Temperature-driven premature ageing of cellular populations in energy-storing tendons: the gap junction connection.

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Athletes who participate in running and jumping have a similar problem to their equine counterparts on the racetrack. Both frequently injure large tendons in their legs that function as 'biological springs' to save muscular effort by stretching and storing energy when the leg bears weight, then rebounding to propel the athlete forward. In people the Achilles tendon (AT) has this function, while in horses it is the superficial digital flexor tendon (SDFT). As more people participate in sports to improve their health and maintain mobility during ageing, more AT injuries are occurring. In Scotland the incidence has increased by 90% over the last 15 years, now comprising >10% of soft tissue injuries. Up to 30% of racehorses suffer similar problems. Tendons heal slowly with scar tissue and never regain their original strength. Such injuries interrupted the international careers of David Beckham and Kelly Holmes, and those of many well-known racehorses including Kicking King, the 2005 Cheltenham Cup winner. Tendon injuries follow an undefined period of accumulation of painless damage to their substance (matrix) during exercise. Tendon cells, called tenocytes, do not repair this 'microdamage' and may be killed, begin to produce the wrong type of collagen, and/or degrade the surrounding matrix, causing a vicious injury cycle. Research over many years has suggested that this degeneration represents acceleration of a normally age-related process. Slowing or preventing this would have a greater impact on preventing injuries, and so improving performance and welfare of athletes than attempting treatment. We think a major factor causing tenocyte injury is high temperature. As tendons stretch and contract, some stored energy is lost as heat that cannot be easily dissipated. The SDFT core reaches at least 45oC during galloping, from its normal temperature of 37-38oC. As the AT functions in a similar way, hyperthermia is also highly likely to occur in its core. In the laboratory, most tenocytes die after 10 min at 45oC, but interestingly this involves transmission of 'death signals' between cells through gap junctions (GJ). GJ are small pores directly connecting neighbouring cells into networks. When GJ are chemically blocked following heating much of the tenocyte death and upregulation of less effective matrix proteins can be prevented. This is exciting, as GJ can be manipulated. We will use cells and tissue from SDFTs to determine when, relative to heating, the tenocytes 'decide' to die or alter their activity. We will measure an array of biomarkers to identify points at which we might reverse these changes e.g. do we need to do something within minutes or hours? Secondly we will explore a drug-free method of intervention: cooling of tendons after exercise e.g. using ice baths is often used as an 'injury prevention' measure. Cooling can close GJ in other cell types, which could prevent spread of death signals during critical post-exercise periods. Mild hypothermia (i.e. 32oC) may also upregulate 'cold-stress proteins' that protect cells. We aim to understand when, for how long, at what levels and how many times temperature differentials should be applied by athletes to reduce tendon microdamage. In the final part of this project we will look at one of the major proteins making up the GJ, called connexin 43 (Cx43). We will investigate methods of targeting the Cx43 molecule i.e. preventing it from being made within tenocytes. This will allow us to better understand how GJ influence cell death/damage following heat shock, but may also facilitate development of drugs to regulate it that could be delivered locally e.g. in gels or ointments. With the upcoming London Olympic Games and Glasgow Commonwealth Games and the likelihood that increasing numbers of people will be inspired to participate in athletic activity, this is a timely opportunity to develop methods of preventing injuries while also improving the welfare of horses.