The role of S100 proteins (MRP8, MRP14) in cellular dynamics of phagocytes

The major calcium-binding proteins expressed in phagocytes are myeloid-related protein 8 (MRP8 [S100A8]) and MRP14 (S100A9). Phagocytes have the ability to migrate rapidly along the activated endothelium to sites of inflammation. MRP14 knockout (ko) cells spontaneously adhered to a greater extend and transmigrated more in number than wildtype cells. Wildtype cells showed a higher basal surface expression of CD49d when compared to the ko. However, after 1 hour of adhesion, the CD49d expression level was downregulated in the wildtype and upregulated in the ko cells. Accordingly, adhesion patterns of wildtype and ko cells reversed in the presence of primary ligands of CD49d. Upon blocking CD49d expression, the adhesion and transmigration ability of the ko cells decreased. Src and Syk kinases were found to be dysregulated in ko cells compared to wildtype cells. Our data precisely indicates that MRP8/14 definitely modulates the function of migrating phagocytes and successfully guides their path along the activated endothelium

Single-Cell Cloning of Breast Cancer Cells Secreting Specific Subsets of Extracellular Vesicles

Extracellular vesicles (EVs) mediate communication in health and disease. Conventional assays are limited in profiling EVs secreted from large populations of cells and cannot map EV secretion onto individual cells and their functional profiles. We developed a high-throughput single-cell technique that enabled the mapping of dynamics of EV secretion. By utilizing breast cancer cell lines, we established that EV secretion is heterogeneous at the single-cell level and that non-metastatic cancer cells can secrete specific subsets of EVs. Single-cell RNA sequencing confirmed that pathways related to EV secretion were enriched in the non-metastatic cells compared with metastatic cells. We established isogenic clonal cell lines from non-metastatic cells with differing propensities for CD81+CD63+EV secretion and showed for the first time that specificity in EV secretion is an inheritable property preserved during cell division. Combined in vitro and animal studies with these cell lines suggested that CD81+CD63+EV secretion can impede tumor formation. In human non-metastatic breast tumors, tumors enriched in signatures of CD81+CD63+EV have a better prognosis, higher immune cytolytic activity, and enrichment of pro-inflammatory macrophages compared with tumors with low CD81+CD63+EVs signatures. Our single-cell methodology enables the direct integration of EV secretion with multiple cellular functions and enables new insights into cell/disease biology