Katja S. Salmela

Characteristics of Helicobacter pylori alcohol dehydrogenase

BACKGROUND Helicobacter pylori shows alcohol dehydrogenase activity, which in the presence of ethanol leads to in vitro production of acetaldehyde, a toxic and highly reactive substance. The present study was undertaken to further define H. pylori-related ethanol and acetaldehyde metabolism by characterizing H. pylori alcohol dehydrogenase and by determining whether the organism possesses aldehyde dehydrogenase. METHODS Cytosolic alcohol and aldehyde dehydrogenase activities were determined spectrophotometrically. Acetaldehyde produced by cytosol during incubation with ethanol was measured by head space gas chromatography. Isoenzyme pattern was studied using isoelectric focusing. RESULTS Significant alcohol dehydrogenase activity was observed at a neutral pH known to occur in gastric mucus. The Km for ethanol oxidation was approximately 100 mmol/L for the two strains tested. Acetaldehyde was formed already from a low ethanol concentration known to prevail in the stomach endogenously. Isoelectric focusing of the enzyme showed activity bands with pI at 7.1-7.3, a pattern different from that of gastric mucosal alcohol dehydrogenase. 4-methylpyrazole inhibited enzyme activity in a competitive manner and suppressed the growth of the organism during culture. Neither Helicobacter strain studied showed aldehyde dehydrogenase activity and can thus not remove acetaldehyde by that pathway. CONCLUSIONS Acetaldehyde production by H. pylori from exogenous or endogenous ethanol may be a pathogenetic mechanism behind mucosal injury associated with the organism.

Alcohol dehydrogenase mediated acetaldehyde production by Helicobacter pylori — a possible mechanism behind gastric injury

Two standard Helicobacter pylori strains showed significant cytosolic alcohol dehydrogenase activity and produced considerable amounts of acetaldehyde when incubated with an ethanol containing solution in vitro. The alcohol dehydrogenase activity of the Helicobacter pylori strains was almost as high as that found in Klebsiella pneumoniae and far greater than that in Escherichia coli or Campylobacter jejuni. The amount of acetaldehyde produced by cytosol prepared from Helicobacter pylori exceeded that by any of the other bacteria studied. The bacterial production of acetaldehyde--a highly toxic and reactive substance--could be an important factor in the pathogenesis of Helicobacter pylori associated gastric injury and increased risk of gastric cancer.

Acetaldehyde and Ethanol Production by Helicobacter pylori

By virtue of possessing alcohol dehydrogenase activity, cytosol prepared from Helicobacter pylori produces toxic acetaldehyde from ethanol in vitro. To approach the in vivo situation in the stomach, we have now investigation whether intact H. pylori--without addition of exogenous nicotinamide adenine dinucleotide--also forms acetaldehyde. Furthermore, to assess the energy metabolism of H. pylori, we determined whether the alcohol dehydrogenase-catalyzed reaction can run in the opposite direction with ethanol as the end-product and thereby yield energy for the organism. Intact H. pylori formed acetaldehyde already at low ethanol concentrations (at 0.5% ethanol, acetaldehyde, 64 +/- 21 and 75 +/- 9 mumol/l (mean +/- SEM) for strains NCTC 11637 and NCTC 11638, respectively). H. pylori produced ethanol in concentrations that can be significant for the energy metabolism of the organism. Acetaldehyde production by H. pylori may be an important factor in the pathogenesis of gastroduodenal diseases associated with the organism. The primary function of H. pylori alcohol dehydrogenase may, however, be alcoholic fermentation and consequent energy production under microaerobic conditions.

Alcohol Metabolism in Helicobacter pylori-infected Stomach

Studies in our laboratory have revealed that Helicobacter pylori exhibits significant cytosolic alcohol dehydrogenase activity and that the enzyme is fully active at ethanol concentrations prevailing in the stomach during alcohol consumption or after alcohol is completely absorbed from the stomach and is available through blood circulation only. Moreover, even the low levels of endogenous ethanol found in the stomach can be oxidized to acetaldehyde by H. pylori alcohol dehydrogenase. The metabolic significance of the enzyme remains as yet unresolved. Under microaerobic conditions, however, the enzyme could be of importance in the energy metabolism of the organism. In the presence of excess ethanol, H. pylori alcohol dehydrogenase produces significant amounts of acetaldehyde. Acetaldehyde is a toxic and reactive compound and could theoretically be a pathogenetic factor in H. pylori-associated gastric injury. Preliminary studies have indicated that acetaldehyde inhibits gastric mucosal regeneration and forms stable adducts with mucosal proteins. Both of these mechanisms could cause gastric injury. The role of H. pylori-related acetaldehyde formation in vivo, however, needs to be established in future studies. In antral human gastric mucosa, H. pylori infection is associated with a significant decrease in alcohol dehydrogenase activity. Similarly, in specific pathogen-free mice with a prolonged infection, gastric alcohol dehydrogenase activity is decreased; however, this is not clearly reflected in the bioavailability of ethanol or the amount of its first pass metabolism.

Colloidal bismuth subcitrate and omeprazole inhibit alcohol dehydrogenase mediated acetaldehyde production by Helicobacter pylori

We have recently shown that Helicobacter pylori possesses marked alcohol dehydrogenase (ADH) activity and is capable--when incubated with an ethanol containing solution in vitro--of producing large amounts of acetaldehyde. In the present study we report that some drugs commonly used for the eradication of H. pylori and for the treatment of gastroduodenal diseases are potent ADH inhibitors and, consequently, effectively prevent bacterial oxidation of ethanol to acetaldehyde. Colloidal bismuth subcitrate (CBS), already at a concentration of 0.01 mM, inhibited H. pylori ADH by 93% at 0.5 M ethanol and decreased oxidation of 22 mM ethanol to acetaldehyde to 82% of control. At concentrations above 5 mM, CBS almost totally inhibited acetaldehyde formation. Omeprazole, a drug also known to suppress growth of H. pylori, also inhibited H. pylori ADH and suppressed bacterial acetaldehyde formation significantly to 69% of control at a drug concentration of 0.1 mM. By contrast, the H2-receptor antagonists ranitidine and famotidine showed only modest effect on bacterial ADH and acetaldehyde production. We suggest that inhibition of bacterial ADH and a consequent suppression of acetaldehyde production from endogenous or exogenous ethanol may be a novel mechanism by which CBS and omeprazole exert their effect both on the growth of H. pylori as well as on H. pylori associated gastric injury.

Binding of acetaldehyde to rat gastric mucosa during ethanol oxidation

Acetaldehyde, the first product of ethanol metabolism, has previously been shown to form potentially harmful adducts with various proteins. The aim of this study was to investigate whether acetaldehyde--either exogenous or metabolically derived--binds to gastric mucosal proteins. Homogenized rat gastric mucosa was incubated with various concentrations of radiolabeled acetaldehyde or ethanol for different time periods. Acetaldehyde-protein adducts were determined by a liquid scintillation counter. In addition, mucosa was incubated with nonlabeled ethanol, and the acetaldehyde formed was measured by using headspace gas chromatography. Incubation of gastric mucosa with (14C)-acetaldehyde led to a concentration- and time-dependent radiolabeling of mucosal proteins. Formation of acetaldehyde adducts occurred relatively rapidly within 30 minutes and even at low acetaldehyde levels (5 micromol/L). Stable adducts represented 77% +/- 5% (mean +/- SEM) of the total adducts formed. In the presence of ethanol, acetaldehyde production and adduct formation took place in a concentration- and time-dependent manner. 4-Methylpyrazole and sodium azide inhibited acetaldehyde production to 7% +/- 1% of control and decreased the amount of acetaldehyde adducts to 55% +/- 8%. Enhanced acetaldehyde formation (to 420% +/- 50%) was clearly reflected in increased adduct formation (550% +/- 110%). In conclusion, both exogenous and endogenous acetaldehyde binds to gastric mucosal proteins in vitro. Gastric mucosal acetaldehyde production and the consequent adduct formation could be a pathogenetic factor behind ethanol-associated gastric injury.

Effect of Bismuth and Nitecapone on Acetaldehyde Production by Helicobacter pylori

BACKGROUND We have recently shown that colloidal bismuth subcitrate inhibits cytosolic alcohol dehydrogenase of Helicobacter pylori as well as acetaldehyde production from excess ethanol. We now extend our studies to bismuth subsalicylate and nitecapone, a novel antiulcer agent. METHODS Cytosol of H. pylori was incubated with 0.1% or 1% ethanol in the presence of different drug concentrations for 2 h, whereafter acetaldehyde formed was analyzed by head space gas chromatography. In addition, we incubated a culture solution containing intact bacteria with the drugs at 1% ethanol. RESULTS Bismuth subsalicylate and nitecapone inhibit acetaldehyde formation from 0.1% ethanol by H. pylori cytosol at drug concentrations theoretically achievable in the stomach after intake of therapeutic doses of these drugs. Furthermore, colloidal bismuth subcitrate, bismuth subsalicylate, and nitecapone also inhibit acetaldehyde production by intact H. pylori, although rather high drug concentrations are required for this to occur. CONCLUSIONS Inhibition of H. pylori acetaldehyde formation may be one of the mechanisms by which bismuth and nitecapone exert their effect in the treatment of H. pylori-related disorders.

Formation of tetrahydroharman (1-methyl-1,2,3,4-tetrahydro-beta-carboline) by Heucobacter pylori in the presence of ethanol and tryptamine

Helicobacter pylori contains alcohol dehydrogenase which oxidizes ethanol to acetaldehyde. In the present study, H. pylori cytosol was incubated in a buffered media at pH 6.0 and 7.4 in the presence of ethanol and tryptamine. Under these conditions, tetrahydroharman (1-methyl-tetrahydro-beta-carboline) was produced as a condensation product of tryptamine and acetaldehyde. At pH 6.0, 20.60 +/- 5.00% of the added tryptamine was converted to tetrahydroharman, while 27.00 +/- 4.80% (mean +/-SD) was converted at pH 7.4. Similar reactions between acetaldehyde and other dietary amines seem likely. Such biogenic alkaloids, if formed in vivo, might contribute to the dysphoric effects of alcohol.

Elevated levels of serum dolichol in aspartylglucosaminuria.

Slightly elevated serum dolichol levels have so far been demonstrated only in alcoholics. We now report two diseases with exceptionally high serum dolichol levels. They are autosomal, recessively inherited lysosomal storage diseases, aspartylglucosaminuria (AGU) and mannosidosis. In 16 patients with AGU the mean serum level of total dolichols (457 +/- 43 ng/ml) was more than two-fold when compared to healthy controls (170 +/- 4 ng/ml). In two patients with mannosidosis the levels were almost two-fold. The percentage distribution of the dolichol homologues with 18, 19 or 20 isoprene units did not differ between the patients and controls. The inclusion of an additional control group excluded the possible influence of mental retardation and imparied moving ability on the results. Elevated serum dolichols in patients with lysosomal storage diseases may reflect a disturbance in lysosomal function and serve as a diagnostic marker. The biochemical mechanisms leading to this phenomenon remain to be established.

ABO and D typing and alloantibody screening in marrow samples: relevance to intraosseous blood transfusion: BLOOD GROUP SEROLOGY IN BM SAMPLES

Blood transfusion through the intraosseous route is gaining popularity in emergency medicine. Pretransfusion peripheral blood (PB) samples are usually not available in these patients, leading to discrepancies in blood group typing and a possible delay in transferring to group‐specific blood products. The aim of this study was to assess the feasibility of ABO and D typing and red blood cell alloantibody screening in marrow (BM) samples.