Summary
The detrimental gelling effect to human tenocytes is noted when performing PRP experiment in small diameter culture well. This condition is avoided when using larger diameter culture wells.
Abstract
Background
Tenocytes of chronic rotator cuff tendon tears are not able to synthesize normal fibrocartilaginous extracellular matrix. Biological strategies are proposed to enhance tissue healing. Platelet-rich plasma (PRP) with different growth factors is believed to be helpful for tenocytes proliferation. Tenocytes from different age, gender of patients and passages has different characteristics. The best-fit PRP for each individual is unknown regarding the protocol of preparation, methods of activation, ratio between different growth factors and role of white blood cell. It needs large number of preparation conditions to screen the personalized best-fit PRP. Small diameter culture wells should be used to screen as much PRP preparation as possible at the same time with limited source of tenocytes from each patient. Results of tenocytes interaction with PRP in smaller culture wells may be different when compared with conventional studies using larger diameter culture wells.
Material
Human tenocytes were isolated from edge of torn human rotator cuff tendons when performing arthroscopic rotator cuff repair. First passage of tenocytes of each individual was used in the following experiment.
Method
Tenocytes were seeded in 4 different commonly used culture plates (96 well, 24 well, 12 well and 6 well) with same seeding density (2x104 cells/cm2). PRP was prepared and added inside each well with adjusted volume according to the diameter of each culture well. Cell proliferation was measured by WST-1 assay.
Results
Tenocytes proliferation was increased in 6 and 12 well culture plate when exposed to PRP. However, it was decreased when exposed to the same condition of PRP (well diameter adjusted) in 24 and 96 well culture plate. The culture medium in small culture wells became gel-like material after PRP was added, which may be responsible for decreased tenocytes proliferation.
Discussion
PRP can enhance tenocytes proliferation by the delivery of various growth factors and cytokines from the a-granules contained in platelets. However, the clinical benefit when applying it for augmentation during rotator cuff repair is still controversial. This may be explained by lacking of standardization of preparation protocol for each individual’s best-fit PRP. To test as much preparation conditions at one time, small diameter culture wells should be used to decrease total tenocytes needed at the same experiment because tenocytes over 3 passages will show phenotypic drift, which will hinder the result of PRP research. The gelling effect of PRP was noted when they were tested in small sized culture wells, which might be detrimental to tenocytes proliferation. The phenomenon disappeared when they were tested in large diameter culture well as published articles. Above condition should be noticed when performing PRP experiment in small diameter culture wells.
Conclusion
The detrimental gelling effect to human tenocytes is noted when performing PRP experiment in small diameter culture well. This condition is avoided when using larger diameter culture wells.