• Development and Reproduction
• /
• v.6 no.2
• /
• pp.117-122
• /
• 2002

Dopamine(DA), norepinephrine(NE), and epinephrine(E) belong to a class of neurotransmitters known as catecholamine (CA) which are synthesized and secreted by mammalian brain and adrenal medulla. CA regulate several behavior patterns connected with breeding, and regulate GnRH-gonadotropin hormone axis' vitality between hypothalamus-pituitary gland linking with reproduction freeze. The present study examined effects of sex steroid hormone on the transcriptional activities of CA biosynthesis enzymes, tyrosine hydroxylase(TH), dopamine $\beta$ -hydroxylase(DBH), and phenylethaolamine-N-methyl transferase(PNMT). Mature female rats were ovariectomized(OVX) and implanted with 17 $\beta$-estradiol(E$_2$: 500 $\mu\textrm{g}$/ml) or sesame oil. Forty-eight hours after implantation all the animals were sacrificed. Total RNAs were extracted immediately and were applied to semi-quantitative reverse transcription-polymerase chain reaction(RT-PCR). The expression level of TH was appeared by hypothalamus > SNc> adrenal medulla orders in OVX＋Oil group, and by SNc > hypothalamus) adrenal medulla orders in OVX+E$_2$ group. Treatment with E$_2$ significantly increased TH expression in SNc and adrenal medulla but in hypothalamus, the reduced TH expression was observed. The expression level of DBH was appeared by adrenal medulla > SNc > hypothalamus orders in OVX＋Oil group and in OVX＋E$_2$ group. Administration of E$_2$ significantly reduced DBH expression in SNc, and increased in adrenal medulla. Two cDNA products, large(PNMT1) and small(PMNTs) species of 110bp difference, were amplified in SNc and hypothalamus, but only PNMTs was observed in adenal medulla. The PNMTs expression level was in the order of adrenal medulla > hypothalamus > SNc in both OVX＋Oil and OVX＋E$_2$ group. The PNMTs expression in SNc and adrenal medulla was significantly increased byE$_2$. The present report demonstrated that estrogen effects on transcriptional activities for CA biosynthethic enzymes were tissue specific in adrenal medulla as well as different region of brain. These results suggest that it might be crucial relationship between the type of estrogen receptor and CA enzyme gene expression.

• Korean Journal of Clinical Laboratory Science
• /
• v.49 no.1
• /
• pp.28-39
• /
• 2017

Persistent organic pollutants (POPs) can affect epigenetic mechanisms and obesity development. Polybrominated diphenyl ethers (PBDEs)-widely used to make flames-are one of the important POPs. Prenatal exposure to endocrine disrupting chemicals (EDCs), such as POPs, may affect global DNA methylation in long interspersed nuclear elements (LINE-1), increasing the risk of obesity later in life. Therefore, pregnant Sprague-Dawley (SD) rats were used to elucidate whether BDE-47 and BDE-209 transferred through placenta and breast milk cause epigenetic changes in LINE-1 and increase genetic susceptibility to obesity as obesogen during the developmental periods. Global DNA methylation in LINE-1 and gene expression related to obesity were measured in dams and offspring, using a methylation-sensitive high resolution melting analysis (MS-HRM) and direct bisulfite sequencing and quantitative real time polymerase chain reaction (qPCR), respectively. The results of MS-HRM showed global DNA hypomethylation patterns in LINE-1 of exposed offspring (2 of total 4) at PND 4, but bisulfite sequencing showed no difference in both the exposed and non-exposed groups. Gene expression in dams related to ${\beta}$-oxidation pathway and those related to adipokines showed different patterns between the two groups. On the contrary, gene expressions of offspring showed a similar pattern. Gene expressions related to ${\beta}$-oxidation pathway and obesity were significantly increased when compared with 'at birth', but not $PPAR-{\alpha}$. In conclusion, this study demonstrated the possibility that co-exposure to BDE-47 and BDE-209-via the placenta and breast milk-may affect epigenetic changes and modulate gene expression levels related to obesity.

• Journal of Chest Surgery
• /
• v.41 no.6
• /
• pp.687-694
• /
• 2008

Background: Although aortic valve sclerosis causes no significant hemodynamic alterations, it is associated with an increased risk of cardiovascular death and myocardial infarction. However, the role of ${\beta}_3$ integrin in aortic valve sclerosis remains unclear. Material and Method: Twenty male New Zealand rabbits were divided into two groups. Group 1 rabbits (n=10) received a normal chow diet, while group 2 (n=10) rabbits received a diet containing 1% cholesterol for 12 weeks. After the rabbits were euthanized, their aortic valves and ascending aortas were excised for analysis. Result: Total serum cholesterol ($2,148.3{\pm}1,012.5\;mg/dL$ versus $53.7{\pm}31.8\;mg/dL$, p<0.05), triglyceride ($240.4{\pm}218.3\;mg/dL$ versus $31.6{\pm}6.4\;mg/dL$, p<0.05), and low density lipoprotein (LDL)-cholesterol($2,065.3{\pm}960.9\;mg/dL$ versus $29.1{\pm}30.9\;mg/dL$, p<0.05) levels were significantly higher in the cholesterol diet group compared with the normal diet group. Myofibroblasts and macrophages were more highly expressed in the aortic valve leaflets of rabbits in the cholesterol diet group than of those in the normal diet group. A real-time polymerase chain reaction revealed decreased ${\beta}_3$ integrin mRNA levels in the hypercholesterolemic aortic valves and aortas. Conclusion: The present study shows that hypercholesterolemia induces aortic valve sclerosis. These findings suggest that alterations in ${\beta}_3$ integrin may playa role in the development of aortic valve sclerosis.

• Clinical and Experimental Pediatrics
• /
• v.52 no.1
• /
• pp.36-43
• /
• 2009

urpose : The renin-angiotensin system (RAS) has been demonstrated to play a major role in regulating blood pressure. Therefore, components of the RAS are likely candidate genes that may predispose an individual to essential hypertension and cardiovascular complications. Among them, the M235T polymorphism of the angiotensinogen gene has been speculated to be associated with elevated circulating angiotensinogen concentrations and essential hypertension. This study aimed to analyze the angiotensinogen M235T polymorphism in hypertensive adolescents and investigate its relationship with cardiovascular risks. Methods : Forty Korean hypertensive adolescents (aged 16-17, systolic $BP{\geq}140 mmHg$ and/or diastolic $BP{\geq}90 mmHg$) and fifty seven normal adolescents were included. Obesity index (OI), body mass index (BMI) were calculated. BP was measured by oscillometric methods in resting state. Polymerase chain reaction (PCR) technique was performed on DNA from the hypertensives subjects to analyze the M235T polymorphism. Serum homocysteine, insulin, renin, aldosterone and angiotensin converting enzyme (ACE) were tested according to each genotype. The carotid intima-media thickness (IMT) and carotid artery diameter, Pulse wave velocity (PWV) and ankle-brachial index (ABI) were measured according to each genotype. Results : Genotype frequencies of T/T, M/T and M/M were 62.5%, 35.0%, 2.5%, respectively in hypertensive adolescents. The results were not significantly different compared to control group. Serum insulin, renin levels, BMI and OI were significantly higher in thoses with the M/M genotype as compared to those with the T/T of M/T genotype. Conclusion : This study showed that the M235T polymorphism was not associated with essential hypertension or any cardiovascular risks. Further clinical research is required to ascertain the relationship between this polymorphism and cardiovascular complications in Korean hypertensive adolescents.

• Asian Pacific Journal of Cancer Prevention
• /
• v.14 no.7
• /
• pp.4421-4426
• /
• 2013

Objective: The present study employed 5-aza-2'-deoxycytidine (5-Aza-CdR) to treat non-small cell lung cancer (NSCLC) cell line A549 to investigate the effects on proliferation and expression of the TFPI-2 gene. Methods: Proliferation was assessed by MTT assay after A549 cells were treated with 0, 1, 5, 10 ${\mu}mol/L$ 5-Aza-CdR, a specific demethylating agent, for 24, 48 and 72h. At the last time point cells were also analyzed by flow cytometry (FCM) to identify any change in their cell cycle profiles. Methylation-specific polymerase chain reaction (MSPCR), real time polymerase chain reaction(real-time PCR) and western blotting were carried out to determine TFPI-2 gene methylation status, mRNA expression and protein expression. Results: MTT assay showed that the growth of A549 cells which were treated with 5-Aza-CdR was significantly suppressed as compared with the control group (0 ${\mu}mol/L$ 5-Aza-CdR). After treatment with 0, 1, 5, 10 ${\mu}mol/L$ 5-Aza-CdR for 72h, FCM showed their proportion in G0/G1 was $69.7{\pm}0.99%$, $76.1{\pm}0.83%$, $83.8{\pm}0.35%$, $95.5{\pm}0.55%$ respectively (P<0.05), and the proportion in S was $29.8{\pm}0.43%$, $23.7{\pm}0.96%$, $15.7{\pm}0.75%$, $1.73{\pm}0.45%$, respectively (P<0.05), suggesting 5-Aza-CdR treatment induced G0/G1 phase arrest. MSPCR showed that hypermethylation in the promoter region of TFPI-2 gene was detected in control group (0 ${\mu}mol/L$ 5-Aza-CdR), and demethylation appeared after treatment with 1, 5, 10 ${\mu}mol/L$ 5-Aza-CdR for 72h. Real-time PCR showed that the expression levels of TFPI-2 gene mRNA were $1{\pm}0$, $1.49{\pm}0.14$, $1.86{\pm}0.09$ and $5.80{\pm}0.15$ (P<0.05) respectively. Western blotting analysis showed the relative expression levels of TFPI-2 protein were $0.12{\pm}0.01$, $0.23{\pm}0.02$, $0.31{\pm}0.02$, $0.62{\pm}0.03$ (P<0.05). TFPI-2 protein expression in A549 cells was gradually increased significantly with increase in the 5-Aza-CdR concentration. Conclusions: TFPI-2 gene promoter methylation results in the loss of TFPI-2 mRNA and protein expression in the non-small cell lung cancer cell line A549, and 5-Aza-CdR treatment could induce the demethylation of TFPI-2 gene promoter and restore TFPI-2 gene expression. These findings provide theoretic evidence for clinical treatment of advanced non-small cell lung cancer with the demethylation agent 5-Aza-CdR. TFPI-2 may be one molecular marker for effective treatment of advanced non-small cell lung cancer with 5-Aza-CdR.

• Development and Reproduction
• /
• v.5 no.2
• /
• pp.175-180
• /
• 2001

The ectopic expression of gonadotropin releasing hormone(GnRH and luteinizing hormone(LH) in several tissues is a quite intriguing phenomenon. Recently, the presence of GnRH and its receptor has been clearly demonstrated in rodents and human mammary gland. In this context, one can postulate that the presence of local circuit composed of GnRH and LH in the gland. The present study was undertaken to elucidate whether there is a correlation between the LH expression in rat mammary gland and physiological status during the process of mammary differentiation. LH contents in mammary gland from cycling to weaning rats were measured by radioimmunoassay(RIA). In cycling rats, changes of the LH level in both serum and mammary gland showed similar pattern as the highest level in proestrus and the lowest level in diestrus II stage. While the serum LH levels were fluctuated from pregnant through involution stage, a sharp decline of mammary LH contents was observed in the lactating rats. This decrement was recovered in involuting rats to the level of proestrus stage. Reverse transcription-polymerase chain reaction (RT-PCR) and Southern blot analyses demonstrated that the transcriptional activities of the mammary LH and GnRH were increased from diestrus I stage to estrus stage, and the increased levels were maintained in pregnant, lactation and involution stages. To test the hypothesis that the alteration in mammary LH expression might be steroid-dependant, ovariectomy(OVX) and steroid supplement model was employed. As expected, supplement of estradiol(E$_2$) after OVX remarkably decreased serum LH level compared to that in serum from vehicle-only treated rats. Likewise, administration of E$_2$ significantly reduced the mammary LH content. The present study demonstrated that (i) the LH expression in mammary gland could be altered by some physiological parameters such as estrous cycle, pregnancy, lactation and involution, and (ii) ovarian steroid especially estrogen seems to be one of major endocrine factors which are responsible for regulation of mammary LH expression.

• Tuberculosis and Respiratory Diseases
• /
• v.43 no.3
• /
• pp.348-358
• /
• 1996

Background : Interleukin-5 (IL-5) is responsible for eosinophilia in allergic diseases. In allergic bronchial asthma, there is a correlation between the extent of eosinophil infiltration in bronchial mucosa and IL-5 concentrations. In addition, IL-2 concentration is elevated in the airways and associated with eosinophilia in symptomatic patients with bronchial asthma. In animal studies, IL-2 can induce eosinophilia by increasing the synthesis of IL-5, however, it is still unknown how IL-2 can induce eosinophila in human being. The aim of this study is to evaluation the effect and mechanism of IL-2 on prolongation of eosinophil survival. Methods : After purifiing the eosinophils from the venous blood of allergic patients with eosinophilia, we measured the survival rates of eosinophils using trypan blue dye exclusion test, and the number of eosinophils with Randolp's solution. We compared the survival rates of eosinophils in the presence of IL-2 or IL-5. Neutralizing antibody for IL-5 was added in IL-2 treated eosinophils to reveal whether IL-2 induced prolongation of eosinophil survival was mediated by IL-5. We checked IL-5 m-RNA expression of lymphocytes in the presence of IL-2 by using Reverse transcription-Polymerase chain reaction (RT-PCR) method to revealed the effect of IL-2 on IL-5 m-RNA expression on lymphocyte. $\alpha$ and $\beta$ IL-2 receptors were measured on eosinophils and lymphocytes with flow-cytometer after stimulated with IL-2. Results : 1) Eosinophil survival rates increased dose dependently on IL-5 and IL-2. 2) The eosinophil survival rates increased by IL-2 were not inhibited by the pretreatment with neutralizing antibody for IL-5. 3) IL-5 m-RNA was not expressed on lymphocytes by the treatment with IL-2 up to 96 hours. 4) IL-2 upregulate the expression of IL-$2R{\alpha}$ on eosinophils, instead of no effect on the expression of IL-$2R{\beta}$. Conclusion: Interleukin-2 had the enhancing effect on the survival rates of eosinophils. The mechanism behind IL-2 induced eosinophilia might be the increment of IL-2 receptors on eosinophils rather than IL-5 synthesis by lymphocytes.

• Tuberculosis and Respiratory Diseases
• /
• v.60 no.6
• /
• pp.663-672
• /
• 2006

Background: $PM_{10}$(Particulate matter with a diameter ($<10{\mu}m$), which is characterized by different environmental conditions, is a complex mixture of organic and inorganic compounds. The Asian dust event caused by meteorological phenomena can also produce unique particulate matter in affected areas. This study investigated the cytokine produced by A549 epithelial cells exposed to particles collected during both the Asian dust pfenomenon and ambient air particles in a non-dusty period. Method: Air samples were collected using a high volume air sampler(Sibata Model HV500F) with an air flow at $500{\ell}/min$ for at least 6 hours. The cytokine messenger RNA(mRNA) was measured using a reverse transcriptase polymerase chain reaction(RT-PCR). The A549 cells were exposed to 10 to $500{\mu}g/m{\ell}$ of a suspension containing $PM_{10}$ for 24 hours. Each was compared with those in the non-exposed control cells. Result: The mRNA levels of interleukin(IL)-$1{\alpha}$, $IL-I{\beta}$, IL-8, and the granulocyte macrophage colony stimulating factor(GM-CSF) increased after veing exposed to $PM_{10}$ in the ambient air particles, compared with those in the non-exposed control cells. The increase in $IL-1{\alpha}$ and IL-8 were dose dependent at a $PM_{10}$ concentration between $100{\mu}g/m{\ell}$ and $500{\mu}g/m{\ell}$. The mRNA level of IL-8 in the A549 epithelial cells was higher during the in the Asian dust period($500{\mu}g/m{\ell}$) than during the non dust period. Conclusion: A549 cells exposed to the $PM_{10}$ collected during the Asian dust period produce more proinflammatory cytokine than during non-dusty period. This cytokine enhances the local inflammatory response in the airways and can also contribute to the systemic component of this inflammatory process.

• Journal of Yeungnam Medical Science
• /
• v.12 no.2
• /
• pp.366-385
• /
• 1995

Epidemiological studies are important in both the prevention and treatment of mycobacterial infections. This study was initiated to establish the pulsed-field gel electrophoresis (PFGE) method, which are not yet extensively studied. The most apprpriate restriction endonucleases included DraI, AsnI, and XbaI. The optimal PFGE condition was different according to the enzymes used. Two stage PFGE was performed, in case of DraI first stage was performed with 10 seconds of initial pulse and 15 seconds of final pulse, while the second stage was performed with 60 seconds of initial pulse and 70 seconds of final pulse. The electrophoresis time for DraI-PFGE was 14 hours for each stage. Electrophoresis was performed for 22 hours, in case of XbaI, with 3 seconds of initial pulse and 12 seconds of final pulse. Electrophoresis was performed for 22 hours, in case of AsnI, with 5 seconds of initial pulse and 25 seconds of final pulse. In all cases the voltage of the electrophoresis was maintained constantly at 200 voltage. Standard mycobacterial strains, which included Mycobacterium bovis BCG, M. tuberculosis, and M. fortuitum, could not be differentiated by PFGE analysis. PFGE analysis was performed to differentiate 9 clinically isolated M. fortuitum strains using AsnI. All M. fortuitum strains showed different genotypes except 2 strains. Cluster analysis divided M. fortuitum strains into 2 large groups. PFGE analysis was performed to further differentiate M. fortuitum isolates using XbaI. The undifferentiated 2 M. fortuitum strains showed different PFGE patterns with Xba I. Cluster analysis of the XbaI-PFGE patterns showed more complex grouping than AsnI-PFGE patterns, which showed that XbaI-PFGE analysis was better than AsnI-PFGE in M. fortuitum genotyping. The top dissimilarity values of AsnI-PFGE and XbaI-PFGE were 0.74 and 0.75, respectively. This value was higher than that of arbitrarily primed polymerase chain reaction (AP-PCR) analysis and lower than that of restriction fragment length polymorphism (RFLP) analysis. This suggested that PFGE can be used as a supportive or alternative genotyping method to RFLP analysis.

• Journal of Marine Life Science
• /
• v.5 no.2
• /
• pp.35-42
• /
• 2020

17β-estradiol (E₂) is a natural hormone secreted by ovary, and continuously discharged from household and livestock wastewater into aquatic environment. Due to its strong estrogenic activity, it has adverse effects on development and reproduction in crustacean as an endocrine disrupting chemical. Although ecdysteroid signaling pathway play a key role in development in crustacean, little information on transcriptional modulation of ecdysteroid-related genes in response to E₂ is available in small crustacean. Here, we investigated the acute toxicity of E₂ to obtain 24-h LC_x values in the brackish water flea Diaphanosoma celebensis. Time-dependent expression patterns of seven ecdysteroid pathway - related genes (CYP314a1, EcRA, EcRB, USP, ERR, Vtg, VtgR) were further examined using quantitative real time reverse transcriptase polymerase chain reaction (qRT-PCR). As results, 24-h LC₅₀ and LC₁₀ values were 9.581 mg/l and 4.842 mg/l, respectively. The mRNA expression of CYP314a1, EcRA, USP, VtgR was significantly up-regulated at 12 or 24 h after exposure to E₂. These findings indicate that E₂ can affect their molting and reproduction by modulating the expression of ecdysteroid pathway - related in D. celebensis. This study will be useful for better understanding of molecular mode of action of endocrine disrupting chemicals on molting process in small crustacean.