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Caenorhabditis elegans (C. elegans) is a robust model organism in cell biology, physiology, pharmacology, and toxicology. It is widely recognized for its short lifespan (about 30 days), rapid life cycle, and genetic similarities to mammals. Known for their utility in lifespan research, compounds identified in C. elegans studies have shown lifespan-extending effects in higher organisms, making them invaluable for aging research. Recent work has highlighted the importance of food source conditions, specifically whether C. elegans is fed live or dead Escherichia coli (E. coli) OP50, and solvents like dimethyl sulfoxide (DMSO) in evaluating compound efficacy and organismal health. In this study, we employed C. elegans health lifespan auto-monitoring system (C-HAS), an automated imaging technology capable of objectively analyzing lifespan and healthspan by tracking movement patterns in real-time. Our results reveal that C. elegans fed dead bacteria, specifically heat-killed (HK) and freeze-dried (Fd) E. coli , display extended lifespan and healthspan compared to those fed live bacteria, reducing the proportion of short-lived, unhealthy nematodes. Moreover, 0.1 % DMSO treatment, a concentration previously reported as not affecting nematode longevity, notably shortens both lifespan and healthspan in C. elegans under dead bacterial conditions, with similar negative effects observed across different dead bacteria types. These findings highlight the importance of considering bacterial food state and DMSO presence when conducting lifespan and healthspan studies in C. elegans. This work provides foundational insights into how specific experimental conditions impact the health quality of C. elegans , advancing our understanding of environmental influences on organismal aging. [Display omitted] • Dead bacterial feeding conditions extend C. elegans lifespan and healthspan. • 0.1 % DMSO shortened lifespan and healthspan under dead bacterial conditions. • C. elegans healthy aging is influenced by dietary state and solvent, particularly DMSO. [ABSTRACT FROM AUTHOR]

Glucagon-like peptide-1 (GLP-1) is a gastrointestinal hormone that stimulates glucose-mediated insulin production by pancreatic beta cells. It is also associated with protective effects in multiple tissues. GLP-1 receptor is highly expressed in pulmonary tissue, hinting possible pulmonary delivery of GLP-1 drugs. However, little is known about the role of GLP-1 signaling in the lung, especially in mucus hypersecretory obstructive lung diseases. Here, we showed that treatment with exendin-4, a clinically available GLP-1 receptor agonist, up-regulates mucin expression in normal airway epithelial cells and in the lung of normal mice, indicating mucus stimulatory effect of GLP-1 under physiological condition. Exendin-4 also increased mucin expression in in vitro cellular and in vivo murine models of obstructive lung diseases via the activation of p38 MAP kinase. Notably, mucin induction in vivo exacerbated key pulmonary abnormalities including emphysematous phenotypes, implying that GLP-1 signaling in the lung is detrimental under pulmonary obstructive condition. Another GLP-1 receptor agonist liraglutide had similar induction of mucin. Together, our studies not only demonstrate novel physiological and pathological roles of GLP-1 in the lung but may also caution against the clinical use of inhaled GLP-1 receptor agonists in the patients with obstructive lung diseases. Image 1 • GLP-1 receptor agonist exendin-4 up-regulates mucin expression in normal airways. • Exendin-4 exacerbates key pulmonary abnormalities in the models of obstructive lung diseases possibly through p38-depndent mucin induction. • Another GLP-1 receptor agonist liraglutide also induced mucin in airway epithelial cells. [ABSTRACT FROM AUTHOR]

Abstract Cystic fibrosis transmembrane regulator (CFTR) is a cyclic AMP-dependent Cl− channel, and its dysfunction, due to CFTR gene mutations, causes the lethal inherited disorder cystic fibrosis (CF). To date, widespread dysregulation of certain coding genes in CF airway epithelial cells is well studied and considered as the driver of pulmonary abnormality. However, the involvement of non-coding genes, novel classes of functional RNAs with little or no protein-coding capacity, in the regulation of CF-associated gene dysregulation is poorly understood. Here, we utilized integrative analyses of human transcriptome array (HTA) and characterized 99 coding and 91 non-coding RNAs that are dysregulated in CFTR-defective CF bronchial epithelial cell line CFBE41o-. Among these genes, the expression level of linc-SUMF1-2 , an intergenic non-coding RNA (lincRNA) whose function is unknown, was inversely correlated with that of WT-CFTR and consistently higher in primary human CF airway epithelial cells (DHBE-CF). Further integrative analyses under linc-SUMF1- knockdown condition determined MXRA5 , SEMA5A , CXCL10 , AK022877, CTGF , MYC, AREG and LAMB3 as both CFTR- and linc-SUMF1-2 -dependent dysregulated gene sets in CF airway epithelial cells. Overall, our analyses reveal linc-SUMF1-2 as a dysregulated non-coding gene in CF as well as CFTR- linc-SUMF1-2 axis as a novel regulatory pathway involved in CF-associated gene dysregulation. Graphical abstract Image 1 Highlights • Transcriptome analysis using CF patients-derived airway epithelial cells. • Linc-SUMF1-2 is highly up-regulated in CFTR-dysfunctional airway epithelia. • Linc-SUMF1-2 modulates transcriptional level of dysregulated genes in airway epithelial cells under CFTR dysfunctional conditions. [ABSTRACT FROM AUTHOR]

Abstract: The Ganoderma lucidum (G. lucidum) is one of the oriental fungi that has been reported to have immunomodulatory properties. Although effect of β-glucans from G. lucidum has been well documented, little is known about how other major bioactive components, the triterpenes, contribute to the immunomodulatory function of G. lucidum. Here, we showed that triterpenes-rich extract of antlered form of G. lucidum (G. lucidum AF) induces TNFα production in monocytic THP-1 cells. Furthermore, the extract also synergized with lipopolysaccharide (LPS) to induce TNFα production in THP-1 cells, suggesting an immunostimulatory role of triterpenes-rich extract of G. lucidum AF. Notably, the extract enhanced LPS-induced phosphorylation of p38 mitogen-activated protein kinase (MAPK), while it suppressed LPS-induced phosphorylation of c-Jun N-terminal kinase (JNK) MAPK. p38 Inhibitor suppressed TNFα production, while JNK inhibitor enhanced TNFα production, implying that synergistic effect of the extract may work by modulating p38 and JNK MAPKs. Moreover, we found that the triterpenes-rich extract of G. lucidum AF contains high amounts of lucidenic acids. Lucidenic acid-A, -F and -D2, which seem to dominantly exist in the extract, were purified from the triterpenes-rich extract. We also identified Lucidenic acid-A and -F as modulators of JNK and p38, respectively. Thus, our data demonstrate that lucidenic acids-rich extract from G. lucidum AF enhances LPS-induced immune responses in monocytic THP-1 cells possibly via the modulation of p38 and JNK MAPKs activation. [Copyright &y& Elsevier]

Abstract: Human breast cancer resistance protein (BCRP)/MXR/ABCG2 is a well-recognized ABC half-transporter that is highly expressed at the apical membrane of many normal tissues and cancer cells. BCRP facilitates disposition of endogenous and exogenous harmful xenobiotics to protect cells/tissues from xenobiotic-induced toxicity. Despite the enormous impact of BCRP in the physiological and pathophysiological regulation of the transport of a wide variety of substrates, little is known about the factors that regulate posttranslational expression of BCRP. Here, we identified Derlin-1, a member of a family of proteins that bears homology to yeast Der1p, as a posttranslational regulator of BCRP expression. Overexpression of Derlin-1 suppressed ER to Golgi transport of wild-type (WT) BCRP that is known to be efficiently trafficked to the plasma membrane. On the other hand, protein expression of N596Q variant of BCRP, N-linked glycosylation-deficient mutant that preferentially undergoes ubiquitin-mediated ER-associated degradation (ERAD), was strongly suppressed by the overexpression of Derlin-1, whereas knockdown of Derlin-1 stabilized N596Q protein, suggesting a negative regulatory role of Derlin-1 for N596Q protein expression. Notably, knockdown of Derlin-1 also stabilized the expression of tunicamycin-induced deglycosylated WT BCRP protein, implying the importance of glycosylation state for the recognition of BCRP by Derlin-1. Thus, our data demonstrate that Derlin-1 is a negative regulator for both glycosylated and non-glycosylated BCRP expression and provide a novel posttranslational regulatory mechanism of BCRP by Derlin-1. [ABSTRACT FROM AUTHOR]

Abstract: Endoplasmic reticulum (ER) stress is involved in a wide range of pathological conditions including neurodegenerative disorders, diabetes mellitus, atherosclerosis, inflammation, and infection. The ability of ER stress to induce an inflammatory response is considered to play a role in the pathogenesis of these diseases. However, its role in regulating the gene expression and function of toll-like receptors (TLRs), host defense receptors that recognize invading pathogens, remains unknown. Here we showed that several well-characterized ER stress inducers (thapsigargin, tunicamycin, and dithiothreitol) increase the expression of TLR2 in epithelial cells. Ligand-responsiveness of TLR2 was also enhanced by ER stress inducers, implying a contributory role of ER stress for the regulation of TLR2-dependent inflammatory responses. Furthermore, there was significant increase of TLR2 mRNA level in the livers of tunicamycin-treated mice and high-fat diet-fed mice, suggesting an impact of ER stress in vivo on the expression of TLR2. Overexpression and knockdown experiments showed the importance of activating transcription factor 4 (ATF4), an ER stress-induced transcription factor, in the induction of TLR2 expression during ER stress. This was confirmed by the increased expression and function of TLR2 during treatment with salubrinal, an activator of ATF4 pathway. Taken together, our study provides further insights into the role of ER stress in enhancing host bacterial response or in exaggerating the inflammatory condition via up-regulating TLR2 expression. [Copyright &y& Elsevier]

Abstract: Toll-like receptor-2 (TLR2) is a pattern recognition receptor that senses many types of bacterial components and activates signaling pathways that induce inflammatory cytokines. A hyperresponsiveness to pathogens caused by increased expression of TLR2 triggers exaggeration of some inflammatory diseases. Here, we showed that curcumin, a well-known anti-inflammatory agent derived from the curry spice turmeric, inhibits TLR2 expression in various TLR2-expressing innate immune cell lines such as monocytic THP-1 cells, neutrophilic-differentiated HL-60 cells. Strong suppression of TLR2 gene expression was specifically observed at concentrations of curcumin in the range 40–100μM. Consistent with decreased expression of TLR2 mRNA, protein expression and ligand-responsiveness of TLR2 were markedly reduced by curcumin treatment. Moreover, curcumin-dependent down-regulation of TLR2 expression and function was also observed in primary peripheral blood monocytes (MC) and polymorphonuclear neutrophils (PMN). Finally, we determined the importance of curcumin-dependent radical generation for the suppressive effect of curcumin on TLR2 expression. Thus, our data demonstrate that curcumin inhibits TLR2 gene expression and function possibly via an oxidative process. [ABSTRACT FROM AUTHOR]

Abstract: Obese adipose tissue is characterized by an excessive production of inflammatory adipokines including tumor necrosis factor-α (TNF-α). TNF-α stimulates free fatty acid (FFA) secretion through adipocyte lipolysis, and increased plasma levels of FFA promote insulin resistance. In this report, we show that hesperetin and naringenin, two citrus flavonoids, inhibit TNF-α-stimulated FFA secretion from mouse adipocytes. These flavonoids block the TNF-α-induced activation of the NF-κB and ERK pathways. Moreover, hesperetin and naringenin prevent TNF-α from downregulating the transcription of two antilipolytic genes, perilipin and PDE3B. These effects are mediated through the inhibition of the ERK pathway. In contrast, the inhibition of the NF-κB pathway by hesperetin and naringenin suppresses the transcription of IL-6, which induces FFA secretion in an autocrine manner. Our results provide novel evidence that hesperetin and naringenin directly inhibit TNF-α-stimulated FFA secretion. These findings may be useful for ameliorating FFA-induced insulin resistance. [Copyright &y& Elsevier]

Abstract: Glucocorticoids are highly effective in the control of many inflammatory and immune diseases. Despite the importance of glucocorticoids in suppressing immune and inflammatory responses, the molecular basis for the inhibitory effect of glucocorticoids on mucin overproduction, a hallmark of chronic respiratory diseases, still remains unclear. Here we show that glucocorticoids markedly inhibit up-regulation of MUC5AC induced by NTHi, a major human bacterial pathogen causing chronic obstructive pulmonary disease and otitis media. Inhibition of NTHi-induced MUC5AC expression by dexamethasone occurs at the level of p38 MAPK via glucocorticoid receptor. Moreover, glucocorticoids up-regulate MKP-1 expression, which in turn leads to p38 dephosphorylation and the subsequent inhibition of NTHi-induced MUC5AC expression. These studies provide new insight into the molecular mechanism underlying glucocorticoid therapy and may lead to novel therapeutic intervention for inhibiting mucin overproduction in patients with NTHi infections. [Copyright &y& Elsevier]

Abstract: Mucin, a major component of mucus, plays a critical role in host mucosal defense response by participating in mucociliary clearance. However, if overproduced, overproduced mucus leads to airway mucus obstruction and conductive hearing loss. Despite extensive studies that focus on investigating how MUC5AC mucin is regulated by one inducer at a time, how MUC5AC is synergistically regulated by multiple factors remains unknown. Here we provide direct evidence for the first time that bacterial pathogen NTHi and human growth factor EGF synergize with each other to potently up-regulate MUC5AC mucin transcription. Moreover, activation of both p38 and ERK is required for synergistic induction of MUC5AC by NTHi and EGF. Finally, PAK2 and PAK4 are differentially involved in this synergistic induction of MUC5AC by acting upstream of p38 and ERK. Our studies bring novel insights into our understanding of synergistic regulation of MUC5AC mucin by both pathological and physiological inducers. [Copyright &y& Elsevier]

Abstract: Curcumin has been reported to correct cystic fibrosis caused by the ΔF508 mutation of the cystic fibrosis transmembrane regulator (CFTR) but its mechanistic action remains unclear. We have recently demonstrated that the ER chaperone calreticulin (CRT) negatively regulates the CFTR cell surface expression and activity. Thus, we aimed at determining whether CRT mediates the effect of curcumin on CFTR. We show here that the treatment with curcumin of Chinese hamster ovary cells suppressed CRT expression and increased wild-type CFTR but did not affect ΔF508 CFTR expression. However, we determined that although curcumin did not augment ΔF508 CFTR expression, it enhanced the functional competence of ΔF508 CFTR induced by 26°C incubation. Knock down of CRT by siRNA at low-temperature had a similar effect. Our findings suggest that the positive effect of curcumin on CFTR expression is mediated through the down-regulation of CRT, a negative regulator of CFTR. [Copyright &y& Elsevier]

Abstract: Myeloid elf-1-like factor (MEF) or Elf4 is an ETS protein known to regulate the basal expression of the anti-microbial peptides, lysozyme and human β-defensin-2, in epithelial cells and activate the transcription of perforin in natural killer cells. The numerous target genes of MEF and its biological functions signify the importance of this Ets transcription factor. Here we show that MEF is modified by conjugation with SUMO-1/-2 (small ubiquitin-related modifier) both in mammalian cells and in Escherichia coli overexpressing human SUMO-1/-2. We identified by point mutation that lysine 657 of MEF is the site for sumoylation. This modification down-regulated MEF activity on lysozyme and perforin promoters, and decreased the lysozyme mRNA expression. Chromatin immuno-precipitation analysis revealed that SUMO-conjugation diminished the recruitment of MEF to the lysozyme promoter, which partly explains the down-regulation of MEF activity by SUMO. These findings contribute to our understanding of the regulation of the ETS factor MEF. [Copyright &y& Elsevier]

Abstract: ATP-binding cassette (ABC) G5 (G5) and ABCG8 (G8) heterodimerize and function as sterol transporter that promote biliary excretion of neutral sterols. Both G5 and G8 interact with a lectin-like chaperone, calnexin (CNX), in the endoplasmic reticulum (ER) but the significance of this interaction remains unclear. Here, we show that not only CNX, but also its homologue calreticulin (CRT), is involved in the biosynthesis of G5/G8 sterol transporter. Both CNX and CRT interacted with immature forms of G5 and G8, and stimulated their productive folding by inhibiting their degradation. Interestingly, CRT predominantly enhanced the cell surface expression of mature G5/G8 whereas CNX did not have a similar effect. Inhibitors of N-glycan processing indicated that quality control of G5 and G8 might be differentially regulated in the ER. These findings clarify the role of CNX and CRT in the biosynthesis and quality control of G5/G8 sterol transporter. [Copyright &y& Elsevier]

Abstract: Lipopolysaccharide (LPS) induces inflammatory activation through TLR4 (toll-like receptor-4)/MD-2 (myeloid differentiation-2)/CD14 (cluster of differentiation-14) complex. Although optimal LPS signaling is required to activate our innate immune systems against gram-negative bacterium, excessive amount of LPS signaling develops a detrimental inflammatory response in gram-negative bacterial infections. Downregulation of surface TLR4 expression is one of the critical mechanisms that can restrict LPS signaling. Here, we found that membrane-anchored CD14 is required for LPS-induced downregulation of TLR4 and MD-2 in CHO cells. Moreover, pretreatment of the cells with sterol-binding agent filipin reduced LPS-induced TLR4 downregulation, suggesting the involvement of caveolae-mediated endocytosis pathway. Involvement of caveolae in LPS-induced TLR4 endocytosis was further confirmed by immunoprecipitation. Thus, our data indicate that caveolae-dependent endocytosis pathway is involved in LPS-induced TLR4 downregulation and that this is dependent on membrane-anchored CD14 expression. [Copyright &y& Elsevier]

Abstract: Hyperthermia, which is used as an adjunctive therapy for cancer, is known to modulate the activity of natural killer (NK) cells in vitro, but its effect in vivo is unclear. In the present study, we used a whole body hyperthermia (WBH) device heated by infrared rays to evaluate the effect of WBH on mice models. We demonstrate here that wild type C57BL/6J mice exposed to 42°C for 60min had reduced NK cell cytolytic activity against YAC-1 target cells as determined by cytolytic assay. This result was confirmed using Rag-2 knockout mice, which possess functional NK but not cytolytic T or NK-T cells. Moreover, WBH decreased the mRNA expression of perforin and granzyme B in spleens of mice. But the expression of TNF cytokines (Fas ligand and TRAIL) was unchanged. These data suggest that the suppression of NK cell activity induced by WBH could be mediated through the perforin/granzyme pathway. [Copyright &y& Elsevier]

Lysozyme protects us from the ever-present danger of bacterial infection. The expression of lysozyme is, in part, regulated by the Ets factor, myeloid elf-1-like factor (MEF). MEF binds to the ETS site of the lysozyme promoter at −46 to −40bp. Closer analysis of the promoter using a series of deletion mutants and point mutants indicated that the region around −75bp is also essential in regulating the activity of lysozyme. The sequences in this region correspond to the Sp1 consensus binding site. Sp1 is known to regulate a variety of house-keeping and tissue-specific genes by itself or with other transcription factors like AP-1 or ETS. We indicate here that Sp1 regulates the lysozyme gene by binding to the GT-core sequences of lysozyme promoter. Treatment with mithramycin A down-regulated the promoter activity and the transfection of anti-sense Sp1 induced a decrease in the endogenous expression of lysozyme. [Copyright &y& Elsevier]

Granulocyte–macrophage colony-stimulating factor (GM-CSF) is a cytokine expressed in the non-small lung carcinoma cells (NSCLC). However, transcriptional regulation of GM-CSF is not well characterized in NSCLC. In this study we found that two cis-acting ETS family consensus sites are important for transcriptional regulation of GM-CSF in A549 human lung carcinoma cells. These two sites are located separately at around −40 and −100 bp from the transcription start site. Results of transient transfection assays with A549 cells indicated that ETS2 had a strong positive effect on GM-CSF promoter activity. Furthermore, this activity was enhanced by protein kinase C activator, phorbol 12-myristate 13-acetate (PMA), in an ETS consensus-dependent manner, while PMA could also enhance the expression level of ETS2. The protein kinase C inhibitors decreased GM-CSF promoter activity induced by the protein kinase C activator PMA. We also found that antisense ETS2 mRNA decreased PMA-induced GM-CSF promoter activity, supporting the possibility that ETS2 is involved in protein kinase C-induced GM-CSF transcriptional function. Endogenous expression of GM-CSF mRNA was increased by ETS2 transfection and the increased expression was further enhanced by PMA. These data indicate that GM-CSF is up-regulated by ETS2, a target of protein kinase C. [Copyright &y& Elsevier]