Biomaterials have played an enormous role in the success of medical devices and drug delivery systems. We discuss here new challenges and directions in biomaterials research. These include synthetic replacements for biological tissues, designing materials for specific medical applications, and materials for new applications such as diagnostics and array technologies.

Designing materials for biology and medicine

BACKGROUND Living cells contain distinct subcompartments to facilitate spatiotemporal regulation of biological reactions. In addition to canonical membrane-bound organelles such as secretory vesicles and endoplasmic reticulum, there are many organelles that do not have an enclosing membrane yet remain coherent structures that can compartmentalize and concentrate specific sets of molecules. Examples include assemblies in the nucleus such as the nucleolus, Cajal bodies, and nuclear speckles and also cytoplasmic structures such as stress granules, P-bodies, and germ granules. These structures play diverse roles in various biological processes and are also increasingly implicated in protein aggregation diseases. ADVANCES A number of studies have shown that membrane-less assemblies exhibit remarkable liquid-like features. As with conventional liquids, they typically adopt round morphologies and coalesce into a single droplet upon contact with one another and also wet intracellular surfaces such as the nuclear envelope. Moreover, component molecules exhibit dynamic exchange with the surrounding nucleoplasm and cytoplasm. These findings together suggest that these structures represent liquid-phase condensates, which form via a biologically regulated (liquid-liquid) phase separation process. Liquid phase condensation increasingly appears to be a fundamental mechanism for organizing intracellular space. Consistent with this concept, several membrane-less organelles have been shown to exhibit a concentration threshold for assembly, a hallmark of phase separation. At the molecular level, weak, transient interactions between molecules with multivalent domains or intrinsically disordered regions (IDRs) are a driving force for phase separation. In cells, condensation of liquid-phase assemblies can be regulated by active processes, including transcription and various posttranslational modifications. The simplest physical picture of a homogeneous liquid phase is often not enough to capture the full complexity of intracellular condensates, which frequently exhibit heterogeneous multilayered structures with partially solid-like characters. However, recent studies have shown that multiple distinct liquid phases can coexist and give rise to richly structured droplet architectures determined by the relative liquid surface tensions. Moreover, solid-like phases can emerge from metastable liquid condensates via multiple routes of potentially both kinetic and thermodynamic origins, which has important implications for the role of intracellular liquids in protein aggregation pathologies. OUTLOOK The list of intracellular assemblies driven by liquid phase condensation is growing rapidly, but our understanding of their sequence-encoded biological function and dysfunction lags behind. Moreover, unlike equilibrium phases of nonliving matter, living cells are far from equilibrium, with intracellular condensates subject to various posttranslational regulation and other adenosine triphosphate–dependent biological activity. Efforts using in vitro reconstitution, combined with traditional cell biology approaches and quantitative biophysical tools, are required to elucidate how such nonequilibrium features of living cells control intracellular phase behavior. The functional consequences of forming liquid condensates are likely multifaceted and may include facilitated reaction, sequestration of specific factors, and organization of associated intracellular structures. Liquid phase condensation is particularly interesting in the nucleus, given the growing interest in the impact of nuclear phase behavior on the flow of genetic information; nuclear condensates range from micrometer-sized bodies such as the nucleolus to submicrometer structures such as transcriptional assemblies, all of which directly interact with and regulate the genome. Deepening our understanding of these intracellular states of matter not only will shed light on the basic biology of cellular organization but also may enable therapeutic intervention in protein aggregation disease by targeting intracellular phase behavior.

Liquid phase condensation in cell physiology and disease.

Considerations and Challenges in Studying Liquid-Liquid Phase Separation and Biomolecular Condensates.

Interest in thermoresponsive polymers has steadily grown over many decades, and a great deal of work has been dedicated to developing temperature sensitive macromolecules that can be crafted into new smart materials. However, the overwhelming majority of previously reported temperature-responsive polymers are based on poly(N-isopropylacrylamide) (PNIPAM), despite the fact that a wide range of other thermoresponsive polymers have demonstrated similar promise for the preparation of adaptive materials. Herein, we aim to highlight recent results that involve thermoresponsive systems that have not yet been as fully considered. Many of these (co)polymers represent clear opportunities for advancements in emerging biomedical and materials fields due to their increased biocompatibility and tuneable response. By highlighting recent examples of newly developed thermoresponsive polymer systems, we hope to promote the development of new generations of smart materials.

New directions in thermoresponsive polymers.

Elastin-like polypeptides (ELPs) undergo a reversible, inverse phase transition. Below their transition temperature (Tt), ELPs are soluble in water, but when the temperature is raised above Tt, phase transition occurs, leading to aggregation of the polypeptide. We demonstrate a method for purification of soluble fusion proteins incorporating an ELP tag. Advantages of this method, termed "inverse transition cycling," include technical simplicity, low cost, ease of scale-up, and capacity for multiplexing. More broadly, the ability to environmentally modulate the physicochemical properties of recombinant proteins by fusion with ELPs will allow diverse applications in bioseparation, immunoassays, biocatalysis, and drug delivery.

Purification of recombinant proteins by fusion with thermally-responsive polypeptides

Temperature of polypeptide inverse temperature transition depends on mean residue hydrophobicity

In general, proteins fold with hydrophobic residues buried, away from water. Reversible protein folding due to hydrophobic interactions results from inverse temperature transitions where folding occurs on raising the temperature. Because homoiothermic animals constitute an infinite heat reservoir, it is the transition temperature, Tt, not the endothermic heat of the transition, that determines the hydrophobically folded state of polypeptides at body temperature. Reported here is a new hydrophobicity scale based on the values of Tt for each amino acid residue as a guest in a natural repeating peptide sequence, the high polymers of which exhibit reversible inverse temperature transitions. Significantly, a number of ways have been demonstrated for changing Tt such that reversibly lowering Tt from above to below physiological temperature becomes a means of isothermally and reversibly driving hydrophobic folding. Accordingly, controlling Tt becomes a mechanism whereby proteins can be induced to carry out isothermal free energy transduction.

Hydrophobicity scale for proteins based on inverse temperature transitions

The complete series of the recommended generic biological tests for materials and devices in contact with tissues and tissue fluids and blood have been carried out by an independent testing laborat...

Michael C. Reid

Papers

Temperature of polypeptide inverse temperature transition depends on mean residue hydrophobicity

Hydrophobicity scale for proteins based on inverse temperature transitions

Biocompatibility of the Bioelastic Materials, Poly(GVGVP) and Its γ-Irradiation Cross-Linked Matrix: Summary of Generic Biological Test Results