Biological hydrogels as selective diffusion barriers

The controlled exchange of molecules between organelles, cells, or organisms and their environment is crucial for life. Biological gels such as mucus, the extracellular matrix (ECM), and the biopolymer barrier within the nuclear pore are well suited to achieve such a selective exchange, allowing passage of particular molecules while rejecting many others. Although hydrogel-based filters are integral parts of biology, clear concepts of how their barrier function is controlled at a microscopic level are still missing. We summarize here our current understanding of how selective filtering is established by different biopolymer-based hydrogels. We ask if the modulation of microscopic particle transport in biological hydrogels is based on a generic filtering principle which employs biochemical/biophysical interactions with the filtered molecules rather than size-exclusion effects.National Institutes of Health (U.S.) (Grant P50GM068763)MIT Start-up FundsGerman Academic Exchange Service (Postdoctoral Fellowship

Biophysical Properties of the Basal Lamina: A Highly Selective Extracellular Matrix

In this chapter, we discuss a specialized version of the extracellular matrix, the basal lamina. We focus on biophysical approaches which helped in identifying the mechanistic principles that allow the basal lamina to act as a selective permeability barrier. We discuss the physicochemical interactions that entail binding of molecules or nanoparticles to the basal lamina matrix and outline physiological scenarios where altered selective permeability properties of the basal lamina might contribute to physiological (mal) function

An adsorption chromatography assay to probe bulk particle transport through hydrogels

Biopolymer-based hydrogels such as mucus and the basal lamina play a key role in biology, where they control the exchange of material between different compartments. They also pose a barrier that needs to be overcome for successful drug delivery. Characterizing the permeability properties of such hydrogels is mandatory for the development of suitable drug delivery vectors and pharmaceutics. Here, we present an experimental method to measure bulk particle transport through hydrogels. We validate our assay by applying it to mucin hydrogels and show that the permeability properties of these mucin hydrogels can be modulated by polymer density and pH, in agreement with previous results obtained from single particle tracking. The method we present here is easy to handle, inexpensive, and high-throughput compatible. It is also a suitable platform for the design and screening of drugs that aim at modifying the barrier properties of hydrogels. This system can also aid in the characterization and development of synthetic gels for a range of biomedical applications.National Institutes of Health (U.S.) (Grant P50-GM068763)National Institutes of Health (U.S.) (Grant P30-ES002109

Micro- and Macrorheological Properties of Isotropically Cross-linked Actin Networks

Cells make use of semi-flexible biopolymers such as actin or intermediate filaments to control their local viscoelastic response by dynamically adjusting the concentration and type of cross-linker molecules. The microstructure of the resulting networks mainly determines their mechanical properties. It remains an important challenge to relate structural transitions to both the molecular properties of the cross-linking molecules and the mechanical response of the network. This can be achieved best by well-defined in vitro model systems in combination with microscopic techniques. Here, we show that with increasing concentrations of the cross-linker HMM (heavy meromyosin) a transition in the mechanical network response occurs. At low cross-linker densities the network elasticity is dominated by the entanglement length of the polymer, while at high HMM densities the cross-linker distance determines the elastic behavior. Using microrheology the formation of heterogeneous networks is observed at low cross-linker concentrations. Micro- and macrorheology both report the same transition to a homogeneous cross-linked phase. This transition is set by a constant average cross-linker distance. Thus, the micro- and macromechanical properties of isotropically cross-linked in vitro actin networks are determined by only one intrinsic network parameter.Comment: 14 pages, 5 figure

The Biophysical Properties of Basal Lamina Gels Depend on the Biochemical Composition of the Gel

The migration of cells within a three-dimensional extracellular matrix (ECM) depends sensitively on the biochemical and biophysical properties of the matrix. An example for a biological ECM is given by reconstituted basal lamina gels purified from the Engelbreth-HolmSwarm sarcoma of mice. Here, we compare four different commercial variants of this ECM, which have all been purified according to the same protocol. Nevertheless, in those gels, we detect strong differences in the migration behavior of leukocyte cells as well as in the Brownian motion of nanoparticles. We show that these differences correlate with the mechanical properties and the microarchitecture of the gels which in turn arise from small variations in their biochemical composition

Mechanical robustness of Pseudomonas aeruginosa biofilms

Biofilms grow on various surfaces and in many different environments, a phenomenon that constitutes major problems in industry and medicine. Despite their importance little is known about the viscoelastic properties of biofilms and how these depend on the chemical microenvironment. Here, we find that the mechanical properties of Pseudomonas aeruginosa (P.a.) biofilms are highly robust towards chemical perturbations. Specifically, we observe that P.a. biofilms are able to fully regain their initial stiffness after yielding is enforced, even in the presence of chemicals. Moreover, only trivalent ions and citric acid significantly affect the biofilm elasticity, the first of which also alters the texture of the material. Finally, our results indicate that biofilm mechanics and bacteria viability inside the biofilm are not necessarily linked which suggests that targeting bacteria alone might not be sufficient for biofilm removal strategies.National Institute of Mental Health (U.S.) (P50-GM068763)National Institute of Mental Health (U.S.) (P30-ES002109)German Academic Exchange Service (DAAD

Rheological Characterization of the Bundling Transition in F-Actin Solutions Induced by Methylcellulose

In many in vitro experiments Brownian motion hampers quantitative data analysis. Therefore, additives are widely used to increase the solvent viscosity. For this purpose, methylcellulose (MC) has been proven highly effective as already small concentrations can significantly slow down diffusive processes. Beside this advantage, it has already been reported that high MC concentrations can alter the microstructure of polymer solutions such as filamentous actin. However, it remains to be shown to what extent the mechanical properties of a composite actin/MC gel depend on the MC concentration. In particular, significant alterations might occur even if the microstructure seems unaffected. Indeed, we find that the viscoelastic response of entangled F-actin solutions depends sensitively on the amount of MC added. At concentrations higher than 0.2% (w/v) MC, actin filaments are reorganized into bundles which drastically changes the viscoelastic response. At small MC concentrations the impact of MC is more subtle: the two constituents, actin and MC, contribute in an additive way to the mechanical response of the composite material. As a consequence, the effect of methylcellulose on actin solutions has to be considered very carefully when MC is used in biochemical experiments

Cervical Mucus Properties Stratify Risk for Preterm Birth

Background: Ascending infection from the colonized vagina to the normally sterile intrauterine cavity is a well-documented cause of preterm birth. The primary physical barrier to microbial ascension is the cervical canal, which is filled with a dense and protective mucus plug. Despite its central role in separating the vaginal from the intrauterine tract, the barrier properties of cervical mucus have not been studied in preterm birth. Methods and Findings: To study the protective function of the cervical mucus in preterm birth we performed a pilot case-control study to measure the viscoelasticity and permeability properties of mucus obtained from pregnant women at high-risk and low-risk for preterm birth. Using extensional and shear rheology we found that cervical mucus from women at high-risk for preterm birth was more extensible and forms significantly weaker gels compared to cervical mucus from women at low-risk of preterm birth. Moreover, permeability measurements using fluorescent microbeads show that high-risk mucus was more permeable compared with low-risk mucus. Conclusions: Our findings suggest that critical biophysical barrier properties of cervical mucus in women at high-risk for preterm birth are compromised compared to women with healthy pregnancy. We hypothesize that impaired barrier properties of cervical mucus could contribute to increased rates of intrauterine infection seen in women with preterm birth. We furthermore suggest that a robust association of spinnbarkeit and preterm birth could be an effectively exploited biomarker for preterm birth prediction.Massachusetts Institute of Technology. Charles E. Reed Faculty Initiative FundBurroughs Wellcome Fund (Preterm Birth Research Grant)National Science Foundation (U.S.). Graduate Research Fellowship Progra