Vitamin B12-impaired metabolism produces apoptosis and Parkinson phenotype in rats expressing the transcobalamin-oleosin chimera in substantia nigra.
BACKGROUND
Vitamin B12 is indispensable for proper brain functioning and cytosolic synthesis of S-adenosylmethionine. Whether its deficiency produces effects on viability and apoptosis of neurons remains unknown. There is a particular interest in investigating these effects in Parkinson disease where Levodopa treatment is known to increase the consumption of S-adenosylmethionine. To cause deprivation of vitamin B12, we have recently developed a cell model that produces decreased synthesis of S-adenosylmethionine by anchoring transcobalamin (TCII) to the reticulum through its fusion with Oleosin (OLEO).


METHODS
Gene constructs including transcobalamin-oleosin (TCII-OLEO) and control constructs, green fluorescent protein-transcobalamin-oleosin (GFP-TCII-OLEO), oleosin-transcobalamin (OLEO-TCII), TCII and OLEO were used for expression in N1E-115 cells (mouse neuroblastoma) and in substantia nigra of adult rats, using a targeted transfection with a Neurotensin polyplex system. We studied the viability and the apoptosis in the transfected cells and targeted tissue. The turning behavior was evaluated in the rats transfected with the different plasmids.


RESULTS
The transfection of N1E-115 cells by the TCII-OLEO-expressing plasmid significantly affected cell viability and increased immunoreactivity of cleaved Caspase-3. No change in propidium iodide uptake (used as a necrosis marker) was observed. The transfected rats lost neurons immunoreactive to tyrosine hydroxylase. The expression of TCII-OLEO was observed in cells immunoreactive to tyrosine hydroxylase of the substantia nigra, with a superimposed expression of cleaved Caspase-3. These cellular and tissular effects were not observed with the control plasmids. Rats transfected with TCII-OLEO expressing plasmid presented with a significantly higher number of turns, compared with those transfected with the other plasmids.


CONCLUSIONS
In conclusion, the TCII-OLEO transfection was responsible for apoptosis in N1E-115 cells and rat substantia nigra and for Parkinson-like phenotype. This suggests evaluating whether vitamin B12 deficit could aggravate the PD in patients under Levodopa therapy by impairing S-adenosylmethionine synthesis in substantia nigra.
Orozco-Barrios
Carlos Enrique
CE
Department of Physiology, Biophysics and Neuroscience, Centro de Investigación y Estudios Avanzados del Instituto Politécnico Nacional (CINVESTAV), Mexico City, Mexico.
Battaglia-Hsu
Shyue-Fang
SF
Arango-Rodriguez
Martha Ligia
ML
Ayala-Davila
Jose
J
Chery
Celine
C
Alberto
Jean-Marc
JM
Schroeder
Henry
H
Daval
Jean-Luc
JL
Martinez-Fong
Daniel
D
Gueant
Jean-Louis
JL
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001522:Behavior, Animal / Q000187:drug effects; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D006801:Humans; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism; D008694:Methamphetamine / Q000494:pharmacology; D051379:Mice; D009336:Necrosis; D010300:Parkinson Disease / Q000473:pathology*; D010940:Plant Proteins / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D021381:Protein Transport / Q000187:drug effects; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D013378:Substantia Nigra / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D014155:Transcobalamins / Q000378:metabolism*; D014162:Transfection; D019076:Transgenes / Q000235:genetics; D014805:Vitamin B 12 / Q000378:metabolism*
D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; D014155:Transcobalamins; D008694:Methamphetamine; D014805:Vitamin B 12