Stimulations of the Culture Medium of Activated Microglia
and TNF-Alpha on a Scrapie-Infected Cell Line Decrease the Cell Viability
and Induce Marked Necroptosis That Also Occurs in the Brains from
the Patients of Human Prion Diseases
posted on 2018-11-06, 00:00authored byYue Ma, Qi Shi, Kang Xiao, Jing Wang, Cao Chen, Li-Ping Gao, Chen Gao, Xiao-Ping Dong
Activation
of microglia and increased expression of TNF-α
are frequently observed in the brains of human and animal prion diseases.
As an important cytokine, TNF-α participates in not only pro-inflammatory
responses but also in cellular communication, cell differentiation,
and cell death. However, the role of TNF-α in the pathogenesis
of prion disease remains ambiguous. In this study, the activities
of a scrapie-infected cell line SMB-S15 and its normal partner SMB-PS
exposed to the supernatant of a LPS-activated microglia cell line
BV2 were evaluated. After it was exposed to the LPS-stimulated supernatant
of BV2 cells, the cell viability of SMB-S15 cells was markedly decreased,
whereas that of the SMB-PS cells remained unchanged. The level of
TNF-α was significantly increased in the LPS-stimulated supernatant
of BV2 cells. Further, we found that the recombinant TNF-α alone
induced the decreased cell viability of SMB-S15 and the neutralizing
antibody for TNF-α completely antagonized the decreased cell
viability caused by the LPS-stimulated supernatant of BV2 cells. Stimulation
with TNF-α induced the remarkable increases of apoptosis-associated
proteins in SMB-PS cells, such as cleaved caspase-3 and RIP1, whereas
an obvious increase of necroptosis-associated protein in SMB-S15 cells,
such as p-MLKL. Meanwhile, the upregulation of caspase-8 activity
in SMB-PS cells was more significant than that of SMB-S15 cells. The
decreased cell viability of SMB-S15 and the increased expression of
p-MLKL induced by TNF-α were completely rescued by Necrostatin-1.
Moreover, we verified that removal of PrPSc propagation
in SMB-S15 cells by resveratrol partially rescues the cell tolerance
to the stimulation of TNF-α. These data indicate that the prion-infected
cell line SMB-S15 is more vulnerable to the stimulations of activated
microglia and TNF-α, which is likely due to the outcome of necroptosis
rather than apoptosis. Furthermore, significant upregulation of p-MLKL,
MLKL, and RIP3 was detected in the post-mortem cortical brains of
the patients of various types of human prion diseases, including sporadic
Creutzfeldt-Jakob disease (sCJD), G114 V-genetic CJD (gCJD), and fatal
familial insomnia (FFI).