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(Circulation. 2002;105:380.)
© 2002 American Heart Association, Inc.

Basic Science Reports

Bone Marrow–Derived Regenerated Cardiomyocytes (CMG Cells) Express Functional Adrenergic and Muscarinic Receptors

Daihiko Hakuno, MD, PhD; Keiichi Fukuda, MD; Shinji Makino, MD; Fusako Konishi, PhD; Yuichi Tomita, MD; Tomohiro Manabe, MD; Yusuke Suzuki, MD; Akihiro Umezawa, MD, PhD; Satoshi Ogawa, MD, PhD

From the Cardiopulmonary Division (D.H., K.F., S.M., F.K., Y.T., T.M., Y.S., S.O.), Department of Internal Medicine, Institute for Advanced Cardiac Therapeutics (K.F.), and the Department of Pathology (A.U.), Keio University School of Medicine, Tokyo, Japan.

Correspondence to Keiichi Fukuda, MD, PhD, Institute for Advanced Cardiac Therapeutics, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582, Japan. E-mail:kfukuda{at}sc.itc.keio.ac.jp

Background— We recently reported that cardiomyocytes could be differentiated from bone marrow mesenchymal stem cells in vitro by 5-azacytidine treatment. In native cardiomyocytes, adrenergic and muscarinic receptors play crucial roles in mediating heart rate, conduction velocity, contractility, and cardiac hypertrophy. We investigated whether these receptors are expressed in differentiated CMG cells, and if so, whether they have downstream signaling systems.

Methods and Results— Reverse transcription–polymerase chain reaction revealed that CMG cells had already expressed _1A-, _1B-, and _1D-adrenergic receptor mRNA before 5-azacytidine treatment, whereas expression of ß₁-, ß₂-adrenergic and M₁-, M₂-muscarinic receptors was first detected at 1 day. Phenylephrine dose-dependently induced phosphorylation of ERK1/2, which was completely inhibited by prazosin, and significantly increased cell size. Isoproterenol augmented cAMP by 38-fold, which was fully inhibited by propranolol. Isoproterenol (10^-7 mol/L) increased the spontaneous beating rate by 47.6% (basal, 127±16 bpm), and propranolol and CGP20712A (ß₁-selective blocker) reduced it by 79.0% and 71.0%, respectively, whereas ICI118551 (ß₂-selective blocker) induced slight reduction. Cell motion, percent shortening, and contractile velocity were increased by 37.5%, 26.9%, and 50.6%, respectively, in response to isoproterenol. Phenylephrine and isoproterenol augmented ANP and BNP gene expressions. Carbachol increased IP₃ by 32-fold, which was markedly inhibited by atropine as well as AFDX116 (M₂-selective blocker) measured by radioimmunoassay.

Conclusions— These findings indicate that CMG cells expressed _1A, _1B, and _1D receptors before differentiation and expressed ß₁, ß₂, M₁, and M₂ receptors after they obtained the cardiomyocyte phenotype. These receptors had functional signal transduction pathways and could modulate cell function.

Key Words: cells • heart rate • receptors, adrenergic, beta • signal transduction • receptors, adrenergic, alpha

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