Non-small cell lung cancer invading the visceral pleura is characterized by a particular richness of mediastinal lymph node (LN) metastases. This may be due to subpleural lymphatic drainage of tumor cells. The aim of this study was to determine mediastinal LN lymphatic drainage from the diaphragmatic pleura. Subpleural lymphatics of 30 adult cadavers and 12 fetuses were injected with a modified Gerota's medium to permit lymph vessels and nodes to be visualized and then dissected. Each stage of the dissection was described and photographed. In 32 cadavers mediastinal visceral LN chains were injected, of which 29 originated from the mediolateral portion of the diaphragm. On the right, injections (n=16) demonstrated lymph vessels (n=20) ascending directly along the inferior pulmonary ligaments (n=8) or after having encircled the inferior vena cava (n=8), and lymph vessels passing between the pulmonary veins (n=4); all these lymphatics were connected to the intertracheobronchial nodes and some ascended along the tracheobronchial LN chains in the upper mediastinum. On the left, injections (n=13) demonstrated lymph vessels (n=16) ascending along the inferior pulmonary ligament (n=5) or along the esophagus (n=11) and connecting to the intertracheobronchial nodes, some of which ascended further in the upper mediastinum (left paratracheobronchial LN chain). These mediastinal LN chains are the same as those that receive lymph from the pulmonary segments. Lymphatic drainage of the diaphragmatic pleura may add to that of the lung involved in cancer and potentially increases lymphatic spread of tumor cells.