Mutually enhanced multi-view information learning for segmentation of lung tumor in CT images.

Objective.The accurate automatic segmentation of tumors from computed tomography (CT) volumes facilitates early diagnosis and treatment of patients. A significant challenge in tumor segmentation is the integration of the spatial correlations among multiple parts of a CT volume and the context relationship across multiple channels.Approach.We proposed a mutually enhanced multi-view information model (MEMI) to propagate and fuse the spatial correlations and the context relationship and then apply it to lung tumor CT segmentation. First, a feature map was obtained from segmentation backbone encoder, which contained many image region nodes. An attention mechanism from the region node perspective was presented to determine the impact of all the other nodes on a specific node and enhance the node attribute embedding. A gated convolution-based strategy was also designed to integrate the enhanced attributes and the original node features. Second, transformer across multiple channels was constructed to integrate the channel context relationship. Finally, since the encoded node attributes from the gated convolution view and those from the channel transformer view were complementary, an interaction attention mechanism was proposed to propagate the mutual information among the multiple views.Main results.The segmentation performance was evaluated on both public lung tumor dataset and private dataset collected from a hospital. The experimental results demonstrated that MEMI was superior to other compared segmentation methods. Ablation studies showed the contributions of node correlation learning, channel context relationship learning, and mutual information interaction across multiple views to the improved segmentation performance. Utilizing MEMI on multiple segmentation backbones also demonstrated MEMI's generalization ability.Significance.Our model improved the lung tumor segmentation performance by learning the correlations among multiple region nodes, integrating the channel context relationship, and mutual information enhancement from multiple views.

Deacetylation of ATG7 drives the induction of macroautophagy and LC3-associated microautophagy.

LC3 lipidation plays an important role in the regulation of macroautophagy and LC3-associated microautophagy. The E1-like enzyme ATG7 is one of the core components that are directly involved in LC3 lipidation reaction. Here, we provide evidence showing that acetylation of ATG7 tightly controls its enzyme activity to regulate the induction of macroautophagy and LC3-associated microautophagy. Mechanistically, acetylation of ATG7 disrupts its interaction with the E2-like enzyme ATG3, leading to an inhibition of LC3 lipidation in vitro and in vivo. Functionally, in response to various different stimuli, cellular ATG7 undergoes deacetylation to induce macroautophagy and LC3-associated microautophagy, which are necessary for cells to eliminate cytoplasmic DNA and degrade lysosome membrane proteins, respectively. Taken together, these findings reveal that ATG7 acetylation acts as a critical rheostat in controlling LC3 lipidation and related cellular processes.Abbreviations: AMPK: AMP-activated protein kinase; ATG: autophagy-related; cGAMP: cyclic GMP-AMP; CGAS: cyclic GMP-AMP synthase; CREBBP/CBP: CREB binding protein; EGF: epidermal growth factor; EGFR: epidermal growth factor receptor; EP300/p300: E1A binding protein p300; IFNB1: interferon beta 1; ISD: interferon stimulatory DNA; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MCOLN1/TRPML1: mucolipin TRP cation channel 1; MEF: mouse embryonic fibroblast; MTOR: mechanistic target of rapamycin kinase; NAM: nicotinamide; PE: phosphatidylethanolamine; PTM: post-translational modification; RB1CC1/FIP200: RB1 inducible coiled-coil 1; SIRT: sirtuin; SQSTM1/p62: sequestosome 1; STING1: stimulator of interferon response cGAMP interactor 1; TSA: trichostatin A; ULK1: unc-51 like autophagy activating kinase 1; WIPI2: WD repeat domain, phosphoinositide interacting 2; WT: wild-type.

Emerging roles of p300/CBP in autophagy and autophagy-related human disorders.

As one of the major acetyltransferases in mammalian cells, p300 (also known as EP300) and its highly related protein CBP (also known as CREBBP), collectively termed p300/CBP, is characterized as a key regulator in gene transcription by modulating the acetylation of histones. In recent decades, proteomic analyses have revealed that p300 is also involved in the regulation of various cellular processes by acetylating many non-histone proteins. Among the identified substrates, some are key players involved in different autophagy steps, which together establish p300 as a master regulator of autophagy. Accumulating evidence has shown that p300 activity is controlled by many distinct cellular pathways to regulate autophagy in response to cellular or environmental stimuli. In addition, several small molecules have been shown to regulate autophagy by targeting p300, suggesting that manipulation of p300 activity is sufficient for controlling autophagy. Importantly, dysfunction of p300-regulated autophagy has been implicated in a number of human disorders, such as cancer, aging and neurodegeneration, highlighting p300 as a promising target for the drug development of autophagy-related human disorders. Here, we focus on the roles of p300-mediated protein acetylation in the regulation of autophagy and discuss implications for autophagy-related human disorders.

Surgical management of acute combined injuries of the ipsilateral wrist and elbow joints.

Background: Combined injuries of ipsilateral wrist and elbow joints are rare in clinical practice, characterized by multiple joint dislocations or/and fractures and varying manifestations. As there are still no clinical guidelines and no consensus on the standard treatment, this study aimed to explore the surgical intervention and complications of this kind of combined injuries. Methods: This retrospective study was conducted in a single center. A total of 13 patients with acute combined injuries of the ipsilateral wrist and elbow joints receiving surgical treatment from August 2013 to May 2016 were retrospectively analyzed. The fracture and joint instability and structural damages were repaired and reconstructed. Results: All 13 patients were followed up for a mean duration of 17 months (range: 14 to 22 months). The X-ray films showed good fracture reduction and joint alignment, no fixation failure, re-displacement, bone nonunion, or ischemic necrosis in all cases. According to the Mayo Elbow Performance Score (MEPS), the excellent and good rate of joint function was 84.6%. According to the Mayo Modified Wrist Score (MMWS), the excellent and good rate of joint function was 76.9%. There were no significant restrictions on elbow and wrist movements. The disabilities of the arm, shoulder, and hand (DASH) score was excellent, with an average of 18.5 points. Conclusions: The key to intervention of combined injuries of the wrist and elbow is to identify the types of injuries and conduct an overall assessment to determine the appropriate surgical methods. Early surgical intervention and rehabilitation exercise are the main principles for the treatment.

Acetylation in the regulation of autophagy.

Post-translational modifications, such as phosphorylation, ubiquitination and acetylation, play crucial roles in the regulation of autophagy. Acetylation has emerged as an important regulatory mechanism for autophagy. Acetylation regulates autophagy initiation and autophagosome formation by targeting core components of the ULK1 complex, the BECN1-PIK3C3 complex, and the LC3 lipidation system. Recent studies have shown that acetylation occurs on the key proteins participating in autophagic cargo assembly and autophagosome-lysosome fusion, such as SQSTM1/p62 and STX17. In addition, acetylation controls autophagy at the transcriptional level by targeting histones and the transcription factor TFEB. Here, we review the current knowledge on acetylation of autophagy proteins and their regulations and functions in the autophagy pathway with focus on recent findings.Abbreviations : ACAT1: acetyl-CoA acetyltransferase 1; ACSS2: acyl-CoA synthetase short chain family member 2; AMPK: AMP-activated protein kinase; ATG: autophagy-related; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CCAR2/DBC1: cell cycle and apoptosis regulator 2; BECN1: beclin 1; CMA: chaperone-mediated autophagy; CREBBP/CBP: CREB binding protein; EP300/p300: E1A binding protein p300; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GSK3: glycogen synthase kinase 3; HDAC6: histone deacetylase 6; HSPA8/HSC70: heat shock protein family A (Hsp70) member 8; KAT2A/GCN5: lysine acetyltransferase 2A; KAT2B/PCAF: lysine acetyltransferase 2B; KAT5/TIP60: lysine acetyltransferase 5; KAT8/MOF: lysine acetyltransferase 8; LAMP2A: lysosomal associated membrane protein 2A; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MTOR: mechanistic target of rapamycin kinase; NBR1: NBR1 autophagy cargo receptor; OPTN: optineurin; PD: Parkinson disease; PE: phosphatidylethanolamine; PIK3C3/VPS34: phosphatidylinositol 3-kinase catalytic subunit type 3; PKM2: pyruvate kinase M1/2; PtdIns3P: phosphatidylinositol-3-phosphate; PTM: post-translational modification; RB1CC1/FIP200: RB1 inducible coiled-coil 1; RUBCN/Rubicon: rubicon autophagy regulator; RUBCNL/Pacer: rubicon like autophagy enhancer; SIRT1: sirtuin 1; SNAP29: synaptosome associated protein 29; SNARE: soluble N-ethylamide-sensitive factor attachment protein receptor; SQSTM1/p62: sequestosome 1; STX17: syntaxin 17; TFEB: transcription factor EB; TP53/p53: tumor protein p53; TP53INP2/DOR: tumor protein p53 inducible nuclear protein 2; UBA: ubiquitin-associated; ULK1: unc-51 like autophagy activating kinase 1; VAMP8: vesicle associated membrane protein 8; WIPI2: WD repeat domain, phosphoinositide interacting 2.

Transcriptome Profile Analysis of Intestinal Upper Villus Epithelial Cells and Crypt Epithelial Cells of Suckling Piglets.

It is well known that the small intestinal epithelial cells of mammals rapidly undergo differentiation, maturation, and apoptosis. However, few studies have defined the physiological state and gene expression changes of enterocytes along the crypt-villus axis in suckling piglets. In the present study, we obtained the intestinal upper villus epithelial cells (F1) and crypt epithelial cells (F3) of 21-day suckling piglets using the divalent chelation and precipitation technique. The activities of alkaline phosphatase, sucrase, and lactase of F1 were significantly higher (p < 0.05) than those of F3. To explore the differences at the gene transcription level, we compared the global transcriptional profiles of F1 and F3 using RNA-seq analysis technology. A total of 672 differentially expressed genes (DEGs) were identified between F1 and F3, including 224 highly expressed and 448 minimally expressed unigenes. Functional analyses indicated that some DEGs were involved in the transcriptional regulation of nutrient transportation (SLC15A1, SLC5A1, and SLC3A1), cell differentiation (LGR5, HOXA5 and KLF4), cell proliferation (PLK2 and TGFB3), transcriptional regulation (JUN, FOS and ATF3), and signaling transduction (WNT10B and BMP1), suggesting that these genes were related to intestinal epithelial cell maturation and cell renewal. Gene Ontology (GO) enrichment analysis showed that the DEGs were mainly associated with binding, catalytic activity, enzyme regulator activity, and molecular transducer activity. Furthermore, KEGG pathway analysis revealed that the DGEs were categorized into 284 significantly enriched pathways. The greatest number of DEGs enriched in signal transduction, some of which (Wnt, Hippo, TGF-beta, mTOR, PI3K-Akt, and MAPK signaling pathways) were closely related to the differentiation, proliferation, maturation and apoptosis of intestinal epithelial cells. We validated the expression levels of eight DEGs in F1 and F3 using qRT-PCR. The present study revealed temporal and regional changes in mRNA expression between F1 and F3 of suckling piglets, which provides insights into the regulatory mechanisms underlying intestinal epithelial cell renewal and the rapid repair of intestinal mucosal damage.

Autophagy regulates rRNA synthesis.

Autophagy has emerged as a key regulator of cell metabolism. Recently, we have demonstrated that autophagy is involved in RNA metabolism by regulating ribosomal RNA (rRNA) synthesis. We found that autophagy-deficient cells display much higher 47S precursor rRNA level, which is caused by the accumulation of SQSTM1/p62 (sequestosome 1) but not other autophagy receptors. Mechanistically, SQSTM1 accumulation potentiates the activation of MTOR (mechanistic target of rapamycin kinase) complex 1 (MTORC1) signaling, which facilitates the assembly of RNA polymerase I pre-initiation complex at ribosomal DNA (rDNA) promoter regions and leads to the activation of rDNA transcription. Finally, we showed that SQSTM1 accumulation is responsible for the increase in protein synthesis, cell growth and cell proliferation in autophagy-deficient cells. Taken together, our findings reveal a regulatory role of autophagy and autophagy receptor SQSTM1 in rRNA synthesis and may provide novel mechanisms for the hyperactivated rDNA transcription in autophagy-related human diseases.Abbreviations: 5-FUrd: 5-fluorouridine; LAP: MAP1LC3/LC3-associated phagocytosis; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MTOR: mechanistic target of rapamycin kinase; PIC: pre-initiation complex; POLR1: RNA polymerase I; POLR1A: RNA polymerase I subunit A; rDNA: ribosomal DNA; RRN3: RRN3 homolog, RNA polymerase I transcription factor; rRNA: ribosomal RNA; SQSTM1/p62: sequestosome 1; TP53INP2: tumor protein p53 inducible nuclear protein 2; UBTF: upstream binding transcription factor.

Autophagy deficiency activates rDNA transcription.

Macroautophagy/autophagy, a highly conserved lysosome-dependent degradation pathway, has been intensively studied in regulating cell metabolism by degradation of intracellular components. In this study, we link autophagy to RNA metabolism by uncovering a regulatory role of autophagy in ribosomal RNA (rRNA) synthesis. Autophagy-deficient cells exhibit much higher 47S precursor rRNA level, which is caused by the accumulation of SQSTM1/p62 (sequestosome 1) but not other autophagy receptors. Mechanistically, SQSTM1 accumulation potentiates the activation of MTOR (mechanistic target of rapamycin kinase) complex 1 (MTORC1) signaling and promotes the assembly of RNA polymerase I pre-initiation complex at ribosomal DNA (rDNA) promoters, which leads to an increase of 47S rRNA transcribed from rDNA. Functionally, autophagy deficiency promotes protein synthesis, cell growth and cell proliferation, both of which are dependent on SQSTM1 accumulation. Taken together, our findings suggest that autophagy deficiency is involved in RNA metabolism by activating rDNA transcription and provide novel mechanisms for the reprogramming of cell metabolism in autophagy-related diseases including multiple types of cancers.Abbreviations: 5-FUrd: 5-fluorouridine; AMPK: AMP-activated protein kinase; ATG: autophagy related; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; ChIP: chromatin immunoprecipitation; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAPK/ERK: mitogen-activated protein kinase; MTOR: mechanistic target of rapamycin kinase; NBR1: NBR1 autophagy cargo receptor; NFKB/NF-κB: nuclear factor kappa B; NFE2L2/NRF2: nuclear factor, erythroid 2 like 2; OPTN: optineurin; PIC: pre-initiation complex; POLR1: RNA polymerase I; POLR1A/RPA194: RNA polymerase I subunit A; POLR2A: RNA polymerase II subunit A; rDNA: ribosomal DNA; RPS6KB1/S6K1: ribosomal protein S6 kinase B1; rRNA: ribosomal RNA; RUBCN/Rubicon: rubicon autophagy regulator; SQSTM1/p62: sequestosome 1; STX17: syntaxin 17; SUnSET: surface sensing of translation; TAX1BP1: Tax1 binding protein 1; UBTF/UBF1: upstream binding transcription factor; WIPI2: WD repeat domain, phosphoinositide interacting 2; WT: wild-type.

Removal of atrazine in catalytic degradation solutions by microalgae Chlorella sp. and evaluation of toxicity of degradation products via algal growth and photosynthetic activity.

Degradation solutions containing atrazine need to be further purified before they are discharged into the aquatic environment. With the objectives of evaluating removal capacity of the microalga Chlorella sp. toward atrazine in degradation solutions and toxicity of the degradation products, we investigated the removal efficiency (RE) and bioaccumulation of atrazine in the microalgae after an 8 d exposure to diluted degraded solutions containing 40 µg/L and 80 µg/L of atrazine as well as degradation products in the present study. Moreover, pure atrazine solutions with similar concentrations were simultaneously inoculated with the microalgae in order to distinguish the influence of the products. The photocatalytic degradation results showed that 31.4% of atrazine was degraded after 60 min, and three degradation products, desisopropyl-atrazine (DIA), desethyl-atrazine (DEA), and desethyl-desisopropyl-atrazine (DEIA) were detected. After an 8-d exposure, 83.0% and 64.3% of atrazine were removed from the degraded solutions containing 40 µg/L and 80 µg/L of atrazine, respectively. In comparison with the control, i.e., pure atrazine solution with equal concentration, Chlorella sp. in the degraded atrazine solution showed lower RE and growth rate. The photosynthetic parameters, especially performance index (PIABS), clearly displayed the differences between treatments. The values of PIABS of Chlorella sp. cultured in degradation atrazine for 8 days were significantly lower (P < 0.01) than that in the corresponding pure atrazine, suggesting potential inhibitory effect of degradation products on the microalgae. Atrazine and the degradation products inhibited algal photosynthesis via depressed light absorption and electron transport, and reduced utilization of light energy via energy dissipation. Our results demonstrated that microalgae Chlorella sp. had an encouraging atrazine removal potential and the degradation products of atrazine may inhibit algal growth and removal capability. This study may be useful for the application of microalgae in herbicide wastewater treatment and understanding algal removal of atrazine in natural aquatic environment.

A novel human anti-TIGIT monoclonal antibody with excellent function in eliciting NK cell-mediated antitumor immunity.

TIGIT is an emerging novel checkpoint target that is expressed on both tumor-infiltrating T cells and NK cells. Some current investigational antibodies targeting TIGIT have also achieved dramatic antitumor efficacy in late clinical research. Most recently, the relevance of NK cell-associated TIGIT signaling pathway to tumors' evasion of the immune system has been clearly revealed, which endows NK cells with a pivotal role in the therapeutic effects of TIGIT blockade. In this article, we describe a novel anti-TIGIT monoclonal antibody, AET2010, which was acquired from a phage-displayed human single-chain antibody library through a cell panning strategy. With emphasis on its regulation of NK cells, we confirmed the excellent ex vivo and in vivo antitumor immunity of AET2010 mediated by the NK-92MI cells. Intriguingly, our work also revealed that AET2010 displays a lower affinity but parallel avidity and activity relative to MK7684, an investigational monoclonal antibody from MSD, implying a reasonable balance of potency and potential side effects for AET2010. Together, these results are promising and warrant further development of AET2010.

Identification of curcumin as a novel natural inhibitor of rDNA transcription.

Ribosomal DNA (rDNA) transcription drives cell growth and cell proliferation via the product ribosomal RNA (rRNA), the essential component of ribosome. Given the fundamental role of rRNA in ribosome biogenesis, rDNA transcription has emerged as one of the effective targets for a number of human diseases including various types of cancers. In this study, we identify curcumin, an ancient drug, as a novel natural inhibitor of rDNA transcription. Curcumin treatment impairs the assembly of the RNA polymerase I preinitiation complex at rDNA promoters and represses rDNA promoter activity, which leads to the decrease of rRNA synthesis. In addition, curcumin treatment stimulates autophagosome formation and promotes autophagic degradation in cells. Mechanistically, curcumin inactivates the mechanistic target of rapamycin complex 1 (mTORC1), the upstream regulator of rDNA transcription and autophagy induction, by inhibiting mTOR lysosomal localization. Functionally, curcumin treatment inhibits protein synthesis, cell growth and cell proliferation. Taken together, these findings identify curcumin as an effective inhibitor of rDNA transcription and provide novel mechanisms for the anticancer properties of curcumin. Abbreviations: Atg: autophagy-related; GFP: green fluorescent protein; LAMP2: lysosomal associated membrane protein 2; LC3: microtubule-associated protein 1 light chain 3; MEF: mouse embryonic fibroblast; mTORC1: mechanistic target of rapamycin complex 1; rDNA: ribosomal DNA; rRNA: ribosomal RNA; TP53INP2: tumor protein p53 inducible nuclear protein 2.

Circular RNA hsa_circ_0003496 Contributes to Tumorigenesis and Chemoresistance in Osteosarcoma Through Targeting (microRNA) miR-370/Krüppel-Like Factor 12 Axis.

BACKGROUND: Osteosarcoma (OS) is the most common primary malignancy of bone with a high incidence in children. Circular RNAs (circRNAs) play crucial roles in the carcinogenesis and chemoresistance of OS. In the current work, we focused on the function and mechanism of hsa_circ_0003496 (circ_0003496) in OS progression and chemoresistance. MATERIALS AND METHODS: The expression levels of circ_0003496, miR-370 and Krüppel-like factor 12 (KLF12) mRNA were measured by quantitative real-time polymerase chain reaction (qRT-PCR). The Cell Counting Kit-8 (CCK-8) assay was used to assess the 50% inhibitory concentration (IC50) value and cell proliferation. Cell migration, invasion and apoptosis were detected by transwell assay and flow cytometry, respectively. Western blot analysis was performed to assess the protein level. Targeted relationships among circ_0003496, miR-370 and KLF12 were validated by dual-luciferase reporter, RNA immunoprecipitation (RIP) and RNA pull-down assays. Animal studies were carried out to observe the role of circ_0003496 in vivo. RESULTS: Our results indicated that circ_0003496 up-regulation was associated with doxorubicin (DXR) resistance of OS. Circ_0003496 knockdown repressed DXR-resistant OS cell proliferation, migration and invasion, and enhanced apoptosis and DXR sensitivity. Circ_0003496 functioned as a sponge of miR-370, and miR-370 mediated the regulatory effect of circ_0003496 depletion on DXR-resistant OS cell progression and DXR sensitivity. KLF12 was a direct target of miR-370, and miR-370 overexpression suppressed cell progression and enhanced DXR sensitivity by KLF12. Moreover, circ_0003496 protected against KLF12 repression through sponging miR-370. Additionally, circ_0003496 knockdown hampered tumor growth and promoted DXR sensitivity in vivo. CONCLUSION: Our present work suggested that the knockdown of circ_0003496 suppressed OS progression and enhanced DXR sensitivity at least partially through modulating KLF12 expression via functioning as a miR-370 sponge, highlighting new opportunities for OS management.

Effects of carbon nanotubes on the toxicities of copper, cadmium and zinc toward the freshwater microalgae Scenedesmus obliquus.

Due to their unique structure and properties, carbon nanotubes (CNTs) released into the aquatic environment can potentially influence the behavior of other coexisting pollutants, thereby altering their toxicity to aquatic organisms. In this study, the toxicities of multi-walled CNTs and three heavy metals, copper (Cu), cadmium (Cd) and zinc (Zn) were determined individually. Following this, CNTs with low concentrations (1 and 5 mg/L) were co-exposed with Cu, Cd or Zn to the microalgae Scenedesmus obliquus, to investigate the effects and underlying mechanisms of CNTs on metal toxicity. Results showed that CNTs, especially at a concentration of 5 mg/L, promoted algae growth and enhanced photosynthetic efficiency via increasing exciton trap efficiency and quantum yield for electron transport. Introduction of CNTs appeared to alleviate the adverse effects of Cu, Cd or Zn on microalgae, indicated by algae growth, total chlorophyll content and photosynthetic indices. However, these effects differed greatly for different metals, depending on both the toxicity of each metal and the exposure period (4 day and 8 day). Enhancement of photosynthesis and interference of metal uptake by CNTs, have a crucial role in the effects of CNTs on metal toxicity.

The bifunctional role of TP53INP2 in transcription and autophagy.

Cells integrate intracellular and extracellular cues to precisely control the balance of anabolic and catabolic processes, which is essential for cells to maintain homeostasis. The nuclear protein TP53INP2 (tumor protein p53 inducible nuclear protein 2) has emerged as one of the key players participating in both anabolic and catabolic processes. In the nucleus including the nucleolus, TP53INP2 binds to multiple transcription-related factors to modulate transcription, such as the transcription of thyroid hormone-related genes and ribosomal DNA. Interestingly, upon nutrient deprivation, TP53INP2 rapidly moves from the nucleus to the cytoplasm and participates in the regulation of macroautophagy/autophagy. By acting as a nutrient status sensor, TP53INP2 switches its role between transcription and autophagy by changing its subcellular localization and helps the cell to cope with environmental changes. ABBREVIATIONS: Atg: autophagy related; LIR: LC3-interacting region; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MTORC1: mechanistic target of rapamycin kinase complex 1; rDNA: ribosomal DNA; TP53INP2: tumor protein p53 inducible nuclear protein 2; UIM: ubiquitin-interacting motif.

To evaluate the toxicity of atrazine on the freshwater microalgae Chlorella sp. using sensitive indices indicated by photosynthetic parameters.

Atrazine is a widely-applied herbicide used primarily to control weeds, which can persist in the ecosystem and exert potential toxicity to phytoplankton in the aquatic environment. In this study, acute toxicity of atrazine on microalgae Chlorella sp. was investigated with different initial cell densities (1 × 105 and 1 × 106 cells mL-1) and exposure periods (4 d and 8 d). Both growth rate and photosynthetic parameters of the microalgae in response of atrazine stress were determined to find out the sensitive indices and toxicological mechanisms. Because of the independence of initial cell density as well as the high sensitivity and reliability, the performance index PIABS was verified as the most convincing photosynthetic parameter for indicating IC50 of atrazine on Chlorella sp., being superior to the traditional parameters of growth rate and FV/FM. The IP amplitude (ΔFIP, fluorescence amplitude of the I-to-P-rise in the OJIP curve) was another sensitive biomarker to reflect atrazine stress. Results from chlorophyll fluorescence transient revealed that atrazine damaged the photosystem II (PS II) reaction center, suppressed the electron transport at the donor and receptor sides, and acted on the absorption, transfer, and utilization of light energy. Our results provide confirmatory references for understanding the toxicity and mechanisms of atrazine on freshwater microalgae.

Analgesic, anti-inflammatory and sedative/hypnotic effects of Icaritin, and its effect on chloride influx in mouse brain cortical cells.

Inflammation and insomnia are medical problems that may severely affect work and health, thereby necessitating strategies for their effective treatment. Icartin (ICT) is a major active monomeric component of icariin  . Studies have revealed that ICT possesses several pharmacological properties such anti-inflammatory, anti-tumor, anti-fibrotic, anti-osteoporotic and neuroprotective effects. The present research was carried out to investigate the anti-inflammatory, analgesic and sedative/hypnotic effects of ICT. The results obtained revealed that ICT exerted a good anti-inflammatory effect related to the downregulations of inflammatory cytokines and the inhibition of COX-2 signaling pathway. Moreover, ICT enhanced Cl- influx in mouse cortical cells in a concentration-dependent manner. These data suggest that ICT exerts a hypnotic effect in mice through a mechanism associated with increased Cl- influx in cortical cells.

TP53INP2 mediates autophagic degradation of ubiquitinated proteins through its ubiquitin-interacting motif.

The tumor protein p53-inducible nuclear protein 2 (TP53INP2) has been reported to participate in autophagy by interacting with autophagosome-localized autophagy-related protein 8 (Atg8) family proteins, including LC3. Here, we uncover a novel function for TP53INP2 in the autophagic degradation of proteins. We identify the ubiquitin-interacting motif (UIM) of TP53INP2 that mediates its binding to ubiquitin and ubiquitinated proteins. TP53INP2 lacking the UIM is able to displace autophagic adaptor p62 from LC3, which leads to accumulation of ubiquitinated proteins in cells. Furthermore, overexpression of TP53INP2 lacking the UIM sensitizes cells to chloroquine treatment. Our findings indicate that TP53INP2 may act as a novel autophagic adaptor through recruiting ubquitinated substrates to autophagosomes for degradation.

Gefitinib with or without Transarterial Infusion Chemotherapy (Cisplatin) for Large Nonsmall Cell Lung Cancer with Epidermal Growth Factor Receptor Mutations.

PURPOSE: To retrospectively investigate the safety and benefit of gefitinib plus transarterial infusion (TAI) therapy as a first-line treatment compared to gefitinib alone for patients with large (>7 cm) nonsmall cell lung cancer (NSCLC) with epidermal growth factor receptor (EGFR) mutations. MATERIALS AND METHODS: Between January 2010 and December 2013, 92 consecutive treatment-naïve patients with large NSCLC with EGFR mutations, who were treated using gefitinib plus TAI (G+T, n = 42) or gefitinib alone (G, n = 50) were reviewed. The primary endpoints were the objective response rate (ORR) and tumor reduction rate. The secondary endpoints were progression-free survival (PFS) and overall survival (OS), and safety was also assessed. RESULTS: The baseline characteristics of the 2 groups were balanced, and no patients experienced treatment-related death. Toxicity outcomes did not differ between the G+T and G groups. The tumor reduction rate in the G+T group was significantly higher than that in the G group (42.9 vs 31.9%, P = .028). The ORR was 83% in the G+T group and 72% in the G group (P = .197). The median PFS was significantly longer in the G+T group than in the G group (14.0 vs 10.0 months, P = .023). The median OS was 30.0 months in the G+T group and 27.0 months in the G group (P = .235). CONCLUSIONS: This study suggests that compared with gefitinib alone, combination therapy with gefitinib plus TAI was well tolerated and potentially improved the tumor reduction rate and PFS in patients with large NSCLC with EGFR mutations.

TP53INP2 contributes to autophagosome formation by promoting LC3-ATG7 interaction.

TP53INP2/DOR (tumor protein p53-inducible nuclear protein 2) contributes to mammalian macroautophagy/autophagy by carrying nuclear deacetylated MAP1LC3/LC3 to the cytoplasm. Here, we report that in the cytoplasm, TP53INP2 further functions in autophagosome biogenesis by promoting LC3B-ATG7 interaction. Cytoplasmic expression of the N-terminal region of TP53INP2, which includes the LC3-interacting region, effectively triggered LC3B-PE production and autophagosome formation. In the cytoplasm, TP53INP2 colocalized to early autophagic membrane structures containing ATG14, ZFYVE1/DFCP1 or WIPI2. While knockdown of TP53INP2 did not affect the formation of these autophagic structures, deletion of BECN1 or Atg5, or mutations preventing TP53INP2 from LC3 interaction, disrupted the membrane binding of TP53INP2. TP53INP2 interacted directly with ATG7 to form a LC3B-TP53INP2-ATG7 complex in the cytoplasm. Loss of TP53INP2-LC3 or TP53INP2-ATG7 interaction significantly reduced LC3B-ATG7 binding. Together, these results suggest that after shifting from the nucleus, cytoplasmic TP53INP2 is targeted to early autophagic membranes accompanied by LC3, where it contributes to autophagosome biogenesis by mediating LC3-ATG7 interaction. Abbreviations: 3-MA, 3-methyladenine; 3NES, 3 repeated nuclear export signal; 3NLS, 3 repeated nuclear localization signal; ACTB, actin beta; ATG, autophagy related; BECN1, Beclin 1; mCherry, monomeric red fluorescent protein mCherry; GFP, green fluorescent protein; GST, glutathione S-transferase; KO, knockout; LC3B/MAP1LC3B, microtubule-associated protein 1 light chain 3 beta; LC3B[G120], LC3B mutant lacking amino acids after glycine 120; LDH, lactate dehydrogenase; LMNB1, lamin B1; LIR, LC3-interacting region; MTORC1, mechanistic target of rapamycin complex 1; PE, phosphatidylethanolamine; PtdIns3K, phosphatidylinositol 3-kinase; PtdIns3P, phosphatidylinositol 3-phosphate; rDNA, ribosomal DNA; RFP, red fluorescent protein; RNAi, RNA interference; SQSTM1, sequestosome 1; TP53INP2, tumor protein p53-inducible nuclear protein 2; TP53INP2[1-28], TP53INP2 mutant containing amino acids 1 to 28; TP53INP2[28-45], TP53INP2 mutant containing amino acids 28 to 45; TP53INP2[LIRΔ], TP53INP2 mutant lacking amino acids 1 to 144; TP53INP2[NLSΔ], TP53INP2 mutant lacking amino acids 145 to 221; TP53INP2W35,I38A, TP53INP2 mutant in which tryptophan 35 and isoleucine 38 are replaced with alanine; TP53INP2W35,I38A[NLSΔ], TP53INP2 mutant lacking amino acids 145 to 221, and tryptophan 35 and isoleucine 38 are replaced with alanine; TP53INP2W35,I38A[Δ1-28],[NLSΔ], TP53INP2 mutant lacking amino acids 1 to 28 and amino acids 145 to 221, and tryptophan 35 and isoleucine 38 are replaced with alanine; TP53INP2[Δ1-28],[NLSΔ], TP53INP2 mutant lacking amino acids 1 to 28 and amino acids 145 to 221; TP53INP2[Δ67-111],[NLSΔ], TP53INP2 mutant lacking amino acids 67 to 111 and amino acids 145 to 221; TP53INP2[Δ112-144],[NLSΔ], TP53INP2 mutant lacking amino acids 112 to 144 and amino acids 145 to 221; TUBB, tubulin beta class I; ULK1, unc-51 like autophagy activating kinase 1; VMP1, vacuole membrane protein 1; WIPI2, WD repeat domain phosphoinositide-interacting 2; WT, wild-type; ZFYVE1/DFCP1, zinc finger FYVE-type containing 1.

mTORC1-Regulated and HUWE1-Mediated WIPI2 Degradation Controls Autophagy Flux.

mTORC1, the major homeostatic sensor and responder, regulates cell catabolism mainly by targeting autophagy. Here, we show that mTORC1 directly controls autophagosome formation via phosphorylation of WIPI2, a critical protein in isolation membrane growth and elongation. mTORC1 phosphorylates Ser395 of WIPI2, directing WIPI2 to interact specifically with the E3 ubiquitin ligase HUWE1 for ubiquitination and proteasomal degradation. Physiological or pharmacological inhibition of mTORC1 in cells promotes WIPI2 stabilization, autophagosome formation, and autophagic degradation. In mouse liver, fasting significantly increases the WIPI2 protein level, while silencing HUWE1 enhances autophagy, and introducing WIPI2 improves lipid clearance. Thus, regulation of the intracellular WIPI2 protein level by mTORC1 and HUWE1 is a key determinant of autophagy flux and may coordinate the initiation, progression, and completion of autophagy.