Folate Receptor Targeted Imaging Using Poly (ethylene glycol)-folate: In Vitro and In Vivo Studies

The aim of this study was to ascertain the folate receptor (FR) targetability by an in vitro study and to acquire FR-targeted images in vivo models by using synthetic folate conjugates. PEG-folate was synthesized and labeled with (99m)Tc and fluorescein isothiocynate (FITC). Cell uptake studies were carried out in KB cells (FR-positive) and A549 cells (FR-negative) using FITC- and the (99m)Tc-labeled conjugates. The radiolabeled conjugate was intravenously injected to KB tumor xenografted mice. After it was injected, gamma images were recorded at 30 min, 1, 2, 3 and 4 hr. Cell uptake studies showed a difference between the KB cells and the A549 cells by flow cytometry analysis and gamma counting. On in vivo images, the tumor-tonormal muscle ratio was greater than 4. It ascertained that the PEG-folate conjugate specifically binds to the FR expressed on tumor cells in vitro. Moreover, it was possible to acquire the FR-targeted gamma images using PEG-folate conjugates in tumor models.

Inhibition of endocytotic functions in Dictyostelium discoideum treated with a carbamate pesticide.

Administration of a carbamate pesticide carbaryl (1-Naphthyl-N-methyl carbamate) at a concentration of 60 and 100 ppm greatly inhibits the endocytotic functions during growth of the cellular slime mold D. discoideum. The ingestion of fluorescien isothiocynate (FITC) labeled E. coli is reduced between 30 and 40% in the treated cells as compared to controls. Similarly, the uptake of FITC-labeled dextran, which has been used as fluid-phase marker for pinocytosis also show 40-50% inhibition in the treated cells. 3H-leucine uptake and incorporation are also inhibited in the treated cells. SDS-PAGE analysis of cytoskeletal proteins shows a higher actin association with the membrane of treated cells. The results demonstrate the detrimental effects of Carbamate on the soil microbe even at a very low concentration and the efficacy of the slime mold cells as a biosensor for the carbamate-induced cytotoxicity.

Determination of macromolecular exchange and PO2 in the microcirculation: a simple system for in vivo fluorescence and phosphorescence videomicroscopy

We have developed a system with two epi-illumination sources, a DC-regulated lamp for transillumination and mechanical switches for rapid shift of illumination and detection of defined areas (250-750 æm²) by fluorescence and phosphorescence videomicroscopy. The system permits investigation of standard microvascular parameters, vascular permeability as well as intra- and extravascular PO2 by phosphorescence quenching of Pd-meso-tetra (4-carboxyphenyl) porphine (PORPH). A Pechan prism was used to position a defined region over the photomultiplier and TV camera. In order to validate the system for in vivo use, in vitro tests were performed with probes at concentrations that can be found in microvascular studies. Extensive in vitro evaluations were performed by filling glass capillaries with solutions of various concentrations of FITC-dextran (diluted in blood and in saline) mixed with different amounts of PORPH. Fluorescence intensity and phosphorescence decay were determined for each mixture. FITC-dextran solutions without PORPH and PORPH solutions without FITC-dextran were used as references. Phosphorescence decay curves were relatively unaffected by the presence of FITC-dextran at all concentrations tested (0.1 æg/ml to 5 mg/ml). Likewise, fluorescence determinations were performed in the presence of PORPH (0.05 to 0.5 mg/ml). The system was successfully used to study macromolecular extravasation and PO2 in the rat mesentery circulation under controlled conditions and during ischemia-reperfusion