Dynamic RGD ligands derived from highly mobile cyclodextrins regulate spreading and proliferation of endothelial cells to promote vasculogenesis.

A thiolated RGD was incorporated into the threaded allyl-ß-cyclodextrins (Allyl-ß-CDs) of the polyrotaxane (PR) through a thiol-ene click reaction, resulting in the formation of dynamic RGD ligands on the PR surface (dRGD-PR). When maintaining consistent RGD density and other physical properties, endothelial cells (ECs) cultured on dRGD-PR exhibited significantly increased cell proliferation and a larger cell spreading area compared to those on the non-dynamic RGD (nRGD-PCL). Furthermore, ECs on dRGD-PR demonstrated elevated expression levels of FAK, p-FAK, and p-AKT, along with a larger population of cells in the G2/M stage during cell cycle analysis, in contrast to cells on nRGD-PCL. These findings suggest that the movement of the RGD ligands may exert additional beneficial effects in promoting EC spreading and proliferation, beyond their essential adhesion and proliferation-promoting capabilities, possibly mediated by the RGD-integrin-FAK-AKT pathway. Moreover, in vitro vasculogenesis tests were conducted using two methods, revealing that ECs cultured on dRGD-PR exhibited much better vasculogenesis than nRGD-PCL in vitro. In vivo testing further demonstrated an increased presence of CD31-positive tissues on dRGD-PR. In conclusion, the enhanced EC spreading and proliferation resulting from the dynamic RGD ligands may contribute to improved in vitro vasculogenesis and in vivo vascularization.

Study on wet oxidation process and mechanism for ethylene spent caustic.

Ethylene spent caustic with strong alkalinity, high chemical oxygen demand (COD) and toxicity could not be directly discharged without pretreatment. In this paper, the composition of spent caustic from a petrochemical enterprise was analyzed. Effects of oxidants, reaction temperature, pressure and residence time on the removal ratio of COD in the spent caustic were investigated. The oxidation reaction mechanism of main organic sulphides in the spent caustic was discussed. Results show that COD of the ethylene spent caustic is ∼ 24.5 g/L, the main compositions are S2- and disulphides including a small amount of ketones and phenolic organics. The removal ratio of S2- in spent caustic is up to 100%, and the removal ratio of COD is over 83% at 180 °C with the pressure of 2 MPa and residence time of 15 min using oxygen as an oxidant. Taking dimethyl disulphide as an example, dimethyl disulphide with HO• firstly produces methyl thiosulphonate and methyl sulphenic acid, then further oxidized to generate methanesulphonic acid, and the C-S bonds in the methanesulphonic acid were broken to generate sulphuric acid and formic acid. So the oxidative degradation products of organic sulphides are sulphuric acid, formic acid and a small amount of acetic acid.

Construction of PARPi Resistance-related Competing Endogenous RNA Network.

Objective: Ovarian cancer is a kind of common gynecological malignancy in women. PARP inhibitors (PARPi) have been approved for ovarian cancer treatment. However, the primary and acquired resistance have limited the application of PARPi. The mechanisms remain to be elucidated. Methods: In this study, we characterized the expression profiles of mRNA and nonconding RNAs (ncRNAs) and constructed the regulatory networks based on RNA sequencing in PARPi Olaparib-induced ovarian cancer cells. Results: We found that the functions of the differentially expressed genes were enriched in "PI3K/AKT signaling pathway," "MAPK signaling pathway" and "metabolic process". The functions of DELs (cis) were enriched in "Human papillomavirus infection""tight junction" "MAPK signaling pathway". As the central regulator of ceRNAs, the differentially expressed miRNAs were enriched in "Human papillomavirus infection" "MAPK signaling pathway" "Ras signaling pathway". According to the degree of interaction, we identified 3 lncRNAs, 2 circRNAs, 7 miRNAs, and 12 mRNA as the key regulatory ceRNA axis, in which miR-320b was the important mediator. Conclusion: Here, we revealed the key regulatory lncRNA (circRNA)-miRNA-mRNA axis and their involved pathways in the PARPi resistant ovarian cancer cells. These findings provide new insights into exploring the ceRNA regulatory networks and developing new targets for PARPi resistance.

Curcumin Ameliorates White Matter Injury after Ischemic Stroke by Inhibiting Microglia/Macrophage Pyroptosis through NF-κB Suppression and NLRP3 Inflammasome Inhibition.

NLRP3 inflammasome-mediated pyroptosis is a proinflammatory programmed cell death pathway, which plays a vital role in functional outcomes after stroke. We previously described the beneficial effects of curcumin against stroke-induced neuronal damage through modulating microglial polarization. However, the impact of curcumin on microglial pyroptosis remains unknown. Here, stroke was modeled in mice by middle cerebral artery occlusion (MCAO) for 60 minutes and treated with curcumin (150 mg/kg) intraperitoneally immediately after reperfusion, followed by daily administrations for 7 days. Curcumin ameliorated white matter (WM) lesions and brain tissue loss 21 days poststroke and improved sensorimotor function 3, 10, and 21 days after stroke. Furthermore, curcumin significantly reduced the number of gasdermin D+ (GSDMD+) Iba1+ and caspase-1+Iba1+ microglia/macrophage 21 days after stroke. In vitro, lipopolysaccharide (LPS) with ATP treatment was used to induce pyroptosis in primary microglia. Western blot revealed a decrease in pyroptosis-related proteins, e.g., GSDMD-N, cleaved caspase-1, NLRP3, IL-1ß, and IL-18, following in vitro or in vivo curcumin treatment. Mechanistically, both in vivo and in vitro studies confirmed that curcumin inhibited the activation of the NF-κB pathway. NLRP3 knocked down by siRNA transfection markedly increased the inhibitory effects of curcumin on microglial pyroptosis and proinflammatory responses, both in vitro and in vivo. Furthermore, stereotaxic microinjection of AAV-based NLRP3 shRNA significantly improved sensorimotor function and reduced WM lesion following curcumin treatment in MCAO mice. Our study suggested that curcumin reduced stroke-induced WM damage, improved functional outcomes, and attenuated microglial pyroptosis, at least partially, through suppression of the NF-κB/NLRP3 signaling pathway, further supporting curcumin as a potential therapeutic drug for stroke.

Current insights into the implications of m6A RNA methylation and autophagy interaction in human diseases.

Autophagy is a conserved degradation process crucial to maintaining the primary function of cellular and organismal metabolism. Impaired autophagy could develop numerous diseases, including cancer, cardiomyopathy, neurodegenerative disorders, and aging. N6-methyladenosine (m6A) is the most common RNA modification in eukaryotic cells, and the fate of m6A modified transcripts is controlled by m6A RNA binding proteins. m6A modification influences mRNA alternative splicing, stability, translation, and subcellular localization. Intriguingly, recent studies show that m6A RNA methylation could alter the expression of essential autophagy-related (ATG) genes and influence the autophagy function. Thus, both m6A modification and autophagy could play a crucial role in the onset and progression of various human diseases. In this review, we summarize the latest studies describing the impact of m6A modification in autophagy regulation and discuss the role of m6A modification-autophagy axis in different human diseases, including obesity, heart disease, azoospermatism or oligospermatism, intervertebral disc degeneration, and cancer. The comprehensive understanding of the m6A modification and autophagy interplay may help in interpreting their impact on human diseases and may aid in devising future therapeutic strategies.

Melatonin Protects Against Ischemic Brain Injury by Modulating PI3K/AKT Signaling Pathway via Suppression of PTEN Activity.

Stroke is one of the leading causes of death and disability worldwide with limited therapeutic options. Melatonin can attenuate ischemic brain damage with improved functional outcomes. However, the cellular mechanisms of melatonin-driven neuroprotection against post-stroke neuronal death remain unknown. Here, distal middle cerebral artery occlusion (dMCAO) was performed in C57BL/6j mice to develop an ischemic stroke in vivo model. Melatonin was injected intraperitoneally immediately after ischemia, and 24 and 48 hours later. Melatonin treatment, with 5 to 20 mg/kg, elicited a dose-dependent decrease in infarct volume and concomitant increase in sensorimotor function. At the molecular level, phosphorylation of PTEN and Akt were increased, whereas PTEN activity was decreased in melatonin treated animals 72 hours after dMCAO. At the cellular level, oxygenglucose deprivation (OGD) challenge of neuronal cell line Neuro-2a (N2a) and primary neurons supported melatonin's direct protection against neuronal cell death. Melatonin treatment reduced LDH release and neuronal apoptosis at various time points, markedly increased Akt phosphorylation in neuronal membrane, but significantly suppressed it in the cytoplasm of post-OGD neurons. Mechanistically, melatonin-induced Akt phosphorylation and neuronal survival was blocked by Wortmannin, a potent PIP3 inhibitor, exposing increased PI3K/Akt activation as a central player in melatonin-driven neuroprotection. Finally, PTEN knock-down through siRNA significantly inhibited PI3K/Akt activation and cell survival following melatonin treatment, suggesting that melatonin protection against ischemic brain damage, is at least partially, dependent on modulation of the PTEN/PI3K/Akt signaling axis.

Cell Models for Birth Defects Caused by Chloroethyl Nitrosourea-Induced DNA Lesions.

ABSTRACT: Birth defects have been linked to administration of alkylating agents during pregnancy. The anti-tumor efficacy of alkylating agents correlate with their ability to induce DNA lesions, especially interstrand crosslinks (ICLs). Yet the role of DNA damages in birth defects remains to be clarified, owing, in part, to a lack of cell models. Here we generate DNA lesions in NIH/3T3 cells to mimic defects in fetus triggered by 3-Bis(2-chloroethyl)-1-nitrosourea (BCNU, carmustine). CCK-8 assay suggests that BCNU-induced cell death was dose-dependent. Alkaline comet tests and γ-H2AX staining confirm DNA ICLs and other forms of DNA damages caused by BCNUs. The cell cycle analysis shows cells arrest in G2/M phase until crosslinks repair is complete. Taken together, all these experiments demonstrate we have successfully established normal cell models for birth defects caused by BCNU-mediated DNA damages. The model can not only guide the development of effective and low-toxicity anticancer drugs, but also be of great significance for the study of neonatal malformation triggered by BCNUs.

Analysis of trace organics and its correlation with COD in condensate from natural gas to hydrogen production.

Qualitative and quantitative analysis of trace organics in the condensate and its correlation with chemical oxygen demand (COD) is the key to the research on the reuse technology of condensate (condensate) from natural gas to hydrogen production process. The contents of anions, COD, total organic carbon (TOC) and total nitrogen (TN) were measured by ion chromatography and the TOC analyzer. Trace organics in the condensate and its correlation with COD was investigated in this paper. Results show that the contents of COD and TOC is 74.1 and 17.81 mg/L, respectively, and the anions in the condensate are mainly Cl-, I-, and SO4 2-, etc. The condensate mainly contains small molecule organics including methanol, ethanol and formic acid with the content of 41.4, 2.1 and 3.2 mg/L, respectively. The spiked recovery of methanol, ethanol and formic acid is 96.1%, 100.2% and 103.9% by high performance liquid chromatography (HPLC) and gas chromatography (GC), respectively. Methanol is the main source of COD in the condensate, and the contribution rate reaches up to 83.8%. The removal of trace methanol can significantly reduce the COD of the condensate. This work might provide basic data for reasonable recovery and utilization of condensate in the hydrogen production process.

Melatonin protects against ischemic stroke by modulating microglia/macrophage polarization toward anti-inflammatory phenotype through STAT3 pathway.

AIMS: Microglia and infiltrated macrophages play important roles in inflammatory processes after ischemic stroke. Modulating microglia/macrophage polarization from pro-inflammatory phenotype to anti-inflammatory state has been suggested as a potential therapeutic approach in the treatment of ischemic stroke. Melatonin has been shown to be neuroprotective in experimental stroke models. However, the effect of melatonin on microglia polarization after stroke and underlying mechanisms remain unknown. METHODS: In vivo, cerebral ischemia was induced by distal middle cerebral artery occlusion (dMCAO) in C57BL/6J mice. Melatonin was injected intraperitoneally (20 mg/kg) at 0 and 24 hours after ischemia. In vitro, the microglial cell line BV2 was stimulated to the pro-inflammatory state with conditioned media (CM) collected from oxygen-glucose deprivation (OGD) challenged neuronal cell line Neuro-2a (N2a). Real-time PCR was utilized to detect the mRNA expression of microglia phenotype markers. Activation of signal transducer and activator of transcription 3 (STAT3) pathway was determined by Western blot of phosphorylated STAT3 (pSTAT3). A neuron-microglia co-culture system was used to determine whether melatonin can inhibit the neurotoxic effect of pro-inflammatory microglia to post-OGD neurons. RESULTS: Melatonin treatment reduced brain infarct and improved neurological functions 3 days after dMCAO, which was accompanied by decreased expression of pro-inflammatory markers and increased expression of anti-inflammatory markers in the ischemic brain. In vitro studies confirmed that melatonin directly inhibited the pro-inflammatory responses in BV2 cells upon exposure to OGD neuron CM. The microglia possessing pro-inflammatory phenotype exacerbated post-OGD N2a cells death, whereas melatonin reduced such neurotoxic effect. Further, melatonin enhanced the otherwise inhibited pSTAT3 expression in BV2 cells treated with OGD neuron CM. STAT3 blockade significantly reduced the effect of melatonin on microglial phenotype shift. CONCLUSION: Melatonin treatment ameliorates brain damage at least partially through shifting microglia phenotype from pro-inflammatory to anti-inflammatory polarity in a STAT3-dependent manner.

A study on the protective effects of CpG oligodeoxynucleotide-induced mucosal immunity against lung injury in a mouse acute respiratory distress syndrome model.

This study aims to determine the feasibility of using oligodeoxynucleotides with unmethylated cytosine-guanine dinucleotide sequences (CpG ODN) as an immunity protection strategy for a mouse model of acute respiratory distress syndrome (ARDS). This is a prospective laboratory animal investigation. Twenty-week-old BALB/c mice in Animal research laboratory were randomized into groups. An ARDS model was induced in mice using lipopolysaccharides (LPSs). CpG ODN was intranasally and transrectally immunized before or after the 3rd and 7th days of establishing the ARDS model. Mice were euthanized on Day 7 after the second immunization. Then, retroorbital bleeding was carried out and the chest was rapidly opened to collect the trachea and tissues from both lungs for testing. CpG ODN significantly improved the pathologic impairment in mice lung, especially after the intranasal administration of 50 µg. This resulted in the least severe lung tissue injury. Furthermore, interleukin-6 (IL-6) and IL-8 concentrations were lower, which was second to mice treated with the rectal administration of 20 µg CpG ODN. In contrast, the nasal and rectal administration of CpG ODN in BALB/c mice before LPS immunization did not appear to exhibit any significant protective effects. The intranasal administration of CpG ODN may be a potential treatment approach to ARDS. More studies are needed to further determine the protective mechanism of CpG ODN.

Melatonin improves functional recovery in female rats after acute spinal cord injury by modulating polarization of spinal microglial/macrophages.

Spinal cord injury (SCI) is a devastating neurological disorder, but few drugs have proven to be effective for its treatment. Neuroinflammation exaggerates the secondary injury subsequent to trauma. Emerging evidence suggests that melatonin may help protect neural tissue against secondary injury after SCI, but the underlying mechanisms remain elusive. Microglial/macrophages polarization plays an important role in regulating immune responses. To examine whether melatonin exerts neuroprotective effects after acute SCI by regulating microglial/macrophages polarization in the spinal cord, we administered intraperitoneal injections of melatonin (50 mg/kg) in female rats immediately after SCI and then daily for seven consecutive days (n = 6). Compared with the vehicle-treated group (n = 6), the melatonin-treated group exhibited a greater Basso, Beattie, and Bresnahan locomotor rating score, smaller spinal cavity, and less cleaved caspase 3 immunofluorescence staining in the injured spinal segments. Real-time PCR data revealed decreases in the expression levels of M1 microglia phenotypic markers and increases in M2 markers in the spinal cord of melatonin-treated SCI rats, as compared to levels in the vehicle-treated group. Melatonin increased the number of CD206+ and Arg1+ cells, decreased the number of CD16+ and iNOS+ cells and reduced the levels of pro-inflammatory cytokines (TNF-α, IL-6, and IL-1ß) in the spinal cord tissue of female SCI rats. Current findings suggest that melatonin may inhibit pro-inflammatory responses and promote M2 polarization of microglial/macrophages in the spinal cord in the early stage of SCI, facilitating functional recovery. Accordingly, melatonin may represent a promising therapeutic candidate for acute SCI.

Microarray expression profile and analysis of circular RNA regulatory network in malignant pleural effusion.

Malignant pleural effusion (MPE) is a common complication of lung cancer. Accumulating evidence has suggested that circular RNAs (circRNAs) play important roles in oncogenesis and progression of cancer. However, the expression pattern of circRNAs in MPE remains largely unknown and awaits investigation. The study was designed to elucidate the potential roles of differentially expressed circRNAs in MPE. Herein, we detected a total of 1350 differentially expressed circRNAs and 1727 differentially expressed mRNAs in lung adenocarcinoma-associated malignant pleural effusion (LA-MPE) compared with tuberculous pleural effusion (TPE) by Clariom D Human Microarray. Among the top 5 up-regulated circRNAs (hsa_circ_0067705, hsa_circ_0025542, hsa_circ_0072793, hsa_circ_0084927, and hsa_circ_0085386), four were verified significantly up-regulated in LA-MPE by qRT-PCR and hsa_circ_0085386 had an increasing trend. CircRNA-miRNA-mRNA network for the top 5 up-regulated circRNAs was constructed and pathway analysis indicated that the enriched mRNA targets involved in PI3K-Akt signaling pathway, Axon guidance, Regulation of actin cytoskeleton and Rap1 signaling pathway were potentially regulated by these aberrantly expressed circRNAs. We generated specific circRNA profiles in LA-MPE for the first time. And analysis of circRNA regulatory network could provide evidence that circRNAs are important in MPE development because they participate in cancer-related pathways by sequestering miRNAs. Our findings suggested that aberrantly expressed circRNAs may be involved in the development of LA-MPE.

Molecular and Bioinformatics Analyses Identify 7 Circular RNAs Involved in Regulation of Oncogenic Transformation and Cell Proliferation in Human Bladder Cancer.

BACKGROUND Circular RNAs (circRNAs) have emerged as important regulators in carcinogenesis and metastasis. However, the knowledge of circRNAs in bladder cancer remains limited. This study aimed to investigate the role and mechanism of circRNAs in the development and progression of bladder cancer. MATERIAL AND METHODS Three pairs of bladder carcinomas (including high- and low-grade tumors) and adjacent normal tissues were collected from patients. The total RNAs were extracted from these samples and subjected to Clariom D microarray assays to detect the differentially expressed circRNAs and mRNAs. The mRNA targets for these circRNAs were predicted by miRanda in combination with stringent differential mRNA filters. The interaction network for the circRNA-mRNA pairs was generated by Cytoscape. RESULTS Among the 1038 circRNAs detected by the Clariom D microarray assay, we identified 7 significantly differentially expressed circRNAs in the tumors (fold change >2, FDR <0.05). Principal component analysis of the differential circRNAs confirmed that the tumor samples were separated from the normal samples. Hierarchical clustering analyses on these RNAs and their predicted mRNA targets showed that the majority of differentially expressed circRNAs and mRNAs had been up-regulated in the bladder tumors. KEGG signaling pathway analysis has indicated that genes involved in cell proliferation, oncogenic transformation, and metastasis are potentially regulated by these circRNAs. CONCLUSIONS The current study provides a molecular basis for further investigating the mechanisms by which circRNAs regulate bladder cancer. The clinical significance of the identified circRNAs is highlighted by their potentials as diagnostic and prognostic biomarkers for bladder cancer patients.

Remote Ischemic Preconditioning-Mediated Neuroprotection against Stroke is Associated with Significant Alterations in Peripheral Immune Responses.

AIMS: Remote ischemic preconditioning (RIPC) of a limb is a clinically feasible strategy to protect against ischemia-reperfusion injury after stroke. However, the mechanism underlying RIPC remains elusive. METHODS: We generated a rat model of noninvasive RIPC by four repeated cycles of brief blood flow constriction (5 min) in the hindlimbs using a tourniquet. Blood was collected 1 h after preconditioning and 3 days after brain reperfusion. The impact of RIPC on immune cell and cytokine profiles prior to and after transient middle cerebral artery occlusion (MCAO) was assessed. RESULTS: Remote ischemic preconditioning protects against focal ischemia and preserves neurological functions 3 days after stroke. Flow cytometry analysis demonstrated that RIPC ameliorates the post-MCAO reduction of CD3(+)CD8(+) T cells and abolishes the reduction of CD3(+)/CD161a(+) NKT cells in the blood. In addition, RIPC robustly elevates the percentage of B cells in peripheral blood, thereby reversing the reduction in the B-cell population after stroke. RIPC also markedly elevates the percentage of CD43(+)/CD172a(+) noninflammatory resident monocytes, without any impact on the percentage of CD43(-)/CD172a(+) inflammatory monocytes. Finally, RIPC induces IL-6 expression and enhances the elevation of TNF-α after stroke. CONCLUSION: Our results reveal dramatic immune changes during RIPC-afforded neuroprotection against cerebral ischemia.

Separation of peptides with an aqueous mobile phase by temperature-responsive chromatographic column.

Peptide separation technology is significant and is still an analytical challenge in proteomic studies. We report a simple preparation method for poly(N-isopropylacrylamide) grafted silica through the copolymerization of N-isopropylacrylamide with acetyl moieties immobilized on the silica surfaces. Differential scanning calorimetry results indicated that the prepared silica exhibited a sharp phase transition at 35.03°C. Silica grafted with poly(N-isopropylacrylamide) was evaluated as a temperature-responsive chromatography medium for the separation of peptides using 0.2 M NaCl solution as a mobile phase. Results indicated that at 10°C, the peptides were not resolved, but baseline separation with prolonged retention time at 50°C was attained. Particularly, a mixture of four peptides was efficiently separated within 8 min. The theoretical plate number of every peptide was more than 2500, and the resolutions were more than 3.40. The increased selectivity of the temperature-responsive column resulted from the temperature-modulated hydrophobic interaction with peptides. The retention times of these peptides were related to their hydrophobicities. This protocol provided a reliable set of chromatographic tool usable across all research and development applications that required isolation and analysis of peptides. It may represent a step forward in the complex analysis of hydrophobic and other proteins.