JCAD deficiency delayed liver regenerative repair through the Hippo-YAP signalling pathway.

BACKGROUND AND AIMS: Liver regeneration retardation post partial hepatectomy (PH) is a common clinical problem after liver transplantation. Identification of key regulators in liver regeneration post PH may be beneficial for clinically improving the prognosis of patients after liver transplantation. This study aimed to clarify the function of junctional protein-associated with coronary artery disease (JCAD) in liver regeneration post PH and to reveal the underlying mechanisms. METHODS: JCAD knockout (JCAD-KO), liver-specific JCAD-KO (Jcad△Hep) mice and their control group were subjected to 70% PH. RNA sequencing was conducted to unravel the related signalling pathways. Primary hepatocytes from KO mice were treated with epidermal growth factor (EGF) to evaluate DNA replication. Fluorescent ubiquitination-based cell cycle indicator (FUCCI) live-imaging system was used to visualise the phases of cell cycle. RESULTS: Both global and liver-specific JCAD deficiency postponed liver regeneration after PH as indicated by reduced gene expression of cell cycle transition and DNA replication. Prolonged retention in G1 phase and failure to transition over the cell cycle checkpoint in JCAD-KO cell line was indicated by a FUCCI live-imaging system as well as pharmacologic blockage. JCAD replenishment by adenovirus reversed the impaired DNA synthesis in JCAD-KO primary hepatocyte in exposure to EGF, which was abrogated by a Yes-associated protein (YAP) inhibitor, verteporfin. Mechanistically, JCAD competed with large tumour suppressor 2 (LATS2) for WWC1 interaction, leading to LATS2 inhibition and thereafter YAP activation, and enhanced expression of cell cycle-associated genes. CONCLUSION: JCAD deficiency led to delayed regeneration after PH as a result of blockage in cell cycle progression through the Hippo-YAP signalling pathway. These findings uncovered novel functions of JCAD and suggested a potential strategy for improving graft growth and function post liver transplantation. KEY POINTS: JCAD deficiency leads to an impaired liver growth after PH due to cell division blockage. JCAD competes with LATS2 for WWC1 interaction, resulting in LATS2 inhibition, YAP activation and enhanced expression of cell cycle-associated genes. Delineation of JCADHippoYAP signalling pathway would facilitate to improve prognosis of acute liver failure and graft growth in living-donor liver transplantation.

The Hippo signaling pathway in development and regeneration.

The Hippo signaling pathway is a central growth control mechanism in multicellular organisms. By integrating diverse mechanical, biochemical, and stress cues, the Hippo pathway orchestrates proliferation, survival, differentiation, and mechanics of cells, which in turn regulate organ development, homeostasis, and regeneration. A deep understanding of the regulation and function of the Hippo pathway therefore holds great promise for developing novel therapeutics in regenerative medicine. Here, we provide updates on the molecular organization of the mammalian Hippo signaling network, review the regulatory signals and functional outputs of the pathway, and discuss the roles of Hippo signaling in development and regeneration.

WWC1/2 regulate spinogenesis and cognition in mice by stabilizing AMOT.

WWC1 regulates episodic learning and memory, and genetic nucleotide polymorphism of WWC1 is associated with neurodegenerative diseases such as Alzheimer's disease. However, the molecular mechanism through which WWC1 regulates neuronal function has not been fully elucidated. Here, we show that WWC1 and its paralogs (WWC2/3) bind directly to angiomotin (AMOT) family proteins (Motins), and recruit USP9X to deubiquitinate and stabilize Motins. Deletion of WWC genes in different cell types leads to reduced protein levels of Motins. In mice, neuron-specific deletion of Wwc1 and Wwc2 results in reduced expression of Motins and lower density of dendritic spines in the cortex and hippocampus, in association with impaired cognitive functions such as memory and learning. Interestingly, ectopic expression of AMOT partially rescues the neuronal phenotypes associated with Wwc1/2 deletion. Thus, WWC proteins modulate spinogenesis and cognition, at least in part, by regulating the protein stability of Motins.

A novel calcium peroxide/attapulgite-Fe(II) process for high concentration phosphate removal and recovery: Efficiency and mechanism.

Phosphorus (P) has been overused in livestock farming, which inevitably results in high-concentration P-containing wastewater. Managing total phosphorus discharge is important to prevent eutrophication in aquatic environments, thus it is critical to develop new technologies for the removal and recovery of high-concentration phosphate. In this study, a novel calcium peroxide/attapulgite (CP/ATP) composite was developed and coupled with Fe(II) for high-concentration phosphate removal and recovery. The results demonstrated that the optimal dosage of the CP/ATP-Fe(II) process was CP/ATP = 0.25 g/L and Fe(II) = 2 mM. The pH effect on phosphate removal was minimal, while phosphate removal efficiency rose by 16.7% with the temperature increased from 10 °C to 25 °C. The co-existing ions exhibited little effect on phosphate removal, and the CP/ATP-Fe(II) process showed effective phosphate removal from the real piggery wastewater. The P content of the precipitates after phosphate removal by this process was as high as 25.82%, indicating its good potential for P recycling. A significant synergistic effect existed in CP/ATP and Fe(II) for phosphate removal, and the SEM-EDS, XRD, Raman and XPS characterization exhibited that the phosphate removal mainly relied on the in-situ-formed Fe(III) and the participation of calcium (Ca) species. Co-precipitation was the predominant mechanism for phosphate removal, and the proportions of Fe(III)-P, Ca-P and Ca-Fe(III)-P in the precipitates were 51.5%, 31.2% and 17.3%, respectively. This study provides a highly efficient process for phosphate removal and recovery from wastewater, and insights into interactions among phosphorus, iron and calcium.

Two Hippo signaling modules orchestrate liver size and tumorigenesis.

The Hippo pathway is a central regulator of organ size and tumorigenesis and is commonly depicted as a kinase cascade, with an increasing number of regulatory and adaptor proteins linked to its regulation over recent years. Here, we propose that two Hippo signaling modules, MST1/2-SAV1-WWC1-3 (HPO1) and MAP4K1-7-NF2 (HPO2), together regulate the activity of LATS1/2 kinases and YAP/TAZ transcriptional co-activators. In mouse livers, the genetic inactivation of either HPO1 or HPO2 module results in partial activation of YAP/TAZ, bile duct hyperplasia, and hepatocellular carcinoma (HCC). On the contrary, inactivation of both HPO1 and HPO2 modules results in full activation of YAP/TAZ, rapid development of intrahepatic cholangiocarcinoma (iCCA), and early lethality. Interestingly, HPO1 has a predominant role in regulating organ size. HPO1 inactivation causes a homogenous YAP/TAZ activation and cell proliferation across the whole liver, resulting in a proportional and rapid increase in liver size. Thus, this study has reconstructed the order of the Hippo signaling network and suggests that LATS1/2 and YAP/TAZ activities are finetuned by HPO1 and HPO2 modules to cause different cell fates, organ size changes, and tumorigenesis trajectories.

Tracing the variation of dissolved organic matter in the two-stage anoxic/aerobic process treating swine wastewater using fluorescence excitation-emission matrix with parallel factor analysis.

Swine wastewater has become one of the main agricultural pollution sources. Quantitative characterization of dissolved organic matter (DOM) is often used in various water bodies, but there are few studies on DOM analysis of swine wastewater. In this study, swine wastewater was treated by a step-feed two-stage anoxic/aerobic (SF-A/O/A/O) process. By using parallel factor (PARAFAC) analysis of fluorescence excitation-emission matrix (EEM), the main components of swine wastewater were aromatic protein-like substances (C1), tryptophan-like substances (C2), fulvic acid-like/humic-like substances (C3) and humic-like substances (C4). Protein-like substances were degraded significantly, while humic-like substances were difficult to be utilized by microorganisms. Fluorescence spectral indexes showed that the characteristics of endogenous input and humus were enhanced. Moreover, several significant correlations between DOM components, fluorescence spectral indexes and water quality indexes were observed. These findings help to understand the biochemical role and the impact of DOM in water quality monitoring and control of swine wastewater.

Stellettin B Sensitizes Glioblastoma to DNA-Damaging Treatments by Suppressing PI3K-Mediated Homologous Recombination Repair.

Glioblastoma (GBM) is the most aggressive type of cancer. Its current first-line postsurgery regimens are radiotherapy and temozolomide (TMZ) chemotherapy, both of which are DNA damage-inducing therapies but show very limited efficacy and a high risk of resistance. There is an urgent need to develop novel agents to sensitize GBM to DNA-damaging treatments. Here it is found that the triterpene compound stellettin B (STELB) greatly enhances the sensitivity of GBM to ionizing radiation and TMZ in vitro and in vivo. Mechanistically, STELB inhibits the expression of homologous recombination repair (HR) factors BRCA1/2 and RAD51 by promoting the degradation of PI3Kα through the ubiquitin-proteasome pathway; and the induced HR deficiency then leads to augmented DNA damage and cell death. It is further demonstrated that STELB has the potential to rapidly penetrate the blood-brain barrier to exert anti-GBM effects in the brain, based on zebrafish and nude mouse orthotopic xenograft tumor models. The study provides strong evidence that STELB represents a promising drug candidate to improve GBM therapy in combination with DNA-damaging treatments.

Distribution of an antioxidant in polypropylene/ethylene-octene copolymer blends studied by atomic force microscopy-infrared.

In this work, the microscopic distribution of antioxidant 1010 (AT1010) in blends of isotactic polypropylene (iPP) with an ethylene-octene copolymer (POE) was investigated in situ using the atomic force microscopy-infrared (AFM-IR) technique. Pellets of an iPP/POE blend containing AT1010 at a mass ratio of 79.5 : 20 : 0.5 were extruded at different screw speeds, and were then injection-molded into plates. The domain size of the POE disperse phase in the pellets was about 1 µm, regardless of the screw speed, and remained unchanged in the injection molding. AFM-IR analyses revealed that AT1010 preferred to stay in the POE disperse phase rather than in the iPP matrix, with a concentration ratio of ∼1.2 in the extruded pellets independent of the screw speed, which was further increased to ∼1.5 in the molded plates. The preferred concentration of AT1010 in the POE was in line with its higher solubility in rubber than in iPP, and the enhanced partition of AT1010 in the molded plates was attributed to a longer processing time in the molten state than that for the extruded pellets, which was verified by AFM-IR analyses of pellets extruded with different residence times.

Transmembrane protein KIRREL1 regulates Hippo signaling via a feedback loop and represents a therapeutic target in YAP/TAZ-active cancers.

The Hippo tumor-suppressor pathway is frequently dysregulated in human cancers and represents a therapeutic target. However, strategies targeting the mammalian Hippo pathway are limited because of the lack of a well-established cell-surface regulator. Here, we show that transmembrane protein KIRREL1, by interacting with both SAV1 and LATS1/2, promotes LATS1/2 activation by MST1/2 (Hippo kinases), and LATS1/2 activation, in turn, inhibits activity of YAP/TAZ oncoproteins. Conversely, YAP/TAZ directly induce the expression of KIRREL1 in a TEAD1-4-dependent manner. Indeed, KIRREL1 expression positively correlates with canonical YAP/TAZ target gene expression in clinical tumor specimens and predicts poor prognosis. Moreover, transgenic expression of KIRREL1 effectively blocks tumorigenesis in a mouse intrahepatic cholangiocarcinoma model, indicating a tumor-suppressor role of KIRREL1. Hence, KIRREL1 constitutes a negative feedback mechanism regulating the Hippo pathway and serves as a cell-surface marker and potential drug target in cancers with YAP/TAZ dependency.

WWC proteins mediate LATS1/2 activation by Hippo kinases and imply a tumor suppression strategy.

YAP and TAZ (YAP/TAZ), two major effectors of the Hippo signaling pathway, are frequently activated in human cancers. The activity of YAP/TAZ is strictly repressed upon phosphorylation by LATS1/2 tumor suppressors. However, it is unclear how LATS1/2 are precisely regulated by upstream factors such as Hippo kinases MST1/2. Here, we show that WWC proteins (WWC1/2/3) directly interact with LATS1/2 and SAV1, and SAV1, in turn, brings in MST1/2 to phosphorylate and activate LATS1/2. Hence, WWC1/2/3 play an organizer role in a signaling module that mediates LATS1/2 activation by MST1/2. Moreover, we have defined a minimum protein interaction interface on WWC1/2/3 that is sufficient to activate LATS1/2 in a robust and specific manner. The corresponding minigene, dubbed as SuperHippo, can effectively suppress tumorigenesis in multiple tumor models. Our study has uncovered a molecular mechanism underlying LATS1/2 regulation and provides a strategy for treating diverse malignancies related to Hippo pathway dysregulation.

Hydroxychloroquine synergizes with the PI3K inhibitor BKM120 to exhibit antitumor efficacy independent of autophagy.

BACKGROUND: The critical role of phosphoinositide 3-kinase (PI3K) activation in tumor cell biology has prompted massive efforts to develop PI3K inhibitors (PI3Kis) for cancer therapy. However, recent results from clinical trials have shown only a modest therapeutic efficacy of single-agent PI3Kis in solid tumors. Targeting autophagy has controversial context-dependent effects in cancer treatment. As a FDA-approved lysosomotropic agent, hydroxychloroquine (HCQ) has been well tested as an autophagy inhibitor in preclinical models. Here, we elucidated the novel mechanism of HCQ alone or in combination with PI3Ki BKM120 in the treatment of cancer. METHODS: The antitumor effects of HCQ and BKM120 on three different types of tumor cells were assessed by in vitro PrestoBlue assay, colony formation assay and in vivo zebrafish and nude mouse xenograft models. The involved molecular mechanisms were investigated by MDC staining, LC3 puncta formation assay, immunofluorescent assay, flow cytometric analysis of apoptosis and ROS, qRT-PCR, Western blot, comet assay, homologous recombination (HR) assay and immunohistochemical staining. RESULTS: HCQ significantly sensitized cancer cells to BKM120 in vitro and in vivo. Interestingly, the sensitization mediated by HCQ could not be phenocopied by treatment with other autophagy inhibitors (Spautin-1, 3-MA and bafilomycin A1) or knockdown of the essential autophagy genes Atg5/Atg7, suggesting that the sensitizing effect might be mediated independent of autophagy status. Mechanistically, HCQ induced ROS production and activated the transcription factor NRF2. In contrast, BKM120 prevented the elimination of ROS by inactivation of NRF2, leading to accumulation of DNA damage. In addition, HCQ activated ATM to enhance HR repair, a high-fidelity repair for DNA double-strand breaks (DSBs) in cells, while BKM120 inhibited HR repair by blocking the phosphorylation of ATM and the expression of BRCA1/2 and Rad51. CONCLUSIONS: Our study revealed that HCQ and BKM120 synergistically increased DSBs in tumor cells and therefore augmented apoptosis, resulting in enhanced antitumor efficacy. Our findings provide a new insight into how HCQ exhibits antitumor efficacy and synergizes with PI3Ki BKM120, and warn that one should consider the "off target" effects of HCQ when used as autophagy inhibitor in the clinical treatment of cancer.

Integrating sulfur, iron(II), and fixed organic carbon for mixotrophic denitrification in a composite filter bed reactor for decentralized wastewater treatment: Performance and microbial community.

Decentralized wastewater treatment in rural areas is an imperative challenge around the world, particularly in developing countries. The composite filter bed reactor is viable for decentralized wastewater treatment, but its performance on nitrogen removal often fluctuates with the unstable influent characteristics and loadings. Here, a composite filter bed reactor integrating sulfur, iron(II), and fixed organic carbon (shaddock peel) was developed and continuously operated under different conditions. The fixed organic carbon source promoted nitrogen removal with an efficiency higher than 90% and reduced effluent sulfate level by 40%, indicating that the integrated electron donors could improve the resistance and stability of the reactor. Moreover, sulfur-oxidizing bacteria (Thiomonas, Sulfuriferula, and Acidithiobacillus), iron-oxidizing bacteria (Ferritrophicum), and denitrifiers (Simplicispira and Hydrogenophaga) were identified in the anoxic/anaerobic layer of the reactor, suggesting that mixotrophic denitrification was stimulated by sulfur, iron(II), and fixed organic carbon. The findings of this study indicate that the developed reactor with the integrated electron donors could be reliable for carbon, nitrogen, and phosphorus removal and promising for the application of decentralized wastewater treatment.

Phosphate sequestration by magnetic La-impregnated bentonite granules: A combined experimental and DFT study.

To use the lanthanum hydroxide (La(OH)3) as a low-cost, highly-efficient, and recyclable adsorbent, it could be embedded on a magnetic substance to improve its physical features and lower the overall cost. Herein, novel millimetric-size magnetic lanthanum-modified bentonite (La-MB) granules were fabricated for P sequestration, and the adsorption performance and mechanisms were systematic studied. The maximum capacity of P uptake by La-MB was up to 48.4 mg/g, which was higher than many previous reported La-based adsorbents. Moreover, the enhanced uptake of P was achieved over a wide pH range (3-9) and in the coexistence of common anions (Cl-, NO3-, and SO42-). Besides, the exhausted La-MB can be effectively regenerated by 5 mol/L NaOH with about 94.5% desorption efficiency and 60.8% uptake capacity remained during 5 cycles. The La-MB also exhibited excellent performance of anti-interference in two kinds of real wastewaters. The postsorption characterization and DFT calculations revealed that the electrostatic interaction and chemical precipitation jointly facilitated phosphate sequestration by La-MB during the rapid sorption phase, while ligand exchange and complexation reaction played more important roles than others during the slow sorption step. The electrostatic interaction not only effectively promoted the ligand exchange, and also further accelerated chemical precipitation via the formation of LaPO4 during the whole process of phosphate uptake. Overall, millimetric La-MB is considered to have great potential for engineering application, and this work also provides new insights into the molecular-level mechanism of phosphate sequestration by La-MB.

Transparent and Scratch-Resistant Antifogging Coatings with Rapid Self-Healing Capability.

Typical antifogging coatings based on hydrophilic polymers are soft and susceptible to mechanical damage. In this paper, an antifogging coating that is both scratch-resistant and self-healing is fabricated by copolymerizing sulfobetaine methacrylate and 2-hydroxyethyl methacrylate in the presence of sulfobetaine-modified silica nanoparticles in one pot. The coating is highly efficient in preventing fog formation at the surface and reducing ice adhesion, and is resistant to fouling by oil and protein, due to the strong hydration ability of the zwitterionic moieties. The composite coating is resistant to scratching and abrasion under normal use conditions to maintain its transparency due to increased hardness by the filled silica nanoparticles and is able to heal completely within several minutes severe scratches and cuts inflicted in harsh conditions, owing to the water-assisted reversibility of the electrostatic and hydrogen-bonding interactions holding together the polymer components and the silica nanoparticles. The multiple desirable properties demonstrated and the simple fabrication process of the coating offers great potential in many practical applications.

Enhanced phosphate sequestration by Fe(iii) modified biochar derived from coconut shell.

In this work, a novel Fe-modified coconut shell biochar (Fe-CSB) was synthesized and utilized to remove phosphate from aqueous solution. Characterization results confirmed that the iron in the Fe(iii)-impregnated CSB existed mainly in the amorphous phase, as ferrihydrite and amorphous hydroxide, which substantially enhanced the phosphate adsorption. Batch experiments indicated that phosphate adsorption on the Fe-CSB was highly dependent on the pH, the humic acid, and temperature, while it was less affected by the nitrate. Phosphate adsorption by the CSB and Fe-CSB could be well described by the pseudo n-th order and Langmuir-Freundlich models. The fitting of the experimental data with the intra-particle diffusion model revealed that surface adsorption and inner-sphere diffusion were involved in the phosphate adsorption process, and that the latter was the rate-controlling step. Batch adsorption experiments and post-adsorption characterization results revealed that the phosphate adsorption by Fe-CSB was primarily governed by four mechanisms: ligand exchange, electrostatic attraction, chemical precipitation, and inner-sphere complexation. This work demonstrated that the modified Fe-CSB is an environmentally friendly and cost-effective bioretention medium and could open up new pathways for the removal of phosphorus from stormwater, as well as solve the problem of waste biomass pollution.

Superhydrophilic Anti-Icing Coatings Based on Polyzwitterion Brushes.

In this work, an anti-icing coating based on superhydrophilic polyzwitterion brushes is reported. Poly(sulfobetaine methacrylate) (PSBMA) brushes were synthesized by surface-initiated atom-transferred radical polymerization to create superhydrophilic films on silicon wafers. The thickness of the PSBMA brushes film increased linearly with the polymerization time, and the film remained superhydrophilic in a nonassociated state when the thickness was less than ∼100 nm. DSC and FTIR analyses revealed that PSBMA contains more nonfreezable bound water than typical polyelectrolytes such as poly(sodium styrenesulfonate) and poly(2-(methacryloyloxy)ethyltrimethylammonium chloride), leading to lower ice adhesion strength than the latter two. At -20 °C, the PSBMA brushes coating demonstrated low ice adhesion strength of 60 kPa, showing a significant reduction in ice adhesion by up to ∼75% compared to uncoated silicon wafer. The optimum PSBMA layer thickness for low ice adhesion was ∼100 nm. These findings suggest that polyzwitterions are excellent candidates for anti-icing coating application.