Gamma-Interferon-Induced Nitric-Oxide Production Reduces Chlamydia-Trachomatis Infectivity in McCoy Cells

McCoy cells, murine-derived cells commonly used for propagation of chlamydiae, were found to be efficient producers of nitric oxide (NO) when primed with murine gamma interferon (IFN-gamma) and then exposed to the second signals provided by Escherichia coli lipopolysaccharide, human interleukin-1 alpha, murine tumor necrosis factor alpha, or Chlamydia trachomatis type H. Murine recombinant IFN-gamma over a range of 0 to 50 U/ml inhibited infectivity of C. trachomatis type H in a dose-dependent fashion in McCoy cells while simultaneously inducing NO production. Quantitation of infectious chlamydia progeny remaining in McCoy cells 48 or 72 h postinfection revealed that IFN-gamma-primed McCoy cells reduced chlamydial inclusion-forming units (expressed as units per milliliter) by 4 log10 units at higher IFN-gamma concentrations (50 U/ml) compared with control values. The magnitude of this antichlamydial effect was directly related to increased synthesis of NO, the production of which was IFN-gamma dose dependent. The antichlamydial effects of IFN-gamma were blocked in a dose-dependent manner by the addition of N-guanidino-monomethyl L-arginine (MLA), an inhibitor of nitric oxide synthesis. These results suggest that although IFN-gamma priming of McCoy cells is required for antichlamydial activity, nitric oxide is a necessary effector molecule involved in the mechanism(s) of IFN-gamma-induced inhibition of chlamydial proliferation in this murine cell line. The ability to block the potent antichlamydial effects of IFN-gamma by inhibition of a specific enzyme, nitric oxide synthase, may give insights into mechanisms by which IFN-gamma and perhaps other cytokines are able to control proliferation of chlamydiae and other intracellular pathogens

Complex coordinated extracellular metabolism: Acid phosphatases activate diluted human leukocyte proteins to generate energy flow as NADPH from purine nucleotide ribose

Complex metabolism is thought to occur exclusively in the crowded intracellular environment. Here we report that diluted enzymes from lysed human leukocytes produce extracellular energy. Our findings involve two pathways: the purine nucleotide catabolic pathway and the pentose phosphate pathway, which function together to generate energy as NADPH. Glucose6P fuel for NADPH production is generated from structural ribose of purine ribonucleoside monophosphates, ADP, and ADP-ribose. NADPH drives glutathione reductase to reduce an oxidized glutathione disulfide-glutathione redox couple. Acid phosphatases initiate ribose5P salvage from purine ribonucleoside monophosphates, and transaldolase controls the direction of carbon chain flow through the nonoxidative branch of the pentose phosphate pathway. These metabolic control points are regulated by pH. Biologically, this energy conserving metabolism could function in perturbed extracellular spaces