Opuntia fícus-indica and Opuntia cochenillifera are species of Cactaceae, found in the arid regions of the planet. They present water, cellulose, hemicellulose, pectins, extractives, ashes and lignins. Here we aimed to study the immunomodulatory action of lignins from these two species against mice splenocytes, since no study for this purpose has yet been reported. The antioxidant activities of these lignins were evaluated by the DPPH, ABTS, NO assays and total antioxidant activity. Cytotoxicity was evaluated through Annexin V-FITC and propidium iodide-PE probs and cell proliferation was determined by CFSE. Immunomodulation studies with Opuntia lignins obtained were performed through investigation of ROS levels, cytosolic calcium release, changes on mitochondrial membrane potential, cytokine production and NO release. Results showed that Opuntia cochenillifera lignin presented more phenolic amount and antioxidant activities than Opuntia ficius-indica. Both lignins showed high cell viability (>96%) and cell proliferation. Activation signal was observed for both lignins with increase of ROS and cytosolic calcium levels, and changes in mitochondrial membrane potential. In addition, lignins induced high TNF-α, IL-6 and IL-10 production and reduced NO release. Therefore, these lignins present great potential to be used as molecules with a proinflammatory profile, being shown as a promising therapeutic agent.