TAF15 exacerbates nonalcoholic steatohepatitis progression by regulating lipid metabolism and inflammation via FASN and p65 NF-κB.

BACKGROUND & AIMS: Nonalcoholic fatty liver disease (NAFLD) consists of a broad spectrum of conditions, and nonalcoholic steatohepatitis (NASH) is the advanced form of NAFLD. TAF15 is a DNA and RNA binding protein and is involved in crucial inflammatory signalling pathways. We aimed to investigate the role of TAF15 in the progression of NASH and the underlying molecular mechanism. METHODS: We generated mice with hepatocyte-specific knockdown and overexpression of TAF15 using a specific adeno-associated virus (AAV). NASH models were established by feeding mice high-fat and high-cholesterol diets and methionine- and choline-deficient diets. Cleavage under targets and tagmentation and dual-luciferase reporter assays were performed to investigate the effect of TAF15 on FASN transcription. Coimmunoprecipitation and immunofluorescence assays were conducted to explore the interaction of TAF15 and p65. In vitro coculture systems were established to study the interactions of hepatocytes, macrophages and HSCs. RESULTS: TAF15 was significantly increased in the livers of mouse NASH models and primary hepatocyte NASH model. Knockdown of TAF15 inhibited steatosis, inflammation and fibrosis, while overexpression of TAF15 promoted NASH phenotypes. Mechanistically, TAF15 bound directly to the promoter region of FASN to facilitate its expression, thereby promoting steatosis. Moreover, TAF15 interacted with p65 and activated the NF-κB signalling pathway, increasing the secretion of proinflammatory cytokines and triggering M1 macrophage polarization. Treatment with the FASN inhibitor orlistat partially reversed the phenotypes. CONCLUSIONS: These results suggested that TAF15 exacerbated NASH progression by regulating lipid metabolism and inflammation via transcriptional activation of FASN and interacting with p65 to activate the NF-κB signalling pathway.

Detrimental role of SIX1 in hepatic lipogenesis and fibrosis of non-alcoholic fatty liver disease.

BACKGROUND AND AIMS: Nonalcoholic fatty liver disease (NAFLD) is the most common liver disease worldwide. Aberrant lipid metabolism and accumulation of extracellular matrix proteins are hallmarks of the disease, but the underlying mechanisms are largely unknown. This study aims to elucidate the key role of sine oculis homeobox homologue 1 (SIX1) in the development of NAFLD. METHODS: Alb-Cre mice were administered the AAV9 vector for SIX1 liver-specific overexpression or knockdown. Metabolic disorders, hepatic steatosis, and inflammation were monitored in mice fed with HFHC or MCD diet. High throughput CUT&Tag analysis was employed to investigate the mechanism of SIX1 in diet-induced steatohepatitis. RESULTS: Here, we found increased SIX1 expression in the livers of NAFLD patients and animal models. Liver-specific overexpression of SIX1 using adeno-associated virus serotype 9 (AAV9) provoked more severe inflammation, metabolic disorders, and hepatic steatosis in the HFHC or MCD-induced mice model. Mechanistically, we demonstrated that SIX1 directly activated the expression of liver X receptor α (LXRα) and liver X receptor ß (LXRß), thus inducing de novo lipogenesis (DNL). In addition, our results also illustrated a critical role of SIX1 in regulating the TGF-ß pathway by increasing the levels of type I and II TGF-ß receptor (TGFßRI/TGFßRII) in hepatic stellate cells (HSCs). Finally, we found that liver-specific SIX1 deficiency could ameliorate diet-induced NAFLD pathogenesis. CONCLUSION: Our findings suggest a detrimental function of SIX1 in the progression of NAFLD. The direct regulation of LXRα/ß and TGF-ß signalling by SIX1 provides a new regulatory mechanism in hepatic steatosis and fibrosis.

Microflow multi-layer diffraction optical element processed by hybrid manufacturing technology.

Traditional planar diffractive optical elements (DOEs) are challenged in imaging systems due to diffraction efficiency and chromatic dispersion. In this paper, we have designed a microfluidic diffractive optical element (MFDOE), which is processed by digital micromirror device (DMD) maskless lithography (DMDML) assisted femtosecond laser direct writing (FsLDW). MFDOE is a combination of photoresist-based multi-layer harmonic diffraction surface and liquid, realizing diffraction efficiency of more than 90% in the visible band. And it shows achromatic characteristics in the two bands of 469 nm (±20 nm) and 625 nm (±20 nm). These results show that MFDOE has good imaging performance.

CARM1 promotes gastric cancer progression by regulating TFE3 mediated autophagy enhancement through the cytoplasmic AMPK-mTOR and nuclear AMPK-CARM1-TFE3 signaling pathways.

BACKGROUND: The role of CARM1 in tumors is inconsistent. It acts as an oncogene in most cancers but it inhibits the progression of liver and pancreatic cancers. CARM1 has recently been reported to regulate autophagy, but this function is also context-dependent. However, the effect of CARM1 on gastric cancer (GC) has not been studied. We aimed to explore whether CARM1 was involved in the progression of GC by regulating autophagy. METHODS: The clinical values of CARM1 and autophagy in GC were evaluated by immunohistochemistry and qRT-PCR. Transmission electron microscopy, immunofluorescence and western blotting were employed to identify autophagy. The role of CARM1 in GC was investigated by CCK-8, colony formation and flow cytometry assays in vitro and a xenograft model in vivo. Immunoprecipitation assays were performed to determine the interaction of CARM1 and TFE3. RESULTS: CARM1 was upregulated in clinical GC tissues and cell lines, and higher CARM1 expression predicted worse prognosis. CARM1 enhanced GC cell proliferation, facilitated G1-S transition and inhibited ER stress-induced apoptosis by regulating autophagy. Importantly, treatment with a CARM1 inhibitor rescued the tumor-promoting effects of CARM1 both in vitro and in vivo. Furthermore, we demonstrated that CARM1 promoted TFE3 nuclear translocation to induce autophagy through the cytoplasmic AMPK-mTOR and nuclear AMPK-CARM1-TFE3 signaling pathways. CONCLUSION: CARM1 promoted GC cell proliferation, accelerated G1-S transition and reduced ER stress-induced apoptosis by regulating autophagy. Mechanistically, CARM1 triggered autophagy by facilitating TFE3 nuclear translocation through the AMPK-mTOR and AMPK-CARM1-TFE3 signaling pathways.

Long-chain unsaturated fatty acids are involved in the viability and itraconazole susceptibility of Aspergillus fumigatus.

The prevalence of invasive aspergillosis with azole resistance is increasing, but the mechanisms underlying the development of resistance and treatment strategies are still limited. The present work is focused on finding a relationship between long-chain unsaturated fatty acids (LCUFAs), Aspergillus fumigatus development, and antifungal resistance. The effects of LCUFAs on antifungal agents in vitro were determined, and the stearic acid desaturase gene (sdeA) of A. fumigatus was characterized. In in vitro antifungal tests, LCUFAs antagonized the antifungal activity of itraconazole by extracting it from media, thereby preventing it from entering cells. The OA auxotrophic phenotype caused by an sdeA deletion confirmed that SdeA was required for OA biosynthesis in A. fumigatus. Furthermore, several low-level sdeA-overexpressing mutants with impaired vegetative growth phenotypes were successfully constructed. Additionally, an sdeA-overexpressing mutant, OEsdeA-5, showed lowered sensitivity levels to itraconazole. Moreover, RNA sequencing of OEsdeA-5 revealed that the altered gene-expression pattern. Through targeted metabolomics, decreased palmitic acid and stearic acid contents, accompanied by higher palmitoleic acid, margaroleic acid, and OA production levels, were found in OEsdeA-5. This study provides a novel insight of understanding of azole resistance and a potential target for drug development.

Involvement of a novel Ca2+-independent C-type lectin from Sinonovacula constricta in food recognition and innate immunity.

Bivalve lectins perform a crucial function in recognition of foreign particles, such as microalgae and pathogenic bacteria. In this study, a novel C-type lectin form Sinonovacula constricta (ScCL) was characterized. The full-length cDNA of ScCL was 1645 bp, encoding a predicted polypeptide of 273 amino acids with one typical carbohydrate-recognition domain. ScCL has the highest similarity and closest phylogenetic relationship with the C-type lectin from Solen grandis. Real-time PCR analysis showed that ScCL was expressed in all tested tissues, with the highest expression in the foot and the lowest expression in hemocytes. Agglutination activity of ScCL was Ca2+-independent. ScCL showed the strongest agglutination on Chlorella vulgaris, the modest agglutination on Platymonas subcordiformis, Nannochloropsis sp., and Thalassiosira pseudonana, the weakest agglutination on Chaetoceros sp., and no agglutination on Isochrysis zhanjiangensis. Meanwhile, agglutination tests and western blot analysis revealed that the recombinant ScCL protein could agglutinate Staphylococcus aureus and Vibrio harveyi, but could not agglutinate Vibrio anguillarum, Bacillus cereus, or Vibrio parahaemolyticus. Furthermore, ScCL had a high binding activity with LPS and mannose, a low binding activity with LTA, and no binding activity with PGN. The expression of ScCL in the gill of S. constricta fed with C. vulgaris and T. pseudonana was significantly increased at 1 and/or 3 h. After injection with S. aureus, the expression of ScCL in the gill was significantly increased at 3, 6, and 24 h. These results indicated that ScCL was involved in food particle recognition and immunity of S. constricta.

Cadmium transcriptionally regulates Scd1 expression in silver pomfret.

Cadmium (Cd) is one of the major contaminants in aquatic ecosystem. Stearoyl-coenzyme A desaturase 1 (Scd1) has been implicated in adaptive responses to environmental stressors. The objectives of this study are (a) to characterize scd1 mRNA from silver pomfret (Pampus argenteus); (b) to investigate the expression and activity of Scd1 in silver pomfret exposed to Cd; and (c) to investigate how Cd modifies scd1 gene transcription in silver pomfret. Results indicated that Scd1 was generally conserved across fish species and scd1 mRNA level was higher by far in the brain and liver, followed by the kidney and intestine. Exposure to Cd led to significant changes of the expression and activity of Scd1 in in the liver and intestine. The liver mRNA abundance of scd1 was significantly lower in the Cd-treated groups than in the control group. The 10 days treatment with 1 mg/L Cd significantly upregulated the intestinal scd1 mRNA level, an approximately 9-fold higher in the 1 mg/L Cd-treated group as compared with the control group. Accordingly, Scd1 activity indices (18:1n-9/18:0) in the liver were significantly decreased in the 0.5 mg/L group compared with the control group, while Scd1 activity indices in the intestine were significantly increased in the 1 mg/L group compared with the control group. Moreover, overexpression of sterol regulatory element binding transcription factor 1 (Srebp1) and peroxisome proliferator-activated receptor γ (Pparγ )in HEK 293T cells produced a 2-fold increment in the activity of the scd1 promoter. Furthermore, srebp1 had a similar expression pattern to scd1 in the liver and intestine of silver pomfret exposed to Cd. These results indicated that Cd could regulate scd1 expression, possibly through the transcriptional factor Srebp1.

O-GlcNAcylation promotes colorectal cancer progression by regulating protein stability and potential catcinogenic function of DDX5.

The RNA helicase p68 (DDX5), a key player in RNA metabolism, belongs to the DEAD box family and is involved in the development of colorectal cancer. Here, we found both DDX5 and O-GlcNAcylation are up-regulated in colorectal cancer. In addition, DDX5 protein level is significantly positively correlated with the expression of O-GlcNAcylation. Although it was known DDX5 protein could be regulated by post-translational modification (PTM), how O-GlcNAcylation modification regulated of DDX5 remains unclear. Here we show that DDX5 interacts directly with OGT in the SW480 cell line, which is the only known enzyme that catalyses O-GlcNAcylation in humans. Meanwhile, O-GlcNAcylation could promote DDX5 protein stability. The OGT-DDX5 axis affects colorectal cancer progression mainly by regulating activation of the AKT/mTOR signalling pathway. Taken together, these results indicated that OGT-mediated O-GlcNAcylation stabilizes DDX5, promoting activation of the AKT/mTOR signalling pathway, thus accelerating colorectal cancer progression. This study not only reveals the novel functional of O-GlcNAcylation in regulating DDX5, but also reveals the carcinogenic effect of the OGT-DDX5 axis in colorectal cancer.

DDX21 promotes gastric cancer proliferation by regulating cell cycle.

DEAD (Asp-Glu-Ala-Asp) cassette helicase 21 (DDX21) is an ATP-dependent RNA helicase that is overexpressed in various malignancies. There is increasing evidence that DDX21 is involved in carcinogenesis and cancer progression by promoting cell proliferation. However, the functional role of DDX21 in gastric cancer is largely unknown. In this study, we observed that DDX21 was significantly up-regulated in gastric cancer tissues compared to paired adjacent normal tissues. The expression of DDX21 was closely related to the pathological stage of gastric cancer. In vitro and in vivo studies had shown that knockdown of DDX21 inhibited gastric cancer cell proliferation, colony formation, G1/S cell cycle transition and xenograft growth, while ectopic expression of DDX21 promoted these cell functions. Mechanically, DDX21 induced gastric cancer cell growth by up-regulating levels of Cyclin D1 and CDK2. Taken together, these results revealed a novel role for DDX21 in the proliferation of gastric cancer cells via the Cyclin D1 and CDK2 pathways. Therefore, DDX21 can be used as a therapeutic target for gastric cancer.

miR-5590-3p inhibited tumor growth in gastric cancer by targeting DDX5/AKT/m-TOR pathway.

Recent evidence suggests that microRNAs play important roles in the negative post-transcriptional regulators with altered expression levels found in gastric cancer (GC). Therefore, we employed explore the anti-cancer miRNA and the potential mechanisms by which miRNAs modulate GC progression. We have predicted GC miRNA expression data sets in TargetScan. miR-5590-3p is higher in adjacent nonmalignant tissue than in cancer tissue in 42 pairs of GC tissues. Functional assays, CCK-8 and colony formation assay, were used to determine the Anti-cancer role of miR-5590-3p in human GC progression. In addition, Ago2-based RIP and dual-luciferase reporter assay were conducted to study the miR-5590-3p as a direct target of DDX5. Next, Xenograft nude mouse models were used to determine the role of miR-5590-3p in GC tumorigenicity in vivo. Upregulation of miR-5590-3p suppressed GC cell proliferation, whereas downregulation of miR-5590-3p promoted GC proliferation in vitro. Furthermore, we identified DDX5 as a direct target of miR-5590-3p, and that the biological function of miR-5590-3p during GC progression in vitro and in vivo is through the DDX5/AKT/m-TOR pathway and downstream cyclinD1 and CDK2 expression. Finally, we confirmed the effect of miR-5590-3p directly targeting DDX5 on the development of gastric cancer through salvage experiments in vivo and in vitro.

Leaf stage-associated resistance is correlated with phytohormones in a pathosystem-dependent manner.

It has been reported in several pathosystems that disease resistance can vary in leaves at different stages. However, how general this leaf stage-associated resistance is, and the molecular mechanism(s) underlying it, remain largely unknown. Here, we investigated the effect of leaf stage on basal resistance, effector-triggered immunity (ETI) and nonhost resistance, using eight pathosystems involving the hosts Arabidopsis thaliana, Nicotiana tabacum, and N. benthamiana and the pathogens Sclerotinia sclerotiorum, Pseudomonas syringae pv. tabaci, P. syringae pv. tomato DC3000, and Xanthomonas oryzae pv. oryzae (Xoo). We show evidence that leaf stage-associated resistance exists ubiquitously in plants, but with varying intensity at different stages in diverse pathosystems. Microarray expression profiling assays demonstrated that hundreds of genes involved in defense responses, phytohormone biosynthesis and signaling, and calcium signaling, were differentially expressed between leaves at different stages. The Arabidopsis mutants sid1, sid2-3, ein2, jar1-1, aba1 and aao3 lost leaf stage-associated resistance to S. sclerotiorum, and the mutants aba1 and sid2-3 were affected in leaf stage-associated RPS2/AvrRpt2+ -conferred ETI, whereas only the mutant sid2-3 influenced leaf stage-associated nonhost resistance to Xoo. Our results reveal that the phytohormones salicylic acid, ethylene, jasmonic acid and abscisic acid likely play an essential, but pathosystem-dependent, role in leaf stage-associated resistance.

Tight regulation of the interaction between Brassica napus and Sclerotinia sclerotiorum at the microRNA level.

MicroRNAs (miRNAs) are multifunctional non-coding short nucleotide molecules. Nevertheless, the role of miRNAs in the interactions between plants and necrotrophic pathogens is largely unknown. Here, we report the identification of the miRNA repertoire of the economically important oil crop oilseed rape (Brassica napus) and those involved in interacting with its most devastating necrotrophic pathogen Sclerotinia sclerotiorum. We identified 280 B. napus miRNA candidates, including 53 novel candidates and 227 canonical members or variants of known miRNA families, by high-throughput deep sequencing of small RNAs from both normal and S. sclerotiorum-inoculated leaves. Target genes of 15 novel candidates and 222 known miRNAs were further identified by sequencing of degradomes from the two types of samples. MiRNA microarray analysis revealed that 68 miRNAs were differentially expressed between S. sclerotiorum-inoculated and uninoculated leaves. A set of these miRNAs target genes involved in plant defense to S. sclerotiorum and/or other pathogens such as nucleotide binding site-leucine-rich repeat (NBS-LRR) R genes and nitric oxygen and reactive oxygen species related genes. Additionally, three miRNAs target AGO1 and AGO2, key components of post-transcriptional gene silencing (PTGS). Expression of several viral PTGS suppressors reduced resistance to S. sclerotiorum. Arabidopsis mutants of AGO1 and AGO2 exhibited reduced resistance while transgenic lines over-expressing AGO1 displayed increased resistance to S. sclerotiorum in an AGO1 expression level-dependent manner. Moreover, transient over-expression of miRNAs targeting AGO1 and AGO2 decreased resistance to S. sclerotiorum in oilseed rape. Our results demonstrate that the interactions between B. napus and S. sclerotiorum are tightly regulated at miRNA level and probably involve PTGS.

Curcusone D, a novel ubiquitin-proteasome pathway inhibitor via ROS-induced DUB inhibition, is synergistic with bortezomib against multiple myeloma cell growth.

BACKGROUND: Ubiquitin-proteasome pathway (UPP) plays a very important role in the degradation of proteins. Finding novel UPP inhibitors is a promising strategy for treating multiple myeloma (MM). METHODS: Ub-YFP reporter assays were used as cellular UPP models. MM cell growth, apoptosis and overall death were evaluated with the MTS assay, Annexin V/PI dual-staining flow cytometry, poly (ADP-ribose) polymerase (PARP) cleavage, and PI uptake, respectively. The mechanism of UPP inhibition was analyzed by western blotting for ubiquitin, in vitro and cellular proteasomal and deubiquitinases (DUBs) activity assays. Cellular reactive oxygen species (ROS) were measured with H2DCFDA. RESULTS: Curcusone D, identified as a novel UPP inhibitor, causes cell growth inhibition and apoptosis in MM cells. Curcusone D induced the accumulation of poly-ubiquitin-conjugated proteins but could not inhibit proteasomal activity in vitro or in cells. Interestingly, the mono-ubiquitin level and the total cellular DUB activity were significantly downregulated following curcusone D treatment. Furthermore, curcusone D could induce ROS, which were closely correlated with DUB inhibition that could be nearly completely reversed by NAC. Finally, curcusone D and the proteasomal inhibitor bortezomib showed a strong synergistic effect against MM cells. CONCLUSIONS: Curcusone D is novel UPP inhibitor that acts via the ROS-induced inhibition of DUBs to produce strong growth inhibition and apoptosis of MM cells and synergize with bortezomib. GENERAL SIGNIFICANCE: The anti-MM molecular mechanism study of curcusone D will promote combination therapies with different UPP inhibitors against MM and further support the concept of oxidative stress regulating the activity of DUBs.

Design, synthesis and biological evaluation of novel tripeptidyl epoxyketone derivatives constructed from ß-amino acid as proteasome inhibitors.

A series of novel tripeptidyl epoxyketone derivatives constructed from ß-amino acid were designed, synthesized and evaluated as proteasome inhibitors. All target compounds were tested for their proteasome inhibitory activities and selected compounds were tested for their anti-proliferation activities against two multiple myeloma (MM) cell lines RPMI 8226 and NCI-H929. Among them, eleven compounds exhibited proteasome inhibitory rates of more than 50% at the concentration of 1 µg/mL and nine compounds showed anti-proliferation activities with IC50 values at low micromolar level. Compound 20h displayed the most potent proteasome inhibitory activities (IC50: 0.11 ± 0.01 µM) and anti-proliferation activities with IC50 values at 0.23 ± 0.01 and 0.17 ± 0.02 µM against two tested cell lines. Additionally, the poly-ubiquitin accumulation in the western blot analysis supported that proteasome inhibition in a cellular system was induced by compound 20h. All these experimental results confirmed that ß-amino acid can be introduced as a building block for the development of proteasome inhibitors.

U2-Net and ResNet50-Based Automatic Pipeline for Bacterial Colony Counting.

In this paper, an automatic colony counting system based on an improved image preprocessing algorithm and convolutional neural network (CNN)-assisted automatic counting method was developed. Firstly, we assembled an LED backlighting illumination platform as an image capturing system to obtain photographs of laboratory cultures. Consequently, a dataset was introduced consisting of 390 photos of agar plate cultures, which included 8 microorganisms. Secondly, we implemented a new algorithm for image preprocessing based on light intensity correction, which facilitated clearer differentiation between colony and media areas. Thirdly, a U2-Net was used to predict the probability distribution of the edge of the Petri dish in images to locate region of interest (ROI), and then threshold segmentation was applied to separate it. This U2-Net achieved an F1 score of 99.5% and a mean absolute error (MAE) of 0.0033 on the validation set. Then, another U2-Net was used to separate the colony region within the ROI. This U2-Net achieved an F1 score of 96.5% and an MAE of 0.005 on the validation set. After that, the colony area was segmented into multiple components containing single or adhesive colonies. Finally, the colony components (CC) were innovatively rotated and the image crops were resized as the input (with 14,921 image crops in the training set and 4281 image crops in the validation set) for the ResNet50 network to automatically count the number of colonies. Our method achieved an overall recovery of 97.82% for colony counting and exhibited excellent performance in adhesion classification. To the best of our knowledge, the proposed "light intensity correction-based image preprocessing→U2-Net segmentation for Petri dish edge→U2-Net segmentation for colony region→ResNet50-based counting" scheme represents a new attempt and demonstrates a high degree of automation and accuracy in recognizing and counting single-colony and multi-colony targets.

Drug Exposure and Susceptibility of Pyrazinamide Correlate with Treatment Response in Pyrazinamide-Susceptible Patients with Multidrug-Resistant Tuberculosis.

Exploring the influence of pyrazinamide exposure and susceptibility on treatment response is crucial for optimizing the management of multidrug-resistant tuberculosis (MDR-TB). This study aimed to investigate the association between pyrazinamide exposure, susceptibility, and response to MDR-TB treatment, as well as find clinical thresholds for pyrazinamide. A prospective multi-center cohort study of participants with MDR-TB using pyrazinamide was conducted in three TB-designated hospitals in China. Univariate and multivariate analyses were applied to investigate the associations. Classification and Regression Tree (CART) analysis was used to identify clinical thresholds, which were further evaluated by multivariate analysis and receiver operating characteristic (ROC) curves. The study included 143 patients with MDR-TB. The exposure/susceptibility ratio of pyrazinamide was associated with two-month culture conversion (adjusted risk ratio (aRR), 1.1; 95% confidence interval (CI), 1.07-1.20), six-month culture conversion (aRR, 1.1; 95% CI, 1.06-1.16), treatment success (aRR, 1.07; 95% CI, 1.03-1.10), as well as culture conversion time (adjusted hazard ratio (aHR) 1.18; 95% CI,1.14-1.23). The threshold for optimal improvement in sputum culture results at the sixth month of treatment was determined to be a pyrazinamide AUC0-24h/MIC ratio of 7.8. In conclusion, the exposure/susceptibility ratio of pyrazinamide is associated with the treatment response of MDR-TB, which may change in different Group A drug-based regimens.

Second-line antituberculosis drug exposure thresholds predictive of adverse events in multidrug-resistant tuberculosis treatment.

OBJECTIVES: This study aimed to investigate the association between drug exposure and adverse events (AEs) during the standardized multidrug-resistant tuberculosis (MDR-TB) treatment, as well as to identify predictive drug exposure thresholds. METHODS: We conducted a prospective, observational multicenter study among participants receiving standardized MDR-TB treatment between 2016 and 2019 in China. AEs were monitored throughout the treatment and their relationships to drug exposure (e.g., the area under the drug concentration-time curve from 0 to 24 h, AUC0-24 h) were analyzed. The thresholds of pharmacokinetic predictors of observed AEs were identified by boosted classification and regression tree (CART) and further evaluated by external validation. RESULTS: Of 197 study participants, 124 (62.9%) had at least one AE, and 15 (7.6%) experienced serious AEs. The association between drug exposure and AEs was observed including bedaquiline, its metabolite M2, moxifloxacin and QTcF prolongation (QTcF >450 ms), linezolid and mitochondrial toxicity, cycloserine and psychiatric AEs. The CART-derived thresholds of AUC0-24 h predictive of the respective AEs were 3.2 mg·h/l (bedaquiline M2); 49.3 mg·h/l (moxifloxacin); 119.3 mg·h/l (linezolid); 718.7 mg·h/l (cycloserine). CONCLUSIONS: This study demonstrated the drug exposure thresholds predictive of AEs for key drugs against MDR-TB treatment. Using the derived thresholds will provide the knowledge base for further randomized clinical trials of dose adjustment to minimize the risk of AEs.

Genome-wide identification and functional analyses of calmodulin genes in Solanaceous species.

BACKGROUND: Calmodulin (CaM) is a major calcium sensor in all eukaryotes. It binds calcium and modulates the activity of a wide range of downstream proteins in response to calcium signals. However, little is known about the CaM gene family in Solanaceous species, including the economically important species, tomato (Solanum lycopersicum), and the gene silencing model plant, Nicotiana benthamiana. Moreover, the potential function of CaM in plant disease resistance remains largely unclear. RESULTS: We performed genome-wide identification of CaM gene families in Solanaceous species. Employing bioinformatics approaches, multiple full-length CaM genes were identified from tomato, N. benthamiana and potato (S. tuberosum) genomes, with tomato having 6 CaM genes, N. benthamiana having 7 CaM genes, and potato having 4 CaM genes. Sequence comparison analyses showed that three tomato genes, SlCaM3/4/5, two potato genes StCaM2/3, and two sets of N. benthamiana genes, NbCaM1/2/3/4 and NbCaM5/6, encode identical CaM proteins, yet the genes contain different intron/exon organization and are located on different chromosomes. Further sequence comparisons and gene structural and phylogenetic analyses reveal that Solanaceous species gained a new group of CaM genes during evolution. These new CaM genes are unusual in that they contain three introns in contrast to only a single intron typical of known CaM genes in plants. The tomato CaM (SlCaM) genes were found to be expressed in all organs. Prediction of cis-acting elements in 5' upstream sequences and expression analyses demonstrated that SlCaM genes have potential to be highly responsive to a variety of biotic and abiotic stimuli. Additionally, silencing of SlCaM2 and SlCaM6 altered expression of a set of signaling and defense-related genes and resulted in significantly lower resistance to Tobacco rattle virus and the oomycete pathogen, Pythium aphanidermatum. CONCLUSIONS: The CaM gene families in the Solanaceous species tomato, N. benthamiana and potato were identified through a genome-wide analysis. All three plant species harbor a small set of genes that encode identical CaM proteins, which may manifest a strategy of plants to retain redundancy or enhanced quantitative gene function. In addition, Solanaceous species have evolved one new group of CaM genes during evolution. CaM genes play important roles in plant disease resistance to a variety of pathogens.

Synthesis and biological evaluation of 3-benzisoxazolyl-4-indolylmaleimides as potent, selective inhibitors of glycogen synthase kinase-3ß.

A series of novel 3-benzisoxazolyl-4-indolyl-maleimides were synthesized and evaluated for their GSK-3ß inhibitory activity. Most compounds exhibited high inhibitory potency towards GSK-3ß. Among them, compound 7j with an IC50 value of 0.73 nM was the most promising GSK-3ß inhibitor. Preliminary structure-activity relationships were examined and showed that different substituents on the indole ring and N¹-position of the indole ring had varying degrees of influence on the GSK-3ß inhibitory potency. Compounds 7c, 7f, 7j-l and 7o-q could obviously reduce Aß-induced Tau hyperphosphorylation by inhibiting GSK-3ß in a cell-based functional assay.

In Vivo Electrochemical Measurement of Glucose Variation in the Brain of Early Diabetic Mice.

Diabetes is a chronic disease caused by a decrease in insulin level or insulin resistance. Diabetes also has detrimental effects on the brain, which can lead to the injury of the blood-brain barrier and influence the glucose transport. In this study, we use in vivo electrochemical measurement to explore the glucose variation in the brain of early diabetic mice. The glucose level in mice brain is measured using a carbon fiber microelectrode modified with the osmium-derivatized polymer and glucose oxidase. The electrode shows an excellent electrochemical performance, antibiofouling ability, and high stability, which can work stably in the mice brain for 2 h. By monitoring the glucose level in the brain of normal and diabetic mice after injection of concentrated glucose solution into the abdominal cavity, it is found that the variation of cerebral glucose decreases by ∼2 fold for diabetic mice. It is proposed that diabetes can downregulate the activity of glucose transporter in the brain and finally inhibit the brain glucose uptake.