• Biomolecules & Therapeutics
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• 제32권2호
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• pp.205-213
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• 2024

Hydroxychavicol, a primary active phenolic compound of betel leaves, previously inhibited bone loss in vivo by stimulating osteogenesis. However, the effect of hydroxychavicol on bone remodeling induced by osteoclasts is unknown. In this study, the anti-osteoclastogenic effects of hydroxychavicol and its mechanism were investigated in receptor activator of nuclear factor kappa-B ligand (RANKL)-induced osteoclasts. Hydroxychavicol reduced the number of tartrate resistance acid phosphatase (TRAP)-positive multinucleated, F-actin ring formation and bone-resorbing activity of osteoclasts differentiated from RAW264.7 cells in a concentration-dependent manner. Furthermore, hydroxychavicol decreased the expression of osteoclast-specific genes, including cathepsin K, MMP-9, and dendritic cell-specific transmembrane protein (DC-STAMP). For mechanistic studies, hydroxychavicol suppressed RANKL-induced expression of major transcription factors, including the nuclear factor of activated T-cells 1 (NFATc1), c-Fos, and c-Jun. At the early stage of osteoclast differentiation, hydroxychavicol blocked the phosphorylation of NF-κB subunits (p65 and Iκβα). This blockade led to the decrease of nuclear translocation of p65 induced by RANKL. In addition, the anti-osteoclastogenic effect of hydroxychavicol was confirmed by the inhibition of TRAP-positive multinucleated differentiation from human peripheral mononuclear cells (PBMCs). In conclusion, hydroxychavicol inhibits osteoclastogenesis by abrogating RANKL-induced NFATc1 expression by suppressing the NF-κB signaling pathway in vitro.

• Molecules and Cells
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• 제41권3호
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• pp.244-255
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• 2018

Low-grade pro-inflammatory state and leptin resistance are important underlying mechanisms that contribute to obesity-associated hypertension. We tested the hypothesis that Astragaloside IV (As IV), known to counteract obesity and hypertension, could prevent obesity-associated hypertension by inhibiting pro-inflammatory reaction and leptin resistance. High-fat diet (HFD) induced obese rats were randomly assigned to three groups: the HFD control group (HF con group), As IV group, and the As IV + ${\alpha}$-bungaratoxin (${\alpha}-BGT$) group (As IV+${\alpha}-BGT$ group). As IV ($20mg{\cdot}Kg^{-1}{\cdot}d^{-1}$) was administrated to rats for 6 weeks via daily oral gavage. Body weight and blood pressure were continuously measured, and NE levels in the plasma and renal cortex was evaluated to reflect the sympathetic activity. The expressions of leptin receptor (LepRb) mRNA, phosphorylated signal transducer and activator of transcription-3 (p-STAT3), phosphorylated phosphatidylinositol 3-kinase (p-PI3K), suppressor of cytokine signaling 3 (SOCS3) mRNA, and protein-tyrosine phosphatase 1B (PTP1B) mRNA, pro-opiomelanocortin (POMC) mRNA and neuropeptide Y (NPY) mRNA were measured by Western blot or qRT-PCR to evaluate the hypothalamic leptin sensitivity. Additionally, we measured the protein or mRNA levels of ${\alpha}7nAChR$, inhibitor of nuclear factor ${\kappa}B$ kinase subunit ${\beta}/nuclear$ factor ${\kappa}B$ ($IKK{\beta}/NF-KB$) and pro-inflammatory cytokines ($IL-1{\beta}$ and $TNF-{\alpha}$) in hypothalamus and adipose tissue to reflect the anti-inflammatory effects of As IV through upregulating expression of ${\alpha}7nAChR$. We found that As IV prevented body weight gain and adipose accumulation, and also improved metabolic disorders in HFD rats. Furthermore, As IV decreased BP and HR, as well as NE levels in blood and renal tissue. In the hypothalamus, As IV alleviated leptin resistance as evidenced by the increased p-STAT3, LepRb mRNA and POMC mRNA, and decreased p-PI3K, SOCS3 mRNA, and PTP1B mRNA. The effects of As IV on leptin sensitivity were related in part to the up-regulated ${\alpha}7nAchR$ and suppressed $IKK{\beta}/NF-KB$ signaling and pro-inflammatory cytokines in the hypothalamus and adipose tissue, since co-administration of ${\alpha}7nAChR$ selective antagonist ${\alpha}-BGT$ could weaken the improved effect of As IV on central leptin resistance. Our study suggested that As IV could efficiently prevent obesityassociated hypertension through inhibiting inflammatory reaction and improving leptin resistance; furthermore, these effects of As IV was partly related to the increased ${\alpha}7nAchR$ expression.

• Journal of Periodontal and Implant Science
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• 제28권4호
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• pp.769-786
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• 1998

The purpose of the present study is to examine the effect of cell proliferation and alkaline phosphatase activity in osteoblastic cells and to compare the bone healing ability of rat calvarial defects between the control group and the safflower seed extract treated group. Osteoblastic cells were obtained from calvariae of a fetal rat. Cells were cultured containing DMEM and safflower seed extract ($10^{-6}g/ml$, $10^{-3}g/ml$) at $37^{\circ}$ with 5% $CO_2$ in 100% humidity for 3 days. MTT was performed to examine the viability of the cells, and alkaline phosphatase activity was analyzed to examine the mineralization in vitro. Rat calvarial defects($5{\times}5mm$) in 250g Sprague-Dawly were made using round bur. Rats were administrated with safflower seed extract(0.35g/kg/day) for experimental periods. Calvarial defects were studied histopathologically and immunohistochemically at 1,4, and 8 weeks. High concentration group($10^{-3}g/ml$) of safflower seed extract significantly increased in the cell proliferation and alkaline phos phatase synthesis in osteoblastic cells. The infiltration of inflammatory cells and osteoclastic activities were decreased in the safflower seed extract treated group as compared with control group. Bone maturation was accelerated in the safflower seed extract treated group as compared to control group. No difference in osteoinductive process was observed between the control and the safflower seed extract treated group. Immunohistochemical observation revealed that protein expression of TGF-$\beta$and osteonectin during early healing phase in the safflower seed extract treated group was slightly increased as compared to control group. These results indicate that safflower seed extract promotes the healing process in bony defect of rat calvariae, and retains a potential applicability as an adjuvant therapeutic modality for regeneration of periodontal bony defect.

• 한국동물학회지
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• 제34권2호
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• pp.159-172
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• 1991

흰쥐 부정소 정자의 성숙 전후에 일어나는 변화를 몇가지 효소를 중심으로 관찰하였고 그 성숙과정중에 일어나는 상피세포, 내강 및 정자 사이의 상호관계를 알아보기 위하여 실험군 별로 몇가지 효소의 활성도와 탄수화물 잔기의 함량을 측정하였으며, 전기영동을 이용하여 각 군의 차이를 관찰하고 이에 대한 웅성호르몬의 관련성을 알아보았다. 1. 부정소 두 정자와 부정소미 정자에서 활성도 측정시 lactate dehydrogenase, glucose-6-phosphatase 및 Na+ -K+ -ATPase의 경우는 유의한 차이가 없었으며, $Mg^2$+-ATPase의 경우만이 부정소미 정자가 부정소두 정자보다 유의성 있게 높은 활성도를 가지는 것으로 나타났다. 또한 부정소두와 부정소미 상피세포 , 내강 및 정자의 세군으로 나누어 각각 탄수화물 잔기를 정량하였을 때 hexosamine은 상피세포에서, sialic acid는 상피세포와 내강액에서 부정소미의 경우가 더 높은 함량이 존재하였으며, 내강액과 정자의 crude membrance fraction을 SDS-PAGE 했을 때 분자량이 33-37 KD 사이에 존재하던 band가 부정소미 내강액과 부정소미 정자의 crude membrance fraction에서 관찰되었으므로 흰쥐에서 정자의 성숙과정과 관련된 부정소내의 여러 변화를 비교하는 자료가 될 수 있었다. 2. 부정소 내강액에서 $\beta$ -glucuronidase와 $\beta$ -glucosidase의 활성도 및 웅성호르몬에 대한 의존성을 측정하였을 때 거세 후 5일째부터 이 두 효소의 활성도가 모두 유의하게 감소하기 시작하였고, tentosterone을 투여하였을 때는 $\beta$ -glucuronidase는 투여 5일, $\beta$ -glucosidase는 투여 10일 후부터 유의하게 증가하였으며 웅성호르몬에 대한 내강액의 의존성을 알아보기 위하여 SDS-PAGE하였을 때 tentosterone투여군의 부정소두에서 분자량이 약 21 KD에 해당하는 band를 새로이 관찰하였다.

• Genomics & Informatics
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• 제6권3호
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• pp.99-109
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• 2008

Protein phosphorylation at tyrosine residues is a key regulatory event that modulates insulin signal transduction. We studied the PTPN1 gene with regard to susceptibility to Korean type 2 diabetes mellitus (T2DM) and its related quantitative traits. A total of seven SNPs [g.36171G>A (rs941798), g.58166G>A (rs3787343), g.58208A>G (rs2909270), g.64840C>T (rs754118), g.69560C>G (rs6020612), g.69866G>A (rs718050), and g.69934T>G (rs3787343)] were selected based on frequency (>0.05), linkage disequilibrium (LD) status, and haplotype tagging status. We studied the seven SNPs in 483 unrelated patients with type 2 diabetes (age: $64{\pm}2.8$ years, onset age: $56{\pm}8.1$ years; 206 men, 277 women) and 1138 nondiabetic control subjects (age: $64{\pm}2.9$; 516 men, 622 women). The SNP rs941798 had protective effects against T2DM with an odds ratio of 0.726 (C.I. $0.541{\sim}0.975$) and p-value=0.034, but none of the remaining six SNPs was associated with T2DM. Also, rs941798 was associated with blood pressure, HDL cholesterol, insulin sensitivity. rs941798 also has been associated with T2DM in previous reports of Caucasian-American and Hispanic-American populations. This is the first report that shows an association between PTPN1 and T2DM in the Korean as well as Asian population.

• Journal of Ginseng Research
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• 제39권1호
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• pp.46-53
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• 2015

Background: Glucocorticoids (GCs) are commonly used in many chemotherapeutic protocols and play an important role in the normal regulation of bone remodeling. However, the prolonged use of GCs results in osteoporosis, which is partially due to apoptosis of osteoblasts and osteocytes. In this study, effects of Korean Red Ginseng (KRG) on GC-treated murine osteoblastic MC3T3-E1 cells and a GC-induced osteoporosis mouse model were investigated. Methods: MC3T3-E1 cells were exposed to dexamethasone (Dex) with or without KRG and cell viability was measured by the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. Realtime polymerase chain reaction was performed to evaluate the apoptotic gene expression; osteogenic gene expression and alkaline phosphatase (ALP) activity were also measured. Western blotting was performed to evaluate the mitogen-activated protein kinase (MAPK) proteins. A GC-induced osteoporosis animal model was used for in vivo study. Results and conclusion: The MTT assay revealed that Korean Red Ginseng (KRG) prevents loss of cell viability caused by Dex-induced apoptosis in MC3T3E1 cells. Real-time polymerase chain reaction data showed that groups treated with both Dex and KRG exhibited lower mRNA levels of caspase-3 and -9, whereas the mRNA levels of Bcl2, IAPs, and XIAP increased. Moreover, groups treated with both Dex and KRG demonstrated increased mRNA levels of ALP, RUNX2, and bone morphogenic proteins as well as increased ALP activity in MC3T3-E1 cells, compared to cells treated with Dex only. In addition, KRG increased protein kinase B (AKT) phosphorylation and decreased c-Jun N-terminal kinase (JNK) phosphorylation. Moreover, microcomputed tomography analysis of the femurs showed that GC implantation caused trabecular bone loss. However, a significant reduction of bone loss was observed in the KRG-treated group. These results suggest that the molecular mechanism of KRG in the GC-induced apoptosis may lead to the development of therapeutic strategies to prevent and/or delay osteoporosis.

• Clinical and Experimental Reproductive Medicine
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• 제28권1호
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• pp.33-40
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• 2001

Objective: This study was to establish the human embryonic stem (ES) cells derived from frozen-thawed blastocyst stage embryo that were destined to be discarded after five years in routine human IVF-ET program. Methods: Frozen-thawed and survived human blastocysts were treated by immunosurgery, and recovered ICM cells were cultured onto STO feeder cell layer and ICM colony was subcultured by mechanical dissociation into clumps. To identify ES cell, alkaline phosphatase staining and expression of Oct4 in replated ICM colonies were examined. Also, to examine the possibility of ES cell differentiation, retinoic acid (RA), basic fibroblast growth factor (b-FGF), nerve growth factor (NGF) were added in culture medium. In addition, to classify the specific cell type, differentiated cells were stained by indirect immunocytochemistry. Results: One ICM colony recovered from frozen-thawed six blastocysts was subcultured, continuously replated during 40 passage culture duration without differentiation. Subcultured colonies were strong positively stained by alkaline phophatase. When the expression of Oct4 in cultured ES colony was examined, Oct4b type is more clearly indicated than Oct4a one although there was not detected in embryoid body or differentiated cells. In differentiated cardiomyocytes from ES colony, cells were beaten regularly (60 times/min). In differentiated neural cells from ES colony, neurofilament (NF) 200 kDa protein, microtubule associated protein (MAP) 2 and ${\beta}$-tubulin of specific marker in neurons, glial fibrillary acidic protein (GFAP) of specific marker in astrocytes and galactocelebrocide (GalC) of specific marker in oligodendrocytes were confirmed by indirect immunocytochemistry. Also, muscle cells were detected by indirect immunocytochemistry. In addition, ES colonies can be successfully cryopreserved. Conclusion: This study suggested that establishment of human ES cells can be successfully derived from frozen-thawed blastocysts that were destined to be discarded, and obtained specific cell types (cardiomyocytes, neurons and muscle cells) through the in vitro differentiation procedures of ES cells.

• 대한수의학회지
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• 제21권2호
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• pp.167-170
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• 1981

Changes in serum glutamic oxaloacetic transaminase (s-GOT) activities, serum alkaline phosphatase activities and serum total protein amounts were investigated on seven Korean native cows having normal estrus cycle of 18~24 days after normal parturition, dividing estrus cycle into estrus (0~1 days), metestrus (2~6 days), diestrus (7~16 days), proestrus (17~20 days). The results were as follows. 1. Serum GOT activities at estrus ranged from 73.5 to 121.5 Ku with a mean of $89.14{\pm}17.16$ Ku, at metestrus 57 to 89 Ku with a mean of $67.01{\pm}10.81$ Ku, at diestrus 54 to 89.5 Ku with a mean of $67.05{\pm}10.05$ Ku and at proestrus 53 to 112 Ku with a mean of $73.00{\pm}20.05$ Ku. The activities were significantly increased at the estrus comparing with other stages (P<0.01). 2. Serum ALP activities at estrus ranged from 8.0 to 10.4 K-Au with a mean of $8.74{\pm}0.83$ K-Au, at metestrus 6.5 to 9.2 K-Au with a mean of$7.74{\pm}0.76$ K-Au, at diestrus 5.6 to 9.0 K-Au with a mean of $7.67{\pm}1.13$ K-Au, at proestrus 6.2 to 9.3 K-Au with a mean of $7.12{\pm}1.18$ K-Au. The significance was not recognized among the stages of estrus cycle. 3. Serum total protein amounts at estrus ranged from 6.45 to 8.0g/10dl with a mean of $7.25{\pm}0.57/100dl$, at metestrus, 6.37 to 7.9g/100dl with a mean of $7.65{\pm}0.50g/100dl$, fat diestrus, 6.56 to 8.67g/100dl, with a mean of $7.53{\pm}0.55g/100dl$ and at proestrus 5.94 to 7.71g/100dl with a mean of $6.54{\pm}0.65g/dl$. There was not significance among the stages of estrus cycle.

• Asian-Australasian Journal of Animal Sciences
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• 제30권2호
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• pp.221-228
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• 2017

Objective: The objective of this experiment was to characterize the mRNA expression profile of type IIb sodium-inorganic phosphate cotransporter (NaPi-IIb) and the biochemical values of serum alkaline phosphatase (AKP), calcium, inorganic phosphorus, tibial ash and minerals of broiler chickens with aging. Methods: A total of 56 one-day-old Arbor Acres male broiler chickens were used. Broiler chickens were weighed and samples were collected weekly from day 1. Results: The result showed that before the growth inflection point, ash, calcium, and phosphorus content in the tibia of broiler chickens increased with growth (before 3 weeks of age), although there were no significant differences in chicks at different ages in the later period of the experiment and weight gain rate was relatively slow at this stage (4 to 6 weeks). NaPi-IIb gene expression in the small intestine in the early growth stage was higher than that in the later growth stage. Expression of calbindin and the vitamin D receptor protein in the intestinal mucosa increased with age in the duodenum and jejunum. Serum AKP activity first increased and subsequently decreased after peaking at 1 week of age, but there was no significant difference after 3 weeks of age. Conclusion: These results show that compared with the early growth stage, the weight-gain rate of broiler chickens in the late growth stage gradually decreased with gradual tibia maturation, along with weaker positive transport of phosphorus in the intestine and reinforced re-absorption of phosphorus in the kidney, which might be the reason that phosphorus requirement in the late growth stage was decreased.

• Parasites, Hosts and Diseases
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• 제58권3호
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• pp.249-255
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• 2020

Toxoplasma gondii, a ubiquitous, intracellular parasite of the phylum Apicomplexa, infects an estimated one-third of the human population as well as a broad range of warm-blooded animals. We have observed that some tyrosine kinase inhibitors suppressed the growth of T. gondii within host ARPE-10 cells. Among them, afatinib, human epithermal growth factor receptor 2 and 4 (HER2/4) inhibitor, may be used as a therapeutic agent for inhibiting parasite growth with minimal adverse effects on host. In this report, we conducted a proteomic analysis to observe changes in host proteins that were altered via infection with T. gondii and the treatment of HER2/4 inhibitors. Secreting proteins were subjected to a procedure of micor basic reverse phase liquid chromatography, nano-liquid chromatography-mass spectrometry, and ingenuity pathway analysis serially. As a result, the expression level of heterogeneous nuclear ribonucleoprotein K, semaphorin 7A, a GPI membrane anchor, serine/threonine-protein phosphatase 2A, and calpain small subunit 1 proteins were significantly changed, and which were confirmed further by western blot analysis. Changes in various proteins, including these 4 proteins, can be used as a basis for explaining the effects of T. gondii infections and HER2/4 inhibitors.