• Journal of Dental Rehabilitation and Applied Science
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• v.19 no.4
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• pp.281-290
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• 2003

Platelet-rich plasma which is made with the newly developed technique concentrating platelets 3-folds or more is also proven to be very effective method to stimulate and accelerate the healing of bone and soft tissue. This study is aimed to investigate the effect of platelet-rich plasma on the attachment of osteoblast. To evaluate the effect on human, human osteoblast cell line was cultured. Platelet-rich plasma was extracted from the blood of a healthy volunteer. The effect on the attachment was evaluated by MTT assay. To evaluate autocrine and paracrine effect on osteoblast, conditioned medium was made and compared with platelet-rich plasma. By western blot analysis, the expression of fibronectin and vitronectin in experimental groups was examined. The results were as following: The cellular attachment of osteoblast cell line increased depending on the concentration of platelet-rich plasma and conditioned medium. The amount of increasing was similar between two groups. The expression of fibronectin and vitronectin in platelet-rich plasma and conditioned medium is more than control group in western blot analysis. These findings imply that platelet-rich plasma enhance the cellular attachment by inducing fibronectin, vitronectin from osteoblast and maximize the cellular attachment by using the autocrine and paracrine effect of platelet-rich plasma.

• Journal of Physiology & Pathology in Korean Medicine
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• v.25 no.1
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• pp.37-47
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• 2011

This study was aimed to investigate the memory enhancing effect of Jeongjihwan against scopolamine-induced amnesia in C57BL/6 mice. To determine the effect of Jeongjihwan on the memory and cognitive function, we have injected scopolamine (1 mg/kg, i.p.) into C57BL/6 mice 30 min before beginning of behavior tests. We have conducted Y-maze, Morris water-maze, passive avoidance and fear conditioning tests to compare learning and memory functions. Scopolamine-induced behavior changes of memory impairment were significantly restored by oral administration of Jeongjihwan (100 or 200 mg/kg/day). To elucidate the molecular mechanism underlying the memory enhancing effect of Jeongjihwan, we have examined the antioxidant defense system and neurotrophic factors. Jeongjihwan treatment attenuated intracellular accumulation of reactive oxygen species and up-regulated mRNA and protein expression of antioxidant enzymes as assessed by RT-PCR and western blot analysis, respectively. Jeongjihwan also increased protein levels of brain-derived neurotrophic factor (BDNF) compared with those in the scopolamine-treated group. Furthermore, as an upstream regulator, the activation of cAMP response element-binding protein (CREB) via phosphorylation was assessed by Western blot analysis. Jeongjihwan elevated the phosphorylation of CREB (p-CREB), which seemed to be mediated partly by extracellular signal-regulated kinase1/2 (ERK1/2) and protein kinase B/Akt. These findings suggest that Jeongjihwan may have preventive and therapeutic potential in the management of amnesia.

• Molecules and Cells
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• v.22 no.1
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• pp.44-50
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• 2006

We investigated the possible role of p38 MAPK and $ET_B$ receptors in ET-1 induction of cyclooxygenase-2 (COX-2) and prostaglandin $E_2$ ($PGE_2$) in cultured feline esophageal smooth muscle cells (ESMC). Confluent layers of ESMC were stimulated with 10 nM ET-1 and expression of COX-1 and COX-2, involvement of receptors, and activation of p38 MAPK, were examined by Western blot analysis. Levels of $PGE_2$ induced by ET-1 were measured by Elisa. Using $ET_A$and $ET_B$ antagonists (BQ-123 and BQ-788, respectively), the contribution of the ET receptors to COX-1 and COX-2 expression induced by ET-1 was determined. Western blot analysis revealed that treatment of ESMC with ET-1 resulted in transient expression of COX-2 and activation of p38 MAPK. Activation of p38 MAPK was maximal after 1 h. SB202190, a p38 MAPK inhibitor, reduced expression of COX-2, but not COX-1. ET-1-induced release of $PGE_2$ was also blocked by SB202190. COX-2 expression was upregulated only via the $ET_B$ receptor, and COX-1 expression was not affected by either antagonist. Taken together, our data suggest that ET-1 causes p38 MAPK-dependent expression of COX-2 by interacting with $ET_B$ receptors on ESMC.

• Journal of Physiology & Pathology in Korean Medicine
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• v.20 no.6
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• pp.1593-1597
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• 2006

We hope to evaluate the effects of Gami-Choakwiyeum (GCKY) on the PPAR-${\gamma}$’ in the OVA induced asthma mouse model. Female BALB/c mice, 8 weeks of age and free of murine specific pathogens were used. Mice were sensitized by intraperitoneal injection of OVA emulsified in aluminum hydroxide in a total volume of 200 ${\mu}{\ell}$ on one day and 14 days. On 21, 22, and 23 days after the initial intraperitoneal injection of OVA, the mice were challenged using an ultrasonic nebulizer. GCKY was administered 7 times by oral gavage at 24 hour intervals fromdays 19 after intraperitoneal injection of OVA. Bronchoalveolar lavage was perfromed 72 hours after the last challenge, and total cell numbers in the BAL fluid were counted. Also, the level of PPAR-${\gamma}$ of normal and OVA-induced asthma moused with/without administration of GCKY were measured by Western blot analysis. For the histologic examination, the specimens were stained with hematoxylin 2 and eosin-Y.(H & E). Numbers of total cells were increased significantly at 72 h after OVA inhalation compared with numbers of total cells in the normal and the administration of GCKY. Especially, the increased numbers of eosinophils in BAL fluids after OVA inhalation were significantly increased. However, the numbers of eosinophils reduced by the administration of GCKY. Western blot analysis revealed that PPAR-${\gamma}$ levels in nuclear level were increased slightly after OVA inhalation compared with the levels in the normal group. After the administration of GCKY, PPAR-${\gamma}$ levels in cytosolic and nuclear levels at 72 h after OVA inhalation were markedly increased. On pathologic examination, there were many acute inflammatory cells around the alveoli, bronchioles, and airway lumen of mice with OVA-induced asthma compared with inflammatory cells in the normal group. However, acute inflammatory cells around alveoli, bronchioles, and airway lumen markedly decreased after administration of GCKY, GCKY can increase a PPAR-${\gamma}$ level and could be an effective treatment in asthma patients through the PPAR-${\gamma}$ mechanism for bronchial asthma.

• Journal of The Korean Society of Integrative Medicine
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• v.11 no.2
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• pp.119-128
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• 2023

Purpose : The fruit of Actinidia polygama has been used in oriental medicine for the treatment of gout, rheumatoid arthritis, and inflammation. Though A. polygama exhibited anti-inflammatory activity in RAW 264.7 cells and carrageenan-induced rat paw edema, the exact mechanism for anti-inflammation was not evaluated yet. In this study, the anti-inflammatory mechanisms of A. polygama ethanol extract (APEE) in lipopolysaccharide (LPS) stimulated RAW 264.7 cells. Methods : WST-1 assay was applied to analyze the cytotoxic effect of APEE in RAW 264.7 cells. The productions of nitric oxide (NO) and prostaglandin (PG) E₂ were analyzed by the Griess reaction and enzyme immunoassay (EIA) assay, respectively. In addition, protein expressions for inducible NO synthase (iNOS) and cyclooxygenase (COX)-2 were measured by Western blot analysis. The activated status of an inflammatory transcription factor, NF-κ B, and its upstream signaling molecules, mitogen-activated protein kinases (MAPKs), was also evaluated by Western blot analysis. Results : As a result, APEE treatment did not exhibit any cytotoxicity until the concentration of 200 ㎍/㎖. APEE treatment significantly inhibited NO and PGE₂ productions as well as their enzymes, iNOS and COX-2 in a dose-dependent manner. The inflammatory transcription factor, NF-κ B, was also attenuated by APEE treatment. In addition, the phosphorylated status of MAPKs such as extracellular regulated kinase (ERK), c-jun NH2 kinase (JNK), and p38, were significantly diminished by APEE treatment in LPS stimulated RAW 264.7 cells. Conclusion : Consequently, APEE treatment significantly attenuated the production of inflammatory mediators and their enzyme expressions in LPS-stimulated RAW 264.7 cells. The inflammatory transcription factor, NF-κ B, and upstream signaling molecules, MAPKs, were also significantly attenuated by APEE treatment in LPS-activated RAW 264.7 cells. These results indicate that APEE might be a candidate to be utilized as a promising candidate for the treatment of inflammatory disorders.

• Journal of Physiology & Pathology in Korean Medicine
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• v.25 no.1
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• pp.109-114
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• 2011

Atractylodis Rhizoma Alba is commonly used herbal medicine and it has been known that has immuno-regualtory effects and anti-cancer effects. The inhibition of osteoclastogenesis is essential for the prevention and treatment of osteoporosis. The aim of this study was to evaluate the effects of Atractylodis Rhizoma Alba on osteoclast differentiation in vitro and on resorbing activity of osteoclast. Osteoclast formation was evaluated in bone marrow cells (BMC) in the presence or absence of Atractylodis Rhizoma Alba. The expression of c-fos, tartrate-resistant acid phosphatase (TRAP), OSCAR, DC-STAMP, cathepsin K, MafB and NFATc1 mRNA in osteoclast precursor were assessed by RT-PCR. The levels of TNF receptor-associated factor-6 (TRAF-6), c-fos and NFATc1 protein were assessed by Western blot analysis. Also the correlation with MAPKs and NF-${\kappa}B$ pathways were measured by using Western blot analysis. With bone resorption study, I tried to evaluate the inhibitory effects of Atractylodis Rhizoma Alba on mature osteoclast function. Atractylodis Rhizoma Alba inhibited the RANKL induced osteoclastic differentiation from bone marrow macrophage in a dose dependant manner without cellular toxicity. Gene expression of c-fos and NFATc1 was significantly down regulated with Atractylodis Rhizoma Alba treatment. Atractylodis Rhizoma Alba markedly inhibited the RANKL-induced osteoclastogenesis through suppression of nuclear factor kappa b (NF-${\kappa}B$) pathway, down stream pathway of p38, ERK and JNK pathway. Taken together, I concluded that Atractylodis Rhizoma Alba have beneficial effect on osteoporosis by inhibition of osteoclast differentiation and by inhibition of functioning osteoclast. Thus I expect that Atractylodis Rhizoma Alba could be a treatment option for osteoporosis.

• Journal of Physiology & Pathology in Korean Medicine
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• v.22 no.5
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• pp.1322-1329
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• 2008

Mulberry has been reported to contain wide range of polyphenols and have chemopreventive activity. However, little has been known regarding the effect of mulberry fruits on cell viability in human glioma cells. The present study was undertaken to examine the effect of mulberry fruit (Mar; Fructus) on cell viability and to determine its underlying mechanism in human glioma cells. Cell viability and cell death were estimated by MTT assay and trypanblue exclusion assay, respectively. Reactive oxygen species (ROS) generation was measured using the fluorescence probe DCFH-DA. The mitochondrial transmembrane potential was measured with $DiOC_6$(3). Bax expression and cytochrome c release were measured by Western blot analysis. Caspase activity was estimated using colorimetric kit. Mori Fructus resulted in apoptotic cell death in a dose- and time-dependent manner. Mori Fructus increased ROS generation and the Mori Fructus-induced cell death was also prevented by antioxidants, suggesting that ROS generation plays a critical role in Mari Fructus-induced cell death. Western blot analysis showed that Mori Fructus treatment caused an increase in Bax expression, which was inhibited by the antioxidant N-acetylcysteine (NAC). Mori Fructus induced depolarization of mitochondrial membrane potential and its effect was inhibited by the antioxidants NAC and catalase. Mori Fructus induced cytochrome c release, which was inhibited by NAC. Caspase activity was stimulated by Mori Fructus and caspase inhibitors prevented the Mori Fructus-induced cell death. These findings suggest that Mori Fructus results in human glioma cell death through ROS-dependent mitochondrial pathway in human glioma cells.

• Journal of Periodontal and Implant Science
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• v.35 no.3
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• pp.747-760
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• 2005

Periodontal therapy has dealt primarily with attempts at arresting progression of disease. however, more recent techniques have focused on regenerating the periodontal ligament having the capacity to regenerate the periodontium. Recombinant human bone morphogenetic protein-7(rhBMP-7) can differentiate the osteoprogenitor cells and induce bone formation. The purpose of this study was to evaluate the effect of BMP-7 on rat periodontal ligament cells differentiation, in vitro. In the control group, cells was cultured with DMEM media. In the experimental groups, cells were cultured with rhBMP-7 in concentration of 10, 25, 50 and 100 ng/ml. Each group was characterized by examining alkaline phosphatase activity at 3 and 5 days of culture and the ability to produce mineralized nodules of rat calvarial cells at 14 days of culture. Synthesis of type I collagen(COL-I), osteocalcin(OCN), and bone sialoprotein(BSP) was evaluated by RT-PCR at 7 days of culture. Activation of Smad proteins and p38 MAP kinase was determined by western blot analysis of the cell lysates. Alkaline phosphatase activity was significantly increased in the concentration of BMP-7 50 ng/ml and 100 ng/ml compared to the control(p<0.05). The mineralized bone nodule formation was greater with addition of 50 ng/ml and 100 ng/ml BMP-7 than the control(p<0.01). In 7 days' culture, the expressions of COL-I, BSP, and OCN was increased by BMP-7 in concentration of 10 $ng/ml{\sim}100$ ng/ml. In western blot analysis, BMP-7 treated culture cells expressed Smad 1,5,8 in dose-dependent manner, whereas BMP-7 did not activate phosphorylated form of p38 MAP kinase. These result suggested that BMP-7 stimulate rat periodontal ligament cells to differentiate toward osteoblast phenotype and increase bone matrix production by activation of BMP-Smad pathway.

• The Korean Journal of Physiology and Pharmacology
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• v.22 no.3
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• pp.349-360
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• 2018

Autophagy has been studied as a therapeutic strategy for cardiovascular diseases. However, insufficient studies have been reported concerning the influence of vascular smooth muscle cells (VSMCs) through autophagy regulation. The aim of the present study was to determine the effects of VSMCs on the regulation of autophagy under in vitro conditions similar to vascular status of the equipped micro-tubule target agent-eluting stent and increased release of platelet-derived growth factor-BB (PDGF-BB). Cell viability and proliferation were measured using MTT and cell counting assays. Immunofluorescence using an $anti-{\alpha}-tubulin$ antibody was performed to determine microtubule dynamic formation. Cell apoptosis was measured by cleavage of caspase-3 using western blot analysis, and by nuclear fragmentation using a fluorescence assay. Autophagy activity was assessed by microtubule-associated protein light chain 3-II (LC-II) using western blot analysis. Levels of intracellular reactive oxygen species (ROS) were measured using $H_2DCFDA$. The proliferation and viability of VSMCs were inhibited by microtubule regulation. Additionally, microtubule-regulated and PDGF-BB-stimulated VSMCs increased the cleavage of caspase-3 more than only the microtubule-regulated condition, similar to that of LC3-II, implying autophagy. Inhibitory autophagy of microtubule-regulated and PDGF-BB-stimulated VSMCs resulted in low viability. However, enhancement of autophagy maintained survival through the reduction of ROS. These results suggest that the apoptosis of conditioned VSMCs is decreased by the blocking generation of ROS via the promotion of autophagy, and proliferation is also inhibited. Thus, promoting autophagy as a therapeutic target for vascular restenosis and atherosclerosis may be a good strategy.

• Journal of Ginseng Research
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• v.45 no.5
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• pp.575-582
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• 2021

Background: In ginseng, there exists a glycolipoprotein complex with a special form of lipid LPAs called Gintonin. The purpose of this study is to show that Gintonin has a therapeutic effect on rheumatoid arthritis through LPA2 receptors. Methods: Fibroblast-like synoviocytes (FLS) were treated with Gintonin and stimulated with interleukin (IL)-1β. The antioxidant effect of Gintonin was measured using MitoSOX and H₂DCFDA experiments. The anti-arthritic efficacy of Gintonin was examined by analyzing the expression levels of inflammatory mediators, phosphorylation of mitogen-activated protein kinase (MAPK) pathways, and translocation of nuclear factor kappa B (NF-κB)/p65 into the nucleus through western blot. Next, after treatment with LPAR2 antagonist, western blot analysis was performed to measure inflammatory mediator expression levels, and NF-κB signaling pathway. Carrageenan/kaolin-induced arthritis rat model was used. Rats were orally administered with Gintonin (25, 50, and 100 mg/kg) every day for 6 days. The knee joint thickness, squeaking score, and weight distribution ratio (WDR) were measured as the behavioral parameters. After sacrifice, H&E staining was performed for histological analysis. Results: Gintonin significantly inhibited the expression of iNOS, TNF-α, IL-6 and COX-2. Gintonin prevented NF-κB/p65 from moving into the nucleus through the JNK and ERK MAPK phosphorylation in FLS cells. However, pretreatment with an LPA2 antagonist significantly reversed these effects of Gintonin. In the arthritis rat model, Gintonin suppressed all parameters that were measured. Conclusion: This study suggests that LPA2 receptor plays a key role in mediating the anti-arthritic effects of Gintonin by modulating inflammatory mediators, the MAPK and NF-κB signaling pathways.