SARS-CoV-2 spike S1 subunit protein-mediated increase of beta-secretase 1 (BACE1) impairs human brain vessel cells.

Increasing evidence suggests incomplete recovery of COVID-19 patients, who continue to suffer from cardiovascular diseases, including cerebral vascular disorders (CVD) and neurological symptoms. Recent findings indicate that some of the damaging effects of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, especially in the brain, may be induced by the spike protein, leading to the disruption of the initial blood-brain barrier (BBB). SARS-CoV-2-infected cells and animals exhibit age-dependent pathogenesis. In this study, we identified endothelial BACE1 as a critical mediator of BBB disruption and cellular senescence induced by the SARS-CoV-2 spike S1 subunit protein. Increased BACE1 in human brain microvascular endothelial cells (HBMVEC) decreases the levels of tight junction proteins, including ZO-1, occludin, and claudins. Moreover, BACE1 overexpression leads to the accumulation of p16 and p21, typical hallmarks of cellular senescence. Our findings show that the SARS-CoV-2 spike S1 subunit protein upregulated BACE1 expression in HBMVECs, causing endothelial leakage. In addition, the SARS-CoV-2 spike S1 subunit protein induced p16 and p21 expression, indicating BACE1-mediated cellular senescence, confirmed by ß-Gal staining in HBMVECs. In conclusion, this study demonstrated that BACE1-mediated endothelial cell damage and senescence may be linked to CVD after COVID-19 infection.

Notch1-mediated inflammation is associated with endothelial dysfunction in human brain microvascular endothelial cells upon particulate matter exposure.

Exposure to atmospheric particulate matter (PM) is an emerging risk factor for the pathogenesis of several diseases in humans, including cerebrovascular diseases. However, the mechanisms underlying PM-induced endothelial dysfunction are currently unclear. In this study, we examined how PM leads to endothelial dysfunction in human brain microvascular endothelial cells (HBMECs). We demonstrated that PM10 exposure (up to 25 µg/mL) increase Notch1 cleavage, and it regulates endothelial dysfunction through NICD-mediated inflammation and senescence. PM10-induced NICD signaling causes increased expression of interleukin-1 beta (IL-1ß) and enhances characteristics of cellular senescence, which leads to increased endothelial permeability in HBMECs. Knockdown of Notch1 by siRNA blocks PM10-induced endothelial dysfunction via the suppression of inflammation and senescence. Furthermore, we found that Notch1-mediated inflammation accelerates endothelial senescence, which eventually leads to endothelial dysfunction. Altogether, our data suggest that Notch1 and NICD are potential target regulators for the prevention of cerebrovascular endothelial dysfunction induced by ambient air pollutants such as PM.

Altered COVID-19 receptor ACE2 expression in a higher risk group for cerebrovascular disease and ischemic stroke.

Coronavirus disease 2019 (COVID-19) is a worldwide pandemic. It has a high transmission rate among humans, and is a threat to global public health. However, there are no effective prophylactics or therapeutics available. It is necessary to identify vulnerable and susceptible groups for adequate protection and care against this disease. Recent studies have reported that COVID-19 has angiotensin-converting enzyme 2 (ACE2) as a functional receptor, which may lead to the development of severe cerebrovascular diseases (CVD), including strokes, in patients with risk factors for CVD such as diabetes and smoking. Thus, the World Health Organization (WHO) advised caution against COVID-19 for smokers and patients with underlying clinical symptoms, including cardiovascular diseases. Here, we observed ACE2 expression in the brain of rat middle cerebral artery occlusion (MCAO) model and evaluated the effects of cigarette smoke extract (CSE) and diabetes on ACE2 expression in vessels. We showed that the levels of ACE2 expression was increased in the cortex penumbra after ischemic injuries. CSE treatment significantly elevated ACE2 expression in human brain vessels. We found that ACE2 expression was upregulated in primary cultured human blood vessels with diabetes compared to healthy controls. This study demonstrates that ACE2 expression is increased in ischemic brains and vessels exposed to diabetes or smoking, makes them vulnerable to COVID-19 infection.

NDP52 associates with phosphorylated tau in brains of an Alzheimer disease mouse model.

We previously showed that NDP52 (also known as calcoco2) plays a role as an autophagic receptor for phosphorylated tau facilitating its clearance via autophagy. Here, we examined the expression and association of NDP52 with autophagy-regulated gene (ATG) proteins including LC3, as well as phosphorylated tau and amyloid-beta (Aß) in brains of an AD mouse model. NDP52 was expressed not only in neurons, but also in microglia and astrocytes. NDP52 co-localized with ATGs and phosphorylated tau as expected since it functions as an autophagy receptor for phosphorylated tau in brain. Compared to wild-type mice, the number of autophagic vesicles (AVs) containing NDP52 in both cortex and hippocampal regions was significantly greater in AD model mice. Moreover, the protein levels of NDP52 and phosphorylated tau together with LC3-II were also significantly increased in AD model mice, reflecting autophagy impairment in the AD mouse model. By contrast, a significant change in p62/SQSTM1 level was not observed in this AD mouse model. NDP52 was also associated with intracellular Aß, but not with the extracellular Aß of amyloid plaques. We conclude that NDP52 is a key autophagy receptor for phosphorylated tau in brain. Further our data provide clear evidence for autophagy impairment in brains of AD mouse model, and thus strategies that result in enhancement of autophagic flux in AD are likely to be beneficial.

Rottlerin Enhances the Autophagic Degradation of Phosphorylated Tau in Neuronal Cells.

Intra-neuronal accumulation of hyper-phosphorylated tau as neurofibrillary tangles (NFT) is a hallmark of Alzheimer's disease (AD). To prevent the aggregation of phosphorylated tau in neurons, decreasing the phosphorylated tau protein levels is important. Here, we examined the biological effects of rottlerin, a phytochemical compound extracted from the Kamala tree, Mallotus philippinensis, on phosphorylated tau levels. Notably, rottlerin decreased the levels of intracellular phosphorylated and total tau. A marked increase in the LC3-II, a hallmark of autophagy, was observed in these cells, indicating that rottlerin strongly induced autophagy. Interestingly, rottlerin induced the phosphorylation of Raptor at S792 through the activation of adenosine-monophosphate activated-protein kinase (AMPK), which likely inhibits the mammalian target of rapamycin complex 1 (mTORC1), thus resulting in the activation of transcription factor EB (TFEB), a master regulator of autophagy. In addition, nuclear factor erythroid 2-related factor 2 (Nrf2) activity increased in the presence of rottlerin. The decrease of phosphorylated tau levels in the presence of rottlerin was ameliorated by the knockdown of TFEB and partially attenuated by the knockout of the Nrf2 gene. Taken together, rottlerin likely enhances the degradation of phosphorylated tau through autophagy activated by TFEB and Nrf2. Thus, our results suggest that a natural compound rottlerin could be used as a preventive and therapeutic drug for AD.

Mitogen-activated protein kinase kinase 1 (MEK1) stabilizes MyoD through direct phosphorylation at tyrosine 156 during myogenic differentiation.

Previously, we reported that mitogen-activated protein kinase kinase 1 (MEK1) activated in the mid-stage of skeletal muscle differentiation promotes myogenic differentiation. To elucidate the molecular mechanism, we investigated an activity of MEK1 for MyoD. Activated MEK1 associates with MyoD in the nucleus of differentiating myoblasts. In vitro kinase assay using active MEK1, a (32)P-labeled protein band corresponding to GST-MyoD was observed but not to mutant GST-MyoD-Y156F. Tyrosine phosphorylation of endogenous MyoD was detected with a specific anti-pMyoD-Y156 antibody; however, this response was blocked by PD184352, a MEK-specific inhibitor. These results indicate that activated MEK1 phosphorylates the MyoD-Y156 residue directly. Interestingly, the protein level of mutant MyoD-Y156F decreased compared with that of wild type but was recovered in the presence of lactacystin, a proteasome inhibitor. The protein level of MyoD-Y156E, which mimics phosphorylation at Tyr-156, was above that of wild type, indicating that the phosphorylation protects MyoD from the ubiquitin proteasome-mediated degradation. In addition, the low protein level of MyoD-Y156F was recovered over that of wild type by an additional mutation at Leu-164, a critical binding residue of MAFbx/AT-1, a Skp, Cullin, F-box (SCF) E3-ubiquitin ligase. The amount of MyoD co-precipitated with MAFbx/AT-1 also was reduced in the presence of active MEK1. Thus, these results suggested that the phosphorylation probably interrupts the binding of MAFbx/AT-1 to MyoD and thereby increases its stability. Collectively, our results suggest that MEK1 activated in differentiating myoblasts stimulates muscle differentiation by phosphorylating MyoD-Y156, which results in MyoD stabilization.

Altered TIMP-3 Levels in the Cerebrospinal Fluid and Plasma of Patients with Alzheimer's Disease.

Tissue inhibitor of metalloproteinase-3 (TIMP-3) is a component of the extracellular environment and is suggested to play an indirect role in regulating Aß production and the pathophysiology of Aß deposition in brains. However, studies on the amount of TIMP-3 in bodily fluids of Alzheimer's disease (AD) patients have not been conducted. Here, we investigated the relationship between fluid TIMP-3 levels and AD pathology. We first showed that the fluid levels of TIMP-3 were lower in AD dementia patients compared with in non-AD patients. ELISA results revealed that plasma levels of TIMP-3 in 65 patients with AD were significantly lower than those in 115 healthy control subjects and 71 mild cognitive impairment (MCI) subjects. Furthermore, we found that cerebrospinal fluid (CSF) level of TIMP-3 was decreased in AD compared with that in healthy control. These data suggest that fluid TIMP-3 levels negatively correlated with progress of cognitive decline. Collectively, our study suggests that alterations of fluid TIMP-3 levels might be associated with AD pathology.

Identification of a pleiotropic effect of ADIPOQ on cardiac dysfunction and Alzheimer's disease based on genetic evidence and health care records.

Observations of comorbidity in heart diseases, including cardiac dysfunction (CD) are increasing, including and cognitive impairment, such as Alzheimer's disease and dementia (AD/D). This comorbidity might be due to a pleiotropic effect of genetic variants shared between CD and AD/D. Here, we validated comorbidity of CD and AD/D based on diagnostic records from millions of patients in Korea and the University of California, San Francisco Medical Center (odds ratio 11.5 [8.5-15.5, 95% Confidence Interval (CI)]). By integrating a comprehensive human disease-SNP association database (VARIMED, VARiants Informing MEDicine) and whole-exome sequencing of 50 brains from individuals with and without Alzheimer's disease (AD), we identified missense variants in coding regions including APOB, a known risk factor for CD and AD/D, which potentially have a pleiotropic role in both diseases. Of the identified variants, site-directed mutation of ADIPOQ (268 G > A; Gly90Ser) in neurons produced abnormal aggregation of tau proteins (p = 0.02), suggesting a functional impact for AD/D. The association of CD and ADIPOQ variants was confirmed based on domain deletion in cardiac cells. Using the UK Biobank including data from over 500000 individuals, we examined a pleiotropic effect of the ADIPOQ variant by comparing CD- and AD/D-associated phenotypic evidence, including cardiac hypertrophy and cognitive degeneration. These results indicate that convergence of health care records and genetic evidences may help to dissect the molecular underpinnings of heart disease and associated cognitive impairment, and could potentially serve a prognostic function. Validation of disease-disease associations through health care records and genomic evidence can determine whether health conditions share risk factors based on pleiotropy.

Curcumin activates Nrf2 through PKCδ-mediated p62 phosphorylation at Ser351.

Curcumin, a phytochemical extracted from Curcuma longa rhizomes, is known to be protective in neurons via activation of Nrf2, a master regulator of endogenous defense against oxidative stress in cells. However, the exact mechanism by which curcumin activates Nrf2 remains controversial. Here, we observed that curcumin induced the expression of genes downstream of Nrf2 such as HO-1, NQO1, and GST-mu1 in neuronal cells, and increased the level of Nrf2 protein. Notably, the level of p62 phosphorylation at S351 (S349 in human) was significantly increased in cells treated with curcumin. Additionally, curcumin-induced Nrf2 activation was abrogated in p62 knockout (-/-) MEFs, indicating that p62 phosphorylation at S351 played a crucial role in curcumin-induced Nrf2 activation. Among the kinases involved in p62 phosphorylation at S351, PKCδ was activated in curcumin-treated cells. The phosphorylation of p62 at S351 was enhanced by transfection of PKCδ expression plasmid; in contrast, it was inhibited in cells treated with PKCδ-specific siRNA. Together, these results suggest that PKCδ is mainly involved in curcumin-induced p62 phosphorylation and Nrf2 activation. Accordingly, we demonstrate for the first time that curcumin activates Nrf2 through PKCδ-mediated p62 phosphorylation at S351.

A splicing variant of TFEB negatively regulates the TFEB-autophagy pathway.

Transcription factor EB (TFEB) is a master regulator of the autophagy-lysosomal pathway (ALP). Here, we cloned a novel splicing variant of TFEB, comprising 281 amino acids (hereafter referred to as small TFEB), and lacking the helix-loop-helix (HLH) and leucine zipper (LZ) motifs present in the full-length TFEB (TFEB-L). The TFEB variant is widely expressed in several tissues, including the brain, although its expression level is considerably lower than that of TFEB-L. Intriguingly, in cells stably expressing small TFEB, the expression profile of genes was inverted compared to that in cells ectopically expressing TFEB-L. In addition, fisetin-induced luciferase activity of promoter containing either coordinated lysosomal expression and regulation (CLEAR) element or antioxidant response element (ARE) was significantly repressed by co-transfection with small TFEB. Moreover, fisetin-mediated clearance of phosphorylated tau or α-synuclein was attenuated in the presence of small TFEB. Taken together, the results suggest that small TFEB is a novel splicing variant of TFEB that might act as a negative regulator of TFEB-L, thus fine tuning the activity of ALP during cellular stress.

ApoE4 attenuates autophagy via FoxO3a repression in the brain.

Apolipoprotein E (ApoE) plays multiple roles in lipid transport, neuronal signaling, glucose metabolism, mitochondrial function, and inflammation in the brain. It is also associated with neurodegenerative diseases, and its influence differs depending on the isoform. In particular, the ε4 allele of APOE is the highest genetic risk factor for developing late-onset Alzheimer's disease (AD). However, the mechanism by which ApoE4 contributes to the pathogenesis of AD remains unclear. We investigated the effect of ApoE4 on autophagy in the human brains of ApoE4 carriers. Compared to non-carriers, the expression of FoxO3a regulating autophagy-related genes was significantly reduced in ApoE4 carriers, and the phosphorylation level of FoxO3a at Ser253 increased in ApoE4 carriers, indicating that FoxO3a is considerably repressed in ApoE4 carriers. As a result, the protein expression of FoxO3a downstream genes, such as Atg12, Beclin-1, BNIP3, and PINK1, was significantly decreased, likely leading to dysfunction of both autophagy and mitophagy in ApoE4 carriers. In addition, phosphorylated tau accumulated more in ApoE4 carriers than in non-carriers. Taken together, our results suggest that ApoE4 might attenuate autophagy via the repression of FoxO3a in AD pathogenesis. The regulation of the ApoE4-FoxO3a axis may provide a novel therapeutic target for the prevention and treatment of AD with the APOE4 allele.

Caspases-2 and -8 are involved in the presenilin1/gamma-secretase-dependent cleavage of amyloid precursor protein after the induction of apoptosis.

The presenilin/gamma-secretase protease cleaves many type-I membrane proteins, including the amyloid beta-protein (Abeta) precursor (APP). Previous studies have shown that apoptosis induces alterations in Abeta production in a caspase-dependent manner. Here, we report that staurosporine (STS)-induced apoptosis induces caspase-8 and/or-2-dependent gamma-secretase activation. Blocking of caspase activity with caspase-8 inhibitor z-IETD-fmk, and caspase-2 inhibitor z-VDVAD-fmk reduced Abeta production by STS in H4 cells expressing the Swedish mutant of APP (HSW) or APP-C99 (H4-C99). There was no inhibitory effect of other caspases (-1, -3, -5, -6, -9) on Abeta production by STS. This finding was further supported by evidence that siRNA transfection, depleting caspase-2 or -8 levels, lowered Abeta production in HSW and H4-C99 cells without affecting expression of APP or gamma-secretase complex. In addition, Abeta production by STS was decreased by JNK inhibitors, SP600125. These results suggest that caspase-2 and/or -8 is involved in presenilin/gamma-secretase activation and Abeta production in apoptosis.

Association between blood lead level and risk of stroke in Korean adults: a cross-sectional study in the Korea National Health and Nutrition Examination Survey 2008-2013.

OBJECTIVES: Although lead is a potential risk factor for cardiovascular diseases such as stroke, research on this association in the Korean population remains limited. Therefore, we aimed to investigate the association between lead level and stroke in Korean adults. DESIGN: A population-based cross-sectional study. SETTING: The Korea National Health and Nutrition Examination Survey 2008-2013, which enrolled a representative sample of the Korean population. PARTICIPANTS: We excluded participants younger than 20 years, missing weight data, pregnant or lactating, and missing blood lead and stroke data. A total of 11 510 participants were included in this analysis. PRIMARY AND SECONDARY OUTCOME MEASUREMENT: The participants were classified by blood lead concentration into the low-level (≤2.189 µg/dL, n=5756) and high-level (>2.189 µg/dL, n=5754) groups. The main outcome, stroke, was assessed by information from physician diagnosis, prevalence of stroke or treatment for stroke. The ORs and 95% CIs were calculated to evaluate the association between blood lead level and stroke using multivariate logistic regression analysis. RESULTS: Although blood lead level was not significantly associated with stroke (OR: 1.30, 95% CI: 0.66-2.58) in the multivariate-adjusted model, in individuals with hypertension, the high-level group was 2.36-fold higher odds of stroke (OR: 2.36, 95% CI: 1.02-5.44) compared to that in the low-level group. No association was observed in individuals with normotension (OR: 0.42, 95% CI: 0.13-1.38, p for interaction=0.007). CONCLUSION: The association between blood lead concentration and stroke may be influenced by hypertension status. Our findings suggest the need for closer attention to lead exposure in patients with hypertension.

Involvement of ADAM10 in acrolein-induced astrocytic inflammation.

Acrolein is a neurotoxin produced through lipid peroxidation in the brain affected by ischemic stroke, which results in neuronal cell injury and inflammation. However the mechanism underlying acrolein-induced brain inflammation remains unclear. Therefore we examined how acrolein leads to astrocytic inflammation. It was found that acrolein increased the levels of NLRP3 and cleaved caspase-1, which led to the maturation of interleukin-1ß (IL-1ß). ELISA assay results, which showed that acrolein increased the secreted IL-1ß, further supported acrolein-induced astrocytic inflammation. Acrolein increased ADAM10 protein levels and the cleavage of N-cadherin. The ADAM10 inhibitor, GI 254023X blocked N-cadherin cleavage by acrolein, suggesting that ADAM10 is an upstream of N-cadherin. Furthermore, we found that acrolein activated p38 MAPK and NF-κB p65, while pretreatment with p38 MAPK inhibitor, SB203580 and GI 254023X inhibited NF-κB p65 activation and NLRP3 inflammasome. This suggests that p38 MAPK mediates the activation of NF-κB p65, which is associated with NLRP3 expression. Finally, we showed that acrolein induced cell toxicity and decrease of EAAT1 expression, suggesting that acrolein may induce a loss of glutamate uptake function. In conclusion, we demonstrate that acrolein induces astrocytic inflammation through NLRP3 inflammasome, which is regulated by ADAM10 and attributed to p38 MAPK-activated NF-κB p65 activity.

Elevation of plasma soluble amyloid precursor protein beta in Alzheimer's disease.

INTRODUCTION: Beta-amyloid is considered to be a pathophysiological marker in Alzheimer's disease (AD). Soluble amyloid precursor proteins (sAPPs) -α (sAPPα) and -ß (sAPPß), which are the byproducts of non-amyloidogenic and amyloidogenic process of APP, respectively, have been repeatedly observed in the cerebrospinal fluids (CSF) of AD patients. The present study focused on the determination of sAPP levels in peripheral blood. METHODS: The plasma protein levels of sAPPα and sAPPß were measured with ELISA. Plasma from 52 AD patients, 98 amnestic mild cognitive impairment (MCI) patients, and 114 cognitively normal controls were compared. RESULTS: The plasma level of sAPPß was significantly increased in AD patients than in cognitively healthy controls. However, no significant change in plasma sAPPα was observed among the three groups. Furthermore, the plasma sAPPß levels significantly correlated with cognitive assessment scales, such as clinical dementia rating (CDR), and mini-mental status examination (MMSE). Interestingly, sAPPα and sAPPß had a positive correlation with each other in blood plasma, similar to previous studies on CSF sAPP. This correlation was stronger in the MCI and AD groups than in the cognitively healthy controls. CONCLUSIONS: These results suggest that individuals with elevated plasma sAPPß levels are at an increased risk of AD; elevation in these levels may reflect the progression of disease.

Elevated Cerebrospinal Fluid and Plasma N-Cadherin in Alzheimer Disease.

N-cadherin is a synaptic adhesion molecule stabilizing synaptic cell structure and function. Cleavage of N-cadherin by γ-secretase produces a C-terminal fragment, which is increased in the brains of Alzheimer disease (AD) patients. Here, we investigated the relationship between fluid N-cadherin levels and AD pathology. We first showed that the cleaved levels of N-cadherin were increased in homogenates of postmortem brain from AD patients compared with that in non-AD patients. We found that cleaved N-cadherin levels in the cerebrospinal fluid were increased in AD dementia compared with that in healthy control. ELISA results revealed that plasma levels of N-cadherin in 76 patients with AD were higher than those in 133 healthy control subjects. The N-cadherin levels in the brains of an AD mouse model, APP Swedish/PS1delE9 Tg (APP Tg) were reduced compared with that in control. The N-terminal fragment of N-cadherin produced by cleavage at a plasma membrane was detected extravascularly, accumulated in senile plaques in the cortex of an APP Tg mouse. In addition, N-cadherin plasma levels were increased in APP Tg mice. Collectively, our study suggests that alteration of N-cadherin levels might be associated with AD pathology.

Oncostatin M induces growth arrest of skeletal muscle cells in G1 phase by regulating cyclin D1 protein level.

Oncostatin M (OSM), an IL-6 family cytokine, either inhibits or enhances the growth of cells depending on cell type. Here, we report that OSM inhibits proliferation of skeletal muscle cells by blocking cell cycle progression from G(1) to S phase. OSM treatment significantly reduced levels of cyclin D1 protein and phosphorylation of retinoblastoma protein (Rb) at Ser-795, a CDK4-specific phosphorylation site. The OSM-induced cyclin D1 reduction correlated with decreased amount of the cyclin D1/p27 Kip1 complex and increased amounts of the CDK2/p27 Kip1 complex, resulting in inhibition of CDK2 activity. Results obtained with lactacystin, a proteasome inhibitor, demonstrated that cyclin D1 reduction occurred through ubiquitin/proteasome proteolysis. In addition, activation of STAT3, but not STAT1, is likely to regulate OSM-induced cyclin D1 reduction. Dominant negative (DN)-STAT3 blocked OSM-induced cyclin D1 reduction, and constitutively active-STAT3 also induced cyclin D1 reduction. These results suggest that OSM arrests skeletal muscle cell growth at the G1/S checkpoint and that this response occurs by an ubiquitin/proteasome-dependent cyclin D1 protein reduction which is regulated by STAT3.

Cigarette smoke extracts and cadmium induce COX-2 expression through γ-secretase-mediated p38 MAPK activation in C6 astroglia cells.

Exposure to cigarette smoke has been implicated in the progression of cerebrovascular and neurological disorders like stroke through inflammation and blood-brain barrier disruption. In this study, we investigated the signaling cascade activated by cigarette smoke extracts (CSE) and cadmium (Cd) resulting in the COX-2 induction in C6 rat astroglia cells. CSE or Cd induced Notch1 cleavage and activated p38 MAPK and CREB signaling pathways in C6 astroglia cells. Knockdown of nicastrin using siRNA or γ-secretase inhibitors, DAPT and L-685,486, reduced Notch1 cleavage and phosphorylation of p38 MAPK and CREB, while phosphorylation of ERK and JNK remained unaffected. Additionally, the blockage of γ-secretase activity did not show any effect on the phosphorylation of AKT, another upstream activator of CREB, indicating that γ-secretase-mediated CREB activation occurs via p38 MAPK. γ-secretase inhibitor also inhibited the CSE and Cd-mediated increase in the expression of COX-2. Furthermore, recombinant overexpression of Notch1 intracellular domain resulted in an increase in the expression of COX-2. Notch signaling induced by CSE and Cd induced apoptosis in C6 cells. Our results demonstrate that CSE exposure activated the p38 MAPK and CREB-mediated induction in COX-2 expression in astrocytes via γ-secretase-mediated Notch1 signaling. Our data provides novel insights into the potential mechanism of pro-inflammatory response activated by exposure to cigarette smoke.

TFEB activates Nrf2 by repressing its E3 ubiquitin ligase DCAF11 and promoting phosphorylation of p62.

Transcriptional factor EB (TFEB) and nuclear factor E2-related factor 2 (Nrf2) play crucial roles in the biological response against cellular stressors; however, their relationship has not yet been investigated. Here, we constructed human neuroglioma cell lines stably expressing TFEB. The expression of Nrf2-response genes, including heme oxygenase (HO)-1, glutathione-s-transferase-mu1 (GSTM1), and p62, was induced in the cell line, independent of oxidative stress. Of note, the protein level of Nrf2 was significantly increased, and its ubiquitinated fraction was reduced in stable cells compared to that in the control cells. Among E3 ubiquitin ligases known to be involved in the ubiquitination of Nrf2, DDB1 and Cullin4 associated factor 11 (DCAF11) was down-regulated at both protein and mRNA levels in stable cells, indicating that the repression of DCAF11 by TFEB may be mainly involved in the stabilization of Nrf2. In addition, the level of phosphorylated p62 at S349 was highly increased in stable cells compared to that in control cells, which could allow it to interfere with the association of Keap1 and Nrf2, thus stabilizing Nrf2. We suggest for the first time that TFEB could activate Nrf2 by increasing its stability under conditions devoid of oxidative stress.

Dietary carbohydrate quality and quantity in relation to the incidence of type 2 diabetes: A prospective cohort study of middle-aged and older Korean adults.

OBJECTIVES: This study aimed to investigate whether dietary glycemic load (GL), glycemic index (GI), and carbohydrate intake were prospectively associated with incident type 2 diabetes mellitus (T2DM) in a middle-aged and older Korean populations. METHODS: Data from the Korean Genome and Epidemiology Study were used. A total of 7294 Korean adults ages 40 y to 69 y and with no previous diagnosis of T2DM or cancer at baseline were followed for 10 y. Dietary GL, GI, and carbohydrate intake were estimated on the basis of participants' responses to a validated, semiquantitative, food-frequency questionnaire at baseline. T2DM was defined according to the World Health Organization and International Diabetes Federation criteria. RESULTS: During 7.7 y (56 377 person-years) of follow-up time, 1259 participants (17.3%) developed T2DM. Grain and its products (particularly refined and whole grains) were the greatest contributors to dietary GL. In the multivariable Cox models, dietary GL was differentially associated with T2DM risk by sex. Men in the highest quintile demonstrated a higher risk of T2DM incidence than did those with the lowest, energy-adjusted, dietary GL (hazard ratio for fifth vs. first quarter = 1.26; 95% confidence interval, 1.05-1.52; P for trend < 0.05) but no association between dietary GL and the risk of T2DM was observed in women. Similar to the findings from the main models, the effect of dietary GL on T2DM incidence according to body mass index, abdominal obesity, and physical activity levels differed substantially by sex. CONCLUSIONS: High GL diets may increase the risk of the development of T2DM in middle-aged and older Korean men but not in women. Nutrition education and emphasis on self-monitoring of dietary carbohydrate quality and quantity of overall diets is necessary in the middle-aged and older Korean populations.