• Journal of Genetic Medicine
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• v.2 no.2
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• pp.65-70
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• 1998

Lysosomal acid lipase (LAL) plays a central role in the intracellular degradation of neutral lipids derived from plasma lipoproteins. In this study, we investigated the missense mutation within exon 2 of human LAL gene changing of codon -6 of prepeptide from threonine to proline. The Thr-6Pro mutation was detected by the HaeIII restriction fragment length polymorphism (RFLP) and single-strand conformation polymorphism (SSCP). We analyzed the mutation in subjects with 221 unrelated randomly selected control samples and 86 patients with familial hypercholesterolemia (FH) in Korea. We observed that mutation is present with high frequency in Korea compared to other populations studied previously. The frequency of PP homozygote in the FH group was observed considerably higher than that of control. However, there was no significant difference of genotype frequency between two groups. These results, together with the fact that plasma lipids and lipoproteins levels between genotypes showed no statistical difference, suggest that the Thr-6Pro mutation in the LAL gene may have no association with the increased risk of FH development.

• BMB Reports
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• v.35 no.6
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• pp.553-561
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• 2002

A gene that encodes a protein homologue to baculoviral IE-1 was identified and sequenced in the genome of the Choristoneura fumiferana granulovirus (ChfuGV). The gene has an 1278 nucleotide (nt) open-reading frame (ORF) that encodes 426 amino acids with an estimated molecular weight of 50.33 kDa. At the nucleotide level, several cis-acting regulatory elements were detected within the promoter region of the ie-1 gene of ChfuGV along with other studied granuloviruses (GVs). Two putative CCAAT elements were detected within the noncoding leader region of this gene; one was located on the opposite strand at -92 and the other at -420 nt from the putative start triplet. Two baculoviral late promoter motifs (TAAG) were also detected within the promoter region of the ie-1 gene of ChfuGV. A single polyadenylation signal, AATAAA, was located 18nt downstream of the putative translational stop codon of ie-1 from ChfuGV. At the protein level, the amino acid sequence data that was derived from the nucleotide sequence in ChfuGV IE-1 was compared to those of the Cydia pomonella granulovirus (CpGV), Xestia c-nigrum granulovirus (XcGV) and Plutella xylostella granulovirus (PxGV). The C-terminal regions of the granuloviral IE-1 sequences appeared to be more conserved when compared to the N-terminal regions. A domain, similar to the basic helix-loop-helix like (bHLH-like) domain in NPVs, was detected at the C-terminal region of IE-1 from ChfuGV (residues 387 to 414). A phylogenetic tree for baculoviral IE-1 was constructed using a maximum parsimony analysis. A phylogenetic estimation demonstrates that ChfuGV IE-1 is most closely related to that of CpGV.

• Molecular & Cellular Toxicology
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• v.1 no.2
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• pp.111-115
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• 2005

Ultraviolet (UV) radiation to mammalian skin is known to alter cellular function via generation of Reactive Oxygen Species (ROS), DNA damage and DNA lesions, such as pyrimidine dimmers and photoproducts, which could lead to DNA mutation if they are not repaired. In this study, we have investigated the reduction of DNA damage and of apoptosis with a particular attention to genetic effect of paeoniflorin in Normal Human Epidermal Keratinocytes (NHEK). After UVB irradiation from $10\;to\;500mJ/cm^{2}$ to NHEK, Mean Tail Moments (MTM) were increased with UVB dose increase. The greatest amount of strand breaks was induced at $500mJ/cm^{2}$ of UVB. Even at the lowest dose of UVB ($10mJ/cm^{2}$), change in MTM was detected (P<0.0001). Pretreated cell with 0.1% paeoniflorin maximally reduced the level of DNA damage to about 21.3%, compared to untreated cell. In the lower concentrations less than 0.01% of paeoniflorin, MTM had a small increase but paeoniflorin still had reductive effects of DNA damage. We measured the apoptosis suppression of paeoniflorin with annexin V flous staining kit. As we observed under the fluorescence microscopy to detect apoptosis in the irradiated cell, the fluorescence intensity was clearly increased in the untreated cell, but decreased in treated cells with paeoniflorin. These results suggest that paeoniflorin reduces the alteration of cell membranes and prevents DNA damage. Therefore, the use of paeoniflorin as a free radical scavenger to reduce the harmful effects of UV lights such as chronic skin damage, wrinkling and skin cancer can be useful to prevent the formation of photooxidants that result in radical damage.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.12
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• pp.6055-6057
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• 2012

The mismatch repair system (MMR) is a post-replicative DNA repair mechanism whose defects can lead to cancer. The MSH3 protein is an essential component of the system. We postulated that MSH3 gene polymorphisms might therefore be associated with prostate cancer (PC). We studied MSH3 codon 222 and MSH3 codon 1036 polymorphisms in a group of Iranian sporadic PC patients. A total of 60 controls and 18 patients were assessed using the polymerase chain reaction and single strand conformational polymorphism. For comparing the genotype frequencies of patients and controls the chi-square test was applied. The obtained result indicated that there was significantly association between G/A genotype of MSH3 codon 222 and G/G genotype of MSH3 codon 1036 with an increased PC risk (P=0.012 and P=0.02 respectively). Our results demonstrated that MSH3 codon 222 and MSH3 codon 1036 polymorphisms may be risk factors for sporadic prostate cancer in the Iranian population.

• Korean Journal of Ecology and Environment
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• v.41 no.3
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• pp.331-337
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• 2008

Polycyclic aromatic hydrocarbons (PAHs) derived from leakage of fossil fuels and incomplete combustion of organic materials have been considered as harmful contaminants in environments. This study evaluated the effect of benzo[k]fluoranthene (BkF), one of the PAHs, using the multiple biomarkers and applied the integration model with those biomarker responses. After 10 days of the exposure at the measured concentrations of BkF (6, 25, and 45 ${\mu}g\;L^{-1}$), the changes of the four biomarkers, that is, 7-ethoxyresorufin-O-deethylase (EROD), DNA single-strand breaks (Comet), acetylcholinesterase (AChE) and vitellogenin (VTG) in the common carp (Cyprinus carpio) were observed. The standardized values of four biomarker responses were computed and integrated as star plots, representing Integrated Biomarker Respnse (IBR) values. DNA damage was induced in a dose-dependent manner, and increased significantly compared with that in the control. EROD and VTG levels were significantly elevated at low concentrations of BkF. On the other hand, AChE activities were not altered by BkF. IBR values increased as the exposure concentrations increased. Thus, the metabolic, endocrine and genetic changes of the biomarker responses in the common carp exposed to BkF should be considered in the case of the ecological risk assessment of the BkF in fish and it can be used as a biomonitoring tool in aquatic ecosystems. In addition, star plots can be used as a useful analysis tool in multibiomarker integration approach.

• Korean Journal of Veterinary Research
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• v.38 no.3
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• pp.565-573
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• 1998

To study the specific tools for the diagnosis of Newcastle disease virus (NDV) in chicken, polymerase chain reaction (PCR) and its presumable conditions were evaluated for the detection of hemagglutinin-neuraminidase (HN) gene of NDV RNA. For these purposes, Kyojeongwon strain of the NDV was propagated in allantoic cavity of SPF embryonating chicken eggs, and viral RNA was extracted from fractionated virus after the allantoic fluids were ultracentrifuged with sucrose gradient. The first-strand cDNA was then made for the HN gene of NDV RNA by reverse transcription at $42^{\circ}C$ for 1 hour using specific primer complementary to the HN gene. The single-stranded cDNA was used as template in the PCR of the HN-DNA, and various conditions of the PCR were evaluated to set up method for the specific detection of the HN-DNA. The PCR conditions promising for the detection of HN gene consist of preheating at $94^{\circ}C$, 5 min, 30 cycles of denaturation at $94^{\circ}C$, 1 min, annealing at $55^{\circ}C$, 1 min and polymerization at $72^{\circ}C$, 2 min, and a cycle of extension at $72^{\circ}C$, 5 min. when NDVs of allantoic fluids without fractionation were applied to the above PCR condition, the HN genes were detected effectively not only from Kyojeongwon but from other velogenic strains such as Herts and a field isolate.

• Journal of Gastric Cancer
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• v.3 no.4
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• pp.206-213
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• 2003

Purpose: The clinical implication of p53 mutation in gastric cancer is still unclear, as shown by the discordant results that continue to be reported in the literature. Materials and Methods: To assess p53 gene mutation, tumor p53 overexpression, and serum anti-p53 antibody, we employed a polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) analysis, an immunohistochemistry using monoclonal antibody DO-7, and an enzymelinked immunosorbent assay (ELISA), respectively. Results: Of 169 surgical specimens of gastric cancer, mutation at exon $5\∼8$ of the p53 was identified in 33 ($19.5\%$) and was significantly correlated with lymph node metastasis. Overexpression of p53 was found in 62 specimens ($36.7\%$) and had a significant correlation with tumor differentiation. Serum anti-p53 antibody was positive in 18 patients ($10.7\%$). Twenty-three of the mutated tumors ($69.7\%$) and 39 of the non-mutated tumors ($28.7\%$) displayed immunoreactivity. Twelve of the immunopositive tumors ($19.4\%$) and 6 of the immunonegative tumors produced anti-p53 antibody. These differences were statistically significant (P<0.001 and P=0.005, respectively). There was no significant difference in survival according to the mutation of p53. Conclusion: Mutation and overexpression of p53 can be easily detected by immunohistochemistry. However, standardization of the immunohistochemical staining method, as well as guidelines for interpreting the stained result, will produce concordant results and thereby improve clinical application.

• Asian-Australasian Journal of Animal Sciences
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• v.18 no.5
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• pp.734-740
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• 2005

The blood samples were collected from dairy cows at the same milking stage. The single-strand conformation polymorphism (PCR-SSCP) method was used to analyze for polymorphism at the 5'flanking region of the hsp70 gene. The mRNA expression levels of HSP70, HSF1, Bcl-2 and Bax-$\alpha$ at different daily-mean-temperature were analyzed by relative quantitative RTPCR. The DNA content, cell phase and the ratio of apoptosis of lymphocytes in peripheral blood of dairy cattle at different daily-meantemperature were determined by FCM. The PCR-SSCP products of primer pair 1 showed polymorphisms and could be divided into four genotypes: aa, ab, ac, cc, with the cis-acting element (CCAAT box) included. Mutations in the hsp70 5'flanking region (468-752 bp) had different effects on mRNA expression of HSP70, HSF1, Bcl-2 and Bax-$\alpha$. The ac genotypic cows showed higher expressions of HSP70mRNA, HSF1mRNA and Bcl-2mRNA/Bax-$\alpha$mRNA and lower ratio of apoptosis. These mutation sites can be used as molecular genetic markers to assist selection for anti-heat stress cows.

• Proceedings of the Korea Society of Environmental Toocicology Conference
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• 2002.10a
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• pp.169-169
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• 2002

The mobilizable amount of transition metals is a fraction of the total amount of the metal from urban particulate matter. Although the fraction is small, some metals (Fe, Cu) are the major participants in a reaction that generates reactive oxygen species (ROS), which can damage various biomolecules. Damaging effects of the metals can be measured by the single strand breakage (SSB) of X174 RFI DNA or the carbonyl formation of protein. In another study, we have shown that more metals are mobilized by PM2.5 than by PM10 in general. DNA SSB of >20% for PM2.5 and >15% for PM10 was observed in the presence of chelator (EDTA or citrate)/reductant (ascorbate), compared to the control (<3%) only with the chelator. The carbonyl formation by both PMs was very similar in the presence of the chelator, regardless of the kind of proteins. Compared to the control in the absence of chelator/reductant, 3.3 times and 4.9 times more carbonyl formation for PM2.5 and PM10, respectively, was obtained with BSA in the presence of chelator/reductant, showing that PM10 induced 33% more damage than PM2.5. However, 4.8 times and 1.9 times more carbonyl formation for PM2.5 and PM10, respectively, was observed with lysozyme in the presence of chelator/reductant, showing that PM2.5 induced 250% more damage than PM10. Although different proteins showed different sensitivities toward ROS, all these results indicate that the degrees of the oxidation of or damage to the biomolecules by the mobilized metals were higher with PM2.5 than with PM10. Therefore, it is expected that more metals mobilized from PM2.5 than from PM10, more damage to the biomolecules by PM2.5 than by PM10. We suggest that when the toxicity of the dust particle is considered, the particle size as well as the mobilizable fraction of the metal should be considered in place of the total amounts.

• The Journal of Korean Society of Virology
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• v.30 no.1
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• pp.29-37
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• 2000

The aim of this study was to investigate viral etiology in dilated cardiomyopathy (DCM) by polymerase chain reaction (PCR) or nested reverse transcription PCR (RT-PCR), and characterize the enteroviral RNA presented in the clinical specimens. Twenty-eight paraffin-embedded heart tissue samples were assayed to detect cytomegalovirus, herpes simplex virus type 1, type 2, parvovirus, adenovirus, and enterovirus (EV) with each specific primer. Of these 28 patients (mean age: 27, M: 24, F: 4), 26 were histologically diagnosed as DCM and 2 as myocardial infarction (MI). Nested RT-PCR detected enteroviral RNA in 7 (26.9%) of 26 patients with DCM, and none of patients with MI. And none of DNA viruses tested were detected from the samples. Amplified products were also genotyped by single-strand conformation polymorphism (SSCP). Three subtypes can be differentiated from 7 clinical specimens. Furthermore, direct sequence analysis was performed to determine whether genetic variation of EV is present in the explanted heart tissues from patients with DCM. Although most of the sequences among the wild isolates have the greatest similarity to those of coxsackievirus B3, there are specific regions of variable sequences (no 490 - no 510). The data suggest that enterovirus may be a major viral pathogen for the DCM in Korea and nucleotide sequence data indicate that coxsackievirus B3 may be a leading etiologic agent of DCM.