Host CD3+ T-cells can significantly modulate phage treatment effects on bacterial bioburden in mouse models.

Wound healing is a complex system including such key players as host, microbe, and treatments. However, little is known about their dynamic interactions. Here we explored the interplay between: (1) bacterial bioburden and host immune responses, (2) bacterial bioburden and wound size, and (3) treatments and wound size, using murine models and various treatment modalities: Phosphate buffer saline (PBS or vehicle, negative control), doxycycline, and two doses of A. baumannii phage mixtures. We uncovered that the interplay between bacterial bioburden and host immune system may be bidirectional, and that there is an interaction between host CD3+ T-cells and phage dosage, which significantly impacts bacterial bioburden. Furthermore, the bacterial bioburden and wound size association is significantly modulated by the host CD3+ T-cells. When the host CD3+ T-cells (x on log10 scale) are in the appropriate range (1.35 < x < = 1.5), we observed a strong association between colony forming units (CFU) and wound size, indicating a hallmark of wound healing. On the basis of the findings and our previous work, we proposed an integrated parallel systems biology model.

Intergenomic and epistatic interactions control free radical mediated pancreatic ß-cell damage.

Alloxan (AL)-generated Reactive Oxygen Species (ROS) selectively destroy insulin-producing pancreatic ß-cells. A previous genome-wide scan (GWS) using a cohort of 296 F2 hybrids between NOD (AL-sensitive) and ALR (AL-resistant) mice identified linkages contributing to ß-cell susceptibility or resistance to AL-induced diabetes on Chromosomes (Chr) 2, 3, 8, and a single nucleotide polymorphism in mt-Nd2 of the mitochondrial genome (mtDNA). AL treatment of congenic and consomic NOD mouse stocks confirmed resistance linked to both the mtDNA and the Chr 8 locus from ALR [NOD.mtALR.ALR-(D8Mit293-D8Mit137)]. To identify possible epistatic interactions, the GWS analysis was expanded to 678 F2 mice. ALR-derived diabetes-resistance linkages on Chr 8 as well as the mt-Nd2 a allele were confirmed and novel additional linkages on Chr 4, 5, 6, 7, and 13 were identified. Epistasis was observed between the linkages on Chr 8 and 2 and Chr 8 and 6. Furthermore, the mt-Nd2 genotype affected the epistatic interactions between Chr 8 and 2. These results demonstrate that a combination of nuclear-cytoplasmic genome interactions regulates ß-cell sensitivity to ROS-mediated ALD.

[Analysis of A Pedigree with Hereditary Coagulation Factor â « Deficiency Caused by Compound Heterozygous Mutations].

OBJECTIVE: To analysis clinical phenotype and potential genetic cause of a family affected with hereditary coagulation factor Ⅻ deficiency. METHODS: The prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (FIB), D-Dimer (D-D), coagulation factor Ⅻ activity (FⅫ:C) and coagulation factor Ⅻ antigen (FⅫ:Ag) were determined for phenotype diagnosis of the proband and his family members(3 generations and 5 people). Targeted capture and whole exome sequencing were performed in peripheral blood sample of the proband. Possible disease-causing mutations of F12 gene were obtained and further confirmed by Sanger sequencing. The corresponding mutation sites of the family members were analyzed afterwards. The online bioinformatics software AutoPVS1 and Mutation Taster was used to predict the effects of mutation sites on protein function. RESULTS: The APTT of the proband was significantly prolonged, reaching 180.9s. FⅫ:C and FⅫ:Ag of the proband was significantly reduced to 0.8% and 4.17%, respectively. The results of whole exome sequencing displayed that there were compound heterozygous mutations in F12 gene of the proband, including the c.1261G＞T heterozygous nonsense mutation in exon 11 (causing p.Glu421*) and the c.251dupG heterozygous frameshift mutation in exon 4 (causing p.Trp85Metfs*53). Both mutations are loss of function mutations with very strong pathogenicity, leading to premature termination of the protein. AutoPVS1 and Mutation Taster software predicted both mutations as pathogenic mutations. The results of Sanger sequencing revealed that c.1261G＞T heterozygous mutation of the proband was inherited from his mother, for which his brother and his daughter were c.1261G＞T heterozygous carriers. Genotype-phenotype cosegregation was observed in this family. CONCLUSION: The c.1261G＞T heterozygous nonsense mutation in exon 11 and the c.251dupG heterozygous frameshift mutation in exon 4 of the F12 gene probably account for coagulation factor Ⅻ deficiency in this family. This study reports two novel pathogenic F12 mutations for the first time worldwide.

Correlation between preoperative sleep disorders and postoperative delayed neurocognitive recovery in elderly patients. / è€å¹´æ‚£è€ æœ¯å‰ç¡çœ éšœç¢ä¸Žæœ¯åŽç¥žç»è®¤çŸ¥æ¢å¤å»¶è¿Ÿçš„ç›¸å ³æ€§.

OBJECTIVES: Perioperative neurocognitive disorders (PND) is one of the important factors affecting the recovery of the elderly after surgery, and sleep disorders are also one of the common diseases of the elderly. Previous studies have shown that the quality of postoperative sleep may be factor affecting postoperative cognitive function, but there are few studies on the relationship between preoperative sleep disorders and postoperative cognitive dysfunction. This study aims to explore the relationship between preoperative sleep disorders and postoperative delayed neurocognitive recovery in elderly patients, and provide references for improving the prognosis and quality of life of patients. METHODS: This study was porformed as a prospective cohort study. Elderly patients (age≥65 years old) underwent elective non-cardiac surgery at Xiangya Hospital of Central South University from October 2019 to January 2020 were selected and interviewed 1 day before the operation. The Mini-Mental State Examination (MMSE) and Montreal Cognitive Assessment Scale (MoCA) were used to assess the patient's baseline cognitive status. Patients with preoperative MMSE scores of less than 24 points were excluded. For patients meeting the criteria of inclusion, Pittsburgh Sleep Quality Index (PSQI) scale was used to evaluate the patients, and the patients were divided into a sleep disorder group and a non-sleep disorder group according to the score. General data of patients were collected and intraoperative data were recorded, such as duration of surgery, anesthetic time, surgical site, intraoperative fluid input, intraoperative blood product input, intraoperative blood loss and drug use. On consecutive 5 days after surgery, Numerical Rating Scale (NRS) was used to evaluate the sleep of the previous night and the pain of the day, which were recorded as sleep NRS score and pain NRS score; Confusion Assessment Method for ICU (CAM-ICU) scale and Confusion Assessment Method (CAM) scale were used to assess the occurrence of delirium. On the 7th day after the operation, the MMSE and MoCA scales were used to evaluate cognitive function of patients. We compared the incidence of postoperative complications, the number of deaths, the number of unplanned ICU patients, the number of unplanned secondary operations, etc between the 2 groups. The baseline and prognosis of the 2 groups of patients were analyzed by univariate and multivariate logistics to analyze their correlation. RESULTS: A total of 105 patients were collected in this study, including 32 patients in the sleep disorder group and 73 patients in the non-sleep disorder group. The general information of the 2 groups, such as age, gender, body mass index, and surgery site, were not statistically significant (all P>0.05). There was no statistically significant difference in the operation time, intraoperative bleeding, the number of delirium, and sleep NRS score at 5 days after operation between the 2 groups (all P>0.05). The incidence of unplanned second surgery after surgery and the NRS pain score on the first day in the sleep disorder group were significantly higher than those in the non-sleep disorder group (P=0.002, P=0.045, respectively). A total of 20 patients (19%) in the 2 groups had postoperative delayed neurocognitive recovery on the 7th day after surgery. Among them, 11 patients (34.4%) in the sleep disorder group, 9 patients (12.3%) in the non-sleep disorder group. The incidence of delayed neurocognitive recovery in the sleep disorder group was significantly higher than that in the non-sleep disorder group (P=0.008). The incidence of postoperative infection in the sleep disorder group was also higher than that in the non-sleep disorder group (P=0.020). After controlling for the confounding factors, preoperative sleep disorders were still independently associated with postoperative delayed neurocognitive recovery (OR=3.330, 95% CI 1.063-10.431, P=0.039). CONCLUSIONS: Preoperative sleep disorders can increase the risk of delayed neurocognitive function recovery in elderly patients. Active treatment of preoperative sleep disorders may improve perioperative neurocognitive function in elderly patients.

Risk factors for postoperative deep venous thrombosis in patients underwent craniotomy. / å¼€é¢ æ‰‹æœ¯åŽæ·±é™è„‰è¡€æ “å½¢æˆçš„å±é™©å› ç´ .

OBJECTIVES: To analyze the risk factors for postoperative deep vein thrombosis (DVT) in neurosurgical patients to provide the basis for the prevention of postoperative DVT. METHODS: A total of 141 patients underwent neurosurgery were enrolled. Thrombelastography (TEG) test was performed before and at the end of surgery. According to whether there was DVT formation after operation, the patients were divided into a thrombosis group and a non-thrombosis group. T-test and rank sum test were used to compare the general clinical characteristics of the 2 groups, such as age, gender, intraoperative blood loss, D-dimer, intraoperative crystal input, colloid input, blood product transfusion, operation duration, length of postoperative hospitalization. The application of chi-square test and rank-sum test were used to compared TEG main test indicators such as R and K values between the 2 groups. Logistic regression was used to analyze the possible risk factors for postoperative DVT in neurosurgical patients. RESULTS: There were significant differences in postoperative TEG index R, clotting factor function, intraoperative blood loss, hypertension or not, length of postoperative hospital stay, and postoperative absolute bed time (all P<0.05). Logistic regression analysis showed hypercoagulability, more intraoperative blood loss and longer postoperative absolute bed time were risk factors for DVT formation after craniotomy. CONCLUSIONS: Hypercoagulability in postoperative TEG test of patients is an important risk factor for the formation of postoperative DVT after neurosurgery, which can predict the occurrence of postoperative DVT to some extent.

Over-expression of poplar NAC15 gene enhances wood formation in transgenic tobacco.

BACKGROUND: NAC (NAM/ATAF/CUC) is one of the largest plant-specific transcription factor (TF) families known to play significant roles in wood formation. Acting as master gene regulators, a few NAC genes can activate secondary wall biosynthesis during wood formation in woody plants. RESULTS: In the present study, firstly, we screened 110 differentially expressed NAC genes in the leaves, stems, and roots of di-haploid Populus simonii×P. nigra by RNA-Seq. Then we identified a nucleus-targeted gene, NAC15 gene, which was one of the highly expressed genes in the stem among 110 NAC family members. Thirdly, we conducted expression pattern analysis of NAC15 gene, and observed NAC15 gene was most highly expressed in the xylem by RT-qPCR. Moreover, we transferred NAC15 gene into tobacco and obtained 12 transgenic lines overexpressing NAC15 gene (TLs). And the relative higher content of hemicellulose, cellulose and lignin was observed in the TLs compared to the control lines containing empty vector (CLs). It also showed darker staining in the culms of the TLs with phloroglucinol staining, compared to the CLs. Furthermore, the relative expression level of a few lignin- and cellulose-related genes was significantly higher in the TLs than that in the CLs. CONCLUSIONS: The overall results indicated that NAC15 gene is highly expressed in the xylem of poplar and may be a potential candidate gene playing an important role in wood formation in transgenic tobacco.

Plasma MIR-212-3p as a biomarker for acute right heart failure with pulmonary artery hypertension.

BACKGROUND: Acute right heart failure occurs in patients with pulmonary artery hypertension (PAH) with exposure to acute inflammation, the mortality rate is very high when right heart failure occurs. Biomarkers that can be used to detect acute right heart failure in patients with pulmonary hypertension need to be studied. METHODS: A PAH rat model was established using monocrotaline, and lipopolysaccharide was used to induce acute right heart failure. The Agilent rat miRNA microarray, Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were used to assess the microRNA expression of PAH rats. The expression of up- and downregulated miRNAs in plasma from PAH patients with acute right heart failure was validated with quantitative reverse transcription polymerase chain reaction (qRT-PCR). Then, the Wilcoxon matched paired test and receiver operating characteristic (ROC) curve analysis were performed. RESULTS: Thirty-three miRNAs were upregulated, and 7 miRNAs were downregulated in plasma of PAH rats with acute right heart failure. In the plasma of PAH patients, the miR-212-3p level was inversely correlated with the level of NT-pro BNP, and the area under the ROC curve was 0.751. CONCLUSIONS: These results suggest that the reduction of the expression of MIR-212-3p may be a biomarker for PAH patients with right heart dysfunction.

Over-Expression of ERF38 Gene Enhances Salt and Osmotic Tolerance in Transgenic Poplar.

Ethylene response factor (ERF) gene family plays an important role in abiotic stress responses. In this study, we isolated a salt-inducible ERF gene, ERF38 (Potri.006G138900.1), from the 84K poplar (Populus alba × Populus glandulosa) and investigated its functions in salt and osmotic tolerance. We identified that ERF38 protein was targeted to nucleus and had no self-activation. Results from yeast-one-hybrid indicated that the ERF38 protein can specifically bind to the dehydration responsive element (DRE). We then successfully transferred the ERF38 gene into the 84K poplar. Under respective salt and polyethylene glycol (PEG)-6000 stresses, four of the physiological traits, including peroxidase (POD) and superoxide dismutase (SOD) activities, soluble protein content, and proline content, increased significantly in the transgenic plants, compared to the wild type. Regarding the other two parameters, hydrogen peroxide (H2O2) and malondialdehyde (MDA) content, their increments in the transgenic lines under the stresses, which were compared to the water control, were significantly low than that of the wild type. In addition, reactive oxygen species (ROS) are scavenged in the transgenic lines under the stresses, but not in the wild type (WT). Interestingly, when challenged with the stresses, expression levels of a few genes associated with POD and SOD metabolism were significantly increased in the transgenic poplars. In all, evidence from morphological, physiological, and biochemical analyses indicated that over-expression of ERF38 gene can improve salt and osmotic tolerance in the transgenic poplar.

Functional characterization of poplar WRKY75 in salt and osmotic tolerance.

The WRKY transcription factor family is one of the most important families in plants, playing a significant role in plant growth and development, as well as in stress responses. However, functional studies on the family in response to abiotic stresses are limited in poplar. In the present study, we cloned a WRKY transcription factor gene PagWRKY75, which was down-regulated during early stages of salt and osmotic stresses. The PagWRKY75 protein belongs to the WRKY IIc subfamily. It is located in the nucleus and can bind to the W box. We obtained transgenic poplar lines with PagWRKY75 overexpression or inhibited expression by RNA interference. Stress treatment experiments indicated that the transgenic poplar lines overexpressing PagWRKY75 were more sensitive to salt and osmotic stresses, compared to wild type. The transgenic lines with PagWRKY75 inhibition displayed opposite effects. Furthermore, our results showed that PagWRKY75 can reduce the ability of reactive oxygen species scavenging and the accumulation of proline under stresses, and positively regulate the water loss rate of leaves. These results indicate that the transcription factor PagWRKY75 can negatively regulate salt and osmotic tolerance by modulating various physiological processes.

[Prognosis in patients underwent craniotomy for aneurysm clipping with cardiovascular diseases].

OBJECTIVE: To analyze the prognostic factors for patients with or without cardiovascular diseases after craniotomy for aneurysm clipping, and to provide evidences for the improvement of perioperative management in these patients.
 Methods: We collected 297 patients who underwent craniotomy for aneurysm clipping in Xiangya Hospital of Central South University from May 2016 to February 2017. The patients were divided into two groups: the cardiovascular disease group and the non-cardiovascular disease group. The perioperative clinical data, neurological function assessments at admission and discharge and Glasgow Outcome Scale (GOS) scores of one-year-follow-up after discharge were analyzed. The primary outcome of this study was the GOS scores collected at one year after discharge. The secondary outcomes were the lengths of their ICU stay, neurological functions at discharge and adverse events morbidity during the hospitalization.
 Results: A total of 241 patients were eventually enrolled. There was no significant difference in their general data between the two groups except for their ages. The GOS scores of the one-year-follow-up were significantly different between the two groups (P=0.007). The lengths of ICU stay, neurological dysfunctions at discharge and adverse events morbidity during hospitalization were also significantly different (P=0.036, P=0.011, P=0.005, respectively). A multivariate logistic regression analysis in which GOS score was the dependent variable with age adjusted also supported the previous results that long-term prognosis was not significantly correlated with the age of patients (P>0.05), but it was correlated with cardiovascular disease and sanity at admission (P=0.001). In patients with cardiovascular diseases, there was significantly different in perioperative mortality and neurological recovery of patients who had or had not cardiovascular events (P=0.006, P=0.001, respectively).
 Conclusion: Undergoing craniotomy for aneurysm clipping, patients with cardiovascular diseases have worse outcomes in both of short and long terms. Perioperative treatments for cardiovascular disease could not only improve postoperative neurological deficits, but also reduce mortality for these patients.

Expression analysis of four pseudo-response regulator (PRR) genes in Chrysanthemum morifolium under different photoperiods.

Genes encoding pseudo-response regulator (PRR) proteins play significant roles in plant circadian clocks. In this study, four genes related to flowering time were isolated from Chrysanthemum morifolium. Phylogenetic analysis showed that they are highly homologous to the counterparts of PRRs of Helianthus annuus and named as CmPRR2, CmPRR7, CmPRR37, and CmPRR73. Conserved motifs prediction indicated that most of the closely related members in the phylogenetic tree share common protein sequence motifs, suggesting functional similarities among the PRR proteins within the same subtree. In order to explore functions of the genes, we selected two Chrysanthemum varieties for comparison; that is, a short-day sensitive Zijiao and a short-day insensitive Aoyunbaixue. Compared to Aoyunbaixue, Zijiao needs 13 more days to complete the flower bud differentiation. Evidence from spatio-temporal gene expression patterns demonstrated that the CmPRRs are highly expressed in flower and stem tissues, with a growing trend across the Chrysanthemum developmental process. In addition, we also characterized the CmPRRs expression patterns and found that CmPRRs can maintain their circadian oscillation features to some extent under different photoperiod treatment conditions. These lines of evidence indicated that the four CmPRRs undergo circadian oscillation and possibly play roles in regulating the flowering time of C. morifolium.

Comprehensive analysis of the three-amino-acid-loop-extension gene family and its tissue-differential expression in response to salt stress in poplar.

The three-amino-acid-loop-extension (TALE) transcription factor gene family is widely present in plants and plays an important role in its growth and development. However, studies on the gene family are limited in poplar. In this study, we investigated 35 TALE gene family members in terms of their evolutionary relationship, classification, physicochemical properties, gene structures, and protein motifs. We divided the genes into four classes, based on their protein sequences similarity. The members from each class share similar gene structures and motif compositions. Evidence from transcript profiling indicated that the majority of the TALE genes exhibited distinct expression patterns over leaf, stem, and root tissues. Out of the 35 genes, 17 genes are highly expressed in stems, suggesting that the TALE gene family may play an important role in secondary growth and wood formation. Furthermore, out of the 35 genes, 11 genes are responsive to salt stress, and the spatio-temporal expression patterns of these 11 genes under salt stress were analysed using RT-qPCR. Yeast two-hybridization analysis indicated that poplar TALE proteins from different classes can form heterodimers. These results lay the foundation for future studies on biological functions of poplar TALE genes.

Characterization of the basic helix-loop-helix gene family and its tissue-differential expression in response to salt stress in poplar.

The basic helix-loop-helix (bHLH) transcription factor gene family is one of the largest gene families and extensively involved in plant growth, development, and stress responses. However, limited studies are available on the gene family in poplar. In this study, we focused on 202 bHLH genes, exploring their DNA and protein sequences and physicochemical properties. According to their protein sequence similarities, we classified the genes into 25 groups with specific motif structures. In order to explore their expressions, we performed gene expression profiling using RNA-Seq and identified 19 genes that display tissue-differential expression patterns without treatment. Furthermore, we also performed gene expression profiling under salt stress. We found 74 differentially expressed genes (DEGs), which are responsive to the treatment. A total of 18 of the 19 genes correspond well to the DEGs. We validated the results using reverse transcription quantitative real-time PCR. This study lays the foundation for future studies on gene cloning, transgenes, and biological mechanisms.

Expression Pattern of ERF Gene Family under Multiple Abiotic Stresses in Populus simonii × P. nigra.

Identification of gene expression patterns of key genes across multiple abiotic stresses is critical for mechanistic understanding of stress resistance in plant. In the present study, we identified differentially expressed genes (DEGs) in di-haploid Populus simonii × P. nigra under respective stresses of NaCl, KCl, CdCl2, and PEG. On the basis of RNA-Seq, we detected 247 DEGs that are shared by the four stresses in wild type poplar, and mRNA abundance of the DEGs were validated in transgenic poplar overexpressing ERF76 gene by RNA-Seq and RT-qPCR. Results from gene ontology analysis indicated that these genes are enriched in significant pathways, such as phenylpropanoid biosynthesis, phenylalanine metabolism, starch and sucrose metabolism, and plant hormone signal transduction. Ethylene response factor (ERF) gene family plays significant role in plant abiotic stress responses. We also investigated expression pattern of ERF gene family under the four stresses. The ERFs and DEGs share similar expression pattern across the four stresses. The transgenic poplar is superior to WT in morphologic, physiological and biochemical traits, which demonstrated the ERF76 gene plays a significant role in stress resistance. These studies will give a rise in understanding the stress response mechanisms in poplar.

In vitro pharmacological and rat pharmacokinetic characterization of LY3020371, a potent and selective mGlu2/3 receptor antagonist.

Metabotropic glutamate 2/3 (mGlu2/3) receptors are of considerable interest owing to their role in modulating glutamate transmission via presynaptic, postsynaptic and glial mechanisms. As part of our ongoing efforts to identify novel ligands for these receptors, we have discovered (1S,2R,3S,4S,5R,6R)-2-amino-3-[(3,4-difluorophenyl)sulfanylmethyl]-4-hydroxy-bicyclo[3.1.0]hexane-2,6-dicarboxylic acid; (LY3020371), a potent and selective orthosteric mGlu2/3 receptor antagonist. In this account, we characterize the effects of LY3020371 in membranes and cells expressing human recombinant mGlu receptor subtypes as well as in native rodent and human brain tissue preparations, providing important translational information for this molecule. In membranes from cells expressing recombinant human mGlu2 and mGlu3 receptor subtypes, LY3020371.HCl competitively displaced binding of the mGlu2/3 agonist ligand [3H]-459477 with high affinity (hmGlu2 Ki = 5.26 nM; hmGlu3 Ki = 2.50 nM). In cells expressing hmGlu2 receptors, LY3020371.HCl potently blocked mGlu2/3 agonist (DCG-IV)-inhibited, forskolin-stimulated cAMP formation (IC50 = 16.2 nM), an effect that was similarly observed in hmGlu3-expressing cells (IC50 = 6.21 nM). Evaluation of LY3020371 in cells expressing the other human mGlu receptor subtypes revealed high mGlu2/3 receptor selectivity. In rat native tissue assays, LY3020371 demonstrated effective displacement of [3H]-459477 from frontal cortical membranes (Ki = 33 nM), and functional antagonist activity in cortical synaptosomes measuring both the reversal of agonist-suppressed second messenger production (IC50 = 29 nM) and agonist-inhibited, K+-evoked glutamate release (IC50 = 86 nM). Antagonism was fully recapitulated in both primary cultured cortical neurons where LY3020371 blocked agonist-suppressed spontaneous Ca2+ oscillations (IC50 = 34 nM) and in an intact hippocampal slice preparation (IC50 = 46 nM). Functional antagonist activity was similarly demonstrated in synaptosomes prepared from epileptic human cortical or hippocampal tissues, suggesting a translation of the mGlu2/3 antagonist pharmacology from rat to human. Intravenous dosing of LY3020371 in rats led to cerebrospinal fluid drug levels that are expected to effectively block mGlu2/3 receptors in vivo. Taken together, these results establish LY3020371 as an important new pharmacological tool for studying mGlu2/3 receptors in vitro and in vivo. This article is part of the Special Issue entitled 'Metabotropic Glutamate Receptors, 5 years on'.

Discovery of (1S,2R,3S,4S,5R,6R)-2-Amino-3-[(3,4-difluorophenyl)sulfanylmethyl]-4-hydroxy-bicyclo[3.1.0]hexane-2,6-dicarboxylic Acid Hydrochloride (LY3020371·HCl): A Potent, Metabotropic Glutamate 2/3 Receptor Antagonist with Antidepressant-Like Activity.

As part of our ongoing efforts to identify novel ligands for the metabotropic glutamate 2 and 3 (mGlu2/3) receptors, we have incorporated substitution at the C3 and C4 positions of the (1S,2R,5R,6R)-2-amino-bicyclo[3.1.0]hexane-2,6-dicarboxylic acid scaffold to generate mGlu2/3 antagonists. Exploration of this structure-activity relationship (SAR) led to the identification of (1S,2R,3S,4S,5R,6R)-2-amino-3-[(3,4-difluorophenyl)sulfanylmethyl]-4-hydroxy-bicyclo[3.1.0]hexane-2,6-dicarboxylic acid hydrochloride (LY3020371·HCl, 19f), a potent, selective, and maximally efficacious mGlu2/3 antagonist. Further characterization of compound 19f binding to the human metabotropic 2 glutamate (hmGlu2) site was established by cocrystallization of this molecule with the amino terminal domain (ATD) of the hmGlu2 receptor protein. The resulting cocrystal structure revealed the specific ligand-protein interactions, which likely explain the high affinity of 19f for this site and support its functional mGlu2 antagonist pharmacology. Further characterization of 19f in vivo demonstrated an antidepressant-like signature in the mouse forced-swim test (mFST) assay when brain levels of this compound exceeded the cellular mGlu2 IC50 value.

Novel bicyclo[3.1.0]hexane analogs as antagonists of metabotropic glutamate 2/3 receptors for the treatment of depression.

Negative modulators of metabotropic glutamate 2 & 3 receptors demonstrate antidepressant-like activity in animal models and hold promise as novel therapeutic agents for the treatment of major depressive disorder. Herein we describe our efforts to prepare and optimize a series of conformationally constrained 3,4-disubstituted bicyclo[3.1.0]hexane glutamic acid analogs as orthosteric (glutamate site) mGlu2/3 receptor antagonists. This work led to the discovery of a highly potent and efficacious tool compound 18 (hmGlu2 IC50 46±14.2nM, hmGlu3 IC50=46.1±36.2nM). Compound 18 showed activity in the mouse forced swim test with a minimal effective dose (MED) of 1mg/kg ip. While in rat EEG studies it exhibited wake promoting effects at 3 and 10mg/kg ip without any significant effects on locomotor activity. Compound 18 thus represents a novel tool molecule for studying the impact of blocking mGlu2/3 receptors both in vitro and in vivo.

Over-expression of poplar transcription factor ERF76 gene confers salt tolerance in transgenic tobacco.

Ethylene response factors (ERFs) belong to a large plant-specific transcription factor family, which play a significant role in plant development and stress responses. Poplar ERF76 gene, a member of ERF TF family, can be up-regulated in response to salt stress, osmotic stress, and ABA treatment. The ERF76 protein was confirmed to be targeted preferentially in the nucleus of onion cell by particle bombardment. In order to understand the functions of ERF76 gene in salt stress response, we conducted temporal and spatial expression analysis of ERF76 gene in poplar. Then the ERF76 cDNA fragment containing an ORF was cloned from di-haploid Populus simonii×P. nigra and transferred into tobacco (Nicotiana tobacum) genome by Agrobacterium-mediated leaf disc method. Under salt stress, transgenic tobacco over-expressing ERF76 gene showed a significant increase in seed germination rate, plant height, root length, and fresh weight, as well as in relative water content (RWC), superoxide dismutase (SOD) activity, peroxidase (POD) activity, and proline content, compared to control tobacco lines. In contrast, transgenic tobacco lines displayed a decrease in malondialdehyde (MDA) accumulation, relative electrical conductivity (REC) and reactive oxygen species (ROS) accumulation in response to salt stress, compared to control tobacco lines. Over all, the results indicated that ERF76 gene plays a critical role in salt tolerance in transgenic tobacco.

A Gene Regulatory Program in Human Breast Cancer.

Molecular heterogeneity in human breast cancer has challenged diagnosis, prognosis, and clinical treatment. It is well known that molecular subtypes of breast tumors are associated with significant differences in prognosis and survival. Assuming that the differences are attributed to subtype-specific pathways, we then suspect that there might be gene regulatory mechanisms that modulate the behavior of the pathways and their interactions. In this study, we proposed an integrated methodology, including machine learning and information theory, to explore the mechanisms. Using existing data from three large cohorts of human breast cancer populations, we have identified an ensemble of 16 master regulator genes (or MR16) that can discriminate breast tumor samples into four major subtypes. Evidence from gene expression across the three cohorts has consistently indicated that the MR16 can be divided into two groups that demonstrate subtype-specific gene expression patterns. For example, group 1 MRs, including ESR1, FOXA1, and GATA3, are overexpressed in luminal A and luminal B subtypes, but lowly expressed in HER2-enriched and basal-like subtypes. In contrast, group 2 MRs, including FOXM1, EZH2, MYBL2, and ZNF695, display an opposite pattern. Furthermore, evidence from mutual information modeling has congruently indicated that the two groups of MRs either up- or down-regulate cancer driver-related genes in opposite directions. Furthermore, integration of somatic mutations with pathway changes leads to identification of canonical genomic alternations in a subtype-specific fashion. Taken together, these studies have implicated a gene regulatory program for breast tumor progression.