- Email: [email protected]
In vivo functional brain imaging and a therapeutic trial of l -arginine in MELAS patients
Biochimica et Biophysica Acta 1820 (2012) 615–618
Contents lists available at ScienceDirect
Biochimica et Biophysica Acta j o u r n a l h o m e p a g e : w w w. e l s ev i e r. c o m / l o c a t e / b b a g e n
In vivo functional brain imaging and a therapeutic trial of L-arginine in MELAS patients☆ Makoto Yoneda a,⁎, Masamichi Ikawa a, Kenichiro Arakawa b, Takashi Kudo c, Hirohiko Kimura d, Yasuhisa Fujibayashi e, Hidehiko Okazawa c a
Department of Neurology, Faculty of Medical Sciences, University of Fukui, Fukui, Japan Department of Cardiology, Faculty of Medical Sciences, University of Fukui, Fukui, Japan Biomedical Imaging Research Center, University of Fukui, Fukui, Japan d Department of Radiology, Faculty of Medical Sciences, University of Fukui, Fukui, Japan e Molecular Imaging Center, National Institute of Radiological Sciences, Chiba, Japan b c
a r t i c l e
i n f o
Article history: Received 7 April 2011 Received in revised form 28 April 2011 Accepted 29 April 2011 Available online 8 May 2011 Keywords: MELAS Functional brain imaging PET MRI L-arginine
a b s t r a c t Background: Mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes (MELAS) is the most common type of mitochondrial disease and is characterized by stroke-like episodes (SEs), myopathy, lactic acidosis, diabetes mellitus, hearing-loss and cardiomyopathy. The causal hypotheses for SEs in MELAS presented to date are angiopathy, cytopathy and neuronal hyperexcitability. L-arginine (Arg) has been applied for the therapy in MELAS patients. Scope of review: We will introduce novel in vivo functional brain imaging techniques such as MRI and PET, and discuss the pathogenesis of SEs in MELAS patients. We will further describe here our clinical experience with L-arg therapy and discuss the dual pharmaceutical effects of this drug on MELAS. Major conclusions: Administration of L-arg to MELAS patients has been successful in reducing neurological symptoms due to acute strokes and preventing recurrences of SEs in the chronic phase. L-Arg has dual pharmaceutical effects on both angiopathy and cytopathy in MELAS. General significance: In vivo functional brain imaging promotes a better understanding of the pathogenesis and potential therapies for MELAS patients. This article is part of a Special Issue entitled Biochemistry of Mitochondria, Life and Intervention 2010. © 2011 Elsevier B.V. All rights reserved.
1. Background Mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes (MELAS), was first described by Pavlakis et al., and is the most common type of mitochondrial disease [1]. While over 40 mutations in the mitochondrial DNA (mtDNA) have been reported in association with MELAS, an A-to-G transition at nucleotide position 3243 (A3243G) in mtDNA is responsible for approximately 80% of the affected patients [2,3]. The clinical features of MELAS consist of strokelike episodes (SEs), myopathy, lactic acidosis, diabetes mellitus, hearing-loss and cardiomyopathy [1]. SEs provoke various neurological symptoms due to the affected brain lesions. These SEs are crucial life-threatening factors that determine the
☆ This article is part of a Special Issue entitled Biochemistry of Mitochondria, Life and Intervention 2010. ⁎ Corresponding author at: Department of Neurology, Faculty of Medical Sciences, University of Fukui, Fukui 910-1193, Japan. Tel.: + 81 776 61 8351; fax: + 81 776 61 8110. E-mail address: [email protected] (M. Yoneda). 0304-4165/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.bbagen.2011.04.018
prognosis of patients with MELAS. These SEs demonstrate unique characteristics different from ordinary ischemic strokes [1]. Stroke-like lesions spread beyond the vascular territory and are expanded to neighboring areas of the brain [4]. In the acute phase (several days after the stroke onset), there is focal vasodilatation and hyper-perfusion whereas, in the chronic phase (several months after the stroke onset), hypo-perfusion and irreversible changes appear [5]. However, in the super-acute phase (within 3 hours after the stroke onset), Koga et al. demonstrated hypoperfusion in the stroke lesion by a single photon emission tomography (SPECT) analysis [6]. Although several casual hypotheses of SEs have been proposed, the pathogenesis of SEs remains obscure. Causal hypotheses for SEs presented to date are angiopathy [7,8], cytopathy [9] and neuronal hyperexcitability [3,10]. Especially, the hypothesis involving angiopathy is one of the most attractive ideas explaining SEs in MELAS patients. Abnormal accumulation of mitochondria was described in the vascular endothelium and smooth muscle cells in small arteries and arterioles of the autopsied brain of a MELAS patient, and “mitochondrial angiopathy” was proposed 25 years ago [7]. A recent electron microscopic study further demonstrated disruption of capillary endothelial tight junctions in the autopsied brain of a patient with MELAS [11]. This
616
M. Yoneda et al. / Biochimica et Biophysica Acta 1820 (2012) 615–618
endothelial barrier disruption can result in impairments of the blood brain barrier and ion homeostasis between extra- and intra-cellular fluid, causing vasogenic edema that was detected on an apparent diffusion coefficient (ADC) map of the affected brain in a MELAS patient on magnetic resonance imaging (MRI) [12]. A physiological study also supported the angiopathy evidenced by a decreased vasodilatation capacity in small arteries examined by the flow mediated vasodilatation (FMD) method [6]. There are several biochemical data supportive of cytopathy in MELAS especially in association with A3243G mutation; i.e. increased unstabletRNA-Leu(UUR), increased ratio of uncharged aminoacyl-tRNALeu(UUR), accumulation of aminoacylation with leucine without any misacylation, deficient taurine modification and processing abnormality of the mitochondrial transcripts [13–16]. These biochemical defects lead cells to energy failure such as enhanced glycolysis, impaired electrontransport and oxidative phosphorylation, finally resulting in decreased ATP production. The recent imaging studies using MRI, magnetic resonance spectroscopy (MRS), SPECT and positron emission tomography (PET) demonstrated the process of SEs or pathophysiology in MELAS patients, as described below. Koga et al. found a decrease in L-arginine (Arg) that is metabolized into citrulline (Cit) and nitric oxide (a strong vasodilator), and relative increase in an arginine metabolite, asymmetrical dimethyl-arginine (ADMA) known as a risk factor for ischemic heart disease, during the acute phase of SEs of MELAS [17]. These biochemical studies suggested a trial of L-Arg to improve symptoms during the acute phase and preventing recurrence of SEs during the chronic phase in MELAS patients [17,18]. In addition to the vasodilatative effect by LArg, we speculated that there is also a pharmaceutical effect on cytopathy in MELAS. We therefore reported impairments in the tricarbonic acid (TCA) cycle metabolism and evaluated it in the hearts of MELAS patients in vivo using 11C-acetate PET [19]. 2. Review In this article, we will introduce novel in vivo functional brain imaging techniques, and discuss the pathogenesis of SEs in MELAS patients. We will further describe here our clinical experience with LArg therapy and discuss the dual pharmaceutical effects of this drug on MELAS.
2.1. In vivo functional brain imaging of SEs and its pathogenesis in MELAS The causal hypotheses for SEs in MELAS presented to date are angiopathy (neural damage due to impaired mitochondria in the blood vessels and inappropriate intracranial hemodynamics) [7,8,12,20], cytopathy (neural damage due to mitochondrial dysfunction itself) [9] and neuronal hyperexcitability (neural damage due to imbalance between energy supply and demand) [3,10]. Recent imaging techniques using MRI, MRS, SPECT and PET have facilitated in vivo evaluation of several pathophysiologic conditions such as brain edema, blood flow, neuronal damages, aerobic and anaerobic energy statuses, mitochondrial membrane potential and oxidative stress [12,21–23]; ADC on MRI can evaluate the proton mobility in the tissues (brain edema) [12]; MRS can evaluate anaerobic metabolism (lactate) and neuronal damages (N-acetylaspartate; NAA); Continuous arterial spin labeling images (CASL) on MRI can evaluate regional cerebral blood flow [23]; SPECT can evaluate the mitochondrial membrane potential or lipid metabolism [21]; PET using 62Cudiacetyl-bis(N 4-methylthiosemicarbazone) (ATSM) or 18F-fluorodeoxyglucose (FDG) can evaluate oxidative stress or glucose metabolism, respectively [22] (Fig. 1). Thus, we have now obtained powerful in vivo imaging tools for investigating the pathogenesis of SEs in MELAS patients. We performed several imaging studies during different stages of SEs in MELAS patients, and raised the following hypothesis as described previously [22]; (i) increased energy demand in brain cells with respiratory defect (i.e. neuronal hyperexcitability) triggered by several factors (ex. stress, infection, etc.), (ii) vasodilatation and hyperemia caused by impaired cerebral blood-vessel cells leading to vasogenic edema (i.e. angiopathy), (iii) overloading of oxygen and enhanced glucose metabolism in the neurons and glial cells through hyperemia, (iv) over-reduction state in impaired respiratory chains burdened with excess electrons due to glycolytic hypermetabolism, (v) enhanced ROS generation (oxidative stress) (i.e. cytopathy), and (vi) neuronal cell death caused mainly by oxidative damage (Fig. 1). The present and future development of imaging techniques is expected to better understand the pathogenesis of mitochondrial diseases (MELAS) and facilitate appropriate evaluation of potential therapies for these patients.
Fig. 1. Applications of functional brain imaging for evaluation of the pathogenesis of stroke-like episodes in MELAS, and the hypothesis underlying the process. ADC, apparent diffusion coefficient; CASL, continuous arterial spin labeling images; FDG, 18F-fluorodeoxyglucose; PET, positron emission tomography; MRS, magnetic resonance spectroscopy; ATSM, diacetyl-bis(N4-methylthiosemicarbazone); NAA, N-acetylaspartate.
M. Yoneda et al. / Biochimica et Biophysica Acta 1820 (2012) 615–618
Fig. 2. Lactate, pH, and arginine and citrulline monitored during L-Arg therapy for a comatose MELAS patient. The pH is indicated by red dots and lines. The level of lactate is indicated by blue dots and lines. The concentration of arginine is indicated by green bars. The concentration of citrulline is indicated by yellow bars. Bars attached to each scale bars in pH, lactate, Arg and Cit indicate individual normal ranges.
2.2. L-Arg therapy targeting for angiopathy: a lesson from a MELAS case rescued from a comatose state We present here a MELAS case that showed successful improvement following intravenous administration of L-Arg, and emphasize the usefulness of this therapy and the effects on angiopathy. A comatose 13-year-old boy was admitted to the emergency unit of the University of Fukui Hospital. This patient was born after normal pregnancy and developed normally in the infantile and child periods. His parents were non-consanguineous and had no signs of mitochondrial disease. This patient started to show recurrent SEs consisting of migraine headaches, vomiting and visual disturbance (homonymous hemianopsia) at the age of 9 years. At the age of 10 years, he was diagnosed as having MELAS after leukocyte DNA examination demonstrated A3243G mutation. His younger sister without any signs of MELAS also had the A3243G mutation. He had been administered vitamins and coenzyme Q10 orally. The detailed clinical course up to this period has been reported previously [12]. When he was admitted to the emergency unit of our hospital at the age of 13 years, he was in a comatose state and needed artificial ventilation support. He presented with recurrent partial complex
617
seizures extending from his left arm to the whole body. Brain stem responses such as light reflex, ciliary reflex, corneal reflex and oculocephalic reflex had completely disappeared. Arterial blood gas analysis demonstrated severe metabolic acidosis under 8 L/min. oxygen inspiration (pH 7.12, pCO2 43 Torr, pO2 254 Torr, BE −15.3). Brain MRI demonstrated marked swelling with multiple abnormal signals in the brain parenchyma. After written permission was obtained from his parents and approval for L-Arg therapy from the ethics committee of the University of Fukui, intravenous administration of L-Arg (0.5 g/kg weight for 30 min) had started for this comatose patient. Fifteen minutes after the L-Arg infusion had started, he showed voluntary movements in his face and extremities. Six hours after the L-Arg infusion, the level of his consciousness improved markedly from deep coma to the level of somnolence. He became alert and no longer required artificial ventilator support 36 hours after the infusion had started. The blood levels of lactate and pH, and the plasma Arg and Cit concentrations were monitored sequentially during L-Arg therapy (Fig. 2). The pH has normalized in 6 hours (indicated by red dots and lines), and the level of lactate became nearly normal in 36 hours (indicated by blue dots and lines). While the Arg and Cit were below the normal levels before L-Arg infusion, the level of Arg increased above the normal range (indicated by green bars) and the level of Cit returned to the nearly normal range (indicated by yellow bars) (Fig. 2). The present case demonstrates that intravenous administration of L-Arg shows a rapid clinical effect on acute stroke in a MELAS patient. The biochemical data supported the pharmaceutical effect of L-Arg as a strong vasodilator or modulator. Along with the acute effect of L-Arg infusion, oral administration of L-Arg prevented the recurrence of SEs in the chronic phase in MELAS patients [17]. We also successfully administered L-Arg to prevent recurrence to three MELAS patients with A3243G mutation who had previously presented with frequent with SEs. 2.3. Another pharmaceutical effect of L-Arg for cytopathy We speculated that another pharmaceutical effect of L-Arg was as an accelerator of the TCA cycle, as well as a vasodilator or modulator of microcirculation (Fig. 3). Recently our study using 11 C-acetate-PET demonstrated that the TCA cycle metabolic rate was markedly suppressed in the hearts of MELAS cardiomyopathic patients [19]. This indicates a shift in energy production to the
Fig. 3. Dual pharmaceutical effect of L-Arg. A. The effect on angiopathy by nitric oxide, a vasodilator. NOS, nitric oxide synthase; TCA, tricarbonic acid. B. The effect on cytopathy, TCA cycle metabolism.
618
M. Yoneda et al. / Biochimica et Biophysica Acta 1820 (2012) 615–618
anaerobic pathway. Remarkably, L-Arg can enhance TCA-cycle metabolism, regardless of its vasodilatation effect, and can be used as a treatment for patients with mitochondrial cardiomyopathy [19]. We expected that the regions of elevated TCA cycle kinetics would match the regions of improved myocardial blood flow (MBF). However, there was no apparent relationship between the TCA cycle kinetics and MBFs, which appears to support our hypothesis that L-Arg enters the TCA cycle by conversion to 2-oxoglutarate and accelerates the TCA cycle rate (elevation of TCA cycle kinetics without increasing MBF), which may rescue MELAS patients from the stress of excessive reduction in energy, with little relevance to the coronary microcirculation (Fig. 3B). Although this pharmaceutical effect on the energy metabolism in MELAS was observed in the heart not in the brain of MELAS patients, this finding clearly demonstrated that L-Arg has dual pharmaceutical effects such as acceleration of the TCA cycle (cytopathy) (Fig. 3B) and vasodilatation effect (angiopathy) (Fig. 3A). 3. Conclusions This article described several novel in vivo functional brain imaging techniques and their application for the evaluation of pathogenic conditions such as circulation, edema, neuronal cell death, aerobic and anaerobic energy statuses, mitochondrial membrane potentials and oxidative stress in MELAS patients. L-Arg has dual pharmaceutical effects on both angiopathy and cytopathy in MELAS. In vivo functional brain imaging is expected to promote further understanding of the pathogenesis of MELAS and facilitate monitoring of the clinical and biochemical effects of potential therapies including L-Arg. Acknowledgments This study was supported by the National Institute of Radiological Sciences, Chiba, Japan. This study was funded in part by Grants-in-Aid for Scientific Research (B) (17209040), Scientific Research on Innovative Areas (2020021) and Young Scientists (B) (21790838) from the Japan Society for the Promotion of Science, and the 21st Century COE Program (Medical Science). References [1] S.G. Pavlakis, P.C. Phillips, S. DiMauro, D.C. De Vivo, L.P. Rowland, Mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes: a distinctive clinical syndrome, Ann Neurol 16 (1984) 481–488. [2] Y. Goto, I. Nonaka, S. Horai, A mutation in the tRNA (Leu) (UUR) gene associated with the MELAS subgroup of mitochondrial encephalomyopathies, Nature 348 (1990) 651–653. [3] T. Iizuka, F. Sakai, Pathophysiology of stroke-like episodes in MELAS: neuron– astrocyte uncoupling in neuronal hyperexcitability, Future Neurol 5 (2010) 61–83.
[4] M. Kuriyama, H. Umezaki, Y. Fukuda, M. Osame, K. Koike, J. Tateishi, A. Igata, Mitochondrial encephalomyopathy with lactate-pyruvate elevation and brain infarctions, Neurology 34 (1884) 72–77. [5] T.I. Gropen, I. Prohovnik, T.K. Tatemichi, M. Hirano, Cerebral hyperemia in MELAS, Stroke 25 (1994) 1873–1976. [6] Y. Koga, Y. Akita, N. Junko, S. Yatsuga, N. Povalko, R. Fukiyama, M. Ishii, T. Matsuishi, Endothelial dysfunction in MELAS improved by L-arginine supplementation, Neurology 66 (2006) 1766–1769. [7] E. Ohama, S. Ohara, F. Ikuta, K. Tanaka, M. Nishizawa, T. Miyatake, Mitochondrial angiopathy in cerebral blood vessels of mitochondrial encephalomyopathy, Acta Neuropathol Berl 74 (1987) 226–233. [8] R. Sakuta, I. Nonaka, Vascular involvement in mitochondrial myopathy, Ann Neurol 25 (1989) 594–601. [9] T. Fujii, T. Okuna, M. Ito, K. Mutoh, Y. Horiguchi, H. Tashiro, H. Mikawa, MELAS of infantile onset: mitochondrial angiopathy or cytopathy? J Neurol Sci 10 (1991) 337–341. [10] T. Iizuka, F. Sakai, N. Suzuki, T. Hata, S. Tsukahara, M. Fukuda, Y. Takiyama, Neuronal hyperexcitability in stroke-like episodes of MELAS syndrome, Neurology 59 (2002) 816–824. [11] M. Matsuzaki, R. Takahashi, T. Nakayama, K. Shishikura, H. Suzuki, Y. Hirayama, M. Osawa, H. Oda, Disruption of endothelial tight junctions in a patient with mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS), Neuropediatrics 41 (2010) 72–74. [12] M. Yoneda, M. Maeda, H. Kimura, A. Fujii, K. Katayama, M. Kuriyama, Vasogenic edema on MELAS: a serial study with diffusion-weighted MR imaging, Neurology 53 (1999) 2182–2184. [13] T. Yasukawa, T. Suzuki, T. Ueda, S. Ohta, K. Watanabe, Modification defect at anticodon wobble nucleotide of mitochondrial tRNAs(Leu)(UUR) with pathogenic mutations of mitochondrial myopathy, encephalopathy, lactic acidosis, and strokelike episodes, J Biol Chem 275 (2000) 4251–4257. [14] M.P. King, Y. Koga, M. Davidson, E.A. Schon, Defects in mitochondrial protein synthesis and respiratory chain activity segregate with the tRNA(Leu(UUR)) mutation associated with mitochondrial myopathy, encephalopathy, lactic acidosis, and strokelike episodes, Mol Cell Biol 12 (1992) 480–490. [15] Y. Koga, M. Yoshino, H. Kato, MELAS exhibits dominant negative effects on mitochondrial RNA processing, Ann Neurol 43 (1998) 835. [16] A. Koga, Y. Koga Test, article sample title placed here, Neuromuscul Disord 13 (2003) 259–262. [17] Y. Koga, Y. Akita, J. Nishioka, S. Yatsuga, N. Povalko, Y. Tanabe, S. Fujimoto, T. Matsuishi, L-arginine improves the symptoms of strokelike episodes in MELAS, Neurology 64 (2005) 710–712. [18] Y. Koga, M. Ishibashi, I. Ueki, S. Yatsuga, R. Fukiyama, Y. Akita, T. Matsuishi, Effects of L-arginine on the acute phase of strokes in three patients with MELAS, Neurology 58 (2002) 827–828. [19] K. Arakawa, T. Kudo, M. Ikawa, N. Morikawa, Y. Kawai, K. Sahashi, J.D. Lee, M. Kuriyama, I. Mityamori, H. Okazawa, M. Yoneda, Abnormal myocardial energyproduction state in mitochondrial cardiomyopathy and acute response to Larginine infusion. C-11 acetate kinetics revealed by positron emission tomography, Circ J 74 (2010) 2702–2711. [20] J. Nishioka, Y. Akita, S. Yatsuga, K. Katayama, T. Matsuishi, M. Ishibashi, Y. Koga, Inappropriate intracranial hemodynamics in the natural course of MELAS, Brain Dev 30 (2008) 100–105. [21] M. Ikawa, Y. Kawai, K. Arakawa, T. Tsuchida, I. Miyamori, M. Kuriyama, M. Tanaka, M. Yoneda, Evaluation of respiratory chain failure in mitochondrial cardiomyopathy by assessments of 99mTc-MIBI washout and 123I-BMIPP/99mTc-MIBI mismatch, Mitochondrion 7 (2007) 164–170 E. [22] M. Ikawa, H. Okazawa, K. Arakawa, T. Kudo, H. Kimura, Y. Fujibayashi, M. Kuriyama, M. Yoneda, PET imaging of redox and energy states in stroke-like episodes of MELAS, Mitochondrion 9 (2009) 144–148. [23] T. Tsujikawa, M. Yoneda, Y. Shimizu, H. Uematsu, M. Toyooka, M. Ikawa, T. Kudo, H. Okazawa, M.K. Uriyama, H. Kimura, Pathophysiologic evaluation of MELAS strokes by serially quantified MRS and CASL perfusion images, Brain Dev 32 (2010) 143–149.
Contents lists available at ScienceDirect
Biochimica et Biophysica Acta j o u r n a l h o m e p a g e : w w w. e l s ev i e r. c o m / l o c a t e / b b a g e n
In vivo functional brain imaging and a therapeutic trial of L-arginine in MELAS patients☆ Makoto Yoneda a,⁎, Masamichi Ikawa a, Kenichiro Arakawa b, Takashi Kudo c, Hirohiko Kimura d, Yasuhisa Fujibayashi e, Hidehiko Okazawa c a
Department of Neurology, Faculty of Medical Sciences, University of Fukui, Fukui, Japan Department of Cardiology, Faculty of Medical Sciences, University of Fukui, Fukui, Japan Biomedical Imaging Research Center, University of Fukui, Fukui, Japan d Department of Radiology, Faculty of Medical Sciences, University of Fukui, Fukui, Japan e Molecular Imaging Center, National Institute of Radiological Sciences, Chiba, Japan b c
a r t i c l e
i n f o
Article history: Received 7 April 2011 Received in revised form 28 April 2011 Accepted 29 April 2011 Available online 8 May 2011 Keywords: MELAS Functional brain imaging PET MRI L-arginine
a b s t r a c t Background: Mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes (MELAS) is the most common type of mitochondrial disease and is characterized by stroke-like episodes (SEs), myopathy, lactic acidosis, diabetes mellitus, hearing-loss and cardiomyopathy. The causal hypotheses for SEs in MELAS presented to date are angiopathy, cytopathy and neuronal hyperexcitability. L-arginine (Arg) has been applied for the therapy in MELAS patients. Scope of review: We will introduce novel in vivo functional brain imaging techniques such as MRI and PET, and discuss the pathogenesis of SEs in MELAS patients. We will further describe here our clinical experience with L-arg therapy and discuss the dual pharmaceutical effects of this drug on MELAS. Major conclusions: Administration of L-arg to MELAS patients has been successful in reducing neurological symptoms due to acute strokes and preventing recurrences of SEs in the chronic phase. L-Arg has dual pharmaceutical effects on both angiopathy and cytopathy in MELAS. General significance: In vivo functional brain imaging promotes a better understanding of the pathogenesis and potential therapies for MELAS patients. This article is part of a Special Issue entitled Biochemistry of Mitochondria, Life and Intervention 2010. © 2011 Elsevier B.V. All rights reserved.
1. Background Mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes (MELAS), was first described by Pavlakis et al., and is the most common type of mitochondrial disease [1]. While over 40 mutations in the mitochondrial DNA (mtDNA) have been reported in association with MELAS, an A-to-G transition at nucleotide position 3243 (A3243G) in mtDNA is responsible for approximately 80% of the affected patients [2,3]. The clinical features of MELAS consist of strokelike episodes (SEs), myopathy, lactic acidosis, diabetes mellitus, hearing-loss and cardiomyopathy [1]. SEs provoke various neurological symptoms due to the affected brain lesions. These SEs are crucial life-threatening factors that determine the
☆ This article is part of a Special Issue entitled Biochemistry of Mitochondria, Life and Intervention 2010. ⁎ Corresponding author at: Department of Neurology, Faculty of Medical Sciences, University of Fukui, Fukui 910-1193, Japan. Tel.: + 81 776 61 8351; fax: + 81 776 61 8110. E-mail address: [email protected] (M. Yoneda). 0304-4165/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.bbagen.2011.04.018
prognosis of patients with MELAS. These SEs demonstrate unique characteristics different from ordinary ischemic strokes [1]. Stroke-like lesions spread beyond the vascular territory and are expanded to neighboring areas of the brain [4]. In the acute phase (several days after the stroke onset), there is focal vasodilatation and hyper-perfusion whereas, in the chronic phase (several months after the stroke onset), hypo-perfusion and irreversible changes appear [5]. However, in the super-acute phase (within 3 hours after the stroke onset), Koga et al. demonstrated hypoperfusion in the stroke lesion by a single photon emission tomography (SPECT) analysis [6]. Although several casual hypotheses of SEs have been proposed, the pathogenesis of SEs remains obscure. Causal hypotheses for SEs presented to date are angiopathy [7,8], cytopathy [9] and neuronal hyperexcitability [3,10]. Especially, the hypothesis involving angiopathy is one of the most attractive ideas explaining SEs in MELAS patients. Abnormal accumulation of mitochondria was described in the vascular endothelium and smooth muscle cells in small arteries and arterioles of the autopsied brain of a MELAS patient, and “mitochondrial angiopathy” was proposed 25 years ago [7]. A recent electron microscopic study further demonstrated disruption of capillary endothelial tight junctions in the autopsied brain of a patient with MELAS [11]. This
616
M. Yoneda et al. / Biochimica et Biophysica Acta 1820 (2012) 615–618
endothelial barrier disruption can result in impairments of the blood brain barrier and ion homeostasis between extra- and intra-cellular fluid, causing vasogenic edema that was detected on an apparent diffusion coefficient (ADC) map of the affected brain in a MELAS patient on magnetic resonance imaging (MRI) [12]. A physiological study also supported the angiopathy evidenced by a decreased vasodilatation capacity in small arteries examined by the flow mediated vasodilatation (FMD) method [6]. There are several biochemical data supportive of cytopathy in MELAS especially in association with A3243G mutation; i.e. increased unstabletRNA-Leu(UUR), increased ratio of uncharged aminoacyl-tRNALeu(UUR), accumulation of aminoacylation with leucine without any misacylation, deficient taurine modification and processing abnormality of the mitochondrial transcripts [13–16]. These biochemical defects lead cells to energy failure such as enhanced glycolysis, impaired electrontransport and oxidative phosphorylation, finally resulting in decreased ATP production. The recent imaging studies using MRI, magnetic resonance spectroscopy (MRS), SPECT and positron emission tomography (PET) demonstrated the process of SEs or pathophysiology in MELAS patients, as described below. Koga et al. found a decrease in L-arginine (Arg) that is metabolized into citrulline (Cit) and nitric oxide (a strong vasodilator), and relative increase in an arginine metabolite, asymmetrical dimethyl-arginine (ADMA) known as a risk factor for ischemic heart disease, during the acute phase of SEs of MELAS [17]. These biochemical studies suggested a trial of L-Arg to improve symptoms during the acute phase and preventing recurrence of SEs during the chronic phase in MELAS patients [17,18]. In addition to the vasodilatative effect by LArg, we speculated that there is also a pharmaceutical effect on cytopathy in MELAS. We therefore reported impairments in the tricarbonic acid (TCA) cycle metabolism and evaluated it in the hearts of MELAS patients in vivo using 11C-acetate PET [19]. 2. Review In this article, we will introduce novel in vivo functional brain imaging techniques, and discuss the pathogenesis of SEs in MELAS patients. We will further describe here our clinical experience with LArg therapy and discuss the dual pharmaceutical effects of this drug on MELAS.
2.1. In vivo functional brain imaging of SEs and its pathogenesis in MELAS The causal hypotheses for SEs in MELAS presented to date are angiopathy (neural damage due to impaired mitochondria in the blood vessels and inappropriate intracranial hemodynamics) [7,8,12,20], cytopathy (neural damage due to mitochondrial dysfunction itself) [9] and neuronal hyperexcitability (neural damage due to imbalance between energy supply and demand) [3,10]. Recent imaging techniques using MRI, MRS, SPECT and PET have facilitated in vivo evaluation of several pathophysiologic conditions such as brain edema, blood flow, neuronal damages, aerobic and anaerobic energy statuses, mitochondrial membrane potential and oxidative stress [12,21–23]; ADC on MRI can evaluate the proton mobility in the tissues (brain edema) [12]; MRS can evaluate anaerobic metabolism (lactate) and neuronal damages (N-acetylaspartate; NAA); Continuous arterial spin labeling images (CASL) on MRI can evaluate regional cerebral blood flow [23]; SPECT can evaluate the mitochondrial membrane potential or lipid metabolism [21]; PET using 62Cudiacetyl-bis(N 4-methylthiosemicarbazone) (ATSM) or 18F-fluorodeoxyglucose (FDG) can evaluate oxidative stress or glucose metabolism, respectively [22] (Fig. 1). Thus, we have now obtained powerful in vivo imaging tools for investigating the pathogenesis of SEs in MELAS patients. We performed several imaging studies during different stages of SEs in MELAS patients, and raised the following hypothesis as described previously [22]; (i) increased energy demand in brain cells with respiratory defect (i.e. neuronal hyperexcitability) triggered by several factors (ex. stress, infection, etc.), (ii) vasodilatation and hyperemia caused by impaired cerebral blood-vessel cells leading to vasogenic edema (i.e. angiopathy), (iii) overloading of oxygen and enhanced glucose metabolism in the neurons and glial cells through hyperemia, (iv) over-reduction state in impaired respiratory chains burdened with excess electrons due to glycolytic hypermetabolism, (v) enhanced ROS generation (oxidative stress) (i.e. cytopathy), and (vi) neuronal cell death caused mainly by oxidative damage (Fig. 1). The present and future development of imaging techniques is expected to better understand the pathogenesis of mitochondrial diseases (MELAS) and facilitate appropriate evaluation of potential therapies for these patients.
Fig. 1. Applications of functional brain imaging for evaluation of the pathogenesis of stroke-like episodes in MELAS, and the hypothesis underlying the process. ADC, apparent diffusion coefficient; CASL, continuous arterial spin labeling images; FDG, 18F-fluorodeoxyglucose; PET, positron emission tomography; MRS, magnetic resonance spectroscopy; ATSM, diacetyl-bis(N4-methylthiosemicarbazone); NAA, N-acetylaspartate.
M. Yoneda et al. / Biochimica et Biophysica Acta 1820 (2012) 615–618
Fig. 2. Lactate, pH, and arginine and citrulline monitored during L-Arg therapy for a comatose MELAS patient. The pH is indicated by red dots and lines. The level of lactate is indicated by blue dots and lines. The concentration of arginine is indicated by green bars. The concentration of citrulline is indicated by yellow bars. Bars attached to each scale bars in pH, lactate, Arg and Cit indicate individual normal ranges.
2.2. L-Arg therapy targeting for angiopathy: a lesson from a MELAS case rescued from a comatose state We present here a MELAS case that showed successful improvement following intravenous administration of L-Arg, and emphasize the usefulness of this therapy and the effects on angiopathy. A comatose 13-year-old boy was admitted to the emergency unit of the University of Fukui Hospital. This patient was born after normal pregnancy and developed normally in the infantile and child periods. His parents were non-consanguineous and had no signs of mitochondrial disease. This patient started to show recurrent SEs consisting of migraine headaches, vomiting and visual disturbance (homonymous hemianopsia) at the age of 9 years. At the age of 10 years, he was diagnosed as having MELAS after leukocyte DNA examination demonstrated A3243G mutation. His younger sister without any signs of MELAS also had the A3243G mutation. He had been administered vitamins and coenzyme Q10 orally. The detailed clinical course up to this period has been reported previously [12]. When he was admitted to the emergency unit of our hospital at the age of 13 years, he was in a comatose state and needed artificial ventilation support. He presented with recurrent partial complex
617
seizures extending from his left arm to the whole body. Brain stem responses such as light reflex, ciliary reflex, corneal reflex and oculocephalic reflex had completely disappeared. Arterial blood gas analysis demonstrated severe metabolic acidosis under 8 L/min. oxygen inspiration (pH 7.12, pCO2 43 Torr, pO2 254 Torr, BE −15.3). Brain MRI demonstrated marked swelling with multiple abnormal signals in the brain parenchyma. After written permission was obtained from his parents and approval for L-Arg therapy from the ethics committee of the University of Fukui, intravenous administration of L-Arg (0.5 g/kg weight for 30 min) had started for this comatose patient. Fifteen minutes after the L-Arg infusion had started, he showed voluntary movements in his face and extremities. Six hours after the L-Arg infusion, the level of his consciousness improved markedly from deep coma to the level of somnolence. He became alert and no longer required artificial ventilator support 36 hours after the infusion had started. The blood levels of lactate and pH, and the plasma Arg and Cit concentrations were monitored sequentially during L-Arg therapy (Fig. 2). The pH has normalized in 6 hours (indicated by red dots and lines), and the level of lactate became nearly normal in 36 hours (indicated by blue dots and lines). While the Arg and Cit were below the normal levels before L-Arg infusion, the level of Arg increased above the normal range (indicated by green bars) and the level of Cit returned to the nearly normal range (indicated by yellow bars) (Fig. 2). The present case demonstrates that intravenous administration of L-Arg shows a rapid clinical effect on acute stroke in a MELAS patient. The biochemical data supported the pharmaceutical effect of L-Arg as a strong vasodilator or modulator. Along with the acute effect of L-Arg infusion, oral administration of L-Arg prevented the recurrence of SEs in the chronic phase in MELAS patients [17]. We also successfully administered L-Arg to prevent recurrence to three MELAS patients with A3243G mutation who had previously presented with frequent with SEs. 2.3. Another pharmaceutical effect of L-Arg for cytopathy We speculated that another pharmaceutical effect of L-Arg was as an accelerator of the TCA cycle, as well as a vasodilator or modulator of microcirculation (Fig. 3). Recently our study using 11 C-acetate-PET demonstrated that the TCA cycle metabolic rate was markedly suppressed in the hearts of MELAS cardiomyopathic patients [19]. This indicates a shift in energy production to the
Fig. 3. Dual pharmaceutical effect of L-Arg. A. The effect on angiopathy by nitric oxide, a vasodilator. NOS, nitric oxide synthase; TCA, tricarbonic acid. B. The effect on cytopathy, TCA cycle metabolism.
618
M. Yoneda et al. / Biochimica et Biophysica Acta 1820 (2012) 615–618
anaerobic pathway. Remarkably, L-Arg can enhance TCA-cycle metabolism, regardless of its vasodilatation effect, and can be used as a treatment for patients with mitochondrial cardiomyopathy [19]. We expected that the regions of elevated TCA cycle kinetics would match the regions of improved myocardial blood flow (MBF). However, there was no apparent relationship between the TCA cycle kinetics and MBFs, which appears to support our hypothesis that L-Arg enters the TCA cycle by conversion to 2-oxoglutarate and accelerates the TCA cycle rate (elevation of TCA cycle kinetics without increasing MBF), which may rescue MELAS patients from the stress of excessive reduction in energy, with little relevance to the coronary microcirculation (Fig. 3B). Although this pharmaceutical effect on the energy metabolism in MELAS was observed in the heart not in the brain of MELAS patients, this finding clearly demonstrated that L-Arg has dual pharmaceutical effects such as acceleration of the TCA cycle (cytopathy) (Fig. 3B) and vasodilatation effect (angiopathy) (Fig. 3A). 3. Conclusions This article described several novel in vivo functional brain imaging techniques and their application for the evaluation of pathogenic conditions such as circulation, edema, neuronal cell death, aerobic and anaerobic energy statuses, mitochondrial membrane potentials and oxidative stress in MELAS patients. L-Arg has dual pharmaceutical effects on both angiopathy and cytopathy in MELAS. In vivo functional brain imaging is expected to promote further understanding of the pathogenesis of MELAS and facilitate monitoring of the clinical and biochemical effects of potential therapies including L-Arg. Acknowledgments This study was supported by the National Institute of Radiological Sciences, Chiba, Japan. This study was funded in part by Grants-in-Aid for Scientific Research (B) (17209040), Scientific Research on Innovative Areas (2020021) and Young Scientists (B) (21790838) from the Japan Society for the Promotion of Science, and the 21st Century COE Program (Medical Science). References [1] S.G. Pavlakis, P.C. Phillips, S. DiMauro, D.C. De Vivo, L.P. Rowland, Mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes: a distinctive clinical syndrome, Ann Neurol 16 (1984) 481–488. [2] Y. Goto, I. Nonaka, S. Horai, A mutation in the tRNA (Leu) (UUR) gene associated with the MELAS subgroup of mitochondrial encephalomyopathies, Nature 348 (1990) 651–653. [3] T. Iizuka, F. Sakai, Pathophysiology of stroke-like episodes in MELAS: neuron– astrocyte uncoupling in neuronal hyperexcitability, Future Neurol 5 (2010) 61–83.
[4] M. Kuriyama, H. Umezaki, Y. Fukuda, M. Osame, K. Koike, J. Tateishi, A. Igata, Mitochondrial encephalomyopathy with lactate-pyruvate elevation and brain infarctions, Neurology 34 (1884) 72–77. [5] T.I. Gropen, I. Prohovnik, T.K. Tatemichi, M. Hirano, Cerebral hyperemia in MELAS, Stroke 25 (1994) 1873–1976. [6] Y. Koga, Y. Akita, N. Junko, S. Yatsuga, N. Povalko, R. Fukiyama, M. Ishii, T. Matsuishi, Endothelial dysfunction in MELAS improved by L-arginine supplementation, Neurology 66 (2006) 1766–1769. [7] E. Ohama, S. Ohara, F. Ikuta, K. Tanaka, M. Nishizawa, T. Miyatake, Mitochondrial angiopathy in cerebral blood vessels of mitochondrial encephalomyopathy, Acta Neuropathol Berl 74 (1987) 226–233. [8] R. Sakuta, I. Nonaka, Vascular involvement in mitochondrial myopathy, Ann Neurol 25 (1989) 594–601. [9] T. Fujii, T. Okuna, M. Ito, K. Mutoh, Y. Horiguchi, H. Tashiro, H. Mikawa, MELAS of infantile onset: mitochondrial angiopathy or cytopathy? J Neurol Sci 10 (1991) 337–341. [10] T. Iizuka, F. Sakai, N. Suzuki, T. Hata, S. Tsukahara, M. Fukuda, Y. Takiyama, Neuronal hyperexcitability in stroke-like episodes of MELAS syndrome, Neurology 59 (2002) 816–824. [11] M. Matsuzaki, R. Takahashi, T. Nakayama, K. Shishikura, H. Suzuki, Y. Hirayama, M. Osawa, H. Oda, Disruption of endothelial tight junctions in a patient with mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS), Neuropediatrics 41 (2010) 72–74. [12] M. Yoneda, M. Maeda, H. Kimura, A. Fujii, K. Katayama, M. Kuriyama, Vasogenic edema on MELAS: a serial study with diffusion-weighted MR imaging, Neurology 53 (1999) 2182–2184. [13] T. Yasukawa, T. Suzuki, T. Ueda, S. Ohta, K. Watanabe, Modification defect at anticodon wobble nucleotide of mitochondrial tRNAs(Leu)(UUR) with pathogenic mutations of mitochondrial myopathy, encephalopathy, lactic acidosis, and strokelike episodes, J Biol Chem 275 (2000) 4251–4257. [14] M.P. King, Y. Koga, M. Davidson, E.A. Schon, Defects in mitochondrial protein synthesis and respiratory chain activity segregate with the tRNA(Leu(UUR)) mutation associated with mitochondrial myopathy, encephalopathy, lactic acidosis, and strokelike episodes, Mol Cell Biol 12 (1992) 480–490. [15] Y. Koga, M. Yoshino, H. Kato, MELAS exhibits dominant negative effects on mitochondrial RNA processing, Ann Neurol 43 (1998) 835. [16] A. Koga, Y. Koga Test, article sample title placed here, Neuromuscul Disord 13 (2003) 259–262. [17] Y. Koga, Y. Akita, J. Nishioka, S. Yatsuga, N. Povalko, Y. Tanabe, S. Fujimoto, T. Matsuishi, L-arginine improves the symptoms of strokelike episodes in MELAS, Neurology 64 (2005) 710–712. [18] Y. Koga, M. Ishibashi, I. Ueki, S. Yatsuga, R. Fukiyama, Y. Akita, T. Matsuishi, Effects of L-arginine on the acute phase of strokes in three patients with MELAS, Neurology 58 (2002) 827–828. [19] K. Arakawa, T. Kudo, M. Ikawa, N. Morikawa, Y. Kawai, K. Sahashi, J.D. Lee, M. Kuriyama, I. Mityamori, H. Okazawa, M. Yoneda, Abnormal myocardial energyproduction state in mitochondrial cardiomyopathy and acute response to Larginine infusion. C-11 acetate kinetics revealed by positron emission tomography, Circ J 74 (2010) 2702–2711. [20] J. Nishioka, Y. Akita, S. Yatsuga, K. Katayama, T. Matsuishi, M. Ishibashi, Y. Koga, Inappropriate intracranial hemodynamics in the natural course of MELAS, Brain Dev 30 (2008) 100–105. [21] M. Ikawa, Y. Kawai, K. Arakawa, T. Tsuchida, I. Miyamori, M. Kuriyama, M. Tanaka, M. Yoneda, Evaluation of respiratory chain failure in mitochondrial cardiomyopathy by assessments of 99mTc-MIBI washout and 123I-BMIPP/99mTc-MIBI mismatch, Mitochondrion 7 (2007) 164–170 E. [22] M. Ikawa, H. Okazawa, K. Arakawa, T. Kudo, H. Kimura, Y. Fujibayashi, M. Kuriyama, M. Yoneda, PET imaging of redox and energy states in stroke-like episodes of MELAS, Mitochondrion 9 (2009) 144–148. [23] T. Tsujikawa, M. Yoneda, Y. Shimizu, H. Uematsu, M. Toyooka, M. Ikawa, T. Kudo, H. Okazawa, M.K. Uriyama, H. Kimura, Pathophysiologic evaluation of MELAS strokes by serially quantified MRS and CASL perfusion images, Brain Dev 32 (2010) 143–149.