Mechanism and Tissue Specificity of Nicotine-mediated Lung S-Adenosylmethionine Reduction

Abstract

We previously reported that chronic nicotine infusion blocks development of Pneumocystis pneumonia. This discovery developed from our work demonstrating the inability of this fungal pathogen to synthesize the critical metabolic intermediate S-adenosylmethionine and work by others showing nicotine to cause lung-specific reduction of S-adenosylmethionine in guinea pigs. We had found nicotine infusion to cause increased lung ornithine decarboxylase activity (rate-controlling enzyme of polyamine synthesis) and hypothesized that S-adenosylmethionine reduction is driven by up-regulated polyamine biosynthesis. Here we report a critical test of our hypothesis; inhibition of ornithine decarboxylase blocks the effect of nicotine on lung S-adenosylmethionine. Further support is provided by metabolite analyses showing nicotine to cause a strong diversion of S-adenosylmethionine toward polyamine synthesis and away from methylation reactions; these shifts are reversed by inhibition of ornithine decarboxylase. Because the nicotine effect on Pneumocystis is so striking, we considered the possibility of tissue specificity. Using laser capture microdissection, we collected samples of lung alveolar regions (site of infection) and respiratory epithelium for controls. We found nicotine to cause increased ornithine decarboxylase protein in alveolar regions but not airway epithelium; we conclude that tissue specificity likely contributes to the effect of nicotine on Pneumocystis pneumonia. Earlier we reported that the full effect of nicotine requires 3 weeks of treatment, and here we show recovery is symmetrical, also requiring 3 weeks after treatment cessation. Because this time frame is similar to pneumocyte turnover time, the shift in polyamine metabolism may occur as new pneumocytes are produced. (2) and in patients with PCP (3, 4). This led to the prediction that deliberate reduction of host AdoMet will reduce susceptibility to PCP. A means for testing this was suggested by data reported 20 years ago: parenteral nicotine administration to guinea pigs causes selective reduction of lung AdoMet (5). Encouraging circumstantial support came from a large clinical study of factors that might relate to rates of PCP relapse in people with AIDS. Smoking was included as one of the parameters, and although no explanation was offered, a significant negative correlation was reported between smoking and PCP relapse (6). We tested whether nicotine would reduce rat lung AdoMet, as reported for guinea pigs (5) and found the effect to be similar (6). Infusion of the R-(ϩ)-nicotine isomer caused a 15-fold reduction in lung AdoMet with no change in plasma or liver AdoMet. When nicotine was evaluated in a rat model of PCP, it was strongly protective (6). Compared with controls infused with saline, rats infused with 475 g of R-(ϩ)-nicotine kg Ϫ1 h Ϫ1 starting at the time of inoculation had 99.9% fewer Pneumocystis organisms in their lungs 21 days post-5inoculation. Pneumocystis is an opportunistic fungus causingUnderstanding the shifts in metabolic pa...

Five lipoxygenase hypomethylation mediates the homocysteine effect on Alzheimer’s phenotype

“…The powder was re-suspended in 100 μl of NKPD buffer (2.68 mM KCl, 1.47 mM KH2PO4, 51.10 mM Na2HPO4, 7.43 mM NaH2PO4, 62 mM NaCl, 1 mM EDTA, and 1 mM dithiothreitol) and sonicated at 40 watts and 70% duty cycle for about 2 min, then clarified by centrifugation at 10,000x g for 15 min. Measurements were performed by HPLC analysis using Waters AccQ.Fluor derivitizing reagents (Waters Corp., MA) as previously described3435. The final concentration of Hcy, SAH or SAM in the samples was always normalized by mg protein.…”

Homocysteine exacerbates β‐amyloid pathology, tau pathology, and cognitive deficit in a mouse model of Alzheimer disease with plaques and tangles

“…Measurements were performed by high-performance liquid chromatography (HPLC) analysis using Waters AccQ.Fluor derivitizing reagents (Waters Corporation, Milford, MA) as previously described. 18,19 For brain samples, the limit of detection is 0.02nmol, and linearity extends to 5,000nmol. All of the samples were analyzed in triplicate, and the amount of Hcy in the samples was normalized by milligrams of protein.…”

Laser microdissection of conifer stem tissues: Isolation and analysis of high quality RNA, terpene synthase enzyme activity and terpenoid metabolites from resin ducts and cambial zone tissue of white spruce (Picea glauca)

“…Protein, enzyme and metabolite analysis has been limited partly because amplification is not possible for these molecules. Microdissected tissues have been successfully analyzed using proteomics [20] and metabolomics techniques [21], but there are few reports (none in plants) of isolation of intact protein samples for enzyme assays [22,23]. LMD has recently been applied successfully for microchemical analysis of stone cells from Norway spruce ( P. abies ) stems [24], but laser-assisted microdissection has not been widely applied to woody plant tissues.…”