scholarly journals Bee Venom Prevents Mucin 5AC Production through Inhibition of AKT and SPDEF Activation in Airway Epithelia Cells

Toxins ◽  
2021 ◽  
Vol 13 (11) ◽  
pp. 773
Author(s):  
Sanga Kim ◽  
Hee-Won Kim ◽  
Seok-Hwan Chang ◽  
Kang-Hyun Leem ◽  
Hae-Jeong Park
Keyword(s):  
Protein Expression ◽  
A549 Cells ◽  
Allergic Diseases ◽  
Bee Venom ◽  
Airway Epithelia ◽  
Akt Phosphorylation ◽  
Akt Activation ◽  
Phosphoinositide 3 Kinase ◽  
Induced Changes ◽  
Mucus Metaplasia

IL-13 induces mucus metaplasia, which causes airway obstruction in asthma. Bee venom (BV) and its components have shown anti-inflammatory effects in allergic diseases such as atopic dermatitis and asthma. In this study, we investigated the effect of BV on IL-13-induced mucus metaplasia through activation of the signal transducer and activator of transcription (STAT6), and regulation of SAM-pointed domain containing Ets-like factor (SPDEF) and forkhead box A2 (FOXA2) in the airway epithelia cell line A549. In A549 cells, BV (1.0 µg/mL) inhibited IL-13 (10 ng/mL)-induced AKT phosphorylation, increase in SPDEF protein expression, and decrease in FOXA2 protein expression—but not STAT6 phosphorylation. BV also prevented the IL-13-induced increase in mucin 5AC (MUC5AC) mRNA and protein expression. Moreover, we observed that inhibition of phosphoinositide 3 kinase (PI3K)/AKT using LY294002 (50 µM) could reverse the alterations in FOXA2 and MUC5AC expression -by IL-13 and BV. However, LY294002 did not affect IL-13- and BV-induced changes in SPDEF expression. These findings indicate that BV inhibits MUC5AC production through the regulation of SPDEF and FOXA2. The inhibition of MUC5AC production through FOXA2 is mediated via the suppression of PI3K/AKT activation by BV. BV may be helpful in the prevention of mucus metaplasia in asthma.

scholarly journals CTLA-4 and PD-1 Receptors Inhibit T-Cell Activation by Distinct Mechanisms

2005 ◽  
Vol 25 (21) ◽  
pp. 9543-9553 ◽  
Author(s):  
Richard V. Parry ◽  
Jens M. Chemnitz ◽  
Kenneth A. Frauwirth ◽  
Anthony R. Lanfranco ◽  
Inbal Braunstein ◽  
...  
Keyword(s):  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Direct Evidence ◽  
Phosphatidylinositol 3 Kinase ◽  
Akt Phosphorylation ◽  
Akt Activation ◽  
Pi3k Activation ◽  
Cellular Phenotypes ◽  
Induced Changes

ABSTRACT CTLA-4 and PD-1 are receptors that negatively regulate T-cell activation. Ligation of both CTLA-4 and PD-1 blocked CD3/CD28-mediated upregulation of glucose metabolism and Akt activity, but each accomplished this regulation using separate mechanisms. CTLA-4-mediated inhibition of Akt phosphorylation is sensitive to okadaic acid, providing direct evidence that PP2A plays a prominent role in mediating CTLA-4 suppression of T-cell activation. In contrast, PD-1 signaling inhibits Akt phosphorylation by preventing CD28-mediated activation of phosphatidylinositol 3-kinase (PI3K). The ability of PD-1 to suppress PI3K/AKT activation was dependent upon the immunoreceptor tyrosine-based switch motif located in its cytoplasmic tail, adding further importance to this domain in mediating PD-1 signal transduction. Lastly, PD-1 ligation is more effective in suppressing CD3/CD28-induced changes in the T-cell transcriptional profile, suggesting that differential regulation of PI3K activation by PD-1 and CTLA-4 ligation results in distinct cellular phenotypes. Together, these data suggest that CTLA-4 and PD-1 inhibit T-cell activation through distinct and potentially synergistic mechanisms.

scholarly journals Peroxynitrite activates the phosphoinositide 3-kinase/Akt pathway in human skin primary fibroblasts

2000 ◽  
Vol 352 (1) ◽  
pp. 219-225 ◽  
Author(s):  
Lars-Oliver KLOTZ ◽  
Stefan M. SCHIEKE ◽  
Helmut SIES ◽  
Nikki J. HOLBROOK
Keyword(s):  
Tyrosine Kinases ◽  
Cellular Response ◽  
Glycogen Synthase ◽  
Nitrogen Monoxide ◽  
Akt Phosphorylation ◽  
Akt Activation ◽  
Acute Response ◽  
Diffusion Limited ◽  
Primary Fibroblasts ◽  
Phosphoinositide 3 Kinase

Peroxynitrite is a potent oxidizing and nitrating species formed in a diffusion-limited reaction between nitrogen monoxide and superoxide. It induces apoptosis through unknown mechanisms and is believed to interfere with receptor tyrosine kinase signalling through nitration of tyrosine residues. One pathway emanating from receptor tyrosine kinases is that leading to activation of the anti-apoptotic kinase Akt. In the present study we provide evidence that peroxynitrite, administered to cells using two different delivery systems, results in the dose- and time-dependent activation of Akt. Akt activation is rapid and followed by phosphorylation of glycogen synthase kinase-3, an established substrate of Akt. Akt activation is inhibited in the presence of the phosphoinositide 3-kinase (PI-3K) inhibitors wortmannin and LY294002, and by treatment with the platelet-derived growth factor (PDGF) receptor (PDGFR) inhibitor AG1295, indicating a requirement for PDGFR and PI-3K in mediating peroxynitrite-induced Akt activation. Accordingly, the PDGFR-A and PDGFR-B isoforms were shown to undergo rapid tyrosine phosphorylation on treatment with peroxynitrite. Prior exposure of cells to peroxynitrite interferes with PDGF-induced Akt phosphorylation. Our findings suggest that Akt activation occurs as an acute response to peroxynitrite treatment and could play an important role in influencing cell survival and/or alter the cellular response to other growth regulatory signals.

scholarly journals Phosphatidylinositol-3,4,5-Trisphosphate Is Required for Insulin-Like Growth Factor 1-Mediated Survival of 3T3-L1 Preadipocytes**This work was supported by a grant (to A.S.) from the Medical Research Council of Canada.

Endocrinology ◽  
2001 ◽  
Vol 142 (1) ◽  
pp. 205-212 ◽  
Author(s):  
AnneMarie Gagnon ◽  
Patti Dods ◽  
Nicolas Roustan-Delatour ◽  
Ching-Shih Chen ◽  
Alexander Sorisky
Keyword(s):  
Cell Death ◽  
Growth Factor ◽  
Mammalian Target Of Rapamycin ◽  
Inhibitory Effect ◽  
Tunel Staining ◽  
Insulin Like Growth Factor ◽  
Tissue Mass ◽  
Akt Phosphorylation ◽  
Dependent Cell ◽  
Phosphoinositide 3 Kinase

Abstract Adipocyte number, a determinant of adipose tissue mass, reflects the balance between the rates of proliferation/differentiation vs. apoptosis of preadipocytes. The percentage of 3T3-L1 preadipocytes undergoing cell death following serum deprivation was reduced by 10 nm insulin-like growth factor (IGF)-1 (from 50.0 ± 0.7% for control starved cells to 27.5 ± 3.1%). TUNEL staining confirmed the apoptotic nature of the cell death. The protective effect of IGF-1 was blocked by phosphoinositide 3-kinase (PI3K) inhibitors, wortmannin, and LY294002, but was unaffected by rapamycin, PD98059, or SB203580, which inhibit mammalian target of rapamycin (mTOR), ERK kinase (MEK1), and p38 MAPK respectively. Exogenous PI(3,4,5)P3 (10 μm), the principal product of IGF-1-stimulated PI3K in 3T3-L1 preadipocytes, had a modest survival effect on its own, reducing cell death from 47.9± 3.4% to 35.6 ± 3.5%. When added to the combination of IGF-1 and LY294002, PI(3,4,5)P3 reversed most of the inhibitory effect of LY294002 on IGF-1-dependent cell survival, protein kinase B/Akt phosphorylation, and caspase-3 activity. Taken together, these results implicate PI(3,4,5)P3 as a necessary signal for the anti-apoptotic action of IGF-1 on 3T3-L1 preadipocytes.

Global Analysis of Protein Expression in A549 Cells After Prolonged Nicotine Exposure by Using Label-free Quantification

2021 ◽  
Vol 41 (8) ◽  
pp. 3833-3842
Author(s):  
SASIKARN KOMKLEOW ◽  
CHURAT WEERAPHAN ◽  
DARANEE CHOKCHAICHAMNANKIT ◽  
PAPADA CHAISURIYA ◽  
CHRIS VERATHAMJAMRAS ◽  
...  
Keyword(s):  
Protein Expression ◽  
Global Analysis ◽  
A549 Cells ◽  
Nicotine Exposure ◽  
Label Free ◽  
Label Free Quantification ◽  
Free Quantification

Daidzein affects steroidogenesis and oestrogen receptor expression in medium ovarian follicles of pigs

2013 ◽  
Vol 61 (1) ◽  
pp. 85-98 ◽  
Author(s):  
Anna Nynca ◽  
Dominika Słonina ◽  
Olga Jablońska ◽  
Barbara Kamińska ◽  
Renata Ciereszko
Keyword(s):  
Protein Expression ◽  
Granulosa Cells ◽  
Oestrogen Receptor ◽  
Receptor Expression ◽  
Progesterone Production ◽  
Progesterone Secretion ◽  
Mrna And Protein Expression ◽  
Induced Changes ◽  
Reproductive Processes

Daidzein, a phytoestrogen present in soybean products used in swine feed, has been demonstrated to affect both reproductive and endocrine functions. The aims of this study were to examine the in vitro effects of daidzein on (1) progesterone (P4) and oestradiol (E2) secretion by porcine luteinised granulosa cells harvested from medium follicles, and (2) the mRNA and protein expression of oestrogen receptors α and β (ERα and ERβ) in these cells. The influence of E2 on P4 secretion and ERα and ERβ expression in the granulosa cells of pigs was also investigated. It was found that daidzein inhibited progesterone secretion by luteinised granulosa cells isolated from medium follicles. In contrast, E2 did not affect progesterone production by these cells. Moreover, daidzein did not alter the granulosal secretion of E2. Both daidzein and E2 decreased mRNA expression of ERα in the cells examined. The expression of ERβ mRNA was not affected by daidzein but was inhibited by E2. ERα protein was not detected while ERβ protein was found in the nuclei of the cells. Daidzein and E2 upregulated the expression of ERβ protein in the cells. In summary, the phytoestrogen daidzein directly affected the porcine ovary by inhibiting progesterone production and increasing ERβ protein expression. Daidzein-induced changes in follicular steroidogenesis and granulosal sensitivity to oestrogens may disturb reproductive processes in pigs.

scholarly journals α-Hederin Inhibits the Growth of Lung Cancer A549 Cells in vitro and in vivo by Increasing SIRT6 Dependent Glycolysis

2020 ◽  
Author(s):  
cong fang ◽  
Yahui Liu ◽  
Lanying Chen ◽  
Yingying Luo ◽  
Yaru Cui ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Mouse Model ◽  
Protein Expression ◽  
Glucose Uptake ◽  
A549 Cells ◽  
Lactate Production ◽  
Tumor Xenograft ◽  
Xenograft Mouse Model ◽  
Atp Generation

Abstract Background: α-hederin an effective component of Pulsatilla chinensis (Bunge) Regel, Studies showed that α-hederin exert many pharmacological activities, However, the effect of α-hederin on metabolism is still unclear. This study aimed to illuminate the role of α-hederin in glucose metabolism in lung cancer cells and investigate the molecular mechanism of α-hederin. Methods: CCK8 and colony formation assays were employed to assess the anti-proliferative effects induced by α-hederin. Glucose uptake, ATP generation, and reduced lactate production were measured using kits, and an A549 tumor xenograft mouse model of lung cancer was used to assess the in vivo antitumor effect of α-hederin (5, 10 mg/kg). Glycolytic-related key enzymes hexokinase 2 (HK2), glucose transporters 1 (GLUT1), pyruvate kinase M2 (PKM2), lactate dehydrogenase A (LDHA), monocarboxylate transporter (MCT4), c-Myc, Hypoxia inducible factor-1α (HIF-1α) and Sirtuin 6 (SIRT6) protein expression were detected by western blotting and immunohistochemical staining and SIRT6 inhibitors was verified in A549 cells. Results: Our results showed that cell proliferation was significantly inhibited by α-hederin in a dose-dependent manner and that α-hederin inhibited glucose uptake and ATP generation and reduced lactate production. Furthermore, α-hederin remarkably inhibited HK2, GLUT1, PKM2, LDHA, MCT4, c-Myc, HIF-1α and activated SIRT6 protein expression. Using inhibitors, we proved that α-hederin inhibits glycolysis by activating SIRT6. Moreover, a tumor xenograft mouse model of lung cancer further confirmed that α-hederin inhibits lung cancer growth via inhibiting glycolysis in vivo. Conclusions: α-hederin inhibits the growth of non-small cell lung cancer A549 cells by inhibiting glycolysis. The mechanism of glycolysis inhibition includes α-hederin activating the expression of the glycolytic related protein SIRT6.

scholarly journals Differentiation and Maturation Effect of All-trans Retinoic Acid on Cultured Fetal RPE and Stem Cell-Derived RPE Cells for Cell-Based Therapy

2021 ◽  
Author(s):  
Tingyu Yan ◽  
Na Yang ◽  
Wei Hu ◽  
Xinxin Zhang ◽  
Xuedong Li ◽  
...  
Keyword(s):  
Stem Cell ◽  
Retinoic Acid ◽  
Protein Expression ◽  
Rpe Cells ◽  
Plating Density ◽  
Cell Based Therapy ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid ◽  
Gene And Protein Expression ◽  
Induced Changes

Abstract Background: Phase I/II clinical trials using fetal retinal pigment epithelium (fRPE), human embryonic stem cell (hESC)-derived RPE, or human induced pluripotent stem cell (hiPSC)-derived RPE as potential sources of materials for cell-based therapy to treat degenerative retinal diseases have been carried out during the past decade. Challenges for successful translational cell-based therapy include cell manufacture, cell quality, cell storage, and cell behavior in vivo. In this study, we investigated the culture-induced changes in passaged fetal RPE, hESC-RPE and hiPSC-RPE cells in vitro and explored the differentiation and maturation effect of all-trans retinoic acid (ATRA) on those RPE cells. Methods: A total of 9 fetal RPE cell lines, hESC-RPE and hiPSC-RPE cell lines were set up using previously described methods. The culture-induced changes in subsequent passages caused by manipulating plating density, dissociation method and repeated passaging were studied by microscope, real-time quantitative PCR, western blot and immunofluorescent assays. Gene and protein expression and functional characteristics of fRPE, hESC-RPE and hiPSC-RPE incubated with ATRA at different concentration were also evaluated.Results: Compared with fRPE, hESC-RPE and hiPSC-RPE showed decreased gene and protein expression of RPE markers. Passage 3 RPE of all three types seeded at a density of 6×105 and 9x105 cells/mL in basal medium maintained pigmented polygonal, cobblestone-like morphology. RPE cells underwent mesenchymal changes showing increased expression of mesenchymal markers including a-SMA, N-cadherin, fibronectin and decreased expression of RPE markers including RPE65, E-cadherin and ZO-1, as a subsequence of low plating density, inappropriate dissociated method, and repeated passaging. fRPE, hESC-RPE and iPSC-RPE treated by ATRA at different concentrations showed increased expression of RPE markers such as RPE65, bestrophin (BEST) and CRALBP, and increased expression of negative complement regulatory proteins (CRP) including complement factor H (CFH), CD46, CD55 and CD59, and increased transepithelial resistance (TER) as well.Conclusion: Although hESC and hiPSC-derived RPE are morphologically similar to fRPE, and also have the tendency to undergo epithelial-to-mesenchymal transition (EMT) changes during the culturing and passaging process in vitro, differences in protein and gene expression among three RPE types exist. Moreover, ATRA can increase RPE markers expression, as well as to increase the expression levels of CRPs gene and protein in fRPE and stem cell-derived RPE.

scholarly journals A p53-Phosphoinositide Signalosome Regulates Nuclear Akt Activation

2021 ◽  
Author(s):  
Mo Chen ◽  
Suyong Choi ◽  
Tianmu Wen ◽  
Changliang Chen ◽  
Narendra Thapa ◽  
...  
Keyword(s):  
Genotoxic Stress ◽  
Akt Pathway ◽  
Mutant P53 ◽  
Tumor Suppressor P53 ◽  
Wild Type ◽  
Akt Activation ◽  
Pi3k Inhibitors ◽  
Phosphoinositide 3 Kinase ◽  
Induced Apoptosis ◽  
Dependent Mechanism

The tumor suppressor p53 and the phosphoinositide 3-kinase (PI3K)-Akt pathway have fundamental roles in regulating cell growth, apoptosis and are frequently mutated in cancer. Here, we show that genotoxic stress induces nuclear Akt activation by a p53-dependent mechanism that is independent from the canonical membrane-localized PI3K-Akt pathway. Upon genotoxic stress a nuclear p53-PI3,4,5P3 complex is generated in regions devoid of membranes by a nuclear PI3K, and this complex recruits all the kinases required to activate Akt and phosphorylate FOXOs, inhibiting DNA damage-induced apoptosis. Wild-type p53 activates nuclear Akt in an on/off fashion upon stress, whereas mutant p53 stimulates high basal Akt activity, indicating a fundamental difference. The nuclear p53-phosphoinositide signalosome is distinct from the canonical membrane-localized pathway and insensitive to PI3K inhibitors currently in the clinic, underscoring its therapeutic relevance.

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