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1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine-resistant, flat-cell PC12 variants having a partial loss of transformed
J K Andersen1, M B Zhang, X H Zhong
1Department of Biological Chemistry, School of Medicine, University of California, Los Angeles 90024.
Abstract:
We have cloned and characterized two variants of PC12 cells. MPT1 cells were selected by their resistance to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), and variant 2068 was isolated nonselectively as a large, flat-cell variant commonly occurring in PC12 cultures. Variant 2068 cells also exhibit resistance to MPTP. Karyotype analysis revealed that these variants are true derivatives of wild-type PC12 cells; however, each variant is tetraploid, whereas the wild-type parent is diploid. The two variants contain an altered level and composition of lactate dehydrogenase isoenzymes, which could account for a previously described difference in lactate metabolism. Both variants exhibit a partial loss of transformed phenotype in culture in that they are nonrefractile, grow in monolayers, and fail to multiply in soft agar. We suggest that this alteration in transformed phenotype may result in altered mitochondria and lactate dehydrogenase and thus account for their resistance to MPTP. Compared with wild-type PC12 cells, MPT1 cells have a decreased level of fos mRNA and an increased level of myc mRNA; the latter results from an increased level of transcription of exon 1 of the myc gene. Studies with hybrid cells obtained by fusing MPT1 cells with wild-type-like cells show that most, but not all, of the parameters of the MPT1 phenotype predominate.
Insights
Two PC12 cell variants, MPT1 and 2068, exhibit resistance to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). These tetraploid cells show altered lactate dehydrogenase and a partial loss of transformed phenotype, contributing to MPTP resistance.
Area of Science:
- Cell Biology
- Neuroscience
- Biochemistry
Background:
- PC12 cells are a widely used model for neuronal differentiation and toxicology.
- 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is a neurotoxin that selectively destroys dopaminergic neurons, serving as a model for Parkinson's disease.
Purpose of the Study:
- To characterize two novel PC12 cell variants, MPT1 and 2068, selected for resistance to MPTP.
- To investigate the underlying molecular and cellular mechanisms contributing to MPTP resistance in these variants.
Main Methods:
- Cell culture and characterization of MPT1 and 2068 variants.
- Karyotype analysis to determine ploidy.
- Lactate dehydrogenase (LDH) isoenzyme analysis.
- Assessment of transformed phenotype markers (soft agar growth, morphology).
- mRNA expression analysis (fos, myc).
- Hybrid cell studies.
Main Results:
- MPT1 and 2068 variants are tetraploid derivatives of PC12 cells and exhibit resistance to MPTP.
- Both variants show altered LDH isoenzyme levels and composition, potentially affecting lactate metabolism.
- A partial loss of transformed phenotype was observed, including non-refractile growth in monolayers and inability to multiply in soft agar.
- MPT1 cells display decreased fos mRNA and increased myc mRNA levels, with enhanced myc exon 1 transcription.
- Hybrid cell studies indicate that MPT1 phenotype traits are largely dominant.
Conclusions:
- The characterized PC12 cell variants (MPT1 and 2068) offer valuable models for studying MPTP neurotoxicity and resistance mechanisms.
- Alterations in lactate metabolism, mitochondrial function, and gene expression (myc, fos) likely contribute to MPTP resistance.
- The loss of transformed phenotype in these variants is associated with MPTP resistance, suggesting a link between cellular transformation and neurotoxin sensitivity.