Flexor Tendons Case Study

2.1.1.a Flexor Tendon and Rotator Cuff

The ability to flex the finger consists of a serial of flexor muscles in the forearm and their tendons are inserted to the bones of finger. The injury of flexor tendon might cause the loss of bending of the fingers or thumb. The flexor digitorum profundus tendon (FDP) attaching to the distal phalanx and the flexor digitorum superficialis tendon banding to middle phalanx well demonstrated the specific type of tendon-to-bone insertion site characterized by the four-zone enthesis.[1] The retinacula (sheath) structures serve as strong fibrous bands wrap around the flexor tendons in order to keep the flexor tendons in place while flexion.

Injuries to flexor tendon remain the most difficult problem among

all the hand surgeries.[14] Currently, the surgeons would approximate the ends of the cut tendon by using special designated stiches. But the surgery might result in hand/finger malfunction with restricted motion based on the fact that the insertion site is not regenerated.
The rotator cuff is a group of tendons and muscles in the shoulder, connecting the upper arm (humorous) to the shoulder blade (scapula).[15] The rotator cuff tendons stabilize the shoulder, the muscle allow it to move.[15] Rotator cuff tears of the shoulder are a common of pain, and it may result in loss of tendon-to-bone enthesis, which leads to malfunction, instability of the rotator cuff. Approximately, 30% of the elder people over 60 year of age are suffering rotator cuff tear pain.[16-17] Although the surgery for repair of rotator cuff tear is also a common one for orthopedic surgeons, the patients are experiencing less expected results since the quality of the resulting tissue differs greatly from the normal.[18]
2.1.1.b Surgical and Mechanical Challenge in Healing of Tendon-to-Bone Enthesis
The tendon-to-bone enthesis possesses a graded connective tissue transition includes unmineralized tissue (i.e., tendon) followed by uncalcified cartilage, calcified cartilage, and mineralized tissue (i.e., bone).

No obvious boundaries are observed between the two distinct materials, tendon and bone. Type I collagen and tenocytes are highly aligned in tendon. In uncalcified fibrocartilage, where collagen type II is of great content, along with rich type III collagen and small amount of type X collagen, decorin, and aggrecan. Similarly, with a great amount of type II collagen, the mineralized cartilage presents significant amounts of collagen type X and scarce levels of aggrecan. Note that the collagen fibers are highly aligned in the direction of tensile force in tendon but less oriented in the insertion site (Figure 2).[4, 19] Additionally, the insertion site possesses a transitional decrease in tissue organization while an increase in mineral content.[4] The complex collagen and mineralization content in this region lead the repair and rehabilitation of tendon-to-bone insertion site more

difficult.
Known as an unregenerated tissue, the functional graded tendon-to-bone insertion site has been a challenge to orthopedic surgeons in repair of tendon tear surgeries, i.e., the repair of flexor tendon injuries.[14] This includes the achievement of mechanical stabilization of tendon-to-bone during healing, the accomplishment of a natural gradation structure from soft to hard tissue while healing, and the realization of a developed transitional area with maintained population of graded cell phenotypes (tenocytes, chondrocytes, and bone related cells) along with graded mechanical structure.[20]
Figure 2. A schematic of the cross-section of a rat supraspinatus tendon-to-bone enthesis. Blue shading indicates the concentration of mineralization in the gradation of tissue. As is shown in the enlargements, the collagen fibers possess less orientation in tendon than in bone. Figure reproduced with permission from the NIH Public Access.[20] [Contacting publisher]
2.1.2 Tissue Engineering Strategies
As is described above, there three requirements justifying the quality of a tissue engineering tendon-to-bone strategy.

Plenty of experiments and tests have been performed in order to regenerate the four zone

region.
Based on the fact that seeding multiple cell types onto specific tissue composite is challenging for clinical usage. Mesenchymal stem cells (MSCs) are used since they can be derived from versatile types of tissues and then differentiate to the target cells phenotypes. Gulotta, et al., hypothesized that the utilization of bone marrow-derived MSCs would improve the biological environment around the repair to promote regeneration of the natural insertion site, and to prevent the formation of scar tissue.[21] In this experiment, they tried to apply MSCs to the healing rotator cuff insertion site of Lewis rats. However, compared to the no addition MSCs animal models, no difference was detected upon the improvement of the structure or the strength of the healing tendon attachment site. Knowing the fact that, spatial changes of cell phenotypes result in changes in tissue composition and structure by the secretion of biological factors. Biological factors might play a key role in regulating cell phenotype and tissue integrity as the gradients in biological factors reflect a.[22] Wang et al. presented a possible solution to the difficulties of culturing several types of cells by utilizing two growth factors, bone morphogenetic protein 2 (BMP-2) and insulin-like growth factor I (IGF-I), as a single concentration gradient or reverse gradient combined the scaffolds.[22] Initially, an alginate gel was fabricated into a cylinder shape with microspheres incorporated as gradients, following the construction of the gradient making system and the delivery efficiency of polylactic-co-glycolic acid (PLGA) and silk fibroin microspheres. On the surface of silk constructs adsorbed with BMP-2 and IGF-I, MSCs were cultured in osteochondral medium. By increasing the content of BMP-2 while reducing IGF-I content, graded increase of calcium, glycosaminoglycan (GAG) deposition, type I, II, and X collagen gene transcriptions were obtained. The obtained gradients have similar trends to the unmineralized fibrocartilage to mineralized fibrocartilage in normal insertion sites.
Other than cell phenotypes and biological factors, which play important roles in realization of a transitional area, biomechanical driving force is also a key challenge in cell differentiation. Biomechanical force, electromagnetic fields, and ultrasound have been proved to play an active part in regulating the differentiation of MSCs.[23-25] Chen et al., has reported that mRNA expressions of tendon related and osteoblast-specific marker genes in human MSCs seeded onto a 6-well plate coated with type I collagen.[26] The results shown that the mRNA expressions of type I, III collagen, and tenascin-C significantly increased in MSCs when underwent 10% stretching for 48 hours. Nevertheless, the effects from stretching still existed after the stretched cells had rested for 48 hours. This is one of the supportive demonstrations of using biological factors to induce the formation of a graded tendon-to-bone attachment with addition of MSCs. Other methodology such as utilizing electrospun PLGA nanofibers as a substrate and applying simulated body fluid to generate mineral gradient is also a alterative solution.[27]
Researchers have postulated that the mechanical loading is of great importance in regulating the graded transition in cell morphology, mineral content, and tissue biomechanical properties.[28-30] Tatara et al. made a hypothesis that muscle unloading would suppress bone formation and enhance bone resorption at the enthesis, and that the unloading-induced bony defects could be rescued by suppressing osteoclast activity.[31] With comparison at enthesis between muscle loading mice and unloading mice, the later resulted in delay of endochondral of ossification at enthesis as well as several defects in bone volume and cancellous bone structure. The study demonstrated that the bone remodeling at developing enthesis requires muscle forces. Another experiment was achieved by Schwarz et al. by measure the mineral content and pattern in murine supraspinatus enthesis development in the absence of muscle[32] This localized paralysis model resulted in joint level deformities and mineralization defects. In addition, significantly decrease in tissue biomechanical properties along with changes in composition and structure. According to the results from Raman spectroscopy and transmission electron microscopy, a further research could be perform by combining the biological signaling with muscle loading.