ABSTRACT
For the first time the immunonutritional role of pyruvate on neutrophils (PMN), free α-keto and amino acid profiles, important reactive oxygen species (ROS) produced [superoxide anion (O(2) (-)), hydrogen peroxide (H(2)O(2))] as well as released myeloperoxidase (MPO) acitivity has been investigated. Exogenous pyruvate significantly increased PMN pyruvate, α-ketoglutarate, asparagine, glutamine, aspartate, glutamate, arginine, citrulline, alanine, glycine and serine in a dose as well as duration of exposure dependent manner. Moreover, increases in O(2) (-) formation, H(2)O(2)-generation and MPO acitivity in parallel with intracellular pyruvate changes have also been detected. Regarding the interesting findings presented here we believe, that pyruvate fulfils considerably the criteria for a potent immunonutritional molecule in the regulation of the PMN dynamic α-keto and amino acid pools. Moreover it also plays an important role in parallel modulation of the granulocyte-dependent innate immune regulation. Although further research is necessary to clarify pyruvate's sole therapeutical role in critically ill patients' immunonutrition, the first scientific successes seem to be very promising.
Subject(s)
Granulocytes/metabolism , Neutrophils/metabolism , Pyruvic Acid , Adult , Granulocytes/drug effects , Granulocytes/immunology , Humans , Hydrogen Peroxide/metabolism , Immunomodulation , Ketoglutaric Acids/metabolism , Male , Middle Aged , Neutrophils/drug effects , Neutrophils/immunology , Nutritional Physiological Phenomena , Peroxidase/metabolism , Pyruvic Acid/metabolism , Pyruvic Acid/pharmacology , Reactive Oxygen Species/metabolism , Superoxides/metabolismABSTRACT
The aim of this study was to determine the effects of alpha-ketoglutarate on neutrophil (PMN), free alpha-keto and amino-acid profiles as well as important reactive oxygen species (ROS) produced [superoxide anion (O(2) (-)), hydrogen peroxide (H(2)O(2))] and released myeloperoxidase (MPO) activity. Exogenous alpha-ketoglutarate significantly increased PMN alpha-ketoglutarate, pyruvate, asparagine, glutamine, asparatate, glutamate, arginine, citrulline, alanine, glycine and serine in a dose as well as duration of exposure dependent manner. Additionally, in parallel with intracellular alpha-ketoglutarate changes, increases in O(2) (-) formation, H(2)O(2)-generation and MPO activity have also been observed. We therefore believe that alpha-ketoglutarate is important for affecting PMN "susceptible free amino- and alpha-keto acid pools" although important mechanisms and backgrounds are not yet completely explored. Moreover, our results also show very clearly that changes in intragranulocytic alpha-ketoglutarate levels are relevant metabolic determinants in PMN nutrition considerably influencing and modulating the magnitude and quality of the granulocytic host defense capability as well as production of ROS.
Subject(s)
Amino Acids/metabolism , Keto Acids/metabolism , Ketoglutaric Acids/pharmacology , Neutrophils/metabolism , Reactive Oxygen Species/metabolism , Adult , Cells, Cultured , Humans , Male , Neutrophils/drug effects , Young AdultABSTRACT
Procedures for the analysis of free alpha-keto acids in human fluids (i.e. plasma, cerebrospinal fluid, urine, etc.) as well as for studying the dynamic free alpha-keto acid pools in differentiated tissues and organ cells have been the subject of growing clinical interest in the study of metabolic regulatory and pathophysiological phenomena. Due to the high instability and polarity of the alpha-keto acids being examined, the development of a quantitative and reproducible analysis of metabolically relevant intracellular alpha-keto acids still presents a substantial methodological challenge. The aim of small sample size, rapid, non-damaging and "metabolism-neutral" cell isolation, careful sample preparation and stability, as well as reproducible analytics technology is not often achieved. Only few of the methods described can satisfy the rigorous demands for an ultra-sensitive, comprehensive and rapid intracellular alpha-keto acid analysis.
Subject(s)
Cell Fractionation/methods , Chromatography, High Pressure Liquid/methods , Keto Acids/analysis , Spectrometry, Fluorescence/methods , Animals , Chromatography, Gas , Humans , Keto Acids/chemistry , Keto Acids/metabolismABSTRACT
We examined the effects of beta-alanine (taurine analogue and taurine transport antagonist), taurine (regarding its role in neutrophil (PMN) immunonutrition) and taurine combined either with L-NAME (inhibitor of *NO-synthase), SNAP (*NO donor), DON (glutamine-analogue and inhibitor of glutamine-requiring enzymes), DFMO (inhibitor of ornithine-decarboxylase) and beta-alanine on neutrophil amino- and alpha-keto acid profiles or important PMN immune functions in order to establish whether taurine transport-, nitric oxide-, glutamine- or ornithine-dependent mechanisms are involved in any of the taurine-induced effects. According to the present findings, the taurine-mediated effect appears to be based primarily on a modulation of important transmembraneous transport mechanisms and only secondarily on directly or indirectly induced modifications in intragranulocytic amino- and alpha-keto acid homoeostasis or metabolism. Although a direct relation to the parallel observed immunological modifications can only be presumed, these results show very clearly that compositional modifications in the free intragranulocytic amino- and alpha keto-acid pools coinciding with changes in intragranulocytic taurine levels are relevant metabolic determinants that can significantly influence the magnitude and quality of the granulocytic immune response.
Subject(s)
Amino Acids/metabolism , Homeostasis/drug effects , Keto Acids/metabolism , Neutrophils/physiology , Taurine/physiology , beta-Alanine/pharmacology , Adult , Diazooxonorleucine/pharmacology , Eflornithine/pharmacology , Humans , Hydrogen Peroxide/metabolism , Male , NG-Nitroarginine Methyl Ester/pharmacology , Neutrophils/immunology , Peroxidase/metabolism , S-Nitroso-N-Acetylpenicillamine/pharmacology , Superoxides/metabolismABSTRACT
We examined the effects of DON [glutamine-analogue and inhibitor of glutamine-requiring enzymes], alanyl-glutamine (regarding its role in neutrophil immunonutrition) and alanyl-glutamine combined with L-NAME, SNAP, DON, beta-alanine and DFMO on neutrophil amino and alpha-keto acid concentrations or important neutrophil immune functions in order to establish whether an inhibitor of *NO-synthase [L-NAME], an *NO donor [SNAP], an analogue of taurine and a taurine transport antagonist [beta-alanine], an inhibitor of ornithine-decarboxylase [DFMO] as well as DON could influence any of the alanyl-glutamine-induced effects. In summary, irrespective of which pharmacological, metabolism-inhibiting or receptor-mediated mechanisms were involved, our results showed that impairment of granulocytic glutamine uptake, modulation of intracellular glutamine metabolisation and/or de novo synthesis as well as a blockade of important glutamine-dependent metabolic processes may led to significant modifications of physiological and immunological functions of the affected cells.
Subject(s)
Amino Acids/metabolism , Dipeptides/metabolism , Homeostasis , Immunocompetence/physiology , Keto Acids/metabolism , Neutrophils/metabolism , Signal Transduction/physiology , Adult , Amino Acids/chemistry , Antibiotics, Antineoplastic/metabolism , Diazooxonorleucine/metabolism , Eflornithine/metabolism , Enzyme Inhibitors/metabolism , Humans , Hydrogen Peroxide/metabolism , Keto Acids/chemistry , Male , NG-Nitroarginine Methyl Ester/metabolism , Neutrophils/chemistry , Neutrophils/cytology , Nitric Oxide Donors/metabolism , Oxidants/metabolism , Peroxidase/metabolism , S-Nitroso-N-Acetylpenicillamine/metabolism , Superoxides/metabolismABSTRACT
We have examined the effects of N(omega)-nitro-L-arginine-methylester-hydrochloride [L-NAME; inhibitor of nitric oxide synthase], S-nitroso-N-acetyl-penicillamine [SNAP; nitric oxide donor], alpha-difluoro-methyl-ornithine [DFMO; inhibitor of ornithine decarboxylase] arginine or ornithine as well as the combination of arginine or ornithine with L-NAME, SNAP or DFMO on intracellular free amino- and alpha-keto acid profiles and the immune function markers superoxide anion and hydrogen peroxide generation as well as released myeloperoxidase activity in neutrophils (PMN). Although the underlying mechanisms still remain unclear, we believe from our results that nitric oxide as well as polyamine-dependent pathways are involved in the signal transmission of free radical molecule, beneficial nutritional therapy or maleficient pharmacological stress-induced alterations in PMN nutrient composition. Relevant changes in intragranulocyte free amino- and alpha-keto acid homeostasis and metabolism, especially, may be one of the determinants in PMN nutrition that positively or negatively influences and modulate neutrophil host defence capability and immunocompetence.
Subject(s)
Amino Acids/metabolism , Keto Acids/metabolism , Neutrophils/metabolism , Nitric Oxide/metabolism , Polyamines/metabolism , Adult , Amino Acids/pharmacology , Eflornithine/pharmacology , Enzyme Activation/drug effects , Humans , Hydrogen Peroxide/metabolism , Male , NG-Nitroarginine Methyl Ester/pharmacology , Neutrophil Activation/drug effects , Neutrophils/drug effects , Penicillamine/analogs & derivatives , Penicillamine/pharmacology , Peroxidase/drug effects , Superoxides/metabolismABSTRACT
The management of tumor-related pain, particularly that which is neuropathic in origin, is both difficult and challenging. According to WHO guidelines, opioids are the first-line analgesics in case of severe tumor-related pain syndromes. Some patients receiving continuous opioid therapy rapidly require increase of the opioid dose. In this case it can be necessary to extend the analgesic regime and to either apply interventional procedures or change the opioid or the route of administration. The case presented is remarkable regarding the severity and persistent nature of the patient's pain despite a variety of commonly employed treatments.
Subject(s)
Analgesics, Opioid/therapeutic use , Morphine/therapeutic use , Neoplasms/physiopathology , Pain/drug therapy , Sufentanil/therapeutic use , Analgesics, Opioid/administration & dosage , Disease Progression , Drug Tolerance , Female , Humans , Injections, Spinal , Middle Aged , Morphine/administration & dosage , Sufentanil/supply & distributionABSTRACT
The objective of this study was to determine the dose as well as duration of exposure-dependent effects of L-alanyl-L-glutamine, arginine or taurine on polymorphonuclear neutrophil (PMN) free alpha-keto acid profiles and, in a parallel study, on PMN immune functions. Exogenous L-alanyl-L-glutamine significantly increased PMN alpha-ketoglutarate, pyruvate PMN superoxide anion (O2-) generation, hydrogen peroxide (H2O2) formation and released myeloperoxidase (MPO) activity. Arginine also led to significant increases in alpha-ketoglutarate, pyruvate, MPO release and H2O2 generation. Formation of O2- on the other hand was decreased by arginine. Incubation with taurine resulted in lower intracellular pyruvate and alpha-ketobutyrate levels, decreased O2- and H2O2 formation and a concomitant significantly increased MPO activity. We therefore believe that considerable changes in PMN free-alpha-keto-acid profiles, induced for example by L-alanyl-L-glutamine, arginine or taurine, may be one of the determinants in cell nutrition that considerably modulates the immunological competence of PMN.
Subject(s)
Arginine/pharmacology , Dipeptides/pharmacology , Keto Acids/immunology , Neutrophils/drug effects , Neutrophils/immunology , Taurine/pharmacology , Adult , Enzyme Activation/drug effects , Humans , Intracellular Fluid/metabolism , Keto Acids/chemistry , Male , Neutrophils/chemistry , Oxidation-Reduction , Peroxidase/drug effects , Time FactorsABSTRACT
We have examined the effects of midazolam, Ro 5-4864 (agonist for "peripheral" [p] benzodiazepine receptors [BR]), PK 11195 (antagonist for pBR), flumazenil (antagonist for "central" BR), naloxone (antagonist for opiate receptors) and the combination of midazolam and Ro 5-4864, PK 11195, flumazenil or naloxone on intracellular amino- and alpha-keto acids and the immune function markers superoxide anion (O(2)(-)), hydrogen peroxide (H(2)O(2)) and released myeloperoxidase (MPO) activity in neutrophils (PMN). Only midazolam and Ro 5-4864 led to significant changes in the dynamic PMN free amino- and alpha-keto acid pools. Concerning PMN immune function markers, midazolam and Ro 5-4864 significantly decreased O(2)(-) and H(2)O(2) formation and released MPO. When midazolam and Ro 5-4864 were applied together they appeared to act additively. Pre-incubation with PK 11195 partially neutralized the midazolam effects whereas flumazenil or naloxone showed no effects. We therefore believe that pBR are involved in the signal transmission of anesthetic-induced cellular metabolic changes in PMN.
Subject(s)
Amino Acids/metabolism , Keto Acids/metabolism , Neutrophils/immunology , Neutrophils/metabolism , Receptors, GABA-A/metabolism , Adult , Benzodiazepinones/pharmacology , Cells, Cultured , Flumazenil/pharmacology , GABA Modulators/pharmacology , GABA-A Receptor Agonists , GABA-A Receptor Antagonists , Humans , Hydrogen Peroxide/metabolism , Isoquinolines/pharmacology , Male , Midazolam/pharmacology , Naloxone/pharmacology , Neutrophils/drug effects , Peroxidase/drug effects , Peroxidase/metabolism , Receptors, GABA-A/drug effects , Superoxides/metabolismABSTRACT
The objective of this study was to determine the effects of ornithine on polymorphonuclear leucocyte (PMN) free amino- and alpha-keto acid profiles, superoxide anion (O2-) generation, hydrogen peroxide (H2O2) formation and released myeloperoxidase activity (MPO). Exogenous ornithine significantly increased PMN asparagine, glutamine, aspartate, glutamate, arginine, citrulline, alanine, alpha-ketoglutarate and pyruvate as intracellular ornithine increased. Concerning PMN immune function markers ornithine increased H2O2-generation and MPO activity while O2- -formation was decreased. We believe therefore that ornithine is important for affecting PMN "susceptible free amino- and alpha-keto acid pool" although the mechanisms are not yet clear. This may be one of the determinants in PMN nutrition considerably influencing and modulating PMN host defense capability.
Subject(s)
Amino Acids/blood , Keto Acids/blood , Neutrophils/immunology , Neutrophils/metabolism , Ornithine/pharmacology , Adult , Cells, Cultured , Dose-Response Relationship, Drug , Humans , Hydrogen Peroxide/blood , Male , Neutrophils/drug effects , Ornithine/blood , Oxidation-Reduction , Peroxidase/blood , Peroxidase/drug effects , Superoxides/bloodABSTRACT
An electrophoresis cell with scanning UV-absorption optics is presented. It allows the measurement of moving reaction boundaries of dilute protein solutions with a high-resolution. The protein profiles in the boundaries can be extrapolated to infinite time after an appropriate transformation of space and time coordinates and then evaluated with respect to association constants. This is demonstrated for the dimer-tetramer equilibrium of haemoglobin.