Complete biosynthesis of QS-21 in engineered yeast.

QS-21 is a potent vaccine adjuvant and remains the only saponin-based adjuvant that has been clinically approved for use in humans1,2. However, owing to the complex structure of QS-21, its availability is limited. Today, the supply depends on laborious extraction from the Chilean soapbark tree or on low-yielding total chemical synthesis3,4. Here we demonstrate the complete biosynthesis of QS-21 and its precursors, as well as structural derivatives, in engineered yeast strains. The successful biosynthesis in yeast requires fine-tuning of the host's native pathway fluxes, as well as the functional and balanced expression of 38 heterologous enzymes. The required biosynthetic pathway spans seven enzyme families-a terpene synthase, P450s, nucleotide sugar synthases, glycosyltransferases, a coenzyme A ligase, acyl transferases and polyketide synthases-from six organisms, and mimics in yeast the subcellular compartmentalization of plants from the endoplasmic reticulum membrane to the cytosol. Finally, by taking advantage of the promiscuity of certain pathway enzymes, we produced structural analogues of QS-21 using this biosynthetic platform. This microbial production scheme will allow for the future establishment of a structure-activity relationship, and will thus enable the rational design of potent vaccine adjuvants.

Human umbilical cord blood mesenchymal stem cells mediate microglia activation and improve anxiety-like behavior in MIA-induced offspring of schizophrenic rats.

Current treatments for schizophrenia (SCZ) remain largely ineffective in one-third of patients. Recent studies using stem cell therapy show a close relationship between stem cell immunomodulatory function and neuroinflammation in SCZ. To better investigate the efficacy of stem cell therapy for SCZ, human umbilical cord blood mesenchymal stem cells (hUC-MSC) with powerful immunomodulatory effects were administered to rats via the tail vein (once a week for 5 consecutive weeks starting from the weaning period) using a maternal immune activation (MIA) rodent model. Open field, PPI, Western blotting, Q-PCR, and immunofluorescence were used to assess the biological effects of repeated tail vein injections of hUC-MSC in offspring rats following the MIA model of SCZ. The results indicated that offspring of MIA rats exhibited schizophrenia-like (SCZ-like) anxiety behavior, with observed microglial activation triggering neuroinflammation. Furthermore, levels of IBA1, HMGB1, and PSD95 were significantly up-regulated, while SYP was significantly down-regulated. It is suggested that hUCB-MSCs may act through HMGB1, Iba1, PSD95, and related pathway molecules to alleviate neuroinflammation and repair synaptic damage by regulating the activity state of microglia. Consequently, this could improve the abnormal behavior observed in MIA offspring rats.

Decreased CNNM2 expression in prefrontal cortex affects sensorimotor gating function, cognition, dendritic spine morphogenesis and risk of schizophrenia.

Genome-wide association studies (GWASs) have reported multiple single nucleotide polymorphisms (SNPs) associated with schizophrenia, yet the underlying molecular mechanisms are largely unknown. In this study, we aimed to identify schizophrenia relevant genes showing alterations in mRNA and protein expression associated with risk SNPs at the 10q24.32-33 GWAS locus. We carried out the quantitative trait loci (QTL) and summary data-based Mendelian randomization (SMR) analyses, using the PsychENCODE dorsolateral prefrontal cortex (DLPFC) expression QTL (eQTL) database, as well as the ROSMAP and Banner DLPFC protein QTL (pQTL) datasets. The gene CNNM2 (encoding a magnesium transporter) at 10q24.32-33 was identified to be a robust schizophrenia risk gene, and was highly expressed in human neurons according to single cell RNA-seq (scRNA-seq) data. We further revealed that reduced Cnnm2 in the mPFC of mice led to impaired cognition and compromised sensorimotor gating function, and decreased Cnnm2 in primary cortical neurons altered dendritic spine morphogenesis, confirming the link between CNNM2 and endophenotypes of schizophrenia. Proteomics analyses showed that reduced Cnnm2 level changed expression of proteins associated with neuronal structure and function. Together, these results identify a robust gene in the pathogenesis of schizophrenia.

Lactate levels in the brain and blood of schizophrenia patients: A systematic review and meta-analysis.

BACKGROUND: The pathophysiological mechanisms of schizophrenia are still unclear. Converging evidence suggests that energy metabolism abnormalities are involved in schizophrenia, and support its role in the pathophysiology of this disease. Lactate plays an important role in energy metabolism. Many studies have reported changes in the levels of lactate in the brain and serum of schizophrenia patients; however, the results from these studies are not consistent. To overcome this limitation, the goal of the present meta-analysis is to analyze the changes in lactate levels in the brain and blood of schizophrenia patients. METHODS: For this systematic review and meta-analysis, we performed a thorough search of relevant literature in the English language, using the MEDLINE, Cochrane, and Embase databases. RESULTS: In the present meta-analysis, 20 studies were scrutinized, including 13 studies on brain lactate levels, which involved 322 schizophrenia patients and 324 healthy individuals as controls. 7 studies on blood lactate levels, involving 234 schizophrenia patients and 238 healthy individuals, were also included. Brain lactate levels were elevated in schizophrenia patients, both in vivo and in post-mortem studies. Nevertheless, blood lactate levels in schizophrenia patients have revealed no statistically significant difference, as compared with control individuals. CONCLUSIONS: In comparison with healthy individuals, schizophrenia patients had higher lactate levels in the brain, rather than in the blood. These findings suggest independent regulatory mechanisms of lactate levels in the brain and peripheral tissues. Abnormal lactate metabolism in the brain may be an important pathological mechanism in schizophrenia.

Efficient 2-O-α-D-glucopyranosyl-sn-glycerol production by single whole-cell biotransformation through combined engineering and expression regulation with novel sucrose phosphorylase from Leuconostoc mesenteroides ATCC 8293.

2-O-α-D-glucopyranosyl-sn-glycerol (2-αGG) is a high value product with wide applications. Here, an efficient, safe and sustainable bioprocesses for 2-αGG production was designed. A novel sucrose phosphorylase (SPase) was firstly identified from Leuconostoc mesenteroides ATCC 8293. Subsequently, SPase mutations were processed with computer-aided engineering, of which the activity of SPaseK138C was 160% higher than that of the wild-type. Structural analysis revealed that K138C was a key functional residue moderating substrate binding pocket and thus influences catalytic activity. Furthermore, Corynebacterium glutamicum was employed to construct microbial cell factories along with ribosome binding site (RBS) fine-tuning and a two-stage substrate feeding control strategy. The maximum production of 2-αGG by these combined strategies reached 351.8 g·L-1 with 98% conversion rate from 1.4 M sucrose and 3.5 M glycerol in a 5-L bioreactor. This was one of the best performance reported in single-cell biosynthesis of 2-αGG, which paved effective ways for industrial-scale preparation of 2-αGG.

Cortical anatomical variations, gene expression profiles, and clinical phenotypes in patients with schizophrenia.

BACKGROUND AND HYPOTHESIS: Schizophrenia (SZ) patients display significant structural brain abnormalities; nevertheless, the genetic mechanisms regulating cortical anatomical variations and their correlation with the disease phenotype are still ambiguous. STUDY DESIGN: We characterized anatomical variation using a surface-based method derived from structural magnetic resonance imaging of patients with SZ and age- and sex-matched healthy controls (HCs). Partial least-squares regression was performed across cortex regions between anatomical variation and average transcriptional profiles of SZ risk genes and all qualified genes from the Allen Human Brain Atlas. The morphological features of each brain region were correlated to symptomology variables in patients with SZ using partial correlation analysis. STUDY RESULTS: A total of 203 SZ and 201 HCs were included in the final analysis. We observed significant variation of 55 regions of cortical thickness, 23 regions of volume, 7 regions of area, and 55 regions of local gyrification index (LGI) between SZ and HC groups. Expression profiles of 4 SZ risk genes and 96 genes from all qualified genes showed a correlation to anatomical variability, however, after multiple comparisons, the correlations were no longer significant. LGI variability in multiple frontal subregions was associated with specific symptoms of SZ, whereas cognitive function involving attention/vigilance was linked to LGI variability across nine brain regions. CONCLUSIONS: Cortical anatomical variation of patients with schizophrenia is associated with gene transcriptome profiles as well as clinical phenotypes.

Abnormalities of Regional Brain Activity in Patients With Schizophrenia: A Longitudinal Resting-State fMRI Study.

BACKGROUND: Evidence from functional and structural research suggests that abnormal brain activity plays an important role in the pathophysiology of schizophrenia (SZ). However, limited studies have focused on post-treatment changes, and current conclusions are inconsistent. STUDY DESIGN: We recruited 104 SZ patients to have resting-state functional magnetic resonance imaging scans at baseline and 8 weeks of treatment with second-generation antipsychotics, along with baseline scanning of 86 healthy controls (HCs) for comparison purposes. Individual regional homogeneity (ReHo), amplitude of low-frequency fluctuations (ALFF), and degree centrality values were calculated to evaluate the functional activity. The Positive and Negative Syndrome Scale (PANSS) and MATRICS Consensus Cognitive Battery were applied to measure psychiatric symptoms and cognitive impairment in SZ patients. RESULTS: Compared with HCs at baseline, SZ patients had higher ALFF and ReHo values in the bilateral inferior temporal gyrus, inferior frontal gyrus, and lower ALFF and ReHo values in fusiform gyrus and precuneus. Following 8 weeks of treatment, ReHo was increased in right medial region of the superior frontal gyrus (SFGmed) and decreased in the left middle occipital gyrus and the left postcentral gyrus. Meanwhile, ReHo of the right SFGmed was increased after treatment in the response group (the reduction rate of PANSS ≥50%). Enhanced ALFF in the dorsolateral of SFG correlated with improvement in depressive factor score. CONCLUSIONS: These findings provide novel evidence for the abnormal functional activity hypothesis of SZ, suggesting that abnormality of right SFGmed can be used as a biomarker of treatment response in SZ.

Efficient biosynthesis of (R)-mandelic acid from styrene oxide by an adaptive evolutionary Gluconobacter oxydans STA.

BACKGROUND: (R)-mandelic acid (R-MA) is a highly valuable hydroxyl acid in the pharmaceutical industry. However, biosynthesis of optically pure R-MA remains significant challenges, including the lack of suitable catalysts and high toxicity to host strains. Adaptive laboratory evolution (ALE) was a promising and powerful strategy to obtain specially evolved strains. RESULTS: Herein, we report a new cell factory of the Gluconobacter oxydans to biocatalytic styrene oxide into R-MA by utilizing the G. oxydans endogenous efficiently incomplete oxidization and the epoxide hydrolase (SpEH) heterologous expressed in G. oxydans. With a new screened strong endogenous promoter P12780, the production of R-MA was improved to 10.26 g/L compared to 7.36 g/L of using Plac. As R-MA showed great inhibition for the reaction and toxicity to cell growth, adaptive laboratory evolution (ALE) strategy was introduced to improve the cellular R-MA tolerance. The adapted strain that can tolerate 6 g/L R-MA was isolated (named G. oxydans STA), while the wild-type strain cannot grow under this stress. The conversion rate was increased from 0.366 g/L/h of wild type to 0.703 g/L/h by the recombinant STA, and the final R-MA titer reached 14.06 g/L. Whole-genome sequencing revealed multiple gene-mutations in STA, in combination with transcriptome analysis under R-MA stress condition, we identified five critical genes that were associated with R-MA tolerance, among which AcrA overexpression could further improve R-MA titer to 15.70 g/L, the highest titer reported from bulk styrene oxide substrate. CONCLUSIONS: The microbial engineering with systematic combination of static regulation, ALE, and transcriptome analysis strategy provides valuable solutions for high-efficient chemical biosynthesis, and our evolved G. oxydans would be better to serve as a chassis cell for hydroxyl acid production.

Potential plasma biomarker panels identification for the diagnosis of first-episode schizophrenia and monitoring antipsychotic monotherapy with the use of metabolomics analyses.

Schizophrenia (SCZ) is a severe mental disorder. Using liquid chromatography mass spectrometry, we performed comprehensive metabolomics analyses of plasma samples from healthy controls (HC) and first episode SCZ patients before and after an acute period of medication. Ten lipid metabolites and 27 soluble small molecules were identified as potential biomarkers associated with the diagnosis and treatment of SCZ. These metabolites were significantly reduced in SCZ, and lipids and sulfate were significantly increased after treatment. Of the metabolites identified, four showed significant correlations with the Positive and Negative Syndrome Scale total scores. A biomarker panel composed of alpha-dimorphecolic, Phosphatidylcholine (PC) (16:0/18:1(11Z)), 1-methylnicotinamide, Phosphatidylethanolamine (PE) (20:2(11Z,14Z)/18:2(9Z,12Z)), sulfate, and L-tryptophan was selected to distinguish SCZ from HC; this provided the maximum classification performance with an AUC of 0.972. A biomarker panel including C16 sphinganine, gamma-linolenic acid, linoleic acid, PC(16:0/18:1(11Z)), PE(20:2(11Z,14Z)/18:2(9Z,12Z)), and sulfate, was selected for discrimination between SCZ before and after medication, and produced the optimal classification performance with an AUC of 0.905. Disturbances in lipid metabolism, sulfation modification, tryptophan metabolism, anti-inflammatory and antioxidant systems, and unsaturated fatty acids metabolism, were identified in SCZ. Our findings could facilitate the development of objective diagnostic or drug treatment monitoring tools for schizophrenia.

Development of a 2-pyrrolidone biosynthetic pathway in Corynebacterium glutamicum by engineering an acetyl-CoA balance route.

2-Pyrrolidone is widely used in the textile and pharmaceutical industries. Here, we established a 2-pyrrolidone biosynthesis pathway in Corynebacterium glutamicum, by expressing glutamate decarboxylase (Gad) mutant and ß-alanine CoA transferase (Act) which activates spontaneous dehydration cyclization of GABA to form 2-pyrrolidone. Also, the 5' untranslated regions (UTR) strategy was used to increase the expression of protein. Furthermore, considering the importance of acetyl-CoA in the 2-pyrrolidone synthesis pathway, the acetyl-CoA synthetase (acsA) gene was introduced to convert acetate into acetyl-CoA thus achieving the recyclability of the economy. Finally, the fed-batch fermentation of the final strain in a 5 L bioreactor produced 10.5 g/L 2-pyrrolidone within 78 h, which increased by 42.5% by altering the level of gene expression. This is the first time to build the basic chemical 2-pyrrolidone from glucose in one step in C. glutamicum.

Abnormal patterns of regional homogeneity and functional connectivity across the adolescent first-episode, adult first-episode and adult chronic schizophrenia.

Functional deficits in schizophrenia (SZ) are observed prior to the onset of psychosis and differ at different stages of SZ. However, there is a paucity of studies focused on adolescent first-episode SZ (AOS), adult first-episode SZ (AFES), and adult chronic SZ (CHSZ). In this study, we investigated regional activity and corresponding functional connectivity alterations that have aimed to compare the three disease stages simultaneously. The subjects comprised 49 patients with AOS, 57 patients with AFES, 51 patients with CHSZ, 41 adolescent healthy controls, and 138 adult healthy controls. We compared regional homogeneity (ReHo) between patients at each disease stage with matched healthy controls. We focused on the shared brain regions that showed significant differences between SZ patients at the three different disease stages and healthy controls. Further analysis was conducted to explore whether the patterns of the whole brain functional connectivity alterations were similar. The putamen and medial frontal gyrus (MFG) showed consistently abnormal patterns in AOS, AFES, and CHSZ. Commonly decreased ReHo values in the MFG and increased ReHo values in the bilateral putamen were found in AOS, AFES, and CHSZ. Functional connectivity of MFG remained common abnormality in different SZ stage. In conclusion, ReHo abnormalities in the MFG and the putamen may be common abnormal patterns of brain function in the three different stages of SZ. The vmPFC-dlPFC FC abnormality common occurs in adolescence and adulthood.. This study may provide a more comprehensive understanding of the neurodevelopmental abnormality across the AOS, AFES, and CHSZ.

Sustainable isomaltulose production in Corynebacterium glutamicum by engineering the thermostability of sucrose isomerase coupled with one-step simplified cell immobilization.

Sucrose isomerase (SI), catalyzing sucrose to isomaltulose, has been widely used in isomaltulose production, but its poor thermostability is still resisted in sustainable batches production. Here, protein engineering and one-step immobilized cell strategy were simultaneously coupled to maintain steady state for long-term operational stabilities. First, rational design of Pantoea dispersa SI (PdSI) for improving its thermostability by predicting and substituting the unstable amino acid residues was investigated using computational analysis. After screening mutagenesis library, two single mutants (PdSIV280L and PdSIS499F) displayed favorable characteristics on thermostability, and further study found that the double mutant PdSIV280L/S499F could stabilize PdSIWT better. Compared with PdSIWT, PdSIV280L/S499F displayed a 3.2°C-higher T m , and showed a ninefold prolonged half-life at 45°C. Subsequently, a one-step simplified immobilization method was developed for encapsulation of PdSIV280L/S499F in food-grade Corynebacterium glutamicum cells to further enhance the recyclability of isomaltulose production. Recombinant cells expressing combinatorial mutant (RCSI2) were successfully immobilized in 2.5% sodium alginate without prior permeabilization. The immobilized RCSI2 showed that the maximum yield of isomaltulose by batch conversion reached to 453.0 g/L isomaltulose with a productivity of 41.2 g/l/h from 500.0 g/L sucrose solution, and the conversion rate remained 83.2% after 26 repeated batches.

The Role of m6A RNA Methylation in Cancer: Implication for Nature Products Anti-Cancer Research.

N6-methyladenosine (m6A) RNA methylation is identified as the most common, abundant and reversible RNA epigenetic modification in messenger RNA (mRNA) and non-coding RNA, especially within eukaryotic messenger RNAs (mRNAs), which post-transcriptionally directs many important processes of RNA. It has also been demonstrated that m6A modification plays a pivotal role in the occurrence and development of tumors by regulating RNA splicing, localization, translation, stabilization and decay. Growing number of studies have indicated that natural products have outstanding anti-cancer effects of their unique advantages of high efficiency and minimal side effects. However, at present, there are very few research articles to study and explore the relationship between natural products and m6A RNA modification in tumorigenesis. m6A is dynamically deposited, removed, and recognized by m6A methyltransferases (METTL3/14, METTL16, WTAP, RBM15/15B, VIRMA, CBLL1, and ZC3H13, called as "writers"), demethylases (FTO and ALKBH5, called as "erasers"), and m6A-specific binding proteins (YTHDF1/2/3, YTHDC1/2, IGH2BP1/2/3, hnRNPs, eIF3, and FMR1, called as "readers"), respectively. In this review, we summarize the biological function of m6A modification, the role of m6A and the related signaling pathway in cancer, such as AKT, NF-kB, MAPK, ERK, Wnt/ß-catenin, STAT, p53, Notch signaling pathway, and so on. Furthermore, we reviewed the current research on nature products in anti-tumor, and further to get a better understanding of the anti-tumor mechanism, thus provide an implication for nature products with anti-cancer research by regulating m6A modification in the future.

The LIM Protein AJUBA is a Potential Oncogenic Target and Prognostic Marker in Human Cancer via Pan-Cancer Analysis.

Purpose : The LIM (Lin-11, Isl1, MEC-3) domain protein AJUBA is involved in multiple biological functions, and its aberrant expression is related to the occurrence and progression of various cancers. However, there are no analytical studies on AJUBA in pan-cancer. Methods: We performed a comprehensive pan-cancer analysis and explored the potential oncogenic roles of AJUBA, including gene expression, genetic mutation, protein phosphorylation, clinical diagnostic biomarker, prognosis, and AJUBA-related immune infiltration based on The Cancer Genome Atlas and Genotype-Tissue Expression databases. Results: The results revealed that the expression of AJUBA highly correlated with poor clinical outcomes in patients with different types of cancer. Meanwhile, AJUBA expression was positively correlated with cancer-associated fibroblasts in many human cancers, such as breast invasive carcinoma, colon adenocarcinoma, brain lower-grade glioma, lung adenocarcinoma (LUAD), and ovarian serous cystadenocarcinoma (OV). Gene ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses showed that AJUBA is mainly involved in protein serine/threonine kinase activity, cell-cell junction, covalent chromatin modification, and Hippo signaling pathway. Conclusion: The pan-cancer study reveals the oncogenic roles of AJUBA and provides a comprehensive understanding of the molecular biological genetic information of AJUBA in various tumors.

[Efficient production of biliverdin through whole-cell biocatalysis using recombinant Escherichia coli].

Biliverdin is an important cellular antioxidant. Traditionally, biliverdin is produced by chemical oxidation of bilirubin, which is a complex process and the final product is of low purity. Here we report an efficient, green and safe process for biotechnological production of biliverdin. A heme oxygenase (HO) gene from Clostridium tetani was screened, and a recombinant strain Escherichia coli BL21/pETDuet-hoCt with the ability of transforming heme into biliverdin was constructed. A biliverdin yield of 32.9 mg/L from 100 mg/L substrate was achieved under pH 7.0 and 35 ℃. In order to improve the supply of reducing power, an NADPH regeneration system using glutamate dehydrogenase (GdhA) was constructed, resulting in a recombinant strain E. coli BL21/pETDuet-gdhAEc-hoCt which was capable of producing 71.5 mg/L biliverdin. Moreover, through introduction of a membrane surface display system, a recombinant strain E. coli BL21/pETDuet-gdhAEc-blc/hoCt was constructed to shorten the transformation time, and the production of biliverdin was further increased to 76.3 mg/L, this is the highest titer of biosynthesized biliverdin reported to date, and the research may thus facilitate the green production of biliverdin.

Efficient D-allulose synthesis under acidic conditions by auto-inducing expression of the tandem D-allulose 3-epimerase genes in Bacillus subtilis.

BACKGROUND: D-allulose, a hexulose monosaccharide with low calorie content and high sweetness, is commonly used as a functional sugar in food and nutrition. However, enzyme preparation of D-allulose from D-frutose was severely hindered by the non-enzymatic browning under alkaline and high-temperature, and the unnecessary by-products further increased the difficulties in separation and extraction for industrial applications. Here, to address the above issue during the production process, a tandem D-allulose 3-epimerase (DPEases) isomerase synergistic expression strategy and an auto-inducible promoter engineering were levered in Bacillus subtilis 168 (Bs168) for efficient synthesis of D-allulose under the acidic conditions without browning. RESULTS: First, based on the dicistron expression system, two DPEases with complementary functional characteristics from Dorea sp. CAG:317 (DSdpe) and Clostridium cellulolyticum H10 (RCdpe) were expressed in tandem under the promoter HpaII in one cell. A better potential strain Bs168/pMA5-DSdpe-RCdpe increases enzyme activity to 18.9 U/mL at acidic conditions (pH 6.5), much higher than 17.2 and 16.7 U/mL of Bs168/pMA5-DSdpe and Bs168/pMA5-RCdpe, respectively. Subsequently, six recombinant strains based on four constitutive promoters were constructed in variable expression cassettes for improving the expression level of protein. Among those engineered strains, Bs168/pMA5-PspoVG-DSdpe-PsrfA-RCdpe exhibited the highest enzyme activity with 480.1 U/mL on fed-batch fermentation process in a 5 L fermenter at pH 6.5, about 2.1-times higher than the 228.5 U/mL of flask fermentation. Finally, the maximum yield of D-allulose reached as high as 163.5 g/L at the fructose concentration (50% w/v) by whole-cell biocatalyst. CONCLUSION: In this work, the engineered recombinant strain Bs168/pMA5-PspoVG-DSdpe-PsrfA-RCdpe was demonstrated as an effective microbial cell factory for the high-efficient synthesis of D-allulose without browning under acidic conditions. Based on the perspectives from this research, this strategy presented here also made it possible to meet the requirements of the industrial hyper-production of other rare sugars under more acidic conditions in theory.

Associations between expression of indoleamine 2, 3-dioxygenase enzyme and inflammatory cytokines in patients with first-episode drug-naive Schizophrenia.

The indoleamine 2,3-dioxygenase (IDO) enzyme is the first rate-limiting enzyme of the tryptophan degradation pathway in which dysfunction of neuroactive metabolites has been implicated in the pathophysiology of schizophrenia. Inflammatory molecules such as pro-inflammatory cytokines could enhance the activity of IDO. There are few studies on the expression of IDO levels and its correlation with levels of inflammatory cytokines in first-episode drug-naive patients with schizophrenia. One hundred inpatients (female = 33, male = 67) with first-episode drug-naive schizophrenia entered a 6-week, double-blind, randomized, placebo-controlled clinical trial. All individuals were assigned celecoxib or placebo combined with risperidone. Serum levels of IDO and six inflammatory cytokines (IL-1ß, IL-6, TNF-α IL-17, IL-4, and INF-γ) were measured. The Positive and Negative Syndrome Scale (PANSS) was used to assess the severity of psychotic symptoms. Compared to healthy subjects, patients had significantly elevated levels of IDO and six cytokines at baseline. Over the 6-week treatment period, the decrease in the levels of IDO and TNF-α and the improvement in the PANSS total score, positive scores, and negative scores in the celecoxib group were significantly greater than in the placebo group. There was a significantly positive correlation between IDO levels and the PANSS negative scores and between IDO levels and TNF-α and IFN-γ levels in the celecoxib group. These findings showed abnormal expression of IDO levels which correlated with negative symptoms and pro-inflammatory cytokine levels in patients with first-episode drug-naive schizophrenia, suggesting the important role of IDO in the pathological mechanism of schizophrenia. Registration number: ChiCTR2000041403.

[Molecular modification and highly efficient expression of L-asparaginase from Rhizomucor miehei].

L-asparaginase hydrolyzes L-asparagine to produce L-aspartic acid and ammonia. It is widely distributed in microorganisms, plants and serum of some rodents, and has important applications in the pharmaceutical and food industries. However, the poor thermal stability, low catalytic efficiency and low yield hampered the further application of L-asparaginase. In this paper, rational design and 5' untranslated region (5'UTR) design strategies were used to increase the specific enzyme activity and protein expression of L-asparaginase derived from Rhizomucor miehei (RmAsnase). The results showed that among the six mutants constructed through homology modeling combined with sequence alignment, the specific enzyme activity of the mutant A344E was 1.5 times higher than the wild type. Subsequently, a food-safe strain Bacillus subtilis 168/pMA5-A344E was constructed, and the UTR strategy was used for the construction of recombinant strain B. subtilis 168/pMA5 UTR-A344E. The enzyme activity of B. subtilis 168/pMA5 UTR-A344E was 7.2 times higher than that of B. subtilis 168/pMA5-A344E. The recombinant strain B. subtilis 168/pMA5 UTR-A344E was scaled up in 5 L fermenter, and the final yield of L-asparaginase was 489.1 U/mL, showing great potential for industrial application.

Chronic developmental exposure to low-dose ([C8mim][PF6]) induces neurotoxicity and behavioural abnormalities in rats.

Ionic liquids (ILs) are widely used for their physical and chemical properties. Toxicological assessments of ILs could help to avoid their threat to human health, but these are rarely reported, and no assessments of IL neurotoxicity in mammals have been performed. Here, we aimed to evaluate the neurotoxicity of chronic 1-octyl-3-methylimidazolium hexafluorophosphate ([C8mim][PF6]) (0, 1 mg/kg) exposure during development on rats. Our results indicated that chronic exposure to low-dose ([C8mim][PF6]) induces behavioural abnormalities, including cognitive deficits, social communication disorders, and sensory gating function impairment. Moreover, rats subjected to chronic ([C8mim][PF6]) exposure showed hypofunction of glutamatergic excitatory synapses, including increased expression of NMDA receptor subunits, increased density and immaturity of dendritic spines, and increased expression of PSD95. Additionally, ([C8mim][PF6]) exposure resulted in hippocampal-specific inflammatory activation, indicated by increased levels of proinflammatory factors, elevated nuclear localisation of NF-κB, and activation of microglia and astrocytes. In conclusion, chronic exposure to low-dose ([C8mim][PF6]) induced neurotoxicity, including damage to glutamatergic excitatory synapses and inflammatory activation, which may illuminate the associated behavioural abnormalities. The results presented here may be helpful for the safe use of ILs in the future.

Weak Association Between the Glutamate Decarboxylase 1 Gene (GAD1) and Schizophrenia in Han Chinese Population.

OBJECTIVES: Schizophrenia (SZ) is a complex psychiatric disorder with high heritability, and genetic components are thought to be pivotal risk factors for this illness. The glutamate decarboxylase 1 gene (GAD1) was hypothesized to be a candidate risk locus for SZ given its crucial role in the GABAergic neurotransmission system, and previous studies have examined the associations of single nucleotide polymorphisms (SNPs) spanning the GAD1 gene with SZ. However, inconsistent results were obtained. We hence examined the associations between GAD1 SNPs and SZ in two independent case-control samples of Han Chinese ancestry. MATERIALS AND METHODS: Two Han Chinese SZ case-control samples, referred as the discovery sample and the replication sample, respectively, were recruited for the current study. The discovery sample comprised of 528 paranoid SZ cases (with age of first onset ≥ 18) and 528 healthy controls; the independent replication sample contained 1,256 early onset SZ cases (with age of first onset < 18) and 2,661 healthy controls. Logistic regression analysis was performed to examine the associations between GAD1 SNPs and SZ. RESULTS: Ten SNPs covering GAD1 gene were analyzed in the discovery sample, and two SNPs showed nominal associations with SZ (rs2241165, P = 0.0181, OR = 1.261; rs2241164, P = 0.0225, OR = 1.219). SNP rs2241164 was also nominally significant in the independent replication sample (P = 0.0462, OR = 1.110), and the significance became stronger in a subsequent meta-analysis combining both discovery and replication samples (P = 0.00398, OR = 1.138). Nevertheless, such association could not survive multiple corrections, although the effect size of rs2241164 was comparable with other SZ risk loci identified in genome-wide association studies (GWAS) in Han Chinese population. We also examined the associations between GAD1 SNPs and SZ in published datasets of SZ GWAS in East Asians and Europeans, and no significant associations were observed. CONCLUSION: We observed weak associations between GAD1 SNPs and risk of SZ in Han Chinese populations. Further analyses in larger Han Chinese samples with more detailed phenotyping are necessary to elucidate the genetic correlation between GAD1 SNPs and SZ.