Related Experiment Video
Updated: Nov 17, 2025
In vivo Dual Substrate Bioluminescent Imaging
Published on: October 11, 2011
Real-time visualization of intratumoral necrosis using split-luciferase reconstitution by protein trans-splicing
Go Kagiya1,2, Ayaka Sato3, Ryohei Ogawa4
1School of Allied Health Sciences, Kitasato University, 1-15-1 Kitasato, Minami-ku, Sagamihara, Kanagawa 252-0373, Japan.
Abstract:
Necrosis, a form of cell death, occurs not only with the development of various diseases but also with a tumor tissue response to cancer treatment. Therefore, pursuing progress for cancer therapy through induction of necrosis may be one of the most effective approaches for cancer eradication. We herein describe the development of a real-time imaging system to visualize intratumoral necrosis. The system is composed of two types of cells expressing either one of two necrosis imaging reporters that consist of a DnaE intein sequence linking to one of two split-luciferase fragments. When necrosis occurs in a tumor composed of both of the cells, the two types of leaked reporters can reconstitute the enzymatic activity as a result of protein trans-splicing and thereby emit bioluminescence in the presence of the substrate. This system, which was constructed with shrimp-derived luciferase, allowed in vitro imaging of necrosis. We further confirmed real-time imaging of intratumoral necrosis caused by physical or chemical tissue disruption, validating its application in in vivo necrosis imaging. Thus, the constructed imaging system could be a powerful tool for the optimization of the therapeutic condition for cancer therapy and for the evaluation of novel anticancer drugs targeting necrosis.
Insights
Researchers developed a novel real-time imaging system to visualize necrosis, a form of cell death, within tumors. This tool aids in optimizing cancer therapy and evaluating new necrosis-targeting drugs.
Area of Science:
- Biomedical Engineering
- Molecular Imaging
- Cancer Research
Background:
- Necrosis, a form of programmed cell death, is implicated in disease development and tumor response to cancer therapy.
- Inducing necrosis is a promising strategy for effective cancer eradication.
- Visualizing intratumoral necrosis is crucial for treatment optimization and drug evaluation.
Purpose of the Study:
- To develop a real-time imaging system for visualizing intratumoral necrosis.
- To create a tool for evaluating cancer therapies that target necrosis.
Main Methods:
- Engineered a necrosis imaging reporter system using split-luciferase fragments and a DnaE intein sequence.
- Co-cultured two cell types, each expressing one part of the reporter system.
- Utilized protein trans-splicing to reconstitute luciferase activity and generate bioluminescence upon necrosis-induced reporter leakage.
Main Results:
- Successfully demonstrated in vitro imaging of necrosis using the developed system.
- Confirmed real-time in vivo imaging of intratumoral necrosis induced by physical and chemical disruption.
- Validated the system's capability for monitoring necrosis in a tumor microenvironment.
Conclusions:
- The developed real-time imaging system provides a powerful tool for visualizing intratumoral necrosis.
- This technology can aid in optimizing therapeutic conditions for cancer treatment.
- The system is valuable for evaluating novel anticancer drugs designed to target necrosis.
