The diversity and
safety of nanofibrillated cellulose (NFC) hydrogels
have gained a vast amount of interest at the pharmaceutical site in
recent years. Moreover, this biomaterial has a high potential to be
utilized as a protective matrix during the freeze-drying of heat-sensitive
pharmaceuticals and biologics to increase their properties for long-term
storing at room temperature and transportation. Since freeze-drying
and subsequent reconstitution have not been optimized for this biomaterial,
we must find a wider understanding of the process itself as well as
the molecular level interactions between the NFC hydrogel and the
most suitable lyoprotectants. Herein we optimized the reconstitution
of the freeze-dried NFC hydrogel by considering critical quality attributes
required to ensure the success of the process and gained insights
of the obtained experimental data by simulating the effects of the
used lyoprotectants on water and NFC. We discovered the correlation
between the measured characteristics and molecular dynamics simulations
and obtained successful freeze-drying and subsequent reconstitution
of NFC hydrogel with the presence of 300 mM of sucrose. These findings
demonstrated the possibility of using the simulations together with
the experimental measurements to obtain a more comprehensive way to
design a successful freeze-drying process, which could be utilized
in future pharmaceutical applications.
The analysis of nanoparticle (NP) dynamics in live cell studies by video tracking provides detailed information on their interactions and trafficking in the cells. Although the video analysis is not yet routinely used in NP studies, the equipment suitable for the experiments is already available in most laboratories. Here, we compare trajectory patterns, diffusion coefficients, and particle velocities of NPs in A549 cells with a rather simple experimental setup consisting of a fluorescence microscope and openly available trajectory analysis software. The studied NPs include commercial fluorescent polymeric particles and two subpopulations of PC-3 cell-derived extracellular vesicles (EVs). As bioderived natural nanoparticles, the fluorescence intensities of the EVs limited the recording speed. Therefore, we studied the effect of the recording frame rate and analysis parameters to the trajectory results with bright fluorescent commercial NPs. We show that the trajectory classification and the apparent particle velocities are affected by the recording frame rate, while the diffusion constants stay comparable. The NP trajectory patterns were similar for all NP types and resembled intracellular vesicular transport. Interestingly, the EV movements were faster than the commercial NPs, which contrasts with their physical sizes and may indicate a greater role of the motor proteins in their intracellular transports.
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