In vitro rejuvenation of osteoarthritis-affected human chondrocytes cultured in a polymer based 3D matrix, evidenced by enhanced miRNA expression

Osteoarthritis is a debilitating, chronic disease that affects joints through progressive inflammation and degradation of the articular cartilage, inducing pain, swelling and immobility. This condition is associated with risk factors such as age, obesity and diabetes that are widespread in our society. Thus, the World Health Organization estimates that the incidence of osteoarthritis is about 10% and 18.0% for men and women over sixty, respectively. The knee and hip joints are the most affected areas, but others, such as those in the hands, can also be impaired with osteoarthritis. Advanced cell treatments including matrix-associated chondrocyte implantation (MACI) and autologous chondrocyte implantation (ACI) have been shown to be effective in treating osteoarthritis-related articular cartilage abnormalities. Some frequently mentioned factors that affects the clinical results in cell-based cartilage repair therapies are cell seeding density during in vitro culture, cell count and the preservation of the hyaline phenotype both after transplantation and during in vitro culture. MicroRNAs are single-stranded noncoding RNA molecules that regulate gene expression by binding to specific sequences in messenger RNAs. It leads to posttranscriptional degradation or repression of the target mRNAs. MiRNA-140 believed to be the most important microRNA affecting cartilage growth in the context of osteoarthritis and cartilage decreased expression of miRNA-140. It has been linked to the altered gene expression associated with disease progression and osteoarthritis. miRNA- 140-5p is a cartilage-specific microRNA and thought to have anti-inflammatory effects.

Studies have claimed that miRNA-140-5p is required for in vitro chondrogenesis and to be significantly suppressed in osteoarthritic cartilage. miRNA-140-3p is its passenger strand that is highly expressed microRNA in healthful cartilage. It exhibits increased expression during in vitro chondrogenesis. Moreover, intra-articular injection of miRNA-140 has been found to reduce osteoarthritis progression by altering extracellular matrix homeostasis and is considered a possible therapeutic strategy for osteoarthritis.

Tissue engineering in thermoreversible gelation polymer (TGP), a 3D Platform enhances in-vitro development of rabbit, bovine, and human chondrocytes and retains the hyaline phenotype for an extended period of time and supports the proliferation of chondrocyte progenitor cells that express pluripotency-related surface glycans.  Moreover, the 3D TGP scaffold has also been shown to have in vivo transplantation potential. The added benefits of the 3D TGP platform in boosting miRNA-140 expression during the in vitro expansion of osteoarthritis -affected human chondrocytes was recently conducted by Shojiro Katoh, Hiroshi Yoshioka, Rajappa Senthilkumar, Senthilkumar Preethy, and led by Dr. Samuel J.K. Abraham in a newly published research paper in Life Sciences.

MicroRNA-based therapies are a new treatment option for osteoarthritis. In an in vitro model of osteoarthritis, forced expression of microRNA-140 increased cartilage healing and decreased cartilage degradation. In another inflammatory model of osteoarthritis, transfection of miRNA-140 significantly suppressed critical inflammatory mediators and helped restore other homeostatic cellular systems, like metabolism. In rats, intra-articular injection of miRNA-140 reduced osteoarthritis progression via altering extracellular matrix homeostasis. Additionally, it has been demonstrated that loss of miRNA-140 contributes to the development of age-related osteoarthritis-like changes. In terms of miRNA-21, its expression is reduced in osteoarthritis cartilage and it stimulates the development of hyaline cartilage.

In vitro and on vivo, human articular chondrocytes were cultured in the 3D-TGP platform and was found to assist maintain the hyaline phenotype required for tissue healing in osteoarthritis. The authors found that chondrocytes generated from osteoarthritis patients preserved their hyaline phenotype as well as the production of pluripotent chondrocyte progenitors. MiRNA-140-3p expression was found to be lower in the supernatant of 3D-TGP-cultured cells than in that of 2D-cultured cells. They investigated miRNA 140-3p in the Phase I-supernatant because it was reported to be the most significant miRNA linked with cartilage homeostasis. In Phase II, they evaluated miRNA 140-3p and 5p, as well as miRNA-21-3p and 5p, in the cell lysate. The 3D-TGP culture had higher expression levels of miRNA 140-3p and 5p, as well as miRNA-21-3p and 5p, in cell lysate than the 2D culture. This data suggest that the miRNA was maintained in the 3D-TGP-cultured cells.

The exact mechanism of how the 3D-TGP environment encourages the enhanced expression of miRNA 21 and miRNA 140 still unknown. When this 3D-TGP platform was previously used for the culture of various types of cells, such as pluripotent stem cells, it was found to provide the necessary extracellular cues that are similar to native in vivo environments resulting in cell maintenance and positive gene expression associated with healthy cells. An equivalent culture methodology in 2D monolayer cultures resulted in unwanted hydrodynamic stresses from the culture vessel, possibly resulting in cell death. Although a great deal of effort is still required to make such discoveries a routine clinical practice, the promise is that improved cell-based therapies will ultimately lead to the repair of damaged or diseased joints, osteoarthritis could be a safer, cheaper and a more effective treatment modality. In a statement to Medicine Innovates Series, the authors quoted: “With an ability to rejuvenate human osteoarthritis affected cartilage derived cells by enhancing their miRNA-140 in the lab, our 3D TGP platform, is ideal for culturing chondrocytes for regenerative medicine applications like ACI and MACI that address cartilage damage, though cell based therapies”.