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Glioblastoma multiforme (GBM) is recognized as the most common and lethal form of central nervous system cancer. Currently used surgical techniques, chemotherapeutic agents, and radiotherapy strategies have done very little in extending the life expectancies of patients diagnosed with GBM. The difficulty in treating this malignant disease lies both in its inherent complexity and numerous mechanisms of drug resistance. In this review, we summarize several of the primary mechanisms of drug resistance. We reviewed available published literature in the English language regarding drug resistance in glioblastoma. The reasons for drug resistance in glioblastoma include drug efflux, hypoxic areas of tumor cells, cancer stem cells, DNA damage repair, and miRNAs. Many potential therapies target these mechanisms, including a series of investigated alternative and plant-derived agents. Future research and clinical trials in glioblastoma patients should pursue combination of therapies to help combat drug resistance. The emerging new data on the potential of plant-derived therapeutics should also be closely considered and further investigated.

BACKGROUND Human glioblastoma is a deadly brain cancer that continues to defy all current therapeutic strategies. We induced apoptosis in human glioblastoma T98G and U87MG cells following treatment with apigenin (APG), (−)-epigallocatechin (EGC), (−)-epigallocatechin-3-gallate (EGCG), and genistein (GST) that did not induce apoptosis in human normal astrocytes (HNA). METHODS Induction of apoptosis was examined using Wright staining and ApopTag assay. Production of reactive oxygen species (ROS) and increase in intracellular free [Ca2+] were measured by fluoresent probes. Analysis of mRNA and Western blotting indicated increases in expression and activities of the stress kinases and cysteine proteases for apoptosis. JC-1 showed changes in mitochondrial membrane potential (ΔΨm) and use of specific inhibitors confirmed activation of kinases and proteases in apoptosis. RESULTS Treatment of glioblastoma cells with APG, EGC, EGCG, or GST triggered ROS production that induced apoptosis with phosphorylation of p38 MAPK and activation of the redox-sensitive JNK1 pathway. Pretreatment of cells with ascorbic acid attenuated ROS production and p38 MAPK phosphorylation. Increases in intracellular free [Ca2+] and activation of caspase-4 indicated involvement of endoplasmic reticulum stress in apoptosis. Other events in apoptosis included overexpression of Bax, loss of ΔΨm, mitochondrial release of cytochrome c and Smac into the cytosol, down regulation of baculoviral inhibitor-of-apoptosis repeat containing proteins, and activation of calpain, caspase-9, and caspase-3. EGC and EGCG also induced caspase-8 activity. APG, EGC, EGCG, or GST did not induce apoptosis in HNA. CONCLUSION Results strongly suggest that flavonoids are potential therapeutic agents for induction of apoptosis in human glioblastoma cells.

Temporal lobe epilepsy (TLE) is the most common form of focal epilepsy. Previous research has demonstrated several trends in human tissue that, undoubtedly, contribute to the development and progression of TLE. In this study we examined resected human hippocampus tissue for a variety of changes including gliosis that may contribute to the development and presentation of TLE. The study subjects consisted of 6 TLE patients and 3 sudden-death controls. Clinicopathological characteristics were evaluated by H&E staining. Immunohistological staining and Western blotting methods were used to analyze the samples. Neuronal hypertrophy was observed in resected epileptic tissue. Immunohistological staining demonstrated that activation of astrocytes was significantly increased in epileptic tissue as compared corresponding regions of the control group. The western blot data also showed increased CX43 and AQP4 in the hippocampus and downregulation of Kir4.1, α-syntrophin, and dystrophinin, which are the key constituents of AQP4 multi-molecular complex. These tissues also demonstrated changes in inflammatory factors (COX-2, TGF-β, NFkB) suggesting that these molecules may play an important role in TLE pathogenesis. In addition we detected increases in metabotropic glutamate receptor (mGluR) 2/3, mGluR5 and kainic acid receptor subunits KA1 (Grik4) and KA2 (Grik5) in patients' hippocampi. We noted increased expression of the α1c subunit comprising Class C L-type Ca2+ channels and calpain expression in these tissues, suggesting that these subunits may have an integral role in TLE pathogenesis. These changes found in the resected tissue suggest that they may contribute to TLE and that the Kainic acid receptor (KAR) and deregulation of GluR2 receptor may play an important role in TLE development and disease course. This study identifies alterations in number of commonly studied molecular targets associated with astrogliosis, cellular hypertrophy, water homeostasis, inflammation, and modulation of excitatory neurotransmission in hippocampal tissue from TLE patients.

BACKGROUND. Garlic‐derived organosulfur compounds such as diallyl sulfide (DAS), diallyl disulfide (DADS), and diallyl trisulfide (DATS) provide significant protection against carcinogenesis. METHODS. Dose‐dependent cytotoxic effects of the garlic compounds (DAS, DADS, and DATS) were tested in human glioblastoma T98G and U87MG cells. Wright staining and ApopTag assay confirmed induction of apoptosis. Measurements showed that production of reactive oxygen species (ROS) and an increase in intracellular free [Ca2+] promoted apoptosis. Western blot analysis indicated that increased expression and activities of the stress kinases and cysteine proteases caused apoptosis. Use of JC‐1 showed changes in mitochondrial membrane potential (Δψm) for mediation of apoptosis. Use of the specific inhibitors monitored the activation of different kinases and proteases in apoptosis. RESULTS. Treatment of glioblastoma cells with garlic compounds triggered production of ROS that induced apoptosis with the phosphorylation of p38 MAPK and activation of the redox‐sensitive JNK1 pathway. Pretreatment of cells with ascorbic acid attenuated ROS production, p38 MAPK phosphorylation, and JNK1 activation. Pretreatment with JNK1 inhibitor I also significantly reduced cell death. Increases in intracellular free [Ca2+], expression of calreticulin, and activation of caspase‐4 indicated involvement of endoplasmic reticulum (ER) stress in apoptosis. Other events in apoptosis included overexpression of Bax, down‐regulation of Bcl‐2 and some BIRC proteins, mitochondrial release of cytochrome c and Smac into the cytosol, and activation of calpain, caspase‐9, and caspase‐3. CONCLUSIONS. Garlic compounds induced apoptosis in glioblastoma cells due toproduction of ROS, increase in ER stress, decrease in Δψm, and activation of stress kinases and cysteine proteases. Cancer 2007. © 2007 American Cancer Society.

The incidence of acute and chronic spinal cord injury (SCI) in the United States is more than 10,000 per year, resulting in 720 cases per million persons enduring permanent disability each year. The economic impact of SCI is estimated to be more than 4 billion dollars annually. Preclinical studies, case reports, and small clinical trials suggest that early treatment may improve neurological recovery. To date, no proven therapeutic modality exists that has demonstrated a positive effect on neurological outcome. Emerging data from recent preclinical and clinical studies offer hope for this devastating condition. This review gives an overview of current basic research and clinical studies for the treatment of SCI.

Neuroblastoma is the most common extracranial solid tumor in children causing death at pre-school age, as no cure has yet been developed. We investigated the proteolytic mechanisms for apoptosis in human malignant (N-type) neuroblastoma SH-SY5Y cells following exposure to flavonoids such as apigenin (APG), (2)-epigallocatechin (EGC), (2)-epigallocatechin-3-gallate (EGCG) and genistein (GST). We found decrease in viability of SH-SY5Y cells with an increase in dose of APG, EGC, EGCG and GST. Predominantly apoptosis occurred following exposure of SH-SY5Y cells to 50 lM APG, 50 lM EGC, 50 lM EGCG and 100 lM GST for 24 hr. Apoptosis was associated with increases in intracellular free [Ca 21 ] and Bax:Bcl-2 ratio, mitochondrial release of cytochrome c and activation of caspase-9, calpain and caspase-3. Induction of apoptosis with APG and GST showed activation of caspase-12 as well. Activation of caspase-3 could cleave the inhibitor-of-caspaseactivated DNase (ICAD) to release and translocate caspase-3-activated DNase (CAD) to the nucleus. Activation of caspase-8 cleaved Bid to truncated Bid (tBid) in cells treated with EGC and EGCG. EGC and EGCG induced apoptosis with caspase-8 activation and mitochondria-mediated pathway, whereas APG and GST caused apoptosis via an increase in intracellular free [Ca 21 ] with calpain activation and mitochondria-mediated pathway. Activation of different proteases for cell death was confirmed using caspase-8 inhibitor II, calpeptin (calpain inhibitor), caspase-9 inhibitor I and caspase-3 inhibitor IV. Thus, plant-derived flavonoids cause cell death with activation of proteolytic activities of calpain and caspases in SH-SY5Y cells, and therefore serve as potential therapeutic agents for controlling the growth of neuroblastoma. ' 2006 Wiley-Liss, Inc.

Glutamate toxicity has been implicated in cell death in neurodegenerative diseases and injuries. Glutamate-induced Ca2+ influx may mediate activation of calpain, a Ca2+-dependent cysteine protease, which in turn may degrade key cytoskeletal proteins. We investigated glutamate-mediated apoptosis of VSC4.1 motoneurons and functional neuroprotection by calpain inhibition. Exposure of VSC4.1 cells to 10 microM glutamate for 24 hr caused significant increases in intracellular free [Ca2+], as determined by fura-2 assay. Pretreatment of cells with 10 or 25 microM calpeptin (a cell-permeable calpain-specific inhibitor) for 1 hr prevented glutamate-induced Ca2+ influx. Western blot analyses showed an increase in Bax:Bcl-2 ratio, release of cytochrome c from mitochondria, and calpain and caspase-3 activities during apoptosis. Cell morphology, as evaluated by Wright staining, indicated predominantly apoptotic features following glutamate exposure. ApopTag assay further substantiated apoptotic features morphologically as well as biochemically. Our data showed that calpeptin mainly prevented calpain-mediated proteolysis and apoptosis and maintained whole-cell membrane potential, indicating functional neuroprotection. The results imply that calpeptin may serve as a therapeutic agent for preventing motoneuron degeneration, which occurs in amyotrophic lateral sclerosis and spinal cord injury. In this investigation, we also examined glutamate receptor subtypes involved in the initiation of apoptosis in VSC4.1 cells following exposure to glutamate. Our results indicated that the N-methyl-D-aspartate (NMDA) receptors contributed more than alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid (AMPA) receptors to glutamate-mediated Ca2+ influx and cell death mechanism. Inhibition of the activities of both NMDA and AMPA receptors protected VSC4.1 cells from glutamate toxicity and preserved whole-cell membrane potential.