• Title/Summary/Keyword: In-vitro techniques

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Development of New Materials of Ginseng by Nanoparticles

  • Yang, Deok Chun;Mathiyalagan, Ramya;Yang, Dong Uk;Perez, Zuly Elizabeth Jimenez;Hurh, Joon;Ahn, Jong Chan
    • Proceedings of the Plant Resources Society of Korea Conference
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    • 2018.04a
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    • pp.3-3
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    • 2018
  • For centuries, Panax ginseng Meyer (Korean ginseng) has been widely used as a medicinal herb in Korea, China, and Japan. Ginsenosides are a class of triterpene saponins and recognized as the bioactive components in Korean ginseng. Ginsenosides, which can be classified broadly as protopanaxadiols (PPD), protopanaxatriols (PPT), and oleanolic acids, have been shown to flaunt a vast array of pharmacological activities such as immune-modulatory, anti-inflammatory, anti-tumor, anti-diabetic, and antioxidant effects. In recent years, a number of ginseng and ginsenoside researches have increasingly gained wide attention owing to its unique pharmacological properties. Although good efficacies of ginsenosides have been reported, lack of target specific delivery into tumor sites, low solubility, and low bioavailability due to modifications in gastro-intestinal environments limit their biomedical application in clinical trials. As a result to this major challenge, nanotechnology and drug delivery techniques play a significant role to solve this problematic issue. Thus, we reported the preparation of poly-ethylene glycol (PEG) and glycol chitosan (GC) functionalized to ginsenoside (Compound K and PPD) conjugates via hydrolysable ester bonds with improved aqueous solubility and pH-dependent drug release. In vitro cytotoxicity assays revealed that PEG-CK, and PPD-CK conjugates exhibited lower cytotoxicity compared to bare CK and PPD in HT29 cells. However, GC-CK conjugates exhibited higher and similar cytotoxicity in HT29 and HepG2 cells. Furthermore, GC-CK-treated RAW264.7 cells did not exhibit significant cell death at higher concentration of treatment which supports the biocompatibility of the polymer conjugates. They also inhibited nitric oxide production in lipopolysaccharide (LPS)-induced RAW64.7 cells. In addition to polymer-ginsenoside conjugates, silver (AgNps) and gold nanoparticles (AuNps) have been successfully synthesized by green chemistry using different m. The biosynthesized nanoparticles demonstrated antimicrobial efficacy, anticancer, anti-inflammatory, antioxidant activity, biofilm inhibition, and anticoagulant effect. Special interest on the effective delivery methods of ginsenoside to treatment sites is the focus of metal nanoparticle research.In short, nano-sizing of ginsenoside results in an increased water solubility and bioavailability. The use of nano-sized ginsenoside and P. ginseng mediated metallic nanoparticles is expected to be effective on medical platform against various diseases in the future.

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Gossypii Semen oil alleviates memory dysfunction in scopolamine-treated mice (면화자 정유의 기억력 손상 완화 효과)

  • Lee, Jihye;Jung, Eun Mi;Lee, Eunhong;Jang, Gwi Yeong;Seo, Kyung Hye;Kim, Mi Ryeo;Jung, Ji Wook
    • The Korea Journal of Herbology
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    • v.36 no.2
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    • pp.1-9
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    • 2021
  • Objectives : Gossypium arboreum (cotton) is traditionally used to treat various health disorders. However, anti-amnesic effect of G. arboreum has not been reported. The objective of this study was to investigate in-vivo the anti-amnesic effects along with in vitro antioxidant and acetylcholinesterase (AChE) inhibition potential in G. arboreum seed essential oil. Methods : The essential oil of G. arboreum obtained by solid phase microextraction (SPME) techniques were identified by gas chromatography-mass spectroscopy (GC-MS). 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) assay were performed to determine the antioxidant activity at various concentrations (312.5, 625, 1250, 2500, 5000, 10000 ㎍/㎖. Y-maze, passive avoidance and Morris water maze tests were carried out to evaluate improved effect on scopolamine (1 mg/kg)-induced memory dysfunction at the dose level of 50, 100 and 200 mg/kg. Donepezil (5 mg/kg) was used as a positive drug control. We performed acetylcholinesterase (AChE) activity assay in ex vivo. Results : Five volatile compounds were identified in G. arboreum. The assays of DPPH and ABTS revealed that G. arboreum increased antioxidant activity in a dose-dependent manner. G. arboreum ameliorated the percent of spontaneous alternation in the Y-maze test, shortened step-through latency in the passive avoidance test, and increased swimming time in the target zone in the Morris water maze test. In addition, G. arboreum inhibited the AChE activity. Conclusions : Based on these findings, G. arboreum may aid in the prevention and treatment of learning and memory-deficit disorders through antioxidant and AChE inhibitory activities.

Acridine Orange Stained Micronucleus Assay in Human B and T-lymphocytes after Low Dose ${\gamma}-irradiation$ (아크리딘 오렌지 형광염색법을 이용한 저선량 감마선 유도 말초혈액 B와 T-림프구 미소핵 분석)

  • Choi, Jeong-Mi;Kim, Hee-Sun;Yang, Kwang-Hee;Kim, Cha-Soon;Lim, Yong-Khi;Kim, Chong-Soon;Woon, Jae-Ho
    • Journal of Radiation Protection and Research
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    • v.29 no.1
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    • pp.9-15
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    • 2004
  • Firstly, we compared the two staining techniques, Giemsa and Acridine orange, to determine micronuclei on samples of cultures of five healthy human peripheral blood lymphocytes after ${\gamma}-irradiation\;(^{137}Cs)$ in dose ranges of 0 to 800cGy. It was found that the Acridine orange staining method gives more reliable results than the usual Giemsa staining method in micronucleus tests. Moreover, the frequency of micronuclei in cytokinesis-blocked human B-lymphocytes was studied after in vitro irradiation in dose ranges of 0 to 50cGy. After setting and separating the B-lymphocytes, the frequency of radiation-induced micronuclei were observed as the end-point markers for the low-dose radiation dosimetry after staining with Giemsa and Acridine orange dyes. The micronuclei frequency in B-lymphocytes was significantly elevated from 10 to 30cGy ${\gamma}-irradiation$. The determination of micronuclei in B-lymphocytes after staining with Acridine orange was higher than that of Giemsa. The frequency of micronuclei in B-lymphocytes was observed to be at least two times higher than those of T-lymphocytes Giemsa in dose increasing. Therefore, the determination of low-dose radiation-induced micronuclei in B-lymphocytes after staining with Acridine orange is likely to have the greatest potential in the estimation of low dose radiation exposure.

Effect of Enamel Matrix Drivatives application on the expression of PDLs17, PDLs22 of cultured human periodontal ligament cells in vitro (Enamel Matrix Derivatives가 사람 치주인대 세포의 특이유전자인 PDLs17, PDLs22의 발현에 끼치는 효과)

  • Han, Geun-A;Jang, Hyun-seon;Kok, Jung-Ki;Park, Ju-Chol;Kim, Heoung-Jung;Kim, Jung-Gwan;Kim, Byung-Ock
    • Journal of Periodontal and Implant Science
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    • v.34 no.2
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    • pp.333-344
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    • 2004
  • The enamel matrix derivative (EMD) has been recently used in the periodontal regenerative techniques. The present study was established to investigate the influence of EMD on human periodontal ligament cells using expression of mRNA of periodontal ligament specific gene (PDLs)17, PDLs22, type I collagen when EMD applied to periodontal ligament cells. Periodontal ligament cells were obtained from a healthy periodontium and cultured in Dulbecco's modified Eagle's medium (DMEM) plus 10% fetal bovine serum and ${\beta}-glycerophosphate$ with ascorbic acid. Test groups were two; One adds EMD in culture media and another added EMD and Dexamethasone (DEX) in culture media. Positive control group added DEX in culture media, and negative control group adds niether of EMD nor DEX. $Emdogain^{(R)}$ (Biora, Sweden, 30 mg/ml) was diluted by 75 ${\mu}g/ml$ concentration to culture media. For reverse transcription-polymerase chain reaction (RT-PCR), total RNA isolated on days 0, 7, 14 and 21. mRNA of PDLs17 was expressed on days 14 and 21 in EMD or DEX group, and expressed on days 7, 14 and 21 in EMD plus DEX group, the other side, expressed on days 21 in negative control group. mRNA of PDLs22 expressed on days 7, 14 and 21 in EMD group, and expressed on days 14 and 21 in DEX group, and expressed on days 7, 14 and 21 in EMD plus DEX group. Negative control group expressed on days 14 and 21. Type I collagen was expressed on all days and all groups. These results indicate that EMD promotes differentiation of periodontal ligament cells, and this is considered to offer basis that can apply EMD to periodontal tissue regeneration technique.

Development of Techniques for Evaluating the Virus Removal Rate using Adenovirus (아데노바이러스를 이용한 바이러스 제거율 평가를 위한 기법 개발)

  • Cho, Yoonjung;Lim, Jaewon;Baek, Dawoon;Lee, Sang-Hoon;Lee, In-Soo;Lee, Hyeyoung;Park, Donghee;Jung, Dongju;Kim, Tae Ue
    • Journal of Korean Society of Water and Wastewater
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    • v.29 no.6
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    • pp.633-641
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    • 2015
  • Waterborne infectious disease is induced by several pathogenic microbes such as bacteria, viruses and protozoans, and the cases caused by viral infection is currently increasing. Water treatment process could reduce the number of virus in the water, but there were many difficulties to completely remove the virus particles from water. Therefore, the membrane separation technology which was reported to effectively remove pollutants from raw water has attracted increasing attention and demand. Since its efficiency has been introduced, demands for evaluation method toward the membrane filtration process are increasing. However, progression of the method development is slow due to the difficulties in cultivation of several waterborne viruses from animal models or cell culture system. To overcome the difficulties, we used adenovirus, one of the commonly isolated pathogenic waterborne viruses which can grow in cell culture system in vitro. The adenovirus used in this study was identified as human adenovirus C strain. The adenovirus was spiked in the raw water and passed through the microfiltration membrane produced by Econity, a Korean membrane company, and then the viral removal rate was evaluated by real-time PCR. In the results, the amount of virus in the filtered water was decreased approximately by 5 log scale. Because coagulant treatment has been known to reduce filtering function of the membrane by inducing fouling, we also investigated whether there was any interference of coagulant. In the results, we confirmed that coagulant treatment did not show significant interference on microfiltration membrane. In this study, we found that waterborne virus can be effectively removed by membrane filtration system. In particular, here we also suggest that real-time PCR method can rapidly, sensitively and quantitatively evaluate the removal rate of virus. These results may provide a standard method to qualifying membrane filtration processes.

Difference of Neuronal Recovery by Incubation Condition after Transient Hypoxia (배양조건에 의한 일과성 저산소상태 후 신경세포회복의 차이)

  • Moon, Soo-Hyeon;Oh, Jae-Inn;Park, Youn-Kwan;Chung, Heung-Sub;Lee, Hoon-Kap;Lee, Ki-Chan
    • Journal of Korean Neurosurgical Society
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    • v.29 no.9
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    • pp.1161-1170
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    • 2000
  • Objective : The transverse hippocampal slice is one of the most commonly studied in vitro models of mammalian brain physiology. However, despite its broad usage, there has been no standardization of slice preparation techniques or recording condition. It is well known that variations in recording conditions can result in profound different effects to neuronal responses. Evoked field potentials, recorded extracellularly, were used to investigate the effects of variations in hippocampal slice preparation protocol on hypoxia responses of CA1 neurones. Material & Methods : Before hypoxic injury, hippocampal slices were incubated for 4 hours. During incubation period, the slices were placed in a incubation chamber($21^{\circ}C$) for recovery from preparation injury and then transferred to recording chamber($34^{\circ}C$) for more recovery and baseline electric recording with current stimulation(0.1Hz). Various time periods in incubation chamber and recording chamber were applied to each experimental group(group 1=60min : 180min, group 2=90min : 150min, group 3=180min : 60min, time in incubation chamber : time in recording chamber) before 10 min hypoxia produced by replacing 95% $O_2$+5% $CO_2$ mixed gas to 95% $N_2$+5% $CO_2$ gas. Calcium, Magnesium ions and several drugs effecting on glutamate receptor also were studied. Recoveries from hypoxic injury of hippocampal slices were estimated by percent recovery of population spike(PS). Statistic analysis of study were performed using paired t-test. Results : The percent recovery of PS after 10min hypoxia was considerably enhanced by increasing the period of current stimulation during incubation period before hypoxic injury. Temperature effect on the result of this experiment was also studied(group 4) but the result from this showed no statistic significance. Low magnesium ion concentration of artificial CSF(Mg-free aCSF) during incubation period enhanced the recovery of PS but low calcium (calcium-free) and high magnesium ion concentration(2mM) reduced it after hypoxic injury. L-glutamate($100{\mu}M$) and AP-5($50{\mu}M$) had no effect on the recovery of PS but CNQX($10{\mu}M$) in artificial CSF during incubation period markedly enhanced the recovery of PS. Co-treatment of AP-5($50{\mu}M$), CNQX($10{\mu}M$) and high magnesium concentration(2mM) enhanced recovery of PS in immediate following period of hypoxic injury but the effect of cotreatment after then decayed rapidly and lost statistic significance. Conclusions : Judging from above results, the condition of baseline recording is important in observing the recovery of population spike after hypoxia, and the time and the condition should be controled more strictly to obtain reliable results.

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Effects of ascorbic acid, citric acid and silver nitrate on the growth of in vitro lily plantlets and reduction of browning (Ascorbic acid, citric acid 및 AgNO3가 나리 기내식물체 생장촉진 및 갈변화 감소에 미치는 영향)

  • Roh, Hee Sun;Lee, Sang Il;Kang, Yun Im;Kim, Mi Seon;Kim, Jong Bo
    • Journal of Plant Biotechnology
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    • v.40 no.4
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    • pp.224-230
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    • 2013
  • Lily is an important cut-flower in Korea and world as well due to it's a variety of flower colors and various sizes of flowers. To develop elite lily cultivars, conventional breeding techniques have been used so far. However, an introduction of tissue culture system in mass propagation of bulbs and regeneration of shoots with a high efficiency is prerequisite at this moment. Especially, growth of bulbs and shoots as well as reduction of browning is critical factor to proliferate bulbs with shoots of superior lines or cultivar in lily. For this purpose, we tried to test whether ascorbic acid, citric acid and silver nitrate in medium to facilitate growth of bulbs and shoots as well as reduction of browning of bulb scales in lily. As a result, ascorbic acid and silver nitrate showed no significant differences compared to control plants in the growth of bulbs and shoots. When bulb scales were treated with $150mg{\cdot}l^{-1}$ of citric acid, formations of shoots and bulbs showed best result. While, bulb scale treated with $100mg{\cdot}l^{-1}$ of citric acid showed the growth of shoot and root as well as increasing of fresh weight compared to other treatments. Regarding the reduction of browning, $150mg{\cdot}l^{-1}$ of ascorbic acid showed the best result with the less than 2%. Although more experiments with several commercial varieties are needed in the future to establish mass propagation of bulbs of lily, results obtained in this study are supposed to provide the basic knowledge and contribution in tissue culture system of lily.

Effects of Compositions of Saponin Fraction from Korean Red Ginseng in the Relaxation of Rabbit and Rat Corpus Cavernosum (토끼와 흰쥐 음경해면체 이완작용에 미치는 홍삼사포닌 분획별 효과)

  • Choi Young Deuk;Park Jin Ah;Choi Hyung Ki;Nam Ki Yeul
    • Journal of Ginseng Research
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    • v.23 no.1 s.53
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    • pp.13-20
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    • 1999
  • We previously reported that Korean red ginseng (KRG) has a relaxation effect on the smooth muscles of corpus cavernosum via nitric oxide (NO) pathway and calcium and potassium channels. However, it is suggested that the active ingredients of KRG might be different depending on the sources of preparation, and there might be differences in actions for different compositions. We first investigated the composition of KRG saponins according to the extractions of the various sources of KRG, then with these extractions the relaxation effects were evaluated in vitro and hemodynamical in vivo using New Zealand white rabbit and rat corpus cavernosum. The total compositions of ginsenoside $(G-Rb_1,\;-Rb_2,\;-Rc,\;-Rd,\;G-Re,\;-Rf,\;-Rg_1)$ in fractionated KRG saponin designated as TS-1, TS-2, TS-3 were $41\%,\;40\%,\;and\;62\%,$ respectively, and the ratios of PD saponin and PT saponin (PD/PT) were 1,55, 1.72, 2.25, and 2.61, the values of which were statistically significant. In vitro studies using the rabbit corpus cavernosal muscle strips, the KRG saponin relaxed cavernosal strips in a dose-dependent manner, and same results were observed in in vivo studies, that KRG saponin increased the intracavernosal pressure in the rat. There was difference in the efficacy according to fractionation techniques. The differences in the total contents of ginsenosides did not affect relaxation, rather PT saponin content was statistically related to the degree of cavernosal relaxation, and this action presumed to be mediated by NO pathway and calcium and potassium channels. In conclusion, KRG exerts relaxation which is a key step in erection via combination of effects on NO system or calcium and potassium channels. The efficacy of this action is different to the sources of ginseng, which is affected by the different composition of ginsenosides $(G-Rb_1,\;-Rb_2,\;-Rc,\;-Rd,\;G-Re,\;-Rf,\;-Rg_1).$ Thus the further studies on the active ingredients such as minor ginsenosides and non-saponin components of red ginseng with maximum potency should be sought.

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Mechanical evaluation of the use of conventional and locking miniplate/screw systems used in sagittal split ramus osteotomy

  • Santos, Zarina Tatia Barbosa Vieira;Goulart, Douglas Rangel;Sigua-Rodriguez, Eder Alberto;Pozzer, Leandro;Olate, Sergio;Albergaria-Barbosa, Jose Ricardo
    • Journal of the Korean Association of Oral and Maxillofacial Surgeons
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    • v.43 no.2
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    • pp.77-82
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    • 2017
  • Objectives: The aim of this study was to compare the mechanical resistance of four different osteosyntheses modeled in two different sagittal split ramus osteotomy (SSRO) designs and to determine the linear loading in a universal testing machine. Materials and Methods: An in vitro experiment was conducted with 40 polyurethane hemimandibles. The samples were divided into two groups based on osteotomy design; Group I, right angles between osteotomies and Group II, no right angles between osteotomies. In each group, the hemimandibles were distributed into four subgroups according to the osteosynthesis method, using one 4-hole 2.0 mm conventional or locking plate, with or without one bicortical screw with a length of 12.0 mm (hybrid technique). Each subgroup contained five samples and was subjected to a linear loading test in a universal testing machine. Results: The peak load and peak displacement were compared for statistical significance using PASW Statistics 18.0 (IBM Co., USA). In general, there was no difference between the peak load and peak displacement related to osteotomy design. However, when the subgroups were compared, the osteotomy without right angles offered higher mechanical resistance when one conventional or locking 2.0 mm plate was used. One locking plate with one bicortical screw showed higher mechanical resistance ($162.72{\pm}42.55N$), and these results were statistically significantly compared to one conventional plate with monocortical screws (P=0.016) and one locking plate with monocortical screws (P=0.012). The difference in peak displacement was not statistically significant based on osteotomy design or internal fixation system configuration. Conclusion: The placement of one bicortical screw in the distal region promoted better stabilization of SSRO. The osteotomy design did not influence the mechanical behavior of SSRO when the hybrid technique was applied.

Autoradiographic Localization of Atdal Natriuretic Peptide Binding Sites in the Pig Ovary (돼지 난소 Atrial Natriuretic Peptide 결합 부위의 자가방사법에 의한 검증)

  • 김성주;김선희
    • The Korean Journal of Zoology
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    • v.38 no.4
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    • pp.523-530
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    • 1995
  • Specific affinity binding sites for atrial natriuretic peptide (ANP) were Investigated in the pig ovarian tissues by in vitro autoradiographic techniques. In the pig ovary, the highest binding sites for 12514abeiled rANP(l~28) were localized in the granulosa cell layer of the forncles. The binding sies on theca layer of the ovarian follicles were mainly localized in the external layer, but none was observed In the Internal layer. In the corpus luteum, the binding site was not observed. The specific bindings of 200 pM of l2Sl4abelled rANP(l~28) to granulosa and theca externa layers were reversed completely by excess concentration (1 ~4) of unlabelled rANP(l~28) but not by 10 ~ of unrelated peptides, human angiotensin II and arginine vasopressin. The binding was also displaced by 1 ~ of desiGIn18, Ser19, Gly2O, leu21, Gly22I ANP(4~2g) (C- ANF) as a spedfic ligand of the ANP clearance receptor. Therefore these results indicate ~hat the biological and the clearance ANP receptors exist in the theca externa and granulosa layer of the pig ovary, and suggest that the ANP receptors may be related with the regulatory lundion of the ovarian follicular development including oocyte maturation.

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