Periodic Reporting for period 2 - THaCH (The effects of hypercholesterolemia on tendon health (Tendon Health and CHolesterol))
Período documentado: 2018-09-01 hasta 2019-08-31
Musculoskeletal disorders are a major cause of occupational disability. The link between high cholesterol and poor tendon function is acknowledged, but the cause of this relationship is still unknown.
Cholesterol can leave the circulation and accrue in tissues such as blood vessels (atherosclerosis) and tendons (xanthomas). We suspected that cholesterol accumulation in tendon tissue negatively affects tendon homeostasis and function and may increase tendon injury risk. To probe at these aspects of tendon health, we investigated avenues spanning cell biology to human movement. Specifically, our research aimed to understand how cholesterol 1) weakens tendons, and 2) affects tendon healing after injury.
We found tendon metabolism was modified with exposure to cholesterol, and in tendons which demonstrated lipid accumulation, tendon properties and behaviour were different. These findings are important for understanding the link between high cholesterol and tendon injury and may be important for several medical fields.
Cholesterol can leave the circulation and accrue in tissues such as blood vessels (atherosclerosis) and tendons (xanthomas). We suspected that cholesterol accumulation in tendon tissue negatively affects tendon homeostasis and function and may increase tendon injury risk. To probe at these aspects of tendon health, we investigated avenues spanning cell biology to human movement. Specifically, our research aimed to understand how cholesterol 1) weakens tendons, and 2) affects tendon healing after injury.
We found tendon metabolism was modified with exposure to cholesterol, and in tendons which demonstrated lipid accumulation, tendon properties and behaviour were different. These findings are important for understanding the link between high cholesterol and tendon injury and may be important for several medical fields.
Our use of inherited hypercholesterolemia models throughout the project have allowed us to conduct a series of experiments to investigate the link between hypercholesterolemia and tendon health. The results from this project demonstrate that tendon health is impacted physiologically and biomechanically by lipid presence.
Animal Studies (1-3)
We bred a cohort of control and genetically modified rats in-house to keep genetic variations minimal. At 12 weeks old, rats received an injury to the right patellar tendon. After 3, 14 or 42 days, animals were euthanized and the patellar, Achilles and tail tendons collected for various investigations. Blood samples were used to quantify cholesterol levels.
Human Studies (3-5)
Using a collaboration with an FH network, we have accessed patients from which we have conducted very relevant and clinically helpful studies. In using this cohort, we have also been able to minimize factors associated with secondary hypercholesterolemia, which may have confused our results or made recruitment difficult with exclusion criteria.
1. The patellar tendon was used to examine the effect of high cholesterol on tendon healing after injury. There was little histological evidence of lipid content in the tendons of either group and no obvious differences in healing speed between groups. No differences in tendon size, strength or collagen content were found between the injured tendons of the control and FH rats. However, there were differences in the gene expression of injured patellar tendon tissue between groups that suggest high cholesterol may modulate tendon inflammation and healing, even with a mild phenotype. This work was recently presented at the Experimental Biology biennial conference and is currently being written up for publication.
2. Achilles tendons were used to investigate regional lipid presence. We found histological evidence of lipid accumulation in the Achilles tendons of both groups that appeared more intense in the experimental vs. control rats. Moreover, lipid staining was almost exclusively found in the matrix between Achilles sub-tendons, rather than between tendon fibres. We quantified the biomechanical properties of the matrix using sub-tendon sliding, an important feature of AT function, and found higher strains in the experimental group. Higher strains = higher injury risk. This work is currently being written up for publication.
3. Rat tail and human semitendinosus tendons were used for extracting tendon cells. In both species we found that cells exposed to cholesterol were less able to move into an injury site and expressed different gene patterns in unstretched and stretched conditions; the differences were greater with increasing cholesterol concentration. These results link cholesterol exposure to inferior wound healing and greater inflammation and tissue turnover, which may lead to increased risk of tendon injury. The human work was recently presented at the American Society for Matrix Biology annual conference and has been submitted for publication; the rat work is a work in progress.
4. 16 FH patients and 16 control participants were recruited for studying the impact of tendon xanthomas on tendon function during walking. Participants walked on a treadmill while movement kinematics and Achilles tendon displacements were recorded. This data allowed us to visualize tendon function and compute certain tendon properties that relate to strength. We found tendon function and some tendon properties differed in FH, which may indicate an increased risk of injury in this population. This work was presented at international conferences and has now been published (doi: 10.1371/journal.pone.0257269).
5. We capitalized on our access to FH patients and investigated tendon xanthoma diagnosis, which are typically diagnosed by eye and feel, crude techniques that do not allow them to be distinguished from conditions with similar physical symptoms e.g. tendinopathy. 30 participants (10 FH, 10 tendinopathy, 10 controls) had their Achilles tendons scanned using non-invasive imaging techniques; scans were analysed for fat and water content, collagen organization and tendon thickness. FH individuals demonstrated a higher water content in combination with higher lipid content. This information could be used to modify existing criteria for clinicians to identify FH and differentiate from other conditions with similar symptoms. This work has now been published (doi: 10.1186/s12891-021-04494-0).
Animal Studies (1-3)
We bred a cohort of control and genetically modified rats in-house to keep genetic variations minimal. At 12 weeks old, rats received an injury to the right patellar tendon. After 3, 14 or 42 days, animals were euthanized and the patellar, Achilles and tail tendons collected for various investigations. Blood samples were used to quantify cholesterol levels.
Human Studies (3-5)
Using a collaboration with an FH network, we have accessed patients from which we have conducted very relevant and clinically helpful studies. In using this cohort, we have also been able to minimize factors associated with secondary hypercholesterolemia, which may have confused our results or made recruitment difficult with exclusion criteria.
1. The patellar tendon was used to examine the effect of high cholesterol on tendon healing after injury. There was little histological evidence of lipid content in the tendons of either group and no obvious differences in healing speed between groups. No differences in tendon size, strength or collagen content were found between the injured tendons of the control and FH rats. However, there were differences in the gene expression of injured patellar tendon tissue between groups that suggest high cholesterol may modulate tendon inflammation and healing, even with a mild phenotype. This work was recently presented at the Experimental Biology biennial conference and is currently being written up for publication.
2. Achilles tendons were used to investigate regional lipid presence. We found histological evidence of lipid accumulation in the Achilles tendons of both groups that appeared more intense in the experimental vs. control rats. Moreover, lipid staining was almost exclusively found in the matrix between Achilles sub-tendons, rather than between tendon fibres. We quantified the biomechanical properties of the matrix using sub-tendon sliding, an important feature of AT function, and found higher strains in the experimental group. Higher strains = higher injury risk. This work is currently being written up for publication.
3. Rat tail and human semitendinosus tendons were used for extracting tendon cells. In both species we found that cells exposed to cholesterol were less able to move into an injury site and expressed different gene patterns in unstretched and stretched conditions; the differences were greater with increasing cholesterol concentration. These results link cholesterol exposure to inferior wound healing and greater inflammation and tissue turnover, which may lead to increased risk of tendon injury. The human work was recently presented at the American Society for Matrix Biology annual conference and has been submitted for publication; the rat work is a work in progress.
4. 16 FH patients and 16 control participants were recruited for studying the impact of tendon xanthomas on tendon function during walking. Participants walked on a treadmill while movement kinematics and Achilles tendon displacements were recorded. This data allowed us to visualize tendon function and compute certain tendon properties that relate to strength. We found tendon function and some tendon properties differed in FH, which may indicate an increased risk of injury in this population. This work was presented at international conferences and has now been published (doi: 10.1371/journal.pone.0257269).
5. We capitalized on our access to FH patients and investigated tendon xanthoma diagnosis, which are typically diagnosed by eye and feel, crude techniques that do not allow them to be distinguished from conditions with similar physical symptoms e.g. tendinopathy. 30 participants (10 FH, 10 tendinopathy, 10 controls) had their Achilles tendons scanned using non-invasive imaging techniques; scans were analysed for fat and water content, collagen organization and tendon thickness. FH individuals demonstrated a higher water content in combination with higher lipid content. This information could be used to modify existing criteria for clinicians to identify FH and differentiate from other conditions with similar symptoms. This work has now been published (doi: 10.1186/s12891-021-04494-0).
Our results show that tendon health is impacted by cholesterol on multiple levels. We have been very fortunate to collaborate with leading scientists and use state-of-the-art imaging techniques and laboratory methods to further our understanding of tendon health and function in very controlled, high cholesterol conditions.
Using a cohort of FH patients for human studies, we have had a much more standardized population than if we had used individuals with secondary hypercholesterolemia (who typically exhibit factors which may confound results). Using FH rats in place of diet-induced hypercholesterolemia has the same advantages for the other objectives. Moreover, as all animals were bred in-house from two sets of parents, we were able to greatly reduce the genetic variability of the animals. The highly-controlled groups we have used in our work make us confident that our results reflect high cholesterol conditions rather than e.g. obesity, diabetes, which allows us to make more confident statements about our results.
We envisage that the information generated from this work will provide avenues for exploring the development of interventions aimed at the preventing or rehabilitating tendon injury. For example, targeting tendon lipid and water content with pharmaceutical and/or exercise interventions may reverse some of the characteristics that influence tendon injury risk.
Our results provide a rationale for investigating tendon function during higher impact exercise to create physical activity recommendations for managing FH safely.
Better knowledge of tendon metabolism in FH armed with better diagnostic options for FH would enable clinicians to use a protective rather than reactive approach to musculoskeletal injury in individuals with high cholesterol. Such strategies would have significant socioeconomic consequences and may change the extent of tendon disease or damage over a patient’s lifetime.
Using a cohort of FH patients for human studies, we have had a much more standardized population than if we had used individuals with secondary hypercholesterolemia (who typically exhibit factors which may confound results). Using FH rats in place of diet-induced hypercholesterolemia has the same advantages for the other objectives. Moreover, as all animals were bred in-house from two sets of parents, we were able to greatly reduce the genetic variability of the animals. The highly-controlled groups we have used in our work make us confident that our results reflect high cholesterol conditions rather than e.g. obesity, diabetes, which allows us to make more confident statements about our results.
We envisage that the information generated from this work will provide avenues for exploring the development of interventions aimed at the preventing or rehabilitating tendon injury. For example, targeting tendon lipid and water content with pharmaceutical and/or exercise interventions may reverse some of the characteristics that influence tendon injury risk.
Our results provide a rationale for investigating tendon function during higher impact exercise to create physical activity recommendations for managing FH safely.
Better knowledge of tendon metabolism in FH armed with better diagnostic options for FH would enable clinicians to use a protective rather than reactive approach to musculoskeletal injury in individuals with high cholesterol. Such strategies would have significant socioeconomic consequences and may change the extent of tendon disease or damage over a patient’s lifetime.