• Toxicological Research
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• 제11권2호
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• pp.175-180
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• 1995

We found that bufalln, one of the prominent components of the bufadlenolides in the Chinese medicine chan'su, has the potent inhibitory effects on growth and proliferation of the cultured bovine aortlc endothelial (BAE) and human umbilical vein endothelial (HUVE) cells. All naturally-occuring bufadienolides used in this study inhibited the cell growth in a dose-dependent manner. Particularly, bufalin among the bufadienolides showed the strongest inhibitory activity for the cell growth. The order of growth inhibition by bufadienolides on BAE cells was as follows: bufalin > gamabufotalln > bufotalln > cinobufagin > cinobufotalin > resibufogenin. The $IC_50$ values (50% inhibition of cell growth) of bufalin as determined by XTT assay were the range of 1-10 nM in BAE and HUVE cells. Bufalin exhibited a higher sensitivity towards cultured bovine aortic endothelial cells than human umbilical vein endothelial cells.

• Journal of Yeungnam Medical Science
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• 제27권2호
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• pp.91-97
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• 2010

암세포의 전이, 죽상경화증, 당뇨성 망막병증과 같은 병적인 과정에서 혈관내피세포는 핵심적인 역할을 담당한다. 죽상경화증의 죽종 형성에 혈관민무늬근육세포가 직접적으로 관여한다. 배꼽정맥, 혈관내벽, 그리고 망막에 있는 이들 내피세포들은 다양한 효소용액들을 이용하여 얻는다. 순수하게 분리된 이들 세포는 내피세포와 관련된 질병의 시험관 내 연구에 있어 중요한 모델이다. 이러한 관점에서 볼 때 대동맥 벽의 중간막에서 분리한 후 배양한 민무늬근육세포도 죽상경화증의 발병을 설명할 수 있다. 이 종설에서는 사람배꼽정맥내피세포(HUVEC),대동맥의 내피세포 및 민무늬근육세포, 그리고 망막미세혈관내피세포(RMEC)의 분리 뿐 만 아니라 이들 세포를 이용한 질병연구에 관한 논문들을 소개하고자 한다.

• Preventive Nutrition and Food Science
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• 제20권4호
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• pp.221-229
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• 2015

Hesperidin has been shown to possess a potential inhibitory effect on vascular formation in endothelial cells. However, the fundamental mechanism for the anti-angiogenic activity of hesperidin is not fully understood. In the present study, we evaluated whether hesperidin has anti-angiogenic effects in mouse embryonic stem cell (mES)-derived endothelial-like cells, and human umbilical vascular endothelial cells (HUVECs), and evaluated their mechanism via the AKT/mammalian target of rapamycin (mTOR) signaling pathway. The endothelial cells were treated with several doses of hesperidin (12.5, 25, 50, and $100{\mu}M$) for 24 h. Cell viability and vascular formation were analyzed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and tube formation assay, respectively. Alteration of the AKT/mTOR signaling in vascular formation was analyzed by western blot. In addition, a mouse aortic ring assay was used to determine the effect of hesperidin on vascular formation. There were no differences between the viability of mES-derived endothelial-like cells and HUVECs after hesperidin treatment. However, hesperidin significantly inhibited cell migration and tube formation of HUVECs (P<0.05) and suppressed sprouting of microvessels in the mouse aortic ring assay. Moreover, hesperidin suppressed the expression of AKT and mTOR in HUVECs. Taken together, these findings suggest that hesperidin inhibits vascular formation by blocking the AKT/mTOR signaling pathways.

• Advances in nano research
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• 제12권5호
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• pp.501-514
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• 2022

This study aimed to investigate the effects and potential mechanisms of Chikusetsusaponin V (CsV) on endothelial nitric oxide synthase (eNOS) and vascular endothelial cell functions. Different concentrations of CsV were added to animal models, bovine aorta endothelial cells (BAECs) and human umbilical vein endothelial cells (HUVECs) cultured in vitro. qPCR, Western blotting (WB), and B ultrasound were performed to explore the effects of CsV on mouse endothelial cell functions, vascular stiffness and cellular eNOS mRNA, protein expression and NO release. Bioinformatics analysis, network pharmacology, molecular docking and protein mass spectrometry analysis were conducted to jointly predict the upstream transcription factors of eNOS. Furthermore, pulldown and ChIP and dual luciferase assays were employed for subsequent verification. At the presence or absence of CsV stimulation, either overexpression or knockdown of purine rich element binding protein A (PURA) was conducted, and PCR assay was employed to detect PURA and eNOS mRNA expressions, Western blot was used to detect PURA and eNOS protein expressions, cell NO release and serum NO levels. Tube formation experiment was conducted to detect the tube forming capability of HUVECs cells. The animal vasodilation function test detected the vasodilation functions. Ultrasonic detection was performed to determine the mouse aortic arch pulse wave velocity to identify aortic stiffness. CsV stimulus on bovine aortic cells revealed that CsV could upregulate eNOS protein levels in vascular endothelial cells in a concentration and time dependent manner. The expression levels of eNOS mRNA and phosphorylation sites Ser1177, Ser633 and Thr495 increased significantly after CsV stimulation. Meanwhile, CsV could also enhance the tube forming capability of HUVECs cells. Following the mice were gavaged using CsV, the eNOS protein level of mouse aortic endothelial cells was upregulated in a concentration- and time-dependent manner, and serum NO release and vasodilation ability were simultaneously elevated whereas arterial stiffness was alleviated. The pulldown, ChIP and dual luciferase assays demonstrated that PURA could bind to the eNOS promoter and facilitate the transcription of eNOS. Under the conditions of presence or absence of CsV stimulation, overexpression or knockdown of PURA indicated that the effect of CsV on vascular endothelial function and eNOS was weakened following PURA gene silence, whereas overexpression of PURA gene could enhance the effect of CsV upregulating eNOS expression. CsV could promote NO release from endothelial cells by upregulating the expression of PURA/eNOS pathway, improve endothelial cell functions, enhance vasodilation capability, and alleviate vessel stiffness. The present study plays a role in offering a theoretical basis for the development and application of CsV in vascular function improvement, and it also provides a more comprehensive understanding of the pharmacodynamics of CsV.

• International Journal of Oral Biology
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• 제33권4호
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• pp.149-154
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• 2008

Porphyromonas gingivalis, a major periodontal pathogen, has been implicated in the initiation and progression of periodontal disease. Endothelial dysfunction (Editor note: Aberrant and dysfunction are somewhat redundant. The authors may want to choose one or the other.) contributes to chronic periodontal inflammation. Using cDNA-representational difference analysis, we found that P.gingivalis lipopolysaccharide differentially induces a number of genes in human microvascular endothelial cells. Among these upregulated genes, we focused on intercellular adhesion molecule-1 (VCAM-1), which is crucial for leukocyte recruitment during vascular inflammation. P. gingivalis LPS significantly increased the expression of vascular cell adhesion molecule-1 (VCAM-1) as well as ICAM-1. Promoter assays revealed that the transcription of these cell adhesion molecules was mainly regulated by nuclear factor-${\kappa}B$ (NF-${\kappa}B$) in endothelial cells. Furthermore, P. gingivalis LPS significantly increased leukocyte adhesiveness to microvascular endothelial cells and to aortic endothelium. Taken together, our results demonstrate that P. gingivalis LPS activates microvascular endothelial cells through NF-${\kappa}B$-dependent expression of cell adhesion molecules.

• Preventive Nutrition and Food Science
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• 제19권4호
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• pp.299-306
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• 2014

Hesperetin has been shown to possess a potential anti-angiogenic effect, including vascular formation by endothelial cells. However, the mechanisms underlying the potential anti-angiogenic activity of hesperetin are not fully understood. In the present study, we evaluated whether hesperetin has anti-angiogenic effects in human umbilical vascular endothelial cells (HUVECs). HUVECs were treated with 50 ng/mL vascular endothelial growth factor (VEGF) to induce proliferation as well as vascular formation, followed by treatment with several doses of hesperetin (25, 50, and $100{\mu}M$) for 24 h. Cell proliferation and vascular formation were analyzed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and tube formation assay, respectively. In addition, cell signaling related to cell proliferation and vascular formation was analyzed by western blot. Furthermore, a mouse aorta ring assay was performed to confirm the effect of hesperetin on vascular formation. Hesperetin treatment did not cause differences in HUVECs proliferation. However, hesperetin significantly inhibited VEGF-induced cell migration and tube formation of HUVECs (P<0.05). Moreover, hesperetin suppressed the expression of ERK, p38 MAPK, and PI3K/AKT in the VEGF-induced HUVECs. In an ex vivo model, hesperetin also suppressed microvessel sprouting of mouse aortic rings. Taken together, the findings suggest that hesperetin inhibited vascular formation by endothelial cells via the inhibition of the PI3K/AKT, ERK and p38 MAPK signaling.

• Journal of Veterinary Science
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• 제21권1호
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• pp.9.1-9.15
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• 2020

Regenerative therapy holds great promise in the development of cures of some untreatable diseases such as cardiovascular diseases, and pluripotent stem cells (PSCs) including induced PSCs (iPSCs) are the most important regenerative seed cells. Recently, differentiation of human PSCs into functional tissues and cells in vitro has been widely reported. However, although porcine reports are rare they are quite essential, as the pig is an important animal model for the in vitro generation of human organs. In this study, we reprogramed porcine embryonic fibroblasts into porcine iPSCs (piPSCs), and differentiated them into cluster of differentiation 31 (CD31)-positive endothelial cells (ECs) (piPSC-derived ECs, piPS-ECs) using an optimized single-layer culture method. During differentiation, we observed that a combination of GSK3β inhibitor (CHIR99021) and bone morphogenetic protein 4 (BMP4) promoted mesodermal differentiation, resulting in higher proportions of CD31-positive cells than those from separate CHIR99021 or BMP4 treatment. Importantly, the piPS-ECs showed comparable morphological and functional properties to immortalized porcine aortic ECs, which are capable of taking up low-density lipoprotein and forming network structures on Matrigel. Our study, which is the first trial on a species other than human and mouse, has provided an optimized single-layer culture method for obtaining ECs from porcine PSCs. Our approach can be beneficial when evaluating autologous EC transplantation in pig models.

• 농업과학연구
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• 제48권2호
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• pp.353-365
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• 2021

Previously, we reported that tissue factor (Tf) was included in the list of differentially expressed genes as an upregulated gene in a rejected porcine heart after xenotransplantation into monkey. In this study, we analyzed that expression of Tf in aortic endothelial cells (pAEC) isolated from alpha 1,3-galactosyltransferase knockout pig in response to allogeneic porcine serum and xenogeneic human serum. The consequence was significant upregulation of Tf expression by responding to human serum compared with porcine serum. To analyze the function of Tf gene as a promoter, we constructed reporter vectors for expression of luciferase linked to 1,246 and 787 base pairs of porcine Tf (pTF1246 and pTF787), and 535 base pairs of human TF (hTF535) sequences including putative promoter regions and AP-1 biding site at the 5' end. The reporter vectors were transfected into pAEC including cytomegalovirus enhancer/chicken β-actin (CAG)-luciferase vector as a control. Luciferase assay showed that all of the promoters were insufficient to express luciferase compared with CAG promoter in basic culture conditions. Notably, pTF1246, pTF787, and hTF535 led to a significant increase of luciferase expression in response to human serum compared with porcine serum while no change of CAG. pTF1246 and pTF787 showed higher expression than hTF535. Taken together, our findings suggest that pTF1246 and pTF787 promoters could mediate target gene expression specifically at xenogeneic stress condition.

• BMB Reports
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• 제42권9호
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• pp.561-567
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• 2009

Although many plant-derived phenolic compounds display antioxidant effects in biological systems, their mechanism of action remains controversial. In this study, the mechanism by which p-coumaric acid (p-CA) performs its antioxidant action was investigated in bovine aortic endothelial cells under oxidative stress due to high levels of glucose (HG) and arachidonic acid (AA), a free fatty acid. p-CA prevented lipid peroxidation and cell death due to HG+AA without affecting the production of reactive oxygen species. The antioxidant effect of p-CA was not decreased by buthionine-(S,R)-sulfoximine, an inhibitor of cellular GSH synthesis. In contrast, pretreatment with p-CA caused the induction of peroxidases that decomposed t-butyl hydroperoxide in a p-CA-dependent manner. Furthermore, the antioxidant effect of p-CA was significantly mitigated by methimazole, which was shown to inhibit the catalytic activity of 'p-CA peroxidases' in vitro. Therefore, it is suggested that the induction of these previously unidentified 'p-CA peroxidases' is responsible for the antioxidant effect of p-CA.

• 동의생리병리학회지
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• 제18권5호
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• pp.1382-1386
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• 2004

This study was performed for the investigation of vasodilatory efficacy and its underlying mechanisms of Samhwangsasim-tang(SST), herbal remedy. SST relaxed vascular strips precontracted with phenylephrine or KCI(51 mM), but the magnitude of relaxation was greater in phenylephrine(PE) induced contraction. The relaxation effects of SST was endothelium-independent. L-NAME, iNOS inhibitor, and methyl en blue(MB), cGMP inhibitor, did not attenuate the relaxation responses of SST. In the absence of extracellular Ca2+, pre-incubation of the aortic rings with SST significantly reduced the contraction by PE, suggesting that the relaxant action of the SST includes inhibition of Ca/sup 2+/ influx and release of Ca/sup 2+/ from intracellular stores (SR). In addition, the cell death was induced by SST in human aortic smooth muscle cells but not that of human umbilical vein endothelial cells. We conclude that in rat thoracic aorta, SST may induce in part vasodilation through inhibition of Ca/sup 2+/ influx and release of Ca/sup 2+/ from intracellular stores.