• Journal of Microbiology and Biotechnology
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• 제25권2호
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• pp.274-279
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• 2015

Receptor activator of nuclear factor-kappa B ligand (RANKL) is a critical factor in osteoclastogenesis. It makes osteoclasts differentiate and multinucleate in bone remodeling. In the present study, RANKL was expressed as a soluble maltose binding protein (MBP)-fusion protein using the Escherichia coli maltose binding domain tag system (pMAL) expression vector system. The host cell E. coli DH5α was cultured and induced by isopropyl β-_D-1-thiogalactopyranoside for rRANKL expression. Cells were disrupted by sonication to collect soluble MBP-fused rRANKL. The MBP-fusion rRANKL was purified with MBP Trap affinity chromatography and treated with Tobacco Etch Virus nuclear inclusion endopeptidase (TEV protease) to remove the MBP fusion protein. Dialysis was then carried out to remove binding maltose from the cleaved rRANKL solution. The cleaved rRANKL was purified with a second MBP Trap affinity chromatography to separate unsevered MBP-fusion rRANKL and cleaved MBP fusion protein. The purified rRANKL was shown to have biological activity by performing in vitro cell tests. In conclusion, biologically active rRANKL was successfully purified by a simple two-step chromatography purification process with one column.

• The Korean Journal of Physiology and Pharmacology
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• 제13권1호
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• pp.1-7
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• 2009

Sodium retention is a hallmark of nephrotic syndrome. We investigated whether sodium retention is associated with changes of natriuretic peptide system at different stages (i.e., a sodium retaining stage and a compensatory stage) of nephrotic syndrome. At day 7 after PAN(puromycin aminonucleoside) injection, the urinary excretion of sodium was decreased, along with the development of ascites and positive sodium balance. The plasma and urinary ANP(atrial natriuretic peptide) immunoreactivities were increased. ANP mRNA expression was increased in the heart and kidney, whereas that of NPR(natriuretic peptide receptor)-A and NPR-C mRNA was decreased in the kidney. The expression of NEP was decreased in the kidney. At day 14, urinary excretion of sodium did not differ from the control. The plasma ANP level and heart ANP mRNA expression returned to their control values. The expression of ANP mRNA in the kidney was increased in association with increased urinary ANP immunoreactivities. The expression of NPR-A in the kidney became normal, whereas that of NPR-C kept decreased. The expression of NEP(neutral endopeptidase) remained decreased. These findings suggest that the increased renal ANP synthesis in association with decreased metabolism via NEP and NPR-C may play a compensatory role against the development of sodium retention in nephrotic syndrome. The decreased of NPR-A expression in the kidney may contribute to the ANP resistance at day 7. The subsequent recovery of NPR-A expression may play a role in promoting sodium excretion in later stage(at day 14).

• 대한한방내과학회지
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• 제21권4호
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• pp.543-548
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• 2000

After Vanherweghem J-L reported the rapidly progressive interstitial renal fibrosis that developed in patients taking the slimming preparation compounded with chinese Herb in Belgium 1993. Chinese Herb Nephropathy(CHN) has become known as a new renal disease, CHN is described as the decrease of urinary renal enzyme, neutral endopeptidase(NEP). N-acetyl-${\beta}$-Dglucosaminidase(NAG). increase of urinary low molecular protein, ${\alpha}$1-microglobulin, ${\beta}$2-microglobulin. clara cell protein(CC16), retinol-binding protein(RBP) in clinical findings, and the proximal tubular atrophy, interstitial fibrosis. urothelial atrophy. glomerular sclerosis in histology, Because CHN was caused by Chinese herb contained in slimming preparation, western medical doctors have thought that all Herb medicine might have caused renal disease and prohibit the taking of any Herb medicine, However. CHN was actually caused by the aristolochic acid contained in some Herb medicines. Aristolochia manshuriensis, Aristolochia fang chi, which is the substitutions of Akebia quinata, Stephania tetrandra has being used in clinical. Aristolochia manshuriensis. Aristolochia fang chi were different with Akebia quinata. Stephania tetrandra in botany, and it have not been classified with medicines in Oriental medicine, That is, the aristolochic acid, not Herb medicines. causes CHN, So, Chinese Herb Nephropathy should be changed to Aristolochic acid Nephropathy.

• Asian Pacific Journal of Cancer Prevention
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• 제14권9호
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• pp.5055-5060
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• 2013

Objective: Twist, a basic helix-loop-helix transcription factor, plays a key role in the metastatic progression of human cancer. Matrix metalloproteinase (MMP)-9 is an endopeptidase that digests basement membrane type IV collagen, therefore being possibly related to tumor progression. It has been reported that Twist and matrix metalloproteinase-9 (MMP-9) are expressed in gastric cancers. However, the exact roles of Twist and MMP-9 in tumor metastasis and prognosis remain unclear. The aim of this study was to casts light on this question. Methods:Twist and MMP-9 expression in tissue sections of 37 gastric carcinomas was evaluated with immunohistochemistry. The staining results were compared with clinicopatholgic features and to patients' outcome. Results: Twist positive expression was significantly increased in gastric cancer cases with lymph node metastasis (P=0.023). But no correlations were found between MMP-9 overexpression and clinicopathologic features, such as recurrence, TNM stage, and lymph node metastasis. Overall survival (OS) was significantly correlated with recurrence, serosa invasion, TNM stages, distant metastasis, and MMP-9 (P=0.027, 0.021, 0.000, 0.024 and 0.036, respectively). Disease-free survival (DFS) was prominently related to recurrence location, serosa invasion and TNM stages (P=0.000, 0.038 and 0.003, respectively). In the Cox regression multivariate analysis, TNM stage, distant metastasis and MMP-9 were significantly associated with prognosis of gastric cancer (P=0.002, 0.019, and 0.032, respectively). Conclusions: This study showed Twist positive expression to be significantly correlated with lymph node metastasis in gastric cancer. MMP-9 overexpression is associated with OS, suggesting that MMP-9 is a prognostic indicator for survival in patients with gastric cancer.

• Journal of Microbiology and Biotechnology
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• 제25권8호
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• pp.1307-1314
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• 2015

Leukemia inhibitory factor (LIF) is a member of the IL-6 cytokine family, having pleiotropic actions such as maintaining stem cell pluripotency and enabling blastocyst implantation. Because the action of LIF is mediated by a ligand-receptor interaction with the LIF receptor (LIF-R), an antagonist for LIF-R has been developed to inhibit LIF-induced signaling. In this study, we present a novel method for the production and purification of an antagonist to human LIF-R (hLA). His-tagged hLA was expressed in E. coli, and simple purification methods without any endopeptidase cleavage were designed. In addition, we determined the optimal temperature conditions for enhancing the production of soluble hLA. Finally, the bioactivity of His-tagged hLA was examined using STAT3 phosphorylation and receptivity of human endometrial ECC-1 cells. Our strategy provides a rapid and efficient method to produce biologically active recombinant hLA.

• The Korean Journal of Physiology and Pharmacology
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• 제15권5호
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• pp.273-277
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• 2011

Nitric oxide (NO) and atrial natriuretic peptide (ANP) may induce vascular relaxation by increasing the production of cyclic guanosine monophosphate (cGMP), an important mediator of vascular tone during sepsis. This study aimed to determine whether regulation of NO and the ANP system is altered in lipopolysaccharide (LPS)-induced kidney injury. LPS (10 $mg{\cdot}kg^{-1}$) was injected in the tail veins of male Sprague-Dawley rats; 12 hours later, the kidneys were removed. Protein expression of NO synthase (NOS) and neutral endopeptidase (NEP) was determined by semiquantitative immuno-blotting. As an index of synthesis of NO, its stable metabolites (nitrite/nitrate, NOx) were measured using colorimetric assays. mRNA expression of the ANP system was determined by real-time polymerase chain reaction. To determine the activity of guanylyl cyclase (GC), the amount of cGMP generated in response to sodium nitroprusside (SNP) and ANP was calculated. Creatinine clearance decreased and fractional excretion of sodium increased in LPS-treated rats compared with the controls. Inducible NOS protein expression increased in LPS-treated rats, while that of endothelial NOS, neuronal NOS, and NEP remained unchanged. Additionally, urinary and plasma NOx levels increased in LPS-treated rats. SNP-stimulated GC activity remained unchanged in the glomerulus and papilla in the LPS-treated rats. mRNA expression of natriuretic peptide receptor (NPR)-C decreased in LPS-treated rats, while that of ANP and NPR-A did not change. ANP-stimulated GC activity reduced in the glomerulus and papilla. In conclusion, enhancement of the NO/cGMP pathway and decrease in ANP clearance were found play a role in the pathogenesis of LPS-induced kidney injury.

• 생약학회지
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• 제36권3호통권142호
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• pp.235-239
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• 2005

Changes in chemical composition of the ethanolic extract of roasted Asparagi Tuber were investigated by HPLC. One dramatically increased peak $(t_R\;14.85 min)$ was isolated by silica gel column chromatograph and identified as 5-hydroxymethyl furfural (HMF) by comparing its $1^H-NMR$ data with that of a commercial standard. HMF content reached its maximum level at $190^{\circ}C$ for 60 minutes. Under these conditions, HMF contents in the roasted Asparagi Tuber was increased about thirteen times (9.26mg/g) over the not-roasted control (0.71 mg/g). No significant differences were found in macrophage-activating, prolyl endopeptidase-inhibiting, antioxidative (DPPH), anti-coagulating (activated partial thromboplastin times), and ACE-inhibiting activities between roasted and not-roasted Asparagi Tuber.

• Food Science and Biotechnology
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• 제15권4호
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• pp.617-624
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• 2006

Various plant phenolics were assessed for (${\beta}$-secretase (BACE1) inhibitory activity in order to screen for anti-dementia agents. Among 39 phenolics, eight compounds, 1,2,3-trigalloyl glucopyranoside, acetonyl geraniin, euphorscopin, furosine, helioscopinin A, helioscopinin B, jolkinin, and rugosin E exhibited strong inhibition of BACE1 with $IC_{50}$ values of $5.87{\times}10^{-8}-54.93{\times}10^{-6}\;M$. Among them, rugosin E was the most potent ($IC_{50}$ $5.87{\times}10^{-8}\;M$). The active compounds were shown to be non-competitive inhibitors by Dixon plot. All the phenolic BACE1 inhibitors except furosin also suppressed prolyl endopeptidase (PEP) activity. However, these phenolic compounds caused less inhibition of ${\alpha}$-secretase (tumor necrosis factor a converting enzyme; TACE) and no significant inhibition of other serine proteases such as trypsin, chymotrypsin, and elastase was seen, demonstrating that they are relatively specific to both BACE1 and PEP. No significant structure-activity relationships were found.

• 치위생과학회지
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• 제21권4호
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• pp.227-232
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• 2021

Background: Cancer-associated fibroblasts (CAFs) are abundant in tumor microenvironments and interact with cancer cells to promote tumor proliferation in oral squamous cell carcinoma (OSCC). Cathepsin D (CTSD) is a soluble lysosomal aspartic endopeptidase involved in tumor proliferation and angiogenesis. In this preliminary study, we observed CTSD expression in OSCC and CAFs, postulating that CTSD might act as a bridge between OSCC and CAFs. Methods: Human epidermal keratinocytes (HEKs), OSCC, and immortalized human normal oral fibroblasts (hTERT-hNOFs) were used in this study. Additionally, we used hTERT-hNOFs transfected with an empty vector, WT (wild-type)-YAP (Yes-associated protein), and YAPS127A (YAP serine 127 to alanine). YAP127A hTERT-hNOFs activated fibroblasts similar to CAFs. To identify CTSD expression between OSCC and CAFs, conditioned medium (CM) was collected from each cell. Protein expression of CTSD was identified by western blotting. Results: To identify the expression of CTSD in fibroblasts stimulated by OSCC, we treated fibroblasts with CM from HEK and OSCC. Results indicated that hTERT-hNOFs with OSCC CM showed a weakly increased expression of CTSD compared to stimulation by HEK CM. This indicates that CAFs, YAPS127 hTRET-hNOFs, overexpress CTSD protein. HEK cells showed no CTSD expression, regardless of treatment with fibroblast CM, whereas OSCC highly expressed CTSD proteins compared with the CTSD expression in HEK cells. We also found that CTSD expression was unaffected by changes in transforming growth factor-β levels. Conclusion: This study proposes that CTSD might have potential as an interacting executor between OSCC and CAFs. Further studies are needed to investigate the role of CTSD in tumor and stromal cells.

• 운동영양학회지
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• 제25권4호
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• pp.38-44
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• 2021

[Purpose] Exercise can prevent conditions such as atrophy and degenerative brain diseases. However, owing to individual differences in athletic ability, exercise supplements can be used to improve a person's exercise capacity. Schisandra chinensis (SC) is a natural product with various physiologically active effects. In this study, we analyzed SC using a pharmacological network and determined whether it could be used as an exercise supplement. [Methods] The active compounds of SC and target genes were identified using the Traditional Chinese Medicine Database and Analysis Platform (TCMSP). The active compound and target genes were selected based on pharmacokinetic (PK) conditions (oral bioavailability (OB) ≥ 30%, Caco-2 permeability (Caco-2) ≥ -0.4, and drug-likeness (DL) ≥ 0.18). Gene ontology (GO) was analyzed using the Cytoscape software. [Results] Eight active compounds were identified according to the PK conditions. Twenty-one target genes were identified after excluding duplicates in the eight active compounds. The top 10 GOs were analyzed using GO-biological process analysis. GO was subsequently divided into three representative categories: postsynaptic neurotransmitter receptor activity (53.85%), an intracellular steroid hormone receptor signaling pathway (36.46%), and endopeptidase activity (10%). SC is related to immune function. [Conclusion] According to the GO analysis, SC plays a role in immunity and inflammation, promotes liver metabolism, improves fatigue, and regulates the function of steroid receptors. Therefore, we suggest SC as an exercise supplement with nutritional and anti-fatigue benefits.