Transfection of hypertrophic cardiac myocytes in vitro with (99)Tc(m)-labeled antisense miR208b oligonucleotide / 南方医科大学学报

ABSTRACT

<p><b>OBJECTIVE</b>To test the efficiency of transfecting (99)Tc(m)-labeled anti-miR208b oligonucleotide into early hypertrophic cardiac myocytes in vitro.</p><p><b>METHODS</b>The anti-oligonucleotide targeting miR208b (AMO) was synthesized and modified with LNA followed by conjugation with N-hydroxysuccinimidyl S-acetyl-meraptoacetyl triglycine (NHS-MAG3) and radiolabeling with (99)Tc(m). NHS-MAG3-LNA-AMO and labeled AMO were purified with Sep-Pak C18 column chromatography, and the former was examined for UV absorption at the 260 nm using Gene Quant DNA/RNA calculator. The labeling efficiency, radiochemical purity, stability and molecular hybridization activity were analyzed. An angiotensin II-induced cell model of hypertrophic cardiac myocytes was transfected with (99)Tc(m)-NHS-MAG3-LNA-AMO via liposome, and the relative expression of miRNA208b and retention ratio of the labeled AMO in early hypertrophic cells were determined.</p><p><b>RESULTS</b>The labeling efficiency and radiochemical purity of the labeled AMO after purification exceeded 84% and 86%, respectively. The radio- chemical purities of the labeled AMO incubated in serum and normal saline for 12 h were both higher than 80%, and the labeled AMO showed a capacity to hybridize with the target gene. In the hypertrophic model of cardiac myocytes, the retention ratio of labeled AMO at 6 h was higher than 20%.</p><p><b>CONCLUSION</b>The (99)Tc(m)-labeled antisense probe can be efficiently transfected into hypertrophic cardiac myocytes in vitro, which provides an experimental basis for subsequent radionuclide imaging studies.</p>