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The role of neurohypophyseal hormones in formation of cardiac disorders after the trauma
J Mol Cell Cardiol 24 (Supplement I) (1992)
P-06-lOPOINT MUTATIONS IN MITOCHONDRIAL HYPERTROPHIC CARDIOMYOPATHY
DNA OF PATIENTS
WITH
Toshihiro Obayashi, Masaaki Takasawa, Kazuki Hattori, Masashi Tanaka, Satoru Sugiyama, Takayuki Ito, Takayuki Ozawa. Department of Internal Medicine II and *Biomedical Chemistry, Faculty of Medicine, University of Nagoya, Nagoya, Japan Mutations in mitochondrial DNA (mtDNA) as well as those in nuclear DNA are suggested to be involved in the etiology of hypertrophic cardiomyopathy (HCM). MtDNA consists of 16,569 base pairs, which is very small compared with nuclear DNA, but nonetheless too large to sequence completely in many patients using conventional radioisotope methods. In this study, we prepared a variety of primer pairs, and developed a rapid and accurate method for total mtDNA sequencing using a fluorescence-based automated sequencing method. We were able to analyze total mtDNA in seven patients with HCM using this method. We found several point mutations in patients with HCM. Although no common point mutations were observed among the patients, some of these point mutations may result in functionally important amino acid replacements. This could result in a significant impairment in ATP production. Point mutations in transfer RNA (tRNA) genes were also observed in four of the seven patients. Two patients had the same point mutation observed in a mitochondrial neuromuscular disease, myopathy, encephalopathy, lactic acidosis and stroke-like episodes (MELAS), which is often associated with cardiac hypertrophy. From these results, point mutations in mtDNA which are almost always associated with significant rise in the rate of deletion might be closely linked with the genesis of HCM, and our method developed here might be useful for the detection of point mutations in mtDNA.
P-06-11 TOLERANCE OF ISOLATED HEART TO HYPOXIA IN CARDIOMYOPATHIC
HAMSTERS Shin-ichi Momomura, Yasushi Nagai, Toshiaki Ogawa, Yukuo Nezu, Tadashi Sato, Hiroshi Yamashita, Takashi Serizawa, Tsuneaki Sugimoto. University of Tokyo and Eisai Laboratory, Tokyo and Ibaragi, Japan To study hemodynamic and metabolic responses of myopathic hearts to hypoxia, 3’P-NMR spectrum was obtained for every 5 minutes in isolated isovolumically beating hearts of 20 week-old control hamsters (C, n=lO) and cadiomyopathic hamsters, BIO 14.6 (B, n=lO). Initial left ventricular end-diastolic pressure (EDP) was set to 10 mmHg, and hearts were perfused with mildly hypoxic Krebs-Henseleit buffer equilibrated with 20% O,, 75% N,, 5% CO, gas mixture for 30 minutes. In B, changes in EDP (AEDP). developed pressure (ADP), ATP (AATP), creatine phosphate (ACP), inorganic phosphate (Alp), and intracellular pH (APHI) with 30 minutes of hypoxia were all mild compared to those in C (mean&SD, * p
P-()&12NOREPINEPHRINE-ST’HULATED PROTO-ONCOGENE EXPRESSION IN THE IDIOPATHIC CARDIOMYOPATHIC HAIISTER HEARTS Tetsuya Kurimoto, Keiko Y-Takihara, Shunzo Onishi*, Junlchi Azuma. Department of Medicine III, *College of Biomedical Technology, Osaka University, Osaka JAPAN. Proto-oncogenes play a crucial role in cellular growth and dlffetentiatlon. Effect of norepinephrine(NE) on proto-oncogene expression was examined in the control(RB) and cardiamyopathic(BI014.6) hamster hearts. The hearts were removed at 0.5, I, 2, 4, 8, and 24 hr after peritoneal injection of NE(3.Omg/kg), or immediately after treadmill running for lhr with or without NE(O.lmg/kg) pretreatment. Northern blot analysis was carried out using c-fos, v-Ha-ras, and I,adrenergic receptor(AR) cDNA as a probe. NE induced c-fos and c-Ha-t-as expression in both RB and 61014.6 hearts. The levels of c-fos and c-Ha-ras mRNA reached their peak at 0.5”lhr and 8hr respectively, and returned to baseline by 24hr. Treadmill running induced c-fos expression only in the NE pretreated BIO14.6 hamster heart. There was no significant difference in the cardiac mRNA encording #,-AR between RB and BIOl4.6 hamsters. These results could lead to better understanding of treating hypertrophic cardlomyopathic patients with I, blocking agents.
S.217
P-06-lOPOINT MUTATIONS IN MITOCHONDRIAL HYPERTROPHIC CARDIOMYOPATHY
DNA OF PATIENTS
WITH
Toshihiro Obayashi, Masaaki Takasawa, Kazuki Hattori, Masashi Tanaka, Satoru Sugiyama, Takayuki Ito, Takayuki Ozawa. Department of Internal Medicine II and *Biomedical Chemistry, Faculty of Medicine, University of Nagoya, Nagoya, Japan Mutations in mitochondrial DNA (mtDNA) as well as those in nuclear DNA are suggested to be involved in the etiology of hypertrophic cardiomyopathy (HCM). MtDNA consists of 16,569 base pairs, which is very small compared with nuclear DNA, but nonetheless too large to sequence completely in many patients using conventional radioisotope methods. In this study, we prepared a variety of primer pairs, and developed a rapid and accurate method for total mtDNA sequencing using a fluorescence-based automated sequencing method. We were able to analyze total mtDNA in seven patients with HCM using this method. We found several point mutations in patients with HCM. Although no common point mutations were observed among the patients, some of these point mutations may result in functionally important amino acid replacements. This could result in a significant impairment in ATP production. Point mutations in transfer RNA (tRNA) genes were also observed in four of the seven patients. Two patients had the same point mutation observed in a mitochondrial neuromuscular disease, myopathy, encephalopathy, lactic acidosis and stroke-like episodes (MELAS), which is often associated with cardiac hypertrophy. From these results, point mutations in mtDNA which are almost always associated with significant rise in the rate of deletion might be closely linked with the genesis of HCM, and our method developed here might be useful for the detection of point mutations in mtDNA.
P-06-11 TOLERANCE OF ISOLATED HEART TO HYPOXIA IN CARDIOMYOPATHIC
HAMSTERS Shin-ichi Momomura, Yasushi Nagai, Toshiaki Ogawa, Yukuo Nezu, Tadashi Sato, Hiroshi Yamashita, Takashi Serizawa, Tsuneaki Sugimoto. University of Tokyo and Eisai Laboratory, Tokyo and Ibaragi, Japan To study hemodynamic and metabolic responses of myopathic hearts to hypoxia, 3’P-NMR spectrum was obtained for every 5 minutes in isolated isovolumically beating hearts of 20 week-old control hamsters (C, n=lO) and cadiomyopathic hamsters, BIO 14.6 (B, n=lO). Initial left ventricular end-diastolic pressure (EDP) was set to 10 mmHg, and hearts were perfused with mildly hypoxic Krebs-Henseleit buffer equilibrated with 20% O,, 75% N,, 5% CO, gas mixture for 30 minutes. In B, changes in EDP (AEDP). developed pressure (ADP), ATP (AATP), creatine phosphate (ACP), inorganic phosphate (Alp), and intracellular pH (APHI) with 30 minutes of hypoxia were all mild compared to those in C (mean&SD, * p
P-()&12NOREPINEPHRINE-ST’HULATED PROTO-ONCOGENE EXPRESSION IN THE IDIOPATHIC CARDIOMYOPATHIC HAIISTER HEARTS Tetsuya Kurimoto, Keiko Y-Takihara, Shunzo Onishi*, Junlchi Azuma. Department of Medicine III, *College of Biomedical Technology, Osaka University, Osaka JAPAN. Proto-oncogenes play a crucial role in cellular growth and dlffetentiatlon. Effect of norepinephrine(NE) on proto-oncogene expression was examined in the control(RB) and cardiamyopathic(BI014.6) hamster hearts. The hearts were removed at 0.5, I, 2, 4, 8, and 24 hr after peritoneal injection of NE(3.Omg/kg), or immediately after treadmill running for lhr with or without NE(O.lmg/kg) pretreatment. Northern blot analysis was carried out using c-fos, v-Ha-ras, and I,adrenergic receptor(AR) cDNA as a probe. NE induced c-fos and c-Ha-t-as expression in both RB and 61014.6 hearts. The levels of c-fos and c-Ha-ras mRNA reached their peak at 0.5”lhr and 8hr respectively, and returned to baseline by 24hr. Treadmill running induced c-fos expression only in the NE pretreated BIO14.6 hamster heart. There was no significant difference in the cardiac mRNA encording #,-AR between RB and BIOl4.6 hamsters. These results could lead to better understanding of treating hypertrophic cardlomyopathic patients with I, blocking agents.
S.217