In vivo monitoring of angiogenesis in a mouse hindlimb ischemia model using fluorescent peptide-based probes.

Vascular endothelial growth factor receptor (VEGFR) and matrix metalloproteinase (MMP) are up-regulated in ischemic tissue and play pivotal roles in promoting angiogenesis. The purpose of the present study was to evaluate two fluorophore-conjugated peptide probes specific to VEGFR and MMP for dual-targeted in vivo monitoring of angiogenesis in a murine model of hindlimb ischemia. To this end, VEGFR-Probe and MMP-Probe were developed by conjugating distinct near-infrared fluorophores to VEGFR-binding and MMP substrate peptides, respectively. VEGFR-Probe exhibited specific binding to VEGFR on HUVECs, and self-quenched MMP-Probe produced strong fluorescence intensity in the presence of MMPs in vitro. Subsequently, VEGFR-Probe and MMP-Probe were successfully utilized for time course in vivo visualization of VEGFR or MMP, respectively. Simultaneous visualization provided information regarding the spatial distribution of these proteins, including areas of co-localization. This dual-targeted in vivo imaging approach will be useful for understanding the detailed mechanism of angiogenesis and for evaluating therapeutic angiogenesis.

Radioisotope imaging of microRNA-9-regulating neurogenesis using sodium iodide sympoter.

Since microRNAs (miRNA, miR) are known to be critical in various cellular processes and diseases, non-invasive molecular imaging system for miRNA is very important for imaging cellular therapy and disease diagnosis. In this study, we developed a radionuclide imaging system for miR-9 using sodium iodide symporter (NIS). During neuronal differentiation of P 19 cells induced by the treatment of retinoic acid (RA), in vitro and in vivo imaging demonstrated that the expression and activity of NIS from the miR-9 NIS reporter gene was clearly repressed by the increased expression and functional activity of miR-9 that bound with the target sequences in the NIS reporter gene and resulted in destabilized the transcriptional activity of NIS gene, compared with the undifferentiated P19 cells. The decreased activity of NIS from the differentiated P19 cells resulted in low uptake of radionuclide and decreased radioisotope signals. The NIS reporter gene-based miRNA imaging system showed a great specificity of imaging miRNA biogenesis during cellular developments. The miRNA NIS reporter gene will provide high sensitive imaging for visualizing miRNA-regulating cellular developments and diseases.

A reverse complementary multimodal imaging system to visualize microRNA9-involved neurogenesis using peptide targeting transferrin receptor-conjugated magnetic fluorescence nanoparticles.

Multimodal imaging systems may eliminate the disadvantages of individual imaging modality by providing complementary information about cellular and molecular activites. In this sutdy, we developed a reverse complementary multimodal imaging system to image microRNAs (miRNA, miR) during neurognesis using transferrin receptor (TfR) and a magnetic fluorescence (MF) nanoparticle-conjugated peptide targeting TfR (MF targeting TfR). Both in vitro and in vivo imaging demonstrated that, in the absence of miR9 during pre-differentiation of P19 cells, the MF targeting TfR nanoparticles greatly targeted TfR and were successfully internalized into P19 cells, resulting in high fluorescence and low MR signals. When the miR9 was highly expressed during neurogenesis of P19 cells, the MF targeting TfR nanoparticles were hardly targeted due to the miR9 function, which represses the expression and functional activity of TfR from the miRNA TfR reproter gene, resulting in low fluorescence and high MR signals. The reverse complementary multimodal miRNA imaging system may serve as a new imaging probe to montior miRNA-involved cellular developments and diseases.

Molecular imaging of a cancer-targeting theragnostics probe using a nucleolin aptamer- and microRNA-221 molecular beacon-conjugated nanoparticle.

MicroRNAs (miRNA, miR) have been reported as cancer biomarkers that regulate tumor suppressor genes. Hence, simultaneous detecting and inhibiting of miRNA function will be useful as a cancer theragnostics probe to minimize side effects and invasiveness. In this study, we developed a cancer-targeting therangostics probe in a single system using an AS1411 aptamer - and miRNA-221 molecular beacon (miR-221 MB)-conjugated magnetic fluorescence (MF) nanoparticle (MFAS miR-221 MB) to simultaneously target to cancer tissue, image intracellularly expressed miRNA-221 and treat miRNA-221-involved carcinogenesis. AS1411 aptamer-conjugated MF (MFAS) nanoparticles displayed a great selectivity and delivery into various cancer cell lines. The miR-221 MB detached from the MFAS miR-221 MB in the cytoplasm of C6 cells clearly imaged miRNA-221 biogenesis and simultaneously resulted in antitumor therapeutic effects by inhibiting miRNA function, indicating a successful astrocytoma-targeting theragnostics. MFAS miRNA MB can be easily applied to other cancers by simply changing a targeted miRNA highly expressed in cancers.