Histidine-containing Dipeptide Carnosine Research Articles

Histidine-Containing Dipeptides Mitigate Salt Stress in Plants by Scavenging Reactive Carbonyl Species.

Reactive oxygen species (ROS) are critical factors that cause damage in salt-stressed plants, but their mechanisms of action in living cells are largely unknown. We investigated the roles of reactive carbonyl species (RCS), i.e., the lipid peroxide-derived α,β-unsaturated aldehydes and ketones, in plant growth retardation under salt stress. When Arabidopsis thaliana Col-0 seeds were exposed to 100 mM NaCl, germination was delayed and the levels of ROS, RCS, and protein carbonylation in the seedlings were increased. Adding the histidine-containing dipeptides carnosine, N-acetylcarnosine, and anserine, which are reported RCS scavengers, restored the germination speed and suppressed the increases in RCS and protein carbonylation but did not affect the ROS level. Increases in the levels of the RCS acrolein, crotonaldehyde, (E)-2-pentenal, and 4-hydroxy-(E)-2-nonenal were positively correlated with the delay of germination and growth inhibition. These RCS, generated downstream of ROS, are thus primarily responsible for the salt-stress symptoms of plants.

CNDP1 knockout in zebrafish alters the amino acid metabolism, restrains weight gain, but does not protect from diabetic complications.

The gene CNDP1 was associated with the development of diabetic nephropathy. Its enzyme carnosinase 1 (CN1) primarily hydrolyzes the histidine-containing dipeptide carnosine but other organ and metabolic functions are mainly unknown. In our study we generated CNDP1 knockout zebrafish, which showed strongly decreased CN1 activity and increased intracellular carnosine levels. Vasculature and kidneys of CNDP1-/- zebrafish were not affected, except for a transient glomerular alteration. Amino acid profiling showed a decrease of certain amino acids in CNDP1-/- zebrafish, suggesting a specific function for CN1 in the amino acid metabolisms. Indeed, we identified a CN1 activity for Ala-His and Ser-His. Under diabetic conditions increased carnosine levels in CNDP1-/- embryos could not protect from respective organ alterations. Although, weight gain through overfeeding was restrained by CNDP1 loss. Together, zebrafish exhibits CN1 functions, while CNDP1 knockout alters the amino acid metabolism, attenuates weight gain but cannot protect organs from diabetic complications.

Effect of carnosine on conditioned passive avoidance response in the norm and under hypoxia conditions

We studied the effect of the histidine-containing dipeptide carnosine in the norm and under hypoxia conditions on the conditioned passive avoidance response in a three-compartment chamber, which consisted of a light compartment, a dark dangerous compartment where the rat received a foot shock, and a dark safety compartment, where the rat did not receive a foot shock. We found that carnosine normalized physiological indices, which were impaired by acute exposure to hypobaric hypoxia, and increased the number of choices of the safety compartment in both intact rats and rats subjected to hypoxia.

Prevention of iron-mediated cytotoxicity of cultured endothelial and renal tubular epithelial cells by carnosine treatment

Epidemiological studies have indicated a positive association between high body iron stores and the risk of type 2 diabetes (T2DM). High ferritin levels in T2DM are closely related to the development of diabetic vascular complications, possibly through the interaction with VEGF. An increased transferrin excretion in diabetic patients with microalbuminuria may contribute to tubulointerstitial injuries due to transferrin reabsorption and thus increased intracellular iron concentrations in proximal tubular epithelial cells (PTEC). Because the histidine-containing dipeptide carnosine has proven its therapeutic efficacy in a variety of in vivo models for T2DM, we assessed: 1) if high glucose (HG) exposure makes cultured endothelial cells and PTEC more susceptible to metal-induced toxicity, 2) if this correlates with increased expression of known iron transporters (DMT1, IREG, TFRC), and 3) if carnosine can prevent toxicity in these cells.

Natural Dipeptides as Mini-Chaperones: Molecular Mechanism of Inhibition of Lens βL-Crystallin Aggregation

The effect of histidine-containing dipeptides-carnosine and N-acetylcarnosine-on preventing and treating of cataracts of various etiologic origins has been demonstrated in many studies in vivo, while the precise molecular mechanism of their action is actually obscure. Cataract has been recently attributed to conformational diseases due to the association of lens structure protein aggregation with cataract pathogenesis. In our study, effect of histidine-containing dipeptides-carnosine, N-acetylcarnosine, and anserine-on the UV induced βL-crystallin aggregation was studied in vitro. It was first demonstrated that N-acetylcarnosine and anserine (10-40 mM) considerably suppressed UV induced aggregation of βL-crystallin, while carnosine exerted no effect. Positive correlation between anti-aggregating activity of the compounds used and their hydrophobicity was obtained. It was revealed that N-acetylcarnosine and anserine inhibited the initial stages of the protein photochemical damage. A decrease in the size of protein aggregates was detected in the presence of N-acetylcarnosine and anserine. UV irradiation of βL-crystallin resulted in a significant increase in the number of protein carbonyl groups, and the dipeptides studied did not affect this process. We suppose that N-acetylcarnosine and anserine inhibit βL-crystallin aggregation via formation of a protein-dipeptide complex that prevents macromolecular conformational changes and ensuing protein aggregation.

Carnosine protects against NMDA-induced neurotoxicity in differentiated rat PC12 cells through carnosine-histidine-histamine pathway and H 1/H 3 receptors

Since the histidine-containing dipeptide carnosine (β-alanyl- l-histidine) is believed to have many physiological functions in the brain, we investigated the neuroprotective effects of carnosine and its mechanisms of action in an in vitro model of neurotoxicity induced by N-methyl- d-aspartate (NMDA) in differentiated PC12 cells. Pretreatment with carnosine increased the viability and decreased the number of apoptotic and necrotic cells measured by MTT and Hoechst 33342 and propidium iodide (PI) double staining assays. Carnosine also can inhibite the glutamate release and increase HDC activity and the intracellular and extracellular contents of carnosine, histidine and histamine detected by high-performance liquid chromatography (HPLC). The protection by carnosine was reversed by α- fluoromethylhistidine, a selective and irreversible inhibitor of histidine decarboxylase (HDC). Pyrilamine and thioperamide, selective central histamine H 1 and H 3 antagonists also significantly reversed the protection of carnosine. Further, the inhibition of glutamate release by carnosine was reversed by thioperamide. Therefore, the protective mechanism of carnosine may not only involve the carnosine-histidine-histamine pathway, but also H 1/H 3 receptors and the effective inhibition of glutamate release. This study indicates that carnosine may be an endogenous protective factor and calls for its further study as a new antiexcitotoxic agent.

LC–ESI-MS/MS determination of 4-hydroxy-trans-2-nonenal Michael adducts with cysteine and histidine-containing peptides as early markers of oxidative stress in excitable tissues

A sensitive, selective, specific and rapid liquid chromatographic-electrospray ionization tandem mass spectrometric assay was developed and validated for the simultaneous determination in skeletal muscle of the Michael adducts between 4-hydroxy-trans-2-nonenal (HNE), one of the most reactive lipid peroxidation-driven unsaturated aldehyde, and glutathione (GSH) and the endogenous histidine-containing dipeptides carnosine (CAR) and anserine (ANS), with the final aim to use conjugated adducts as specific and unequivocal markers of lipid peroxidation. Samples (skeletal muscle homogenates from male rats) were prepared by protein precipitation with 1 vol. of a HClO(4) solution (4.2%; w/v) containing H-Tyr-His-OH as internal standard. The supernatant, diluted (1:1, v/v) in mobile phase, was separated on a Phenomenex Sinergy polar-RP column with a mobile phase of water-acetonitrile-heptafluorobutyric acid (9:1:0.01, v/v/v) at a flow rate of 0.2 ml/min, with a run time of 12 min. Detection was on a triple quadrupole mass spectrometer equipped with an ESI interface operating in positive ionization mode. The acquisitions were in multiple reaction monitoring (MRM) mode using the following precursor-->product ion combinations: H-Tyr-His-OH (IS): m/z 319.2--> 156.5+301.6; GS-HNE: m/z 464.3--> 179.1+308.0; CAR-HNE: m/z 383.1--> 110.1+266.6; ANS-HNE: m/z 397.2--> 109.1+126.1. The method was validated over the concentration ranges 1.5-90 (GS-HNE) and 0.4-40 (CAR-HNE, ANS-HNE) nmoles/g wet tissue, and the LLOQ were 1.25 and 0.33 pmoles injected respectively. The intra- and inter-day precisions (CV%) were <7.38% (<or=10.90% at the LLOQs); intra- and inter-assay accuracy (RE%) was within +/-7.0% for all the concentrations (<or=18% at the LLOQs). The method was applied to quantitate peptide-HNE Michael adducts in rat skeletal muscles exposed to oxidative stress to endogenously generate HNE, and the results indicate that CAR-HNE can be considered as an early, specific and stable marker of lipid peroxidation in excitable tissues.

Profiling histidine‐containing dipeptides in rat tissues by liquid chromatography/electrospray ionization tandem mass spectrometry

The histidine-containing dipeptides carnosine (CAR) and structurally related anserine (ANS) and homocarnosine (HCAR), widely distributed in vertebrate organisms, have recently been proposed as endogenous quenchers for highly cytotoxic alpha,beta-unsaturated aldehydes generated by peroxidation. A sensitive, selective, specific and rapid liquid chromatographic/electrospray ionization tandem mass spectrometric assay was developed and validated for the simultaneous determination of these peptides in biological matrices in order to establish their plasma/tissue distribution. Samples (plasma or tissue homogenates from male rats) were prepared by protein precipitation with HClO(4) (1 : 1, v/v) containing H-Tyr-His-OH as internal standard. The supernatant was separated on a Phenomenex Sinergy polar-RP column with a mobile phase of water-acetonitrile-heptafluorobutyric acid (9 : 1 : 0.01, v/v/v) at a flow-rate of 0.2 ml min(-1), with a run time of 10 min. Detection was effected on an ion trap mass spectrometer equipped with an electrospray ionization interface operating in positive ionization mode. The acquisitions were in the multiple reaction monitoring mode using the following precursor --> product ion combinations: H-Tyr-His-OH (internal standard) m/z 319 --> 301; CAR m/z 227 --> 210 + 209; ANS m/z 241 --> 224 + 197 + 170; HCAR m/z 241 --> 156. The method was validated over the concentration range 15-1000 nmol g(-1) and the limit of quantification (LOQ) and limit of detection (LOD) were 12.5 and 4.2 pmol injected, respectively. The intra- and inter-day precisions were <10% (< or =17.47% at the LOQ) and the intra- and inter-assay accuracies were within +/-10% for all concentrations. The mapping profile in rat tissue gave the following results: the highest concentrations of CAR and ANS were found in skeletal muscles (soleus, gastrocnemius, tibialis), followed by the heart, cerebellum and brain (ANS below the LOQ). HCAR was found only in the brain and cerebellum. No histidine-containing dipeptides were detectable in plasma, liver, kidney and lung.

トリ胸肉抽出物(CBEX^&lt;TM&gt;)の経口摂取が高強度間欠的運動パフォーマンスに及ぼす影響

The histidine-containing dipeptide carnosine (p-alanyl-L-histidine) exists widely in the skeletal muscle of vertebrates. The biological role of carnosine has not been well established, but it is believed to act as a cytosolic buffering agent. We prepared an extract of chicken breast meat that is an abundant source of carnosine and its related dipeptide anserine (β-alanyl-1-methyl-L-histidine). The purpose of this study was to examine the effects of extract (CBEX^<TM>) supplementation on high-intensity intermittent exercise performance. Eight male subjects performed two experimental trials (CBEX and placebo treatment). Each trial consisted of two intermittent cycle ergometer sprints separated by a 20-min recovery period. Intermittent exercise consisted often 5-s maximal sprints with 25-s of recovery between each sprint. The subjects were given 190 g of experimental soup containing either 40 g of CBEX or no CBEX 30 min before each trial. Anserine and its related amino acid in plasma were detectable 30 min after CBEX supplementation. However, plasma carnosine was not detectable. In the CBEX group, the subjects exhibited high power during the latter half of intermittent exercise compared with the placebo group. These results suggest that pre-exercise CBEX supplementation improves intermittent exercise performance, possibility by restraining the decrease of intracellular pH and thereby delaying the onset of fatigue.

The Effect of Sprint Training on Skeletal Muscle Carnosine in Humans

It has been suggested that histidine-containing dipeptide carnosine (β-alanyl-L-histidine), which is believed to act as a cytosolic buffering agent, is present predominantly in skeletal muscle. The purpose of this study was to investigate the effects of sprint training (30-s maximal cycle ergometer sprinting) on muscle carnosine concentration. Six untrained males trained 2 days per week for 8 weeks on an electronic-braked cycle ergometer. Muscle biopsy samples were taken from the vastus lateralis before and two days after the last training session and were analyzed for carnosine concentration by the use of an amino acid autoanalyzer. The carnosine concentration was significantly increased after sprint training (P < 0.05). The mean power during 30-s maximal cycle ergometer sprinting was significantly increased following training. When dividing the 30-s sprinting into 6 phases (0-5, 6-10, 11-15, 16-20, 21-25, 26-30 s), the magnitude of increase in mean power was significantly larger for the last 2 phases than the first phase (P < 0.05). These results suggest that the increases in skeletal muscle carnosine concentration following sprint training may be associated with the increase in sustainability of high power during 30-s maximal cycle ergometer sprinting.

High Level of Skeletal Muscle Carnosine Contributes to the Latter Half of Exercise Performance during 30-s Maximal Cycle Ergometer Sprinting.

The histidine-containing dipeptide carnosine (beta-alanyl-L-histidine) has been shown to significantly contribute to the physicochemical buffering in skeletal muscles, which maintains acid-base balance when a large quantity of H(+) is produced in association with lactic acid accumulation during high-intensity exercise. The purpose of the present study was to examine the relations among the skeletal muscle carnosine concentration, fiber-type distribution, and high-intensity exercise performance. The subjects were 11 healthy men. Muscle biopsy samples were taken from the vastus lateralis at rest. The carnosine concentration was determined by the use of an amino acid autoanalyzer. The fiber-type distribution was determined by the staining intensity of myosin adenosinetriphosphatase. The high-intensity exercise performance was assessed by the use of 30-s maximal cycle ergometer sprinting. A significant correlation was demonstrated between the carnosine concentration and the type IIX fiber composition (r=0.646, p<0.05). The carnosine concentration was significantly correlated with the mean power per body mass (r=0.785, p<0.01) during the 30-s sprinting. When dividing the sprinting into 6 phases (0-5, 6-10, 11-15, 16-20, 21-25, 26-30 s), significant correlations were observed between the carnosine concentration and the mean power per body mass of the final 2 phases (21-25 s: r=0.694, p<0.05; 26-30 s: r=0.660, p<0.05). These results indicated that the carnosine concentration could be an important factor in determining the high-intensity exercise performance.

Molecular mechanisms of regulation of the activity of sarcoplasmic reticulum Ca-release channels (ryanodine receptors), muscle fatigue, and Severin's phenomenon.

In this short review of the literature and our own data the characteristics of structural organization of sarcoplasmic reticulum Ca-release channels (ryanodine receptors) in different types of muscles, the participation of other sarcoplasmic reticulum proteins in excitation-contraction coupling and Ca-release channel operation, and the regulation of the channel activity by endogenous low molecular weight compounds are analyzed. Special attention is given to changes that occur in muscle cells during exhausting work and to the role of sarcoplasmic reticulum Ca-release channels in the loss of muscle contractile activity during the development of fatigue. It is concluded that the protection of muscle fibers against fatigue in the presence of the histidine-containing dipeptide carnosine, called in the literature "Severin's phenomenon", is primarily connected with modulation of sarcoplasmic reticulum Ca-release channel activity by carnosine.

Histidine-containing dipeptides as endogenous regulators of the activity of sarcoplasmic reticulum Ca-release channels

It is shown that histidine-containing dipeptide carnosine ( β-alanyl- l-histidine), which is present in skeletal muscles in millimolar concentrations, decreases the rate of Ca 2+ accumulation by the heavy fraction of sarcoplasmic reticulum from rabbit skeletal muscles. This effect results from the ability of carnosine to induce a rapid Ca 2+ release from the heavy sarcoplasmic reticulum vesicles via activation of the ruthenium red-sensitive Ca-channels. The effect of carnosine is dose-dependent that indicates the presence of saturable site(s) for carnosine in the molecules of Ca-channels. The C 0.5 value for carnosine (the concentration that induces the half-maximal Ca 2+ release) is 8.7 mM. The 1 N-methylated derivative of carnosine, i.e., anserine, also induces a rapid Ca 2+ release with the half-maximal effect at 2.7 mM. Conversely, neither histidine nor β-alanine (both separately and in the mixture) cause Ca 2+ release. In addition, carnosine increases the sensitivity of Ca-channels to their well-known activators (caffeine, AMP, and Ca 2+) and decreases inhibitory effect of low concentrations of Mg 2+. It is concluded that carnosine as a component of skeletal muscles can be an endogenous regulator of the sarcoplasmic reticulum Ca-channel activity.

Effect of carnosine on the activity of lipoxygenase isolated from rabbit reticulocytes

The effects of the histidine-containing dipeptide carnosine and of histidine on the activity of 15-lipoxygenase from rabbit reticulocytes are studied. It is shown that low concentrations of carnosine (0.5–4 mM) activate oxidation of arachidonic acid by lipoxygenase, while high concentrations (>4 mM) inhibit the enzyme activity. Histidine elicits no activatory effect, although its inhibitory activity is comparable to that of carnosine.

Natural histidine-containing dipeptide carnosine as a potent hydrophilic antioxidant with membrane stabilizing function. A biomedical aspect.

A review on the distribution and biological effects of carnosine and a hypothesis for its biological mechanisms of action are presented. Carnosine and its structural and functional relative, anserine, were found in skeletal muscles at the beginning of the century. Their effects on muscle-working capacity, on the stability of membrane-bound enzymes, as well as their potent immunomodulating property, could not be explained by their pH-buffering capacity or formation of the secondary metabolites histidine and beta-alanine alone. This article suggests that the basis for the biological activities of carnosine and relative compounds is their potent antioxidant and membrane-protecting activity. The plausible chemical mechanism of this activity is discussed, and data regarding the usage of carnosine as a drug for treatment of immunodeficiency are summarized.

Detection of pork in processed meal: experimental comparison of methodology: W.N. Sawaya, T. Saeed, M. Mameesh, E. El-Rayes, A. Husain, S. Ali and H. Abdul Rahman, Food Chem. 37, 201–219

It is shown that histidine-containing dipeptide carnosine (β-alanyl-l-histidine), which is present in skeletal muscles in millimolar concentrations, decreases the rate of Ca2+ accumulation by the heavy fraction of sarcoplasmic reticulum from rabbit skeletal muscles. This effect results from the ability of carnosine to induce a rapid Ca2+ release from the heavy sarcoplasmic reticulum vesicles via activation of the ruthenium red-sensitive Ca-channels. The effect of carnosine is dose-dependent that indicates the presence of saturable site(s) for carnosine in the molecules of Ca-channels. The C0.5 value for carnosine (the concentration that induces the half-maximal Ca2+ release) is 8.7 mM. The 1N-methylated derivative of carnosine, i.e., anserine, also induces a rapid Ca2+ release with the half-maximal effect at 2.7 mM. Conversely, neither histidine nor β-alanine (both separately and in the mixture) cause Ca2+ release. In addition, carnosine increases the sensitivity of Ca-channels to their well-known activators (caffeine, AMP, and Ca2+) and decreases inhibitory effect of low concentrations of Mg2+. It is concluded that carnosine as a component of skeletal muscles can be an endogenous regulator of the sarcoplasmic reticulum Ca-channel activity.