Fibrinogen-cellular prion protein complex formation on astrocytes

Abstract

Traumatic brain injury (TBI) is one of the most common neurological disorders causing memory reduction, particularly short-term memory (STM). We showed that during TBI-induced inflammation, increased blood content of fibrinogen (Fg) enhanced vascular protein transcytosis and deposition of extravasated Fg in vasculo-astrocyte interfaces. In addition, we found that deposition of cellular prion protein (PrPC) was also increased in the vasculo-astrocyte endfeet interface. However, association of Fg and PrPC was not confirmed. Presently, we aimed to define whether Fg can associate with PrPC on astrocytes and cause their activation. Cultured mouse brain astrocytes MBA were treated with medium alone (control), Fg (2 mg/ml or 4 mg/ml), 4 mg/ml of Fg in the presence of a function-blocking anti-PrPC peptide or anti-mouse IgG, function-blocking anti-PrPC peptide, or anti-mouse IgG alone. After treatment, either cell lysates were collected and analyzed via Western blot or co-immunoprecipitation was performed, or astrocytes were fixed and their activation was assessed with immunohistochemistry. Results showed that Fg dose-dependently activated astrocytes, increased expressions of PrPC and tyrosine (tropomyosin) receptor kinase B (TrkB), and PrP gene. Blocking the function of PrPC reduced these effects. Co-immunoprecipitation demonstrated Fg and PrPC association. Since it is known that prion protein has a greater effect on memory reduction than amyloid beta, and that activation of TrkB is involved in neurodegeneration, our findings confirming the possible formation of Fg-PrPC and Fg-induced overexpression of TrkB on astrocytes suggest a possible triggering mechanism for STM reduction that was seen previously during mild-to-moderate TBI.

Fibrinogen Interaction with Astrocyte ICAM-1 and PrPC Results in the Generation of ROS and Neuronal Death

“…The data showed that Fg was co-localized and thus strongly associated with astrocyte ICAM-1 and PrP C , suggesting that it is possibly binding to these receptors on the surface of astrocytes. This is the first time a Fg specific interaction with astrocytes was visualized, confirming our previous finding that Fg could be associated with ICAM-1 [ 21 ] and PrP C with high specificity [ 16 , 36 , 39 ]. Thus, the increased respective PLA signals can be the result of a Fg interaction with over-expressed astrocyte ICAM-1 and PrP C .…”

“…Our observations of Fg-induced increases in astrocytic IL-6, CXCL-10 and CCL2 along with previously shown overexpression of tyrosine receptor kinase B [ 21 ] and PrP gene [ 16 ], suggest that Fg induces activation of astrocytes with functional polarization towards their neurotoxic, A1 phenotype [ 44 , 45 ], potentially exacerbating inflammatory conditions typically found during TBI [ 46 ].…”

“…We used this method to observe interactions between Fg and its receptors ICAM-1 and PrP C on the surface of astrocytes. In a preliminary study, the use of a higher content of Fg (2 or 4 mg/mL), similar to those used in our previous studies [ 16 , 21 ], resulted in signal saturation. Therefore, we used lower doses of Fg to detect if there was an association of Fg with its astrocyte receptors.…”

“…PrP C has been implicated in memory reduction [ 14 , 15 ]. While we have found that Fg can interact with astrocytic PrP C [ 16 ], no direct visual evidence of this association was provided. Therefore, we aimed to investigate if Fg can associate with ICAM-1 and PrP C on the surface of astrocytes using proximity ligation assay (PLA) and define if these interactions cause activation of astrocytic pro-inflammatory markers resulting in neuronal apoptosis and death.…”

The Effects of Fibrinogen’s Interactions with Its Neuronal Receptors, Intercellular Adhesion Molecule-1 and Cellular Prion Protein

“…Cellular prion protein (PrP C ) is a glycosylphosphatidylinositol-anchored glycoprotein abundantly expressed on the surfaces of neurons [ 13 ], while intercellular adhesion molecule-1 (ICAM-1) is a transmembrane glycoprotein that is constitutively expressed at low levels in unstimulated endothelial cells [ 14 ] and neurons [ 15 , 16 ]. Both PrP C [ 17 ] and ICAM-1 [ 18 , 19 ] are known Fg receptors. An increase in ICAM-1 expression was attributed to stimulation by inflammatory cytokines and oxidant species [ 20 , 21 ].…”