Articular cartilage: Chemical, Physical, and Tribological Properties

A KSV CAM100 computerized tensiometer was used to measure the contact angle of cartilage samples. A drop of the 0.155M saline solution was deposited on the air-dry cartilage surface. The tests on the normal, partial and completely depleted cartilage samples were repeated five times.

The measurements were performed using a sliding pin-on-disc tribotester T-11 manufactured by the NISTR, Poland. The tests were conducted at room temperature, at a speed of 1 mm/s during 5 minutes, and under a load of 15 N (1.2 MPa) which corresponded to thephysiological lubrication condition.

Prior to the friction tests, the lubricants were prepared using the Britton-Robinson universal buffer solution and its pH values were measured. The samples were equilibrated with each buffer solution under a load for 5 minutes and the results of (f)as a function of pH are given in Figure 2. A total number of five tests were conducted using fresh samples for each experimental set-up with at least four repetitions per specimen pair, from which the mean and standard deviation were calculated.

Figure 2.Friction coefficient f vs. charge density for bovine cartilage (AC) obtained in the buffer solution). For isoelectric point IEP, molecule has equal charge distribution (H3N+ (CH2)n PO4- -R1R2). Standard deviation SD of (f ) 11 to 15 (%).

The wetted surfaces (pH~7.3) of the phospholipid membranes are negatively charged (–PO₄^-). Biosurface wettability can be measured relative to differences in the charge density of the functional phosphate (–PO₄^-) groups. In this regard, the wettability of a hydrated surface is characterized by the concentration of charged anionic phosphate (–PO₄^-) groups that become deactivated when the surface is dehydrated. The dehydration of the phospholipid bilayer surface activates hydrophobic groups, R (CH₂)_n- due to the formation of a hydrophobic monolayer ³. The hydrophilic surface in the course of dehydration, say via air-drying, undergoes a slow increase in the wettability contact angle indicating conformational changes in the surface (flip-flop) of phospholipidic molecules (Figure 1). Poor lubrication in animal joints, particularly on the articular surface of cartilage, can be attributed to deterioration of the bilayer surface, where the wettability or contact angle (θ) changes from 100° (healthy) to less than 70° (unhealthy) ^{4, 5}. The smart-surface of articular cartilage constitution of the superficial phospholipid bilayer in (a) aqueous electrolyte solution and (b) air-dry conditions. A change in surface energy leads to conformational changes in the surface of the bovine patella from bilayer (super hydrophilic ~0° contact angle) to monolayer (hydrophobic ~100°).

Figure 1.The wettability contact angle (o) as a function of air-drying time (a) in aqueous electrolyte and (b) air-dry conditions; (c) AC surface depleted in (chloroform/methanol (2:1,v/v). Curve (1) after 17 min, Curve (2) 7 min, Curve (3) 3 min; Curve (4) normal (untreated) AC surface; (n = 7, error bars = 95% confidence limit). (d) Book cover “Articular cartilage: Lamellar-repulsive lubrication of natural joints” 1.

Figure 2 presents a typical plot of afriction test between two normal surfaces (AC/AC), and these results indicate that the friction on cartilage surfaces is largely dependent upon the charge density of the tissue. It is evident that friction coefficient increases (curve part a), to reach a maximum at the isoelectric point, IEP, and then decreases gradually while sliding (curve part b) takes place over the pH range of the test. The cartilage surface carrying the positive charge, that changes to negative, can be attributed to the proton transfer reaction. Equation 1 depicts a cartilage surface carrying positive charges; after IEP, while negative charges can be attributed to the proton transfer reaction⁹.

-NH₃⁺ + OH^- → -NH₂ + H₂O; -PO₄H + OH^- → -PO₄^- + H₂O (1)

Curve part (a) friction increase, pH 2 → 4.5: -NH₃⁺ → -NH₂(surface losingcharge)

The maximum point, IEP, pH 4.5, no net electrical charge, H₂N(CH₂)_nPO₄H-R₁R_{2 ⇔}

H₃N⁺(CH₂)_n PO₄^- -R₁R₂ (2)

Curve part (b), friction decrease, pH 4.5 → 6.8: -PO₄H → -PO₄^- (gaining negative charge)

Curve part (c), constantfriction, pH ~ 7 to 9.0 (-PO₄^-, surface is negatively charged)

By varying the charge density, the friction coefficient can be varied by about one order of magnitude over the experimental range studied. Cartilage’s extremely low friction is significantly dependent on the electrostatic interaction between two cartilage surfaces ^{6, 7}. In an animal’s body, where the tissues slide over each other, the lubrication mechanism has been referred to as a “lamellar-repulsive”, because the surfaces coated with PL bilayers and lamellar phases negatively charged on articular surface with synovial fluid support this mechanism⁸.

The amphoteric PLs are the main solid-phase components on the surface of articular cartilage (AC). Cartilage phospholipidic membrane has been shown to undergo conformational reorientation when wettability changed from the wet (~ 0°) super hydrophilic to dry-air (103°) hydrophobic state (named smart material), Figure 1.

It has been well established that low friction is supported by the PLs bilayers mechanism which essentially consists of a surface amorphous layer (SAL) surrounded by a 0.155 M electrolyte synovial fluid (SF) of pH ~7.4 with high-molecular-weight charged biomacromolecules. The cartilage implication from osteoarthritis disease by a gradual erosion of the surface amorphous layer has shown anincreased friction coefficient¹⁰,¹¹.

The surfaces, coated with PL bilayers and a lamellar structure negatively charged on the articular surface with synovial fluid, have been referred to as a “lamellar-repulsive” mechanism. The role, played by hydration or structural force, is believed to arise from a strongly bound and oriented first layer of the water molecules on charged surfaces. The water molecule allows participating in strong polar (electrostatic charge - dipole or hydrogen - bonding) interactions. The short-range repulsion often observed between biological surfaces is not due to the layered structure of water but to entropic repulsion. The cartilage surfaces experience weak van der Waals attractive forces and much stronger short-range repulsive forces due to hydration repulsion. Hydration repulsion dominants the interaction between charged cartilage surfaces at nanometer separations and ultimately prevents the sticking together of cartilage surfaces, even at high load. A hydration water layer strongly binds to the negative charge cartilage surface, when in contact with synovial fluid components (charged biomacromolecules, PL lamellar aggregates, and liposomes), acts to reduce the friction between cartilage surfaces ^{1, 12}.

In this work, we demonstrated experimentally that the smart cartilage biomaterial at varies pH is sensitive to friction and introduces a novel concept in joint lubrication on charged surfaces. The cartilage surface was characterized using wettability test fresh and depleted AC samples.