Validation of laser scanning confocal microscopy as a diagnostic method for lymphedema using a rat model

Most previous diagnostic methods for lymphedema are invasive. Laser scanning confocal microscopy (LSCM) combines laser and computer image processing technology, is capable of increasing the resolution of optical microscopy by 30-40%, and boasts a comparable resolution to that of histological examination. We constructed the rat tail lymphedema model to simulate secondary lymphedema and to validate the noninvasive technique of in vivo reflectance LSCM for the diagnosis of lymphedema. The rat tail lymphedema model was constructed by cutting and ligating the lymphatic vessels in the rat tail. Lymphedema in the postoperative rat tail was assessed by a comprehensive range of methods including the change of rat tail diameter, lymphocytic radionuclide imaging, LSCM, and immunohistochemistry using a specific lymphatic vessel marker, prospero homeobox protein 1 (PROX1). The noninvasive LSCM method along with other techniques validated the rat tail lymphedema model. LSCM was used to perform qualitative and quantitative evaluation of the state and extent of lymphedema in the rat tail model. Receiver operating characteristic (ROC) curve analysis, which provided an area under the curve (AUC) value of 0.861, supported the feasibility of using LSCM as a reliable diagnostic technique for lymphedema. The rat tail lymphedema model can be successfully constructed by cutting and ligating the lymphatic vessels in the rat tail. Although LSCM cannot replace the method of skin biopsy examination, it offers a painless and noninvasive alternative for diagnosing lymphedema. Thus, LSCM can potentially be adopted in clinical practice as a supporting method to be used in combination with other techniques.