• Development and Reproduction
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• v.11 no.3
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• pp.167-177
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• 2007

In the present study, we isolated three human adult stem cells including adipose tissue-derived stem cells(HAD), umbilical cord-derived stem cells(HUC), and amnion-derived stem cells(HAM) and analysed their characteristics. And we examined whether HAD could be used as therapeutical cells for the heart diseases. Both HAM and HUC appeared very similar morphology but HAD was different. Doubling time of HUC was most fast, but total doubling numbers of HUC was same with HAM. Total doubling numbers of HAD was much more than others. Expression patterns of genes and proteins of three human adult stem cells were very similar. Also they were differentiated into adipocytes, osteocytes, and chondrocytes. In addition, they expressed many cardiomyocyte-related genes. But expression pattern of genes is a little different. When HAD were cultivated in the presence or absence of various combinations of BMP and FGF after 5-azacytidine expose for 24 h, expression of Cmlc-1, and ${\alpha}1c$ genes was significantly increased. However, expression of troponin T, troponin I and Kv4.3 genes was not changed. Based on these observations, it is suggested that HAD, HUC, and HAM might be used as potentially therapeutical cells for clinical application.

• Clinical and Experimental Pediatrics
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• v.45 no.12
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• pp.1551-1558
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• 2002

Purpose : Loss of hippocampal interneurons in dentate gyrus has been reported in patients with severe temporal lobe epilepsy and in animals treated with kainic acid(KA). Interneurons contain $Ca^{2+}$- binding protein parvalbumin(PV). The effects of kainic acid on parvalbumin-immunoreactive (PV-IR) interneurons in dentate gyrus were investigated in organotypic hippocampal slice cultures. Methods : Cultured hippocampal slices from postnatal day nine C57/BL6 mice were exposed to $10{\mu}M$ KA, and were observed at 0, 8, 24, 48, 72 hours after a one hour KA exposure. Neuronal injury was determined by morphologic changes of PV-IR interneuron in dentate gyrus. Results : Transient(1 hour) exposure of hippocampal explant cultures to KA produced marked varicosities in dendrites of PV-IR interneuron in dentate gyrus and the shaft of interbeaded dendrite is often much thinner than those in control. The presence of varicosities in dendrites was reversible with KA washout. The dendrites of KA treated explants were no longer beaded at 8, 24, 48 and 72 hours after KA exposure. The number of cells in PV-IR interneurons in dentate gyrus was decreased at 0, 8 hours after exposure. But there was no significant difference in 24, 48 and 72 hours recovery group compared with control group. Conclusion : The results suggested that loss of PV-IR interneurons in dentate gyrus is transient, and is not accompanied by PV-IR interneuronal cell death.

• The Korean Journal of Zoology
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• v.18 no.2
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• pp.51-60
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• 1975

The present study was performed histologically and histochemically to observe the cutaneous mucous glands in the frog, Rana nigromaculata during metamorphosis. The cutaneous thssues including dermal plicae in the dorsal portions of the frog tadpoles at each metamorphosis stage were fixed in 10% buffered formalin at$4^{\circ}C$, embedded in paraffin wax, sectioned 4 $\mu$m thickness and stained with periodic acid-Schiff(PAS) and alcian blue (AB) at both pH 2.5 and pH 1.0. The results of observation were as follows: 1. The developments of cutaneous mucous glands of the frog tadpole were begun with appearance of gland cell nest in the dermis at metamorphosis XV stage and significant numerical increases could be seen at metamorphosis XX, XXIII and XXIV stages. 2. This cutaneous mucous gland of the frog tadpole could be divided into two types; A-type glands showed strong positivities to both PAS and AB at pH 2.5 in the gland body cells and to PAS in the gland neck cells, and B-type glands at AB pH 2.5 in the gland body cells. 3. In the A-type mucous glands, the reactivities of the glandular epithelial cells to both PAS and AB stain could be first seen at the metamorphosis XIX stage of frog tadpole. The reactivities of the glandular epithelial cells to both PAS and AB pH 2.5 were gradually increased according to the process of metamorphosis after XX stage of metamorphosis. 4. The B-type mucous glands were first seen at the XX stage and the reactivity of the glandular epithelial cells to AB at pH 2.5 was gradually increased according to the process of metamorphosis after XX stage. 5. The A-type and the B-type mucous glands were in the ratio of 99 : 1, 7 : 3 and 5.5 : 4.5 for each of metamorphosis XX, XXI-XXII and XXIII-XXV stages. 6. The remarkable development of the cutaneous mucous glands of the frog tadpoles might be needed to maintain water and electrolyte balances according to the change of way from aquatic life to amphibious.

• Clinical and Experimental Pediatrics
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• v.49 no.6
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• pp.677-685
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• 2006

Purpose : This study aimed to investigate whether exogenous thyroxine($T_4$) treatment to alcohol-fed dams would ameliorate the detrimental effects of alcohol on the postnatal development of neuropeptide-Y(NPY)-containing neurons of the cerebral cortex and hippocampus of the offspring. Methods : Time-pregnant rats were divided into three groups. An alcohol-fed group A received 35 calories of liquid alcohol diet daily from gestation day 6; control group B was fed a liquid diet in which dextrin replaced alcohol isocalorically; and alcohol＋$T_4$ group C received 35 calories of liquid alcohol diet and exogenous thyroxine subcutaneously. The features of the growth and maturation of rat brain tissue were observed at 0, 7, 14, 21 and 28 postnatal days via immunohistochemistry. Results : Group C showed prominent NPY immunoreactivity in the cerebral cortex compared to group A and B at P7. In group C, NPY-containing neurons were widely distributed in the all layers of cerebral cortex after P14. Also, numerical decreases of NPY-containing neuron were not found according to increasing age in group C. A decrease of NPY-containing neurons, however, was clearly observed in group A compared to group C at P28. In the hippocampus, similar patterns appeared in groups B and C after P7. Especially, in groups B and C, NPY-containing fibers formed plexus in the cerebral cortex and hippocampus at P14. Conclusion : These results suggest that the increase of NPY synthesis caused by maternal administration of exogenous thyroxine may convalesce fetal alcohol effects, one of the effects of the dysthyroid state following maternal alcohol abuse.

• Clinical and Experimental Pediatrics
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• v.45 no.7
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• pp.875-883
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• 2002

Purpose : Airways eosinophilia and increased IgE, characteristic features of asthma, result from a predominant Th2 response. In this study, we investigated the effect of CpG oligodeoxynucleotides (ODNs) on the inhibition of airways eosinophilia in mice with established airway inflammation. We also investigated the immunological mechanisms involved. Methods : Groups of BALB/c mice were sensitized intradermally with ovalbumin(OVA). At week 10, airway inflammation was induced by intranasal challenge of the mice with OVA. At week 14, the mice were challenged intranasally again with OVA in the presence and without the presence of CpG ODNs. Mice with saline administration served as negative controls. Bronchoalveolar lavage fluids(BALF) were obtained and eosinophils were counted. Th1 and Th2 cytokines in the spleen cell cultures were measured by ELISA. Serum OVA-specific IgE and IgG2a antibodies were also measured by ELISA. Results : BALF eosinophils were significantly inhibited in the CpG ODNs-treated mice(P<0.01). IgE and IgG2a levels increased significantly in both CpG ODNs-treated and untreated groups as compared to the negative control group; there was, however, no significant difference between the two groups four days after intranasal administration of CpG ODNs. Cytokine analysis revealed decreased production of IL-4, IL-5, and IL-13 and increased production of IL-12 in the CpG ODNs-treated group as compared to the untreated group. Interestingly, $IFN-{\gamma}$ levels were not upregulated in the CpG ODNs-treated group. Conclusion : CpG ODNs vaccination is a potentially useful approach for reversing airways eosinophilia in mice with established airways inflammation.

• Tuberculosis and Respiratory Diseases
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• v.57 no.5
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• pp.425-433
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• 2004

Background : Induction of oral tolerance (OT) has been known to prevent allergic inflammation in acute asthma model within 4 weeks. However it is remained whether induction of OT may effectively prevent allergic inflammation in chronic asthma model over 4 weeks. We observed the effect of induction of OT on allergic inflammation and airway remodeling in chronic asthma model up to 8 weeks. Methods : 5-week-old female BALB/c mice divided into 4 groups-control group, asthma group, low dose OT group, and high dose OT group. To induce oral tolerance mice were fed ovalbumin (OVA) before sensitization with OVA and aluminum hydroxide-1 mg for 6 consecutive days in the low dose OT group and 25 mg once in the high dose OT group. Mice in the asthma group were fed phosphate buffered saline instead of OVA. After sensitization followed by repeated challenge with aerosolized 1% OVA during 6 weeks, enhanced pause (Penh), inflammatory cells, IL-13, and IFN-${\gamma}$ levels in bronchoalveolar lavage (BAL) fluids as well as OVA-specific IgE, IgG1, and IgG2a levels in serum were measured. In addition the degree of goblet cell hyperplasia and peribronchial fibrosis were observed from lung tissues by PAS and Masson's trichrome stain. Results : Both OT groups showed a significant decrease in Penh, inflammatory cells, IL-13, and IFN-${\gamma}$ levels in BAL fluids as well as OVA-specific IgE, IgG1, and IgG2a levels in serum compared with the asthma group (P<0.05). In addition, the degree of goblet cell hyperplasia and peribronchial fibrosis were significantly attenuated in both OT groups compared with the asthma group (P<0.01). Conclusion : These results suggest that induction of OT may effectively prevent allergic inflammation as well as airway remodeling even in chronic asthma model up to 8 weeks.

• Korean Journal of Animal Reproduction
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• v.21 no.2
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• pp.157-165
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• 1997

포유류 배반포배의 epiblast는 내세포괴에 포함되어 있으며, 이 epiblast cells이 배 및 태아의 생식세포와 일반 체세포로 분화된다 (Beddington, 1986; Lawson 등, 1991). 그런데 조기에 발달된 부화배반포기 배가 지연발생된 부화배반포기 배보다도 많은 epiblast cells을 가지고 있다고 한다(Talbot 등, 1995). 그래서 본 연구에서는 체외수정 유래의 배반포배의 발육속도에 따른 내세포괴/영양배엽세포의 비율 및 배아간세포 확립 효율을 비교하여 발달일령 간에 차이가 있는지를 규명하고자 하였다. 공시한 소의 난포란을 TCM-199에 0.5$\mu\textrm{g}$/ml FSH, 5$\mu\textrm{g}$/ml LH, 10% FBS, 100 units/ml penicilin, 및 100$\mu\textrm{g}$/ml streptomycin을 첨가하여 39$^{\circ}C$, 5% CO2 조건하에서 24시간 동안 체외성숙한 후, 5$\mu\textrm{g}$/ml heparin으로 수정능이 획득된 1$\times$106 sperm/ml의 정자로 체외수정을 유도하였다. 체외수정 후 18~20시간에 과립막세포를 vortexing으로 제거하여 얻은 모든 체외수정란을 3mg/ml BSA, 20${\mu}\ell$/ml NEM amino acids, 40${\mu}\ell$/ml BME amino acids, 10mM glycine 및 1mM alanine이 함유된 CR1aa 배양액에서 BRL 단층세포와 공배양을 실시하였다. 수정후 7, 8 및 9일재 (체외수정일 : 0일)에 확장배반포기까지 발달한 수정란을 이중염색 및 배아간세포의 확립 실험에 공시하였다. 체외수정후 24시간에 분할된 총 1,145개의 수정란이 7, 8 및 9일째에 후기 배반포기까지 각각 222(15.6%), 103(7.2%) 및 52(3.6%)로 발달하여 총 377개 (26.4%)가 발달하였다. 내세포괴/영양배엽세포의 비율은 7일 및 8일째 배반포배에서 각각 47.2$\pm$11.9/95.1$\pm$24.4개 (33/67%) 및 40.3$\pm$12.4/83.3$\pm$26.9개 (33/67%)로서 9일째 배반포배의 19.3$\pm$8.1/62.3$\pm$23.1개 (24/76%) 보다 유의적(P<0.05)으로 높았다. ES-like cells을 확립하기 위하여 후기배반포기 배를 mouse embryonic fibroblast 단층 공배양기에 옮긴 후 5일에 내세포괴의 부착 여부를 판정하고, 10일에 배아간세포의 확립 여부를 판정하였다. 그 결과 7, 8 및 9일째의 배반포기배의 각각 47.7% (82/172), 30.9%(22/71) 및 15.6%(5/32)에서 배아간세포가 확립되었다. 이상의 결과에서 배반포기까지의 발육이 빠른 수정란에서 영양배엽에 대한 내세포괴세포의 비율이 높았고 배아간세포의 확립율도 높다는 사실을 입증할 수 있었으며, 이와 같은 결과에서 체외수정란 유래 배반포배의 질을 결정할 수 있을 것으로 생각된다.

• Korean Journal of Animal Reproduction
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• v.17 no.2
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• pp.149-157
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• 1993

A new technique was established to maximize the numbers of follicular oocytes recovered from the ovaries obtained at the slaughter house. And their further developmental capacity was demonstrated. There recovery techniques were used; aspiration (ASP, control), slicing (SLC) and slicing combining aspiration (ASP+SLC). Recovered oocytes were cultured in TCM 199+15% FCS+gonadotrophins in an atmosphere of 5% CO$_2$ in air at 39$^{\circ}C$ for 24 h. The nuclear maturation was detemined with chromo-some configuration by rapid staining. And cytoplasmic maturation was examined by the formation of female pronuclei with parthenogenetic activation of the matured oocyte after 18 h of co-culture with granulosa cell monolayer. Total 1,641 bovine follicular oocytes recovered from 245 ovaries. The number of oocytcs per ovary was 1.87 in ASP, 11.05 in SLC and 7.88 in ASP+SLC, respectively. SLC would yield 5.9 folds increase, compared with ASP. The rate of maturation were 92.9% in ASP, 79.1% in SLC and 71.7% in ASP+SLC, respectively. Although the maturation rate in ASP was the highest, metaphase II oocytes per ovary in SLC was 5 times higher than that of ASP. The rates of pronuclei formation upon ethanol activation were 75% in ASP, 67% in SLC and 62.5% in ASP+SLC, respectively. The results demonstrate that it should be possible to maximize the number of the follicular oocyte from the ovary for mass production of bovine embryos. Thus the established technique may provide efficient supply of bovine embryos for biochemical and molecular study of early bovine embryos.

• KSBB Journal
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• v.19 no.2
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• pp.118-124
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• 2004

Melanocytes synthesize melanin within discrete organelle termed melanosomes which are transferred to the surrounding keratinocytes and can be produced in varying sizes, numbers and densities. Skin whitening products have become increasingly popular in the past few years. The most successful natural skin whitening agents are: Arbutin, Vitamin C, Kojic acid, Mulberry, which are all tyrosinase inhibitors. In this work, melanoston, a melanogenesis inhibitor isolated from yeast was studied to understand its mechanism of melanogenesis inhibition. It was found that melanoston was not a tyrosinase inhibitor, while when melanoston was applied to the B16 melanoma cell culture media, the intracellular tyrosinase activity was decreased by more than 30％, When B16 melanoma was stimulated with ${\alpha}$-MSH, cell morphololgy was dramatically changed to have lots of dendrites on the cell membrane surface. On the other hand, B16 was treated with ${\alpha}$-MSH and melanoston, simultaneously, the change of cell morphology was not so great. This inhibition effect of melanoston was found to be related to the inhibition of intracellular activation and transportation of tyrosinase, which was observed by immunostaining of B16 melanoma using anti-tyrosinase antibody. From these results, melanoston was regarded as an inhibitor to the differentiation of melanoma cells.

• Journal of the Society of Cosmetic Scientists of Korea
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• v.31 no.1 s.49
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• pp.25-33
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• 2005

Melanocytes synthesize melanin within discrete organelle termed melanosomes which are transferred to the surrounding keratinocytes and can be produced in varying sizes, numbers and densities. Skin whitening products have become increasingly popular in the past few years. The most successful natural skin whitening agents are: arbutin, vitamin C, kojic acid, and mulberry, which are all tyrosinase inhibitors. In this work, melanoston, a melanogenesis inhibitor isolated from yeast was studied to understand its mechanism of melanogenesis inhibition. It was found that melanoston was not a tyrosinase inhibitor, while when melanoston was applied to the B16 melanoma cell culture media, the intracellular tyrosinase activity was decreased by more than $30\%$. When B16 melanoma was stimulated with $\alpha$-MSH, cell morphololgy was dramatically changed to have lots of dendrites on the cell membrane surface. On the other hand, B16 was treated with $\alpha$-MSH and melanoston, simultaneously, the change of cell morphologv was not so great. This inhibitory effect of melanoston was found to be related to the inhibition of intracellar activation and transportation of tyrosinase, which was observed by irmmunostaining of B16 melanoma using anti-tyrosinase antibody. From these results, melanoston was regarded as an inhibitor to the differentiation of melanoma cells.