scholarly journals Mitogen-Activated Protein Kinase-Dependent Stimulation of Proliferation of Rat Lactotrophs in Culture by 3′,5′-Cyclic Adenosine Monophosphate*

Endocrinology ◽  
1999 ◽  
Vol 140 (6) ◽  
pp. 2850-2858 ◽  
Author(s):  
Shinichi Suzuki ◽  
Isao Yamamoto ◽  
Jun Arita
Keyword(s):  
Protein Kinase ◽  
Kinase Inhibitor ◽  
Mitogen Activated Protein Kinase ◽  
Dopamine Receptor Agonist ◽  
Intracellular Camp ◽  
Pituitary Cells ◽  
Dependent Manner ◽  
Stimulatory Action ◽  
Mitogen Activated Protein ◽  
Brdu Labeling

Abstract Intracellular cAMP regulates cell proliferation as a second messenger of extracellular signals in a number of cell types. We investigated, by pharmacological means, whether an increase in intracellular cAMP levels changes proliferation rates of lactotrophs in primary culture, whether there are interactions between signal transduction pathways of cAMP and the growth factor insulin, and where the dopamine receptor agonist bromocriptine acts in the cAMP pathway to inhibit lactotroph proliferation. Rat anterior pituitary cells, cultured in serum-free medium, were treated with cAMP-increasing agents, followed by 5-bromo-2′-deoxyuridine (BrdU) to label proliferating pituitary cells. BrdU-labeling indices indicative of the proliferation rate of lactotrophs were determined by double immunofluorescence staining for PRL and BrdU. Treatment with forskolin (an adenylate cyclase activator) or (Bu)2cAMP (a membrane-permeable cAMP analog) increased BrdU-labeling indices of lactotrophs in a dose- and incubation time-dependent manner. The cAMP-increasing agents were also effective in increasing BrdU-labeling indices in populations enriched for lactotrophs by differential sedimentation. The stimulatory action of forskolin was observed, regardless of concentrations of insulin that were added in combination with forskolin. Inhibition of the action of endogenous cAMP by H89 or KT5720, a protein kinase A inhibitor, attenuated an increase in BrdU-labeling indices by insulin treatment. On the other hand, the specific mitogen-activated protein kinase inhibitor PD98059, which was effective in blocking the mitogenic action of insulin, markedly suppressed the forskolin-induced increase in BrdU-labeling indices. (Bu)2cAMP antagonized not only inhibition of BrdU labeling indices but also changes in cell shape induced by bromocriptine treatment, although forskolin did not have such an antagonizing effect. These results suggest that: 1) intracellular cAMP plays a stimulatory role in the regulation of lactotroph proliferation; 2) cAMP and insulin/mitogen-activated protein kinase signalings require each other for their mitogenic actions; and 3) the antimitogenic action of bromocriptine is, at least in part, caused by inhibition of cAMP production.

CCK activates p90rsk in rat pancreatic acini through protein kinase C

1997 ◽  
Vol 272 (3) ◽  
pp. G401-G407 ◽  
Author(s):  
M. J. Bragado ◽  
A. Dabrowski ◽  
G. E. Groblewski ◽  
J. A. Williams
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Kinase Inhibitor ◽  
Threshold Concentration ◽  
Mitogen Activated Protein Kinase ◽  
Maximal Effect ◽  
Dependent Manner ◽  
Pancreatic Acini ◽  
Kinase C ◽  
Mitogen Activated Protein

The presence of the 90-kDa ribosomal S6 protein kinase (p90(rsk)) in isolated rat pancreatic acini was demonstrated by Western blotting and immunoprecipitation with anti-p90(rsk). Cholecystokinin (CCK) activated p90(rsk) activity in a time- and dose-dependent manner and increased its phosphorylation. The threshold concentration of CCK was 10 pM and the maximal effect was seen at 1 nM. An increase in p90(rsk) was observed 1 min after 1 nM CCK stimulation, reaching a maximum at 10 min, when p90(rsk) activity was increased 5.4-fold. Carbachol and bombesin, but not vasoactive intestinal peptide, also activated p90(rsk). CCK-induced activation of p90(rsk) appears to be mediated by protein kinase C (PKC), since 12-O-tetradecanoylphorbol-13-acetate increased p90(rsk) activity 5.3-fold. GF-109293X, a potent inhibitor of PKC, strongly inhibited CCK-evoked p90(rsk) activity. Treatment of acini with ionomycin or 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid had no effect, indicating that mobilization of intracellular Ca2+ by CCK is not important in p90(rsk) activation. Although there were some quantitative differences in the extent of inhibition, the specific inhibitors [rapamycin, wortmannin, mitogen-activated protein kinase (MAPK) kinase inhibitor PD98059, and GF-109293X] had parallel effects on p90(rsk) and p42(mapk) activities, consistent with a model in which p90(rsk) can be regulated in acini by MAPK.

scholarly journals Mechanically activated Piezo1 channels of cardiac fibroblasts stimulate p38 mitogen-activated protein kinase activity and interleukin-6 secretion

2019 ◽  
Vol 294 (46) ◽  
pp. 17395-17408 ◽  
Author(s):  
Nicola M. Blythe ◽  
Katsuhiko Muraki ◽  
Melanie J. Ludlow ◽  
Vasili Stylianidis ◽  
Hamish T. J. Gilbert ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Kinase Activity ◽  
Kinase Inhibitor ◽  
Cardiac Fibroblasts ◽  
Ruthenium Red ◽  
Mitogen Activated Protein Kinase ◽  
Protein Kinase Activity ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Mitogen Activated Protein

Piezo1 is a mechanosensitive cation channel with widespread physiological importance; however, its role in the heart is poorly understood. Cardiac fibroblasts help preserve myocardial integrity and play a key role in regulating its repair and remodeling following stress or injury. Here we investigated Piezo1 expression and function in cultured human and mouse cardiac fibroblasts. RT-PCR experiments confirmed that Piezo1 mRNA in cardiac fibroblasts is expressed at levels similar to those in endothelial cells. The results of a Fura-2 intracellular Ca2+ assay validated Piezo1 as a functional ion channel that is activated by its agonist, Yoda1. Yoda1-induced Ca2+ entry was inhibited by Piezo1 blockers (gadolinium and ruthenium red) and was reduced proportionally by siRNA-mediated Piezo1 knockdown or in murine Piezo1+/− cells. Results from cell-attached patch clamp recordings on human cardiac fibroblasts established that they contain mechanically activated ion channels and that their pressure responses are reduced by Piezo1 knockdown. Investigation of Yoda1 effects on selected remodeling genes indicated that Piezo1 activation increases both mRNA levels and protein secretion of IL-6, a pro-hypertrophic and profibrotic cytokine, in a Piezo1-dependent manner. Moreover, Piezo1 knockdown reduced basal IL-6 expression from cells cultured on softer collagen-coated substrates. Multiplex kinase activity profiling combined with kinase inhibitor experiments and phosphospecific immunoblotting established that Piezo1 activation stimulates IL-6 secretion via the p38 mitogen-activated protein kinase downstream of Ca2+ entry. In summary, cardiac fibroblasts express mechanically activated Piezo1 channels coupled to secretion of the paracrine signaling molecule IL-6. Piezo1 may therefore be important in regulating cardiac remodeling.

scholarly journals Phosphorylation and activation of p42 and p44 mitogen-activated protein kinase are required for the P2 purinoceptor stimulation of endothelial prostacyclin production

1996 ◽  
Vol 320 (1) ◽  
pp. 221-226 ◽  
Author(s):  
Viral PATEL ◽  
Colin BROWN ◽  
Adele GOODWIN ◽  
Neil WILKIE ◽  
Michael R BOARDER
Keyword(s):  
Protein Kinase ◽  
Tyrosine Phosphorylation ◽  
Kinase Activity ◽  
Kinase Inhibitor ◽  
Mitogen Activated Protein Kinase ◽  
Dependent Manner ◽  
Western Blots ◽  
Mitogen Activated Protein ◽  
Two Peaks ◽  
Stimulation Of

Extracellular ATP and ADP, released from platelets and other sites stimulate the endothelial production of prostacyclin (PGI2) by acting on G-protein-coupled P2Y1 and P2Y2 purinoceptors, contributing to the maintenance of a non-thrombogenic surface. The mechanism, widely described as being dependent on elevated cytosolic [Ca2+], also requires protein tyrosine phosphorylation. Here we show that activation of both these P2 receptor types leads to the tyrosine phosphorylation and activation of both the p42 and p44 forms of mitogen-activated protein kinase (MAPK). 2-Methylthio-ATP and UTP, selectively activating P2Y1 and P2Y2 purinoceptors respectively, and ATP, a non-selective agonist at these two receptors, stimulate the tyrosine phosphorylation of both p42mapk and p44mapk, as revealed by Western blots with an antiserum specific for the tyrosine-phosphorylated forms of the enzymes. By using separation on Resource Q columns, peptide kinase activity associated with the phosphorylated MAPK enzymes distributes into two peaks, one mainly p42mapk and one mainly p44mapk, both of which are stimulated by ATP with respect to kinase activity and phospho-MAPK immunoreactivity. Stimulation of P2Y1 or P2Y2 purinoceptors leads to a severalfold increase in PGI2 efflux; this was blocked in a dose-dependent manner by the selective MAPK kinase inhibitor PD98059. This drug also blocked the agonist-stimulated increase in phospho-MAPK immunoreactivity for both p42mapk and p44mapk but left the phospholipase C response to P2 agonists essentially unchanged. Olomoucine has been reported to inhibit p44mapk activity. Here we show that in the same concentration range olomoucine inhibits activity in both peaks from the Resource Q column and also the agonist stimulation of 6-keto-PGF1, but has no effect on agonist-stimulated phospho-MAPK immunoreactivity. These results provide direct evidence for the involvement of p42 and p44 MAPK in the PGI2 response of intact endothelial cells: we have shown that both the endothelial P2Y purinoceptors are linked to activation of MAPK, and that activation of this pathway is a requirement for the stimulation by ATP/ADP of endothelial PGI2 production.

Mitochondrial ROS initiate phosphorylation of p38 MAP kinase during hypoxia in cardiomyocytes

2002 ◽  
Vol 282 (6) ◽  
pp. L1324-L1329 ◽  
Author(s):  
Andre Kulisz ◽  
Ningfang Chen ◽  
Navdeep S. Chandel ◽  
Zuohui Shao ◽  
Paul T. Schumacker
Keyword(s):  
Kinase Inhibitor ◽  
Anion Channel ◽  
P38 Map Kinase ◽  
Proximal Region ◽  
Mitogen Activated Protein Kinase ◽  
Ros Generation ◽  
Dependent Manner ◽  
Adaptive Responses ◽  
Mitochondrial Ros ◽  
Mitogen Activated Protein

The p38 mitogen-activated protein kinase (MAPK) is phosphorylated in response to oxidative stress. Mitochondria in cardiomyocytes increase their generation of reactive oxygen species (ROS) during hypoxia (1–5% O2). These ROS participate in signal transduction pathways involved in adaptive responses, including ischemic preconditioning and gene transcription. The present study therefore tested the hypothesis that hypoxia induces p38 MAPK phosphorylation by augmenting mitochondrial ROS generation. In cardiomyocytes, phosphorylation of p38 was observed in a Po 2-dependent manner during hypoxia. This response was inhibited by rotenone, thenoyltrifluoroacetone, and myxothiazol, inhibitors of mitochondrial complexes I, II, and III, respectively. A similar inhibition was observed in the cells pretreated with anion channel inhibitor DIDS, which may block ROS release from mitochondria. During normoxia, increases in mitochondrial ROS elicited by azide (1–2 mM) or by the mitochondrial inhibitor antimycin A caused increased phosphorylation of p38. Brief treatment with exogenous H2O2 during normoxia also induced phosphorylation of p38 as hypoxia, but this effect was not abolished by myxothiazol or DIDS. The antioxidant N-acetyl-cysteine abolished the p38 response to hypoxia, presumably by scavenging H2O2, but the mitogen extracellular receptor kinase inhibitor PD-98059 did not inhibit p38 phosphorylation during hypoxia. Thus physiological hypoxia leads to p38 phosphorylation through a mechanism that requires electron flux in the proximal region of the mitochondrial electron transport chain, which suggests that either H2O2 or superoxide participates in activating that process.

scholarly journals Ghrelin exerts a proliferative effect on a rat pituitary somatotroph cell line via the mitogen-activated protein kinase pathway

2004 ◽  
pp. 233-240 ◽  
Author(s):  
AM Nanzer ◽  
S Khalaf ◽  
AM Mozid ◽  
RC Fowkes ◽  
MV Patel ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Protein Kinase ◽  
Cell Line ◽  
Kinase Inhibitor ◽  
Thymidine Incorporation ◽  
Mapk Pathway ◽  
Mitogen Activated Protein Kinase ◽  
Gh3 Cells ◽  
Rat Pituitary ◽  
Mitogen Activated Protein

OBJECTIVES: Ghrelin is a brain-gut peptide with GH-releasing and appetite-inducing activities and a widespread tissue distribution. Ghrelin is the endogenous ligand of the GH secretagogue receptor type 1a (GHS-R1a), and both ghrelin and the GHS-R1a are expressed in the pituitary. There are conflicting data regarding the effects of ghrelin on cell proliferation. A positive effect on proliferation and activation of the mitogen-activated protein kinase (MAPK) pathway has been found in hepatoma, adipose, cardiomyocyte and prostate cell lines. However, ghrelin has also been shown to have anti-proliferative effects on breast, lung and thyroid cell lines. We therefore examined the effect of ghrelin on the rat pituitary cell line GH3. METHODS: RT-PCR was used for the detection of GHS-R1a and pre-proghrelin mRNA expression in GH3 cells. The effect of ghrelin on cell proliferation was studied using [(3)H]thymidine incorporation; cell counting and the activation of the MAPK pathway were studied using immunoblotting and inhibitors of the extracellular signal-regulated kinase 1 and 2 (ERK 1/2), protein kinase C (PKC) and tyrosine phosphatase pathways. RESULTS: GHS-R1a and ghrelin mRNA expression were detected in GH3 cells. Ghrelin, at 10(-10) to 10(-6) M concentrations, significantly increased [(3)H]thymidine incorporation (at 10(-9) M, 183+/-13% (means+/-s.e.m.) compared with untreated controls), while 12-phorbol 13-myristate acetate (PMA) at 10(-7) M (used as a positive control) caused a 212+/-14% increase. A reproducible stimulatory effect of desoctanoyl ghrelin was also observed on [(3)H]thymidine incorporation (135+/-5%; P<0.01 at 10(-9) M compared with control), as well as on the cell count (control 6.8 x 10(4)+/-8.7 x 10(3) cells/ml vs desoctanoyl ghrelin (10(-9) M) 1.04 x 10(5)+/-7.5 x 10(3) cells/ml; P<0.01). Ghrelin caused a significant increase in phosphorylated ERK 1/2 in immunoblotting, while desoctanoyl ghrelin showed a smaller but also significant stimulatory effect. The positive effect of ghrelin and desoctanoyl ghrelin on [(3)H]thymidine incorporation was abolished by the MAPK kinase inhibitor U0126, the PKC inhibitor GF109203X and the tyrosine kinase inhibitor tyrphostin 23, suggesting that the ghrelin-induced cell proliferation of GH3 cells is mediated both via a PKC-MAPK-dependent pathway and via a tyrosine kinase-dependent pathway. This could also be clearly demonstrated by Western blot analysis, where a transient increase in ERK 1/2 phosphorylation by ghrelin was attenuated by all three inhibitors. CONCLUSION: We have shown a novel role for ghrelin in stimulating the proliferation of a somatotroph pituitary tumour cell line, suggesting that ERK activation is involved in mediating the effects of ghrelin on cell proliferation. Desoctanoyl ghrelin showed a similar effect. As ghrelin has been shown to be expressed in both normal and adenomatous pituitary tissue, locally produced ghrelin may play a role in pituitary tumorigenesis via an autocrine/paracrine pathway.

scholarly journals Platyphyllenone Induces Autophagy and Apoptosis by Modulating the AKT and JNK Mitogen-Activated Protein Kinase Pathways in Oral Cancer Cells

2021 ◽  
Vol 22 (8) ◽  
pp. 4211
Author(s):  
Yen-Tze Liu ◽  
Hsin-Yu Ho ◽  
Chia-Chieh Lin ◽  
Yi-Ching Chuang ◽  
Yu-Sheng Lo ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Oral Cancer ◽  
Protein Expression ◽  
Caspase 3 ◽  
Therapeutic Option ◽  
Mapk Signaling ◽  
Mitogen Activated Protein Kinase ◽  
Dependent Manner ◽  
Cancer Proliferation ◽  
Mitogen Activated Protein

Platyphyllenone is a type of diarylheptanoid that exhibits anti-inflammatory and chemoprotective effects. However, its effect on oral cancer remains unclear. In this study, we investigated whether platyphyllenone can promote apoptosis and autophagy in SCC-9 and SCC-47 cells. We found that it dose-dependently promoted the cleavage of PARP; caspase-3, -8, and -9 protein expression; and also led to cell cycle arrest at the G2/M phase. Platyphyllenone up-regulated LC3-II and p62 protein expression in both SCC-9 and SCC-47 cell lines, implying that it can induce autophagy. Furthermore, the results demonstrated that platyphyllenone significantly decreased p-AKT and increased p-JNK1/2 mitogen-activated protein kinase (MAPK) signaling pathway in a dose-dependent manner. The specific inhibitors of p-JNK1/2 also reduced platyphyllenone-induced cleavage of PARP, caspase-3, and caspase -8, LC3-II and p62 protein expression. These findings are the first to demonstrate that platyphyllenone can induce both autophagy and apoptosis in oral cancers, and it is expected to provide a therapeutic option as a chemopreventive agent against oral cancer proliferation.

scholarly journals Phosphothreonine Lyase Promotes p65 Degradation in a Mitogen-Activated Protein Kinase/Mitogen- and Stress-Activated Protein Kinase 1-Dependent Manner

2018 ◽  
Vol 87 (1) ◽  
Author(s):  
Mingyu Hou ◽  
Wenhui Wang ◽  
Feizi Hu ◽  
Yuanxing Zhang ◽  
Dahai Yang ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Pathogenic Bacteria ◽  
Critical Role ◽  
Nuclear Factor Κb ◽  
Mapk Signaling ◽  
Mitogen Activated Protein Kinase ◽  
Dependent Manner ◽  
Content Type ◽  
Catalytic Active Site ◽  
Mitogen Activated Protein

ABSTRACT Bacterial phosphothreonine lyases have been identified to be type III secretion system (T3SS) effectors that irreversibly dephosphorylate host mitogen-activated protein kinase (MAPK) signaling to promote infection. However, the effects of phosphothreonine lyase on nuclear factor κB (NF-κB) signaling remain largely unknown. In this study, we detected significant phosphothreonine lyase-dependent p65 degradation during Edwardsiella piscicida infection in macrophages, and this degradative effect was blocked by the protease inhibitor MG132. Further analysis revealed that phosphothreonine lyase promotes the dephosphorylation and ubiquitination of p65 by inhibiting the phosphorylation of mitogen- and stress-activated protein kinase-1 (MSK1) and by inhibiting the phosphorylation of extracellular signal-related kinase 1/2 (ERK1/2), p38α, and c-Jun N-terminal kinase (JNK). Moreover, we revealed that the catalytic active site of phosphothreonine lyase plays a critical role in regulating the MAPK-MSK1-p65 signaling axis. Collectively, the mechanism described here expands our understanding of the pathogenic effector in not only regulating MAPK signaling but also regulating p65. These findings uncover a new mechanism by which pathogenic bacteria overcome host innate immunity to promote pathogenesis.

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