• Journal of Applied Biological Chemistry
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• v.66
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• pp.236-243
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• 2023

Platelets act a fundamental role in primary- and secondary-hemostasis, however, platelet activation may cause thrombosis simultaneously. Therefore, control of platelet aggregation is crucial in preventing thrombosis-mediated diseases. Recently, the development of insect materials is attracting attention. Among the highly nutritious functional food sources, insects such as two-spotted cricket (Gryllus bimaculatus). Gryllus bimaculatus (G. bimaculatus) contains high protein and unsaturated fatty acids and has been registered as a food material September 2015 by the Ministry of Food and Drug Safety of Korea. In this study, we examined whether G. bimaculatus extract (GBE) inhibits platelet aggregation, intracellular calcium mobilization, thromboxane A₂ production and glycoprotein IIb/IIIa (integrin αIIb/β3) activation. We investigated whether GBE can regulate signaling molecules, such as 1, 4, 5-triphosphate receptor type I, extracellular signal-regulated kinase, cytosolic phospholipase A₂, mitogen-activated protein kinases p38, vasodilator-stimulated phosphoprotein, phosphatidylinositol-3 kinase, Akt, glycogen synthase kinase-3α/β, and SYK. Taken together, GBE is a potential therapeutic drug candidate to prevent platelet-related thrombosis and cardiovascular disease.

• Journal of Korean Medicine for Obesity Research
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• v.24 no.1
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• pp.25-40
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• 2024

Objectives: In the present study, we investigated the effects of clean-diabetes mellitus 3 (C-DM3), a herbal formula with Trichosanthis Radix, Coptidis Rhizoma, Crataegi Fructus, and Cinnamomi Cortex, on the pathological and serological symptoms of diabetes and its related molecular mechanisms in diet-induced obese mice. Methods: We prepared an obese mouse model using a high-fat diet for 8 weeks and then administered the C-DM3 extract for 4 weeks. The changes of pathological and serological biomarkers for diabetes assessment were measured in the mice and histological changes were observed in the liver and pancreas tissues. We also identified the main compounds in the C-DM3 extract using high pressure liquid chromatography (HPLC) and analyzed the molecular mechanism of the disease condition by network pharmacological analysis. Results: In the in vivo, the administration of C-DM extract to obese mice significantly reduced body weight gain, fatty liver symptoms, and muscle loss, and decreased the levels of fasting blood glucose, insulin, aspertate aminotransferase, triglycerides, and low-density lipoprotein-cholesterol. In addition, C-DM extract significantly increased the phosphorylation of insulin receptor substrate 1, protein kinase b (AKT), phosphoinositide 3-kinase (PI3K), adenosine monophosphate-activated protein kinase, and glucose transporter 4 in all pancreatic and liver tissues, with inhibition of histopathological changes in obese mice. HPLC analysis identified hyperoside, berberine, epiberberine, columbamin, coptisine, coumarin, jatrorrhizine, and citric acid as the main compounds. In the network pharmacological analysis, the molecular targets of C-DM3 extract on obesity and diabetes were shown as the insulin, AKT, PI3K, and mitogen-activated protein kinase pathways with the regulation of inflammatory molecules interleukin 6 (IL-6), jun proto-oncogene, and IL-1β, which matched our in vivo targets. Conclusions: Based on these results, C-DM3 extract is expected to be effective in improving obesity and preventing diabetic progression.

• Journal of Life Science
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• v.30 no.2
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• pp.137-146
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• 2020

Calcium is one of the important secondary signaling molecules in plant cells. Calcium-dependent protein kinases (CDPK)-the sensor proteins of Ca²⁺ and phosphorylating enzymes-are the most abundant serine/threonine kinases in plant cells. They convert and transmit signals in response to various stimuli, resulting in specific responses in plants. In rice, 31 CDPK gene families have been identified, which are mainly involved in plant growth and development and are known to play roles in response to various stress conditions. However, little is known about the biochemical characteristics of CDPK proteins. In this study, OsCPK11-a CDPK in rice-was partially purified, and its biochemical characteristics were found. Partially purified OsCPK11 from rice seedlings was obtained by three-step column chromatography that involved anion exchange chromatography consisting of DEAE, hydrophobic interaction chromatography consisting of phenyl-Sepharose, and gel filtration chromatography consisting of Sephacryl-200HR. An in vitro kinase assay using partially purified OsCPK11 was also performed. This partially purified OsCPK11 had a molecular weight of 54 kDa and showed a strong hydrophobic interaction with the hydrophobic resin. In vitro kinase assay showed that the OsCPK11 also had Ca²⁺-dependent autophosphorylation activity. The OsCPK11 phosphorylated histone III-S, and the optimum pH for its kinase activity was found to be 7.5~8.0. The native OsCPK11 shared several biochemical characteristics with recombinant OsCPK11 studied previously, and both had Ca²⁺-dependent autophosphorylation activity and favored histone III-S as a substrate for kinase activity, which also had a Ca²⁺-dependence.

• Applied Microscopy
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• v.35 no.3
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• pp.121-128
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• 2005

It has been known that ras signaling transduction leads to cell proliferation and migration including various adaptor molecules. Dynamin protein has been implicated in the formation of nascent vesicles in both the endocytic and secretory pathways. Dynamin was classified into three isoforms: dynamin I is only expressed in neuronal tissue, dynamin II is expressed ubiquitously in all tissue but that of dynamin III is confined to testis. We have reported in previous study that Grb2, binding to ras, was associated with dynamin II in NIH3T3 cells. Therefore we have tried to identify the relative expression of dynamin II according to overexpressed ras protein in ras oncogene transfected cells (NIH3T3 (ras)). For the detection of differential expression of dynamin II, we have used immunofluorescent staining and western blot methods in NIH3T3 and NIH3T3 (ras) cells. Next we have described the morphological differences between NIH3T3 and NIH3T3 (ras) cells using SEM and TEM. From these experiments dynamin II was highly expressed in NIH3T3 (ras) cells. NIH3T3 cells was transformed to more spindle shape with many cell process by transfection of ras oncogene. Moreover dynamin II was more concentrated in endocytotic membrane of the NIH3T3 (ras) cells compared to that of NIH3T3 cells. The present results suggested that dynamin II may involve the intermediate messenger in Ras signaling transduction pathway.

• Clinical and Experimental Reproductive Medicine
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• v.37 no.3
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• pp.207-216
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• 2010

Objective: This study was performed to clarify the role of HomeoboxA (HOXA) and its related signaling molecules in the decidualization of primary cultured endometrial cells. Methods: Human endometrial tissues were obtained by curettage of hysterectomy specimens from patients with conditions other than endometrial diseases. Tissues were minced and digested with Trypsin-EDTA for 20 min, $37^{\circ}C$. Cells were cultured with DMEM/F12 medium in $37^{\circ}C$, 5% $CO_2$ incubator for 24 hrs. Cells were treated with HOXA10 siRNA and added transforming growth factor (TGF)-${\beta}1$ (10 ng/mL) for 48 hrs to induces decidualization in vitro. Reverse transcription polymerase chain reaction analysis was accomplished to observe the expression of HOXA10, prolactin, cyclooxygenase (COX)-2, peroxisome proliferatoractivated receptor (PPAR)-$\gamma$, and wingless-type MMTV integration site family (Wnt). Results: HOXA10 expression was increased (1.8 fold vs. non-treated control) in TGF-${\beta}1$ treated cells. Decidualization marker, prolactin, was significantly increased in TGF-${\beta}1$ treated cells compared with HOXA10 siRNA treated cells. Endometrial cell differentiation marker, COX-2 was down-regulated by HOXA10 siRNA even if cells were treated with TGF-${\beta}1$. Wnt4 was down-regulated by treated with HOXA10 siRNA, this expression patters was not changed by TGF-${\beta}1$. Expression of PPAR-$\gamma$ was down regulated by TGF-${\beta}1$ in regardless of HOXA10 siRNA treatment. Conclusion: TGF-${\beta}1$ which is induced by progesterone in endometrial epithelial cells may induces stromal cell decidualization via HOXA10 and Wnt signaling cascade.

• Development and Reproduction
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• v.13 no.1
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• pp.1-11
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• 2009

Implantation itself is governed by an array of endocrine, paracrine and autocrine modulators, of embryonic and maternal origin. Window of implantation is the unique temporal and spatial expression of factors allows the embryo to implant via signaling, appositioning, attachment, and invasion in a specific time frame of $2{\sim}4$ days. When the embryo has arrived in the uterine cavity, a preprogrammed sequence of events occurs, which involves the production and secretion of a multitude of biochemical factors such as cytokines, growth factors, and adhesion molecules by the endometrium and the embryo, thus leading to the formation of a receptive endometrium. Cytokines such as LIF, CSF-1, and IL-1 have all been shown to play important roles in the cascade of events that leads to implantation. Integrin, L-selectin ligands, glycodelin, mucin-1, HB-EGF and pinopodes are involved in appositioning and attachment. The embryo also produces cytokines and growth factors (ILs, VEGF) and receptors for endometrial signals such as LIF, CSF-1, IGF and HB-EGF. The immune system and angiogenesis play an important role. The usefulness of these factors to assess endometrial receptivity and to estimate the prognosis for pregnancy in natural and artificial cycles remains to be proven. Integrins, pinopodes, glycodelin and LIF (from biopsies) are promising candidates; from uterine flushings, glycodelin and LIF are also candidates. The ideal serum marker is not available, but VEGF, glycodelin and CSF have some clinical implications. Further evaluation that includes larger groups of infertile women and fertile controls are needed to elucidate whether their presence in plasma, flushing fluid, or endometrial samples can be used as some kind of a screening tool to assess endometrial function and prognosis for pregnancy before and after artificial reproductive therapy. A better understanding of their function in human implantation may lead to therapeutic intervention, thereby improving the success rate in reproduction treatment. New molecular techniques are becoming available for measuring both embryonic and endometrial changes prior to and during implantation. The use of predictive sets of markers may prove to be more reliable than a single marker. Ultimately, the aim is to use these tools to increase implantation in artificial cycles and consequently improve live-birth rates.

• Journal of Life Science
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• v.23 no.11
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• pp.1397-1403
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• 2013

Fructus Sophorae, the dried ripe fruit of Styphnolobium japonicum (L.), is an herbal ingredient used in traditional Oriental medicine. This study was carried out to investigate the effects of Fructus Sophorae extracts (FSE) on immune modulation in a murine RAW 264.7 macrophage model. As immune response parameters, the production of prostaglandin $E_2$ ($PGE_2$) and tumor necrotic $factor-{\alpha}$ ($TNF-{\alpha}$) were evaluated. Our data revealed that FSE increased the macrophage activation and the production of $PGE_2$ and $TNF-{\alpha}$, which was consistently correlated with upregulation of cyclooxygenase-2 (COX-2) and $TNF-{\alpha}$ expression at both transcriptional and translational levels. On comparative cytokine protein array, FSE significantly increased several cytokines, which was associated with phosphorylation of mitogen- activated protein kinases (MAPKs), including extracellular signal-regulated kinase (ERK), p38 MAPK and c-Jun N-terminal kinase (JNK), and Akt in RAW 264.7 cells. However, each inhibitor of these molecules attenuated the FSE-induced $PGE_2$ production. These results indicate that FSE activated macrophages through the activation of MAPKs and phosphatidylinositol-3-kinase (PI3K)/Akt signaling pathways in RAW 264.7 macrophages. These findings suggest that FSE may provide a promising source of an immunoenhancing agent.

• Tuberculosis and Respiratory Diseases
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• v.82 no.1
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• pp.42-52
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• 2019

Background: Transforming growth factor ${\beta}$ (TGF-${\beta}$), retinoic acid (RA), p38 mitogen-activated protein kinase (MAPK), and MEK signaling play critical roles in cell differentiation, proliferation, and apoptosis. We investigated the effect of RA and the role of these signaling molecules on the phosphorylation of Smad2/3 (p-Smad2/3) induced by TGF-${\beta}1$. Methods: A549 epithelial cells and CCD-11Lu fibroblasts were incubated and stimulated with or without all-trans RA (ATRA) and TGF-${\beta}1$ and with MAPK or MEK inhibitors. The levels of p-Smad2/3 were analyzed by western blotting. For animal models, we studied three experimental mouse groups: control, bleomycin, and bleomycin+ATRA group. Changes in histopathology, lung injury score, and levels of TGF-${\beta}1$ and Smad3 were evaluated at 1 and 3 weeks. Results: When A549 cells were pre-stimulated with TGF-${\beta}1$ prior to RA treatment, RA completely inhibited the p-Smad2/3. However, when A549 cells were pre-treated with RA prior to TGF-${\beta}1$ stimulation, RA did not completely suppress the p-Smad2/3. When A549 cells were pre-treated with MAPK inhibitor, TGF-${\beta}1$ failed to phosphorylate Smad2/3. In fibroblasts, p38 MAPK inhibitor suppressed TGF-${\beta}1$-induced p-Smad2. In a bleomycin-induced lung injury mouse model, RA decreased the expression of TGF-${\beta}1$ and Smad3 at 1 and 3 weeks. Conclusion: RA had inhibitory effects on the phosphorylation of Smad induced by TGF-${\beta}1$ in vitro, and RA also decreased the expression of TGF-${\beta}1$ at 1 and 3 weeks in vivo. Furthermore, pre-treatment with a MAPK inhibitor showed a preventative effect on TGF-${\beta}1$/Smad phosphorylation in epithelial cells. As a result, a combination of RA and MAPK inhibitors may suppress the TGF-${\beta}1$-induced lung injury and fibrosis.

• Korean Journal of Plant Resources
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• v.34 no.1
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• pp.23-30
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• 2021

Hemistepta lyrata Bunge (HL) has been used as a folk remedy to treat cancer, inflammation, bleeding, hemorrhoids and fever, and leaves and young shoots have been used as famine food. Nevertheless, the biological activities and underlying mechanisms of the anti-inflammatory effects remain unclear. In this study, it was undertaken to explore the functions of the aerial part of HL as a suppressor of inflammation by using RAW 264.7 cells. As immune response parameters, the productions of as nitric oxide (NO) and prostaglandin E2 (PGE2), cytokines such tumor necrotic factor (TNF)-α and interleukin (IL)-6 were evaluated. Although the release of TNF-α remained unchanged in HL-treated RAW 264.7 cells, the productions of NO, PGE2 and IL-6 were significantly increased at concentrations with no cytotoxicity. Furthermore, HL significantly attenuated the mitogen-activated protein kinases (MAPK) pathway including decreasing the phosphorylation of the extracellular signal-regulated kinase (ERK1/2) and p38 mitogen-activated protein kinases. Collectively, this study provides evidence that HL inhibits the production of major pro-inflammatory molecules in LPS-stimulated RAW 264.7 cells via suppression of ERK and P38 MAPK signaling pathways. These findings suggest that the beneficial therapeutic effects of HL may be attributed partly to its ability to modulate immune functions in macrophages.

• Journal of Apiculture
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• v.34 no.3
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• pp.245-254
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• 2019

We investigated the anti-tumor effects and molecular mechanism of Brazil, China and Korean regional propolis on HeLa ovarian cancer cell line. Each propolis extracts was prepared by ethanol extraction method. Cytotoxicity of propolis extracts was determinated by EZ-cytox cell viability assay. To necessity of anti-tumor effect and molecular mechanism of propolis, we must be adjusting propolis concentration. Due to 100 ㎍/mL of propolis extract were reduced cell viability to less than 50%, we adjusted all of propolis concentration to 100 ㎍/mL. By Western blotting analysis, we confirmed that anti-tumor mechanism of Brazil, China and Korea regional propolis has significantly difference. All of propolis was activated apoptosis related molecules such as PARP, caspase-3. However, cell proliferation signaling molecules including Akt1, ERK and Bcl-2 were reduced the protein expression level. Especially, the expression of tumor suppressor protein p53 was significantly increased in propolis-treated group such as Gyeonggi, Chungbuk, Chungnam, Jeonbuk, Gyeongnam and China. The phosphorylation of Bax which as apoptosis indicator was appeared in propolis-treated group such as Gyeonggi, Gangwon, Chungnam, Gyeongbuk, China. In this results showed that the regional propolis has completely different mechanism in anti-tumor. Thus, propolis extracts may be useful source of functional materials on anti-cancer and it will be able to choose the suitable propolis for cancer therapy by analyzing individual characteristics.