• 제목/요약/키워드: intact cells

검색결과 444건 처리시간 0.03초

Genetic Transformation of the Yeast Dekkera/Brettanomyces bruxellensis with Non-Homologous DNA

  • Miklenic, Marina;Stafa, Anamarija;Bajic, Ana;Zunar, Bojan;Lisnic, Berislav;Svetec, Ivan-Kresimir
    • Journal of Microbiology and Biotechnology
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    • 제23권5호
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    • pp.674-680
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    • 2013
  • Yeast Dekkera/Brettanomyces bruxellensis is probably the most common contaminant in wineries and ethanol production processes. The considerable economic losses caused by this yeast, but also its ability to produce and tolerate high ethanol concentrations, make it an attractive subject for research with potential for industrial applications. Unfortunately, efforts to understand the biology of D. bruxellensis and facilitate its broader use in industry are hampered by the lack of adequate procedures for delivery of exogenous DNA into this organism. Here we describe the development of transformation protocols (spheroplast transformation, LiAc/PEG method, and electroporation) and report the first genetic transformation of yeast D. bruxellensis. A linear heterologous DNA fragment carrying the kanMX4 sequence was used for transformation, which allowed transformants to be selected on plates containing geneticin. We found the spheroplast transformation method using 1M sorbitol as osmotic stabilizer to be inappropriate because sorbitol strikingly decreases the plating efficiency of both D. bruxellensis spheroplast and intact cells. However, we managed to modify the LiAc/PEG transformation method and electroporation to accommodate D. bruxellensis transformation, achieving efficiencies of 0.6-16 and 10-20 transformants/${\mu}g$ DNA, respectively. The stability of the transformants ranged from 93.6% to 100%. All putative transformants were analyzed by Southern blot using the kanMX4 sequence as a hybridization probe, which confirmed that the transforming DNA fragment had integrated into the genome. The results of the molecular analysis were consistent with the expected illegitimate integration of a heterologous transforming fragment.

평택 희곡리 출토목재 부후형태의 전자현미경적 관찰 (Electron Microscopic Observation on the Decay of the Waterlogged Woods Excavated from Higokri, Pyungtaek)

  • 김수철;박원규
    • 보존과학회지
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    • 제23권
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    • pp.67-72
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    • 2008
  • 경기도 평택 희곡리 토탄층(약 5,700~5,900년전(前))에서 수침상태로 출토된 물푸레나무속 1종과 오리나무속 1종을 대상으로 전자현미경 관찰을 통하여 수종에 따른 분해정도와 형태를 조사하였다. 관찰결과 두 수종 모두 연부후와 박테리아에 의한 세포벽 공격형태와 분해형태는 거의 유사하였다. $S_2$층부터 복합세포간층 방향으로 점진적으로 불규칙하게 분해하는 침식형 박테리아와 목재세포벽을 분해하여 $S_2$층에 터널을 만드는 터널형 박테리아가 관찰되었다. 침식형 박테리아는 종종 연부후균에 의한 공동과 유사하지만 크기가 작은 공동을 형성하였다. 도관요소를 제외한 다른 세포들의 세포벽은 극심히 분해되었지만 세포벽의 외형은 수분으로 인해 그 고유한 형태를 유지하고 있었다. 해양천공충의 식흔 흔적이 발견되지 않고 연부후균과 박테리아에 의한 분해 형태만 발견되어 평택희곡리 토탄지역 형성 당시인 5,700~5,900여년전이 온난화에 의한 홀로세 기후최적기 기간중이기 때문에 서해안 해수면 상승에 의해 해수에 침수되었을 것이라는 가설은 사실이 아닌 것으로 해석되었다.

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Modulation of Presynaptic GABA Release by Oxidative Stress in Mechanically-isolated Rat Cerebral Cortical Neurons

  • Hahm, Eu-Teum;Seo, Jung-Woo;Hur, Jin-Young;Cho, Young-Wuk
    • The Korean Journal of Physiology and Pharmacology
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    • 제14권3호
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    • pp.127-132
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    • 2010
  • Reactive oxygen species (ROS), which include hydrogen peroxide ($H_2O_2$), the superoxide anion (${O_2}^-{\cdot}$), and the hydroxyl radical ($OH{\cdot}$), are generated as by-products of oxidative metabolism in cells. The cerebral cortex has been found to be particularly vulnerable to production of ROS associated with conditions such as ischemia-reperfusion, Parkinson's disease, and aging. To investigate the effect of ROS on inhibitory GABAergic synaptic transmission, we examined the electrophysiological mechanisms of the modulatory effect of $H_2O_2$ on GABAergic miniature inhibitory postsynaptic current (mIPSCs) in mechanically isolated rat cerebral cortical neurons retaining intact synaptic boutons. The membrane potential was voltage-clamped at -60 mV and mIPSCs were recorded and analyzed. Superfusion of 1-mM $H_2O_2$ gradually potentiated mIPSCs. This potentiating effect of $H_2O_2$ was blocked by the pretreatment with either 10,000-unit/mL catalase or $300-{\mu}M$ N-acetyl-cysteine. The potentiating effect of $H_2O_2$ was occluded by an adenylate cyclase activator, forskolin, and was blocked by a protein kinase A inhibitor, N -(2-[p-bromocinnamylamino] ethyl)-5-isoquinolinesulfonamide hydrochloride. This study indicates that oxidative stress may potentiate presynaptic GABA release through the mechanism of cAMP-dependent protein kinase A (PKA)-dependent pathways, which may result in the inhibition of the cerebral cortex neuronal activity.

백작약 열수추출물의 피부노화 억제 효과(I) -염증 및 피부장벽 손상 완화 효과를 중심으로- (Inhibitory Effects of Peonia japonica Water Extract on Skin Aging (I) - Focussed on Alleviative Effects of Inflammation and Skin Barrier Damage -)

  • 권미화;민경진;김영철
    • Environmental Analysis Health and Toxicology
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    • 제24권2호
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    • pp.159-167
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    • 2009
  • To investigate the alleviative effects of Peonia japonica water extract(PJWE) on inflammation and skin barrier damage, both the irradiation of UVB and the application of squalene monohydroperoxide (Sq-OOH) to the backs of hairless mice were performed for 4 weeks. And at the same time experimental materials were applied topically. The skin erythema indices for the positive control (PC, 0.01% retinoic acid) and experimental (E, PJWE) groups were lower than that of the control (C) group. Whereas both the lipid and water capacities for the PC and E groups were higher than those of the C group. Epidermis and dermis of the C group were remarkably thickened in comparison with the PC and E groups. Relatively much less number of inflammatory cells, including lymphocytes, neutrophils and macrophages were found in dermis of the PC and E groups compared with the C group. Lipid lamellae of the C group were broken severely showing an irregular arrangement and lipid content was much reduced. Whereas those of the PC and E groups were almost intact with a regular arrangement, which were similar to that of the N group. Taken the results all together, it was confirmed that PJWE could be effective natural herbal material for the alleviation of inflammation and skin barrier damage in hairless mice skin which were induced by UVB and Sq-OOH.

Mechanism of Acetylcholine-induced Endothelium-dependent Relaxation in the Rabbit Carotid Artery by M3-receptor Activation

  • Song, Yong-Jin;Kwon, Seong-Chun
    • The Korean Journal of Physiology and Pharmacology
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    • 제8권6호
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    • pp.313-317
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    • 2004
  • The present study were designed to characterize the action mechanisms of acetylcholine (ACh)-induced endothelium-dependent relaxation in arteries precontracted with high $K^+$(70 mM). For this, we simultaneously measured both muscle tension and cytosolic free $Ca^{2+}$ concentration $([Ca^{2+}]_i)$, using fura-2, in endothelium-intact, rabbit carotid arterial strips. In the artery with endothelium, high $K^+$ increased both $[Ca^{2+}]_i$ and muscle tension whereas ACh $(10{\mu}M)$ significantly relaxed the muscle and increased $[Ca^{2+}]_i$. In the presence of $N^G$-nitro-L-arginine (L-NAME, 0.1 mM), ACh increased $[Ca^{2+}]_i$ without relaxing the muscle. In the artery without endothelium, high $K^+$ increased both $[Ca^{2+}]_i$ and muscle tension although ACh was ineffective. 4-DAMP (10 nM) or atropine $(0.1{\mu}M)$ abolished ACh-induced increase in $[Ca^{2+}]_i$ and relaxation. The increase of $[Ca^{2+}]_i$ and vasorelaxation by ACh was siginificantly reduced by either $3{\mu}M$ gadolinium, $10{\mu}M$ lanthanum, or by $10{\mu}M$ SKF 96365. These results suggest that in rabbit carotid artery, ACh-evoked relaxation of 70 mM $K^+$-induced contractions appears to be mediated by the release of NO. ACh-evoked vasorelaxation is mediated via the $M_3$ subtype, and activation of the $M_3$ subtype is suggested to stimulate nonselective cation channels, leading to increase of $[Ca^{2+}]_i$ in endothelial cells.

탈세포화 기법을 이용한 돼지 바이오 스캐폴드: 환자 맞춤형 장기재생을 위한 역분화 줄기세포 동물모델 (Decellularized Bioscaffold of Pig Organs: A Tool for Patient-specific Organogenesis Using Induced Pluripotent Stem Cells)

  • 박경미;곽호현;남현숙;박인철;전용환;박성민;이승태;우제석;우흥명
    • 한국임상수의학회지
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    • 제28권1호
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    • pp.57-62
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    • 2011
  • The shortage of transplantable kidneys has many efforts to regenerate bioartificial kidneys using transgenic animals and diverse kinds of scaffolds which are important tools for cell seeding. However, there are many limitations for clinical applications so far. Recently, decellularized bioscaffolds using animal organs come into spotlight because of its many superior advantages. In current study, we produced decellularized kidney bioscaffolds of pig which is an attractive animal as a clinical model for human. We decellularized pig kidneys with 1% SDS detergent solution using peristaltic pump systems for 12h. After decellularization process, the kidney bioscaffolds preserved intact 3D morphology including glomerular structure and almost DNA from pig was entirely removed. In addition, this process could preserve micro vascular network which is necessary for cell survival. Although, additional studies for recellularization and transplantation should be required, the decellular vascularized kidney bioscaffolds might have many potentials for kidney regeneration.

Efficacy of an LED toothbrush on a Porphyromonas gingivalis biofilm on a sandblasted and acid-etched titanium surface: an in vitro study

  • Lee, Hae;Kim, Yong-Gun;Um, Heung-Sik;Chang, Beom-Seok;Lee, Si Young;Lee, Jae-Kwan
    • Journal of Periodontal and Implant Science
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    • 제48권3호
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    • pp.164-173
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    • 2018
  • Purpose: The aim of this study was to evaluate the antimicrobial effect of a newly devised toothbrush with light-emitting diodes (LEDs) on Porphyromonas gingivalis attached to sandblasted and acid-etched titanium surfaces. Methods: The study included a control group, a commercial photodynamic therapy (PDT) group, and 3 test groups (B, BL, and BLE). The disks in the PDT group were placed in methylene blue and then irradiated with a diode laser. The B disks were only brushed, the BL disks were brushed with an LED toothbrush, and the BLE disks were placed into erythrosine and then brushed with an LED toothbrush. After the different treatments, bacteria were detached from the disks and spread on selective agar. The number of viable bacteria and percentage of bacterial reduction were determined from colony counts. Scanning electron microscopy was performed to visualize bacterial alterations. Results: The number of viable bacteria in the BLE group was significantly lower than that in the other groups (P<0.05). Scanning electron microscopy showed that bacterial cell walls were intact in the control and B groups, but changed after commercial PDT and LED exposure. Conclusions: The findings suggest that an LED toothbrush with erythrosine treatment was more effective than a commercial PDT kit in reducing the number of P. gingivalis cells attached to surface-modified titanium in vitro.

동종 탈회골의 크기가 유도골 형성에 미치는 영향에 관한 실험적 연구 (EXPERIMENTAL STUDY OF EFFECT ON INDUCED OSTEOGENESIS ACCORDING TO THE SIZE OF DEMINERALIZED ALLOGENEIC BONE)

  • 방만혁;엄인웅;이동근;민승기
    • Maxillofacial Plastic and Reconstructive Surgery
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    • 제17권4호
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    • pp.350-364
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    • 1995
  • As early as 1889, treatment of ostemyelitis was reported using xenogeneic demineralized bone. In 1965, Urist discovered that demineralized long bone fragment, even when implanted in nonskeletal tissue, would stimulate osteogenesis. The clinical use of demineralized bone of Oral and Maxillofacial surgery is not new. The demineralized bone implants were used for 1) interposition within osteotomy gaps, cystic detects, alveolar clefts ; 2) augmentation, over intact bone surfaces ; 3) construction of new bone within soft tissue. Demineralized bone grafts invokes a induced osteogenesis which is the transformation of host cells into osteoblasts. Demineralized bone has identified several factors that modulate the osteogeneic response : sterilization method, recipient age, particle size etc. Especially, pulverization of bone matrix may enhance its osteoinductive properties, to allow rapid, efficient bridging of large defects. the purpose of the present report was to describe the potential efficacy of demineralized allogeneic bone powder of skull of rabbits as a particle size ; 212 ${\mu}m$, 710 ${\mu}m$, 1 mm each other. Microscopic finding in our experimental studies shown that 710 ${\mu}m$ demineralized bone powder is the most potent osteogenic response, and then 212 ${\mu}m$, 1 mm size. Densitometric analysis shown that density of all group was continue to increase until 4 weeks after operation, and then continue to decrease.

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연구개 및 구개수에 발생한 편평세포암종 (SQUAMOUS CELL CARCINOMA OF THE SOFT PALATE AND UVULA)

  • 김창룡;조규승;김기영;이성훈;이승호;박문성;류선열
    • Maxillofacial Plastic and Reconstructive Surgery
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    • 제18권4호
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    • pp.673-678
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    • 1996
  • 독특한 해부학적 구조와 상기도에서 수행하는 기능으로 인해 수술후에 발음이나 연하에 심각한 문제를 야기할 수 있는 연구개 및 구개수에 발생한 편평세포암종 환자에서, 저자들은 술전 항암화학요법과 외과적 절제술 그리고 6000 cGy의 방사선 조사를 시행하여 치료하였다. 2년 이상 경과한 현재까지 재발의 소견은 관찰되지 않고 비교적 양호한 기능을 보이고 있다.

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구강의 퇴행성 신경집종: 2예 보고 (Ancient schwannoma in oral cavity: a report of two cases)

  • 김나래;정동해;박대송;김동우;이상칠;김성용;임호용;염학열;김현민
    • Journal of the Korean Association of Oral and Maxillofacial Surgeons
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    • 제37권6호
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    • pp.530-534
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    • 2011
  • This paper reports two cases of schwannomas arising from the oral cavity. One is an intraoral ancient schwannoma located at the left cheek, which evolved over a period of 13 years. The tumor was a well-demarcated buccal mass, which was located in the left lower first premolar area, with an obliterated the buccal vestibule, leaving the overlying mucosa intact. The second case was a central intraosseous schwannoma located from the left lower 1st molar periapical area to the left 3rd molar periapical area. Pathologically, the first mass was composed of the spindle shaped tumor cells with wavy nuclei beneath the fibroconnective tissue of the gingiva but second case mass was not. Occasional nuclear pleomorphism was observed but mitosis or necrosis was absent. There were Antoni A and B areas along with strong, diffuse staining with the S-100 protein. Ancient schwannomas were diagnosed. Schwannoma is a slow-growing benign tumor, and an ancient schwannoma that shows cellular atypism is a variant of a schwannoma caused by purely degenerative changes. To date, only limited cases of ancient schwannomas in the oral cavity have been reported.