Combined toxicity of polyethylene microplastics and nickel oxide nanoparticle on earthworm (Eisenia andrei): oxidative stress responses, bioavailability and joint effect.

The co-occurrence of heavy metals and microplastics (MPs) is an emerging issue that has attracted considerable attention. However, the interaction of nickel oxide nanoparticle (nano-NiO) combined with MPs in soil was poorly researched. Here, experiments were conducted to study the influence of nano-NiO (200 mg/kg) and polyethylene (PE) MPs with different concentrations (0.1, 1, and 10%) and sizes (13, 50, and 500 µm) on earthworms for 28 days. Compared to control, the damage was induced by PE and nano-NiO, which was evaluated by biomarker Integrated Biomarker Response index: version 2 (IBRv2) based on six biomarkers including SOD, POD, CAT, MDA, AChE, Na+/K+-ATPase and cellulase. The majority of the chosen biomarkers showed significant but complicated responses with increasing contaminant concentrations after 28 days of exposure. Moreover, the joint effect was assessed as antagonism by the effect addition index (EAI). Overall, this work expands our understanding of the combined toxicity of PE and nano-NiO in soil ecosystems.

Liquiritigenin Induces Cell Cycle Arrest and Apoptosis in Lung Squamous Cell Carcinoma.

Liquiritigenin (LQ), as a dihydroflavone monomer compound extracted from Glycyrrhiza uralensis Fisch, has been demonstrated to show anti-tumor effects in multiple human cancers, including lung adenocarcinoma. Our study aimed to explore its role in lung squamous cell carcinoma (LSCC) development and the related mechanism. The effects of LQ on SK-MES-1 and NCI-H520 cell proliferation, cell cycle, and apoptosis were investigated. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and colony formation assays revealed that LQ inhibited LSCC cell viability and proliferation in a dose- and time-dependent manner. Flow cytometry analysis demonstrated that LQ promoted G2/M cell cycle arrest, cell apoptosis, and loss of mitochondrial membrane potential. In vivo assays showed that LQ administration suppressed tumor growth in nude mice. Additionally, LQ treatment reduced the levels of phosphorylated PI3K, AKT, and mTOR levels in LSCC cells. Pretreatment with the PI3K inhibitor LY294002 antagonized the LQ-mediated effects on cell proliferation, cell cycle arrest, and apoptosis in LSCC cells. Collectively, LQ induces cell cycle arrest and apoptosis in LSCC by inactivating the PI3K/AKT/mTOR pathway.

N6-methyladenosine demethyltransferase FTO mediated m6A modification of estrogen receptor alpha in non-small cell lung cancer tumorigenesis.

Fat mass and obesity-associated protein (FTO), which is closely linked with obesity and dietary intake, plays an important role in diet-related metabolic diseases. However, the underlying mechanism of the N6-methyladenosine (m6A) demethyltransferase FTO in tumor development and progression remains largely unexplored. Here, we demonstrated that FTO expression was largely lower in non-small cell lung cancer (NSCLC) samples than in adjacent healthy tissues, and its expression negatively correlated with poor prognosis. Gain- and loss-of-function assays revealed that FTO inhibited NSCLC tumor cell growth and metastasis in vitro and in vivo. Mechanistically, estrogen receptor alpha (ESR1) is a target of FTO, and increased FTO expression significantly impaired the m6A levels of ESR1 mRNA. There were two clear m6A modification sites (5247A and 5409A) in the 3' untranslated region (3'UTR) of ESR1, and FTO could decrease their methylation. Moreover, the m6A readers YTHDF1 and IGF2BP3 recognized and bound the m6A sites in ESR1 mRNA, thereby enhancing its stability and facilitating tumor growth. We also showed that ESR1 has good diagnostic value for NSCLC. In conclusion, we uncovered an important mechanism of epitranscriptomic regulation by the FTO-YTHDF1-IGF2BP3-ESR1 axis and identified the potential of m6A-dependent therapeutic strategies for NSCLC.

Therapeutic potential of HUC-MSC-exos primed with IFN-γ against LPS-induced acute lung injury.

Objectives: Human umbilical cord mesenchymal stem cells (HUC-MSCs) are pluripotent stem cells with anti-inflammatory and immunomodulatory properties used in the treatment of acute lung injury (ALI). However, the treatment of ALI using exosomes derived from HUC-MSCs (HUC-MSC-exos) primed with interferon-gamma (IFN-γ-exos) has not been described. This study investigated the effects of IFN-γ-exos on ALI. Materials and Methods: IFN-γ primed and unprimed HUC-MSC-exos (IFN-γ-exos and CON-exos, respectively) were extracted, identified, and traced. A549 cells and mice subjected to lipopolysaccharide (LPS)-induced inflammation were treated with IFN-γ-exos or CON-exos. Viability; interleukin (IL)-1ß, IL-6, tumor necrosis factor (TNF)-α, and reactive oxygen species (ROS) levels; NF-κB p65, and NLRP3 expression and histology and lung injury scores were measured in cell, supernatant or lung tissue. Results: Indoleamine 2,3-dioxygenase (IDO) mRNA expression was elevated in HUC-MSCs primed with 5 ng/mL IFN-γ (P<0.001), and IFN-γ-exos and CON-exos were successfully extracted. LPS-induced inflammation resulted in decreased cell viability in A549 cells, and increased IL-1ß, IL-6, TNF-α and ROS levels and NF-κB p65 and NLRP3 expression in A549 cells and mice(P<0.05 to P<0.001). Treatment with IFN-γ-exos and CON-exos increased cell viability and decreased the concentrations of IL-1ß, and ROS, expression of NF-κB p65 and NLRP3, and the lung injury score, and these effects were more obvious for IFN-γ-exos(P<0.05 to P<0.001). Conclusion: IFN-γ-exos reduced oxidative stress and inflammatory responses in LPS-induced A549 cells and mice. The result demonstrated the therapeutic potential of IFN-γ-exos in LPS-induced ALI.

Meta-analysis of the impact of laser interstitial hyperthermia on wound healing complications in brain tumors.

High-grade gliomas (HGGs) may be amenable to the neurosurgical technique known as laser interstitial thermal therapy (LITT), which delivers thermal energy to interstitial brain injuries and wounds with pinpoint accuracy. The purpose of this extensive meta-analysis was to evaluate the effects of LITT on wound complications among patients who have brain tumours. Diverse conclusions emerge from a systematic review of pertinent studies, necessitating a comprehensive examination. The meta-analysis, performed utilizing the meta library provided by the R package meta, reveals an initial significant overall effect (RR: -2.1262, 95% CI [-2.7466, -1.5059], p < 0.0001) accompanied by considerable heterogeneity among studies (I2 = 61.13%). Following analyses that specifically examined the incidence of wounds, a complex correlation was found (RR: 0.0471, 95% CI [0.0264, 0.0842], p < 0.0001), indicating that LITT has a discernible but insignificant effect on the occurrence of wounds. Although the meta-analysis emphasizes a notable decrease in wound complications subsequent to LITT treatment, additional research is warranted due to constraints in standardized reporting, data accessibility, and small sample sizes. The results of this study underscore the need for exhaustive protocols to analyse wound complications in patients with brain tumours undergoing LITT.

Staphylococcus aureus lysate induces an IgE response via memory B cells in nasal polyps.

BACKGROUND: Locally increased IgE levels plays a pathologic role in chronic rhinosinusitis with nasal polyps (CRSwNP). OBJECTIVE: This study aimed to investigate whether Staphylococcus aureus could induce aberrant IgE synthesis in CRSwNP and the potential mechanisms involved. METHODS: Total IgE, IL-4, IL-5, and IL-13 concentrations in the supernatants of the cultures stimulated with S aureus lysate were assessed by ELISA. S aureus-induced cellular responses were investigated by single-cell RNA sequencing. Flow cytometry and quantitative reverse transcription PCR were used to analyze B-cell subsets and stimulated cell ε-germline transcript expression, respectively. IgE-positive B-cell and germinal center localization were assessed by immunohistochemistry and immunofluorescence. RESULTS: S aureus lysate induced IgE production in the supernatants of nasal polyp (NP) tissues but not in those of healthy nasal mucosa. Moreover, IgE levels increased from days 2 to 4 after stimulation, paralleling the enhanced ε-germline transcript, IL-5, and IL-13 expression. Single-cell RNA sequencing revealed that there were increased IL-5 and IL-13 in group 2 innate lymphoid cells and identified a clonal overlap between unstimulated memory B cells and S aureus-stimulated plasma cells. The enriched IgE within NPs was mainly produced by IgE-negative memory B cells. Cellular evidence indicated that the IgE memory response to S aureus might also exist in the peripheral blood of CRSwNP patients. The S aureus-induced IgE memory response was associated with elevated IgE levels in NPs, asthma, and postoperative CRSwNP recurrence. CONCLUSIONS: S aureus induced an IgE response via IgE-negative memory B cells in CRSwNP patients, possibly contributing to CRSwNP development.

Synergistic toxicity of some food additives used in non-alcoholic beverages on renal tubular epithelial cells.

Evidences supported many food additives (FAs) possess toxicity to human health due to chronic excessive exposure. Global hygienic standards strictly limit the dosage of each FA and mixture of the same functional FAs. However, the synergetic effects caused by the combination of FAs with different functions require careful evaluation. In the present study, the content of each FA in beverages was determined by HPLC-UV-Vis detection. The cytotoxic effects of selected typical FAs alone or their combination were evaluated in human renal tubular epithelial cells. Mathematical Modeling and bioinformatics methods were employed to evaluate the toxicity of FAs and to predict the key target proteins of FAs on renal tubular cell toxicity, which were verified by western blot. The results indicated above 5 FAs were used in each surveyed beverage. The content of each FA and the respective ratios of the same functional FAs in each beverage did not exceed the maximum permitted level. But it was intensively shown that the significant synergistic cytotoxicity for the combination of FAs with lower concentration. The intercellular signaling transduction pathways including JNK/STAT, PI3P/AKT, and MAPK pathways, which could also be activated by PDGF signaling, were predicted to be involved in Fas-induced cytotoxicity. The increased expression of p-STAT3, p-JNK and p-AKT was associated with renal tubular injury. The current study implied the synergistic cytotoxic effect caused by multiple FAs at no toxic dosages via activated cellular transduction pathways regulating cell survival and apoptosis function, which warning of the synergistic toxic effects of different types of FAs.

Mechanism of aconitine mediated neuronal apoptosis induced by mitochondrial calcium overload caused by MCU.

Aconitine is a crucial toxic component in Chinese herbal medicines such as Aconitum, Aconitum coreanum, and Aconitum soongaricum. The poisoning symptoms of the central nervous system and cardiovascular system caused by it are relatively common in China, and there are many studies on cardiovascular system diseases caused by aconitine. However, the specific mechanism of neurotoxicity induced by aconitine is still unclear. This study explored the effect and mechanism of mitochondrial calcium uniporter on mitochondrial energy metabolism disorder in aconitine poisoning hippocampal neurons. The results showed that after treatment with 400µmol/L aconitine, mitochondrial energy metabolism was abnormal in rat hippocampal neuron cells, the expression of MCU in mitochondria was up-regulated, calcium overload in mitochondria, ATP production decreased, and mitochondrial membrane potential Changes, increased expression of the apoptosis gene Cleaved-Caspase-3. After treatment with the MCU agonist spermine, mitochondrial energy metabolism was significantly abnormal, and cell apoptosis was increased considerably. However, pretreatment with calcium ion channel inhibitor Ruthenium Red (RR) effectively promoted the generation of ATP, thereby improving mitochondrial energy metabolism disorders and reducing cell apoptosis. These results suggest that aconitine induces mitochondrial energy metabolism dysfunction in hippocampal neurons, which may be related to the increased expression of MCU.

A Spectrophotometric Turbidity Assay to Study Liquid-Liquid Phase Separation of UBQLN2 In Vitro.

Liquid-liquid phase separation (LLPS) is hypothesized to be the underlying mechanism for how membraneless organelles or biomolecular condensates form inside both prokaryotic and eukaryotic cells. Protein LLPS is a biophysical process during which proteins demix from homogeneous solution to form protein-dense droplets with liquid-like properties. Disruptions to LLPS, such as changes to material properties of condensates or physicochemical parameters for LLPS onset, are implicated in neurodegenerative diseases and cancer. Therefore, it is essential to determine the physicochemical parameters that promote protein LLPS. Here, we present our UV-Vis spectrophotometric turbidity assay to characterize the temperature and concentration dependence of LLPS for UBQLN2, a protein that undergoes LLPS via homotypic interactions in vitro and forms stress-induced condensates in cells. Mutations in UBQLN2 cause amyotrophic lateral sclerosis (ALS) and disrupt UBQLN2 LLPS. We present a detailed expression and purification protocol for a C-terminal construct of UBQLN2 and how we use microscopy to image UBQLN2 LLPS. We use our UV-Vis assay to construct temperature-concentration phase diagrams for wild-type and mutant UBQLN2 constructs to determine the effects of domain deletions and/or mutations on UBQLN2 phase separation.

Cloning and functional analysis of a pacifastin-like protein from the sea cucumber, Apostichopus japonicus.

Pacifastin family proteins play a crucial role in regulating innate immune responses such as phagocytosis in invertebrates. However, the function of the Ajpacifastin-like counterpart in the sea cucumber Apostichopus japonicus remains elusive. In this study, the pacifastin gene of A. japonicus was cloned, characterized and named Ajpacifastin-like. The open reading frame of Ajpacifastin-like is 1497 bp in length and encodes a polypeptide containing 498 amino acid residues. Structural analysis revealed that the protein encoded by Ajpacifastin-like contains two pacifastin light chain domains (amino acids 287-322 and amino acids 376-407). Real-time reverse transcriptase PCR showed that Ajpacifastin-like mRNA is ubiquitously expressed in all tissues examined, with the highest expression in muscle. Ajpacifastin-like mRNA expression was significantly upregulated to 3.27-fold after challenge with Vibrio splendidus for 24 h. To explore the function of the Ajpacifastin-like protein in the immune response of A. japonicus, dsRNA interference with Ajpacifastin-like expression and with the expression of its postulated target gene was performed. Flow cytometry analysis showed that the rate of phagocytosis by coelomocytes increased to 1.21-fold in individuals treated with specific Ajpacifastin-like siRNA. However, rate of phagocytosis by coelomocytes decreased to 86% in individuals treated with Ajphenoloxidase siRNA. These results show that the Ajpacifastin-like gene is ubiquitously expressed in almost all tissues and that Ajpacifastin-like protein acts as an immunomodulatory factor via phenoloxidase to mediate phagocytosis by coelomocytes in pathogen-challenged A. japonicus.

Mechanistic insights into enhancement or inhibition of phase separation by different polyubiquitin chains.

Ubiquitin-binding shuttle UBQLN2 mediates crosstalk between proteasomal degradation and autophagy, likely via interactions with K48- and K63-linked polyubiquitin chains, respectively. UBQLN2 comprises self-associating regions that drive its homotypic liquid-liquid phase separation (LLPS). Specific interactions between one of these regions and ubiquitin inhibit UBQLN2 LLPS. Here, we show that, unlike ubiquitin, the effects of multivalent polyubiquitin chains on UBQLN2 LLPS are highly dependent on chain types. Specifically, K11-Ub4 and K48-Ub4 chains generally inhibit UBQLN2 LLPS, whereas K63-Ub4, M1-Ub4 chains, and a designed tetrameric ubiquitin construct significantly enhance LLPS. We demonstrate that these opposing effects stem from differences in chain conformations but not in affinities between chains and UBQLN2. Chains with extended conformations and increased accessibility to the ubiquitin-binding surface promote UBQLN2 LLPS by enabling a switch between homotypic to partially heterotypic LLPS that is driven by both UBQLN2 self-interactions and interactions between multiple UBQLN2 units with each polyubiquitin chain. Our study provides mechanistic insights into how the structural and conformational properties of polyubiquitin chains contribute to heterotypic LLPS with ubiquitin-binding shuttles and adaptors.

Applicability of Anticancer Drugs for the Triple-Negative Breast Cancer Based on Homologous Recombination Repair Deficiency.

Triple-negative breast cancer (TNBC) is a highly aggressive disease with historically poor outcomes, primarily due to the lack of effective targeted therapies. Here, we established a drug sensitivity prediction model based on the homologous recombination deficiency (HRD) using 83 TNBC patients from TCGA. Through analyzing the effect of HRD status on response efficacy of anticancer drugs and elucidating its related mechanisms of action, we found rucaparib (PARP inhibitor) and doxorubicin (anthracycline) sensitive in HR-deficient patients, while paclitaxel sensitive in the HR-proficient. Further, we identified a HRD signature based on gene expression data and constructed a transcriptomic HRD score, for analyzing the functional association between anticancer drug perturbation and HRD. The results revealed that CHIR99021 (GSK3 inhibitor) and doxorubicin have similar expression perturbation patterns with HRD, and talazoparib (PARP inhibitor) could kill tumor cells by reversing the HRD activity. Genomic characteristics indicated that doxorubicin inhibited tumor cells growth by hindering the process of DNA damage repair, while the resistance of cisplatin was related to the activation of angiogenesis and epithelial-mesenchymal transition. The negative correlation of HRD signature score could interpret the association of doxorubicin pIC50 with worse chemotherapy response and shorter survival of TNBC patients. In summary, these findings explain the applicability of anticancer drugs in TNBC and underscore the importance of HRD in promoting personalized treatment development.

Analysis of Mutations and Dysregulated Pathways Unravels Carcinogenic Effect and Clinical Actionability of Mutational Processes.

Somatic mutations accumulate over time in cancer cells as a consequence of mutational processes. However, the role of mutational processes in carcinogenesis remains poorly understood. Here, we infer the causal relationship between mutational processes and somatic mutations in 5,828 samples spanning 34 cancer subtypes. We found most mutational processes cause abundant recurrent mutations in cancer genes, while exceptionally ultraviolet exposure and altered activity of the error-prone polymerase bring a large number of recurrent non-driver mutations. Furthermore, some mutations are specifically induced by a certain mutational process, such as IDH1 p.R132H which is mainly caused by spontaneous deamination of 5-methylcytosine. At the pathway level, clock-like mutational processes extensively trigger mutations to dysregulate cancer signal transduction pathways. In addition, APOBEC mutational process destroys DNA double-strand break repair pathway, and bladder cancer patients with high APOBEC activity, though with homologous recombination proficient, show a significantly longer overall survival with platinum regimens. These findings help to understand how mutational processes act on the genome to promote carcinogenesis, and further, presents novel insights for cancer prevention and treatment, as our results showing, APOBEC mutagenesis and HRD synergistically contributed to the clinical benefits of platinum-based treatment.

Combined homologous recombination repair deficiency and immune activation analysis for predicting intensified responses of anthracycline, cyclophosphamide and taxane chemotherapy in triple-negative breast cancer.

BACKGROUND: Triple-negative breast cancer (TNBC) is a clinically aggressive disease with abundant variants that cause homologous recombination repair deficiency (HRD). Whether TNBC patients with HRD are sensitive to anthracycline, cyclophosphamide and taxane (ACT), and whether the combination of HRD and tumour immunity can improve the recognition of ACT responders are still unknown. METHODS: Data from 83 TNBC patients in The Cancer Genome Atlas (TCGA) was used as a discovery cohort to analyse the association between HRD and ACT chemotherapy benefits. The combined effects of HRD and immune activation on ACT chemotherapy were explored at both the genome and the transcriptome levels. Independent cohorts from the Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) and Gene Expression Omnibus (GEO) were adopted to validate our findings. RESULTS: HRD was associated with a longer ACT chemotherapy failure-free interval (FFI) with a hazard ratio of 0.16 (P = 0.004) and improved patient prognosis (P = 0.0063). By analysing both HRD status and ACT response, we identified patients with a distinct TNBC subtype (ACT-S&HR-P) that showed higher tumour lymphocyte infiltration, IFN-γ activity and NK cell levels. Patients with ACT-S&HR-P had significantly elevated immune inhibitor levels and presented immune activation associated with the increased activities of both innate immune cells and adaptive immune cells, which suggested treatment with immune checkpoint blockade as an option for this subtype. Our analysis revealed that the combination of HRD and immune activation enhanced the efficiency of identifying responders to ACT chemotherapy (AUC = 0.91, P = 1.06e-04) and synergistically contributed to the clinical benefits of TNBC patients. A transcriptional HRD signature of ACT response-related prognostic factors was identified and independently validated to be significantly associated with improved survival in the GEO cohort (P = 0.0038) and the METABRIC dataset (P < 0.0001). CONCLUSIONS: These findings highlight that HR deficiency prolongs FFI and predicts intensified responses in TNBC patients by combining HRD and immune activation, which provides a molecular basis for identifying ACT responders.

Structure, dynamics and functions of UBQLNs: at the crossroads of protein quality control machinery.

Cells rely on protein homeostasis to maintain proper biological functions. Dysregulation of protein homeostasis contributes to the pathogenesis of many neurodegenerative diseases and cancers. Ubiquilins (UBQLNs) are versatile proteins that engage with many components of protein quality control (PQC) machinery in cells. Disease-linked mutations of UBQLNs are most commonly associated with amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), and other neurodegenerative disorders. UBQLNs play well-established roles in PQC processes, including facilitating degradation of substrates through the ubiquitin-proteasome system (UPS), autophagy, and endoplasmic-reticulum-associated protein degradation (ERAD) pathways. In addition, UBQLNs engage with chaperones to sequester, degrade, or assist repair of misfolded client proteins. Furthermore, UBQLNs regulate DNA damage repair mechanisms, interact with RNA-binding proteins (RBPs), and engage with cytoskeletal elements to regulate cell differentiation and development. Important to the myriad functions of UBQLNs are its multidomain architecture and ability to self-associate. UBQLNs are linked to numerous types of cellular puncta, including stress-induced biomolecular condensates, autophagosomes, aggresomes, and aggregates. In this review, we focus on deciphering how UBQLNs function on a molecular level. We examine the properties of oligomerization-driven interactions among the structured and intrinsically disordered segments of UBQLNs. These interactions, together with the knowledge from studies of disease-linked mutations, provide significant insights to UBQLN structure, dynamics and function.

Clonal tumor mutations in homologous recombination genes predict favorable clinical outcome in ovarian cancer treated with platinum-based chemotherapy.

OBJECTIVE: Platinum-based chemotherapy remains the first-line treatment for ovarian carcinoma by inducing DNA damage. The therapeutic impact of clonal and subclonal somatic mutations in DNA damage repair (DDR) pathways remains unexplored. METHODS: We performed an integrated analysis to infer the clonality of somatic deleterious mutations in 385 ovarian carcinomas treated with platinum-based chemotherapy. The Kaplan-Meier method was performed for visualization and the differences between survival curves were calculated by log-rank test. Proportional hazards models were used to estimate relative hazards for platinum-free interval (PFI), progression-free survival (PFS) and overall survival (OS). RESULTS: We found that somatic deleterious mutations in DDR pathways exhibited widespread clonal heterogeneity, and that patients with DDR clonal mutations exhibited a "hypermutator phenotype". Clonal somatic mutations in homologous recombination repair (HRR) pathway were significantly associated with better OS (HR = 0.19 (95% CI, 0.06-0.59), P = 0.0044) and PFS (HR = 0.20 (95% CI, 0.08-0.49), P = 0.0005) than HRR wild-type, while HRR subclonal mutations were not associated with prognosis. Moreover, HRR clonal mutations were associated with significantly higher chemotherapy sensitive rate (P = 0.0027) and longer PFI (HR = 0.20 (95% CI, 0.08-0.49), P = 0.0005) than HRR wild-type, while HRR subclonal mutations were not. We validated our findings using an independent cohort of 93 ovarian cancer patients that received platinum-based chemotherapy. CONCLUSIONS: HRR clonal mutations, but not subclonal mutations, were associated with improved survival, chemotherapy response, and genome instability compared with HRR wild-type.

High-fat diet induces dry eye-like ocular surface damages in murine.

PURPOSE: A high-fat diet leads to dysfunction in multiple systems of the body. Herein we investigate the effects of a high-fat diet on the ocular surface using a murine model. METHODS: Four-week-old male C57BL/6 mice were fed with a standard-fat diet (10 kcal% fat, SFD) or a high-fat diet (60 kcal% fat, HFD) for 1 or 3 months. Phenol red thread test was used to detect tear production, oregon green dextran (OGD) staining was performed to assess corneal epithelial permeability, and PAS staining was conducted to ascertain the presence of conjunctival goblet cells. Squamous metaplasia in the ocular surface and corneal epithelial barrier function were detected by immunofluorescent staining, zymography and Western blot analysis. Oxidative stress related protein expression was evaluated by immunostaining and Western blot analysis. Corneal and conjunctival cell apoptosis was determined by TUNEL assay and caspase-3 expression. RESULTS: A HFD induced obvious ocular surface damages, including decreased tear production, notable OGD staining and distinct goblet cell loss. It also resulted in corneal epithelial barrier dysfunction and significant squamous metaplasia of the corneal and conjunctival epithelia. The HFD also upregulated key factors that regulate oxidative stress in the ocular surface, and upregulated cell apoptosis in ocular surface epithelial cells. CONCLUSIONS: A HFD induces dry eye-like ocular surface damages in mice via the activation of oxidative stress and an induction of apoptosis in the cells of the ocular surface.

Identification of Dysregulated Competitive Endogenous RNA Networks Driven by Copy Number Variations in Malignant Gliomas.

Gliomas represent 80% of malignant brain tumors. Because of the high heterogeneity, the oncogenic mechanisms in gliomas are still unclear. In this study, we developed a new approach to identify dysregulated competitive endogenous RNA (ceRNA) interactions driven by copy number variation (CNV) in both lower-grade glioma (LGG) and glioblastoma multiforme (GBM). By analyzing genome and transcriptome data from The Cancer Genome Atlas (TCGA), we first found out the protein coding genes and long non-coding RNAs (lncRNAs) significantly affected by CNVs and further determined CNV-driven dysregulated ceRNA interactions by a customized pipeline. We obtained 13,776 CNV-driven dysregulated ceRNA pairs (including 3,954 mRNAs and 306 lncRNAs) in LGG and 262 pairs (including 221 mRNAs and 11 lncRNAs) in GBM, respectively. Our results showed that most of the ceRNA interactions were weakened by CNVs in both LGG and GBM, and many CNV-driven genes shared the same ceRNAs in the dysregulated ceRNA networks. Functional analysis indicated that the CNV-driven ceRNA network involved in some important mechanisms of tumorigenesis, such as cell cycle, p53 signaling pathway and TGF-beta signaling pathway. Further investigation of the ceRNA pairs in the communities from the dysregulated ceRNA network revealed more detailed biological functions related to the oncogenesis of malignant gliomas. Moreover, by exploring the association of CNV-driven ceRNAs with prognosis and histological subtype, we found that the copy number status of MTAP, KLHL9, and ELAVL2 related to the overall survival in LGG and showed high correlation with histological subtype. In conclusion, this study provided new insight into the molecular mechanisms and clinical biomarkers in gliomas.

Ubiquitylomes Analysis of the Whole blood in Postmenopausal Osteoporosis Patients and healthy Postmenopausal Women.

OBJECTIVES: To determine the mechanisms of ubiquitination in postmenopausal osteoporosis and investigate the ubiquitinated spectrum of novel targets between healthy postmenopausal women and postmenopausal osteoporosis patients, we performed ubiquitylome analysis of the whole blood of postmenopausal women and postmenopausal osteoporosis patients. METHODS: To obtain a more comprehensive understanding of the postmenopausal osteoporosis mechanism, we performed a quantitative assessment of the ubiquitylome in whole blood from seven healthy postmenopausal women and seven postmenopausal osteoporosis patients using high-performance liquid chromatography fractionation, affinity enrichment, and liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). To examine the ubiquitylome data, we performed enrichment analysis using an ubiquitylated amino acid motif, Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway. RESULTS: Altogether, 133 ubiquitinated sites and 102 proteins were quantified. A difference of more than 1.2 times is considered significant upregulation and less than 0.83 significant downregulation; 32 ubiquitinated sites on 25 proteins were upregulated and 101 ubiquitinated sites on 77 proteins were downregulated. These quantified proteins, both with differently ubiquitinated sites, participated in various cellular processes, such as cellular processes, biological regulation processes, response to stimulus processes, single-organism and metabolic processes. Ubiquitin conjugating enzyme activity and ubiquitin-like protein conjugating enzyme activity were the most highly enriched in molecular function of upregulated sites with corresponding proteins, but they were not enriched in downregulated in sites with corresponding proteins. The KEGG pathways analysis of quantified proteins with differentiated ubiquitinated sites found 13 kinds of molecular interactions and functional pathways, such as glyoxylate and decarboxylate metabolism, dopaminergic synapse, ubiquitin-mediated proteolysis, salivary secretion, coagulation and complement cascades, Parkinson's disease, and hippo signaling pathway. In addition, hsa04120 ubiquitin-mediated proteolysis was the most highly enriched in proteins with upregulated sites, hsa04610 complement and coagulation cascades was the most highly enriched in proteins with downregulated ubiquitinated sites, and hsa04114 Oocyte meiosis was the most highly enriched among all differential proteins. CONCLUSION: Our study expands the understanding of the spectrum of novel targets that are differentially ubiquitinated in whole blood from healthy postmenopausal women and postmenopausal osteoporosis patients. The findings will contribute toward our understanding of the underlying proteostasis pathways in postmenopausal osteoporosis and the potential identification of diagnostic biomarkers in whole blood.

Discovery and Identification of Serum Succinyl-Proteome for Postmenopausal Women with Osteoporosis and Osteopenia.

OBJECTIVE: For the purpose of providing evidence for the treatment of osteoporosis and osteopenia, this study retrospectively identified succinylation-modified sites and proteins in postmenopausal women, and bioinformatics analysis were performed. METHODS: From January 2016 to June 2018, a total of 30 postmenopausal women aged from 55 to 70 years old were assigned to three groups: 10 cases with osteoporosis; 10 cases with osteopenia; and 10 cases with normal bone mass. Subsequently, the serum samples were collected from all cases for succinyl-proteome. Measures comprised label-free quantitative analysis, succinylation enrichment techniques, the liquid chromatograph-mass spectrometer/mass spectrometer (LC-MS/MS) methods, and bioinformatics. RESULTS: A total of 113 succinylation sites on 35 proteins were identified based on quantitative information. The variation of the different multiple folds were more than 1.2 times as a significant increase for up-regulated and less than 1/1.2 times as a significant decrease for down-regulated. Among the quantified succinylation sites, 66 were up-regulated and 11 down-regulated in the Osteopenia/Normal comparison group, 24 were up-regulated and 44 down-regulated in the Osteoporosis/Osteopenia comparison group, 45 were up-regulated and 32 down-regulated in the Osteoporosis/Normal comparison group. Among the quantified succinylation proteins, 24 were up-regulated and 7 down-regulated in the Osteopenia/Normal comparison group, 15 were up-regulated and 20 down-regulated in the Osteoporosis/Osteopenia comparison group, 20 were up-regulated and 17 down-regulated in the Osteoporosis/Normal comparison group. The percentage of proteins differed in immune response, signaling pathway, proteolysis, lymphocyte, leukocyte, and cell activation. Four differentially expressed proteins (apolipoprotein A-I, apolipoprotein A-II, hemoglobin subunit alpha, and haptoglobin) contained quantitative information; they were mediated with receptors, factors, mechanisms, that related to bone metabolism. Hemoglobin subunit alpha was screened for diagnosis of osteopenia. CONCLUSIONS: The succinyl-proteome experimental data indicated that apolipoprotein A-I, apolipoprotein A-II, hemoglobin subunit alpha, and haptoglobin were valuable for diagnosis and treatment in postmenopausal women with osteoporosis and osteopenia.