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Nitroglycerine has been used clinically in the treatment of angina for 130 years, yet important details on the mechanism of action, biotransformation, and the associated phenomenon of nitrate tolerance remain unanswered. The biological activity of organic nitrates can be said to be nitric oxide mimetic, leading to recent, exciting progress in realizing the therapeutic potential of nitrates. Unequivocally, nitroglycerine and most other organic nitrates, including NO-NSAIDs, do not behave as NO donors in the most fundamental action: in vitro activation of sGC to produce cGMP. The question as to whether the biological activity of nitrates results primarily or exclusively from NO donation will not be satisfactorily answered until the location, the apparatus, and the mechanism of reduction of nitrates to NO are defined. Similarly, the therapeutic potential of nitrates will not be unlocked until this knowledge is attained. Aspects of the therapeutic and biological activity of nitrates are reviewed in the context of the chemistry of nitrates and the elusive efficient 3e− reduction required to generate NO. [Copyright &y& Elsevier]

Post-hypoxic injuries in plants are primarily caused by bursts of reactive oxygen species and acetaldehyde. In agreement with previous studies, we found accumulations of acetaldehyde in rice during re-aeration following submergence. During re-aeration, acetaldehyde-oxidizing aldehyde dehydrogenase (ALDH) activity increased, thereby causing the acetaldehyde content to decrease in rice. Interestingly, re-aerated rice plants showed an intense mitochondrial ALDH2a protein induction, even though ALDH2a mRNA was submergence induced and declined upon re-aeration. This suggests that rice ALDH2a mRNA is accumulated in order to quickly metabolize acetaldehyde that is produced upon re-aeration. [Copyright &y& Elsevier]

Many mitochondrial matrix and inner-membrane proteins are synthesized in the cytosol as precursor proteins with an N-terminal presequence, and are imported into the mitochondria. Although no distinct sequence homology has been found among mitochondrial presequences, Tom20, a general import receptor in the outer mitohcondrial membrane, binds to presequences, and distinguishes mitochondrial proteins from non-mitochonrial proteins. The recently determined structure of the cytosolic domain of Tom20 (ΔTom20) in a complex with the presequence of rat aldehyde dehydrogenase (ALDH) showed that a short stretch of the presequence forms an amphiphilic helix, and its hydrophobic surface interacts with the hydrophobic-binding groove of Tom20. The following NMR analyses revealed a common five-residue pattern for Tom20 binding in five different presequences. To refine the common amino acid motif for the recognition by Tom20, we introduced a new peptide library approach in this study: we prepared a mixture of ALDH presequence variants, tethered these peptides to ΔTom20 in a competitive manner by an intermolecular disulfide bond, and determined the relative affinities by MALDI-TOF mass spectrometry. We successfully deduced a refined, common motif for the recognition by Tom20, and found that the segment consisting of residues 14–20 of the ALDH presequence was locally optimized in the sequence space, with respect to Tom20 binding. [Copyright &y& Elsevier]

Recent studies have suggested that mitochondrial aldehyde dehydrogenase (aldehyde:NAD(P)+ oxidoreductase, EC 1.2.1.3) (ALDH2) plays essential roles in pollen development in plants. Rice (Oryza sativa L.) ALDH2 is encoded by at least two ALDH2 genes, one of which (ALDH2a) was previously identified. In this study, to understand the roles of ALDH2 in rice, we isolated and characterized a cDNA clone encoding another rice ALDH2 (ALDH2b). An in vitro ALDH assay indicated that ALDH2b possesses an NAD+-linked activity for oxidation of acetaldehyde, glycolaldehyde and propionaldehyde. Northern blot and immunoblot analyses revealed that ALDH2b was constitutively present in all the organs examined, whereas ALDH2a was expressed in leaves of dark-grown seedlings and panicles. By RFLP linkage mapping, the ALDH2a and ALDH2b genes were mapped to the long arm of chromosome 2 and the short arm of chromosome 6, respectively. We suggest that the rice ALDH2a and ALDH2b genes are orthologues of maize mitochondrial ALDH genes, rf2b and rf2a, respectively. [Copyright &y& Elsevier]

Transient up-regulation of ALDH3A1, CYP1A1 and CYP1B1 transcription by transient exposure to aryl hydrocarbon receptor (AhR) ligands, e.g. 3-methylcholanthrene, is via transient transactivation of xenobiotic responsive elements (XRE) present in the 5′-upstream regions of these genes. Others have shown that AhR ligand-mediated induction of increased CYP1A1 levels in cultured human breast (adeno)carcinoma cell lines is apparently estrogen receptor (ER)-dependent, i.e. it was observed in ER+ cell lines but not in ER− cell lines, whereas AhR ligand-mediated induction of increased CYP1B1 levels is ER-independent, i.e. it was observed in both ER+ and ER− cell lines. The present investigation established that transient, AhR ligand/XRE-mediated induction of increased ALDH3A1 levels in human breast (adeno)carcinoma cell lines was, like that of CYP1A1 and unlike that of CYP1B1, apparently ER-dependent. Thus, transient exposure to 3-methylcholanthrene induced increased levels of ALDH3A1 in five cultured human breast (adeno)carcinoma cell lines that were documented as being ER+, viz., MCF-7/0, MCF-7/OAP, T-47D, ZR-75-1 and MDA-MB-468, but failed to induce increased levels of this enzyme in four cultured human breast (adeno)carcinoma cell lines that have been historically viewed as being ER−, viz., MDA-MB-231, SK-BR-3, HS-578-T and MDA-MB-435. Somewhat at odds with the foregoing, transient exposure to 3-methylcholanthrene also induced increased levels of ALDH3A1 and CYP1A1 in cultured, essentially ER−, human breast epithelial MCF-10A cells. These cells, like cultured human breast (adeno)carcinoma cells, are immortal, but unlike the latter, are not tumorigenic. Transient induction of increased ALDH3A1 levels can also be effected by agents that are not AhR ligands, viz., electrophiles such as catechol, and thus, cannot up-regulate ALDH3A1 transcription via transactivation of a 5′-upsteam region XRE. Rather, they are thought to up-regulate ALDH3A1 transcription via transient transactivation of an electrophile responsive element (EpRE) that is putatively also present in the 5′-upstream region of this gene. Electrophile-initiated/EpRE-mediated induction of increased ALDH3A1 levels was found to be ER-independent. Thus, catechol transiently induced increased levels of ALDH3A1 in the five ER+ human breast (adeno)carcinoma cell lines, the four ER− human breast (adeno)carcinoma cell lines, and the ER−, immortal but not tumorigenic, human breast epithelial cell line. [Copyright &y& Elsevier]

The first primary structure of a plant aminoaldehyde dehydrogenase (AMADH, EC 1.2.1.19) is reported. The enzyme of pea (Pisum sativum) seedlings subjected to our study oxidises ω-aminoaldehydes to the corresponding ω-amino acids. Although pea does not accumulate betaine aldehyde as a compatible osmolyte, the N-terminal sequence of a purified pea AMADH resembles those of plant betaine aldehyde dehydrogenases (BADHs). On the basis of an anticipated pea AMADH homology to these enzymes, degenerated oligonucleotide primers were designed and used for PCR amplification. Two cDNA fragments were obtained in initial 5′ RACE experiments. Subsequent 5′and 3′ RACE performed with specific non-degenerated primers provided two putative cDNAs of the plant BADH family. Both encoded protein sequences (AMADH1 and AMADH2) are highly homologous to those of plant BADHs. They show 81% identity and 92% in mutual alignment. As a deduced product of the first cDNA, AMADH1 completely matches the N-terminal sequence of pea AMADH analysed previously by Edman degradation. AMADH 2 represents a putative AMADH or BADH that has not yet been isolated and characterised. We also tried to identify essential amino acid residues of a purified pea AMADH by both determination of its dissociation constants and evaluation of inhibition effects of specific modification reagents. From our results, it is clear that there are Cys (pK = 8.0) and Glu/Asp residues at the active site participating in the catalysis. This is in accordance with the presence of the conserved Glu and Cys active site regions of plant BADHs in both AMADH1 and AMADH2. [Copyright &y& Elsevier]

Antiepileptic therapy with a broad spectrum drug felbamate (FBM) has been limited due to reports of hepatotoxicity and aplastic anemia associated with its use. It was proposed that a bioactivation of FBM leading to formation of α,β-unsaturated aldehyde, atropaldehyde (ATPAL) could be responsible for toxicities associated with the parent drug. Other members of this class of compounds, acrolein and 4-hydroxynonenal (HNE), are known for their reactivity and toxicity. It has been proposed that the bioactivation of FBM to ATPAL proceeds though a more stable cyclized product, 4-hydroxy-5-phenyltetrahydro-1,3-oxazin-2-one (CCMF) whose formation has been shown recently. Aldehyde dehydrogenase (ALDH) and glutathione transferase (GST) are detoxifying enzymes and targets for reactive aldehydes. This study examined effects of ATPAL and its precursor, CCMF on ALDH, GST and cell viability in liver, the target tissue for its metabolism and toxicity. A known toxin, HNE, which is also a substrate for ALDH and GST, was used for comparison. Interspecies difference in metabolism of FBM is well documented, therefore, human tissue was deemed most relevant and used for these studies. ATPAL inhibited ALDH and GST activities and led to a loss of hepatocyte viability. Several fold greater concentrations of CCMF were necessary to demonstrate a similar degree of ALDH inhibition or cytotoxicity as observed with ATPAL. This is consistent with CCMF requiring prior conversion to the more proximate toxin, ATPAL. GSH was shown to protect against ALDH inhibition by ATPAL. In this context, ALDH and GST are detoxifying pathways and their inhibition would lead to an accumulation of reactive species from FBM metabolism and/or metabolism of other endogenous or exogenous compounds and predisposing to or causing toxicity. Therefore, mechanisms of reactive aldehydes toxicity could include direct interaction with critical cellular macromolecules or indirect interference with cellular detoxification mechanisms. [Copyright &y& Elsevier]

In wheat, non-phosphorylating, NADP-dependent glyceraldehyde-3-phosphate dehydrogenase (GAPN) was found to be encoded by one gene giving rise to a single protein. However, Western blots revealed two different subunits of about 58 and 60 kDa in endosperm and shoots. The latter was attributed to in vivo phosphorylation of shoot GAPN. No modification occurred in leaves, where the enzyme is composed by a single 58 kDa polypeptide. GAPN partially purified from shoots and endosperm was dephosphorylated in vitro with alkaline phosphatase. Phosphorylated GAPN exhibited similar affinity for substrates but a lower Vmax compared to the non-phosphorylated enzyme. Results suggest that reversible phosphorylation of GAPN could regulate NADPH production in the cytosol of heterotrophic plant cells. [Copyright &y& Elsevier]

The inability to target cancer stem cells (CSC) may be a significant factor contributing to treatment failure. We have developed a strategy to target the CSC populations in melanoma and squamous cell carcinoma using CSC lysate-pulsed dendritic cells (DCs). The CSC-DC vaccine was administered in the adjuvant setting after localized radiation therapy of established tumors. Using mouse models we demonstrated that DCs pulsed with CSCs enriched by virtue of their expression of the CSC marker ALDH (termed CSC-DC) significantly inhibited tumor growth, reduced development of pulmonary metastases and prolonged survival. The effect was associated with downregulation of chemokine (C-C motif) receptors CCR7 and CCR10 in tumor cells and decreased expression of the chemokine (C-C motif) ligands CCL21, CCL27 and CCL28 in lung tissue. The CSC-DC vaccine significantly reduced ALDHhighCSC frequency in primary tumors. Direct targeting of CSCs was demonstrated by the specific binding of IgG produced by ALDHhighCSC-DC vaccine-primed B cells to ALDHhighCSCs, resulting in lysis of these target CSCs in the presence of complement. These data suggest that the CSC-DC vaccine approach may be useful in the adjuvant setting where local and systemic relapse are high after conventional treatment of cancers. [ABSTRACT FROM PUBLISHER]

Lung cancer remains a challenging disease. It is responsible for the high cancer mortality rates in the US and worldwide. Elucidation of the molecular mechanisms operative in lung cancer is an important first step in developing effective therapies. Accumulating evidence over the last 2 decades suggests a critical role for Signal Transducer and Activator of Transcription 3 (STAT3) as a point of convergence for various signaling pathways that are dysregulated in the disease. In this review, we discuss possible molecular mechanisms involving STAT3 in lung tumorigenesis based on recent literature. We consider possible roles of STAT3 in cancer cell proliferation and survival, in the tumor immune environment, and in epigenetic regulation and interaction of STAT3 with other transcription factors. We also discuss the potential role of STAT3 in tumor suppression, which complicates strategies of targeting STAT3 in cancer therapy. [ABSTRACT FROM PUBLISHER]