Prenatal folate deficiency impairs sociability and memory/recognition in mice offspring.

Folate is essential for the normal growth and development of the fetus. Folic acid supplementation during the fetal period affects postnatal brain development and reduces the incidence of mental disorders in animal and human studies. However, the association between folate deficiency (FD) during pregnancy and developmental disorders in children remains poorly understood. In this study, we investigated whether prenatal FD is associated with neurodevelopmental disorders in offspring. ICR mice were fed a control diet (2 mg folic acid/kg diet) or a folate-deficient diet (0.3 mg folic acid/kg diet) from embryonic day 1 until parturition. We evaluated locomotor activity, anxiety, grooming, sociability and learning memory in male offspring at 7-10 weeks of age. No differences were found in locomotor activity or anxiety in the open field test, nor in grooming time in the self-grooming test. However, sociability, spatial memory, and novel object recognition were impaired in the FD mice compared with control offspring. Furthermore, we measured protein expression levels of the NMDA and AMPA receptors, as well as PSD-95 and the GABA-synthesizing enzymes GAD65/67 in the frontal cortex and hippocampus. In FD mice, expression levels of AMPA receptor 1 and PSD-95 in both regions were reduced compared with control mice. Moreover, NMDA receptor subunit 2B and GAD65/67 were significantly downregulated in the frontal cortex of prenatal FD mice compared with the controls. Collectively, these findings suggest that prenatal FD causes behavioral deficits together with a reduction in synaptic protein levels in the frontal cortex and hippocampus.

Perilla Pomace, a By-product of Oil Extraction, Is Rich in Nutrients and Can Favorably Modulate Lipid Metabolism in Sprague-Dawley Rats.

Perilla pomace, a by-product of oil extraction, is rich in nutrients, such as proteins, but it has not been used for purposes other than livestock feeding. The aim of this study was to determine how perilla pomace modulates glucose and lipid metabolism in Sprague-Dawley rats. Dried perilla pomace was added to diet at a concentration of 16%. One experimental group was administered perilla oil equivalent to that in the perilla pomace. After four weeks, the animals were euthanized, and biochemical parameters were measured. Two experiments were conducted using a low-fat (7% by weight) and a high-fat (21% by weight) diet. Regardless of the level of fat in the diets, no differences in food intake were found among the groups. In the low-fat diet-fed rats (Experiment 1), epididymal adipose tissue weight was slightly, but not significantly, lower in perilla pomace-fed rats than in those fed the control diet. Hepatic triglyceride and cholesterol levels were significantly reduced by perilla pomace compared to those in the control group. Serum lipid profiles (triglycerides and cholesterol) were similar to those in the liver, without statistically significant differences. Perilla pomace significantly diminished hepatic fatty acid synthase (FAS) activity. In high-fat diet-fed rats (Experiment 2), pomace did not significantly lower epididymal adipose tissue weight. Hepatic cholesterol levels were lower in rats on the perilla oil than in control rats. The activity of hepatic enzymes involved in fat oxidation was significantly higher in rats fed the perilla pomace than in those fed the control diet. Collectively, these results show that perilla pomace favorably modulates fat metabolism, and the specific effects depend on the fat content in the diet.

5-Fluorouracil exacerbates cefepime-induced convulsions in pentylenetetrazol-kindled mice.

OBJECTIVE: The antibiotics cefepime and meropenem are recommended for the treatment of neutropenia. However, cefepime has been found to be associated with both peripheral and central adverse events such as renal impairment and seizures, respectively. Previous studies showed that cefepime exacerbated convulsions in corneal kindled mouse models of epilepsy. However, its involvement in chemotherapy-induced side effects is unknown. METHODS: In this study, we examined the convulsive potential of cefepime (500 mg/kg) and meropenem (500 mg/kg) in pentylenetetrazol (PTZ)-kindled mice using an electroconvulsive shock test with low-intensity stimulus currents. Then, the effects of 5-fluorouracil (5-FU, 200 and 400 mg/kg, i.p.) treatment, a model of chemotherapy-induced side effects, were investigated in the PTZ-kindled mouse model. RESULTS: In fully PTZ-kindled mice, intravenous administration of cefepime (500 mg/kg) or meropenem (500 mg/kg) did not elicit any convulsions in the electroconvulsive shock test with low-intensity stimulus currents. In the PTZ-kindled mice treated with 5-FU (200 mg/kg), intravenous administration of cefepime (500 mg/kg) exacerbated the convulsions that occurred within 1 min in the electroconvulsive shock test, and the mice subsequently developed convulsive status epilepticus. However, intravenous administration of meropenem (500 mg/kg) did not produce such effects. CONCLUSION: These findings suggest that the combination of 5-FU and cefepime exacerbates early-onset convulsive seizures and elicits delayed-onset convulsive status epilepticus. Additionally, 5-FU treatment increases the risk of induction of neurotoxic side effects by cefepime.

Change in mutation frequency at a TP53 hotspot during culture of ENU-mutagenised human lymphoblastoid cells.

Mutations in oncogenes or tumour suppressor genes cause increases in cell growth capacity. In some cases, fully malignant cancer cells develop after additional mutations occur in initially mutated cells. In such instances, the risk of cancer would increase in response to growth of these initially mutated cells. To ascertain whether such a situation might occur in cultured cells, three independent cultures of human lymphoblastoid GM00130 cells were treated with N-ethyl-N-nitrosourea to induce mutations, and the cells were maintained for 12 weeks. Mutant frequencies and spectra of the cells at the MspI and HaeIII restriction sites located at codons 247-250 of the TP53 gene were examined. Mutant frequencies at both sites in the gene exhibited a declining trend during cell culture and reached background levels after 12 weeks; this was also supported by mutation spectra findings. These results indicate that the mutations detected under our assay conditions are disadvantageous to cell growth.

Iso-α-Acids, Bitter Components in Beer, Suppress Inflammatory Responses and Attenuate Neural Hyperactivation in the Hippocampus.

Due to the growth in aging populations worldwide, prevention and therapy for age-related cognitive decline and dementia are in great demand. We previously demonstrated that long-term intake of iso-α-acids, which are hop-derived bitter compounds found in beer, prevent Alzheimer's pathology in a rodent model. On the other hand, the effects of iso-α-acids on neural activity in Alzheimer's disease model mice have not been investigated. Here, we demonstrated that short-term intake of iso-α-acids suppresses inflammation in the hippocampus and improves memory impairment even after disease onset. Importantly, we demonstrated that short-term administration of iso-α-acids attenuated the neural hyperactivation in hippocampus. In 6-month-old 5 × FAD mice exhibiting hippocampus inflammation and memory impairment, oral administration of iso-α-acids for 7 days reduced inflammatory cytokines, including MIP-1α and soluble Aß and improved object memory in the novel object recognition test. In 12-month-old J20 mice, intake of iso-α-acids for 7 days also suppressed inflammatory cytokines and soluble Aß in the brain. Manganese-enhanced magnetic resonance imaging (MEMRI) of hippocampi of J20 mice showed increased manganese compared with wild type mice, but iso-α-acids canceled this increased MEMRI signal in J20 mice, particularly in the hippocampus CA1 and CA3 region. Taken together, these findings suggest that short-term intake of iso-α-acids can suppress hippocampus inflammation even after disease onset and improve hyper neural activity in Alzheimer's disease model mice.

TRPV1 mediates the anticonvulsant effects of acetaminophen in mice.

OBJECTIVE: Acetaminophen is one of the most commonly used analgesic and antipyretic drugs. It has been reported that acetaminophen has anticonvulsant effects in several animal models of seizure. An active metabolite of acetaminophen, AM404, inhibits the uptake of the endocannabinoid anandamide. However, the mechanism of the anticonvulsant effect of acetaminophen is unknown. METHODS: This study was performed to examine whether or not acetaminophen can protect against pentylenetetrazol-induced kindling in mice and to investigate the precise mechanisms of the anticonvulsant effect of acetaminophen using the fully kindled mouse models. RESULTS: Repeated administration of acetaminophen significantly delayed the progression of seizure severity induced by pentylenetetrazol. Additionally, acetaminophen showed a dose-dependent anticonvulsant activity against fully pentylenetetrazol-kindled seizures. AM404 also exhibited a dose-dependent anticonvulsant activity in fully kindled animals. The anticonvulsant activity of acetaminophen was antagonized by capsazepine and AMG9810, two transient receptor potential vanilloid-1 (TRPV1) antagonists. However, the transient receptor potential ankyrin 1 (TRPA1) antagonist HC030031 and CB1 receptor antagonist AM251 had no effect. CONCLUSION: These findings suggest that acetaminophen has an anticonvulsant effect in pentylenetetrazol-kindled mouse models and TRPV1 mediates the anticonvulsant action.

Environmental enrichment alleviates cognitive and behavioral impairments in EL mice.

Epilepsy in children is occasionally associated with comorbidities, such as cognitive impairment, behavioral disturbances, and social deficits. These neurobehavioral comorbidities are closely related to environmental factors and the severity of the seizures. Previous studies show that environmental enrichment has a beneficial effect in animal models of temporal lobe epilepsy following systemic chemoconvulsant administration. However, the effect of environmental enrichment on behavioral impairments in the EL mouse, a genetic model of human idiopathic epilepsy, remains unknown. In the present study, we examined the effect of environmental enrichment on cognitive and behavioral impairments in this murine model. The EL mice, under standard laboratory conditions, exhibited impairments in spatial memory in the Morris water maze test, hyperactivity and impaired habituation in the open-field test, and a deficit in social novelty preference in the three-chamber social approach test, compared with control DDY mice, a genetically related nonepileptic strain. These impairments in EL mice were ameliorated by exposure to an enriched environment. These findings suggest that environmental enrichment effectively ameliorates cognitive and behavioral deficits in EL mice.

Tau Depletion in APP Transgenic Mice Attenuates Task-Related Hyperactivation of the Hippocampus and Differentially Influences Locomotor Activity and Spatial Memory.

Hippocampal hyperactivity, ascribed to amyloid ß (Aß)-induced imbalances in neural excitation and inhibition, is found in patients with mild cognitive impairment, a prodromal stage of Alzheimer's disease (AD). To better understand the relationship between hippocampal hyperactivity and the molecular triggers of behavioral impairments in AD, we used Mn-enhanced MRI (MEMRI) to assess neuronal activity after subjecting mice to a task requiring spatial learning and memory. Depletion of endogenous tau in an amyloid precursor protein (APP) transgenic (J20) mouse line was shown to ameliorate hippocampal hyperactivity in J20 animals, tau depletion failed to reverse memory deficits associated with APP/Aß overproduction. On the other hand, deletion of tau alleviated the hyperlocomotion displayed by APP transgenics, suggesting that the functional effects of Aß-tau interactions reflect the temporal appearance of these molecules in individual brain areas.

Anticonvulsant effects of acetaminophen in mice: Comparison with the effects of nonsteroidal anti-inflammatory drugs.

OBJECTIVE: The appropriate use of analgesic drugs based on their degree of analgesia and adverse effects is important for pain management. Although it has been reported that AM404, a metabolite of acetaminophen, has anticonvulsant effects in several animal seizure models, little is known about the relation between acetaminophen and seizures. We therefore investigated the effects of acetaminophen on seizure susceptibility in several mouse seizure and epilepsy models and compared the effects with those of nonsteroidal anti-inflammatory drugs (NSAIDs). METHODS: Anticonvulsant activity was evaluated in ICR mice using maximum electroshock-induced seizure tests and acute pentylenetetrazol-induced seizure tests. Electrical kindling via corneal stimulation and pentylenetetrazol administration were used to establish animal kindling epilepsy models. Proconvulsive activity was examined using an electroconvulsive shock test with low-stimulus currents. RESULTS: Acetaminophen showed slight, but not statistically significant, anticonvulsant activity in both the maximum electroshock-induced seizure test (300-600mg/kg i.p.) and acute pentylenetetrazol-induced seizure test (100-600mg/kg i.p.). In contrast, acetaminophen exhibited significant anticonvulsant effects in corneal electroshock-kindled and pentylenetetrazol-kindled mice (ED50 values: 251 and 310mg/kg i.p., respectively). When the proconvulsive effects of NSAIDs were examined in the low-current electroconvulsive shock-induced seizure model, the nonselective cyclooxygenase (COX)-1 and COX-2 inhibitors indomethacin, diclofenac, and loxoprofen induced dose-dependent proconvulsant activity. Celecoxib, a COX-2 selective inhibitor, had no proconvulsant activity. CONCLUSION: These findings suggest that acetaminophen has a significant anticonvulsant effect and that its profile is completely different from that of NSAIDs.

Species-specific roles of sulfolipid metabolism in acclimation of photosynthetic microbes to sulfur-starvation stress.

Photosynthetic organisms utilize sulfate for the synthesis of sulfur-compounds including proteins and a sulfolipid, sulfoquinovosyl diacylglycerol. Upon ambient deficiency in sulfate, cells of a green alga, Chlamydomonas reinhardtii, degrade the chloroplast membrane sulfolipid to ensure an intracellular-sulfur source for necessary protein synthesis. Here, the effects of sulfate-starvation on the sulfolipid stability were investigated in another green alga, Chlorella kessleri, and two cyanobacteria, Synechocystis sp. PCC 6803 and Synechococcus elongatus PCC 7942. The results showed that sulfolipid degradation was induced only in C. kessleri, raising the possibility that this degradation ability was obtained not by cyanobacteria, but by eukaryotic algae during the evolution of photosynthetic organisms. Meanwhile, Synechococcus disruptants concerning sqdB and sqdX genes, which are involved in successive reactions in the sulfolipid synthesis pathway, were respectively characterized in cellular response to sulfate-starvation. Phycobilisome degradation intrinsic to Synechococcus, but not to Synechocystis, and cell growth under sulfate-starved conditions were repressed in the sqdB and sqdX disruptants, respectively, relative to in the wild type. Their distinct phenotypes, despite the common loss of the sulfolipid, inferred specific roles of sqdB and sqdX. This study demonstrated that sulfolipid metabolism might have been developed to enable species- or cyanobacterial-strain dependent processes for acclimation to sulfate-starvation.

Gangliosides and hearing.

Severe auditory impairment observed in GM3 synthase-deficient mice and humans indicates that glycosphingolipids, especially sialic-acid containing gangliosides, are indispensable for hearing. Gangliosides associate with glycoproteins to form membrane microdomains, the composition of which plays a special role in maintaining the structural and functional integrity of hair cells. These microdomains, also called lipid rafts, connect with intracellular signaling and cytoskeletal systems to link cellular responses to environmental cues. During development, ganglioside species are expressed in distinctive spatial and temporal patterns throughout the cochlea. In both mice and humans, blocking particular steps of ganglioside metabolism produces distinctive neurological and auditory phenotypes. Thus each ganglioside species may have specific, non-overlapping functions within the cochlea, central auditory network, and brain.

Modification of Tau by 8-Nitroguanosine 3',5'-Cyclic Monophosphate (8-Nitro-cGMP): EFFECTS OF NITRIC OXIDE-LINKED CHEMICAL MODIFICATION ON TAU AGGREGATION.

Neurofibrillar tangles caused by intracellular hyperphosphorylated tau inclusion and extracellular amyloid ß peptide deposition are hallmarks of Alzheimer's disease. Tau contains one or two cysteine residues in three or four repeats of the microtubule binding region following alternative splicing of exon 10, and formation of intermolecular cysteine disulfide bonds accelerates tau aggregation. 8-Nitroguanosine 3',5'-cyclic monophosphate (8-nitro-cGMP) acts as a novel second messenger of nitric oxide (NO) by covalently binding cGMP to cysteine residues by electrophilic properties, a process termed protein S-guanylation. Here we studied S-guanylation of tau and its effects on tau aggregation. 8-Nitro-cGMP exposure induced S-guanylation of tau both in vitro and in tau-overexpressed HEK293T cells. S-guanylated tau inhibited heparin-induced tau aggregation in a thioflavin T assay. Atomic force microscopy observations indicated that S-guanylated tau could not form tau granules and fibrils. Further biochemical analyses showed that S-guanylated tau was inhibited at the step of tau oligomer formation. In P301L tau-expressing Neuro2A cells, 8-nitro-cGMP treatment significantly reduced the amount of sarcosyl-insoluble tau. NO-linked chemical modification on cysteine residues of tau could block tau aggregation, and therefore, increasing 8-nitro-cGMP levels in the brain could become a potential therapeutic strategy for Alzheimer's disease.

Assessment of Pulp Regeneration Induced by Stem Cell Therapy by Magnetic Resonance Imaging.

INTRODUCTION: This study was designed to evaluate the usefulness of magnetic resonance imaging (MRI) to assess the regeneration of pulp tissue. METHODS: Mobilized dental pulp stem cells and granulocyte colony-stimulating factor with collagen were transplanted into mature pulpectomized teeth for pulp regeneration (n = 4). The controls consisted of pulpectomized teeth with or without collagen and normal teeth with intact pulp tissue (n = 4, each). The signal intensity (SI) of MRI using T2 sequences was compared after the extraction of teeth in dogs. MRI was correlated with the corresponding histologic findings. RESULTS: Pulp tissue was fully regenerated 90 days after cell transplantation. On the other hand, the root canal was empty in the control collagen-transplanted teeth at 90 days. The SI of the normal teeth was significantly higher than that of nonvital pulpectomized teeth and the controls of collagen transplanted teeth at 90 days. The stem cell transplanted teeth showed a gradual decrease in the SI until 180 days at which time the SI was similar to that in the normal teeth and significantly higher than that in the teeth transplanted with collagen alone without the stem cells. CONCLUSIONS: The changes in the SI of the pulplike tissue were consistent with the histologic findings, showing the potential usefulness of the noninvasive method to serially access the efficacy of pulp regenerative therapy.

Toxic tau oligomer formation blocked by capping of cysteine residues with 1,2-dihydroxybenzene groups.

Neurofibrillary tangles, composed of hyperphosphorylated tau fibrils, are a pathological hallmark of Alzheimer's disease; the neurofibrillary tangle load correlates strongly with clinical progression of the disease. A growing body of evidence indicates that tau oligomer formation precedes the appearance of neurofibrillary tangles and contributes to neuronal loss. Here we show that tau oligomer formation can be inhibited by compounds whose chemical backbone includes 1,2-dihydroxybenzene. Specifically, we demonstrate that 1,2-dihydroxybenzene-containing compounds bind to and cap cysteine residues of tau and prevent its aggregation by hindering interactions between tau molecules. Further, we show that orally administered DL-isoproterenol, an adrenergic receptor agonist whose skeleton includes 1,2-dihydroxybenzene and which penetrates the brain, reduces the levels of detergent-insoluble tau, neuronal loss and reverses neurofibrillary tangle-associated brain dysfunction. Thus, compounds that target the cysteine residues of tau may prove useful in halting the progression of Alzheimer's disease and other tauopathies.

Ganglioside GM3 is essential for the structural integrity and function of cochlear hair cells.

GM3 synthase (ST3GAL5) is the first biosynthetic enzyme of a- and b-series gangliosides. Patients with GM3 synthase deficiency suffer severe neurological disability and deafness. Eight children (ages 4.1 ± 2.3 years) homozygous for ST3GAL5 c.694C>T had no detectable GM3 (a-series) or GD3 (b-series) in plasma. Their auditory function was characterized by the absence of middle ear muscle reflexes, distortion product otoacoustic emissions and cochlear microphonics, as well as abnormal auditory brainstem responses and cortical auditory-evoked potentials. In St3gal5(-/-) mice, stereocilia of outer hair cells showed signs of degeneration as early as postnatal Day 3 (P3); thereafter, blebs devoid of actin or tubulin appeared at the region of vestigial kinocilia, suggesting impaired vesicular trafficking. Stereocilia of St3gal5(-/-) inner hair cells were fused by P17, and protein tyrosine phosphatase receptor Q, normally linked to myosin VI at the tapered base of stereocilia, was maldistributed along the cell membrane. B4galnt1(-/-) (GM2 synthase-deficient) mice expressing only GM3 and GD3 gangliosides had normal auditory structure and function. Thus, GM3-dependent membrane microdomains might be essential for the proper organization and maintenance of stereocilia in auditory hair cells.

Glycoconjugates in the mammalian auditory system.

Cell-surface glycoconjugates, such as proteoglycans, glycoproteins, and glycosphingolipids have been suggested to serve important functions in hearing because of their variety and their specific expression patterns during the development and maturation of cochlea. However, there has been no definitive proof regarding their involvement in auditory functions. In this study, we provide an overview of the expression of glycoconjugates in auditory systems and consider their possible involvement in hearing functions. We include our recent findings regarding deafness in ganglioside (sialic acid containing glycosphingolipids)-deficient mice, and address the importance of functional glycobiology in auditory systems.

Mice lacking ganglioside GM3 synthase exhibit complete hearing loss due to selective degeneration of the organ of Corti.

The ganglioside GM3 synthase (SAT-I), encoded by a single-copy gene, is a primary glycosyltransferase for the synthesis of complex gangliosides. In SAT-I null mice, hearing ability, assessed by brainstem auditory-evoked potentials (BAEP), was impaired at the onset of hearing and had been completely lost by 17 days after birth (P17), showing a deformity in hair cells in the organ of Corti. By 2 months of age, the organ of Corti had selectively and completely disappeared without effect on balance or motor function or in the histology of vestibule. Interestingly, spatiotemporal changes in localization of individual gangliosides, including GM3 and GT1b, were observed during the postnatal development and maturation of the normal inner ear. GM3 expressed in almost all regions of cochlea at P3, but at the onset of hearing it distinctly localized in stria vascularis, spiral ganglion, and the organ of Corti. In addition, SAT-I null mice maintain the function of stria vascularis, because normal potassium concentration and endocochlear potential of endolymph were observed even when they lost the BAEP completely. Thus, the defect of hearing ability of SAT-I null mice could be attributed to the functional disorganization of the organ of Corti, and the expression of gangliosides, especially GM3, during the early part of the functional maturation of the cochlea could be essential for the acquisition and maintenance of hearing function.