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Major Accomplishments
·
Conceived
and spearheaded a project to investigate synaptic mRNAs in neurodegenerative
diseases such as Alzheimer’s Disease (AD) (5).
Patients and brain area (frontal cortex) were selected to investigate early Aß induced molecular change
at the synapse that
may be reversible. The novel idea of isolating synaptic mRNAs from patients
using synaptoneurosome preparations and obtaining
mRNA expression profiles generated a unique set of synaptic mRNAs modified in
patients with incipient AD. Many of these genes bear consensus sequences in
the 3`UTR which regulate translation (5).
Dysregulation of synaptic mRNA may be an
important component of neurological diseases, not only neurodegenerative but
also developmental (as in Fragile X) and psychiatric (schizophrenia).
·
Discovered
unique localization of a monoclonal antibody to neurons of the spinocerebellar system in human and mouse CNS (2,3). Determined that spinocerebellar
neurons in the spinal cord (spinal border neurons and Clarke’s column) are
targeted in amyotrophic lateral sclerosis albeit at a slower rate than motorneurons (2). Isolated and characterized the target
protein as a chondroitin sulfate proteoglycan, somataglycan
(3). Determined that somataglycan is localized to
horizontal neurons in the retina (4).
·
Developed
a technique for isolating large neurons such as spinal or A4 motorneurons from flash frozen CNS tissue obtained at
autopsy. Isolated mRNA from ~ 200 neurons and used it to generate cDNA libraries via the SMART cDNA
library construction kit. The libraries were used to generate double-stranded
DNA and then, via an assymetric PCR technique, biotinylated cDNA. The labeled cDNA
was used for expression profiling on Superarrays .
Techniques
Biochemistry
Protein
purification and analysis Gel filtration, affinity chromatography, DEAE anion
exchange chromatography,immunoaffinity
purification, differential centrifugation using discontinous
sucrose gradients, preparation and running of gradient polyacrylamide gels,
Western immunoblots, glycosidase and lyase digests of glycoconjugates,
ELISA, protein assay, ligand binding assay
Cell culture:
Clonal cell line culture: Maintenance of glial,neuronal and fibroblast cell lines, viral plaque
assay
Primary cultures: Primary cortical cultures from mice grown in a
microfluidic platform. Co-cultures of spinal cord and muscle from embryonic
mice, in vitro sensitization of thymocytes
or spleen cells for hybridoma preparation, feeder
cell and conditioned medium preparation maintenance of hybridoma
cell lines , including sub-cloning, B-cell cloning
Immunocytochemistry:
Immunofluorescence and immunoperoxidase including
double labeling. Knowledge of anatomy and fine structure of the nervous system,
especially the spinal cord, frontal cortex and hippocampus Tissue preparation
for cryostat sectioning
Microscopy and Imaging:
Installation of digital imaging system consisting of Zeiss Axiophot microscope with a Spot digital camera hooked up
to a Dell Precision 530 workstation. Preparation of digital images for
publication using Photoshop and slides for presentation using Powerpoint.
Molecular :
Amplification of mRNA from small sample of enriched neuronal sub-populations
(~100cells) using magnetic beads and SMART system. PCR, RT-PCR, asymetric PCR for probe preparation. Agarose
gel, Southern, Superarray and preparation of
samples for microarray. Labeling with 32P or biotin. cDNA library screening. Isolation of DNA clone from
phagemid and purification Labelling
of clone with biotin [in situ hybridization]
Manuscript Preparation: All
aspects including writing, editing and figure preparation. Figures prepared
digitally and submitted as tif files.Electronic
manuscript prepared and published at the 1st Electronic Glycoconjugate
Conference [1995] and to PLOS 2009.
Computing : HTML see-ITP
pages , Administrator
of computer systems, XP including all hardware, software and network
troubleshooting. Expertise in Photoshop, Illustrator, Powerpoint,
Word, Excel, Access, Image-Pro Plus, Partek
Microarray Suite(5), Ingenuity Pathway Analysis (5)
.
Publications
1. Kozlowski, M.A., C.Williams, D.R. Hinton, C.A.
Miller.[1989] Heterotopic neurons in the spinal cords of patients with ALS.
Neurology 39:644-648.
2. Williams, C., M. Kozlowski, D.R. Hinton, C.A. Miller.[1990]
Degeneration of spinocerebellar tract neurons in
ALS. Annals of Neurol. 27: 215-225.
3. Williams C., D.R. Hinton, C. A. Miller.[1994] Somataglycan-S: A neuronal surface proteoglycan defines
the spinocerebellar system. J.Neurochem.
4 ,1615-1630.
4. Williams C., M.Villegas, C.A. Miller. [1998]
Chondroitin Sulfate Proteoglycans Specific to Retinal Horizontal Neurons J.Comp. Neurology,390:268-277 (PDF)
5. Williams C, Mehrian
Shai R, Wu Y, Hsu Y-H, Sitzer
T, Spann B, McCleary C, Mo Y, Miller CA. Transcriptome analysis
of synaptoneurosomes identifies neuroplasticity
genes overexpressed in incipient Alzheimer’s disease. PLoS ONE 2009;
4(3):e4936. http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0004936
6. Vu M.G., Li, Q., Fisher, B., Nacca, A., Leahy, R.M., Walsh, J.P., Mukherjee, J.,
Williams, C., Jakowec, M.W., and Petzinger, G.M. Exercise elevates dopamine D2 receptor in
a mouse model of Parkinson's disease: In vivo imaging with [18F]fallypride. Movement Disorders
2010; 25,
2777-2784.
1.
Abstracts
1.Miller,C.A., D.R. Hinton, C.Williams,
V.Henderson.[1986] Neuronal subsets in the human
central nervous system defined by monoclonal antibodies. J. Neuropath. Exp.
Neurol. 45:345.
2.Williams,C., D.R. Hinton, C.A. Miller.[1986]
Neuronal subsets in human spinal cord and in murine spinal cord-muscle
cultures identified by monoclonal antibodies Muscle and Nerve 9:247.
3. Williams,C., C,A. Miller, D.R. Hinton.[1986]
Neuronal subsets identified by monoclonal antibodies in human spinal cord and
in murine spinal cord-muscle cultures. J. Neuropath. Exp. Neurol. 45: 364.
4. Henderson,V.W., D.R. Hinton,C.Williams,
C.A. Miller.[1986] Organization of the human nervous system revealed by
monoclonal antibodies. Neurology 36: 253.
5. Williams C., M. Kozlowski, C.A. Miller.[1987]
Spinal border cells in ALS. J. Neuropath. Exp. Neurol. 46: 6. Miller, C.A.,M. Kozlowski, C.Williams, D.R.
Hinton.[1988] Ectopic spinal cord neurons in ALS. J. Neuropath. Exp. Neurol.
47:8.
7. Williams, C., D.R. Hinton, C.A.Miller.[1989] Spinocerebellar tract neurons detected in murine spinal
cord by monoclonal antibody 6A2. J. Neuropath.Exp.
Neurol. 48:363.
8. Williams, C., L.R. Reiter, G. Pegolo, S. Benzer, C.A.Miller.[1990] Glycoconjugates
associated with spinocerebellar neurons are
identified by MAb 6A2. J. Neuropath. Expt. Neurol.
49:278. 9. Williams, C., L.R. Reiter, D.R. Hinton, S. Benzer,
C.A. Miller.[1991] A neuronal surface sub-population
specific proteoglycan in mouse and Drosophila. Soc. Neurosci.
Abstr. 17:575
10. Williams, C.,E. Pomerantz,
R.A. Atkinson,W.K.Engel, C.A. Miller.[1992] Serum Igs bind to surfaces of isolated motorneurons.
J. Neuropath.Expt. Neurol. 51:
11. Atkinson, R.A., C. Williams,M.Nawrot,W.K.
Engel, C.A. Miller.[1993] Sera of patients with lower motorneuron[LMN]
degeneration bind to surface proteoglycan of LMN. J.Neuropath.
Expt. Neurol. 52:292.
12. Williams C., M.Villegas, C.A.Miller.[1994] Somataglycan: An
interneuron specific proteoglycan. Glycotechnology
13. LeMay L.G., Williams C. and Miller CA.[1995] Somataglycan from motor
cortex of human brain binds neurotrophins NGF,NT-3,
NT-4/5. J.Neuropath.Exp.Neurol., 54:443
14. LeMay L.G., Williams C. and Miller CA.[1995] Somataglycan from motor
cortex of human brain binds nerve growth factor and neurotrophin-4/5. J. Neurochem. 64:95A 15.
16. LeMay LG, Williams C, Miller CA [1995] Somataglycan-S from motor cortex of human brain binds neurotrophins; NGF, NT-3, NT 4/5. J Neuropath Exp Neurol 54(3):443.
17. Williams C., Miller CA [1995] Somataglycan
isolated from human cerebral cortex: Possible molecular diversity conferred
by glycoaminoglycans. 1st Electronic Glycoconjugate Conference (EGC-1).
18. Atkinsons RD, C. Williams, Engel WK, Miller CA.
[1995] Serum autoantibodies other than anti-GM-1 in the pathogenesis of
progressive muscular atrophy. Soc for Neurosci 21:1006.
19. Atkinson RD, C. Williams, Engel WK, Miller CA [1996] Progressive muscular
atrophy autoantibodies bind to lower motorneurons.
J Neuropath Exp Neurol
55(5):104.
20. Williams C., Villegas M, Atkinson RA, Miller CA [1998] Chondroitin
sulfate proteoglycan specific to retinal horizontal neurons. J Comp Neurol 390:268-277.
21. Williams C., Villegas M, Miller CA. [1998] Human motorneuron
cells lines [NSC] express Trk receptors. J
Neuropath & Exp Neurol
57:486.69.
22. Rose AT, Salas E, C. Williams, Mack W, Miller CA [2001] Vascular
proliferation in aging and Alzheimer’s disease. J Neuropath & Exp Neurology 60(5):544.165.
23. Del Villar K, C. Williams, Miller CA. [2004]
DENN/MADD Down-regulation and synaptic terminal dysfunction in AD. J
Neuropath & Exp Neurol
63(552):170.
24. Williams C., Miller CA.[2004] mRNA expression in
Alzheimer’s disease-affected synaptosomes. The
Cytoskeleton & Synaptic Function, San Diego, CA, October.
25. Lin C, C. Williams, Zhou L, Sonnen J, McCleary C, Spann BM, Miller CA. [2005] Activation of
neurogenesis genes and synaptic terminal regeneration in AD. J Neuropath Exp Neurol 64(5):39.
26. Williams C., Mehrian Shai
R, Wu Y, Miller CA. [2006] Expression of mRNAs Regulating Local Translation
in Synaptic Terminals of MCI and Early AD Brain. Alzheimer's & Dementia:
The Journal of the Alzheimer's Association July
2006 (Vol. 2, Issue 3 (Supplement), Pages S505-S506).
27. Williams C, Miller CA. Comparison of GluR2 regulation in 3xTg-AD
mice and incipient AD patients. AANP
Annual Meeting, Philadelphia, PA, June 9-14, 2010. (Vol. 2, Issue 3 (Supplement), Pages
S505-S506)
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