• 통합자연과학논문집
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• 제12권4호
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• pp.127-132
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• 2019

Programmed cell death 4 (PDCD4) is a novel tumor suppressor that function in the nucleus and the cytoplasm and appears to be involved in the regulation of transcription and translation. Stress granules (SGs) are cytoplasmic foci at which untranslated mRNAs accumulate when cells exposed to environmental stresses. Since PDCD4 has implicated in translation repression through direct interaction with eukaryotic translation initiation factor 4A (eIF4A), we here investigated if PDCD4 has a functional role in the process of SG assembly under oxidative stresses. Using immunofluorescence microscopy, we found that PDCD4 is localized to SGs under oxidative stresses. Next, we tested if knockdown of PDCD4 has an effect on the assembly of SG using PDCD4-specific siRNA. Interestingly, SG assembly was accelerated and this effect was caused by sensitization of phosphorylation of eIF2α and dephosphorylation of eIF4E binding protein (4E-BP). These results suggest that PDCD4 has an effect on SG dynamics and possibly involved in cap-dependent translation repression under stress conditions.

• BMB Reports
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• 제43권3호
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• pp.205-211
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• 2010

Sprouty (Spry) proteins have previously been suggested as negative regulators of the MAPK pathway through interaction with Raf-1. However, the molecular basis of this inhibition has not been elucidated. In this study, we used cells expressing FLAGtagged Raf-1 with point mutations at known phosphorylation sites to reveal that activation of Raf-1 mutants does not correlate with their degree of interaction with Spry2. The association of Raf-1 with Spry2 in intact cells was further corroborated by immunofluorescence colocalization. Additionally, there was no significant change observed in the strength of interaction between Raf-1 mutants and Spry2 after paclitaxel treatment despite differences in the activation levels of these mutants. Thus, our study provides the evidence that Spry2 does not directly regulate Raf-1 kinase activity, but instead acts as a scaffolding protein that assists interactions between Raf-1 kinase and its direct regulators.

• 대한미생물학회지
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• 제21권1호
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• pp.163-170
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• 1986

Balb/c mouse spleen cells in vitro sensitized against ICR spleen cells were cultured in conditioned media(CM). The CM was produced by ICR spleen cells stimulated with Concanavalin-A(Con-A), and sensitized lymphoid cells were grown in CM. ICR mouse spleen cells were appeared to be a good generator of IL-2. Optimal growth was seen in growth medium containing 20% fetal calf serum. and 25% CM. When cultures were initiated at 1, 5, $10{\times}10^4\;cells/ml$, the cells were increased in numbers by about 20, 13, 5-fold, respectively, every 9 days. Such growth pattern was sustained for about 4-6 weeks and thereafter the cell growth was diminished gradually. Direct immunofluorescence indicated that 93% of the lymphoid cells grown in CM(for 10 days) expressed Thyl surface antigen. And the cells grown in CM were cytotoxic to the sesitizing ICR mouse spleen cells though cytotoxicity level was not high. According to these results, the cells grown in CM were considered to be cytotoxic T lymphocytes. The lymphoid cells grown for 20 days were nearly unresponsive to Con-A and therefore dependent only IL-2 to be used for IL-2 assay.

• 대한수의학회지
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• 제49권1호
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• pp.73-78
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• 2009

Discoid lupus erythematosus (DLE) is an immune-mediated skin disease which requires histopathology and immunohistopathology in both dogs and humans. A 10-month-old, intact female Alaskan Malamute presented for depigmentation, swelling, alopecia, erythema, and crusting on the bridge of the nose and the nasal planum. Cytological examination of nasal lesions revealed numerous cocci and neutrophils. Histopathological features included of infiltration of mononuclear cells at the dermoepidermal junction. Direct immunofluorescence tests and immunohistochemistry exhibited positive IgG, IgM, IgA, CD3, CD18, and CD79a on the epidermal basement membranes and around adnexal glands. This case indicates both T cells and B cells are related to mechanism of canine DLE. This case report describes advanced diagnostic tests and clinical outcome with immune suppressive therapy in a rare juvenile canine DLE case.

• Animal Bioscience
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• 제34권4호
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• pp.533-538
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• 2021

Objective: Pluripotent stem cell-derived lymphatic endothelial cells (LECs) show great promise in their therapeutic application in the field of regenerative medicine related to lymphatic vessels. We tested the approach of forced differentiation of mouse embryonal stem cells into LECs using biodegradable poly lactic-co-glycolic acid (PLGA) nanospheres in conjugation with growth factors (vascular endothelial growth factors [VEGF-A and VEGF-C]). Methods: We evaluated the practical use of heparin-conjugated PLGA nanoparticles (molecular weight ~15,000) in conjugation with VEGF-A/C, embryoid body (EB) formation, and LEC differentiation using immunofluorescence staining followed by quantification and quantitative real-time polymerase chain reaction analysis. Results: We showed that formation and differentiation of EB with VEGF-A/C-conjugated PLGA nanospheres, compared to direct supplementation of VEGF-A/C to the EB differentiation media, greatly improved yield of LYVE1(+) LECs. Our analyses revealed that the enhanced potential of LEC differentiation using VEGF-A/C-conjugated PLGA nanospheres was mediated by elevation of expression of the genes that are important for lymphatic vessel formation. Conclusion: Together, we not only established an improved protocol for LEC differentiation using PLGA nanospheres but also provided a platform technology for the mechanistic study of LEC development in mammals.

• Animal cells and systems
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• 제15권4호
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• pp.259-267
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• 2011

This study was undertaken to produce a $F_{ab}$ fragment of a human monoclonal antibody reactive to oxidized and carbamylated low-density lipoprotein (oxLDL and cLDL) using phage display technology. An analysis of DNA sequences of this $F_{ab}$, termed plaque 15,16-46 $F_{ab}$, revealed that the rearranged $V_H$ was highly mutated. Complementarity-determining regions of the $V_H$ showed a very high R/S ratio and contained many positively charged amino acids. In direct binding and competitive ELISA, the $F_{ab}$ reacted strongly with both MDA-LDL and Cu-oxLDL forms of oxLDL, and also showed high affinity for cLDL. Immunofluorescence and immunohistochemical analyses showed that this $F_{ab}$ positively stained atherosclerotic aortic plaques in $ApoE^{-/-}$ mice as well as those in patients with atherosclerosis. The $F_{ab}$ also showed positive staining in placental decidua from patients with preeclampsia. It is suggested that the plaque 15,16-46 $F_{ab}$ against oxLDL and cLDL might possibly be applicable for developing a diagnostic reagent for both human and rodent animal research to detect and characterize atherosclerotic disease progression in atherosclerotic lesions as well as exploring the pathogenesis of atherogenic diseases such as preeclampsia.

• International Journal of Oral Biology
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• 제37권2호
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• pp.77-83
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• 2012

We investigated the pulpal response to direct pulp capping in rat molar teeth using mineral trioxide aggregate (MTA) and calcium hydroxide (CH). A palatal cavity was prepared in rat maxillary molar teeth. Either MTA or CH was placed on the exposed pulp and all cavities were restored with composite. Rats were sacrificed for histological evaluation after 12 hours and at 2, 7, 14 and 21 days. In both the MTA and CH groups, reparative dentin formation was clearly observed on histology after 14 days. The MTA-capped pulps were found to be mostly free from inflammation, and hard tissue of a tubular consistent barrier was observed. In contrast, in CH-capped teeth, excessive formation of reparative dentin toward residual pulp was evident. The pulpal cell response beneath the reparative dentin layer was examined by immunofluorescence using antibodies against DSP. After 2 days, a few DSP immunopositive cells, most of which showed a cuboidal shape, appeared beneath the predentin layer. At 7 days, DSP-immunopositive cells with columnar odontoblast-like cells were seen beneath the newly formed hard tissues. At 14 and 21 days, DSP was more abundant in the vicinity of the odontoblastic process along the dentinal tubules than in the mineralized reparative dentin. The CH group showed strong expression patterns in terms of DSP immunoreactivity. Our results thus indicate that MTA may be a more effective pulp capping material as it induces the differentiation of odontoblast-like cells and the formation of reparative dentin without the loss of residual pulp functions.

• Animal cells and systems
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• 제15권3호
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• pp.211-218
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• 2011

Fragmentation in human pre-implantation embryos has been suggested as the process of apoptosis. We have previously demonstrated a direct relationship between the increased reactive oxygen species (ROS) and apoptosis in human pre-implantation embryos. ROS is known to suppress the function of mitochondria in which steroidogenic acute regulatory protein (StAR) and peripheral-type benzodiazepine receptor (PBR) are presented. Therefore, the purpose of this study was to examine the expression of StAR and PBR in human pre-implantation embryos and to evaluate whether reduction of these proteins is associated with apoptosis. Apoptosis was detected by annexin V-fluorescein isothiocyanate (FITC) and mitochondrial membrane potential was measured by 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethyl-benzimidazolyl-carbocyanine iodide (JC-1). Immunofluorescence staining and Western blotting were applied to examine the expression of StAR and PBR in the embryos. Lipid droplets in the embryos were stained with Oil Red O. The fragmented pre-implantation embryos were stained with annexin V-FITC, but not the normal ones. The mitochondria with active membrane potential were present less in the fragmented embryos compared with the non-fragmented embryos. We also confirmed that both StAR and PBR were expressed in the embryos and their expression levels were lower in the fragmented ones. In addition, the number and size of lipid droplets were increased in the fragmented embryos. The present study provides evidence that reduction of StAR and PBR in human pre-implantation embryos is associated with an increase in the lipid droplets leading to apoptosis.

• Childhood Kidney Diseases
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• 제10권1호
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• pp.40-44
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• 2006

[ $Henoch-Sch{\"{o}}nlein$ ] 자반증은 소혈관의 혈관염으로 오는 전신적 혈관 장애로 자반, 위장 증세, 관절 증세, 신 증세를 주 증세로 하는 질환이다. HSP의 흔한 피부증상은 주로 궁둥이와 하지의 자반 증상이며 성인에서는 출혈성 수포를 동반하는 경우가 흔하나 소아에서는 드문 것으로 알려져 있다. 저자들은 관절통과 복통을 호소하는 환아에서 출혈성 수포가 동반되어 있어 피부조직 검사를 하였고 진피의 leukocytoclastic vasculitis와 면역 형광법상 소혈관의 IgA와 fibrinogen의 침착을 관찰하였다. HSP로 진단하여 스테로이드 치료를 하였고 수일 내로 관절통과 복통이 호전되어 문헌고찰과 함께 보고하는 바이다.

• International Journal of Stem Cells
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• 제16권1호
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• pp.93-107
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• 2023

Background and Objectives: Chronic periodontitis can lead to alveolar bone resorption and eventually tooth loss. Stem cells from exfoliated deciduous teeth (SHED) are appropriate bone regeneration seed cells. To track the survival, migration, and differentiation of the transplanted SHED, we used super paramagnetic iron oxide particles (SPIO) Molday ION Rhodamine-B (MIRB) to label and monitor the transplanted cells while repairing periodontal bone defects. Methods and Results: We determined an appropriate dose of MIRB for labeling SHED by examining the growth and osteogenic differentiation of labeled SHED. Finally, SHED was labeled with 25 ㎍ Fe/ml MIRB before being transplanted into rats. Magnetic resonance imaging was used to track SHED survival and migration in vivo due to a low-intensity signal artifact caused by MIRB. HE and immunohistochemical analyses revealed that both MIRB-labeled and unlabeled SHED could promote periodontal bone regeneration. The colocalization of hNUC and MIRB demonstrated that SHED transplanted into rats could survive in vivo. Furthermore, some MIRB-positive cells expressed the osteoblast and osteocyte markers OCN and DMP1, respectively. Enzyme-linked immunosorbent assay revealed that SHED could secrete protein factors, such as IGF-1, OCN, ALP, IL-4, VEGF, and bFGF, which promote bone regeneration. Immunofluorescence staining revealed that the transplanted SHED was surrounded by a large number of host-derived Runx2- and Col II-positive cells that played important roles in the bone healing process. Conclusions: SHED could promote periodontal bone regeneration in rats, and the survival of SHED could be tracked in vivo by labeling them with MIRB. SHED are likely to promote bone healing through both direct differentiation and paracrine mechanisms.