Assessment of TCR signal strength of antigen-specific memory CD8+ T cells in human blood.

Assessment of the quality and the breadth of antigen (Ag)-specific memory T cells in human samples is of paramount importance to elucidate the pathogenesis and to develop new treatments in various diseases. T-cell receptor (TCR) signal strength, primarily controlled by TCR affinity, affects many fundamental aspects of T-cell biology; however, no current assays for detection of Ag-specific CD8+ T cells can assess their TCR signal strength in human samples. Here, we provide evidence that interferon regulatory factor 4 (IRF4), a transcription factor rapidly upregulated in correlation with TCR signal strength, permits the assessment of the TCR signal strength of Ag-specific CD8+ T cells in human peripheral blood mononuclear cells (PBMCs). Coexpression of IRF4 and CD137 sensitively detected peptide-specific CD8+ T cells with extremely low background in PBMCs stimulated for 18 hours with MHC class I peptides. Our assay revealed that human memory CD8+ T cells with high-affinity TCRs have an intrinsic ability to highly express CD25. Furthermore, HIV-specific CD8+ T cells in chronic HIV+ subjects were found to display primarily low-affinity TCRs with low CD25 expression capacity. Impairment in the functions of HIV-specific CD8+ T cells might be associated with their suboptimal TCR signals, as well as impaired responsiveness to interleukin-2.

Novel nanocomposite biomaterial to differentiate bone marrow mesenchymal stem cells to the osteogenic lineage for bone restoration.

BACKGROUND/OBJECTIVE: As the bone engineering field moves away from nonviable implants to more biocompatible and natural structures, nanomedicine has emerged as a superior tool for developing implantable materials. METHODS: Here, we describe the fabrication and testing of a nanocomposite structure composed of chitosan and a biocompatible thermoplastic (PMMA). RESULTS: Our nanocomposite material displayed morphologically similar characteristics to an extracted murine femur during microscopic and spectroscopic analysis as seen through SEM and FTIR. Crosslinking our nanocomposite enhanced structural and strength characteristics significantly above the noncrosslinked sample, mimicking the strength of an extracted mammalian bone. When cocultured with bone marrow mesenchymal stem cells, the composite material proved to be osteoinductive and osteogenic via DAPI and actin staining, differentiating BMSCs into the osteogenic lineage and promoting mineral deposition. Nodule formation, indicative of mineralization during BMSC differentiation, was confirmed spectroscopically via FTIR and autofluorescence of the nodule. CONCLUSION: These encouraging results show promise for in vivo implantation of our novel scaffold that is both biocompatible and biomimetic in strength and composition.