Development of vitamin C irrigation saline to promote graft healing in anterior cruciate ligament reconstruction

SummaryGraft healing in anterior cruciate ligament (ACL) reconstruction is often slow and compromised. There are numerous treatment strategies targeting at the biological healing process in ACL reconstruction. Most of these treatments attempt to promote graft incorporation at the tendon to bone interface, although the intra-articular mid substance would be more problematic. As intraoperative injection of vitamin C has been reported to be able to promote tendon healing, we speculate that vitamin C supplementation may also work for graft healing enhancement in ACL reconstruction. The purpose of this study is to formulate a vitamin C-supplemented saline for intraoperative irrigation, in order to promote biological healing in ACL reconstruction. The study hypothesis was tested using 114 rats. During ACL reconstruction, the knee joint of the rats were irrigated with either normal saline, or saline supplemented with 3 mg/mL, 10 mg/mL, or 30 mg/mL vitamin C. Inflammatory response was measured by serum C-reactive protein enzyme-linked immunosorbent assay at Day 1, Day 4, and Day 7 postoperation. Functional recovery was evaluated by gait analysis at preinjury, and Week 4 and Week 6 postoperation. Knee samples were harvested at Week 6 postoperation for anterior–posterior (A–P) knee laxity test and graft pull-out test. Histology was performed at Day 1, Day 4, Day 7, and Day 42 for scoring. Vitamin C supplementation at all doses significantly reduced serum C-reactive protein level as compared to saline control at Day 1 (p = 0.029) postoperation. At 6 weeks postoperation, ACL reconstruction with 3 mg/mL vitamin C irrigation led to a better restoration of A–P knee laxity as compared to saline irrigation (p = 0.032), whereas no positive effect was observed in the groups with 10 mg/mL and 30 mg/mL vitamin C irrigation. Vitamin C irrigation did not affect ultimate load of the graft complex. Histological analysis revealed that at Week 6 postoperation, graft deterioration was significantly reduced in 3 mg/mL and 10 mg/mL vitamin C groups, but no significant difference was observed in graft incorporation among different groups at Day 42 postoperation. Although no significant difference was detected in the gait parameters among experimental groups, increased percentage of rats with asymmetric walking gait (with limb idleness index > 1.3) was found in the groups of 10 mg/mL or 30 mg/mL vitamin C irrigation. The results showed that intraoperative supplementation of low dose (3 mg/mL) vitamin C effectively reduced graft deterioration and improved restoration of A–P knee laxity at 6 weeks postoperation, but higher doses of vitamin C irrigation may have side effects that lead to a higher chance of gait asymmetry. This study reveals the potential use of surgical irrigation saline as a delivery method for bioactive agents to promote healing, which might be a cost-effective and translatable approach for biological modulation for orthopaedic surgeries.