Inflammatory biomarkers with focus on Calprotectin (S100A8/S100A9) and S100A12 (EN-RAGE) : method development and application in acute radiation proctitis and rheumatoid arthritis patients
Abstract
Assessments of diseases are usually performed by a combination of clinical
symptoms, signs and objective measurements. Biochemical markers of inflammation
are important in evaluation of treatment of acute and chronic diseases. In this thesis
we have focused on two inflammatory diseases: acute radiation proctitis and
rheumatoid arthritis. We examined established and new biomarkers, with focus on
two closely related proteins, calprotectin (S100A8/S100A9) and S100A12 (ENRAGE).
These and other biomarkers were compared to clinical symptoms and signs
with the aim of defining their usefulness in monitoring the course of the diseases.
Characterizing the changes induced by radiation therapy to the pelvic region showed
incongruent time profiles in symptoms, histological scoring of inflammation and
biochemical measurements. Compared to the examination before start of radiation
therapy, biopsies from rectal mucosa showed a maximal inflammation at the first
examination during radiation therapy, while symptoms were at the most intense
towards the end of treatment. Vitamin E concentrations in blood were significantly
lower during treatment but returned to normal within a month. Calprotectin and
lactoferrin concentrations in stool samples increased during therapy and were the
most promising biomarkers for radiation proctitis in our study.
Development of an ELISA immunoassay for the quantification of S100A12 in blood
showed that calcium concentration influenced the estimated S100A12 concentrations.
Various experiments led to the conclusions that the antibodies used in this assay
could only recognize S100A12 when calcium were above a threshold and that
S100A12 were found in many molecular sized complexes (oligomers) in blood.
S100A12 was found to bind to heparin and this will be further examined in the future.
Calprotectin and CRP in blood correlated with disease activity of rheumatoid
arthritis. S100A12 was associated with a more severe disease course in terms of
extra-articular disease manifestations and the presence of cardiovascular disease. S100A12 correlated with rheumatoid factor and anti-CCP and may thus be of
prognostic value.
In summary, radiation proctitis occurs in biopsies before symptoms are evident. This
is important knowledge for studies with interventions to prevent development of late
radiation enteropathy. Stool markers are at present the best suited objective means of
assessing interventions. The quantification of S100A12 in blood samples are
recommended to be done in serum samples, since both anticoagulants EDTA and
heparin may influence conformational changes of S100A12 protein structure and
hence the measured concentrations. The usefulness of S100A12 as a biomarker in
rheumatoid arthritis and other inflammatory diseases warrants further examinations.