Journal: Acta Neuropathologica Communications

Article Title: Trichuris suis induces human non-classical patrolling monocytes via the mannose receptor and PKC: implications for multiple sclerosis

doi: 10.1186/s40478-015-0223-1

Figure Lengend Snippet: PKC signals downstream upon TsSP-MR interaction. Quantification of flow cytometry analysis of the expression levels of ( a ) CCR2, ( b ) LFA-1 avidity and ( c ) LFA-1 affinity on human monocytes. These cells were cultured in the presence or absence of TsSP (40 μg/ml, 16 h), the PKC inhibitor Bisindolylmaleimide I (GF109203, 2 μM) and blocking antibodies for the MR or its isotype control (both 10 μg/ml). ( b ) and ( c ) are expressed as the percentage of avidity/affinity compared to total LFA-1. Cell supernatants were used to determine the levels of ( d ) TNF-α and ( e ) IL-10 secretion by enzyme-linked immunosorbent assay analysis. f PKC activation was determined by Western Blotting using phosphoPKC antibodies compared to GAPDH expression and images were subsequently quantified using ImageJ. Experiments were performed in triplicate using cells derived from 4 different human donors and the results are presented as the mean ( a-c ); the mean percentage ( d, e ) or the mean PKC phosphorylation +/− SEM. * p < 0.05, ** p < 0.01, *** p < 0.001 as determined by ANOVA and Students t test

Article Snippet: The following primary antibodies (in TSM/0,05 % Tween/5 % BSA) were used: PhosphoPKC (Cell signaling, Beverly, MA, USA, #9371S) and GAPDH (Santa Cruz Biotechnology, Heidelberg, Germany, #sc-32233) as a loading control.

Techniques: Flow Cytometry, Expressing, Cell Culture, Blocking Assay, Control, Enzyme-linked Immunosorbent Assay, Activation Assay, Western Blot, Derivative Assay, Phospho-proteomics