Chromosomal editing of Corynebacterium glutamicum ATCC 13032 to produce gamma-aminobutyric acid.

Corynebacterium glutamicum has been used as a sustainable microbial producer for various bioproducts using cheap biomass resources. In this study, a high GABA-producing C. glutamicum strain was constructed by chromosomal editing. Lactobacillus brevis-derived gadB2 was introduced into the chromosome of C. glutamicum ATCC 13032 to produce gamma-aminobutyric acid and simultaneously blocked the biosynthesis of lactate and acetate. GABA transport and degradation in C. glutamicum were also blocked to improve GABA production. As precursor of GABA, l-glutamic acid synthesis in C. glutamicum was enhanced by introducing E. coli gdhA encoding glutamic dehydrogenase, and the copy numbers of gdhA and gadB2 were also optimized for higher GABA production. The final C. glutamicum strain CGY705 could produce 33.17 g/L GABA from glucose in a 2.4-L bioreactor after 78 h fed-batch fermentation. Since all deletion and expression of genes were performed using chromosomal editing, fermentation of the GABA-producing strains constructed in this study does not need supplementation of any antibiotics and inducers. The strategy used in this study has potential value in the development of GABA-producing bacteria.

Sponging of glutamate at the outer plasma membrane surface reveals roles for glutamate in development.

Plants use electrical and chemical signals for systemic communication. Herbivory, for instance, appears to trigger local apoplasmic glutamate accumulation, systemic electrical signals, and calcium waves that travel to report insect damage to neighboring leaves and initiate defense. To monitor extra- and intracellular glutamate concentrations in plants, we generated Arabidopsis lines expressing genetically encoded fluorescent glutamate sensors. In contrast to cytosolically localized sensors, extracellularly displayed variants inhibited plant growth and proper development. Phenotypic analyses of high-affinity display sensor lines revealed that root meristem development, particularly the quiescent center, number of lateral roots, vegetative growth, and floral architecture were impacted. Notably, the severity of the phenotypes was positively correlated with the affinity of the display sensors, intimating that their ability to sequester glutamate at the surface of the plasma membrane was responsible for the defects. Root growth defects were suppressed by supplementing culture media with low levels of glutamate. Together, the data indicate that sequestration of glutamate at the cell surface either disrupts the supply of glutamate to meristematic cells and/or impairs localized glutamatergic signaling important for developmental processes.

Decitabine Enhances Acute Myeloid Leukemia Cell Apoptosis through SH3BGRL Upregulation.

BACKGROUND: SH3-domain-binding glutamic acid-rich protein-like protein (SH3BGRL) is downregulated in acute myeloid leukemia (AML). Clinically, DNA demethylating drug decitabine (DAC) combined with traditional chemotherapies reveals better efficacy on AML patients than the conventional chemotherapies alone. Our previous results revealed that human SH3-domain-binding glutamic acid-rich protein-like protein (SH3BGRL) plays a tumor suppressive role in AML but whether there is a connection between DAC and SH3BGRL expression remains elusive. METHODS: Here, we tentatively treated AML cell lines U937, MV4, and HL-60 with DAC and Western Blots, RT-PCR was used to detect the expression of SH3BGRL. Cell proliferation and apoptosis were determined using Annexin V/7- AAD staining. Real-time RT-PCR and Western blot were used to determine the expression of SH3BGRL mRNA and protein. Methylation-specific PCR was used to quantify the DNA methylation in AML cell lines. RESULTS: DAC had cytotoxicity in HL-60, MV4, and U937. In U937 cell lines, treatment with DAC showed the upregulation of cleaved caspase3, PARP, and SH3BGRL. Upon treatment, up-regulation of SH3BGRL mRNA and protein was dose-dependent and this activity was partially inhibited in endogenous SH3BGRL knockdown cell lines. CONCLUSION: Thus, our results demonstrated a possibly cytotoxic role of DAC on AML cells by upregulation of SH3BGRL expression at epigenetic modulation level and the methylation status in the SH3BGRL promoter region could be a supplemental diagnostic marker to the precise administration of DAC to AML patients.

Localized Prediction of Glutamate from Whole-Brain Functional Connectivity of the Pregenual Anterior Cingulate Cortex.

Local measures of neurotransmitters provide crucial insights into neurobiological changes underlying altered functional connectivity in psychiatric disorders. However, noninvasive neuroimaging techniques such as magnetic resonance spectroscopy (MRS) may cover anatomically and functionally distinct areas, such as p32 and p24 of the pregenual anterior cingulate cortex (pgACC). Here, we aimed to overcome this low spatial specificity of MRS by predicting local glutamate and GABA based on functional characteristics and neuroanatomy in a sample of 88 human participants (35 females), using complementary machine learning approaches. Functional connectivity profiles of pgACC area p32 predicted pgACC glutamate better than chance (R2 = 0.324) and explained more variance compared with area p24 using both elastic net and partial least-squares regression. In contrast, GABA could not be robustly predicted. To summarize, machine learning helps exploit the high resolution of fMRI to improve the interpretation of local neurometabolism. Our augmented multimodal imaging analysis can deliver novel insights into neurobiology by using complementary information.SIGNIFICANCE STATEMENT Magnetic resonance spectroscopy (MRS) measures local glutamate and GABA noninvasively. However, conventional MRS requires large voxels compared with fMRI, because of its inherently low signal-to-noise ratio. Consequently, a single MRS voxel may cover areas with distinct cytoarchitecture. In the largest multimodal 7 tesla machine learning study to date, we overcome this limitation by capitalizing on the spatial resolution of fMRI to predict local neurotransmitters in the PFC. Critically, we found that prefrontal glutamate could be robustly and exclusively predicted from the functional connectivity fingerprint of one of two anatomically and functionally defined areas that form the pregenual anterior cingulate cortex. Our approach provides greater spatial specificity on neurotransmitter levels, potentially improving the understanding of altered functional connectivity in mental disorders.

Glutamatergic function in a genetic high-risk group for psychosis: A proton magnetic resonance spectroscopy study in individuals with 22q11.2 deletion.

Glutamatergic dysregulation is one of the leading theories regarding the pathoaetiolopy of schizophrenia. Meta-analysis of magnetic resonance spectroscopy studies in schizophrenia shows increased levels of glutamate and glutamine (Glx) in the medial frontal cortex and basal ganglia in clinical high-risk groups for psychosis and increased glutamine levels in the thalamus, but it is unclear if this is also the case in people at genetic high risk for psychosis. The aim of this study was to investigate glutamatergic function in the anterior cingulate cortex, striatum and thalamus in carriers of a genetic variant (22q11.2 deletion) associated with a high risk for psychosis. 53 volunteers (23 22q11.2 deletion carriers and 30 controls) underwent proton magnetic resonance spectroscopy imaging and neuropsychological assessments for prodromal psychotic symptoms, schizotypy, anxiety, depression and FSIQ. We did not find any difference between groups in Glx in the anterior cingulate cortex, striatum or thalamus (Glx: t(50)=-1.26, p = 0.21; U = 251, z = -0.7, p = 0.49; U = 316, z= -0.26, p = 0.79, respectively). No correlation was detected between Glx levels in any region and symptomatology or FSIQ. Our findings indicate that glutamatergic function is not altered in people at genetic high risk of psychosis due to the 22q11.2 deletion, which could suggest that this is not the mechanism underlying psychosis risk in 22q11.2 deletion carriers.

Inhibition of Anaplerotic Glutaminolysis Underlies Selenite Toxicity in Human Lung Cancer.

Large clinical trials and model systems studies suggest that the chemical form of selenium dictates chemopreventive and chemotherapeutic efficacy. Selenite induces excess ROS production, which mediates autophagy and eventual cell death in non-small cell lung cancer adenocarcinoma A549 cells. As the mechanisms underlying these phenotypic effects are unclear, the clinical relevance of selenite for cancer therapy remains to be determined. The authors' previous stable isotope-resolved metabolomics and gene expression analysis showed that selenite disrupts glycolysis, the Krebs cycle, and polyamine metabolism in A549 cells, potentially through perturbed glutaminolysis, a vital anaplerotic process for proliferation of many cancer cells. Herein, the role of the glutaminolytic enzyme glutaminase 1 (GLS1) in selenite's toxicity in A549 cells and in patient-derived lung cancer tissues is investigated. Using [13 C6 ]-glucose and [13 C5 ,15 N2 ]-glutamine tracers, selenite's action on metabolic networks is determined. Selenite inhibits glutaminolysis and glutathione synthesis by suppressing GLS1 expression, and blocks the Krebs cycle, but transiently activates pyruvate carboxylase activity. Glutamate supplementation partially rescues these anti-proliferative and oxidative stress activities. Similar metabolic perturbations and necrosis are observed in selenite-treated human patients' cancerous lung tissues ex vivo. The results support the hypothesis that GLS1 suppression mediates part of the anti-cancer activity of selenite both in vitro and ex vivo.

A Systematic Review and Meta-Analysis of the Diagnostic Performance of BRAF V600E Immunohistochemistry in Thyroid Histopathology.

Immunohistochemistry (IHC) in evaluating thyroid surgical specimens may facilitate diagnostic and prognostic evaluation, with potential therapeutic implications. We performed a systematic review and meta-analysis examining the analytic validity of IHC in detecting BRAFV600E mutations in thyroid cancer (primary or metastatic). We screened citations from three electronic databases (until December 20, 2018), supplemented by a hand search of authors' files and cross-references of reviews. Citations and full-text papers were independently reviewed in duplicate, and consensus was achieved on inclusion of papers. Two reviewers independently critically appraised and abstracted data from included papers. Random-effect meta-analyses were conducted for sensitivity and specificity estimates. We reviewed 1499 unique citations and 93 full-text articles. We included 1 systematic review and 30 original articles. The published review (from 2015) needed to be updated as there were multiple subsequent original studies. The pooled sensitivity of IHC in detecting a BRAFV600E mutation was 96.8% (95% confidence interval [CI] at 94.1%, 98.3%) (29 studies, including 2659 BRAFV600E mutant tumors). The IHC pooled specificity was 86.3% (95% CI 80.7%, 90.4%) (28 studies, including 1107 BRAFV600E wild-type specimens). These meta-analyses were subject to statistically significant heterogeneity, partly explained by antibody type (sensitivity and specificity) and tissue/tumor type (specificity). In conclusion, BRAF IHC is highly sensitive and reasonably specific in detecting the BRAFV600E mutation; however, there is some variability in analytic performance.

Membralin deficiency dysregulates astrocytic glutamate homeostasis leading to ALS-like impairment.

Mechanisms underlying motor neuron degeneration in amyotrophic lateral sclerosis (ALS) are yet unclear. Specific deletion of the ER-component membralin in astrocytes manifested postnatal motor defects and lethality in mice, causing the accumulation of extracellular glutamate through reducing the glutamate transporter EAAT2. Restoring EAAT2 levels in membralin KO astrocytes limited astrocyte-dependent excitotoxicity in motor neurons. Transcriptomic profiles from mouse astrocytic membralin KO motor cortex indicated significant perturbation in KEGG pathway components related to ALS, including downregulation of Eaat2 and upregulation of Tnfrsf1a. Changes in gene expression with membralin deletion also overlapped with mouse ALS models and reactive astrocytes. Our results shown that activation of TNF receptor (TNFR1)-NFκB pathway known to suppress Eaat2 transcription was upregulated with membralin deletion. Further, reduced membralin and EAAT2 levels correlated with disease progression in spinal cord from SOD1-mutant mouse models, and reductions in membralin/EAAT2 were observed in human ALS spinal cord. Importantly, overexpression of membralin in SOD1G93A astrocytes decreased TNFR1 levels and increased EAAT2 expression, and improved motor neuron survival. Importantly, upregulation of membralin in SOD1G93A mice significantly prolonged mouse survival. Together, our study provided a mechanism for ALS pathogenesis where membralin limited glutamatergic neurotoxicity, suggesting that modulating membralin had potentials in ALS therapy.

Re-evaluation of cyanophycin synthesis in Corynebacterium glutamicum and incorporation of glutamic acid and lysine into the polymer.

Corynebacterium glutamicum was only examined in the early 2000s as a possible microorganism for the production of the polyamide cyanophycin (multi-L-arginyl-poly-[L-aspartic acid], CGP). CGP is a potential precursor for the synthesis of polyaspartic acid and CGP-derived dipeptides which may be of use in peptide-based clinical diets, as dietary supplements, or in livestock feeds. In the past, C. glutamicum was disregarded for CGP production due to low CGP contents and difficulties in isolating the polymer. However, considering recent advances in CGP research, the capabilities of this organism were revisited. In this study, several cyanophycin synthetases (CphA) as well as expression vectors and cultivation conditions were evaluated. The ability of C. glutamicum to incorporate additional amino acids such as lysine and glutamic acid was also examined. The strains C. glutamicum pVWEx1::cphAΔ1 and C. glutamicum pVWEx1::cphABP1 accumulated up to 14% of their dry weight CGP, including soluble CGP containing more than 40 mol% of the alternative side-chain amino acid lysine. The soluble, lysine-rich form of the polymer was not detected in C. glutamicum in previous studies. Additionally, an incorporation of up to 6 mol% of glutamic acid into the backbone of CGP synthesized by C. glutamicum pVWEx1::cphADh was detected. The strain accumulated up to 17% of its dry weight in soluble CGP. Although glutamic acid had previously been found to replace arginine in the side chain, this is the first time that glutamic acid was found to substitute aspartic acid in the backbone.

Impact of curcumin on sirtuins: A review.

Curcumin is a bioactive phytochemical that modulates several physiological and cellular processes leading to therapeutic effects against different diseases. Sirtuins are highly conserved nicotine adenine dinucleotide-dependent proteins that regulate the activity of target enzymes and transcription factors by deacetylation. Curcumin possesses both antioxidant and anti-inflammatory properties and has been shown to increase sirtuin-1 (SIRT1) by activating small molecules. Upregulation of SIRT1 by curcumin has been reported to confer protective effects against a range of neurological disorders including glutamate excitotoxicity, ß-amyloid-induced cell death in cortical neurons, cerebral ischemic damage, and stroke. Activation of AMPK and SIRT1 by curcumin has also been noted to mediate the protective effects of curcumin against ischemia/reperfusion injury, cardiac fibrosis, diabetes, and lipid metabolism abnormalities. These protective effects of SIRT1 activation are partly mediated by the deacetylation of p53 and reduction of apoptosis. In this review, we summarize the role of SIRT1 in mediating the pharmacological effects of curcumin in several diseases.

Determinants of the VP1/2A junction cleavage by the 3C protease in foot-and-mouth disease virus-infected cells.

The foot-and-mouth disease virus (FMDV) capsid precursor, P1-2A, is cleaved by FMDV 3C protease to yield VP0, VP3, VP1 and 2A. Cleavage of the VP1/2A junction is the slowest. Serotype O FMDVs with uncleaved VP1-2A (having a K210E substitution in VP1; at position P2 in cleavage site) have been described previously and acquired a second site substitution (VP1 E83K) during virus rescue. Furthermore, introduction of the VP1 E83K substitution alone generated a second site change at the VP1/2A junction (2A L2P, position P2' in cleavage site). These virus adaptations have now been analysed using next-generation sequencing to determine sub-consensus level changes in the virus; this revealed other variants within the E83K mutant virus population that changed residue VP1 K210. The construction of serotype A viruses with a blocked VP1/2A cleavage site (containing K210E) has now been achieved. A collection of alternative amino acid substitutions was made at this site, and the properties of the mutant viruses were determined. Only the presence of a positively charged residue at position P2 in the cleavage site permitted efficient cleavage of the VP1/2A junction, consistent with analyses of diverse FMDV genome sequences. Interestingly, in contrast to the serotype O virus results, no second site mutations occurred within the VP1 coding region of serotype A viruses with the blocked VP1/2A cleavage site. However, some of these viruses acquired changes in the 2C protein that is involved in enterovirus morphogenesis. These results have implications for the testing of potential antiviral agents targeting the FMDV 3C protease.

N-acetylcysteine modulates glutamatergic dysfunction and depressive behavior in Huntington's disease.

Glutamatergic dysfunction has been implicated in the pathogenesis of depressive disorders and Huntington's disease (HD), in which depression is the most common psychiatric symptom. Synaptic glutamate homeostasis is regulated by cystine-dependent glutamate transporters, including GLT-1 and system xc- In HD, the enzyme regulating cysteine (and subsequently cystine) production, cystathionine-γ-lygase, has recently been shown to be lowered. The aim of the present study was to establish whether cysteine supplementation, using N-acetylcysteine (NAC) could ameliorate glutamate pathology through the cystine-dependent transporters, system xc- and GLT-1. We demonstrate that the R6/1 transgenic mouse model of HD has lower basal levels of cystine, and showed depressive-like behaviors in the forced-swim test. Administration of NAC reversed these behaviors. This effect was blocked by co-administration of the system xc- and GLT-1 inhibitors CPG and DHK, showing that glutamate transporter activity was required for the antidepressant effects of NAC. NAC was also able to specifically increase glutamate in HD mice, in a glutamate transporter-dependent manner. These in vivo changes reflect changes in glutamate transporter protein in HD mice and human HD post-mortem tissue. Furthermore, NAC was able to rescue changes in key glutamate receptor proteins related to excitotoxicity in HD, including NMDAR2B. Thus, we have shown that baseline reductions in cysteine underlie glutamatergic dysfunction and depressive-like behavior in HD and these changes can be rescued by treatment with NAC. These findings have implications for the development of new therapeutic approaches for depressive disorders.

ENDOCRINE TUMORS: BRAF V600E mutations in papillary craniopharyngioma.

Papillary craniopharyngioma (PCP) is an intracranial tumor that results in high levels of morbidity. We recently demonstrated that the vast majority of these tumors harbor the oncogenic BRAF V600E mutation. The pathologic diagnosis of PCP can now be confirmed using mutation specific immunohistochemistry and targeted genetic testing. Treatment with targeted agents is now also a possibility in select situations. We recently reported a patient with a multiply recurrent PCP in whom targeting both BRAF and MEK resulted in a dramatic therapeutic response with a marked anti-tumor immune response. This work shows that activation of the MAPK pathway is the likely principal oncogenic driver of these tumors. We will now investigate the efficacy of this approach in a multicenter phase II clinical trial. Post-treatment resection samples will be monitored for the emergence of resistance mechanisms. Further advances in the non-invasive diagnosis of PCP by radiologic criteria and by cell-free DNA testing could someday allow neo-adjuvant therapy for this disease in select patient populations.

Acidic Residues in the Hfq Chaperone Increase the Selectivity of sRNA Binding and Annealing.

Hfq facilitates gene regulation by small non-coding RNAs (sRNAs), thereby affecting bacterial attributes such as biofilm formation and virulence. Escherichia coli Hfq recognizes specific U-rich and AAN motifs in sRNAs and target mRNAs, after which an arginine patch on the rim promotes base pairing between their complementary sequences. In the cell, Hfq must discriminate between many similar RNAs. Here, we report that acidic amino acids lining the sRNA binding channel between the inner pore and rim of the Hfq hexamer contribute to the selectivity of Hfq's chaperone activity. RNase footprinting, in vitro binding and stopped-flow fluorescence annealing assays showed that alanine substitution of D9, E18 or E37 strengthened RNA interactions with the rim of Hfq and increased annealing of non-specific or U-tailed RNA oligomers. Although the mutants were less able than wild-type Hfq to anneal sRNAs with wild-type rpoS mRNA, the D9A mutation bypassed recruitment of Hfq to an (AAN)4 motif in rpoS, both in vitro and in vivo. These results suggest that acidic residues normally modulate access of RNAs to the arginine patch. We propose that this selectivity limits indiscriminate target selection by E. coli Hfq and enforces binding modes that favor genuine sRNA and mRNA pairs.

Characterization and Regulation of the Amino Acid Transporter SNAT2 in the Small Intestine of Piglets.

The sodium-dependent neutral amino acid transporter 2 (SNAT2), which has dual transport/receptor functions, is well documented in eukaryotes and some mammalian systems, but has not yet been verified in piglets. The objective of this study was to investigate the characteristics and regulation of SNAT2 in the small intestine of piglets. The 1,521-bp porcine full cDNA sequence of SNAT2 (KC769999) from the small intestine of piglets was cloned. The open reading frame of cDNA encodes 506 deduced amino acid residues with a calculated molecular mass of 56.08 kDa and an isoelectric point (pI) of 7.16. Sequence alignment and phylogenetic analysis revealed that SNAT2 is highly evolutionarily conserved in mammals. SNAT2 mRNA can be detected in the duodenum, jejunum and ileum by real-time quantitative PCR. During the suckling period from days 1 to 21, the duodenum had the highest abundance of SNAT2 mRNA among the three segments of the small intestine. There was a significant decrease in the expression of SNAT2 mRNA in the duodenal and jejunal mucosa and in the expression of SNAT2 protein in the jejunal and ileal mucosa on day 1 after weaning (P < 0.05). Studies with enterocytes in vitro showed that amino acid starvation and supplementation with glutamate, arginine or leucine enhanced, while supplementation with glutamine reduced, SNAT2 mRNA expression (P < 0.05). These results regarding the characteristics and regulation of SNAT2 should help to provide some information to further clarify its roles in the absorption of amino acids and signal transduction in the porcine small intestine.

Bioenergetic modulation with dichloroacetate reduces the growth of melanoma cells and potentiates their response to BRAFV600E inhibition.

BACKGROUND: Advances in melanoma treatment through targeted inhibition of oncogenic BRAF are limited owing to the development of acquired resistance. The involvement of BRAFV600E in metabolic reprogramming of melanoma cells provides a rationale for co-targeting metabolism as a therapeutic approach. METHODS: We examined the effects of dichloroacetate (DCA), an inhibitor of pyruvate dehydrogenase kinase, on the growth and metabolic activity of human melanoma cell lines. The combined effect of DCA and the BRAF inhibitor vemurafenib was investigated in BRAFV600E -mutated melanoma cell lines. Vemurafenib-resistant cell lines were established in vitro and their sensitivity to DCA was tested. RESULTS: DCA induced a reduction in glycolytic activity and intracellular ATP levels, and inhibited cellular growth. Co-treatment of BRAFV600E-mutant melanoma cells with DCA and vemurafenib induced a greater reduction in intracellular ATP levels and cellular growth than either compound alone. In addition, melanoma cells with in vitro acquired resistance to vemurafenib retained their sensitivity to DCA. CONCLUSIONS: These results suggest that DCA potentiates the effect of vemurafenib through a cooperative attenuation of energy production. Furthermore, the demonstration of retained sensitivity to DCA in melanoma cells with acquired resistance to vemurafenib could have implications for melanoma treatment.

Astrocyte elevated gene-1 is a novel modulator of HIV-1-associated neuroinflammation via regulation of nuclear factor-κB signaling and excitatory amino acid transporter-2 repression.

Astrocyte elevated gene-1 (AEG-1), a novel human immunodeficiency virus (HIV)-1 and tumor necrosis factor (TNF)-α-inducible oncogene, has generated significant interest in the field of cancer research as a therapeutic target for many metastatic aggressive tumors. However, little is known about its role in astrocyte responses during HIV-1 central nervous system (CNS) infection and whether it contributes toward the development of HIV-associated neurocognitive disorders (HAND). Therefore, in this study, we investigated changes in AEG-1 CNS expression in HIV-1-infected brain tissues and elucidated a potential mechanism of AEG-1-mediated regulation of HAND. Immunoblotting and immunohistochemical analyses of HIV-1 seropositive and HIV-1 encephalitic human brain tissues revealed significantly elevated levels of AEG-1 protein. Immunohistochemical analyses of HIV-1 Tat transgenic mouse brain tissues also showed a marked increase in AEG-1 staining. Similar to in vivo observations, cultured astrocytes expressing HIV-1 Tat also revealed AEG-1 and cytokine up-regulation. Astrocytes treated with HAND-relevant stimuli, TNF-α, interleukin (IL)-1ß, and HIV-1, also significantly induced AEG-1 expression and nuclear translocation via activation of the nuclear factor (NF)-κB pathway. Co-immunoprecipitation studies demonstrated IL-1ß- or TNF-α-induced AEG-1 interaction with NF-κB p65 subunit. AEG-1 knockdown decreased NF-κB activation, nuclear translocation, and transcriptional output in TNF-α-treated astrocytes. Moreover, IL-1ß treatment of AEG-1-overexpressing astrocytes significantly lowered expression of excitatory amino acid transporter 2, increased expression of excitatory amino acid transporter 2 repressor ying yang 1, and reduced glutamate clearance, a major transducer of excitotoxic neuronal damage. Findings from this study identify a novel transcriptional co-factor function of AEG-1 and further implicate AEG-1 in HAND-associated neuroinflammation.

Transmissions of serotonin, dopamine, and glutamate are required for the formation of neurotoxicity from Al2O3-NPs in nematode Caenorhabditis elegans.

In this study, we investigated genetic mechanisms of neurotransmitters in regulating the formation of adverse effects on locomotion behavior in Al2O3 nanoparticles (NPs)-exposed Caenorhabditis elegans. Al2O3-NPs exposure caused the decrease of locomotion behavior with head thrash and body bend as endpoints. Interestingly, the neurotransmitters of glutamate, serotonin, and dopamine were required for the adverse effects of Al2O3-NPs on locomotion behavior in nematodes. Glutamate transporter EAT-4, serotonin transporter MOD-5, and dopamine transporter DAT-1 might serve as the molecular targets of Al2O3-NPs for neurotoxicity formation. Moreover, the behavioral response of nematodes to Al2O3-NPs exposure was primarily mediated by non-NMDA glutamate receptors GLR-2 and GLR-6, ionotropic serotonin receptor MOD-1, and D1-like dopamine receptor DOP-1. Therefore, Al2O3-NPs exposure influences locomotion behavior of nematodes primarily by impinging on their glutamatergic, serotoninergic, and dopaminergic systems. Our data will shed light on questions surrounding the involvement of neurotransmitters in mediating the adverse behavioral effects from Al2O3-NPs.

Glutamatergic transmission in schizophrenia: from basic research to clinical practice.

PURPOSE OF REVIEW: The past 20 years have seen the glutamatergic hypothesis go from theory to phase III trials of novel mechanism antipsychotics. RECENT FINDINGS: We review the recent literature on glutamatergic theory, covering assessment and genetic studies, as well as drug development in animals and humans. SUMMARY: Although evidence continues to accumulate in support of glutamate hypotheses, further research continues to be required and interactions with other key systems need to be explored.

The Akt inhibitor MK2206 synergizes, but perifosine antagonizes, the BRAF(V600E) inhibitor PLX4032 and the MEK1/2 inhibitor AZD6244 in the inhibition of thyroid cancer cells.

PURPOSE: The purpose of the study was to explore optimal combinations of currently actively developed drugs for dually targeting the Ras → Raf → MAPK kinase (MEK) → MAPK/ERK (MAPK) and the phosphatidylinositol 3-kinase/Akt pathways as effective treatments for thyroid cancer. EXPERIMENTAL DESIGN: We tested the combinations of the Akt inhibitors MK2206 or perifosine with the BRAF(V600E) inhibitor PLX4032 or the MEK1/2 inhibitor AZD6244 in thyroid cancer cells harboring both the BRAF(V600E) and PIK3CA mutations. RESULTS: We found that MK2206 could potently, when used alone, and synergistically, when combined with either PLX4032 or AZD6244, inhibit thyroid cancer cell growth with all the combination index values lower than 1. Perifosine could potently inhibit thyroid cancer cell growth when used alone, but a strong antagonism occurred between this drug and PLX4032 or AZD6244 in the inhibition of thyroid cancer cell growth with all combination index values higher than 1. Combinations of MK2206 with PLX4032 or AZD6244 dramatically enhanced G1 cell cycle arrest induced by each drug alone. However, G2 cell cycle arrest uniquely induced by perifosine alone and G1 cell cycle arrest induced by PLX4032 or AZD6244 were both reversed by combination treatments, providing a mechanism for their antagonism. All these drugs could correspondingly inhibit the MAPK and phosphatidylinositol 3-kinase/Akt signalings, confirming their expected target effects. CONCLUSIONS: We demonstrated, unexpectedly, opposite outcomes of MK2206 and perifosine in their combinational treatments with BRAF(V600E)/MEK inhibitors in thyroid cancer cells. The data may help appropriate selection of these prominent drugs for clinical trials of combination therapies for thyroid cancer.