Two-pronged approach: Therapeutic effect of biological scaffold combined with immune intervention and β-cell...

Type 1 diabetes mellitus (T1DM) is an autoimmune disease characterized by β-cell damage and absolute insulin deficiency. We consider combining immune intervention and β-cell replacement by biological scaffold to treat T1DM. Zinc transporter 8 (ZnT8) is known to be a pancreatic islet-specific autoantigen. Studies have shown that ZnT8(107-115)/HLA-A2 dimers can be used as antigen-specific immunosuppressants for T1DM. Mesenchymal stem cells (MSCs) can be induced to differentiate into insulin-producing cells (IPCs) under certain conditions in vitro. In recent years, the development of biomaterials has provided a more suitable three-dimensional microenvironment for cell transplantation. Our research group previously prepared gelatin/polylactic acid (PLLA/G) nanofiber scaffold by electrospinning technology, fixed GLP-1 analogues on the scaffold by surface modification with polydopamine (pDA) and confirmed the scaffold can promote bone mesenchymal stem cells (BMSCs) proliferation and improve cell survival rate. In addition, the scaffold can promote the differentiation of BMSCs into IPCs. Subsequently, ZnT8(107-115)/HLA-A2 dimer was constructed and loaded on the scaffold and confirmed the scaffold loaded with immunosuppressants can inhibit the proliferation and cytotoxicity of specific CD8⁺ T cells in vitro. This study used the above scaffold to co-load ZnT8(107-115)/HLA-A2 dimer and IPCs and implanted the scaffold into T1DM mice to study the blood glucose control and immunomodulatory effects on T1DM.

First, the PLLA/G scaffold was modified with pDA and fixed with Liraglutide (LIR) to obtain the PLLA/G-pDA-LIR scaffold. Rat BMSCs were loaded on the scaffold and the 'three-step induction method' was used to induce differentiation in vitro. The insulin expression of IPCs was detected by Dithizone (DTZ) staining, glucose stimulate insulin secretion (GSIS) in vitro and intraperitoneal glucose tolerance test (IPGTT) in vivo. Then, non obese diabetes mice were modelled with T1DM and randomly divided into 5 groups. Blank control group was not treated; negative control group underwent sham surgery; positive control group was injected with IPCs through tail vein; single load scaffold group was subcutaneously transplanted with PLLA/G-pDA-LIR scaffold loaded with IPCs; double load scaffold group was subcutaneously transplanted with PLLA/G-pDA-LIR scaffold loaded with IPCs and ZnT8(107-115)/HLA-A2 dimer. Blood glucose and body weight were measured weekly before and after transplantation. At 2, 4 and 6 W after transplantation, some mice were taken from each group to detect serum insulin and C-peptide, spleen lymphocyte subsets and Tregs and pancreatic Th1/Th2 cell inflammatory factors.

The results of insulin expression in induced differentiated IPCs show that cells with insulin expression can be obtained through the "three-step induction method", and transplanting IPCs can effectively reduce blood glucose and improve glucose tolerance. Compared with non-scaffold induced IPCs, fixing Liraglutide scaffolds can improve the insulin expression level of IPCs and promote the induction of differentiation. The results after scaffold transplantation showed that compared with the positive control group and the single load scaffold group, the blood glucose was significantly reduced, and the serum insulin and C-peptide were significantly increased and lasted longer. In addition, the pathogenic T cells and inflammatory factors in the double load scaffold group were significantly reduced, and Tregs and anti-inflammatory factors were significantly increased.

The results show that the biological scaffold with IPCs and ZnT8(107-115)/HLA-A2 dimers has the dual functions of controlling blood glucose and regulating immunity, and may effectively treat T1DM.