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The aims of this study were to distinguish between behavioral compensation and behavioral recovery and to determine the role of stroke lesions and the optimal timing of rehabilitation in true recovery.Single pellet reaching test has been performed to analyze both quantitative and qualitative measures of forelimb function in a stroke animal model with lesions in the motor cortex, somatosensory cortex, or sensorimotor cortex. The four gestures of compensatory movement patterns that comprised a reach were head lift, limb withdrawal, pellet chasing, and phantom grasp.Functional recovery improved in all the stroke groups after rehabilitation ( P0.001). However, the compensatory movement patterns of the motor cortex and somatosensory cortex stroke groups initially increased and subsequently decreased ( P = 0.0054), whereas those of the sensorimotor cortex stroke group increased and persisted ( P = 0.0063). In the sensorimotor cortex stroke group, compensatory movement patterns significantly decreased when training was initiated 5 and 14 days after stroke ( P = 0.0083, P = 0.0226, respectively), while they increased and persisted when training was initiated 1 day after stroke.These findings suggest that true recovery by task-specific training after stroke depends, probably, on the lesion size and the timing of rehabilitation.

Inflammation, oxidative stress, and apoptosis are thought to be important causes of chronic obstructive pulmonary disease (COPD). We investigated the effect of YPL-001 (under phase 2a study, ClinicalTrials.gov identifier NCT02272634), a drug derived from Pseudolysimachion rotundum var. subintegrum, on cigarette smoke extract (CSE)-induced inflammation, the anti-oxidative pathway, and apoptosis in human lung epithelial cells and on CSE-induced emphysema in mice. YPL-001 suppressed CSE-induced expression of IL8 mRNA and protein. This was due to the reduction in NF-κB transcriptional activity by YPL-001, which resulted from the blockade of acetylation of the NF-κB subunit p65 (Lys310). Histone deacetylases (HDACs) prevent gene transcription by condensing the DNA structure and affecting NF-κB nuclear binding. YPL-001 alone increased HDAC2 activity and enhanced CSE-induced activation of HDAC2. YPL-001-induced suppression of NF-κB transcriptional activity might be caused by increased HDAC2 activity. YPL-001 increased nuclear factor (erythroid-derived 2)-like 2 (Nrf2) expression via both degradation of its inhibitory protein, Kelch-like ECH-associated protein 1, and an increase in de novo protein synthesis. YPL-001 increased the DNA binding activity of Nrf2. Consequently, YPL-001 upregulated the expression of Nrf2-targeted anti-oxidant genes such as NAD(P)H quinone dehydrogenase 1 and heme oxygenase 1. Moreover, YPL-001 significantly suppressed CSE-induced apoptotic cell death. In vivo study showed that CSE-induced emphysematous changes, neutrophilic inflammation, protein leakage into bronchoalveolar space, and lung cell apoptosis in mice were suppressed by YPL-001 treatment. Taken together, these results suggest that YPL-001 is a good therapeutic candidate for the treatment of COPD by blocking inflammation and apoptosis and activating the anti-oxidative pathway.

Cereblon (CRBN) has been shown to play an essential role in regulating inflammatory response and endoplasmic reticulum stress, thus mediating the development of various diseases. However, little is known about the roles of CRBN in chronic obstructive pulmonary disease (COPD) pathogenesis. We found that the protein levels of CRBN in lung homogenates from patients with COPD were lower than those from never smokers and smokers. The CRBN protein level was positively correlated with the forced expiratory volume in 1 s (FEV1)/forced vital capacity (FVC). To investigate the role of CRBN in modulating elastase-induced emphysema, we used Crbn knockout (KO) mice. Elastase-induced emphysematous changes were significantly aggravated in Crbn KO mice. Neutrophil infiltration, lung cell injury, and protein leakage into the bronchoalveolar space were more severe in Crbn KO mice than in wild-type (WT) mice. Furthermore, Crbn KO resulted in the elevated release of neutrophilic chemokines and inflammatory cytokines in lung epithelial cells and macrophages. The transcriptional activity of nuclear factor-κB (NF-κB) was significantly increased in Crbn knocked-down cells. In conclusion, Crbn deficiency might be involved in the development of emphysema by enhancing NF-κB activation, suggesting that targeting CRBN might be an effective therapeutic approach for the treatment of COPD.

Background: Lung epithelial cells serve as first line of defense against various inhaled pollutant particles. This study focused on assessing the impact of organic extracts of PM2.5 collected in Seoul, South Korea on primary human lung epithelial cells and identifying the relevant components and sources which induced lung epithelial cell injury.Results: We used primary lung epithelial cells isolated directly from healthy donors and evaluated the effects of organic compounds of twelve selected, seasonal PM2.5 on inflammation, cellular aging, and macroautophagy in primary lung epithelial cells. Organic extracts of PM2.5 specifically induced neutrophilic chemokine, interleukin-8, via extracellular signal regulated kinase activation. Moreover, PM2.5 significantly increased the expression of aging markers (p16, p21, and p27) and activated macroautophagy. Average mass concentrations, OC and EC, had no significant correlations with PM2.5 effects. However, regression analysis showed that polycyclic aromatic hydrocarbons and n-alkanes were the most relevant components of PM2.5 correlated with neutrophilic inflammation. Vegetative detritus and residential bituminous coal combustion sources were strongly correlated with neutrophilic inflammation, aging and macroautophagy activation.Conclusions: These data suggest that the chemical composition of PM2.5 is crucially important to determine the adverse health effects of PM2.5. Our study provides encouraging evidence to regulate the harmful components of PM2.5 in Seoul.

Inflammation, oxidative stress, and protease–antiprotease imbalance have been suggested to be a pathogenic triad in chronic obstructive pulmonary disease (COPD). However, it is not clear how proteases interact with components of inflammatory pathways. Therefore, this study aimed to evaluate the effect of neutrophil elastase (NE) on lipopolysaccharide (LPS)-induced interleukin 8 (IL-8) production and determine the molecular mechanism in human bronchial epithelial cells (HBECs). Immortalized bronchial epithelial cells and primary HBECs were used to investigate the impact of NE on LPS-induced IL-8 production. The molecular mechanism by which NE modulated LPS-induced IL-8 production was confirmed in elastase-treated C57BL/6 mice and primary HBECs obtained from COPD patients and healthy controls. The results showed that NE treatment synergistically augmented LPS-induced IL-8 production in both immortalized bronchial epithelial cells and primary HBECs. NE partially degraded peroxisome proliferator-activated receptor gamma (PPARγ), which is known to regulate IL-8 production in the nucleus. Treatment with a PPARγ agonist and overexpression of PPARγ reversed the NE-induced synergistic increase in LPS-induced IL-8 production. Moreover, PPARγ levels were lower in lung homogenates and lung epithelial cells from elastase-treated mice than in those from saline-treated mice. In accordance with the findings in mice, PPARγ levels were lower in primary HBECs from COPD patients than in those from healthy never-smokers or healthy smokers. In conclusion, a vicious cycle of mutual augmentation of protease activity and inflammation resulting from PPARγ degradation plays a role in the pathogenesis of COPD., Lung disease: getting to the root of inflammation Tracing how a protein-degrading enzyme called neutrophil elastase (NE) interacts with cytokines, molecules that trigger inflammation, has revealed a potential new treatment target for chronic obstructive pulmonary disease (COPD). COPD is a progressive disease in which chronic inflammation damages lung function. Although it was known that cytokines play key roles in COPD, the detailed molecular mechanism was not well understood. Chul-Gyu Yoo at Seoul National University College of Medicine in South Korea and co-workers investigated the role that NE plays in inflammation in COPD. They found that NE degraded peroxisome proliferator-activated receptor gamma (PPARγ), a transcription factor that keeps cytokine production in check. Patients with COPD showed low levels of PPARγ, and boosting PPARγ or suppressing NE in cultured cells showed potential for reducing inflammation. These results highlight PPARγ as a candidate for development of COPD treatments.

Pulmonary fibrosis is a progressive and lethal lung disease characterized by the proliferation and differentiation of lung fibroblasts and the accumulation of extracellular matrices. Since pulmonary fibrosis was reported to be associated with adenosine monophosphate-activated protein kinase (AMPK) activation, which is negatively regulated by cereblon (CRBN), we aimed to determine whether CRBN is involved in the development of pulmonary fibrosis. Therefore, we evaluated the role of CRBN in bleomycin (BLM)-induced pulmonary fibrosis in mice and in transforming growth factor-beta 1 (TGF-β1)-induced differentiation of human lung fibroblasts. BLM-induced fibrosis and the mRNA expression of collagen and fibronectin were increased in the lung tissues of wild-type (WT) mice; however, they were significantly suppressed in Crbn knockout (KO) mice. While the concentrations of TGF-β1/2 in bronchoalveolar lavage fluid were increased via BLM treatment, they were similar between BLM-treated WT and Crbn KO mice. Knockdown of CRBN suppressed TGF-β1-induced activation of small mothers against decapentaplegic 3 (SMAD3), and overexpression of CRBN increased it. TGF-β1-induced activation of SMAD3 increased α-smooth muscle actin (α-SMA) and collagen levels. CRBN was found to be colocalized with AMPKα1 in lung fibroblasts. CRBN overexpression inactivated AMPKα1. When cells were treated with metformin (an AMPK activator), the CRBN-induced activation of SMAD3 and upregulation of α-SMA and collagen expression were significantly suppressed, suggesting that increased TGF-β1-induced activation of SMAD3 via CRBN overexpression is associated with AMPKα1 inactivation. Taken together, these data suggest that CRBN is a profibrotic regulator and maybe a potential target for treating lung fibrosis., Lung disease: Putting the brakes on fibrosis Interventions that target a regulatory protein called cereblon could help reduce the damage inflicted on the lungs by idiopathic pulmonary fibrosis (IPF). This incurable and generally fatal condition is associated with the accumulation of scar tissue in the lungs, which leads to the gradual loss of respiratory function. Researchers led by Kyoung-Hee Lee at Seoul National University Hospital in South Korea have now identified cereblon as a potentially important contributor to this scarring process. They found that cereblon regulates a complex metabolic pathway that ultimately contributes to production of fibrosis-related proteins in a mouse model of IPF. Genetically modified animals that lacked the gene encoding cereblon showed reduced accumulation of these proteins in their lungs. These results suggest that cereblon-inhibiting agents could potentially control the progression of IPF and help preserve lung function.

Herein, cluster analysis was applied to evaluate the spatiotemporal variations in water quality variables of a river. The analysis was performed using the data obtained from 15 monitoring stations during 2007-2018 in the Yeongsan River, Republic of Korea. The spatiotemporal analysis successfully clustered the annual water quality variables temporally into years of poor water quality (2007-2012) and good water quality (2013-2018), and spatially into stations observing bad water quality (midstream) and good water quality (upstream and downstream). For the spatial cluster analysis results before and after a large river engineering project, the water quality was grouped into four clusters according to regional effects and water pollutant sources. The clustering analysis results clearly reflected changes in the water quality along the river due to the project. Overall, this study demonstrates that cluster analysis can be effectively used for evaluating spatiotemporal variations in river water quality.

Yun-Jeong Jeong,1,* Kyoung-Hee Lee,2,* Jisu Woo,2 Ji Yeon Kim,2 Chang-Hoon Lee,2,3 Chul-Gyu Yoo2,3 1Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, Dongguk University Ilsan Hospital, Ilsan Dong-gu, Goyang-si, Gyeonggi-do, Korea; 2Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Korea; 3Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Korea*These authors contributed equally to this workCorrespondence: Chul-Gyu Yoo, 101 Daehakno, Jongno-gu, Seoul, 03080, Korea, Tel +82-2-2072-3760, Fax +82-2-762-9662, Email cgyoo@snu.ac.krBackground: Macroautophagy plays an important role in the pathogenesis of chronic obstructive pulmonary disease (COPD), but the role of chaperone-mediated autophagy (CMA) has not been investigated. We investigated if and how CMA is involved in the pathogenesis of COPD.Methods: We measured the level of lysosome-associated membrane protein-2A (LAMP-2A), which is a critical component of CMA that functions as a receptor for cytosolic substrate proteins, in total lung tissues and primary human bronchial epithelial cells (HBECs) from healthy never smokers, smokers, and COPD patients. We assessed the effects of LAMP-2A knock-down on cigarette smoke extract (CSE)-induced aging, cell cycle arrest, and apoptosis in BEAS-2B cells and the expression levels of apoptosis hallmarks in primary HBECs and lung tissue sections.Results: We found that the protein levels of LAMP-2A in lung homogenates and primary HBECs from smokers and COPD patients were lower than those from never smokers. In addition, its level in primary HBECs was negatively correlated with years of smoking. CSE caused degradation of LAMP-2A protein via the lysosomal pathway by activating macroautophagy. Knock-down of LAMP-2A markedly enhanced CSE-induced expression of senescence markers such as p16, p21, p27, and p53. G2/M cell cycle arrest, up-regulation of cyclin B1, and apoptosis in BEAS-2B cells. Apoptosis was increased in CSE-treated primary HBECs and in lung tissues from smokers and COPD patients.Conclusion: Cigarette smoke-induced down-regulation of LAMP-2A is involved in acceleration of aging and apoptosis of lung epithelial cells, which might at least partially contribute to COPD pathogenesis.Keywords: COPD, LAMP-2A, lung epithelial cells, aging, apoptosis

This study examined the distribution of pharmaceuticals in Yeongsan River and at point sources (PSs) in the associated water system, and performed a risk assessment based on our findings. The samples included effluents collected from three sewage treatment plants (PS1, PS2, and PS3) and two industrial complexes (PS4 and PS5) as well as surface water collected from seven mainstreams and 11 tributaries of the river. The target pharmaceuticals were acetylsalicylic acid, carbamazepine, clarithromycin, naproxen, sulfamethazine, sulfamethoxazole, sulfathiazole, and trimethoprim, which were detected by liquid chromatography–tandem mass spectrometry. All pharmaceuticals except acetylsalicylic acid and sulfathiazole were found in PS1, PS2, and PS3 samples, whereas acetylsalicylic acid, carbamazepine, sulfamethazine, and sulfamethoxazole were found in PS4, most of the pharmaceuticals were not present in PS5. The rank order of pharmaceutical concentration in surface water was carbamazepine (97.2%, 0.2067 μg/L) > sulfamethoxazole (88.9%, 0.1132 μg/L) > naproxen (51.4%, 0.0516 μg/L) > clarithromycin (43.1%, 0.0427 μg/L). The distribution of pharmaceuticals in the Yeongsan River at PSs and non-PSs differed, and higher concentrations of human pharmaceuticals were detected in upstream and midstream areas whereas higher concentrations of animal pharmaceuticals were found downstream. Hazard quotients (HQs) evaluated at each sites based on mean concentration and 95% upper confidence limits (95% UCLs) were all less than one, indicating a low risk of toxicity. The findings of this study are expected to be useful for risk assessment of aquatic ecosystems.

Background Despite the high disease burden of chronic obstructive pulmonary disease (COPD) and risk of acute COPD exacerbation, few COPD biomarkers are available. As developmental endothelial locus-1 (DEL-1) has been proposed to possess beneficial effects, including anti-inflammatory effects, we hypothesized that DEL-1 could be a blood biomarker for COPD. Objective To elucidate the role of plasma DEL-1 as a biomarker of COPD in terms of pathogenesis and for predicting acute exacerbation. Methods Cigarette smoke extract (CSE) or saline was intratracheally administered to wild-type (WT) and DEL-1 knockout (KO) C57BL/6 mice. Subsequently, lung sections were obtained to quantify the degree of emphysema using the mean linear intercept (MLI). Additionally, plasma DEL-1 levels were compared between COPD and non-COPD participants recruited in ongoing prospective cohorts. Using negative binomial regression analysis, the association between the plasma DEL-1 level and subsequent acute exacerbation risk was evaluated in patients with COPD. Results In the in vivo study, DEL-1 KO induced emphysema (KO saline vs. WT saline; P = 0.003) and augmented CSE-induced emphysema (KO CSE vs. WT CSE; P P = 0.04), especially non-COPD never smokers (P = 0.019). During 1.2 ± 0.3 years, patients with COPD in the lowest quartile of Log(DEL-1) demonstrated an increased risk of subsequent acute exacerbation, compared with those in the highest quartile of Log(DEL-1) (adjusted incidence rate ratio, 3.64; 95% confidence interval, 1.03–12.9). Conclusion Low DEL-1 levels are associated with COPD development and increased risk of subsequent COPD acute exacerbation. DEL-1 can be a useful biomarker in patients with COPD.

Plant somatic cells can be reprogrammed into pluripotent cell mass, called callus, through a two-step in vitro tissue culture method. Incubation on callus-inducing medium triggers active cell proliferation to form a pluripotent callus. Notably, DNA methylation is implicated during callus formation, but a detailed molecular process regulated by DNA methylation remains to be fully elucidated. Here, we compared genome-wide DNA methylation profiles between leaf and callus tissues in Arabidopsis using whole-genome bisulphite-sequencing. Global distribution of DNA methylation showed that CHG methylation was increased, whereas CHH methylation was reduced especially around transposable element (TE) regions during the leaf-to-callus transition. We further analysed differentially expressed genes around differentially methylated TEs (DMTEs) during the leaf-to-callus transition and found that genes involved in cell cycle regulation were enriched and also constituted a coexpression gene network along with pluripotency regulators. In addition, a conserved DNA sequence analysis for upstream cis-elements led us to find a putative transcription factor associated with cell fate transition. CIRCADIAN CLOCK-ASSOCIATED 1 (CCA1) was newly identified as a regulator of plant regeneration, and consistently, the cca1lhy mutant displayed altered phenotypes in callus proliferation. Overall, these results suggest that DNA methylation coordinates cell cycle regulation during callus formation, and CCA1 may act as a key upstream coordinator at least in part in the processes.

Although Alzheimer&rsquo, s disease (AD)-like pathology is frequently found in patients with post-stroke dementia, little is known about the effects of aerobic exercise on the modifications of tau and related proteins. Therefore, we evaluated the effects of aerobic exercise on the phosphorylation and acetylation of tau and the expressions of tau-related proteins, after middle cerebral artery occlusion (MCAO) stroke. Twenty-four Sprague&ndash, Dawley rats with MCAO infarction were used in this study. The rehabilitation group (RG) received treadmill training 40 min/day for 12 weeks, whereas the sedentary group (SG) did not receive any type of training. Functional tests, such as the single pellet reaching task, rotarod, and radial arm maze tests, were performed monthly for 3 months. In ipsilateral cortices in the RG and SG groups, level of Ac-tau was lower in the RG, whereas levels of p-tauS396, p-tauS262, and p-tauS202/T205 were not significantly lower in the RG. Level of phosphorylated glycogen synthase kinase 3-beta Tyr 216 (p-GSK3&beta, Y216) was lower in the RG, but levels of p-AMPK and phosphorylated glycogen synthase kinase 3-beta Ser 9 (p-GSK3&beta, S9) were not significantly lower. Levels of COX-2 and BDNF were not significantly different between the two groups, while SIRT1 significantly decreased in ipsilateral cortices in RG. In addition, aerobic training also improved motor, balance, and memory functions. Rehabilitation with aerobic exercise inhibited tau modification, especially tau acetylation, following infarction in the rat MCAO model, which was accompanied with the improvement of motor and cognitive functions.

BACKGROUND Recent evidence indicates brain ischemia is associated with accumulations of abnormal tau and related proteins. However, the effects of aerobic training on these proteins have not been evaluated. OBJECTIVE We aimed to evaluate the effect of aerobic exercise on the phosphorylation and acetylation of tau and on the expressions of tau related proteins in a rat stroke model and to compare the effects of aerobic exercise with those observed in our previous study on task specific training (TST). METHODS Twenty-four Sprague- Dawley rats with photothrombotic cortical infarction were used in the current study. The rehabilitation group (RG) received treadmill training 40 min/day for 28 days, whereas the sedentary group (SG) did not receive any type of training. Functional tests such as the single pellet reaching task, rotarod, and radial arm maze tests were performed weekly for 4 weeks post-infarction. RESULTS Levels of p-taus396 and p-AMPK were found to be lower in ipsilateral cortices in the RG than in the SG (p

Although inflammation, oxidative stress, and protease-antiprotease imbalance have been referred to as a pathogenic triad in chronic obstructive pulmonary disease (COPD), little is known about how they interact. The objectives of this study were to elucidate the effect of cigarette smoke extract (CSE) on the neutrophil elastase (NE)-induced inflammatory response and its molecular mechanism in bronchial epithelial cells. We observed that NE activated extracellular signal-regulated kinase (ERK) and induced IL-8 production. Blocking ERK activation using a MEK inhibitor (U0126) suppressed NE-induced IL-8 secretion and knockdown of proteinase-activated receptor 2 (PAR2) using siRNAs inhibited both NE-induced ERK activation and subsequent IL-8 release, suggesting that NE-induced IL-8 production is dependent on PAR2-mediated ERK activation. Interestingly, pre-exposure to CSE markedly enhanced NE-induced IL-8 production. As PAR2 acts as a receptor for NE, we next investigated the effect of CSE on PAR2 expression as a molecular mechanism for the increased IL-8 production induced by NE in CSE exposed cells. CSE, but not NE, increased the expression of PAR2 mRNA and surface membrane protein. Inhibition of p38 MAPK reduced PAR2 expression induced by CSE while inhibition of the ERK and Akt pathway had no effect. Consequently, p38 inhibition significantly abrogated CSE-induced enhancement of IL-8 production in NE-treated cells. Of note, we observed increased PAR2 levels in lung homogenates and lung epithelial cells from CSE-treated mice and from both smokers and patients with COPD. Taken together, these results suggest that CSE upregulates PAR2 in normal human bronchial epithelial cells, thereby enhancing the inflammatory response to NE., Lung disease: How cigarette smoking enhances lung inflammation A study of the effect of cigarette smoke extract (CSE) on lung lining cells clarifies the interaction between CSE and an enzyme implicated in causing chronic obstructive pulmonary disease (COPD). The enzyme elastase is released by neutrophils and is known to induce inflammation, which causes lung irritation in COPD. Chul-Gyu Yoo and colleagues at Seoul National University in South Korea exposed cultured human lung cells to CSE. This exposure activated specific molecular signaling processes by which the elastase enzyme-induced inflammation was enhanced. Of particular note was an increase in the levels of a cell surface receptor protein in CSE-treated cells. This protein was increased in lungs of smokers and COPD patients. The research helps to understand the role of cigarette smoking in the pathogenesis of COPD.

A dual isotopes approach and the Bayesian isotope mixing model were applied to trace nitrogen pollution sources and to quantify their relative contribution to river water quality. We focused on two points to enhance the applicability of the method: 1) Direct measurement on the end-members to distinguish “sewage” and “manure” which used to be grouped in one pollution source as their isotope ranges overlap; 2) The Lagrangian sampling method was applied to consider the transport of nitrogen pollutants in a long river so that any fractionation process can be dealt with in the given Bayesian modeling framework. The results of the analysis confirmed the NO3− isotope composition in the river of interest to be within the range of NO3− with origins in “NH4+ in fertilizer”, “Soil N”, and “Manure and sewage” pollution. This suggests that nitrogen pollution is mostly attributed to anthropogenic sources. The δ18O NO3 value follows the range +2.5∼+15.0‰, implying that NO3− in the river is mainly derived from nitrification, and possible nitrification in groundwater or waterfront other than surface water. The ratio of the concentration of δ15N NO3 to that of δ18O NO3, and the corresponding regression equation indicates that the denitrification effect in surface water was insignificant during the study period. From the results of the contribution ratio of each source, improving the water quality of the discharge from the sewage treatment plants was proved to be the key factor to reduce nitrogen pollution in the river.

Patients with chronic obstructive pulmonary disease (COPD) are susceptible to infection owing to the impaired immune function of alveolar macrophages. This is presumed to be caused, at least partially, by cigarette smoke (CS), which is a major risk factor for COPD. Although CS has been reported to inhibit Toll-like receptor (TLR) function and phagocytosis in macrophages, the molecular mechanism of CS-mediated impairment of macrophage immune function has not been completely elucidated. We investigated the effects of CS extracts (CSE) on macrophage immune function and its molecular mechanism. We assessed lipopolysaccharide (LPS, TLR4 ligand)-, Pam3CSK4 (TLR2 ligand)-, or CpG-oligodeoxynucleotide (TLR9 ligand)-induced IL-6, TNF-α, and IL-1β production in macrophages. Upregulation of IL-6, TNF-α, and IL-1β mRNA and protein by TLR ligands was suppressed on treatment with CSE. However, LPS-induced MAP kinase activation, IκBα degradation, and nuclear translocation of NF-κB were not impeded by CSE. In contrast, CSE significantly suppressed NF-κB transcriptional activity in the nucleus. We found that p300, which acetylates RelA/p65 at lysine 310, and acetyl-p65 (K310) were downregulated upon CSE treatment. Knock-down of p300 suppressed LPS-induced acetylation of NF-κB p65 and production of inflammatory cytokine. To summarize, these results suggest that CSE impair cytokine response by decreasing the expression levels of p300.

Although stroke elicits progressive cognitive decline and is a leading cause of dementia, molecular interplay between stroke and Alzheimer's disease (AD) pathology has not been fully elucidated. Furthermore, studies on the effects of post-stroke rehabilitation on AD pathology are limited. We evaluated the acute effect of stroke on tau modification, and the molecular effects of task-specific training (TST) on tau modification using a model of photochemically-induced thrombosis (PIT)-induced cortical infarction. Following PIT in the dominant side of sensorimotor cortex, the rehabilitation group received 4-weeks of TST rehabilitation once daily by single pellet reaching training, whereas the sedentary control group did not received any type of training. Cortical expression levels of proteins related to tau modification were evaluated on post-stroke day 1 (PSD1) and 28; functional tests were also evaluated performed every week. The expression levels of acetyl-tau, phosphorylated-tau (p-tau), cyclooxygenase-2 and Akt-mTORC1-p70S6K pathway in infarcted cortices on PSD1 were significantly greater, whereas the expression levels of p-AMPK were significantly lower than in the paired contralateral sides. TST rehabilitation for 4 weeks greatly improved functional motor performance but not memory, which concurred with the down-regulations of ipsilateral p-AMPK, cyclooxygenase-2, Akt-mTORC1-p70S6K pathway, and p-tau in rehabilitation group. PIT-induced cortical infarction was found to induce cortical tau modification through the Akt-mTORC1-p70S6K activation, and to suppress the expression of AMPK-related proteins. TST rehabilitation greatly improved motor function, but not memory, and suppressed p-tau expression and neuroinflammation. Nevertheless, the role of TST-mediated regulation of tau hyperphosphorylation required further clarification.

An imbalance between oxidative stress and antioxidant activity plays an important role in the pathogenesis of chronic obstructive pulmonary disease (COPD). Cigarette smoke, a major risk factor of COPD, induces cellular oxidative stress, but levels of antioxidants such as heme oxygenase-1 (HO-1) are reduced in individuals with severe COPD. In this study, we evaluated the molecular mechanism of reduced HO-1 expression in human bronchial epithelial cells. We found that cigarette smoke extract (CSE) increases HO-1 levels via activation of NFE2-related factor 2 (Nrf2). However, pretreating cells with the protease neutrophil elastase (NE) suppressed the CSE-induced expression of HO-1 mRNA and protein. NE also decreased the sirtuin 1 (SIRT1) level, but did not inhibit CSE-induced nuclear translocation and DNA-binding activity of Nrf2. Transfection of cells with a Myc/His-tagged SIRT1 expression vector completely blocked the NE-mediated suppression of HO-1 expression. We further noted that the NE-induced down-regulation of SIRT1 was not due to decreased transcription or proteasomal/lysosomal degradation or loss of solubility. Immunofluorescence staining revealed that NE enters the cell cytoplasm, and we observed that NE directly cleaved SIRT1 in vitro, indicating that SIRT1 levels are decreased via direct degradation by internalized NE. Of note, we observed decreased SIRT1 levels in NE-treated primary human bronchial epithelial cells and in lung homogenates from both smokers and patients with COPD. In conclusion, NE suppresses CSE-induced HO-1 expression by cleaving SIRT1. This finding indicates the importance of cross-talk between oxidative stress and protease responses in the pathogenesis of COPD.

Background and objective Alveolar macrophages of patients with COPD display impaired cytokine release and diminished phagocytosis. COPD exacerbations exhibit immune dysfunction towards the respiratory pathogens. CS and CSE were reported to aggravate bacterial infections in COPD patients. Methods MARCO is highly expressed in lungs and is involved in pathogen clearance. We investigated the effect of CSE on MARCO expression and its regulatory mechanisms. After relevant siRNA transfection and treatment with CSE and/or LPS, we measured the levels of MARCO by q-RT PCR, immunoblotting and flow cytometry. Immunofluorescence staining and immunoprecipitation were used to evaluate the mechanism. Results CSE decreased LPS-induced expression of MARCO mRNA and protein. Upregulation of MARCO by LPS was Nrf2-dependent. Nrf2 knockdown significantly suppressed LPS-induced increase in MARCO transcripts. CSE did not block nuclear translocation of Nrf2 in LPS-treated cells, but rather CSE itself strongly accumulated Nrf2 in the nucleus through the degradation of its cytoplasmic inhibitor, KEAP1. However, CSE markedly suppressed LPS-induced Nrf2 acetylation. Histone acetyltransferase p300/CBP directly acetylates Nrf2, which augments promoter-specific DNA binding of Nrf2. Our results reveal CSE-induced polyubiquitinylation and subsequent degradation of p300 via the proteasome. Pretreatment with proteasome inhibitors completely blocked CSE-induced degradation of p300 and suppression of MARCO expression. Conclusion These findings suggest that CSE decreases MARCO expression via the proteasomal degradation of p300 in macrophages, which may be in part responsible for impaired bacterial phagocytosis.

Background Previous studies report that apoptosis and autophagy are involved in the pathogenesis of emphysema, and macroautophagy is one of the processes regulating the apoptosis pathway. However, few studies have evaluated whether chaperone-mediated autophagy (CMA) contributes to the regulation of apoptosis. In this study, we investigated the impact of autophagy, including both macroautophagy and CMA, on the apoptosis in bronchial epithelial cells. Methods Cigarette smoke extract (CSE) was injected intratracheally into C57BL/6 mice, and emphysema and apoptosis were evaluated in the lungs. After treatment with CSE, apoptosis, macroautophagy, and CMA were measured in BEAS2-B cells, and the impact of autophagy on the apoptosis was evaluated following knockdown of autophagy-related genes by short interfering RNAs (siRNAs). Results Intratracheal CSE injection resulted in the development of emphysema and an increase in apoptosis in mice. CSE increased the apoptosis in BEAS2-B cells, and also elevated the expression of proteins related to both macroautophagy and CMA in BEAS2-B cells. The knockdown experiment with siRNAs showed that macroautophagy increases apoptosis in BEAS2-B cells, while CMA suppresses apoptosis. Conclusion The intratracheal injection of CSE induces pulmonary emphysema and an increase in apoptosis in mice. CSE also induces apoptosis, macroautophagy, and CMA of bronchial epithelial cells. Macroautophagy and CMA regulate apoptosis in opposite directions.

Summary Brassinosteroids (BRs) regulate a variety of physiological processes in plants via extensive crosstalk with diverse biological signaling networks. Although BRs are known to reciprocally regulate circadian oscillation, the molecular mechanism underlying BR-mediated regulation of circadian clock remains unknown. Here, we demonstrate that the BR-activated transcription factor bri1-EMS-SUPPRESSOR 1 (BES1) integrates BR signaling into the circadian network in Arabidopsis. BES1 repressed expression of CIRCADIAN CLOCK-ASSOCIATED 1 (CCA1) and LATE ELONGATED HYPOCOTYL (LHY) at night by binding to their promoters, together with TOPLESS (TPL). The repression of CCA1 and LHY by BR treatment, which occurred during the night, was compromised in bes1-ko and tpl-8 mutants. Consistently, long-term treatment with BR shortened the circadian period, and BR-induced rhythmic shortening was impaired in bes1-ko and tpl-8 single mutants and in the cca1-1lhy-21 double mutant. Overall, BR signaling is conveyed to the circadian oscillator via the BES1/TPL-CCA1/LHY module, contributing to gating diurnal BR responses in plants., Graphical Abstract, Highlights • Circadian rhythm is shortened in the presence of phytohormone BR • BR signaling is relayed to circadian clock through BES1 • BES1/TPL complex binds to CCA1/LHY promoter in the dark, repressing expression • The BES1/TPL-CCA1/LHY module shapes circadian gating of BR signaling, Chronobiology; Plant Biology

IκB, a cytoplasmic inhibitor of nuclear factor-κB (NF-κB), is reportedly degraded via the proteasome. However, we recently found that long-term incubation with proteasome inhibitors (PIs) such as PS-341 or MG132 induces IκBα degradation via an alternative pathway, lysosome, which results in NF-κB activation and confers resistance to PI-induced lung cancer cell death. To enhance the anti-cancer efficacy of PIs, elucidation of the regulatory mechanism of PI-induced IκBα degradation is necessary. Here, we demonstrated that PI upregulates nuclear factor (erythroid-derived 2)-like 2 (Nrf2) via both de novo protein synthesis and Kelch-like ECH-associated protein 1 (KEAP1) degradation, which is responsible for IκBα degradation via macroautophagy activation. PIs increased the protein level of light chain 3B (LC3B, macroautophagy marker), but not lysosome-associated membrane protein 2a (Lamp2a, the receptor for chaperone-mediated autophagy) in NCI-H157 and A549 lung cancer cells. Pretreatment with macroautophagy inhibitor or knock-down of LC3B blocked PI-induced IκBα degradation. PIs up-regulated Nrf2 by increasing its transcription and mediating degradation of KEAP1 (cytoplasmic inhibitor of Nrf2). Overexpression of dominant-negative Nrf2, which lacks an N-terminal transactivating domain, or knock-down of Nrf2 suppressed PI-induced LC3B protein expression and subsequent IκBα degradation. Thus, blocking of the Nrf2 pathway enhanced PI-induced cell death. These findings suggest that Nrf2-driven induction of LC3B plays an essential role in PI-induced activation of the IκB/NF-κB pathway, which attenuates the anti-tumor efficacy of PIs.