• KSBB Journal
• /
• v.20 no.5 s.94
• /
• pp.363-370
• /
• 2005

Neural stem cells (NSCs) of the central nervous system (CNS) have raised a great interest not only for their importance in basic neural development but also for their therapeutic potentials in neurologically degenerative diseases such as Parkinson's, Alzheimer and stroke. During the CNS development, two molecular cascades determine specification of midbrain dopamine system. In one pathway, FGF-8, sonic hedgehog and transcription factor Nurr1 specify dopamine neurotransmitter phenotype. In the other, transcription factors $Lm{\times}lb\;and\;Pt{\times}3$ are required for induction of dopaminergic neurons. In Nurr1 knockout mouse, tyrosine hydroxylase (TH) positive cells fail to appear in substantia nigra, indicating that Nurr1 is essential in specification of dopaminergic cell phenotype. In this study, we used the immortalized human NSCs retrovirally transduced with Nurr1 gene to probe the Nurr1 mediated mechanism to induce dopamine phenotype. While Nurr1 over-expression alone did not generate dopamine phenotype in NSCs, applications of retinoid and forskolin induced expression of TH and AADC mRNAs. In addition, co-cultures of Nurr1 expressing NSCs with human astrocytes induced a marked increase of TH expression. In this co-culture system, the addition of retinoid and forskolin dramatically increased expression of TH. These results indicate that the immortalized human NSCs with Nurr1 gene could have a clinical utility for cell replacement for the Parkinson patients.

• Journal of the Society of Cosmetic Scientists of Korea
• /
• v.31 no.1 s.49
• /
• pp.43-49
• /
• 2005

Recently, according as people who have sensitive skin increase, we've been giving more importance to the safety of cosmetics. Especially, preservative is known to be one of the main stimuli which cause side-effects of cosmetics. However, there have been few reports describing cell cytotoxicity, skin penetration, oil-aqueous phase partition, anti-microbial activity of preservatives and their correlation with skin irritation. The study is aimed to develop low irritable preservative system with phenoxyethanol, one of the most commonly used preservatives in cosmetics, considering various factors mentioned above. According to our results of cell cytotoxicity against human normal fibroblasts by means of MTT assay, phenoxyethanol showed the lowest cytotoxicity when compared to other preservatives tested (cytotoxicity: pro-pylparaben > butylparaben > ethylparaben > methylparaben > triclosan > phenoxyethanol), but human patch test for assessing shin primary irritation revealed that phenoxyethanol has higher skin irritation than methylparaben and triclosan. We performed in vitro skin penetration test using horizontal Franz diffusion cells with skin membrane prepared from hairless mouse (5 ${\~}$ 8 weeks, male) to evaluate the rate of skin penetration of preservatives. From the results, we found that the higher irritable property of phenoxyethanol in human skin correlates with its predominant permeability (skin penetration: phenoxyethanol > methylparaben > ethylparaben > propylparaben > butylfaraben > triclosan). Therefore, we made an effort to reduce skin permeability of phenoxyethanol and found that not only the rate of skin penetration of phenoxyethanol but also its skin irritation is dramatically reduced in formulas containing oils with low polarity. In the experiments to investigate the effect of oil polarity on the oil-aqueous phase partition of phenoxyethanol, more than $70\%$ of phenoxyethanol was partitioned in aqueous phase in formulas containing oils with low polarity, while about $70 {\~} 90\%$ of phenoxyethanol was partitioned in oil phase in formulas containing oils with high polarity. Also, in aqueous phase phenoxyethanol showed greater anti-microbial activity. Conclusively, it appears that we can develop less toxic preservative system with reduced use dosage of phenox-yethanol and its skin penetration by changing oil composition in formulas.

• Molecules and Cells
• /
• v.26 no.5
• /
• pp.443-453
• /
• 2008

The Bric-a-brac, Tramtrack, Broad-complex (BTB) domain is a protein-protein interaction domain that is found in many zinc finger transcription factors. BTB containing proteins play important roles in a variety of cellular functions including regulation of transcription, regulation of the cytoskeleton, protein ubiquitination, angiogenesis, and apoptosis. Here, we report the cloning and characterization of a novel human gene, KLHL31, from a human embryonic heart cDNA library. The cDNA of KLHL31 is 5743 bp long, encoding a protein product of 634 amino acids containing a BTB domain. The protein is highly conserved across different species. Western blot analysis indicates that the KLHL31 protein is abundantly expressed in both embryonic skeletal and heart tissue. In COS-7 cells, KLHL31 proteins are localized to both the nucleus and the cytoplasm. In primary cultures of nascent mouse cardiomyocytes, the majority of endogenous KLHL31 proteins are localized to the cytoplasm. KLHL31 acts as a transcription repressor when fused to GAL4 DNA-binding domain and deletion analysis indicates that the BTB domain is the main region responsible for this repression. Overexpression of KLHL31 in COS-7 cells inhibits the transcriptional activities of both the TPA-response element (TRE) and serum response element (SRE). KLHL31 also significantly reduces JNK activation leading to decreased phosphorylation and protein levels of the JNK target c-Jun in both COS-7 and Hela cells. These results suggest that KLHL31 protein may act as a new transcriptional repressor in MAPK/JNK signaling pathway to regulate cellular functions.

• Development and Reproduction
• /
• v.7 no.1
• /
• pp.49-56
• /
• 2003

There have been reports that administrated high-dose gonadotropin-releasing hormone-agonist(GnRH-Ag) suppresses endogenous gonadotropin production and inhibits function of ovary. In human IVF-ET program, however, GnRH-Ag is employed in large amounts during superovulation induction resulting to luteal phase defects which must be supported with progesterone. To elucidate the reason of luteal phase defects by GnRH-Ag, the aim of this study was to investigate the apoptosis changes in the ovary and the hormonal changes in the serum after GnRH-Ag and PMSG administration in adult mice in a method similar to human superovualtion induction. GnRH-Ag(10 ${\mu}$g) or saline was injected every 12h beginning 48h prior to PMSG injection until 48h at)or PMSG injection when blood sampling and ovary collection was performed. In results, the ovary weight in the GnRH-Ag only injection group was significantly lower when compared with the other two groups, PMSG only or PMSC + GnRH-Ag injection. The ratio of preantral follicles in the ovary are increased in the GnRH-Ag only group, while the ratio of antral follicles are decreased and the corpus luteum ratio is increased in the PMSG + GnRH-Ag group. The proportion of all follicles showing apoptosis in the GnRH-Ag only in.iection group was seen to be more than twice the proportion seen in the PMSC only injection group, and such increased apoptosis is decreased after addition of PMSC. The serum levels of both estradiol and progesterone were significantly lower in the CnRH-hg only group compared to those in the other two groups. When the administration of GnRH-Ag were followed by PMSG in;ection, however, estradiol concentration was completely recovered compared to the serum level of PMSG group, but not progesterone level. In conclusion the use of GnRH-Ag in human IVF-ET program may induce the apoptosis and the suppression of hormone production by ovary leading to luteal phase defects, thus adequate progesterone support seems to be necessary against them.

• Journal of Animal Science and Technology
• /
• v.50 no.6
• /
• pp.753-762
• /
• 2008

The primers for RT-PCR and RACE-PCR were designed by aligning the pig genomic sequence and the human complement factor B(CFB) coding sequence(CDS) from the GenBank. Each PCR product was amplified in pig cDNA and sequencing was carried out. The CDS length of pig CFB gene was determined to be 2298 bp. In addition, the pig CDS was more longer than human and mouse orthologs because of insertion and deletion. The identities of porcine nucleotide sequences with those of human and mice were 84% and 80%, and the identities of amino acids were 79% to 77%, respectively. Three complement control protein(CCP) domains, one Von Willebrand factor A(VWFA) domain and a serine protease domain, that are revealed typically in mammals, were found in the pig CFB gene. Based on the CDSs determined, the primers were designed in intron regions for amplification of entire length of exons. In amplification and direct sequencing with genomic DNAs of six pig breeds, three cSNPs(coding single nucleotide polymorphisms) were identified and verified as missense mutations. Using the Multiplex-ARMS method, we genotyped and verified the mutations identified from direct sequencing. To demonstrate recrudescence, we performed both direct sequencing and Multiplex-ARMS with two randomly selected DNA samples. The genotype of each sample exhibited the same results using both methods. Therefore, three cSNPs were identified from pig CFB gene and that can be used for haplotype analysis of the swine leukocyte antigen(SLA) class III region. Moreover, the results indicate that the Multiplex-ARMS method should be powerful for genotyping of genes in the SLA region.

• The korean journal of orthodontics
• /
• v.26 no.1 s.54
• /
• pp.83-94
• /
• 1996

Substance P is one of the neuropeptide which presents highly in tension site of periodontal ligament during the orthodontic tooth movement. It has bnn also hon as one of the neuropeptides which cause neurogenic inflammation in various tissues and organs. However, there is no report about the effect of substance P on major extracellular matrix protein, collagen production. The purpose of this study was to evaluate the collagen production by substance P in human periodontal ligament cell. The collagenase-digestion method was used to evaluate collagen production and also used Northern blot hybridization for the evaluation of collagen mRNA level. This study also Included in terms of prostanglandins and gelatinase production with respect to collagen production. For the collagen degradation, zymography was used to estimate denatured collagen degradation. Dose-dependent effect of substance P on noncollagen protein, collagen, and percent collagen was that substance P increased noncollagen protein synthesis, but decreased collagen sytnsis. So the percent collagen, which determined by relative collagen production against total protein production, w3s decreased from $7\%\;to\;3.6\%$. This inhibitory effect of substance P on collagen production was disappeared when cells were treated concomitantly with indomethacin. It means that substance P-induced inhibitory effect on collagen production was due at least in part to the production of prostaglandins. To evaluate whether substance P-induced inhibitory effect on collagen production is correspond to the steady-state levels of procollagen mRNA, Northern blot hybridization was performed and it showed that substance P has no effect on the steady-slate level of ${\alpha}1(I)$ procollagen mRNA. It means that the inhibitory effect of substance P on collagen production was due to the change of a certain mechanism after posttranscription. In this context, gelatinase production by substance P in periodontal ligament cells was evaluated by zymography. Zymogram showed that substance P has no effect on gelatinase production in periodontal ligament cells. To explore wheter substance P-induced inhibitory effect on collagen production is selevtive in periodontal ligament cells or not, MC3T3-E1 cells which originated from mouse calvaria was used. It showed that substance P has no effect on collagen production in MC3T3-E1 cells. Taken together, substance P inhibits collagen production in human periodontal ligament cells. This effect was not due to the change of the steady-state level of procollagen mRNA and gelatinase production, but due at least in part to the change of prostaglandins production.

• Korean Journal of Food Science and Technology
• /
• v.40 no.6
• /
• pp.674-679
• /
• 2008

Scutellaria radix (SR) has been utilized as a traditional medicine for a variety of diseases including Rheumatoid arthritis and its major flavonoids - baicalein, baicalin, and wogonin - have been reported to exert beneficial health effects, including anti-bacterial, anti-viral, anti-inflammatory, and free-radical scavenging. However, the mechanisms underlying this effect remain poorly understood. The principal objective of this study was to determine the effect of SR on osteoblast and osteoclast cells. SR extract was prepared using 70% ethanol solvent. Osteoblastic MC3T3-E1 cells and osteoclast precursor Raw 264.7 macrophage cells were utilized. SR extract increased MC3T3-E1 cell proliferation and stimulated alkaline phosphatase activity dose-dependently, 152.0% of the control at concentration $1{\mu}g/mL$. Additionally, SR extract ($1{\mu}g/mL$) stimulated Bone nodule formation activity in MC3T3-E1 cells, approximately 223.3% of the control, 20 days after the exposure. In addition, SR extract significantly reduced the number of tartrate-resistant acid phosphatase-positive (TRAP+) multinucleated cells from Raw 264.7 cells. In conclusion, SR extract stimulates the proliferation and bioactivities of boneforming osteoblasts, and inhibits the activities of bone-resorbing osteoclasts to a certain degree.

• Korean Journal of Microbiology
• /
• v.41 no.1
• /
• pp.24-37
• /
• 2005

This study was conducted to analyze the molecular epidemiological properties and to select the most efficient and reliable PCR method on 116 of Staphylococcus aureus (S. aureus) isolates from Korean cattle, black goat, pig, dog, chicken, mouse and also human clinical cases from hospital. The distribution patterns of SSG [species specific genes; coagulase (coa), protein A (spa), nuclease (nuc) and aroA (RsaI) gene] were analyzed by PCR method. Among the SSGs, the nuc-gene was found in all strains $(100\%)$ tested and followed by coa-gene $(87.9\%)$, spa-gene $(91.4\%)$ and aroA-gene $(26.7\%)$, in order. The genetic subtyping by RFLP method was performed on the coa [AluI] and aroA-gene [RsaI] PCR products. The mecA-gene PCR and PCR-RFLP techniques were chosen to detect and verify of MRSA strains. Only the human strains $(12.1\%)$ were detected the positive mecA-gene products (533 bp), which were divided into two specific bands [201 & 332 bp] by HhaI enzyme digestion. On coa-gene and spa-gene typing, coa-gene was typed with ten kinds of genotype and coa-3 type were determined as the most predominant genotype, while spa-gene was divided into eleven kinds of genotype and also spa-7 type were selected the most prevalent genotype based on their genetic variations. On the aroA and coa-gene subtyping by PCR-RFLP, aroA-gene products were discriminated with only seven types of genotype, while coa-gene products were further divided into an eleven genotype, respectively. In comparison of SID values of five PCR based typing methods, the coa-PCR-RFLP (SID0.894) was evaluated the most efficient and reliable tools and followed by coa-PCR (SID0.883) and aroA-PCR-RFLP (SID0.462), in order. In conclusion, we could determined that the coa-PCR-RFLP method was the most suitable genetic analysis tool for S. aureus and MRSA strains from domestic animals and humans.

• The Korean Journal of Nuclear Medicine
• /
• v.38 no.6
• /
• pp.528-531
• /
• 2004

Purpose: PET has some disadvantage in the imaging of small animal due to poor resolution. With the advent of microPET scanner, it is possible to image small animals. However, the image quality was not good enough as human image. Due to larger brain, cat brain imaging was superior to mouse or rat. In this study, we established the cat brain infarction model and evaluate it and its temporal charge using microPET scanner. Materials and Methods: Two adult male cats were used. Anesthesia was done with xylazine and ketamine HCl. A burr hole was made at 1cm right lateral to the bregma. Collagenase type IV 10 ${\mu}l$ was injected using 30 G needle for 5 minutes to establish the infarction model. $^{18}F$-FDG microPET (Concorde Microsystems Inc., Knoxville, TN) scans were performed 1, 11 and 32 days after the infarction. In addition, $^{18}F$-FDG PET scans were performed using human PET scanner (Gemini, Philips medical systems, CA, USA) 13 and 47 days after the infarction. Results: Two cat brain infarction models were established. The glucose metabolism of an infarction lesion improved with time. An infarction lesion was also distinguishable in the human PET scan. Conclusion: We successfully established the cat brain infarction model and evaluated the infarcted lesion and its temporal change using $^{18}F$-FDG microPET scanner.

• Development and Reproduction
• /
• v.4 no.2
• /
• pp.215-220
• /
• 2000

Most of endocrine disrupters (EDs) have been reported to exhibit estrogenic or anti-androgenic activity and thereby may disrupt reproductive development in human or wildlife. This study was performed to investigate the effects of estrogen (E$_2$), bisphenol (BP) and octylphenol (OP) on the mouse Leydig cell line (TM3). TM3 originated from testis of 11~13-daly-old BALB/c nu/＋ mice was cultured in DMEM supplemented with 10％ FBS alone or medium with estrogen (E$_2$), bisphenol (BP) and octylphenol (OP; 1 pM, 1 nM, 1 $\mu$M, 1 mM, respectively) for 48 hours. After culture, total cell number and viability were assessed by heamocyto-meter and trypan blue stain. Expression of cytochrome P450scc (CYPscc) mRNA whose product is involved in steroid hormone biosynthesis and estrogen receptor $\alpha$(ER $\alpha$) mRNA were detected by RT-PCR. As a result, treatment of TM3 with E$_2$, BP and OP(1 mM, respectively) significantly decreased the viability but not all of groups as high as 1 $\mu$M. Exposure of TM3 to OP significantly reduced the total cell number but not E$_2$ or BP. The expression of CYPscc mRNA was slightly reduced in BP (1 nM, 1 $\mu$M) and significantly decreased in OP (1 nM, 1 $\mu$M) treated TM3, except E$_2$ group. But the expression of ER $\alpha$ mRNA was sightly increased in all treated groups. In conclusion, BP and OP (high concentration) might inhibit steroidogenesis by decreasing the CYPscc mRNA expression in the mouse testis. These results suggest that BP and OP might impair spermatogenesis and subsequently disturb testicular function.