Analysis of Keratinocytic Exosomes from Diabetic and Nondiabetic Mice by Charge Detection Mass Spectrometry.

Unresolved inflammation compromises diabetic wound healing. Recently, we reported that inadequate RNA packaging in murine wound-edge keratinocyte-originated exosomes () leads to persistent inflammation [Zhou, X. 2020, 14(10), 12732-12748]. Herein, we use charge detection mass spectrometry (CDMS) to analyze intact isolated from a 5 day old wound-edge tissue of diabetic mice and a heterozygous nondiabetic littermate control group. In CDMS, the charge () and mass-to-charge ratio (/) of individual exosome particles are measured simultaneously, enabling the direct analysis of masses in the 1-200 MDa range anticipated for exosomes. These measurements reveal a broad mass range for from ∼10 to >100 MDa. The and values for these exosomes appear to fall into families (subpopulations); a statistical modeling analysis partially resolves ∼10-20 subpopulations. Complementary proteomics, immunofluorescence, and electron microscopy studies support the CDMS results that from diabetic and nondiabetic mice vary substantially. Subpopulations having high (>650) and high (>44 MDa) are more abundant in nondiabetic animals. We propose that these high and particles may arise from differences in cargo packaging. The veracity of this idea is discussed in light of other recent CDMS results involving genome packaging in vaccines, as well as exosome imaging experiments. Characterization of intact exosome particles based on the physical properties of and provides a new means of investigating wound healing and suggests that CDMS may be useful for other pathologies.