Gene expression profiling of blood cells from rats treated with different fractions of a standarized aequeous willow bark extract – evidence for immuno- and neuromodulatory activities

Willow bark extracts (WB) are known for their anti-inflammatory activities. Their efficacy is primarily attributed to the content of salicin and its derivates. However, WB have a substantial content of polyphenols known to possess also antioxidant, neuroprotective and regulatory effects on signalling pathways. We used the gene microarray technique (Agilent whole Genome Rat array) to analyse the expression of the complete rat genome (˜ 41 000 genes) which may be modulated by WB or its different fractions. The WB STW 33-I was sequentially separated into five fractions of different polarity, using toluene, ethyl acetate, n-butanol and ethanol in addition to the aqueous extract. 84 rats were treated with these fractions (30mg/kg). Blood samples (3ml) of treated and untreated rats (n=12) were collected in PAX-gene collection tubes. RNA was isolated and the gene modulation was determined in two to three animals per group.

After filtering the data to remove genes showing no differences between the differentially treated samples, an ANOVA analysis was performed. The resulting gene set was clustered both hierarchically and by applying SOTA. The analysis revealed groups with a consistent gene expression according to the treatment. 1143 genes were identified as differentially regulated. They included genes for AMP-activated protein kinases, hyaluronoglucosidase 6, chondroitin sulfate N-acetylgalactosamyltransferase 2, H2-Ea (histocompatibility class II antigen) or Gria2, a glutamate receptor, activated in a variety of normal neurophysiologic processes. WB appears to be more than an anti-inflammatory agent. Microarray expression profiling will support us in understanding the mode of action of phytopreparations.