Chondrocyte Thrombomodulin Protects against Osteoarthritis.

Osteoarthritis (OA) is a prevalent form of arthritis that affects over 32.5 million adults worldwide, causing significant cartilage damage and disability. Unfortunately, there are currently no effective treatments for OA, highlighting the need for novel therapeutic approaches. Thrombomodulin (TM), a glycoprotein expressed by chondrocytes and other cell types, has an unknown role in OA. Here, we investigated the function of TM in chondrocytes and OA using various methods, including recombinant TM (rTM), transgenic mice lacking the TM lectin-like domain (TMLeD/LeD), and a microRNA (miRNA) antagomir that increased TM expression. Results showed that chondrocyte-expressed TM and soluble TM [sTM, like recombinant TM domain 1 to 3 (rTMD123)] enhanced cell growth and migration, blocked interleukin-1ß (IL-1ß)-mediated signaling and protected against knee function and bone integrity loss in an anterior cruciate ligament transection (ACLT)-induced mouse model of OA. Conversely, TMLeD/LeD mice exhibited accelerated knee function loss, while treatment with rTMD123 protected against cartilage loss even one-week post-surgery. The administration of an miRNA antagomir (miR-up-TM) also increased TM expression and protected against cartilage damage in the OA model. These findings suggested that chondrocyte TM plays a crucial role in counteracting OA, and miR-up-TM may represent a promising therapeutic approach to protect against cartilage-related disorders.

High-fat diet reduces novelty-induced expression of activity-regulated cytoskeleton-associated protein.

Chronic high-fat-diet (HFD) consumption can lead to the development of brain insulin resistance, which then exerts deleterious effects on learning and memory. Activity-regulated cytoskeleton-associated protein (Arc) is a memory-related protein, and its expression can be induced by insulin stimulation. In HFD-fed animals, their basal Arc protein levels in cerebral cortex and hippocampus are reduced. However, the effects of HFD on novelty-induced Arc protein expression that is important for cognitive function is still unknown. In the present study, after feeding HFD (60% kcal from fat) for 5 weeks, mice developed brain insulin resistance and had a significant reduction in the novelty-induced but not the basal Arc protein levels in their hippocampi. Further experiments were performed in primary rat hippocampal neurons. The results show that, under the condition of neuronal insulin resistance, acute insulin stimulation induced less activation of the phosphatidylinositol 3-kinase/protein kinase B/p70 ribosomal S6 kinase (PI3K/Akt/p70S6K) pathway, resulting in reduced induction of Arc protein expression. Accordingly, it is suggested that following HFD feeding, the reduction in novelty-induced Arc protein expression in animal's hippocampus is probably related to a suppressed activation of the PI3K/Akt/p70S6K pathway due to the existence of brain insulin resistance.

The Cholinergic Signaling Responsible for the Expression of a Memory-Related Protein in Primary Rat Cortical Neurons.

Cholinergic dysfunction in the brain is closely related to cognitive impairment including memory loss. In addition to the degeneration of basal forebrain cholinergic neurons, deficits in the cholinergic receptor signaling may also play an important role. In the present study, to examine the cholinergic signaling pathways responsible for the induction of a memory-related postsynaptic protein, a cholinergic agonist carbachol was used to induce the expression of activity-regulated cytoskeleton associated protein (Arc) in primary rat cortical neurons. After pretreating neurons with various antagonists or inhibitors, the levels of carbachol-induced Arc protein expression were detected by Western blot analysis. The results show that carbachol induces Arc protein expression mainly through activating M1 acetylcholine receptors and the downstream phospholipase C pathway, which may lead to the activation of the MAPK/ERK signaling pathway. Importantly, carbachol-mediated M2 receptor activation exerts negative effects on Arc protein expression and thus counteracts the enhanced effects of M1 activation. Furthermore, it is suggested for the first time that M1-mediated enhancement of N-methyl-D-aspartate receptor (NMDAR) responses, leading to Ca(2+) entry through NMDARs, contributes to carbachol-induced Arc protein expression. These findings reveal a more complete cholinergic signaling that is responsible for carbachol-induced Arc protein expression, and thus provide more information for developing treatments that can modulate cholinergic signaling and consequently alleviate cognitive impairment. J. Cell. Physiol. 231: 2428-2438, 2016. © 2016 Wiley Periodicals, Inc.

Nonlethal dose of silver nanoparticles attenuates TNF-α-induced hepatic epithelial cell death through HSP70 overexpression.

Silver nanoparticles (Ag-nps) have been widely used in various biomedical products. Compared with its hazardous effects extensively being studied, rare attention has been paid to the potential protective effect of Ag-nps to human health. The present study was designed to evaluate the protective effects of Ag-nps and heat shock treatment on tumor necrosis factor-α (TNF-α)-induced cell damage in Clone 9 cells. Clone 9 cells were pretreated with nonlethal concentration of Ag-nps (1 µg/ml) or heat shock, and then cell damages were induced by TNF-α (1 ng/ml). Protective effects of Ag-nps administration or heat shock treatment were determined by examining the TNF-α-induced changes in cell viabilities. The results showed that the intensity of cytotoxicity produced by TNF-α was alleviated upon treatment with nonlethal concentration of Ag-nps (1 µg/ml). Similar protective effects were also found upon heat shock treatment. These data demonstrate that Ag-nps and heat shock treatment were equally capable of inducing heat shock protein 70 (HSP70) protein expression in Clone 9 cells. The results suggest that clinically Ag-nps administration is a viable strategy to induce endogenous HSP70 expression instead of applying heat shock. In conclusion, our study for the first time provides evidence that Ag-nps may act as a viable alternative for HSP70 induction clinically.

Insulin can induce the expression of a memory-related synaptic protein through facilitating AMPA receptor endocytosis in rat cortical neurons.

Learning and memory depend on long-term synaptic plasticity including long-term potentiation (LTP) and depression (LTD). Activity-regulated cytoskeleton-associated protein (Arc) plays versatile roles in synaptic plasticity mainly through inducing F-actin formation, underlying consolidation of LTP, and promoting AMPA receptor (AMPAR) endocytosis, underlying LTD. Insulin can also induce LTD by facilitating the internalization of AMPARs. In neuroblastoma cells, insulin induced a dramatic increase in Arc mRNA and Arc protein levels, which may underlie the memory-enhancing action of insulin. Thus, a hypothesis was made that, in response to insulin, increased AMPAR endocytosis leads to enhanced Arc expression, and vice versa. Primary cultures of neonatal Sprague-Dawley rat cortical neurons were used. Using Western-blot analysis and immunofluorescent staining, our results reveal that inhibiting AMPAR-mediated responses with AMPAR antagonists significantly enhanced whereas blocking AMPAR endocytosis with various reagents significantly prevented insulin (200 nM, 2 h)-induced Arc expression. Furthermore, via surface biotinylation assay, we demonstrate that acute blockade of new Arc synthesis after insulin stimulation using Arc antisense oligodeoxynucleotide prevented insulin-stimulated AMPAR endocytosis. These findings suggest for the first time that an interaction exists between insulin-stimulated AMPAR endocytosis and insulin-induced Arc expression.

Motor skill learning enhances the expression of activity-regulated cytoskeleton-associated protein in the rat cerebellum.

Motor skill learning is essential for environmental adaptations during everyday life. It has been shown that the cerebellum plays an important role in both the adaptation of eye movements and the motor skill learning. However, the neuronal substrates responsible for consolidation of complex motor skills rather than simple reflexes are still uncertain. Because the induction of immediate-early genes activity-regulated cytoskeleton-associated protein (Arc) and zinc finger binding protein clone 268 (Zif268) has been regarded as a marker for recent neuronal activity, therefore, in the present study, a rat paradigm of motor skill learning was used to investigate the protein expression of Arc and zif268 in the cerebellum after motor skill learning. Rats were trained to traverse the runway apparatus for 5 days. Protein samples were collected from the cerebellar cortices 1 hour after the training on days 1, 3, and 5, and analyzed by western blotting. The results showed that the expression of Arc, but not zif268, was significantly increased in the cerebellum following motor skill learning. These findings suggest that motor skill learning induces Arc expression in the cerebellum, which may play a role in acquiring complex motor skills.

Both PKMζ and KIBRA are closely related to reference memory but not working memory in a T-maze task in rats.

Protein kinase M zeta (PKMζ) and the kidney and brain protein (KIBRA) play important roles in various forms of memories. However, whether they are involved in performing the T-maze task is still unknown. In this study, the delayed nonmatch-to-sample (DNMS) task in a T-maze was given to rats. The percentage of correct choices denoting the performance accuracy was calculated and the protein levels of PKMζ and KIBRA in rat's prefrontal cortex were measured. The results showed significantly increased performance accuracy after the training phase, which was maintained on the next day in groups with a delay of 10 s but not 30 s, indicating that 30 s is too long for rats to maintain working memory. As for the expressions of PKMζ and KIBRA, significant increases were observed 1 day after the training phase, indicating that the formation of reference memory accompanies an increase in PKMζ and KIBRA. No significant difference was found among groups with various delay intervals, indicating that the expressions of PKMζ and KIBRA exert no effects on the performance of working memory. These results provide the first evidence that KIBRA as well as PKMζ is closely related to reference memory but not working memory in rats.

Exercise prevents the increased anxiety-like behavior in lactational di-(2-ethylhexyl) phthalate-exposed female rats in late adolescence by improving the regulation of hypothalamus-pituitary-adrenal axis.

Both the detrimental effects of early life adversity and the beneficial effects of exercise on the hypothalamic-pituitary-adrenal (HPA) axis have been reported. Early life exposure to di-(2-ethylhexyl)-phthalate (DEHP) may impair the development of endocrine system. In this study, we investigated the effects of lactational DEHP exposure on stress responses in late adolescent female rats and examined the protective role of treadmill running. Sprague-Dawley dams were fed with DEHP (10mg/kg per day) or vehicle during lactation. After weaning, the female offspring rats were trained to exercise on a treadmill for 5 weeks and then stressed by exploring on an elevated plus maze. The activities of HPA axis were evaluated by measuring the plasma levels of ACTH and corticosterone, the expressions of adrenal enzymes cholesterol side-chain cleavage enzyme (CYP11A1) and cytochrome P-450 11ß-hydroxylase (CYP11B1), and the expression of hypothalamic glucocorticoid receptors (GR). The results demonstrate that DEHP-exposed rats exhibited enhanced anxiety-like behaviors. Increased hypothalamic GR and plasma ACTH levels, but decreased adrenal CYP11A1 and corticosterone levels, were observed in DEHP-exposed animals under stressed condition. Importantly, in DEHP-exposed animals, exercise during childhood-adolescence reduced anxiety-like behaviors by normalizing stress-induced alterations in ACTH level and adrenal CYP11A1 expression. The findings of this study suggest that treadmill running may provide beneficial effects on ameliorating the dysregulation of HPA axis in lactational DEHP-exposed adolescent female rats.

Baicalein, an active component of Scutellaria baicalensis Georgi, prevents lysophosphatidylcholine-induced cardiac injury by reducing reactive oxygen species production, calcium overload and apoptosis via MAPK pathways.

BACKGROUND: Lysophosphatidylcholine (lysoPC), a metabolite from membrane phospholipids, accumulates in the ischemic myocardium and plays an important role in the development of myocardial dysfunction ventricular arrhythmia. In this study, we investigated if baicalein, a major component of Huang Qui, can protect against lysoPC-induced cytotoxicity in rat H9c2 embryonic cardiomyocytes. METHODS: Cell viability was detected by the MTT assay; ROS levels were assessed using DCFH-DA; and intracellular free calcium concentrations were assayed by spectrofluorophotometer. Cell apoptosis and necrosis were evaluated by the flow cytometry assay and Hoechst staining. Mitogen-Activated Protein Kinases (MAPKs), which included the ERK, JNK, and p38, and the apoptotic mechanisms including Bcl-2/Bax, caspase-3, caspase-9 and cytochrome c pathways were examined by Western blot analysis. The activation of MAPKs was examined by enzyme-linked immunosorbent assay. RESULTS: We found that lysoPC induced death and apoptosis of H9c2 cells in a dose-dependent manner. Baicalein could prevent lysoPC-induced cell death, production of reactive oxygen species (ROS), and increase of intracellular calcium concentration in H9c2 cardiomyoctes. In addition, baicalein also inhibited lysoPC-induced apoptosis, with associated decreased pro-apoptotic Bax protein, increased anti-apoptotic Bcl-2 protein, resulting in an increase in the Bcl-2/Bax ratio. Finally, baicalein attenuated lysoPC-induced the expression of cytochrome c, casapase-3, casapase-9, and the phosphorylations of ERK1/2, JNK, and p38. LysoPC-induced ERK1/2, JNK, and p38 activations were inhibited by baicalein. CONCLUSIONS: Baicalein protects cardiomyocytes from lysoPC-induced apoptosis by reducing ROS production, inhibition of calcium overload, and deactivations of MAPK signaling pathways.

Histone deacetylase inhibitor attenuates the effects of 27-hydroxycholesterol on the rat brain.

Hypercholesterolemia is a risk factor for Alzheimer's disease (AD). Plasma cholesterol does not pass the blood-brain barrier whereas its metabolite 27-hydroxycholesterol (27-OHC) can enter the brain. High 27-OHC in the brain has been suggested to mediate hypercholesterolemia-induced impairments of learning and memory through promoting amyloid-ß accumulation and facilitating synaptic disruption. In AD brains, the activity of histone deacetylase (HDAC) is elevated. Treating AD animals with HDAC inhibitors decreases amyloid-ß levels and synaptic damages, which leads to memory improvement. Whether HDAC activity is involved in the actions of 27-OHC is still uncertain. In this study, 4 weekly injections of 27-OHC/vehicle were given to rats followed by 3 daily injections of HDAC inhibitor trichostatin (TSA)/vehicle. The results of Morris water maze test reveal that all rats have intact spatial learning ability during the 5-d training phase. However, the behavioral performance during the probe trial was impaired by 27-OHC treatment, which was improved by adding TSA treatments. Furthermore, 27-OHC treatments reduced the hippocampal levels of acetylated histone H3, acetylated α tubulin, insulin-degrading enzyme and postsynaptic protein PSD-95, indicating that 27-OHC treatments may induce enhanced HDAC activity, decreased amyloid-ß clearance and synaptic disruption. All reduced levels returned to the basal levels by adding TSA treatments. These findings support our hypothesis that HDAC activity is enhanced following long-term exposure to excess 27-OHC.

Enhanced expression of activity-regulated cytoskeleton-associated protein in the medial prefrontal cortex is involved in working memory performance.

Working memory (WM) is a cognitive function important for guiding the on-going or upcoming behavior. A memory-related protein Arc (activity-regulated cytoskeleton-associated protein) is implicated in long-term memory consolidation. Recent evidence further suggests the involvement of hippocampal Arc in spatial WM. The medial prefrontal cortex (mPFC) is a key brain region mediating WM. However, the role of mPFC Arc in WM is still uncertain. To investigate whether mPFC Arc protein is involved in WM performance, delayed non-match to sample (DNMS) T-maze task was performed in rats with or without blocking new synthesis of mPFC Arc. In DNMS task, a 10-s or 30-s delay between the sample run and the choice run was given to evaluate WM performance. To block new Arc protein synthesis during the DNMS task, Arc antisense oligodeoxynucleotides (ODNs) were injected to the bilateral mPFC. The results show that, in rats without surgery for cannula implantation and subsequent intracerebral injection of ODNs, WM was functioning well during the DNMS task with a delay of 10 s but not 30 s, which was accompanied with a significantly increased level of mPFC Arc protein, indicating a possible link between enhanced Arc protein expression and the performance of WM. After preventing the enhancement of mPFC Arc protein expression with Arc antisense ODNs, rat's WM performance was impaired. These findings support enhanced mPFC Arc protein expression playing a role during WM performance.

Amyloid-beta interrupts the PI3K-Akt-mTOR signaling pathway that could be involved in brain-derived neurotrophic factor-induced Arc expression in rat cortical neurons.

The deposition of amyloid-beta (Abeta) contributes to the pathogenesis of Alzheimer's disease. Even at low levels, Abeta may interfere with various signaling cascades critical for the synaptic plasticity that underlies learning and memory. Brain-derived neurotrophic factor (BDNF) is well known to be capable of inducing the synthesis of activity-regulated cytoskeleton-associated protein (Arc), which plays a fundamental role in modulating synaptic plasticity. Our recent study has demonstrated that treatment of fibrillar Abeta at a nonlethal level was sufficient to impair BDNF-induced Arc expression in cultured rat cortical neurons. In this study, BDNF treatment alone induced the activation of the phosphatidylinositol 3-kinase-Akt-mammlian target of rapamycin (PI3K-Akt-mTOR) signaling pathway, the phosphorylation of eukaryotic initiation factor 4E binding protein (4EBP1) and p70 ribosomal S6 kinase (p70S6K), the dephosphorylation of eukaryotic elongation factor 2 (eEF2), and the expression of Arc. Interrupting the PI3K-Akt-mTOR signaling pathway by inhibitors prevented the effects of BDNF, indicating the involvement of this pathway in BDNF-induced 4EBP1 phosphorylation, p70S6K phosphorylation, eEF2 dephosphorylation, and Arc expression. Nonlethal Abeta pretreatment partially blocked these effects of BDNF. Double- immunofluorescent staining in rat cortical neurons further confirmed the coexistence of eEF2 dephosphorylation and Arc expression following BDNF treatment regardless of the presence of Abeta. These results reveal that, in cultured rat cortical neurons, Abeta interrupts the PI3K-Akt-mTOR signaling pathway that could be involved in BDNF-induced Arc expression. Moreover, this study also provides the first evidence that there is a close correlation between BDNF-induced eEF2 dephosphorylation and BDNF-induced Arc expression. (c) 2009 Wiley-Liss, Inc.

Functional changes on ascending auditory pathway in rats caused by germanium dioxide exposure: an electrophysiological study.

The semiconductor element, germanium (Ge), is essential for the manufacture of modern integrated circuits. Because of its anti-tumor and immunomodulative effects, Ge-containing compounds are also used as health-promoting ingredients in food. However, some histological studies have shown the toxic effects of Ge-containing compounds on various organs, including the central nervous system. Even now, the effect of germanium on auditory system function is not completely clear. To clarify this question, brainstem auditory evoked potentials (BAEPs) were applied to examine the effect of germanium dioxide (GeO(2)) on the ascending auditory pathway. Since the voltage-gated sodium channel is important to neuron activation and nerve conduction, the effect of GeO(2) on voltage-gated sodium channels was also examined. The result revealed GeO(2) elevated the BAEPs threshold dose-dependently. GeO(2) also prolonged latencies and interpeak latencies (IPLs) of BAEPs, but the amplitudes of suprathreshold intensities (90dB) did not show any obvious change. In addition, the results of whole cell patch clamp studies indicated GeO(2) reduced inward sodium current. These results suggest the toxic effect of GeO(2) on the conduction of the auditory system, and that inhibitory effect of GeO(2) on the voltage-gated sodium channels might play a role in GeO(2)-induced abnormal hearing loss.

Modifier of cell adhesion regulates N-cadherin-mediated cell-cell adhesion and neurite outgrowth.

Modifier of cell adhesion (MOCA) is a member of the dedicator of cytokinesis 180 family of proteins and is highly expressed in CNS neurons. MOCA is associated with Alzheimer's disease tangles and regulates the accumulation of amyloid precursor protein and beta-amyloid. Here, we report that MOCA modulates cell-cell adhesion and morphology. MOCA increases the accumulation of adherens junction proteins, including N-cadherin and beta-catenin, whereas reducing endogenous MOCA expression lowers cell-cell aggregation and N-cadherin expression. MOCA colocalizes with N-cadherin and actin in areas of cell-cell and cell substratum contact and is expressed in neuronal processes. MOCA accumulates during neuronal differentiation, and its expression enhances NGF-induced neurite outgrowth and morphological complexity. We conclude that MOCA regulates N-cadherin-mediated cell-cell adhesion and neurite outgrowth.

Amyloid-beta at sublethal level impairs BDNF-induced arc expression in cortical neurons.

Alzheimer's disease (AD) is characterized by progressive memory loss and cognitive dysfunction that probably due to a deficit in synaptic plasticity. One member of neurotrophins, brain-derived neurotrophic factor (BDNF), is known to be involved in the hippocampal long-term potentiation (LTP), a cellular model for learning and memory. Moreover, activity-regulated cytoskeleton-associated gene (Arc), an immediate early gene, is found to be a downstream effector of the BDNF signaling cascade. Inhibition of Arc protein synthesis impairs both the maintenance of LTP and the consolidation of long-term memory. In addition, the formation of senile plaques is a pathological feature in AD and mainly consists of the deposition of amyloid-beta (Abeta), a proteolytic product of amyloid precursor protein. Several studies concerning neurobehavioral performance have suggested that Abeta at sublethal levels interfere with the signaling cascades critical for synaptic plasticity and thus lead to the cognitive impairment in early stage of AD. Whether the BDNF-mediated Arc synthesis is impaired by sublethal Abeta in early AD is still unclear. Therefore, in the present study, primary cultures of neonatal rat cortical neurons were used to evaluate the effect of sublethal Abeta on the BDNF-induced Arc protein expression. Consistent with the literature, Arc, an indicator of synaptic plasticity, was induced by BDNF (25 ng/ml) in both dose- and time-dependent manners. After treating cultures with sublethal Abeta (5 microM), a significant suppression was observed on the level of BDNF-induced Arc protein expression. This result indicates that Abeta at sublethal level impairs the BDNF-mediated signaling in cortical neurons and thus underlies the deficits of synaptic plasticity occurred at the early stage of AD before significant neuronal loss.

Germanium dioxide induces mitochondria-mediated apoptosis in Neuro-2A cells.

Germanium (Ge) is commonly used in the semiconductor industry as well as health-promoting and medical field. Biologically, germanium possesses erythropoietic, anti-microbial, anti-tumor, anti-amyloidosis, and immunomodulative effects. However, toxic effects of Ge-containing compounds on kidney, muscle, neuronal cells, and nerves have been reported. Mitochondrial dysfunction was found to be involved in the pathogenesis of GeO(2)-induced nephropathy and myopathy. Since it is well known that mitochondria play a major role in apoptosis triggered by many stimuli, an effort was made to examine whether the Ge-induced neurotoxicity occurs through mitochondria-mediated apoptosis. A mouse neuroblastoma cell line, Neuro-2A, was used in the present study. After incubating with 0.1-800microM of GeO(2) for 0-72h, the cell viability of Neuro-2A cells was inhibited in a dose- and time-dependent manner. Further analysis showed that aside from the changes in the nuclear morphology responsible for apoptosis, the release of cytochrome c, the loss of mitochondrial membrane potential, the translocation of Bax, and the reduction of Bcl-2 expression were also observed in Neuro-2A cells after GeO(2) treatment. These results indicate that the mitochondria-mediated apoptosis is involved in this in vitro model of GeO(2)-induced neurotoxicity.

Lead-induced attenuation in the aggregation of acetylcholine receptors during the neuromuscular junction formation.

Lead (Pb2+) toxicity is more common in children and is associated with cognitive deficits, which may reflect lead-induced changes in central synaptic development and function. Aside from neurotoxicity, lead exposure may also impact mature neuromuscular junction (NMJ) and cause muscle weakness. NMJ is known as a peripheral cholinergic synapse and its signaling cascades responsible for development are similar to those for the central synapses. However, the effect of lead exposure on the formation of NMJ in mammals is unclear. In the present study, a NG108-15/C2C12 coculture model was used to measure the acetylcholine receptor (AChR) aggregates formed on the myotubes which was an early hallmark for the NMJ formation. AChR aggregates were identified by alpha-bungarotoxin under fluorescent microscope. Single dose of lead acetate with final concentrations at 10(-3), 10(-1), or 10 microM was applied to dishes at the beginning of coculturing. Following 3-day exposure, although NG108-15 cells could extend long neurites to nearby myotubes, obvious dose-dependent attenuation in AChR aggregation was shown. The averaged area of an AChR aggregate, the averaged number of AChR aggregates per myotube, and the total area of AChR aggregates per myotube were all significantly decreased. In addition, the distribution percentages of various sizes of AChR aggregates showed that almost half of the AChR aggregates were formed with a size of 2-5 microm2 regardless of lead exposure. After treating 10 microM of lead acetate, significantly more AChR aggregates ranged from 2 to 20 microm2 were formed and significantly less AChR aggregates larger than 20 microm2 were formed. These results indicated that lead exposure reduced the extent of AChR aggregation concerning both the size and number of AChR aggregates and large AChR aggregates could hardly be formed after acute high-level lead exposure. No significant change was found in the total amount of AChRs on the myotubes after lead exposure, which indicated that the attenuation of AChR aggregation was not caused by reducing the synthesis of AChRs but by remaining dispersed pattern of AChRs on the myotubes. These data suggest that lead exposure exerts detrimental effects on the formation of NMJ.

Neurobehavioral changes in Taiwanese lead-exposed workers.

OBJECTIVE: The dysfunction of the central nervous system (CNS) caused by lead exposure was evaluated. METHODS: Workers who have been exposed to lead for at least 3 years and have been informed of having elevated blood lead levels (BLLs) were recruited. According to their current BLLs, 33 and 28 males were assigned to the medium (40-80 microg/dL) and low (<40 microg/dL) BLLs groups, respectively. Sixty-two nonexposed healthy men served as the control group. Their neurobehaviors were examined by a computerized evaluation system. RESULTS: Significantly impaired neurobehavioral functions were shown in the medium BLLs group, which included slow performance of psychomotor tasks, impaired processing of visual-spatial information, reduced memory and learning functions, low performance accuracy, slow execution of responses, and poor attentional control. CONCLUSIONS: Subtle CNS dysfunction could be detected from lead-exposed workers who have no obvious neurologic and cognitive deficits.

Implication of nNOS in the enlargement of AChR aggregates but not the initial aggregate formation in a novel coculture model.

The aggregation of nicotinic acetylcholine receptors (AChRs) is an early hallmark of the formation of neuromuscular junction (NMJ), and nitric oxide is recently known to play an important role. In many NMJ studies, nerve-muscle coculture model was used, and NG108-15 cells, a neuroblastoma x glioma hybrid cell line, were the most frequently used nerve cells. However, possible contributions from glial cells could not be excluded. In this study, Neuro-2a neuroblastoma cells were used instead of [corrected] coculture with myotubes, and the relationship between AChR aggregation and spatiotemporal expression and activation of nNOS (neuronal nitric oxide synthase) was examined. Upon coculture, AChR aggregates were observed by FITC-conjugated alpha-bungarotoxin, and double labeling of AChRs and neurofilament showed that the neurites of a Neuro-2a cell innervated several myotubes. After treating the cocultures with single dose of L-NAME at the end of 1-day [corrected] coculturing, only slight effect on AChR aggregation could be found indicating that nNOS is not related to the initial formation of AChR aggregates. In contrast, when L-NAME treatment was given at the end of a 3-day coculturing, the day just before reaching the maximum extent of AChR aggregation, new AChR aggregates were hardly formed and the preformed AChR aggregates were even dispersed indicating that the enlargement of AChR aggregates is highly dependent on the nNOS activity. Double-labeling study of nNOS and AChR further showed that the coupling of membranous nNOS to regions nearby the AChR aggregates was essential for the enlargement of AChR aggregates. These results not only revealed the spatiotemporal relationship between AChR aggregation and nNOS activity but also verified the feasibility and usefulness of using Neuro-2a cells in a coculture model.

Increased vulnerability of auditory system to noise exposure in mdx mice.

OBJECTIVES: Dystrophin is a cytoskeletal protein mainly found just beneath the sarcolemma. Lack of dystrophin is known to be the cause of Duchenne muscular dystrophy (DMD). Other tissues, including the brain, retina, and cochlear hair cells, also express dystrophin. Recently, a gene (Xp21.2) associated with sensorineural hearing impairment has been mapped within the localization site for dystrophin in two families. Thus, it is reasonable to assume that dystrophin may play a role in auditory function. However, animal studies have produced conflicting results. STUDY DESIGN: An attempt was made to clarify the differences between the auditory systems of dystrophin-deficient mdx mice and control B-10 mice by exposure to noise. METHODS: In the present study, mdx mice and B-10 mice were used. Animals were exposed daily to noise for 1 month, and their auditory functions were evaluated by recording the brainstem auditory evoked potentials (BAEPs). RESULTS: Before noise exposure, the mdx mouse demonstrated normal BAEP threshold when compared with the B-10 mouse. After 1 month of noise exposure, the B-10 mouse showed no apparent change in hearing threshold and BAEP latencies. In contrast, significantly increased hearing threshold and prolonged BAEP peak and interpeak latencies were observed in the mdx mouse after noise exposure. CONCLUSIONS: These results indicate that the mdx mice are more vulnerable to noise damage. This involves not only the peripheral auditory system, but also the brainstem central auditory pathway. Therefore, a significant role for dystrophin in the auditory system, especially under noise stress, is suggested.