Carnosine (beta-alanyl-L-histidine, CAR) and related peptides are histidine-containing dipeptides particularly abundant in excitable tissues such as nervous system and skeletal muscle. Although their biochemical role is still unknown, some evidence indicates that these endogenous compounds can act as quenchers of reactive and cytotoxic carbonyl species (RCS). We firstly reported the structural evidence and ex vivo data supporting this hypothesis. As a first step, we investigated the reaction mechanism of CAR as quencher of alpha,beta-unsaturated aldehydes such as 4-hydroxy-trans-2,3-nonenal (HNE) [1] and acrolein (ACR) [2]. It was found that CAR efficiently detoxifies alpha,beta-unsaturated aldehydes through an auto-catalytic mechanism involving both the beta-alanine group and the nucleophilic imidazole ring, forming a stable Michael adduct between the C3 of the aldehyde and the N of the histidine group. The efficacy of CAR and related peptides such as anserine (ANS) as detoxifying agents of HNE was then demonstrated in biological matrices such as spontaneously oxidized rat skeletal muscle, by detecting the corresponding HNE-Michael adducts by liquid chromatography/electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS)[3]. The ability of CAR to quench ,β-unsaturated aldehydes was then demonstrated ex vivo, by detecting the HNE Michael adduct and correlated metabolites by liquid chromatography-tandem mass spectrometric (LC-MS/MS) approach, based on the precursor ion scanning technique using a triple-stage quadrupole. In particular, the MS approach was applied to urines from Zucker obese rats, a nondiabetic animal model characterized by obesity and hyperlipidemia, where RCS formation plays a key role in the development of renal and cardiac dysfunction. By this approach, the carnosine-HNE Michael adduct was identified together to the two following His metabolites: His-1,4-dihydroxynonane (His-DHN), His-4-hydroxynonanoic acid (His-HNA)[4]. The biological effect of CAR as quencher of RCS was then investigated in Zucker rats supplemented with CAR or with its enantiomer stable to carnosinase (beta-alanyl-D-histidine, D-CAR) (30 mg/kg in drinking water) for 24 weeks. CAR was found to greatly reduce obesity-related diseases in obese Zucker rats, significantly restraining the development of dyslipidaemia, hypertension and renal injury. We believe that the biological effects of CAR in Zucker obese rat are mediated by a direct carbonyl quenching mechanism and this hypothesis is firstly sustained by the in vivo capacity of supplemented CAR to quench RCS, as demonstrated by detecting CAR-HNE Michael adduct in the urine of obese rats, following CAR treatment. Further evidence suggesting a RCS-quenching mechanism is that the diseases inhibited by CAR have a common pathogenetic route, i.e. the oxidative/carbonyl stress, which represents the molecular target of CAR itself. The efficacy of D-CAR was found to be superimposable to that of CAR thus excluding a pro-histamine and receptor-mediated mechanism and confirming a RCS quenching mechanism [5]. In conclusion, among the several biological properties so far reported for CAR and related peptides, their ability to quench and detoxify cytotoxic RCS and in particular alpha,beta-unsaturated aldehydes should also be considered not only to better explain the biochemical role of these dipeptides but also to design novel bioactive drugs. [1] Aldini G et al. Biochem Biophys Res Commun. 298(5):699-706 (2002). [2] Carini M et al. J Mass Spectrom. 38(9):996-1006 (2003). [3] Orioli M et al. J Chromatogr B 827(1):109-18 (2005). [4] Orioli M et al. Anal Chem. 79(23):9174-84 (2007). [5] Aldini G et al. J Cell Mol Med. 15(6):1339-1354 (2011).

Carnosine and related dipeptides as quenchers of reactive carbonyl species / G. Aldini, M. Orioli, M. Carini - In: Atti del congresso: Carnosine in exercise and disease[s.l] : Medicongress, 2011. (( convegno Carnosine in exercise and disease tenutosi a Ghent (Belgium) nel 2011.

Carnosine and related dipeptides as quenchers of reactive carbonyl species

G. Aldini;M. Orioli;M. Carini
2011

Abstract

Carnosine (beta-alanyl-L-histidine, CAR) and related peptides are histidine-containing dipeptides particularly abundant in excitable tissues such as nervous system and skeletal muscle. Although their biochemical role is still unknown, some evidence indicates that these endogenous compounds can act as quenchers of reactive and cytotoxic carbonyl species (RCS). We firstly reported the structural evidence and ex vivo data supporting this hypothesis. As a first step, we investigated the reaction mechanism of CAR as quencher of alpha,beta-unsaturated aldehydes such as 4-hydroxy-trans-2,3-nonenal (HNE) [1] and acrolein (ACR) [2]. It was found that CAR efficiently detoxifies alpha,beta-unsaturated aldehydes through an auto-catalytic mechanism involving both the beta-alanine group and the nucleophilic imidazole ring, forming a stable Michael adduct between the C3 of the aldehyde and the N of the histidine group. The efficacy of CAR and related peptides such as anserine (ANS) as detoxifying agents of HNE was then demonstrated in biological matrices such as spontaneously oxidized rat skeletal muscle, by detecting the corresponding HNE-Michael adducts by liquid chromatography/electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS)[3]. The ability of CAR to quench ,β-unsaturated aldehydes was then demonstrated ex vivo, by detecting the HNE Michael adduct and correlated metabolites by liquid chromatography-tandem mass spectrometric (LC-MS/MS) approach, based on the precursor ion scanning technique using a triple-stage quadrupole. In particular, the MS approach was applied to urines from Zucker obese rats, a nondiabetic animal model characterized by obesity and hyperlipidemia, where RCS formation plays a key role in the development of renal and cardiac dysfunction. By this approach, the carnosine-HNE Michael adduct was identified together to the two following His metabolites: His-1,4-dihydroxynonane (His-DHN), His-4-hydroxynonanoic acid (His-HNA)[4]. The biological effect of CAR as quencher of RCS was then investigated in Zucker rats supplemented with CAR or with its enantiomer stable to carnosinase (beta-alanyl-D-histidine, D-CAR) (30 mg/kg in drinking water) for 24 weeks. CAR was found to greatly reduce obesity-related diseases in obese Zucker rats, significantly restraining the development of dyslipidaemia, hypertension and renal injury. We believe that the biological effects of CAR in Zucker obese rat are mediated by a direct carbonyl quenching mechanism and this hypothesis is firstly sustained by the in vivo capacity of supplemented CAR to quench RCS, as demonstrated by detecting CAR-HNE Michael adduct in the urine of obese rats, following CAR treatment. Further evidence suggesting a RCS-quenching mechanism is that the diseases inhibited by CAR have a common pathogenetic route, i.e. the oxidative/carbonyl stress, which represents the molecular target of CAR itself. The efficacy of D-CAR was found to be superimposable to that of CAR thus excluding a pro-histamine and receptor-mediated mechanism and confirming a RCS quenching mechanism [5]. In conclusion, among the several biological properties so far reported for CAR and related peptides, their ability to quench and detoxify cytotoxic RCS and in particular alpha,beta-unsaturated aldehydes should also be considered not only to better explain the biochemical role of these dipeptides but also to design novel bioactive drugs. [1] Aldini G et al. Biochem Biophys Res Commun. 298(5):699-706 (2002). [2] Carini M et al. J Mass Spectrom. 38(9):996-1006 (2003). [3] Orioli M et al. J Chromatogr B 827(1):109-18 (2005). [4] Orioli M et al. Anal Chem. 79(23):9174-84 (2007). [5] Aldini G et al. J Cell Mol Med. 15(6):1339-1354 (2011).
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