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Obesity is a world-wide problem, especially the child obesity, with the complication of various metabolic diseases. Child obesity can be developed as early as the age between 2 and 6. The expansion of fat mass in child age includes both hyperplasia and hypertrophy of adipose tissue, suggesting the importance of proliferation and adipogenesis of preadipocytes. The changed composition of gut microbiota is associated with obesity, revealing the roles of lipopolysaccharide (LPS) on manipulating adipose tissue development. Studies suggest that LPS enters the circulation and acts as a pro-inflammatory regulator to facilitate pathologies. Nevertheless, the underlying mechanisms behind LPS-modulated obesity are yet clearly elucidated. This study showed that LPS enhanced the expression of cyclooxygenase-2 (COX-2), an inflammatory regulator of obesity, in preadipocytes. Pretreating preadipocytes with the scavenger of reactive oxygen species (ROS) or the inhibitors of NADPH oxidase or p42/p44 MAPK markedly decreased LPS-stimulated gene expression of COX-2 together with the phosphorylation of p47

Background Trichomoniasis is the most common non-viral sexually transmitted disease caused by the protozoan parasite Trichomonas vaginalis. Metronidazole (MTZ) is a widely used drug for the treatment of trichomoniasis; however, increased resistance of the parasite to MTZ has emerged as a highly problematic public health issue. Methods We conducted iTRAQ-based analysis to profile the proteomes of MTZ-sensitive (MTZ-S) and MTZ-resistant (MTZ-R) parasites. STRING and gene set enrichment analysis (GESA) were utilized to explore the protein-protein interaction networks and enriched pathways of the differentially expressed proteins, respectively. Proteins potentially related to MTZ resistance were selected for functional validation. Results A total of 3123 proteins were identified from the MTZ-S and MTZ-R proteomes in response to drug treatment. Among the identified proteins, 304 proteins were differentially expressed in the MTZ-R proteome, including 228 upregulated and 76 downregulated proteins. GSEA showed that the amino acid-related metabolism, including arginine, proline, alanine, aspartate, and glutamate are the most upregulated pathways in the MTZ-R proteome, whereas oxidative phosphorylation is the most downregulated pathway. Ten proteins categorized into the gene set of oxidative phosphorylation were ATP synthase subunit-related proteins. Drug resistance was further examined in MTZ-S parasites pretreated with the ATP synthase inhibitors oligomycin and bafilomycin A1, showing enhanced MTZ resistance and potential roles of ATP synthase in drug susceptibility. Conclusions We provide novel insights into previously unidentified proteins associated with MTZ resistance, paving the way for future development of new drugs against MTZ-refractory trichomoniasis.

Recently, many studies have indicated that ambient air particulate matter (PM) can increase the risk of oral cancer. The most common malignant tumor in the oral cavity is oral squamous cell carcinoma (OSCC). Usually, cancer cell migration/invasion is the most important cause of cancer mortality. Matrix metalloproteinase-2 (MMP-2) and MMP-9 have been shown to play important roles in regulating metastasis and the tumor microenvironment. Here, we studied the anti-cancer effects of surfactin, a cyclic lipopeptide generated by Bacillus subtilis, on cancer cell migration and invasion. Surfactin suppressed PM-promoted cell migration and invasion and colony formation of SCC4 and SCC25 human oral squamous cell carcinoma cell lines. We observed that PM induced MMP-2 and MMP-9 expression, which was inhibited by surfactin. Transfection with p65, p50, c-Jun, c-Fos, p85, p110, Akt, mammalian target of rapamycin (mTOR), or interleukin-6 (IL-6) siRNA markedly inhibited PM-induced MMP-2 and MMP-9 expression. Moreover, surfactin could reduce Akt, mTOR, p65, and c-Jun activation and IL-6 secretion induced by PM. Finally, we proved that transfection with Akt, p65, or c-Jun siRNA significantly inhibited PM-induced IL-6 release. Taken together, these results suggest that surfactin functions as a suppressor of PM-induced MMP2/9-dependent oral cancer cell migration and invasion by inhibiting the activation of phosphoinositide 3-kinase (PI3K)/Akt/mTOR and PI3K/Akt/nuclear factor-κB (NF-κB) and activator protein-1 (AP-1)/IL-6 signaling pathways.

Recently, ambient air particulate matter (PM) has been shown to increase the risk of oral cancer. The most common malignant tumor in the oral cavity is oral squamous cell carcinoma (OSCC). Recent studies have revealed that surfactin, a cyclic lipopeptide generated by Bacillus subtilis, has anti-inflammatory and anti-cancer properties. However, the exact anti-cancer effects of surfactin on human OSCC and underlying molecular mechanisms remain largely unknown. In the present study, we found that treatment of SCC4 and SCC25 cells (human OSCC cell lines) with surfactin reduced the viability of SCC4 and SCC25 cells by induction of apoptosis. Surfactin-induced apoptosis was associated with caspase activation and poly(ADP-ribose) polymerase (PARP) cleavage and was regulated by the mitochondrial pathway, exemplified by mitochondrial depolarization, mitochondrial-derived reactive oxidative species (ROS) production, cytochrome c release, up-regulation of Bad and Bax, and down-regulation of Bcl-2. Surfactin induced NADPH oxidase-dependent ROS generation, which appeared essential for the activation of the mitochondrial pathway. Surfactin-induced mitochondrial-derived ROS generation was associated with JNK1/2 activation. After treatment with surfactin, ROS caused JNK1/2-dependent cell death of SCC4 and SCC25 cells. Taken together, our findings suggest that surfactin induces mitochondria associated apoptosis of human OSCC cell lines, and surfactin may be a potential chemotherapeutic agent for future OSCC treatment.

Trichomonas vaginalis is the causative agent of trichomoniasis, the most prevalent non-viral sexually transmitted infection worldwide. Metronidazole (MTZ) is the mainstay of anti-trichomonal chemotherapy; however, drug resistance has become an increasingly worrying issue. Additionally, the molecular events of MTZ-induced cell death in T. vaginalis remain elusive. To gain insight into the differential expression of genes related to MTZ resistance and cell death, we conducted RNA-sequencing of three paired MTZ-resistant (MTZ-R) and MTZ-sensitive (MTZ-S) T. vaginalis strains treated with or without MTZ. Comparative transcriptomes analysis identified that several putative drug-resistant genes were exclusively upregulated in different MTZ-R strains, such as ATP-binding cassette (ABC) transporters and multidrug resistance pumps. Additionally, several shared upregulated genes among all the MTZ-R transcriptomes were not previously identified in T. vaginalis, such as 5′-nucleotidase surE and Na+-driven multidrug efflux pump, which are a potential stress response protein and a multidrug and toxic compound extrusion (MATE)-like protein, respectively. Functional enrichment analysis revealed that purine and pyrimidine metabolisms were suppressed in MTZ-S parasites upon drug treatment, whereas the endoplasmic reticulum-associated degradation (ERAD) pathway, proteasome, and ubiquitin-mediated proteolysis were strikingly activated, highlighting the novel pathways responsible for drug-induced stress. Our work presents the most detailed analysis of the transcriptional changes and the regulatory networks associated with MTZ resistance and MTZ-induced signaling, providing insights into MTZ resistance and cell death mechanisms in trichomonads.

The three most common sexually transmitted infections (STIs) are Chlamydia trachomatis (CT), Neisseria gonorrhoeae (GC) and Trichomonas vaginalis (TV). The prevalence of these STIs in Taiwan remains largely unknown and the risk of STI acquisition affected by the vaginal microbiota is also elusive. In this study, a total of 327 vaginal swabs collected from women with vaginitis were analyzed to determine the presence of STIs and the associated microorganisms by using the BD Max CT/GC/TV molecular assay, microbial cultures, and 16S rRNA sequencing. The prevalence of CT, TV, and GC was 10.8%, 2.2% and 0.6%, respectively. A culture-dependent method identified that Escherichia coli and Streptococcus agalactiae (GBS) were more likely to be associated with CT and TV infections. In CT-positive patients, the vaginal microbiota was dominated by L. iners, and the relative abundance of Gardnerella vaginalis (12.46%) was also higher than that in TV-positive patients and the non-STIs group. However, Lactobacillus spp. was significantly lower in TV-positive patients, while GBS (10.11%), Prevotella bivia (6.19%), Sneathia sanguinegens (12.75%), and Gemella asaccharolytica (5.31%) were significantly enriched. Using an in vitro co-culture assay, we demonstrated that the growth of L. iners was suppressed in the initial interaction with TV, but it may adapt and survive after longer exposure to TV. Additionally, it is noteworthy that TV was able to promote GBS growth. Our study highlights the vaginal microbiota composition associated with the common STIs and the crosstalk between TV and the associated bacteria, paving the way for future development of health interventions targeting the specific vaginal bacterial taxa to reduce the risk of common STIs.

The proliferation and adipogenesis of preadipocytes played important roles in the development of adipose tissue and contributed much to the processes of obesity. On the other hand, lipopolysaccharide (LPS), also known as endotoxin, is a key outer membrane component of gram-negative bacteria in the gut microbiota, and has a dominant role in linking inflammation to high-fat diet-induced metabolic syndrome. Studies suggested the potential roles of LPS in hepatic steatosis and in obese mice models. However, the molecular mechanisms underlying LPS-regulated obesity remained largely unknown. Here we reported that LPS stimulated expression of cyosolic phospholipase A2 (cPLA2), one of inflammation regulators of obesity, in the preadipocytes. Pretreatment the inhibitors of JAK2, STAT3, STAT5 or AMPK significantly reduced LPS-increased mRNA and protein expression of cPLA2 together with phosphorylation of JAK2, STAT3, STAT5 and AMPK, separately. Similarly, transfection of siRNA against JAK2 or AMPK abolished expression of cPLA2 and phosphorylation of JAK2 or AMPK together with downregulated expression of JAK2 and AMPK protein. LPS enhanced activation of STAT3 and STAT5 via JAK2-dependent manner in the preadipocytes. Transfection of JAK2 or AMPK siRNA further proofed the independence of JAK2 and AMPK in LPS-treated preadipocytes. In addition, LPS-increased DNA synthesis, cell numbers and cell viability of preadipocytes were attenuated by AACOCF3, AG490, BML-275, cPLA2 siRNA, JAK2 siRNA or AMPK siRNA. Attenuation JAK2/STAT or AMPK-dependent cPLA2 expression reduced LPS-mediated adipogenesis of preadipocytes. Stimulation of arachidonic acid or AMPK activator, A-769662, increased cell numbers and cell viability and promoted differentiation of preadipocytes. Collectively, these results indicated that LPS increased preadipocytes proliferation and adipogenesis via JAK/STAT and AMPK-dependent cPLA2 expression. The mechanisms of LPS-stimulated cPLA2 expression may be a link between bacteria and obesity and provides the molecular basis for preventing metabolic syndrome or hyperplasic obesity.

Objectives To investigate the anticancer effects and underlying mechanisms of surfactin on human oral squamous cell carcinoma (OSCC). Materials and methods The capacity of surfactin to induce apoptosis, autophagy, and cell cycle arrest of two different human OSCC cell lines was investigated by cell viability, acridine orange staining, and cell cycle regulatory protein expression, respectively. The signaling network underlying these processes were determined by the analysis of reactive oxygen species (ROS) generation, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity, endoplasmic reticulum (ER) stress-related protein levels, calcium release, mitogen-activated protein kinases activation and cell cycle regulatory protein expression through corresponding reagents and experiments under various experimental conditions using specific pharmaceutical inhibitors or small interfering RNAs. Results Surfactin was able to induce apoptosis through NADPH oxidase/ROS/ER stress/calcium-downregulated extracellular signal-regulated kinases 1/2 pathway. Surfactin could also lead to autophagy that shared the common regulatory signals with apoptosis pathway until calcium node. Cell cycle arrest at G2 /M phase caused by surfactin was demonstrated through p53 and p21 accumulation combined p34cdc2 , phosphorylated p34cdc2 and cyclin B1 inhibition, which was regulated by NADPH oxidase-derived ROS. Conclusion Surfactin could induce apoptosis, autophagy, and cell cycle arrest in ROS-dependent manner, suggesting a multifaced anticancer agent for OSCC.

Particulate matter (PM), a major air pollutant, may be associated with adverse cardiovascular effects. Reactive oxygen species- (ROS-) dependent proinflammatory cytokine production, such as interleukin-6 (IL-6), is a possible underlying mechanism. Carbon monoxide- (CO-) releasing molecule-2 (CORM-2) which liberates exogenous CO can exert many beneficial effects, particularly anti-inflammation and antioxidant effects. The purpose of this study was to explore the protective effects and underpinning mechanisms of CORM-2 on PM-induced aorta inflammation. Here, human aortic vascular smooth muscle cells (HASMCs) were utilized as in vitro models for the assessment of signaling pathways behind CORM-2 activities against PM-induced inflammatory responses, including Toll-like receptors (TLRs), NADPH oxidase, ROS, nuclear factor-kappa B (NF-κB), and IL-6. The modulation of monocyte adherence and HASMC migration, that are two critical cellular events of inflammatory process, along with their regulators, including intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and matrix metalloproteinase-2 (MMP-2) and MMP-9, in response to PM by CORM-2, were further evaluated. Finally, mice experiments under different conditions were conducted for the in vivo evaluation of CORM-2 benefits on the expression of inflammatory molecules including IL-6, ICAM-1, VCAM-1, MMP-2, and MMP-9. Our results found that PM could induce aorta inflammation in vitro and in vivo, as evidenced by the increase of IL-6 expression that was regulated by the TLR2 and TLR4/NADPH oxidase/ROS/NF-κB signaling pathway, thereby promoting ICAM-1- and VCAM-1-dependent monocyte adhesion and MMP-2- and MMP-9-dependent HASMC migration. Importantly, our experimental models demonstrated that CORM-2-liberated CO effectively inhibited the whole identified PM-induced inflammatory cascade in HASMCs and tissues. In conclusion, CORM-2 treatment may elicit multiple beneficial effects on inflammatory responses of aorta due to PM exposure, thereby providing therapeutic value in the context of inflammatory diseases of the cardiovascular system.

Abdominal aortic aneurysm (AAA) is a common inflammatory vascular disease. Angiotensin II (Ang II) involves in AAA progression by promoting the proliferation and migration of vascular smooth muscle cells, the degradation of extracellular matrices, and the generation of ROS to lead to vascular inflammation. Carbon monoxide releasing molecule-2 (CORM-2) is known to exert anti-inflammatory and antioxidant activities. However, it remains unclear whether CORM-2 can suppress Ang II-induced vascular inflammation to prevent AAA progression. Therefore, this study aimed to investigate the vasoprotective effects of CORM-2 against Ang II-induced inflammatory responses of human aortic smooth muscle cells (HASMCs) and the underlying mechanisms of those effects. The results showed that Ang II induced inflammatory responses of HASMCs via NADPH oxidase- and mitochondria-derived ROS/NF-κB/IL-6/Jak2/Stat3 pathway which was attenuated by the pretreatment with CORM-2. Additionally, CORM-2 further exhibited anti-inflammatory activities in Ang II-stimulated HASMCs, as indicated by the reduction of monocyte adhesion to HASMCs and migration of HASMCs via the suppression of ICAM-1 and VCAM-1 as well as MMP-2 and MMP-9 levels, respectively. Moreover, Ang II-induced COX-2-mediated PGE

Introduction Exposure to airborne particulate matter (PM) increases the proportion of oral inflammatory diseases. During the formation of inflammatory conditions, the nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3) inflammasome activation plays an important regulator. Carbon monoxide (CO) arising from heme degradation, catalyzed particularly by heme oxygenase-1 (HO-1), has been shown to own cytoprotective effects including anti-inflammation and antioxidant. Here, we determined the novel mechanisms of carbon monoxide releasing molecule-2 (CORM-2) on PM-induced inflammatory responses in human oral keratinocytes (HOKs). Methods The effects of CORM-2 on the expression of various inflammatory proteins induced by PM were determined by Western blot, real-time PCR, promoter assay, and ELISA. The involvement of signaling molecules in these responses was studied by using the selective pharmacological inhibitors and siRNAs. Results We proved that PM enhanced C-reactive protein (CRP) levels, NLRP3 inflammasome and caspase-1 activation, and IL-1β release, which were reduced by preincubation with CORM-2. Transfection with PKCα siRNA and preincubation with the ROS scavenger (N-acetyl-cysteine, NAC), an inhibitor of NADPH oxidase (diphenyleneiodonium, DPI), or the mitochondria-specific superoxide scavenger (MitoTEMPO) inhibited PM-mediated inflammatory responses. In addition, PM-regulated PKCα and NADPH oxidase activation as well as NADPH oxidase- and mitochondria-derived ROS generation were inhibited by CORM-2, but not inactivate CORM-2 (iCORM-2) pretreatment. At the end, we confirmed that CORM-2 improved PM-induced inflammatory responses via the induction of Nrf2 activation and HO-1 expression. Conclusion We suggest that CORM-2 inhibits PM-induced inflammatory responses in HOKs via the inhibition of PKCα/ROS/NLRP3 inflammasome activation combined with the induction of Nrf2/HO-1 expression.

Staphylococcus aureus (S. aureus) can lead to many life-threatening diseases. It has the ability to invade normal endovascular tissue. The molecular mechanisms and pathological changes of endothelial cells after S. aureus infection are of interest, but the basic understanding of how S. aureus destroys this barrier is not clear. Here, we showed that S. aureus enhanced COX-2 expression and prostaglandin E2 (PGE2) secretion in human aortic endothelial cells (HAECs). In addition, S. aureus induced PGE2/interleukin-6 (IL-6)/matrix metallopeptidase-9 (MMP-9)-dependent cell migration. S. aureus-induced COX-2, IL-6, and MMP-9 levels were inhibited by transfection with siRNA of Toll-like receptor 2 (TLR2), p38, p42, p44, p50, or p65. S. aureus also induced p38 MAPK, ATF2, ERK1/2, and NF-κB p65 activation. Interestingly, we proved that S. aureus decreased intracellular generation of reactive oxygen species (ROS), which suggests that the inhibition of ROS production promoted inflammatory responses. Finally, we showed that S. aureus enhanced a variety of biomarkers of inflammation in cardiovascular diseases. However, the free radical scavenger (MCI-186) or antioxidant (N-acetyl-L-cysteine, NAC) markedly enhanced S. aureus-induced COX-2 mRNA levels in the aorta tissues. Taken together, these findings established that S. aureus promoted aorta inflammation via activation of p38 MAPK, ERK1/2, and NF-κB and inhibition of ROS generation.

Pemphigus foliaceus (PF) is a chronic autoimmune blistering dermatosis (ABD) that often involves the face, scalp and trunk. Kaposi's varicelliform eruption (KVE) is a viral vesicular dermatosis that rarely complicates ABD. In addition, KVE infrequently causes ocular involvement including herpes simplex keratitis (HSK). Timely diagnosis of KVE may be difficult in patients with PF due to their overlapping clinical features. In this report, we described a 79-year-old female with PF complicated by KVE and bilateral HSK. This report reminds physicians that KVE should be considered in patients with refractory PF and other ABD.

HO-1 (heme oxygenase-1), an antioxidant enzyme, induced by rosiglitazone (PPAR ligands) can be a potential treatment of inflammation. However, the mechanisms of rosiglitazone-induced HO-1 expression in human pulmonary alveolar epithelial cells (HPAEpiCs) remain largely unknown. In this study, we found that upregulation of HO-1 in vitro or in vivo by rosiglitazone attenuated VCAM-1 gene expression and monocyte adhesion to HPAEpiCs challenged with lipopolysaccharide (LPS). The inhibitory effects of rosiglitazone on LPS-mediated responses were reversed by transfection with HO-1 siRNA. LPS-induced VCAM-1 expression was mediated through NF-κB activation which was attenuated by rosiglitazone via suppressing p65 activation and translocation into the nucleus. Moreover, pretreatment with the inhibitor of PKCs (H7), PKCα (Gö6976), AMPKα (Compound C), p38 MAPKα (p38i VIII), SIRT1 (Sirtinol), or PPARγ (T0070907) and transfection with siRNA of PKCα, AMPKα, p38 MAPKα, SIRT1, or PPARγ abolished the rosiglitazone-induced HO-1 expression in HPAEpiCs. Further studies indicated that rosiglitazone stimulated SIRT1 deacetylase leading to PGC1α translocation from the cytosol into the nucleus, promoting fragmentation of NCoR and phosphorylation of PPARγ. Subsequently, PPARγ was activated by phosphorylation of PKCα, AMPKα, p38 MAPKα, and SIRT1, which turned on transcription of HO-1 gene by binding to PPAR response element (PPRE) and enhancing PPARγ promoter activity. These results suggested that rosiglitazone-induced HO-1 expression is mediated through PKCα/AMPKα/p38 MAPKα/SIRT1-dependent deacetylation of Ac-PGC1α and fragmentation of NCoR/PPARγ activation in HPAEpiCs. Up-regulation of HO-1 protected against the inflammatory responses triggered by LPS, at least in part, through attenuation of NF-κB.

Background and Purpose Haem oxygenase-1 (HO-1) could provide cytoprotection against various inflammatory diseases. However, the mechanisms underlying the protective effect of CO-releasing molecule-2 (CORM-2)-induced HO-1 expression against TNF-α-induced inflammatory responses in human pulmonary alveolar epithelial cells (HPAEpiCs) remain unknown. Experimental Approach CORM-2-induced HO-1 protein and mRNA expression, and signalling pathways were determined by Western blot and real-time PCR, coupled with respective pharmacological inhibitors or transfection with siRNAs. The effect of CORM-2 on TNF-α-induced increase in leukocyte counts in BAL fluid and VCAM-1 expression in lung was determined by cell counting and Western blot analysis. Key Results CORM-2 attenuated the TNF-α-induced pulmonary haematoma, VCAM-1 expression and increase in leukocytes through an up-regulation of HO-1 in mice; this effect of CORM-2 was reversed by the HO-1 inhibitor zinc protoporphyrin IX. Furthermore, CORM-2 increased HO-1 protein and mRNA expression as well as the phosphorylation of PYK2, PKCα and ERK1/2 (p44/p42 MAPK) in HPAEpiCs; these effects were attenuated by their respective pharmacological inhibitors or transfection with siRNAs. Inhibition of PKCα by Go6976 or Go6983 attenuated CORM-2-induced stimulation of PKCα and ERK1/2 phosphorylation but had no effect on PYK2 phosphorylation. Moreover, inhibition of PYK2 by PF431396 reduced the phosphorylation of all three protein kinases. Finally, PYK2/PKCα/ERK1/2-mediated stimulation of activator protein 1 was shown to play a key role in CORM-2-induced HO-1 expression via an up-regulation of c-Fos mRNA. Conclusions and Implications CORM-2 activates a PYK2/PKCα/ERK1/2/AP-1 pathway leading to HO-1 expression in HPAEpiCs. This HO-1/CO system might have potential as a therapeutic target in pulmonary inflammation.

Background: Mevastatin (MVS), a 3-hydroxy-3-methylglutaryl coenzyme, a reductase (HMG-CoA) inhibitor, has anti-inflammatory effects potentially via up-regulation of heme oxygenase-1 (HO-1). However, the mechanisms underlying MVS-induced HO-1 expression remain largely unknown in human pulmonary alveolar epithelial cells (HPAEpiCs). Methods: HO-1 and intercellular adhesion molecule (ICAM)-1 expression were determined using real-time PCR, Western blotting, and promoter reporter analyses. The signaling components were investigated using pharmacological inhibitors or specific small interfering RNA (siRNA)s. Interaction between Nrf2 and the antioxidant response element (ARE) binding site for the HO-1 promoter was determined by chromatin immunoprecipitation (ChIP) assay. Results: Upregulation of HO-1 by MVS attenuated the tumor necrosis factor (TNF)-&alpha, stimulated ICAM-1 expression associated with THP-1 adhesion to HPAEpiCs. These inhibitory effects of HO-1 were reversed by tin protoporphyrin (SnPP)IX or by transfection with HO-1 siRNA. MVS-induced HO-1 expression was mediated via NADPH oxidase (Nox)-derived reactive oxygen species (ROS) generation. Activation of Nox2/ROS further stimulated the phosphorylation of p47phox, proto-oncogene tyrosine-protein kinase (c-Src), platelet-derived growth factor receptor (PDFGR)&alpha, protein kinase B (Akt), and Nrf2, which were inhibited by siRNAs. Pretreatment with pharmacological inhibitors, including diphenyleneiodonium (DPI), apocynin (APO), N-acetyl-L-cysteine (NAC), PP1, AG1296, or LY294002, reduced the MVS-activated Nrf2 nuclear-translocation binding to the ARE on the HO-1 promoter. Conclusions: MVS-induced HO-1 is, at least in part, mediated through a p47phox/Nox2/ROS-dependent activation of c-Src/PDGFR&alpha, /PI3K/Akt-regulated Nrf2/ARE axis and suppresses the TNF-&alpha, mediated inflammatory responses in HPAEpiCs.

Background Chronic rhinosinusitis (CRS) is a multi-factorial disorder that causes systemic symptoms beyond rhinologic symptoms alone. A possible association between autonomic nervous system (ANS) dysfunction and CRS has been identified; however, few studies have confirmed this observation. In this study, we prospectively measured changes in ANS dysfunction symptoms following functional endoscopic sinus surgery (FESS) and explored the impact of ANS dysfunction on surgical outcomes of CRS. Methodology Patients diagnosed with CRS who consented to surgical intervention were included prospectively. All patients completed the Sino-nasal Outcome Test-22 (SNOT-22) and the 31-item Composite Autonomic Symptom Score (COMPASS 31) questionnaires before the operation and during the follow-up period. Clinical demographic data, Lund-Mackay, and modified Lund-Kennedy scores were recorded and measured. Results A total of 102 patients were enrolled. The median SNOT-22 and COMPASS 31 scores significantly improved following FESS from 43.0 to 14.0 and 21.0 to 11.2 (all P less than 0.001), respectively. FESS led to a significant reduction in the prevalence of various ANS dysfunction symptoms. In multivariate analyses, revision surgeries (odds ratio [OR] 5.012, 95% confidence interval [CI] 1.52416.489; P=0.008), CRS with nasal polyps (OR 4.071, 95% CI 1.454-11.40; P=0.008), and higher Pre-FESS COMPASS 31 scores (OR 1.043, 95% CI 1.003-1.084; P=0.036) were independent risk factors for uncontrolled inflammation following FESS. Conclusions ANS dysfunction symptoms are prevalent in CRS and higher preoperative COMPASS 31 scores correspond with poor surgical outcomes. Following FESS, the majority of ANS dysfunction symptoms can be alleviated. Further investigations are required to explore the possible mechanism of how ANS is involved in the pathogenesis of CRS.

Obesity is associated with metabolic endotoxemia, reactive oxygen species (ROS), chronic inflammation, and obese kidney fibrosis. Although the fat&ndash, intestine&ndash, kidney axis has been documented, the pathomechanism and therapeutic targets of obese kidney fibrosis remain unelucidated. To mimic obese humans with metabolic endotoxemia, high-fat-diet-fed mice (HF group) were injected with lipopolysaccharide (LPS) to yield the obese kidney fibrosis&ndash, metabolic endotoxemia mouse model (HL group). Therapeutic effects of ROS, cytosolic phospholipases A2 (cPLA2) and cyclooxygenase-2 (COX-2) inhibitors were analyzed with a quantitative comparison of immunohistochemistry stains and morphometric approach in the tubulointerstitium of different groups. Compared with basal and HF groups, the HL group exhibited the most prominent obese kidney fibrosis, tubular epithelial lipid vacuoles, and lymphocyte infiltration in the tubulointerstitium. Furthermore, inhibitors of nonspecific ROS, cPLA2 and COX-2 ameliorated the above renal damages. Notably, the ROS-inhibitor-treated group ameliorated not only oxidative injury but also the expression of cPLA2 and COX-2, indicating that ROS functions as the upstream signaling molecule in the inflammatory cascade of obese kidney fibrosis. ROS acts as a key messenger in the signaling transduction of obese kidney fibrosis, activating downstream cPLA2 and COX-2. The given antioxidant treatment ameliorates obese kidney fibrosis resulting from a combined high-fat diet and LPS&mdash, ROS could serve as a potential therapeutic target of obese kidney fibrosis with metabolic endotoxemia.

Psoriasis is a chronic inflammatory skin disease that affects about 2% of the general population. Activation of the Absent in Melanoma 2 (AIM2) inflammasome is crucial for immune defense, but it can also cause inflammatory and autoimmune diseases, including psoriasis. We currently lack an AIM2 inflammasome inhibitor that could be used therapeutically. Here, we show that EFLA 945, a safe product of red grape vine leaf extracts, can restrict AIM2 inflammasome activation. Mechanistically, EFLA945 prevents DNA entry into THP-1-derived macrophages, and thereby inhibits cytoplasmic DNA-dependent apoptosis-associated speck-like protein containing a CARD (ASC) oligomerization, caspase-1 activation, and the secretion of interleukin (IL)-1β and IL-18. The major phytochemicals of EFLA 945, resveratrol and peonidin 3-O-glucoside (P3G), appear to be the potential bioactive compounds responsible for its ability to restrict AIM2-dependent IL-1β secretion. Importantly, in an in vivo mouse model, EFLA 945 attenuates imiquimod (IMQ)-induced psoriasis-related pro-inflammatory responses in topical psoriatic skin, including caspase-1 activation, IL-1β maturation, and IL-17 production, and decreases the severity of psoriasis. Together, these results demonstrate that the safe natural product, EFLA 945, can restrict the AIM2 inflammasome activation through preventing DNA entry and may prove beneficial for treating psoriasis.

Obesity is a worldwide epidemic problem and correlates to varieties of acute or chronic lung diseases such as acute respiratory distress syndrome, chronic obstructive pulmonary disease, and pulmonary fibrosis. An increase of leptin, a kind of adipokine, in lean mice plasma has been found to impair immune responses and facilitate the infection of Klebsiella pneumoniae, resulting in increased pneumonia severity. Also, a higher leptin level is found in exhaled breath condensates of obese or asthmatic subjects, compared to healthy ones, suggesting that leptin is involved in the occurrence or exacerbation of lung injury. In previous studies, we showed that leptin stimulated cytosolic phospholipase A2-&alpha, (cPLA2&alpha, ) gene expression in lung alveolar type II cells via mitogen-activated protein kinase (MAPK) and nuclear factor-kappa B (NF-&kappa, B)-activated coactivator p300. Herein, we show that the in vivo application of leptin in the respiratory system upregulated the expression of inflammatory proteins cPLA2&alpha, and cyclooxygenase-2 (COX-2) together with leukocyte infiltration. Treatment with an ROS scavenger (N-acetylcysteine, NAC), an NADPH oxidase inhibitor (apocynin), or an activating protein (AP)-1 inhibitor (tanshinone IIA) attenuated leptin-mediated cPLA2&alpha, /COX-2 expression and leukocyte recruitment in the lung. Leptin increased intracellular oxidative stress in a leptin receptor (OB-R) and NADPH oxidase-dependent manner, leading to the phosphorylation of the AP-1 subunit c-Jun. In summation, leptin increased lung cPLA2&alpha, /COX-2 expression and leukocyte recruitment via the NADPH oxidase/ROS/AP-1 pathway. Understanding the inflammatory effects of leptin on the pulmonary system provides opportunities to develop strategies against lung injury related to metabolic syndrome or obesity.

Exposure to airborne particulate matter (PM) not only causes lung inflammation and chronic respiratory diseases, but also increases the incidence and mortality of cardiopulmonary diseases. The nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3) inflammasome activation has been shown to play a critical role in the formation of many chronic disorders. On the other hand, carbon monoxide (CO) has been shown to possess anti-inflammatory and antioxidant effects in many tissues and organs. Here, we investigated the effects and mechanisms of carbon monoxide releasing molecule-2 (CORM-2) on PM-induced inflammatory responses in human pulmonary alveolar epithelial cells (HPAEpiCs). We found that PM induced C-reactive protein (CRP) expression, NLRP3 inflammasome activation, IL-1β secretion, and caspase-1 activation, which were inhibited by pretreatment with CORM-2. In addition, transfection with siRNA of Toll-like receptor 2 (TLR2) or TLR4 and pretreatment with an antioxidant (N-acetyl-cysteine, NAC), the inhibitor of NADPH oxidase (diphenyleneiodonium, DPI), or a mitochondria-specific superoxide scavenger (MitoTEMPO) reduced PM-induced inflammatory responses. CORM-2 also inhibited PM-induced NADPH oxidase activity and NADPH oxidase- and mitochondria-derived ROS generation. However, pretreatment with inactivate CORM-2 (iCORM-2) had no effects on PM-induced inflammatory responses. Finally, we showed that CORM-2 inhibited PM-induced CRP, NLRP3 inflammasome, and ASC protein expression in the lung tissues of mice and IL-1β levels in the serum of mice. PM-enhanced leukocyte count in bronchoalveolar lavage fluid in mice was reduced by CORM-2. The results of this study suggested a protective role of CORM-2 in PM-induced lung inflammation by inhibiting the TLR2 and TLR4/ROS-NLRP3 inflammasome-CRP axial.

Adiponectin, an adipokine, accumulated in lung system via T-cadherin after allergens/ozone challenge. However, the roles of adiponectin on lung pathologies were controversial. Here we reported that adiponectin stimulated expression of inflammatory proteins, cytosolic phospholipase A2 (cPLA2), cyclooxygenase-2 (COX-2), and production of reactive oxygen species (ROS) in human alveolar type II A549 cells. AdipoR1/2 involved in adiponectin-activated NADPH oxidase and mitochondria, which further promoted intracellular ROS accumulation. Protein kinase C (PKC) may involve an adiponectin-activated NADPH oxidase. Similarly, p300 phosphorylation and histone H4 acetylation occurred in adiponectin-challenged A549 cells. Moreover, adiponectin-upregulated cPLA2 and COX-2 expression was significantly abrogated by ROS scavenger ( N-acetylcysteine) or the inhibitors of NADPH oxidase (apocynin), mitochondrial complex I (rotenone), PKC (Ro31-8220, Gö-6976, and rottlerin), and p300 (garcinol). Briefly, we reported that adiponectin stimulated cPLA2 and COX-2 expression via AdipoR1/2-dependent activation of PKC/NADPH oxidase/mitochondria resulting in ROS accumulation, p300 phosphorylation, and histone H4 acetylation. These results suggested that adiponectin promoted lung inflammation, resulting in exacerbation of pulmonary diseases via upregulating cPLA2 and COX-2 expression together with intracellular ROS production. Understanding the adiponectin signaling pathways on regulating cPLA2 and COX-2 may help develop therapeutic strategies on pulmonary diseases.

Background: Interactions and early warning effects of non-hepatic alkaline phosphatase (NHALP) and high-sensitivity C-reactive protein (hs-CRP) on the progression of vertebral fractures (VFs) in patients with rheumatoid arthritis (RA) remain unclear. We aim to explore whether serum concentrations of NHALP and hs-CRP could serve as a promising dual biomarker for prognostic assessment of VF progression. Methods: Unadjusted and adjusted hazard ratios (aHRs) of VF progression were calculated for different categories of serum NHALP and hs-CRP using the Cox regression model in RA patients. The modification effect between serum NHALP and hs-CRP on VF progression was determined using an interaction product term. Results: During 4489 person-years of follow-up, higher NHALP (&gt, 125 U/L) and hs-CRP (&gt, 3.0 mg/L) were robustly associated with incremental risks of VF progression in RA patients (aHR: 2.2 (95% confidence intervals (CIs): 1.2&ndash, 3.9) and 2.0 (95% CI: 1.3&ndash, 3.3) compared to the lowest HR category, respectively). The interaction between NHALP and hs-CRP on VF progression was statistically significant (p &lt, 0.05). In the stratified analysis, patients with combined highest NHALP and hs-CRP had the greatest risk of VF progression (aHR: 4.9 (95% CI: 2.5&ndash, 9.6)) compared to the lowest HR group (NHALP &lt, 90 U/L and hs-CRP &lt, 1 mg/L). Conclusions: In light of underdiagnoses of VFs and misleading diagnosis by single test, NHALP and hs-CRP could serve as compensatory biomarkers to predict subclinical VF progression in RA patients.

Background: Interactions and joint effects of galectin-3 and vascular cell adhesion molecule 1 (VCAM-1) on risks of all-cause and cardiovascular (CV) mortality remain unclear in patients with maintenance hemodialysis (MHD). Methods: Unadjusted and adjusted hazard ratios (aHRs) of mortality risks were analyzed between higher and lower concentration groups of serum galectin-3 and VCAM-1. The modification effect between serum galectin-3 and VCAM-1 on mortality risk was investigated using an interaction product term. Results: During follow-up, galectin-3 and VCAM-1 were associated with incremental risks of all-cause mortality (aHR: 1.038 (95% confidence interval (CI): 1.001&ndash, 1.077) and 1.002 (95% CI: 1.001&ndash, 1.003), respectively). Nonetheless, VCAM-1 but not galectin-3 predicted CV mortality (aHR: 1.043 (95% CI: 0.993&ndash, 1.096) and 1.002 (95% CI: 1.001&ndash, 1.003), respectively). In the interaction analysis, patients with combined higher galectin-3 (&gt, 29.5 ng/mL) and VCAM-1 (&gt, 1546.9 ng/mL) were at the greatest risk of all-cause and CV mortality (aHR: 4.6 (95% CI: 1.6&ndash, 13.4), and 4.2 (95% CI: 1.3&ndash, 14.4), respectively). The interactions between galectin-3 and VCAM-1 with respect to all-cause and CV mortality were statistically significant (p &lt, 0.01 and &lt, 0.05, respectively). Conclusion: Galectin-3 and VCAM-1 could serve as a promising dual biomarker for prognostic assessment, considering their joint effects on pathogenesis of leukocyte trafficking and atherothrombosis.

Molecular mechanisms and pathological features of p-Cresyl sulfate (PCS)-induced uremic lung injury (ULI) in chronic kidney disease (CKD) remain unclear. We analyzed pleural effusions (PE) from CKD and non-CKD patients for uremic toxins, reactive oxygen species (ROS), and chemotactic cytokines. Correlations between PE biomarkers and serum creatinine were also studied. Cell viability and inflammatory signaling pathways were investigated in PCS-treated human alveolar cell model. To mimic human diseases, CKD-ULI mouse model was developed with quantitative comparison of immunostaining and morphometric approach. PE from CKD patients enhance expressions of uremic toxins, hydroxyl radicals, and IL-5/IL-6/IL-8/IL-10/IL-13/ENA-78/GRO &alpha, /MDC/thrombopoietin/VEGF. PE concentrations of ENA-78/VEGF/IL-8/MDC/PCS/indoxyl sulphate correlate with serum creatinine concentrations. In vitro, PCS promotes alveolar cell death, cPLA2/COX-2/aquaporin-4 expression, and NADPH oxidase/mitochondria activation-related ROS. Intracellular ROS is abrogated by non-specific ROS scavenger N-acetyl cysteine (NAC), inhibitors of NADPH oxidase and mitochondria-targeted superoxide scavenger. However, only NAC protects against PCS-induced cell death. In vivo, expressions of cPLA2/COX2/8-OHdG, resident alveolar macrophages, recruited leukocytes, alveolar space, interstitial edema and capillary leakage increase in lung tissues of CKD-ULI mice, and NAC pretreatment ameliorates alveolar&ndash, capillary injury. PCS causes alveolar&ndash, capillary injury through triggering intracellular ROS, downstream prostaglandin pathways, cell death, and activating leukocytes to release multiplex chemoattractants and extracellular ROS. Thus PCS and nonspecific ROS serve as potential therapeutic targets.

Staphylococcus aureus (S. aureus) can often lead to many life-threatening diseases. It has the ability to invade normal endovascular tissue. Acute inflammation and its resolution are important to ensure bacterial clearance and limit tissue injury. Carbon monoxide (CO) has been shown to exert anti-inflammatory effects in various tissues and organ systems. In our study, we investigated the effects and the mechanisms of carbon monoxide releasing molecule-2 (CORM-2) on S. aureus-induced inflammatory responses in human aortic endothelial cells (HAECs). We proved that S. aureus induced cyclooxygenase-2 (COX-2)/prostaglandin E2 (PGE2)/interleukin-6 (IL-6)/matrix metallopeptidase-9 (MMP-9) expression and cell migration, which were decreased by CORM-2. Moreover, CORM-2 had no effects on TLR2 mRNA levels in response to S. aureus. Interestingly, we proved that S. aureus decreased intracellular ROS generation, suggesting that the inhibition of ROS further promoted inflammatory responses. However, CORM-2 significantly inhibited S. aureus-induced inflammation by increasing intracellular ROS generation. S. aureus-induced NF-κB activation was also inhibited by CORM-2. Finally, we proved that S. aureus induced levels of the biomarkers of inflammation in cardiovascular diseases, which were inhibited by CORM-2. Taken together, these results suggest that CORM-2 inhibits S. aureus-induced COX-2/PGE2/IL-6/MMP-9 expression and aorta inflammatory responses by increasing the ROS generation and reducing the inflammatory molecules levels.

Staphylococcus aureus (S. aureus) can lead to many life-threatening diseases. It has the ability to invade normal endovascular tissue. The molecular mechanisms and pathological changes of endothelial cells after S. aureus infection are of interest, but the basic understanding of how S. aureus destroys this barrier is not clear. Here, we showed that S. aureus enhanced COX-2 expression and prostaglandin E

Hyperplasia or hypertrophy of adipose tissues plays a crucial role in obesity, which is accompanied by the release of leptin. Recently, obesity was determined to be associated with various pulmonary diseases including asthma, acute lung injury, and chronic obstructive pulmonary disease. However, how obesity contributes to pulmonary diseases and whether leptin directly regulates lung inflammation remains unclear. We used cell and animal models to study the mechanisms of leptin mediation of pulmonary inflammation. We found that leptin activated de novo synthesis of cytosolic phospholipase A2-α (cPLA2-α) in vitro in the lung alveolar type II cells, A549, and in vivo in ICR mice. Upregulated cPLA2-α protein was attenuated by pretreatment with an OB-R blocking antibody, U0126, SB202190, SP600125, Bay11-7086, garcinol, and p300 siRNA, suggesting roles of p42/p44 MAPK, p38 MAPK, JNK1/2, NF-κB, and p300 in leptin effects. Leptin enhanced the activities of p42/p44 MAPK, p38 MAPK, JNK1/2, and p65 NF-κB in a time-dependent manner. Additional studies have suggested the participation of OB-R, p42/p44 MAPK, and JNK1/2 in leptin-increased p65 phosphorylation. Furthermore, p300 phosphorylation and histone H4 acetylation were reduced by blockage of OB-R, p42/p44 MAPK, p38 MAPK, JNK1/2, and NF-κB in leptin-stimulated cells. Similarly, blockage of the MAPKs/NF-κB/p300 cascade significantly inhibited leptin-mediated cPLA2-α mRNA expression. Our data as a whole showed that leptin contributed to lung cPLA2-α expression through OB-R-dependent activation of the MAPKs/NF-κB/p300 cascade.

Experimental and clinical studies have pointed out the lipid-induced renal damage, and statins may have pleiotropic effects on renoprotection. We reported a girl with X-linked Alport syndrome whose late steroid-responsive nephrotic syndrome (NS) was resolved by atorvastatin. She had been in a nephrotic condition despite of prednisone therapy 60 mg/day for 8 weeks. Renal biopsy dispicted extreme foamy appearance of tubular epithelial cells with detachment led to luminal obliteration. Atorvastatin was started on the ninth week of prednisone therapy due to severe hypercholesterolemia. Partial remission of NS was dramatically achieved with unchanged dosage of prednisone at the end of the twelfth week. Our case provides a pathology-based evidence to support the use of statins in profoundly hyperlipidemic patients with NS. In patients with NS and profound hyperlipidemia, early initiation of statin therapy is required in combination with immunosuppressive therapy.

Haem oxygenase-1 (HO-1) could provide cytoprotection against various inflammatory diseases. However, the mechanisms underlying the protective effect of CO-releasing molecule-2 (CORM-2)-induced HO-1 expression against TNF-α-induced inflammatory responses in human pulmonary alveolar epithelial cells (HPAEpiCs) remain unknown.CORM-2-induced HO-1 protein and mRNA expression, and signalling pathways were determined by Western blot and real-time PCR, coupled with respective pharmacological inhibitors or transfection with siRNAs. The effect of CORM-2 on TNF-α-induced increase in leukocyte counts in BAL fluid and VCAM-1 expression in lung was determined by cell counting and Western blot analysis.CORM-2 attenuated the TNF-α-induced pulmonary haematoma, VCAM-1 expression and increase in leukocytes through an up-regulation of HO-1 in mice; this effect of CORM-2 was reversed by the HO-1 inhibitor zinc protoporphyrin IX. Furthermore, CORM-2 increased HO-1 protein and mRNA expression as well as the phosphorylation of PYK2, PKCα and ERK1/2 (p44/p42 MAPK) in HPAEpiCs; these effects were attenuated by their respective pharmacological inhibitors or transfection with siRNAs. Inhibition of PKCα by Gö6976 or Gö6983 attenuated CORM-2-induced stimulation of PKCα and ERK1/2 phosphorylation but had no effect on PYK2 phosphorylation. Moreover, inhibition of PYK2 by PF431396 reduced the phosphorylation of all three protein kinases. Finally, PYK2/PKCα/ERK1/2-mediated stimulation of activator protein 1 was shown to play a key role in CORM-2-induced HO-1 expression via an up-regulation of c-Fos mRNA.CORM-2 activates a PYK2/PKCα/ERK1/2/AP-1 pathway leading to HO-1 expression in HPAEpiCs. This HO-1/CO system might have potential as a therapeutic target in pulmonary inflammation.

Up-regulation of cyclooxygenase (COX)-2 and prostaglandin E2 (PGE2) are implicated in lung inflammation. Adenosine 5'-triphosphate (ATP) has been shown to act via activation of P2 purinoceptors, leading to COX-2 expression in various inflammatory diseases. The mechanisms of ATP-induced COX-2 expression and PGE2 release remain unclear. We showed that pretreatment with the inhibitors of P2 receptors (PPADS and Suramin), Gq protein (GPA2A), phosphatidylcholine-phospholipase C (PC-PLC; D609), phosphoinositide-phospholipase C (PI-PLC; ET-18-OCH3), Ca(2+)/calmodulin-dependent protein kinase II (CaMKII; KN62), protein kinase C (PKC; Gö6976, Ro-318220, GF109203X, and rottlerin), MEK1/2 (PD98059), p38 MAPK (SB202190), and nuclear factor-kappaB (NF-κB; Bay11-7082) and the intracellular calcium chelator (BAPTA/AM) or transfection with siRNAs of these molecules and cPLA2 reduced ATPγS-induced COX-2 expression or PGE2 production in A549 cells. In addition, ATPγS-induced elevation of intracellular Ca(2+) concentration was attenuated by PPADS, Suramin, D609, or ET-18-OCH3. ATPγS-induced p38 MAPK, p42/p44 MAPK, and NF-κB p65 activation were inhibited by Gö6976, Ro-318220, GF109203X, or rottlerin. ATPγS also induced cPLA2 phosphorylation and activity, which were reduced via inhibition of P2 receptors, PKCs, p38 MAPK, and p42/p44 MAPK. ATPγS-induced cPLA2 expression was inhibited by SB202190, PD98059, or Bay11-7082. In the in vitro study, we established that ATPγS induced PGE2 generation via a cPLA2/COX-2-dependent pathway. In the in vivo study, we found that ATPγS induced COX-2 mRNA expression in the lungs and leukocyte (mainly eosinophils and neutrophils) count in bronchoalveolar lavage (BAL) fluid in mice via a P2 receptors-dependent signaling pathway. We concluded that ATPγS may induce lung inflammation via a cPLA2/COX-2/PGE2-dependent pathway.

Tumor necrosis factor-alpha (TNF-alpha) triggers activation of cytosolic phospholipase A2 (cPLA2) and then enhancing the synthesis of prostaglandin (PG) in inflammatory diseases. However, the detailed mechanisms of TNF-alpha induced cPLA2 expression were not fully defined in human pulmonary alveolar epithelial cells (HPAEpiCs). We found that TNF-alpha-stimulated increases in cPLA2 mRNA (5.2 folds) and protein (3.9 folds) expression, promoter activity (4.3 folds), and PGE2 secretion (4.7 folds) in HPAEpiCs, determined by Western blot, real-time PCR, promoter activity assay and PGE2 ELISA kit. These TNF-alpha-mediated responses were abrogated by the inhibitors of NADPH oxidase [apocynin (APO) and diphenyleneiodonium chloride (DPI)], ROS [N-acetyl cysteine, (NAC)], NF-kappaB (Bay11-7082) and transfection with siRNA of ASK1, p47phox, TRAF2, NIK, IKKalpha, IKKbeta, or p65. TNF-alpha markedly stimulated NADPH oxidase activation and ROS including superoxide and hydrogen peroxide production which were inhibited by pretreatment with a TNFR1 neutralizing antibody, APO, DPI or transfection with siRNA of TRAF2, ASK1, or p47phox. In addition, TNF-alpha also stimulated p47phox phosphorylation and translocation in a time-dependent manner. On the other hand, TNF-alpha induced TNFR1, TRAF2, ASK1, and p47phox complex formation in HPAEpiCs, which were attenuated by a TNF-alpha neutralizing antibody. We found that pretreatment with NAC, DPI, or APO also attenuated the TNF-alpha-stimulated IKKalpha/beta and NF-kappaB p65 phosphorylation, NF-kappaB (p65) translocation, and NF-kappaB promoter activity in HPAEpiCs. Finally, we observed that TNF-alpha-stimulated NADPH oxidase activation and ROS generation activates NF-kappaB through the NIK/IKKalpha/beta pathway. Taken together, our results demonstrated that in HPAEpiCs, up-regulation of cPLA2 by TNF-alpha is, at least in part, mediated through the cooperation of TNFR1, TRAF2, ASK1, and NADPH oxidase leading to ROS generation and ultimately activates NF-kappaB pathway.

The d-galactose (d-gal)-injected animal model, which is typically established by administering consecutive subcutaneous d-gal injections to animals for approximately six or eight weeks, has been frequently used for aging research. In addition, this animal model has been demonstrated to accelerate aging in the brain, kidneys, liver and blood cells. However, studies on aging in male reproductive organs that have used this animal model remain few. Therefore, the current study aimed to optimize a model of male reproductive aging by administering d-gal injections to male mice and to determine the possible mechanism expediting senescence processes during spermatogenesis. In this study, C57Bl/6 mice were randomized into five groups (each containing 8–10 mice according to the daily intraperitoneal injection of vehicle control or 100 or 200 mg/kg dosages of d-gal for a period of six or eight weeks). First, mice subjected to d-gal injections for six or eight weeks demonstrated considerably decreased superoxide dismutase activity in the serum and testis lysates compared to those in the control group. The lipid peroxidation in testis also increased in the d-gal-injected groups. Furthermore, the d-gal-injected groups exhibited a decreased ratio of testis weight/body weight and sperm count compared to the control group. The percentages of both immotile sperm and abnormal sperm increased considerably in the d-gal-injected groups compared to those of the control group. To determine the genes influenced by the d-gal injection during murine spermatogenesis, a c-DNA microarray was conducted to compare testicular RNA samples between the treated groups and the control group. The d-gal-injected groups exhibited RNA transcripts of nine spermatogenesis-related genes (Cycl2, Hk1, Pltp, Utp3, Cabyr, Zpbp2, Speer2, Csnka2ip and Katnb1) that were up- or down-regulated by at least two-fold compared to the control group. Several of these genes are critical for forming sperm-head morphologies or maintaining nuclear integration (e.g., cylicin, basic protein of sperm head cytoskeleton 2 (Cylc2), casein kinase 2, alpha prime interacting protein (Csnka2ip) and katanin p80 (WD40-containing) subunit B1 (Katnb1)). These results indicate that d-gal-injected mice are suitable for investigating male reproductive aging.

BACKGROUND AND PURPOSE Lipopolysaccharide (LPS)-induced expression of cyclooxygenase-2 (COX-2) and cytosolic phospholipase A(2) (cPLA(2) ) has been implicated in several respiratory diseases. HuR is known to enhance the expression of genes by binding to 3'-untranslated region (3'-UTR) of mRNA and stabilizing mRNA. However, the exact mechanisms by which HuR affects the stability of mRNA and modulates LPS-induced COX-2 and cPLA(2) expression in human tracheal smooth muscle cells (HTSMCs) are not known. EXPERIMENTAL APPROACH The expression of prostaglandin E(2) (PGE(2) ) was measured by ELISA, and pro-inflammatory proteins were determined by use of a promoter assay, PCR or Western blot analysis. Overexpression of siRNAs to knock down the target components was used to manipulate the expression of HuR. Release of reactive oxygen species (ROS) was detected by fluorescence dye. The activation of signalling components was assessed by comparing phosphorylation levels, localization of protein kinases or coimmunoprecipitation assay. KEY RESULTS LPS induced COX-2 and cPLA(2) expression via post-translational regulation of mRNA stabilization, which were attenuated by transfection with HuR siRNA in HTSMCs. In addition, LPS-stimulated NADPH oxidase activation and ROS generation were attenuated by the NADPH oxidase inhibitors diphenyleneiodonium chloride (DPI) and apocynin (APO). Generation of ROS induced phosphorylation of p42/p44 mitogen-activated protein kinase (MAPK), p38 MAPK and JNK1/2, which was attenuated by DPI and APO and the ROS scavenger N-acetylcysteine. CONCLUSIONS AND IMPLICATIONS These results suggested that in HTSMCs, LPS-induced COX-2 and cPLA(2) expression is mediated through NADPH oxidase/ROS-dependent MAPKs associated with HuR accumulation in the cytoplasm. Activated MAPKs may regulate the nucleocytoplasmic shuttling of HuR, and thus induce the cytoplasmic accumulation of HuR.

A remarkable synergetic effect between the multi-graphene platelets (MGPs) and multi-walled carbon nanotubes (MWCNTs) in improving the mechanical properties and thermal conductivity of epoxy composites is demonstrated. Stacking of individual two-dimensional MGPs is effectively inhibited by introducing one-dimensional MWCNTs. Long and tortuous MWCNTs can bridge adjacent MGPs and inhibit their aggregation, resulting in a high contact area between the MGP/MWCNT structures and the polymer matrix. Scanning electron microscope images of the fracture surfaces of the epoxy matrix showed that MWCNT/MGP hybrid nanofillers exhibited higher solubility and better compatibility than individual MWCNTs and MGPs did. The tensile strength of GD400-MWCNT/MGP/epoxy composites was 35.4% higher than that of the epoxy alone, compared to only a 0.9% increase in tensile strength for MGP/epoxy composites over the epoxy compound. Thermal conductivity increased by 146.9% using GD400-MWCNT/MGP hybrid fillers and 23.9% for MGP fillers, compared to non-derivatised epoxy.