Skip to main content
Figure 4 | Journal of Neuroinflammation

Figure 4

From: Heat shock protein 60: an endogenous inducer of dopaminergic cell death in Parkinson disease

Figure 4

Hsp60 and inflammatory processes mediated by microglial cells. (A) Left: microglial cells exposed to Alexa647-conjugated hHsp60 (red label) or Alexa647-conjugated BSA (used as negative control) with 4’6-diamidino-2-phenylindole (DAPI) (blue label) counterstaining of cell nuclei. Scale bar = 50 μm. Right: confocal laser scanning microscopy of microglial cells that were exposed to Alexa647-conjugated Hsp60 (red label), then stained with an α-tubulin antibody (green label) and counterstained with DAPI (blue label). Scale bar = 10 μm. (B) Measure of cell surface binding of hHsp60 using extracellular FACS analysis of microglial cells labeled with an anti-Hsp60 monoclonal antibody (SPA-810, StressGen, San Diego, CA, USA) and a phycoerythrin-labeled secondary antibody. (C) Microglial cells were treated with hHsp60 at a concentration of 10 μg/ml, after 1, 2, 4, 6 and 24 hours respectively. Lipopolysaccharide (LPS) treatment for 6 hours for cytokine release respectively 24 hours for nitric oxide (NO)-release in a concentration of 0.1 μg/ml was used as positive control. (D) In addition, microglial cells were exposed to different hHsp60 preparations (native hHsp60, hHsp60 denatured by trypsinization or boiling and hHsp60 pre-incubated with Hsp60-specific antibody or isotype control antibody), after 12 and 24 hours, respectively. lipopolysaccharide (LPS) treatment for 12 hours in a concentration of 0.1 μg/ml was used as positive control, PBS as buffer control.

Back to article page