Effect of heat on synthesis of gelatinases and pro-inflammatory cytokines in equine tendinocytes.

This study investigates how heat exposure influences the production of certain enzymes and inflammatory markers (gelatinases and cytokines) in horse tendon cells. The results found that such exposure prompts these cells to produce more inflammatory markers, which subsequently increases the production of gelatinases.

Research Objective and Hypothesis

• The researchers aimed to ascertain whether heat has an effect on the production of matrix metalloproteinases (MMP-2 and -9, also called gelatinases) and pro-inflammatory cytokines (TNF alpha and IL-1beta) in tendon tissue in laboratory conditions.
• Their hypothesis was that exposure to heat stimulates these tendinocytes to synthesize pro-inflammatory cytokines, which then leads to an increased production of gelatinases.

Methodology

• Tendinocytes isolated from horse superficial digital flexor tendons were cultured for the study, with all tests performed on cells that had been passaged 3 or 4 times.
• The cells were exposed to different degrees of heat (37 to 45℃) for varying amounts of time (0 to 60 minutes).

Results

• The survival rate of cells decreased significantly with a rise in temperature and duration of exposure, especially at 42 and 45 degrees C. However, exposure at 40 degrees C showed little impact on cell survival and morphological changes.
• ProMMP-2 and -9 were found to be the only two MMPs remaining in the cultured tendinocytes’ supernatant, including that of unexposed cells.
• The addition of TNFalpha and IL-1beta in the culture medium of tendinocytes drastically increased the production of proMMP-9.
• Heating the tendinocytes at 40 degrees C resulted in a three-fold rise in synthesis of proMMP-9 within a short duration.
• Post-heat exposure (at 30 and 60 minutes), only TNFalpha was detected in the tendinocytes while IL-1beta remained below detectable levels.
• Upon cooling the heated cells from 40 degrees C to 37 degrees C, the synthesis of cellular proMMP-9 notably decreased. Furthermore, the levels of proMMP-9 were significantly reduced in cells exposed to lower temperatures of 20 degrees C and 5 degrees C.

Conclusion

• The findings confirmed the researchers’ hypothesis: that increased heat in horse tendons stimulates tendinocytes to produce pro-inflammatory cytokines, which consequently enhances the synthesis of gelatinases.