Isoproterenol과 Melatonin의 첨가가 돼지 단위발생란의 체외발달에 미치는 영향

정다운,윤윤진,박희성 한국수정란이식학회 2016 한국동물생명공학회지 Vol.31 No.1

In this study, to improve the in vitro development of various cells including cloned embryos, the effects that isoproterenol and melatonin have on in vitro development of porcine parthenogenetic oocytes were investigated. Parthenogenetic activation was induced with electrical stimulation, BSA and 6-DMAP treatment. 10-7 M of melatonin and isoproterenol (10-10, 10-12 and 10-14 M) were supplemented for in vitro maturation (IVM) and in vitro culture (IVC) medium, with different concentrations. When isoproterenol and melatonin were supplemented in IVM medium with different concentrations, there was no significant (P<0.05) difference of maturation rate in the treatment groups as well as in that of only melatonin. As isoproterenol and melatonin were supplemented in IVM medium with different concentrations, blastocyst rates of isoproterenol 10-12 M treatment group (37.1%) were significantly (P<0.05) higher than control group (26.0%). Isoproterenol and melatonin were supplemented in IVC medium with different concentrations, then the cleavage rate of 10-12 M isoproterenol treatment group (82.2%) was significantly (P<0.05) higher than the group that melatonin was only supplemented (70.9%). There was no difference of blastocyst rate between the treatment groups. When isoproterenol and melatonin were supplemented for IVM+IVC medium with different concentrations, the cleavage rate of 10-12 M isoproterenol treatment group (92.5%) was significantly (P<0.05) higher than the control group (82.8%) and the group that melatonin was only treated (81.6%). The blastocyst rate of 10-12 M as 45.6% was significantly (P<0.05) higher than control group (25.2%) and melatonin treatment group (31.2%). The cell number of blastocyst in 10-12 M isoproterenol treatment group 35.5±3.4 was significantly (P<0.05) highest. The results of this study showed that the development rate of IVC when both isoproterenol and melatonin were supplemented was higher than when melatonin was only supplemented. Therefore, it is concluded that isoproterenol is rather effective in the activation of melatonin. 10-7 M melatonin and 10-12 M isoproterenol were considered suitable concentration.

Isoproterenol Increases RANKL Expression in a ATF4/NFATc1-Dependent Manner in Mouse Osteoblastic Cells

Baek, Kyunghwa,Park, Hyun-Jung,Baek, Jeong-Hwa,Kim, Hyung-Ryong MDPI 2017 INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES Vol.18 No.10

<P>Sympathetic nervous system stimulation-induced β-adrenergic signal transduction is known to induce bone loss and increase of osteoclast activity. Although isoproterenol, a nonspecific β-adrenergic receptor agonist, has been shown to increase receptor activator of NF-κB ligand (RANKL), the details of the regulatory mechanisms remain unclear. In the present study, we investigated the role of the nuclear factor of activated T-cells (NFAT) in isoproterenol-induced RANKL expression in C2C12 and in primary cultured mouse calvarial cells. Isoproterenol increased nuclear factor of activated T-cells cytoplasmic 1 (NFATc1) and RANKL expressions at both mRNA and protein levels and increased NFAT reporter activity. NFATc1 knockdown blocked isoproterenol-mediated RANKL expression. Isoproterenol also promoted cAMP response element-binding protein 1 (CREB1) and activating transcription factor 4 (ATF4) phosphorylation. Isoproterenol-mediated transcriptional activation of NFAT was blocked by protein kinase A (PKA) inhibitor H89. Isoproterenol-induced CREB1, ATF4, NFATc1, and RANKL expressions were suppressed by H89. Mutations in cAMP response element-like or NFAT-binding element suppressed isoproterenol-induced <I>RANKL</I> promoter activity. Chromatin immunoprecipitation analysis demonstrated that isoproterenol increased NFAT-binding and ATF4-binding activities on the mouse <I>RANKL</I> promoter, but did not increase CREB1-binding activity. Association of NFATc1 and ATF4 was not observed in a co-immunoprecipitation study. ATF4 knockdown suppressed isoproterenol-induced NFAT binding to the <I>RANKL</I> promoter, whereas NFATc1 knockdown did not suppress isoproterenol-induced ATF4 binding to the <I>RANKL</I> promoter. ATF4 knockdown suppressed isoproterenol-induced expressions of NFATc1 and RANKL. These results suggest that isoproterenol increases RANKL expression in an ATF4/NFATc1-dependent manner.</P>

Deficiency of iNOS Does Not Prevent Isoproterenol-induced Cardiac Hypertrophy in Mice

박소영 대한약리학회 2009 The Korean Journal of Physiology & Pharmacology Vol.13 No.3

We investigated whether deficiency of inducible nitric oxide synthase (iNOS) could prevent isoproterenol-induced cardiac hypertrophy in iNOS knockout (KO) mice. Isoproterenol was continuously infused subcutaneously (15 mg/kg/day) using an osmotic minipump. Isoproterenol reduced body weight and fat mass in both iNOS KO and wild-type mice compared with saline-infused wild-type mice. Isoproterenol increased the heart weight in both iNOS KO and wild-type mice but there was no difference between iNOS KO and wild-type mice. Posterior wall thickness of left ventricle showed the same tendency with heart weight. Protein level of iNOS in the left ventricle was increased in isoproterenol-infused wild-type mice. The gene expression of interleukin-6 (IL-6) and transforming growth factor-β (TGF-β) in isoproterenol-infused wild-type was measured at 2, 4, 24, and 48-hour and isoproterenol increased both IL-6 (2, 4, 24, and 48-hour) and TGF-β (4 and 24-hour). Isoproterenol infusion for 7 days increased the mRNA level of IL-6 and TGF-β in iNOS KO mice, whereas the gene expression in wild-type mice was not increased. Phosphorylated form of extracellular signal-regulated kinases (pERK) was also increased by isoproterenol at 2 and 4-hour but was not increased at 7 days after infusion in wild-type mice. However, the increased pERK level in iNOS KO mice was maintained even at 7 days after isoproterenol infusion. These results suggest that deficiency of iNOS does not prevent isoproterenol- induced cardiac hypertrophy and may have potentially harmful effects on cardiac hypertrophy.

Deficiency of iNOS Does Not Prevent Isoproterenol-induced Cardiac Hypertrophy in Mice

Cha, Hye-Na,Hong, Geu-Ru,Kim, Yong-Woon,Kim, Jong-Yeon,Dan, Jin-Myoung,Park, So-Young The Korean Society of Pharmacology 2009 The Korean Journal of Physiology & Pharmacology Vol.13 No.3

We investigated whether deficiency of inducible nitric oxide synthase (iNOS) could prevent isoproterenol-induced cardiac hypertrophy in iNOS knockout (KO) mice. Isoproterenol was continuously infused subcutaneously (15 mg/kg/day) using an osmotic minipump. Isoproterenol reduced body weight and fat mass in both iNOS KO and wild-type mice compared with saline-infused wild-type mice. Isoproterenol increased the heart weight in both iNOS KO and wild-type mice but there was no difference between iNOS KO and wild-type mice. Posterior wall thickness of left ventricle showed the same tendency with heart weight. Protein level of iNOS in the left ventricle was increased in isoproterenol-infused wild-type mice. The gene expression of interleukin-6 (IL-6) and transforming growth factor-${\beta}$ (TGF-${\beta}$) in isoproterenol-infused wild-type was measured at 2, 4, 24, and 48-hour and isoproterenol increased both IL-6 (2, 4, 24, and 48-hour) and TGF-${\beta}$ (4 and 24-hour). Isoproterenol infusion for 7 days increased the mRNA level of IL-6 and TGF-${\beta}$ in iNOS KO mice, whereas the gene expression in wild-type mice was not increased. Phosphorylated form of extracellular signal-regulated kinases (pERK) was also increased by isoproterenol at 2 and 4-hour but was not increased at 7 days after infusion in wild-type mice. However, the increased pERK level in iNOS KO mice was maintained even at 7 days after isoproterenol infusion. These results suggest that deficiency of iNOS does not prevent isoproterenol-induced cardiac hypertrophy and may have potentially harmful effects on cardiac hypertrophy.

Deficiency of iNOS Does Not Prevent Isoproterenol-induced Cardiac Hypertrophy in Mice

Hye-Na Cha,Geu-Ru Hong,Yong-Woon Kim,Jong-Yeon Kim,Jin-Myoung Dan,So-Young Park 대한생리학회-대한약리학회 2009 The Korean Journal of Physiology & Pharmacology Vol.13 No.3

We investigated whether deficiency of inducible nitric oxide synthase (iNOS) could prevent isoproterenol-induced cardiac hypertrophy in iNOS knockout (KO) mice. Isoproterenol was continuously infused subcutaneously (15 mg/kg/day) using an osmotic minipump. Isoproterenol reduced body weight and fat mass in both iNOS KO and wild-type mice compared with saline-infused wild-type mice. Isoproterenol increased the heart weight in both iNOS KO and wild-type mice but there was no difference between iNOS KO and wild-type mice. Posterior wall thickness of left ventricle showed the same tendency with heart weight. Protein level of iNOS in the left ventricle was increased in isoproterenol-infused wild-type mice. The gene expression of interleukin-6 (IL-6) and transforming growth factor-Ղ (TGF-Ղ) in isoproterenol-infused wild-type was measured at 2, 4, 24, and 48-hour and isoproterenol increased both IL-6 (2, 4, 24, and 48-hour) and TGF-Ղ (4 and 24-hour). Isoproterenol infusion for 7 days increased the mRNA level of IL-6 and TGF-Ղ in iNOS KO mice, whereas the gene expression in wild-type mice was not increased. Phosphorylated form of extracellular signal-regulated kinases (pERK) was also increased by isoproterenol at 2 and 4-hour but was not increased at 7 days after infusion in wild-type mice. However, the increased pERK level in iNOS KO mice was maintained even at 7 days after isoproterenol infusion. These results suggest that deficiency of iNOS does not prevent isoproterenol- induced cardiac hypertrophy and may have potentially harmful effects on cardiac hypertrophy.

토끼 경동맥 평활근에서 Isoproterenol의 이완작용에 미치는 Phorbol Esters의 효과

김법영,이영우 대한신경외과학회 1995 Journal of Korean neurosurgical society Vol.24 No.7

토끼 경동맥에서 phorbol estersr가 isoporterenol에 의한 혈관 긴장도의 조절에 미치는 protein kinase C(PKC)의 작용을 밝히기 위하여 경동맥환에서 등척성 수축을 기록하여 phorbol esters인 phorbol 12,13-di-butyrate(PDBu)와 phorbol 12-myristate 13-acetate(PMA)의 효과를 관찰하였다. β-Adrenergic agonist인 isoproterenol은 phenylephrine에 의한 수축을 이완하였으나 phorbol esters에 의한 수축에는 영향을 미치지 아니하였다. Isoproterenol에 의한 혈관 이완작용은 내피세포의 제거나 methylene blue나 nitro-L-arginine의 전처치에 의하여 감소하였다. 수축을 야기하지 않는 농도는 phorbol esters의 전처치는 내피세포 의존성 및 비의존성으로 isoproterenol에 의해 야기된 혈관이완을 억제하였다. Isoproterenol에 의해 야기된 혈관 이완작용에 대한 PDBu의 억제작용은 수축반응에서 보인 양상과는 달리 PMA의 억제작용보다 낮은 효과를 보였으나 PDBu의 수축효과는 PMA보다 더 강하게 나타났다. PMA는 forskolin의 농도반응에 영향을 미치지 않았다. 그리고 protein kinase C inhibitor인 staurosporine은 isoprocerenol에 의해 야기된 이완과 수축반응에 대한 이들 약물의 작용을 억제하였다. 이상과 같은 결과는 isoproterenol에 의해 야기된 혈관 이완작용은 protein kinase C의 활성제인 phorbol esters에 의해 억제되며, 이는 수축반응에 각기 다른 isozyme이 관여하는 것으로 사료된다. The effects of phorbol esters were studied in rabbit carotid artery to evaluate the action of protein kinase C on the regulation of vascular tone by isoproterenol. The vascular rings. 2 mm in width, were myographied isometrically in an isolated organ bath and the effects of phorbol 12,13-dibutyrate(PDBu) and phorbol 12-myristate 13-acetate(PMA) were determined. Isoproterenol a beta adrenergic agonist, relaxed the vessel which was precontracted by phenylephrine, but not that by phorbol esters. The action of isoproterenol was attenuated by removal of endothelium or pretreatment with methylene blue or nitro-L-arginine. The pretreatment with phorbol esters at concentrations which did not induce contraction, decreased isoproterenol-induced relaxation of vascular rings with or without endothelium. The action of PDBu on isoproterenol-induced relaxation was less effective than that of PMA, unlike those observed in contractile response. but the contractile effect of the former was more potent than that of the latter. PMA did not affect relaxant effect of forskolin, an activator of adenyl cyclase. Staurosporine, a protein kinase C inhibitor, inhibited the action of these drugs on both isoproterenol-induced relaxation and the contractile response. These results suggest that the relaxation induced by isoproterenol was reduced by the activation of protein kinase C. which may be isozyme different from that involved in contractile response.

Isoproterenol 투여로 유발된 심근세포 손상에 미치는 diltiazem의 영향

김현,장대영,라봉진,김호덕,Kim, Hyun,Chang, Dae-Yung,Rah, Bpng-Jin,Kim, Ho-Dirk 한국현미경학회 1997 Applied microscopy Vol.27 No.2

It has been demonstrated that majority of cells in the mammalian body such as myocytes and epithelial cells of skin and intestine respond to mechanical force or environmental factors and exhibit partial disruption of cell membrane, i. e., cell wounding, even in a physiological condition. Myocardial cells are rather apt to be wounded than other cells since they are definitely exposed to mechanical stress by contraction-relaxation and blood flow. However, the mechanism how myocardial cells protect themselves against cell wounding is not yet clarified. On this background, the present study was performed to elucidate whether albumin leakage is related to cell wounding and to assess whether diltiazem, a potent calcium channel blocker, is beneficial in isoproterenol-induced cell wounding in the heart. Hearts isolated from New Zealand White rabbits ($1.5\sim2.0kg$ body weight, n=20) were perfused with Tyrode solution by Langendorff technique. After stabilization of baseline hemodynamics, the hearts were subjected to bolus administration of isoproterenol and diltiazem as following order: $1.6{\mu}M$ isoproterenol at zero min (the beginning point): $16{\mu}M$ diltiazem at 20min; $1.6{\mu}M$ isoproterenol at 25min; $16{\mu}M$ isoproterenol at 45 min; $160{\mu}M$ diltiazem at 65 min; $16{\mu}M$ isoproterenol at 70 min. During all experiments, the left ventricular function was recorded, albumin leakage in the coronary effluents was analyzed by electrophoresis and Western blot, and myocardial cell membranes were examined by conventional transmission electron microscopy. Data were analyzed by t-test and linear regression test. Isoproterenol significantly increased the inotropic and chronotropic contractions, coronary flow, and frequency of arrhythmia, however, diltiazem did not influence on hemodynamics except decrease in the frequency of arrhythmia and a slight decrease in contractility. Isoproterenol also resulted partial disruption of myocardial cell membrane and inclose in albumin leakage, while diltiazem pretreatment showed number of electron-dense plaques in the cell membrane and a tendency of decrease in albumin leakage. These results indicate that albumin leakage may be an indirect index of cell wounding in the heart and diltiazem nay be beneficial to protect myocardial cells against isoproterenol-induced cell wounding. It is likely that diltiazem promotes resealing process of the cell membrane.

Isoproterenol에 의한 자궁근 이완의 기전 : 4-aminopyridine-sensitive K⁺ 채널의 개방

김기하,이영재,조명행,이문한,전보권,류판동,Kim, Ki-ha,Lee, Young-jae,Cho, Myung-haing,Lee, Mun-han,Chun, Boe-gwon,Ryu, Pan-dong 대한수의학회 1996 大韓獸醫學會誌 Vol.36 No.1

Activation of $K^+$ channels induces relaxation of smooth muscles by reducing electrical excitability and cytosolic free $Ca^{2+}$ level. ${\beta}$-adrenergic agonist isoproterenol is known to induce relaxation of the uterine smooth muscle by membrane hyperpolarization and $K^+$ efflux. Recently it is suggested that the activity of $Ca^{2+}$-activated $K^+$ channel was increased by isoproterenol in the uterine myocytes isolated from myometrium of the pregnant rat. However, the type of $K^+$ channel mediating the relaxant effect of isopreterenol in the tissue level has not yet studied. In this work, we investigated the type of $K^+$ channels involved in the isoproterenol-induced relaxation of uterine smooth muscle by measuring the integrated insometric tension of the estrogen-treated isolated nonpregnant rat uterus. Contraction of uterine tissue was induced by oxytocin (0.2nM, 2~3 contractions/min) or high KCl(20~80mM). The result are as follows : 1. Isoproterenol($10^{-10}{\sim}10^{-4}M$) inhibited oxytocin-induced contraction of isolated rat uterus($EC_{50}=1.17{\times}10^{-10}M$). 2. Isoproterenol($10^{-10}{\sim}10^{-4}M$) effectively inhibited uterine contraction induced by low KCl(20~40mM) but little those induced by high KCl(60~80mM). 3. Relaxant effect of isoproterenol($10^{-10}{\sim}10^{-4}M$) on 0.2nM oxytocin-induced contraction was effectively reduced by 4-aminopyridine(3, 10mM) but little by TEA(10~30mM), $Ba^{2+}$($1{\sim}30{\mu}M$) and glibenclamide($100{\mu}M$). Our data suggest that the relaxant effect of isoproterenol is mediated by the $K^+$ channel(s) which can be blocked by 4-aminopyridine.

Isoproterenol 및 Melatonin의 첨가농도와 난포란의 등급에 따른 돼지 단위발생란의 체외 발달

김도연,정현화,정다운,박희성 한국동물생명공학회(구 한국동물번식학회) 2017 Reproductive & Developmental Biology(Supplement) Vol.41 No.2

일반적으로 Isoproterenol은 melatonin의 합성과정에 있어서 melatonin을 활성화 시켜 그 활성도를 향상시키는 것으로 알려져 있다. Melatonin은 포유동물의 송과선에서 분비되 는 호르몬으로서 특히 난포액에서 높은 농도로 검출되며, 난자의 체외배양의 경우, 강력 한 활성산소가 청소부 작용을 하여 난자의 체외발달율을 개선시키는 역할을 한다. 따라 서 본 연구에서는 isoproterenol과 melatonin의 적정농도 확인과 난포란의 quality가 낮은 3등급 난자의 체외발달율 개선효과 여부를 검토하고자 melatonin 10—7∼11—11 및 isoproterenol 10—10∼10—14몰 농도로 체외성숙 배양액에 첨가하고 난포란의 등급을 구분하여 단위발생을 유기하였다. 뿐만 아니라 체외 배양액에 첨가하여 등급이 낮은 돼지 난포란 유래 단위발생란의 분할율 및 배반포기 발달율을 조사하였다. 도축장으로부터 채집한 돼지 난소의 난포에서 난포란을 채취하여 난구세포층이 3∼5층 이상 충실하게 부착되어 있는 난포란을 Grade І(1∼2등급), 난구세포층이 1∼2층 이하 및 부분적으로 나화되어 있는 난포란을 Grade III(3∼4등급)으로 등급을 구분하여 전기자극법 으로 자극을 가한 다음, 2 mM의 6-DMAP 용액으로 2시간 동안 처리하여 단위발생을 유 도하였다. Isoproterenol(—10, —12, —14몰)과 melatonin(—7, —9, —11몰)을 체외성숙 및 체 외 배양시에 각각 병용첨가하였다. 체외성숙 배양액은 TCM-199 배양액을 기본배양액으 로 하여 각종 호르몬을 첨가하여 사용하였으며, 체외배양액은 PZM-5에 3 mg FAF-BSA 를 첨가하여 38.5℃, CO2 배양기에서 7일간 배양을 실시하였다. 체외 성숙 및 배양시에 melatonin 10—10몰 및 isoproterenol 10—12몰을 첨가하였을 때, 3등급 난자의 체외 성숙율 이 79.3%로서 1등급(82.6%) 난포란과 차이가 없을 정도로 개선되었으며, 배반포기로의 체외발달율은 melatonin 10—10몰 및 isoproterenol 10—12몰이 67.0%(1등급) 및 50.2%(3등 급)로서 1등급이 유의적(p<0.05)으로 높은 발달율을 보였으나, 다른 처리구에서는 1, 3등 급 간에 차이가 없었지만 개선되는 경향을 보였다. 이상의 결과로 볼 때, melatonin과 isoproterenol의 첨가농도는 melatonin 10—10몰 및 isoproterenol 10—12이 적정한 농도인 것으로 판단되며, 1, 3등급 간에 차이가 없음을 확인 하였다. 따라서 이들을 첨가하면 난포란의 활용도 제고에 크게 기여할 수 있을 것으로 생각된다. Key words)

Isoproterenol이 백서 자궁수축에 미치는 영향

김영인(YI Kim),김생기(SK Kim),이춘근(CK Lee) 대한산부인과학회 1983 Obstetrics & Gynecology Science Vol.26 No.11

Pharmacological analysis of the inhibitory effect of isoproterenol on the pregnanr and non-pregnant strips of rat uterus, and isoproterenot induced desensitization has been made on the basis of 봭-adrenergic receptors. 1. Inhibitory effect of isoproterenol in the pregnant uterus was greater than in the non-pregnant uterus, while the rate of spontaneous contraction was higher in the non-pregnant uterus. 2. Inderal inhibited the inbibitory effect of isoproterenol in both preparations, resulting in the parallel shifts of does responsive curves to the right. 3. Isoprotereno1-induced desensitization. 4. Isoprotereno1-induced desensitization was not prevented by the treatment of isoprotereno1, inderal or pyrogallol. From the above result, it is suggested that difference of inhibitory effect of isop roterenol in the non-pregnant uterus and in the pregnant uterus, and isoproterenol induced desensitization were not due to the alteration of β-adrenergic receptor.