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Abstract
Polymer-based multilayer microencapsulation technology represents one of the promising strategies for intracellular drug delivery, however, membrane processes involved in vehicle internalisation are not fully understood. Here we employed a scanning probe microscopy technique called Scanning Ion Conductance Microscopy (SICM) to study these complex processes at nanoscale resolution in real time. We were able to image topography simultaneously with local elastic modulus throughout the whole course of microcapsule internalisation in A549 cell culture without disrupting the internalisation process. The imaging revealed that capsules triggered the formation of membrane protrusions in their vicinity, which is an important but not a sufficient step towards full capsule internalisation. A crucial aspect appeared to be nanoscale restructuring of these protrusions into smooth thin layers extending over the surface of capsules. Simultaneous mapping of elastic modulus during capsule internalisation allowed monitoring the structural changes during extension of the membrane sheets over the surface of the capsule and the subsequent post-internalisation phenomenon of capsule buckling. To our knowledge these are the first experimental data capturing the interactions between the cellular membrane and microcapsules in their whole complexity with nanoscale resolution. The methodology established here has the potential to provide new insights into interactions at the interface between the nanostructured materials and cellular membrane under physiological conditions.
Highlights
Usability of many chemical substances with a significant potential for biomedical applications is limited by their poor solubility in water or limited stability in the physiological environment
The hopping mode of Scanning Ion Conductance Microscopy (SICM) proved to be capable of capturing freshly produced polyelectrolyte capsules adhered to the bottom of the tissue culture dish (Fig. 1b) as well as live internalisation of the polyelectrolyte capsules with high resolution (Fig. 1c)
The duration of internalisation showed weak, statistically insignificant dependence on the diameter of microcapsules (R = 0.315, p > 0.05, Fig. S1b†). This was in agreement with the results obtained using confocal fluorescence imaging where 52% (17 out of 33) of capsules appeared to be internalized after one-hour incubation (Fig. S1c and d†), suggesting that SICM imaging had not negatively affected the rate of capsule internalisation
Summary
Usability of many chemical substances with a significant potential for biomedical applications is limited by their poor solubility in water or limited stability in the physiological environment. A detailed understanding of physico-chemical and mechanical interactions between capsules and livings cells is required for specific targeting, effective delivery, and elimination of any potential toxic side effects. This has been largely limited by capabilities of available imaging techniques and the 16902 | Nanoscale, 2018, 10, 16902–16910. SICM uses reduction in ionic current through the probe represented by an electrolyte-filled glass nanopipette immersed in a saline solution to detect proximity of the sample surface.[12,13] This technique has been previously used for high-resolution scanning of biological samples of complexity similar to what can be expected in the case of microcapsules interacting with cells,[14,15] and for mapping mechanical properties at high resolution.[16,17]
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