Resveratrol inhibits Lin28A expression and induces its degradation via the proteasomal pathway in NCCIT cells.

Lin28A is an oncoprotein overexpressed in several cancer types such as testicular, ovarian, colon, breast and lung cancers. As a pluripotency factor that promotes tumorigenesis, Lin28A is associated with more undifferentiated and aggressive tumors phenotypes. Moreover, Lin28A is a highly stable protein that is difficult to downregulate. The compound resveratrol (RSV) has anticancer effects. The present study aimed to elucidate the mechanisms underlying the downregulation of Lin28A protein expression by RSV in the NCCIT cell line. NCCIT cells were treated with different concentrations of RSV to investigate its effects on Lin28A expression. The mRNA expression levels of Lin28A and ubiquitin-specific protease 28 (USP28) were assessed using reverse transcription-quantitative PCR. Western blot analysis was employed to evaluate the protein levels of Lin28A, USP28 and phosphorylated Lin28A. In addition, in some experiments, cells were treated with a MAPK/ERK pathway inhibitor, and other experiments involved transfecting cells with small interfering RNAs targeting USP28. The results demonstrated that RSV significantly reduced Lin28A expression by destabilizing the protein; this effect was mediated by the ability of RSV to suppress the expression of USP28, a deubiquitinase that normally protects Lin28A from ubiquitination and degradation. Additionally, RSV inhibited phosphorylation of Lin28A via the MAPK/ERK pathway; this phosphorylation event has previously been shown to enhance the stability of Lin28A by increasing its half-life. This resulted in Lin28A degradation through the proteasomal pathway in NCCIT cells. The results provide further evidence of the anticancer activity of RSV, and identified Lin28A and USP28 as promising therapeutic targets. As a stable oncoprotein, downregulating Lin28A expression is challenging. However, the present study demonstrated that RSV can overcome this hurdle by inhibiting USP28 expression and MAPK/ERK signaling to promote Lin28A degradation. Furthermore, elucidating these mechanisms provides avenues for developing targeted cancer therapies.

Effect of irisin on ovarian phosphatidylinositol-3-kinase/protein kinase B signaling pathway and mitogen-activated protein kinase/extracellular signal-regulated kinase pathways of rats with polycystic ovary syndrome.

OBJECTIVE: To investigate the independent effects of irisin on insulin resistance (IR) in ovary of polycystic ovary syndrome (PCOS) and explore possible pathways. METHODS: We established PCOS medel using Poretsky L's method, then PCOS rats were randomly divided into model group (M) and irisin group (I), and normal rats (N) were used as the control. Then rats in the group I were injected with recombinant irisin. Then the levels of circulating fasting blood glucose (FBG), fasting insulin (FINS), homeostasis model assessment of IR (HOMA-IR) and PI3K/AKT and MAPK/ERK pathways in each group were observed, as well as the effects of irisin on the levels of circulating HOMA-IR and PI3K/AKT and MAPK/ERK pathways in ovary of PCOS rats were evaluated. RESULTS: Compared with normal group, levels of FBG, FINS, and HOMA-IR of model group were significantly increased (p < 0.001, p < 0.001, and p < 0.001, respectively), levels of average optical density by IHC of p-PI3K, PI3K, p-AKT, and AKT (p = 0.015, p = 0.010, p = 0.005, and p = 0.009, respectively) and levels of mRNA concentration of PI3K and AKT (p = 0.001, and p = 0.005, respectively) were decreased, while the levels of average optical density of p-ERK, ERK (p = 0.011, and p = 0.013, respectively) and level of mRNA concentration of ERK (p < 0.001) were increased in ovary. After irisin intervention, compared with model group, levels of FBG, FINS, and HOMA-IR of rats in irisin group were significantly decreased (p = 0.001, p < 0.001, and p < 0.001, respectively), levels of average optical density by IHC of p-PI3K, PI3K, p-AKT, and AKT (p = 0.030, p = 0.024, p = 0.012, and p = 0.025, respectively) and levels of mRNA concentration of PI3K and AKT (p = 0.002, and p = 0.003, respectively) were significantly increased, while the levels of average optical density of p-ERK, ERK (p = 0.004, and p = 0.026, respectively) and level of mRNA concentration of ERK (p = 0.001) were significantly decreased. CONCLUSION: Our study demonstrated that irisin could not only improve circulating insulin resistance, but may also improve ovarian IR through an increase in the activity of PI3K/AKT signaling and a decrease of MAPK/ERK signaling.

EIF3B affects the invasion and metastasis of hepatocellular carcinoma cells via the TGFBI/MAPK/ERK pathway.

INTRODUCTION AND OBJECTIVES: To study the effect of eukaryotic initiation factor 3B (EIF3B) on the invasion and migration of hepatocellular carcinoma (HCC) and its potential mechanism. MATERIALS AND METHODS: The clinical significance of EIF3B expression was studied with The Cancer Genome Atlas (TCGA) and Gene Expression Profiling Interaction Analysis datasets. Immunohistochemical staining and western blotting were used to examine EIF3B expression in cell lines and tissues from HCC patients. The scratch assay and transwell assay were used to measure the invasion and metastasis of different HCC cell lines in vitro. The molecular mechanism of EIF3B was determined using RNA-seq and identification of dysregulated signaling pathways. Western blotting was used to verify the alterations of EIF3B signaling functioned in the promotion of HCC progression. RESULTS: Elevated expression of EIF3B in HCC correlated significantly with aggressive clinicopathologic characteristics, including advanced tumor grade and poor prognosis. Studies with cultured cells indicated that EIF3B knockdown inhibited HCC cell invasion and metastasis by depressing the epithelial-mesenchymal transition (EMT). EIF3B also activated the TGFBI/MAPK/ERK signaling pathway by increasing the levels of pMEK and pERK. CONCLUSIONS: Our results indicate that EIF3B functions as an oncogene in HCC that accelerates cell invasion, metastasis, and the EMT by stimulation of the TGFBI/MAPK/ERK signaling pathway. EIF3B is a potential target for the treatment of HCC.

Evodiamine inhibits proliferation and induces apoptosis of nasopharyngeal carcinoma cells via the SRC/ERBB2-mediated MAPK/ERK signaling pathway.

This study aimed to investigate the effect and potential mechanism of evodiamine (EVO) on proliferation and apoptosis of nasopharyngeal carcinoma (NPC) cells. EVO inhibited proliferation, blocked cell cycle progression, and induced apoptosis of NPC cells. There are 27 known anti-NPC targets of EVO, of which eight are core targets, namely SRC, ERBB2, STAT3, MAPK8, NOS3, CXCL8, APP, and HDAC1. Molecular docking analysis showed that the binding of EVO with its key targets (SRC, ERBB2) was good. EVO also reduced the expression of SRC and ERBB2, the key proteins p-MEK and p-ERK1/2 of the MAPK/ERK signaling pathway, and the downstream proteins PCNA and XIAP. EVO inhibited the growth of NPC xenografts in nude mice and reduced the expression levels of SRC, ERBB2, ERK1/2, p-ERK1/2, PCNA and XIAP in NPC tissue. When the MAPK/ERK signaling pathway was activated by epidermal growth factor (EGF), the expression levels of PCNA and XIAP increased, the cell proliferation index increased, and the apoptosis rate decreased in the EGF + EVO treatment group compared to treatment with EVO alone. These changes indicated that the inhibitory effect of EVO on proliferation and apoptosis of NPC cells was related to the down-regulation of SRC and ERBB2 expression, and further inhibition of the MAPK/ERK signaling pathway.

Sodium Tungstate Promotes Neurite Outgrowth and Confers Neuroprotection in Neuro2a and SH-SY5Y Cells.

Sodium tungstate (Na2WO4) normalizes glucose metabolism in the liver and muscle, activating the Mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) pathway. Because this pathway controls neuronal survival and differentiation, we investigated the effects of Na2WO4 in mouse Neuro2a and human SH-SY5Y neuroblastoma monolayer cell cultures. Na2WO4 promotes differentiation to cholinergic neurites via an increased G1/G0 cell cycle in response to the synergic activation of the Phosphatidylinositol 3-kinase (PI3K/Akt) and ERK1/2 signaling pathways. In Neuro2a cells, Na2WO4 increases protein synthesis by activating the mechanistic target of rapamycin (mTOR) and S6K kinases and GLUT3-mediated glucose uptake, providing the energy and protein synthesis needed for neurite outgrowth. Furthermore, Na2WO4 increased the expression of myocyte enhancer factor 2D (MEF2D), a member of a family of transcription factors involved in neuronal survival and plasticity, through a post-translational mechanism that increases its half-life. Site-directed mutations of residues involved in the sumoylation of the protein abrogated the positive effects of Na2WO4 on the MEF2D-dependent transcriptional activity. In addition, the neuroprotective effects of Na2WO4 were evaluated in the presence of advanced glycation end products (AGEs). AGEs diminished neurite differentiation owing to a reduction in the G1/G0 cell cycle, concomitant with lower expression of MEF2D and the GLUT3 transporter. These negative effects were corrected in both cell lines after incubation with Na2WO4. These findings support the role of Na2WO4 in neuronal plasticity, albeit further experiments using 3D cultures, and animal models will be needed to validate the therapeutic potential of the compound.

Antrodia camphorata Supplementation during Early Life Alters Gut Microbiota and Inhibits Young-Onset Intestinal Tumorigenesis in APC1638N Mice Later in Life.

Young-onset colorectal cancer is an increasing concern worldwide due to the growing prevalence of Westernized lifestyles in childhood and adolescence. Environmental factors during early life, particularly early-life nutrition, significantly contribute to the increasing incidence. Recently, there have been reports of beneficial effects, including anti-inflammation and anti-cancer, of a unique fungus (Antrodia camphorate, AC) native to Taiwan. The objective of this study is to investigate the impact of AC supplementation in early life on the development of young-onset intestinal tumorigenesis. APC1638N mice were fed with a high-fat diet (HF) at 4-12 weeks of age, which is equivalent to human childhood/adolescence, before switching to a normal maintenance diet for an additional 12 weeks up to 24 weeks of age, which is equivalent to young to middle adulthood in humans. Our results showed that the body weight in the HF groups significantly increased after 8 weeks of feeding (p < 0.05). Following a switch to a normal maintenance diet, the change in body weight persisted. AC supplementation significantly suppressed tumor incidence and multiplicity in females (p < 0.05) and reduced IGF-1 and Wnt/ß-catenin signaling (p < 0.05). Moreover, it altered the gut microbiota, suppressed inflammatory responses, and created a microenvironment towards suppressing tumorigenesis later in life.

Thymol Protects against 5-Fluorouracil-Induced Hepatotoxicity via the Regulation of the Akt/GSK-3ß Pathway in In Vivo and In Silico Experimental Models.

BACKGROUND: 5-fluorouracil (5-FU) is a widely used, highly effective chemotherapeutic agent. However, its therapeutic efficacy is often limited by associated adverse effects, with hepatotoxicity being frequently reported with 5-FU therapy. Thymol is a monoterpene found in thyme (Thymus vulgaris L., Lamiaceae) and is known for its antioxidant, anti-apoptotic, and anticancer activities. This study aimed to explore the hepatoprotective activity of thymol against 5-FU-induced liver injury. METHODS: Rats received two intraperitoneal doses of 5-FU (150 mg/kg) either alone or in combination with thymol at doses of 60 mg/kg or 120 mg/kg. Liver enzymes, oxidative stress, and apoptotic markers, in addition to histopathological changes, were assessed. RESULTS: 5-FU induced marked liver injuries as evidenced by elevated liver enzymes and histopathological changes, in addition to abnormalities of oxidative and apoptotic markers. The administration of thymol ameliorated the 5-FU-induced oxidative damage through increasing hepatic antioxidants and lowering lipid peroxidation. Apoptotic response markers such as Bax, Bcl-2, Bax/Bcl-2 ratio, and PARP were also improved. Furthermore, Western blotting analysis showed that thymol modulated the 5-FU-induced changes in the expression of Akt/GSK-3ß and p44/42 MAPK (ERK1/2) signaling pathways. CONCLUSIONS: Our research is the first to shed light on thymol's potential protective effect against 5-FU- induced hepatotoxicity by inhibiting oxidative and apoptotic pathways and modulating the Akt/ GSK-3ß as well as p44/42 MAPK (ERK1/2) signaling pathways.

Combined swimming with melatonin protects against behavioural deficit in cerebral ischemia-reperfusion injury induced rats associated with modulation of Mst1- MAPK -ERK signalling pathway.

BACKGROUND: The inconvenience of social and behavioural deficits after cerebral ischaemia reperfusion (I/R) injury is still not well documented. AIM: We aimed to study the protective effect of preconditioning swimming exercise combined with melatonin against cerebral I/R induced injury. METHODOLOGY: Sixty rats were allocated into 6 groups; groups I and II served as control. Groups 3,4,5,6 subjected to bilateral carotid ligation for 30 minutes (min.) followed by reperfusion. Group 3 left untreated while groups 4 and 6; underwent swimming exercise 30 min/day, five days a week for three weeks before the surgery. Groups 5 and 6 treated with melatonin 30 minutes before the operation, then, all rats in groups 4, 5,6 were subjected to I/R. After that, groups 5 and 6 treated with 2nd dose of melatonin 30 minutes after reperfusion. RESULTS: Combined strategy exhibited the most neuroprotective effect through prevention of cerebral I/R induced inflammation, oxidative stress and apoptosis with subsequent improvement in socio behaviour deficits and enhanced Glial cell proliferative capacity. CONCLUSION: The protective contribution of combined strategy is associated with modulation in Macrophage-stimulating 1/mitogen-activated protein kinase/extracellular signal-regulated kinase (MST1/MAPK/ERK) pathway which may explain, at least in part, its protective potential.

Preconditioning swimming exercise combined with melatonin protected against cerebral I/R induced socio-behavioural deficit.Cerebral I/R induced pathophysiological alterations in Prefrontal cortical neurons (PFC) are prevented by combined Preconditioning swimming exercise and melatoninCombined Preconditioning swimming exercise/melatonin in cerebral I/R modulates MST1/MAPK/ERK signalling pathwayCombined Preconditioning swimming exercise/melatonin inhibit inflammation, oxidative stress and apoptosis, thus enhance Glial cell proliferative capacity in I/R induced injury in PFC neurons.

Suppressive effect of isofraxidin on the overexpression of IL-6 and its molecular mechanism in a TPA-treated human hepatocellular carcinoma cell line, HuH-7.

Interleukin-6 (IL-6) is a pleiotropic cytokine that has many biological activities, including inflammation, hematopoiesis, bone metabolism, embryonic development, and other fundamental processes. Recently, IL-6 has been widely recognized as an important pro-inflammatory cytokine involved in cytokine storm pathogenesis during severe inflammatory diseases, such as coronavirus disease 2019 (COVID-19). Therefore, IL-6 is considered to be a therapeutic target for inhibiting cytokine storm. In the present study, we investigated the suppressive effect of isofraxidin, a major coumarin compound of Acanthopanax senticosus, on the overexpression of IL-6 and its molecular mechanism. The expression of IL-6 mRNA was measured using quantitative real-time PCR, and intracellular signaling molecules were detected using western blotting. When the HuH-7 human hepatocellular carcinoma cell line and HepG2 human hepatoblastoma cell line were treated with 12-O-tetradecanoylphorbol 13-acetate (TPA), a marked induction of IL-6 mRNA expression was observed in HuH-7 cells compared with HepG2 cells. Isofraxidin significantly suppressed TPA-induced IL-6 mRNA expression in HuH-7 cells in a dose-dependent manner. Furthermore, isofraxidin inhibited TPA-induced phosphorylation of ERK1/2 in a dose-dependent manner. Similarly, the MAPK/ERK inhibitor U0126 suppressed TPA-induced IL-6 mRNA expression. However, isofraxidin had no effects on TPA-induced phosphorylation of SAPK/JNK, Akt (Ser473), and STAT3 (Tyr705), nuclear translocation of NF-κB p65, and degradation of IκB. Taken together, isofraxidin suppresses TPA-induced overexpression of IL-6 mRNA by selectively inhibiting the activation of the MAPK/ERK pathway in HuH-7 cells, indicating that isofraxidin may be an effective anti-inflammatory agent for treating cytokine storm.

Two-polarized roles of transcription factor FOSB in lung cancer progression and prognosis: dependent on p53 status.

BACKGROUND: Activator protein-1 (AP-1) represents a transcription factor family that has garnered growing attention for its extensive involvement in tumor biology. However, the roles of the AP-1 family in the evolution of lung cancer remain poorly characterized. FBJ Murine Osteosarcoma Viral Oncogene Homolog B (FOSB), a classic AP-1 family member, was previously reported to play bewilderingly two-polarized roles in non-small cell lung cancer (NSCLC) as an enigmatic double-edged sword, for which the reasons and significance warrant further elucidation. METHODS AND RESULTS: Based on the bioinformatics analysis of a large NSCLC cohort from the TCGA database, our current work found the well-known tumor suppressor gene TP53 served as a key code to decipher the two sides of FOSB - its expression indicated a positive prognosis in NSCLC patients harboring wild-type TP53 while a negative one in those harboring mutant TP53. By constructing a panel of syngeneically derived NSCLC cells expressing p53 in different statuses, the radically opposite prognostic effects of FOSB expression in NSCLC population were validated, with the TP53-R248Q mutation site emerging as particularly meaningful. Transcriptome sequencing showed that FOSB overexpression elicited diversifying transcriptomic landscapes across NSCLC cells with varying genetic backgrounds of TP53 and, combined with the validation by RT-qPCR, PREX1 (TP53-Null), IGFBP5 (TP53-WT), AKR1C3, and ALDH3A1 (TP53-R248Q) were respectively identified as p53-dependent transcriptional targets of FOSB. Subsequently, the heterogenous impacts of FOSB on the tumor biology in NSCLC cells via the above selective transcriptional targets were confirmed in vitro and in vivo. Mechanistic investigations revealed that wild-type or mutant p53 might guide FOSB to recognize and bind to distinct promoter sequences via protein-protein interactions to transcriptionally activate specific target genes, thereby creating disparate influences on the progression and prognosis in NSCLC. CONCLUSIONS: FOSB expression holds promise as a novel prognostic biomarker for NSCLC in combination with a given genetic background of TP53, and the unique interactions between FOSB and p53 may serve as underlying intervention targets for NSCLC.

Update on the Genetics of Osteogenesis Imperfecta.

Osteogenesis imperfecta (OI) is a heterogeneous heritable skeletal dysplasia characterized by bone fragility and deformity, growth deficiency, and other secondary connective tissue defects. OI is now understood as a collagen-related disorder caused by defects of genes whose protein products interact with collagen for folding, post-translational modification, processing and trafficking, affecting bone mineralization and osteoblast differentiation. This review provides the latest updates on genetics of OI, including new developments in both dominant and rare OI forms, as well as the signaling pathways involved in OI pathophysiology. There is a special emphasis on discoveries of recessive mutations in TENT5A, MESD, KDELR2 and CCDC134 whose causality of OI types XIX, XX, XXI and XXI, respectively, is now established and expends the complexity of mechanisms underlying OI to overlap LRP5/6 and MAPK/ERK pathways. We also review in detail new discoveries connecting the known OI types to each other, which may underlie an eventual understanding of a final common pathway in OI cellular and bone biology.

The use of matrine to inhibit osteosarcoma cell proliferation via the regulation of the MAPK/ERK signaling pathway.

Background: Matrine is an alkaloid extracted from Sophorus beans of the legume family, and it has significant effects and a variety of pharmacological activities. Osteosarcoma(OS) is a common malignant bone tumor that is characterized by high incidence and rapid progression. There have been some preliminary studies on the therapeutic effect of matrine on OS, but the specific mechanism remains unclear. Objective: The aim of this study was to investigate the antitumor effect of matrine on HOS cells and the underlying molecular mechanism. Methods: The effects of matrine on the proliferation, apoptosis and cell cycle progression of HOS cells were determined by CCK-8 assay, TUNEL assay and flow cytometry in vitro. Wound healing and Transwell invasion assays were used to observe the effect of matrine on the migration and invasion of HOS cells. The mechanism underlying the antitumor effect of matrine on HOS cells was investigated by Western blotting. Results: Matrine significantly inhibited HOS cell proliferation, promoted HOS cell apoptosis, and arrested HOS cells in the G1 phase of the cell cycle. Both wound healing and Transwell invasion assays showed that matrine inhibited HOS cell migration and invasion. Western blotting results showed that matrine inhibited the activation of the MAPK/ERK signaling pathway. We found that matrine also downregulated Bcl-2 expression, which may be related to protein synthesis inhibition. Conclusion: Matrine can inhibit the proliferation of HOS cells, arrest HOS cells in the G1 phase, and promote HOS cell apoptosis through the MAPK/ERK signaling pathway.

Glypican-3 deficiency in liver cancer upregulates MAPK/ERK pathway but decreases cell proliferation.

Glypican-3 (GPC3) is overexpressed in hepatocellular carcinomas and hepatoblastomas and represents an important therapeutic target but the biologic importance of GPC3 in liver cancer is unclear. To date, there are limited data characterizing the biological implications of GPC3 knockout (KO) in liver cancers that intrinsically express this target. Here, we report on the development and characterization of GPC3-KO liver cancer cell lines and compare to them to parental lines. GPC3-KO variants were established in HepG2 and Hep3B liver cancer cell lines using a lentivirus-mediated CRISPR/Cas9 system. We assessed the effects of GPC3 deficiency on oncogenic properties in vitro and in murine xenograft models. Downstream cellular signaling pathway changes induced by GPC3 deficiency were examined by RNAseq and western blot. To confirm the usefulness of the models for GPC3-targeted drug development, we evaluated the target engagement of a GPC3-selective antibody, GC33, conjugated to the positron-emitting zirconium-89 (89Zr) in subcutaneous murine xenografts of wild type (WT) and KO liver cancer cell lines. Deletion of GPC3 significantly reduced liver cancer cell proliferation, migration, and invasion compared to the parental cell lines. Additionally, the tumor growth of GPC3-KO liver cancer xenografts was significantly slower compared with control xenografts. RNA sequencing analysis also showed GPC3-KO resulted in a reduction in the expression of genes associated with cell cycle regulation, invasion, and migration. Specifically, we observed the downregulation of components in the AKT/NFκB/WNT signaling pathways and of molecules related to cell cycle regulation with GPC3-KO. In contrast, pMAPK/ERK1/2 was upregulated, suggesting an adaptive compensatory response. KO lines demonstrated increased sensitivity to ERK (GDC09994), while AKT (MK2206) inhibition was more effective in WT lines. Using antibody-based positron emission tomography (immunoPET) imaging, we confirmed that 89Zr-GC33 accumulated exclusively in GPC3-expression xenografts but not in GPC3-KO xenografts with high tumor uptake and tumor-to-liver signal ratio. We show that GPC3-KO liver cancer cell lines exhibit decreased tumorigenicity and altered signaling pathways, including upregulated pMAPK/ERK1/2, compared to parental lines. Furthermore, we successfully distinguished between GPC3+ and GPC3- tumors using the GPC3-targeted immunoPET imaging agent, demonstrating the potential utility of these cell lines in facilitating GPC3-selective drug development.

KLRB1 expression is associated with lung adenocarcinoma prognosis and immune infiltration and regulates lung adenocarcinoma cell proliferation and metastasis through the MAPK/ERK pathway.

Background: Lung cancer is the most common primary malignant tumor of the lung, and as one of the malignant tumors that pose the greatest threat to the health of the population, the incidence rate has remained high in recent years. Previous studies have shown that KLRB1 is transcriptionally repressed in lung adenocarcinoma and correlates with lung adenocarcinoma prognosis. The objective of this study is to investigate the intrinsic mechanisms by which KLRB1 affects the malignant phenotypes of lung adenocarcinoma such as immune infiltration, proliferation, growth and metastasis. Methods: We assessed the expression levels of KLRB1 in publicly available databases and investigated its associations with clinical and pathological variables. Enrichment analysis was subsequently conducted to investigate possible signaling pathways and their associated biological functions. Statistical analysis, including Spearman correlation and the application of multigene prediction models, was utilized to assess the relationship between the expression of KLRB1 and the infiltration of immune cells. The diagnostic and prognostic value of KLRB1 was evaluated using Kaplan-Meier survival curves, diagnostic receptor operating characteristic (ROC) curves, histogram models, and Cox regression analysis. Specimens from lung adenocarcinoma (LUAD) patients were collected, the expression level of KLRB1 was detected by protein blotting analysis, and the expression level of KLRB1 was detected at the mRNA level by real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR). Small interfering RNA (siRNA) was used to silence gene expression, and Transwell, Cell Counting Kit-8 (CCK-8) and colony formation assays were subsequently performed to analyze the effects of KLRB1 on LUAD cell migration, invasion and proliferation. Results: KLRB1 expression was lower in lung cancer tissue than in surrounding healthy tissue. Genes differentially expressed in the low and high KLRB1 expression groups were found to be significantly enriched in pathways related to immunity. KLRB1 exerted an impact on the MAPK/ERK signaling pathway, thereby modulating the growth and proliferation of LUAD cells. KLRB1 expression is linked to prognosis, immune infiltration, and cell migration and proliferation in LUAD. Conclusions: The evidence revealed a correlation between KLRB1 and both prognosis and immune infiltration in LUAD patients.

Repurposing Anticancer Drugs Targeting the MAPK/ERK Signaling Pathway for the Treatment of Respiratory Virus Infections.

Respiratory virus infections remain a significant challenge to human health and the social economy. The symptoms range from mild rhinitis and nasal congestion to severe lower respiratory tract dysfunction and even mortality. The efficacy of therapeutic drugs targeting respiratory viruses varies, depending upon infection time and the drug resistance engendered by a high frequency of viral genome mutations, necessitating the development of new strategies. The MAPK/ERK pathway that was well delineated in the 1980s represents a classical signaling cascade, essential for cell proliferation, survival, and differentiation. Since this pathway is constitutively activated in many cancers by oncogenes, several drugs inhibiting Raf/MEK/ERK have been developed and currently used in anticancer treatment. Two decades ago, it was reported that viruses such as HIV and influenza viruses could exploit the host cellular MAPK/ERK pathway for their replication. Thus, it would be feasible to repurpose this category of the pathway inhibitors for the treatment of respiratory viral infections. The advantage is that the host genes are not easy to mutate such that the drug resistance rarely occurs during short-period treatment of viruses. Therefore, in this review we will summarize the research progress on the role of the MAPK/ERK pathway in respiratory virus amplification and discuss the potential of the pathway inhibitors (MEK inhibitors) in the treatment of respiratory viral infections.

SIRT6 Inhibits the Proliferation and Collagen Synthesis of Keloid Fibroblasts through MAPK/ERK Pathway.

BACKGROUND: Keloid, a fibroproliferative disorder, significantly impacts patients' quality of life, yet effective therapies remain elusive. This study explored the role of silent information regulator 6 (SIRT6) in modulating the proliferation, invasion, and collagen synthesis of keloid fibroblasts. METHODS: Keloid and normal skin specimens were collected, and fibroblasts were isolated from the keloid tissue. SIRT6 recombinant adenovirus (Ad) was constructed to infect keloid fibroblasts to overexpress SIRT6. This study entails three groups: Control group, adenovirus-Negative Control (Ad-NC) group, and Ad-SIRT6 group. SIRT6 protein and mRNA levels were measured via Western blotting and Quantitative reverse transcription polymerase chain reaction (qRT-PCR), respectively. Cell viability was determined using 5-ethynyl-2'-deoxyuridine (EdU) assay. Flow cytometry was exploited to measure cell apoptosis. To investigate cell migration, wound healing assay and Transwell assay were employed. Western blotting was also utilized to study the expression levels of apoptotic proteins, collagen deposition-related proteins, and Mitogen-Activated Protein Kinases (MAPK)/extracellular regulated protein kinases (ERK) pathway-related proteins. RESULTS: Compared to the control and Ad-NC groups, the Ad-SIRT6 group exhibited significantly elevated SIRT6 level; diminished cell proliferation, migration and invasion; reduced protein levels of α-smooth muscle actin (α-SMA), collagen I, collagen III, phospho SMAD Family Member 3 (p-Smad3), transforming growth factor-ß 1 (TGF-ß1), and MAPK/ERK pathway proteins (phospho extracellular signal-regulated protein kinase 1/2 (p-ERK1/2), phospho MAP kinase-ERK kinase (p-MEK) and phospho-c-Raf (p-c-Raf)). Treatment with epidermal growth factor (EGF), an MAPK/ERK pathway agonists, reversed the inhibitory effect of SIRT6 on cell activity and inhibited apoptosis in keloid fibroblasts. CONCLUSION: SIRT6 overexpression in keloid fibroblasts attenuates proliferation, invasion, and collagen synthesis, while fostering apoptosis, likely through the suppression of MAPK/ERK pathway activity. This suggests a potential therapeutic target for keloid treatment.

MARK4 promotes the malignant phenotype of gastric cancer through the MAPK/ERK signaling pathway.

BACKGROUND: Microtubule affinity regulating kinase 4 (MARK4), which is overexpressed in various tumors, is involved in the regulation of cell division, proliferation, migration, and the cell cycle, and has been considered a potential marker for cancer; however, its mechanism of action in gastric cancer (GC) remains unclear. This study aimed to investigate the role of MARK4 in the proliferation, migration, and invasion of GC cell through the MAPK/ERK signaling pathway by targeting MARK4 knockdown. METHODS: Using The Cancer Genome Atlas data and clinical information, MARK4 expression and its relationship with prognosis were analyzed. Possible pathways involving MARK4 were explored using enrichment analysis. Western blotting and real-time quantitative polymerase chain reaction were used to detect MARK4 expression in GC. After targeted transfection of siRNA, the transfection efficiency of the experimental group was detected in AGS and HGC-27 cells. The effects of knockdown MARK4 on the proliferation, migration, and invasion of GC cells were verified using CCK-8, colony formation, wound healing, and transwell assays. Finally, the relationship between MARK4, the MAPK/ERK pathway, and epithelial-mesenchymal transition in GC was verified by western blotting. RESULTS: MARK4 expression was upregulated in GC and associated with poor prognosis in patients with GC. Enrichment analysis showed that MARK4 was involved in the activation of the MAPK signaling pathway. Western blotting results indicated that MARK4 overexpression promoted the proliferation, migration, and invasion of GC cells through the MAPK/ERK pathway. CONCLUSION: MARK4 expression was upregulated in GC and promoted the proliferation, migration, and invasion of GC cells through the MAPK/ERK pathway.

ZNF300 promotes proliferation and migration of hepatocellular carcinoma by upregulating c-MYC gene expression.

BACKGROUND: Hepatocellular carcinoma (HCC) is the most common primary malignant tumor of the liver. Currently, the treatments of HCC are limited to surgical resection and liver transplantation, and there is no effective systemic therapy. OBJECTIVES: To investigate the regulatory mechanism of zinc finger protein 300 (ZNF300) in hepatocellular carcinoma (HCC). METHODS: The expressions of ZNF300 in HCC tissue samples and HCC cell lines (Hep3B, Huh7, SNU-387) were detected. ZNF300 overexpression vector (ZNF300) or shRNAZNF300 (shZNF300) was transfected into HCC cells to increase or inhibit ZNF300 expression. 5-Ethynyl-2'-deoxyuridine assay (EdU), cell counting kit-8 assay (CCK-8) and transwell invasion assay were conducted to evaluate the proliferation, viability, migration, and invasion of HCC cells respectively. The expressions of tumor migration and invasion related proteins (matrix metallopeptidase 2 (MMP-2) and MMP-9), c-MYC, and MAPK/ERK signaling pathway related molecules (p-ERK1/2, ERK1/2, p-P38, P38) were determined by western blotting. Hep3B cells transfected with shZNF300 were subcutaneously injected into nude mice to perform tumor xenograft experiment. Tumor volume and weight were measured. RESULTS: ZNF300 was upregulated in HCC tissues and cells. The expressions of MMP-2 and MMP-9 were increased in HCC cells after transfecting with ZNF300 but reduced in HCC cells transfected with shZNF300. Downregulation of ZNF300 inhibited HCC cell proliferation, migration, and invasion, while overexpression of ZNF300 showed the opposite effects. Moreover, the expressions of c-MYC and MAPK/ERK signaling pathway related molecules were increased after overexpression of ZNF300 but reduced after downregulating ZNF300. In tumor xenograft experiment, downregulation of ZNF300 reduced tumor volume and weight. CONCLUSION: The present study proved that downregulation of ZNF300 inhibited HCC growth by reducing c-MYC expression and MAPK/ERK signaling pathway.

SPRED2 Is a Novel Regulator of Autophagy in Hepatocellular Carcinoma Cells and Normal Hepatocytes.

Sprouty-related enabled/vasodilator-stimulated phosphoprotein homology 1 domain containing 2 (SPRED2) is an inhibitor of the mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) pathway and has been shown to promote autophagy in several cancers. Here, we aimed to determine whether SPRED2 plays a role in autophagy in hepatocellular carcinoma (HCC) cells. The Cancer Genome Atlas (TCGA) Liver Cancer Database showed a negative association between the level of SPRED2 and p62, a ubiquitin-binding scaffold protein that accumulates when autophagy is inhibited. Immunohistochemically, accumulation of p62 was detected in human HCC tissues with low SPRED2 expression. Overexpression of SPRED2 in HCC cells increased the number of autophagosomes and autophagic vacuoles containing damaged mitochondria, decreased p62 levels, and increased levels of light-chain-3 (LC3)-II, an autophagy marker. In contrast, SPRED2 deficiency increased p62 levels and decreased LC3-II levels. SPRED2 expression levels were negatively correlated with translocase of outer mitochondrial membrane 20 (TOM20) expression levels, suggesting its role in mitophagy. Mechanistically, SPRED2 overexpression reduced ERK activation followed by the mechanistic or mammalian target of rapamycin complex 1 (mTORC1)-mediated signaling pathway, and SPRED2 deficiency showed the opposite pattern. Finally, hepatic autophagy was impaired in the liver of SPRED2-deficient mice with hepatic lipid droplet accumulation in response to starvation. These results indicate that SPRED2 is a critical regulator of autophagy not only in HCC cells, but also in hepatocytes, and thus the manipulation of this process may provide new insights into liver pathology.

hMAGEA2 Accelerates the Progression of Prostate Cancer via the EFNA3-Erk1/2 Signaling Pathway.

BACKGROUND/AIM: Human melanoma-associated antigen A2 (hMAGEA2) family members play several roles in many types of cancer and have been explored as potential prognostic markers. In this study, we investigated the molecular mechanism underlying hMAGEA2-mediated tumorigenesis of prostate cancer. MATERIALS AND METHODS: Immunohistochemistry and western blot were used to assess protein expression whereas microarray and quantitative reverse transcription-PCR determined mRNA expression. CCK-8 assay was used to determine cell proliferation. Colony formation assay was used to examine tumorigenesis. Migration and invasion were examined using a transwell assay. Propidium iodide (PI)/Annexin V double staining was performed to measure apoptosis. Transcriptional activity was measured using Dual-luciferase reporter assay. RESULTS: hMAGEA2 was highly over-expressed in human prostate cancer tissues compared to benign prostatic hyperplasia tissues. To elucidate its biological function in prostate cancer, we established two stable hMAGEA2-knockdown prostate cancer cell lines, PC3M and 22RV1, and found that they presented significantly decreased proliferation, anchorage-independent colony formation, migration, and invasion. As hMAGEA2 knockdown suppressed prostate cancer cell growth, we examined its potential influence on tumor apoptosis. hMAGEA2-knockdown cell lines displayed early apoptosis. Moreover, knockdown of hMAGEA2 resulted in the down-regulation of EFNA3 expression. Luciferase assay showed that hMAGEA2 bound to the EFNA promoter region and regulated its transcription. Down-regulation of EFNA3 expression led to decreased Ras/Braf/MEK/Erk1/2 phosphorylation and, consequently, inhibited prostate cancer progression. CONCLUSION: hMAGEA2 promotes prostate cancer growth, metastasis, and tumorigenesis by regulating the EFNA3-Erk1/2 signaling pathway, indicating its potential as a therapeutic marker for prostate cancer.