Studies on compounds promoting the in vitro transformation of Trypanosoma brucei from bloodstream to procyclic forms

In vitro differentiation of pleomorphic blood-stream forms of Trypanosoma brucei to procyclic culture forms occurred rapidly and at high rates at 27 degrees C in a culture medium containing 1 mM cis-aconitate as the transformation-inducing agent. Citrate was required at a much higher concentration (10 mM) to produce a similar transformation rate. The highest percentage of transformed cells was obtained when bloodstream-form trypanosomes were treated with pronase in the absence of a feeder-cell layer. However, under these conditions, the amount of procyclic forms obtained after 72 h was lower than that obtained in the presence of cis-aconitate. Trypsin was also capable of inducing transformation in the absence of a feeder-cell layer, but this treatment again resulted in low numbers of transformed cells. Blood-stream-form trypanosomes were incapable of taking up citrate to any significant extent and the citrate content of these stages was negligible. After 72 h of exposure to citrate (3 mM), intracellular levels of this compound remained very low (< 1 nmol (10(9) cells)-1), increasing in established procyclic stages to approximately 1.7 nmol (10(9) cells)-1. These observations suggest that the tricarboxylic acid (TCA)-cycle metabolite-dependent transformation may be initiated externally to the trypanosome cell membrane. The ability of both citrate and cis-aconitate to bind calcium and, thus, to reduce the concentration of this cation in the culture medium was found not to be responsible for the triggering effect on trypanosome transformation.(ABSTRACT TRUNCATED AT 250 WORDS)