• Molecules and Cells
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• v.27 no.4
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• pp.467-473
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• 2009

Our previous study suggested that OsBWMK1, a gene which encodes a member of the rice MAP kinase family, generates transcript variants which show distinct expression patterns in response to environmental stresses. The transcript variants are generated by alternative splicing and by use of alternative promoters. To test whether the two alternative promoters, pOsBWMK1L (promoter for the OsBWMK1L splice variant) and pOsBWMK1S (promoter for the OsBWMK1S splice variant), are biologically functional, we analyzed transgenic plants expressing GUS fusion constructs for each promoter. Both pOsBWMK1L and pOsBWMK1S are biologically active, although the activity of pOsBWMK1S is lower than that of pOsBWMK1L. Histochemical analysis revealed that pOsBWMK1L is constitutively active in most tissues at various developmental stages in rice and Arabidopsis, whereas pOsBWMK1S activity is spatially and temporally restricted. Furthermore, the expression of pOsBWMK1S::GUS was upregulated in response to hydrogen peroxide, a plant defense signaling molecule, in both plant species. These results suggest that the differential expression of OsBWMK1 splice variants is the result of alternative promoter usage and, moreover, that the mechanisms controlling OsBWMK1 gene expression are conserved in both monocot and dicot plants.

• Journal of Life Science
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• v.29 no.11
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• pp.1200-1207
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• 2019

Ultraviolet radiation exposure is a major cause of extrinsic skin aging, which leads to skin hyperpigmentation. Loganin, a major iridoid glycoside obtained from Corni fructus, has anti-inflammatory, anti-diabetic, and neuroprotective effects. In this study, we investigated the mechanisms underlying the anti-melanogenic effects of loganin in B16F10 melanocytes treated with ${\alpha}$-melanocyte stimulating hormone (${\alpha}-MSH$) and 3-isobutyl-1-methylxanthine (IBMX). Anti-melanogenic activity was measured by treating cells with loganin at concentrations between 1 and $20{\mu}m$. Cell viability assays confirmed that doses of loganin up to $20{\mu}m$ were not cytotoxic. Loganin significantly and dose-dependently decreased intracellular melanin production. We also investigated potential molecular signaling pathways for the anti-melanogenesis effects of loganin. Western blotting showed that treatment with ${\alpha}-MSH$ and IBMX increased the phosphorylation of cAMP response element-binding protein (CREB) and the gene expressions of microphthalmia-associated transcription factor (MITF) and tyrosinase. Addition of loganin suppressed these increases, while promoting the phosphorylation of extracellular signal regulated kinase (ERK) and the anti-melanogenesis response. Our data therefore indicated that loganin could attenuate the increased melanin synthesis induced by ${\alpha}-MSH$ and IBMX treatment of B16F10 melanocytes. This attenuation appears to occur by downregulation of CREB phosphorylation and MITF and tyrosinase gene expression and upregulation of ERK phosphorylation. These finding suggests that loganin could be a valuable candidate for treatment of skin diseases related to hyperpigmentation.

• Animal Bioscience
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• v.34 no.6
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• pp.975-984
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• 2021

Objective: Inbreeding depression of reproduction is a major concern in the conservation of native chicken genetic resources. Here, based on the successful development of strongly inbred (Sinb) and weakly inbred (Winb) Langshan chickens, we aimed to evaluate inbreeding effects on reproductive traits and identify candidate genes involved in inbreeding depression of reproduction in Langshan chickens. Methods: A two-sample t-test was performed to estimate the differences in phenotypic values of reproductive traits between Sinb and Winb chicken groups. Three healthy chickens with reproductive trait values around the group mean values were selected from each of the groups. Differences in ovarian and hypothalamus transcriptomes between the two groups of chickens were analyzed by RNA sequencing (RNA-Seq). Results: The Sinb chicken group showed an obvious inbreeding depression in reproduction, especially for traits of age at the first egg and egg number at 300 days (p<0.01). Furthermore, 68 and 618 differentially expressed genes (DEGs) were obtained in the hypothalamus and ovary between the two chicken groups, respectively. In the hypothalamus, DEGs were mainly enriched in the pathways related to vitamin metabolism, signal transduction and development of the reproductive system, such as the riboflavin metabolism, Wnt signaling pathway, extracellular matrix-receptor interaction and focal adhesion pathways, including stimulated by retinoic acid 6, serpin family F member 1, secreted frizzled related protein 2, Wnt family member 6, and frizzled class receptor 4 genes. In the ovary, DEGs were significantly enriched in pathways associated with basic metabolism, including amino acid metabolism, oxidative phosphorylation, and glycosaminoglycan degradation. A series of key DEGs involved in folate biosynthesis (gamma-glutamyl hydrolase, guanosine triphosphate cyclohydrolase 1), oocyte meiosis and ovarian function (cytoplasmic polyadenylation element binding protein 1, structural maintenance of chromosomes 1B, and speedy/RINGO cell cycle regulator family member A), spermatogenesis and male fertility (prostaglandin D2 synthase 21 kDa), Mov10 RISC complex RNA helicase like 1, and deuterosome assembly protein 1) were identified, and these may play important roles in inbreeding depression in reproduction. Conclusion: The results improve our understanding of the regulatory mechanisms underlying inbreeding depression in chicken reproduction and provide a theoretical basis for the conservation of species resources.

• Journal of Life Science
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• v.29 no.9
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• pp.964-971
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• 2019

Hepatic lipid accumulation and insulin resistance increases in patients with non-alcoholic fatty liver disease. Piperine is a major compound found in black pepper (Piper nigrum) and long pepper (P. longum). Piperine has been used in fine chemical for its anti-cancer, anti-obesity, anti-diabetic, anti-inflammatory and anti-oxidant properties. However, the signaling-based mechanism of piperine and its role as an inhibitor of lipogenesis and insulin resistance in human hepatocyte cells remains ill-defined. In the present study, we explored the effects of piperine on lipid accumulation and insulin resistance, and explored the potential underlying molecular mechanisms in palmitate-treated HepG2 cells. Piperine treatment resulted in a significant reduction of triglyceride content. Furthermore, piperine treatment decreased palmitate-treated intracellular lipid deposition by inhibiting the lipogenic target genes, sterol-regulatory-element-binding protein 1c (SREBP-1c) and fatty acid synthase (FAS); whereas the expression of carnitine palmitoyl transferase (CPT-1) and phosphorylation of acetyl coenzyme A carboxylase (ACC) gene involved in fatty acid oxidation was increased. Moreover, piperine also inhibited the phosphorylation of insulin receptor substrate (IRS)-1 (Ser307). Piperine treatment modulated palmitate-treated lipid accumulation and insulin resistance in HepG2 cells with concomitant reduction of lipogenic target genes, such as SREBP-1 and FAS, and induction of CPT-1-ACC and phosphorylation of IRS-1 (Tyr632)-Akt pathways. Therefore, piperine represents a promising treatment for the prevention of lipid accumulation and insulin resistance.

• Biomolecules & Therapeutics
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• v.27 no.5
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• pp.474-483
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• 2019

Vascular endothelial growth factor (VEGF) plays a pivotal role in pathologic ocular neovascularization and vascular leakage via activation of VEGF receptor 2 (VEGFR2). This study was undertaken to evaluate the therapeutic mechanisms and effects of the tetrapeptide Arg-Leu-Tyr-Glu (RLYE), a VEGFR2 inhibitor, in the development of vascular permeability and choroidal neovascularization (CNV). In cultured human retinal microvascular endothelial cells (HRMECs), treatment with RLYE blocked VEGF-A-induced phosphorylation of VEGFR2, Akt, ERK, and endothelial nitric oxide synthase (eNOS), leading to suppression of VEGF-A-mediated hyper-production of NO. Treatment with RLYE also inhibited VEGF-A-stimulated angiogenic processes (migration, proliferation, and tube formation) and the hyperpermeability of HRMECs, in addition to attenuating VEGF-A-induced angiogenesis and vascular permeability in mice. The anti-vascular permeability activity of RLYE was correlated with enhanced stability and positioning of the junction proteins VE-cadherin, ${\beta}$-catenin, claudin-5, and ZO-1, critical components of the cortical actin ring structure and retinal endothelial barrier, at the boundary between HRMECs stimulated with VEGF-A. Furthermore, intravitreally injected RLYE bound to retinal microvascular endothelium and inhibited laser-induced CNV in mice. These findings suggest that RLYE has potential as a therapeutic drug for the treatment of CNV by preventing VEGFR2-mediated vascular leakage and angiogenesis.

• Biomolecules & Therapeutics
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• v.27 no.5
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• pp.492-501
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• 2019

Nitrogen-containing heterocycles such as quinoline, quinazolinones and indole are scaffolds of natural products and have broad biological effects. During the last years those structures have been intensively synthesized and modified to yield new synthetic molecules that can specifically inhibit the activity of dysregulated protein kinases in cancer cells. Herein, a series of newly synthesized isoquinolinamine (FX-1 to 8) and isoindoloquinazolinone (FX-9, FX-42, FX-43) compounds were evaluated in regards to their anti-leukemic potential on human B- and T- acute lymphoblastic leukemia (ALL) cells. Several biological effects were observed. B-ALL cells (SEM, RS4;11) were more sensitive against isoquinolinamine compounds than T-ALL cells (Jurkat, CEM). In SEM cells, metabolic activity decreased with $10{\mu}M$ up to 26.7% (FX-3), 25.2% (FX-7) and 14.5% (FX-8). The 3-(p-Tolyl) isoquinolin-1-amine FX-9 was the most effective agent against B- and T-ALL cells with IC50 values ranging from 0.54 to $1.94{\mu}M$. None of the tested compounds displayed hemolysis on erythrocytes or cytotoxicity against healthy leukocytes. Anti-proliferative effect of FX-9 was associated with changes in cell morphology and apoptosis induction. Further, influence of FX-9 on PI3K/AKT, MAPK and JAK/STAT signaling was detected but was heterogeneous. Functional inhibition testing of 58 kinases revealed no specific inhibitory activity among cancer-related kinases. In conclusion, FX-9 displays significant antileukemic activity in B- and T-ALL cells and should be further evaluated in regards to the mechanisms of action. Further compounds of the current series might serve as templates for the design of new compounds and as basic structures for modification approaches.

• Journal of the Korean Society of Radiology
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• v.13 no.1
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• pp.125-132
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• 2019

Potassium cyanate (KCN) is an inorganic reagent and can induce the post-translation carbamylation of proteins. The carbamylated reaction in the body is involved in cell death in various diseases. According the results in our previous study, KCN enhances the radiosensitivity of human colorectal cancer cell line, HCT 116 cells. However, it was not enough to confirm the mechanism that KCN works in these cells. To determinated the mechanisms of KCN in the cells with increased radiosensitivity, HCT 116 cells were treated KCN with low-dose gamma-radiation. And then, we examined alteration of the cell cycle, cell proliferation, cytokine level and the activation of cell signaling protein. As a result, cell cycle arrest and cell death were induced by the activation of caspase-3 and PARP in the irradiated cells with KCN treatment. These changes of the irradiated cell with KCN treatment were induced by the release of $TNF-{\alpha}$ via $NF-{\kappa}B$ activation. In conclusions, enhanced radio-sensitivity mediated by KCN induced cell death and it occurs by $NF-{\kappa}B$-dependent $TNF-{\alpha}$ production.

• Journal of Life Science
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• v.29 no.5
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• pp.514-523
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• 2019

We investigated the gene expression patterns and the mechanisms of action of the apoptotic response by microarray analysis of human keratinocyte HaCaT cells treated with ginsenoside Rb1, a saponin of Panax ginseng C. A. Meyer. Genes related to apoptosis, the G2/M transition of the mitotic cell cycle, cell division, mitotic nuclear division, and intracellular protein transport were 2-fold up-regulated in HaCaT cells treated with the ginsenoside Rb1, whereas genes related to DNA repair, regeneration fission, and extracellular matrix organization were 2-fold down-regulated. Apoptosis signaling may be mediated by FAS and PLA2G4A, and pathway analysis indicated that STAT3 might be an upstream regulator of these genes. The activity of FAS and PLA2G4A was verified by qPCR, which showed that FAS was increased about 2-fold in HaCaT cells treated with $10{\mu}g/ml$ of ginsenoside Rb1 for 24 hr, PLA2G4A was increased about twice after 6 hours, and gene expression was increased more than 2-fold after 24 hr. Knockdown of STAT3 with siRNA decreased FAS expression and increased PLA2G4A expression but only FAS was passed from the upstream regulator STAT3. These results indicate that STAT3, which is an upstream regulator, induces apoptosis via FAS during treatment with ginsenoside Rb1.

• Biomolecules & Therapeutics
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• v.29 no.6
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• pp.685-696
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• 2021

In this study, we investigated the inhibitory effect of 5-aminolevulinic acid (ALA), a heme precursor, on inflammatory and oxidative stress activated by lipopolysaccharide (LPS) in RAW 264.7 macrophages by estimating nitric oxide (NO), prostaglandin E2 (PGE2), cytokines, and reactive oxygen species (ROS). We also evaluated the molecular mechanisms through analysis of the expression of their regulatory genes, and further evaluated the anti-inflammatory and antioxidant efficacy of ALA against LPS in the zebrafish model. Our results indicated that ALA treatment significantly attenuated the LPS-induced release of pro-inflammatory mediators including NO and PGE2, which was associated with decreased inducible NO synthase and cyclooxygenase-2 expression. ALA also inhibited the LPS-induced expression of pro-inflammatory cytokines, such as tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6, reducing their extracellular secretion. Additionally, ALA abolished ROS generation, improved the mitochondrial mass, and enhanced the expression of heme oxygenase-1 (HO-1) and the activation of nuclear translocation of nuclear factor-E2-related factor 2 (Nrf2) in LPS-stimulated RAW 264.7 macrophages. However, zinc protoporphyrin, a specific inhibitor of HO-1, reversed the ALA-mediated inhibition of pro-inflammatory cytokines production and activation of mitochondrial function in LPS-treated RAW 264.7 macrophages. Furthermore, ALA significantly abolished the expression of LPS-induced pro-inflammatory mediators and cytokines, and showed strong protective effects against NO and ROS production in zebrafish larvae. In conclusion, our findings suggest that ALA exerts LPS-induced anti-inflammatory and antioxidant effects by upregulating the Nrf2/HO-1 signaling pathway, and that ALA can be a potential functional agent to prevent inflammatory and oxidative damage.

• Journal of Ginseng Research
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• v.45 no.2
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• pp.295-304
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• 2021

Background: Acute promyelocytic leukemia (APL) is a hematopoietic malignancy driven by promyelocytic leukemia-retinoic acid receptor A (PML-RARA) fusion gene. The therapeutic drugs currently used to treat APL have adverse effects. 20(S)-ginsenoside Rh2 (GRh2) is an anticancer medicine with high effectiveness and low toxicity. However, the underlying anticancer mechanisms of GRh2-induced PML-RARA degradation and apoptosis in human APL cell line (NB4 cells) remain unclear. Methods: Apoptosis-related indicators and PML-RARA expression were determined to investigate the effect of GRh2 on NB4 cells. Z-VAD-FMK, LY294002, and C 87, as inhibitors of caspase, and the phosphatidylinositol 3-kinase (PI3K) and tumor necrosis factor-α (TNF-α) pathways were used to clarify the relationship between GRh2-induced apoptosis and PML-RARA degradation. Results: GRh2 dose- and time-dependently decreased NB4 cell viability. GRh2-induced apoptosis, cell cycle arrest, and caspase3, caspase8, and caspase9 activation in NB4 cells after a 12-hour treatment. GRh2-induced apoptosis in NB4 cells was accompanied by massive production of reactive oxygen species, mitochondrial damage and upregulated Bax/Bcl-2 expression. GRh2 also induced PML/PML-RARA degradation, PML nuclear bodies formation, and activation of the downstream p53 pathway in NB4 cells. Z-VAD-FMK inhibited caspase activation and significantly reversed GRh2-induced apoptosis and PML-RARA degradation. GRh2 also upregulated TNF-α expression and inhibited Akt phosphorylation. LY294002, an inhibitor of the PI3K pathway, enhanced the antitumor effects of GRh2, and C 87, an inhibitor of the TNF-α pathway, reversed NB4 cell viability, and GRh2-mediated apoptosis in a caspase-8-dependent manner. Conclusion: GRh2 induced caspase-dependent PML-RARA degradation and apoptosis in NB4 cells via the Akt/Bax/caspase9 and TNF-α/caspase8 pathways.