Wyniki wyszukiwania

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The role of alpha-synuclein in regulation of cyclin dependent kinase 5

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Czapski A. G., Gassowska M., Kazmierczak A., Adamczyk A.

Acta Neurobiologiae Experimentalis

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2013

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tom 73

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nr 1

Alpha-Synuclein (ASN), a small cytosolic protein enriched in synaptic terminals, was implicated in the pathomechanism of several neurodegenerative disorders called alpha-synucleinopathies. ASN was shown to be a main component of characteristic intraneuronal protein aggregates called Lewy bodies (LB) and Lewy neurites (LN), observed i.a. in Parkinson’s disease, dementia with LBs and in the LB variant of Alzheimer’s disease. Recent studies demonstrated that ASN may exist also in the extracellular space. Low-molecular ASN aggregates distributed in the brain parenchyma likely may be more toxic than ASN in LB, however, the exact mechanism of cytotoxicity of extracellular ASN is not fully understood. Our previous studies demonstrated the significant impact of extracellular ASN on calcium homeostasis. ASN evoked deregulation of intracellular calcium concentration leading in consequence to enhancement of nitric oxide synthesis. Deregulation of calcium homeostasis affects other calcium-dependent enzymes, including Calpains. The aim of the present study was to investigate the involvement of Calpaindependent activation of Cyclin Dependent Kinase 5 (Cdk5) in molecular mechanism of extracellular ASN cytotoxicity. The activation of Cdk5 is regulated by binding of regulatory subunits p35 and p39. Deregulation of calcium homeostasis may induce the Calpainmediated breakdown of Cdk5/p35 into Cdk5/p25 leading to overactivation of Cdk5. In our studies we used rat Pheochromocytoma PC12 cells incubated with exogenous ASN (10 µM) in the presence of Calpain inhibitor Calpeptin (10 µM) and Cdk5 inhibitors Roscovitine (10 µM) and BML-259 (10 µM). Our results indicated that incubation of PC12 cells in the presence of extracellular ASN (10 µM) for 48 h evoked cell death, and Cdk5 inhibitors efficiently prevented ASN toxicity, indicating an important role of Cdk5 in molecular mechanism of ASN toxicity. The level of Cdk5 protein was unchanged, but phosphorylation of Cdk5 at Tyr15 was significantly increased, suggesting that the enzymatic activity of Cdk5 is increased in ASN-treated cells. The presence of p25 protein was observed, what suggests that Calpain-dependent proteolysis of p35 occurred in ASN-treated cells. Calpeptin, an inhibitor of Calpains, prevented ASN-induced cell death, confirming the important role of Calpain activation in mechanism of ASN toxicity. In summary, our results demonstrated that alteration of calcium homeostasis evoked by extracellular ASN induce Calpain-dependent overactivation of Cdk5. These molecular processes may be involved in ASN-evoked cell death in vitro and probably also in neurodegenerative disorders.

2

Neuropathic alterations of the myenteric plexus neurons following subacute intraperitoneal administration of salsolinol

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Kurnik M., Gil K., Gajda M., Thor P., Bugajski A.

Folia Histochemica et Cytobiologica

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2015

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tom 53

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nr 1

3

Role of the unfolded protein response in the pathogenesis of Parkinson’s disease

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Varma D., Sen D.

Acta Neurobiologiae Experimentalis

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2015

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tom 75

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nr 1

Parkinson's disease is the second most common neurodegenerative disease which affects almost 1% of the population above the age of 60. It is is characterized by loss of dopaminergic neurons in the striatum and substantia nigra, coupled with the formation of intracellular Lewy bodies in degenerating neurons. Recent evidence suggests endoplasmic reticulum stress as a common and prominent occurrence in the progression of Parkinson's disease pathogenesis in the affected human brain. One of the cellular defense mechanism to combat endoplasmic reticulum stress due to excessive protein accumulation is through activation of the unfolded protein response pathway. In this review we focus on the impact and role of this unfolded protein response as a causative factor of Parkinson's disease leading to neurodegeneration.

4

Expression of alpha-synuclein in different brain parts of adult and aged rats

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Adamczyk A., Solecka J., Strosznajder J. B.

Journal of Physiology and Pharmacology

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2005

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tom 56

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nr 1

The synucleins are a family of presynaptic proteins that are abundant in neurons and include alpha-, ß-, and -synuclein. alpha-Synuclein (ASN) is involved in several neurodegenerative age-related disorders but its relevance in physiological aging is unknown. In the present study we investigated the expression of ASN mRNA and protein in the different brain parts of the adult (4-month-old) and aged (24-month-old) rats by using RT-PCR technique and Western blot, respectively. Our results indicated that mRNA expression and immunoreactivity of ASN is similar in brain cortex, hippocampus and striatum but markedly lower in cerebellum comparing to the other brain parts. Aging lowers ASN mRNA expression in striatum and cerebellum by about 40%. The immunoreactivity of ASN in synaptic plasma membranes (SPM) from aged brain cortex, hippocampus and cerebellum is significantly lower comparing to adult by 39%, 24% and 65%, respectively. ß-synuclein (BSN) was not changed in aged brain comparing to adult. Age-related alteration of ASN may affect the nerve terminals structure and function.

5

Alpha-synuclein A30P point-mutation generates age-dependent nigrostriatal deficiency in mice

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Plaas M., Karis A., Innos J., Rebane E., Baekelandt V., Vaarmann A., Luuk H., Vasar E., Koks S.

Journal of Physiology and Pharmacology

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2008

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tom 59

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nr 2

Lewy bodies are mainly composed of alpha-synuclein (SNCA) and specific mutations in SNCA gene are related to familial forms of Parkinson's disease (PD). The purpose of our study was to generate a mouse line with A30P knock-in point mutation in SNCA gene and to test if a single point-mutation is able to turn otherwise normal SNCA into a toxic form. The behavioral profile of SNCA A30P mice was followed for 16 months. Generally, these mice are healthy and viable without any obvious abnormalities. Starting from the age of 13 months mice developed a significant deficit in motor performance tests related to nigrostriatal function (ink-test and beam walk). In other tests (motility boxes, rotarod) mice continuously performed normally. Moreover, SNCA A30P mice expressed the altered sensitivity to VMAT2 inhibitor reserpine, possibly reflecting a functional deficiency of dopamine. Indeed, mice at 15 months of age had significantly reduced levels of dopamine and its major metabolite DOPAC in the striatum, and reduced levels of dopamine in the mesolimbic system. The present study confirms that SNCA plays an important role in the development of PD and an insertion of a single point mutation is sufficient to generate age-related decline in specific motor performance. The generated mouse line has a potential to become a model for PD with comparable time course and phenotype.

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The role of sphingosine kinases/sphingosine-1-phosphate in the regulation of alpha-synuclein and amyloid beta precursor protein secretion. Implications for neurodegenerative disorders

80%

Jesko H., Okada T., Nakamura S., Strosznajder R. P.

Acta Neurobiologiae Experimentalis

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2013

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tom 73

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nr 1

Sphingolipid deregulation may be an important factor of age-related neuronal stress vulnerability. Current data suggests potential links between sphingosine kinases (SphK1&2), their product sphingosine1-phosphate (S1P) and age-related protein conformation diseases. The aim of this study was to investigate a possible role of SphKs in alpha-synuclein (ASN) and amyloid beta (ABeta) precursor protein (APP) level and secretion. The studies were carried out using human SH-SY5Y neuroblastoma cell line stably transfected with the human gene for α-synuclein (ASNwt). Sphingosine kinase inhibitor (SKI) significantly increased ASN secretion in concentration-dependent manner. S1P also displayed similar influence. Neither compound exerted any significant effect on the ASN protein level. S1P may act via cell surface receptors or as an intracellular second messenger. The similar effect of S1P and SphK inhibitors on ASN secretion may suggest that the regulation of its release is critically dependent on the varied (intra)cellular targets of SphKs and downstream signaling pathways. We have found that stable human ASNwt expression in SH-SY5Y cells caused a three-fold, significant increase of the cellular APP level. In ASN-transfected cells S1P enhanced APP secretion and reduced its intracellular level. This could be linked to the recently reported effect of S1P on secretase beta activity. Inhibition of SphKs significantly decreased APP secretion. In summary our data indicates that endogenous ASN regulates APP level in SH-SY5Y cells and that sphingolipids play a crucial role in the secretion of ASN and APP. These processes may have significant impact on neuronal survival and health.

7

Extracellular alpha-synuclein contributes to gsk-3beta-dependent Tau phosphorylation in PC12 dopaminergic cells: Its role in pathomechanism of Parkinson’s disease

80%

Gassowska M., Czapski G. A., Adamczyk A.

Acta Neurobiologiae Experimentalis

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2013

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tom 73

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nr 1

alpha-Synuclein (ASN) play important role in pathogenesis of Parkinson’s disease (PD) and other neurodegenerative disorders. Novel and most interesting data showed elevated tauopathy in PD and suggested relationship between ASN and Tau protein. However, the mechanism of ASN-evoked Tau protein modification is not fully elucidated. In this study, we investigated the role of glycogen synthase kinase-3β (Gsk-3β) and cyclin-dependent kinase 5 (Cdk5) in ASN-evoked Tau modification in dopaminergic PC12 cells. We used real-time quantitative PCR (qRT-PCR) analysis to assess Gsk3β gene expression and Western blot technique to analyse protein phosphorylation. The presence of apoptotic cells was assessed by Hoechst 33258 fluorescent staining, and cell viability was determined by the 2-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) colorimetric assay. Our data showed that exogenously added ASN (10 μM) increases Tau phosphorylation on Ser396 and specific Gsk-3β inhibitor (SB-216763, 10 µM) opposite to Cdk5 inhibitor protects cells against Tau hyperphosphorylation. Western blot analysis showed that ASN affected Gsk-3β via increasing of protein level and activation of this enzyme. From immunochemical studies, was found that ASN treatment leads to significant increase in GSK-3β immunoreactivity by about 20%. GSK-3β activity evaluated by its phosphorylation status assay showed that ASN significantly increased the phosphorylation of this enzyme at Tyr216 with parallel decrease in phosphorylation at Ser9, indicative of stimulation of GSK-3β activity. ASN-induced apoptotic processes leads to decrease of PC12 cells viability, the apoptotic cells determined by phase contrast together with Hoechst 33258 fluorescent staining, indicated significantly increase of apoptosis in the presence of ASN. SB-216763 prevented ASN-induced cytotoxicity and enhanced PC12 cell viability. In conclusion, all these findings suggested that extracellular ASN is involved in Gsk-3β-dependent Tau modulation and its proapoptotic effect might be mediated at least in part by the Gsk-3β catalysed Tau hyperphosphorylation and impairment of cytoskeleton stability. GSK-3β inhibitors may offer promising tool against ASN-induced Tau modification and cytotoxicity in neurodegenerative disorders. Supported by statutory theme 9.

Ograniczanie wyników

The role of alpha-synuclein in regulation of cyclin dependent kinase 5

Neuropathic alterations of the myenteric plexus neurons following subacute intraperitoneal administration of salsolinol

Role of the unfolded protein response in the pathogenesis of Parkinson’s disease

Expression of alpha-synuclein in different brain parts of adult and aged rats

Alpha-synuclein A30P point-mutation generates age-dependent nigrostriatal deficiency in mice

The role of sphingosine kinases/sphingosine-1-phosphate in the regulation of alpha-synuclein and amyloid beta precursor protein secretion. Implications for neurodegenerative disorders

Extracellular alpha-synuclein contributes to gsk-3beta-dependent Tau phosphorylation in PC12 dopaminergic cells: Its role in pathomechanism of Parkinson’s disease