Combined ascorbic acid and T3 produce better healing compared to bone marrow mesenchymal stem cells in an Achilles tendon injury rat model: a proof of concept study

Abstract

Background This pilot study aimed to ascertain whether the local application of ascorbic acid (AA), of T 3 , and of rat (r) bone marrow mesenchymal stem cells (BMSCs), alone or in all possible combinations, promoted healing after an Achilles tendon injury in a rat model. Methods An Achilles tendon defect was produced in 24 6–8-week-old male inbred Lewis rats. The animals were then randomly divided into eight groups of three rats each. The tendon defect was filled with 50 μL of phosphate-buffered saline (PBS) containing (1) 50 μg/mL AA (AA group), (2) 10 −7 M T 3 (T 3 group), (3) 4 × 10 6 rBMSCs (rBMSC group), (4) 50 μg/mL AA + 10 −7 M T 3 (AA + T 3 group), (5) 4 × 10 6 rBMSCs + 50 μg/mL AA (rBMSC + AA group), (6) 4 × 10 6 rBMSCs + 10 −7 M T 3 (rBMSC + T 3 group), (7) 4 × 10 6 rBMSCS + 50 μg/mL AA + 10 −7 M T 3 (rBMSC + AA + T 3 group), and (8) PBS only (control group: CTRL). All treatments were administered by local injection immediately after the tendons had been damaged; additionally, AA was injected also on the second and fourth day from the first injection (for groups 1, 4, 5, and 7), and T 3 was injected again every day for 4 days (for groups 2, 4, 6, and 7). At 30 days from initial treatment, tendon samples were harvested, and the quality of tendon repair was evaluated using histological and histomorphological analysis. The structure and morphology of the injured Achilles tendons were evaluated using the modified Svensson, Soslowsky, and Cook score, and the collagen type I and III ratio was calculated. Results The group treated with AA combined with T 3 displayed the lowest Svensson, Soslowsky, and Cook total score value of all tissue sections at histopathological examination, with fiber structure close to regular orientation, normal-like tendon vasculature, and no cartilage formation. AA + T 3 also showed the highest collagen I and the lowest collagen III values compared to all other treatments including the CTRL. Conclusion There are potential benefits using a combination of AA and T 3 to accelerate tendon healing.

Supporting Cell-Based Tendon Therapy: Effect of PDGF-BB and Ascorbic Acid on Rabbit Achilles Tenocytes In Vitro

“…As a cofactor for prolyl hydroxylase and lysyl hydroxylase, AA plays a pivotal role in the synthesis of collagen [63,64] and, as such, can play an important role in the healing of connective tissue. Recently, a proof of concept study showed that AA led to better healing in a rat Achilles tendon injury model, particularly in combination with thyroid hormone T3 [65]; with a significant increase in cell proliferation, more regular fiber structure and higher collagen I to collagen III ratio, indicating an accelerated healing process [21]. Moreover, AA supplementation also stimulated Achilles tendon healing in rats after complete rupture [46].…”

Extracellular vesicles from rat-bone-marrow mesenchymal stromal/stem cells improve tendon repair in rat Achilles tendon injury model in dose-dependent manner: A pilot study

“…RBMS/SCs were isolated from the tibia and femur bone-marrow from three 6 to 8-week-old male inbred Lewis rats (ENVIGO, Wyton, UK), as recently described [3]. In brief, bone marrow cells at second passage (P2) were used unless otherwise stated.…”

“…The analysis was performed using the CytExpert Software (Beckman Coulter). The stem-cell characteristics of bone-MSCs were routinely confirmed by their trilineage differentiation abilities (adipocytes, osteoblasts, and chondrocytes) [3]. Adipocytes were visualized by Oil Red-O staining, osteoblasts evidenced by Alizarin red, and chondrocytes were visualized with Alcian Blue.…”

“…Adult, bone marrow-derived mesenchymal stromal/stem cells (BMSCs), are multipotent stem cells which have been widely studied to treat tissue defects, and are generally considered to be a promising alternative to the current therapeutic approach to tendon injuries [2], although contrasting results have also been obtained. Ectopic ossification, calcification and the higher risk of adhesions formation [3,4], as well as the inherent difficulties in quality control before administration [3,4], are among potential problems when using BMSCs for tendon healing.…”

“…Cell-free delivery of bioactive cargos by EV induces the same beneficial responses as stemcell transplantation, offering remarkable benefits over conventional cell-therapy: for example, EVs avoid the risk of tumorigenesis, and heterotopic ossification and calcification [3,4] and are immunologically unresponsive agents [7,8]. Finally EVs play a role in tendon-healing by modulating inflammatory responses [9,10,11].…”