Extracellular Signal-related Kinase Inhibitor Research Articles

Chronic Kidney Disease-associated Lung Injury Is Mediated by Phosphate-induced MAPK/AKT Signaling.

Chronic kidney disease (CKD) is associated with systemic phosphate elevations, called hyperphosphatemia. Translational studies have shown that hyperphosphatemia contributes to CKD-associated inflammation and injury in various tissues, including the kidney, heart, liver, and parathyroid gland. Mechanisms underlying pathologic actions of elevated phosphate on cells are not well understood but seem to involve uptake of phosphate through sodium-phosphate cotransporters and phosphate-induced signaling via fibroblast growth factor receptor (FGFR) 1. Clinical studies indicate CKD patients are more likely to develop inflammatory and restrictive lung diseases, such as fibrotic interstitial lung diseases, and here we aimed to determine whether hyperphosphatemia can cause lung injury. We found that a mouse model of CKD and hyperphosphatemia, induced by an adenine-rich diet, develops lung fibrosis and inflammation. Elevation of systemic phosphate levels by administration of a high-phosphate diet in a mouse model of primary lung inflammation and fibrosis, induced by bleomycin, exacerbated lung injury in the absence of kidney damage. Our in vitro studies identified increases of proinflammatory cytokines in human lung fibroblasts exposed to phosphate elevations. Phosphate activated extracellular signal related kinase (ERK) 1/2 and protein kinase B (PKB/AKT) signaling, and pharmacological inhibition of ERK, AKT, FGFR1, or sodium-phosphate cotransporters prevented phosphate-induced proinflammatory cytokine upregulation. Additionally, inhibition of FGFR1 or sodium-phosphate cotransporters decreased the phosphate-induced activation of ERK and AKT. Our study suggests that phosphate can directly target lung fibroblasts and induce an inflammatory response and that hyperphosphatemia in CKD and non-CKD models contributes to lung injury. Phosphate-lowering strategies might protect from CKD-associated lung injury.

ERK/MAPK Signalling Pathway Regulates MMP2 through ETS1 in Renal Clear Cell Carcinoma.

The c-ETS-1 (ETS1) expression is high in clear cell renal cell carcinoma (ccRCC) tissues; however, how it impacts ccRCC is currently unknown. The online STRING web source was used to construct a protein network interacting with ETS1. The Cell Counting Kit-8 was used to detect the cell viability. A clonogenic assay, a wound-healing assay, and a Transwell assay were used to detect cell proliferation, invasion and migration abilities. Western blot was used to detect the expression of proteins. The data showed the expression of ETS1 in ccRCC tissues to be significantly increased compared to adjacent tissues (p<0.05). The positive expression of ETS1 in ccRCC patients aged 20-100 was statistically significant compared to adjacent normal tissues (p<0.05). The grade of ETS1 positive expression (1-4) and lymph node metastasis (N1) in ccRCC were significantly higher than those in adjacent normal tissues (p<0.05). The tumour stage (stages 1-4) in ccRCC patients with positive ETS1 expression was significantly higher than that in adjacent normal tissues (p<0.05). Knockdown of ETS1 and PERK inhibitors significantly inhibited the proliferation, migration and invasion of ccRCC cells. Knockdown of ETS1 inhibited MMP-2 expression, and an extracellular signal-related kinase (ERK) inhibitor inhibited both ETS1 and MMP-2 expression. A high expression of ETS1 is associated with the progression of ccRCC. This study suggests that ETS1 promotes proliferation by increasing MMP2 expression in ccRCC, and combined knockdown of ETS1 and inhibition of ERK can significantly inhibit the proliferation, migration and invasion of ccRCC. ETS1 may be a therapeutic and prognostic target for renal cell carcinoma.

Inhibition of oxidative stress-induced epithelial-mesenchymal transition in retinal pigment epithelial cells of age-related macular degeneration model by suppressing ERK activation

IntroductionEpithelial-mesenchymal transition (EMT) of retinal pigment epithelial (RPE) cells is related to the pathogenesis of various retinopathies including age-related macular degeneration (AMD). Oxidative stress is the major factor that induces degeneration of RPE cells associated with the etiology of AMD. ObjectivesSodium iodate (NaIO3) generates intracellular reactive oxygen species (ROS) and is widely used to establish a model of AMD due to the selective induction of retinal degeneration. This study was performed to clarify the effects of multiple NaIO3-stimulated signaling pathways on EMT in RPE cells. MethodsThe EMT characteristics in NaIO3-treated human ARPE-19 cells and RPE cells of the mouse eyes were analyzed. Multiple oxidative stress-induced modulators were investigated and the effects of pre-treatment with Ca2+ chelator, extracellular signal-related kinase (ERK) inhibitor, or epidermal growth factor receptor (EGFR) inhibitor on NaIO3-induced EMT were determined. The efficacy of post-treatment with ERK inhibitor on the regulation of NaIO3-induced signaling pathways was dissected and its role in retinal thickness and morphology was evaluated by using histological cross-sections and spectral domain optical coherence tomography. ResultsWe found that NaIO3 induced EMT in ARPE-19 cells and in RPE cells of the mouse eyes. The intracellular ROS, Ca2+, endoplasmic reticulum (ER) stress marker, phospho-ERK, and phospho-EGFR were increased in NaIO3-stimulated cells. Our results showed that pre-treatment with Ca2+ chelator, ERK inhibitor, or EGFR inhibitor decreased NaIO3-induced EMT, interestingly, the inhibition of ERK displayed the most prominent effect. Furthermore, post-treatment with FR180204, a specific ERK inhibitor, reduced intracellular ROS and Ca2+ levels, downregulated phospho-EGFR and ER stress marker, attenuated EMT of RPE cells, and prevented structural disorder of the retina induced by NaIO3. ConclusionsERK is a crucial regulator of multiple NaIO3-induced signaling pathways that coordinate EMT program in RPE cells. Inhibition of ERK may be a potential therapeutic strategy for the treatment of AMD.

Narciclasine suppresses oral cancer metastasis by modulating cathepsin B and extracellular signal–related kinase pathways

Oral cancer is a malignancy with unfavorable prognosis due to its high rates of recurrence and lymph node metastasis. Narciclasine is extracted from Narcissus species (Amaryllidaceae), which have antitumor and anti-inflammatory properties. However, the antitumor properties of narciclasine toward oral cancer remain unclear. The present study explored the antimetastatic effects of narciclasine in oral cancer as well as the underlying molecular mechanisms. We treated three oral cancer cell lines with noncytotoxic concentrations of narciclasine and discovered a dose-dependent antimetastatic effect. Mitogen-activated protein kinase (MAPK) pathways, including extracellular signal–related kinase (ERK), p38, and c-Jun N-terminal kinase (JNK), were regulated by narciclasine. We further discovered the ERK pathway to directly affect narciclasine-induced metastasis inhibition by combining treatment with narciclasine and ERK inhibitor. Furthermore, downregulation of cathepsin B (CTSB) in the SAS and SCC-47 cell lines revealed the critical role of CTSB in the antimetastatic effect of narciclasine. Our findings indicate that narciclasine inhibits oral cancer metastasis by regulating the ERK pathway and CTSB. This study provides evidence of the mechanism of narciclasine-induced inhibition oral cancer metastasis and suggests potential targets for use in oral cancer treatment.

Neuromedin U Induces Activation of Peripheral Group 2 Innate Lymphoid Cells through the ERK Pathway in Allergic Rhinitis Patients

Introduction: In allergic diseases, group 2 innate lymphoid cells (ILC2s) play critical roles. Neuromedin U (NMU), a highly conserved multifunctional neuropeptide, is secreted by cholinergic neurons and involved in asthma pathogenesis by amplifying lung inflammation driven by ILC2s. However, the precise effects of NMU on ILC2s in allergic rhinitis (AR) and related diseases remain unclear. Methods: A total of 15 patients with persistent AR and 8 healthy controls (HCs) were enrolled in the study. Visual analog scale (VAS) scores are used to assess the severity of clinical symptoms in AR patients. The percentages of ILC2s in peripheral blood mononuclear cells (PBMCs) were enumerated using flow cytometry. The soluble or intracellular cytokines (IL-5 and IL-13) in PBMCs or sorted ILC2s were assessed in response to various stimuli with IL-33, NMU, IL-33 combined with extracellular signal-related kinase (ERK) inhibitor or NMU combined with ERK inhibitor in the presence of IL-2. Results: We confirmed the proportion of circulating ILC2s was significantly higher in AR patients than in HCs. ILC2s levels were found to be positively related to VAS scores. We also discovered that the release of IL-5 and IL-13 in AR patients’ PBMCs stimulated by NMU (p < 0.0001 and p < 0.0001, respectively) or IL-33 (p = 0.002; p = 0.044, respectively) was significantly higher than in HCs. In AR patients, NMU stimulated PBMCs or ILC2s to generate greater inflammatory factors IL-5 and IL-13 compared to IL-33. Furthermore, we observed that NMU-promoted ILC2s activation and proliferation functions were restricted when the ERK pathway was inhibited. Conclusion: NMU effectively activated ILC2s in AR patients to produce Th2-type cytokines, and this activation can be prevented by ERK pathway inhibitors. Our findings shed new light on the neuro-immune mechanism of AR and offer new insights into its prevention and treatment.

Resolvin D1 Alleviates Mechanical Allodynia via ALX/FPR2 Receptor Targeted Nod-like Receptor Protein 3/Extracellular Signal-Related Kinase Signaling in a Neuropathic Pain Model

The current study aimed to investigate the role and underlying mechanism of Resolvin D1 (RvD1) alleviating spinal nerve ligation (SNL)-induced neuropathic pain (NP) and its interplay with regulatory cascades of Nod-like Receptor Protein 3 (NLRP3) inflammasome. Sprague-Dawley male rat models of SNL-stimulated NP were established, which were pre-treated with different doses of RvD1, WRW4 (ALX/FPR2 inhibitor) or U0126 (ERK inhibitor) for three successive days following the operation. Pain behavior was assessed by measuring changes in the mechanical sensitivity of the hind paws during an observation period of seven consecutive days. The spinal cord (SC) and dorsal root ganglions (DRGs) tissues were collected on postoperative day 7. Immunohistochemistry (IHC) and Western blot were performed to determine the expression levels of NLRP3 inflammasome complex, ALX/FPR2 receptor and extracellular signal-related kinase (ERK). The pro-inflammatory mediators (IL-1β and IL-18) were measured by enzyme-linked immunosorbent assay (ELISA). The results showed that RvD1 could alleviate mechanical allodynia significantly in the SNL-induced NP rat models. Also, RvD1 inhibited the expression of p-ERK, the NLRP3 inflammasomes complex and its corresponding downstream pro-inflammatory mediators which were significantly enhanced in the SC and DRGs of the rat SNL models. While these changes were partially reversed by pre-administration of WRW4 and further strengthened by co-treated with U0126. Our results suggest that RvD1 dependent on ALX/FPR2 may have an analgesic and anti-inflammatory influence on SNL-induced NP driven by inhibiting NLRP3 inflammasome via ERK signaling pathway. These data also provide strong support for the recent modulation of neuro-inflammatory priming and highlight the potential for specialized pro-resolving mediators (SPMs) as novel therapeutic avenues for NP.

Oncostatin M Induces Lipolysis and Suppresses Insulin Response in 3T3-L1 Adipocytes.

Oncostatin M (OSM) is an immune cell-derived cytokine that is upregulated in adipose tissue in obesity. Upon binding its receptor (OSMR), OSM induces the phosphorylation of the p66 subunit of Src homology 2 domain-containing transforming protein 1 (SHC1), called p66Shc, and activates the extracellular signal-related kinase (ERK) pathway. Mice with adipocyte-specific OSMR deletion (OsmrFKO) are insulin resistant and exhibit adipose tissue inflammation, suggesting that intact adipocyte OSM–OSMR signaling is necessary for maintaining adipose tissue health. How OSM affects specific adipocyte functions is still unclear. Here, we examined the effects of OSM on adipocyte lipolysis. We treated 3T3-L1 adipocytes with OSM, insulin, and/or inhibitors of SHC1 and ERK and measured glycerol release. We also measured phosphorylation of p66Shc, ERK, and insulin receptor substrate-1 (IRS1) and the expression of lipolysis-associated genes in OSM-exposed 3T3-L1 adipocytes and primary adipocytes from control and OsmrFKO mice. We found that OSM induces adipocyte lipolysis via a p66Shc-ERK pathway and inhibits the suppression of lipolysis by insulin. Further, OSM induces phosphorylation of inhibitory IRS1 residues. We conclude that OSM is a stimulator of lipolysis and inhibits adipocyte insulin response. Future studies will determine how these roles of OSM affect adipose tissue function in health and disease.

Zinc antagonizes common carp (Cyprinus carpio) intestinal arsenic poisoning through PI3K/AKT/mTOR signaling cascade and MAPK pathway

Arsenic (As) pollution is a serious and longstanding problem, which has obvious threaten to aquatic organisms. The study aimed to explore the mitigation effect of natural antioxidant zinc (Zn) on As toxicity in the foregut and midgut of common carp (Cyprinus carpio L.), and in-depth disclose related signal cascade. Carps were treated with Zn2+ (1 mg/L) and/or As3+ (2.83 mg/L) for a period of 30 days. Under As exposure, the foregut and midgut showed obvious burst of reactive oxygen species (ROS) and breakdown of antioxidant system. What followed is the activation of the endogenous and exogenous apoptotic pathways, and the rise of autophagy level prompted by the increase in LC3 II and the down-regulation of p62. Mitochondrial swelling, cristae fragmentation and autophagosomes were observed under the electron microscope, which also means the occurrence of apoptosis and autophagy. In addition, As induced the activation of p38 mitogen-activated protein kinase (MAPK), c-Jun N-terminal kinase (JNK) and the inhibition of extracellular signal-related kinase (ERK) in MAPK signaling, and up-regulated the level of autophagy through the inhibition of the phosphatidylinositol 3 kinase (PI3K)/AKT/ mammalian target of rapamycin (mTOR) signaling cascade. However, Zn supplementation has clearly reversed the above phenomenon, and it basically has no effect on foregut and midgut. In conclusion, this study shows that Zn can alleviate the damage caused by subchronic As exposure, which provides a reference for the use of Zn preparations in aquaculture.

Chemokine receptor CCR7 induces intracellular signaling that inhibits apoptosis of mature dendritic cells

AbstractAcquisition of CCR7 expression is an important phenotype change during dendritic cell (DC) maturation that endows these cells with the capability to migrate to lymph nodes. We have analyzed the possible role of CCR7 on the regulation of the survival of DCs. Stimulation with CCR7 ligands CCL19 and CCL21 inhibits apoptotic hallmarks of serum-deprived DCs, including membrane phosphatidylserine exposure, loss of mitochondria membrane potential, increased membrane blebs, and nuclear changes. Both chemokines induced a rapid activation of phosphatidylinositol 3′-kinase/Akt1 (PI3K/Akt1), with a prolonged and persistent activation of Akt1. Interference with PI3K, Gi, or G protein βγ subunits abrogated the effects of the chemokines on Akt1 activation and on survival. In contrast, inhibition of extracellular signal-related kinase 1/2 (Erk1/2), p38, or c-Jun N-terminal kinase (JNK) was ineffective. Nuclear factor–κB (NFκB) was involved in the antiapoptotic effects of chemokines because inhibition of NFκB blunted the effects of CCL19 and CCL21 on survival. Furthermore, chemokines induced down-regulation of the NFκB inhibitor IκB, an increase of NFκB DNA-binding capability, and translocation of the NFκB subunit p65 to the nucleus. In summary, in addition to its well-established role in chemotaxis, we show that CCR7 also induces antiapoptotic signaling in mature DCs.

101P HIF1-α depletion overcomes resistance to oxaliplatin in colorectal cancer via ERK signalling pathway

Chemoresistance is a major reason for treatment failure and results in poor prognosis in patients with advanced colorectal cancer (CRC). Hypoxia-inducible factor-1 (HIF1-α) is a regulator of the transcriptional response to oxygen deprivation in cancer cells and activates the transcription of genes that are involved in angiogenesis, cell survival and invasion. However, the effect of HIF1-α activation on oxaliplatin resistance in CRC is unclear. This study aimed to investigate the effects and potential mechanism of HIF1-α on oxaliplatin resistance in CRC. Oxaliplatin-resistant cells were generated from HCT116 cells. Cell Counting Kit-8, flow cytometry, Transwell assay were used to compare the characteristics of oxaliplatin resistant HCT116 (HCT116_OxR) cells and the corresponding parental HCT116 cells. The expression of HIF1-α, extracellular signal-related kinase (ERK), and transforming growth factor β (TGFβ) were confirmed by RT-PCR and western blotting in HCT116_OxR and HCT116 cells. Next, we evaluated the combination efficacy of inhibitors of HIF1-α (YC-1), ERK (Ravoxertinib), and TGFβ1 (SB431542) with oxaliplatin by in vitro and in vivo experiments. We found that cell viability of HCT116_OxR was higher than that in parental cells in the presence of oxaliplatin. The relative expression of HIF1-α was significantly increased in HCT116_OxR cells compared with the parental HCT116 cells. Downregulation of HIF1-α in HCT116_OxR cells increased oxaliplatin sensitivity, and diminished cell survival and invasion. Significantly, ERK signaling in HCT116_OxR cells is dependent on TGFβ1, promoting oxaliplatin resistance by activating HIF1-α. Inhibition of ERK by Ravoxertinib or TGFβ1 by SB431542 efficiently overcame oxaliplatin resistance in vitro and in vivo. HIF1-α activation is regulated by TGFβ1/ERK axis in HCT116_OxR cells. Hence, HIF1-α, ERK, and TGFβ1 are potential therapeutic targets for overcoming oxaliplatin resistance in patients with CRC.

Growth differentiation factor 11 promotes differentiation of MSCs into endothelial-like cells for angiogenesis.

Growth differentiation factor 11 (GDF11) is a member of the transforming growth factor‐β super family. It has multiple effects on development, physiology and diseases. However, the role of GDF11 in the development of mesenchymal stem cells (MSCs) is not clear. To explore the effects of GDF11 on the differentiation and pro‐angiogenic activities of MSCs, mouse bone marrow–derived MSCs were engineered to overexpress GDF11 (MSCGDF11) and their capacity for differentiation and paracrine actions were examined both in vitro and in vivo. Expression of endothelial markers CD31 and VEGFR2 at the levels of both mRNA and protein was significantly higher in MSCGDF11 than control MSCs (MSCVector) during differentiation. More tube formation was observed in MSCGDF11 as compared with controls. In an in vivo angiogenesis assay with Matrigel plug, MSCGDF11 showed more differentiation into CD31+ endothelial‐like cells and better pro‐angiogenic activity as compared with MSCVector. Mechanistically, the enhanced differentiation by GDF11 involved activation of extracellular‐signal‐related kinase (ERK) and eukaryotic translation initiation factor 4E (EIF4E). Inhibition of either TGF‐β receptor or ERK diminished the effect of GDF11 on MSC differentiation. In summary, our study unveils the function of GDF11 in the pro‐angiogenic activities of MSCs by enhancing endothelial differentiation via the TGFβ‐R/ERK/EIF4E pathway.

EglN3 hydroxylase stabilizes BIM-EL linking VHL type 2C mutations to pheochromocytoma pathogenesis and chemotherapy resistance

Despite the discovery of the oxygen-sensitive regulation of HIFα by the von Hippel-Lindau (VHL) protein, the mechanisms underlying the complex genotype/phenotype correlations in VHL disease remain unknown. Some germline VHL mutations cause familial pheochromocytoma and encode proteins that preserve their ability to down-regulate HIFα. While type 1, 2A, and 2B VHL mutants are defective in regulating HIFα, type 2C mutants encode proteins that preserve their ability to down-regulate HIFα. Here, we identified an oxygen-sensitive function of VHL that is abolished by VHL type 2C mutations. We found that BIM-EL, a proapoptotic BH3-only protein, is hydroxylated by EglN3 and subsequently bound by VHL. VHL mutants fail to bind hydroxylated BIM-EL, regardless of whether they have the ability to bind hydroxylated HIFα or not. VHL binding inhibits BIM-EL phosphorylation by extracellular signal-related kinase (ERK) on serine 69. This causes BIM-EL to escape from proteasomal degradation, allowing it to enhance EglN3-induced apoptosis. BIM-EL was rapidly degraded in cells lacking wild-type VHL or in which EglN3 was inactivated genetically or by lack of oxygen, leading to enhanced cell survival and chemotherapy resistance. Combination therapy using ERK inhibitors, however, resensitizes VHL- and EglN3-deficient cells that are otherwise cisplatin-resistant.

Hypoxia-Induced Epithelial-To-Mesenchymal Transition Mediates Fibroblast Abnormalities via ERK Activation in Cutaneous Wound Healing.

Previous studies described the involvement of extracellular signal-related kinase (ERK) in systemic fibrotic diseases, but the role of ERK in cutaneous scarring is unknown. Although hypoxia drives tissue fibrosis by activating hypoxia-inducible factor-1α (HIF-1α), the specific roles of hypoxia and associated ERK phosphorylation in abnormal fibroblast activity during cutaneous scarring are unclear. Here, we investigated whether pathologic myofibroblast-like keloid fibroblast activity is promoted by hypoxia-induced epithelial–mesenchymal transition mediated by ERK activation. ERK phosphorylation was significantly increased in keloid tissue and fibroblasts. Human dermal fibroblasts cultured under hypoxia (1% O2) expressed phosphorylated ERK and exhibited activation of p38 mitogen-activated protein kinase signaling. Hypoxic human dermal fibroblasts showed increased protein and mRNA levels of epithelial–mesenchymal transition markers. Furthermore, administration of an ERK inhibitor (SCH772984) reduced the hypoxia-induced elevation of collagen type I levels in human dermal fibroblasts. Therefore, ERK may be a promising therapeutic target in profibrogenic diseases.

Silencing KRT16 inhibits keratinocyte proliferation and VEGF secretion in psoriasis via inhibition of ERK signaling pathway.

Psoriasis is a multisystem disease affecting about 2% of the population, while keratin16 (KRT16) has been reported to participate in psoriasis. However, the specific mechanism of KRT16 in psoriasis was inadequately investigated. The objective of the study was to elucidate the mechanism by which siRNA-mediated silencing of KRT16 affects keratinocyte proliferation and vascular endothelial growth factor (VEGF) secretion in psoriasis through the extracellular signal-related kinase (ERK) signaling pathway. Psoriasis-related core gene KRT16 was screened out. Then, the expression of KRT16, VEGF, and ERK signaling pathway-related genes was detected in psoriatic patients. To further investigate the mechanism of KRT16, keratinocytes in psoriatic patients were treated with KRT16 siRNA or/and ERK inhibitor (PD98059) to detect the changes in related gene expression and cell survival. KRT16 was involved in psoriasis development. The expression levels of KRT16, p-ERK1/2, and VEGF in lesion tissues are significantly elevated. Keratinocytes treated with KRT16-siRNA and KRT16-siRNA + PD98059 exhibited reduced KRT16, p-ERK1/2, and VEGF expression. The cell survival rate in cells treated with KRT16-siRNA, PD98059, and KRT16-siRNA + PD98059 reduced significantly. These findings indicate that silencing KRT16 inhibits keratinocyte proliferation and VEGF secretion in psoriasis via inhibition of ERK signaling pathway, which provides a basic theory in the treatment of psoriasis.

Regorafenib reverses HGF-induced sorafenib resistance by inhibiting epithelial-mesenchymal transition in hepatocellular carcinoma.

Sorafenib resistance is one of the major obstacles towards achieving a better outcome in patients with advanced hepatocellular carcinoma (HCC), in which aberrant activation of the hepatocyte growth factor (HGF)/mesenchymal‐epithelial transition pathway is frequently observed. Here, we report that HCC cells develop sorafenib resistance following HGF stimulation. Furthermore, HGF activates the downstream extracellular signal‐related kinase (ERK) and signal transducer and activator of transcription 3 (STAT3) pathway and induces epithelial–mesenchymal transition (EMT) by up‐regulating Snail in HCC cells. Inhibition of ERK and STAT3 abolished the rescue effect of HGF by down‐regulating Snail and EMT. Moreover, phosphoinositide 3‐kinase/Akt was also activated in HGF‐treated HCC cells, although it had no effect on Snail expression. Notably, we also found that regorafenib reversed HGF‐induced sorafenib resistance by inhibiting ERK and STAT3, and subsequently down‐regulating Snail and EMT. Taken together, our results indicate that HGF induces sorafenib resistance by activating phosporylated (P)‐ERK/Snail/EMT and P‐STAT3/Snail/EMT pathways. Inhibition of P‐ERK and P‐STAT3 by regorafenib can block HGF‐induced EMT, thereby reversing HGF‐induced sorafenib resistance.

Effects of IGFBP3 gene silencing mediated inhibition of ERK/MAPK signaling pathway on proliferation, apoptosis, autophagy, and cell senescence in rats nucleus pulposus cells.

Disc degeneration is the common life-threatening disease characterized by flank pain. The gene expression of insulin-like growth factor binding protein 3 (IGFBP3) is increased in patients with disc degeneration, however, its mechanism is still unknown. This study aimed to investigate the influence of IGFBP3 gene silencing mediated inhibition of extracellular signal-related kinase (ERK)/mitogen-activated protein kinase (MAPK) signaling on proliferation, apoptosis, autophagy, and cell senescence in rats nucleus pulposus (NP) cells. The expression of IGFBP3 in disc NP of patients was assessed by real-time PCR (RT-PCR) and western blot. RT-PCR, transwell assay, immunohistochemical staining, SA-β-Gal staining, and western blot were performed to explore the molecular mechanism of IGFBP3 in NP cell migration and invasion. In this study, IGFBP3 was highly expressed in disc NP of patients. With RT-PCR, transwell assay, immunohistochemical staining, SA-β-Gal staining, and western blot, downregulated IGFBP3 could inhibit NP cells' migration and invasion by targeting the ERK/MAPK signaling pathway. Our findings revealed that the inhibition of the ERK/MAPK pathway was mediated by IGFBP3 silencing that had effects on proliferation, apoptosis, autophagy, and cell senescence. Furthermore, our findings suggested the underlying mechanism of disc degeneration.

Wheat Sprout Extract Stimulates Cell Proliferation via Phosphorylation of ERK and Akt in Human Dermal Papilla Cells

We investigated the effect of Wheat Sprout MeOH extract (WSM) on cell proliferation, and analyzed the molecular mechanism in human hair dermal papilla cells (HHDPCs). After exposure to WSM in HHDPCs, cell proliferation was increased compared with minoxidil-treated cells. Treatment with WSM led to phosphorylation of extracellular signal-related kinase (ERK) and Akt. Moreover, WSM-mediated increase of cell proliferation was attenuated by PD98059, ERK inhibitor, and LY294002, Akt inhibitor. These results suggest that WSM induced cell proliferation in HHDPCs through activation of ERK and Akt.

Wnt5a induces catabolic signaling and matrix metalloproteinase production in human articular chondrocytes.

Aberrant Wnt signaling may contribute to osteoarthritis (OA) but the Wnt family members involved have not been fully identified. The purpose of this study was to investigate the role of Wnt5a as a potential mediator of cartilage destruction in OA. Immunohistochemistry to detect Wnt5a was performed using normal and OA human articular cartilage. Cultured normal human chondrocytes were treated with fibronectin fragments (FN-f) as a catabolic stimulus or recombinant Wnt5a protein with or without pretreatment using a panel of signaling inhibitors. Expression of Wnt5a, anabolic genes and catabolic genes were determined by quantitative real-time PCR. Production of Wnt5a protein and matrix metalloproteinases (MMPs) as well as activation of signaling proteins were analyzed by immunoblotting. Wnt5a was present in human articular cartilage with OA changes and its expression and secretion were increased in FN-f stimulated chondrocytes. FN-f stimulated Wnt5a production through the c-Jun N-terminal kinase (JNK) and extracellular signal-related kinase (ERK) pathways. Wnt5a reduced aggrecan gene expression after 48h of treatment. Wnt5a seemed to promote MMP1, -3, and -13 expression as well as MMP1 and MMP13 protein production in normal human chondrocytes. Wnt5a inhibitor peptides did not affect FN-f induced MMP production. Wnt5a activated β-catenin independent signaling including calmodulin-dependent protein kinase II (CaMKII), JNK, p38, ERK1/2, p65 and Akt. Inhibition of JNK, p38, ERK, PI-3 kinase and CaMKII by specific signaling inhibitors suppressed Wnt5a mediated MMP1 and MMP13 production. Wnt5a is present in human OA cartilage and can promote chondrocyte catabolic activity through non-canonical Wnt signaling, which suggests a potential role in OA.

Galectin-7 promotes the invasiveness of human oral squamous cell carcinoma cells via activation of ERK and JNK signaling.

Galectin-7 is a member of the β-galactoside-binding protein family, and is highly expressed in oral squamous cell carcinoma (OSCC). The aim of the present study was to investigate the effects of manipulating galectin-7 expression on the biological phenotype of human OSCC cells and the associated molecular mechanisms. Knockdown of endogenous galectin-7 via small interfering RNA (siRNA) was performed and cell proliferation, apoptosis, migration, and invasion were subsequently assessed. The data indicated that galectin-7 silencing had no impact on the proliferation or apoptosis of OSCC cells. However, compared with non-transfected cells, percentage wound closure was significantly lower in galectin-7-silenced cells following 24 h incubation, indicating decreased cell migration. Furthermore, Matrigel invasion assays demonstrated that galectin-7 knockdown significantly reduced the number of invaded cells, compared with the number in non-transfected cells. Western blot analysis indicated that galectin-7 overexpression resulted in a significant increase in the expression of the proteins matrix metalloproteinase (MMP)-2 and MMP-9. The invasive abilities of cells overexpressing galectin-7 significantly decreased following co-transfection with MMP-2- or MMP-9-specific siRNA. Increasing galectin-7 expression significantly enhanced the phosphorylation of extracellular signal-related kinase (ERK) 1/2 and c-Jun N-terminal kinase (JNK) 1/2. Pharmacological inhibition of ERK or JNK activity significantly suppressed the invasiveness of galectin-7-overexpressing cells and abrogated the upregulation of MMP-2 and MMP-9. Taken together, the results of the current study provide novel evidence for the pro-invasive activity of galectin-7 in OSCC cells, which is associated with activation of ERK and JNK signaling and the induction of MMP-2 and MMP-9.

Rhesus rotavirus VP6 regulates ERK-dependent calcium influx in cholangiocytes

The Rhesus rotavirus (RRV) induced murine model of biliary atresia (BA) is a useful tool in studying the pathogenesis of this neonatal biliary obstructive disease. In this model, the mitogen associated protein kinase pathway is involved in RRV infection of biliary epithelial cells (cholangiocytes). We hypothesized that extracellular signal-related kinase (ERK) phosphorylation is integral to calcium influx, allowing for viral replication within the cholangiocyte. Utilizing ERK and calcium inhibitors in immortalized cholangiocytes and BALB/c pups, we determined that ERK inhibition resulted in reduced viral yield and subsequent decreased symptomatology in mice. In vitro, the RRV VP6 protein induced ERK phosphorylation, leading to cellular calcium influx. Pre-treatment with an ERK inhibitor or Verapamil resulted in lower viral yields. We conclude that the pathogenesis of RRV-induced murine BA is dependent on the VP6 protein causing ERK phosphorylation and triggering calcium influx allowing replication in cholangiocytes.