sepharose and Brain-Neoplasms

Systemic drug delivery to the central nervous system (CNS) has been historically impeded by the presence of the blood brain barrier rendering many therapies inefficacious to any cancer cells residing within the brain. Therefore, local drug delivery systems are being developed to overcome this shortfall. Here we have manufactured polymeric microneedle (MN) patches, which can be anchored within a resection cavity site following surgical removal of a tumour such as isocitrate dehydrogenase wild type glioblastoma (GBM). These MN patches have been loaded with polymer coated nanoparticles (NPs) containing cannabidiol (CBD) or olaparib (OLA) and applied to an in vitro brain simulant and ex vivo rat brain tissue to assess drug release and distance of penetration. MN patches loaded with methylene blue dye were placed into a cavity of 0.6 % agarose to simulate brain tissue. The results showed that clear channels were generated by the MNs and the dye spread laterally throughout the agarose. When loaded with CBD-NPs, the agarose showed a CBD concentration of 12.5 µg/g at 0.5 cm from the MN insertion site. Furthermore, high performance liquid chromatography of ex vivo brain tissue following CBD-NP/MN patch insertion showed successful delivery of 59.6 µg/g into the brain tissue. Similarly, OLA-NP loaded MN patches showed delivery of 5.2 µg/g OLA into agarose gel at 0.5 cm distance from the insertion site. Orbitrap secondary ion mass spectrometry (OrbiSIMS) analysis confirmed the presence of OLA and the MN patch at up to 6 mm away from the insertion site following its application to a rat brain hemisphere. This data has provided insight into the capabilities and versatility of MN patches for use in local brain drug delivery, giving promise for future research.

Topics: Administration, Cutaneous; Animals; Brain; Brain Neoplasms; Drug Delivery Systems; Glioblastoma; Nanoparticles; Needles; Rats; Sepharose

Glioblastoma (GBM) is the most malignant primary brain tumor and contains tumorigenic cancer stem cells (CSCs), which support the progression of tumor growth. The selection of CSCs and facilitation of the brain tumor niches may assist the development of novel therapeutics for GBM. Herein, hydrogel materials composed of agarose and hydroxypropyl methyl cellulose (HMC) in different concentrations were established and compared to emulate brain tumor niches and CSC microenvironments within a label-free system. Human GBM cell line, U-87 MG, was cultured on a series of HMC-agarose based culture system. Cell aggregation and spheroids formation were investigated after 4 days of culture, and 2.5% HMC-agarose based culture system demonstrated the largest spheroids number and size. Moreover, CD133 marker expression of GBM cells after 6 days of culture in 2.5% HMC-agarose based culture system was 60%, relatively higher than the control group at only 15%. Additionally, cells on 2.5% HMC-agarose based culture system show the highest chemoresistance, even at the high dose of 500 µM temozolomide for 72 h, the live cell ratio was still > 80%. Furthermore, the results also indicate that the expression of ABCG2 gene was up-regulated after culture in 2.5% HMC-agarose based culture system. Therefore, our results demonstrated that biomimetic brain tumor microenvironment may regulate GBM cells towards the CSC phenotype and expression of CSC characteristics. The microenvironment selection and spheroids formation in HMC-agarose based culture system may provide a label-free CSC selection strategy and drug testing model for future biomedical applications.

Topics: AC133 Antigen; Antineoplastic Agents, Alkylating; Apoptosis; Brain Neoplasms; Cell Line, Tumor; Cell Survival; Drug Resistance, Neoplasm; Drug Screening Assays, Antitumor; Glioblastoma; Humans; Hydrogels; Hypromellose Derivatives; Neoplastic Stem Cells; Sepharose; Stem Cell Niche; Temozolomide; Tissue Engineering; Tissue Scaffolds; Tumor Microenvironment

Systemic delivery of 5-aminolevulinic acid leads to enhanced fluorescence image contrast in many tumors due to the increased accumulation of protoporphyrin IX (PpIX), a fluorescent porphyrin that is associated with tumor burden and proliferation. The value of PpIX-guided resection of malignant gliomas has been demonstrated in prospective randomized clinical studies in which a twofold greater extent of resection and improved progression-free survival have been observed. In low-grade gliomas and at the diffuse infiltrative margins of all gliomas, PpIX fluorescence is often too weak to be detected with current low-resolution surgical microscopes that are used in operating rooms. However, it has been demonstrated that high-resolution optical-sectioning microscopes are capable of detecting the sparse and punctate accumulations of PpIX that are undetectable via conventional low-power surgical fluorescence microscopes. To standardize the performance of high-resolution optical-sectioning devices for future clinical use, we have developed an imaging phantom and methods to ensure that the imaging of PpIX-expressing brain tissues can be performed reproducibly. Ex vivo imaging studies with a dual-axis confocal microscope demonstrate that these methods enable the acquisition of images from unsectioned human brain tissues that quantitatively and consistently correlate with images of histologically processed tissue sections.

Topics: Algorithms; Aminolevulinic Acid; Brain Neoplasms; Diagnostic Imaging; Disease-Free Survival; Glioma; Humans; Microscopy, Confocal; Microscopy, Fluorescence; Optics and Photonics; Phantoms, Imaging; Protoporphyrins; Sepharose; Signal-To-Noise Ratio

The indications for stereotactic biopsies or implantation of probes for local chemotherapy in diffuse brainstem tumors have recently come under debate. The quality of performing these procedures significantly depends on the precision of the probes' placement in the brainstem. The authors evaluated the precision of brainstem probe positioning using a navigated frameless stereotactic system in an experimental setting.. Using the VarioGuide stereotactic system, 33 probes were placed into a specially designed model filled with agarose. In a second experimental series, 8 anatomical specimens were implanted with a total of 32 catheters into the pontine brainstem using either a suboccipital or a precoronal entry point. Before intervention in both experimental settings, a thin-sliced CT scan for planning was obtained and fused to volumetric T1-weighted MR imaging data. After the probe positioning procedures, another CT scan and an MR image were obtained to compare the course of the catheters versus the planned trajectory. The deviation between the planned and the actual locations was measured to evaluate the precision of the navigated intervention.. Using the VarioGuide system, mean total target deviations of 2.8 +/- 1.2 mm on CT scanning and 3.1 +/- 1.2 mm on MR imaging were detected with a mean catheter length of 151 +/- 6.1 mm in the agarose model. The catheter placement in the anatomical specimens revealed mean total deviations of 1.95 +/- 0.6 mm on CT scanning and 1.8 +/- 0.7 mm on MR imaging for the suboccipital approach and a mean catheter length of 59.5 +/- 4.1 mm. For the precoronal approach, deviations of 2.2 +/- 1.2 mm on CT scanning and 2.1 +/- 1.1 mm on MR imaging were measured (mean catheter length 85.9 +/- 4.7 mm).. The system-based deviation of frameless stereotaxy using the VarioGuide system reveals good probe placement in deep-seated locations such as the brainstem. Therefore, the authors believe that the system can be accurately used to conduct biopsies and place probes in patients with brainstem lesions.

Topics: Biopsy; Brain Neoplasms; Brain Stem; Catheterization; Child; Glioma; Humans; Magnetic Resonance Imaging; Models, Anatomic; Neuronavigation; Reproducibility of Results; Sepharose; Tomography, X-Ray Computed

Brain invasion prevents complete surgical extirpation of malignant gliomas; however, invasive cells from distant, histologically normal brain previously have not been isolated, cultured, and characterized. To evaluate invasive human malignant glioma cells, the authors established cultures from gross tumor and histologically normal brain. Three men and one woman, with a mean age of 67 years, underwent two frontal and two temporal lobectomies for tumors, which yielded specimens of both gross tumor and histologically normal brain. Each specimen was acquired a minimum of 4 cm from the gross tumor. The specimens were split: a portion was sent for neuropathological evaluation (three glioblastomas multiforme and one oligodendroglioma) and a portion was used to establish cell lines. Morphologically, the specimens of gross tumor and histologically normal brain were identical in three of the four cell culture pairs. Histochemical staining characteristics were consistent both within each pair and when compared with the specimens sent for neuropathological evaluation. Cultures demonstrated anchorage-independent growth in soft agarose and neoplastic karyotypes. Growth rates in culture were greater for histologically normal brain than for gross tumor in three of the four culture pairs. Although the observed increases in growth rates of histologically normal brain cultures do not correlate with in vivo behavior, these findings corroborate the previously reported stem cell potential of invasive glioma cells. Using the radial dish assay, no significant differences in motility between cultures of gross tumor and histologically normal brain were found. In summary, tumor cells were cultured from histologically normal brain acquired from a distance greater than 4 cm from the gross tumor, indicating the relative insensitivity of standard histopathological identification of invasive glioma cells (and hence the inadequacy of frozen-section evaluation of resection margins). Cell lines derived from gross tumor and histologically normal brain were usually histologically identical and demonstrated equivalent motility, but had different growth rates.

Topics: Aged; Brain; Brain Neoplasms; Cell Adhesion; Cell Division; Cell Movement; Cells, Cultured; Coloring Agents; Culture Media; Female; Frontal Lobe; Glioblastoma; Glioma; Histocytochemistry; Humans; Karyotyping; Male; Neoplasm Invasiveness; Oligodendroglioma; Sepharose; Stem Cells; Temporal Lobe; Tumor Cells, Cultured

Autoantibodies from the MRL/lpr mice react with numerous proteins on neuronal cell surfaces. The purpose of this study was to isolate and characterize a population of autoantibodies reactive preferentially or exclusively with nervous system tissue. Using a purified plasma membrane preparation from brain cortex of balb/c mice coupled to diaminopropylamine agarose gel, we affinity-isolated antineuronal antibodies from pooled MRL/lpr immunoglobulins. The isolated immunoglobulins reacted with brain cortex plasma membranes and neuroblastoma cells (but not liver, kidney, or fibroblasts) by Western blot and indirect immunofluorescence with confocal microscopy. By Western blot, the epitopes in the brain cortex were proteins of apparent molecular weights 101, 63, 53, 43, 39, and 33, kd; the epitopes in the neuroblastoma cells were 63, 57, and 53 kd. Lectin column isolation revealed that the 101 and 63 kd epitopes were glycosylated. Indirect immunofluorescence revealed that the antibodies bound to the cell soma more intensely than to the cell processes of viable cultured neuroblastoma cells. The cell surface localization of this binding was confirmed by confocal microscopy. Within the central nervous system the antibodies bound more intensely to primary cultures of isolated neurons from fetal cortex than to hippocampal or neostriatal cells. With these antibodies we can begin studies of their potential pathogenic effects.

Topics: Animals; Antibody Specificity; Antigen-Antibody Reactions; Autoantibodies; Autoimmune Diseases; Blotting, Western; Brain Neoplasms; Cell Membrane; Cerebral Cortex; Chromatography, Affinity; Disease Models, Animal; Female; Fibroblasts; Fluorescent Antibody Technique; Gels; Immunosorbent Techniques; Kidney; Liver; Liver Neoplasms, Experimental; Lupus Erythematosus, Systemic; Membrane Glycoproteins; Mice; Mice, Inbred BALB C; Mice, Mutant Strains; Molecular Weight; Nerve Tissue Proteins; Neuroblastoma; Neurons; Organ Specificity; Sepharose

We combined a vector decomposition technique with Gaussian probability thresholding in feature space to segment normal brain tissues, tumors, or other abnormalities on dual-echo MR images. The vector decomposition technique assigns to each voxel a fractional volume for each of two tissues. A probability threshold, based on an assumed Gaussian probability density function describing random noise, isolates a region in feature space for fractional volume calculation that minimizes contamination from other tissues. The calculated fractional volumes are unbiased estimates of the true fractional volumes. The contrast-to-noise ratio (CNR) between tissues on the segmented images is the same as the Euclidean norm of CNRs in the original images. The method is capable of segmenting more than two tissues from a set of dual-echo images by sequentially analyzing different pairs of tissues. The model is analyzed mathematically and in experiments with a phantom. Two clinical examples are presented.

Topics: Adenocarcinoma; Algorithms; Artifacts; Brain; Brain Neoplasms; Cerebrospinal Fluid; Copper; Copper Sulfate; Cysts; Gels; Humans; Image Enhancement; Image Processing, Computer-Assisted; Magnetic Resonance Imaging; Models, Biological; Models, Structural; Models, Theoretical; Multiple Sclerosis; Probability; Sepharose

Type beta transforming growth factor (beta-TGF) is a potent regulator of cell growth and differentiation. The human glioblastoma cell line, T-MGI, was growth inhibited by beta-TGF under anchorage independent conditions. The antiproliferative effect of beta-TGF was potentiated to nearly total arrest by low doses of retinoic acid (RA) or tumor necrosis factor (TNF), while epidermal growth factor, platelet-derived growth factor, interleukin-2, and gamma interferon did not have this potentiating effect. The potentiation of the beta-TGF effect by RA and TNF could not be explained by modulation of the epidermal growth factor receptor, the beta-TGF receptor, or the TNF receptor. beta-TGF alone and in combination with RA or TNF were further tested on primary cultures from freshly resected human glioma biopsies (n = 13). There was great individual variation in sensitivity to beta-TGF, RA, or TNF. The astrocytoma and oligodendroglioma cells were inhibited to various degrees by beta-TGF or TNF, while most of the glioblastomas were not sensitive to these agents. Most of the biopsies were stimulated by RA. RA or TNF did not potentiate the growth inhibitory effect of beta-TGF on biopsy cells. We therefore think it unlikely that beta-TGF in combination with RA or TNF will be effective agents in the treatment of gliomas.

Topics: Adjuvants, Immunologic; Antineoplastic Combined Chemotherapy Protocols; Brain Neoplasms; Cell Line; Clone Cells; Drug Screening Assays, Antitumor; ErbB Receptors; Glioma; Growth Inhibitors; Humans; Iodine Radioisotopes; Peptides; Sepharose; Transforming Growth Factors; Tretinoin; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha

Type beta transforming growth factor (B-TGF) is a potent regulator of cell growth and differentiation. Growth of many human tumour cell lines are inhibited by B-TGF. The effect of B-TGF on proliferation of clonogenic cells from 12 human glioma biopsies was registrated in a thymidine incorporation assay. B-TGF appeared to be a potent growth inhibitor for some gliomas, while it had no effect on others. Maximum inhibition was about 60%. Though not significant, glioblastomas appeared to be less sensitive to inhibition by B-TGF than astrocytomas and oligodendrogliomas. Very little is known about the growth inhibitory action of B-TGF. The negative autocrine growth theory for cancer cells postulates that a reduced production, or production of a defective growth inhibitor normally found in the cell, may account for the autonomous nature of some cancer cells. In view of this theory, we searched for B-TGF in protein extracts from a glioblastoma cell line, T-MGl, whose growth was inhibited by B-TGF. Protein extract from T-MGl cells was analysed for B-TGF activity using a soft agar colony formation assay with normal rat kidney fibroblasts. B-TGF was not found in the extract. Since, according to the literature B-TGF has been found in all other cell lines examined, we believe that there may be a lack of B-TGF or an altered B-TGF in the gliomas whose growth are inhibited by B-TGF. This problem will be studied further.

Topics: Brain Neoplasms; Cell Division; Cell Line; Clone Cells; Culture Media; Glioma; Humans; Sepharose; Transforming Growth Factors