Neurosurgical Approaches to Brain Tissue Harvesting for the Establishment of Cell Cultures in Neural Experimental Cell Models

Abstract

In recent decades, cell biology has made rapid progress. Cell isolation and cultivation techniques, supported by modern laboratory procedures and experimental capabilities, provide a wide range of opportunities for in vitro research to study physiological and pathophysiological processes in health and disease. They can also be used very efficiently for the analysis of biomaterials. Before a new biomaterial is ready for implantation into tissues and widespread use in clinical practice, it must be extensively tested. Experimental cell models, which are a suitable testing ground and the first line of empirical exploration of new biomaterials, must contain suitable cells that form the basis of biomaterial testing. To isolate a stable and suitable cell culture, many steps are required. The first and one of the most important steps is the collection of donor tissue, usually during a surgical procedure. Thus, the collection is the foundation for the success of cell isolation. This article explains the sources and neurosurgical procedures for obtaining brain tissue samples for cell isolation techniques, which are essential for biomaterial testing procedures.

Keywords: biomaterial testing; brain; cell isolation; experimental cell models; neurosurgery.

Figure 1

(A) The primary culture of human astrocytes in low-density culture. Individual polygonally shaped cells are evident. Images were taken at ×50 magnification on Zeiss Axiovert 40 inverted microscope. Scale bar = 200 µm. (B) The immunocytochemical characterization of human astrocytes. The cell morphology was appreciated with orange fluorescent phalloidin conjugate, selectively binding to actin filaments (red). In low-density cultures, astrocytes show a polygonal shape with actin filaments adjacent to the cell membrane. Nuclei were counter-stained with 4′,6-diamidino-2-phenylindole (DAPI) blue. Images were taken at ×10 magnification on EVOS FL fluorescence microscope. Scale bar = 400 µm.

Figure 2

Various resection specimens obtained during brain surgeries. (A) In open surgery and in gross resections, abundant tissue is obtained that can be used for further processing in the cell laboratory, as in open glioblastoma surgery. (B) The resection specimen of glioma obtained from open biopsy or smaller keyhole approach. (C) The glioma sample from the needle biopsy. (D) The biopsy needle with the tissue sample. The images were taken during routine neurosurgical procedures and the tissue was also collected for the purpose of cell culture isolation. The ethical approval number was KME RS 0120-565/2020/5.

Figure 3

(A) Glioblastoma tissue specimen prepared for transport to the cell laboratory for cell isolation and pathology for histopathological evaluation. The sample destined for pathology is preserved in formaldehyde solution (left), and the tissue for the cell laboratory is transported in saline (right). (B) The goal is to transport the tissue on ice and as quickly as possible to reduce cell death.

Figure 4

The culture of glioblastoma cells isolated from the resection specimen. A characteristic polymorphic cell appearance with sparse cytoplasm and differently shaped nuclei are seen. Nicon Diaphot 300 inverted microscope. Scale bar = 100 µm.

Figure 5

(A) Brain exposure in open glioma surgery. (B) The primary brain tumour is clearly visible on the surface. The tumour tissue is more swollen and tan white. (C) The resection cavity after tumour removal. The drainage vein below the tumour was left in place because it is essential to maintain normal brain drainage. All images were taken during routine neurosurgical procedures at our medical centre with patient and ethical committee approval.

Figure 6

Keyhole craniotomy for primary tumours, which is a modification of the conventional craniotomy. (A) The patient is positioned, and the neuronavigation is prepared for guidance. Because less anatomy is visible on the surface, image guidance is essential. (B) The insertion of the instruments for tumour resection through the keyhole craniotomy. With this technique, the morbidity associated with access is minimized, as is the complication rate.

Figure 7

(A) Frame-assisted stereotactic biopsy for deep-seated brain lesions. The stereotactic arch with the attached biopsy guidance introductor is visible. (B) The insertion of the biopsy needle. (C) Frameless stereotactic biopsy. The trajectory is adjusted during the procedure according to neuronavigational panning. (D) The biopsy needle for frameless stereotactic biopsy and needle length adjustment.

Figure 8

(A) The neuroendoport in the position that provides a tubular corridor through the brain to deep-seated lesions. The instruments, the bipolar and aspirator, are then inserted through the neuroendoport into the lesion, with visualization through the operating microscope. (B) Alternatively, the endoscope can be used, which is inserted here through the expandable neuroendoport. The neurosurgical instruments follow next. (C) Intraventricular tumour resection through the expandable neuroendoport. The arrows indicate the lower edge of the neuroendoport corridor.

Figure 9

Neuroendoscopy. (A) The neuroendocoscope is first navigated. (B) During the procedure, the surgical field is observed on the monitor. The exact position of the tip is controlled by a second monitor, which is coupled with the neuronavigation system. (C) Full endoscopy with two working channels. (D) Endoscopic view during tumour resection.