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PDF file - 75KB, Supplemental Figure S1. Inactivation of NR4A1 induces ER stress and autophagy in vitro and in vivo.

PDF file - 36KB, Supplemental Table S2. Proteins showing differential expression in Panc1 cells by transfection of siNR4A1.

PDF file - 33KB, Supplemental Figure S2. Inactivation of NR4A1 induces DNA fragmentation and ER Ca2+ release in Panc-1 cells.

PDF file - 50KB, Supplemental Figure S5. Effect of siNR4A2 on induction of apoptosis and ER stress in pancreatic cancer cells.

PDF file - 20KB, Supplemental Table S1. List of real-time PCR primer sequences.

PDF file - 53KB, Supplemental Figure S4. DIM-C-pPhOH induces ER stress through inactivation of nuclear NR4A1.

PDF file - 125KB, Supplemental Table S3. mRNAs showing differential expression in Panc1 cells by transfection of siNR4A1.

PDF file - 37KB, Supplemental Figure S3. Top 10 significant enriched canonical pathways altered by transfection of Panc-1 cells with siNR4A1.

NR4A1 (Nur77, TR3) is an orphan nuclear receptor that is overexpressed in pancreatic cancer and exhibits pro-oncogenic activity. RNA interference of NR4A1 expression in Panc-1 cells induced apoptosis and subsequent proteomic analysis revealed the induction of several markers of endoplasmic reticulum stress, including glucose-related protein 78 (GRP78), CCAAT/enhancer-binding protein-homologous protein (CHOP), and activating transcription factor-4 (ATF-4). Treatment of pancreatic cancer cells with the NR4A1 antagonist 1,1-bis(3′-indolyl)-1-(p-hydroxyphenyl)methane (DIM-C-pPhOH) gave similar results. Moreover, both NR4A1 knockdown and DIM-C-pPhOH induced reactive oxygen species (ROS), and induction of ROS and endoplasmic reticulum stress by these agents was attenuated after cotreatment with antioxidants. Manipulation of NR4A1 expression coupled with gene expression profiling identified a number of ROS metabolism transcripts regulated by NR4A1. Knockdown of one of these transcripts, thioredoxin domain containing 5 (TXNDC5), recapitulated the elevated ROS and endoplasmic reticulum stress; thus, demonstrating that NR4A1 regulates levels of endoplasmic reticulum stress and ROS in pancreatic cancer cells to facilitate cell proliferation and survival. Finally, inactivation of NR4A1 by knockdown or DIM-C-pPhOH decreased TXNDC5, resulting in activation of the ROS/endoplasmic reticulum stress and proapoptotic pathways.Implications: The NR4A1 receptor is pro-oncogenic, regulates the ROS/endoplasmic reticulum stress pathways, and inactivation of the receptor represents a novel pathway for inducing cell death in pancreatic cancer. Mol Cancer Res; 12(4); 527–38. ©2014 AACR.

Supplementary Figures from Oncogenic transformation confers a selective susceptibility to the combined suppression of the proteasome and autophagy

The proteasome and the autophagy systems are two evolutionarily conserved mechanisms for degrading intracellular materials. They are functionally coupled and suppression of the proteasome promotes autophagy. Although suppression of the proteasome leads to cell death, suppression of autophagy can be either prodeath or prosurvival. To understand the underlining mechanism of this dichotomy and its potential clinical implications, we treated various transformed and nontransformed human cells with proteasome inhibitors. We found that whether autophagy served a prosurvival role in this scenario was contingent on the cellular oncogenic status. Thus, autophagy suppression enhanced apoptosis induced by proteasome inhibitors in transformed cells, but not in nontransformed cells. Oncogenic transformation enhanced the ability of cells to initiate autophagy in response to stress, reflecting a stronger dependence of transformed cells on autophagy for survival. Indeed, a combined use of bortezomib, the only Food and Drug Administration–approved proteasome inhibitor for clinical use, and chloroquine, which inhibits autophagy by disturbing lysosomal functions, suppressed tumor growth more significantly than either agent alone in a xenograft model. These findings indicate that suppression of both intracellular degradation systems could constitute a novel strategy for enhanced cancer control in a tumor-specific way. [Mol Cancer Ther 2009;8(7):2036–45]

In the current work we show that the profibrotic actions of TGF-β are mediated, at least in part, through a metabolic maladaptation in glutamine metabolism and how the inhibition of glutaminase 1 (GLS1) reverses pulmonary fibrosis. GLS1 was found to be highly expressed in fibrotic vs normal lung fibroblasts and the expression of profibrotic targets, cell migration, and soft agar colony formation stimulated by TGF-β required GLS1 activity. Moreover, knockdown of SMAD2 or SMAD3 as well as inhibition of PI3K, mTORC2, and PDGFR abrogated the induction of GLS1 by TGF-β. We further demonstrated that the NAD-dependent protein deacetylase, SIRT7, and the FOXO4 transcription factor acted as endogenous brakes for GLS1 expression, which are inhibited by TGF-β. Lastly, administration of the GLS1 inhibitor CB-839 attenuated bleomycin-induced pulmonary fibrosis. Our study points to an exciting and unexplored connection between epigenetic and transcriptional processes that regulate glutamine metabolism and fibrotic development in a TGF-β-dependent manner.

Female researchers may have experienced more difficulties than their male counterparts since the COVID-19 outbreak because of gendered housework and childcare. Using Microsoft Academic Graph data from 2016 to 2020, this study examined how the proportion of female authors in academic journals on a global scale changed in 2020 (net of recent yearly trends). We observed a decrease in research productivity for female researchers in 2020, mostly as first authors, followed by last author position. Female researchers were not necessarily excluded from but were marginalised in research. We also identified various factors that amplified the gender gap by dividing the authors' backgrounds into individual, organisational and national characteristics. Female researchers were more vulnerable when they were in their mid-career, affiliated to the least influential organisations, and more importantly from less gender-equal countries with higher mortality and restricted mobility as a result of COVID-19., 22 pages, 5 figures

This study explores how digital technologies can benefit older adults’ cognitive health. The need for cognition (NFC) refers to an individual’s tendency to engage in and enjoy cognitive endeavors and is an explanatory variable for behavioral outcomes. To date, little scholarly literature has examined how NFC can be affected by our behavior; therefore, this study explored whether practicing digital skills can help to improve or maintain NFC. Utilizing age as an instrumental variable, we analyzed data from 8,824 respondents (mean age = 49.43) to the 2015 Korea Media Panel survey and discovered that enhanced digital skills led to stronger NFC. This causal effect was stronger for the older cohort (baby boomers and older, who were born before 1963). These findings suggest that older adults’ NFC may not necessarily decline with age if they practice more digital skills. This study could ultimately contribute to the transformation of the older population from a digital minority to a digital beneficiary.

The increasing amount of information online makes it challenging to judge what to believe or discredit. Fact-checking unverified claims shared on platforms, like social media, can play a critical role in correcting misbeliefs. The current study demonstrates how the effect of fact-checking can vary by several factors. We show that fact-checking helps self-correct one's views among young adults. However, this effect is weaker for individuals who perceived the claim negatively at first. Furthermore, borderline messages like “Lack of Evidence” can be perceived as false rather than neutral. We explain these biases via human cognitive mechanisms that avoid risk and uncertainty.

Pathogenic fibrotic diseases, including Idiopathic Pulmonary Fibrosis (IPF), have some of the worst prognoses and affect millions of people worldwide. With unclear etiology and minimally effective therapies, two-thirds of IPF patients die within 2-5 years from this progressive interstitial lung disease. Transforming Growth Factor Beta (TGFβ) and Insulin-like Growth Factor-1 (IGF-1) are known to promote fibrosis; however, myofibroblast specific upregulation of IGF-1 in the initiation and progression of TGFβ-induced fibrogenesis and IPF have remained unexplored. To address this, the current study (1) documents the upregulation of IGF-1 via TGFβ in myofibroblasts and fibrotic lung tissue, as well as its correlation with decreased pulmonary function in advanced IPF; (2) identifies IGF-1’s C1 promoter as mediating the increase in IGF-1 transcription by TGFβ in pulmonary fibroblasts; (3) determines that SMAD2 and mTOR signaling are required for TGFβ-dependent Igf-1 expression in myofibroblasts; (4) demonstrates IGF-1R activation is essential to support TGFβ-driven profibrotic myofibroblast functions and excessive wound-healing; and (5) establishes the effectiveness of slowing the progression of murine lung fibrosis with the IGF-1R inhibitor OSI-906. These findings expand our knowledge of IGF-1’s role as a novel fibrotic-switch, bringing us one step closer to understanding the complex biological mechanisms responsible for fibrotic diseases and developing effective therapies.

전투적 여성주의를 표방한 온라인 커뮤니티 메갈리아의 정체성에 대해 온라인 여성주의에서부터 혐오사이트에 이르기까지 다양한 해석이 존재한다. 본 연구는 익명상황에서 형성되는 사회적 정체성의 분화에 주목하여 상이한 해석이 도출된 원인을 탐색한다. 글쓴이 ID 공개 여부로 구분되는 익명성 수준(완전익명, 부분익명)에 따라 게시물에 나타난 담론 내용의 차이를 파악하고 내부갈등의 전개 양상을 재구성하기 위해 2015년에서 2016년에 이르는 메갈리아의 생애에 걸친 전체게시물 162,893건을 분석하였다. 자연어처리 기법(토픽모델링, Word2Vec)을 활용한 분석결과에 따르면 전체 게시물에서 성차별과 연관있는 주제의 비중이 높았으며, 내부 갈등과 연관된 주제의 비중은 크지 않았다. 한편 완전익명 게시물에서는 메갈리아 방언을 이용한 분노 표출 토픽들이 배타적인 선호를 보였으며, 부분익명 게시물에서는 성차별 반대 캠페인 관련 토픽들이 배타적 선호를 보였다. 이러한 담론의 분화는 게시물의 대다수를 차지하는 완전익명 게시글에 주목하느냐, 혹은 주요 글쓴이에 주목하며 부분익명 게시글의 흐름을 파악하느냐에 따라 메갈리아의 정체성을 전혀 다르게 파악하게 한다. 더불어 익명성 수준에 상관없이 공유되는 주제 내에서도 정체성간 사용하는 단어가 배타적으로 나뉘거나 공유단어의 활용맥락이 달라지는 방식으로 갈등이 발전한다. 두 가지 방식이 모두 관찰된 성소수자 논쟁의 경우 구성원의 대거 이탈과 커뮤니티의 쇠락을 야기했다. 이러한 연구결과를 통해 온라인 커뮤니티에서 사용자들이 익명성 수준과 같은 구조적 여건에 반응하여 정체성을 형성하고 상호작용하는 방식과, 커뮤니티의 생애과정 전개를 통합적으로 이해하는 분석틀의 발전에 기여하길 기대한다.

Metabolic dysregulation in fibroblasts is implicated in the profibrotic actions of transforming growth factor-β (TGF-β). Here, we present evidence that hexokinase 2 (HK2) is important for mediating the fibroproliferative activity of TGF-β both in vitro and in vivo. Both Smad-dependent and Smad-independent TGF-β signaling induced HK2 accumulation in murine and human lung fibroblasts through induction of the transcription factor c-Myc. Knockdown of HK2 or pharmacological inhibition of HK2 activity with Lonidamine decreased TGF-β-stimulated fibrogenic processes, including profibrotic gene expression, cell migration, colony formation, and activation of the transcription factors YAP and TAZ, with no apparent effect on cellular viability. Fibroblasts from patients with idiopathic pulmonary fibrosis (IPF) exhibited an increased abundance of HK2. In a mouse model of bleomycin-induced lung fibrosis, Lonidamine reduced the expression of genes encoding profibrotic markers (collagenΙα1, EDA-fibronectin, α smooth muscle actin, and connective tissue growth factor) and stabilized or improved lung function as assessed by measurement of peripheral blood oxygenation. These findings provide evidence of how metabolic dysregulation through HK2 can be integrated within the context of profibrotic TGF-β signaling.

Transforming growth factor-beta (TGFβ) is an enigmatic protein with various roles in healthy tissue homeostasis/development as well as the development or progression of cancer, wound healing, fibrotic disorders, and immune modulation, to name a few. As TGFβ is causal to various fibroproliferative disorders featuring localized or systemic tissue/organ fibrosis as well as the activated stroma observed in various malignancies, characterizing the pathways and players mediating its action is fundamental. In the current study, we found that TGFβ induces the expression of the immunoinhibitory molecule Programed death-ligand 1 (PD-L1) in human and murine fibroblasts in a Smad2/3- and YAP/TAZ-dependent manner. Furthermore, PD-L1 knockdown decreased the TGFβ-dependent induction of extracellular matrix proteins, including collagen Iα1 (colIα1) and alpha-smooth muscle actin (α-SMA), and cell migration/wound healing. In addition to an endogenous role for PD-L1 in profibrotic TGFβ signaling, TGFβ stimulated-human lung fibroblast-derived PD-L1 into extracellular vesicles (EVs) capable of inhibiting T cell proliferation in response to T cell receptor stimulation and mediating fibroblast cell migration. These findings provide new insights and potential targets for a variety of fibrotic and malignant diseases.

This study investigated whether and how classroom-level bullies’ friendship networks, which reflect the popularity of bullies, were associated with the self-esteem of middle school students in the classroom. We analyzed survey data from 2,444 students in 101 classrooms from 22 middle schools in Korea. Respondents reported their friends’ and bullies’ names from their classroom. For each student, we generated the number of close friends that a respondent reported (outdegree) and the number of classmates who nominated him or her as a close friend (indegree). Bullies’ popularity was measured by the sum of all the bullies’ indegrees in a classroom. The findings showed that students in a classroom of bullies with larger popularity had a lower level of self-esteem. Moreover, bullies’ popularity was harmful to nonvictims’ self-esteem as well as victims’ self-esteem. This study suggests that even indirect exposure to bullying in a classroom can decrease the self-esteem of nonvictims by leading to negative emotions such as anger, anxiety, fear, and/or guilty feelings.

Chronic alcohol consumption causes hepatic steatosis, which is characterized by a considerable increase in free fatty acid (FFA) and triglyceride levels. To identify the possible proteins involved in the progression to alcoholic hepatosteatosis, we performed proteomic analysis on livers of mice exposed to alcohol. 2D-based proteomic analysis revealed that EtOH exposure in mice changed the expression of 43 proteins compared with that in mice fed a normal diet (ND). The most notable protein changes were proteins involved in Met metabolism and oxidative stress, most of which were significantly downregulated in alcohol-exposed animals. Although non-alcoholic fatty liver disease (NAFLD) and alcoholic liver disease (ALD) seem to share the same molecular processes, the difference between these conditions is still unclear. To address this question, we explored the features of alcoholic hepatosteatosis that were different compared with those of methionine and choline deficient (MCD) diet-induced mice with nonalcoholic liver damage. Although most of the differentially expressed proteins associated with ALD did not significantly differ from those of NAFLD, nine proteins showed considerably different patterns. Of these, ornithine aminotransferase, vitamin D binding protein, and phosphatidylethanolamine-binding protein were considerably upregulated in ALD mice, compared to that in NAFLD and ND mice. However, other proteins including inorganic pyrophosphatase were differentially regulated in MCD mice; however, they did not differ significantly between the alcoholic model and ND control mice. These results suggested that the identified proteins might be useful candidate markers to differentiate ALD from NAFLD. J. Cell. Biochem. 118: 1189-1200, 2017. © 2016 Wiley Periodicals, Inc.

TGF-β plays a central role in the pathogenesis of fibroproliferative disorders. Defining the exact underlying molecular basis is therefore critical for the development of viable therapeutic strategies. Here, we show that expression of the facilitative glucose transporter 1 (GLUT1) is induced by TGF-β in fibroblast lines and primary cells and is required for the profibrotic effects of TGF-β. In addition, enhanced GLUT1 expression is observed in fibrotic areas of lungs of both patients with idiopathic pulmonary fibrosis and mice that are subjected to a fibrosis-inducing bleomycin treatment. By using pharmacologic and genetic approaches, we demonstrate that up-regulation of GLUT1 occurs via the canonical Smad2/3 pathway and requires autocrine activation of the receptor tyrosine kinases, platelet-derived and epidermal growth factor receptors. Engagement of the common downstream effector PI3K subsequently triggers activation of the MEK and mammalian target of rapamycin complex 2, which cooperate in regulating GLUT1 expression. Of note, inhibition of GLUT1 activity and/or expression is shown to impair TGF-β-driven fibrogenic processes, including cell proliferation and production of profibrotic mediators. These findings provide new perspectives on the interrelation of metabolism and profibrotic TGF-β signaling and present opportunities for potential therapeutic intervention.-Andrianifahanana, M., Hernandez, D. M., Yin, X., Kang, J.-H., Jung, M.-Y., Wang, Y., Yi, E. S., Roden, A. C., Limper, A. H., Leof, E. B. Profibrotic up-regulation of glucose transporter 1 by TGF-β involves activation of MEK and mammalian target of rapamycin complex 2 pathways.

It is well known that marital status is significantly associated with mortality risk. Little is known, however, regarding whether and how the effects of marital status are moderated by one's own family structure in childhood. The purposes of this study are to examine whether marital status (i.e., family structure in adulthood) and living with both biological parents in childhood (i.e., family structure in childhood) are associated with mortality risk, and whether and how the effects of marital status vary depending on family structure in childhood and gender. We analyze the risk of death in five waves of the General Social Survey (GSS) from 1994 through 2002 after linking the GSS data to death certificate data from the National Death Index through 2008. The findings indicate that being widowed increases the risk of mortality, while living with both parents in childhood lowers it. Interestingly, analysis of the interaction between marital status and family structure in childhood reveals that the disadvantage of widowhood in terms of mortality is significantly stronger for those who lived with both parents in childhood than for those who did not. Subsample analysis by gender shows that the moderating effect of living with both parents is largely equal across men and women, though statistically more robust for men. These findings suggest that living with both parents during childhood may increase vulnerability to marital disruptions due to unwanted life events such as spousal loss. Childhood advantages, ironically, may form more stressful contexts of spousal loss by lowering one's adaptability or immunity to adulthood hardships, especially when the hardships in adulthood are characteristically opposite from the childhood advantages.

Immune checkpoint proteins such as programmed death-ligand 1 (PD-L1) retain general immune homeostasis and self-tolerance. PD-L1 binds to programmed cell death protein 1 (PD-1) and inhibits T cell activation or protects tumor cells from T cell-mediated killing. PD-L1 has recently been shown to be up-regulated in invasive lung fibroblasts by p53 and focal adhesion kinase (FAK), any potential role of transforming growth factor-beta (TGFβ) in PD-L1 action is unknown in fibrosis. TGFβ is a mysterious protein with various roles in healthy tissue homeostasis/development as well as the development or progression of cancer, fibrotic disorders, and immune modulation, et al. In the current study, we showed that TGFβ induces the expression of the immunoinhibitory molecule PD-L1 in human and murine fibroblasts in a Smad2/3- and YAP/TAZ-dependent manner. Furthermore, PD-L1 knockdown decreased the TGFβ-dependent induction of extracellular matrix proteins, including collagen Ia1 (colIa1) and alpha-smooth muscle actin (α-SMA), and cell migration/wound healing. In addition to an endogenous role for PD-L1 in profibrotic TGFβ signaling, TGFβ stimulated-human lung fibroblast-derived PD-L1 into extracellular vesicles (EVs) capable of inhibiting T-cell proliferation in response to T cell receptor stimulation and mediating fibroblast cell migration. These findings provide new insights and potential targets for a variety of fibrotic and malignant diseases.

We evaluated the clinical course and significance of postoperative subdural fluid collection (SFC) and identified the patients who were at risk of developing postoperative chronic subdural hematoma (CSDH) after the clipping of unruptured intracranial aneurysms (UIAs).Between January 2012 and June 2014, we retrospectively reviewed 298 patients with UIAs treated by microsurgical clipping. Among them, 257 patients were enrolled in the present study. Subdural lesions (SDLs) were defined as SFC at 1-month follow-up computed tomography (CT) and a CSDH at any time within 1 month after the clipping of UIAs. We examined the volume changes, Hounsfield unit (HU) values, and the end results of SFC in serial CT scans.The incidence of postoperative CSDH that needed burr hole surgery was 2.5%. Changes in SFC volume that occurred within 1 week of surgery were a risk factor for the occurrence of SDL at the 1-month follow-up CT (odds ratio 34.039; P0.001). The corrected average HU value of SCF (cut-off value: 11.9, with a sensitivity of 83.3% and specificity of 73.7%) on postoperative day 7 was an independent risk factor for development of a CSDH at the 1-month follow-up CT (odds ratio 19.261; P = 0.003).SDLs seen during 1-month follow-up may be associated with the occurrence of increased SFC volume within a week after the clipping of UIAs. The corrected average HU value of the SFC on postoperative day 7 was the only risk factor for the development of CSDHs at 1-month follow-up CT.

Evidence is provided that the fibroproliferative actions of TGF-β are dependent on a metabolic adaptation that sustains pathologic growth. Specifically, profibrotic TGF-β signaling is shown to require fatty acid synthase (FASN), an essential anabolic enzyme responsible for the de novo synthesis of fatty acids. With the use of pharmacologic and genetic approaches, we show that TGF-β-stimulated FASN expression is independent of Smad2/3 and is mediated via mammalian target of rapamycin complex 1. In the absence of FASN activity or protein, TGF-β-driven fibrogenic processes are reduced with no apparent toxicity. Furthermore, as increased FASN expression was also observed to correlate with the degree of lung fibrosis in bleomycin-treated mice, inhibition of FASN was examined in a murine-treatment model of pulmonary fibrosis. Remarkably, inhibition of FASN not only decreased expression of profibrotic targets, but lung function was also stabilized/improved, as assessed by peripheral blood oxygenation.-Jung, M.-Y., Kang, J.-H., Hernandez, D. M., Yin, X., Andrianifahanana, M., Wang, Y., Gonzalez-Guerrico, A., Limper, A. H., Lupu, R., Leof, E. B. Fatty acid synthase is required for profibrotic TGF-β signaling.

Sorting nexin 9 (SNX9) is shown to differentiate Smad3 from Smad2 nuclear delivery by mediating the association of phosphorylated Smad3 with importin 8 and the nuclear membrane. While the absence of SNX9 had negligible effects on transforming growth factor β receptor activity or Smad2 signaling, Smad3-dependent targets and phenotypes were inhibited., Transforming growth factor β (TGFβ) is a pleiotropic protein secreted from essentially all cell types and primary tissues. While TGFβ’s actions reflect the activity of a number of signaling networks, the primary mediator of TGFβ responses are the Smad proteins. Following receptor activation, these cytoplasmic proteins form hetero-oligomeric complexes that translocate to the nucleus and affect gene transcription. Here, through biological, biochemical, and immunofluorescence approaches, sorting nexin 9 (SNX9) is identified as being required for Smad3-dependent responses. SNX9 interacts with phosphorylated (p) Smad3 independent of Smad2 or Smad4 and promotes more rapid nuclear delivery than that observed independent of ligand. Although SNX9 does not bind nucleoporins Nup153 or Nup214 or some β importins (Imp7 or Impβ), it mediates the association of pSmad3 with Imp8 and the nuclear membrane. This facilitates nuclear translocation of pSmad3 but not SNX9.

Previously our research found that caffeic acid (CA) has antioxidant activity in vitro and in vivo. However, the antioxidant mechanism of CA has not been clearly demonstrated. We investigated the protective mechanism of CA on oxidative stress in HepG2 cells and primary hepatocytes. We focused on the effects of CA on glutathione (GSH) synthesis and its mechanisms. Reporter gene assay, transient transfection, quantitative reverse transcription-polymerase chain reaction and western blot were performed to confirm the mechanisms of antioxidant capacity. CA increased GSH level and γ-glutamylcysteine ligase (γ-GCL) activity. γ-Glutamylcysteine ligase catalytic subunit (GCLC) mRNA and protein levels were significantly increased following treatment with CA, whereas no changes were observed in the γ-GCL modifier subunit levels in HepG2 and rat hepatocytes. CA phosphorylated JNK, and activated the AP-1 transcription factor. These results show that the intracellular GSH levels and GCLC increased through the JNK/AP-1 pathways by CA treatment.