• Journal of Pharmaceutical Investigation
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• 제40권6호
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• pp.333-337
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• 2010

Combining cell- and gene-based therapy is a promising therapeutic strategy in regenerative medicine. The aim of this study was to develop genetically modified mesenchymal stem cell (MSC) aggregates using a poly(ethylene glycol) (PEG) hydrogel micro-well array technique. Stable PEG hydrogel micro-well arrays with diameters of 200 to $500\;{\mu}m$ were fabricated and used to generate genetically engineered MSC aggregates. Rat bone marrow-derived MSCs were transfected with a green fluorescent protein (GFP) plasmid as a reporter gene, and aggregated by culturing in the PEG hydrogel micro-well arrays. The resultant cell aggregates had a mean diameter of less than $200\;{\mu}m$, and maintained the mesenchymal phenotype even after genetic modification and cell aggregation. Transplantation of MSC aggregates that are genetically modified to express therapeutic or cell-survival genes may be a potential therapeutic approach for regenerative medicine.

• 한국동물생명공학회지
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• 제39권1호
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• pp.19-30
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• 2024

Background: Although an understanding of the proliferation and differentiation of fish female germline stem cells (GSCs) is very important, an appropriate threedimensional (3D) research model to study them is not well established. As a part of the development of stable 3D culture system for fish female GSCs, we conducted this study to establish a 3D aggregate culture system of ovarian cells in marine medaka, Oryzias dancena. Methods: Ovarian cells were separated by Percoll density gradient centrifugation and two different cell populations were cultured in suspension to form ovarian cell aggregates to find suitable cell populations for its formation. Ovarian cell aggregates formed from different cell populations were evaluated by histology and gene expression analyses. To evaluate the media supplements, ovarian cell aggregate culture was performed under different media conditions, and the morphology, viability, size, gene expression, histology, and E2 secretion of ovarian cell aggregates were analyzed. Results: Ovarian cell aggregates were able to be formed well under specific culture conditions that used ultra-low attachment 96 well plate, complete mESM2, and the cell populations from top to 50% layers after separation of ovarian cells. Moreover, they were able to maintain minimal ovarian function such as germ cell maintenance and E2 synthesis for a short period. Conclusions: We established basic conditions for the culture of O. dancena ovarian cell aggregates. Additional efforts will be required to further optimize the culture conditions so that the ovarian cell aggregates can retain the improved ovarian functions for a longer period of time.

• Molecules and Cells
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• 제42권6호
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• pp.480-494
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• 2019

Aggregates of disease-causing proteins dysregulate cellular functions, thereby causing neuronal cell loss in diverse neurodegenerative diseases. Although many in vitro or in vivo studies of protein aggregate inhibitors have been performed, a therapeutic strategy to control aggregate toxicity has not been earnestly pursued, partly due to the limitations of available aggregate models. In this study, we established a tetracycline (Tet)-inducible nuclear aggregate (${\beta}23$) expression model to screen potential lead compounds inhibiting ${\beta}23$-induced toxicity. High-throughput screening identified several natural compounds as nuclear ${\beta}23$ inhibitors, including peucedanocoumarin III (PCIII). Interestingly, PCIII accelerates disaggregation and proteasomal clearance of both nuclear and cytosolic ${\beta}23$ aggregates and protects SH-SY5Y cells from toxicity induced by ${\beta}23$ expression. Of translational relevance, PCIII disassembled fibrils and enhanced clearance of cytosolic and nuclear protein aggregates in cellular models of huntingtin and ${\alpha}$-synuclein aggregation. Moreover, cellular toxicity was diminished with PCIII treatment for polyglutamine (PolyQ)-huntingtin expression and ${\alpha}$-synuclein expression in conjunction with 6-hydroxydopamine (6-OHDA) treatment. Importantly, PCIII not only inhibited ${\alpha}$-synuclein aggregation but also disaggregated preformed ${\alpha}$-synuclein fibrils in vitro. Taken together, our results suggest that a Tet-Off ${\beta}23$ cell model could serve as a robust platform for screening effective lead compounds inhibiting nuclear or cytosolic protein aggregates. Brain-permeable PCIII or its derivatives could be beneficial for eliminating established protein aggregates.

• Journal of Plant Biotechnology
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• 제34권1호
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• pp.19-24
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• 2007

자색고구마 (Ipomoea batatas cv. Jami) 잎 조직배양으로 유도된 캘러스는 계대배양시 생장량이 감소하였다. 빠르고 간편하게 생장하는 습관성캘러스를 $1{\mu}M\;2,4-D$ 첨가된 MSBM에서 계대배양 없이 6개월간 지속배양하여 선발하였다. 습관성캘러스는 생장조절제가 첨가되지 않는 MSBM의 고체, 액체 배지에서 계대배양 횟수의 제한 없이 빠르게 지속생장이 가능하였다. 암조건의 액체배양 6 개월 후에는 자색생합성 부위가 생겼으며, 이를 2년 여간 분리하여 자색습관성세포괴를 선발하였다. 안토시아닌 함량은 자미의 괴근에서 추출되는 1.5 mg/ml에 근접한 1.0 mg/ml으로 나타났다. 자색습관성 세포괴는 안토시아닌 기내대량에 효과적 세포주로 사용 될 수 있을 것으로 기대한다.

• Journal of Microbiology and Biotechnology
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• 제12권5호
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• pp.735-739
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• 2002

Large quantities of porcine hepatocyte aggregates with various degrees of aggregation (DA) could be obtained by controlling the suspension periods (0,9,24, and 48 h), and by entrapping the hepatocyte aggregates in model materials of encapsulation such as Ca-alginate and type-I collagen gels. The effects of DA on liver-specific functions of hepatocytes were evaluated in order to obtain optimum DA for the cell source of bioartificial liver (BAL) systems. Irregular rugged aggregates (size $75 \pm 28$ $\mu\textrm{m}$) farmed by 24 h of suspension culturing showed peak viability and hepatic functions such as ammonia removal and albumin secretion in the two types of entrapment systems, thus offering themselves as a stable cell source of a BAL system for hepatic functions and scale-up.

• Journal of Korean Neurosurgical Society
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• 제56권5호
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• pp.383-389
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• 2014

Objective : Neural tissue transplantation has been a promising strategy for the treatment of Parkinson's disease (PD). However, transplantation has the disadvantages of low-cell survival and/or development of dyskinesia. Transplantation of cell aggregates has the potential to overcome these problems, because the cells can extend their axons into the host brain and establish synaptic connections with host neurons. In this present study, aggregates of human brain-derived neural stem cells (HB-NSC) were transplanted into a PD animal model and compared to previous report on transplantation of single-cell suspensions. Methods : Rats received an injection of 6-OHDA into the right medial forebrain bundle to generate the PD model and followed by injections of PBS only, or HB-NSC aggregates in PBS into the ipsilateral striatum. Behavioral tests, multitracer (2-deoxy-2-[$^{18}F$]-fluoro-D-glucose ([$^{18}F$]-FDG) and [$^{18}F$]-N-(3-fluoropropyl)-2-carbomethoxy-3-(4-iodophenyl)nortropane ([$^{18}F$]-FP-CIT) microPET scans, as well as immunohistochemical (IHC) and immunofluorescent (IF) staining were conducted to evaluate the results. Results : The stepping test showed significant improvement of contralateral forelimb control in the HB-NSC group from 6-10 weeks compared to the control group (p<0.05). [$^{18}F$]-FP-CIT microPET at 10 weeks posttransplantation demonstrated a significant increase in uptake in the HB-NSC group compared to pretransplantation (p<0.05). In IHC and IF staining, tyrosine hydroxylase and human ${\beta}2$ microglobulin (a human cell marker) positive cells were visualized at the transplant site. Conclusion : These results suggest that the HB-NSC aggregates can survive in the striatum and exert therapeutic effects in a PD model by secreting dopamine.

• Journal of Plant Biology
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• 제32권4호
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• pp.247-253
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• 1989

Protoplasts were enzymatically isolated from suspension culture of sweet potato. High yields of single protoplasts were produced from nonembryogenic cell aggregates. However, most protoplasts obtained from embryogenic cell clumps were spontaneously fused during enzyme treatment; a small portion of them remained single. Upon transfer to Murashige and Skoog's(MS) liquid medium supplemented with 0.1 mg/1 6-benzyladenine(BA) and 1 mg/12,4-dichlorophenoxyacetic acid(2,4-D), protoplasts from nonembryogenic cell aggregates sustained cell divisions to form cellus. Upon subculture onto MS media with 0.2 mg/12,4-D or without growth regulators, the callus did not give rise to any organs. On the other hand, first cell division of single protoplasts from embryogenic cell clumps was sporadically observed.

• Computers and Concrete
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• 제8권5호
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• pp.525-539
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• 2011

This paper describes the development of a fiber reinforced concrete (FRC) unit cell for analyzing concrete structures by executing a multiscale analysis procedure using the theory of homogenization. This was achieved through solving a periodic unit cell problem of the material in order to evaluate its macroscopic properties. Our research describes the creation of an FRC unit cell through the use of concrete paste generic information e.g. the percentage of aggregates, their distribution, and the percentage of fibers in the concrete. The algorithm presented manipulates the percentage and distribution of these aggregates along with fiber weight to create a finite element unit cell model of the FRC which can be used in a multiscale analysis of concrete structures.

• The Korean Journal of Physiology and Pharmacology
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• 제13권4호
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• pp.273-279
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• 2009

The accumulation of ${\beta}$-amyloid (A${\beta}$) aggregates is a characteristic of Alzheimer's disease (AD). Furthermore, these aggregates have neurotoxic effects on cells, and thus, molecules that inhibit A${\beta}$ aggregate formation could be valuable therapeutics for AD. It is well known that aggregation of A${\beta}$ depends on its hydrophobicity, and thus, in order to increase the hydrophilicity of A${\beta}$, we considered using citrate, an anionic surfactant with three carboxylic acid groups. We hypothesized that citrate could reduce hydrophobicity and increase hydrophilicity of A${\beta}_{1-40}$ molecules via hydrophilic/electrostatic interactions. We found that citrate significantly inhibited A${\beta}_{1-40}$ aggregation and significantly protected SH-SY5Y cell line against A${\beta}_{1-40}$ aggregates-induced neurotoxicity. In details, we examined the effects of citrate on A${\beta}_{1-40}$ aggregation and on A${\beta}_{1-40}$ aggregates-induced cytotoxicity, cell viability, and apoptosis. Th-T assays showed that citrate significantly inhibited A${\beta}_{1-40}$ aggregation in a concentration-dependent manner (Th-T intensity: from 91.3% in 0.01 mM citrate to 82.1% in 1.0 mM citrate vs. 100.0% in A${\beta}_{1-40}$ alone). In cytotoxicity and viability assays, citrate reduced the toxicity of A${\beta}_{1-40}$ in a concentration-dependent manner, in which the cytotoxicity decreased from 107.5 to 102.3% as compared with A${\beta}_{1-40}$ aggregates alone treated cells (127.3%) and the cell viability increased from 84.6 to 93.8% as compared with the A${\beta}_{1-40}$ aggregates alone treated cells (65.3%). Furthermore, Hoechst 33342 staining showed that citrate (1.0 mM) suppressed A${\beta}_{1-40}$ aggregates-induced apoptosis in the cells. This study suggests that citrate can inhibit A${\beta}_{1-40}$ aggregation and protect neurons from the apoptotic effects of A${\beta}_{1-40}$ aggregates. Accordingly, our findings suggest that citrate administration should be viewed as a novel neuroprotective strategy for AD.

• 대한약리학회지
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• 제23권2호
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• pp.101-111
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• 1987

중추신경계 특히 뇌내(腦內)의 무수카린성 콜린 수용체 (mAchR)에 대한 수용체 특성의 연구의 하나로, 물리 화학적 성상에 다른 두 종류의 콜린길항제를 사용하여 서로 다른 두 형태의 조직에서 약물의 작용양상 및 다른 약물과의 상호작용을 정초하였다. 실험동물로는 흰쥐를 일정기간 규정사료를 사육하였고, 사용한 Radioactive ligands는 $(^3H)$ QNB와 $(^3H)$ NMS였으며 그외에 다른 수종의 길항제 또는 효능제와의 치환작용을 brain homogenates와 intact brain cell aggregates에서 관찰하여 다음과 같은 결과를 얻었다. 1. $(^3H)$ QNB와 $(^3H)$ NMS는 모두 질량작용의 법칙에 비례하여 수용체와의 결합에서 높은 친화력과 포화를 보였으며 또한 높은 결합 능력을 나타내었다. 더욱이 homogenates 제제와 intact cell aggregates제제에서의 결과 사이에는 유사한 점이 많았다. 2. Homogenates제제를 사용한 실험에서, 제 3 급아민콜린길항제인 QNB, atropine과 scopolamine 또는 제 4 급 암모늄골린 길항제인 methylatropine과 methylscopolamine을 사용하여 위의 radioactive ligands와의 치환작용을 검토하였다. $(^3H)$ NMS 실험군에서는 제 3 급아민 및 제 4 급 암모늄길항제 모두가 구조의 구별없이 질량작용의 법칙에 따라 치환되었으나 $(^3H)$ QNB 실험군에서는 제 4급 암모늄콜린 길항제들을 단일성(unity)이 아닌 높고 낮은 두 종류의 친화도를 가진 결합부위의 양상을 나타내었다. 또 비특이성 콜린길항제인 pirenzepine을 사용한 실험군에서는 두 ligands을 모두 치환시켰고 서로 다른 결합부위가 있음을 보였다. 3. Intact cell aggregates 제제를 사용한 실험에서, $(^3H)$ NMS와 $(^3H)$ QNB 모두 homogenates 제제에서와 같은 양상의 반응을 보였다. 또 $(^3H)$ NMS를 radioligand로 하여 수종의 콜린길항제와 수종의 콜린 효능제를 사용하여 약물 상호작용으로 수용체의 성질을 검토하였다. 그 결과 콜린 길항제들은 질량작용의 법칙에 따라 치환되었으나 콜린 효능제 투여군에서는 높고 낮은 두 종류의 다른 친화력의 결항부위를 나타내었다. 4. 위의 실험의 결과로,(a) 친유성콜린 길항제인 $(^3H)$ QNB는 친수성 콜린길항제인 $(^3H)$ NMS보다 훨씬 높은 결합능력을 보였으며 이것으로 수용체 특히 mAchR의 존재 장소 또는 mAchR의 형상의 일부는 세포막 표면 뿐 아니라 세포막내의 어떤 부위와도 관계가 되는 것으로 간주되는데 이것이 $(^3H)$ QNB가 $(^3H)$ NMS보다 높은 최대 결합능력 $(B_{max})$을 나타낼 이유이다. (b) 두 종류의 다른 제제에서 우리는 같은 양상의 결과를 관찰하었기에 결점이 많은 homogenates 제제보다는 intact cell aggregates 제제를 수용체 연구에 대한 새로운 실험모형(experiment model)으로 사용할 수 있는 가능성을 제시하고자 한다.