cyanine-dye-3 and Carcinoma--Lewis-Lung

To microscopically analyze the chemotherapeutic response of tumors using in vivo staining based on an annexinV-Cy5.5 probe and independently asses their apoptotic count using quantitative histological analysis.. Lewis Lung Carcinomas cells, that are sensitive (CS-LLC) and resistant (CR-LLC) to chemotherapy were implanted in nude mice and grown to tumours. Mice were treated with cyclophosphamide and injected with a Cy5.5-annexinV fluorescent probe. In vivo imaging was performed using Fluorescence Molecular Tomography. Subsequently tumours were excised and prepared for histology. The histological tumour sections were stained for apoptosis using a terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) assay. A minimum of ten tissue sections were analyzed per tumour for apoptosis quantification by TUNEL staining and corresponding Cy5.5 distribution.. We detected higher levels of apoptosis and corresponding higher levels of Cy5.5 fluorescence in the CS-LLC vs. the CR-LLC tumours. The cell count rate on CS-LLC sections over CR-LLC was found to be approximately 2 :1 where the corresponding area observed on Cy5.5 distribution measurements revealed a approximately 1.7 :1 ratio of CS-LLC over CR-LLC. These observations are consistent with the higher apoptotic index expected from the CS-LLC cell line.. Quantitative analysis of histological slices revealed higher fluorescence and higher apoptotic count in the CS-LLC tumour images compared to the CR-LLC tumour images. These observations demonstrate that the annexinV-Cy5.5 probe sensed the chemotherapeutic effect of cyclophospamide and further confirmed in vivo FMT measurements.

Topics: Animals; Annexin A5; Annexins; Apoptosis; Carbocyanines; Carcinoma, Lewis Lung; Cell Count; Cell Line, Tumor; Cyclophosphamide; Drug Resistance, Neoplasm; Feasibility Studies; Female; Fluorescent Dyes; Immunohistochemistry; In Situ Nick-End Labeling; Mice; Mice, Nude; Neoplasm Transplantation; Tomography, Optical

We designed a fluorescent peptide-magnetic nanoparticle conjugate that images E-selectin expression in mouse xenograft models of Lewis lung carcinoma (LLC) by fluorescence reflectance imaging. It was synthesized by attaching the E-selectin-binding peptide (ESBP; CDSDSDITWDQLWDLMK) to a CLIO(Cy5.5) nanoparticle to yield ESBP-CLIO(Cy5.5). Internalization by activated human umbilical vein endothelial cells (HUVECs) was rapid and mediated by E-selectin, indicated by the lack of uptake of nanoparticles bearing similar numbers of a scrambled peptide (Scram). To demonstrate the specificity of E-selectin targeting to ESBP-CLIO(Cy5.5) in vivo, we coinjected ESBP-CLIO(Cy5.5) and Scram-CLIO(Cy3.5) and demonstrated a high Cy5.5/Cy3.5 fluorescence ratio using the LLC. Histology showed that ESBP-CLIO was associated with tumor cells as well as endothelial cells, but fluorescence-activated cell sorter analysis showed a far less expression of E-selectin on LLC than on HUVECs. Using immunohistochemistry, we demonstrated E-selectin expression in both endothelial cells and cancer cells in human prostate cancer specimens. We conclude that ESBP-CLIO(Cy5.5) is a useful probe for imaging E-selectin associated with the LLC tumor, and that E-selectin is expressed not only on endothelial cells but also on LLC cells and human prostate cancer specimens.

Topics: Animals; Carbocyanines; Carcinoma, Lewis Lung; Cell Line; Cell Line, Tumor; Cell Nucleus; Cell Separation; E-Selectin; Edetic Acid; Endothelial Cells; Endothelium, Vascular; Flow Cytometry; Humans; Immunohistochemistry; Interleukin-1; Male; Mice; Microscopy, Confocal; Microscopy, Fluorescence; Nanostructures; Nanotechnology; Neoplasm Transplantation; Peptides; Platelet Endothelial Cell Adhesion Molecule-1; Prostatic Neoplasms; Sensitivity and Specificity; Substrate Specificity; Time Factors; Umbilical Veins