A non-classical type of alveolar macrophage response to Trichinella spiralis infection.

Studies of arginase expression and activity in guinea pig alveolar macrophages during Trichinella spiralis infection, prompted by earlier observation of innate lung response to the parasite, showed the macrophages to express both activity and protein of arginase type I. In cultured macrophages part of the enzyme was found to be always released to the extracellular medium. Whereas BCG in vivo treatment, alone or preceded by T. spiralis infection, stimulated arginase activity, T. spiralis infection alone affected the enzyme distribution between intracellular and extracellular fractions, and properties (K(m) and V(max)), rather than total (intracellular + extracellular) activity, with TGF-beta apparently responsible for a part of the effect. Anti-TGF-beta antibody treatment of the animals influenced both arginase activation by Mn(2+) and dependence of the enzyme-catalysed reaction on pH. Whereas T. spiralis infection activated guinea pig alveolar macrophages by the type II macrophage activation, as indicated by constant arginase expression, associated with previously demonstrated lack of stimulation of nitric oxide production, BCG treatment invoked an alternative type of activation mechanism, reflected by stimulation of macrophage arginase, but not iNOS, activity.

Trichinella spiralis thymidylate synthase: cDNA cloning and sequencing, and developmental pattern of mRNA expression.

The persistent expression of thymidylate synthase activity has previously been demonstrated not only in adult forms, but also in non-developing muscle larvae of Trichinella spiralis and T. pseudospiralis, pointing to an unusual pattern of cell cycle regulation, and prompting further studies on the developmental pattern of T. spiralis thymidylate synthase gene expression. The enzyme cDNA was cloned and sequenced, allowing the characterization of a single open reading frame of 307 amino acids coding for a putative protein of 35,582 Da molecular weight. The amino acid sequence of the parasite enzyme was analysed, the consensus phylogenetic tree built and its stability assessed. The aa sequence identity with thymidylate synthase was confirmed by the enzymatic activity of the recombinant protein expressed in E. coli. As compared with the enzyme purified from muscle larvae, it showed apparently similar Vmax value, but higher Km(app) values desscribing interactions with dUMP (28.8 microM vs. 3.9 microM) and (6RS,alphaS)-N(5,10)-methylenetetrahydrofolate (383 microM vs. 54.7 microM). With the coding region used as a probe, thymidylate synthase mRNA levels, relative to 18S rRNA, were found to be similar in muscle larvae, adult forms and newborn larvae, in agreement with muscle larvae cells being arrested in the cell cycle.