Acta Neurol zyxwvutsrqponmlkj Scand 1999: 100: 317-321 Printed in UK. All rights reserved Coorviaht zyxw & Munbaaurd 1999 zy ACTA NEUROLOGICA SCANDINAVICA ISSN 0001-6314 zyx Upregulation of Bax protein and increased DNA degradation in ALS spinal cord motor neurons Ekegren T, Grundstrom E, Lindholm D, Aquilonius S-M. Upregulation of Bax protein and increased DNA degradation in ALS spinal cord zyxw T. Ekegren', E. Grundstrom', D. Lindholm*, S.-M. Aquilonius' motor neurons. Acta Neurol Scand 1999: 100: 3 17-32 1. zyxwvu 0 Munksgaard 1999. Objectives - To investigate if degeneration of motor neurons in amyotrophic lateral sclerosis (ALS) is related to altered levels of the apoptosis regulating proteins Bcl-2 and Bax. In addition, immuno- reactivity of the cysteine protease ICH-1L and detection of motor neurons with DNA fragmentation, indicative of apoptosis, was also studied. Material and methods - The immunoreactivity of Bcl-2, Bax and ICH-1L were compared in ALS and control spinal cord motor neurons by immunohistochemical analysis and motor neurons with DNA fragmentation were identified by the TUNEL-method. Results - The results demonstrate an increased expression of Bax in the ALS material as compared to controls but no change in Bcl-2 and ICH-1L expressions. Moreover, a larger proportion of motor neurons stained positive for TUNEL in ALS spinal cords. Conclusion - Present study suggest an upregulation of the cell death promoting protein Bax and increased DNA degradation, indicative of apoptosis, in spinal motor neurons of ALS patients. Amyotrophic lateral sclerosis (ALS) is a neurode- generative disease clinically characterized by pro- gressive weakness and wasting of muscles caused by loss of motor neurons in the spinal cord, brain stem and motor cortex. Even though many hypotheses regarding the neurodegeneration in ALS have been proposed the full etiology is still unknown. Lately, attention has been drawn to apoptotic cell death in ALS. Thus, there have been reports on increased DNA degradation (1, 2) as well as expression of Ley, an antigen characteristic of cells undergoing apoptosis (3), in spinal motor neurons of ALS patients. In line with this, altered mRNA expres- sions of the apoptotic cell death genes bcl-2 (B-cell lymphoma/leukemia-2) and bax (Bcl-2-associated X protein) have been reported in ALS spinal cord motor neurons (4). Alterations in gene expressions of bcl-2 and bax might not be reflected at the protein Department of Neuroscience 'Neurology, University Hospital. and 2Developmental Neuroscience. Biomedical Center. Uppsala. Sweden Key words amyotrophic lateral sclerosis (ALS). 8cl-2. Bax. ICH-1 L, neurodegeneration. TUNEL T i t t Ekegren. Department of Neuroscience. Neurology, University Hospital, S-751 85 Uppsala, Sweden Accepted for publication May 18, 1999 levels. Indeed this may also indicate a posttran- scriptional regulation of the bcl-2 gene (5, 6). Bcl-2 is the founder member of a family of regulating proteins, functioning as a central repres- sor of cell death. Another family member, Bax, on the contrary works as an accelerator of cell death, Bax can act both as a homodimer and a hetero- dimer. In the homodimeric state Bax functions as a cell death promoter whereas in the heterodimeric state, attached to Bcl-2, it represses Bcl-2's anti- apoptotic effect. This led to a model in which the ratio of Bcl-2 to Bax determines if cells will survive or die from an apoptotic stimuli (4, 7, 8). ICH-lL (Ice and ced-3 homolog, the long protein) is a cysteine protease in the ICE/CED-3 family which induces apoptosis when overexpressed (9). The morphological characteristics of apoptosis are cell shrinkage, chromatin condensation, DNA fragmen- z 317