ZIF-8-Loading Cascade Nucleic Acid Amplification Circuit For Intracellular MicroRNA Imaging

Currently,cancer is a serious threat to human life and health due to its easy metastasis and incurable characteristics.However,the cure rate for early-stage tumors can reach more than 80%.The survival time of patients can be prolonged if early screening can be performed by the detection of specific biomarkers in cancer,and effective intervention and treatment can be taken as soon as possible before the development of the disease.Mi RNA has been shown to be dysregulated in many cancers,and different mi RNA expression profiles can be classified and analyzed with different hypofractionated tumors.Highly sensitive and specific detection of mi RNAs in tumors has great potential for early diagnosis and precision therapy applications.However,the small mi RNA content,high homology and complex cellular environment make mi RNA detection a difficult research problem,and the accuracy of intracellular mi RNA imaging signals has not been verified.Therefore,we constructed a ZIF-8-loading nucleic acid amplification detection system for precise in situ imaging of intracellular mi RNAs.The specific work is as follows:1.Mechanistic validation of a recognition hairpin binding catalytic hairpin assembly-hybrid chain reaction nucleic acid amplification system(R-CHA-HCR).Mi RNA triggers the opening of hairpin H7 to expose DNA initiator sequences,which in turn triggers the CHA-HCR cascade amplification reaction.The feasibility of signal amplification of R-CHA-HCR was investigated,and the reaction mechanism of CHA-CHR was verified by a dual-signal amplification system.The R-CHA-HCR cascade system can be used for sensitive and specific recognition of mi RNAs.2.Construction of acid-responsive R-CHA-HCR@ZIF-8 nanomaterials.The R-CHA-HCR@ZIF-8 nanomaterials were obtained by one-step encapsulation of the cascade nucleic acid hairpin inside the ZIF-8 nanostructure.The biomineralization process enables the obtainment of structurally stable R-CHA-HCR@ZIF-8 nanomaterials with the ability to respond to the weak acidic microenvironment of tumors.The ZIF-8 nanomaterials can effectively protect the internal nucleic acids from nuclease degradation,and the R-CHA-HCR@ZIF-8 nanomaterials can be used for in vitro mi RNA detection in weakly acidic solutions.Therefore,the successful preparation of R-CHA-HCR@ZIF-8 nanomaterials provides a solid foundation for intracellular and in vivo mi RNA imaging.3.ZIF-8-based nucleic acid amplification system for mi RNA imaging in living cells and in vivo.The imaging performance of R-CHA-HCR@ZIF-8 nanomaterials was evaluated at the cellular and in vivo levels.Using breast cancer cells(MCF-7)as a cellular model,the delivery and protection of ZIF-8 nanomaterials against nucleic acids were verified.R-CHA-HCR@ZIF-8 nanomaterials can be effectively taken up by cells and used for the monitoring of mi RNAs in different cells.The accuracy of the fluorescent signal of the intracellular cascade reaction was verified by the construction of a dual-signal amplification system.Using MCF-7 tumor-bearing mice as an animal model,it showed that R-CHA-HCR@ZIF-8 nanomaterials can be effectively used for in vivo mi RNA imaging in tumors,providing a promising nanoplatform for early cancer detection and prognosis.