• Animal cells and systems
• /
• 제5권3호
• /
• pp.253-262
• /
• 2001

Ligand-receptor clustering event is the most important step in leukocyte adhesion and spreading on endothelial cells. Intercellular adhesion molecule-1 (ICAM-1) has been shown to enhance leukocyte adhesion, but the molecular event during the process of adhesion is unclear. To visualize the dynamics of ICAM-1 movement during adhesion, we have engineered stable Chinese hamster ovary cell lines expressing ICAM-1 fused to a green fluorescent protein (IC1_GFP/CHO) and examined them under the fluorescence microscopy. The transfection of IC1_GFP alone in these cells was sufficient to support the adhesion of K562 cells that express $\alpha$L$\beta$2 (LFA-1) integrin stimulated by CBR LFA-1/2 mAb. This phenomenon was mediated by ICAM-1-LFA-1 interactions, as an mAb that specifically inhibits ICAM-1-LFA-1 interaction (RRl/l) completely abolished the adhesion of LFA-1＊ cells to IC1_ GFP/CHO cells. We found that the characteristic of adhesion was followed almost immediately (~10 min) by the rapid accumulation of ICAM-1 on CHO cells at a tight interface between the two cells. Interestingly, ICI_GFP/CHO cells projected plasma membrane and encircled approximately half surface of LFA-1+ cells, as defined by confocal microscopy. This unusual phenomenon was also confirmed on HUVEC after adhesion of LFA-1＊ cells. Neither cytochalasin D nor 2,3-butanedione 2-monoxime an inhibitor of myosin light chain kinase blocked LFA-1-mediated ICAM-1 clustering, indicating that actin cytoskeleton and myosin-dependent contractility are not necessary for ICAM-1 clustering. Taken together, we suggest that leukocyte adhesion to endothelial cells induces specialized form of ICAM-1 clustering that is distinct from immunological synapse mediated by T cell interaction with antigen presenting cells.

• Journal of Ginseng Research
• /
• 제41권3호
• /
• pp.419-427
• /
• 2017

Background: Glycosylation of natural compounds increases the diversity of secondary metabolites. Glycosylation steps are implicated not only in plant growth and development, but also in plant defense responses. Although the activities of uridine-dependent glycosyltransferases (UGTs) have long been recognized, and genes encoding them in several higher plants have been identified, the specific functions of UGTs in planta remain largely unknown. Methods: Spatial and temporal patterns of gene expression were analyzed by quantitative reverse transcription (qRT)-polymerase chain reaction (PCR) and GUS histochemical assay. In planta transformation in heterologous Arabidopsis was generated by floral dipping using Agrobacterium tumefaciens (C58C1). Protein localization was analyzed by confocal microscopy via fluorescent protein tagging. Results: PgUGT72AL1 was highly expressed in the rhizome, upper root, and youngest leaf compared with the other organs. GUS staining of the promoter: GUS fusion revealed high expression in different organs, including axillary leaf branch. Overexpression of PgUGT72AL1 resulted in a fused organ in the axillary leaf branch. Conclusion: PgUGT72AL1, which is phylogenetically close to PgUGT71A27, is involved in the production of ginsenoside compound K. Considering that compound K is not reported in raw ginseng material, further characterization of this gene may shed light on the biological function of ginsenosides in ginseng plant growth and development. The organ fusion phenotype could be caused by the defective growth of cells in the boundary region, commonly regulated by phytohormones such as auxins or brassinosteroids, and requires further analysis.

• Journal of Ginseng Research
• /
• 제43권4호
• /
• pp.645-653
• /
• 2019

Background: Cytochrome P450 enzymes catalyze a wide range of reactions in plant metabolism. Besides their physiological functions on primary and secondary metabolites, P450s are also involved in herbicide detoxification via hydroxylation or dealkylation. Ginseng as a perennial plant offers more sustainable solutions to herbicide resistance. Methods: Tissue-specific gene expression and differentially modulated transcripts were monitored by quantitative real-time polymerase chain reaction. As a tool to evaluate the function of PgCYP736A12, the 35S promoter was used to overexpress the gene in Arabidopsis. Protein localization was visualized using confocal microscopy by tagging the fluorescent protein. Tolerance to herbicides was analyzed by growing seeds and seedlings on Murashige and Skoog medium containing chlorotoluron. Results: The expression of PgCYP736A12 was three-fold more in leaves compared with other tissues from two-year-old ginseng plants. Transcript levels were similarly upregulated by treatment with abscisic acid, hydrogen peroxide, and NaCl, the highest being with salicylic acid. Jasmonic acid treatment did not alter the mRNA levels of PgCYP736A12. Transgenic lines displayed slightly reduced plant height and were able to tolerate the herbicide chlorotoluron. Reduced stem elongation might be correlated with increased expression of genes involved in bioconversion of gibberellin to inactive forms. PgCYP736A12 protein localized to the cytoplasm and nucleus. Conclusion: PgCYP736A12 does not respond to the well-known secondary metabolite elicitor jasmonic acid, which suggests that it may not function in ginsenoside biosynthesis. Heterologous overexpression of PgCYP736A12 reveals that this gene is actually involved in herbicide metabolism.

• ALGAE
• /
• 제34권2호
• /
• pp.71-90
• /
• 2019

A new encrusting red alga was found growing abundantly on glass debris items that drifted ashore along the coasts of Oregon and Washington. These included discarded fluorescent tubes, incandescent light bulbs, capped liquor bottles, and ball-shaped fishing-net floats. Field collections and unialgal cultures of the alga revealed that it consisted of two morphological phases: a young loosely aggregated turf and a mature consolidated mucilaginous crust. The turf phase consisted of a basal layer of globose cells that produced erect, rarely branched, uniseriate to multiseriate filaments up to $500{\mu}m$ long with closely spaced cells lacking pit-plugs. These filaments expanded in size from their bases to their tips and released single cells as spores. At maturity, a second phase of growth occurred that produced a consolidated crust, up to $370{\mu}m$ thick. It consisted of a basal layer of small, tightly appressed ellipsoidal-to-elongate cells that generated a mucilaginous perithallial matrix containing a second type of filament with irregularly spaced cells often undergoing binary division. At the matrix surface, the original filaments continued to grow and release spores but often also eroded. Individual cells, examined using confocal microscopy and SYBR Green staining, were found to contain a central nucleus, a single highly lobed peripheral chloroplast without a pyrenoid, and numerous chloroplast nucleoids. Morphological data from field and culture isolates and molecular data (rbcL, psbA, and SSU) show that this alga is a new genus and species which we name Viator vitreocola, "a traveller on glass."

• 센서학회지
• /
• 제31권3호
• /
• pp.175-179
• /
• 2022

The physical properties of biomaterials are important for their isolation and separation from body fluids. In particular, the precise evaluation of the multi-physical properties of single biomolecules is essential in that the correlation between physical and biological properties of specific biomolecule. However, the majority of scientific equipment, can only determine specific-physical properties of single nanoparticles, making the evaluation of the multi-physical properties difficult. The improvement of analytical techniques for the evaluation of multi-physical properties is therefore required in various research fields. In this study, we developed a motion-tracking algorithm to evaluate the multi-physical properties of single-nanoparticles by analyzing their behavior. We observed the Brownian motion and electric-field-induced drift of fluorescent nanoparticles injected in a microfluidic chip with two electrodes using confocal microscopy. The proposed algorithm is able to determine the size of the nanoparticles by i) removing the background noise from images, ii) tracking the motion of nanoparticles using the circular-Hough transform, iii) extracting the mean squared displacement (MSD) of the tracked nanoparticles, and iv) applying the MSD to the Stokes-Einstein equation. We compared the evaluated size of the nanoparticles with the size measured by SEM. We also determined the zeta-potential and surface-charge density of the nanoparticles using the extracted electrophoretic velocity and the Helmholtz-Smoluchowski equation. The proposed motion-tracking algorithm could be employed in various fields related to biomaterial analysis, such as exosome analysis.

• Journal of Yeungnam Medical Science
• /
• 제23권1호
• /
• pp.26-35
• /
• 2006

전염증성 사이토카인의 분비를 조절하는 전사인자인 NF-${\kappa}B$는 염증성 장질환 환자의 대장 점막에서 발현이 증가되어 있다고 보고하고 있으며 이를 억제하여 대장의 염증을 억제하려는 연구가 진행 중이다. 루테올린은 다양한 한약제에 포함되어 있는 플라보노이드 중 하나로 항염증 및 항산화작용이 있다고 알려져 있으며 LPS 자극된 대식세포에서 TNF-${\alpha}$ 분비의 억제 뿐만 아니라 전사인자인 NF-kB의 발현을 억제하여 항염증작용을 하는 것으로 알려져 있다. 본 연구에서는 DSS을 이용한 염증성 장질환 모델에서 루테올린의 장염의 치료효과를 보고자 하였다. C57BL/6 NF-${\kappa}B^{EGFP}$ 쥐에게 2.5% DSS를 투여하여 장염을 유발하였으며 치료군(n=6)에서는 매일 루테올린(1 mg/kg, vol 0.1 ml)을 비위관을 통해 경구투여 하였으며, 비치료군(n=6)에서는 매일 같은 양의 vehicle(vol 0.1 ml)를 투여하여 정상대조군(n=6)과 비교하였다. 실험 기간 동안 질병활성도를 기록하였으며, 투약 6일 후 모든 쥐를 희생하여 대장을 분리하여 길이를 측정하고, 조직검사를 시행하였다. 또한 대장점막조직을 배양하여 m IL-12 p40의 분비를 측정하였고, 공초점 형광현미경하에서 EGFP 발현의 정도를 관찰하였다. 질병의 활성도의 관찰에서 치료군에서 2.7점, 비치료군에서 2.5점으로 양군 사이에 유의한 차이가 없어 루테올린에 대한 장염예방효과는 없었다. 또한 치료군과 비치료군에서 대장점막의 m IL-12 p40의 분비 각각 $535.2{\pm}198.2pg/ml$, $412.5{\pm}48.2pg/ml$ 로 양군 사이의 차이는 없었다. 흥미롭게도 EGFP 발현은 치료군에서 오히려 높게 나타났으며, 공초점 형광현미경관찰에서 대부분의 고유근층하 대식세포에서 증가되어 있음을 알 수 있었다. 이상의 결과로 루테올린의 경구 투여는 DSS 유발 염증성 장질환 모델에서 대장 점막 염증을 억제하는 대체 요법으로써의 치료효과는 관찰할 수 없었으며, 향후 항염증작용이 있다고 알려져있는 플라보노이드 물질의 개발에 신중함이 있어야 할 것으로 사료된다.

• 폴리머
• /
• 제29권6호
• /
• pp.543-548
• /
• 2005

이중층 미립구는 단일층 미립구에 비해서 낮은 초기 방출량과 국소 지항성, 및 약물 방출량 제어 등의 장점 을 갖고 있다. 그러나 포접 방식이 까다롭고 2단계 이상의 제조 과정이 필요하며 특히 미립구 제조 시 크기 조절이 어렵다는 단점을 갖고 있다. 따라서 본 연구에서는 락타이드글리콜라이드 공중합체(PLGA)와 덱스트란의 서로 다른 고분자를 초고주파 분쇄 유무에 따른 수중유형(O/W) 용매 증발법을 이용하여 이중층 미립구를 제조하였다. 또한 PLGA의 농도에 따라 미립구 크기의 변화를 연구하였다. 이중층 미립구는 전자주사현미경, 동초점 형광 레이저현미경(CFLM), 캠스코프를 이용하여 조사하였다. 제조된 이중층 미립구의 외부층이 매끄러운 구형의 형태를 나타냄을 확인할 수 있었으며, 절단시 내부층과 외부층의 형태를 확실히 구분할 수 있었다. 이에 외부층과 내부층의 구성 물질을 확인하고자 플루오르신-5-이소시아네이트-덱스트란(FITC-덱스트란)을 이용해 CFLM을 관찰한 결과 형광을 띠는 덱스트란으로 구성된 내부층과 형광을 띠지 않는 PLGA 외부층을 관찰하였다. 또한 PLGA의 함량에 따른 미립구의 크기는 전체적으로 증가하는 경향을 확인하였다. 이와 같은 결과로부터 비교적 간단한 수중유형 용매 증발법을 이용하여 PLGA와 덱스트란의 이중층 미립구의 제조가 가능함을 확인할 수 있었다.

• Journal of Periodontal and Implant Science
• /
• 제29권2호
• /
• pp.415-433
• /
• 1999

Implant stability is the key to long-term successful outcome for osseointegrated implants. To evaluate the initial healing response of bone around HA-coated implants without primary bone contact. 21 HA-coated thread type implants(STERI-OSS?) were placed in the femurs of 5 mongrel dogs, about 1-year old. Implants, 8 mm in length and 3.8mm(experimental 1group), 5.0mm(experimental 2group) and 6.0mm(control group) in diameter, were inserted after 3 holes of 6.0mm in diameter and 10mm in depth were prepared in the surgical sites each dog. Implants were supported by only nonresorbable membrane($Teflon^{(R)}$), in order to prevent the ingrowth of upper soft tissue into the gap between bone and implant, and to maintain each implant to be positioned in the center of the drilled hole. 9 implants with different diameters were inserted in 3 dogs for histologic observation, and 12 implants were inserted in 2 dogs for mobility test and removal torque test. Fluorescent dyes were injected for the observation of new bone formation in order of $Terramycin^{(R)}$, Arizarin $Red^{(R)}$, and $Calcein^{(R)}$ at an interval of 2 weeks. 3 dogs were sacrificed for histologic observation at 4, 8, and 12-week after placement. Light microscopy and confocal laser scanning microscopy were used to qualitatively characterize the bone around HA-coated implant. 2 dogs were sacrificed for mobility test($Periotest^{(R)}$, Simens AG, Bensheim, Germany) and removal torque test($Autograph^{(R)}$ AGS-1000D series, Japan) at 8 and 12-week after placement The results were as follows: 1. Histologic observation showed that osseointegration occurred to both control and experimental groups as time lapse, but delayed bone healing was revealed in 3.8mm group (experimental 1group), compared to contrtol group and 5.0mm group (experimental 2group). 2. The mobility test showed that the experimental groups had no distinguishable movement during experimental periods of 8 and 12-week, and there was no difference in mobility depending on the gap between bone and implant, and time lapse. 3. The removal torque forces were increased depended on the gaps decreasing between bone and implant, and time lapse. The results suggest that HA-coated implant without primary bone contact, based on guided bone regeneration could obtain its stability in all experimental groups as time lapse, but bone healing was delayed in experimental group of 3.8mm. And the results suggested that studies on correlationship between mobility test and removal torque test for implant stability would be necessary.

• 대한화장품학회지
• /
• 제34권4호
• /
• pp.311-316
• /
• 2008

본 연구에서는 모발의 내부 및 외부구조에 자외선이 미치는 영향에 관하여 조사를 수행하였다. 자외선이 조사된 모발의 형태학적 및 화학적 구조의 변화를 알아보기 위해 주사전자현미경(SEM), 투과전자현미경(TEM), 공초점레이져주사현미경(CLSM) 등을 사용하였다. SEM상에서 자외선이 조사된 모발은 자외선에 의한 화학적 산화반응에 의한 표면이 거칠고 부풀어진 형태를 보였다. 이황화결합(disulfide bond)의 산화에 의한 시스테인산(cysteic acid)의 분해로 팽윤부위가 관찰되었다. 또한 자외선 조사에 의해 표면색의 변화를 나타내었다. TEM 분석결과 버진헤어(virgin hair)의 경우에는 매끄러운 표면구조를 나타내었지만, 자외선 조사모발에 있어서는 거칠고 모수질층이 갈라진 형태를 보여주었다. 모피질층에 존재하는 멜라닌 입자의 크기가 감소하고 소실되는 것이 관찰 되었다. CLSM에 의해 3차원적 구조의 입체영상을 얻을 수 있었으며, 광학분할에 의한 버진헤어는 강한 형광을 나타내었지만, 자외선 조사모발은 낮은 형광을 보여주었다. 이는 버진헤어의 경우 자동형광을 나타낼 수 있는 방향족 아미노산이 많이 존재하는데 비해 자외선 조사모발은 방향족 아미노산이 파괴되어 낮게 나온 것으로 여겨진다.

• Journal of Pharmaceutical Investigation
• /
• 제31권4호
• /
• pp.241-256
• /
• 2001

Alginate microspheres, containing fluorescein isothiocyanate-bovine serum albumin (FITC-BSA) or green fluorescent protein (GFP) were prepared and used as a model drug to develop the oral vaccine delivery system. The alginate microspheres were coated with poly-L-lysine or chitosan. Two methods, w/o-emulsion and spray, were used to prepare alginate microspheres. To optimize preparation conditions, effects of several factors on the particle size and particle morphology of microsphere, and loading efficiency of model antigen were investigated. In both preparation methods, the particle size and the loading efficiency were enhanced when the concentration of sodium alginate increased. In the w/o-emulsion preparation method, as the concentration of Span 80 was increased from 0.5% to 2%, the particle size was decreased, but the loading efficiency was increased. The higher the emulsification speed was, the smaller the particle size and loading efficiency were. The concentration of calcium chloride did not show any effect on the particle size and loading efficiency. In the spray preparation method, the particle size was increased as the nozzle pressure $(from\;1\;kgf/m^2\;to\;3\;kgf/m^2)$ and spray rate was raised. Increasing calcium chloride concentration (<7%) decreased the particle size, in contrast to no effect of calcium chloride concentration on the w/o-emulsion preparation method. Alginate microspheres prepared by two methods were different in the particle size and loading efficiency, the particle size of microspheres prepared by the spray method was about $2-6\;{\mu}m$, larger than that prepared by the w/o emulsion method $(about\;2{\mu}m)$, and the loading efficiency was also higher with spray method. Furthermore, drying process for the microspheres prepared by the spray was simpler and easier, compared with the w/o emulsion preparation. Therefore, the spray method was chosen to prepare alginate microspheres for further experiments. Release pattern of FITC-BSA in alginate microspheres was evaluated in simulated intestinal fluid and PBS (phosphate buffered saline). Dissolution rate of FITC-BSA from alginate/chitosan microsphere was lower than that from alginate microsphere and alginate/poly-L-lysine microsphere. By confocal laser scanning microscope, it was revealed that alginate/FITC-poly-L-lysine microspheres were present in close apposition epithelium of the Peyer's patches of rabbits following inoculation into lumen of intestine, which proved that microspheres could be taken up by Peyer's patch. In conclusion, it is suggested that alginate microsphere prepared by spray method, showing a particle size of & $10\;{\mu}m$ and a high loading efficiency, can be used as a model drug for the development of oral vaccine delivery system.