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Harish C. Pant - One of the best experts on this subject based on the ideXlab platform.
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Squid Giant Axon Contains Neurofilament Protein mRNA but does not Synthesize Neurofilament Proteins.
Cellular and molecular neurobiology, 2016Co-Authors: Harold Gainer, Shirley B. House, Dong Sun Kim, Hemin Chin, Harish C. PantAbstract:When isolated squid giant axons are incubated in radioactive amino acids, abundant newly synthesized Proteins are found in the axoplasm. These Proteins are translated in the adaxonal Schwann cells and subsequently transferred into the giant axon. The question as to whether any de novo Protein synthesis occurs in the giant axon itself is difficult to resolve because the small contribution of the Proteins possibly synthesized intra-axonally is not easily distinguished from the large amounts of the Proteins being supplied from the Schwann cells. In this paper, we reexamine this issue by studying the synthesis of endogenous Neurofilament (NF) Proteins in the axon. Our laboratory previously showed that NF mRNA and Protein are present in the squid giant axon, but not in the surrounding adaxonal glia. Therefore, if the isolated squid axon could be shown to contain newly synthesized NF Protein de novo, it could not arise from the adaxonal glia. The results of experiments in this paper show that abundant 3H-labeled NF Protein is synthesized in the squid giant fiber lobe containing the giant axon's neuronal cell bodies, but despite the presence of NF mRNA in the giant axon no labeled NF Protein is detected in the giant axon. This lends support to the glia-axon Protein transfer hypothesis which posits that the squid giant axon obtains newly synthesized Protein by Schwann cell transfer and not through intra-axonal Protein synthesis, and further suggests that the NF mRNA in the axon is in a translationally repressed state.
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pin1 dependent prolyl isomerization modulates the stress induced phosphorylation of high molecular weight Neurofilament Protein
Journal of Biological Chemistry, 2008Co-Authors: Parvathi Rudrabhatla, Niranjana D Amin, Yali Zheng, Sashi Kesavapany, Wayne Albers, Harish C. PantAbstract:Aberrant phosphorylation of neuronal cytoskeletal Proteins is a key pathological event in neurodegenerative disorders such as Alzheimer disease (AD) and amyotrophic lateral sclerosis, but the underlying mechanisms are still unclear. Previous studies have shown that Pin1, a peptidylprolyl cis/trans-isomerase, may be actively involved in the regulation of Tau hyperphosphorylation in AD. Here, we show that Pin1 modulates oxidative stress-induced NF-H phosphorylation. In an in vitro kinase assay, the addition of Pin1 substantially increased phosphorylation of NF-H KSP repeats by proline-directed kinases, Erk1/2, Cdk5/p35, and JNK3 in a concentration-dependent manner. In vivo, dominant-negative (DN) Pin1 and Pin1 small interfering RNA inhibited epidermal growth factor-induced NF-H phosphorylation. Because oxidative stress plays an important role in the pathogenesis of neurodegenerative diseases, we studied the role of Pin1 in stressed cortical neurons and HEK293 cells. Both hydrogen peroxide (H(2)O(2)) and heat stresses induce phosphorylation of NF-H in transfected HEK293 cells and primary cortical cultures. Knockdown of Pin1 by transfected Pin1 short interference RNA and DN-Pin1 rescues the effect of stress-induced NF-H phosphorylation. The H(2)O(2) and heat shock induced perikaryal phospho-NF-H accumulations, and neuronal apoptosis was rescued by inhibition of Pin1 in cortical neurons. JNK3, a brain-specific JNK isoform, is activated under oxidative and heat stresses, and inhibition of Pin1 by Pin1 short interference RNA and DN-Pin1 inhibits this pathway. These results implicate Pin1 as a possible modulator of stress-induced NF-H phosphorylation as seen in neurodegenerative disorders like AD and amyotrophic lateral sclerosis. Thus, Pin1 may be a potential therapeutic target for these diseases.
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Neurofilament Protein synthesis and phosphorylation
Journal of Neurocytology, 2000Co-Authors: Philip Grant, Harish C. PantAbstract:Neurofilament Proteins, a major intermediate filament component of the neuronal cytoskeleton, are organized as 10 nm thick filaments in axons and dendrites. They are large, abundantly phosphorylated Proteins with numerous phosphate acceptor sites, up to 100 in some cases, organized as numerous repeat motifs. Together with other cytoskeletal components such as microtubules, MAPs, actin and plectin-like linking molecules, they make up a dynamic lattice that sustains neuronal function from neuronal “birthday” to apoptotic cell death. The activity of the neuronal cytoskeleton is regulated by phosphorylation, dephosphorylation reactions mediated by numerous associated kinases, phosphatases and their regulators. Factors regulating multisite phosphorylation of NFs are topographically localized, with maximum phosphorylation of NF Proteins consigned to axons. Phosphorylation defines the nature of NF interactions with one another and with other cytoskeletal components such as microtubules, MAPs and actin. To understand how these functional interactions are regulated by phosphorylation we attempt to identify the relevant kinases and phosphatases, their specific targets and the factors modulating their activity. As an initial working model we propose that NF phosphorylation is regulated topographically in neurons by compartment-specific macromolecular complexes of substrates, kinases and phosphatases. This implies that axonal complexes differ structurally and functionally from those in cell bodies and dendrites. Such Protein assemblies, by virtue of conformational changes within Proteins, facilitate ordered, sequential multisite phosphorylations that modulate dynamic cytoskeletal interactions.
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integrin α1β1 mediated activation of cyclin dependent kinase 5 activity is involved in neurite outgrowth and human Neurofilament Protein h lys ser pro tail domain phosphorylation
The Journal of Neuroscience, 2000Co-Authors: Bingsheng Li, Lei Zhang, Jianguo Gu, Niranjana D Amin, Harish C. PantAbstract:Cellular adhesion to the extracellular matrix is mediated by a diverse class of α/β heterodimeric receptors known as integrins, which transduce signals to activate multiple intracellular signal transduction pathways within the cells. The signaling pathway linking integrins to mediate neuronal process outgrowth is not well understood. Here, we have provided evidence that intracellular signaling by the α1β1 integrin-induced activation of cyclin-dependent kinase 5 (cdk5) is involved in neurite outgrowth and human Neurofilament Protein H (hNF-H) Lys-Ser-Pro (KSP) tail domain phosphorylation in differentiated human SH-SY5Y cells. The integrin α1 and β1 monoclonal antibodies and BL-1, a specific cdk5 inhibitor, inhibited these effects. We also demonstrated that cdk5 activity and hNF-H KSP tail domain phosphorylation were increased in cdk5/p35 and hNF-H tail domain co-transfected HEK293 cells grown on laminin. This increased hNF-H tail domain phosphorylation was triggered by cdk5 activation. Taken together, these results indicated that cdk5 may play an important role in promoting neurite outgrowth and hNF-H tail KSP domain phosphorylation through the integrin α1β1signaling pathway.
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integrin α1β1 mediated activation of cyclin dependent kinase 5 activity is involved in neurite outgrowth and human Neurofilament Protein h lys ser pro tail domain phosphorylation
The Journal of Neuroscience, 2000Co-Authors: Lei Zhang, Niranjana D Amin, Harish C. PantAbstract:Cellular adhesion to the extracellular matrix is mediated by a diverse class of alpha/beta heterodimeric receptors known as integrins, which transduce signals to activate multiple intracellular signal transduction pathways within the cells. The signaling pathway linking integrins to mediate neuronal process outgrowth is not well understood. Here, we have provided evidence that intracellular signaling by the alpha(1)beta(1) integrin-induced activation of cyclin-dependent kinase 5 (cdk5) is involved in neurite outgrowth and human Neurofilament Protein H (hNF-H) Lys-Ser-Pro (KSP) tail domain phosphorylation in differentiated human SH-SY5Y cells. The integrin alpha(1) and beta(1) monoclonal antibodies and BL-1, a specific cdk5 inhibitor, inhibited these effects. We also demonstrated that cdk5 activity and hNF-H KSP tail domain phosphorylation were increased in cdk5/p35 and hNF-H tail domain co-transfected HEK293 cells grown on laminin. This increased hNF-H tail domain phosphorylation was triggered by cdk5 activation. Taken together, these results indicated that cdk5 may play an important role in promoting neurite outgrowth and hNF-H tail KSP domain phosphorylation through the integrin alpha(1)beta(1) signaling pathway.
Lars Rosengren - One of the best experts on this subject based on the ideXlab platform.
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biomarker evidence of axonal injury in neuroasymptomatic hiv 1 patients
PLOS ONE, 2014Co-Authors: Jan Jessen Krut, Lars Rosengren, Richard W Price, Lars Hagberg, Tomas Mellberg, Dietmar Fuchs, Staffan Nilsson, Henrik Zetterberg, Magnus GisslenAbstract:Prevalence of neurocognitive impairment in HIV-1 infected patients is reported to be high. Whether this is a result of active HIV-related neurodegeneration is unclear. We examined axonal injury in HIV-1 patients by measuring the light subunit of Neurofilament Protein (NFL) in CSF with a novel, sensitive method.With a cross-sectional design, CSF concentrations of Neurofilament Protein light (NFL) (marker of neuronal injury), neopterin (intrathecal immunoactivation) and CSF/Plasma albumin ratio (blood-brain barrier integrity) were analyzed on CSF from 252 HIV-infected patients, subdivided into untreated neuroasymptomatics (n = 200), HIV-associated dementia (HAD) (n = 14) and on combinations antiretroviral treatment (cART) (n = 85), and healthy controls (n = 204). 46 HIV-infected patients were included in both treated and untreated groups, but sampled at different timepoints. Furthermore, 78 neuroasymptomatic patients were analyzed before and after treatment initiation. While HAD patients had the highest NFL concentrations, elevated CSF NFL was also found in 33% of untreated neuroasymptomatic patients, mainly in those with blood CD4+ cell counts below 250 cells/μL. CSF NFL concentrations in the untreated neuroasymptomatics and treated groups were equivalent to controls 18.5 and 3.9 years older, respectively. Neopterin correlated with NFL levels in untreated groups while the albumin ratio correlated with NFL in both untreated and treated groups. Increased CSF NFL indicates ongoing axonal injury in many neuroasymptomatic patients. Treatment decreases NFL, but treated patients retain higher levels than controls, indicating either continued virus-related injury or an aging-like effect of HIV infection. NFL correlates with neopterin and albumin ratio, suggesting an association between axonal injury, neuroinflammation and blood-brain barrier permeability. NFL appears to be a sensitive biomarker of subclinical and clinical brain injury in HIV and warrants further assessment for broader clinical use.
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antiretroviral treatment reduces increased csf Neurofilament Protein nfl in hiv 1 infection
Neurology, 2007Co-Authors: Asa Mellgren, Lars Rosengren, Richard W Price, Lars Hagberg, Bruce J Brew, Magnus GisslenAbstract:Objective: Increased levels of the light-chain Neurofilament Protein (NFL) in CSF provide a marker of CNS injury in several neurodegenerative disorders and have been reported in the AIDS dementia complex (ADC). We examined the effects of highly active antiretroviral treatment (HAART) on CSF NFL in HIV-1–infected subjects with and without ADC who underwent repeated lumbar punctures (LPs). Method: NFL was measured by ELISA (normal reference value Results: Twenty-one of the subjects had increased CSF NFL at baseline, with a median level of 780 ng/L and an intraquartile range (IQR) of 480 to 7300. After 3 months of treatment, NFL concentrations had fallen to normal in 48% (10/21), and the median decreased to 340 ng/L (IQR p Conclusion: HAART seems to halt the neurodegenerative process(es) caused by HIV-1, as shown by the significant decrease in CSF NFL after treatment initiation. CSF NFL may serve as a useful marker in monitoring CNS injury in HIV-1 infection and in evaluating CNS efficacy of antiretroviral therapy.
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elevated cerebrospinal fluid Neurofilament light Protein concentrations predict the development of aids dementia complex
The Journal of Infectious Diseases, 2007Co-Authors: Magnus Gisslen, Richard W Price, Lars Hagberg, Bruce J Brew, Paola Cinque, Lars RosengrenAbstract:The light subunit of Neurofilament Protein (NFL) is a sensitive indicator of central nervous system axonal injury. We retrospectively identified 9 subjects participating in a longitudinal cohort study who developed acquired immunodeficiency syndrome dementia complex (ADC) and who had had a lumbar puncture performed within 2 years before presentation. Elevated cerebrospinal fluid (CSF) NFL concentrations were found in 7 (78%) of the 9 case patients who later developed ADC, compared with 9 (33%) of 27 CD4 cell count-matched HIV-1-infected control subjects. By contrast, no differences were found in CSF HIV-1 RNA or neopterin concentrations between the 2 groups. CSF NFL may prove to be a useful predictive marker for ADC.
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csf Neurofilament Protein nfl a marker of active hiv related neurodegeneration
Journal of Neurology, 2007Co-Authors: Sahra Abdulle, Lars Rosengren, Asa Mellgren, Richard W Price, Lars Hagberg, Bruce J Brew, Magnus Gisslen, Paola CinqueAbstract:The light subunit of the Neurofilament Protein (NFL), a major structural component of myelinated axons, is a sensitive indicator of axonal injury in the central nervous system (CNS) in a variety of neurodegenerative disorders. Cerebrospinal fluid (CSF) NFL concentrations were measured by ELISA (normal < 250 ng/l) in archived samples from 210 HIV-infected patients not taking antiretroviral treatment: 55 with AIDS dementia complex (ADC), 44 with various CNS opportunistic infections/tumours (CNS OIs), 95 without neurological symptoms or signs, and 16 with primary HIV infection (PHI). The effect of highly active antiretroviral treatment (HAART) was studied by repeated CSF sampling in four of the ADC patients initiating treatment. CSF NFL concentrations were significantly higher in patients with ADC (median 2590 ng/l, IQR 780–7360) and CNS OIs (2315 ng/l, 985–7390 ng/l) than in neuroasymptomatic patients (<250 ng/l, <250–300) or PHI (<250 ng/l, <250–280), p < 0.001. Among patients with ADC, those with more severe disease (stage 2–4) had higher levels than those with milder disease (stage 0.5–1), p < 0.01. CSF NFL declined during HAART to the limit of detection in parallel with virological response and neurological improvement in ADC.CSF NFL concentrations were higher in neuroasymptomatic patients with lower CD4-cell strata than higher, p < 0.001. This increase was less marked than in the ADC patients and noted in 26/58 neuroasymptomatic patients with CD4 counts <200/μl compared to 1/37 with CD4-cells ≥200/μl. The findings of this study support the value of CSF NFL as a useful marker of ongoing CNS damage in HIV infection. Markedly elevated CSF NFL concentrations in patients without CNS OIs are associated with ADC, follow the grade of severity, and decrease after initiation of effective antiretroviral treatment. Nearly all previously suggested CSF markers of ADC relate to immune activation or HIV viral load that do not directly indicate brain injury. By contrast NFL is a sensitive marker of such injury, and should prove useful in evaluating the presence and activity of ongoing CNS injury in HIV infection.
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Neurofilament Protein in cerebrospinal fluid a marker of white matter changes
Journal of Neuroscience Research, 2001Co-Authors: Magnus Sjogren, Lars Rosengren, Kaj Blennow, M Blomberg, Michael Jonsson, L O Wahlund, Ake Edman, Karin Lind, Anders WallinAbstract:The objective of this study was to compare cerebrospinal fluid (CSF) levels of the light subtype of the Neurofilament Proteins (NFL), tau, and beta-amyloid42 (Abeta42) in individuals with moderate or severe white matter changes (WMC) and in those with mild or no WMC. Twenty-two patients with Alzheimer's disease (AD), nine patients with subcortical vascular dementia (SVD), and 20 normal controls were included in the study. The occurrence of WMC was evaluated by a neuroradiologist using the Blennow-Wallin scale. Thirty-seven subjects had no or only punctate WMC; 14 had moderate to severe WMC. Both diagnostic group and WMC, but not gender or apolipoProteinE E4 inheritance, contributed to the variance in the CSF levels of tau, NFL, and Abeta42. In patients with moderate to severe WMC, CSF NFL (P < 0.01), but not CSF tau or CSF Abeta42, was increased also after correction for age, gender, and degree of cognitive impairment. A comparison between patients and controls with any signs of WMC and those without such signs yielded a similar result: CSF NFL (P < 0.001) was increased in the group with signs of WMC. As in numerous previous studies, we found that CSF tau was increased in AD (P < 0.001) compared with controls. Furthermore, CSF NFL was increased in both AD and SVD compared with controls (P < 0.001 for both). Although diagnostic group seems to be a stronger predictor of the variance found in CSF NFL, a clear association between the presence of WMC and increased CSF NFL was found. Because NFL is located mainly in large myelinated axons, increased CSF NFL in individuals with WMC probably reflects axonal degeneration.
Magnus Gisslen - One of the best experts on this subject based on the ideXlab platform.
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biomarker evidence of axonal injury in neuroasymptomatic hiv 1 patients
PLOS ONE, 2014Co-Authors: Jan Jessen Krut, Lars Rosengren, Richard W Price, Lars Hagberg, Tomas Mellberg, Dietmar Fuchs, Staffan Nilsson, Henrik Zetterberg, Magnus GisslenAbstract:Prevalence of neurocognitive impairment in HIV-1 infected patients is reported to be high. Whether this is a result of active HIV-related neurodegeneration is unclear. We examined axonal injury in HIV-1 patients by measuring the light subunit of Neurofilament Protein (NFL) in CSF with a novel, sensitive method.With a cross-sectional design, CSF concentrations of Neurofilament Protein light (NFL) (marker of neuronal injury), neopterin (intrathecal immunoactivation) and CSF/Plasma albumin ratio (blood-brain barrier integrity) were analyzed on CSF from 252 HIV-infected patients, subdivided into untreated neuroasymptomatics (n = 200), HIV-associated dementia (HAD) (n = 14) and on combinations antiretroviral treatment (cART) (n = 85), and healthy controls (n = 204). 46 HIV-infected patients were included in both treated and untreated groups, but sampled at different timepoints. Furthermore, 78 neuroasymptomatic patients were analyzed before and after treatment initiation. While HAD patients had the highest NFL concentrations, elevated CSF NFL was also found in 33% of untreated neuroasymptomatic patients, mainly in those with blood CD4+ cell counts below 250 cells/μL. CSF NFL concentrations in the untreated neuroasymptomatics and treated groups were equivalent to controls 18.5 and 3.9 years older, respectively. Neopterin correlated with NFL levels in untreated groups while the albumin ratio correlated with NFL in both untreated and treated groups. Increased CSF NFL indicates ongoing axonal injury in many neuroasymptomatic patients. Treatment decreases NFL, but treated patients retain higher levels than controls, indicating either continued virus-related injury or an aging-like effect of HIV infection. NFL correlates with neopterin and albumin ratio, suggesting an association between axonal injury, neuroinflammation and blood-brain barrier permeability. NFL appears to be a sensitive biomarker of subclinical and clinical brain injury in HIV and warrants further assessment for broader clinical use.
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antiretroviral treatment reduces increased csf Neurofilament Protein nfl in hiv 1 infection
Neurology, 2007Co-Authors: Asa Mellgren, Lars Rosengren, Richard W Price, Lars Hagberg, Bruce J Brew, Magnus GisslenAbstract:Objective: Increased levels of the light-chain Neurofilament Protein (NFL) in CSF provide a marker of CNS injury in several neurodegenerative disorders and have been reported in the AIDS dementia complex (ADC). We examined the effects of highly active antiretroviral treatment (HAART) on CSF NFL in HIV-1–infected subjects with and without ADC who underwent repeated lumbar punctures (LPs). Method: NFL was measured by ELISA (normal reference value Results: Twenty-one of the subjects had increased CSF NFL at baseline, with a median level of 780 ng/L and an intraquartile range (IQR) of 480 to 7300. After 3 months of treatment, NFL concentrations had fallen to normal in 48% (10/21), and the median decreased to 340 ng/L (IQR p Conclusion: HAART seems to halt the neurodegenerative process(es) caused by HIV-1, as shown by the significant decrease in CSF NFL after treatment initiation. CSF NFL may serve as a useful marker in monitoring CNS injury in HIV-1 infection and in evaluating CNS efficacy of antiretroviral therapy.
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elevated cerebrospinal fluid Neurofilament light Protein concentrations predict the development of aids dementia complex
The Journal of Infectious Diseases, 2007Co-Authors: Magnus Gisslen, Richard W Price, Lars Hagberg, Bruce J Brew, Paola Cinque, Lars RosengrenAbstract:The light subunit of Neurofilament Protein (NFL) is a sensitive indicator of central nervous system axonal injury. We retrospectively identified 9 subjects participating in a longitudinal cohort study who developed acquired immunodeficiency syndrome dementia complex (ADC) and who had had a lumbar puncture performed within 2 years before presentation. Elevated cerebrospinal fluid (CSF) NFL concentrations were found in 7 (78%) of the 9 case patients who later developed ADC, compared with 9 (33%) of 27 CD4 cell count-matched HIV-1-infected control subjects. By contrast, no differences were found in CSF HIV-1 RNA or neopterin concentrations between the 2 groups. CSF NFL may prove to be a useful predictive marker for ADC.
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csf Neurofilament Protein nfl a marker of active hiv related neurodegeneration
Journal of Neurology, 2007Co-Authors: Sahra Abdulle, Lars Rosengren, Asa Mellgren, Richard W Price, Lars Hagberg, Bruce J Brew, Magnus Gisslen, Paola CinqueAbstract:The light subunit of the Neurofilament Protein (NFL), a major structural component of myelinated axons, is a sensitive indicator of axonal injury in the central nervous system (CNS) in a variety of neurodegenerative disorders. Cerebrospinal fluid (CSF) NFL concentrations were measured by ELISA (normal < 250 ng/l) in archived samples from 210 HIV-infected patients not taking antiretroviral treatment: 55 with AIDS dementia complex (ADC), 44 with various CNS opportunistic infections/tumours (CNS OIs), 95 without neurological symptoms or signs, and 16 with primary HIV infection (PHI). The effect of highly active antiretroviral treatment (HAART) was studied by repeated CSF sampling in four of the ADC patients initiating treatment. CSF NFL concentrations were significantly higher in patients with ADC (median 2590 ng/l, IQR 780–7360) and CNS OIs (2315 ng/l, 985–7390 ng/l) than in neuroasymptomatic patients (<250 ng/l, <250–300) or PHI (<250 ng/l, <250–280), p < 0.001. Among patients with ADC, those with more severe disease (stage 2–4) had higher levels than those with milder disease (stage 0.5–1), p < 0.01. CSF NFL declined during HAART to the limit of detection in parallel with virological response and neurological improvement in ADC.CSF NFL concentrations were higher in neuroasymptomatic patients with lower CD4-cell strata than higher, p < 0.001. This increase was less marked than in the ADC patients and noted in 26/58 neuroasymptomatic patients with CD4 counts <200/μl compared to 1/37 with CD4-cells ≥200/μl. The findings of this study support the value of CSF NFL as a useful marker of ongoing CNS damage in HIV infection. Markedly elevated CSF NFL concentrations in patients without CNS OIs are associated with ADC, follow the grade of severity, and decrease after initiation of effective antiretroviral treatment. Nearly all previously suggested CSF markers of ADC relate to immune activation or HIV viral load that do not directly indicate brain injury. By contrast NFL is a sensitive marker of such injury, and should prove useful in evaluating the presence and activity of ongoing CNS injury in HIV infection.
Ronald K H Liem - One of the best experts on this subject based on the ideXlab platform.
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Neurofilament Protein levels quantitative analysis in essential tremor cerebellar cortex
Neuroscience Letters, 2012Co-Authors: Elan D Louis, Ronald K H Liem, Rachel Babij, Etty Cortes, Jean Paul G Vonsattel, Phyllis L FaustAbstract:Essential tremor (ET) is among the most prevalent neurological diseases. A substantial increase in the number of Purkinje cell axonal swellings (torpedoes) has been identified in ET brains. We recently demonstrated that torpedoes in ET contain an over-accumulation of disorganized Neurofilament (NF) Proteins. This now raises the question whether NF Protein composition and/or phosphorylation state in cerebellar tissue might differ between ET cases and controls. We used a Western blot analysis to compare the levels and phosphorylation state of NF Proteins and α-internexin in cerebellar tissue from 47 ET cases versus 26 controls (2:1 ratio). Cases and controls did not differ with respect to the cerebellar levels of NF-light (NF-L), NF-medium (NF-M), NF-heavy (NF-H), or α-internexin. However, SMI-31 levels (i.e., phosphorylated NF-H) and SMI-32 levels (i.e., non-phosphorylated NF-H) were significantly higher in ET cases than controls (1.28±0.47 vs. 1.06±0.32, p=0.02; and 1.38±0.75 vs. 1.00±0.42, p=0.006). Whether the abnormal phosphorylation state that we observed is a cause of defective axonal transport and/or function of NFs in ET is not known. NF abnormalities have been demonstrated in several neurodegenerative diseases. Regardless of whether these Protein aggregates are the cause or consequence of these diseases, NF abnormalities have been shown to be an important factor in the cellular disruption observed in several neurodegenerative diseases. Therefore, further analyses of these NF abnormalities and their mechanisms are important to enhance our understanding of disease pathogenesis in ET.
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phosphorylation of the high molecular weight Neurofilament Protein nf h by cdk5 and p35
Journal of Biological Chemistry, 1996Co-Authors: Dongming Sun, Conrad L Leung, Ronald K H LiemAbstract:Abstract The high molecular weight Neurofilament Protein (NF-H) is highly phosphorylated in the axon. The phosphorylation sites have been identified as KSP (Lys-Ser-Pro) repeats in the tail domain of NF-H. These KSP sequences are present more than 50 times in the NF-H tail, and most of these sites are normally phosphorylated in vivo. These KSP sites can be further divided into two separate consensus sequences, KSPXK and KSPXY (where Y is not K). The extensive phosphorylation of NF-H has been proposed to play a critical role in the determination of axonal diameter. Recent studies have shown that Cdk5, a kinase related to the cell cycle-dependent kinase Cdc2, is expressed in the brain and associates with the cytoskeleton. In vitro phosphorylation studies have shown that Cdk5 in conjunction with its activator, p35, is able to phosphorylate histone H1, dephosphorylated NF-H, as well as a synthetic peptide with the repetitive KSP motif. We have cloned the cDNAs for rat Cdk5 and p35 by reverse transcription-polymerase chain reaction and cDNA library screening and studied the phosphorylation of NF-H both in vivo and in vitro. By transient transfection assays, we have shown that NF-H can only be extensively phosphorylated in the presence of both Cdk5 and p35. This phosphorylation can be inhibited by a Cdk5-dominant negative mutant, an observation which further supports that Cdk5 is a kinase that is able to phosphorylate NF-H. By immunoprecipitating Cdk5 and p35 from the transfected cells, we have been able to show that the KSPXK repeats are the preferred phosphorylation sites for Cdk5, while the KSPXY repeats are not directly phosphorylated by Cdk5 and p35.
In H. Park - One of the best experts on this subject based on the ideXlab platform.
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Multilineage Potential of Stable Human Mesenchymal Stem Cell Line Derived from Fetal Marrow
2013Co-Authors: Atsushi Nagai, Woo K. Kim, Hong J. Lee, Han S. Jeong, Kwang S. Kim, Seok H. Hong, In H. Park, Seung U. KimAbstract:Human bone marrow contains two major cell types, hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs). MSCs possess self-renewal capacity and pluripotency defined by their ability to differentiate into osteoblasts, chondrocytes, adipocytes and muscle cells. MSCs are also known to differentiate into neurons and glial cells in vitro, and in vivo following transplantation into the brain of animal models of neurological disorders including ischemia and intracerebral hemorrhage (ICH) stroke. In order to obtain sufficient number and homogeneous population of human MSCs, we have clonally isolated permanent and stable human MSC lines by transfecting primary cell cultures of fetal human bone marrow MSCs with a retroviral vector encoding v-myc gene. One of the cell lines, HM3.B10 (B10), was found to differentiate into neural cell types including neural stem cells, neurons, astrocytes and oligodendrocytes in vitro as shown by expression of genetic markers for neural stem cells (nestin and Musashi1), neurons (Neurofilament Protein, synapsin and MAP2), astrocytes (glial fibrillary acidic Protein, GFAP) and oligodendrocytes (myelin basic Protein, MBP) as determined by RT-PCR assay. In addition, B10 cells were found to differentiate into neural cell types as shown by immunocytochical demonstration of nestin (for neural stem cells), Neurofilament Protein and b-tubulin III (neurons) GFAP (astrocytes), and galactocerebroside (oligodendrocytes). Following brain transplantation in mouse ICH stroke model, B10 human MSCs integrate into host brain, survive, differentiate into neurons and astrocytes and induce behavioral improvement in the ICH animals. B10 human MSC cell line is not only a useful tool for the studies of organogenesis and specifically for the neurogenesis, but also provides a valuable source of cells for cel
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Neuronal phenotype and neuronal differentiation of B10 cells.
2013Co-Authors: Atsushi Nagai, Woo K. Kim, Hong J. Lee, Han S. Jeong, Kwang S. Kim, Seok H. Hong, In H. Park, Seung U. KimAbstract:Gene expression of stem cell markers and neuronal markers was examined by RT-PCR. B10 cells reproduce genetic features of primary human bone marrow MSCs by expressing ABCG2 and Oct-4 as general stem cell markers, Mash 1 and Otx 2 as neurogenic bHLH genes, nestin as a cell type specific marker for neural stem cells, and low-molecular weight Neurofilament Protein (NF-L), intermediate molecular weight NF (NF-M) and high molecular weight NF (NF-H) as specific markers for neurons. B10 cells were grown in medium containing basic FGF for 2 weeks, In these conditions, B10 cells were demonstrated to differentiate into neurons.
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multilineage potential of stable human mesenchymal stem cell line derived from fetal marrow
PLOS ONE, 2007Co-Authors: Atsushi Nagai, Woo K. Kim, Hong J. Lee, Han S. Jeong, Kwang S. Kim, Seok H. Hong, In H. ParkAbstract:Human bone marrow contains two major cell types, hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs). MSCs possess self-renewal capacity and pluripotency defined by their ability to differentiate into osteoblasts, chondrocytes, adipocytes and muscle cells. MSCs are also known to differentiate into neurons and glial cells in vitro, and in vivo following transplantation into the brain of animal models of neurological disorders including ischemia and intracerebral hemorrhage (ICH) stroke. In order to obtain sufficient number and homogeneous population of human MSCs, we have clonally isolated permanent and stable human MSC lines by transfecting primary cell cultures of fetal human bone marrow MSCs with a retroviral vector encoding v-myc gene. One of the cell lines, HM3.B10 (B10), was found to differentiate into neural cell types including neural stem cells, neurons, astrocytes and oligodendrocytes in vitro as shown by expression of genetic markers for neural stem cells (nestin and Musashi1), neurons (Neurofilament Protein, synapsin and MAP2), astrocytes (glial fibrillary acidic Protein, GFAP) and oligodendrocytes (myelin basic Protein, MBP) as determined by RT-PCR assay. In addition, B10 cells were found to differentiate into neural cell types as shown by immunocytochical demonstration of nestin (for neural stem cells), Neurofilament Protein and β-tubulin III (neurons) GFAP (astrocytes), and galactocerebroside (oligodendrocytes). Following brain transplantation in mouse ICH stroke model, B10 human MSCs integrate into host brain, survive, differentiate into neurons and astrocytes and induce behavioral improvement in the ICH animals. B10 human MSC cell line is not only a useful tool for the studies of organogenesis and specifically for the neurogenesis, but also provides a valuable source of cells for cell therapy studies in animal models of stroke and other neurological disorders.