• Archives of Pharmacal Research
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• v.20 no.3
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• pp.225-233
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• 1997

A murine leukemia x LM fibroblast hybrid cell line with immune augmenting properties stimulated resistance to Mycobacterium avium complex (MAC) in mouse peritoneal macrophages, and in immune deficient beige mice (C57BL/6/bgj/bgj). The proliferation of MAC in mouse peritoneal macrophages was inhibited by medium conditioned by the growth of the hybrid cells (hybrid cell-CM). Under similar circumstances, media conditioned by the growth of LM cells (LM cell-CM), a mouse fibroblast cell line used as one parent in forming the hybrid cell, was exhibited no inhibitory effect. Treatment of mouse peritoneal macrophages with hybrid cell-CM, but not with LM cell-CM, stimulated the expression of each of four previously described macrophage activation antigens, suggesting that the hybrid cells formed immunomodulators in addition to those formed by LM cells. Furthermore, the morphology of the macrophages following treatment with hybrid cell-CM was clearly distinguishable from that following exposure of the cells to LM cell-CM. The therapeutic effects of hybrid cells on the progression of MAC-infection were indicated by the prolonged survival of MAC-infected immune-deficient beige mice. One hundred percent of treated animals survived more than 60 days, while untreated animals died in approximately 22 days.

• Reproductive and Developmental Biology
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• v.31 no.2
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• pp.109-113
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• 2007

This study was conducted to evaluate the effect of cytochalasin B (CB) treatment in the activation medium on the development of somatic cell nuclear transfer (SCNT) rat embryos. Fetal fibroblast cells were isolated from a Day 14.5 fetus, and the oocytes for recipient cytoplasm were recovered from 4-week old Sprague Dawley rats. After enucleation and nuclear injection, the reconstructed oocytes were immediately exposed to activation medium consisting of 10 mM $SrCl_2$ with or without CB for 4 hr, and formation of pseudo-pronucleus (PPN) was checked at 18 hr after activation. Then, they were transferred into day 1 pseudopregnant recipients (Hooded Wistar) or cultured for 5 days to check their developmental competence in vivo or in vitro. The number of PPN was not affected by CB treatment during the activation. However, CB treatment supported pre-implantation development of rat SCNT embryos. Embryos generated by the procedures of SCNT were also capable of implanting, with 1 implantation scar found from a recipient following the transfer of 87 SCNT embryos to four foster mothers. The result of the present study shows that rat SCNT embryo can develop to post-implantation stage following treatment with CB.

• Journal of Periodontal and Implant Science
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• v.31 no.2
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• pp.277-285
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• 2001

Periodontal disease is characterized by inflammation and subsequent loss and/or damage to tooth-supporting tissues such as bone, cementum,and periodontal ligament. Periodontal ligament and cementum are the key tissues in the initial process of regeneration following periodontal disease. Therefore, studies on cementoblasts, which form cementum are emphasized. It is still unclear which cells cementoblast differentiate from. This study was conducted under the hypothesis that PDL fibroblast can differentiate into either cementoblast or osteoblast depending on the conditions of surrounding tissue. Clinically, with excessive traction force of orthodontic appliances or excessive occlusion hypercementosis is observed, and this has been confirmed histologically. Consequently, activation of cementoblast can be expected in rats when mechanical stimuli are given to PDL fibroblast. Therefore, the purpose of this article is to prove that PDL fibroblast differentiates into cementoblast in rats under mechanical stimuli using histologic and molecular methods. In this study, twenty rats were given hard diet. Ten of them were sacrificed after 1 week, and the others were sacrificed after two weeks. Slides were made from tooth specimen, and they were studied under the microscope. In addition, PDL fibroblast and cementum from the extracted teeth were analyzed with Northern blotting. In histologic examination, as time passed, PDL fibroblast migrated to the dentin side, differentiated into cementoblast, and formed new cementum. In Northern blotting, it was found that mRNA expression of cementoblast-specific proteins such as BSP, OC, OPN, and type I collagen were more prominent in rats sacrificed after 2 weeks of hard-diet than rats sacrificed after 1 week. From these findings we can conclude that PDL fibroblast can differentiate into cementoblast under mechanical stimuli. We think that 'Rat Models' used in this study will be beneficial to future studies regarding cementoblast.

• Proceedings of the KSAR Conference
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• 2001.10a
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• pp.60-61
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• 2001

To produce reconstituted rabbit embryos with fetal fibroblasts, the present study was evaluated the efficiencies of the activation conditions as assessments of subsequent development and chromosome in the embryos. New Zealand White rabbits were used throughout the study. Fetal fibroblasts collected from 22-d of fetuses were cultured in DMEM＋10% FBS in 5% CO₂ in air. The culture was maintained for 10 passages. In every passage half of cell suspension were kept in frozen. (omitted)

• Proceedings of the Korean Environmental Sciences Society Conference
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• 2020.10a
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• pp.219-219
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• 2020

Kaempferol (3,4',5,7-tetrahydroxyflavone), a flavonoid found in beans, broccoli, garlic, etc., has been used in natural medicine as an anti-inflammatory and antioxidant. This experiment was carried out to evaluate the anti-apoptotic effect of kaempferol in 12-O-tetradecanoylphorbol 13-acetate (TPA)-treated Normal Human Dermal Fibroblast (NHDF). Kaempferol inhibited the production of intracellular Reactive Oxygen Species (ROS) induced by TPA in NHDF. Kaempferol significantly blocks the phosphorylation of extracellular signal-regulated kinase responsible for the activation of nuclear factor-kappa B. In addition, kaempferol significantly attenuated the expression of Bax and cleaved caspase-3 as regulated by the phosphorylation of nuclear factor-kappa B during its blockage of TPA-induced apoptotic cell death. These findings suggest that kaempferol protects the apoptotic signaling pathway induced by TPA through modulating intracellular ROS in NHDF.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.109-109
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• 2003

Even though success in birth of live offspring from nuclear transfer(NT) using somatic cells in many species, detailed information on processes or mechanisms of development are not well known. Cytoplasm of bovine oocyte has been known to support the development of nuclear transferred embryos using nuclear donor cells from different species. Therefore, interspecies NT might be used to find answers of some questions in basic aspect of nuclear transfer In this study, we examined the developmental potential of reconstructed embryos when bovine oocyte as a cytoplasm recipient and mouse embryonic fibroblast as a nuclear donor were used. The nuclear transfer units were aliocated in Group 1 (murine block media and normal media) and Group 2. (bovine block media and normal media). NT units were not blocked at 2-cell stage regardless of types of medium. On mouse media, poor development of interspecies NT units was observed compared to bovine media. However, as NT units cultured in bovine normal medium, embryos developed over 8-cell stage. Further studies performed to increase the developmental rate in condition of antioxidant treatment. Despite low development, bovine-murine interspecies nuclear transferred embryos could develop to blastocysts and they showed that blastocyts rate of antioxidant group was superior to those of non-antioxidant group. Next, we investigated gene expression pattern which is carried out for zygotic activation. The Xist gene is expressed in female mouse embryo after zygotic activation of 4-cell stage. But interspecies nuclear transferred embryos do not express Xist gene at 4-cell stage. As a result, it is suggested that the bovine cytoplasm controls the early preimplantation development in interspecies NT However, the development of later stages might require genomic control from transferred donor nucleus. Therefore, even though the involvement of several other factors such as mitochondrial incompatibility, effective development of embryos produced by interspecies NT requires proper genomic activation of donor nucleus after overcoming the cytoplasmic control of recipient oocytes.

• Proceedings of the KSAR Conference
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• 2001.03a
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• pp.22-22
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• 2001

To produce reconstituted rabbit embryos with fetal fibroblasts, the present study was evaluated the efficiencies of the different fusion and activation conditions as assessments of subsequent development and chromosome in the embryos. New Zealand White rabbits were used throughout the study. Fetal fibroblasts collected from 22-d of fetuses were cultured in DMEM ＋ 10% FBS in 5% $CO_2$ in air. The culture was maintained for 10 passages. In every passage half of cell suspension were kept In frozen. From rabbits treated with FSH in 30% PVP solution and hCG, oocytes were surgically collected from oviducts at 14 h post-hCG injection and stripped off their cumulus cells by re-pipetting in a 300 IU hyaluronidase solution. Oocytes with an extruded first polar body and dense cytoplasm were enucleated by micromanipulation in Ham's F-10 medium＋7.5 g/$m\ell$ cytochalasin B. Euncleation was confirmed under a fluorescence microscope after staining with 5 g/$m\ell$ bisbenzimide for 2 min. Each enucleated oocyte was injected with a fetal fibroblast into a perivitelline space. Reconstructed eggs were compared fusion rates either at 2.0 ㎸/cm or 1.6 ㎸/cm(60 sec, double pulses). After fusion, all eggs were activated with the combination of 5 M ionomycin (5 min) and 10 g/$m\ell$ cycloheximide (CHX, 3h), and cultured in CRlaa medium and transferred into TCM199＋10% FBS on day 3. Although there was not significantly differ in fusion rate between treatments (60%, 2.0 ㎸/cm vs. 79.4%, 1.6 ㎸/cm), none of them in the eggs fused with 2.0 ㎸/cm developed to blastocyst. In comparison of development and chromosome status between different activation treatments (Group 1; 5 M ionomycin/10 g/$m\ell$ CHX, Group 2; 5 M ionomycin/5 g/$m\ell$ CHX ＋ 2 mM DMAP after fusion with 1.6 ㎸/cm), there were not differ in cleavage and development rates (67.3% and 28.9% in Group 1; 67% and 33% in Group 2). All out of 8 embryos evaluated in Group 1 appeared a normal diploid chromosome sets and mean number of cells (Mean SEM) on day 4.5 of culture was 141.5 23.15 (n=8). It can be concluded that the use of cycloheximide has not happened in chromosome abnormalities, and fetal fibroblasts can be used for cloning in rabbit.

• Korean Journal of Clinical Laboratory Science
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• v.56 no.1
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• pp.43-51
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• 2024

[⁶⁸Ga]Ga-FAPI-04 is a promising radiopharmaceutical that binds specifically to fibroblast activation protein, which is overexpressed in more than 90% of malignant epithelial tumors but not in normal healthy tissue. This study aimed to develop an efficient method for producing ⁶⁸Ga-labelled FAPI-04 using a cassette-based automated synthesizer. [⁶⁸Ga]GaCl₃ was eluted from an Eckert & Ziegler Medical germanium-68/gallium-68 generator using 2.5 mL of 0.1 M HCl. The synthesis of the [⁶⁸Ga]Ga-FAPI-04 was performed using different concentrations of HEPES (1~2.5 M; 4-(2-hydroxyethyl) piperazine-1-ethanesulfonic acid) in 3~10 minutes; amounts of FAPI-04 precursor (5~50 ㎍) and reaction temperature (25℃~100℃) were optimized on the BIKBox^® synthesizer. The labeling efficiency of [⁶⁸Ga]Ga-FAPI-04 was greater than 96% (decay corrected) using 25 ㎍ FAPI-04 synthesized in 10 minutes at 100℃ in 2 M HEPES (pH 3.85), and its stability was greater than 99% at 6 hours. The total synthesis time of [⁶⁸Ga]Ga-FAPI-04 was 32.4 minutes, and the product met all quality control criteria. In this study, we developed and optimized a labeling method using [⁶⁸Ga]Ga-FAPI-04 using a cassette-based synthesizer. The devised method is expected to be useful for supplying [⁶⁸Ga]Ga-FAPI-04 for diagnosis in clinical practice.

• Journal of Physiology & Pathology in Korean Medicine
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• v.25 no.3
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• pp.533-539
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• 2011

Sulfuretin is one of the main flavonoids produced by Rhusverniciflua. Sulfuretin has been shown to exhibit many pharmacological activities including anti-oxidant, anti-obesity, anti-inflammatory and anti-mutagenic activities. However, the anti-skin photoaging effects of sulfuretin has not yet been reported. In the present study, we investigated the inhibitory effect of sulfuretin on the expression levels of MMP-1 and -3 in the human dermal fibroblast cells. Western blot analysis and real-time PCR revealed sulfuretin inhibited UVB-induced MMP-1 and -3 expressions in a dose-dependent manner. UVB-induced MAPK/NF-${\kappa}B$/p50 activation and MMP expression were completely blocked by pretreatment of sulfuretin. Taken together, sulfuretin could prevent UVB-induced MMP expressions through inhibition of MAPK/NF-${\kappa}B$/p50 activation.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.5
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• pp.1833-1837
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• 2015

An aristolactam-type alkaloid, isolated from Orophea enterocarpa, is enterocarpam-III (10-amino-2,3,4,6-tetramethoxyphenanthrene-1-carboxylic acid lactam). It is cytotoxic to various human and murine cancer cell lines; however, the molecular mechanisms remain unclear. The aims of this study were to investigate cytotoxic effects on and mechanism (s) of human cancer cell death in human hepatocellular carcinoma HepG2 and human invasive breast cancer MDA-MB-231 cells compared to normal murine fibroblast NIH3T3 cells. Cell viability was determined by MTT assay to determine $IC_{10}$, $IC_{20}$ and $IC_{50}$ levels, reactive oxygen species (ROS) production with 2',7'-dichlorohydrofluorescein diacetate and the caspase-3, -8 and -9 activities using specific chromogenic (p-nitroaniline) tetrapeptide substrates, viz., DEVD-NA, IETD-NA and LEHD-NA and employing a microplate reader. Mitochondrial transmembrane potential (MTP) was measured by staining with 3, 3'-dihexyloxacarbocyanine iodide ($DiOC_6$) and using flow cytometry. The compound was cytotoxic to HepG2 and MDA-MB-231 cells with the $IC_{50}$ levels of $26.0{\pm}4.45$ and $51.3{\pm}2.05{\mu}M$, respectively. For murine normal fibroblast NIH3T3 cells, the $IC_{50}$ concentration was $81.3{\pm}10.1{\mu}M$. ROS production was reduced in a dose-response manner in HepG2 cells. The caspase-9 and -3 activities increased in a concentration-dependent manner, whereas caspase-8 activity did not alter, indicating the intrinsic pathway activation. Enterocarpam-III decreased the mitochondrial transmembrane potential (MTP) dose-dependently in HepG2 cells, suggesting that the compound induced HepG2 cell apoptosis via the mitochondrial pathway. In conclusion, enterocarpam-III inhibited HepG2 and MDA-MB-231 cell proliferation and induced human HepG2 cells to undergo apoptosis via the intrinsic (mitochondrial) pathway and induction of caspase-9 activity.