Introduction: : Recurrence of glioblastoma multiforme (GBM) after total resection is speculated to originate from precursor cells that pre-existed in surrounding brain tissue. To simulate human GBM, the authors studied morphology and metabolism of rat GBM.

Methods: 1) C6 rat GBM cells cultured in plate wells and brain implant were morphologically observed, and then histochemically stained. 2) Plate cells were treated with temozolomide. 3) Flow cytometry was applied to C6 cells for mitochondrial volume and oxidative metabolism evaluation, 4) Mitochondrial morphology of C6-cells was studied by EM.

Results: 1) Both plate-culture and implant began to grow spindle-form cells, followed by syncytium formation, then by rapid transformation to multiform undifferentiated cells. Spindle cells were morphologically identical to human spongioblasts, which Bailey and Cushing claimed to be the origin of human glioblastoma (1926). CD15-stained spindle cells were found in both culture groups. 2) After temozolomide treatment of plate-cells, multiform cells were mostly destroyed, but spindle cells survived, and grew to malignancy. 3A) Spindle cell mitochondria were greater in volume than multiform cells, and the higher the concentration, the greater the mitochontrial volume in spindle cells. 3B) Superoxide production was greater in spindle cells than in multiform cells. 6) EM of spindle cells showed normal appearance of their mitochondria.

Conclusions: 1) Spindle cells are capable of GBM production, as human GBM precursor. 2) After TMZ treatment, spindle cells are still malignancy-productive. 3) Greater mitochondrial volume of spindle cells over multiform cells suggests higher metabolism of the former. The higher superoxide production of spindle cells over undiffirentiated cells proves the above postulation. 4) Normal EM appearance of spindle cell mitochondria supports their high oxidative metabolism. 4) Our previously reported CD15 and CD133 stained stem-like cells demonstrated in the histologically normal brain adjacent to the tumor margin prove our original precursor speculation. They must be targeted by treatment.

Patient Care: We will improve patient care by applying therapeutic agents vigorously to precursor cells, particularly to those surrounding GBM mass after surgical resection. Finding suppressors of hypermetabolic precursors will add to the GBM eradication.

Learning Objectives: "By the conclusion of this session, participants should be able to: 1) understand the concept of GBM precursor. Describe the importance of Experimental rat GBM has the same morphological and molecular biological behavior, 2) Discuss, in small groups, the importance of experimental results and conclusion 3) Identify an effective treatment targeted to the spindle cells in experimental animal model, and to precursor cells in humans.

References: • Bailay P, Cushing H. (1926) A classification of the tumors of the glioma groups on a histogenic basis with a correlation study of prognosis. Lippincott, Philadelphia. • Yamada SM, Sun L, Yamada: A Role of Spindle Cells in Glioblastoma Growth- Preliminary Report. Society of Pediatric Neuro-Oncology, in San Diego, May 7 & 8, 2015 • Yamada SM, Sun L, Yamada, Kawamoto M: Spindle cells in early C6-GBM growth are stem cells - Functional study with mitochondrial volume for therapeutic purpose. Society of Neuro-Oncology, San Antonio, November 19-21, 2015 • Yamada S, Yamada SM, Sun L, Kawamoto M: Spindle cells expressed by stem cells as the Origin of rat glioblastoma – Equivalent to CD133-positive cells adjacent to human GBM, AANS, Chicago, May 1-5, 2016