Diagnostic and Therapeutic Approaches for Glioblastoma and Neuroblastoma Cancers Using Chlorotoxin Nanoparticles.

Glioblastoma multiforme (GB) and high-risk neuroblastoma (NB) are known to have poor therapeutic outcomes. As for most cancers, chemotherapy and radiotherapy are the current mainstay treatments for GB and NB. However, the known limitations of systemic toxicity, drug resistance, poor targeted delivery, and inability to access the blood-brain barrier (BBB), make these treatments less satisfactory. Other treatment options have been investigated in many studies in the literature, especially nutraceutical and naturopathic products, most of which have also been reported to be poorly effective against these cancer types. This necessitates the development of treatment strategies with the potential to cross the BBB and specifically target cancer cells. Compounds that target the endopeptidase, matrix metalloproteinase 2 (MMP-2), have been reported to offer therapeutic insights for GB and NB since MMP-2 is known to be over-expressed in these cancers and plays significant roles in such physiological processes as angiogenesis, metastasis, and cellular invasion. Chlorotoxin (CTX) is a promising 36-amino acid peptide isolated from the venom of the deathstalker scorpion, Leiurus quinquestriatus, demonstrating high selectivity and binding affinity to a broad-spectrum of cancers, especially GB and NB through specific molecular targets, including MMP-2. The favorable characteristics of nanoparticles (NPs) such as their small sizes, large surface area for active targeting, BBB permeability, etc. make CTX-functionalized NPs (CTX-NPs) promising diagnostic and therapeutic applications for addressing the many challenges associated with these cancers. CTX-NPs may function by improving diffusion through the BBB, enabling increased localization of chemotherapeutic and genotherapeutic drugs to diseased cells specifically, enhancing imaging modalities such as magnetic resonance imaging (MRI), single-photon emission computed tomography (SPECT), optical imaging techniques, image-guided surgery, as well as improving the sensitization of radio-resistant cells to radiotherapy treatment. This review discusses the characteristics of GB and NB cancers, related treatment challenges as well as the potential of CTX and its functionalized NP formulations as targeting systems for diagnostic, therapeutic, and theranostic purposes. It also provides insights into the potential mechanisms through which CTX crosses the BBB to bind cancer cells and provides suggestions for the development and application of novel CTX-based formulations for the diagnosis and treatment of GB and NB in the future.

Fractions of Hoslundia opposita Vahl and hoslundin induced apoptosis in human cancer cells via mitochondrial-dependent reactive oxygen species (ROS) generation.

BACKGROUND: Cancer remains one of the leading causalities of several morbidity and mortality with negative impact on global economy due to low workforce and management/treatment cost. A number of conventional therapies have been explored in the management/treatment of cancer including chemotherapeutic intervention, radiotherapy, and surgery. Among these treatment modalities, chemotherapy remains the most popular first line of intervention in management/treatment of cancer, and natural products have been implicated as the main source of antineoplastic agents with phenomenal efficacy. However, current antineoplastic agents suffer from lack of selectivity and specificity necessitating the need for further research in the search for novel anticancer drug molecules. METHODS: In this present study, the anticancer activity of Hoslundia opposita leaves extracts were tested against a number of cell lines including human hepatoma cell line (HepG2), human breast cancer cell lines (MDA-MB-231), intestinal epithelial cell lines (Caco-2), and human keratinocyte HACAT cell lines. A bio-guided fractionation assay and the structural elucidation of the pure isolate (hoslundin) was conducted by 1D and 2D NMR spectroscopy. The cell viability, colony formation, and apoptotic activities were investigated using MTT assay, clonogenic assay, and caspase - 3 and - 7 kits respectively. Flow cytometry was employed in assessing the altered cell cycle expression. The production of the intracellular reactive oxygen species (ROS) levels and the reduction of the mitochondrial membrane potential (MMP) was determined at the cellular level using fluorescent probe dyes dihydro-fluorescin diacetate (DCFH-DA) and tetramethylrhodamin (TMRE), respectively. RESULTS: The H. opposita fractions and its pure isolate (hoslundin) demonstrated a potent cytocidal activity against the tumorigenic cells (HepG2, MDA-MB-231, Caco-2) at concentration ranging from 25 to 100 µg/mL. The inhibition of the colony formation was significantly observed in HepG2 cell lines. More so, the cellular viability of the normal cells (HaCaT) was relatively unchanged in the presence of H. opposita fractions and its isolate proving the selectivity of the compounds towards tumourigenic cells. The H. opposita fractions and hoslundin exerted their anticancer activity via cell cycle arrest with the accumulation of the DNA content at the S-phase, activation of apoptosis in the caspase 3,7 activities and depolarized mitochondrial membrane potential mediated by mitochondrial-dependent ROS generation in the treated tumor cells. CONCLUSION: The anticancer activities of Hoslundia opposita Vahl and hoslundin exhibited significant efficacy against tumor cells and well tolerated in the presence of normal cells making them a potential antineoplastic agent.

Comparative Study of the Antioxidant Constituents, Activities and the GC-MS Quantification and Identification of Fatty Acids of Four Selected Helichrysum Species.

Helichrysum Mill. (Asteraceae) is a plant genus comprising distinctively of aromatic plants of about 500-600 species. Since most of these plants have not been previously studied, extensive profiling helps to validate their folkloric uses and determine their potential value as sources of plant-derived drug candidates. This study, therefore, aims to investigate the antioxidant activity (DPPH, NO, FRAP); total antioxidant capacity, total phenolic, total flavonoid, and fatty acid compositions of the aqueous acetone extracts from four Helichrysum plants namely, Helichrysum pandurifolium, Helichrysum foetidum, Helichrysum petiolare, and Helichrysum cymocum. The results obtained showed that the H. cymocum extract had the best DPPH radical scavenging activity (IC50 = 11.85 ± 3.20 µg/mL) and H. petiolare extract had the best nitric oxide scavenging activity (IC50 = 20.81 ± 3.73 µg/mL), while H. pandurifolium Schrank extract (0.636 ± 0.005 µg/mL) demonstrated the best ferrous reducing power, all of which are comparable with results from ascorbic acid used as the standard. The IC50 values of the radical scavenging activity ranged from 11.85-41.13 µg/mL (DPPH), 20.81-36.19 µg/mL (NO), and 0.505-0.636 µg/mL (FRAP), for all the plants studied. The H. petiolare has the highest total antioxidant capacity (48.50 ± 1.55 mg/g), highest total phenolic content (54.69 ± 0.23 mg/g), and highest total flavonoid content (56.19 ± 1.01 mg/g) compared with other species. The fatty acid methyl esters were analysed using gas chromatography-mass spectrometry (GC-MS). The results obtained showed variations in the fatty acid composition of the plant extracts, with H. petiolare having the highest saturated fatty acid (SFA) content (7184 µg/g) and polyunsaturated fatty acid (PUFA) content (7005.5 µg/g). In addition, H. foetidum had the highest monounsaturated fatty acid (MUFA) content (1150.3 µg/g), while H. cymocum had the highest PUFA:SFA ratio of 1.202. In conclusion, the findings from this study revealed that H. pandurifolium Schrank, H. foetidum, H. petiolare, and H. cymocum are repositories of natural bioactive compounds with potential health-promoting benefits that need to be investigated, for both their antioxidant activity in a number of disease conditions and for further exploration in drug discovery and development projects.

Medicinal Properties and In Vitro Biological Activities of Selected Helichrysum Species from South Africa: A Review.

The genus Helichrysum Mill comprises hundreds of species that are mostly flowering perennial shrubs. Some of these plants that belong to the Helichrysum species are used in traditional medicine to treat cough, back pain, diabetes, asthma, digestive problems, menstrual pain, chest pain, kidney disorders, skin disorders, wounds, open sores, among other conditions, but, only a few scientific studies are reported in the literature with sufficient information that validates the acclaimed folkloric benefits of these plants. This review, therefore, provides a comprehensive update of the available information on the cytotoxicity, genotoxicity, anti-proliferative, anti-bacterial, anti-fungal, anti-viral, anti-HIV, anti-malarial, anti-ulcerogenic, anti-tyrosinase, anti-inflammatory, and anti-oxidant activities of selected Helichrysum species of interest: H. petiolare, H. cymocum, H. foetidum, and H. pandurifolium Schrank, using scientific databases as well as electronic and print sources. The ethnobotanical and morphological characteristics as well as the phytochemical composition and biological activities of these plants are elucidated. The scientific rationale for their current use is discussed based on the evidence in the literature. This review highlights the putative use of the Helichrysum species as a reliable source of bioactive compounds for the production of standard commercial drugs to treat many ailments, including those reported in folkloric uses. Further research on the many plants in the genus Helichrysum is recommended to explore their economic importance both as edible crops and medicinal botanicals.

Neuroprotective Activities of Crossyne flava Bulbs and Amaryllidaceae Alkaloids: Implications for Parkinson's Disease.

Parkinson's disease (PD) is one of the most common neurodegenerative diseases and affects approximately 6.3 million people worldwide. To date, the treatment of PD remains a challenge, as available treatment options are known to be associated with serious side effects; hence, the search for new treatment strategies is critical. Extracts from the Amaryllidaceae plant family as well as their alkaloids have been reported to have neuroprotective potentials. This study, therefore, investigated the biological activities of Crossyne flava and its isolated alkaloids in an in vitro MPP+ (1-methyl-4-phenylpyridinium) PD model using SH-SY5Y cells. The effects of the total extract as well as the four compounds isolated from Crossyne flava (i.e., pancratinine B (1), bufanidrine (2), buphanisine (3), and epibuphanisine (4)) were evaluated for cell viability, neuroprotection, levels of reactive oxygen species (ROS), adenosine triphosphate activity (ATP), and caspase 3/7 activity in SH-SY5Y cells. The results obtained showed that pre-treatment with both the extract and the isolated compounds was effective in protecting the SH-SY5Y cells from MPP+-induced neurotoxicity and inhibited ROS generation, ATP depletion as well as apoptosis induction in the SH-SY5Y cells. The results of this study show that the Amaryllidaceae plant family may be a source of novel compounds for the treatment of neurodegenerative diseases, which validates the reported traditional uses.

Design, synthesis, and evaluation of 3,7-substituted coumarin derivatives as multifunctional Alzheimer's disease agents.

Multitarget directed ligands (MTDLs) are emerging as promising treatment options for Alzheimer's disease (AD). Coumarin derivatives serve as a good starting point for designing MTDLs due to their inherent inhibition of monoamine oxidase (MAO) and cholinesterase enzymes, which are complicit in AD's complex pathophysiology. A preliminary series of 3,7-substituted coumarin derivatives were synthesised and evaluated for enzyme inhibitory activity, cytotoxicity as well as neuroprotective ability. The results indicated that the compounds are weak cholinesterase inhibitors with five compounds demonstrating relatively potent inhibition and selectivity towards MAO-B with IC50 values between 0.014 and 0.498 hx00B5;µM. Significant neuroprotective effects towards MPP+-compromised SH-SY5Y neuroblastoma cells were also observed, with no inherent cytotoxicity at 10 µM for all compounds. The overall results demonstrated that substitution of the phenylethyloxy moiety at the 7-position imparted superior general activity to the derivatives, with the propargylamine substitution at the 3-position, in particular, displaying the best MAO-B selectivity and neuroprotection.

Inhibition of γH2AX, COX-2 and regulation of antioxidant enzymes in MPP+-exposed SH-SY5Y cells pre-treated with rutin.

Many plant-derived bioactive compounds such as rutin are reportedly effective in attenuating neuronal death in most neurodegenerative disorders. Parkinson's disease (PD) is characterized by the gradual degeneration of dopaminergic neurons in the substantia nigra of the midbrain, and has previously been modelled in-vitro through the specific neurotoxic activity of 1-methyl-4-phenylpyridinium (MPP+) on dopaminergic neurons. Rutin is a bioflavonoid with multiple pharmacological effects, and this study investigated the neuroprotective effects of rutin in the human dopaminergic SH-SY5Y cell line using the neurotoxin MPP+. SH-SY5Y cells pretreated with rutin, were exposed to MPP+ and evaluated for cell viability, nitric oxide (NO), reactive oxygen species (ROS) and antioxidant enzymes activities. In addition, western blot techniques were used to determine the protein expression levels of γH2AX and COX-2. Rutin significantly attenuated MPP+-induced loss of cell viability, mitigated ROS and NO production and inhibited the disruption of antioxidant enzymes activity. It was also observed that rutin significantly reduced protein expression levels of γH2AX and COX-2 in SH-SY5Y cells treated with MPP+. Taken together, findings from this study tend to suggest that rutin is a promising neuroprotective compound for the treatment of PD through its effects on some of the mechanisms that characterize this neurodegenerative disease.

Cytotoxic and Apoptotic Induction Potential of Extracts from Fermented Citrullus vulgaris Thunb. Seeds on Cervical and Liver Cancer Cells.

The anti-cancer activities of many fermented foods and beverages are now scientifically established. Ogiri-egusi is a condiment prepared from fermentation of Citrullus vulgaris (melon) seeds and consumed in many countries of West Africa. Its anti-oxidative and anti-hyperlipidaemic properties have been reported. This study investigated the anti-cancer activities of the aqueous and methanolic extracts from ogiri-egusi. Cytotoxicity was investigated using the MTT and colony-formation inhibition assays while flow-cytometer based Apopercentage assay was used to quantify apoptosis in extracts-treated cervical and liver cancer and normal human fibroblast cells. The inhibitory concentration responsible for killing 50% of cells after 24 h by the aqueous extract in KMST-6, HeLa, and Hep-G2 cells were estimated at 1.610, 1.020, and 1.507 mg/mL respectively. While these values reduced with increasing incubation time in cancer cells it increased in the non-cancer cell. Furthermore, the extract significantly induced apoptosis in HeLa (97 ± 0.18%) and Hep-G2 (73 ± 6.73%) cells. These findings were corroborated by cells morphologic presentations and inhibition of colony formation assay. These findings suggest that the aqueous extract from fermented Citrullus vulgaris seeds might be a nutraceutical with potential anti-cancer properties.

In Vitro Evaluation of the Anti-Diabetic Potential of Aqueous Acetone Helichrysum petiolare Extract (AAHPE) with Molecular Docking Relevance in Diabetes Mellitus.

Diabetes mellitus (DM) is a chronic metabolic condition that can lead to significant complications and a high fatality rate worldwide. Efforts are ramping up to find and develop novel α-glucosidase and α-amylase inhibitors that are both effective and potentially safe. Traditional methodologies are being replaced with new techniques that are less complicated and less time demanding; yet, both the experimental and computational strategies are viable and complementary in drug discovery and development. As a result, this study was conducted to investigate the in vitro anti-diabetic potential of aqueous acetone Helichrysum petiolare and B.L Burtt extract (AAHPE) using a 2-NBDG, 2-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl) amino)-2-deoxy-d-glucose uptake assay. In addition, we performed molecular docking of the flavonoid constituents identified and quantified by liquid chromatography-mass spectrometry (LC-MS) from AAHPE with the potential to serve as effective and safe α-amylase and α-glucosidase inhibitors, which are important in drug discovery and development. The results showed that AAHPE is a potential inhibitor of both α-amylase and α-glucosidase, with IC50 values of 46.50 ± 6.17 (µg/mL) and 37.81 ± 5.15 (µg/mL), respectively. This is demonstrated by a significant increase in the glucose uptake activity percentage in a concentration-dependent manner compared to the control, with the highest AAHPE concentration of 75 µg/mL of glucose uptake activity being higher than metformin, a standard anti-diabetic drug, in the insulin-resistant HepG2 cell line. The molecular docking results displayed that the constituents strongly bind α-amylase and α-glucosidase while achieving better binding affinities that ranged from ΔG = -7.2 to -9.6 kcal/mol (compared with acarbose ΔG = -6.1 kcal/mol) for α-amylase, and ΔG = -7.3 to -9.0 kcal/mol (compared with acarbose ΔG = -6.3 kcal/mol) for α-glucosidase. This study revealed the potential use of the H. petiolare plant extract and its phytochemicals, which could be explored to develop potent and safe α-amylase and α-glucosidase inhibitors to treat postprandial glycemic levels in diabetic patients.

Regulation of AKT/AMPK signaling, autophagy and mitigation of apoptosis in Rutin-pretreated SH-SY5Y cells exposed to MPP.

Accumulating evidence suggest that apoptosis, autophagy and dysregulation of signaling pathways are common mechanisms involved in Parkinson's disease (PD) pathogenesis, and thus development of therapeutic agents targeting these mechanisms may be useful for the treatment of this disease. Although rutin (a bioflavonoid) is reported to have pharmacological benefits such as antioxidant, anti-inflammatory and antitumor activities, there are very few reports on the activity of this compound in 1-methyl-4-phenylpyridinium (MPP+)-induced PD models. Accordingly, we investigated the effects of rutin on apoptosis, autophagy and cell signaling markers (AKT/AMPK) in SH-SY5Y cells exposed to MPP+. Results show reduced changes in nuclear morphology and mitigation of caspase 3/7 and 9 activities in rutin pre-treated cells exposed to MPP+. Likewise, rutin regulated cell signaling pathways (AKT/AMPK) and significantly decreased protein expression levels of cleaved PARP, cytochrome c, LC3-II and p62. Also, rutin significantly increased protein expression levels of full-length caspase 3 in SH-SY5Y cells treated with MPP+. Transmission electron microscope (TEM) images demonstrated a reduction in autophagosomes in rutin-pretreated SH-SY5Y cells exposed to MPP+. These results provide experimental support for rutin's neuroprotective activity against MPP+-induced toxicity in SH-SY5Y cells, which is as a promising therapeutic agent for clinical trials in humans.

Sleep Deprivation and Neurological Disorders.

Sleep plays an important role in maintaining neuronal circuitry, signalling and helps maintain overall health and wellbeing. Sleep deprivation (SD) disturbs the circadian physiology and exerts a negative impact on brain and behavioural functions. SD impairs the cellular clearance of misfolded neurotoxin proteins like α-synuclein, amyloid-ß, and tau which are involved in major neurodegenerative diseases like Alzheimer's disease and Parkinson's disease. In addition, SD is also shown to affect the glymphatic system, a glial-dependent metabolic waste clearance pathway, causing accumulation of misfolded faulty proteins in synaptic compartments resulting in cognitive decline. Also, SD affects the immunological and redox system resulting in neuroinflammation and oxidative stress. Hence, it is important to understand the molecular and biochemical alterations that are the causative factors leading to these pathophysiological effects on the neuronal system. This review is an attempt in this direction. It provides up-to-date information on the alterations in the key processes, pathways, and proteins that are negatively affected by SD and become reasons for neurological disorders over a prolonged period of time, if left unattended.

An Insight into the Mechanism of Holamine- and Funtumine-Induced Cell Death in Cancer Cells.

Holamine and funtumine, steroidal alkaloids with strong and diverse pharmacological activities are commonly found in the Apocynaceae family of Holarrhena. The selective anti-proliferative and cell cycle arrest effects of holamine and funtumine on cancer cells have been previously reported. The present study evaluated the anti-proliferative mechanism of action of these two steroidal alkaloids on cancer cell lines (HT-29, MCF-7 and HeLa) by exploring the mitochondrial depolarization effects, reactive oxygen species (ROS) induction, apoptosis, F-actin perturbation, and inhibition of topoisomerase-I. The apoptosis-inducing effects of the compounds were studied by flow cytometry using the APOPercentageTM dye and Caspase-3/7 Glo assay kit. The two compounds showed a significantly greater cytotoxicity in cancer cells compared to non-cancer (normal) fibroblasts. The observed antiproliferative effects of the two alkaloids presumably are facilitated through the stimulation of apoptosis. The apoptotic effect was elicited through the modulation of mitochondrial function, elevated ROS production, and caspase-3/7 activation. Both compounds also induced F-actin disorganization and inhibited topoisomerase-I activity. Although holamine and funtumine appear to have translational potential for the development of novel anticancer agents, further mechanistic and molecular studies are recommended to fully understand their anticancer effects.

Neuroprotective Activities of Boophone haemanthoides (Amaryllidaceae) Extract and Its Chemical Constituents.

Parkinson's disease (PD) is a neurodegenerative condition that progresses as age increases, and some of its major symptoms include tremor and postural and movement-related difficulties. To date, the treatment of PD remains a challenge because available drugs only treat the symptoms of the disease or possess serious side effects. In light of this, new treatment options are needed; hence, this study investigates the neuroprotective effects of an organic Boophone haemanthoides extract (BHE) and its bioactive compounds using an in vitro model of PD involving the toxin 1-methyl-4-phenylpyridinium (MPP+) and SH-SY5Y neuroblastoma cells. A total of seven compounds were isolated from BHE, viz distichamine (1), 1α,3α-diacetylnerbowdine (2), hippadine (3), stigmast-4-ene-3,6-dione (4), cholest-4-en-3-one (5), tyrosol (6), and 3-hydroxy-1-(4'-hydroxyphenyl)-1-propanone (7). Six compounds (1, 2, 4, 5, 6 and 7) were investigated, and five showed neuroprotection alongside the BHE. This study gives insight into the bioactivity of the non-alkaloidal constituents of Amaryllidaceae, since the isolated compounds and the BHE showed improved cell viability, increased ATP generation in the cells as well as inhibition of MPP+-induced apoptosis. Together, these findings support the claim that the Amaryllidaceae plant family could be a potential reserve of bioactive compounds for the discovery of neuroprotective agents.

4-Oxatricyclo[5.2.1.02,6]dec-8-ene-3,5-dione Derivatives as NMDA Receptor- and VGCC Blockers with Neuroprotective Potential.

The impact of excitotoxicity mediated by N-methyl-D-aspartate (NMDA) receptor overactivation and voltage gated calcium channel (VGCC) depolarization is prominent among the postulated processes involved in the development of neurodegenerative disorders. NGP1-01, a polycyclic amine, has been shown to be neuroprotective through modulation of the NMDA receptor and VGCC, and attenuation of MPP+-induced neurotoxicity. Recently, we reported on the calcium modulating effects of tricycloundecene derivatives, structurally similar to NGP1-01, on the NMDA receptor and VGCC of synaptoneurosomes. In the present study, we investigated novel 4-oxatricyclo[5.2.1.02,6]dec-8-ene-3,5-dione derivatives for their cytotoxicity, neuroprotective effects via attenuation of MPP+-induced neurotoxicity and calcium influx inhibition abilities through the NMDA receptor and VGCC using neuroblastoma SH-SY5Y cells. All compounds, in general, showed low or no toxicity against neuroblastoma cells at 10-50 µM concentrations. At 10 µM, all compounds significantly attenuated MPP+-induced neurotoxicity as evident by the enhancement in cell viability between 23.05 ± 3.45% to 53.56 ± 9.29%. In comparison to known active compounds, the derivatives demonstrated mono or dual calcium modulating effect on the NMDA receptor and/or VGCC. Molecular docking studies using the NMDA receptor protein structure indicated that the compounds are able to bind in a comparable manner to the crystallographic pose of MK-801 inside the NMDA ion channel. The biological characteristics, together with results from in silico studies, suggest that these compounds could act as neuroprotective agents for the purpose of halting or slowing down the degenerative processes in neuronal cells.

Open and rearranged norbornane derived polycyclic cage molecules as potential neuroprotective agents through attenuation of MPP+- and calcium overload-induced excitotoxicity in neuroblastoma SH-SY5Y cells.

The neuroprotective effects of closed polycyclic cage molecules such as NGP1-01, memantine and amantadine have been extensively explored. These effects are mostly linked to the antagonism of the N-methyl-d-aspartate (NMDA) receptor- and the blockage of voltage gated calcium channels (VGCC). The synthesis of structurally related open and rearranged cage derivatives has been studied in depth. However, very little is known on their neuroprotective effects. In this study, a series of open and rearranged polycyclic cage molecules containing a norbornane derived scaffold were synthesised and evaluated for cytotoxicity, neuroprotection and calcium blocking effects via the NMDA receptor and VGCC on neuroblastoma cells at a 10 µM concentration. All compounds showed negligible cytotoxicity and were able to significantly attenuate MPP+-induced neurotoxicity between 26.07 ± 12.50% to 48.42 ± 0.76%, with compound 14 showing the best neuroprotective effect. In comparison to known NMDA receptor antagonists, all compounds demonstrated moderate to excellent calcium blocking effects of 26.50 ± 2.28 to 72.95 ± 3.38%. Docking studies suggest that these compounds are able to show significant NMDA receptor channel blocking ability since they bind in a comparable manner to the crystallographic pose of MK-801 inside the NMDAR ion channel. Some compounds were also able to attenuate calcium influx through VGCC channels between 21.28 ± 3.69% to 50.34 ± 7.67%. Compound 4 and 15 showed the highest inhibition of calcium influx at the VGCC and NMDA receptor, respectively. The compounds exhibiting good cytotoxicity-, neuroprotective- and calcium blocking profiles could potentially act as neuroprotective agents to clinically benefit people suffering from neurodegenerative disorders.

Differential Sensitivity of Two Endothelial Cell Lines to Hydrogen Peroxide Toxicity: Relevance for In Vitro Studies of the Blood-Brain Barrier.

Oxidative stress (OS) has been linked to blood-brain barrier (BBB) dysfunction which in turn has been implicated in the initiation and propagation of some neurological diseases. In this study, we profiled, for the first time, two endothelioma cell lines of mouse brain origin, commonly used as in vitro models of the blood-brain barrier, for their resistance against oxidative stress using viability measures and glutathione contents as markers. OS was induced by exposing cultured cells to varying concentrations of hydrogen peroxide and fluorescence microscopy/spectrometry was used to detect and estimate cellular glutathione contents. A colorimetric viability assay was used to determine changes in the viability of OS-exposed cells. Both the b.End5 and bEnd.3 cell lines investigated showed demonstrable content of glutathione with a statistically insignificant difference in glutathione quantity per unit cell, but with a statistically significant higher capacity for the b.End5 cell line for de novo glutathione synthesis. Furthermore, the b.End5 cells demonstrated greater oxidant buffering capacity to higher concentrations of hydrogen peroxide than the bEnd.3 cells. We concluded that mouse brain endothelial cells, derived from different types of cell lines, differ enormously in their antioxidant characteristics. We hereby recommend caution in making comparisons across BBB models utilizing distinctly different cell lines and require further prerequisites to ensure that in vitro BBB models involving these cell lines are reliable and reproducible.

Attenuation of Endoplasmic Reticulum Stress, Impaired Calcium Homeostasis, and Altered Bioenergetic Functions in MPP+-Exposed SH-SY5Y Cells Pretreated with Rutin.

Parkinson's disease (PD) is a common neurodegenerative disorder that affects approximately 1% of the population over the age of 65 years. While treatment options for PD are limited, reports show that plant-derived bioactive compounds such as rutin possess numerous pharmacological benefits, including antioxidant and antiapoptotic activities. This study aimed to investigate the potential role of rutin in MPP+-treated SH-SY5Y neuroblastoma cells, an established cell model of PD. Our findings reveal increased concentrations of Ca2+ and endoplasmic reticulum (ER) stress as well as impaired mitochondrial membrane potential and bioenergetic status in SH-SY5Y cells treated with MPP+ only. This is demonstrated by a significant reduction in the expression levels of BiP, significantly reduced basal respiration, maximal respiration, and spare respiratory capacity as well as a significant increase in the expression levels of CHOP; however, these effects were significantly attenuated following pretreatment with rutin. Also, rutin significantly improved basal and compensatory glycolysis as a response to an impaired oxidative phosphorylation system triggered by MPP+, characterized by deficient ATP production. In conclusion, our findings provide the first evidence on the ability of rutin to maintain Ca2+ homeostasis, inhibit ER stress, and protect the mitochondria in MPP+-treated SH-SY5Y cells.

GRP78/BIP/HSPA5 as a Therapeutic Target in Models of Parkinson's Disease: A Mini Review.

Parkinson's disease (PD) is a common neurodegenerative disorder characterized by selective loss of dopamine neurons in the substantia nigra pars compacta of the midbrain. Reports from postmortem studies in the human PD brain, and experimental PD models reveal that endoplasmic reticulum (ER) stress is implicated in the pathogenesis of PD. In times of stress, the unfolded or misfolded proteins overload the folding capacity of the ER to induce a condition generally known as ER stress. During ER stress, cells activate the unfolded protein response (UPR) to handle increasing amounts of abnormal proteins, and recent evidence has demonstrated the activation of the ER chaperone GRP78/BiP (78 kDa glucose-regulated protein/binding immunoglobulin protein), which is important for proper folding of newly synthesized and partly folded proteins to maintain protein homeostasis. Although the activation of this protein is essential for the initiation of the UPR in PD, there are inconsistent reports on its expression in various PD models. Consequently, this review article aims to summarize current knowledge on neuroprotective agents targeting the expression of GRP78/BiP in the regulation of ER stress in experimental PD models.

Design, synthesis and evaluation of pentacycloundecane and hexacycloundecane propargylamine derivatives as multifunctional neuroprotective agents.

The multifactorial pathophysiology of neurodegenerative disorders remains one of the main challenges in the design of a single molecule that may ultimately prevent the progression of these disorders in affected patients. In this article, we report on twelve novel polycyclic amine cage derivatives, synthesized with or without a propargylamine function, designed to possess inherent multifunctional neuroprotective activity. The MTT cytotoxicity assay results showed the SH-SY5Y human neuroblastoma cells to be viable with the twelve compounds, particularly at concentrations less than 10 µM. The compounds also showed significant neuroprotective activity, ranging from 31% to 61% at 1 µM, when assayed on SH-SY5Y human neuroblastoma cells in which neurodegeneration was induced by MPP+. Calcium regulation assays conducted on the same cell line showed the compounds to be significant VGCC blockers with activity ranging from 26.6% to 51.3% at 10 µM; as well as significant NMDAr antagonists with compound 5 showing the best activity of 88.3% at 10 µM. When assayed on human MAO isoenzymes, most of the compounds showed significant inhibitory activity, with compound 5 showing the best activity (MAO-B: IC50 = 1.70 µM). Generally, the compounds were about 3-52 times more selective to the MAO-B isoenzyme than the MAO-A isoenzyme. Based on the time-dependency studies conducted, the compounds can be defined as reversible MAO inhibitors. Several structure activity relationships were derived from the various assays conducted, and the compounds' possible putative binding modes within the MAO-B enzyme cavity were assessed in silico.

Design, synthesis and evaluation of indole derivatives as multifunctional agents against Alzheimer's disease.

A series of indole derivatives was designed and synthesised to improve on activity and circumvent pharmacokinetic limitations experienced with the structurally related compound, ladostigil. The compounds consisted of a propargylamine moiety (a known MAO inhibitor and neuroprotector) at the N1 position and a ChE inhibiting diethyl-carbamate/urea moiety at the 5 or 6 position of the indole ring. In order to prevent or slow down the in vivo hydrolysis and deactivation associated with the carbamate function of ladostigil, a urea moeity was incorporated into selected compounds to obtain more metabolically stable structures. The majority of the synthesised compounds showed improved MAO-A inhibitory activity compared to ladostigil. The compounds possessing the propargylamine moiety showed good MAO-B inhibitory activity with 6 and 8 portraying IC50 values between 14-20 fold better than ladostigil. The ChE assay results indicated that the compounds have non-selective inhibitory activities on eeAChE and eqBuChE regardless of the type or position of substitution (IC50: 2-5 µM). MAO-A and MAO-B docking results showed that the propargylamine moiety was positioned in close proximity to the FAD cofactor suggesting that the good inhibitory activity may be attributed to the propargylamine moiety and irreversible inhibition as confirmed in the reversibility studies. Docking results also indicated that the compounds have interactions with important amino acids in the AChE and BuChE catalytic sites. Compound 6 was the most potent multifunctional agent showing better inhibitory activity than ladostigil in vitro on all enzymes tested (hMAO-A IC50 = 4.31 µM, hMAO-B IC50 = 2.62 µM, eeAChE IC50 = 3.70 µM, eqBuChE IC50 = 2.82 µM). Chemical stability tests confirmed the diethyl-urea containing compound 6 to be more stable than its diethyl-carbamate containing counterpart compound 8. Compound 6 also exerted significant neuroprotection (52.62% at 1 µM) against MPP+ insult to SH-SY5Y neural cells and has good in silico predicted ADMET properties. The favourable neuronal enzyme inhibitory activity, likely improved pharmacokinetic properties in vivo and the potent neuroprotective ability of compound 6 make it a promising compound for further development.