Caveolin-1 identified as a key mediator of acute lung injury using bioinformatics and functional research

Acute lung injury (ALI) is a potentially life-threatening, devastating disease with an extremely high rate of mortality. The underlying mechanism of ALI is currently unclear. In this study, we aimed to confirm the hub genes associated with ALI and explore their functions and molecular mechanisms using bioinformatics methods. Five microarray datasets available in GEO were used to perform Robust Rank Aggregation (RRA) to identify differentially expressed genes (DEGs) and the key genes were identified via the protein-protein interaction (PPI) network. Lipopolysaccharide intraperitoneal injection was administered to establish an ALI model. Overall, 40 robust DEGs, which are mainly involved in the inflammatory response, protein catabolic process, and NF-κB signaling pathway were identified. Among these DEGs, we identified two genes associated with ALI, of which the CAV-1/NF-κB axis was significantly upregulated in ALI, and was identified as one of the most effective targets for ALI prevention. Subsequently, the expression of CAV-1 was knocked down using AAV-shCAV-1 or CAV-1-siRNA to study its effect on the pathogenesis of ALI in vivo and in vitro. The results of this study indicated that CAV-1/NF-κB axis levels were elevated in vivo and in vitro, accompanied by an increase in lung inflammation and autophagy. The knockdown of CAV-1 may improve ALI. Mechanistically, inflammation was reduced mainly by decreasing the expression levels of CD3 and F4/80, and activating autophagy by inhibiting AKT/mTOR and promoting the AMPK signaling pathway. Taken together, this study provides crucial evidence that CAV-1 knockdown inhibits the occurrence of ALI, suggesting that the CAV-1/NF-κB axis may be a promising therapeutic target for ALI treatment.Subject terms: Cell signalling, Respiratory tract diseases     

Age‐related memory vulnerability to interfering stimuli is caused by gradual loss of MAPK‐dependent protection in Drosophila

Age‐related memory impairment (AMI) is a common phenomenon across species. Vulnerability to interfering stimuli has been proposed to be an important cause of AMI. However, the molecular mechanisms underlying this vulnerability‐related AMI remain unknown. Here we show that learning‐activated MAPK signals are gradually lost with age, leading to vulnerability‐related AMI in Drosophila. Young flies (2‐ or 3‐day‐old) exhibited a significant increase in phosphorylated MAPK levels within 15 min after learning, whereas aged flies (25‐day‐old) did not. Compared to 3‐day‐old flies, significant 1 h memory impairments were observed in 15‐, 20‐, and 30‐day‐old flies, but not in 10‐day‐old flies. However, with post‐learning interfering stimuli such as cooling or electric stimuli, 10‐day‐old flies had worse memory performance at 1 h than 3‐day‐old flies, showing a premature AMI phenomenon. Increasing learning‐activated MAPK signals through acute transgene expression in mushroom body (MB) neurons restored physiological trace of 1 h memory in a pair of MB output neurons in aged flies. Decreasing such signals in young flies mimicked the impairment of 1 h memory trace in aged flies. Restoring learning‐activated MAPK signals in MB neurons in aged flies significantly suppressed AMI even with interfering stimuli. Thus, our data suggest that age‐related loss of learning‐activated neuronal MAPK signals causes memory vulnerability to interfering stimuli, thereby leading to AMI.     

Quantitative regulation of the thermal stability of enveloped virus vaccines by surface charge engineering to prevent the self-aggregation of attachment glycoproteins

The development of thermostable vaccines can relieve the bottleneck of existing vaccines caused by thermal instability and subsequent poor efficacy, which is one of the predominant reasons for the millions of deaths caused by vaccine-preventable diseases. Research into the mechanism of viral thermostability may provide strategies for developing thermostable vaccines. Using Newcastle disease virus (NDV) as model, we identified the negative surface charge of attachment glycoprotein as a novel determinant of viral thermostability. It prevented the temperature-induced aggregation of glycoprotein and subsequent detachment from virion surface. Then structural stability of virion surface was improved and virus could bind to and infect cells efficiently after heat-treatment. Employing the approach of surface charge engineering, thermal stability of NDV and influenza A virus (IAV) vaccines was successfully improved. The increase in the level of vaccine thermal stability was determined by the value-added in the negative surface charge of the attachment glycoprotein. The engineered live and inactivated vaccines could be used efficiently after storage at 37°C for at least 10 and 60 days, respectively. Thus, our results revealed a novel surface-charge-mediated link between HN protein and NDV thermostability, which could be used to design thermal stable NDV and IAV vaccines rationally.     

Design,synthesis, and biological evaluation of novel carbazole derivatives as potent DNMT1 inhibitors with reasonable PK properties

The DNA methyltransferases (DNMTs) were found in mammals to maintain DNA methylation. Among them, DNMT1 was the first identified, and it is an attractive target for tumour chemotherapy. DC_05 and DC_517 have been reported in our previous work, which is non-nucleoside DNMT1 inhibitor with low micromolar IC₅₀ values and significant selectivity towards other S-adenosyl-_L-methionine (SAM)-dependent protein methyltransferases. In this study, through a process of similarity-based analog searching, a series of DNMT1 inhibitors were designed, synthesized, and evaluated as anticancer agents. SAR studies were conducted based on enzymatic assays. And most of the compounds showed strong inhibitory activity on human DNMT1, especially WK-23 displayed a good inhibitory effect on human DNMT1 with an IC₅₀ value of 5.0 µM. Importantly, the pharmacokinetic (PK) profile of WK-23 was obtained with quite satisfying oral bioavailability and elimination half-life. Taken together, WK-23 is worth developing as DNMT1-selective therapy for the treatment of malignant tumour.     

Association between long-term exposure to ambient air pollution and COVID-19 severity: a prospective cohort study

Background:The tremendous global health burden related to COVID-19 means that identifying determinants of COVID-19 severity is important for prevention and intervention. We aimed to explore long-term exposure to ambient air pollution as a potential contributor to COVID-19 severity, given its known impact on the respiratory system.Methods:We used a cohort of all people with confirmed SARS-CoV-2 infection, aged 20 years and older and not residing in a long-term care facility in Ontario, Canada, during 2020. We evaluated the association between long-term exposure to fine particulate matter (PM_2.5), nitrogen dioxide (NO₂) and ground-level ozone (O₃), and risk of COVID-19-related hospital admission, intensive care unit (ICU) admission and death. We ascertained individuals’ long-term exposures to each air pollutant based on their residence from 2015 to 2019. We used logistic regression and adjusted for confounders and selection bias using various individual and contextual covariates obtained through data linkage.Results:Among the 151 105 people with confirmed SARS-CoV-2 infection in Ontario in 2020, we observed 8630 hospital admissions, 1912 ICU admissions and 2137 deaths related to COVID-19. For each interquartile range increase in exposure to PM_2.5 (1.70 μg/m³), we estimated odds ratios of 1.06 (95% confidence interval [CI] 1.01–1.12), 1.09 (95% CI 0.98–1.21) and 1.00 (95% CI 0.90–1.11) for hospital admission, ICU admission and death, respectively. Estimates were smaller for NO₂. We also estimated odds ratios of 1.15 (95% CI 1.06–1.23), 1.30 (95% CI 1.12–1.50) and 1.18 (95% CI 1.02–1.36) per interquartile range increase of 5.14 ppb in O₃ for hospital admission, ICU admission and death, respectively.Interpretation:Chronic exposure to air pollution may contribute to severe outcomes after SARS-CoV-2 infection, particularly exposure to O₃.

By November 2021, COVID-19 had caused more than 5 million deaths globally¹ and more than 29 400 in Canada.² The clinical manifestations of SARS-CoV-2 infection range from being asymptomatic to multiple organ failure and death. Identifying risk factors for COVID-19 severity is important to better understand etiological mechanisms and identify populations to prioritize for screening, vaccination and medical treatment. Risk factors for severity of COVID-19 include male sex, older age, pre-existing medical conditions and being from racialized communities.^3–5 More recently, ambient air pollution has been implicated as a potential driver of COVID-19 severity.^6–10Long-term exposure to ambient air pollution, a major contributor to global disease burden,¹¹ could increase the risk of severe COVID-19 outcomes by several mechanisms. Air pollutants can reduce individuals’ pulmonary immune responses and antimicrobial activities, boosting viral loads.⁸ Air pollution can also induce chronic inflammation and overexpression of the alveolar angiotensin-converting enzyme 2 (ACE) receptor,⁷ the key receptor that facilitates SARS-CoV-2 entry into cells.^12,13 Exposure to air pollution contributes to chronic conditions, such as cardiovascular disease, that are associated with unfavourable COVID-19 prognosis, possibly owing to persistent immune activation and excessive amplification of cytokine development.¹⁰ Thus, greater exposure to long-term air pollution may lead to severe COVID-19 outcomes.Reports exist of positive associations between long-term exposure to particulate matter with diameters equal to or smaller than 2.5 or 10 μm (PM_2.5 and PM₁₀), ground-level ozone (O₃) and nitrogen dioxide (NO₂), and metrics of COVID-19 severity (e.g., mortality and case fatality rate).^8–10 However, most studies to date have used ecological and cross-sectional designs, owing to limited access to individual data, which leads to ambiguity in interpreting the results, thus hindering their influence on policy. ^6,14 Ecological designs do not allow for disentangling the relative impacts of air pollution on individual susceptibility to infection and disease severity.¹⁴ Residual confounding by factors such as population mobility and social interactions is also problematic. Therefore, a cohort study with data on individuals with SARS-CoV-2 is a more appropriate design.^6,14 Studies that have used individual data were conducted in specific subpopulations^15,16 or populations with few severe cases,¹⁷ or had limited data on individual exposure to air pollutants.¹⁸ In Canada, 1 ecological study found a positive association between long-term exposure to PM_2.5 and COVID-19 incidence,¹⁹ but no published study has explored the association between air pollution and COVID-19 severity.We aimed to examine the associations between long-term exposure to 3 common air pollutants (PM_2.5, NO₂ and O₃) and key indicators of COVID-19 severity, including hospital admission, intensive care unit (ICU) admission and death, using a large prospective cohort of people with confirmed SARS-CoV-2 infection in Ontario, Canada, in 2020. The air contaminants PM_2.5, NO₂ and O₃ are regularly monitored by the Canadian government, and are key pollutants that are considered when setting air-quality policies. They originate from varying sources (NO₂ is primarily emitted during combustion of fuel, O₃ is primarily formed in air by chemical reactions of nitrogen oxides and volatile organic compounds, and PM_2.5 can be emitted during combustion or formed by reactions of chemicals like sulphur dioxide and nitrogen oxides in air) and they may affect human health differently.^20,21,22     

17β‐Oestradiol facilitates M2 macrophage skewing and ameliorates arrhythmias in ovariectomized female infarcted rats

Epidemiological studies have suggested a lower incidence of arrhythmia‐induced sudden cardiac death in women than in men. 17β‐oestradiol (E2) has been reported to have a post‐myocardial infarction antiarrhythmic effect, although the mechanisms have yet to be elucidated. We investigated whether E2‐mediated antioxidation regulates macrophage polarization and affects cardiac sympathetic reinnervation in rats after MI. Ovariectomized Wistar rats were randomly assigned to placebo pellets, E2 treatment, or E2 treatment +3‐morpholinosydnonimine (a peroxynitrite generator) and followed for 4 weeks. The infarct sizes were similar among the infarcted groups. At Day 3 after infarction, post‐infarction was associated with increased superoxide levels, which were inhibited by administering E2. E2 significantly increased myocardial IL‐10 levels and the percentage of regulatory M2 macrophages compared with the ovariectomized infarcted alone group as assessed by immunohistochemical staining, Western blot and RT‐PCR. Nerve growth factor colocalized with both M1 and M2 macrophages at the magnitude significantly higher in M1 compared with M2. At Day 28 after infarction, E2 was associated with attenuated myocardial norepinephrine levels and sympathetic hyperinnervation. These effects of E2 were functionally translated in inhibiting fatal arrhythmias. The beneficial effect of E2 on macrophage polarization and sympathetic hyperinnervation was abolished by 3‐morpholinosydnonimine. Our results indicated that E2 polarized macrophages into the M2 phenotype by inhibiting the superoxide pathway, leading to attenuated nerve growth factor‐induced sympathetic hyperinnervation after myocardial infarction.     

Hemin with Peroxidase Activity Can Inhibit the Oxidative Damage Induced by Ultraviolet A

Excessive reactive oxygen species (ROS), a highly reactive substance that contains oxygen, induced by ultraviolet A (UVA) cause oxidative damage to skin. We confirmed that hemin can catalyze the reaction of tyrosine (Tyr) and hydrogen peroxide (H₂O₂). Catalysis was found to effectively reduce or eliminate oxidative damage to cells induced by H₂O₂ or UVA. The scavenging effects of hemin for other free-radical ROS were also evaluated through pyrogallol autoxidation, 1,1-diphenyl-2-picrylhydrazyl radical (DPPH·)-scavenging assays, and phenanthroline–Fe²⁺ assays. The results show that a mixture of hemin and tyrosine exhibits strong scavenging activities for H₂O₂, superoxide anion (O₂⁻·), DPPH·, and the hydroxyl radical (·OH). Furthermore, the inhibition of oxidative damage to human skin keratinocyte (HaCaT) cells induced by H₂O₂ or UVA was evaluated. The results show that catalysis can significantly reduce the ratio of cell apoptosis and death and inhibit the release of lactate dehydrogenase (LDH), as well as accumulation of malondialdehyde (MDA). Furthermore, the resistance to apoptosis was found to be enhanced. These results show that the mixture of hemin and tyrosine has a significantly protective effect against oxidative damage to HaCaT cells caused by UVA, suggesting it as a protective agent for combating UVA damage.     

ATM phosphorylates histone H2AX in response to DNA double-strand breaks

A very early step in the response of mammalian cells to DNA double-strand breaks is the phosphorylation of histone H2AX at serine 139 at the sites of DNA damage. Although the phosphatidylinositol 3-kinases, DNA-PK (DNA-dependent protein kinase), ATM (ataxia telangiectasia mutated), and ATR (ATM and Rad3-related), have all been implicated in H2AX phosphorylation, the specific kinase involved has not yet been identified. To definitively identify the specific kinase(s) that phosphorylates H2AX in vivo, we have utilized DNA-PKcs-/- and Atm-/- cell lines and mouse embryonic fibroblasts. We find that H2AX phosphorylation and nuclear focus formation are normal in DNA-PKcs-/- cells and severely compromised in Atm-/- cells. We also find that ATM can phosphorylate H2AX in vitro and that ectopic expression of ATM in Atm-/- fibroblasts restores H2AX phosphorylation in vivo. The minimal H2AX phosphorylation in Atm-/- fibroblasts can be abolished by low concentrations of wortmannin suggesting that DNA-PK, rather than ATR, is responsible for low levels of H2AX phosphorylation in the absence of ATM. Our results clearly establish ATM as the major kinase involved in the phosphorylation of H2AX and suggest that ATM is one of the earliest kinases to be activated in the cellular response to double-strand breaks.     

The PRiMA-linked Cholinesterase Tetramers Are Assembled from Homodimers: HYBRID MOLECULES COMPOSED OF ACETYLCHOLINESTERASE AND BUTYRYLCHOLINESTERASE DIMERS ARE UP-REGULATED DURING DEVELOPMENT OF CHICKEN BRAIN*

Acetylcholinesterase (AChE) is anchored onto cell membranes by the transmembrane protein PRiMA (proline-rich membrane anchor) as a tetrameric globular form that is prominently expressed in vertebrate brain. In parallel, the PRiMA-linked tetrameric butyrylcholinesterase (BChE) is also found in the brain. A single type of AChE-BChE hybrid tetramer was formed in cell cultures by co-transfection of cDNAs encoding AChE_T and BChE_T with proline-rich attachment domain-containing proteins, PRiMA I, PRiMA II, or a fragment of ColQ having a C-terminal GPI addition signal (Q_N-GPI). Using AChE and BChE mutants, we showed that AChE-BChE hybrids linked with PRiMA or Q_N-GPI always consist of AChE_T and BChE_T homodimers. The dimer formation of AChE_T and BChE_T depends on the catalytic domains, and the assembly of tetramers with a proline-rich attachment domain-containing protein requires the presence of C-terminal “t-peptides” in cholinesterase subunits. Our results indicate that PRiMA- or ColQ-linked cholinesterase tetramers are assembled from AChE_T or BChE_T homodimers. Moreover, the PRiMA-linked AChE-BChE hybrids occur naturally in chicken brain, and their expression increases during development, suggesting that they might play a role in cholinergic neurotransmission.