Tendon injuries in sheep that were treated with stem cell grafts achieved, in just two months, a diameter and hardness similar to the original healthy tendon, according to the results of a study (“Adipose micro-grafts enhance tendinopathy healing in ovine model: An in vivo experimental perspective study”) released in Stem Cells Translational Medicine (SCTM). These findings suggest that the treatment, which uses autologous adipose micrografts (AAMGs, stem cells derived from fat taken from the recipient) presents a safe, reliable, and relatively fast way to promote tendon healing, according to the researchers.

Due to overuse or age-related degeneration, tendon injuries have become a common clinical problem. Damaged tendons heal slowly, and current treatments often can’t manage the pain. They also are unable to restore the tendon’s original structure and functionality.

“Not only does the patient suffer, but the increased incidence rate and ineffective treatments of tendon and other musculoskeletal disorders have resulted in a rise of up to $874 billion from 2000 to 2015, representing an important socioeconomic burden on healthcare worldwide; hence, an effective and affordable therapeutic plan is surely imperative,” said Francesco De Francesco, MD, a member of the Reconstructive Surgery and Hand Surgery unit at AOU “Ospedali Riuniti” (United Hospitals) of Ancona, Italy.

De Francesco was co-senior author of the new study, which was a multi-institutional collaboration involving colleagues from his university as well as from the University of Camerino, University of Parma, Polytechnic University of Marche, and the University of Ferrara.

In a previous study on rats, AAMGs and stromal vascular fraction (SVF) improved tendon healing in 60 percent to 70 percent of treated animals. The purpose of this new study reported in SCTM was to evaluate the effects of AAMG in sheep with tendinopathy, as larger animals are more comparable to humans than are rodents.

“In Europe, approximatively 100,000 to 500,000 tendon repairs are performed every year. These procedures are associated with a considerable rate of postoperative complications (from 6% to 11%). Autologous micro-grafts (AAMG) and stromal vascular fraction (SVF) have been shown to improve tendon healing in 60% to 70% of treated rodents,” write the investigators.

“The purpose of this study was to evaluate the effects of AAMG in a sheep model with tendinopathy. We used sheep models because, as a large animal, they are more comparable to humans. The hypothesis was that SVF injection would improve tendon healing compared with the control group, reducing inflammatory and matrix degrading, while increasing anti-inflammatory expression and collagen synthesis in the early stage of tendon injury. Sixteen Apennine sheep aged 2 to 5 years underwent 500 UI type I collagenase injections into both common calcaneal tendons (CCT) to induce tendinopathy.

“After 15 days (T0), one CCT in every ovine underwent randomly to 2.5 mL of AAMG obtained by mechanical disruption and the contralateral CCTs received no treatment. Clinical, ecographic, and sonographic evaluations were performed after 4 weeks (T1) and 8 weeks (T2). Histological, immunohistochemical, real-time polymerase chain reaction (RT-PCR), and biomechanical evaluations were performed at T2. At T2, the treated group showed a final tendon diameter (9.1 ± 1.4 mm) and a hardness expression (62%) that were similar to the original healthy tendon (8.1 ± 1.1 mm; 100%), with a significant recovery compared with the control group (9.5 ± 1.7 mm; 39%).

“Moreover, histological analysis of the treated group revealed an improvement in the fiber orientation score, fiber edema score, infiltrative-inflammatory process, and necrosis score (4.3 ± 3.3) compared with control group (8.8 ± 2.9). Immunohistochemically, the treated group showed high expression of collagen 1, Factor VIII and significantly low expression of collagen 3. These data were confirmed by RT-PCR analysis.

“The study findings suggested that AAMGs obtained through mechanical disruption present a safe, efficient, and reliable technique, enhancing tendon healing.

Alternative to enzymatic digestion to isolate the SVF

“This is also the first study on an animal model employing a mechanical fat breakdown system (Rigenera Technology, Rigenera®, HBW, Turin, Italy) as an alternative to enzymatic digestion to isolate the SVF. The process consists of gently disaggregating adipose tissue using a particular micro-blade grid and a filter for cells within a sterile capsule. This device leads to the generation of a micrograft suspension that is ready for use and rich in SVF, extracellular matrix fragments and growth factors, and facilitates and enhances the regenerative potential of the isolated tissue fragments,” De Francesco explained.

This procedure is able to maintain the microenvironment of the perivascular niche, while at the same time removing any pro-inflammatory factors. The residual SVF contain pericytes that are able to gradually convert into activated adipose stem cells.

“The resulting AAMG has a great anti-inflammatory and healing effect when applied to musculoskeletal disorders. Moreover, the harvesting procedure is easier, faster, safer, and more reliable and with less morbidity to the donor site than harvesting bone marrow or platelet-rich plasma,” added De Francesco.

“Two-months post-inoculation, data gained from our analyses showed that in the group treated with SVF the tendon diameter and hardness were similar to that of uninjured tendons. Additionally, we observed positive effects in matrix composition in the treated tendons and in collagen deposits, as well as noted improved blood vessel formation within the lesion sites,” reported co-senior author Michele Riccio, MD, director of the Reconstructive Surgery and Hand Surgery Unit at AOU.

“Our findings suggest that the beneficial effects of tendon repair induced by SVF is attributable to the maintenance and induction of tendon fiber organization, rather than an increase in a pool of cells as part of the healing process. This further indicates that SVFs represent a safe, reliable, and more effective treatment for tendinopathy, with a lower rate of post-intervention complications, than current therapies. We believe it strengthens the rationale for their use as a tendinopathy treatment in humans.”

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