Hydrophilic and negatively charged natural cartilage surface is covered by phospholipids bilayers. These phospholipids have demonstrated efficient lubrication, load processing, and semipermeability for nutrient transport. We examined a bovine cartilage (BC) surface using atomic force microscope. The study was performed using cartilage samples with healthy surfaces and completely depleted surface phospholipids. Our results demonstrated that it is possible to recreate a potentially viable layer of phospholipids on the surface of degenerated cartilage. However, further studies will be required to advance the resurfacing idea developed in this paper for the potential treatment of osteoarthritis and other related orthopedic joint conditions.

Objectives: We attempt to answer degradation of phospholipid bilayers on cartilage surface by antibodies’ β2-glycoprotein I (β2-GP I) (-NH3+) by binding to negatively charged phospholipid (–PO4-) as a main cause of phospholipid syndrome.

Methods:The articular cartilage samples used in this study were obtained from the patellae of 3-4 year old bovine animals.  We examined a bovine cartilage (BC) surface using atomic force microscope (AFM).

Results:Surface cartilage phospholipids have demonstrated efficient lubrication, load processing, and semipermeability for nutrient transport. It was shown that the phospholipid syndrome after an appearance in the joints leads to the devastation of phospholipid bilayers and inactivation of their molecules.

Conclusion: Normal cartilage surface and cartilage surface damaged by osteoarthritis (OA) and rheumatoid arthritis (RA) which is considered antiphospholipid syndrome.  Deactivated PL molecule has no ability to form bilayers and liposomes or be adsorbed by lubricin and hylurunon molecules.

Articular cartilage (AC) is one of the tissues which, in contact with two surfaces and a changing load in the reciprocating movement, participate in the process of friction and lubrication.  The mechanism of boundary joint lubrication with phospholipids (PLs) has been described as a lamellar-electrostatic mechanism involving negatively charged phospholipid bilayers [1]. Low friction is supported by electrostatic repulsion of negatively charged phospholipid bilayers. Types of phospholipid adsorbed onto the surface of cartilage were identified: phosphatidylcholine (41%), phosphatidylethanolamine (27%) and sphingomyelin (32%) were the major components of the lipid bilayers on the normal cartilage surface [2].

In a healthy joint with a normal biological concentration of PLs in SF,a low friction coefficient might be expected. The phospholipids content in SF during joint inflammation, osteoarthritis (OA) and rheumatoid arthritis (RA) is significantly higher (2 to 3 times) [3] above the normal concentration of phospholipids, and has a poor boundary-lubricating ability as compared with that of a healthy cartilage. However, this can be related to the degradation of the phospholipid bilayers system. Additionally, phospholipid molecules lose their surface active properties to form vesicles, lamellar phases, and bilayers spontaneously. The active role played by PLs in OA and RA SF as compared with that of control SF and their functions in cartilage boundary lubrication remains still poorly understood [4, 5].  In this paper, we attempt to answer degradation of phospholipid bilayers on cartilage surface by antibodies’ β2-glycoprotein I (β2-GP I) (-NH3+) by binding to negatively charged phospholipid (–PO4-) [1] as a main cause of phospholipid syndrome.  

The articular cartilage samples used in this study were obtained from the patellae of 3-4 year old bovine animals harvested from the local abattoir and stored at -20oC until required for testing. The samples were thawed out in continuous running water at room temperature and kept in saline solution (0.15M sodium chloride) prior to testing. A stainless-steel punch was used to cut osteochondral plugs (n = 20), containing full thickness articular cartilage-bone laminate and trimmed into specimen of 5 mm by 5 mm.  The bony layer underlying the cartilage was dabbed with a paper towel and immediately glued onto a Petri dish using fast-drying Loctite® 454 glue. The Petri dish was mounted onto the AFM sample holder, ready for AFM measurements. During gluing, the articular surface was moistened repeatedly with drops of saline solution to keep surface intact. The glued sample was submerged in saline solution ready for AFM imaging using the SMENA® head of the NT-MDT P47 Solver scanning probe microscope (SPM) (NT-MDT).  The surface imaging was done using methods described elsewhere in the literature [6]. In order to simulate the loss of cartilage surface lipids, an artificial lipid extraction process was used (delipidization).

Figure

Cartilage destruction in most rheumatic diseases and osteoarthritis has generally been accepted as a mechanism of deactivation of bilayers Fig. 1 [3].   At a pH around 7 amino acids from (β2-GPI) (arginine, lysine and tryptophan) have hydrogen donor atoms in their side chains (-NH3+) and acid–base interaction occurs between protonated amino acid group (-NH3+) and the phospholipid (–PO4-) group: (-NH3+) + (–PO4-) → (-NH3+-PO4-) that is strong enough to deactivate the PLs bilayer surface.  β2-Glycoprotein I (β2-GPI) is a protein that circulates in blood at variable levels (50–500 μg mL−1) with a molecular weight of 50 kDa. β2-Glycoprotein I (β2-GP I) can exist in (a) closed conformation and (b) the open hockey stick-like conformation β2-GP I in its hockey stick-like conformation is a strongly adhesive protein, and binds to different receptors on cells. Binding of β2-GP I to anionic charged phospholipid (–PO4-) groups at pH ~ 7.4, results in a change in conformation and exposure of the epitope for the autoantibodies.

 Continued deterioration of articular cartilage leads to an exposure of the subchondral bone and a more generalized synovial change, Fig.2.  To understand the processes leading to cartilage failure, it is important to look at the cellular processes and biochemical structure of the normal cartilage.

Figure 3 displays 3D topographical image of osteoarthritis (OA) and rheumatoid arthritis (RA): (a) normal healthy cartilage after image processing, showing the nanostructural arrangement of the surface amorphous layer with several peaks and troughs, (b) delipidized cartilage after image processing, showing the loss of the membranous overlay (surface amorphous layer) of the articular surface [4]. 

It has been shown that a lamellar-electrostatic mechanism involving phospholipid bilayers facilitates almost frictionless lubrication of the joint. Cartilage wear and joint degeneration begin from the destruction of phospholipid bilayers on the cartilage surface with the participation of the phospholipid syndrome.  Joint degeneration leads to the destruction of bilayers on the cartilage surface and immobilization of the joint. Synovial fluid with its composition of macromolecules is insufficient to withstand the load and control friction in the joints. It was shown that the phospholipid syndrome after an appearance in the joints leads to the devastation of phospholipid bilayers and inactivation of their molecules. Moreover, it has been experimentally shown that the joint cartilage belongs to intelligent materials and is exposed to destruction, which is graphically expressed in Fig. 1 to 3.

Fig. 4.  Tribochemical (A) Tree of lamellar-repulsive mechanism of joints lubrication

(B) phospholipid bilayer of cartilage in wet (hydrophilic ~0o wettability) and air-dry condition (hydrophobic ~104o wettability) picture from book cover [1]. 

 A tree of ‘lamellar-repulsive mechanism’ of joints lubrication is shown in Fig.4 and is based on lamellar surface amorphous multilayer (SAL) formed on articular hydrophilic surface. SAL membrane is providing boundary lamellar-repulsive hydration lubrication.  The lamellar-repulsive mechanisms is supported by phospholipid lamellar phases and charged macromolecules from synovial fluid between charged cartilage surfaces.