Transfection of hypertrophic cardiac myocytes in vitro with 99Tcm-labeled antisense miR208b oligonucleotide
- VernacularTitle:99Tcm标记反义miR208b寡核苷酸及其转染离体肥大心肌细胞的实验研究
- Author:
Jin WANG
1
;
Huijuan FENG
;
Yangwei OU
;
Yungang SUN
;
Juqing WU
;
Pan CHEN
Author Information
1. 南方医科大学珠江医院核医学科
- Keywords:
miRNA208b;
9 Tcm;
anti-miR oligonucleotide;
NHS-MAG3;
myocardial hypertrophy;
liposome
- From:
Journal of Southern Medical University
2015;(9):1316-1319
- CountryChina
- Language:Chinese
-
Abstract:
Objective To test the efficiency of transfecting 9 Tcm-labeled anti-miR208b oligonucleotide into early hypertrophic cardiac myocytes in vitro. Methods 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 9 Tcm. 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 9 Tcm-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. Results 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%. Conclusion The 9 Tcm-labeled antisense probe can be efficiently transfected into hypertrophic cardiac myocytes in vitro, which provides an experimental basis for subsequent radionuclide imaging studies.
