Soybean lipoxygenase-mediated oxygenation of monounsaturated fatty acids to enones.
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Carbon Radioisotopes↗
Biomedical subjects
M J Novak
Publications and source records attributed to M J Novak.
Toxicology update: the cardiotoxicity of the oxidative stress metabolites of catecholamines (aminochromes).
This toxicology update reviews the oxidative stress metabolites of catecholamines, postulated to be the biochemical initiators of cardiotoxicity. A brief overview of catecholamine metabolism is provided with several noteworthy historical observations relating to the autoxidation and rearrangement of epinephrine. The basic chemical and physical properties of adrenochrome and adrenolutin are discussed. The autoxidative, enzymatic and cellular basis for the transformation of catecholamines to oxidative metabolites is reviewed. Mechanisms seeking to account for the observed cardiotoxic changes in isolated heart perfusion studies and in vivo models are described.
Catecholamines↗
In vitro effects of anionic sulfur compounds on the spectrophotometric properties of native DNA.
Several anionic sulfur compounds are recognized as efficacious pretreatments for sulfur mustard (HD) poisoning. Our intent was to see if pretreatment compounds had a direct effect on DNA, a site where HD damage is thought to occur. A modification of the method of Szinicz et al. (Arzneim.-Forsch. 1981; 31: 1,713-1,717) was used to analyze the UV/VIS spectrum (205-400 nm) (n = 6) of calf thymus DNA (10-15 x 10(3) kDa) in the absence or presence of increasing concentrations of sodium thiosulfate, sodium 2-aminoethanethiosulfonate (thiotaurine), sodium metabisulfite or sodium sulfate. All compounds produced concentration-dependent absorbance decreases primarily at 212 nm, but also at 259 nm, with the exception of sodium sulfate. For example, 8.36 x 10(-4) M sodium thiosulfate reduced the absorbance of DNA at 212 nm by >60%. The kinetics of sulfur compounds on native DNA need further study. We propose that these anionic sulfur compounds interact with DNA possibly by changing the topology of this macromolecule. Effects may be due to interactions of these sulfur compounds at higher concentrations with DNA, with resulting ligand-DNA supercoiling. This process could protect against HD intoxication, which is caused in part by the uncoiling of DNA.
Animals↗
Effects of taurine and thiosulfate on the spectrophotometric properties of RNA and DNA.
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Animals↗
Gamma interferon is not required for mucosal cytotoxic T-lymphocyte responses or heterosubtypic immunity to influenza A virus infection in mice.
Heterosubtypic immunity (HSI) is defined as cross-protection against influenza virus of a different serotype than the virus initially encountered and is thought to be mediated by influenza virus-specific cytotoxic T lymphocytes (CTL). Since gamma interferon (IFN-gamma) stimulates cytotoxic cells, including antigen-specific CTL which may control virus replication by secretion of antiviral cytokines such as tumor necrosis factor alpha and IFN-gamma, we have investigated the mechanism of HSI by analyzing the role of IFN-gamma for HSI in IFN-gamma gene-deleted (IFN-gamma(-/-)) mice. It has been reported that IFN-gamma is not required for recovery from primary infection with influenza virus but is important for HSI. Here, we conclusively show that IFN-gamma is not required for induction of secondary influenza virus-specific CTL responses in mediastinal lymph nodes and HSI to lethal influenza A virus infection. Although T helper 2 (Th2)-type cytokines were upregulated in the lungs of IFN-gamma(-/-) mice after virus challenge, either Th1- or Th2-biased responses could provide heterosubtypic protection. Furthermore, titers of serum-neutralizing and cross-reactive antibodies to conserved nucleoprotein in IFN-gamma(-/-) mice did not differ significantly from those in immunocompetent mice. These results indicate that lack of IFN-gamma does not impair cross-reactive virus-specific immune responses and HSI to lethal infection with influenza virus. Our findings provide new insight for the mechanisms of HSI and should be valuable in the development of protective mucosal vaccines against variant virus strains, such as influenza and human immunodeficiency virus.
Adaptation, Physiological↗
Diverse effects of neutrophil integrin occupation on respiratory burst activation.
Integrin occupation can alter the function of neutrophils (PMN), but the mechanism(s) involved is still unclear. This study demonstrated that the occupation of PMN integrins (especially those of the beta(3) subfamily) strongly enhances TNF stimulation of the respiratory burst but down-regulates that induced by PMA, fMLP, Con A, and serum treated zymosan. Treatment of PMN with genistein, staurosporine, and wortmannin, inhibitors of tyrosine kinases, protein kinase C, and phosphotidylinostol 3-kinase (PI 3-kinase) respectively, completely blocked the TNF-stimulated respiratory burst in PMN. Genistein and wortmannin enhanced the PMA-stimulated respiratory burst but only in cells adherent to RGD peptide. These findings suggest that PMN integrins (beta(3) subfamily) can generate signals that regulate the PMN agonist responses, probably through the activities of tyrosine kinases and PI 3-kinase.
Antibodies↗
Beta2 integrin-dependent phosphorylation of protein-tyrosine kinase Pyk2 stimulated by tumor necrosis factor alpha and fMLP in human neutrophils adherent to fibrinogen.
Tumor necrosis factor alpha and fMLP can activate a broad range of cellular functions in neutrophils adherent to biological surfaces. These functions are mediated by integrins and involve the activation of tyrosine kinases. Here, we report that Pyk2, a member of the focal adhesion kinase family, was present in human neutrophils and was rapidly phosphorylated and activated following tumor necrosis factor alpha and fMLP stimulation in an adhesion-dependent manner. Tyrosine phosphorylation of Pyk2 was attenuated by beta2 integrin blocking with specific antibodies. The tyrosine phosphorylation of Pyk2 was downstream of protein kinases Lyn, Syk and protein kinase C and cytoskeletal organization. The activation of Pyk2 may play a role in adhesion/cytoskeleton-associated neutrophils function.
CD18 Antigens↗
Poliovirus replicons encoding the B subunit of Helicobacter pylori urease elicit a Th1 associated immune response.
The development of a vaccine for Helicobacter pylori is a key strategy for reducing the worldwide prevalence of H. pylori infection. Although immunization with recombinant B subunit of H. pylori urease (ureB) has yielded promising results, for the most part, these studies relied on the use of strong adjuvant, cholera toxin, precluding the use in humans. Thus, the development of new vaccine strategies for H. pylori is essential. Previous studies from our laboratory have described a vaccine vector based on poliovirus in which foreign genes are substituted for the poliovirus capsid genes. The genomes encoding foreign proteins (replicons) are encapsidated into authentic poliovirions by providing the capsids in trans. To test the utility of replicons as a vaccine vector for H. pylori, a replicon was constructed which encodes ureB. Expression of ureB in cells from the replicon was demonstrated by metabolic labeling followed by immunoprecipitation with anti-urease antibodies. To investigate the immunogenicity of the replicons, mice containing the transgene for the receptor for poliovirus were immunized via the intramuscular route. Mice given three doses of replicons did not develop substantial antibodies to ureB as determined by Western blot analysis using lysates from H. pylori. In contrast, mice given two doses of replicon followed by a single injection of recombinant ureB developed serum antibodies to ureB which were predominately IgG2a. Splenic lymphocytes from mice immunized with replicons alone, or replicons plus recombinant ureB produced abundant interferon-gamma and no detectable interleukin-4 upon stimulation with recombinant ureB. These results establish that poliovirus replicons encoding H. pylori ureB are immunogenic and induce primarily a T helper 1 associated immune response.
Animals↗
Heterosubtypic immunity to lethal influenza A virus infection is associated with virus-specific CD8(+) cytotoxic T lymphocyte responses induced in mucosa-associated tissues.
Heterosubtypic immunity, defined as cross-reactive immune responses to influenza virus of a different serotype than the virus initially encountered, was investigated in association with virus-specific cytotoxic T lymphocyte (CTL) responses induced in systemic and mucosa-associated lymph nodes after immunization via different routes. Mice immunized by the pulmonary route with live nonpathogenic influenza virus, strain Udorn (H3N2), survived challenge with mouse-adapted pathogenic influenza virus, strain PR/8/34 (H1N1). These mice developed strong heterosubtypic CTL responses in spleen, cervical lymph nodes (CLN), and mediastinal lymph nodes (MLN). Alternately, only 20% of mice immunized intravenously, intraperitoneally, or intranasally survived the challenge; all of these developed CTL responses in spleen and CLN, but not in MLN. Direct correlation between short-term and long-term memory heterosubtypic CTL responses induced in MLN and host recovery after lethal infection indicates that these CTL responses may play an important role in heterosubtypic immunity. Furthermore, induction and maintenance of memory CTL in regional mucosa-associated lymphoid tissues are highly dependent on mucosal immunization. The results implicate the mechanism of heterosubtypic immunity and should be an important consideration in the development of protective mucosal vaccines against variant strains of influenza and HIV.
Adaptation, Biological↗
New minimal substrate structural requirements in the enzymatic peroxidation of alkenes with soybean lipoxygenase.
A carboxylic or a charged head group in fatty acid analogs is not an essential structural requirement for binding and catalysis in soybean lipoxygenase-1 catalyzed oxidations.
Acetates↗
Recombinant viruses as vectors for mucosal immunity.
The development and characterization of viral based vaccine vectors is extremely active research field. Much of this work has been facilitated by developments in molecular biology that allow work with large plasmid-based vectors, as well as the use of PCR. Several different vector systems are now available using RNA viruses and DNA viruses. Each vector system has its own strengths and weaknesses. Due to the differences and diversity between the viruses used as vectors, it is doubtful that a single system will be useful for all desired vaccines. However, the further development of existing, as well as potentially new systems, will provide a repertoire for vaccinologists to design the recombinant vaccine which will generate an optimal humoral and immune response for protection against infection or disease caused by pathogens that infect via mucosal surfaces.
Administration, Intranasal↗
Src-family kinase-p53/ Lyn p56 plays an important role in TNF-alpha-stimulated production of O2- by human neutrophils adherent to fibrinogen.
Stimulation of neutrophil function by TNFalpha is largely dependent on beta2 integrins. It has also been proposed that src-family kinases are involved in this process. However, the functions of src-like kinases in human neutrophils still remain to be determined. In the present study, we used the new src-family kinase specific inhibitor PP1 [4-Amino-5-(4-methylphenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine] to investigate the role src-kinases play in TNFalpha stimulation of neutrophil function. Our results demonstrated that, in neutrophils adherent to fibrinogen, PP1 inhibited TNFalpha-stimulated superoxide production and protein tyrosine phosphorylation in a dose-dependent manner. In in vitro kinase assays, PP1 profoundly inhibited the activation of p53/56lyn but not p59hck or p72syk. Only slight inhibition was found of p58c-fgr. These data indicate that p53/56lyn plays an important role in TNFalpha-mediated stimulation of PMN function.
Fibrinogen↗
Necrotizing ulcerative periodontitis.
In patients with no known systemic disease or immune dysfunction, necrotizing periodontitis (NUP) appears to share many of the clinical and etiologic characteristics of necrotizing ulcerative gingivitis (NUG) except that patients with NUP demonstrate loss of clinical attachment and alveolar bone at affected sites. In these patients, NUP may be a sequela of a single or multiple episodes of NUG or may be the result of the occurrence of necrotizing disease at a previously periodontitis-affected site. The existence of immune dysfunction may predispose patients to NUG and NUP, especially when associated with an infection of microorganisms frequently associated with periodontal disease such as Treponema and Selenomonas species, Fuscobacterium nucleatum, Prevotella intermedia, and Porphyromonas gingivalis. The role of immune dysfunction is exemplified by the occasionally aggressive nature of necrotic forms of periodontal disease seen in patients with HIV infection or malnutrition, both of which may impact host defenses. Clinical studies of HIV-infected patients have shown that patients with NUP are 20.8 times more likely to have CD4+ cell counts below 200 cells/mm3. However, these same studies have demonstrated that most patients with CD4+ cell counts below 200 cells/mm do not have NUP, suggesting that other factors, in addition to immunocompromisation, are involved. Further studies are needed to define the complex interactions between the microbial, or viral, etiology of necrotic lesions and the immunocompromised host. It is, therefore, recommended that NUG and NUP be classified together under the grouping of necrotizing periodontal diseases based on their clinical characteristics.
Disease Progression↗
In vitro oxidation of the hydrolysis product of sulfur mustard, 2,2'-thiobis-ethanol, by mammalian alcohol dehydrogenase.
The mechanism of vesication from sulfur mustard remains unknown in spite of 80 years of investigation. We recently reported sulfur mustard-related inhibition of one or more protein (serine/threonine) phosphatases in tissue cytosol in vitro, suggesting a mechanism common to other vesicants such as cantharidin and Lewisite. Our investigation showed that this inhibition was related to the concentration of 2,2'-thiobis-ethanol (thiodiglycol), the hydrolysis product of sulfur mustard, rather than to the concentration of mustard itself. Related work showed an increase in the rate of NAD (but not NADP) reduction upon the addition of thiodiglycol to mouse liver cytosol. This result provided evidence that metabolism beyond thiodiglycol may be contributing to protein phosphatase inhibition. This observation indicated that metabolism involving one or more dehydrogenases may be necessary to produce the ultimate inhibitor of the protein phosphatases. We report here that thiodiglycol is a substrate for horse liver alcohol dehydrogenase (Km = 3.68+/-0.45 mM and Vmax = 0.22 +/-0.01 micromol min(-1) mg protein(-1)) and for pyridine nucleotide-linked enzymes in mouse liver and human skin cytosol. The alcohol dehydrogenase-specific inhibitor 4-methylpyrazole inhibited the oxidation of thiodiglycol by the pure horse liver enzyme as well as by the enzymes in human skin and mouse liver cytosol, indicating that the activity in the tissue preparations is also alcohol dehydrogenase.
Alcohol Dehydrogenase↗
Influenza virus-infected epithelial cells present viral antigens to antigen-specific CD8+ cytotoxic T lymphocytes.
We have investigated the mechanisms involved in the clearance of viral infection at the epithelium level by analyzing the activity of influenza virus-specific cytotoxic T lymphocytes (CTL) against virus-infected CMT-93 intestinal epithelial cells. Epithelial cells infected with live influenza virus effectively present viral antigens and were lysed by both homotypic and heterotypic influenza virus-specific CD8+ T cells. These results shed new light on the control of viral infection through the elimination of virus-infected epithelial cells by virus-specific CTL and demonstrate that CMT-93 cells furnish an appropriate model for in vitro evaluation of CTL activity against virus-infected epithelial cells.
Animals↗
Early-onset periodontitis.
The group of periodontal diseases known as the early-onset periodontal diseases are defined by the age of onset of periodontal destruction, distribution of lesions, association of disease with specific microbial infections, and identification of characteristic alterations, in the host response. Significant progress has recently been made in our understanding of the etiology of juvenile and rapidly progressive periodontitis. Considerable evidence points to a familial pattern of disease; both localized and generalized forms of disease may be observed in the same family. The exact mode of inheritance remains unclear, and disease may be the result of a complex interplay between genetically determined alterations of the host response and a specific bacterial challenge. Both neutrophil function and immunoglobulin response are altered and appear to be characteristic of an immunologic hyperresponsiveness. Bacterial colonization by Actinobacillus actinomycetemcomitans (serotype b) and/or Porphyromonas gingivalis appears to be the primary initiator of disease. Evidence suggests that lesion distribution may be a function of the nature of the infecting agent and the characteristics of the immune response.
Adolescent↗
Functional properties of nonhuman primate antibody to Porphyromonas gingivalis.
The nonhuman primate (NHP) serves as a useful model for examining the host-parasite interactions in Porphyromonas gingivalis-associated periodontal disease. This study determined the influence of NHP sera on (i) the direct killing of P. gingivalis, (ii) P. gingivalis-induced superoxide anion (O2-) release from human polymorphonuclear leukocytes (PMNs), and (iii) the ability of PMNs to bind and phagocytize P. gingivalis. Three types of NHP sera were utilized: (i) normal or baseline sera; (ii) sera obtained after ligature-induced periodontitis; and (iii) sera obtained following active immunization with formalinized P. gingivalis. All assays were performed with or without the addition of human complement. Significantly more (P < 0.01) direct killing of P. gingivalis occurred with immunized sera and complement than with any of the other treatments. The sera from ligature-induced periodontitis NHPs had significantly less (P < 0.03) killing capacity than the baseline sera, which contained natural antibody produced to P. gingivalis colonization. Sera from immunized NHPs were used to opsonize P. gingivalis and caused significantly greater (P < 0.01) levels of O2- release from PMNs. Finally, the sera from immunized NHPs significantly enhanced (P < 0.009) the uptake of P. gingivalis by PMNs, although binding of the bacteria to PMNs was similar among all three serum types. Active immunization of NHPs with P. gingivalis elicited a functional antibody that enhanced direct killing, positively influenced the activation of PMNs, and enhanced the ability of PMNs to phagocytize P. gingivalis. Moreover, antibody produced as a sequela of progressing periodontitis appeared to lack these functions. A wide variability in functional capacity of the sera from individual NHPs, which may contribute to an individual's susceptibility to P. gingivalis-induced disease, was noted. This variability suggested that results from functional tests of serum antibody may aid in predicting host susceptibility to disease and response to therapy.
Adult↗
Structural and (patho)physiological diversity of PAF.
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Animals↗