Hyaluronic acid is a naturally occurring linear polysaccharide. It is a polymer of disaccharide units composed of β-1,4-D-glucuronic acid and N-acetyl-D-glucosamine linked together by β-1,3-glycosidic bonds. Hyaluronic acid has a molecular weight ranging from 1,000 to 10,000,000 daltons.
Hyaluronic acid is the only non-sulfated glucosaminoglycan (GAG) found in the extracellular matrix (ECM) of higher animals.
Biomaterials derived from hyaluronic acid are used, for example, in regeneration of cartilage and skin. Also, they play key roles in drug delivery or surgery. Chemically unmodified hyaluronic acid is utilized as the aid for the delivery of ophthalmologic drug to improve the absorption of drugs and proteins at the mucosal tissue.
Further, with regard to the treatment of osteoarthritis, hyaluronic acid improves lubrication at the joint surface and thus reduces pain. Arthritis reduces the production of hyaluronic acid and the increased breakdown by proteases further reduces hyaluronic acid in the joint. As a result, the articular damage can be aggravated because external impact cannot be adequately absorbed or distributed at the joint.
Since approved by the FDA in 1997, the injection of hyaluronic acid is widely used. With compositions similar to those of the joint fluid, hyaluronic acid solution regains viscosity and elasticity when injected into the joint and improves lubrication and absorbs impact, thereby protecting the joint cartilage and preventing further damage.
Also, it is reported to have anti-inflammatory effect and inhibit the breakdown of chondrocytes by several in vitro researches. Based on this, it can be inferred that hyaluronic acid can be utilized as scaffold for cell delivery in the process of cell transplantation therapy of chondrocytes for cartilage tissue regeneration.
The in vivo metabolism of hyaluronic acid is regulated by hyaluronic acid synthase (HA synthase) and hyaluronidase. Hyaluronic acid synthase is an integral membrane enzyme, which synthesizes hyaluronic acid polymers and excretes them out of the cell. In mammals, HAS1, HAS2 and HAS3 isoenzymes are found.
Hyaluronidase is an enzyme that breaks down hyaluronic acid. Depending on the activation pH, it can be classified into neutral hyaluronidase and acidic hyaluronidase. Neutral hyaluronidase (PH-20) is specifically found in the testis (sperm) and shows activity in the physiological pH.
Acidic hyaluronidase (Hyal1-4) is found in body fluids as well as in a variety of human organs, including spleen, cartilage, skin, eyes, liver, kidney, bladder, placenta, etc., and shows activity around at pH 3. Since this enzyme is present in the lysosome, hyaluronic acid is transported into the cytoplasm by endocytosis and degraded.
The endocytosis of hyaluronic acid is mediated by the cell surface receptors. The primary cell surface receptor for HA is CD44. CD44 is a transmembrane glycoprotein and binds primarily to hyaluronic acid. It is expressed in most of human cell membranes. The hyaluronic acid fragments degraded at the lysosome induce different signal transfers depending on their size, affecting cell proliferation and differentiation.
Various porous scaffolds are used in the field of tissue engineering. Chondrocytes cultured in 2D condition such as petridish cannot maintain their chondrocytic phenotype, resulting in synthesis of type I collagen rather than type II.
In contrast, chondrocytes cultured in 3D condition maintain their characters and functions, resulting in spherical cell shape. That is, a 3-dimensional scaffold helps the cell attachment and cartilage formation in vitro and in vivo.
The use of porous scaffolds is very useful in that chondrocytes can be cultured under a restricted condition with significantly reduced necrosis of the tissue.
Thus, as one of new treatments for joint cartilage damage, the technique of culturing stem or progenitor cells derived from the cartilage, bone marrow or periosteum in vitro using biodegradable scaffolds and transplanting them into lesion site is attempted instead of direct injection of autologous chondrocytes.
This technique is advantageous over the method of using cell suspension since necrosis of the cartilage tissue to serve as cell sources is significantly reduced and the cells can be incubated under a restricted condition. Further, it is also adequate considering that cell attachement is improved by a scaffold.
In addition to cell delivery, a scaffold can serve as mold to fill lesion site of tissue. An ideal scaffold is required to be non-immunogenic, non-toxic, biocompatible, biodegradable and easy to manipulate.
The pore size of the scaffold is important. An inadequate pore size may result in the restricted nutrient supply to the cells being cultured in the scaffold or ineffective removal of the wastes from the cells.
If the pore size is too small, the cells may not be completely filled inside the scaffold, which negatively affects the generation of the tissue. In contrast, if the pore size is too large, not all the extracellular matrix proteins synthesized by cells are accumulated. That is, the proteoglycan and collagen accumulated in the scaffold can be found only a part of the total degree of actually synthesized ECM.
Accordingly, even though chondrocytes cultured in a porous scaffold in a metabolically active state, the scaffold may deem as inadequate for the growth of chondrocytes.
Thus, the control of the pore size is important in improving the retention rate of newly synthesized extracellular matrix molecules inside the scaffold.
Also, in case the sponge type scaffold with minimal porosity required for survival, growth and differentiation of cells is in solid state, it comes into problem that the surgical operation is the only way of delivering the entire scaffold including cells to a disease site.