Tatiana Avsievich

Reconstitution of synthetic cells holds potential to advance synthetic biology, biomanufacturing, and therapeutics. Microfluidic generation of cell‐sized liposomes via double emulsion templating offers precise control over composition and formation process, yet the principles underlying solvent‐mediated dewetting remain poorly understood. Using a solvent combination of hexanol and paraffin oil, we demonstrate that solvent‐mediated dewetting liposome generation entails both solvent removal and the application of mechanical stimuli. Solvent removal suffices to induce the morphological transition from double emulsions to partially dewetted liposomes exhibiting low and high budding angles of the residual oil pockets. This transition is driven by relaxation of monolayer and membrane tensions, arising from the increased lipid packing density at the liposome interfaces during solvent depletion. While dewetting kinetics and intermediate stages are governed by solvent removal rate, complete dewetting is not spontaneous. Using optical tweezers, we identify tethering between the liposome and oil pocket and characterize the mechanical force required for liposome detachment. By integrating these principles, a predictive, high‐throughput approach for generating biocompatible, surfactant‐free liposomes is provided. These findings establish a mechanistic framework for liposome dewetting and, through similarities to lipid droplet morphogenesis, offer a protocell platform that could further the understanding of biological budding processes.

Optical tweezers (OTs) are innovative instruments utilized for the manipulation of microscopic biological objects of interest. Rapid improvements in precision and degree of freedom of multichannel and multifunctional OTs have ushered in a new era of studies in basic physical and chemical properties of living tissues and unknown biomechanics in biological processes. Nowadays, OTs are used extensively for studying living cells and have initiated far-reaching influence in various fundamental studies in life sciences. There is also a high potential for using OTs in haemorheology, investigations of blood microcirculation and the mutual interplay of blood cells. In fact, in spite of their great promise in the application of OTs-based approaches for the study of blood, cell formation and maturation in erythropoiesis have not been fully explored. In this review, the background of OTs, their state-of-the-art applications in exploring single-cell level characteristics and bio-rheological properties of mature red blood cells (RBCs) as well as the OTs-assisted studies on erythropoiesis are summarized and presented. The advance developments and future perspectives of the OTs’ application in haemorheology both for fundamental and practical in-depth studies of RBCs formation, functional diagnostics and therapeutic needs are highlighted.

In the framework of novel medical paradigm the red blood cells (RBCs) have a great potential to be used as drug delivery carriers. This approach requires an ultimate understanding of the peculiarities of mutual interaction of RBC influenced by nano-materials composed the drugs. Optical tweezers (OT) is widely used to explore mechanisms of cells’ interaction with the ability to trap non-invasively, manipulate and displace living cells with a notably high accuracy. In the current study, the mutual interaction of RBC with polymeric nano-capsules (NCs) is investigated utilizing a two-channel OT system. The obtained results suggest that, in the presence of NCs, the RBC aggregation in plasma satisfies the ‘cross-bridges’ model. Complementarily, the allocation of NCs on the RBC membrane was observed by scanning electron microscopy (SEM), while for assessment of NCs-induced morphological changes the tests with the human mesenchymal stem cells (hMSC) was performed. The combined application of OT and advanced microscopy approaches brings new insights into the conception of direct observation of cells interaction influenced by NCs for the estimation of possible cytotoxic effects.

Optical Tweezers (OT), as a revolutionary innovation in laser physics, has been extremely useful in studying cell interaction dynamics at a single-cell level. The reversible aggregation process of red blood cells (RBCs) has an important influence on blood rheological properties, but the underlying mechanism has not been fully understood. The regulating effects of low-level laser irradiation on blood rheological properties have been reported. However, the influence of pulsed laser irradiation, and the origin of laser irradiation effects on the interaction between RBCs remain unclear. In this study, RBC interaction was assessed in detail with OT. The effects of both continuous and pulsed low-level He–Ne laser irradiation on RBC aggregation was investigated within a short irradiation period (up to 300 s). The results indicate stronger intercellular interaction between RBCs in the enforced disaggregation process, and both the cell contact time and the initial contact area between two RBCs showed an impact on the measured disaggregation force. Meanwhile, the RBC aggregation force that was independent to measurement conditions decreased after a short time of pulsed He–Ne laser irradiation. These results provide new insights into the understanding of the RBC interaction mechanism and laser irradiation effects on blood properties.

Life sciences that focus on improving the quality and standard of life have attracted worldwide attention in fundamental sciences and have promoted the development of novel tools to reveal the mechanism behind biological activities. Advances in single-cell level methods have further deepened the cell biology study, and helped unravel the different structures and function of living cells on a microscopic and molecular level. Among these methods is the Optical Tweezers (OT), a significant achievement of laser physics, which has been widely applied to understand cell interaction dynamics with the ability to non-invasively trap, manipulate and displace a living cell or part of it with highly accurate positioning of the cells. The reversible aggregation of red blood cells (RBC) that strongly influences the hemodynamic mechanisms and blood microcirculation may serve not only as an indicator of disease, but also as a factor affecting the pathological process. In this study, a two-channel OT system combined with a chopper-modulated laser irradiating system was applied to investigate the RBC aggregation mechanism and to reveal the influence of the low-level pulsed He-Ne laser short-time irradiation on this process. A proportional relationship between the interaction area of RBC and the aggregation force was obtained, verified the applicability of the depletion layer model to the RBC aggregation process in plasma. More importantly, a regulating effect of low-level He-Ne laser irradiation on this process was discovered. A statistically significant decrease (p < 0.05) in RBC aggregation forces was observed following 120 s laser light with 225 Hz pulse frequency. This observation brings new insights into conception of the regulating effects occurring during the laser light interaction with blood.

As a promising drug delivery system, itself or coupled with red blood cells (RBC), nanoparticles (NP) should be studied in frames of their interaction at the cellular level. Experiments were performed on RBC in autologous blood plasma incubated with different NP – TiO2, ZnO, nanodiamonds and polymeric nanocapsules. RBC aggregates formation in RBC suspension was observed with conventional microscopy, while quantitative interaction force measurements between individual RBC was assessed with optical tweezers. Scanning electron microscopy (SEM) imaging demonstrated NP localization and RBC membrane modifications upon binding with NP. Among tested NP, nanodiamonds caused increasing the size of aggregates in RBC suspensions, RBC interaction force increase and strong membrane surface modifications, comparing to other tested NP and control sample. Nanocapsules do not cause any adverse effects on RBC properties, confirming biocompatibility and applicability for drug delivery purposes. Optical tweezers combined with SEM imaging serves as fast informative assessment of NP effects on RBC.

Optical tweezers (OT) is a unique Nobel Prize winning technology that has been widely used in studying cell interaction dynamics at a single-cell level with highly accurate manipulating of living cells and the ability to detect ultra-low intercellular forces. The reversible aggregation process of red blood cells (RBCs) that strongly influences the blood rheological properties thus being critical for blood microcirculation has long been research studied. However, in spite of plentiful researches dealing with RBC aggregation behavior based on an average response of a large number of cells, the detailed mechanism and the applicability of the two coexistent yet mutually opposed models to this reversible process require additional information. In this study, a two-channel optical tweezers system is utilized to reveal the influence of the cell interaction time (0-300 sec) on the RBC (dis)aggregation dynamics. The results show that for RBC enforced disaggregation in autologous plasma, the longer the two RBCs adhere to each other, the stronger the intercellular interaction is, and the lower the degree that a certain optical pulling force can separate two cells, whereas no significant effect of cell interaction time on RBC aggregation process was observed. This observation indicates that the RBC aggregation and disaggregation in autologous plasma are governed by different mechanisms and that the hysteresis effect, namely the dependence of the disaggregation force on RBC interaction history, is a significant feature that needs special consideration in researches related to the RBC disaggregation process.