• Clinical and Experimental Reproductive Medicine
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• 제39권2호
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• pp.58-67
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• 2012

Objective: Previously, we identified that transketolase (Tkt), an important enzyme in the pentose phosphate pathway, is highly expressed at 2 hours of spontaneous maturation in oocytes. Therefore, this study was performed to determine the function of Tkt in meiotic cell cycle regulation, especially at the point of germinal vesicle breakdown (GVBD). Methods: We evaluated the loss-of-function of Tkt by microinjecting Tkt double-stranded RNAs (dsRNAs) into germinal vesicle-stage oocytes, and the oocytes were cultured in vitro to evaluate phenotypic changes during oocyte maturation. In addition to maturation rates, meiotic spindle and chromosome rearrangements, and changes in expression of other enzymes in the pentose phosphate pathway were determined after Tkt RNA interference (RNAi). Results: Despite the complete and specific knockdown of Tkt expression, GVBD occurred and meiosis was arrested at the metaphase I (MI) stage. The arrested oocytes exhibited spindle loss, chromosomal aggregation, and declined maturation promoting factor and mitogen-activated protein kinase activities. The modified expression of two enzymes in the pentose phosphate pathway, Prps1 and Rbks, after Tkt RNAi and decreased maturation rates were amended when ribose-5-phosphate was supplemented in the culture medium, suggesting that the Tkt and pentose phosphate pathway are important for the maturation process. Conclusion: We concluded that Tkt and its associated pentose phosphate pathway play an important role in the MI-MII transition of the oocytes' meiotic cell cycle, but not in the process of GVBD.

• Journal of Microbiology and Biotechnology
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• 제25권2호
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• pp.296-301
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• 2015

Chronic kidney diseases are based on uncontrolled immunological and inflammatory responses to pathophysiological renal circumstances such as glomerulonephritis, which is caused by immunological mechanisms of glomerular inflammation with increased production of renal pro-inflammatory cytokines. Pentoxifylline (PTX) exhibits anti-inflammatory properties by inhibiting cytokine and chemokine production through aggregation of erythrocytes and thrombocytes. We synthesized a series of 1,7-substituted methylxanthine derivatives by the Traube purine reaction, and the formation of purine ring was completed through nitrosation, a reduction of the nitroso to the amine by catalytic hydrogenation as derivatives of PTX. Then we studied biological activities such as renal anti-inflammatory effects of the synthesized compounds in the production of cytokines such as nitric oxide (NO), interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α) and of chemokines such as monocyte chemoattractant protein-1 and IL-8 in Raw 264.7 and HK-2 cells. Renal antiinflammatory activities of this novel series of N-1 and N-7-substituted methylxanthine showed that the N-7 methyl-group-substituted analogs (S7b) showed selective 61% and 77% inhibition of the production of NO and IL-8. The other replacement of the N-1-(CH₂)₄COCH₃ roup, as in the case of compound S6c, also showed an effective 50% and 77% inhibition of TNF-α and IL-8 production in LPS-stimulated Raw 264.7 and HK-2 cells.

• Bulletin of the Korean Chemical Society
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• 제28권12호
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• pp.2283-2287
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• 2007

Amyloid fibril formation of amyloid β/A4 protein (Aβ) is critical to understand the pathological mechanism of Alzheimer's disease and develop controlling strategy toward the neurodegenerative disease. For this purpose, dequalinium (DQ) has been employed as a specific modifier for Aβ aggregation and its subsequent cytotoxicity. In the presence of DQ, the final thioflavin-T binding fluorescence of Aβ aggregates decreased significantly. It was the altered morphology of Aβ aggregates in a form of the bundles of the fibrils, distinctive from normal single-stranded amyloid fibrils, and the resulting reduced β-sheet content that were responsible for the decreased fluorescence. The morphological transition of Aβ aggregates assessed with atomic force microscope indicated that the bundle structure observed with DQ appeared to be resulted from the initial multimeric seed structure rather than lateral association of preformed single-stranded fibrils. Investigation of the seeding effect of the DQ-induced Aβ aggregates clearly demonstrated that the seed structure has determined the final morphology of Aβ aggregates as well as the aggregative kinetics by shortening the lag phase. In addition, the cytotoxicity was also varied depending on the final morphology of the aggregates. Taken together, DQ has been considered to be a useful chemical probe to control the cytotoxicity of the amyloid fibrils by influencing the seed structures which turned out to be central to develop therapeutic strategy by inducing the amyloid fibrils in different shapes with varied toxicities.

• Asian-Australasian Journal of Animal Sciences
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• 제19권10호
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• pp.1508-1513
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• 2006

Myostatin (MSTN) and insulin-like growth factors (IGFs) are a known inhibitor and stimulators of proliferation and differentiation of muscle cells, respectively. The present study was performed to investigate the relationship of MSTN-induced growth inhibition to expression of the IGF system components and myogenin, a muscle cell-specific transcription factor, in rat L6 myoblasts. The L6 cells transfected with a green fluorescent protein-MSTN plasmid expression construct had a 47% less cell number than mock-transfected cells after 3-d serum-free culture, accompanied by delayed differentiation which was suggested by inhibited aggregation of cells. Moreover, cells transfected with the expression construct had decreased expression of IGF-II and myogenin genes, but not IGF-I or its receptor genes, as examined by reverse transcription-polymerase chain reaction. The reduced mitosis of the L6 cells transfected with the MSTN-expression construct increased following an addition of either IGF-I or IGF-II to the culture medium, but not to the level of mock-transfected cells. By contrast, myogenin gene expression in these cells increased after the addition of either IGF to the level of mock-transfected cells. Collectively, these results suggest that the inhibitory effect of MSTN on L6 cell proliferation and differentiation is likely to be partly mediated by serially suppressed expression of IGF-II and myogenin genes, not IGF-I gene.

• BMB Reports
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• 제43권3호
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• pp.176-181
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• 2010

The primary sigma factor ($\sigma^{A}$) of Staphylococcus aureus, a potential drug target, was little investigated at the structural level. Using an N-terminal histidine-tagged $\sigma^{A}$ (His-$\sigma^{A}$), here we have demonstrated that it exits as a monomer in solution, possesses multiple domains, harbors primarily $\alpha$-helix and efficiently binds to a S. aureus promoter DNA in the presence of core RNA polymerase. While both N- and C-terminal ends of His-$\sigma^{A}$ are flexible in nature, two Trp residues in its DNA binding region are buried. Upon increasing the incubation temperature from 25$^{\circ}$ to 40$^{\circ}C$, $\sim$60% of the input His-$\sigma^{A}$ was cleaved by thermolysin. Aggregation of His-$\sigma^{A}$ was also initiated rapidly at 45$^{\circ}C$. From the equilibrium unfolding experiment, the Gibbs free energy of stabilization of His-$\sigma^{A}$ was estimated to be +0.70 kcal $mol^{-1}$. The data together suggest that primary sigma factor of S. aureus is an unstable protein. Core RNA polymerase however stabilized $\sigma^{A}$ appreciably.

• Journal of Microbiology and Biotechnology
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• 제20권3호
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• pp.542-549
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• 2010

In the present study, overexpression, purification, and characterization of Aeropyrum pernix K1 chaperonin B in E. coli were investigated. The chaperonin $\beta$-subunit gene (ApCpnB, 1,665 bp ORF) from the hyperthermophilic archaeon A. pernix K1 was amplified by PCR and subcloned into vector pET21a. The constructed pET21a-ApCpnB (6.9 kb) was transformed into E. coli BL21 Codonplus (DE3). The transformant cell successfully expressed ApCpnB, and the expression of ApCpnB (61.2 kDa) was identified through analysis of the fractions by SDS-PAGE (14% gel). The recombinant ApCpnB was purified to higher than 94% by using heat-shock treatment at $90^{\circ}C$ for 20 min and fast protein liquid chromatography on a HiTrap Q column step. The purified ApCpnB showed ATPase activity and its activity was dependent on temperature. In the presence of ATP, ApCpnB effectively protected citrate synthase (CS) and alcohol dehydrogenase (ADH) from thermal aggregation and inactivation at $43^{\circ}$ and $50^{\circ}$, respectively. Specifically, the activity of malate dehydrogenase (MDH) at $85^{\circ}$ was greatly stabilized by the addition of ApCpnB and ATP. Coexpression of pro-carboxypeptidase B (pro-CPB) and ApCpnB in E. coli BL21 Codonplus (DE3) had a marked effect on the yield of pro-CPB as a soluble and active form, speculating that ApCpnB facilitates the correct folding of pro-CPB. These results suggest that ApCpnB has both foldase and holdase activities and can be used as a powerful molecular machinery for the production of recombinant proteins as soluble and active forms in E. coli.

• The Journal of Korean Physical Therapy
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• 제21권2호
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• pp.109-116
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• 2009

Purpose: $17\beta$-estradiol is the most active endogenous estrogen, which is related to favorable changes in the plasma lipid profile, to relaxation of the coronary vessels, and to a decrease in platelet aggregation and vascular smooth muscle cell migration. However, although the beneficial effect of estrogens on plasma lipoproteins (ie, lowering low-density lipoprotein and increasing high-density lipoprotein cholesterol) contributes to cardiovascular protection, it does not fully account for the protective effect, particularly in the application of physical therapy, including low frequency electrical stimulation. Methods: The aim of this study was to demonstrate the inhibition of stressors, such as endothelin-1 (ET-1), serotonin (5-hydroxytryptamine, 5-HT), prostaglandin $F2\alpha$ ($PGF2\alpha$), and a protein kinase C (PKC) activator 12-deoxyphorbol 13-isobutyrate (DPB), induced isometric tension by $17\beta$-estradiol in vascular smooth muscle strips, respectively. In addition, the effects of low frequency electrical stimulation at the meridian points (CV-3, -4, Ki-12, SP-6, LR-3, BL-25, -28, -32, -52) on the indirect antihypertensive effect were examined by monitoring the changes in the serum $17\beta$-estradiol concentration in healthy volunteers. Results: Isometric tension analysis showed that the responses of inhibited tension by $17\beta$-estradiol were similar to the same stressors in rat aortic smooth muscle strips. Furthermore, although the continued amplitude modulation (AM) type of electrical stimulation was not increased significantly by electrical stimulation, the current of the frequency modulation (FM) type of low frequency electrical stimulation increased the serum $17\beta$-estradiol concentration in normal volunteers. Conclusion: These results, in part, suggest that $17\beta$-estradiol has the capacity to supress stressor-induced muscle tension, and electrical stimulation, particularly current of the FM type, has a modulatory effect on the sex steroid hormones, particularly $17\beta$-estradiol, in healthy volunteers.

• 자연과학논문집
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• 제19권1호
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• pp.45-55
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• 2008

아세틸콜린에스터라아제(AChE)저해제에 의한 아세틸콜린 분해의 억제는 알츠하이머 질병의 환자들을 위한 치료 방법 중의 하나이다. 새로운 AChE 저해제를 개발하여 항 치매성 대체 의약품생산에 응용하기 위해, 전보에서는 다양한 곡류와 두류로부터 AChE 저해활성이 우수한 시료로 수수를 선발하여 추출 최적조건을 조사하였고 영양성 및 생리기능성 등을 조사하였다. 본 연구에서는 AChE 저해제를 함유한 수수 메탄올 추출물의 물리화학적 성질과 안정성을 조사하였다. 수수 메탄올추출물은 물, 메탄올과 DMSO등에 잘 녹았고, 215nm과 282nm의 최대흡수파장을 갖고 있었다. 또한, 추출물은 20-$100^{\circ}C$와 pH 2.0-10.0에서 1시간동안 안정 하였다. 수수 메탄올추출물을 이용하여 시제품을 제조한 후 $20^{\circ}C$와 $40^{\circ}C$에서 8주간 저장하였을 때 모두 생균수와 pH에 변화가 없이 안정하였고 기호도도 우수하게 유지되었다.

• Bulletin of the Korean Chemical Society
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• 제33권1호
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• pp.227-232
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• 2012

Galectins are generally believed to be potential candidates for use in the development of novel antiinflammatory agents or as selective modulators of the immune response. In particular, galectin-9 exhibits some of the extracellular functions, including cell aggregation, adhesion, chemoattraction, activation, and apoptosis. Tl-galectin (Tl-gal, galectin-9 homologue gene) was isolated from an adult worm of the Toxascaris leonina. The full-length Tl-gal gene, which was incorporated into pET-28a, was overexpressed in E. coli and purified by nickel affinity and gel filtration chromatographies. The purified Tl-gal was crystallized using the hangingdrop vapor-diffusion method. The crystal belonged to the tetragonal space group $P4_1$, with unit-cell parameters of a = b = $75.7\AA$ and c = $248.4\AA$. The crystals were obtained at $20^{\circ}C$ and diffracted to a resolution of $3.0\AA$. The asymmetric unit contained four molecules of Tl-gal, which gave a crystal volume per protein mass (Vm) of $2.8\AA^3Da^{-1}$ and a solvent content of 54.1%.

• Korean Chemical Engineering Research
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• 제46권4호
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• pp.722-726
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• 2008

실리카 microsphere는 HPLC를 위한 흡착 충진제와 같은 용도로 사용하기에 적합한 혁신적인 소재로 널리 알려져 있다. microsphere을 기능성고분자나 금속, 생리활성 물질과 같은 특정한 성질을 지닌 물질로 표면 개질시키므로 다양한 용도로 활용할 수 있다. 콜라겐은 생체조직을 구성하는 기본 단백질로 생체적합성이 뛰어난 물질로 주목받고 있는 기능성 소재이다. 본 연구에서는 50% 이상의 세공부피를 지닌 다공성 silica microsphere를 고분자 응집법인 PICA 법을 이용하여 colloidal silica로부터 제조하고 콜라겐 hydrogel을 사용하여 표면 개질시키므로 생체적합성을 증진시키는 방법을 연구하였다. 실리카/콜라겐 microsphere 에 은 나노입자를 담지시킨 microsphere 복합체를 만들고 특성을 조사하므로 생체소재로의 활용 가능성을 조사하였다.