Regulatory T cells (Tregs) are T lymphocytes that perform immunomodulatory functions. By providing accurate, efficient, safe and controlled regulation of systemic immunity, Tregs play a key role in the treatment of autoimmune diseases and immune rejection after organ transplantation. However, Tregs have clinical limitations such as off-targeting and functional phenotype instability. Genetic and biomedical engineering are promising strategies to promote Treg active targeting and chemotaxis to the inflammation site, and to maintain Treg Forkhead box protein 3 (Foxp3) expression during immune-regulation. Herein, Treg anti-inflammatory mechanisms, engineering tactics and bulk production methods are systematically summarized. By anti-inflammatory cytokine secretion, Granzyme B release or effector T cell metabolism intervention, Tregs can inhibit effector T cell proliferation and activation, or even kill immune cells. Moreover, expressing genetically modified receptors (CARs, TCRs and CXCRs) or conjugating nanomaterials onto cell surface, can help enhance targeted recognition and maintain anti-inflammatory function of Tregs. Artificial antigen-presenting-cells or costimulatory molecules can stimulate Treg cell proliferation and activation. By prospecting the application of biomedical-engineered Tregs in autoimmune diseases, organ transplantation and other inflammatory diseases, this work aims to inspire and promote the clinical application of Tregs adoptive therapy.