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Characterization of Beta₃-Adrenoceptors in Human Internal Mammary Artery and Putative Involvement in Coronary Artery Bypass Management

Bertrand Rozec, MD^_*,, Sabrina Serpillon, PhD^_*, Gilles Toumaniantz, PhD^_*, Camille Sèze, MSc^_*, Yohann Rautureau, PhD, Olivier Baron, MD, Jacques Noireaud, PhD^_* and Chantal Gauthier, PhD^_*,||,_*

^_* L’Institut du Thorax, INSERM UMR533, Faculté de Médecine, Nantes, France
Département d’anesthésie et de réanimation chirurgicale, CHRU G et R Laënnec, Nantes, France
Department of Veterinary Basic Science, Royal Veterinary College, London, United Kingdom
Service de chirurgie thoracique et cardiovasculaire, CHRU G et R Laënnec, Nantes, France
^|| Faculté des Sciences et Techniques, Université de Nantes, Nantes, France

View larger version (68K): [in a new window]   Figure 1 Detection by reverse transcription (RT) polymerase chain reaction of human 1A-adrenoceptors (1A-ARs), ß1-adrenoceptors (ß1-AR), ß2-ARs, and ß3-ARs, and hormone-sensitive lipase (LIPE) gene transcripts in human internal mammary artery (IMA) and white adipose tissue. Amplified cDNA fragments were separated on a 2% agarose gel and visualized by staining with ethidium bromide. Sizes of the polymerase chain reaction products are indicated on the middle of the panels. This experiment is representative of three investigations performed on three vessels obtained from different patients. ADNg = genomic DNA; AR-AS = reverse strand; AR-S = forward strand; hfat = human fat; M = 100 pb ladder (Biolabs, New England); NRT = none RT.

View larger version (31K): [in a new window]   Figure 2 Western blot analysis of human beta3-adrenoceptor (ß3-AR) expression in internal mammary artery with or without endothelium. The membrane was incubated without (left panel) or with antihuman ß3-AR antibody (Ab) (right panel) and revealed by chemiluminescence detection procedure. This experiment is representative of five investigations performed on five vessels obtained from different patients. With endo. = with endothelium; Without endo. = without endothelium.

View larger version (67K): [in a new window]   Figure 3 Immunohistochemistry analysis of human beta3-adrenoceptors (hß3-AR) expression in internal mammary artery. Adjacent 10-µm thick sections were incubated without (A, C, E, a, c, and e) or with anti-hß3-AR antibody (Ab) (B, D, F, b, d, and f) revealed by peroxydase-conjugated second antiserum. This experiment is representative of three investigations performed on three vessels obtained from different patients. Black arrowhead shows interesting areas at low magnitude (micrographs x20) that were further observed at high magnitude (micrographs x60).

View larger version (21K): [in a new window]   Figure 4 Effect of beta3-adrenoceptor (ß3-AR) agonists in human internal mammary arteries (IMA). (A) Typical records of relaxant effect of SR 58611A, a preferential ß3-AR agonist, and acetylcholine (1 µmol/l) in human IMA rings. (B) Concentration-response curves to SR 58611A performed in the absence or in the presence of 10 µmol/l nadolol, an ß1 and ß2-AR antagonist, and 3 µmol/l L-748,337, a human ß3-AR antagonist, in human IMA rings constricted with 1 µmol/l phenylephrine. The mean curves resulting from subtraction of the spontaneous relaxation of control vessels are shown. Results are expressed as percentage of relaxation from the contraction level induced by phenylephrine. Each point is the mean value of six experiments, and error bars represent SEM. *p < 0.05 and **p < 0.01 indicate significant differences from response to SR 58611A alone.

View larger version (18K): [in a new window]   Figure 5 Effect of NG-monomethyl-L-arginine monoacetate (L-NMMA) treatment or endothelium removal on SR 58611A-induced vasodilation in human internal mammary artery rings constricted with 1 µm phenylephrine (PE). Concentration-response curves to SR 58611A were performed in the absence or in the presence of 100 µmol/l L-NMMA, an inhibitor of nitric oxide synthases, or after endothelium removal. The mean curves resulting from subtraction of the spontaneous relaxation of control vessels are shown. Results are expressed as percentage of relaxation from the contraction level induced by PE. Each point is the mean value of six experiments, and error bars represent SEM. **p < 0.01 indicates significant differences from response to SR 58611A alone.