A Sequential Micro-Immunotherapy Medicine Increases Collagen Deposition in Human Gingival Fibroblasts and in an Engineered 3D Gingival Model under Inflammatory Conditions

Periodontal therapies use immune mediators, but their side effects can increase with dosage. Micro-immunotherapy (MI) is a promising alternative that employs immune regulators at low and ultralow doses to minimize adverse effects. In this study, the effects of 5 capsules and the entire 10-capsule sequence of the sequential MI medicine (MIM-seq) were tested in two in vitro models of periodontitis. Firstly, human gingival fibroblasts (hGFs) exposed to interleukin (IL)-1 & beta; to induce inflammation were treated with five different capsules of MIM-seq for 3 days or with MIM-seq for 24 days. Subsequently, MIM-seq was analyzed in a 3D model of human tissue equivalent of gingiva (GTE) under the same inflammatory stimulus. Simultaneously, a non-IL-1 & beta;-treated control and a vehicle were included. The effects of the treatments on cytotoxicity, collagen deposition, and the secreted levels of IL-1 & alpha;, IL-6, prostaglandin E2 (PGE2), matrix metalloproteinase-1 (MMP-1), and tissue inhibitor of metalloproteinases-1 (TIMP-1) were evaluated. None of the tested items were cytotoxic. The complete sequence of MIM-seq decreased PGE2 release and restored collagen deposition levels induced by IL-1 & beta; treatment in hGFs exposed to IL-1 & beta;. MIM-seq treatment restored collagen production levels in both models. These promising preclinical findings suggest that MIM-seq should be further investigated for periodontitis treatment.