High-Dose Fluorescein Reveals Unusual Confocal Endomicroscope Imaging of Low-Grade Glioma

Abstract

Background: Fluorescence-guided brain tumor surgery using fluorescein sodium (FNa) for contrast is effective in high-grade gliomas. However, the effectiveness of this technique for visualizing noncontrast-enhancing and low-grade gliomas is unknown. This report is the first documented case of the concurrent use of wide-field fluorescence-guided surgery and confocal laser endomicroscopy (CLE) with high-dose FNa (40 mg/kg) for intraoperative visualization of tumor tissue cellularity in a nonenhancing glioma. Case Description: A patient underwent fluorescence-guided surgery for a left frontal lobe mass without contrast enhancement on magnetic resonance imaging. The patient received 40 mg/kg FNa intravenously at the induction of anesthesia. Surgery was performed under visualization with a Yellow 560 filter and white-light wide-field imaging. Intraoperative CLE produced high-quality images of the lesion 1.5 h after FNa injection. Frozen-section analysis demonstrated findings comparable to those of intraoperative CLE visualization and consistent with World Health Organization (WHO) glioma grades II-III. The patient recovered without complications. Analysis of the permanent histologic sections identified the tumor as an anaplastic oligodendroglioma, IDH-mutant, 1p/19q co-deleted, consistent with WHO grade III because of discrete foci of hypercellularity and increased mitotic figures, but large regions of the lesion were low grade. Conclusions: The use of high-dose FNa in this patient with a nonenhancing borderline low-grade/high-grade glioma produced actionable wide-field fluorescence imaging using the operating microscope and improved CLE visualization of tumor cellularity. Higher doses of FNa for intraoperative CLE imaging and possible simultaneous wide-field fluorescence surgical guidance in nonenhancing gliomas merit further investigation.

Keywords: confocal laser endomicroscopy; fluorescein sodium; fluorescence-guided surgery; low-grade glioma; nonenhancing glioma; oligodendroglioma.

Figure 1

(A–E) Preoperative and (F–I) postoperative magnetic resonance imaging (MRI) of a patient with a nonenhancing lesion (A–I, arrows) in the left precentral gyrus. Preoperative MRIs shown are (A) axial diffusion-weighted, (B) axial T2-weighted, (C) axial T1-weighted, (D) sagittal T1-weighted with contrast, and (E) coronal T1-weighted with contrast. Postoperative MRIs shown are (F) axial T1-weighted with contrast, (G) coronal T1-weighted with contrast, (H) coronal diffusion-weighted, and (I) axial diffusion-weighted. Used with permission from Barrow Neurological Institute, Phoenix, Arizona.

Figure 2

Intraoperative tumor visualization. Exposure during tumor resection with (A) Yellow 560 filter and (B) standard white light. (C) Photograph of intraoperative frozen-tissue section slide interpreted as grade II glioma. (D) Confocal laser endomicroscopy (CLE) image showing subtle discrete cellular aggregates (arrows) within a relatively widespread area of cells interpreted as consistent with lower-grade glioma 1.5 h after injection of 40 mg/kg fluorescein sodium (FNa). (E) A vessel 1.5 h after injection of 40 mg/kg FNa showing irregular endothelium (top arrow), with a focus of cells indicating pleomorphism relative to surrounding cells (center arrow) and what appears to be a mitotic figure (bottom arrow). (F) Scanning electron microscopy (SEM) image showing glioma cell morphology with multiple small vesicles and protrusions. (G) CLE image of another tumor area demonstrates small bright dots that likely represent cell surface vesicles with fluorescein, which corresponds well to the SEM microscopy of the cells. These bright dots were visible throughout the tumor on CLE images. Scale 50 μm (D,E,G). Panels (A–E,G) are used with permission from Barrow Neurological Institute, Phoenix, Arizona. Panel (F) is adapted with permission from Lv et al. (22), CC-BY 4.0.

Figure 3

Good correlation of the (A) confocal laser endomicroscopy image collocated with the (B) hematoxylin-eosin–stained frozen section interpreted as grade II glioma. Scale bars, 50 μm. Used with permission from Barrow Neurological Institute, Phoenix, Arizona.

Figure 4

(A–D) Confocal laser endomicroscopy (CLE) images showing areas of low-grade glioma (A,C,D) that reveal distinct strand-like structures or fibers throughout the tumor (scale 50 μm). (B) Image from the same area stained for neurofilaments (arrows) also shows these structures, which are likely swollen axons resulting from tumor edema (scale bar, 20 μm). We have not previously observed these detailed structures on CLE images with low-dose fluorescein sodium (FNa) staining. Their notable appearance here is likely related to the high dose of FNa used in this case. Used with permission from Barrow Neurological Institute, Phoenix, Arizona.