Measurement of Intercellular Cohesion by Tissue Surface Tensiometry
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Intercellular adhesion plays a vital role in many biological processes including embryonic development, malignant invasion, and wound healing, and can be manipulated to generate complex structures in tissue engineering applications. Accurate measurement of the strength of intercellular adhesion is not trivial and requires methods rooted in sound physical principles. Tissue surface tensiometry (TST) rigorously quantifies intercellular cohesive energy of 3D tissue-like aggregates under physiological conditions. TST utilizes a custom-built tensiometer to compress 3D spheroids between parallel plates. The resistance to the applied force and changes in aggregate geometry are applied to the Young-Laplace equation, generating a measurement of apparent surface tension. We describe all components comprising the tensiometer and provide step by step instructions of all the key steps involved in generating spherical aggregates. We explain how tissue surface tension is calculated and provide a statistical analysis of a sample data set from 12 aggregates.
Key wordsCell adhesion Tissue liquidity Surface tension Tissue surface tensiometry Parallel plate compression
This work was supported by NIH grant CA118755 to RAF. The author also acknowledges the seminal contribution of Malcolm S. Steinberg, Ph.D., to the development of this method.
Aggregate rounding up: Here, an “irregular” fragment of embryonic chick liver was extirpated from a 3.5-day-old embryo, and placed onto the lower compression plate (LCP). The LCP had been pre-coated with poly-HEMA, preventing adhesion of the fragment to the plate. The tissue fragment was incubated overnight and filmed in real time. The movie is run at 8× speed. Note the rounding-up behavior and the significant movement of the aggregate on the plate, indicating little adhesion to the substrate (MOV 44708 kb).
Real-time chart recorder tracing as an aggregate is undergoing compression and force relaxation. The pen deflects from the zero force position at chart recorder position 10 to approximately position 52, representing an applied weight of 4.2 mg. The aggregate undergoes a fast relaxation phase in the first 45 s after compression, whereupon the force tracing begins to level off (MOV 77785 kb).
Real-time chart recorder tracing as an aggregate is decompressed after having reached force and shape equilibrium. This aggregate was under compression for approximately 2 h and 20 min. By this time, the force tracing had leveled off at the 1.9 mg position, indicating that the aggregate had reached equilibrium. Upon decompression, the chart recorder pen deflected back towards the zero force balance position. Accordingly, the F value recorded for application to the Young-Laplace equation was 1.9 mg (MOV 48674 kb).
The elastic response. An embryonic chick aggregate composed of myocardium when compressed and immediately decompressed assumes its original shape (MOV 29802 kb).
The viscous liquid response. An aggregate of embryonic chick myocardium if compressed and allowed to reach shape and force equilibrium does not assume its original spherical shape upon decompression. Note that rounding up takes place in a more gradual manner as depicted in time lapse (MOV 245564 kb).
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