rats, so as to reveal its partial mechanisms in treating VD. Male SD rats were randomly divided into sham operation, model, and EA groups (n=10 rats in each group). The VD model was established by permanent ligation of bilateral common carotid arteries. Rats of the EA group were treated with EA at "Baihui" (GV20), "Dazhui" (GV14) and bilateral "Shenshu" (BL23) for 30 min, once a day for 4 weeks. Morris water maze was used to evaluate the learning and memory ability of rats before modeling, 4 weeks after modeling and after intervention. Transmission electron microscopy (TEM) was used to observe the ultrastructural changes of hippocampal CA1 neurons. The level of ROS in hippocampus was detected by DCFH-DA fluorescence probe. The expressions of NLRP3, autophagy-related protein Beclin1 and microtubule-associated protein 1 light chain 3 (LC3) were measured by Western blot. In comparison with the sham operation group, the average escape latency of rats in the model group was prolonged (P<0.01), and the times of crossing the original platform were reduced (P<0.05), the level of ROS, the expression levels of LC3-Ⅱ/LC3-Ⅰ ratio, Beclin1 and NLRP3 proteins in hippocampus were increased (P<0.01, P<0.05) in the model group. After EA intervention, the average escape latency of rats was significantly shortened (P<0.01), and the times of crossing the original platform were increased (P<0.05), the level of ROS, the expression levels of LC3-Ⅱ/LC3-Ⅰ ratio, Beclin1 and NLRP3 proteins in hippocampus were decreased (P<0.01, P<0.05) in the EA group compared with those of the model group. Outcomes of TEM showed that CA1 neurons in the hippocampus were damaged, chromatin aggregation, mitochondria pyknosis, cristae structure disorder, rough endoplasmic reticulum expanded and degranulated, the number of free ribosomes decreased, and autophagy could be seen in the model group, which were milder in the EA group. EA at GV20, GV14 and BL23 can improve the learning and memory abilities of VD rats, alleviate the ultrastructural damage of neurons in hippocampal CA1 area, and repair the damaged neurons. The mechanism may be related to the reduction of ROS level, LC3-Ⅱ/LC3-Ⅰ ratio, NLRP3 and Beclin1 protein expression, the decrease of neuronal autophagy, inhibition of activation of NLRP3 inflammasome and alleviation of central inflammatory response.

Subchronic Acrylamide Exposure Activates PERK-eIF2α Signaling Pathway and Induces Synaptic Impairment in Rat Hippocampus.

Acrylamide (ACR), a well-documented neurotoxicant to humans, is extensively found in starchy foods. More than 30% of the typical daily calorie intake comes from ACR-containing foods. Epidemiological and toxicological studies have found that ACR exposure is associated with mild cognitive change in men and experimental animals. However, there is limited information on the mechanisms by which ACR exposure induces memory deficits. The aberrant activation of the PKR-like ER kinase (PERK)-eukaryotic initiation factor 2α (eIF2α) signaling pathway is emerging as a major common theme in cognitive decline. The present study is designed to explore the effect of subchronic ACR exposure on the PERK signaling and the synaptic impairment to elucidate the potential mechanism of ACR-induced cognitive dysfunction in rat. ACR exposure at 5 and 10 (mg/kg)/day by gavage for 14 weeks results in gait abnormality and cognitive impairment in rats, which were accompanied by neuronal loss, glial cell proliferation, and synaptic ultrastructure damage in the hippocampus. ACR reduced the expression of phosphorylated cAMP response element-binding protein (P-CREB), brain-derived neurotrophic factor (BDNF), and synaptic vesicle proteins synapsin-1 and synaptophysin synthesis. ACR also excessively activates the PERK-eIF2α signaling, resulting in overexpression of C/EBP homologous protein (CHOP) and activating transcription factor 4 (ATF4). This work helps to propose a possible mechanism of subchronic exposure of ACR-induced neurotoxicity.