Anthrax Protective Antigen 63 (PA63): Toxic Effects in Neural Cultures and Role in Gulf War Illness (GWI)

Effie-Photini C Tsilibary; Eric P Souto; Marian Kratzke; Lisa M James; Brian E Engdahl; Apostolos P Georgopoulos

doi:10.1177/2633105520931966

Anthrax Protective Antigen 63 (PA63): Toxic Effects in Neural Cultures and Role in Gulf War Illness (GWI)

Neurosci Insights. 2020 Jun 30:15:2633105520931966. doi: 10.1177/2633105520931966. eCollection 2020.

Authors

Effie-Photini C Tsilibary^{1

2}, Eric P Souto¹, Marian Kratzke^{1

2}, Lisa M James^{1

2

3}, Brian E Engdahl^{1

2

4}, Apostolos P Georgopoulos^{1

2

3

5}

Affiliations

¹ Brain Sciences Center, Department of Veterans Affairs Health Care System, Minneapolis, MN, USA.
² Department of Neuroscience, Medical School, University of Minnesota, Minneapolis, MN, USA.
³ Department of Psychiatry, Medical School, University of Minnesota, Minneapolis, MN, USA.
⁴ Department of Psychology, University of Minnesota, Minneapolis, MN, USA.
⁵ Department of Neurology, Medical School, University of Minnesota, Minneapolis, MN, USA.

Abstract

Protective antigen (PA) 63 (PA63) is a protein derived from the PA83 component contained in the anthrax vaccine. The anthrax vaccine ("Biothrax") was administered together with other vaccines to Gulf War veterans, about 35% of whom later developed a multisymptom disease (Gulf War Illness [GWI]), with prominent neurological/cognitive/mood symptoms, among others. The disease has been traditionally attributed to exposures to toxic chemicals during the war but other factors could be involved, including vaccines received. Of these, the anthrax vaccine is the most toxic. Here, we assessed directly the PA63 toxin's harmful effects on cultured neuroblastoma 2A (N2A) cells with respect to cell spreading, process formation, apoptosis, and integrity of cell membrane, cytoskeleton, and mitochondria. We found that, when added in N2A cultures, PA63 toxin led to decreased cell spreading and cell aggregation, leading to apoptosis. The mechanisms of PA63-induced cell damage included compromised cell membrane permeability indicated by enhanced access of propidium iodide in cells. In addition, signaling pathways leading to organization of N2A cytoskeleton were negatively affected, as both actin and microtubular networks were compromised. Finally, the mitochondrial membrane potential was impaired in specific assays. Altogether, these alterations led to apoptosis as a collective toxic effect of PA63 which was substantially reduced by the concomitant addition of specific antibodies against PA63.

Keywords: Gulf War Illness; N2A cultures; anthrax PA63; apoptosis; cytoskeleton; membrane permeability; mitochondrial membrane potential.