Neuroblastoma is a tumor stemming from neural crest cells that affects the nervous system (Riley 2004). Neuroblastoma tumors typically grow in young children under the age of four. In fact, 8-10 % of cancers found in children are neuroblastoma tumors (Riley 2004). The severity of the cancer ranges from highly malignant to spontaneous regression (Riley 2004). This range in the severity of the cancer may be due to the unique ability of the neuroblastoma cells to differentiate. Human neuroblastoma cells are phenotypically variable. N-type cells are small and rounded with cytoplasmic projections, and have weak substrate adherence (Biedler et al. 1997). Alternatively, S-type cells are larger, lack cell processes, and are highly adherent to substrates (Biedler et al. 1997). A third type, the I-type, is a phenotypic intermediate between the S and N-types. Aside from their morphological differences, phenotypically distinct cells can further be distinguished from each other by their biochemical characteristics. Many enzymes and immune response surface proteins expressed in the N-type are not expressed in the S- types (Biedler et al 1997). Additionally, S- type cells do not have neuronal activity (Biedler et al. 1997). The physical and biochemical properties in cell neuroblastoma cell types are important and research on differential cell characteristics may lead to a more complete cure in children who display more severe cases of the cancer.
Figure 1- Differential gene expression of FAS gene isoforms in neuroblastoma N-type SH-SY5Y (N), S-type SH-EP1 (S), and I- type BE (2)-C (I) cell lines and a negative control (-).
Figure 2- Differential gene expression of PDK1 gene splice variants in neuroblastoma N-type SH-SY5Y (N), S-type SH-EP1 (S), and I- type BE (2)-C (I) cell lines and a negative control (-).
Figure 3- Differential gene expression of TP53I3 gene splice variants in neuroblastoma N-type SH-SY5Y (N), S-type SH-EP1 (S), and I- type BE (2)-C (I) cell lines and a negative control (-).
Figure 4- Splice variants amplified during RT-PCR. The 200 bp isoform of PDK-1 is lacking a piece of intron 3. The 265 bp isoform of TP53I3 lacks exon 4.
Alternatively spliced mRNAs are functionally different from one another, so it is important to recognize what variants are expressed and the consequences of that expression. Overall, alternative splicing was displayed in the neuroblastoma cells as well as differential expression of variants.
The malignant N-type SH-SY5Y cells appeared to have higher isoform expression ratios based on band intensity of the different variants.
The malignant I-type BE (2)-C is a phenotypic intermediate between the N and S cell types. Expression ratios of splicing variants in the I-type were low and variants were expressed more equally.
Lower expression ratios of slice variants S and I-type in the Tp53I3 gene were observed, which means that the more easily degraded 265 bp gene variant (Nicholls et al. 2004) is more equally expressed than the 462 bp variant.
Future research should include a more intensive look at the functional consequences of varying expression in neuroblastoma cell lines.
I would like to thank Joe Frezzo and Leleesha Samaraweera for their help throughout the course. I would also like to thank Leleesha Samaraweera for providing neuroblastoma cell types and Bo Liu for providing primer sets. I would also like to acknowledge Dr. Berish Rubin, Dr. John Wehr, and Dr. Sylvia Anderson for their assistance and guidance.
|This document was last modified 05/09/2007.|
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