• BMB Reports
• /
• v.49 no.10
• /
• pp.566-571
• /
• 2016

Forskolin (FSK), an adenylyl cyclase activator, has recently been shown to enhance nucleotide excision repair (NER) upon UV exposure. However, our study revealed that this effect was detected in human skin epithelial ARPE19 cells only in growing cells, but not in non-cycling cells. When the cells were grown at low density (70% confluence), FSK was capable of stimulating cAMP responsive element binding (CREB) phosphorylation, a marker for FSK-stimulated PKA activation, and resulted in a significant increase of NER activity compared to control treatment. However, cells grown under 100% confluent conditions showed neither FSK-induced CREB phosphorylation nor the resulting NER enhancement. These findings indicate that cellular growth is critical for FSK-induced NER enhancement and suggest that cellular growth conditions should be considered as a variable while evaluating a reagent's pharmacotherapeutic efficacy.

• Journal of Microbiology and Biotechnology
• /
• v.33 no.12
• /
• pp.1576-1586
• /
• 2023

Vaccination is the most effective method for preventing the spread of the influenza virus. Cell-based influenza vaccines have been developed to overcome the disadvantages of egg-based vaccines and their production efficiency has been previously discussed. In this study, we investigated whether treatment with forskolin (FSK), an adenylyl cyclase activator, affected the output of a cell-based influenza vaccine. We found that FSK increased the propagation of three influenza virus subtypes (A/H1N1/California/4/09, A/H3N2/Mississippi/1/85, and B/Shandong/7/97) in Madin-Darby canine kidney (MDCK) cells. Interestingly, FSK suppressed the growth of MDCK cells. This effect could be a result of protein kinase A (PKA)-Src axis activation, which downregulates extracellular signal-regulated kinase (ERK)1/2 activity and delays cell cycle progression from G1 to S. This delay in cell growth might benefit the binding and entry of the influenza virus in the early stages of viral replication. In contrast, FSK dramatically upregulated ERK1/2 activity via the cAMP-PKA-Raf-1 axis at a late stage of viral replication. Thus, increased ERK1/2 activity might contribute to increased viral ribonucleoprotein export and influenza virus propagation. The increase in viral titer induced by FSK could be explained by the action of cAMP in assisting the entry and binding of the influenza virus. Therefore, FSK addition to cell culture systems could help increase the production efficiency of cell-based vaccines against the influenza virus.

• The Korean Journal of Physiology and Pharmacology
• /
• v.25 no.3
• /
• pp.189-195
• /
• 2021

Fluoxetine (FLX), a selective serotonin reuptake inhibitor antidepressant, exhibits various other mechanisms of action in numerous cell types and has been shown to induce cell death in cancer cells, paving the way for its potential use in cancer therapy. The aim of this study was to determine the off-target effects of the anti-depressant drug FLX, on the human ovarian granulosa tumor COV434 cells stimulated by forskolin (FSK), by measuring the real-time kinetics of intracellular cyclic AMP (cAMP), ATP level, cytoplasmic calcium ([Ca2+]cyt) and survival of COV434 cells. We show that incubating COV434 cells with FLX (between 0.6 and 10 μM) induces a decrease in intracellular cAMP response to FSK, a drop in ATP content and stimulates cytoplasmic Ca2+ accumulation in COV434 cells. Only the highest concentrations of FLX (5-10 μM) diminished cell viability. The present report is the first to identify an action mechanism of FLX in human tumor ovarian cells COV434 cells and thus opening the way to potential use of fluoxetine as a complementary tool, in granulosa tumor treatments.

• Fisheries and Aquatic Sciences
• /
• v.1 no.2
• /
• pp.153-158
• /
• 1998

Effects of cyclic (c) AMP and G-protein related reagents (3-isobutyl-l-methyxanthine (IBMX), Forskolin (FSK), cholera toxin (CTX), and pertussis toxin (PTX≫ on estradiol-17$\beta$ induced vitellogenin (VTG) induction were examined in primary hepatocyte cultures in rainbow trout Oncorhynchus mykiss. The addition of IBMX, FSK, or CTX to the incubation medium markedly increased VTG production, while PTX was not effective in stimulating the production. It is well known that cAMP regulates phosphorylation and dephosphorylation through mediation of protein kinase A. These results suggest that VTG production is highly dependent on cAMP state in hepatocytes because of its highly phosphorylated nature.

• The Korean Journal of Physiology and Pharmacology
• /
• v.7 no.2
• /
• pp.79-84
• /
• 2003

We have previously reported that expression of the somatostatin receptor subtypes, sst1-5, is differentially regulated by growth hormone (GH)-releasing hormone (GHRH) and forskolin (FSK), in vitro. GHRH binds to membrane receptors selectively located on pituitary somatotropes, activates adenylyl cyclase (AC) and increases sst1 and sst2 and decreases sst5 mRNA levels, without significantly altering the expression of sst3 and sst4. In contrast FSK directly activates AC in all pituitary cell types and increases sst1 and sst2 mRNA levels and decreases sst3, sst4 and sst5 expression. Two explanations could account for these differential effects: 1) GHRH inhibits sst3 and sst4 expression in somatotropes, but this inhibitory effect is masked by expression of these receptors in unresponsive pituitary cell types, and 2) FSK inhibits sst3 and sst4 expression levels in pituitary cell types other than somatotropes. To differentiate between these two possibilities, somatotropes were sequentially labeled with monkey anti-rat GH antiserum, biotinylated goat anti-human IgG, and streptavidin-PE and subsequently purified by fluorescent-activated cell sorting (FACS). The resultant cell population consisted of 95% somatotropes, as determined by GH immunohistochemistry using a primary GH antiserum different from that used for FACS sorting. Purified somatotropes were cultured for 3 days and treated for 4 h with vehicle, GHRH (10 nM) or FSK ($10{\mu}M$). Total RNA was isolated by column extraction and specific receptor mRNA levels were determined by semi-quantitative multiplex RT-PCR. Under basal conditions, the relative expression levels of the various somatostatin receptor subtypes were sst2＞sst5＞sst3=sst1＞ sst4. GHRH treatment increased sst1 and sst2 mRNA levels and decreased sst3, sst4 and sst5 mRNA levels in purified somatotropes, comparable to the effects of FSK on purified somatotropes and mixed pituitary cell cultures. Taken together, these results demonstrate that GHRH acutely modulates the expression of all somatostatin receptor subtypes within GH-producing cells and its actions are likely mediated by activation of AC.

• The Korean Journal of Physiology and Pharmacology
• /
• v.13 no.6
• /
• pp.503-510
• /
• 2009

To elucidate the mechanism of cyclic nucleotides, such as adenosine 3',5'-cyclic monophosphate (cAMP) and guanosine 3',5'-cyclic monophosphate (cGMP), in the regulation of human gastric motility, we examined the effects of forskolin (FSK), isoproterenol (ISO) and sodium nitroprusside (SNP) on the spontaneous, high $K^+$ and acetylcholine (ACh)-induced contractions of corporal circular smooth muscle in human stomach. Gastric circular smooth muscle showed regular spontaneous contraction, and FSK, ISO and SNP inhibited its phasic contraction and basal tone in a concentration-dependent manner. High $K^+$ (50 mM) produced sustained tonic contraction, and ACh $(10\;{\mu}M)$ produced initial transient contraction followed by later sustained tonic contraction with superimposed phasic contractions. FSK, ISO and SNP inhibited high $K^+$-induced tonic contraction and also ACh-induced phasic and tonic contraction in a reversible manner. Nifedipine $(1\;{\mu}M)$, inhibitor of voltage-dependent L-type calcium current $(VDCC_L)$, almost abolished ACh-induced phasic contractions. These findings suggest that FSK, ISO and SNP, which are known cyclic nucleotide stimulators, inhibit smooth muscle contraction in human stomach partly via inhibition of $VDCC_L$.

• Journal of Ginseng Research
• /
• v.45 no.5
• /
• pp.555-564
• /
• 2021

Background: Ginsenosides of Panax ginseng are used to enhance skin health and beauty. The present study aimed to investigate the potential use of ginsenoside Rf (Rf) from Panax ginseng as a new anti-pigmentation agent. Methods: The anti-melanogenic effects of Rf were explored. The transcriptional activity of the cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) and the expression levels of tyrosinase, microphthalmia-associated transcription factor (MITF), and tyrosinase-related proteins (Tyrps) were evaluated in melanocytes and UV-irradiated ex vivo human skin. Results: Rf significantly inhibited Forskolin (FSK) or UV-stimulated melanogenesis. Consistently, cellular tyrosinase activity and levels of MITF, tyrosinase, and Tyrps were downregulated. Furthermore, Rf suppressed MITF promoter activity, which was stimulated by FSK or CREB-regulated transcription coactivator 3 (CRTC3) overexpression. Increased CREB phosphorylation and protein kinase A (PKA) activity induced by FSK were also mitigated in the presence of Rf. Conclusion: Rf can be used as a reliable anti-pigmentation agent, which has a scientifically confirmed and reproducible action mechanism, via inhibition of CREB/MITF pathway.

• Archives of Pharmacal Research
• /
• v.27 no.4
• /
• pp.396-401
• /
• 2004

The effects of lipopolysaccharide (LPS) and several cytokines or the c-fos and c-jun mRNA expression were examined in primary cultured astrocytes. Either LPS (500 ng/mL) or inter-feron-$\gamma$ (IFN-$\gamma$ 5 ng/mL) alone increased the level of c-fos mRNA (1 h). However, tumor necro-sis factor-$\alpha$ (TNF-$\alpha$; 10 ng/mL) or interleukin-4 (IL-1$\beta$: 5 ng/mL) alone showed no significant induction of the level of c-fos mRNA. TNF-$\alpha$ showed a potentiating effect in the regulation of LPS-induced c-fos mRNA expression, whereas LPS showed an inhibitory action against IFN-Y-induced c-fos mRNA expression. LPS, but not TNF-$\alpha$, IL-1$\beta$ and IFN-$\gamma$, increased the level of c-jun mRNA (1 h). TNF-$\alpha$ and IFN-$\gamma$ showed an inhibitory action against LPS-induced c-jun mRNA expression. Both phorbol 12-myristate 13-acetate (PMA; 2.5 mM) and forskolin (FSK, 5 mM) increased the c-fos and c-jun mRNA expressions. In addition, the level of c-fos mRNA was expressed in an antagonistic manner when LPS was combined with PMA. When LPS was co-treated with either PMA or FSK, it showed an additive interaction for the induction of c-jun mRNA expression. Our results suggest that LPS and cytokines may be actively involved in the regulation of c-fos and c-jun mRNA expressions in primary cultured astrocytes. Moreover, both the PKA and PKC pathways may regulate the LPS-induced c-fos and c-jun mRNA expressions in different ways.

• The Korean Journal of Physiology and Pharmacology
• /
• v.11 no.5
• /
• pp.207-213
• /
• 2007

This study was designed to characterize ureteral smooth muscle motility and also to study the effect of forskolin(FSK) and isoproterenol(ISO) on smooth muscle contractility in murine ureter. High $K^+$(50 mM) produced tonic contraction by $0.17{\pm}0.06mN$(n=19). Neuropeptide and neurotransmitters such as serotonin($5{\mu}M$), histamine($20{\mu}M$), and carbarchol(CCh, $10{\sim}50{\mu}M$) did not produce significant contraction. However, CCh($50{\mu}M$) produced slow phasic contraction in the presence of 25 mM $K^+$. Cyclopiazonic acid(CPA, $10{\mu}M$), SR $Ca^{2+}$-ATPase blocker, produced tonic contraction(0.07 mN). Meanwhile, inhibition of mitochondria by protonophore carbnylcyanide m-chlorophenylhydrazone(CCCP) also produced weak tonic contraction(0.01 mN). The possible involvement of $K^+$ channels was also pursued. Tetraethyl ammonium chloride(TEA, 10 mM), glibenclamide($10{\mu}M$) and quinidine($20{\mu}M$) which are known to block $Ca^{2+}$-activated $K^+$ channels($K_{Ca}$ channel), ATP-sensitive $K^+$ channels($K_{ATP}$) and nonselective $K^+$ channel, respectively, did not elicit any significant effect. However, $Ba^{2+}$($1{\sim}2mM$), blocker of inward rectifier $K^+$ channels($K_{IR}$ channel), produced phasic contraction in a reversible manner, which was blocked by $1{\mu}M$ nicardipine, a blocker of dehydropyridine-sensitive voltage-dependent L-type $Ca^{2+}$ channels($VDCC_L$) in smooth muscle membrane. This $Ba^{2+}$-induced phasic contraction was significantly enhanced by $10{\mu}M$ cyclopiazonic acid(CPA) in the frequency and amplitude. Finally, regulation of $Ba^{2+}$-induced contraction was studied by FSK and ISO which are known as adenylyl cyclase activator and $\beta$-adrenergic receptor agonist, respectively. These drugs significantly suppressed the frequency and amplitude of $Ba^{2+}$-induced contraction(p<0.05). These results suggest that $Ba^{2+}$ produces phasic contraction in murine ureteral smooth muscle which can be regulated by FSK and $\beta$-adrenergic stimulation.