Binding, internalization, and degradation of high density lipoprotein by cultured normal human fibroblasts

Recently, it was demonstrated that doxorubicin (Dox.HCl), a chemotherapeutic agent, could be photoactivated by Cerenkov radiation (CR). The objective of the present work was to develop a multimodal chemotherapy-radiotherapy-photodynamic therapeutic system based on reconstituted high-density lipoprotein (rHDL) loaded with Dox.HCl and 177Lu-DOTA. 177Lu acts as a therapeutic radionuclide and CR source. The system can be visualized by nuclear imaging. Fluorescence microscopy showed that rHDL-Dox specifically recognized cancer cells (T47D) that are positive for SR-B1 receptors. Encapsulated Dox.HCl was released into the cells and produced reactive oxygen species when irradiated with a 450-nm laser (photodynamic effect). The same effect occurred when Dox.HCl was irradiated by 177Lu CR. Through in vitro experiments, it was confirmed that the addition of 177Lu-DOTA to the rHDL-Dox nanosystem did not affect the specific recognition of SR-B1 receptors expressed in cells, or the cellular internalization of 177Lu-DOTA. The toxicity induced by the rHDL-Dox/177Lu nanosystem in cell lines with high (T47D and PC3), poor (H9C2) and almost-zero (human fibroblasts (FB)) expression of SR-B1 was evaluated in vitro and confirmed the synergy of the combined chemotherapy-radiotherapy-photodynamic therapeutic effect; this induced toxicity was proportional to the expression of the SR-B1 receptor on the surface of the cells used. The HDL-Dox/177Lu nanosystem experienced uptake by tumor cells and the liver-both tissues with high expression of SR-B1 receptors-but not by the heart. 177Lu CR offered the possibility of imparting photodynamic therapy where laser light could not reach.

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The main lytic factor of Trypanosoma brucei brucei in normal human serum is not high density lipoprotein.

Journal of Experimental Medicine ◽

10.1084/jem.183.3.1023 ◽

1996 ◽

Vol 183 (3) ◽

pp. 1023-1029 ◽

Cited By ~ 58

Author(s):

J Raper ◽

V Nussenzweig ◽

S Tomlinson

Keyword(s):

High Density Lipoprotein ◽

Human Serum ◽

Trypanosoma Brucei ◽

Gel Filtration ◽

Density Lipoprotein ◽

High Density ◽

Lytic Activity ◽

Normal Human Serum ◽

Trypanosoma Brucei Brucei ◽

Normal Human

Natural immunity of humans to the cattle pathogen Trypanosoma brucei brucei has been attributed to the presence in normal human serum (NHS) of lytic factors for the parasites. We and others have shown that NHS contains two trypanolytic factors (herein termed TLF1 and TLF2) that can be separated by gel filtration. TLF1 copurifies with a subclass of high density lipoprotein (HDL), whereas TLF2 has a much higher molecular weight and does not appear to be a lipoprotein. We find that the trypanolytic activity of purified TLF1 is totally inhibited by exogenous haptoglobin (Hp) at concentrations (0.1 mg/ml) lower than those present in NHS (0.2-2 mg/ml). In contrast, exogenous Hp (up to 2.5 mg/ml) has no effect on the lytic activity of either NHS or isolated TLF2. Hp-depleted sera from patients with intravascular hemolysis is severalfold more trypanolytic than NHS. These sera contain only TLF1, and their lytic activity is totally abolished upon the addition of Hp (0.1 mg/ml). When NHS containing different Hp allotypes is fractionated by gel filtration, TLF1 activity is either revealed or remains masked, depending on whether it coelutes with Hp. Masked TLF1 activity in the column fractions is revealed if Hp is removed by density gradient ultracentrifugation. We conclude that endogenous Hp inhibits TLF1 activity, and that TLF2 is the main trypanolytic factor in NHS.

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