Construction and Evaluation of Theranostic Near-infrared Fluorescent Probe for Targeting Inflammatory Brain Edema

doi:10.12300/j.issn.1674-5817.2023.166

Abstract

Abstract:

Objective A novel compound based on near-infrared fluorescence (NIRF) probe was prepared to achieve dynamic monitoring of an inflammatory brain edema model in mice and real-time evaluation of therapeutic effects through in vivo imaging. Methods The NIRF probe IR-783 was chemically linked with clinical brain edema therapeutic drug furosemide (FSM) to obtain the new compound, IR-783-FSM. The ultraviolet fluorescence properties of the compound were evaluated using an ultraviolet spectrophotometer. The uptake of the compound by mouse macrophage cells RAW 264.7 was detected with in vitro cellular experiments. Its cytotoxicity was evaluated through CCK8 assays. A brain edema model was established in BALB/c mice via intraperitoneal injection of lipopolysaccharide (LPS), confirmed by HE staining and dry-wet weight methods for brain tissues. The mice in the brain edema model were divided into control group, IR-783, and IR-783-FSM treatment groups, receiving intraperitoneal injections of PBS, IR-783, and IR-783-FSM, respectively. Real-time in vivo fluorescence imaging was then performed. The mice in each group were euthanized after 10 hours. Ex vivo brain imaging and dry-wet weight measurements were performed to observe the NIRF imaging characteristics and therapeutic effects of IR-783-FSM on brain edema model. Results The newly synthesized compound, IR-783-FSM, retained the excellent near-infrared fluorescence characteristics of IR-783. It could target mouse macrophages with an IC₅₀ of 48.82 μmol/L. A brain edema model could be successfully constructed with intraperitoneal injection of LPS, with significantly higher brain tissue water content compared to the control group (P<0.01). In vivo imaging showed that IR-783-FSM had a significantly stronger fluorescence signal in the brain edema model than IR-783. Compared to the control group, the brain water content was significantly reduced in the 2, 5, and 8 mmol/L IR-783-FSM treatment groups (P<0.01). Conclusion The newly synthesized NIRF probe IR-783-FSM facilitates dynamic monitoring of brain edema and real-time evaluation of therapeutic effects.

Key words: Brain edema, Lipopolysaccharide, Near-infrared fluorescence, Furosemide, Theranostics, Mice

CLC Number:

R-332

Jing QIN,Yong ZHAO,Caiqin ZHANG,et al. Construction and Evaluation of Theranostic Near-infrared Fluorescent Probe for Targeting Inflammatory Brain Edema[J]. Laboratory Animal and Comparative Medicine, 2024, 44(3): 243-250. DOI: 10.12300/j.issn.1674-5817.2023.166.

Figures/Tables 5

Figure 1 Structure and characterization of the IR-783-FSM probeNote：A, Structure of IR-783-FSM; B, Nuclear magnetic resonance hydrogen spectrum of IR-783-FSM; C, Fluorescence intensity of IR-783-FSM in different solvents (in the left graph, the solvents from left to right are DMSO, H2O, and PBS; in the right graph, the solvents from left to right are H2O and PBS); D, Fluorescence spectra (left) and absorbance spectra (right) of IR-783-FSM.

Figure 2 In vitro cellular uptake ability and cytotoxicity of IR-783-FSM probe in mouse macrophagesNote：Near-infrared fluorescence microscopy was used to observe the uptake ability of mouse macrophages RAW 264.7 to IR-783-FSM probe (Red fluorescence indicated IR-783-FSM probe. Blue fluorescence indicated the nucleus marked by DAPI. Merged plots represented the entry of the IR-783-FSM probe into cells). B, The viability of mouse macrophages RAW 264.7 treated with IR-783-FSM probe was assessed by CCK8 assay.

Figure 3 The mouse model of lipopolysaccharide-induced brain edema was successfully constructedNote：A, After intraperitoneal injection of lipopolysaccharide (LPS), mice appeared lethargic (left) with diarrhea (right). B, HE staining was used to observe the pathological sections of brain tissues in mice after model construction with LPS intraperitoneal injection (left image indicated the blank group injected with normal saline, and the right indicated the mice injected with LPS, while the arrows indicated marked widening of the perivascular spaces, indicating cerebral edema).

Figure 4 Fluorescence imaging results of IR-783 and IR-783-FSM in a mouse model of brain edemaNote： A, In vivo and ex vivo near-infrared fluorescence imaging 10 h after intraperitoneal injections of PBS, IR-783, and IR-783-FSM; B, Real-time near-infrared fluorescence imaging of IR-783-FSM in a mouse model of brain edema; C, Quantitative analysis of near-infrared fluorescence imaging of IR-783-FSM in a mouse model of brain edema (ROI represents regions of interest).

Table 1 Changes of brain water content in mice with brain edema treated with IR-783-FSM

给药分组（n=5） Grouping and administration (n=5)	脑组织含水量/% Brain water content/%
PBS	79.511±0.483
5 mmol/L IR-783	78.364±0.949
2 mmol/L IR-783-FSM	77.988±0.124^**
5 mmol/L IR-783-FSM	77.276±0.816^**
8 mmol/L IR-783-FSM	77.364±0.949^**

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