OBJECTIVEMachado-Joseph disease (MJD)/Spinocerebellar ataxia type 3 (SCA3) is an autosomal dominant neurodegenerative disorder caused by an expansion of polyglutamine tract near the C-terminus of the MJD1 gene product, ataxin-3. The precise mechanism of the MJD/SCA3 pathogenesis remains unclear. A growing body of evidence demonstrates that phosphorylation plays an important role in the pathogenesis of many neurodegenerative diseases. However, few kinases are known to phosphorylate ataxin-3. The present study is to explore whether ataxin-3 is a substrate of casein kinase 2 (CK2).

METHODSThe interaction between ataxin-3 and CK2 was identified by glutathione S-transferase (GST) pull-down assay and co-immunoprecipition assay. The phosphorylation of ataxin-3 by CK2 was measured by in vitro phosphorylation assays. Results (1) Both wild type and expanded ataxin-3 interacted with CK2alpha and CK2beta in vitro. (2) In 293 cells, both wild type and expanded ataxin-3 interacted with CK2beta, but not CK2alpha. (3) CK2 phosphorylated wild type and expanded ataxin-3.

CONCLUSIONAtaxin-3 is a substrate of protein kinase CK2.

Ataxin-3 ; Casein Kinase II ; metabolism ; Cell Line, Transformed ; Glutathione Transferase ; metabolism ; Humans ; Immunoprecipitation ; methods ; Nerve Tissue Proteins ; metabolism ; Nuclear Proteins ; metabolism ; Phosphorylation ; Repressor Proteins ; metabolism ; Transfection ; methods

OBJECTIVEMachado-Joseph disease (MJD), also known as spinocerebellar ataxia type 3 (SCA3), is a dominant neurodegenerative disorder caused by an expansion of the polyglutamine (polyQ) tract in MJD-1 gene product, ataxin-3 (AT3). This disease is characterized by the formation of intraneuronal inclusions, but the mechanism underlying their formation is still poorly understood. The present study is to explore the relationship between wild type (WT) AT3 and polyQ expanded AT3.

METHODSMouse neuroblastoma (N2a) cells or HEK293 cells were co-transfected with WT AT3 and different truncated forms of expanded AT3. The expressions of WT AT3 and the truncated forms of expanded AT3 were detected by Western blotting, and observed by an inverted fluorescent microscope. The interactions between AT3 and different truncated forms of expanded AT3 were detected by immunoprecipitation and GST pull-down assays.

RESULTSUsing fluorescent microscope, we observed that the truncated forms of expanded AT3 aggregate in transfected cells, and the full-length WT AT3 is recruited onto the aggregates. However, no aggregates were observed in cells transfected with the truncated forms of WT AT3. Immunoprecipitation and GST pull-down analyses indicate that WT AT3 interacts with the truncated AT3 in a polyQ length-dependent manner.

CONCLUSIONWT AT3 deposits in the aggregation that was formed by polyQ expanded AT3, which suggests that the formation of AT3 aggregation may affect the normal function of WT AT3 and increase polyQ protein toxicity in MJD.

Animals ; Ataxin-3 ; Blotting, Western ; Cell Line ; Immunoprecipitation ; Machado-Joseph Disease ; metabolism ; Mice ; Microscopy, Fluorescence ; Nuclear Proteins ; genetics ; metabolism ; Peptides ; metabolism ; Transcription Factors ; genetics ; metabolism ; Transfection

OBJECTIVETo evaluate the clinical effects of carotid endarterectomy for carotid stenosis and occlusion.

METHODSFrom August 2005 to November 2008 moderate and severe carotid stenosis or occlusion were found in 16 patients by Doppler ultrasonography (DUS), MRA, CTA, DSA. The stenosis degree ranged from 60% to 99% in 14 patients and complete occlusion in 2 patients. Twelve patients underwent standard carotid endarterectomy (sCEA) in whom 2 patients were placed carotid shunt and 1 patient underwent carotid patch angioplasty. Four patients underwent eversion carotid endarterectomy (eCEA). All operations were performed by microscope.

RESULTSThere was no stroke, transient ischemic attack and mortality perioperatively and during follow-up from 1 month to 3 years. The ICA flow detected by follow-up duplex scan and MRA was unobstructed. The primary cerebral ischemic symptoms were obviously improved or disappeared after operation. The postoperative complications included one case of upper gastrointestinal hemorrhage and one case of hoarseness and bucking, which disappeared after medical treatment.

CONCLUSIONSCEA is an effective way for treating carotid stenosis. Different operative methods and techniques deal with different carotid lesions to achieve better effect. Microsurgical technique is useful for exposure of high ICA bifurcation and avoid effectively cranial nerve injury and other complications.

Adult ; Aged ; Carotid Stenosis ; surgery ; Endarterectomy, Carotid ; methods ; Female ; Follow-Up Studies ; Humans ; Male ; Microsurgery ; Middle Aged ; Treatment Outcome