Biomedical subjects

D Offen

Publications and source records attributed to D Offen.

At least 19 recordsLinked to original sources

PubMed2005

Increased survival and migration of engrafted mesenchymal bone marrow stem cells in 6-hydroxydopamine-lesioned rodents.

Parkinson's disease is characterized by the loss of dopaminergic neurons in the substantia nigra. Attempted replacement of these neurons by stem cells has proved inconclusive. Bone marrow mesenchymal stem cells (MSC) are multipotent, differentiating into a variety of cells, including neuron-like cells. We used the 6-hydroxydopamine (6-OHDA) animal model of Parkinson's disease to assess migration and differentiation of transplanted MSC. We found in rodents that transplanted MSC survive better in the 6-OHDA-induced damaged hemisphere compared to the unlesioned side. Moreover, contralaterally engrafted MSC migrated through the corpus callosum to populate the striatum, thalamic nuclei and substantia nigra of the 6-OHDA-lesioned hemisphere. In conclusion, we demonstrate that 6-OHDA-induced damage increases the viability of transplanted MSC and attracts these cells from the opposite hemisphere.

Animals↗

PubMed2005

Integral therapeutic potential of bone marrow mesenchymal stem cells.

Bone marrow derived mesenchymal stem cells (MSC) are adult stem cells that reside within the bone marrow compartment. In the traditional developmental model, adult stem cells are able to differentiate only to the tissue in which they reside. Recent data have challenged the committed fate of the adult stem cells, presenting evidence for their multi-lineage differentiation potential. In addition, potential therapeutic benefits of MSC administration have been the main concern of much research, including clinical trials. These studies promote adult stem cell therapy by shedding some light on the therapeutic potential of MSC and their mechanism of action. Many doubts have found their way into MSC research. They question MSC potency and beneficial contribution. However, these obstacles should not arrest but set a challenge to MSC researchers to examine their achievements under a magnifying glass. Therapeutic benefits of MSC exogenous delivery do not run counter to its possible participation in endogenous repair. Several reports imply MSC involvement in physiological repair but no explicit data support this hypothesis. This review tries to put MSC research into perspective. Possible therapeutic applications of MSC therapy for damaged tissue replacement, tissue engineering and the underlying repair mechanisms will be discussed. In addition, reported data about MSC possible involvement in physiological multiple tissue repair, their homing to injury and site-specific differentiation will be presented.

Animals↗

PubMed2004

Polymorphism in candidate genes: implications for the risk and treatment of idiopathic Parkinson's disease.

Idiopathic Parkinson's disease (IPD) is a progressive neurodegenerative disorder for which no restorative or neuroprotective therapy is available. Interest has recently been directed to association studies on polymorphisms of various genes, mainly those related to dopamine metabolism and transport, and their effect on response to PD, which includes primarily levodopa and dopaminomimetics. Approximately 15-20% of patients with PD do not respond to levodopa, and the majority of those who do respond develop adverse fluctuations in motor response, primarily levodopa-induced dyskinesias. This review summarizes the influence of polymorphisms in various genes on the relative risk of IPD and on levodopa efficacy. It focuses on the importance of well-designed polymorphism studies that include large samples of patients with IPD and tightly matched controls and use identical methodologies. Valid data on such polymorphisms might increase the efficacy of levodopa, decrease its side effects, and reduce the occurrence of levodopa-induced dyskinesias. They might also provide a novel diagnostic tool for PD.

Humans↗

PubMed2001

Protective effect of insulin-like-growth-factor-1 against dopamine-induced neurotoxicity in human and rodent neuronal cultures: possible implications for Parkinson's disease.

Parkinson's disease (PD) is characterized by a progressive loss of 70-80% of dopaminergic (DA) neurons in the substantia nigra. High concentrations of DA were suggested to induce oxidative stress and selective neurodegeneration. We evaluated the effect of insulin-like-growth-factor-1 (IGF-1) on DA toxicity in neuronal cultures. IGF-1 (0.5 microg/ml) suppressed cell death induced by exposure to DA (0.3 mM) after 2 and 4 days, in a rat cerebellar culture. Similarly, IGF-1 (0.5 and 1.0 microg/ml) antagonized DA (0.125 and 0.250 mM) neurotoxicity in a human neuroblastoma cell line (SK-N-SH). Flowcytometric analysis of neuroblastoma cells treated with DA (0.5 mM) showed increased apoptosis, which was significantly reduced by IGF-1. The effect of IGF-1 was associated with increased Bcl-2 expression as indicated by flowcytometry and Western blot analysis. We suggest that IGF-1 possesses a neuroprotective effect against DA-induced toxicity, and may have a potential role in the treatment of PD.

Animals↗

PubMed2001

Glutaredoxin protects cerebellar granule neurons from dopamine-induced apoptosis by dual activation of the ras-phosphoinositide 3-kinase and jun n-terminal kinase pathways.

Glutaredoxin 2 (Grx2) from Escherichia coli protects cerebellar neurons from dopamine-induced apoptosis via nuclear factor kappa B (NF-kappaB) activation, which is mediated by the expression of redox factor-1 (Ref-1). An analysis of the mechanisms underlying Grx2 protective activity revealed dual activation of signal transduction pathways. Grx2 significantly activated the Ras/phosphoinositide 3-kinase/Akt/NF-kappaB cascade in parallel to the Jun N-terminal kinase (JNK)/AP1 cascade. Dopamine, in comparison, down-regulated both pathways. Treatment of neurons with Ref-1 antisense oligonucleotide reduced the ability of Grx2 to activate Akt and AP-1 but had no effect on the phosphorylation of JNK1/2, suggesting that Akt/NF-kappaB and AP-1 are regulated by Ref-1. Exposure of the neurons to JNK1/2 antisense oligonucleotide in the presence of Grx2 significantly reduced AP-1 and NF-kappaB DNA binding activities and abolished Grx2 protection. These results demonstrate that dual activation of Ras/phosphoinositide 3-kinase and AP-1 cascades, which are mediated by Ref-1, is an essential component of the Grx2 mechanism of action.

Animals↗

PubMed2001

Bax ablation prevents dopaminergic neurodegeneration in the 1-methyl- 4-phenyl-1,2,3,6-tetrahydropyridine mouse model of Parkinson's disease.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) damages dopaminergic neurons in the substantia nigra pars compacta (SNpc) as seen in Parkinson's disease. Here, we show that the pro-apoptotic protein Bax is highly expressed in the SNpc and that its ablation attenuates SNpc developmental neuronal apoptosis. In adult mice, there is an up-regulation of Bax in the SNpc after MPTP administration and a decrease in Bcl-2. These changes parallel MPTP-induced dopaminergic neurodegeneration. We also show that mutant mice lacking Bax are significantly more resistant to MPTP than their wild-type littermates. This study demonstrates that Bax plays a critical role in the MPTP neurotoxic process and suggests that targeting Bax may provide protective benefit in the treatment of Parkinson's disease.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗

PubMed2001

Glutaredoxin protects cerebellar granule neurons from dopamine-induced apoptosis by activating NF-kappa B via Ref-1.

The neurotransmitter dopamine (DA) induces apoptosis via its oxidative metabolites. This study shows that glutaredoxin 2 (Grx2) from Escherichia coli and human glutaredoxin could protect cerebellar granule neurons from DA-induced apoptosis. E. coli Grx2, which catalyzes glutathione-disulfide oxidoreduction via its -Cys-Pro-Tyr-Cys- active site, penetrates into cerebellar granule neurons and exerts its activity via NF-kappaB activation. Analysis of single and double cysteine to serine substitutions in the active site of Grx2 showed that both cysteine residues were essential for activity. Although DA significantly reduced NF-kappaB binding activity, Grx2 could stimulate the binding of NF-kappaB to DNA by: (i) translocating NF-kappaB from the cytoplasm to the nucleus after promoting the phosphorylation and degradation of I-kappaBalpha, and (ii) activating the binding of pre existing nuclear NF-kappaB. The DNA binding activity of NF-kappaB itself was essential for neuronal survival. Overexpression of I-kappaB dominant negative gene (I-kappaB-DeltaN) in granule neurons significantly reduced their viability, irrespective of the presence of Grx2. Ref-1 expression was down-regulated by DA but up-regulated by Grx2, while treatment of neurons with Ref-1 antisense oligonucleotide reduced the ability of Grx2 to activate NF-kappaB binding activity. These results show that Grx2 exerts its anti apoptotic activity through the activation of Ref-1, which then activates NF-kappaB.

Amino Acid Substitution↗

PubMed2001

Oxidative stress induced-neurodegenerative diseases: the need for antioxidants that penetrate the blood brain barrier.

Oxidative stress (OS) has been implicated in the pathophysiology of many neurological, particularly neurodegenerative diseases. OS can cause cellular damage and subsequent cell death because the reactive oxygen species (ROS) oxidize vital cellular components such as lipids, proteins, and DNA. Moreover, the brain is exposed throughout life to excitatory amino acids (such as glutamate), whose metabolism produces ROS, thereby promoting excitotoxicity. Antioxidant defense mechanisms include removal of O(2), scavenging of reactive oxygen/nitrogen species or their precursors, inhibition of ROS formation, binding of metal ions needed for the catalysis of ROS generation and up-regulation of endogenous antioxidant defenses. However, since our endogenous antioxidant defenses are not always completely effective, and since exposure to damaging environmental factors is increasing, it seems reasonable to propose that exogenous antioxidants could be very effective in diminishing the cumulative effects of oxidative damage. Antioxidants of widely varying chemical structures have been investigated as potential therapeutic agents. However, the therapeutic use of most of these compounds is limited since they do not cross the blood brain barrier (BBB). Although a few of them have shown limited efficiency in animal models or in small clinical studies, none of the currently available antioxidants have proven efficacious in a large-scale controlled study. Therefore, any novel antioxidant molecules designed as potential neuroprotective treatment in acute or chronic neurological disorders should have the mandatory prerequisite that they can cross the BBB after systemic administration.

Animals↗

PubMed2001

6-Hydroxydopamine increases ubiquitin-conjugates and protein degradation: implications for the pathogenesis of Parkinson's disease.

One of the hallmarks of Parkinson's disease (PD) is pathological structure, termed Lewy body, containing inclusions of ubiquitinated proteins in the dopaminergic neurons in the substantia nigra. The mechanism leading to the formation of these aggregates is unclear, although it has been shown that mutations in alpha-synuclein or in the ubiquitin-related enzyme UCH-L1 might induce such protein aggregation. We, therefore, examined the possible role of 6-hydroxydopamine (6-OHDA), a dopaminergic neurotoxin used in PD experimental models, in causing protein degradation and its association with the ubiquitin system. Using antiubiquitin antibodies we found that exposure of SH-SY5Y neuroblastoma and PC-12 cell lines to 6-OHDA increased the levels of free ubiquitin and ubiquitin-conjugated proteins, in a dose-dependent manner. Furthermore, metabolic labeling with 35S-methionine, demonstrated that 6-OHDA markedly increased protein degradation, as indicated by the secretion of protein metabolites to the medium. Inhibition of the proteasome activity by the specific inhibitor MG132, attenuated the protein degradation induced by 6-OHDA and potentiated its toxicity. Administration of the antioxidant N-acetylcysteine to the 6-OHDA-treated cells, increased cell survival and reduced protein degradation. In conclusion, our findings suggest that 6-OHDA toxicity is associated with protein degradation and ubiquitin-proteasome system activation.

Acetylcysteine↗

PubMed2001

Activation of nuclear transcription factor kappa B (NF-kappaB) is essential for dopamine-induced apoptosis in PC12 cells.

The etiology of Parkinson's disease is still unknown, though current investigations support the notion of the pivotal involvement of oxidative stress in the process of neurodegeneration in the substantia nigra (SN). In the present study, we investigated the molecular mechanisms underlying cellular response to a challenge by dopamine, one of the local oxidative stressors in the SN. Based on studies showing that nuclear factor kappa B (NF-kappaB) is activated by oxidative stress, we studied the involvement of NF-kappaB in the toxicity of PC12 cells following dopamine exposure. We found that dopamine (0.1-0.5 m M) treatment increased the phosphorylation of the IkappaB protein, the inhibitory subunit of NF-kappaB in the cytoplasm. Immunoblot analysis demonstrated the presence of NF-kappaB-p65 protein in the nuclear fraction and its disappearance from the cytoplasmic fraction after 2 h of dopamine exposure. Dopamine-induced NF-kappaB activation was also evidenced by electromobility shift assay using radioactive labeled NF-kappaB consensus DNA sequence. Cell-permeable NF-kappaB inhibitor SN-50 rescued the cells from dopamine-induced apoptosis and showed the importance of NF-kappaB activation to the induction of apoptosis. Furthermore, flow cytometry assay demonstrated a higher level of translocated NF-kappaB-p65 in the apoptotic nuclei than in the unaffected nuclei. In conclusion, our findings suggest that NF-kappaB activation is essential to dopamine-induced apoptosis in PC12 cells and it may be involved in nigral neurodegeneration in patients with Parkinson's disease.

Animals↗

PubMed2000

Vasoactive intestinal peptide (VIP) prevents neurotoxicity in neuronal cultures: relevance to neuroprotection in Parkinson's disease.

Vasoactive intestinal peptide (VIP) provides neuroprotection against beta-amyloid toxicity in models of Alzheimer's disease. A superactive analogue, stearyl-Nle17-VIP (SNV) is a 100-fold more potent than VIP. In primary neuronal cultures, VIP protective activity may be mediated by femtomolar-acting glial proteins such as activity-dependent neurotrophic factor (ADNF), activity-dependent neuroprotective protein (ADNP), peptide derivatives ADNF-9 (9aa) and NAP (8aa), respectively. It has been hypothesized that beta-amyloid induces oxidative stress leading to neuronal cell death. Similarly, dopamine and its oxidation products were suggested to trigger dopaminergic nigral cell death in Parkinson's disease. We now examined the possible protective effects of VIP against toxicity of dopamine, 6-hydroxydopamine (6-OHDA) and 1-methyl-4-phenylpyridinium ion (MPP+) in neuronal cultures [rat pheochromocytoma (PC12), human neuroblastoma (SH-SY5Y) and rat cerebellar granular cells]. Remarkably low concentrations of VIP (10(-16)-10(-8) M), ADNF-9 and NAP (10(-18)-10(-10) M) protected against dopamine and 6-OHDA toxicity in PC12 and neuroblastoma cells. VIP (10(-11)-10(-9) M) and SNV (10(-13)-10(-11) M), protected cerebellar granule neurons against 6-OHDA. In contrast, VIP did not rescue neurons from death associated with MPP+. Since dopamine toxicity is linked to the red/ ox state of the cellular glutathione, we investigated neuroprotection in cells depleted of reduced glutathione (GSH). Buthionine sulfoximine (BSO), a selective inhibitor of glutathione synthesis, caused a marked reduction in GSH in neuroblastoma cells and their viability decreased by 70-90%. VIP, SNV or NAP (over a wide concentration range) provided significant neuroprotection against BSO toxicity. These results show that the mechanism of neuroprotection by VIP/SNV/NAP may be mediated through raising cellular resistance against oxidative stress. Our data suggest these compounds as potential lead compounds for protective therapies against Parkinson's disease.

1-Methyl-4-phenylpyridinium↗

PubMed2000

Apoptosis as a general cell death pathway in neurodegenerative diseases.

Neurodegenerative processes are generally characterized by the long-lasting course of neuronal death and the selectivity of the neuronal population or brain structure involved in the lesion. Two main common forms of cell death that have been described in neurons as in other vertebrate tissues i.e., necrosis and apoptosis. Necrosis is the result of cellular "accidents", such as those occurring in tissues subjected to chemical trauma. The necrotizing cells swell, rupture and provoke an inflammatory response. Apoptosis, on the other hand, is dependent on the cell's "decision" to commit suicide and die, and therefore is referred to as "programmed cell death" (PCD). The course of apoptotic death is characterized by a massive morphological change, including cell shrinkage, nuclear (chromosome) condensation and DNA degradation. Activation of PCD in an individual cell is based on its own internal metabolism, environment, developmental background and its genetic information. Such a situation occurs in most of the neurodegenerative disorders such as Alzheimer's, Parkinson's and Huntington's diseases and amyotrophic lateral sclerosis (ALS). In these pathological situations, specific neurons undergo apoptotic cell death characterized by DNA fragmentation, increased levels of pro-apoptotic genes and "apoptotic proteins" both, in human brain and in experimental models. It is of utmost importance to conclusively determine the mode of cell death in neurodegenerative diseases, because new "anti-apoptotic" compounds may offer a means of protecting neurons from cell death and of slowing the rate of cell degeneration and illness progression.

Alzheimer Disease↗

PubMed2000

The molecular mechanism of dopamine-induced apoptosis: identification and characterization of genes that mediate dopamine toxicity.

Parkinson's disease (PD) is a progressive neurological disorder caused by rather selective degeneration of the dopaminergic (DA) neurons in the substantia nigra. Though subject to intensive research, the etiology of this nigral neuronal loss is still enigmatic and treatment is basically symptomatic. The current major hypothesis suggests that nigral neuronal death in PD is due to excessive oxidative stress generated by auto- and enzymatic oxidation of the endogenous neurotransmitter dopamine (DA), the formation of neuromelanin and presence of high concentrations of iron. We have found that DA toxicity is mediated through its oxidative metabolites. Whereas thiol-containing antioxidants provided marked protection against DA toxicity, ascorbic acid accelerated DA-induced death. Using the differential display approach, we sought to isolate and characterize genes whose expression is altered in response to DA toxicity. We found an upregulation of the collapsin response mediator protein (CRM) and TCP-1delta in sympathetic neurons, which undergo dopamine-induced apoptosis. The isolation of these genes led us to examine the expression and activity of CRM and TCP-1delta related genes. Indeed, we found a significant induction of mRNAs of the secreted collapsin-1 and the mitochondrial stress protein HSP60. Antibodies directed against collapsin-1 provided marked and prolonged protection of several neuronal cell types from dopamine-induced apoptosis. In a parallel study, using antisense technology, we found that inhibition of TCP-1delta expression significantly reduced DA-induced neuronal death. These findings suggest a functional role for collapsin-1 and TCP-1delta as positive mediators of DA-induced neuronal apoptosis.

Animals↗

PubMed2000

Levodopa--an exotoxin or a therapeutic drug?

Auto-oxidation of levodopa generates toxic metabolites, such as free radicals, semiquinones and quinones. In vitro, levodopa is a powerful toxin that is lethal to cultures of neurones. This raises the concern that levodopa may also be toxic in vivo, and that chronic treatment with levodopa could induce further damage to nigrostriatal neurones in patients with Parkinson's disease, accelerating the natural predetermined rate of disease progression. Although a few animal studies have shown that chronic levodopa may be toxic in vivo, most others report that it is not. The few available clinical studies also indicate that the course of Parkinson's disease is not accelerated by chronic systemic treatment with levodopa.

Animals↗

PubMed2000

Haloperidol-induced neurotoxicity--possible implications for tardive dyskinesia.

Tardive dyskinesia (TD) is one of the major side effects of long term neuroleptic treatment. The pathophysiology of this disabling and commonly irreversible movement disorder is still obscure. The traditional concept of supersensitivity of striatal dopamine receptors as the mechanism involved in the development of TD is not satisfying, and current studies have focused on the role of neuroleptic-induced neuronal toxicity in the development of TD. We performed a series of experiments to gain a better understanding on the mechanisms involved in induction of TD. We have evaluated the direct neurotoxic effect of haloperidol (HP), a widely--used neuroleptic drug, and its three metabolites, in mouse neuronal cultures and in PC-12 cells. We found that the features of HP-induced cell death were apoptotic rather than necrotic, as indicated by different DNA-staining methods and specific caspases inhibitors. Moreover, cotreatment with antioxidants such as vitamin E and N-acetylcysteine (NAC) significantly protected the cultures. Further studies on the mechanisms underlying HP-induced toxicity may lead to the development of new neuroprotective therapeutic strategies.

Acetylcysteine↗

PubMed2000

Mice overexpressing Bcl-2 in their neurons are resistant to myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE).

Multiple sclerosis (MS) is an inflammatory disease of the central nervous system (CNS) characterized by destruction of myelin. Recent studies have indicated that axonal damage is involved in the pathogenesis of the progressive disability of this disease. To study the role of axonal damage in the pathogenesis of MS-like disease induced by myelin oligodendrocyte glycoprotein (MOG), we compared experimental autoimmune encephalomyelitis (EAE) in wild-type (WT) and transgenic mice expressing the human bcl-2 gene exclusively in neurons under the control of the neuron-specific enolase (NSE) promoter. Our study shows that, following EAE induction with pMOG 35-55, the WT mice developed significant clinical manifestations with complete hind-limb paralysis. In contrast, most of the NSE-bcl-2 mice (16/27) were completely resistant, whereas the others showed only mild clinical signs. Histological examination of CNS tissue sections showed multifocal areas of perivascular lymphohistiocytic inflammation with loss of myelin and axons in the WT mice, whereas only focal inflammation and minimal axonal damage were demonstrated in NSE-bcl-2 mice. No difference could be detected in the immune potency as indicated by delayed-type hypersensitivity (DTH) and T-cell proliferative responses to MOG. We also demonstrated that purified synaptosomes from the NSE-bcl-2 mice produce significantly lower level of reactive oxygen species (ROS) following exposure to H2O2 and nitric oxide (NO) than WT mice. In conclusion, we demonstrated that the expression of the antiapoptotic gene, bcl-2, reduces axonal damage and attenuates the severity of MOG-induced EAE. Our results emphasize the importance of developing neuroprotective therapies, in addition to immune-specific approaches, for treatment of MS.

Animals↗

PubMed2000

Developmental changes in antioxidant enzymes and oxidative damage in kidneys, liver and brain of bcl-2 knockout mice.

While programmed cell death is induced by a variety of internal and external stimuli, including reactive oxygen species, the anti-apoptotic protein Bcl-2 is involved in opposing cell death and affects the antioxidant status of cells. Since the exact mechanism of its action is uncertain, in this study we examined the role of Bcl-2 using a loss of function model, Bcl-2 knockout mice. The consequence of Bcl-2 knockout was assessed in kidneys, liver and brain, using protein carbonyls and cellular levels of antioxidant enzymes as markers of oxidative stress. Kidney extracts from 8 days-old Bcl-2-knockout mice had 59% higher content of protein carbonyls relative to the wild type, but similar levels of oxidized proteins at the age of 30 days. By marked contrast, in liver and brain, levels of protein carbonyls were similar at 8 days but by 30 days the liver of knockout animals (and brains, as we have shown previously) show 36% higher protein carbonyls. Measures of glutathione reductase (GRX), glutathione transferase (GST) and catalase revealed significantly higher levels in kidneys of 8 days old Bcl-2-knockout mice compared to wild type. By 30 days activities of glutathione-related enzymes and catalase increased and abolished the differences between the knockout and wild type. At 8 days, in liver there were no significant differences in activities of all enzymes between the mice, however by 30 days, the specific activity of GRX was significantly higher in Bcl-2-knockout mice, relative to controls. From day 8 to day 30 there was an increase in liver catalase activity that resulted in significantly higher levels in Bcl-2-knockout animals. Catalase activity in brains of Bcl-2-knockout, 8 days old mice was significantly higher compared to the wild type, and significantly lowers at 30 days. Taken together our findings indicate that Bcl-2 knockout results in significant perturbations of oxidative metabolism and antioxidant status of in kidney, liver and brain. Such changes are tissue specific with respect to age, magnitude and type of enzyme affected.

Animals↗

PubMed1999

Dopamine-melanin is actively phagocytized by PC12 cells and cerebellar granular cells: possible implications for the etiology of Parkinson's disease.

Neuromelanin in the substantia nigra may be associated with the pathogenesis of nigral cell death in Parkinson's disease. We used synthetic dopamine-melanin (DA-M) as a model compound for neuromelanin and examined its toxic effects on mice cerebellar granule cells and a rat pheochromocytoma cell line (PC12). The DA-M and dopamine-treated cells showed an accumulation of black deposits when examined by light microscopy. Electron microscopy revealed different stages of DA-M phagocytosis, starting with DA-M binding, engulfment of the particles and the formation of phagosomes located in the cytoplasm. Using absorption assays, we found that NaN3 and low temperature inhibit the internalization of DA-M, pointing to an energy-dependent phagocytosis mechanism. These results suggest that neuromelanin can be phagocytised by neuronal cells which may thus be subjected to its toxic effects. These findings may contribute to our understanding of the formation and disposition of neuromelanin and its possible role in the etiology of Parkinson's disease.

Animals↗