Simple SummaryPreoperative determination of glioma invasion borders remains crucial for neuroradiology. Diffusion kurtosis imaging (DKI) is one of the promising tools as it reflects complex tissue microstructure. Pseudo-continuous arterial spin labeling (pCASL) perfusion is a reliable method to distinguish the most malignant tumor part. In 50 high-grade glioma patients, we demonstrated significant differences between the DKI values in peritumoral white matter (normal-appearing on conventional MRI) and unaffected contralateral hemisphere white matter, which may indicate possible infiltration of normal-appearing peritumoral white matter by glioma cells. The study demonstrated the presence of tumor cells within the edema zone in all gliomas. Tumor cells and tumor stem-like cells were detected in some samples of normal-appearing white matter surrounding glioblastomas. DKI and cerebral flow values showed correlations with quantitative neuropathological markers (proliferative or antiapoptotic activity) in gliomas. Thus, DKI can shed light on tumor-associated brain matter changes and is potentially capable of predicting morphological tumor properties.Abstract(1) Purpose: To determine the borders of malignant gliomas with diffusion kurtosis and perfusion MRI biomarkers. (2) Methods: In 50 high-grade glioma patients, diffusion kurtosis and pseudo-continuous arterial spin labeling (pCASL) cerebral blood flow (CBF) values were determined in contrast-enhancing area, in perifocal infiltrative edema zone, in the normal-appearing peritumoral white matter of the affected cerebral hemisphere, and in the unaffected contralateral hemisphere. Neuronavigation-guided biopsy was performed from all affected hemisphere regions. (3) Results: We showed significant differences between the DKI values in normal-appearing peritumoral white matter and unaffected contralateral hemisphere white matter. We also established significant (p< 0.05) correlations of DKI with Ki-67 labeling index and Bcl-2 expression activity in highly perfused enhancing tumor core and in perifocal infiltrative edema zone. CBF correlated with Ki-67 LI in highly perfused enhancing tumor core. One hundred percent of perifocal infiltrative edema tissue samples contained tumor cells. All glioblastoma samples expressed CD133. In the glioblastoma group, several normal-appearing white matter specimens were infiltrated by tumor cells and expressed CD133. (4) Conclusions: DKI parameters reveal changes in brain microstructure invisible on conventional MRI, e.g., possible infiltration of normal-appearing peritumoral white matter by glioma cells. Our results may be useful for plotting individual tumor invasion maps for brain glioma surgery or radiotherapy planning.