Mitochondria-mediated self-cycling nanoreactor enabling uninterrupted oxidative damage for enhanced chemodynamic therapy.

Chemodynamic therapy (CDT), which employs intracellular H2O2 to produce toxic hydroxyl radicals to kill cancer cells, has received great attention due to its specificity to tumors. However, the relatively insufficient endogenous H2O2 and the short-lifetime and limited diffusion distance of •OH compromise the therapeutic efficacy of CDT. Mitochondria, which play crucial roles in oncogenesis, are highly vulnerable to elevated oxidative stress. Herein, we constructed a mitochondria-mediated self-cycling system to achieve high dose of •OH production through continuous H2O2 supply. Cinnamaldehyde (CA), which can elevate H2O2 level in the mitochondria, was loaded in Cu(II)-containing metal organic framework (MOF), termed as HKUST-1. After actively targeting mitochondria, the intrinsic H2O2 in mitochondria of cancer cells could induce degradation of MOF, releasing the initial free CA. The released CA further triggered the upregulation of endogenous H2O2, resulting in the subsequent adequate release of CA and the final burst growth of H2O2. The cycle process greatly promoted the Fenton-like reaction between Cu2+ and H2O2 and induced long-term high oxidative stress, achieving enhanced chemodynamic therapy. In a word, we put forward an efficient strategy for enhanced chemodynamic therapy.

Comprehensive analysis of the miRNA-mRNA regulatory network involved in spontaneous recovery of an H2O2-induced zebrafish cataract model.

OBJECTIVE: To identify the hub miRNAs and mRNAs contributing to the spontaneous recovery of an H2O2-induced zebrafish cataract model. METHODS: Zebrafishes were divided into three groups, i.e., Group A, which included normal control fish (day 0), and Groups B and C, where fish were injected with 2.5% hydrogen peroxide into the anterior chamber and reared for 14 and 30 days, respectively. Fish eyes were examined by stereomicroscope photography and optical coherence tomography (OCT). RNA profiles of fish lenses were detected by RNA sequencing. Differentially expressed genes (DEGs) and differentially expressed miRNAs (DEmiRs) were identified among three groups. The DEGs and DEmiRs, which changed in opposite positions between "B vs. A" and "C vs. B" were defined as ODGs (opposite positions changed DEGs) and ODmiRs (opposite positions changed DEmiRs). Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes pathway (KEGG) analysis were carried out by R language. The protein-protein interaction network (PPI) was constructed using STRING. Potential targets of miRNAs were obtained using miRanda. miRNA-mRNA networks were constructed by Cytoscape. RESULTS: The fish lens opacity formed on day 14 and recovered to transparent on day 30 after injection. Compared to group B, 1366 DEGs and 54 DEmiRs were identified in group C. "C vs. B" DEGs were enriched in gene clusters related to development and oxidative phosphorylation. Target genes of DEmiRs were enriched in clusters such as development and cysteine metabolism. Among three groups, 786 ODGs and 27 ODmiRs were identified, and 480 ODGs were predicted as targets of ODmiRs. Target ODGs were enriched in pathways related to methionine metabolism, ubiquitin, sensory system development, and structural constituents of the eye lens. In addition, we established an ODmiRs-ODGs regulation network. CONCLUSION: We identified several hub mRNAs and altered miRNAs in the formation and reversal of zebrafish cataracts. These hub miRNAs/mRNAs could be potential targets for the non-surgical treatment of ARC.

Actomyosin-driven motility and coalescence of phase-separated viral inclusion bodies are required for efficient replication of a plant rhabdovirus.

Phase separation has emerged as a fundamental principle for organizing viral and cellular membraneless organelles. Although these subcellular compartments have been recognized for decades, their biogenesis and mechanisms of regulation are poorly understood. Here, we investigate the formation of membraneless inclusion bodies (IBs) induced during the infection of a plant rhabdovirus, tomato yellow mottle-associated virus (TYMaV). We generated recombinant TYMaV encoding a fluorescently labeled IB constituent protein and employed live-cell imaging to characterize the intracellular dynamics and maturation of viral IBs in infected Nicotiana benthamiana cells. We show that TYMaV IBs are phase-separated biomolecular condensates and that viral nucleoprotein and phosphoprotein are minimally required for IB formation in vivo and in vitro. TYMaV IBs move along the microfilaments, likely through the anchoring of viral phosphoprotein to myosin XIs. Furthermore, pharmacological disruption of microfilaments or inhibition of myosin XI functions suppresses IB motility, resulting in arrested IB growth and inefficient virus replication. Our study establishes phase separation as a process driving the formation of liquid viral factories and emphasizes the role of the cytoskeletal system in regulating the dynamics of condensate maturation.

Association of Neutrophil-to-Lymphocyte Ratio with Nutrition in Patients with Various Types of Malignant Tumors: A Multicenter Cross-Sectional Study.

Aim: Neutrophil-to-lymphocyte ratio (NLR) is an index of systemic inflammation. This study is to clarify the role of NLR in body functional status, nutritional risk and nutritional status in the course of tumor. Methods: A multi-center cross-sectional study of patients with various types of malignant tumors was accrued from the whole country. There were 21,457 patients with completed clinical data, biochemical indicators, physical examination, the Patient-Generated Subjective Global Assessment (PG-SGA) and Nutrition Risk Screening 2002 (NRS2002) survey. Logistic regression analysis was used to figure out the influencing factors of NLR, and four models were established to evaluate the influence of NLR on body functions, nutritional risks and nutritional status. Results: Male patients, TNM stage IV, total bilirubin, hypertension and coronary atherosclerotic heart disease (CAHD) were independent predictors of NLR >2.5. BMI, digestive systemic tumors and triglyceride negatively affect NLR in multivariable logistic regression. NLR was an independent predictor of Karnofsky Performance Scale (KPS), fat store deficit in all degrees, moderate and severe muscle deficit, mild fluid retention and PG-SGA grade. Conclusion: Male patients and those with hypertension and CAHD are prone to systemic inflammation. Systemic inflammation significantly degrades body function status and nutritional status, increases nutritional risk and influences fat and muscle metabolism in patients with malignant tumor. Improving the intervenable indicators such as elevating albumin and pre-albumin, decreasing total bilirubin and enhancing nutrition support are imperative. Obesity and triglyceride behave like anti-systemic inflammation, which is misleading due to reverse causation in the course of malignancy.

Immunotherapy for HER-2 positive breast cancer.

Immunotherapy is a developing treatment for advanced breast cancer. Immunotherapy has clinical significance for the treatment of triple-negative breast cancers and human epidermal growth factor receptor-2 positive (HER2+) breast cancers. As a proved effective passive immunotherapy, clinical application of the monoclonal antibodies trastuzumab, pertuzumab and T-DM1 (ado-trastuzumab emtansine) has significantly improved the survival of patients with HER2+ breast cancers. Immune checkpoint inhibitors that block programmed death receptor-1 and its ligand (PD-1/PD-L1) have also shown benefits for breast cancer in various clinical trials. Adoptive T-cell immunotherapies and tumor vaccines are emerging as novel approaches to treating breast cancer, but require further study. This article reviews recent advances in immunotherapy for HER2+ breast cancers.

Age-related cataract: GSTP1 ubiquitination and degradation by Parkin inhibits its anti-apoptosis in lens epithelial cells.

PURPOSE: Oxidative stress-induced apoptosis of lens epithelial cells (LECs) contributes to the pathogenesis of age-related cataract (ARC). The purpose of this research is to underlie the potential mechanism of E3 ligase Parkin and its oxidative stress-associated substrate in cataractogenesis. METHODS: The central anterior capsules were obtained from patients with ARC, Emory mice, and corresponding controls. SRA01/04 cells were exposed to H2O2 combined with cycloheximide (a translational inhibitor), MG-132 (a proteasome inhibitor), chloroquine (an autophagy inhibitor), Mdivi-1 (a mitochondrial division inhibitor), respectively. Co-immunoprecipitation was employed to detect protein-protein interactions and ubiquitin-tagged protein products. Levels of proteins and mRNA were evaluated by western blotting and quantitative RT-PCR assays. RESULTS: Glutathione-S-transferase P1 (GSTP1) was identified as a novel Parkin substrate. Compared with corresponding controls, GSTP1 was significantly decreased in the anterior lens capsules obtained from human cataracts and Emory mice. Similarly, GSTP1 was declined in H2O2-stimulated SRA01/04 cells. Ectopic expression of GSTP1 mitigated H2O2-induced apoptosis, whereas silencing GSTP1 aggregated apoptosis. In addition, H2O2 stimulation and Parkin overexpression could promote the degradation of GSTP1 through the ubiquitin-proteasome system, autophagy-lysosome pathway, and mitophagy. After co-transfection with Parkin, the non-ubiquitinatable GSTP1 mutant maintained its anti-apoptotic function, while wildtype GSTP1 failed. Mechanistically, GSTP1 might promote mitochondrial fusion through upregulating Mitofusins 1/2 (MFN1/2). CONCLUSION: Oxidative stress induces LECs apoptosis via Parkin-regulated degradation of GSTP1, which may provide potential targets for ARC therapy.

Ubiquitination of Ku70 by Parkin promotes apoptosis of lens epithelial cells.

Oxidative damage-triggered apoptosis in lens epithelial cells is considered as a main risk factor in the pathogenesis of age-related cataracts. Ku70 is a key factor in the DNA repair process of double-strand breaks. In the present study, we aimed to investigate the role of Ku70 and its related E3 ubiquitin ligase in lens epithelial cell apoptosis. The levels of Ku70 in the anterior lens capsules of human cataracts and Emory mice were lower compared to controls. H2 O2 treatment resulted in decreased expression of Ku70 through accelerating Ku70 ubiquitination. Parkin, an E3 ubiquitin ligase, could interact with Ku70 and promote the ubiquitination and degradation of this protein. In addition, ubiquitinated Ku70 was regulated by ubiquitin-proteasome, autophagy-lysosome and mitophagy pathways. Ectopic expression of Ku70 protected SRA01/04 cells from H2 O2 -induced apoptosis, whereas silencing Ku70 exhibited the opposite trend. Co-transfected with Parkin non-ubiquitinatable Ku70 mutant could maintain its anti-apoptosis ability, whereas wild-type Ku70 failed. Moreover, Ku70 might facilitate mitochondrial fusion by increasing the expression of Mitofusin 1/2. The present study revealed that Parkin-mediated Ku70 ubiquitination facilitated H2 O2 -induced lens epithelial cell apoptosis through alleviating mitochondrial fusion, which could provide potential targets for age-related cataract treatment.

COF-based artificial probiotic for modulation of gut microbiota and immune microenvironment in inflammatory bowel disease.

Conventional strategies for treating inflammatory bowel disease merely relieve inflammation and excessive immune response, but fail to solve the underlying causes of IBD, such as disrupted gut microbiota and intestinal barrier. Recently, natural probiotics have shown tremendous potential for the treatment of IBD. However, probiotics are not recommended for IBD patients, as they may cause bacteremia or sepsis. Herein, for the first time, we constructed artificial probiotics (Aprobiotics) based on artificial enzyme-dispersed covalent organic frameworks (COFs) as the "organelle" and a yeast shell as the membrane of the Aprobiotics to manage IBD. The COF-based artificial probiotics, with the function of natural probiotics, could markedly relieve IBD by modulating the gut microbiota, suppressing intestinal inflammation, protecting the intestinal epithelial cells, and regulating immunity. This nature-inspired approach may aid in the design of more artificial systems for the treatment of various incurable diseases, such as multidrug-resistant bacterial infection, cancer, and others.

Role of SUMOylation of STAT1 in tubular epithelial­mesenchymal transition induced by high glucose.

Tubulointerstitial fibrosis (TIF) is an important pathological change that occurs during the development of diabetic kidney disease. The epithelial­mesenchymal transition (EMT) of renal tubular epithelial cells is a manifestation of TIF. STAT1, a member of the STAT family of transcription factors, can be modified by the small ubiquitin­related modifier (SUMO), thus affecting the activity of STAT1. The present study investigated the role of STAT1 SUMOylation in high glucose­induced tubular EMT by western blotting, immunocytochemistry, immunofluorescence, co­immunoprecipitation and dual luciferase reporter analysis. The results indicated that in the process of high glucose­induced EMT, STAT1 activation protected the cells from EMT. However, high glucose also increased the SUMOylation of STAT1, which prevented STAT1 from exerting an effective protective role by inhibiting its activity.

Exosomal microRNA-222-3p increases UVB sensitivity of lens epithelium cells by suppressing MGMT.

BACKGROUND: Age-related cataract (ARC) is a leading cause of blindness worldwide with multiple pathogenic factors. Oxidative damage of lens epithelium cells (LECs) is one of the well-accepted pathogenesis of ARC which can be regulated by DNA repair genes (DRGs). The present research aimed to clarify the regulatory mechanism of exosomal microRNAs (miRNAs) on DRGs in LECs. METHODS: The LECs oxidative damage model was established by UVB-irradiation on SRA01/04 (human lens epithelium cell line). Exosomes from UVB-irradiated cells (UVB-exo) and exosomes from normal control cells (NC-exo) were collected from the culture medium. To explore the functions of LECs exosomes, SRA01/04 were incubated with UVB-exo/NC-exo. Then, we detected SRA01/04 proliferation, viability and apoptosis respectively using 5'-ethynyl-2'-deoxyuridine (EdU), cell-counting kit-8 (CCK-8) and TdT-mediated dUTP Nick-End Labeling (TUNEL) assay. Next, the miRNA expression profiles of UVB-exo and NC-exo were identified by miRNA microarrays. RNA expression in exosomes, cells, and clinical samples was verified by qRT-PCR. The location and expression of MGMT and CD63 proteins were detected by immunofluorescence and western blot. The 3'UTR regulation of miR-222-3p to MGMT was verified by luciferase analyses. RESULTS: MGMT down-regulated while miR-222-3p up-regulated in LECs sub-central anterior capsule from ARC lenses. MGMT and miR-222-3p expressions in central and peripheral LECs from anterior lens capsules were differential. UVB-exo can transport the up-regulated miR-222-3p from oxidative-damaged LECs to normal LECs, which could suppress MGMT expression and increase UVB sensitivity of LECs. CONCLUSIONS: Findings on exosomal miRNA functions provided novel insights into pathogenesis of ARC. Exosomal miR-222-3p can be a potential target for prevention and cure of ARC.

miR-125a-3p regulates apoptosis by suppressing TMBIM4 in lens epithelial cells.

PURPOSE: To explore the regulatory effect of miR-125a-3p on lens epithelial cells (LECs) under ultraviolet radiation B (UVB) irradiation. METHODS: The expression of miR-125a-3p in age-related cataract (ARC) specimens and cell models was detected by qRT-PCR. UVB was utilized to establish DNA damage model of LECs. Cell count kit-8 was applied in detecting cell viability. Cell apoptosis ratio was analyzed by flow cytometry. Dual luciferase reports were applied to analyze the mechanism between miRNA and target genes. Nanoparticle tracking analysis, and Western blot were used to identify whether the exosomes were typical exosomes. RESULTS: miR-125a-3p was upregulated in ARC tissues and LECs treated with UVB. Knockdown of miR-125a-3p in LECs significantly decreased apoptosis and increased viability of UVB-irradiated LECs. We predicted that miR-125a-3p could regulate transmembrane Bax inhibitor motif containing 4 (TMBIM4) by the bioinformatics databases TargetScan, miRBase, and miRWalk. Luciferase reporter assays demonstrated that miR-125a-3p may suppress TMBIM4 protein translation by binding to 3'UTR of TMBIM4 mRNA. Overexpression of miR-125a-3p decreased TMBIM4, which suggested that miR-125a-3p could inhibit TMBIM4. Moreover, knockdown of TMBIM4 decreased cell viability and enhanced cell apoptosis during UVB irradiation. In addition, the exosome secretion of LECs irradiated by UVB was enhanced, and the expression of miR-125a-3p was high. Cell viability was significantly decreased, and cell apoptosis was increased during UVB-exos treatment. CONCLUSION: This study indicated that miR-125a-3p regulated apoptosis by suppressing TMBIM4 in LECs under oxidative damage, providing a new idea for clinical therapeutic target of cataract.

SYVN1-mediated ubiquitination and degradation of MSH3 promotes the apoptosis of lens epithelial cells.

The pathology of age-related cataract (ARC) mainly involves the misfolding and aggregation of proteins, especially oxidative damage repair proteins, in the lens, induced by ultraviolet-B (UVB). MSH3, as a key member of the mismatch repair family, primarily maintains genome stability. However, the function of MSH3 and the mechanism by which cells maintain MSH3 proteostasis during cataractogenesis remains unknown. In the present study, the protein expression levels of MSH3 were found to be attenuated in ARC specimens and SRA01/04 cells under UVB exposure. The ectopic expression of MSH3 notably impeded UVB-induced apoptosis, whereas the knockdown of MSH3 promoted apoptosis. Protein half-life assay revealed that UVB irradiation accelerated the decline of MSH3 by ubiquitination and degradation. Subsequently, we found that E3 ubiquitin ligase synoviolin (SYVN1) interacted with MSH3 and promoted its ubiquitination and degradation. Of note, the expression and function of SYVN1 were contrary to those of MSH3 and SYVN1 regulated MSH3 protein degradation via the ubiquitin-proteasome pathway and the autophagy-lysosome pathway. Based on these findings, we propose a mechanism for ARC pathogenesis that involves SYVN1-mediated degradation of MSH3 via the ubiquitin-proteasome pathway and the autophagy-lysosome pathway, and suggest that interventions targeting SYVN1 might be a potential therapeutic strategy for ARC.

XRCC5 downregulated by TRIM25 is susceptible for lens epithelial cell apoptosis.

Exposure of the lens to UVB can lead to oxidative stress, which would result in age-related cataract (ARC) formation. In this study, we investigate the regulatory mechanism of tripartite motif containing 25 (TRIM25) in ARC. The protein level of TRIM25 was elevated in ARC specimens and UVB-exposed SRA01/04 cells. Bioinformatic analysis indicated that X-ray repair cross complementing 5 (XRCC5) might interact with TRIM25, and the interaction was validated via immunoprecipitation. TRIM25 interacted with XRCC5 and ubiquitinated it for degradation. Further studies showed that XRCC5 overexpression notably repressed UVB-induced apoptosis, while XRCC5 knockdown promoted apoptosis. Of note, ubiquitination of XRCC5 mediated by TRIM25 overexpression facilitated apoptosis. Attenuation of XRCC5 ubiquitination by mutant with substitution of lysine residues with arginine residues rescued its anti-apoptosis effect. Moreover, we observed that TRIM25-mediated XRCC5 degradation was reversed by proteasome inhibitor MG-132 or lysosome inhibitor 3-MA. In conclusion, TRIM25 mediates ubiquitination of XRCC5 to regulate the function and degradation of XRCC5, suggesting that interventions targeting TRIM25 might be a promising therapeutic strategy for ARC.

MicroRNA Let-7c-5p-Mediated Regulation of ERCC6 Disrupts Autophagic Flux in Age-Related Cataract via the Binding to VCP.

Purpose: DNA damage contributes to the pathogenesis of age-related cataract (ARC) and is repaired through the nucleotide excision repair (NER) pathway, which includes ERCC6. Evidence has demonstrated that defective autophagy leads to lens organelle degradation and cataract. This study aimed to investigate the effects of ERCC6 on autophagy and determine its mechanisms in ARC.Methods: The clinical case-control study comprised 30 patients with ARC and 30 age-matched controls who received transparent lens extraction. Transmission electron microscopy was used to assess the ultrastructure of autophagic vesicles in lens anterior capsule tissues and lens epithelial cell line (SRA01/04). Real-time polymerase chain reaction and western blot analyses were performed to measure relative gene expression levels. Gene expression levels and localization were assessed by immunofluorescence. A coimmunoprecipitation assay was used to investigate the relationship between CSB which encoded by ERCC6 and VCP. ERCC6-siRNA and let-7 c-5p mimic were used to alter the expression of ERCC6 and let-7 c-5p.Results: Autophagy induction occurred in lens anterior capsule tissues of patients with ARC and in UVB-induced SRA01/04 cells, where the number of LC3B puncta was increased. Consistent with this result, the expression of beclin1 (BECN1) and LC3B, in addition to that of p62, was increased. Additionally, ERCC6 expression decreased, and silencing ERCC6 induced increases in the expression of BECN1, LC3B and p62. Moreover, CSB interacted with VCP, and let-7 c-5p induced dysregulation of autophagy by targeting ERCC6.Conclusion: In ARC, Let-7 c-5p-mediated downregulation of ERCC6 might prevent the degradation of autophagic vacuoles. CSB binds to VCP, inducing autophagosomes to combine with lysosomes and be degraded.

Profiling and Integrated Analysis of the ERCC6-regulated circRNA-miRNA-mRNA Network in Lens Epithelial Cells.

Purpose: To explore the regulatory role of ERCC6 in the circRNA-miRNA-mRNA network using a cellular ERCC6 overexpression model (OE-ERCC6) in lens epithelial cells.Methods: The expression profiles of circRNAs, miRNAs and mRNAs were determined by RNA-seq, and a regulatory circRNA-miRNA-mRNA network was constructed via bioinformatics. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were used for the functional annotation of circRNA host genes, differentially expressed (DE) genes, and miRNA targets.Results: The DE molecules between the OE-ERCC6 and control groups included 269 circRNAs, 241 miRNAs and 3500 mRNAs. We validated 5 selected DE reads of circRNAs (hsa_circ_0001009, hsa_circ_0002024, hsa_circ_0004592, hsa_circ_0001900 and hsa_circ_0001017). Subsequent bioinformatics analysis revealed that the DE circRNAs are mainly involved in oxidative stress- and cell death-related signaling pathways. Finally, a circRNA-miRNA-mRNA network focusing on DNA damage and cell death, which involved 5 circRNAs, 13 miRNAs and 107 mRNAs, was constructed.Conclusion: We constructed a circRNA-miRNA-mRNA network that is regulated by ERCC6. DE circRNAs have the potential to become therapeutic targets related to the lens lesions observed in ARC. The establishment of related in vivo and in vitro models could be a future direction to confirm these hypotheses.

Ginsenoside Rg1 Prevents H2O2-induced Lens Opacity.

RESULTS: Exposure of 0.2 mM H2O2 in lenses resulted in obvious cloudiness and typical pathological changes of cataract such as rupture of the lens capsule, degenerative lens epithelial cells (LECs), etc. Rg1 effectively prevented lens opacity caused by H2O2. After Rg1 treatment, lens WSP content, the level of SOD, total GSH, and reduced GSH were increased, while the level of MDA and oxidized GSH were decreased. In addition, MDA concentration of lens by Rg1 treatment only was found to be lower than the controls. Rg1 attenuated H2O2-induced cell injury at the concentration of 0.4 mM that it elevated cell activity, and peaked at 0.6 mM. CONCLUSIONS: This study demonstrated that Rg1 might have the capability to protect lens against oxidative stress-induced cataract, at least by local administration.Abbreviations: LECs: lens epithelial cells; Rg1: Ginsenoside Rg1; SD: Sprague-Dawley; ROS: reactive oxygen species; SOD: Superoxide Dismutase; GSH: glutathione; MDA: Malonediadehyde; H2O2: Hydrogen peroxide.

Ganoderic acid A protects lens epithelial cells from UVB irradiation and delays lens opacity.

A contributory role of oxidative stress and protection by antioxidant nutrients have been suspected in cataract formation. Ganoderic acid A (GAA), an effective lanostane triterpene, is widely reported as an antioxidant. The aim of this study is to investigate the potential effects of GAA on cataract formation. After lens epithelial cells (LECs) were exposed to UVB radiation for different periods, cell viability, apoptosis-related protein levels, malondialdehyde (MDA) and superoxide dismutase (SOD) activities were monitored. We found that cell viability, the Bcl-2/Bax ratio and SOD activity were increased, while Cleaved caspase-3 levels and MDA activity were decreased compared with those in UVB-impaired LECs after GAA treated. Furthermore, GAA activated PI3K/AKT in UVB-impaired LECs and effectively delayed the occurrence of lens opacity in vitro. In conclusion, these findings demonstrated that GAA exhibited protective functions in SRA01/04 cells and rat lenses against UVB-evoked impairment through elevating cell viability and antioxidant activity, inhibiting cell apoptosis, activating the PI3K/AKT pathway and delaying lens opacity.

A Smart Nanoparticle-Laden and Remote-Controlled Self-Destructive Macrophage for Enhanced Chemo/Chemodynamic Synergistic Therapy.

Macrophages are known to penetrate tumor central hypoxic areas and hold great potential in cancer drug delivery. However, it remains a big challenge for current macrophage-based drug delivery systems (MDDSs) to prevent premature drug leakage and sufficiently release the therapeutics in tumor sites. Moreover, these MDDSs would encounter drug resistance and a hypoxic microenvironment in solid tumors, which further compromised their therapeutic efficacy. Herein, by internalizing a smart nanoparticle (doxorubicin (DOX)-loaded mesoporous carbon nanosphere wrapped with MnO2 shell) into macrophages, a macrophage vehicle (MMDM) is developed for enhanced chemo/chemodynamic synergistic therapy. The resulting MMDM could avoid premature drug leakage-induced cell dysfunction and maximally maintain cell viability. After accumulating in tumor tissues, the MMDM could be destroyed under a near-infrared laser to sufficiently release the nanoparticle out of the carrier macrophages. The released nanoparticle could then decompose H2O2 to generate O2 in the tumor microenvironment to relieve tumor hypoxia. Meanwhile, the MnO2 shell of the nanoparticle is reduced to Mn2+ by intracellular glutathione, triggering the release of DOX and subsequently resulting in an enhanced Mn2+-mediated Fenton-like reaction. This study provides an intriguing strategy to macrophage-based delivery systems for enhanced chemo/chemodynamic synergistic therapy.

Identification and Characterization of N6-Methyladenosine CircRNAs and Methyltransferases in the Lens Epithelium Cells From Age-Related Cataract.

Purpose: To explore the involvement of N6-methyladenosine (m6A) modification in circular RNAs (circRNAs) and relevant methyltransferases in the lesion of lens epithelium cells (LECs) under the circumstances of age-related cataract (ARC). Methods: LECs were collected from normal subjects and patients with cortical type of ARC (ARCC). M6A-tagged circRNAs and circRNAs expression were analyzed by m6A-modified RNA immunoprecipitation sequencing (m6A-RIP-seq) and RNA sequencing (RNA-seq). Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were used to predict possible functions of the m6A-circRNAs. Expression of m6A-related methyltransferase and demethytransferase was measured by quantitative real-time polymerase chain reaction. Expression and location of AlkB homolog 5 RNA demethylase (ALKBH5), a key component of m6A demethytransferase, were determined by Western blot and immunostaining. Results: All 4646 m6A peaks within circRNAs had different abundances, with 2472 enriched and 2174 subdued. The level of m6A abundance in total circRNAs was decreased in the LECs from ARCCs in comparison with the controls. We also found that the expression of highly m6A-tagged circRNAs was mostly decreased in comparison with non-m6A-tagged circRNAs. The bioinformatics analysis predicted the potential functions of m6A modified circRNAs and the relevant pathways that may be associated with m6A modified circRNAs. Among five major methyltransferases, ALKBH5 was significantly upregulated in LECs of ARCCs. Conclusions: Our data provided novel evidence regarding the involvement of circRNAs m6A modifications in ARC. The altered expression of methyltransferases in lens tissue might selectively change the epigenetic profile of lens genome through regulating genes that host the circRNAs, thus enhance the susceptibility to ARC. The results might provide a new insight in the molecular target of ARC pathogenesis.

Long non-coding RNA nuclear paraspeckle assembly transcript 1 protects human lens epithelial cells against H2O2 stimuli through the nuclear factor kappa b/p65 and p38/mitogen-activated protein kinase axis.

BACKGROUND: Long non-coding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (NEAT1) plays a regulatory role in many biological processes; however, its role in cataracts has yet to be illuminated. This study aimed to investigate the protective role of NEAT1 in hydrogen peroxide (H2O2)-treated human lens epithelial cells (HLECs) and its underlying molecular mechanism. METHODS: HLECs (SRA01/04) were treated with 300 µM H2O2 to mimic cataract in vitro. Cell viability was detected by performing an MTT assay and EdU staining. Flow cytometry was carried out to detect apoptosis of HLECs. DNA damage was examined using γ-H2A histone family member X staining. and reactive oxygen species (ROS) production was measured using 2',7'dichlorofluorescin diacetate staining. The expression levels of lncRNA and proteins were detected with quantitative real-time polymerase chain reaction and western blot, respectively. RESULTS: The expression of NEAT1 was observed to be increased in H2O2-treated HLECs and age-related cataract (ARC) tissues. Knockdown NEAT1 strongly protected against H2O2-induced cell death and also regulated the expression of cleaved caspase-3, B-cell lymphoma 2, and Bcl-2-associated X protein. Further, knockdown NEAT1 also significantly suppressed H2O2-induced intracellular ROS production and malondialdehyde (MDA) content, but elevated the glutathione (GSH) activity of H2O2-treated cells. Also, it is demonstrated that si-NEAT1 greatly inhibited H2O2-induced phosphorylation of NF-кB p65 and p38 MAPK. CONCLUSIONS: This study confirmed that knockdown NEAT1 attenuated H2O2-induced damage in HLECs, and inhibited the oxidative stress and apoptosis of HLECs via regulating nuclear factor-kappa B (NF-κB) p65 and p38 MAPK signaling. It may provide a potential target for clinical treatment of cataracts.