Nuclear factor-kappaB Signaling Pathway Research Articles

Ubiquitin-specific protease 4 predicts an unfavorable prognosis and promotes malignant behaviors in vitro in pancreatic cancer

Ubiquitin-specific protease 4 (USP4), has been reported to participate in the progression of various cancers due to its role in post-translational modulation. However, the prognostic significance and mechanism of USP4 in pancreatic cancer (PC) have not been well elucidated before. In the present study, we found that USP4 expression was higher in PC tissues than that in adjacent normal tissues and PC patients with high level of USP4 expression have a poor prognosis via immunohistochemistry and bioinformatics analyses. In vitro study showed that knockdown of USP4 inhibited PC cells proliferation, migration and invasion. Mechanistically, USP4 can activate nuclear factor kappa-B signaling pathway via stabilizing TNF receptor associated factor 6 at its protein level to promote the ability of proliferation, migration and invasion of PC cells. The results of this study revealed that USP4 plays a tumor-promoting role in PC and can be used as a prognostic indicator and therapeutic target for patients with resected PC.

Amorphous silica nanoparticles induce inflammation via activation of NLRP3 inflammasome and HMGB1/TLR4/MYD88/NF-kb signaling pathway in HUVEC cells

Silica nanoparticles (SiO2 NPs) are extensively applied in various field, which increased their health risks to humans. SiO2 NPs were reported to enter into blood through inhalation and meanwhile, the potential use of SiO2 NPs as drug carriers in vivo allows them to present in blood circulation to induce inflammation of vascular endothelial cells which can be closely related with cardiovascular diseases, whilst the intrinsic mechanism has not been well understood. In this study, we found a regulation of signal axis induced by amorphous SiO2 NPs that triggers pro-inflammatory responses in human umbilical vein endothelial cells (HUVECs). HUVECs exposed with SiO2 NPs generate excess amount of reactive oxygen species (ROS) and lactate dehydrogenase (LDH), together with the up-regulation of cell inflammatory factors [interleukin-1 beta (IL-1β), interleukin-6 (IL-6), tumor necrotic factor-α (TNF-α)] and cell adhesion molecules [intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1)]. In addition, SiO2 NPs were found to promote the translocation and release of high-mobility group box 1 (HMGB1) from nucleus to cytoplasm, which was demonstrated to be regulated by ROS and NOD-like receptor pyrin domain containing 3 (NLRP3) inflammasome. Subsequently, toll-like receptor 4 (TLR4) could bind with HMGB1, up-regulate the expression of myeloid differentiation factor 88 (MyD88) and then activate nuclear factor kappa-B (NF-κB) signaling pathway, ultimately induced the inflammatory response of HUVECs. Overall, out results revealed the related signaling pathways of cell inflammation induced by amorphous SiO2 NPs, which provided new insights in understanding SiO2 NPs-induced cytotoxicity and offered safety guidance for further nanomaterial application.

Aβ40 Promotes the Osteoblastic Differentiation of Aortic Valve Interstitial Cells through the RAGE Pathway.

Amyloid beta (Aβ) peptide 40 enhances the activation of receptor for advanced glycation end products (RAGE) in immune-inflammatory diseases. RAGE exhibits several effects in the setting of numerous cardiovascular events. We hypothesized that the Aβ40/RAGE pathway is involved in the osteoblastic differentiation of the valvular interstitial cell (VIC) phenotype, and RAGE knockout intervention could reduce the calcification of aortic valve interstitial cells (AVICs) by inhibiting the extracellular-regulated kinase1/2 (ERK1/2)/nuclear factor kappa-B (NF-κB) signaling pathway. To test this hypothesis, the activation of Aβ40/RAGE pathway in human calcific AVs was evaluated with immunohistochemical staining. Cultured calcific VIC models were used in vitro. The VICs were stimulated using Aβ40, with or without RAGE small interfering ribonucleic acid (siRNA), and ERK1/2 and NF-κB inhibitors for analysis. Our data revealed that Aβ40 induced the ERK1/2/NF-κB signaling pathway and osteoblastic differentiation of AVICs via the RAGE pathway in vitro.

Interleukin-6 promotes migration and extracellular matrix synthesis in retinal pigment epithelial cells.

Proliferative vitreoretinopathy (PVR) is the most common cause of surgical failure in the rhegmatogenous retinal detachment (RD) treatment. Retinal pigment epithelial (RPE) cell proliferation, migration, and the synthesis of extracellular matrix (ECM) are intrinsic to the formation of a PVR membrane. High level of interleukin-6 (IL-6) has been found in the vitreous of PVR patients, while the role of IL-6 in RPE cells remaining further characterized. In the present study, we evaluated the potential regulatory effects of IL-6 on cell migration, ECM components, and transforming growth factor β2 (TGF-β2) expression in RPE cells. Furthermore, cell counting kit-8 (CCK‑8) assay was used to investigate cell proliferation activity. We found that IL-6 promoted fibronectin (Fn) and type I collagen (COL-1), TGF-β2 expression in RPE cells, also stimulate RPE cell migration effectively. Moreover, the induction of IL-6 activated the Janus kinase/signal transducers and activators of transcription (JAK/STAT3) and the nuclear factor kappa-B (NF-κB) signaling pathways significantly. Simultaneously, both JAK/STAT3 and NF-κB pathways inhibitors, WP1066 and BAY11-7082, alleviated IL-6-induced biological effects, respectively. However, it was noted that IL-6 had little effect on α-smooth muscle actin (α-SMA) expression. Collectively, our results reveal that IL-6 promotes RPE cell migration and ECM synthesis via activating JAK/STAT3 and NF-κB signaling pathways, which may play a crucial role in PVR formation.

LncRNA MALAT1 Enhances ox-LDL-Induced Autophagy through the SIRT1/MAPK/NF-κB Pathway in Macrophages.

Atherosclerosis is the main cause of cardiovascular and cerebrovascular diseases. In advanced atherosclerotic plaque, macrophage apoptosis coupled with inflammatory cytokine secretion promotes the formation of necrotic cores. It has also been demonstrated that the long-noncoding Ribonucleic Acid (lnc RNA) metastasis-associated lung adenocarcinoma transcript 1 (MALAT1), with its potent function on gene transcription modulation, maintains oxidized low-density lipoprotein (ox-LDL)- induced macrophage autophagy (i.e., helps with cholesterol efflux). It also showed that MALAT1 activated Sirtuin 1 (SIRT1), which subsequently inhibited the mitogen-activated protein kinase (MAPK) and nuclear factor kappa-B (NF-κB) signaling pathways. ox-LDL has been used to incubate human myeloid leukemia mononuclear cells (THP-1)-derived macrophages to establish an in vitro foam cell model. Quantitative reverse-transcription polymerase chain reaction and Western blot analyses confirmed the increased expression level of MALAT1 and the autophagy-related protein Microtubuleassociated protein light chain 3 (LC-3), beclin-1. The small interfering RNA study showed a significant decrease in autophagy activity and an increase in apoptotic rate when knocking down MALAT1. Further study demonstrated that MALAT1 inhibited the expression of MAPK and NF-κB (p65) by upregulating SIRT1.

Expression of p52, a non-canonical NF-kappaB transcription factor, is associated with poor ovarian cancer prognosis

BackgroundThe canonical and non-canonical nuclear factor-kappaB (NF-κB) signaling pathways have key roles in cancer, but studies have previously evaluated only the association of canonical transcription factors and ovarian cancer survival. Although a number of in vitro and in vivo studies have demonstrated mechanisms by which non-canonical NF-κB signaling potentially contributes to ovarian cancer progression, a prognostic association has yet to be shown in the clinical context.MethodsWe assayed p65 and p52 (major components of the canonical and non-canonical NF-κB pathways) by immunohistochemistry in epithelial ovarian tumor samples; nuclear and cytoplasmic staining were semi-quantified by H-scores and dichotomized at median values. Associations of p65 and p52 with progression-free survival (PFS) and overall survival (OS) were quantified by Hazard Ratios (HR) from proportional-hazards regression.ResultsAmong 196 cases, median p52 and p65 H-scores were higher in high-grade serous cancers. Multivariable regression models indicated that higher p52 was associated with higher hazards of disease progression (cytoplasmic HR: 1.54; nuclear HR: 1.67) and death (cytoplasmic HR: 1.53; nuclear HR: 1.49), while higher nuclear p65 was associated with only a higher hazard of disease progression (HR: 1.40) in unadjusted models. When cytoplasmic and nuclear staining were combined, p52 remained significantly associated with increased hazards of disease progression (HR: 1.91, p = 0.004) and death (HR: 1.70, p = 0.021), even after adjustment for p65 and in analyses among only high-grade serous tumors.ConclusionsThis is the first study to demonstrate that p52, a major component of non-canonical NF-κB signaling, may be an independent prognostic factor for epithelial ovarian cancer, particularly high-grade serous ovarian cancer. Approaches to inhibit non-canonical NF-κB signaling should be explored as novel ovarian cancer therapies are needed.

Tripartite Motif-Containing 46 Promotes Viability and Inhibits Apoptosis of Osteosarcoma Cells by Activating NF-B Signaling Through Ubiquitination of PPAR.

Osteosarcoma (OS), the most common bone cancer, causes high morbidity in children and young adults. TRIM46 is a member of the family of tripartite motif (TRIM)-containing proteins that serve as important regulators of tumorigenesis. Here we investigate the possible role of TRIM46 in OS and the underlying molecular mechanism. We report an increase in the expression of TRIM46 in OS and its association with tumor size, Enneking’s stage, and patient prognosis. TRIM46 knockdown inhibits OS cell viability and cell cycle progression and induces apoptosis, while TRIM46 overexpression exerts inverse effects, which are inhibited by peroxisome proliferator-activated receptor alpha (PPARα) overexpression and the nuclear factor kappa B (NF-κB) inhibitor, pyrrolidine dithiocarbamate (PDTC). Furthermore, TRIM46 negatively regulates PPARα expression via ubiquitination-mediated protein degradation and modification. PPARα overexpression also inactivates NF-κB signaling and NF-κB promoter activity in OS cells overexpressing TRIM46. Moreover, TRIM46 knockdown inhibits tumor growth and induces apoptosis of OS cells in vivo. TRIM46 acts as an oncogene in OS by interacting with and ubiquitinating PPARα, resulting in the activation of NF-κB signaling pathway. Thus, TRIM46 may be a potential biomarker of carcinogenesis.

Anti-Inflammatory Effect of Simonsinol on Lipopolysaccharide Stimulated RAW264.7 Cells through Inactivation of NF-κB Signaling Pathway.

Simonsinol is a natural sesqui-neolignan firstly isolated from the bark of Illicium simonsii. In this study, the anti-inflammatory activity of simonsinol was investigated with a lipopolysaccharide (LPS)-stimulated murine macrophages RAW264.7 cells model. The results demonstrated that simonsinol could antagonize the effect of LPS on morphological changes of RAW264.7 cells, and decrease the production of nitric oxide (NO), tumor necrosis factor α (TNF-α), and interleukin 6 (IL-6) in LPS-stimulated RAW264.7 cells, as determined by Griess assay and enzyme-linked immunosorbent assay (ELISA). Furthermore, simonsinol could downregulate transcription of inducible nitric oxide synthase (iNOS), TNF-α, and IL-6 as measured by reverse transcription polymerase chain reaction (RT-PCR), and inhibit phosphorylation of the alpha inhibitor of NF-κB (IκBα) as assayed by Western blot. In conclusion, these data demonstrate that simonsinol could inhibit inflammation response in LPS-stimulated RAW264.7 cells through the inactivation of the nuclear transcription factor kappa-B (NF-κB) signaling pathway.

Diclofenac Inhibits Phorbol Ester-Induced Gene Expression and Production of MUC5AC Mucin via Affecting Degradation of IkBα and Translocation of NF-kB p65 in NCI-H292 Cells

In this study, diclofenac, a non-steroidal anti-inflammatory drug, was investigated for its potential effect on the gene expression and production of airway MUC5AC mucin. The human respiratory epithelial NCI-H292 cells were pretreated with diclofenac for 30 min and stimulated with phorbol 12-myristate 13-acetate (PMA), for the following 24 h. The effect of diclofenac on PMA-induced nuclear factor kappa B (NF-kB) signaling pathway was also investigated. Diclofenac suppressed the production and gene expression of MUC5AC mucins, induced by PMA through the inhibition of degradation of inhibitory kappa Bα (IkBα) and NF-kB p65 nuclear translocation. These results suggest diclofenac regulates the gene expression and production of mucin through regulation of NF-kB signaling pathway, in human airway epithelial cells.

MicroRNA-212-5p regulates the inflammatory response of periodontal ligament cells by targeting myeloid differentiation factor 88

ObjectiveThis study was aimed to investigate the effects of microRNA-212-5p (miR-212-5p) and myeloid differentiation factor 88 (Myd88) in Porphyromonas gingivalis lipopolysaccharide (P. gingivalis LPS)-challenged human periodontal ligament cells (PDLCs). MethodsThe levels of miR-212-5p and Myd88 in PDLCs were determined via quantitative real-time Polymerase Chain Reaction. Cell viability and pro-inflammatory cytokines were assessed using MTT assay and enzyme-linked immunosorbent assay. Bioinformatics and luciferase reporter gene assay were employed to explore the interaction between miR-212-5p and Myd88. Moreover, the activation of p38 mitogen-activated protein kinase (MAPK) and nuclear factor kappa-B signal pathways were testified via western blot analysis. ResultsMiR-212-5p was downregulated following stimulation of P. gingivalis LPS. Overexpression of miR-212-5p elevated the cell viability and reduced the secretion of pro-inflammatory cytokines. Myd88 is a target of miR-212-5p. Notably, pcDNA3.1-Myd88 reversed the anti-inflammatory effect of miR-212-5p overexpression. Likewise, miR-212-5p mimic inhibited the activation of p38 MAPK and nuclear factor kappa-B, whereas pcDNA3.1-Myd88 abrogated the effect of miR-212-5p mimic. ConclusionsMiR-212-5p inhibited the inflammatory response in PDLCs by targeting Myd88, which may involve the inactivation of p38 MAPK and nuclear factor kappa-B.

Abstract A15: Noncanonical NF-kappaB signaling is associated with poor ovarian cancer prognosis

Abstract Objective: Mediated by a family of transcription factors, the canonical and noncanonical nuclear factor-kappaB (NF-kB) signaling pathways have key roles in inflammation, innate immunity, and cancer development and progression. As prior ovarian cancer studies have evaluated only canonical NF-kB transcription factors in relation to survival, this study was undertaken to evaluate both canonical and noncanonical NF-kB pathway components in tumor samples from epithelial ovarian cancer cases from the Vanderbilt Tissue Repository for Ovarian Cancer (TROC). Methods: We abstracted clinical data from electronic medical records and assayed p65 (representing the canonical NF-kB pathway) and p52 (representing the noncanonical NF-kB pathway) by immunohistochemistry for 197 TROC samples. Nuclear and cytoplasmic staining in tumor cells was semiquantified by H-scores and dichotomized at median values. Associations with progression-free survival (PFS) and overall survival (OS) were quantified by hazard ratios (HR) and 95% confidence intervals (CI) using Cox proportional-hazards regression. Results: Median cytoplasmic and nuclear p65 and p52 H-scores were significantly higher among high-grade and serous cases; median p52 H-scores were also significantly higher among late-stage cases and patients treated with chemotherapy (all Wilcoxon rank sum p<0.05). In unadjusted regression models, higher nuclear p65 and higher nuclear p52 were associated with approximately 40% and 70% shorter PFS and OS, respectively, but these associations were attenuated by adjustment for age, stage, tumor type, race, chemotherapy, and residual disease. In multivariable adjusted models, higher cytoplasmic p52 was associated with more than 50% shorter PFS (HR 1.68, 95% CI 1.18-2.39) and OS (HR 1.53, 95% CI 1.06-2.21). Furthermore, when cytoplasmic and nuclear staining were combined, and mutually adjusted models included both p52 and p65, only p52 remained associated with worse PFS (HR 1.57, 95% CI 1.10-2.37) and OS (HR 1.51, 95% CI 1.05-2.15). Conclusions: This is the first study to demonstrate that p52, representing noncanonical NF-kB signaling, may be an independent prognostic factor for epithelial ovarian cancer. Approaches to inhibit noncanonical NF-kB signaling should be explored as novel therapies for ovarian cancer are needed. Citation Format: Demetra Hufnagel, Andrew J. Wilson, Jamie Saxon, Dineo Khabele, Timothy Blackwell, Marta A. Crispens, Fiona Yull, Alicia Beeghly-Fadiel. Noncanonical NF-kappaB signaling is associated with poor ovarian cancer prognosis [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr A15.

Biomimetic osteogenic peptide with mussel adhesion and osteoimmunomodulatory functions to ameliorate interfacial osseointegration under chronic inflammation

Bone endoprosthesis in patients with systemic chronic inflammation frequently leads to poor osseointegration after implantation mainly due to the increase in pro-inflammatory cytokines that induce bone resorption and impair bone formation. In this work, peptide-coated implants are designed to create a beneficial bone immune microenvironment around prostheses to promote interfacial osteogenesis. By taking advantage of the spontaneous and stable coordination chemistry, Ti-based implants are coated with the mussel-inspired peptide to mitigate lipopolysaccharide (LPS)-induced inflammation by up-regulating the M2 phenotype of macrophages. In addition, the peptide coating increases the bone-implant contact (BIC) by nearly 3 times resulting in suppressed osteoclastogenesis and promoted osteogenesis by inhibiting the nuclear factor kappa-B (NF-κB) signalling pathway. Our findings indicate that biomimetic peptides with osteoimmunomodulatory bioactivity can be incorporated into Ti-based prostheses to facilitate bone regeneration in patients with chronic inflammatory diseases.

Ziyuglycoside II Inhibits Rotavirus Induced Diarrhea Possibly via TLR4/NF-κB Pathways.

Rotavirus (RV) induced diarrhea has been a major reason affecting children healthy under 5 years old especially in developing countries. Although specific vaccines have preventive effects, antiviral therapy is essential for the diarrhea patients. Ziyuglycoside II is a traditional Chinese herb which has been proven to possess anti-virus effects. This study aimed to investigate the roles of Ziyuglycoside II in rotavirus-induced diarrhea and the underlying molecular mechanism. We found that normal MA104 cells treated with RV became swollen and gather together. However, Ziyuglycoside II treatment inhibited cell growth in a dose- and time dependent manner and suppressed RV replication. Moreover, Ziyuglycoside II reversed RV-induced downregulation of anti-inflammatory cytokine interleukin (IL)-10 and upregulation of pro-inflammatory factors, such as interferon-γ (IFN-γ), IL-1β, IL-6, and tumor necrosis factor (TNF-α). Moreover, Ziyuglycoside II administration and ribavirin blocked toll-like receptor 4 (TLR4)/nuclear factor kappa-B (NF-κB) signaling pathway both in mRNA and protein level, which was paralleled with immunohistochemical assay. Additionally, Ziyuglycoside II administration improved diarrhea symptoms and decreased diarrhea scores. Ziyuglycoside II and ribavirin inhibited the apoptosis of small intestine epithelial cells induced by RV. Taken together, RV treatment induced diarrhea. Ziyuglycoside II administration inhibited TLR4/NF-κB pathway and inflammatory response and improved RV-induced diarrhea. The inhibitory effects of Ziyuglycoside II on RV-induced diarrhea predicted Ziyuglycoside II may be a promising drug for diarrhea.

Dihydromyricetin attenuates Escherichia coli lipopolysaccharide-induced ileum injury in chickens by inhibiting NLRP3 inflammasome and TLR4/NF-\u03baB signalling pathway

Lipopolysaccharide (LPS) as a major component of Escherichia coli cell wall can cause inflammation and cell death. Dihydromyricetin (ampelopsin, DHM) is a natural flavonoid compound with anti-inflammatory, anti-oxidant and anti-bacterial effects. The preventive effects of DHM against ileum injury remain unclear. Here, we explored the protective role of DHM against LPS-induced ileum injury in chickens. In this study, DHM significantly attenuated LPS-induced alteration in diamine oxidase, malondialdehyde, reduced glutathione, glutathione peroxidase and superoxide dismutase levels in chicken plasma and ileum. Histology evaluation showed that the structure of blood vessels in ileum was seriously fragmented and presence of necrotic tissue in the lumen in the LPS group. Scanning electron microscopic observation revealed that the surface of the villi was rough and uneven, the structure was chaotic, and the normal finger shape was lost in the LPS group. In contrast, 0.05% and 0.1% DHM treatment partially alleviated the abnormal morphology. Additionally, DHM maintained the barrier function by restoring the protein expression of occludin, claudin-1 and zonula occludens protein-1. DHM inhibited apoptosis through the reduction of the expression of bax and caspase-3 and restored the expression of bcl-2. Importantly, DHM could reduce ileum NLR family pyrin domain-containing 3 (NLRP3), caspase-1, interleukin (IL)-1β and IL-18 expression to protect tissues from pyroptosis and inhibited toll-like receptor 4 (TLR4)/nuclear factor kappa-B (NF-κB) signalling pathway. In summary, DHM attenuated the ileum mucosal damage, oxidative stress and apoptosis, maintained barrier function, inhibited NLRP3 inflammasome and TLR4/NF-κB signalling pathway activation triggered by Escherichia coli LPS.

Downregulation of hsa_circ_0005243 induces trophoblast cell dysfunction and inflammation via the \u03b2-catenin and NF-\u03baB pathways

BackgroundGestational diabetes mellitus (GDM) is a common complication in pregnancy that poses a serious threat to the health of both mother and child. While the specific etiology and pathogenesis of this disease are not fully understood, it is thought to arise due to a combination of insulin resistance, inflammation, and genetic factors. Circular RNAs (circRNAs) are a special kind of non-coding RNA that have attracted significant attention in recent years due to their diverse activities, including a potential regulatory role in pregnancy-related diseases, such as GDM.MethodsWe previously reported the existence of a novel circRNA, hsa_circ_0005243, which was identified by RNA sequencing. In this study, we examined its expression in 20 pregnant women with GDM and 20 normal pregnant controls using quantitative reverse transcription PCR analysis. Subsequent in vitro experiments were conducted following hsa_circ_0005243 knockdown in HTR-8/SVneo cells to examine the role of hsa_circ_0005243 in cell proliferation and migration, as well as the secretion of inflammatory factors such as tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6). Finally, we examined the expression of β-catenin and nuclear factor kappa-B (NF-κB) signaling pathways to assess their role in GDM pathogenesis.ResultsExpression of hsa_circ_0005243 was significantly reduced in both the placenta and plasma of GDM patients. Knockdown of hsa_circ_0005243 in trophoblast cells significantly suppressed cell proliferation and migration ability. In addition, increased secretion of inflammatory factors (TNF-α and IL-6) was observed after hsa_circ_0005243 depletion. Further analyses showed that knockdown of hsa_circ_0005243 reduced the expression of β-catenin and increased nuclear NF-κB p65 nuclear translocation.ConclusionsDownregulation of hsa_circ_0005243 may be associated with the pathogenesis of GDM via the regulation of β-catenin and NF-κB signal pathways, suggesting a new potential therapeutic target for GDM.

Light-Switchable Yolk-Mesoporous Shell UCNPs@MgSiO3 for Nitric Oxide-Evoked Multidrug Resistance Reversal in Cancer Therapy.

Gas therapy has emerged as a forceful strategy for augmenting the effects of chemotherapeutic drugs against cancer cells. However, it remains extremely challenging to effectively deliver gas into tissues of interest and unravel its underlying mechanisms. Herein, we designed a near-infrared (NIR) light-switchable nitric oxide (NO) delivery nanosystem for high-efficacy multidrug resistance (MDR) reversal in cancer therapy based on a yolk-shell upconverting nanoparticles@magnesium silica (UCNP@MgSiO3). The internal hollow cavity and flower-like mesoporous shell of UCNPs@MgSiO3 not only enabled a significantly high encapsulation capacity for the NO precursor (BNN6) and doxorubicin (DOX) but also allowed the enhanced cellular uptake, resulting in NIR-triggered NO generation and low pH-triggered DOX release in cancer cells. Mechanistically, intracellular NO can downregulate the drug efflux-related P-glycoprotein and adenosine 5'-triphosphate-binding cassette transporters, thereby increasing the DOX accumulation in the cell nuclei. Such combination therapy of NO and DOX induced the apoptosis of MDR cells and completely inhibited in vivo MDR tumor growth. We further elucidated the therapy mechanism via proteomic profiling, showcasing the downregulation of the ubiquitin-proteasome pathway and nuclear factor kappa-B signaling pathway in the NO-treated MDR cells. Therefore, our findings develop a promising nanoscale gas/drug delivery paradigm for fighting MDR tumors and providing molecular insights into cancer therapy.

MON-721 Crude Protein of Pyropia Yezoensis Protects Against Tumor Necrosis Factor-á-Induced Myotube Atrophy by Regulating the Mitogen-Activated Protein Kinase and Nuclear Factor-Kappab Signaling Pathways in C2C12 Myotubes

Proinflammatory cytokines induce ubiquitin-proteasome-dependent proteolysis by activating intracellular factors in skeletal muscle, leading to muscle atrophy. Therefore, we investigated the protective effect of Pyropia yezoensis crude protein (PYCP) on tumor necrosis factor (TNF)-α-induced muscle atrophy in vitro. Mouse skeletal muscle C2C12 myotubes were treated for 48 h with TNF-α (20 ng/mL) in the presence or absence of PYCP (25, 50, and 100 μg/mL). PYCP at concentrations up to 100 μg/mL did not affect cell viability. Exposure to TNF-α for 48 h significantly decreased the diameter of myotubes, which was increased by treatment with 25, 50, and 100 μg/mL PYCP. PYCP inhibited TNF-α-induced intracellular reactive oxygen species accumulation in C2C12 myotubes. In addition, PYCP significantly reduced the levels of phosphorylated p38 and JNK. Moreover, by inhibiting the degradation of inhibitor of kappaB-α, PYCP significantly suppressed the TNF-α-induced increased transcriptional activity and nuclear translocation of nuclear factor-kappaB (NF-κB). Furthermore, PYCP inhibited E3-ubiquitin ligases in TNF-α-treated C2C12 myotubes. In conclusion, PYCP ameliorated TNF-α-induced muscle atrophy by inhibiting the mitogen-activated protein kinase-mediated NF-κB pathway, indicating that it has therapeutic potential for related disorders.

USP8 protects against lipopolysaccharide-induced cognitive and motor deficits by modulating microglia phenotypes through TLR4/MyD88/NF-κB signaling pathway in mice

Ubiquitin-specific protease 8 (USP8) regulates inflammation in vitro; however, the mechanisms by which USP8 inhibits neuroinflammation and its pathophysiological functions are not completely understood. In this study, we aimed to determine whether USP8 exerts neuroprotective effects in a mouse model of lipopolysaccharide (LPS)-induced cognitive and motor impairment. We commenced intracerebroventricular USP8 administration 7 days prior to i.p. injection of LPS (750 μg/kg). All treatments and behavioral experiments were performed once per day for 7 consecutive days. Behavioral tests and pathological/biochemical assays were performed to evaluate LPS-induced hippocampal damage. USP8 attenuated LPS-induced cognitive and motor impairments in mice. Moreover, USP8 downregulated several pro-inflammatory cytokines [nitric oxide (NO), tumor necrosis factor α (TNF-α), prostaglandin E2 (PGE2), and interleukin-1β (IL-1β)] in the serum and brain, and the relevant protein factors [inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX-2)] in the brain. Furthermore, USP8 upregulated the anti-inflammatory mediators interleukin (IL)-4 and IL-10 in the serum and brain, and promoted a shift from pro-inflammatory to anti-inflammatory microglial phenotypes. The LPS-induced microglial pro-inflammatory phenotype was abolished by TLR4 inhibitor and in TLR4−/− mice; these effects were similar to those of USP8 treatment. Mechanistically, we found that USP8 increased the expression of neuregulin receptor degradation protein-1 (Nrdp1), potently downregulated the expression of TLR4 and myeloid differentiation primary response protein 88 (MyD88) protein, and inhibited the phosphorylation of IκB kinase (IKK) β and kappa B-alpha (IκBα), thereby reducing nuclear translocation of p65 by inhibiting the activation of the nuclear factor-kappaB (NF-κB) signaling pathway in LPS-induced mice. Our results demonstrated that USP8 exerts protective effects against LPS-induced cognitive and motor deficits in mice by modulating microglial phenotypes via TLR4/MyD88/NF-κB signaling.

RAGE-dependent NF-kB inflammation processes in the capsule of frozen shoulders

The etiology of frozen shoulder (FS) remains uncertain. Advanced glycation end-products (AGEs) cause the cross-linking and stabilization of collagen and are increased in FS. The purpose of this study was to elucidate the pathogenesis of FS by evaluating the receptor of AGE (RAGE)-dependent pathways. Tissue samples of the coracohumeral ligament (CHL) and anterior inferior glenohumeral ligament (IGHL) were collected from 33 patients with FS, with severe stiffness, and 25 with rotator cuff tears (RCTs) as controls. Gene expression levels of RAGE, high-mobility group box 1 (HMGB1), Toll-like receptor 2 (TLR2), TLR4, nuclear factor-kappa B (NF-kB), and cytokines were evaluated using a quantitative real-time polymerase chain reaction. The immunoreactivities of carboxymethyllysine (CML), pentosidine, and RAGE were also evaluated. CML and pentosidine were further evaluated using high-performance liquid chromatography. Gene expression levels of RAGE, HMGB1, TLR2, TLR4, and NF-kB were significantly greater in the CHLs and IGHLs from the FS group than in those from the RCT group. Immunoreactivities of RAGE and CML were stronger in the CHLs and IGHLs from the FS group than in those from the RCT group. Pentosidine was weakly immunostained in the CHLs and IGHLs from the FS group. CML using high-performance liquid chromatography was significantly greater in the CHLs and IGHLs from the FS group than in those from the RCT group. AGEs and HMGB1 might play important roles in the pathogenesis of FS by binding to RAGE and activating NF-kB signaling pathways. Suppression of these pathways could be a treatment option for FS.

S-allylmercaptocysteine ameliorates lipopolysaccharide-induced acute lung injury in mice by inhibiting inflammation and oxidative stress via nuclear factor kappa B and Keap1/Nrf2 pathways

The garlic-derived organosulfur compound S-allylmercaptocysteine (SAMC) has been reported to exhibit anti-inflammatory and anti-oxidative activities, whereas its potential therapeutic effect on lipopolysaccharide (LPS)-induced acute lung injury (ALI) is unknown. In this study, we focused on exploring the therapeutic effects of SAMC on LPS-induced ALI mice and the involvement of underlying molecular mechanisms. BalB/c mice were treated with SAMC (10, 30 and 60 mg/kg) or positive control N-acetylcysteine (NAC, 500 mg/kg) by gavage after intratracheal instillation of LPS for 30 min and were sacrificed 24 h after LPS administration. Our results indicate that the treatment with SAMC not only ameliorated the histological changes but also decreased LPS-triggered lung edema. Moreover, SAMC displayed an anti-inflammatory effect through reducing inflammatory cells infiltration, myeloperoxidase (MPO) formation and inhibiting pro-inflammatory cytokines/mediator production including tumor necrosis factor alpha (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX2) via suppressing the activation of nuclear factor-kappaB (NF-κB) signaling pathway. Furthermore, SAMC attenuated oxidative stress evoked by LPS via diminishing malondialdehyde (MDA) formation and reversing glutathione (GSH) and superoxide dismutase (SOD) depletion. Meanwhile, SAMC up-regulated expressions of endogenous antioxidant/detoxifying proteins including heme oxygenase-1 (HO-1) and NAD(P)H: quinone oxidoreductase 1(NQO1) through reversing the suppression of Kelch-like ECH-associated protein 1 (Keap1)/nuclear factor erythroid-2 related factor 2 (Nrf2) signaling pathway. Our results demonstrate that SAMC effectively attenuated LPS-induced ALI which was largely dependent upon inhibition of inflammation and oxidative stress via NF-κB and Keap1/Nrf2 signaling pathways.