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Michael Kirsch

Publications and source records attributed to Michael Kirsch.

At least 19 recordsLinked to original sources
PubMed2006

On the mechanism of the ascorbic acid-induced release of nitric oxide from N-nitrosated tryptophan derivatives: scavenging of NO by ascorbyl radicals.

During the past years, there has been increasing interest in endogenous nitric oxide storage compounds. Recently, we briefly reported on the ascorbate-dependent release of nitric oxide ((.)NO) from N-nitrosotryptophan derivatives. In the present study, the underlying mechanism of (.)NO release is studied in more detail, primarily utilizing N-acetyl-N-nitrosotryptophan (NANT) as a model compound. The initial rate of the ascorbate-induced release of nitric oxide has been found to correspond to the rate of NANT decay. In this process, N-acetyltryptophan (NAT) is produced almost quantitatively. The final yield of nitrite amounted to around 90 % with respect to the applied amount of NANT. However, the total release of nitric oxide was only 60 %, as determined by using an FNOCT-4(fluorescent nitric oxide cheletropic trap number 4) assay. Besides nitric oxide, a second volatile product, dinitrogen oxide (N(2)O), has been identified by using (15)N NMR spectrometry, strongly indicating the intermediacy of nitroxyl (HNO). The formation of intermediate ascorbyl radical anions during the NANT-ascorbate reaction has been monitored by using ESR spectrometry. Unexpectedly, it was found that the primary oxidized product of vitamin C, dehydroascorbic acid (DHA), efficiently consumes nitric oxide. Since ESR spectrometry further revealed that ascorbyl radical anions are also generated during the spontaneous decay of DHA, the DHA-nitric oxide reaction is related to recombination of (.)NO with the thus formed ascorbyl radical anions. A conclusively established mechanism of the NANT-ascorbate reaction is presented, with O-nitrosoascorbate as a key intermediate, as additionally supported by CBS-QB3 calculations. The present study suggests that vitamin C and its oxidation products can chemically counterbalance endogenous nitric oxide levels.

Ascorbic Acid↗
PubMed2006

GSK-3 mediates differentiation and activation of proinflammatory dendritic cells.

The key components of the intracellular molecular network required for the expression of a specific function of dendritic cells (DCs) are as yet undefined. Using an in vitro model of human monocyte-derived DC differentiation, this study investigates the role of glycogen synthase kinase 3 (GSK-3), a multifunctional enzyme critical for cellular differentiation, apoptosis, self-renewal, and motility, in this context. We demonstrate that GSK-3 (1) inhibits macrophage development during differentiation of DCs, (2) is constitutively active in immature DCs and suppresses spontaneous maturation, and (3) acquires a proinflammatory functional status mediating high levels of IL-12, IL-6, and TNF-alpha secretion, and partially inhibits IL-10 in the context of DC activation. In particular, GSK-3 enhances IL-12p35 mRNA expression and thus the production of the proinflammatory cytokine IL-12p70 by integrating the activities of other kinases priming GSK-3 targets and the inhibitory effects of Akt-1. GSK-3 may therefore act as a key integrator of activating and inhibitory pathways involved in proinflammatory DC differentiation and activation.

Cell Differentiation↗
PubMed2006

Adaptive functional differentiation of dendritic cells: integrating the network of extra- and intracellular signals.

Phenotypic maturation, cytokine secretion, and migration are distinct functional characteristics of dendritic cells (DCs). These functions are independently regulated by a number of extracellular variables, such as type, strength, and persistence of an array of soluble and membrane-bound mediators. Since the exact composition of these variables in response to infection may differ between individuals, the intracellular signaling pathways activated by these extracellular networks may more closely correlate with DC function and predict the course of adaptive immunity. We found that activation of p38 kinase (p38K), extracellular signal-related kinase 1/2 (ERK1/2), and phosphatidylcholine-specific phospholipase C (PC-PLC) enhanced cytokine secretion, whereas p38K, cyclic adenosine monophosphate (cAMP), and PC-PLC enhanced migration. In contrast, phosphatidylinositol 3-kinase (PI3K)/Akt-1 and cAMP inhibited cytokine secretion while ERK1/2 inhibited migration. Migration and cytokine secretion further differed in their sensitivity to inhibition over time. However, although DCs could be manipulated to express migration, cytokine secretion, or both, the level of activation or persistence of intracellular pathway signaling was not predictive. Our results suggest a modular organization of function. We hypothesize that the expression of specific DC functions integrates a large variety of activating and inhibitory variables, and is represented by the formation of a functional unit of molecular networks-the signal response module (SRM). The combined activities of these modules define the functional outcome of DC activation.

Cell Differentiation↗
PubMed2006

15N CIDNP investigations of the peroxynitric acid nitration of L-tyrosine and of related compounds.

Peroxynitric acid (O2NOOH) nitrates L-tyrosine and related compounds at pH 2-5. During reaction with O2(15)NOOH in the probe of a 15N NMR spectrometer, the NMR signals of the nitration products of L-tyrosine, N-acetyl-L-tyrosine, 4-fluorophenol and 4-methoxyphenylacetic acid appear in emission indicating a nitration via free radicals. Nuclear polarizations are built up in radical pairs [15NO2* , PhO*]F or [15NO2* , ArH*+]F formed by diffusive encounters of 15NO2 with phenoxyl-type radicals PhO or with aromatic radical cations ArH*+. Quantitative 15N CIDNP investigations with N-acetyl-L-tyrosine and 4-fluorophenol show that the radical-dependent nitration is the only reaction pathway. During the nitration reaction, the 15N NMR signal of 15NO3- also appears in emission. This is explained by singlet-triplet transitions in radical pairs [15NO2* , 15NO3*]S generated by electron transfer between O2(15)NOOH and H15NO2 formed as a reaction intermediate. During reaction of peroxynitric acid with ascorbic acid, 15N CIDNP is again observed in the 15N NMR signal of 15NO3- showing that ascorbic acid is oxidized by free radicals. In contrast to this, O2(15)NOOH reacts with glutathione and cysteine without the appearance of 15N CIDNP, indicating a direct oxidation without participation of free radicals.

Ascorbic Acid↗
PubMed2006

Non-oxygen-forming pathways of hydrogen peroxide degradation by bovine liver catalase at low hydrogen peroxide fluxes.

Heme catalases are considered to degrade two molecules of H(2)O(2) to two molecules of H(2)O and one molecule of O(2) employing the catalatic cycle. We here studied the catalytic behaviour of bovine liver catalase at low fluxes of H(2)O(2) (relative to catalase concentration), adjusted by H(2)O(2)-generating systems. At a ratio of a H(2)O(2) flux (given in microM/min(- 1)) to catalase concentration (given in microM) of 10 min(- 1) and above, H(2)O(2) degradation occurred via the catalatic cycle. At lower ratios, however, H(2)O(2) degradation proceeded with increasingly diminished production of O(2). At a ratio of 1 min(- 1), O(2) formation could no longer be observed, although the enzyme still degraded H(2)O(2). These results strongly suggest that at low physiological H(2)O(2) fluxes H(2)O(2) is preferentially metabolised reductively to H(2)O, without release of O(2). The pathways involved in the reductive metabolism of H(2)O(2) are presumably those previously reported as inactivation and reactivation pathways. They start from compound I and are operative at low and high H(2)O(2) fluxes but kinetically outcompete the reaction of compound I with H(2)O(2) at low H(2)O(2) production rates. In the absence of NADPH, the reducing equivalents for the reductive metabolism of H(2)O(2) are most likely provided by the protein moiety of the enzyme. In the presence of NADPH, they are at least in part provided by the coenzyme.

Animals↗
PubMed2006

Enhanced oxidation of NAD(P)H by oxidants in the presence of dehydrogenases but no evidence for a superoxide-propagated chain oxidation of the bound coenzymes.

Recently we demonstrated that lactate dehydrogenase (LDH)-bound NADH is oxidized by O2, H2O2, HNO2 and peroxynitrite predominantly via a chain radical mechanism which is propagated by superoxide. Here we studied both whether other dehydrogenases also increase their coenzymes' reactivity towards these oxidants and whether a chain radical mechanism is operating. Almost all dehydrogenases increased the oxidation of their physiological coenzymes by at least one of the oxidants. The oxidation of NADH or NADPH depended both on the binding dehydrogenase and the applied oxidant and in some cases the reactions were remarkably fast. The highest rate constant (k = 370 M-1 s-1) was found for the reaction of HNO2 with NADH bound to alcohol dehydrogenase. Regardless of the applied oxidant, superoxide dismutase failed to inhibit the oxidation of protein-bound NADH and NADPH. We therefore conclude that several dehydrogenases increase the oxidation of NADH and/or NADPH by the employed set of oxidants in bimolecular reactions, but, unlike LDH, do not mediate a O2*(-) dependent chain radical mechanism.

Animals↗
PubMed2006

Preoperative application of selective angiographic embolization in the treatment of focal nodular hyperplasia.

A focal nodular hyperplasia (FNH) rarely requires surgical intervention unless complications or clinical symptoms come into play. In 2 female patients, ages 35 and 44, with symptomatic tumors of that entity (10 and 14 cm, respectively, in size) we performed a laparoscopic liver resection. Directly before undergoing this operation they underwent a selective angiographic embolization of the arterial inflow for control of hemorrhage by application of polyvinyl alcohol particles to the afferent arteries. This procedure led to a complete interruption in arterial inflow and the subsequent laparoscopic resection itself could be carried out as planned with minimal blood loss as well an uncomplicated postoperative course. The patients demonstrated a high degree of subjective satisfaction with the procedure. A reduction in arterial inflow through selective embolization is a useful tool in the preparation for laparoscopic resection of large, well-vascularized liver tumors.

Adult↗
PubMed2006

Detection of N-nitrosomelatonin and other N-nitrosotryptophan derivatives by transnitrosation of APF and DAF-2.

S-nitrosothiols can be analyzed with some simple detection procedures and this fact strongly accelerated the understanding of the biological impact of S-nitrosothiols. Unfortunately, such simple analytic methods are presently missing for low molecular weight N-nitrosotryptophan derivatives like N-nitrosomelatonin (NOMela). Here we demonstrate that commercially available primary aromatic amines, i.e. aminophenylfluorescein (APF) and 4,5-diaminofluorescin (DAF-2), can be used for a quantitative determination of NOMela. Under optimized conditions (e.g. pH 11) of the assays, the lifetime of N-nitrosotryptophan derivatives is largely prolonged and the reactivity of S-nitrosothiols with aromatic amines can be safely ignored. The influence of reactive nitrogen oxide species like N2O3 is additionally limited at the alkaline pH and may be further decreased by working under hypoxic conditions. As a result of these optimal conditions, the APF assay has a detection limit for NOMela of about 25 nm but this assay fails to detect protein-bound N-nitrosotryptophan residues. The DAF-2 assay, however, might be used for a qualitative analysis of such residues. Due to the high efficacy of the APF assay it is safely demonstrated that in regard to peroxynitrite, N2O3 is about 50-fold more effective in nitrosating melatonin at physiological pH.

Aniline Compounds↗
PubMed2005

Catecholamine-induced release of nitric oxide from N-nitrosotryptophan derivatives: a non-enzymatic method for catecholamine oxidation.

In recent years, interest in the physiological functions of S-nitrosothiols has strongly increased owing to the potential of these compounds to release nitric oxide. In contrast, little is known about similar functions of N-nitrosated (N-terminal-blocked) tryptophan derivatives, which can be also formed at physiological pH. Utilizing N-acetyl-N-nitrosotryptophan (NANT) and N-nitrosomelatonin (NOMela) as model compounds, we have studied their reaction with catechol and catecholamines such as epinephrine and dopamine. In these reactions, NANT was quantitatively converted to N-acetyltryptophan (NAT), and nitric oxide was identified as a volatile product. During this process, ortho-semiquinone-type radical anions deriving from catechol and dopamine, were detected by ESR spectrometry. The catechol radical concentration was about eight times higher under normoxia than under hypoxia and a similar relationship was found for the decay rates of NANT under these conditions. An epinephrine-derived oxidation product, namely adrenochrome, but not a catechol-derived one, was identified. These observations strongly indicate that N-nitrosotryptophan derivatives transfer their nitroso-function to an oxygen atom of the catecholamines, and that the so-formed intermediary aryl nitrite may decompose homolytically with release of nitric oxide, in addition to a competing hydrolysis reaction to yield nitrite and the corresponding catechol. These conclusions were supported by quantum chemical calculations performed at the CBS-QB3 level of theory. Since nitric oxide is non-enzymatically released from N-nitrosotryptophan derivatives on reaction with catecholamines, there might be a possibility for the development of epinephrine-antagonizing drugs in illnesses like hypertension and pheochromocytoma.

Catecholamines↗
PubMed2005

Generation of peroxynitrite from reaction of N-acetyl-N-nitrosotryptophan with hydrogen peroxide over a wide range of pH values.

The novel reaction of N-acetyl-N-nitrosotryptophan (NANT) with hydrogen peroxide to yield peroxynitrite is demonstrated. Quantum chemical calculations performed at CBS-QB3 level of theory predicted that the reaction of N-nitrosoindole with both H(2)O(2) and its corresponding anion is thermodynamically feasible. At pH 13, the formation of peroxynitrite from the bimolecular reaction of NANT with H(2)O(2) is unequivocally demonstrated by (15)N NMR spectrometry. In order to prove the intermediacy of peroxynitrite from the NANT-H(2)O(2) system at neutral (7.4) and acidic pH (4.5), the characteristic pattern of CIDNP (chemically induced dynamic nuclear polarization) signals were recorded, i.e. enhanced absorption in the (15)N NMR signal of nitrate and emission in the (15)N NMR signal of nitrite. Most interestingly, the NANT-H(2)O(2) system nitrated N-acetyltyrosine at pH 4 via recombination of freely diffusing nitrogen dioxide and tyrosyl radicals, but nitration was negligible at pH 7.4. Since the combination between NANT and H(2)O(2) is slow, endogenous N-nitrosotryptophan residues cannot act as a "carrier" for peroxynitrite.

Hydrogen Peroxide↗
PubMed2005

Enzyme-independent nitric oxide formation during UVA challenge of human skin: characterization, molecular sources, and mechanisms.

Many of the local UV-induced responses including erythema and edema formation, inflammation, premature aging, and immune suppression can be influenced by nitric oxide synthase (NOS)-produced NO which is known to play a pivotal role in cutaneous physiology. Besides NOS-mediated NO production, UV radiation might trigger an enzyme-independent NO formation in human skin by a mechanism comprising the decomposition of photo-reactive nitrogen oxides. Therefore, we have examined the chemical-storage forms of potential NO-generating agents, the mechanisms and kinetics of their decomposition, and their biological relevance. In normal human skin specimens we find nitrite and S-nitrosothiols (RSNO) at concentrations 25- or 360-fold higher than those found in plasma of healthy volunteers. UVA irradiation of human skin leads to high-output formation of bioactive NO due to photo-decomposition of RSNO and nitrite which represents the primary basis for NO formation during UVA exposure. Interestingly, reduced thiols strongly augment photo-decomposition of nitrite and are essential for maximal NO release. The enzyme-independent NO formation found in human skin opens a completely new field in cutaneous physiology and will extend our understanding of mechanisms contributing to skin aging, inflammation, and cancerogenesis.

Animals↗
PubMed2005

Initiation of a superoxide-dependent chain oxidation of lactate dehydrogenase-bound NADH by oxidants of low and high reactivity.

In cells, NADH and NADPH are mainly bound to dehydrogenases such as lactate dehydrogenase (LDH). In cell-free systems, the binary LDH-NADH complex has been demonstrated to produce reactive oxygen species via a chain oxidation of NADH initiated and propagated by superoxide. We studied here whether this chain radical reaction can be initiated by oxidants other than LDH largely increased the oxidation of NADH (but not of NADPH) by O(2), H(2)O(2) and during the intermediacy of HNO(2). LDH also increased the oxidation of NADH by peroxynitrite. The increases in NADH oxidation were completely prevented by superoxide dismutase (SOD). In contrast, the nitrogen dioxide-dependent oxidation of NADH and NADPH was decreased by LDH in a SOD-independent manner. These experimental data strongly indicate that oxidation of LDH-bound NADH can be induced from reaction of either weak oxidants with LDH-bound NADH or of strong oxidants with free NADH thus yielding which is highly effective to propagate the chain. Our results underline the importance of SOD in terminating superoxide-dependent chain reactions in cells under oxidative stress.

Animals↗
PubMed2005

First insights into regiospecific transnitrosation reactions between tryptophan derivatives: melatonin as an effective target.

Melatonin, a derivative of the essential amino acid tryptophan, has been portrayed as a hormone, a tissue factor, an autocoid, a paracoid, and a vitamin with antioxidative capabilities. In the present study a novel reaction which cannot be attributed to any of these suggested features, i.e. the transfer of the nitroso-function from N-nitrosotryptophan derivatives to melatonin, is unequivocally demonstrated. In the lipophilic buffer dimethylsulfoxide reaction of N-acetyl-N-nitrosotryptophan (NANT) with melatonin was very slow (k = 1.5 x 10(-6)/m/s), but reversible as shown by 15N-NMR spectrometry. These measurements demonstrated also that the thermodynamical equilibrium lies on the side of N-nitrosomelatonin (NOMela). Quantum-chemical calculations performed with the third-generation density functional B97-2 additionally predicted that this is also the case in an aqueous environment. In fact, reaction of melatonin with either NANT or N-nitrosotryptophan located at the endothelin-1 fragment 16-21 yielded NOMela with a rate constant of 1.7 +/- 0.5/m/s as shown by capillary zone electrophoresis. Interestingly, the known reactive nitrogen oxide species scavenger, piperazine, did not inhibit the NANT-dependent nitrosation of melatonin, thus very strongly indicating a direct transnitrosation reaction. All of these capabilities are known from the reaction of S-nitrosothiols with thiolate anions and are believed to be highly important in the transport and targeting of nitric oxide.

Electrophoresis, Capillary↗
PubMed2005

Combined immunoradiotherapy induces long-term remission of CNS relapse of peripheral, diffuse, large-cell lymphoma after allogeneic stem cell transplantation: case study.

Relapse of peripheral non-Hodgkin's lymphoma (NHL) in the central nervous system commonly has a poor prognosis. Graft-versus-leukemia effects (GvL) contribute substantially to eradication of hematological malignancies after allogeneic stem cell transplantation. Few data are available describing GvL activity within the brain. We report the case of a man allografted for peripheral NHL. On day +83 after transplantation a CNS relapse of the lymphoma occurred. The brain was irradiated with 44 Gy, anti-CD20 antibodies were given, and the immunosuppression was withdrawn. Subsequently, limited-stage, chronic graft-versus-host disease occurred. The lymphoma regressed completely, and the patient has been in continuous complete remission for 30 months. The favorable course suggests substantial contribution of immunomodulation to excellent outcome.

Adult↗
PubMed2004

Formation of S-nitrosothiols from regiospecific reaction of thiols with N-nitrosotryptophan derivatives.

S-nitrosothiols transport nitric oxide in vivo, and so-called transnitrosation reactions (i.e. the transfer of the nitroso function from nitrosothiol to thiolate) are believed to be involved in this process. In the present study we examined the N-nitrosotryptophan derivative-dependent nitrosation of thiols, a hitherto ignored possibility for the formation of S-nitrosothiols. The corresponding products were identified by (15)N-NMR spectrometry. The fact that the reaction proceeded under hypoxic conditions as well as in non-aqueous solution strongly indicated the occurrence of a transnitrosation reaction. Interestingly, S-nitrosothiols could only very slowly transnitrosate N-terminal-blocked tryptophan derivatives like melatonin in non-aqueous solution but did not induce such a reaction in water. The indole moiety of the N-nitrosotryptophan derivatives was fully restituted during the reaction with thiols, as demonstrated by both capillary zone electrophoresis and fluorescence spectroscopy. A determination of the Arrhenius parameters demonstrated that the corresponding rate constants were comparable with the ones known for the transfer of the nitroso function from nitrosothiol to thiolate. Thus, N-nitrosotryptophan-dependent nitrosation of thiols may occur in vivo and might offer the possibility of developing a new class of vasodilative drugs.

Cysteine↗