Attenuation of fragment inflammation response in macrophages by chondroitin Sulphate

Attenuation of hyaluronan fragment induced inflammatory response in macrophages by chondroitin Sulphate

Article specifications

This study was published in 2015 in journal of Osteoarthritis and cartilage (IF 2015:4.165) by Spanish molecular physiologists. In this study researchers determine whether chondroitin sulphate CS could similarly ameliorate an HA fragment mediated inflammatory response and whether any CS inflammatory inhibition was acting at the level of the inflammasome.  Cell culture: The THP-1 human monocytic cell line was grown in RPMI 1640 these cells were grown to a density of 1.5 x10‏6 cells/ml. After 2 days the media was replaced with media without the phorbol ester and grown for an additional 2 days. Cells were primed with a very small amount (10 ng/ml) of LPS for 24 hours with CS in various physiologically relevant concentrations (0, 10-200 mg/ml). After 24 hours, cells were washed with PBS and media replaced with serum free Opti-MEM along with the previously mentioned concentrations of CS and various molecular weights (sizes) (ULMW=7.5 kDA, LMW=29 kDa, MMW=289 kDA, and HMW=1.54 x10‏6 kDa) and concentrations (0, 1, 10, and 100 ug/ml) of HA fragments were added to the media. After a further 24 hours, cell viability was assessed and the media harvested. Following a PBS wash, water was added to the remaining cells and plates were subjected to 3 sonication/freeze/thaw cycles in order to release the cell contents. Caspase-1 experiments were carried out in 25cm2 flasks and activity was measured on fresh lysate derived from 2x10‏6 cells. Biochemical measurements included: Media and cell contents were analyzed for IL-1b and proIL-1b, Caspase-1 activity, Cell viability.

Results

The results of the study showed that HA fragments (<289 kDa) induced an inflammatory response in THP-1 macrophages which could be attenuated by CS. An anti-inflammatory effect at the level of the inflammasome would be expected to decrease intracellular caspase-1 activity thereby decreasing the ratio of IL-1b to proIL-1b.