• International Journal of Oral Biology
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• 제45권1호
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• pp.15-24
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• 2020

The fruit of Chaenomeles sinensis (Thouin) Koehne (Chaenomelis Fructus) known as "Mo-Gua" in Korea has been commonly used in traditional medicine to treat inflammatory diseases, such as sore throat. However, its effect on bone metabolism has not been elucidated yet. Here, we examined the effect of Chaenomelis Fructus ethanol extract (CF-E) on receptor activator of nuclear factor (NF)-κB ligand (RANKL)-mediated osteoclast differentiation and formation. CF-E considerably inhibited osteoclast differentiation and tartrate-resistant acid phosphatase-positive multinuclear cell formation from bone marrow-derived macrophages and osteoclast precursor cells in a dose-dependent manner. In addition, the formation of actin rings and resorption pits were significantly suppressed in CF-E-treated osteoclasts as compared with the findings in non-treated control cells. Consistent with these phenotypic inhibitory results, the expressions of osteoclast differentiation marker genes (Acp5, Atp6v0d2, Oscar, CtsK, and Tm7sf4) and Nfatc1, a pivotal transcription factor for osteoclastogenesis, were markedly decreased by CF-E treatment. The inhibitory effect of CF-E on RANKL-induced osteoclastogenesis was associated with the suppression of NFATc1 expression, not by regulation of mitogen-activated protein kinases and NF-κB activation but by the inactivation of phospholipase C gamma 1 and 2. These results indicate that CF-E has an inhibitory effect on osteoclast differentiation and formation, and they suggest the possibility of CF-E as a traditional therapeutic agent against bone-resorptive diseases, such as osteoporosis, rheumatoid arthritis, and periodontitis.

• Archives of Pharmacal Research
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• 제24권2호
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• pp.126-135
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• 2001

Quinacrine (QU), a phospholipase-A2 (PLA-2) inhibitor has been used clinically as a chemotherapeutic adjuvant. To understand the mechanisms leading to its chemotherapeutic effect, we have investigated QU-induced apoptotic signaling pathways in human cervical squamous carcinoma HeLa cells. In this study, we found that QU induced cytochrome c-dependent apoptotic signaling. The release of pro-apoptotic cytochrome c was QU concentration- and time-dependent, and preceded activation of caspase-9 and -3. Flow cytometric FACScan analysis using fluorescence intensities of $DiOC_6$/ demonstrated that QU-induced cytochrome c release was independent of mitochondrial permeability transition (MPT), since the concentrations of QU that induced cytochrome c release did not alter mitochondrial membrane potential (${\blacktriangle}{\Psi}_m$). Moreover, kinetic analysis of caspase activities showed that cytochrome c release led to the activation of caspase-9 and downstream death effector caspase-3, Caspase-3 inhibitor (Ac-DEVD-CHO) partially blocked QU-induced apoptosis, suggesting the importance of caspase-3 in this apoptotic signaling mechanism. Supplementation with arachidonic acid (AA) sustained caspase-3 activation induced by QU. Using inhibitors against cellular arachidonate metabolism of lipooxygenase (Nordihydroxyguaiaretic Acid, NDGA) and cyclooxygenase (5,8,11,14-Eicosatetraynoic Acid, ETYA) demonstrated that QU-induced apoptotic signaling may be dependent on its role as a PLA-2 inhibitor. Interestingly, NDCA attenuated QU-induced cytochrome c release, caspase activity as well as apoptotic cell death. The blockade of cytochrome c release by NDCA was much more effective than that attained with cyclosporin A (CsA), a MPT inhibitor. ETYA was not effective in blocking cytochrome c release, except under very high concentrations. Caspase inhibitor z-VAD blocked the release of cytochrome c suggesting that this signaling event is caspase dependent, and caspase-8 activation may be upstream of the mitochondrial events. In summary, we report that QU induced cytochrome c-dependent apoptotic signaling cascade, which may be dependent on its role as a PLA-2 inhibitor. This apoptotic mechanism induced by QU may contribute to its known chemotherapeutic effects.

• Journal of Ginseng Research
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• 제24권4호
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• pp.168-175
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• 2000

Xenopus oocytes를 이용하여 인삼의 유효 성분으로 알려진 Ginseng total saponin(GTS)의 신호 전달 기작을 two electrode voltage clamp 방법을 이용하여 연구하였다. GTS는 세포 바깥에 처리했을 때 -2OmV보다 더 positive한 voltage에서 커다란 outward current를 유도하였다. 그러나, 세포 안쪽에 GTS를 injection할 경우 아무런 효과가 없는 것으로 나타났다. GTS처리에 의한 outward current유발 효과는 GTS 투여 용량에 의존적인 것으로 나타났다(EC$_{50}$ : 4.4 $\mu\textrm{g}$/ml). GTS의 작용은 $Ca^{2+}$-activated Cl- channel blocker인 niflumic acid에 의하여 차단되었다. 칼슘 chelator인 BAPIA와 IP$_3$ 수용체 길항제인 heparin을 세포내 injection에 의하여 차단되었다. 또한 active phospholipase C inhibitor(PLC)인U-73122를 세포 바깥에 전처리할 경우에도 GTS의 작용이 부분적으로 억제되는 것으로 나타났다. 백일해 독소를 전처리할 경우GTS의 작용은 억제되지 않은 것으로 나타났으나, GTP analog인 GTP${\gamma}$S를 세포내 injection할 경우 GTS의 작용은 억제되는 것으로 나타났다. 이러한 연구 결과는 GTS가 oocytes세포막 성분과 상호 작용에 의하여 $Ca^{2+}$-activated Cl- channel이 열리도록 하고, 이 과정에 PLC활성 및 백일해 독소에 민감하지 않은 G단백질활성 및 IP3에 민감한 세포내 $Ca^{2+}$-activated로부터 칼슘 방출을 유도하는 것으로 나타났다났다

• 한국식품과학회지
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• 제50권6호
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• pp.671-679
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• 2018

퉁퉁마디는 식용 염생식물로서 이에 대한 유용 기능성 탐색 및 생리활성 평가에 대한 연구가 지속적으로 진행되고 있다. 본 연구에서는 퉁퉁마디 70% 메탄올 추출물을 극성에 따라 분획하여 얻은 Fr.H, Fr.E, Fr.EA, Fr.B 및 Fr.W의 5분획의 NO제거 및 iNOS 발현, 아라키돈산 대사에 관련된 효소 $cPLA_2$, COX-2, 5-LOX 등의 발현과 활성화에 대한 효과를 분석하여 항염작용과 메커니즘을 검토하였다. 이 중 Fr.EA가 가장 높은 폴리페놀 및 플라보노이드 성분 함량을 나타내었고 nitrite/아질산의 제거에 있어서도 가장 뛰어난 활성을 보였다. 하지만 LPS로 자극한 RAW264.7 큰포식세포 배양계에서는 Fr.E가 가장 우수한 NO 제거활성과 iNOS발현억제 활성을 나타내었고 이어 Fr.H와 Fr.EA의 활성 순을 나타냈다. Fr.E는 또한 LPS로 자극한 RAW264.7 배양 세포계에서 $cPLA_2$와 COX-2의 발현억제에 가장 큰 효과를 나타내었으며 이들의 억제를 위한 $IC_{50}$는 각각 33.4과 $27.9{\mu}g/mL$로 나타났다. Fr.E는 $cPLA_2$의 활성화에 중요한 영향을 미치는 ERK1/2의 인산화도 농도의존적으로 저해하였다. Fr.H와 Fr.EA도 $80{\mu}g/mL$ 이하 농도에서 iNOS와 아라키돈산 대사효소들의 발현을 농도의존적으로 억제하였다. 하지만 가장 극성 분획인 Fr.W는 모든 활성평가 시스템에서 거의 효과를 나타내지 않았다. 본 연구는 퉁퉁마디가 iNOS와 아라키돈산 대사효소계를 효과적으로 억제하여 항염증작용을 할 수 있다는 것을 보고하고 있으며, 특히 Fr.E와 Fr.H와 같은 소수성 분획들이 우수한 활성을 나타내어 향후 이를 타깃으로 하는 관련 기능성 소재로서의 활용이 기대된다.

• Asian-Australasian Journal of Animal Sciences
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• 제30권4호
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• pp.462-469
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• 2017

Objective: The aim of this study is to identify genomic regions or genes controlling growth traits in pigs. Methods: Using a panel of 54,148 single nucleotide polymorphisms (SNPs), we performed a genome-wide Association (GWA) study in 562 pure Yorshire pigs with four growth traits: average daily gain from 30 kg to 100 kg or 115 kg, and days to 100 kg or 115 kg. Fixed and random model Circulating Probability Unification method was used to identify the associations between 54,148 SNPs and these four traits. SNP annotations were performed through the Sus scrofa data set from Ensembl. Bioinformatics analysis, including gene ontology analysis, pathway analysis and network analysis, was used to identify the candidate genes. Results: We detected 6 significant and 12 suggestive SNPs, and identified 9 candidate genes in close proximity to them (suppressor of glucose by autophagy [SOGA1], R-Spondin 2 [RSPO2], mitogen activated protein kinase kinase 6 [MAP2K6], phospholipase C beta 1 [PLCB1], rho GTPASE activating protein 24 [ARHGAP24], cytoplasmic polyadenylation element binding protein 4 [CPEB4], GLI family zinc finger 2 [GLI2], neuronal tyrosine-phosphorylated phosphoinositide-3-kinase adaptor 2 [NYAP2], and zinc finger protein multitype 2 [ZFPM2]). Gene ontology analysis and literature mining indicated that the candidate genes are involved in bone, muscle, fat, and lung development. Pathway analysis revealed that PLCB1 and MAP2K6 participate in the gonadotropin signaling pathway and suggests that these two genes contribute to growth at the onset of puberty. Conclusion: Our results provide new clues for understanding the genetic mechanisms underlying growth traits, and may help improve these traits in future breeding programs.

• The Plant Pathology Journal
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• 제38권1호
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• pp.12-24
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• 2022

In this study, we conducted whole-genome sequencing with six species of Pectobacterium composed of seven strains, JR1.1, BP201601.1, JK2.1, HNP201719, MYP201603, PZ1, and HC, for the analysis of pathogenic factors associated with the genome of Pectobacterium. The genome sizes ranged from 4,724,337 bp to 5,208,618 bp, with the GC content ranging from 50.4% to 52.3%. The average nucleotide identity was 98% among the two Pectobacterium species and ranged from 88% to 96% among the remaining six species. A similar distribution was observed in the carbohydrate-active enzymes (CAZymes) class and extracellular plant cell wall degrading enzymes (PCWDEs). HC showed the highest number of enzymes in CAZymes and the lowest number in the extracellular PCWDEs. Six strains showed four subsets, and HC demonstrated three subsets, except hasDEF, in type I secretion system, while the type II secretion system of the seven strains was conserved. Components of human pathogens, such as Salmonella pathogenicity island 1 type type III secretion system (T3SS) and effectors, were identified in PZ1; T3SSa was not identified in HC. Two putative effectors, including hrpK, were identified in seven strains along with dspEF. We also identified 13 structural genes, six regulator genes, and five accessory genes in the type VI secretion system (T6SS) gene cluster of six Pectobacterium species, along with the loss of T6SS in PZ1. HC had two subsets, and JK2.1 had three subsets of T6SS. With the GxSxG motif, the phospholipase A gene did locate among tssID and duf4123 genes in the T6SSa cluster of all strains. Important domains were identified in the vgrG/paar islands, including duf4123, duf2235, vrr-nuc, and duf3396.

• 한국잠사곤충학회지
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• 제52권1호
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• pp.6-9
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• 2014

서양종 꿀벌로부터 봉독채집장치를 이용하여 채집한 정제봉독의 pH 안정성에 대하여 평가하고자 멜리틴의 함량, 단백질의변화, 항균력, 세포증식률을 측정하였다. 정제봉독을 pH2에서 pH9까지 24시간 동안 처리한 후 멜리틴 함량과 단백질의 변화, S. aureus, P. acnes, S. mutans와 E. coli에 대한 항균력, 피부세포인 HDF에 대한 세포증식률에 미치는 영향을 분석하였다. pH2에서부터 pH9까지 각각 처리한 정제봉독의 멜리틴 및 단백질 성분의 변화는 확인되지 않았다. 봉독의 주요 약리효과인 항균력에 있어서도 시험균주마다 다소 차이가 있었으나 S. aureus, P. acnes, S. mutans와 E. coli 모두 산도에 따른 유의할 만한 변화는 확인되지 않았다. 또한 피부세포 증식률에 있어서도 산도에 따른 변화는 확인되지 않았다. 이러한 결과로 미루어 봉독의 pH처리는 멜리틴 함량, 단백질의 변화, 항균력 및 세포증식률에 영향을 주지 않는 것으로 확인되어 봉독은 산도의 변화에 있어 안정성이 매우 높은 것으로 사료된다.

• 대한약리학회지
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• 제32권2호
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• pp.243-257
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• 1996

CCK and cholinergic agonist stimulate enzyme release from the pancreatic acini via G-protein-mediated activation of phospholipase C, In contrast secretin and related peptides increase the level of cAMP and activate cAMP-dependent protein kinase. Camostat, a synthetic protease inhibitor, causes pancreatic hypertrophy and hyperplasia by increasing the CCK release. In this study, the secretagogue-induced changes of intracellular proteins were examined in the dispersed pancreatic acini of rats with or without camostat treatment. Camostat(FOY-305, 200 mg/kg, p.o.) was given for 4 days twice daily and the dispersed acini were prepared at 12 bouts after last treatment. The profiles of Intracellular phosphoproteins were analyzed by two-dimensional gel electrophoresis after incubating the acini with $^{32}P$. The amylase release from the dispersed acini was measured. The pancreatic weight was increased to 126% of control, while amylase activity per mg acinar protein decreased to 41% of control, The maximum response of amylase release from dispersed acini to CCK-8 or carbachol was markedly decreased(65% or 46% of control, respectively). The group of intracellular proteins(24 kD, pI $4.5{\sim}8.5$) was increased in quantity by camostat. CCK-8 or secretin increased phosphorylation of a protein(34 kD, pI 4.7) in camostat-treated as well as control rats. CCK-8 increased tyrosine phosphoryiation in the acini of control rats. However, in camostat-treated rats, the basal level of tyrosine phosphorylation was increased and it was rather decreased by CCK-8. Secretin had no effect on the level of tyrosine phosphorylation in acini. These results indicate that both phospholipase C and adenylate cyclase induce phosphorylation of an intracellular acinar protein(34 kD, pI 4.7) and camostat treatment increases the basal level of tyrosine phosphorylation in acinar cells. And these results suggest that not only serine/threonine protein kinase but also protein tyrosine kinase/phosphatase are involved in the process of CCK receptor mediated stimulation-secrelion coupling.

• KSBB Journal
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• 제10권3호
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• pp.343-348
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• 1995

LeI은 스테로이드를 통울에 투여하였을 때 분비가 촉진되어 항염증성 효과를 나타내는 calcium 의 존성 phospholipid 결합 단백질이다. S. cerevisiae는 대장균과 통물세포의 장점을 모두 가지고 있으므로 동물세포 유래의 이종 단백질의 분비 생산에 많이 이용되고 있다. 본 연구에서는 GAL10 promoter­p ppL-LCI유전자 LCI terminator로 구성된 pYGLPT5 로 LCI을 S. cerevisiae SEY2102에서 발현 분비시키고 각 분획으로 나누어 LCI양을 비교한 결과 protoplast 68.6 %. periplasmic 24 %, culture supernatant 7.4%로 분포하였다. pYGLPT5로 형질전환된 S. cereviswe 2102를 유가 배양한 결과, 최종적인 LCI의 생산량은 약 $500mg/\ell$ 였다. LCI은 N 말단 부근에 $CA^{++}$ 결합부위가 있으므로 이를 이용하여 hydroxylapatite column chromatography로 정 제하 였다. 배지로 분비된 34kDa LCI을 ultrafiltration 과 hydroxylapatite column chromatography 의 두 단계로 순도 99% 이상으로 정제할 수 있었다.

• Pediatric Gastroenterology, Hepatology & Nutrition
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• 제24권5호
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• pp.455-469
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• 2021

Purpose: The rs641738 C>T in membrane-bound O-acyltransferase domain-containing protein 7 (MBOAT7) is implicated, along with the rs738409 C>G polymorphism in patatin-like phospholipase domain-containing protein 3 (PNPLA3), in nonalcoholic fatty liver disease (NAFLD). The association of these polymorphisms and NAFLD are investigated in Hispanic children with obesity. Methods: Obese children with and without NAFLD were enrolled at a pediatric tertiary care health system and genotyped for MBOAT7 rs641738 C>T and PNPLA3 rs738409 C>G. NAFLD was characterized by the ultrasonographic presence of hepatic steatosis along with persistently elevated liver enzymes. Genetic variants and demographic and biochemical data were analyzed for the effects on NAFLD. Results: Among 126 enrolled subjects, 84 in the case group had NAFLD and 42 in the control group did not. The two groups had similar demographic distribution. NAFLD was associated with abnormal liver enzymes and elevated triglycerides and cholesterol (p<0.05). Children with NAFLD had higher percentage of PNPLA3 GG genotype at 70.2% versus 31.0% in non-NAFLD, and lower MBOAT7 TT genotype at 4.8% versus 16.7% in non-NAFLD (p<0.05). PNPLA3 rs738409 C>G had an additive effect in NAFLD; however, MBOAT7 rs641738 C>T had no effects alone or synergistically with PNPLA3 polymorphism. NAFLD risk increased 3.7-fold in subjects carrying PNPLA3 GG genotype and decreased in MBOAT7 TT genotype. Conclusion: In Hispanic children with obesity, PNPLA3 rs738409 C>G polymorphism increased the risk for NAFLD. The role of MBOAT7 rs641738 variant in NAFLD is less evident.