RESUMO
Principal component analysis (PCA) as a machine-learning technique could serve in disease diagnosis and prognosis by evaluating the dynamic morphological features of exosomes via Cryo-TEM-imaging. This hypothesis was investigated after the crude isolation of similarly featured exosomes derived from the extracellular vehicles (EVs) of immature dendritic cells (IDCs) JAWSII. It is possible to identify functional molecular groups by FTIR, but the unique physical and morphological characteristics of exosomes can only be revealed by specialized imaging techniques such as cryo-TEM. On the other hand, PCA has the ability to examine the morphological features of each of these IDC-derived exosomes by considering software parameters such as various membrane projections and differences in Gaussians, Hessian, hue, and class to assess the 3D orientation, shape, size, and brightness of the isolated IDC-derived exosome structures. In addition, Brownian motions from nanoparticle tracking analysis of EV IDC-derived exosomes were also compared with EV IDC-derived exosome images collected by scanning electron microscopy and confocal microscopy. Sodium-Dodecyl-Sulphate-Polyacrylamide-Gel-Electrophoresis (SDS-PAGE) was performed to separate the protein content of the crude isolates showing that no considerable protein contamination occurred during the crude isolation technique of IDC-derived-exosomes. This is an important finding because no additional purification of these exosomes is required, making PCA analysis both valuable and novel.
RESUMO
Electrospray is a promising technique to scale-up production of microparticles and nanoparticles. In this study, electrospraying was used in order to produce candidate biopatches (CPH) by using chitosan, poly(ethylene glycol) (PEG) and hyaluronic acid (HA). Four different ratios of polymer blend compositions (CPH1, CPH2, CPH3 and CPH4) were tested by dissolving in 2% acetic acid solution (Ac.A.). The HA amount in each blend was kept the same to designate the optimum surface with different chitosan/PEG ratios for electrospray process. Fourier-transform infrared (FTIR) microscopy, scanning electron microscopy (SEM), and atomic force microscopy (AFM) studies showed that obtained patches had highly adhesive surfaces with the aid of heterogeneously distributed micro- and nano-particles. Additionally, video images of FTIR microscopy and AFM images proved that all surfaces have similar heterogeneity except CPH2. The most homogenous surface was obtained by CPH3. Patches were directly subjected to antibacterial tests against ten different types of gram positive and gram negative bacteria using disc diffusion assay (Kirby-Bauer method). Extraordinarily there was no antibacterial property of patches coated with microparticles. Finally, biocompatibility studies were performed by using mouse fibroblast L929 cell lines (ATTC number CCL-1) to test cell adhesion and proliferation properties of the patches. Results of 72 h viability tests proved the electrospray of ternary blends had displayed good biocompatibility; in particular, CPH3 had the highest cell viability.
