Uptake and distribution of haematoporphyrin derivative in the unicellular eukaryote Paramecium

Haematoporphyrin derivative (HpD) uptake, intracellular distribution and turnover were examined in a free-living protozoan cell, Paramecium aurelia, which had been demonstrated to internalize lipoproteins. A 10 min incubation in HpD completed with low-density lipoprotein (LDL) resulted in the appearance of distinct red-fluorescing vesicles, together with a diffuse fluorescence of the cytoplasm. Fluorescein labelling of LDL demonstrated the intracellular localization of HpD and LDL within the same vesicles. Pretreatment of Paramecium with the β-adrenergic antagonist l-propranolol, which blocked its phagocytotic activity, resulted in an absence of red-fluorescing vesicles; thus these were proved to be endosomes. Fluorescence emission recorded in the endosomes was characterized by a band at about 660-670 nm which was attributed to the partially unfolded oligomers; this emission was present during maintenance of the cells in drug-free culture medium for up to 120 min. Propranolol-pretreated cells exhibited only a diffuse cytoplasmic fluorescence characterized by an emission band at 630 nm, which was attributed to the monomers; this disappeared rapidly on washing. These results suggest the following: (i) HpD monomers enter Paramecium via transmembrane influx and/or fluid phase uptake; (ii) HpD oligomers are mainly internalized via receptor-mediated endocytosis; (iii) the extent of the endocytotic process is increased when HpD is completed with LDL; (iv) after internalization, aggregate species undergo a disaggregating process which accounts for the persistence of the intracellular fluorescence.