• 한국산학기술학회논문지
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• 제20권8호
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• pp.477-484
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• 2019

포유류에 존재하는 Collectin-Placenta 1 (CL-P1)을 포함한 여러 종류의 scavenger 수용체는 주로 내피 세포, 대식 세포 및 평활근 세포 표면에 발현되는 분자이다. 이들 분자는 산화 된 저밀도 지질 단백질 (oxLDL)에 결합하여 처리 할 수 있는 세포 표면 당 단백질이다. 이들 분자 중 케르세틴이 CL-P1 활성화에 어떤 영향을 미치는가를 확인하였다. 케르세틴은 산화 반응을 담당하는 자유 라디칼의 제거제 역할을 하여 산화를 중지시키는 항산화제로 알려져 있다. 본 논문에서는 마우스 CL-P1 유전자 promoter 부분의 전사 시작 점부터 -500 번째 염기까지의 단편을 DNA 중합효소를 이용하여 클로닝 하였다. 그 후에 대식세포 계열인 RAW264.7 및 섬유아세포계열의 NIH3T3 세포에 도입하여 케르세틴이 CL-P1 유전자 발현에 어떠한 영향을 미치는지에 대한 연구를 하였다. 이 부위에는 세포주기 조절 인자인 E2F 결합부위를 비롯해서 여러 종류의 전사 인자가 결합하는 염기서열이 다수 위치하고 있다. 이러한 500염기 단편을 pGL4.10 기본 벡터 및 프로모터에 연결시킨 후 세포에 도입시켰다. 그리고 배양 중에 케르세틴을 처리하여 유전자 발현양을 형광 색소 발현기법으로 측정하였다. 그 결과 유전자 발현이 시작되는 앞쪽 부분의 -250에서 -350사이의 염기들이 CL-P1 단백질을 만드는데 중요하다는 것을 확인하였다. 그 중에서도 E2F결합 부위가 결정적인 것 이라는 것을 DNA 돌연변이 실험을 통해 확인 하였다. 또한 부착 세포인 RAW264.7 배양액에 케르세틴을 첨가 한 결과, 배양용기 표면에서 탈락하는 현상을 확인하였다. 즉 발현된 CL-P1단백질이 케르세틴에 의해 세표 표면의 부착 분자에도 영향을 주는 것을 확인하였다.

• Journal of Nutrition and Health
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• 제57권2호
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• pp.196-210
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• 2024

본 연구에서는 정향과 마가목 복합물 (SS)의 항비만 효과를 알아보기 위해 실험을 진행하였다. SS 투여는 3T3-L1 세포 내 TG와 TC가 유의적으로 감소하는 효과를 나타냈으며, 지질 합성 관련 유전자와 지방산 산화 관련 유전자 발현을 조절하는 효과를 보여주었다. 비만이 유도된 C57BL/6 mice에서 SS 투여는 혈청 내 leptin 호르몬 수치를 감소시켰으며, AMPK/ACC/SREBP-1 경로를 경유하여 TG 합성을 억제하였다. 또한, 조직병리학적 분석을 통해 지질 축적과 지방세포의 크기가 감소된 것을 확인하였다. 따라서 SS는 비만의 예방과 치료를 위한 잠재력을 갖춘 소재로 사료된다.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 2007년도 Proceedings of The Convention
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• pp.79-92
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• 2007

Oxidative stress have known to be a risk factor for the degenerative processes and closely related to a lot of diseases. It is well established that antioxidants are good in protection and therapeutic means against oxidative damage. There is increasing interest in natural antioxidants and many natural antioxidants have been found and utilized as the possible protection for various diseases and skin aging. We have screened natural antioxidant agents for cosmeceuticals, nutraceuticals, and drugs as therapeutic and preventive means against oxidative stress, and have developed a number of novel antioxidants from various natural sources. A novel melanin synthesis inhibitor, Melanocin A, isolated from the metabolite of a fungal strain Eupenicillium shearii F80695 inhibited mushroom tyrosinase and melanin biosynthesis of B16 melanoma cells with $IC_{50}$ value of 9.0 nM and MIC value of $0.9\;{\mu}M$, respectively. Melanocin A also exhibited potent antioxidant activity by scavenging of DPPH and superoxide anion radicals. UV was found to increase the level of hydrogen peroxides and other reactive oxygen species (ROS) in skin tissues. This increase in ROS may not only alter the structure and function of many genes and proteins directly but may also modulate their expressions through signal transduction pathways and, ultimately, lead to skin damage. We investigated the effect of Melanocin A on UV-induced premature skin aging. Firstly, the effect of Melanocin A on UV-induced matrix metalloproteinase (MMP)-9 expression in an immortalized human keratinocyte cell line, HaCaT in vitro was investigated. Acute UV irradiation induced MMP-9 expression at both the mRNA and protein levels and Melanocin A suppressed this expression in a dose-dependent manner. We then investigated UV-induced skin changes in hairless mice in vivo by Melanocin A. Chronic exposure of hairless mouse dorsal skin to UV increased skin thickness and induced wrinkle formation and the gelatinase activities of MMP-2 and MMP-9. Moreover, Melanocin A significantly suppressed UV-induced morphologic skin changes and MMP-2 and MMP-9 expression. These results show that Melanocin A can prevent the harmful effects of UV that lead to skin aging. Therefore, we suggest that Melanocin A should be viewed as a potential therapeutic agent for preventing and/or treating premature skin aging. Terrein is a bioactive fungal metabolite isolated from Penicillium species. Terrein has a relatively simple structure and can be easily synthesized. However, the biologic effects of terrein are comparatively unknown. We found for the first time that terrein potently inhibit melanin production in melanocytes and has a strong hypopigmentary effect in a spontaneously immortalized mouse melanocyte cell line, Mel-Ab. Treatment of Mel-Ab cells with terrein (10-100 mM) for 4 days significantly reduced melanin levels in a dose-dependent manner. In addition, terrein at the same concentration also reduced tyrosinase activity. We then investigated whether terrein influences the extracellular signal-regulated protein kinase (ERK) pathway and the expression of microphthalmia-associated transcription factor (MITF), which is required for tyrosinase expression. Terrein was found to induce sustained ERK activation and MITF down-regulation, and luciferase assays showed that terrein inhibits MITF promoter activity in a dose-dependent manner. To elucidate the correlation between ERK pathway activation and a decreased MITF transcriptional level, PD98059, a specific inhibitor of the ERK pathway, was applied before terrain treatment and found to abrogate the terrein-induced MITF attenuation. Terrein also reduced the tyrosinase protein level for at least 72 h. These results suggest that terrain reduces melanin synthesis by reducing tyrosinase production via ERK activation, and that this is followed by MITF down-regulation.

• Archives of Pharmacal Research
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• 제25권6호
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• pp.910-916
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• 2002

Endotoxin tolerance reduces the capacity of monocytes to produce proinflammatory cytokines, results in cellular immune paralysis, and down-regulates the production of helper T (Th)1 type cytokines with a shift toward a Th2 cytokine response. Prostaglandin (PG)E$_2$ in the immune system also results in macrophage inactivation and the suppression of Th1 activation and the enhancement of Th2 activation. However, the inhibitory effects of PGE$_2$ on the altered polarization of the Th cell and macrophage interleukin (IL)-6 production characterized in part by cellular immune paralysis in a state of endotoxin tolerance is unclear. This study was undertaken, using indomethacin, to investigate the role of endogenous PGE$_2$ on the Th cytokines and macrophage IL-6 production in a state of endotoxin tolerance compared to those with endotoxemia mice, wherein, in this latter case, the increased production of proinflammatory cytokines and PGE$_2$ is exhibited. Endotoxemia was induced by injection of lipopolysaccharide (LPS; 10 mg/kg in saline) i.p. once in BALB/c mice, and endotoxin tolerance was induced by pretreatment with LPS (1 mg/kg in saline) injected i.p. daily for two consecutive days and then with LPS 10 mg/kg on day 4. Splenocytes or macrophages were obtained from endotoxemia and endotoxin tolerance models pretreated with indomethacin, and then cytokine production was induced by Con A-stimulated splenocytes for the Th cytokine assays and LPS-stimulated macrophages for the IL-6 assay. Our results showed that endotoxemia led to significantly reduced IL-2 and IL-4 production, to significantly increased IL-6 production, whereas interferon $(IFN)-{\gamma}$ production was not affected. Indomethacin in the case of endotoxemia markedly attenuated $IFN-{\gamma}$ and IL-6 production and didnt reverse IL-2 and IL-4 production. Endotoxin tolerance resulted in the significantly reduced production of IL-2 and $IFN-{\gamma}$ and the significantly increased production of IL-4 and IL-6. Indomethacin in endotoxin tolerance greatly augmented IL-2 production, significantly decreased IL-4 production, and slightly attenuated IL-6 production. These findings indicate that endogenous PGE$_2$ may mediate the suppressed Th1 type immune response, with a shift toward a Th2 cytokine response in a state of endotoxin tolerance, whereas endotoxemia may be regulated differentially. Also, endogenous PGE$_2$ may mediate macrophage IL-6 production in the case of endotoxemia to a greater extent than in the case of endotoxin tolerance.

• Journal of Animal Science and Technology
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• 제47권6호
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• pp.947-954
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• 2005

경구로 섭취된 생균제가 소화관 효소의 작용을 받은 경우 장관면역계의 macrophage에 미치는 영향을 조사하기 위하여 돼지의 장내에서 분리된 lactobacilli를 pepsin과 pancreatin과 같은 소화관 효소로 분해한 균체분해산물에 의해 RAW 264.7 murine macrophage에서 유도되는 NO, TNF-$\alpha$ 및 IL-6의 생성을 측정하였다. Macrophage에서 유도되는 NO, TNF-$\alpha$ 및 IL-6는 균종과 균체분해산물의 양에 따라 달랐다. NO의 생성수준은 10～150 ug/ml의 분해물에서 관찰 되었으나 TNF-$\alpha$와 IL-6는 50～300 ug/ml의 고농도처리에서 나타났다. 6개의 Lactobacillus strains 중에서 3149와 3156 균주는 다른 균주에 비해 TNF-$\alpha$와 IL-6 유도능이 높았다. 또한 소화관 효소에 의해 분해된 lactobacilli의 균체분해산물의 성분이 macrophage 활성 증가와 관련이 있으며 생체내에서 숙주면역계를 조절할 것으로 사료된다. 본 시험에서 사용한 방법은 소화관 면역계에 미치는 유산균의 효과를 규명하는데 유익할 것이다.

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2003년도 학술발표대회 발표논문초록집
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• pp.54-54
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• 2003

The activation of protooncogenes or the inactivation of their gene products may be a specific and effective functional study for human neoplasia. To examine this possibility, we have used the tetracycline regulatory system to generate transgenic mice that conditionally express the HccR-2 protooncogene in vivo. The new human cervical cancer protooncogene (HccR-2) was detected from cervical cancer cell line. To elucidate its biological functions, we generated transgenic mice that expressed the HccR-2 gene. The sustained expression of the HccR-2 transgene culminated chronic neutrophilic leukemia (CNL). CNL is a rare chronic myeloproliferative disorder that presents as a sustained, mature neutrophilic leukocytosis with few or no circulating immature granulocytes, the absence of peripheral blood monocytosis, basophilia, or eosinophilia, and infiltration of neutrophils at the liver, spleen and kidney. Mice expressing the HccR-2 and tetracycline-transactivating protein (tTa) transgene were found to have altered myeloid development that was characterized by increased percentages of mature neutrophil and band form neutrophil in the peripheral blood, liver and spleen. Activation of the transgene causes CNL. In our model, expression of HccR-2 transgene mice was similar in many respects to the human CNL. This model will be valuable not only for investigating the biological properties of the HccR-2 and other protooncogenes in vivo but also for analyzing the mechanism involved in the progression of CNL.

• The Korean Journal of Physiology and Pharmacology
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• 제21권3호
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• pp.309-316
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• 2017

Transient receptor potential vanilloid 3 (TRPV3) is a non-selective cation channel with modest permeability to calcium ions. It is involved in intracellular calcium signaling and is therefore important in processes such as thermal sensation, skin barrier formation, and wound healing. TRPV3 was initially proposed as a warm temperature sensor. It is activated by synthetic small-molecule chemicals and plant-derived natural compounds such as camphor and eugenol. Schisandra chinensis (Turcz.) Baill (SC) has diverse pharmacological properties including antiallergic, anti-inflammatory, and wound healing activities. It is extensively used as an oriental herbal medicine for the treatment of various diseases. In this study, we investigated whether SC fruit extracts and seed oil, as well as four compounds isolated from the fruit can activate the TRPV3 channel. By performing whole-cell patch clamp recording in HEK293T cells overexpressing TRPV3, we found that the methanolic extract of SC fruit has an agonistic effect on the TRPV3 channel. Furthermore, electrophysiological analysis revealed that ${\gamma}$-schisandrin, one of the isolated compounds, activated TRPV3 at a concentration of $30{\mu}M$. In addition, ${\gamma}$-schisandrin (${\sim}100{\mu}M$) increased cytoplasmic $Ca^{2+}$ concentrations by approximately 20% in response to TRPV3 activation. This is the first report to indicate that SC extract and ${\gamma}$-schisandrin can modulate the TRPV3 channel. This report also suggests a mechanism by which ${\gamma}$-schisandrin acts as a therapeutic agent against TRPV3-related diseases.

• 한국발생생물학회:학술대회논문집
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• 한국발생생물학회 2003년도 제3회 국제심포지움 및 학술대회
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• pp.83-83
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• 2003

The lutropin/choriogonadotropin receptor (LHR) is a member of the rhodopsin-like subfamily of G protein coupled receptor (GPCRs), that has been shown to mediate the internalization of its two naturally occurring agonist, lutropin and choriogonadotropin (CG). The clustered agonist-receptor complex is internalized by a dynamin-dependent pathway and traverses the endosomal compartment without agonist dissociation Dissociation of the agonist-receptor complex occurs in the lysosomes, where both the agonist and receptor are degrade. Recently, constitutively activating mutations of the receptor have been identified that are associated with familial male-precocious puberty (FMPP). A FMPP is a form of sexual precocious puberty in boys in which testosterone levels are elevated independent of changes in luteinizing hormone-releasing hormone and serum luteinizing hormone levels, We have now analyzed two naturally occurring, constitutively active mutants of the human LHR. These mutations were introduced into the rat LHR (rLHR) and are designated L435R and D556Y. Cells expressing rLHR-D556Y bind human choriogonadotropin (hCG) with normal affinity, exhibit a 25-fold increase in basal cAMP and respond to hCG with a normal increase in cAMP accumulation. Cells expressing rLHR-L435R also bind hCG with normal affinity, exhibit a 47-fold increase in basal cAMP, and do not respond to hCG with a further increase in cAMP accumulation. This mutation enhances the internalization of the free and agonist-occupied receptors ~2- and ~17- fold, respectively We conclude that the state of activation of the rLHR can modulate its basal and/or agonist-stimulated internalization. Since the internalization of hCG is involved in the termination of hCG actions, we suggest that the lack of responsiveness detected in cells expressing rLHR-L435R is due to the fast rate of internalization of the bound hCG. The finding that membranes expressing rLHR-L435R respond to hCG with an increase in adenylyl cyclase activity supports this suggestion. Autonomous Leydig cell activity in FMPP is caused by a constitutively activating LH/CGR.

• 한국식품저장유통학회지
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• 제30권3호
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• pp.483-491
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• 2023

본 연구에서는 제주도에서 자생한 병풀을 수집해 스마트팜과 노지에서 재배하고 이를 이용하여 주요성분 및 각질형성세포 활성화에 미치는 영향을 확인 및 비교하였다. 스마트팜 재배 병풀과 노지 재배 병풀의 유전자 확인을 통한 종 분석을 위해, 핵 속의 ITS DNA와 엽록체의 psbA-H DNA를 증폭하여 염기서열을 분석한 후 NCBI 유전자 은행에서 보고된 식물들의 DNA와 비교하였다. 스마트팜 재배 병풀과 노지 재배 병풀의 ITS DNA 염기서열은 유전자 은행의 MH768338.1번 Centella asiatica와 일치하고 엽록체 psbA-H DNA 또한 유전자 은행의 JQ425422.1번 C. asiatica와 일치하였다. 스마트팜 재배 병풀추출물(SEE)과 노지 재배 병풀추출물(FEE)의 triterpene은 HPLC에 의해 분석되었으며, SEE의 madecassoside, asiaticoside, madecassic acid, asiatic acid 함량은 각각 59.31±0.94 mg/g, 46.38±2.26 mg/g, 6.21±1.47 mg/g, 7.04±1.93 mg/g으로 분석되었다. 반면, FEE는 각각 24.38±1.31 mg/g, 21.28±1.44 mg/g, 3.11±1.05 mg/g, 5.40±1.26 mg/g으로 측정되어 SEE가 FEE보다 더 높은 triterpene을 갖는 것이 확인되었다. 사람 각질형성세포에 대한 SEE와 FEE의 독성은 실험된 농도 내에서 관찰되지 않았으며, 스크래치가 유발된 세포 내 회복은 SEE가 FEE보다 더 높은 회복능을 보였다. 따라서, 본 실험 결과 triterpene 함량이 더 높은 스마트팜 재배 병풀이 건강기능식품 소재로서 더 효과적이라고 판단된다.

• International Journal of Oncology
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• 제55권1호
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• pp.203-210
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• 2019

To overcome the poor prognosis of patients with ovarian cancer, attempting to target ovarian cancer with effective antitumor compounds has been conducted for numerous years. Although the 3,4-dihydroquinazoline derivative KYS05090S was known to exert antitumor effects in A549 and ovarian cancer cells by inhibition of T-type Ca²⁺ channels, the complete underlying antitumor mechanism of this compound remains unclear. Thus, in the present study, the potential apoptotic mechanism of KYS05090S was elucidated in SKOV3 and OVCAR3 ovarian cancer cells. KYS05090S exerted significant cytotoxicity in SKOV3 and OVCAR3 ovarian cancer cells, and also increased the number of apoptotic bodies, and the number of terminal deoxynucleotidyl transferase dUTP nick end labeling positive cells and the sub-G1 population as a feature of apoptosis. Consistently, KYS05090S induced cleavage of poly(ADP-ribose) polymerase and caspase-9/3 in ovarian cancer cells. Notably, KYS05090S attenuated the expression of anti-apoptotic proteins, including cyclin D1 and B-cell lymphoma-2 (Bcl-2), and reduced the phosphorylation of Janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) in ovarian cancer cells. Additionally, KYS05090S blocked the nuclear translocation of STAT3 and suppressed the signaling of JAK2/STAT3 in interleukin-6-treated SKOV3 cells, as a STAT3 activator. Overall, these observations indicated that inhibition of JAK2/STAT3 signaling and activation of caspase-9/3 are critically involved in the effects of KYS05090S on apoptosis in ovarian cancer types, and the compound may be beneficial as a potent antitumor agent.