Abstract Interleukin‐15 (IL‐15) is a pleiotropic cytokine that plays a key regulatory role in immune responses and inflammation by promoting the proliferation and survival of natural killer (NK) cells and CD8 + T cells. However, the large‐scale production of bioactive recombinant human interleukin‐15 (rhIL‐15) remains challenging due to the limitations of conventional expression systems. In this study, we developed a transgenic silkworm bioreactor for the efficient production of bioactive rhIL‐15 in the posterior silk glands (PSG) using the fibroin heavy chain (FibH) expression system. The human IL‐15 coding sequence was fused with the N‐terminal domain of FibH and introduced into the silkworm genome via piggyBac ‐mediated germline transformation. The recombinant protein was successfully expressed in PSG and secreted into the cocoons, achieving a yield of 14.18 mg/g cocoon shell. Transgenic silkworms exhibited normal growth and cocoon traits comparable to those of wild‐type controls. The purified rhIL‐15 effectively promoted mouse cytotoxic T lymphocyte line (CTLL‐2) cell proliferation, increased intracellular ATP levels, inhibited apoptosis, and upregulated the expression of downstream target genes, including Myc , Pim1 , and Bcl2 . Importantly, its bioactivity was comparable to that of a commercial IL‐15 standard. Collectively, these findings demonstrate that the silkworm silk gland bioreactor represents a robust and potentially cost‐effective platform for the production of bioactive rhIL‐15. Furthermore, the resulting functionalized silk fibers may hold promise for applications in immunomodulatory biomaterials.
