Extensive investigations in outer space have revealed not only how life adapts to the space environment, but also that interesting biophysical phenomena occur. These phenomena affect human health and other life forms (animals, plants, bacteria, and fungi), and to ensure the safety of future human space exploration need to be further investigated. This calls for joint research efforts between biologists and physicists, as these phenomena present cross-disciplinary barriers. Various national organizations provide useful forums for bridging this gap. Additional discussion avenues and database resources are helpful for facilitating the interdisciplinary investigations of these phenomena. In this paper, we present the newly established Space Life Investigation Database (SpaceLID, https://bidd.group/spacelid/ ) which provides information about biophysical phenomena occurring in space. Examples obtained using the database are given while discussing the underlying causes of these phenomena and their implications for the physiology and health of life in space.

Starting with a model for a product-activated enzymatic reaction proposed for glycolytic oscillations, we show how more complex oscillatory phenomena may develop when the basic model is modified by addition of product recycling into substrate or by coupling in parallel or in series two autocatalytic enzyme reactions. Among the new modes of behavior are the coexistence between two stable types of oscillations (birhythmicity), bursting, and aperiodic oscillations (chaos). On the basis of these results, we outline an empirical method for finding complex oscillatory phenomena in autonomous biochemical systems, not subjected to forcing by a periodic input. This procedure relies on finding in parameter space two domains of instability of the steady state and bringing them close to each other until they merge. Complex phenomena occur in or near the region where the two domains overlap. The method applies to the search for birhythmicity, bursting and chaos in a model for the cAMP signalling system of Dictyostelium discoideum amoebae.

While multicompartment models have long been used to study the biophysics of neurons, it is still challenging to infer the parameters of such models from data including uncertainty estimates. Here, we performed Bayesian inference for the parameters of detailed neuron models of a photoreceptor and an OFF- and an ON-cone bipolar cell from the mouse retina based on two-photon imaging data. We obtained multivariate posterior distributions specifying plausible parameter ranges consistent with the data and allowing to identify parameters poorly constrained by the data. To demonstrate the potential of such mechanistic data-driven neuron models, we created a simulation environment for external electrical stimulation of the retina and optimized stimulus waveforms to target OFF- and ON-cone bipolar cells, a current major problem of retinal neuroprosthetics.

There is a growing appreciation in the fields of cell biology and developmental biology that cells collectively process information in time and space. While many powerful molecular tools exist to observe biophysical dynamics, biologists must find ways to quantitatively understand these phenomena at the systems level. Here, we present a guide for the application of well-established information theory metrics to biological datasets and explain these metrics using examples from cell, developmental and regenerative biology. We introduce a novel computational tool named after its intended purpose, calcium imaging, (CAIM) for simple, rigorous application of these metrics to time series datasets. Finally, we use CAIM to study calcium and cytoskeletal actin information flow patterns between Xenopus laevis embryonic animal cap stem cells. The tools that we present here should enable biologists to apply information theory to develop a systems-level understanding of information processing across a diverse array of experimental systems.

Carbonic anhydrases (CA, EC 4.2.1.1) catalyze the hydration of carbon dioxide and take part in many essential physiological processes. In humans, 15 CAs are characterized, including the only secreted isoenzyme CA VI. CA VI has been linked to specific processes in the mouth, namely bitter taste perception, dental caries, and maintenance of enamel pellicle, and implicated in several immunity-related phenomena. However, little is known of the mechanisms of the above. In this study, we characterized human CA VI purified from saliva and milk with biophysical methods and measured their enzyme activities and acetazolamide inhibition. Size-exclusion chromatography showed peaks of salivary and milk CA VI corresponding to hexameric state or larger at pH 7.5. At pH 5.0 the hexamer peaks dominated. SDS- PAGE of milk CA VI protein treated with a bifunctional crosslinker further confirmed that a majority of CA VI is oligomers of similar sizes in solution. Mass spectrometry experiments confirmed that both of the two putative N-glycosylation sites, Asn67 and Asn256, are heterogeneously glycosylated. The attached glycans in milk CA VI were di- and triantennary complex-type glycans, carrying both a core fucose and 1 to 2 additional fucose units, whereas the glycans in salivary CA VI were smaller, seemingly degraded forms of core fucosylated complex- or hybrid-type glycans. Mass spectrometry also verified the predicted signal peptide cleavage site and the terminal residue, Gln 18, being in pyroglutamate form. Thorough characterization of CA VI paves way to better understanding of the biological function of the protein.

Certain G-quadruplex forming guanine-rich oligonucleotides (GROs), including AS1411, are endowed with cancer-selective antiproliferative activity. They are known to bind to nucleolin protein, resulting in the inhibition of nucleolin-mediated phenomena. However, multiple nucleolin-independent biological effects of GROs have also been reported, allowing them to be considered promising candidates for multi-targeted cancer therapy. Herein, with the aim of optimizing AS1411 structural features to find GROs with improved anticancer properties, we have studied a small library of AS1411 derivatives differing in the sequence length and base composition. The AS1411 derivatives were characterized by using circular dichroism and nuclear magnetic resonance spectroscopies and then investigated for their enzymatic resistance in serum and nuclear extract, as well as for their ability to bind nucleolin, inhibit topoisomerase I, and affect the viability of MCF-7 human breast adenocarcinoma cells. All derivatives showed higher thermal stability and inhibitory effect against topoisomerase I than AS1411. In addition, most of them showed an improved antiproliferative activity on MCF-7 cells compared to AS1411 despite a weaker binding to nucleolin. Our results support the hypothesis that the antiproliferative properties of GROs are due to multi-targeted effects.

Helium (4He) ion beam therapy provides favorable biophysical characteristics compared to currently administered particle therapies, i.e., reduced lateral scattering and enhanced biological damage to deep-seated tumors like heavier ions, while simultaneously lessened particle fragmentation in distal healthy tissues as observed with lighter protons. Despite these biophysical advantages, raster-scanning 4He ion therapy remains poorly explored e.g., clinical translational is hampered by the lack of reliable and robust estimation of physical and radiobiological uncertainties. Therefore, prior to the upcoming 4He ion therapy program at the Heidelberg Ion-beam Therapy Center (HIT), we aimed to characterize the biophysical phenomena of 4He ion beams and various aspects of the associated models for clinical integration.

Circulating tumor cells (CTCs) survive in the bloodstream and then seed and invade to foster tumor metastasis. The arrest of cancer cells is favored by permissive flow forces and geometrical constraints. Through the use of high-throughput microfluidic devices designed to mimic capillary-sized vessels, we applied pressure differences to cancer cells (MCF-7 cell line) and recorded the cell traverse-vessel behaviors. Our results showed that cancer cells transform from a Newtonian droplet state to an adhesion/migration state when cancer cells traverse artificial vessels. To explain these phenomena, a modified Newtonian droplet model was also proposed. These phenomena and the modified model may reveal how CTCs in the blood seed and invade vessels under suitable conditions.

The functions performed by the concentric shells of multilayered dsRNA viruses require specific protein interactions that can be directly explored through their mechanical properties. We studied the stiffness, breaking force, critical strain and mechanical fatigue of individual Triple, Double and Single layered rotavirus (RV) particles. Our results, in combination with Finite Element simulations, demonstrate that the mechanics of the external layer provides the resistance needed to counteract the stringent conditions of extracellular media. Our experiments, in combination with electrostatic analyses, reveal a strong interaction between the two outer layers and how it is suppressed by the removal of calcium ions, a key step for transcription initiation. The intermediate layer presents weak hydrophobic interactions with the inner layer that allow the assembly and favor the conformational dynamics needed for transcription. Our work shows how the biophysical properties of the three shells are finely tuned to produce an infective RV virion.

The functional and biophysical properties of a sustained, or "persistent," Na(+) current (I(NaP)) responsible for the generation of subthreshold oscillatory activity in entorhinal cortex layer-II principal neurons (the "stellate cells") were investigated with whole-cell, patch-clamp experiments. Both acutely dissociated cells and slices derived from adult rat entorhinal cortex were used. I(NaP), activated by either slow voltage ramps or long-lasting depolarizing pulses, was prominent in both isolated and, especially, in situ neurons. The analysis of the gating properties of the transient Na(+) current (I(NaT)) in the same neurons revealed that the resulting time-independent "window" current (I(NaTW)) had both amplitude and voltage dependence not compatible with those of the observed I(NaP), thus implying the existence of an alternative mechanism of persistent Na(+)-current generation. The tetrodotoxin-sensitive Na(+) currents evoked by slow voltage ramps decreased in amplitude with decreasing ramp slopes, thus suggesting that a time-dependent inactivation was taking place during ramp depolarizations. When ramps were preceded by increasingly positive, long-lasting voltage prepulses, I(NaP) was progressively, and eventually completely, inactivated. The V(1/2) of I(NaP) steady state inactivation was approximately -49 mV. The time dependence of the development of the inactivation was also studied by varying the duration of the inactivating prepulse: time constants ranging from approximately 6.8 to approximately 2.6 s, depending on the voltage level, were revealed. Moreover, the activation and inactivation properties of I(NaP) were such as to generate, within a relatively broad membrane-voltage range, a really persistent window current (I(NaPW)). Significantly, I(NaPW) was maximal at about the same voltage level at which subthreshold oscillations are expressed by the stellate cells. Indeed, at -50 mV, the I(NaPW) was shown to contribute to >80% of the persistent Na(+) current that sustains the subthreshold oscillations, whereas only the remaining part can be attributed to a classical Hodgkin-Huxley I(NaTW). Finally, the single-channel bases of I(NaP) slow inactivation and I(NaPW) generation were investigated in cell-attached experiments. Both phenomena were found to be underlain by repetitive, relatively prolonged late channel openings that appeared to undergo inactivation in a nearly irreversible manner at high depolarization levels (-10 mV), but not at more negative potentials (-40 mV).

Cell-cell fusion is fundamental to a multitude of biological processes ranging from cell differentiation and embryogenesis to cancer metastasis and biomaterial-tissue interactions. Fusogenic cells are exposed to biochemical and biophysical factors, which could potentially alter cell behavior. While biochemical inducers of fusion such as cytokines and kinases have been identified, little is known about the biophysical regulation of cell-cell fusion. Here, we designed experiments to examine cell-cell fusion using bulk metallic glass (BMG) nanorod arrays with varying biophysical cues, i.e. nanotopography and stiffness. Through independent variation of stiffness and topography, we found that nanotopography constitutes the primary biophysical cue that can override biochemical signals to attenuate fusion. Specifically, nanotopography restricts cytoskeletal remodeling-associated signaling, which leads to reduced fusion. This finding expands our fundamental understanding of the nanoscale biophysical regulation of cell fusion and can be exploited in biomaterials design to induce desirable biomaterial-tissue interactions.

Tetracaine (TTC) is a local anesthetic broadly used for topical and spinal blockade, despite its systemic toxicity. Encapsulation in nanostructured lipid carriers (NLC) may prolong TTC delivery at the site of injection, reducing such toxicity. This work reports the development of NLC loading 4% TTC. Structural properties and encapsulation efficiency (%EE > 63%) guided the selection of three pre-formulations of different lipid composition, through a 23 factorial design of experiments (DOE). DLS and TEM analyses revealed average sizes (193-220 nm), polydispersity (< 0.2), zeta potential |- 21.8 to - 30.1 mV| and spherical shape of the nanoparticles, while FTIR-ATR, NTA, DSC, XRD and SANS provided details on their structure and physicochemical stability over time. Interestingly, one optimized pre-formulation (CP-TRANS/TTC) showed phase-separation after 4 months, as predicted by Raman imaging that detected lack of miscibility between its solid (cetyl palmitate) and liquid (Transcutol) lipids. SANS analyses identified lamellar arrangements inside such nanoparticles, the thickness of the lamellae been decreased by TTC. As a result of this combined approach (DOE and biophysical techniques) two optimized pre-formulations were rationally selected, both with great potential as drug delivery systems, extending the release of the anesthetic (> 48 h) and reducing TTC cytotoxicity against Balb/c 3T3 cells.

The hydrophobic effect is essential for many biophysical phenomena and processes. It is governed by a fine-tuned balance between enthalpy and entropy contributions from the hydration shell. Whereas enthalpies can in principle be calculated from an atomistic simulation trajectory, calculating solvation entropies by sampling the extremely large configuration space is challenging and often impossible. Furthermore, to qualitatively understand how the balance is affected by individual side chains, chemical groups, or the protein topology, a local description of the hydration entropy is required. In this study, we present and assess the new method "Per|Mut", which uses a permutation reduction to alleviate the sampling problem by a factor of N! and employs a mutual information expansion to the third order to obtain spatially resolved hydration entropies. We tested the method on an argon system, a series of solvated n-alkanes, and solvated octanol.

The concurrent assessment of principal sonoporation factors has been accomplished in a single systemic study. Microbubble sonodestruction dynamics and cavitation spectral characteristics, ultrasound scattering and attenuation, were examined in relation to the intracellular delivery of anticancer drug, bleomycin. Experiments were conducted on Chinese hamster ovary cells coadministered with Sonovue microbubbles. Detailed analysis of the scattering and attenuation temporal functions culminated in quantification of metrics, inertial cavitation dose and attenuation rate, suitable for cavitation control. The exponents, representing microbubble sonodestruction kinetics were exploited to derive dosimetric, microbubble sonodestruction rate. High intracorrelation between empirically-attained metrics defines the relations which indicate deep physical interdependencies within inherent phenomena. Subsequently each quantified metric was validated to be well-applicable to prognosticate the efficacy of bleomycin delivery and cell viability, as indicated by strong overall correlation (R2 > 0.85). Presented results draw valuable insights in sonoporation dosimetry and contribute towards the development of universal sonoporation dosimetry model. Both bleomycin delivery and cell viability reach their respective plateau levels by the time, required to attain total microbubble sonodestruction, which accord with scattering and attenuation decrease to background levels. This suggests a well-defined criterion, feasible through signal-registration, universally employable to set optimal duration of exposure for efficient sonoporation outcome.

A long-standing problem is how memories can be stored for very long times despite the volatility of the underlying neural substrate, most notably the high turnover of dendritic spines and synapses. To address this problem, here we are using a generic and simple probabilistic model for the creation and removal of synapses. We show that information can be stored for several months when utilizing the intrinsic dynamics of multi-synapse connections. In such systems, single synapses can still show high turnover, which enables fast learning of new information, but this will not perturb prior stored information (slow forgetting), which is represented by the compound state of the connections. The model matches the time course of recent experimental spine data during learning and memory in mice supporting the assumption of multi-synapse connections as the basis for long-term storage.

Three-dimensional (3D) cell cultures represent fundamental tools for the comprehension of cellular phenomena both in normal and in pathological conditions. In particular, mechanical and chemical stimuli play a relevant role on cell fate, cancer onset and malignant evolution. Here, we use mechanically-tuned alginate hydrogels to study the role of substrate elasticity on breast adenocarcinoma cell activity. The hydrogel elastic modulus (E) was measured via atomic force microscopy (AFM) and a remarkable range (150-4000 kPa) was obtained. A breast cancer cell line, MCF-7, was seeded within the 3D gels, on standard Petri and alginate-coated dishes (2D controls). Cells showed dramatic morphological differences when cultured in 3D versus 2D, exhibiting a flat shape in both 2D conditions, while maintaining a circular, spheroid-organized (cluster) conformation within the gels, similar to those in vivo. Moreover, we observed a strict correlation between cell viability and substrate elasticity; in particular, the number of MCF-7 cells decreased constantly with increasing hydrogel elasticity. Remarkably, the highest cellular proliferation rate, associated with the formation of cell clusters, occurred at two weeks only in the softest hydrogels (E = 150-200 kPa), highlighting the need to adopt more realistic and a priori defined models for in vitro cancer studies.

Microorganisms often form complex multicellular assemblies such as biofilms and colonies. Understanding the interplay between assembly expansion, metabolic yield, and nutrient diffusion within a freely growing colony remains a challenge. Most available data on microorganisms are from planktonic cultures, due to the lack of experimental tools to control the growth of multicellular assemblies. Here, we propose a method to constrain the growth of yeast colonies into simple geometric shapes such as cylinders. To this end, we designed a simple, versatile culture system to control the location of nutrient delivery below a growing colony. Under such culture conditions, yeast colonies grow vertically and only at the locations where nutrients are delivered. Colonies increase in height at a steady growth rate that is inversely proportional to the cylinder radius. We show that the vertical growth rate of cylindrical colonies is not defined by the single-cell division rate, but rather by the colony metabolic yield. This contrasts with cells in liquid culture, in which the single-cell division rate is the only parameter that defines the population growth rate. This method also provides a direct, simple method to estimate the metabolic yield of a colony. Our study further demonstrates the importance of the shape of colonies on setting their expansion. We anticipate that our approach will be a starting point for elaborate studies of the population dynamics, evolution, and ecology of microbial colonies in complex landscapes.

PHEPS (pH-dependent Protein Electrostatics Server) is a web service for fast prediction and experiment planning support, as well as for correlation and analysis of experimentally obtained results, reflecting charge-dependent phenomena in globular proteins. Its implementation is based on long-term experience (PHEI package) and the need to explain measured physicochemical characteristics at the level of protein atomic structure. The approach is semi-empirical and based on a mean field scheme for description and evaluation of global and local pH-dependent electrostatic properties: protein proton binding; ionic sites proton population; free energy electrostatic term; ionic groups proton affinities (pK(a,i)) and their Coulomb interaction with whole charge multipole; electrostatic potential of whole molecule at fixed pH and pH-dependent local electrostatic potentials at user-defined set of points. The speed of calculation is based on fast determination of distance-dependent pair charge-charge interactions as empirical three exponential function that covers charge-charge, charge-dipole and dipole-dipole contributions. After atomic coordinates input, all standard parameters are used as defaults to facilitate non-experienced users. Special attention was given to interactive addition of non-polypeptide charges, extra ionizable groups with intrinsic pK(a)s or fixed ions. The output information is given as plain-text, readable by 'RasMol', 'Origin' and the like. The PHEPS server is accessible at http://pheps.orgchm.bas.bg/home.html.

Brain inflammation plays a key role in neurological disease. Although much research has been conducted investigating inflammatory events in animal models, potential differences in human brain versus rodent models makes it imperative that we also study these phenomena in human cells and tissue.

Skin is the largest human organ with easily noticeable biophysical manifestations of aging. As human tissues age, there is chronological accumulation of biophysical changes due to internal and environmental factors. Skin aging leads to decreased elasticity and the loss of dermal matrix integrity via degradation. The mechanical properties of the dermal matrix are maintained by fibroblasts, which undergo replicative aging and may reach senescence. While the secretory phenotype of senescent fibroblasts is well studied, little is known about changes in the fibroblasts biophysical phenotype. Therefore, we compare biophysical properties of young versus proliferatively aged primary fibroblasts via fluorescence and traction force microscopy, single-cell atomic force spectroscopy, microfluidics, and microrheology of the cytoskeleton. Results show senescent fibroblasts have decreased cytoskeletal tension and myosin II regulatory light chain phosphorylation, in addition to significant loss of traction force. The alteration of cellular forces is harmful to extracellular matrix homeostasis, while decreased cytoskeletal tension can amplify epigenetic changes involved in senescence. Further exploration and detection of these mechanical phenomena provide possibilities for previously unexplored pharmaceutical targets against aging.