Knockdown of Bcl-3 alleviates psoriasis and dyslipidemia comorbidity by regulating Akt pathway

Background Psoriasis is considered as an inflammatory skin disease accompanied by dyslipidemia comorbidity. B-cell leukemia-3 (Bcl-3) belongs to IκB (inhibitor of nuclear factor kappa B [NF-κB]) family, and regulates inflammatory response through associating with NF-κB. The role of Bcl-3 in psoriasis was investigated in this study. Methods Apolipoprotein E (ApoE)-deficient mice were treated with imiquimod to induce psoriasis and dyslipidemia. Mice were injected intradermally in the back with lentiviral particles encoding Bcl-3 small hairpin RNA (shRNA). Hematoxylin and eosin were used to detect pathological characteristics. The blood lipid levels were determined by automatic biochemical analyzer, and inflammation was assessed by enzyme-linked-immunosorbent serologic assay and real-time quantitative reverse transcription polymerase chain reaction. Results Bcl-3 was elevated in imiquimod-induced ApoE-deficient mice. Injection with lentiviral particles encoding Bcl-3 shRNA reduced Psoriasis area and severity index (PASI) score in ApoE-deficient psoriatic mice. Knockdown of Bcl-3 also ameliorated imiquimod-induced psoriasiform skin lesions in ApoE-deficient mice. Moreover, loss of Bcl-3 enhanced expression of loricrin, an epidermal barrier protein, reduced expression of proliferating cell nuclear antigen (PCNA) and lectin-like oxidized LDL (oxLDL) receptor-1 (LOX-1) in imiquimod-induced ApoE-deficient mice. The enhanced levels of blood lipid in ApoE-deficient mice were attenuated by silencing of Bcl-3 with increase of high-density lipoprotein, and reduction of total cholesterol, triglycerides, and low-density lipoprotein cholesterol. Knockdown of Bcl-3 attenuated imiquimod-induced decrease of transforming growth factor beta (TGF-β), and increase of Interleukin (IL)-17A, IL-23, IL-6, and tumor necrosis factor-α (TNF-α) in ApoE-deficient mice. Protein expression of phospho-Akt (p-Akt) and p-GSK3β in ApoE-deficient psoriatic mice was decreased by silencing of Bcl-3 (AU)

CRYAB reduces cigarette smoke-induced inflammation, apoptosis, and oxidative stress by retarding PI3K/Akt and NF-kB signaling pathways in human bronchial epithelial cells

Background: Chronic obstructive pulmonary disease (COPD) is a familiar airway disease characterized by chronic immune response in the lungs. More and more evidences have assured that cigarette smoking is the primary reason for the progression of COPD, but its related regulatory mechanism requires further clarification. The α-B-crystallin (CRYAB) has been identified to exhibit vital functions in different diseases, and is down-regulated in the alveoli of mice mediated by cigarette smoke extract (CSE).Methods: The messenger RNA expression of CRYAB was assessed by reverse transcription--quantitative polymerase chain reaction. The proteins’ expressions were tested using Western blot method. The cytotoxicity was measured by lactate dehydrogenase assay. The levels of malondialdehyde, superoxide dismutase, catalase, myeloperoxidase, tumor necrosis factor-α, interleukin (IL)-1β, and IL-6 were assessed through enzyme-linked-immunosorbent serologic assay (ELISA).Results: In this study, it was discovered that the expression of CRYAB was markedly decreased with the increased time of cigarette smoking. Moreover, CRYAB overexpression increased cell viability and decreased cell apoptosis induced by cigarette smoke. In addition, the strengthened oxidative stress and inflammation mediated by CSE treatment was relieved after overexpression of CRYAB. Eventually, results OF Western blot method confirmed that CRYAB retarded the activation of phosphatidylinositol 3-kinase–Ak strain transforming (PI3K–Akt) and nuclear factor kappa B (NF-κB) signaling pathways.Conclusion: Our results manifested that CRYAB reduced cigarette smoke-induced inflammation, apoptosis, and oxidative stress in normal and diseased bronchial epithelial (NHBE) and human bronchial epithelial (BEAS-2B) cells by suppressing PI3K/Akt and NF-κB signaling pathways, which highlighted the functioning of CRYAB in preventing or treating COPD (AU)

Disruption of the IL-33-ST2-AKT signaling axis impairs neurodevelopment by inhibiting microglial metabolic adaptation and phagocytic function.

To accommodate the changing needs of the developing brain, microglia must undergo substantial morphological, phenotypic, and functional reprogramming. Here, we examined whether cellular metabolism regulates microglial function during neurodevelopment. Microglial mitochondria bioenergetics correlated with and were functionally coupled to phagocytic activity in the developing brain. Transcriptional profiling of microglia with diverse metabolic profiles revealed an activation signature wherein the interleukin (IL)-33 signaling axis is associated with phagocytic activity. Genetic perturbation of IL-33 or its receptor ST2 led to microglial dystrophy, impaired synaptic function, and behavioral abnormalities. Conditional deletion of Il33 from astrocytes or Il1rl1, encoding ST2, in microglia increased susceptibility to seizures. Mechanistically, IL-33 promoted mitochondrial activity and phagocytosis in an AKT-dependent manner. Mitochondrial metabolism and AKT activity were temporally regulated in vivo. Thus, a microglia-astrocyte circuit mediated by the IL-33-ST2-AKT signaling axis supports microglial metabolic adaptation and phagocytic function during early development, with implications for neurodevelopmental and neuropsychiatric disorders.

Close association between the synergistic toxicity of zearalenone-deoxynivalenol combination and microRNA221-mediated PTEN/PI3K/AKT signaling in HepG2 cells.

Mycotoxins can impart different types of combined toxicity to humans and animals, therefore, it is critical to understand the underlying mechanisms to eliminate the harm. Herein a combination of zearalenone (ZEA) at 2 µM and deoxynivalenol (DON) at 0.1 µM decreased cell viability and increased ROS level in HepG2 cells, suggesting synergistic toxicity exerted by ZEA and DON even at their low toxic concentrations. Moreover, apoptosis and inflammatory response were promoted after the co-exposure of ZEA and DON, indicated by the increased expression of BAX, Caspase-3, IL-1ß and IL-6 genes. Such synergistic toxicity was closely associated with miR-221-mediated PTEN/PI3K/AKT signal pathway, with a negative regulatory relationship between PTEN and PI3K/AKT signaling. MiR-221 could influence cell viability and ROS level to counter the combined toxicity of ZEA and DON through targeting directly PTEN gene. This study demonstrated the toxicological impact of mycotoxin interactions on cells, and critical role of the interplay between miRNAs and PTEN in monitoring the synergistic toxicity of mycotoxin mixture.

Polycyclic aromatic hydrocarbons in bone homeostasis.

Prolonged exposure to polycyclic aromatic hydrocarbons (PAHs) may result in autoimmune diseases, such as rheumatoid arthritis (RA) and osteoporosis (OP), which are based on an imbalance in bone homeostasis. These diseases are characterized by bone erosion and even a disruption in homeostasis, including in osteoblasts and osteoclasts. Current evidence indicates that multiple factors affect the progression of bone homeostasis, such as genetic susceptibility and epigenetic modifications. However, environmental factors, especially PAHs from various sources, have been shown to play an increasingly prominent role in the progression of bone homeostasis. Hence, it is essential to investigate the effects and pathogenesis of PAHs in bone homeostasis. In this review, recent progress is summarized concerning the effects and mechanisms of PAHs and their ligands and receptors in bone homeostasis. Moreover, strategies based on the effects and mechanisms of PAHs in the regulation of the bone balance and alleviation of bone destruction are also reviewed. We further discuss the future challenges and perspectives regarding the roles of PAHs in autoimmune diseases based on bone homeostasis.

Dihydroartemisinin inhibits Lewis Lung carcinoma progression by inducing macrophages M1 polarization via AKT/mTOR pathway.

Preclinical and clinical data show a close relationship between high infiltration of tumor-associated macrophages (TAMs) and a poor prognosis in most types of tumors, thus targeting TAMs stands out as promising anticancer immunotherapies. Recent studies have demonstrated the anti-tumor effects of artemisinin via enhancing anti-tumor immunity within tumor microenvironment, but the underlying mechanism is still not clear. In the present study we uncovered an important role of dihydroartemisinin (DHA) in regulating intratumoral TAM polarization and anti-tumor immune responses in mouse Lewis Lung carcinoma model. We found that DHA inhibited Lewis Lung carcinoma progress, moderately decreased the frequencies of TAMs within tumor stroma, and significantly increased CD86 expression while decreased CD206 expression on TAMs which indicates the role of DHA in polarizing TAMs into a M1-like phenotype. Then, our in vitro data confirmed that DHA dose-dependently promoted macrophage M1 phenotype transition by increasing M1 phenotype-related molecules, meanwhile decreasing the expression of M2 phenotype-related molecules. In addition, DHA increased proinflammatory cytokine production, enhanced the phagocytic capacity while decreased anti-inflammatory cytokine production. Finally, in order to prove that AKT/mTOR signaling potentially mediated DHA-induced macrophage differentiation, we used rapamycin to specifically block the activity of mTOR and stimulated macrophages under M1 stimuli. Our data clearly showed that rapamycin significantly decreased DHA-induced M1-related phenotypes and proinflammatory cytokine expression. In summary, our study highlighted DHA as one of future potential therapeutic options for the development of novel anticancer immunotherapies in lung cancer.

Jolkinolide B inhibits proliferation or migration and promotes apoptosis of MCF-7 or BT-474 breast cancer cells by downregulating the PI3K-Akt pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: The diterpenoids extracted from Euphorbia kansui S.L. Liou ex S.B.Ho, Euphorbia fischeriana Steud. have good antitumor effects. Jolkinolide B has anti-breast cancer effect, but it is unclear whether it has different therapeutic effects between luminal A subtype and luminal B subtype breast cancer. AIM OF THE STUDY: This study investigated the Jolkinolide B has different therapeutic, important targets and pathways effects between luminal A subtype and luminal B subtype breast cancer. MATERIALS AND METHODS: We used bioinformatics to predict the biological process and molecular mechanism of Jolkinolide B in treating two types of breast cancer. Then, in vitro, cultured MCF-7 cells and BT-474 cells were divided into control group, PI3K inhibitor + control group, Jolkinolide B group and PI3K inhibitor + Jolkinolide B group. The CCK-8 assay, Flow cytometric analysis and Transwell cell migration assay was used to detect the cell proliferation, apoptosis, and migration in each group, respectively. ELISA was used to measure the content of Akt and phosphorylated Akt (p-Akt) in cell lysis buffer. RESULTS: Compared to luminal A breast cancer, Jolkinolide B had more targets, proliferation, migration processes and KEGG pathways when treating luminal B subtype breast cancer. Jolkinolide B significantly prolonged the survival time of luminal B subtype breast cancer patients. Compared to the control group, the cell proliferation absorbance value (A value) and migration number of the two kinds of breast cancer cells in the Jolkinolide B group were decreased (P < 0.01, n = 6), and the number of apoptotic cells was increased (P < 0.01, n = 6). Compared to the Jolkinolide B group, the A value and migration number of the two types of breast cancer cells were significantly decreased in the PI3K inhibitor + Jolkinolide B group (P < 0.01, n = 6), and the number of apoptotic cells was significantly increased (P < 0.01, n = 6). In addition, compared to MCF-7 cells, the A value and migration number of BT-474 cells stimulated with Jolkinolide B were significantly decreased (P < 0.01, n = 6), and the number of apoptotic cells was significantly increased (P < 0.01, n = 6). Akt and p-Akt protein levels in the two breast cancer cell lines in the Jolkinolide B group were all decreased (P < 0.01, n = 6), especially in BT-474 cells stimulated by Jolkinolide B. CONCLUSION: Jolkinolide B regulates the luminal A and luminal B subtypes of breast cancer through PI3K-Akt, EGFR and other pathways. Jolkinolide B has more significant therapeutic effect on luminal B subtype breast cancer. In vitro, experiments verified that Jolkinolide B significantly inhibited the proliferation and migration activity of BT-474 breast cancer cells by downregulating the PI3K-Akt pathway.

Tricin attenuates the progression of LPS-induced severe pneumonia in bronchial epithelial cells by regulating AKT and MAPK signaling pathways

Background Pneumonia is a continuous and widespread disease with higher incidence, the effects of it on human life can be fearful. Tricin has been demonstrated to take part in the progression and development of diseases. However, the function of Tricin and its related regulatory pathways remain unclear. This study was planned to investigate the effects of Tricin on severe pneumonia.Methods The cell viability was detected through CCK-8 assay. The TNF-α, IL-1β and IL-6 levels were assessed through ELISA and RT-qPCR. The levels of MDA, SOD and GSH were tested through corresponding commercial kits. The protein expressions were examined through western blot.Results In our study, the lipopolysaccharide (LPS) was firstly used to stimulate cell model for severe pneumonia. We discovered that Tricin had no toxic effects on BEAS-2B cells and the decreased cell viability induced by LPS was relieved by a dose-dependent Tricin treatment. Additionally, through ELISA and RT-qPCR, it was uncovered that Tricin reduced the LPS-induced inflammation through regulating TNF-α, IL-1β and IL-6. Furthermore, Tricin relieved LPS-induced oxidative stress through reducing MDA level and enhancing SOD and GSH levels. Finally, it was demonstrated that Tricin retarded LPS-activated AKT and MAPK pathways.Conclusion Our findings revealed that Tricin attenuated the progression of LPS induced severe pneumonia through modulating AKT and MAPK signaling pathways. This discovery might afford one novel sight for the treatment of severe pneumonia (AU)

TMEM176B Regulates AKT/mTOR Signaling and Tumor Growth in Triple-Negative Breast Cancer.

TMEM176B is a member of the membrane spanning 4-domains (MS4) family of transmembrane proteins, and a putative ion channel that is expressed in immune cells and certain cancers. We aimed to understand the role of TMEM176B in cancer cell signaling, gene expression, cell proliferation, and migration in vitro, as well as tumor growth in vivo. We generated breast cancer cell lines with overexpressed and silenced TMEM176B, and a therapeutic antibody targeting TMEM176B. Proliferation and migration assays were performed in vitro, and tumor growth was evaluated in vivo. We performed gene expression and Western blot analyses to identify the most differentially regulated genes and signaling pathways in cells with TMEM176B overexpression and silencing. Silencing TMEM176B or inhibiting it with a therapeutic antibody impaired cell proliferation, while overexpression increased proliferation in vitro. Syngeneic and xenograft tumor studies revealed the attenuated growth of tumors with TMEM176B gene silencing compared with controls. We found that the AKT/mTOR signaling pathway was activated or repressed in cells overexpressing or silenced for TMEM176B, respectively. Overall, our results suggest that TMEM176B expression in breast cancer cells regulates key signaling pathways and genes that contribute to cancer cell growth and progression, and is a potential target for therapeutic antibodies.

Cross-regulation of notch/AKT and serum/glucocorticoid regulated kinase 1 (SGK1) in IL-4-stimulated human macrophages.

Notch signaling regulates the responses of macrophages to different stimuli in a context-dependent manner. The roles of Notch signaling in proinflammatory macrophages are well characterized, whereas its involvement, if any, in IL-4-stimulated macrophages (M(IL-4)) is still unclear. We observed that Notch signaling is functional in human M(IL-4). We performed transcriptome analysis of the Notch1 intracellular domain (NIC1)-overexpressing human monocytic cell line THP-1 with or without IL-4 stimulation to understand the global impact of Notch signaling in M(IL-4). The results revealed that NIC1-overexpressing THP-1 upregulated proinflammatory-associated genes and target genes of IL-4 signaling. We identified serum/glucocorticoid regulated kinase 1 (SGK1) as one of the genes increased by NIC1 overexpression in M(IL-4). To dissect the signaling pathway leading to SGK1 upregulation, we pretreated THP-1-derived macrophages with specific inhibitors of Notch (DAPT), AKT (LY294002) or ERK (U0126). Among these inhibitors, only LY294002 decreased the SGK1 mRNA levels in M(IL-4), indicating that the AKT pathway plays a key role in SGK1 transcription in M(IL-4). Furthermore, treatment of THP-1-derived macrophages with the SGK1 inhibitor (GSK650394) suppressed AKT phosphorylation, but not STAT6, in response to IL-4, indicating that SGK1 positively regulates AKT pathway in M(IL-4). Finally, GSK650394 treatment of human M(IL-4) increased the levels of PPARG mRNA and its protein, indicating a negative role of SGK1 in M(IL-4) function. Overall, we report that the Notch signaling and AKT pathways cooperatively regulate SGK1 expression in M(IL-4) where SGK1, in turn, plays an important role in suppressing IL-4-induced PPARγ expression.

MiRNA-494-3p Regulates Bupivacaine-Induced Neurotoxicity by the CDK6-PI3K/AKT Signaling.

Bupivacaine (BUP) is a long-acting amide local anesthetic that may induce strong neurotoxicity and neurological complications. In this study, we elucidate the influence of microRNA-494-3p (miR-494-3p) in BUP-induced neurotoxicity in primary mouse hippocampal neuronal cells. In this study, primary hippocampal neurons were isolated from neonatal C57BL/6 mice. The isolated neurons were treated with various doses of BUP. MTT assay was conducted to analyze neuronal viability. Gene expression measurement was done by RT-qPCR. The impact of miR-494-3p in BUP-mediated neural injury was examined using TUNEL, flow cytometry, western blotting, and ROS activity detection. The regulatory relationship between miR-494-3p and cyclin-dependent kinases 4 and 6 (CDK6) was identified using a luciferase reporter assay. BUP treatment led to neurotoxicity and miR-494-3p upregulation in primary cultured hippocampal neurons. Functionally, miR-494-3p depletion alleviated neuronal apoptosis and oxidative damage induced by BUP. We verified that miR-494-3p targeted and negatively modulated CDK6. MiR-494-3p depletion also activated PI3K/AKT signaling by elevating CDK6 expression in BUP-treated neurons. Furthermore, CDK6 knockdown or PI3K/AKT inactivation attenuated the neuroprotective role of miR-494-3p depletion. Silencing miR-494-3p exerts neuroprotective function in hippocampal neuronal cells against BUP-induced injury by the CDK6-PI3K/AKT pathway.

Senescence marker protein 30 (SMP30) protects against high glucose-induced apoptosis, oxidative stress and inflammatory response in retinal ganglion cells by enhancing Nrf2 activation via regulation of Akt/GSK-3ß pathway.

Senescence marker protein 30 (SMP30) is an aging-related protein that participates in the regulation of tissue damage under various pathological conditions. However, the role of SMP30 in mediating high glucose (HG)-induced injury of retinal ganglion cells (RGCs) has not been fully determined. We found that SMP30 expression declined during HG stimulation in RGCs. Cellular functional studies showed that the up-regulation of SMP30 dramatically prohibited HG-evoked apoptosis, oxidative stress and inflammatory response in RGCs. Mechanism research reported that SMP30 overexpression led to the enhancement of nuclear factor erythroid 2-related factor (Nrf2) activation in HG-stimulated RGCs. Moreover, SMP30 overexpression enhanced the phosphorylation of Akt and glucogen synthase kinase-3ß (GSK-3ß), and the suppression of Akt markedly abolished SMP30-mediated Nrf2 activation in HG-stimulated RGCs. Additionally, the suppression of Nrf2 substantially reversed SMP30-overexpression-induced anti-HG injury effects in RGCs. Overall, these findings suggest that SMP30 protects against HG injury of RGCs by potentiating Nrf2 through regulation of the Akt/GSK-3ß pathway. Our work underscores that SMP30/Akt/GSK-3ß/Nrf2 may exert a vital role in mediating the injury of RGCs during diabetic retinopathy.

Bixin protects mice against bronchial asthma though modulating PI3K/Akt pathway.

Accumulating evidence has implicated the potential of natural compounds in treatment of asthma. Bixin is a natural food coloring isolated from the seeds of Bixa Orellana, which possesses anti-tumor, anti-inflammatory and antioxidative properties. Nevertheless, its therapeutic effect in asthma has not been elucidated. Our present study demonstrated that administration of Bixin suppressed allergic airway inflammation and reversed glucocorticoids resistance, as well as alleviated airway remodeling and airway hyperresponsiveness (AHR) in asthmatic mice. In vitro studies showed that Bixin treatment could inhibit the development of epithelial-mesenchymal transition (EMT) mediated by transforming growth factor beta (TGF-ß) signaling. Importantly, Bixin antagonized activation of phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) pathway both in vitro and in vivo. Above all, our findings reveal that Bixin functions as a potent antagonist of PI3K/Akt signaling to protect against allergic asthma, highlighting a novel strategy for asthma treatment based on natural products.

Wild-Type KRAS Allele Effects on Druggable Targets in KRAS Mutant Lung Adenocarcinomas.

KRAS mutations are one of the most common oncogenic drivers in non-small cell lung cancer (NSCLC) and in lung adenocarcinomas in particular. Development of therapeutics targeting KRAS has been incredibly challenging, prompting indirect inhibition of downstream targets such as MEK and ERK. Such inhibitors, unfortunately, come with limited clinical efficacy, and therefore the demand for developing novel therapeutic strategies remains an urgent need for these patients. Exploring the influence of wild-type (WT) KRAS on druggable targets can uncover new vulnerabilities for the treatment of KRAS mutant lung adenocarcinomas. Using commercially available KRAS mutant lung adenocarcinoma cell lines, we explored the influence of WT KRAS on signaling networks and druggable targets. Expression and/or activation of 183 signaling proteins, most of which are targets of FDA-approved drugs, were captured by reverse-phase protein microarray (RPPA). Selected findings were validated on a cohort of 23 surgical biospecimens using the RPPA. Kinase-driven signatures associated with the presence of the KRAS WT allele were detected along the MAPK and AKT/mTOR signaling pathway and alterations of cell cycle regulators. FoxM1 emerged as a potential vulnerability of tumors retaining the KRAS WT allele both in cell lines and in the clinical samples. Our findings suggest that loss of WT KRAS impacts on signaling events and druggable targets in KRAS mutant lung adenocarcinomas.

Host defense against Neospora caninum infection via IL-12p40 production through TLR2/TLR3-AKT-ERK signaling pathway in C57BL/6 mice.

Neospora caninum is an intracellular parasite which can cause neosporosis and significant economic losses in both dairy and beef industries worldwide. A better understanding of the immune response by host cells against N. caninum could help to design better strategies for the prevention and treatment of neosporosis. Although previous studies have shown TLR2/TLR3 were involved in controlling N. caninum infection in mice, the precise mechanisms of the AKT and MAPK pathways controlled by TLR2/TLR3 to regulate N. caninum-induced IL-12p40 production and the role of TLR2/TLR3 in anti-N. caninum infection in bovine macrophages remain unclear. In the present study, TLR2-/- mice displayed more parasite burden and lower level of IL-12p40 production compared to TLR3-/- mice. N. caninum could activate AKT and ERK signaling pathways in WT mouse macrophages, which were inhibited in TLR2-/- and TLR3-/- mouse macrophages. In N. caninum-infected WT mouse macrophages, AKT inhibitor or AKT siRNA could decrease the phosphorylation of ERK. AKT or ERK inhibitors reduced the production of IL-12p40 and increased the number of parasites. The productions of ROS, NO, and GBP2 were significantly reduced in TLR2-/- and TLR3-/- mouse macrophages. Supplementation of rIL-12p40 inhibited N. caninum proliferation and rescued the productions of IFN-γ, NO, and GBP2 in WT, TLR2-/-, and TLR3-/- mouse macrophages. In bovine macrophages, the expressions of TLR2, TLR3, and IL-12p40 mRNA were significantly enhanced by N. caninum, and N. caninum proliferation was inhibited by TLR2/TLR3 agonists. Taken together, the proliferation of N. caninum in mouse macrophages was controlled by the TLR2/TLR3-AKT-ERK signal pathway via increased IL-12p40 production, which in turn lead to the productions of NO, GBP2, and IFN-γ during N. caninum infection. And in bovine macrophages, TLR2 and TLR3 contributed to inhibiting N. caninum proliferation via increased IL-12p40 production.

Lactobacillus rhamnosus GG Promotes Early B Lineage Development and IgA Production in the Lamina Propria in Piglets.

Gut microbes play an important role in the development of host B cells. It has been controversial whether GALT is the development site of B cells in pigs. By investigating the relationship between gut microbes and the development of B cells in the GALT of piglets, we found, to our knowledge for the first time, that early B cells exist in the gut lamina propria (LP) in pigs at different ages. We further used Lactobacillus rhamnosus GG (LGG) to treat piglets. The results showed that LGG promotes the development of the early B lineage, affects the composition of the Ig CDR3 repertoires of B cells, and promotes the production of IgA in the intestinal LP. Additionally, we found that the p40 protein derived from LGG can activate the EGFR/AKT and NF-κB signaling pathways, inducing porcine intestinal epithelial cells (IPEC-J2) to secrete a proliferation-inducing ligand (APRIL), which promotes IgA production in B cells. Finally, we identified ARF4 and DIF3 as candidates for p40 receptors on IPEC-J2 by GST pull-down, liquid chromatography-mass spectrometry/mass spectrometry analysis, and coimmunoprecipitation. In conclusion, LGG could promote early B cell differentiation and development in the intestinal LP in piglets and might contribute to promoting IgA production via secretion of p40, which interacts with the membrane receptors on IPEC-J2 and induces them to secrete APRIL. Our study will provide insight to aid in better utilization of probiotics to increase human health.

Medical ozone induces proliferation and migration inhibition through ROS accumulation and PI3K/AKT/NF-kB suppression in human liver cancer cells in vitro

Background Hepatocellular carcinoma is one of the most common malignancies and leading cancer-associated deaths worldwide. Ozone has been proposed as a promising therapeutic agent in the treatment of various disorders. Purpose The purpose of this paper is to assess the potential anticancer effects of the ozone on liver cancer cells. Method The liver cancer cell line of bel7402 and SMMC7721 was used in this study. Proliferation was evaluated using the CCK-8 and the colony formation assay. Wond healing assay and transwell assay without Matrigel were used to evaluate their migration ability. Flow cytometry was used for cell cycle analysis and reactive oxygen species (ROS) determination. Glutathione detection kit was used for measurement of glutathione level. Protein expression was estimated by western blot analysis. Results Ozone treatment inhibited liver cancer cell proliferation, colony formation. Ozone induced G2/M phase cell cycle arrest, which could be elucidated by the change of protein levels of p53, p21, Cyclin D1, cyclin B1, cdc2, and CDK4. We also found that ozone treatment inhibited migration ability by inhibiting EMT-relating protein. Ozone also induced ROS accumulation and decreased glutathione level decreased, which contributed to the inactivation of the PI3K/AKT/NF-κB pathway. Finally, we found that pre-treatment of liver cancer cells with N-acetylcysteine resisted ozone-induced effects. Conclusions Ozone restrains the proliferation and migration potential and EMT process of liver cancer cells via ROS accumulation and PI3K/AKT/NF-κB suppression (AU)

Bcl2-associated athanogene 4 promotes the invasion and metastasis of gastric cancer cells by activating the PI3K/AKT/NF-κB/ZEB1 axis.

Bcl2-associated athanogene 4 (BAG4) has been found to be aberrantly expressed in several types of human cancers. However, little is known about its expression, role, and clinical significance in gastric cancer (GC). In this study, we aimed to address these issues and to explore the underlying mechanisms. The expression level of BAG4, measured by immunohistochemistry, was significantly higher in GC tissues than in paired normal tissues. Elevated BAG4 expression was positively correlated with T stage, lymph node metastasis, and tumor size of GC and was associated with unfavorable outcomes of the patients. The overexpression of BAG4 promoted the in vitro invasion and in vivo metastasis of GC cells, and opposite results were observed after silencing of BAG4. Silencing of BAG4 significantly reduced the phosphorylation of PI3K, AKT, and p65, whereas overexpression of BAG4 markedly enhanced the phosphorylation of these molecules. At the same time, manipulating BAG4 expression resulted in the corresponding changes in p65 nuclear translocation and ZEB1 expression. Luciferase reporter and chromatin immunoprecipitation assays verified that p65 binds to the promoter of ZEB1 to upregulate its transcription. Our results demonstrate that BAG4 plays an oncogenic role in the invasion and metastasis of GC cells by activating the PI3K/AKT/NF-κB/ZEB1 axis to induce epithelial-mesenchymal transition.

Expression of activator proteins of SHH/GLI and PI3K/Akt/mTORC1 signaling pathways in human gliomas is associated with high grade tumors.

Recent findings have demonstrated a synergic crosstalk between SHH/GLI and PI3K/Akt/mTORC1 signaling in glioblastoma progression cells in vitro and in tumors in mice, but it is not known if this also occurs in human gliomas. We then aimed to investigate the expression of key proteins of these pathways in different human gliomas. The expression of PTEN, phospho-Akt (Ser473), phospho-S6K1 (Thr389), SHH, GLI1, GLI2 and GLI3 was assessed by immunohistochemistry in gliomas and in control brain tissues. The pattern of expression of each protein was established according to glioma type, glioma grade and to cell type; the relative expression of each protein was used to perform statistical analyses. We found that the expression of proteins of both signaling pathways differs between normal brain and glioma tissues. For instance, normal astrocytes had a different protein expression pattern compared with reactive and tumoral astrocytes. Interestingly, we detected a recurrent pattern of expression of GLI3 in oligodendrocytes and of phospho-S6K1 in mitotic neoplastic cells. We also identified differences of cell signaling according to glioma type: oligodendrogliomas and ependymomas are related with the expression of SHH/GLI proteins. Finally, we detected that high grade gliomas statistically correlate with the expression of GLI1 and GLI2, and that GLI1, GLI2, phospho-Akt and phospho-S6K1 are more expressed in patients with less survival, suggesting that activation of these cell signaling influences glioma outcome and patient survival. In summary, our results show that proteins of PI3K/Akt/mTORC1 and SHH/GLI pathways are differentially expressed in human gliomas according to tumor type and grade, and suggest that the activation of these signaling networks is associated with glioma progression.

Platelet-derived growth factor (PDGF)-BB protects dopaminergic neurons via activation of Akt/ERK/CREB pathways to upregulate tyrosine hydroxylase.

AIMS: The neurotropic growth factor PDGF-BB was shown to have vital neurorestorative functions in various animal models of Parkinson's disease (PD). Previous studies indicated that the regenerative property of PDGF-BB contributes to the increased intensity of tyrosine hydroxylase (TH) fibers in vivo. However, whether PDGF-BB directly modulates the expression of TH, and the underlying mechanism is still unknown. We will carefully examine this in our current study. METHOD: MPTP-lesion mice received PDGF-BB treatment via intracerebroventricular (i.c.v) administration, and the expression of TH in different brain regions was assessed by RT-PCR, Western blot, and immunohistochemistry staining. The molecular mechanisms of PDGF-BB-mediated TH upregulation were examined by RT-PCR, Western blot, ChIP assay, luciferase reporter assay, and immunocytochemistry. RESULTS: We validated a reversal expression of TH in MPTP-lesion mice upon i.c.v administration of PDGF-BB for seven days. Similar effects of PDGF-BB-mediated TH upregulation were also observed in MPP+ -treated primary neuronal culture and dopaminergic neuronal cell line SH-SY5Y cells. We next demonstrated that PDGF-BB rapidly activated the pro-survival PI3K/Akt and MAPK/ERK signaling pathways, as well as the downstream CREB in SH-SY5Y cells. We further confirmed the significant induction of p-CREB in PDGF-BB-treated animals in vivo. Using a genetic approach, we demonstrated that the transcription factor CREB is critical for PDGF-BB-mediated TH expression. The activation and nucleus translocation of CREB were promoted in PDGF-BB-treated SH-SY5Y cells, and the enrichment of CREB on the promoter region of TH gene was also increased upon PDGF-BB treatment. CONCLUSION: Our data demonstrated that PDGF-BB directly regulated the expression of TH via activating the downstream Akt/ERK/CREB signaling pathways. Our finding will further support the therapeutic potential of PDGF-BB in PD, and provide the possibility that targeting PDGF signaling can be harnessed as an adjunctive therapy in PD in the future.