Radical Scavenging Is Not Involved in Thymoquinone-Induced Cell Protection in Neural Oxidative Stress Models.

Thymoquinone (TQ), an active compound from Nigella sativa seeds, is often described as a pharmacologically relevant compound with antioxidative properties, while the synthesis of TQ in the plant via oxidations makes it inapplicable for scavenging radicals. Therefore, the present study was designed to reassess the radical scavenging properties of TQ and explore a potential mode of action. The effects of TQ were studied in models with mitochondrial impairment and oxidative stress induced by rotenone in N18TG2 neuroblastoma cells and rotenone/MPP+ in primary mesencephalic cells. Tyrosine hydroxylase staining revealed that TQ significantly protected dopaminergic neurons and preserved their morphology under oxidative stress conditions. Quantification of the formation of superoxide radicals via electron paramagnetic resonance showed an initial increase in the level of superoxide radicals in the cell by TQ. Measurements in both cell culture systems revealed that the mitochondrial membrane potential was tendentially lowered, while ATP production was mostly unaffected. Additionally, the total ROS levels were unaltered. In mesencephalic cell culture under oxidative stress conditions, caspase-3 activity was decreased when TQ was administered. On the contrary, TQ itself tremendously increased the caspase-3 activity in the neuroblastoma cell line. Evaluation of the glutathione level revealed an increased level of total glutathione in both cell culture systems. Therefore, the enhanced resistance against oxidative stress in primary cell culture might be a consequence of a lowered caspase-3 activity combined with an increased pool of reduced glutathione. The described anti-cancer ability of TQ might be a result of the pro-apoptotic condition in neuroblastoma cells. Our study provides evidence that TQ has no direct scavenging effect on superoxide radicals.

From the tyrosine hydroxylase hypothesis of Parkinson's disease to modern strategies: a short historical overview.

A time span of 60 years covers the detection of catecholamines in the brain, their function in movement and correlation to Parkinson's disease (PD). The clinical findings that orally given L-DOPA can alleviate or even prevent akinesia gave great hope for the treatment of PD. Attention focused on the role of tyrosine hydroxylase (TH) as the rate-limiting enzyme in the formation of catecholamines. It became evident that the enzyme driven formation is lowered in PD. Such results could only be obtained from studying human brain samples demonstrating the necessity for human brain banks. Originally, a TH enzyme deficiency was suspected in PD. Studies were conducted on the enzyme properties: its induction and turnover, the complex regulation starting with cofactor requirements as tetrahydrobiopterin and ferrous iron, and the necessity for phosphorylation for activity as well as inhibition by toxins or regulatory feedback inhibition by catecholamines. In the course of time, it became evident that neurodegeneration and cell death of dopaminergic neurons is the actual pathological process and the decrease of TH a cophenomenon. Nevertheless, TH immunochemistry has ever since been a valuable tool to study neuronal pathways, neurodegeneration in various animal models of neurotoxicity and cell cultures, which have been used as well to test potential neuroprotective strategies.

The Reassessed Impact of Nicotine against Neurotoxicity in Mesencephalic Dopaminergic Cell Cultures and Neuroblastoma N18TG2 Cells.

Neuroprotective effects of nicotine are still under debate, so further studies on its effectiveness against Parkinson's disease are required. In our present study, we used primary dopaminergic cell cultures and N18TG2 neuroblastoma cells to investigate the effect of nicotine and its neuroprotective potential against rotenone toxicity. Nicotine protected dopaminergic (tyrosine hydroxylase immunoreactive) neurons against rotenone. This effect was not nAChR receptor-dependent. Moreover, the alkaloid at a concentration of 5 µM caused an increase in neurite length, and at a concentration of 500 µM, it caused an increase in neurite count in dopaminergic cells exposed to rotenone. Nicotine alone was not toxic in either cell culture model, while the highest tested concentration of nicotine (500 µM) caused growth inhibition of N18TG2 neuroblastoma cells. Nicotine alone increased the level of glutathione in both cell cultures and also in rotenone-treated neuroblastoma cells. The obtained results may be helpful to explain the potential neuroprotective action of nicotine on neural cell cultures.

Autophagy inhibitor 3-methyladenine could not modulate rotenone neurotoxicity in primary mesencephalic cell culture / El inhibidor de la autofagia 3-metiladenina no logra modular la neurotoxicidad de la rotenona en el cultivo primario de células mesencefálicas

Dysregulated autophagy, whether excessive or downregulated, has been thought to be associated with neurodegenerative disorders including Parkinson's disease. Accordingly, the present study was carried out to investigate whether 3-methyladenine, an autophagy inhibitor, can modulate the effects of rotenone on dopaminergic neurons in primary mesencephalic cell culture. Cultures were prepared from embryonic mouse mesencephala at gestation day 14. Four groups of cultures were treated on the 10th DIV for 48 h as follows: the first was kept as an untreated control, the second was treated with 3-methyladenine alone (1, 10, 100, 200 mM), the third was treated with 20 nM rotenone and the fourth was co-treated with 20 nM rotenone and 3-methyladenine (1, 10, 100, 200 mM). On the 12th DIV, cultured cells were stained immunohistochemically against tyrosine hydroxylase and culture media were used to measure the levels of lactate dehydrogenase. 3methyladenine had no effects on both the survival of dopaminergic neurons and the release of lactate dehydrogenase. Rotenone significantly decreased the number of dopaminergic neurons and increased the levels of lactate dehydrogenase in the culture media. When cultures concomitantly treated with 3-methyladenine and rotenone, 3-methyladenine had no effect against rotenone-induced dopaminergic cell damage and lactate dehydrogenase release into the culture medium. In conclusion, the autophagy inhibitor 3-methyladenine could not modulate rotenone-induced dopaminergic cell damage in primary mesencephalic cell culture.

Se estima que la autofagia desregulada, ya sea excesiva o con baja regulación, está asociada con trastornos neurodegenerativos, incluyendo la enfermedad de Parkinson. En consecuencia, el se realizó este estudio para investigar si la 3metiladenina, un inhibidor de la autofagia,puede modular los efectos de la rotenona en las neuronas dopaminérgicas en el cultivo primario de células mesencefálicas. Los cultivos se prepararon a partir de mesencéfalo de ratón embrionario el día 14 de gestación. Cuatro grupos de cultivos se trataron en el 10º DIV durante 48 h de la siguiente manera: el primer grupo se mantuvo como un control no tratado, el segundo se trató con 3-metiladenina sola (1, 10, 100, 200 mM), el tercer grupo se trató con rotenona 20 nM y el cuarto se trató conjuntamente con rotenona 20 nM y 3-metiladenina (1, 10, 100, 200 mM). En el 12º DIV; las células cultivadas fueron tratadas mediante tinción inmunohistoquímica en tirosina hidroxilasa y se usaron medios de cultivo para medir los niveles de lactato deshidrogenasa. La 3-metiladenina no tuvo efectos tanto en la supervivencia de las neuronas dopaminérgicas como en la liberación de lactato deshidrogenasa. La rotenona disminuyó significativamente el número de neuronas dopaminérgicas y se observó un aumento de los niveles de lactato deshidrogenasa en los medios de cultivo. Cuando los cultivos tratados concomitantemente con 3-metiladenina y rotenona, la 3metiladenina no tuvo efecto contra el daño celular dopaminérgico inducido por la rotenona y la liberación de lactato deshidrogenasa en el medio de cultivo. En conclusión, el inhibidor de la autofagia 3-metiladenina no moduló el daño celular dopaminérgico inducido por la rotenona en el cultivo celular mesencefálico primario.

Minocycline protects against acrylamide-induced neurotoxicity and testicular damage in Sprague-Dawley rats.

This study investigated the protective effects of minocycline against acrylamide (ACR)-induced neurotoxicity and testicular damage in Sprague-Dawley rats. Forty rats were divided into five groups (eight rats each). Group I received saline (0.5 mL/rat) daily for 10 days and served as the untreated control group. Group II received ACR (30 mg/kg body weight (b.w.)) daily for 10 days. Group III received ACR (30 mg/kg b.w.) daily for 10 days and subsequently minocycline (60 mg/kg b.w.) for five days. Group IV received ACR (30 mg/kg b.w.) daily for 10 days followed by saline for five days and served as the control group for the ACR-minocycline-treated group. Group V received minocycline (60 mg/kg b.w.) for five days. All treatments were administered orally. Rats in group I and V showed normal locomotor behavior and normal histology of the brain and testes. Administration of ACR (Group II and IV) resulted in weight loss and gait abnormalities. Furthermore, neuronal degeneration in the hippocampus and cerebellum and degeneration of the seminiferous tubular epithelium with formation of spermatid giant cells were observed. Ultrastructurally, ACR specifically damaged spermatogonia and spermatocytes. Acrylamide was also seen to cause a significant increase of malondialdehyde levels in the brain and testes. Treatment of ACR-administered rats with minocycline (Group III) significantly alleviated the loss of body weight and improved locomotor function. Minocycline also ameliorated neuronal degeneration and seminiferous tubular damage and decreased malondialdehyde concentrations. In conclusion, minocycline protects against neurotoxic effects of acrylamide and seminiferous tubular damage. Decreasing lipid peroxidation by minocycline might play a role in such protection.

Ultrastructural changes of extraocular muscles in strabismus patients.

Strabismus is an ocular disorder characterized by partial or complete inability to keep eye alignment. It represents a very common ocular problem at ophthalmology clinics worldwide. The current study aimed to show the most encountered ultrastructural changes in extraocular muscles (EOMs) collected from patients with different forms of strabismus. Nine specimens of EOMs were collected from five patients during strabismus correction surgery and processed for light and electron microscopy examinations. Histologically, skeletal muscle fibers in normal EOMs appeared tight and normally arranged with clear striations. In strabismic muscles, the fibers appeared disarranged, and atrophied, swollen and disintegrated in some situations. By transmission electron microscopy, normal EOMs were formed of skeletal muscle fibers with intact basal membrane and sarcolemma, tightly aligned myofibrils with well-arranged sarcomeres, Z line and H zone, and normally distributed mitochondria. On the other hand, strabismic EOMs revealed vacuolation and degeneration of myofibrils, accumulation of lipid droplets, subsarcolemmal inclusions and clustering of mitochondria. EOMs obtained from a Down syndrome patient with V-pattern infantile esotropia showed extensive vacuolation and disintegration of myofibrils, and extra- and intracellular deposition of collagen fibers. Interestingly, some skeletal muscle cells exhibited features of autophagic cell death with a trial of engulfing process by neighboring cells. In conclusion, our study traces some characteristic ultrastructural changes in strabismic EOMs, most notably, extensive vacuolation, clustering of mitochondria, degeneration of myofibrils and autophagic changes. These changes might be emphasized as possibly secondary to strabismus.

Neurotoxic effects of acrylamide on dopaminergic neurons in primary mesencephalic cell culture.

INTRODUCTION: Exposure to acrylamide is increasing worldwide as a result of its heavy use in industry and formation in carbohydrate-rich food cooked at high temperature. Despite its neurotoxicity, no studies have shown its toxic effects on dopaminergic neurons yet. Therefore, the current study was carried out to show whether acrylamide adversely affects primary cultured dopaminergic neurons. MATERIAL AND METHODS: Acrylamide (0.001, 0.01, 0.1, 1, 2 mM) was added to two different groups of primary mesencephalic cell cultures on the 9th day in vitro for 24 and 48 h, respectively. Moreover, a group of cultures was treated with lower concentrations of acrylamide (0.01, 0.05, 0.1, 0.5 mM) on the 6th day in vitro for 5 consecutive days to investigate its long-term effects on dopaminergic neurons. Following each treatment, culture media were obtained for measuring lactate dehydrogenase, and cultured cells were stained immunocytochemically against tyrosine hydroxylase and neuronal nuclear antigens. RESULTS: Treatment of cultures with acrylamide for 48 h significantly reduced the number of dopaminergic neurons, adversely altered the morphology of the surviving neurons and increased levels of lactate dehydrogenase in the culture media. Similar treatment of cultures with acrylamide also resulted in lower numbers of total neuronal cells as shown by a reduced expression of the neuronal nuclear antigen. Prolonged treatment of cultures with lower concentrations of acrylamide slightly reduced the survival of dopaminergic neurons but increased the release of lactate dehydrogenase into the culture media as well. CONCLUSIONS: The current study shows, for the first time, neurotoxicity of acrylamide on dopaminergic neurons in the primary mesencephalic cell culture.

Rotenone: from modelling to implication in Parkinson's disease.

Rotenone ([2R-(2α,6aα,12aα)]-1,2,12,12a-tetrahydro-8,9-dimethoxy-2-(1-methylethenyl)-[1]benzopyran[3,4-b]furo [2,3-h][1]benzopyran-6(6aH)-one) is a naturally occurring compound derived from the roots and stems of Derris, Tephrosia, Lonchocarpus and Mundulea plant species. Since its discovery at the end of the 19th century, rotenone has been widely used as a pesticide for controlling insects, ticks and lice, and as a piscicide for management of nuisance fish in lakes and reservoirs. In 2000, Betarbet et al. reproduced most of the behavioural, biochemical and pathological features of Parkinson's disease (PD) in rotenone-treated rats. Since that time, rotenone has received much attention as it would be one of the environmental neurotoxins implicated in etiopathogenesis of PD. Moreover, it represents a common experimental model to investigate the underlying mechanisms leading to PD and evaluate the new potential therapies for the disease. In the current general review, we aimed to address recent advances in the hazards of the environmental applications of rotenone and discuss the updates on the rotenone model of PD and whether it is implicated in the etiopathogenesis of the disease.

Long-term neurotoxic effects of domoic acid on primary dopaminergic neurons.

Domoic acid, an excitatory neurotoxin produced by certain algae, reaches the food chain through accumulation in some sea organisms. To investigate its long-term neurotoxicity on dopaminergic neurons, prepared primary mesencephalic cell cultures were exposed to different concentrations of domoic acid (0.1, 1, 10, 100 µM) on the 8th day in vitro (DIV) for 4 days. On the 12th DIV, culture media were collected for measurement of lactate dehydrogenase and cultured cells were subjected to immunohistochemistry against tyrosine hydroxylase, neuronal nuclear antigen and glial fibrillary acidic protein, and fluorescence staining using H2DCFDA, JC-1 and Hoechst 33342 dyes. Moreover, roles of AMPA/KA and NMDA receptors in domoic acid neurotoxicity were also investigated. Domoic acid significantly decreased the number of dopaminergic neurons and adversely affected their morphology, and slightly reduced the expression of neuronal nuclear antigen and glial fibrillary acidic protein. Co-treatment of cultures with domoic acid and the AMPA/KA or NMDA receptor antagonists NBQX and MK-801 rescued significant number of dopaminergic neurons. Domoic acid significantly decreased red:green fluorescence ratio of JC-1 and did not affect production of reactive oxygen species and apoptotic cell death. In conclusions, the present study reveals that long-term treatment of primary mesencephalic cell culture with domoic acid significantly destroyed dopaminergic neurons. This effect appears to be attributed to activation of AMPA/KA and NMDA receptors and mitochondrial damage.

Radiation-induced damage to lacrimal glands: an ultrastructural study in Sprague Dawley rats.

Injury to lacrimal glands represents a major health problem after radiation therapy of the head and neck malignancies. Accordingly, this study aimed to investigate significant ultrastructural changes of lacrimal glands and some of their underlying mechanisms following the exposure to different fractionated doses of irradiation. In this study, 28 Sprague Dawley (SD) rats were assigned to four groups (seven rats each): Group I acted as control and received no irradiation. Groups II-IV received fractionated irradiation of 5 Gy (100 cGy/fraction daily for 5 days), 9 Gy (300 cGy/fraction daily for 3 days), and 20 Gy (one fraction), respectively. One month after the experiment, examination of lacrimal glands with transmission electron microscopy (TEM) demonstrated dose-dependent ultrastructural changes in the lacrimal acinar and intralobular ductal epithelial cells. In the acinar cells, there were swollen rough endoplasmic reticulum, irregularly shaped nuclei with chromatin condensation, mitochondrial damage, and retention of secretory granules. Intaralobular ductal epithelial cells showed loss of surface microvilli and damage to mitochondria. In addition to the potential direct effects of irradiation on lacrimal acinar and intralobular ductal epithelial cells, damage to blood vessels and nerve endings seemed to mediate some of the underlying mechanisms of these irradiation-induced ultrastructural changes. In conclusion, using TEM reveals that lacrimal gland is highly sensitive to even small doses of irradiation therapy; in addition, swelling of rough endoplasmic reticulum and aberrant nuclei are the most encountered structural changes. Damage to blood vessels and nerve endings might mediate some of the underlying mechanisms of irradiation-induced secondary injury in lacrimal glands.

Neurotoxic effects of domoic acid on dopaminergic neurons in primary mesencephalic cell culture.

INTRODUCTION: Domoic acid is a potent marine neurotoxin produced by certain species of the diatom genus Pseudonitzschia. To our knowledge, there are no studies that have investigated neurotoxic effects of domoic acid on dopaminergic neurons. Accordingly, the present study was carried out to investigate the potential neurotoxic effects of domoic acid on dopaminergic neurons in primary mesencephalic cell culture. MATERIAL AND METHODS: Cultures prepared from embryonic mouse mesencephala (total of 250 embryos) were treated with different concentrations of domoic acid (0.1, 1, 10, 100 µM) on the 10th DIV for 48 h. On the 12th DIV, culture media were used for measurement of lactate dehydrogenase and cultured cells were subjected to immunostaining for tyrosine hydroxylase, neuronal nuclear antigen and glial fibrillary acidic protein, and fluorescence staining using H2DCFDA, JC-1 and DAPI stains. Moreover, roles of AMPA/KA and NMDA receptors in domoic acid neurotoxicity were also investigated. RESULTS: Domoic acid significantly decreased the number of dopaminergic neurons, decreased the expression of neuronal nuclear antigen and slightly affected astrocyte populations, and increased the release of lactate dehydrogenase into the culture media. AMPA/KA receptor antagonist NBQX but not NMDA receptor antagonist MK-801 significantly inhibited the neurotoxic effect of domoic acid on dopaminergic neurons. H₂DCFDA, JC-1 and DAPI fluorescence staining, respectively, revealed that DomA slightly raised ROS production, and significantly decreased mitochondrial membrane potential and increased apoptotic cell death of cultured cells. CONCLUSION: The current study presents for the first time the neurotoxic effects of domoic acid on dopaminergic neurons and this effect appears to be attributed to activation of AMPA/KA receptors on dopaminergic neurons.

Ultrastructural changes induced by Solanum incanum aqueous extract on HCT 116 colon cancer cells.

Medicinal plants have recently gained increasing scientific interest as an important source of molecules with different therapeutic potentials. Accordingly, the present study was carried out to investigate ultrastructural changes induced by the aqueous extract of Solanum incanum (SI) fruit on human colorectal carcinoma cell line (HCT 116 cells). Examination of SI-treated HCT 116 cells with transmission electron microscopy (TEM) demonstrated numerous ultrastructural changes in the form of loss of the surface microvilli, mitochondrial damage and dilatation of cristae, and formation of autophagic vacuoles and increasing numbers of lipid droplets. Also, majority of the treated cells showed nuclear shrinkage with chromatin condensation and nucleolar changes. Moreover, some cells showed focal areas of cytoplasmic degeneration associating with formation of myelin figures and fatty globules. In conclusion, TEM was able to verify cytotoxicity of SI aqueous extract against HCT 116 colon cancer cells.

Ultrastructural changes of kidney in Schistosoma mansoni-infected mice.

Schistosomiasis is the second threatening parasitic disease after malaria and among Schistosoma spp., Schistosoma mansoni (S. mansoni) affects about 100 million people in tropic regions in Africa and South America. The current study was carried out to investigate ultrastructural changes of the kidney in mice infected with cercariae of S. mansoni, in which 20 Swiss albino mice of 60-day-old were assigned into two groups (10 each). Control group received 1 ml normal saline by intraperitoneal route. Model group were intraperitoneally infected with 1 ml normal saline containing 40 cercariae of S. mansoni/mouse. After 60 days of infection, specimens from the kidneys of both control and infected mice were obtained and processed for transmission electron microscopy (TEM) examination. The main ultrastructural changes were observed in both glomeruli and tubules. Glomerular findings included irregular thickening and splitting of the glomerular basement membrane (GBM), flattening and effacement of the foot processes of podocytes, and proliferation of mesangial cells. Tubular changes were in the form of swelling, atrophy and vacuolation of tubular epithelial cells, and presence of autophagic vacuoles. In conclusion, adopting TEM shows a number of ultrastructural changes in the kidneys of mice infected with cercariae of S. mansoni, most notably thickening and splitting of GBM and flattening and effacement of foot processes of podocytes and tubular autophagic vacuoles. These changes are still unraveled well in the literature.

Recent Advances on the Role of Neurogenesis in the Adult Brain: Therapeutic Potential in Parkinson's and Alzheimer's Diseases.

BACKGROUND: Generation of nascent functional neurons from neural stem cells in the adult brain has recently become largely accepted by the neuroscience community. In adult mammals including humans, the process of neurogenesis has been well documented in two brain regions; the subventricular zone of the lateral ventricles and the subgranular zone in the dentate gyrus of the hippocampus. METHOD: Some evidence has indicated neurogenesis in other regions of the adult mammalian brain such as the neocortex, cerebellum, striatum, amygdala and hypothalamus. These discoveries question a long standing dogma on nervous system regeneration and provide medical science with potential new strategies to harness the process of neurogenesis for treating neurological disabilities and neurodegenerative diseases. CONCLUSION: In this current review, we address the most recent advances on the role of neurogenesis in the adult brain and therapeutic potential in the two most common neurodegenerative disorders, Parkinson's and Alzheimer's diseases.

THC (Δ9-Tetrahydrocannabinol) Exerts Neuroprotective Effect in Glutamate-affected Murine Primary Mesencephalic Cultures Through Restoring Mitochondrial Membrane Potential and Anti-apoptosis Involving CB1 Receptor-dependent Mechanism.

Aging-related neurodegenerative diseases, such as Parkinson's disease (PD) or related disorders, are an increasing societal and economic burden worldwide. Δ9-Tetrahydrocannabinol (THC) is discussed as a neuroprotective agent in several in vitro and in vivo models of brain injury. However, the mechanisms by which THC exhibits neuroprotective properties are not completely understood. In the present study, we investigated neuroprotective mechanisms of THC in glutamate-induced neurotoxicity in primary murine mesencephalic cultures, as a culture model for PD. Glutamate was administered for 48 h with or without concomitant THC treatment. Immunocytochemistry staining and resazurin assay were used to evaluate cell viability. Furthermore, superoxide levels, caspase-3 activity, and mitochondrial membrane potential were determined to explore the mode of action of this compound. THC protected dopaminergic neurons and other cell types of primary dissociated cultures from glutamate-induced neurotoxicity. Moreover, THC significantly counteracted the glutamate-induced mitochondrial membrane depolarization and apoptosis. SR141716A, a CB1 receptor antagonist, concentration-dependently blocked the protective effect of THC in primary mesencephalic cultures. In conclusion, THC exerts anti-apoptotic and restores mitochondrial membrane potential via a mechanism dependent on CB1 receptor. It strengthens the fact that THC has a benefit on degenerative cellular processes occurring, among others, in PD and other neurodegenerative diseases by slowing down the progression of neuronal cell death. Copyright © 2016 John Wiley & Sons, Ltd.

Comparable Neuroprotective Effects of Pergolide and Pramipexole on Ferrous Sulfate-Induced Dopaminergic Cell Death in Cell Culture.

BACKGROUND: Dopamine agonists are utilized clinically as an initial treatment in younger Parkinson's disease patients to delay the side effects associated with commencement of levodopa medication. These agonists also serveas adjunctive therapeutics with levodopa to lower the incidence of adverse motor symptoms in advanced stages of the disease. OBJECTIVES: To compare the neuroprotective effects of the dopamine agonists pergolide and pramipexole on ferrous sulfate-induced neurotoxicity in dopaminergic neurons from primary mesencephalic cell culture. METHODS: Pergolide (0.001-1 µM) and pramipexole (0.01-200 µM) were administered to 8 day primary murine mesencephalic cultures for 24 h. in the presence or absence of desferal, sulpiride or cycloheximide. Ferrous sulfate (450 µM) was then added for 24 hrs. Lactate dehydrogenase was assayed in the supernatant, glutathione concentrations measured in cell lysates and fixed cells were stained for tyrosine hydroxylase. RESULTS: Ferrous sulphate induced neurotoxity in cultures (p<0.0001) was abolished in the presence of the iron chelator desferal (p<0.008). Both pergolide (p<0.0001) and pramipexole (p<0.0001) significantly protected dopaminergic neurons against ferrous sulfate induced neurotoxicity and pramipexole helped preserve neurite morphology. Pramipexole treatment significantly reduced lactate dehydrogenase release (p<0.0001) as a measure of cellular injury. The dopamine receptor antagonist sulpiride (p<0.0001) and the protein synthesis inhibitor cycloheximide (p<0.0001) reduced the neuroprotective effects of pergolide indicating the involvement receptor stimulation and de novo protein synthesis in pergolide-mediated neuroprotection. Pramipexole also significantly reversed the decrease in cellular glutathione concentrations induced by ferrous sulfate (p<0.001). CONCLUSION: Both pergolide and pramipexole protect dopaminergic neurons against the neurotoxicity of ferrous sulfate. Pergolide specifically protects dopaminergic neurons through activation of dopamine receptors and de novo protein synthesis whereas pramipexole shows an overall effect through an antioxidant mechanism.

Ultrastructural pathology of human liver in Rift Valley fever.

Rift Valley fever (RVF) is a zoonotic disease that primarily affects ruminant animals and can also cause fatal disease in humans. In the current report, we present the ultrastructural changes in the liver of a man aged 60 years who died from RVF in the Aseer Central Hospital, Abha, Saudi Arabia. The main hepatic changes by transmission electron microscopy included the presence of 95-115 nm electron-dense particles consistent with RVF virions, nuclear condensation, vacuolar degeneration, lipid droplet accumulation and mitochondrial damage and dilation. There were also viral inclusion bodies with electron-dense aggregates, dilation of intercellular spaces, damage of sinusoidal microvilli with widening of space of Disse, dilation of bile canaliculi and increasing number of phagolysosomes.

Ultrastructural Changes of the Smooth Muscle in Esophageal Atresia.

Esophageal atresia (EA) with or without tracheo-esophageal fistula (TEF) is a relatively rare congenital anomaly. Despite the advances in the management techniques and neonatal intensive care, esophageal dysmotility remains a very common problem following EA/TEF repair. Our current study aimed to describe the most significant ultrastructural changes of the smooth muscle cells (SMCs) trying to highlight some of the underlying mechanisms of esophageal dysmotility following EA/TEF repair. Twenty-three biopsies were obtained from the tip of the lower esophageal pouch (LEP) of 23 patients during primary repair of EA/TEF. Light microscopic examination was performed with hematoxylin and eosin (HE), and Van Gieson's stains. Ultrastructural examination was done using transmission electron microscopy (TEM). Histopathological examination showed distortion of smooth muscle layer and deposition of an abundant amount of fibrous tissue in-between smooth muscles. Using TEM, SMCs exhibited loss of the cell-to-cell adhesion, mitochondrial vacuolation, formation of myelin figures, and apoptotic fragmentation. There were also plasmalemmal projections and formation of ghost bodies. Interestingly, SMCs were found extending pseudopodia-like projections around adjacent collagen fibers. Engulfed collagen fibers by SMCs underwent degradation within autophagic vacuoles. Degeneration of SMCs and deposition of abundant extracellular collagen fibers are prominent pathological changes in LEP of EA/TEF. These changes might contribute to the pathogenesis of esophageal dysmotility in patients who have survived EA/TEF.

Rapamycin protects dopaminergic neurons against rotenone-induced cell death in primary mesencephalic cell culture.

INTRODUCTION: Parkinson's disease is the most common movement disorder, characterized by a progressive and extensive loss of dopaminergic neurons in the substantia nigra pars compacta and their terminals in the striatum. So far, only symptomatic treatment is available, and no cure or disease-modifying drugs exist. The present study was designed to investigate the neuroprotective effect of rapamycin, an autophagy inducer, on dopaminergic neurons against rotenone-induced cell death in primary mesencephalic cell culture. MATERIAL AND METHODS: Primary mesencephalic cell cultures were prepared from embryonic mouse mesencephala (OFI/SPF, Vienna, Austria) at gestation day 14. Four sets of cultures were treated as follows: one was run as an untreated control, a second one was treated with 20 nM rotenone on the 10th day in vitro (DIV) for 48 h, a third one was co-treated with 20 nM rotenone and rapamycin (1, 10, 100, 1000 nM) on the 10th DIV for 48 h, and a fourth one was treated with rapamycin alone (1, 10, 100, 1000 nM) on the 10th DIV for 48 h. On the 12th DIV, cultures were subjected to immunohistochemistry against tyrosine hydroxylase and to fluorescence staining using LysoTracker Deep Red, JC-1 and DAPI stains. RESULTS: Exposure of such cultures to 20 nM rotenone on the 10th DIV for 48 h reduced the number of dopaminergic neurons by 41% and increased the release of lactate dehydrogenase (LDH) by 178% above untreated controls. Rapamycin (1, 10, 100, 1000 nM) added together with rotenone from the 10th to 12th DIV spared dopaminergic neurons by 17% and reduced the release of LDH by 64% at the concentration of 100 nM compared to rotenone-treated cultures. Activation of an autophagic process by rapamycin was demonstrated by LysoTracker Deep Red fluorescent dye, as indicated by a shift to increased red fluorescence. Rapamycin also significantly elevated the mitochondrial membrane potential (Δψm), as shown by an increase of the red:green fluorescence ratio of JC-1. Increased apoptotic cell death due to rotenone was lowered by rapamycin, as shown by the blue-fluorescent DAPI nucleic acid stain. CONCLUSIONS: Our study indicates for the first time that rapamycin, known as an autophagy inducer, protected dopaminergic neurons against rotenone-induced cell death in primary mesencephalic cell culture.

Recent advances in autophagy-based neuroprotection.

Macroautophagy is a highly regulated intracellular process that, under certain circumstances, delivers cytoplasmic components to the lysosomes for degradation. It consists of several sequential steps including initiation and nucleation, double membrane formation and elongation, formation and maturation of autophagosomes and finally autophagosomes/lysosomes fusion and degradation of intra-autophagosomal contents by lysosomal enzymes. After decades of considering autophagy as a cell death pathway, it has recently been shown to have a survival function through clearing of protein aggregates and damaged cytoplasmic organelles in response to a variety of stress conditions. Most recently, there is increasing evidence from literature revealing that autophagy induction may combat neurodegeneration. In the light of this, our current review tried to address the recent advances in the role of induced autophagy in neuroprotection with a particular focus on its contribution in the most common neurodegenerative disorders like Alzheimer's disease, Parkinson's disease and Huntington's disease.