Inguinal Lymph Nodes Flap Transfer Promotes Lymphatic Channel Formation in the Rat Sciatic Nerve

INTRODUCTION: The potential role of the lymphatic system in peripheral nerve regeneration is explained by its ability to prevent edema, thereby reducing the risk of intraneural compartment syndrome and subsequent demyelination. Additionally, its anti-inflammatory properties and contribution to neovascularization help create a supportive environment for nerve regeneration. However, the presence and functional significance of the lymphatic system in peripheral nerves remain poorly understood. OBJECTIVES: Our primary objective is to investigate the presence and characteristics of the lymphatic structures within the normal rat sciatic nerve. The secondary objective is to develop a lymphatic tissue transfer model and evaluate its ability to promote lymphatic channel formation within the nerve. METHODS: Immunofluorescence (IF) staining was performed on Wister Lewis rat sciatic nerve using podoplanin (PDPN), a specific marker for lymphatic endothelium, along with CD31, an endothelial marker for blood vessels that is also expressed in lymphatic vessels [1]. For lymphatic tissue transfer, rat inguinal lymph nodes (LN) were harvested with adipose tissues as a pedicled SIEA flap [2] and positioned around the transected and repaired sciatic nerve, ensuring close proximity of the LN to the neural repair site. Eight weeks post-surgery, IF staining was performed on the nerve covered by the inguinal LN flap. The number of PDPN- and/or CD31-positive areas within a 500 um square region was counted by five independent volunteers who were not associated with this project. The averages were then compared among three groups: normal nerve (control), transected and repaired nerve (neurorrhaphy only), and transected and repaired nerve covered by the inguinal LN flap (LN flap coverage). RESULTS: IF staining of the normal rat sciatic nerve revealed that PDPN/CD31 double-positive areas adjacent to CD31 single-positive areas on the outer side of the nerve fibers, specifically within the fibrofatty tissue surrounding the epineurium. Counting by five independent volunteers demonstrated that the average number of PDPN/CD31 double-positive areas was significantly higher in the LN flap coverage group than in the control (45.6 vs. 2.6, p = 0.0002) and the neurorrhaphy-only groups (45.6 vs. 2.0, p = 0.0002). DISCUSSION: Our results confirm the presence of lymphatic vessels adjacent to blood vessels in the fibrofatty tissue of the normal rat sciatic nerve. Considering that lymphatic channels were nearly absent within the nerve fibers in the control and neurorrhaphy-only groups, the significant increase observed in the LN flap coverage group suggests that the transferred inguinal LN facilitated lymphatic channel formation, potentially supporting nerve regeneration. CONCLUSION: The inguinal LN flap transfer significantly promotes lymphatic channel formation within sciatic nerve fibers. This suggests that the transferred LN contributes to lymphatic channel induction, potentially enhancing nerve regeneration and tissue repair following peripheral nerve injury.